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TECHNICAL REPORTS SERIES No. 159

Californium-252 in Teaching and Research

E.J. Hall and H.H. Rossi

\ S f J I N T E R N A T I O N A L A T O M I C ENERGY A G E N C Y . V I E N N A , 1974

CALIFORNIUM-25 2 IN TEACHING AND RESEARCH

The f o l l o w i n g States are Members of the International A t o m i c Energy A g e n c y :

AFGHANISTAN HAITI PAKISTAN ALBANIA HOLY SEE PANAMA ALGERIA HUNGARY PARAGUAY ARGENTINA ICELAND PERU AUSTRALIA INDIA PHILIPPINES AUSTRIA INDONESIA POLAND BANGLADESH IRAN PORTUGAL BELGIUM IRAQ ROMANIA BOLIVIA IRELAND SAUDI ARABIA BRAZIL ISRAEL SENEGAL BULGARIA ITALY SIERRA LEONE BURMA IVORY C O A S T SINGAPORE BYELORUSSIAN SOVIET JAMAICA SOUTH AFRICA

SOCIALIST REPUBLIC JAPAN SPAIN CAMEROON JORDAN SRI LANKA C A N A D A KENYA SUDAN CHILE KHMER REPUBLIC SWEDEN COLOMBIA KOREA, REPUBLIC OF SWITZERLAND COSTA RICA KUWAIT SYRIAN ARAB REPUBLIC CUBA LEBANON THAILAND CYPRUS LIBERIA TUNISIA CZECHOSLOVAK SOCIALIST LIBYAN ARAB REPUBLIC TURKEY

REPUBLIC LIECHTENSTEIN UGANDA DENMARK LUXEMBOURG UKRAINIAN SOVIET SOCIALIST DOMINICAN REPUBLIC MADAGASCAR REPUBLIC ECUADOR MALAYSIA UNION OF SOVIET SOCIALIST EGYPT, ARAB REPUBLIC OF MALI REPUBLICS EL SALVADOR MEXICO UNITED KINGDOM OF GREAT ETHIOPIA MONACO BRITAIN AND NORTHERN FINLAND MONGOLIA IRELAND FRANCE MOROCCO UNITED STATES OF AMERICA GABON NETHERLANDS URUGUAY GERMAN DEMOCRATIC REPUBLIC NEW ZEALAND VENEZUELA GERMANY, FEDERAL REPUBLIC OF NIGER VIET-NAM GHANA NIGERIA YUGOSLAVIA GREECE NORWAY ZAIRE, REPUBLIC OF GUATEMALA ZAMBIA

The A g e n c y ' s Statute was approved on 23 October 1956 by the C o n f e r e n c e on the Statute of the IAEA he ld at Uni ted Nat ions Headquarters , N e w York; i t entered into force on 2 9 July 1 9 5 7 . The Headquarters of the A g e n c y are s i tuated in V i e n n a . Its principal o b j e c t i v e is "to a c c e l e r a t e and e n l a r g e the contr ibut ion of a t o m i c energy to p e a c e , h e a l t h and prosperity throughout the world".

( C ) I A E A , 1 9 7 4

Permiss ion to reproduce or translate the i n f o r m a t i o n c o n t a i n e d in this p u b l i c a t i o n m a y be obta ined by writ ing to the Internat ional A t o m i c Energy A g e n c y , Kärntner Ring 1 1 , P . O . Box 5 9 0 , A - 1 0 1 1 V i e n n a , Austria.

Printed by the IAEA in Austria August 1974

T E C H N I C A L R E P O R T S SERIES No. 159

CALIFORNIUM-252 IN TEACHING AND RESEARCH

E . J . H a l l a n d H . H . R o s s i

Rad io log ica l R e s e a r c h L a b o r a t o r y , Col lege of P h y s i c i a n s and Surgeons of

Columbia Un ive r s i t y , New York , N. Y. , United S ta tes of A m e r i c a

INTERNATIONAL ATOMIC ENERGY AGENCY VIENNA, 1974

CALIFORNIUM-252 IN TEACHING AND RESEARCH IAEA, VIENNA, 1974

S T I / D O C / 1 0 / 1 5 9

IS BN-9 2 -0 -1151 74-8

FOREWORD

In h i s opening s t a t e m e n t at the f i f t een th G e n e r a l C o n f e r e n c e of the IAEA held in Vienna in 1971, D r . Seaborg, then c h a i r m a n of the USAEC, announced the o f f e r of the United Sta tes to lend c a l i f o r n i u m - 2 5 2 s o u r c e s to the IAEA. The p u r p o s e of th i s loan was to m a k e neu t ron s o u r c e s ava i l ab le to u n i v e r s i t i e s of the M e m b e r S ta tes f o r u s e in educa t iona l p r o g r a m m e s . The s o u r c e s , in the f o r m of s m a l l need le s des igned f o r m e d i c a l u s e in r ad i a t i on t h e r a p y , w e r e judged highly su i tab le f o r d idac t ic app l i ca t ions b e c a u s e of t h e i r s m a l l s ize , l imi ted ac t iv i ty and wel l documented r a d i o l o g i c a l p a r a m e t e r s .

In May 1973 the D i r e c t o r Gene ra l of the IAEA, D r . Sigvard Eklund, announced the ava i l ab i l i ty of the ca l i fo rn ium s o u r c e s to the M e m b e r S ta tes . To da te , n u m e r o u s s o u r c e s have been lent to u n i v e r s i t i e s in Cos t a Rica , Ghana, India, Uruguay , J a p a n , I s r a e l , the United Kingdom, the F e d e r a l Republ ic of G e r m a n y , Swi tze r land , the Czechos lovak Soc ia l i s t Republ ic and South A f r i c a , and addi t iona l appl ica t ions f o r loan a r e be ing p r o c e s s e d . It i s an t i c ipa ted that the loan p r o g r a m m e will be t e r m i n a t e d in 1975 once a l l the ava i l ab le s o u r c e s have been d i s t r ibu ted .

It h a s b e c o m e c l e a r that m a n y u n i v e r s i t i e s needed guidance in the f o r m of an i n s t r u c t i o n a l sy l l abus , a l a b o r a t o r y m a n u a l f o r e x p e r i m e n t s u t i l i z ing the s o u r c e s and f o r the s a f e ty p r e c a u t i o n s inhe ren t in the p r o p e r handl ing of n e u t r o n - e m i t t i n g r ad ionuc l i de s . P r o f e s s o r s E . J . Hall and H . H . R o s s i of Columbia Un ive r s i t y , New York , g r ac ious ly under took the t a sk of wr i t i ng such a book on behalf of the IAEA. Although s o m e i n f o r m a t i o n on the educa t iona l u s e s of c a l i f o rn ium has p r e v i o u s l y been r e p o r t e d (e. g. ' P r o g r a m to eva lua te the educa t iona l u s e s of a c a l i f o r n i u m - 2 5 2 s o u r c e in a n u c l e a r s c i ence and eng inee r ing p r o g r a m ' by P r o f e s s o r G. E ichho lz , Georg ia Ins t i tu te of Technology , TID-25752, 1970), t h e r e h a s been no expos i t ion of the d o s i m e t r i c , r ad iob io log ica l and m i c r o d o s i m e t r i c s u b j e c t s which a r e of such g r e a t c u r r e n t i n t e r e s t . The p r e s e n t publ ica t ion con-s t i t u t e s a s ign i f ican t unde r t ak ing of c r i t i c a l ins ight into a d i f f icu l t sub jec t by two e x p e r t s who have been r e s p o n s i b l e f o r m a n y a d v a n c e s in the f i e l d s of neu t ron d o s i m e t r y , m i c r o d o s i m e t r y and radiobiology.

Guidance on the c o n s t r u c t i o n of a p ro to type u s e and s t o r a g e f ac i l i t y i s p rov ided in an appendix. Although each u s e r m u s t b e a r the r e s p o n s i b i l i t y f o r des igning and c o n s t r u c t i n g h i s own fac i l i ty , the appendix m a y s e r v e to p rov ide u s e f u l i n f o r m a t i o n on th i s impor t an t f i r s t s tep . The con ta ine r was des igned by the staff of the D o s i m e t r y Sect ion of the IAEA ' s Div is ion of L i fe Sc iences , n a m e l y R. Abed in -Zadeh , H . H . E i s e n l o h r (Section Head) , J . H a i d e r and I . A . L e r c h (Cf-252 P r o j e c t Leade r ) and cons t ruc t ed in the A g e n c y ' s Se ibe r sdo r f workshop .

CONTENTS

P A R T 1: INSTRUCTIONAL SYLLABUS

C h a p t e r 1 Ob jec t i ve s and scope 3 C h a p t e r 2 Rad ia t ions and t h e i r i n t e r ac t i ons 5 C h a p t e r 3 D o s i m e t r y 11 C h a p t e r 4 Radia t ion qual i ty and m i c rodo s i m e t r y 23 C h a p t e r 5 Bio log ica l t e s t s y s t e m s 27 C h a p t e r 6 Re la t ive b io log ica l e f f e c t i v e n e s s 35 C h a p t e r 7 E f f e c t of m o l e c u l a r oxygen 41 C h a p t e r 8 Cel l age and r ad io sens i t i v i t y 47 C h a p t e r 9 R e p a i r of sub le tha l damage and the d o s e - r a t e e f f ec t . . . 53 C h a p t e r 10 La te e f f e c t s 59 Chapter; 11 Radia t ion p r o t e c t i o n 65

P A R T 2: LABORATORY MANUAL

C h a p t e r 12 G e n e r a l i n f o r m a t i o n f o r l a b o r a t o r y e x p e r i m e n t s 71 C h a p t e r 13 E x p e r i m e n t s with a GM counter 73 Chap t e r 14 E x p e r i m e n t s with l a r g e ioniza t ion c h a m b e r s 77 C h a p t e r 15 Advanced e x p e r i m e n t s in d o s i m e t r y and

m i c r o d o s i m e t r y 83 C h a p t e r 16 E x p e r i m e n t s with T r a d e s c a n t i a 85 C h a p t e r 17 E x p e r i m e n t s with r o o t m e r i s t e m s 91 C h a p t e r 18 M a m m a l i a n c e l l s in cu l tu re 101 C h a p t e r 19 R e s p o n s e of m o u s e j e juna l c r y p t c e l l s 109 C h a p t e r 20 Rad ia t ion - induced m a m m a r y t u m o u r s in the r a t I l l

Appendix I P h y s i c a l c h a r a c t e r i s t i c s of c a l i f o r n i u m - 2 5 2 115 Appendix II D e s c r i p t i o n of c a l i f o r n i u m - 2 5 2 s o u r c e s and

the p ro to type use and s t o r a g e fac i l i ty 119

R e f e r e n c e s 133 Bib l iography 137

P a r t 1

INSTRUCTIONAL SYLLABUS

C h a p t e r 1

O B J E C T I V E S AND SCOPE

1 .1 USE O F CALIFORNIUM-252

This publ ica t ion has been p r e p a r e d to p r o m o t e the s a f e in t roduct ion of the m a n - m a d e nucl ide c a l i f o r n i u m - 2 5 2 into the teaching p r o g r a m s of u n i v e r s i t i e s and r e s e a r c h i n s t i t u t e s in M e m b e r S ta tes of the In te rna t iona l A tomic E n e r g y Agency. The m o s t i m p o r t a n t f e a t u r e of t h i s nucl ide is tha t it r e p r e s e n t s a p o r t a b l e s o u r c e of n e u t r o n s . Teach ing d e p a r t m e n t s and r e s e a r c h g roups tha t do not have a c c e s s to l a r g e and expens ive a c c e l e r a t o r f a c i l i t i e s can now extend t h e i r scope to include n e u t r o n s because of the ava i lab i l i ty of c a l i f o r n i u m - 2 52 p roduced by the United S ta tes Atomic E n e r g y C o m m i s s i o n .

The u s e of any ionizing r a d i a t i o n inevi tably involves the poss ib i l i ty of a h a z a r d . T h e r e is i n c r e a s i n g evidence ind ica t ing tha t s m a l l d o s e s of neu-t r o n s a r e ve ry m u c h m o r e h a z a r d o u s than s m a l l d o s e s of X - r a y s , and so it i s i m p e r a t i v e tha t any po ten t i a l u s e r of c a l i f o rn ium-252 be ful ly acquain ted with the phys ica l and rad iob io log ica l p r o p e r t i e s of the nucl ide , and educa ted conce rn ing the need f o r adequate p r o t e c t i o n m e a s u r e s .

1. 2 SYLLABUS AND MANUAL

The book c o n s i s t s e s s e n t i a l l y of two p a r t s . The f i r s t ou t l ines an i n s t r u c t i o n a l sy l labus or c o u r s e of study which, if fol lowed, wil l p rov ide the r e a d e r with a bas i c u n d e r s t a n d i n g of r ad i a t i on p h y s i c s , and me thods of m e a s u r i n g the quant i ty and qual i ty of d i f f e r en t t ypes of r ad ia t ion . In addi t ion, the biological e f f e c t s of r ad i a t i on a r e r ev i ewed . P a r t i c u l a r e m p h a s i s i s p laced upon p r o t e c t i o n c o n s i d e r a t i o n s , which a r e unusual ly complex in the c a s e of c a l i f o r n i u m - 2 5 2 , s ince the e m i s s i o n f r o m t h i s nucl ide i s a m i x t u r e of g a m m a r a y s and n e u t r o n s .

The second p a r t of the book i s a l a b o r a t o r y m a n u a l d e s c r i b i n g a r a n g e of e x p e r i m e n t s tha t m a y be p e r f o r m e d in p h y s i c s and in radiobiology. In the d e s c r i p t i o n of the e x p e r i m e n t s , the de t a i l s given a r e a s c o m p r e h e n s i v e and a s fu l l a s i s p r a c t i c a l in a publ ica t ion of t h i s s o r t . The r a n g e of the e x p e r i m e n t s d i s c u s s e d i s so wide tha t the a u t h o r s have not been able to c a r r y out eve ry e x p e r i m e n t d e s c r i b e d . A publ ica t ion at t h i s level cannot hope to be a s comple te and a s c o m p r e h e n s i v e a s , f o r example , a high-school l a b o r a t o r y m a n u a l d e s c r i b i n g e x p e r i m e n t s in c l a s s i c a l phys i c s . The a im of the manua l i s to guide p o s t g r a d u a t e u n i v e r s i t y s tuden t s in the s a f e conduct of i n s t ruc t i ve e x p e r i m e n t s involving th i s mixed g a m m a - r a y and neu t ron e m i t t e r .

In addit ion to the two m a i n p a r t s , the book a l so h a s two appendices : Appendix I g ives the phys ica l c h a r a c t e r i s t i c s of c a l i f o rn ium-252 , and Appendix II d e s c r i b e s the s o u r c e s and gives guidance on the cons t ruc t ion of a p ro to type use and s t o r a g e fac i l i ty .

3

4 CHAPTER 2

1 . 3 PROBLEMS AND LIMITATIONS

T h e r e have been two m a j o r p r o b l e m s in the p r e p a r a t i o n of the m a n u a l . F i r s t , t he c a l i f o r n i u m - 2 52 s o u r c e s ava i lab le unde r the In te rna t iona l Atomic E n e r g y Agency loan p r o g r a m a r e l imi t ed both in n u m b e r and in r ad ioac t i ve content . F o r the m o s t p a r t they a r e s o u r c e s which a r e not su i tab le f o r human c l in ica l app l i ca t ions , e i the r because they a r e of an obso le te design, o r b e c a u s e they have decayed to too low a r a d i o a c t i v e content . The r a d i o -b io log ica l e x p e r i m e n t s , in p a r t i c u l a r , a r e s e v e r e l y r e s t r i c t e d because of t he low d o s e - r a t e s ava i l ab le f r o m t h e s e s o u r c e s . Second, t he level of s c i en t i f i c sophis t i ca t ion , a s wel l as the ava i lab i l i ty of complex equipment , v a r i e s g r ea t l y amongs t t he v a r i o u s ins t i tu t ions that have e x p r e s s e d an i n t e r e s t to p a r t i c i p a t e in the ca l i f o rn ium-2 52 loan p r o g r a m .

B e c a u s e of t h e s e f a c t o r s , both the sy l l abus and l a b o r a t o r y m a n u a l c a t e r to a wide r a n g e of i n t e r e s t s . This i s at once i t s s t r eng th and i t s w e a k n e s s . F o r d e p a r t m e n t s with an advanced p r o g r a m in r ad i a t i on phys i c s and r ad ia t ion biology, s o m e of t he conten ts of the book wil l be redundant , and many of t he e x p e r i m e n t s d e s c r i b e d wil l be too s imp le . F o r o ther ins t i tu t ions which have not a s ye t developed such p r o g r a m s in r a d i a t i o n phys i c s and radiobio logy, o r w h e r e l a b o r a t o r y f a c i l i t i e s a r e l imi ted , some of the e x p e r i m e n t s d e s c r i b e d m a y be i m p o s s i b l e to c a r r y out.

The c o v e r a g e of r ad io log ica l phys i c s and r ad i a t i on p ro t ec t i on f o c u s e s on f i r s t p r i n c i p l e s and on m a t e r i a l not eas i ly a c c e s s i b l e in publ ished f o r m . V a r i o u s r e p o r t s of the In te rna t iona l C o m m i s s i o n on Radia t ion Uni t s and M e a s u r e m e n t s which a r e r ead i l y ava i lab le and r a t h e r inexpens ive should be consu l ted by both i n s t r u c t o r s and s tuden ts . T h e s e and o ther key w o r k s a r e given in the R e f e r e n c e s and Bib l iography at the end of the book.

C h a p t e r 2

RADIATIONS AND THEIR INTERACTIONS

2 . 1 ELECTROMAGNETIC

Unti l r e c e n t y e a r s m o s t r a d i a t i o n e x p e r i m e n t s with b io logica l s y s t e m s have involved X or g a m m a r a y s . T h e s e a r e f o r m s of e l e c t r o m a g n e t i c r ad i a t ion . X and g a m m a r a y s do not d i f f e r f r o m one ano ther in n a t u r e o r in p r o p e r t i e s ; t h e i r des igna t ion r e f l e c t s m e r e l y the way in which they o r ig ina te . X - r a y s a r e p roduced e x t r a n u c l e a r l y , and g a m m a r a y s a r e p roduced i n t r a -n u c l e a r ly. In p r a c t i c a l t e r m s , t h i s m e a n s tha t X - r a y s a r e g e n e r a t e d in an e l e c t r i c a l device which a c c e l e r a t e s e l e c t r o n s to high energy and then s tops t h e m abrup t ly in a t a r g e t , usua l ly m a d e of tungs ten . P a r t of the ki -ne t i c ene rgy of the e l e c t r o n s i s conver ted into X - r a y s . G a m m a r a y s , on the o the r hand, a r e emi t t ed by r a d i o a c t i v e i so topes ; they r e p r e s e n t e x c e s s ene rgy which i s given off as uns t ab le nuc le i decay into m o r e s t ab le f o r m s .

E l e c t r o m a g n e t i c r ad i a t i on can be c o n s i d e r e d to cons i s t of waves o r of p a r t i c l e s . The p a r t i c l e s a r e t e r m e d photons . The r e l a t i o n between wave-length X and photon ene rgy E i s

X = (1 /E) X 12 .4 X 10"7

if X is in m e t r e s (m) and E in e l e c t r o n - v o l t s (eV). The d i f f e r e n c e between ioniz ing and non - ion i z ing -e l ec t romagne t i c r a d i a -

t ions i s the ene rgy of the individual photons . As f a r a s biological e f f e c t s a r e conce rned , e l e c t r o m a g n e t i c r a d i a t i o n s a r e usual ly cons ide red to be ionizing if the photon energy i s in e x c e s s of about 125 eV, which c o r r e s p o n d s to a wavelength s h o r t e r than about lO"10 m .

2 . 2 CHARGED PARTICLES

C h a r g e d p a r t i c l e r a d i a t i o n s u s e d expe r imen ta l l y include e l e c t r o n s , p ro tons , a lpha p a r t i c l e s , nega t ive pi m e s o n s , and heavy ions.

E l e c t r o n s a r e negat ive ly cha rged p a r t i c l e s which can be a c c e l e r a t e d to high ene rgy , and to a speed c lose to tha t of l ight, by m e a n s of an e l e c t r i c a l device such a s a b e t a t r o n .

P r o t o n s a r e pos i t ive ly cha rged p a r t i c l e s having a m a s s a l m o s t two thousand t i m e s tha t of an e l ec t ron . B e c a u s e of t h e i r m a s s they r e q u i r e m o r e complex and m o r e expens ive equipment fo r a c c e l e r a t i o n to u s e f u l e n e r g i e s .

Alpha p a r t i c l e s a r e nuc le i of he l ium a t o m s and cons i s t of two p ro tons and two n e u t r o n s in c lose a s soc ia t ion . They have a net pos i t ive c h a r g e and can t h e r e f o r e be a c c e l e r a t e d in l a r g e e l e c t r i c a l dev ices s i m i l a r to t hose used fo r p r o t o n s , but they a r e a l so emi t t ed dur ing the decay of some r a d i o -ac t ive i so topes .

Heavy cha rged ions a r e nuc le i of e l e m e n t s such a s n i t rogen , ca rbon , boron, neon o r a rgon which a r e pos i t ive ly cha rged s ince s o m e o r a l l of t h e i r p l a n e t a r y e l e c t r o n s have been s t r i p p e d f r o m t h e m . To be u s e f u l in r a d i o -biology they m u s t be a c c e l e r a t e d to e n e r g i e s of hundreds or thousands of m i l l i ons of vo l t s , and they can t h e r e f o r e be p roduced in only a v e r y l imi ted n u m b e r of l a b o r a t o r i e s .

5

6 CHAPTER 2

2 . 3 NEUTRONS

Neu t rons a r e p a r t i c l e s with a m a s s s i m i l a r to that of p ro tons , but they c a r r y no e l e c t r i c a l cha rge . B e c a u s e they a r e e l e c t r i c a l l y n e u t r a l they cannot be a c c e l e r a t e d in an e l e c t r i c a l device . They a r e p roduced when a charged p a r t i c l e , such a s a deu te ron , is a c c e l e r a t e d to high energy and then m a d e to impinge upon a su i tab le t a r g e t m a t e r i a l . Neu t rons a r e a l so emi t ted a s a by-product when heavy nucle i undergo f i s s ion , i. e. spl i t up to f o r m two s m a l l e r nuc le i . In p r a c t i c a l t e r m s t h e r e a r e t h r e e u s e f u l s o u r c e s of n e u t r o n s .

F i s s i o n n e u t r o n s . N e u t r o n s with a wide r a n g e of e n e r g i e s a r e p roduced in abundance ins ide n u c l e a r r e a c t o r s by the f i s s i o n of u r a n i u m - 2 3 5 . Some nuc le i , in p a r t i c u l a r ca l i fo rn ium-252 , undergo spontaneous f i s s i o n and s m a l l s ea l ed tubes containing t h i s nucl ide cons t i tu te a convenient po r t ab l e s o u r c e of n e u t r o n s . However , they a l so emi t g a m m a r a d i a t i o n tha t o r i g i n a t e s in the f i s s i o n p r o d u c t s . F i s s i o n n e u t r o n s have a wide r a n g e of e n e r g i e s a round 1 MeV.

Cyc lo t ron-produced neu t rons . A cyc lo t ron i s an e l e c t r i c a l device capable of a c c e l e r a t i n g pos i t ive ly charged p a r t i c l e s , such a s deu t e rons , to an energy of many mi l l i ons of vo l t s . If the e n e r g e t i c deu te ron i s m a d e to impinge on a t a r g e t of be ry l l i um, t h e p r o t o n i s ' s t r i pped ' f r o m the deu te ron , leaving a neu t ron which c a r r i e s much of the kinet ic ene rgy of the o r ig ina l deuteron . F o r example , if d e u t e r o n s a r e a c c e l e r a t e d to 50 MeV and m a d e to impinge upon a be ry l l i um t a r g e t , a s p e c t r u m of n e u t r o n s i s p roduced which has a m o d a l ene rgy of about 25 MeV. Cyc lo t rons a r e cos t ly to build and only a l imi t ed n u m b e r a r e ava i lab le .

14-MeV d-»T n e u t r o n s . If a beam of deu t e rons , a c c e l e r a t e d to an energy of t he o r d e r of 300 keV, i s incident on a t r i t i u m t a r g e t , n e u t r o n s a r e p r o -duced which a r e e s s en t i a l l y monoene rge t i c . The r e a c t i o n i s d(t, n)4He. The energy of t he neu t ron produced (about 14 MeV) i s much g r e a t e r than the k ine t ic ene rgy of the a c c e l e r a t e d deu t e ron . Th i s e x t r a ene rgy i s ava i lab le b e c a u s e the i n t e r n a l binding energy of the he l ium nuc leus i s l e s s t han that of the i n t e r a c t i n g p a r t i c l e s . This i s a v e r y a t t r a c t i v e method f o r gene ra t ing n e u t r o n s , b e c a u s e it i s r e l a t i v e l y inexpens ive and involves a r e l a t i v e l y s m a l l mach ine . At the p r e s e n t t i m e the s e r i o u s d rawback of th is me thod of neu t ron p roduc t ion i s the low d o s e - r a t e , that i s l i m i t e d b e c a u s e of t echn ica l d i f f i cu l t i e s a s s o c i a t e d with the cons t ruc t ion of t a r g e t s capable of r e t a in ing t r i t i u m u n d e r in t ense b e a m s .

2 . 4 ABSORPTION O F RADIATION

Radia t ions a r e usua l ly c l a s s i f i e d a s d i r ec t l y o r ind i rec t ly ionizing. Al l of the cha rged p a r t i c l e s l i s t ed above a r e d i r ec t l y ionizing; tha t i s , p r o -vided they have suf f ic ien t k ine t ic energy , they can d i r ec t l y d i s rup t the a tomic s t r u c t u r e of the m a t t e r t h rough which they p a s s , and p roduce phys ica l , chemica l and biological changes . E l e c t r o m a g n e t i c r a d i a t i o n s (X and g a m m a r a y s ) , t o g e t h e r with n e u t r o n s a r e ind i rec t ly ionizing. When they a r e ab so rbed they t r a n s f e r ene rgy to p r o d u c e f a s t - m o v i n g s e c o n d a r y cha rged p a r t i c l e s .

RADIATIONS AND THEIR INTERACTIONS 7

Since g a m m a r a y s and n e u t r o n s a r e the p r i n c i p a l r a d i a t i o n s emi t t ed by en-capsu la ted ca l i fo rn ium-252 s o u r c e s , t h e i r i n t e r a c t i o n s wil l be d e s c r i b e d m o r e fu l ly .

The p r o c e s s by which g a m m a - r a y photons a r e a b s o r b e d depends upon the ene rgy of the photons concerned , and upon the chemica l compos i t ion of the a b s o r b i n g m a t e r i a l . A fu l l d e s c r i p t i o n can be found in any bas i c r ad io log ica l p h y s i c s t e s t (Johns 1964). F o r p r e s e n t p u r p o s e s , however , we wil l cons ide r t he only impor t an t p r o c e s s (the Compton effect ) which o c c u r s when g a m m a r a y s of about 1 MeV a r e abso rbed by b io log ica l m a t e r i a l .

In the Compton e f fec t the g a m m a - r a y photon i n t e r á c t s with an o r b i t a l e l e c t r o n of one of t he a t o m s of the a b s o r b e r . P a r t of the photon ene rgy i s given to the e l e c t r o n a s k ine t ic ene rgy ; t he photon, with r e d u c e d ene rgy , cont inues on i t s way, usua l ly de f lec ted f r o m i t s o r ig ina l d i r ec t ion . In p l ace of the inc ident photon, t h e r e i s now a f a s t e l e c t r o n and a photon of r e d u c e d energy , which may go on to t ake p a r t in f u r t h e r i n t e r a c t i o n s . When g a m m a r a y s i n t e r a c t with t i s s u e , the net r e s u l t i s the p roduc t ion of a l a r g e n u m b e r of f a s t e l e c t r o n s , many of which have the abi l i ty to ionize a t o m s of the ab-s o r b e r , b r e a k c h e m i c a l bonds , and in i t i a te a chain of events which u l t ima te ly l eads to b io log ica l damage .

F o r n e u t r o n s , too, the m e c h a n i s m of abso rp t i on depends on e n e r g y . F o r f a s t n e u t r o n s , in the energy r a n g e f r o m 100 keV to 20 MeV, e l a s t i c s c a t t e r i n g i s the dominant p r o c e s s . The incident neu t ron co l l ides with the nuc leus of an a tom of the a b s o r b e r ; p a r t of i t s ene rgy i s given to the nuc leus involved, and the r e m a i n d e r i s r e t a i n e d by the de f lec ted neu t ron , which m a y u n d e r g o f u r t h e r i n t e r a c t i o n s .

When n e u t r o n s f r o m ca l i fo rn ium-252 a r e ab so rbed in sof t t i s s u e s , the i n t e r a c t i o n between incident n e u t r o n s and hydrogen nucle i , which a r e of c o u r s e s ingle p ro tons , i s the dominant p r o c e s s of energy t r a n s f e r . Hydro-gen i s t he m o s t abundant a tom in sof t t i s s u e , and the co l l i s ion c r o s s - s e c t i o n fo r hydrogen i s l a r g e . The r e c o i l p r o t o n s se t in mot ion in th i s way l o s e ene rgy by exci ta t ion and ionizat ion a s they p a s s through the biological m a -t e r i a l . Co l l i s ions with o ther nuc le i (oxygen, ca rbon , e t c . ) a l so r e s u l t in r e c o i l s which, however , c a r r y only a s m a l l f r a c t i o n of the ene rgy abso rbed .

In each e l a s t i c col l i s ion n e u t r o n s lo se typica l ly half t h e i r energy , with the r e s u l t t ha t a f t e r s o m e 20 co l l i s ions the ene rgy is of the o r d e r of a few e l e c t r o n - v o l t s . In t h i s r ange two c a p t u r e p r o c e s s e s become i m p o r t a n t . In one of t h e s e , a n i t r ogen nuc leus c a p t u r e s the neu t ron and e m i t s a p ro ton having an ene rgy of about 700 keV. In the o the r cap tu re r e a c t i o n the neu t ron combines with a p ro ton and a p e n e t r a t i n g (2. 2 MeV) g a m m a photon i s emi t t ed . B e c a u s e of t h i s p r o c e s s , t i s s u e exposed to an uncon tamina ted neu t ron beam i s n e v e r t h e l e s s exposed to g a m m a r a d i a t i o n a s wel l . However , in m o s t i n s t a n c e s the b iological e f fec t of t h i s r a d i a t i o n i s m i n o r .

To sum up the p r o c e s s e s of abso rp t ion : g a m m a r a y photons i n t e r a c t with the o rb i t a l e l e c t r o n s of a t o m s of t he a b s o r b i n g m a t e r i a l , and se t in mot ion f a s t e l e c t r o n s . Neu t rons i n t e r a c t with the nuc le i of a t o m s of the a b s o r b i n g m a t e r i a l , and in the m o s t i m p o r t a n t p r o c e s s se t in mot ion f a s t r e c o i l p ro tons . E l e c t r o n s and p ro tons both c a r r y unit e l e c t r i c a l c h a r g e , but d i f f e r in m a s s by a f a c t o r of n e a r l y 2000. Consequent ly , t he p a t t e r n of ion iza t ions and exc i t a t ions a long the t r a c k s of e l e c t r o n s and p ro tons d i f f e r s m a r k e d l y , and t h i s accounts f o r subs t an t i a l d i f f e r e n c e s of b io log ica l e f f ec t s be tween g a m m a r a y s and neu t rons .

8 CHAPTER 2

2. 5 IONIZATIONS AND ENERGY DEGRADATION As has been shown, e x p o s u r e to d i r ec t ly or ind i rec t ly ionizing r a d i a t i o n s

r e s u l t s in the a p p e a r a n c e of cha rged p a r t i c l e s in the i r r a d i a t e d m a t t e r . T h e s e p a r t i c l e s i n t e r a c t with a tomic e l e c t r o n s d i sp lac ing t h e m into h igher o r b i t s (exci tat ion) or r e m o v i n g t h e m f r o m the p a r e n t a tom (ionization). In the c a s e of ioniza t ion , the e j ec t ed e l ec t ron f r equen t ly has suf f ic ien t ene rgy to p r o d u c e addi t ional ionizat ion. The p r o c e s s of ioniza t ion may be d e s c r i b e d by the equat ion

A * A+ + e"

The e l e c t r o n so p roduced may b e c o m e a t tached to ano ther a tom B to f o r m a negat ive ion:

B + e" > B~

The ions p roduced , A+ and B , a r e of ten t e r m e d an ion p a i r . The l i f e t ime of ion p a i r s i s e x t r e m e l y v a r i a b l e , but a s a r u l e it i s qui te sho r t ( l e s s t han 10"10 s).

An i n t e r m e d i a t e s t ep between ionizat ion o r exc i ta t ion and the a l t e r a t i o n of m o l e c u l e s of b iological i m p o r t a n c e i s usua l ly the p roduc t ion of f r e e r a d i c a l s . A f r e e r a d i c a l does not c a r r y e l e c t r i c cha rge , but has e x c e s s ene rgy and i s highly r e a c t i v e because of t he p r e s e n c e of an unpa i r ed e l ec t ron . An example of ioniza t ion and a subsequent f o r m a t i o n of a f r e e r a d i c a l o c c u r s when ionizing r a d i a t i o n i n t e r a c t s with w a t e r . The in i t ia l ionizat ion r e p r e -sented by

H 2 0 + H 2 0 + + e"

can be fol lowed by i n t e r a c t i o n between the ion and ano ther w a t e r mo lecu l e which l e a d s to the p roduc t ion of the highly r e a c t i v e hydroxyl r a d i c a l OH*

H 2 0 + + H 2 0 + HgO+ + OH-

The ex i s t ence of a g r e a t v a r i e t y of o ther r e a c t i o n s r e n d e r s the s i tua t ion v e r y m u c h m o r e complex , even in p u r e w a t e r . It i s even m o r e compl ica ted when the a b s o r b i n g m a t e r i a l c o n s i s t s of l a r g e biological m o l e c u l e s . T h e r e a r e two i m p o r t a n t po in ts t o r e m e m b e r conce rn ing f r e e r a d i c a l s . In the f i r s t p l a c e f r e e r a d i c a l s have a l i f e t ime of the o r d e r of 10"5 s , which i s m u c h l onge r t han tha t of the ion p a i r s f r o m which they a r e f o r m e d . They c a r r y a g r e a t dea l of ene rgy and can b r eak c h e m i c a l bonds; they a l m o s t a lways act a s an i n t e r m e d i a r y between the c r e a t i o n of ion p a i r s and the f ina l c h e m i c a l p r o d u c t s . Second, it i s thought tha t the oxygen ef fec t , which wil l be d i s c u s s e d in Chap t e r 7, o p e r a t e s at the f r e e r a d i c a l level .

The chain of events , f r o m the incident g a m m a - r a y photon or f a s t neu t ron , to the f ina l o b s e r v e d b io logica l changes , may be d e s c r i b e d a s fo l lows:

F a s t neu t ron G a m m a photon + +

Recoi l p ro ton F a s t e l e c t r o n 4 *

Ionizat ions & exc i t a t ions + F r e e r a d i c a l s +

B i o m o l e c u l a r changes + Biological d a m a g e

RADIATIONS AND THEIR INTERACTIONS 9

2. 6 PHYSICAL QUANTITIES AND THEIR UNITS

The fol lowing i s a s imp l i f i ed condensa t ion of m a t e r i a l given in Repor t 19 of the In t e rna t iona l C o m m i s s i o n on Radia t ion Uni ts and M e a s u r e m e n t s (ICRU 1971) which i s an e s s e n t i a l r e f e r e n c e . Other pe r t i nen t data may be ob ta ined f r o m Attix, Roesch and Tochi l in (1968), ICRU Repor t 13 (1969), and Hine and Browne l l (1956).

S e v e r a l of the quan t i t i e s u s e d in r ad io log ica l phys i c s have spec ia l uni t s , i. e. un i t s tha t apply to one o r a few quan t i t i e s only. On the o ther hand the In t e rna t iona l Sys tem (SI) u t i l i z e s the s a m e b a s e un i t s or combina t ions the reof f o r a l l phys i ca l quan t i t i e s . At p r e s e n t , spec i a l un i t s a r e of f a r g r e a t e r p r a c t i c a l i m p o r t a n c e , but it a p p e a r s l ikely tha t t he SI s y s t e m will be adopted in r ad io log i ca l p h y s i c s a s it ha s been in m o s t o the r b r anches of phys i c s . The base un i t s of the SI s y s t e m which a r e of i m p o r t a n c e h e r e a r e t h e s e c o n d (s) f o r t i m e , the m e t r e (m) f o r length, the k i l o g r a m (kg) f o r m a s s , and t h e a m p e r e (A) f o r e l e c t r i c a l c u r r e n t . Der ived un i t s of the SI s y s t e m which wi l l be r e f e r r e d to a r e the coulomb (C) f o r e l e c t r i c c h a r g e and the joule (J) f o r energy .

The fol lowing phys i ca l quan t i t i e s a r e of i m p o r t a n c e in succeed ing c h a p t e r s :

1. The f luence 0 , which i s the n u m b e r of p a r t i c l e s t r a v e r s i n g a s m a l l s p h e r e at the point of i n t e r e s t , divided by the s p h e r e c r o s s - s e c t i o n .

2. The flux dens i ty o r f luence r a t e cp , which i s the f luence p e r unit t i m e . 3. The energy f luence Y , which i s a p roduc t of f luence and p a r t i c l e

ene rgy . 4. The energy f lux dens i ty or energy f luence r a t e ip , which i s ene rgy

f luence p e r unit t i m e .

Quan t i t i e s 1 - 4 apply to both d i r ec t l y and ind i r ec t ly ionizing p a r t i c l e s . One can t h u s speak of a f luence of X - r a y s which in i r r a d i a t e d m a t t e r p r o -duces a f luence of s econda ry e l e c t r o n s .

5. The a b s o r b e d dose D is the energy a b s o r b e d p e r uni t m a s s of i r r a d i a t e d m e d i u m .

6. The a b s o r b e d d o s e - r a t e D i s the a b s o r b e d dose de l i ve red p e r unit t i m e .

Quan t i t i e s 5 and 6 a r e a l so appl ied to both d i r ec t ly and ind i rec t ly ioniz ing r a d i a t i o n s , al though the ene rgy i s a b s o r b e d f r o m charged p a r t i c l e s only.

A s p e c i a l unit of a b s o r b e d dose i s the r a d which equa ls 10" 2 J -kg" 1 . B e c a u s e of t he low r a d i a t i o n l e v e l s involved in m o s t of t he e x p e r i m e n t s c o n s i d e r e d h e r e the m o s t f r equen t l y employed unit of a b s o r b e d d o s e - r a t e wi l l be the m r a d - h " 1 .

The fol lowing quan t i t i e s apply t o ind i rec t ly ioniz ing r ad ia t ion only:

7. The k e r m a K is the ene rgy t r a n s f e r r e d f r o m ind i r ec t ly ioniz ing r ad i a t i on to d i r e c t l y ioniz ing r ad i a t i on p e r unit m a s s of i r r a -diated m a t e r i a l .

8. The k e r m a r a t e K is t he k e r m a p e r uni t t i m e .

The un i t s fo r k e r m a and k e r m a r a t e a r e the s a m e a s t h o s e fo r ab-s o r b e d dose and a b s o r b e d d o s e - r a t e .

10 CHAPTER 2

9. The m a s s a t tenua t ion coef f ic ien t n /p i s the f r a c t i o n of ind i rec t ly ionizing p a r t i c l e s i n t e r a c t i n g in a m e d i u m p e r unit d i s t ance t r a -v e r s e d , divided by the densi ty p of the m e d i u m .

The fol lowing quan t i t i e s apply to d i r e c t l y ioniz ing p a r t i c l e s only:

10. The to ta l m a s s s topping power S/p i s the ene rgy los t by a cha rged p a r t i c l e p e r uni t d i s t ance t r a v e r s e d , divided by the densi ty p of the m e d i u m .

11. The l i nea r ene rgy t r a n s f e r (LET) L A i s the energy los t by a cha rged p a r t i c l e in co l l i s ions with ene rgy t r a n s f e r l e s s than A , p e r uni t d i s t ance t r a v e r s e d .

L „ ( s o m e t i m e s s imply denoted L) i s equal to the p roduc t of m a s s s topping power and densi ty (S/p X p = S).

12. The a v e r a g e ene rgy expended in the p roduc t ion of an ion p a i r W is the e n e r g y expended by c h a r g e d p a r t i c l e s in a gas , divided by the n u m b e r of ion p a i r s p roduced .

The fol lowing a r e the s t o c h a s t i c quan t i t i e s of m i c r o d o s i m e t r y :

13. The spec i f i c ene rgy z i s the ene rgy depos i t ed in a reg ion , divided by the m a s s in that r e g i o n .

14. The l ineal ene rgy y i s the ene rgy depos i ted in a reg ion , divided by the m e a n d i a m e t e r of the r e g i o n .

The m e a n i n g of the t e r m 1 s t o c h a s t i c quant i ty ' and the s ign i f i cance of t h e s e quan t i t i e s i s d i s c u s s e d in Chap te r 4. The uni t of z i s the r a d . The unit of y i s keV/ium.

C h a p t e r 3 DOSIMETRY

3 .1 . O B J E C T I V E S

Although the t e r m ' d o s i m e t r y ' l i t e r a l l y m e a n s m e a s u r e m e n t of dose , it i s u sua l l y given a w i d e r connotat ion. Thus one speaks of d o s i m e t r i c ca lcu la t ions in which the abso rbed dose i s ca lcula ted f o r v a r i o u s po in ts in a phan tom that i s i r r a d i a t e d u n d e r defined g e o m e t r i c a l condi t ions by a s o u r c e hav ing known e m i s s i o n s . E x a m p l e s of such ca l cu la t ions a r e given in NCRP R e p o r t 38 (1971).

D o s i m e t r y m a y a l s o be conce rned with the m e a s u r e m e n t of quan t i t i e s o the r than a b s o r b e d dose , in p a r t i c u l a r that of k e r m a . K e r m a r e p r e s e n t s e n e r g y t r a n s f e r r e d f r o m ind i r ec t ly ionizing r ad i a t i on to d i r e c t l y ionizing r ad i a t i on p e r uni t m a s s of a spec i f i ed m a t e r i a l . It i s def ined f o r both f r e e space o r ins ide a sol id phantom. Excep t f o r a thin t r a n s i t i o n r e g i o n in which m o r e s e c o n d a r i e s a r e p roduced than a r e a b s o r b e d , the k e r m a f o r the phantom m a t e r i a l and the a b s o r b e d dose a r e in e s s e n c e equal at each point , p rov ided that the p e n e t r a t i o n of i nd i r ec t ly ionizing r ad i a t i on i s much g r e a t e r than that of the d i r e c t l y ionizing s e c o n d a r i e s . T h i s condit ion of r a d i a t i o n equ i l i b r ium i s wel l m e t f o r the r a d i a t i o n s emi t t ed by c a l i f o r n i u m - 2 52, and the concept of t i s s u e k e r m a is of l i t t le u t i l i ty ins ide t i s s u e equivalent phan toms . It i s , howeve r , of cons ide r ab l e i n t e r e s t in f r e e space w h e r e the t i s s u e k e r m a a t s o m e d i s t ance f r o m a s o u r c e i s one of the m o s t i m p o r t a n t c h a r a c t e r i z a t i o n s . T h i s i s s o m e t i m e s loose ly cal léd the ' a i r dose ' o r ' t i s s u e dose in f r e e a i r ' . Th i s i n c o r r e c t u s a g e i s quite u n d e r s t a n d a b l e , s ince t i s s u e k e r m a in f r e e a i r i s typica l ly m e a s u r e d by the s a m e d e v i c e s which a r e u s e d to d e t e r m i n e the a b s o r b e d dose in phan toms . T h e s e a r e u sua l l y ioniza t ion c h a m b e r s , o r o the r i n s t r u m e n t s having wa l l s that a r e not much t h i c k e r than r e q u i r e d to e s t ab l i sh r ad i a t i on equ i l ib r ium. The dose r e c e i v e d by the sens i t i ve e l emen t of such i n s t r u m e n t s ( e . g . the gas in an ioniza t ion c h a m b e r ) can of ten be cons ide red to be equal to the k e r m a .

The b io logica l e f f ec t of r a d i a t i o n depends not only on r ad i a t i on quant i ty but a l s o on r a d i a t i o n qual i ty . T h i s m e a n s tha t in addit ion to the a b s o r b e d dose , the n a t u r e and ene rgy of t he cha rged p a r t i c l e s that de l i ve r th i s dose a r e of c o n s i d e r a b l e i m p o r t a n c e . Qual i ty can be spec i f ied with i n c r e a s i n g amount of de ta i l .

(1) Iden t i f i ca t ion of the f r a c t i o n of the a b s o r b e d dose that i s due to v a r i o u s t ypes of i n d i r e c t l y ioniz ing r a d i a t i o n s (in the c a s e of c a l i f o r n i u m - 2 5 2 , a s e p a r a t e a s s e s s m e n t of the a b s o r b e d doses due to neu t rons and g a m m a rays ) .

(2) A spec i f i ca t i on of the a b s o r b e d dose de l i ve red by c h a r g e d p a r t i c l e s a t each L E T , i. e. d(L), the d i s t r ibu t ion of a b s o r b e d dose in L E T .

(3) A spec i f i ca t i on of the d i s t r ibu t ion of e n e r g y i n c r e m e n t s in v o l u m e s of v a r i o u s s i ze within the i r r a d i a t e d m a t e r i a l . In th is f o r m of d e s c r i p t i o n the m i c r o s c o p i c concep ts of l inea l e n e r g y and spec i f i c e n e r g y a r e employed .

In a c c o r d a n c e with c u r r e n t u s a g e , i tem (1) will be c o n s i d e r e d p a r t of convent ional d o s i m e t r y , while i t e m s (2) and (3) wil l be c o n s i d e r e d in C h a p t e r 4.

1 1

12 CHAPTER 2

TABLE I. ATOMIC COMPOSITION ( P E R C E N T OF MASS) OF MAJOR CONSTITUENTS OF VARIOUS MATERIALS

H C N 0

Tissue (ICRU 1964, 1969) 10.1 11.1 2 . 6 76 .2

T issue-Equiva lent P l a s t i c (Shonka, A150)* 10.1 7 7 . 6 3 . 5 5 .2

T issue-Equiva lent Gas (Rossi 6 Fa i 1 la ( 1 9 5 6 ) , 64. CH., 3 2 . 4 1 C0 2 , 3 . 2 1 N 2 " * ) h 10.2 45 .6 3 . 5 4 0 . 7

Water 11.1 0 0 88 .9

T issue-Equiva lent L iquid (Goodman (1969 ) , 65.6? Water , 26.8% g l y c e r o l , 7 . 6 Urea) 10.2 12.0 3 . 6 74 .2

Polystyrene 7 . 7 92 .3 0 0

Polyethylene 14.3 85 .7 0 0

Methyl Methacylate ( L u c i t e , Perspex) 8 60 0 32

*The balance of the m a t e r i a l cons is ts o f calcium f l u o r i d e

The values given f o r these gases are percent p a r t i a l pressures

3. 2. D E T E C T O R S AND DOSIMETERS

If a device capable of indica t ing the p r e s e n c e of r ad i a t i on is exposed to v a r i o u s i n t ens i t i e s of t h i s r ad i a t i on , i t s r e s p o n s e m a y f r e q u e n t l y be p r o p o r t i o n a l to in tens i ty , and if a su i tab le ca l i b r a t i on f a c t o r i s p rov ided , the i n s t r u m e n t r e a d i n g m a y s e r v e to indica te any r ad i a t i on quanti ty including a b s o r b e d dose . However , if the r ad ia t ion i s changed, e i t h e r in e n e r g y o r compos i t ion , a d i f f e r e n t c a l i b r a t i o n f a c t o r m a y be r e q u i r e d . If th i s i s the c a s e , the i n s t r u m e n t i s sa id to be e n e r g y - d e p e n d e n t o r qua l i ty -dependent . In p r i nc ip l e , a d o s i m e t e r h a s no such dependence but the t e r m ' d o s i m e t e r ' i s o f ten employed f o r dev ices which have qual i ty o r e n e r g y dependence , which i s accep tab le . O t h e r r ad i a t i on m e a s u r i n g i n s t r u m e n t s a r e u sua l l y r e f e r r e d to a s d e t e c t o r s . In m a n y c a s e s the c l a s s i f i c a t i o n of a given i n s t r u m e n t i s a m a t t e r of opinion.

ICRP Repo r t 20 (1971) con ta ins d e s c r i p t i o n s of a wide v a r i e t y of i n s t r u m e n t s that can be used f o r d o s i m e t r y of g a m m a r a y s and neu t rons . The only type of d o s i m e t e r which wil l be d i s c u s s e d h e r e in any de ta i l is the ioniza t ion c h a m b e r , b e c a u s e it can be made to have m i n i m a l dependence on r ad i a t i on ene rgy and type, and b e c a u s e it i s p e r h a p s the s i m p l e s t r ad i a t i on i n s t r u m e n t .

DOSIMETRY 1 3

FIG. la . Large ionization Chamber (P type): drawing of T. E. model . This chamber is of guard ring (3-electrode) design and consists of T. E. plastic and polyethylene. It is entirely heat-sealed. The plug with the toroidal rubber gasket ('O-ring') is used as a temporary seal immediate ly after filling with T. E. gas and the heat-sealed cover is applied minutes after filling. Other versions of this chamber uti l ize O-rings instead of heat seals. These types require re-f i l l ing after a few months (because of gas leakage). The model illustrated changes in sensitivity by less than 10°lo per year. This chamber has been extensively employed in radiation protection measurements.

3. 3. THE IONIZATION CHAMBER

In an ionizat ion c h a m b e r des igned f o r d o s i m e t r y , the ionizat ion o b s e r v e d in the gas occupying a cavi ty i s a m e a s u r e of the e n e r g y abso rbed p e r uni t m a s s of the s u r r o u n d i n g wal l m a t e r i a l . When the cavi ty i s such that the cha rged p a r t i c l e s in the wal l t r a v e r s e it with negl igible expendi tu re of t h e i r e n e r g y , the e n e r g y depos i t ion DG , p e r unit m a s s of gas , and Dw , p e r unit m a s s of wal l , a r e in the r a t i o of the s topping p o w e r s in the two m e d i a . T h u s

D G / D W = (S/pfe/ (S/p)w

1 4 CHAPTER 2

FIG. lb. Large ionization chamber (P type): external appearance of T. E. model .

I 10 CENTIMETRES

FIG. l c . Large ionization chamber: external appearance of aluminium model . This chamber is filled with argon.

DOSIMETRY 1 5

FIG. 2a. M e d i u m - s i z e ion iza t ion chamber (B type): drawing of T. E. mode l . This is a condenser type (2 -e lec trode chamber) which c a n be charged in a standard e l ec trometer (Victoreen) . It has been e m p l o y e d in radiobio log ica l dosimetry.

1 6 CHAPTER 2

j incuts

FIG. 2b. M e d i u m - s i z e ion iza t ion chamber (B type): photograph of chamber, e l e c t rometer and associated

equipment .

DISC CHAMBER

g g g l POLYSTYRENE LUCITE

TEFLON | g | § ALUMINIUM

g¡¡¡¡ TISSUE-EQUIVALENT PLASTIC

FIG. 3a. D i s c - t y p e chamber: drawing of chamber . This is a 3 - e l e c t r o d e chamber operated in continuous

gas f low. T h e c o l l e c t i n g e l ec trode is a reinforced thin (about i m m ) disc .

DOSIMETRY 1 7

T I S S U E EQU i V A L E N T P L A S T I C D I S C S TH 5 CH NE SS I N MM

• • • • • . 25 0 . 5 1 0 . 7 6 2 . 2

FIG. 3b. Disc-type chamber: external appearance of chamber, ln this version one wall has been reduced to 0,14 mm and the discs shown are employed to study build-up phenomena. The long gas and electric lines are provided to permit immersion into a liquid phantom.

D g i s p ropo r t i ona l to J , the ionizat ion p e r uni t m a s s of gas , and the p ropo r t i ona l i t y f a c t o r i s We w h e r e W is a v e r a g e e n e r g y expended p e r ioniza t ion , and e i s the e l e c t r o n i c c h a r g e . T h u s

D w = JWe(S/p) w / (S /p ) G

Thi s equat ion, known a s the B r a g g - G r a y T h e o r e m , is of fundamen ta l i m p o r t a n c e in r ad ia t ion d o s i m e t r y . I ts b a s i c a s s u m p t i o n , to the e f fec t that the cavi ty r e p r e s e n t s a negl igible d i s t u r b a n c e to the flow of charged p a r t i c l e s in the m e d i u m , i s not e a s i l y fu l f i l l ed at low rad ia t ion l eve l s , such a s those l ike ly to be encoun te red in work with s m a l l c a l i f o r n i u m - 2 5 2 s o u r c e s . However , an i m p o r t a n t ex tens ion of the t h e o r e m r e l a t e s to the c a s e when wall and gas have the s a m e a tomic compos i t ion . With th i s condit ion the equat ion b e c o m e s

D w = JWe

b e c a u s e the r a t i o of m a s s s topping p o w e r s i s 1.

I

I cm

GAP ADJUSTMENT RING

POLYSTYRENE

ALUMINIUM

1 7 / / / / / / / / / A STAINLESS STEEL

LUCITE

KEL-F

TISSUE-EQUIVALENT PLASTIC

FIG.4. Multiplication chamber. (a) Drawing of aluminium

chamber. This is a 3-electrode gas flow chamber. The base of the collecting volume is defined by the end face of the central electrode. The annular co-planar electrode is the guard ring. The chamber is surrounded by a Lucite envelope for insulation.

(b) T .E. model: detail of electrode structure. Gas ascends in a groove on the right, emerges between collector and guard, and leaves at left .

(c) T .E . and aluminium models inside liquid phantom. A remotely operated source manipulator is moving a 252Cf needle near one of the chambers.

20 CHAPTER 10

H o w e v e r , a f a r m o r e i m p o r t a n t c o n s e q u e n c e of e q u a l c o m p o s i t i o n of w a l l and g a s , i s t ha t a s m a l l c a v i t y i s no l o n g e r r e q u i r e d , s i n c e t h e r e i s e x a c t c o m p e n s a t i o n of e n e r g y l o s s of s e c o n d a r i e s o r i g i n a t i n g in the wa l l b y p r o d u c t i o n of s e c o n d a r i e s in the g a s . C h a m b e r s o p e r a t i n g u n d e r t h e s e c o n d i t i o n s a r e t e r m e d h o m o g e n e o u s c h a m b e r s .

S ince the m a t e r i a l of p r i n c i p a l i n t e r e s t to r a d i o b i o l o g y i s t i s s u e , c h a m b e r s h a v i n g w a l l s and g a s of t i s s u e - e q u i v a l e n t m a t e r i a l (T. E . c h a m b e r s ) a r e of p r i m e i n t e r e s t ( R o s s i and F a i l l a , 1956). A l though t r u e t i s s u e e q u i -v a l e n c e c a n b e a c h i e v e d , i t i s u s u a l l y s u f f i c i e n t t o d e a l w i th m a t e r i a l s c o n t a i n i n g c o r r e c t p r o p o r t i o n s of h y d r o g e n and n i t r o g e n , wi th t he r e m a i n i n g two p r i n c i p a l e l e m e n t s (oxygen and c a r b o n ) in m o r e o r l e s s a r b i t r a r y p r o p o r t i o n s . T a b l e I s h o w s t h e c o n t e n t of t h e s e p r i n c i p a l e l e m e n t s in a v a r i e t y of s u b s t a n c e s .

In t h e d o s i m e t r y of g a m m a r a y s h a v i n g e n e r g i e s in e x c e s s of a f e w h u n d r e d k e V , c h a m b e r s w h o s e w a l l and g a s a r e c o n s t i t u t e d of l i gh t e l e m e n t s ( a t o m i c n u m b e r l e s s t h a n a b o u t 15) wi l l be found to h a v e a n e n e r g y d e p e n d e n c e w h i c h i s l e s s t h a n a f e w p e r c e n t . H e n c e , a v a r i e t y of m a t e r i a l s c a n b e u s e d in t he c o n s t r u c t i o n of i o n i z a t i o n c h a m b e r s f o r t h e s e r a d i a t i o n s . H o w e v e r , in t he c a s e of n e u t r o n s , e s p e c i a l l y t h o s e p r o d u c e d by c a l i f o r n i u m - 2 5 2 , h y d r o g e n c o n t e n t i s of p r i m a r y i m p o r t a n c e , and the r e s p o n s e of h o m o g e n e o u s h y d r o g e n o u s c h a m b e r s i s l a r g e l y p r o p o r t i o n a l t o h y d r o g e n c o n t e n t .

P l a s t i c s h a v e b e e n d e v e l o p e d which not on ly a r e t i s s u e - e q u i v a l e n t w i th r e s p e c t t o h y d r o g e n and n i t r o g e n c o n t e n t , bu t a l s o e x h i b i t a d e q u a t e c o n d u c t i v i t y f o r u s e in i o n i z a t i o n c h a m b e r s . F i g u r e s 1 - 3 show e x a m p l e s of s u c h c h a m b e r s , a l l of wh ich a r e o p e r a t e d e i t h e r wi th a c o n t i n u o u s f low of t i s s u e - e q u i v a l e n t g a s , o r a r e f i l l e d wi th t h i s g a s and p r o v i d e d wi th h e r m e t i c s e a l s . In t he l a t t e r m o d e it i s not on ly n e c e s s a r y t h a t a l l s e a l s b e c o m p l e t e l y t i g h t , but a l s o t h a t t h e s e c h a m b e r s be o c c a s i o n a l l y r e f i l l e d b e c a u s e of a b s o r p t i o n of t he g a s by m a t e r i a l s in the c h a m b e r ( p a r t i c u l a r l y p l a s t i c ) .

An i m p o r t a n t m o d i f i c a t i o n of t i s s u e - e q u i v a l e n t i o n i z a t i o n c h a m b e r s ( a s w e l l a s of o t h e r t y p e s ) i s i n t e r n a l g a s m u l t i p l i c a t i o n ( B e n g t s s o n , G o o d m a n & R o s s i , 1969). In s u c h c h a m b e r s , v o l t a g e s m u c h in e x c e s s of t h o s e r e q u i r e d f o r s a t u r a t i o n a r e a p p l i e d , in o r d e r t o a c h i e v e a m p l i f i c a t i o n of the ion c u r r e n t by i n e l a s t i c c o l l i s i o n s in the g a s . S u c c e s s f u l o p e r a t i o n r e q u i r e s a p p r o p r i a t e d e s i g n of t he e l e c t r o d e s , wh ich shou ld a i m a t a u n i f o r m p a r a l l e l , c y l i n d r i c a l o r s p h e r i c a l e l e c t r i c f i e l d t h r o u g h o u t m o s t of t he c h a m b e r v o l u m e . A n e x a m p l e of a p a r a l l e l p l a t e c h a m b e r i s shown in F i g . 4. T h i s c h a m b e r , w h i c h "-"as a v o l u m e of on ly a b o u t 2 m m 3 , w a s u s e d to o b t a i n t he d a t a of T a b l e s IV and V in A p p e n d i x I. T h e v o l u m e is s o s m a l l t h a t a c c u r a t e m e a s u r e m e n t s could be m a d e a f ew m i l l i m e t r e s f r o m a c a l i f o r n i u m - 2 5 2 s o u r c e ( R o s s i & C o l v e t t , 1972; C o l v e t t , R o s s i & K r i s h n a s w a m y , 1972).

I o n i z a t i o n c h a m b e r s c a n be c a l i b r a t e d ' a b s o l u t e l y ' , u t i l i z i n g t he B r a g g - G r a y r e l a t i o n g i v e n e a r l i e r i n t h i s c h a p t e r . T h i s r e q u i r e s d e t e r -m i n a t i o n s of t he m a s s of t he c o n t a i n e d g a s , and the c h a r g e c o l l e c t e d , a s w e l l a s k n o w l e d g e of W. F o r t h e t i s s u e - e q u i v a l e n t g a s h a v i n g a c o m p o s i t i o n s h o w n in T a b l e I , W i s a p p r o x i m a t e l y 29 eV f o r g a m m a r a y s , and 30.5 eV f o r n e u t r o n s . A f a r s i m p l e r m o d e of c a l i b r a t i o n i n v o l v e s i r r a d i a t i o n of t he c h a m b e r t o a known e x p o s u r e of g a m m a r a d i a t i o n , a s m a y f o r i n s t a n c e b e a c c o m p l i s h e d b y p l a c i n g the c h a m b e r a t a n a c c u r a t e l y d e t e r m i n e d d i s t a n c e f r o m a s m a l l s o u r c e c o n t a i n i n g r a d i u m , o r a n o t h e r g a m m a -

DOSIMETRY 21

e m i t t i n g n u c l i d e . T h e e x p o s u r e r a t e c o n s t a n t (which i s 8 .25 R • m 2 • h"1 • C i " 1

f o r u n s h i e l d e d r a d i u m ) t h e n e s t a b l i s h e s the e x p o s u r e . T h e n u m e r i c a l v a l u e of the a b s o r b e d d o s e r e c e i v e d b y the g a s i s a p p r o x i m a t e l y 0 .96 t i m e s the e x p o s u r e . T h i s e s t a b l i s h e s the g a m m a s e n s i t i v i t y of the c h a m b e r . T h e n e u t r o n s e n s i t i v i t y i s a p p r o x i m a t e l y 4% l e s s . In m i x e d f i e l d s , s u c h a s t h o s e e n c o u n t e r e d in c a l i f o r n i u m - 2 5 2 w o r k , a W c o r r e c t i o n of 2% shou ld be a d e q u a t e .

3 . 4 . S E P A R A T E A S S E S S M E N T O F N E U T R O N AND GAMMA D O S E S

B e c a u s e of i t s a l m o s t u n i f o r m r e s p o n s e to a l l t y p e s of i o n i z i n g r a d i a t i o n s , the t i s s u e - e q u i v a l e n t i o n i z a t i o n c h a m b e r m a y r e a d i l y be e m p l o y e d to d e t e r m i n e t o t a l a b s o r b e d d o s e . It i s u s e f u l , t h e r e f o r e , f o r s e p a r a t e d e t e r m i n a t i o n s of n e u t r o n and g a m m a d o s e s on ly in tha t it g i v e s t h e s u m of s u c h d o s e s , wi th the r e m a i n i n g r e q u i r e m e n t t h a t one of t h e s e m u s t be d e t e r m i n e d s e p a r a t e l y . It i s g e n e r a l l y m o r e d i f f i c u l t t o c o n s t r u c t d o s i m e t e r s wh ich f u l l y r e s p o n d to n e u t r o n s and h a v e n e g l i g i b l e g a m m a - r a y s e n s i t i v i t y , p a r t i c u l a r l y w h e n the n e u t r o n l e v e l i s too low to p e r m i t t he e m p l o y m e n t of t h r e s h o l d d e t e c t o r s . M o s t of t he i n s t r u m e n t s s u i t a b l e f o r s u c h d e t e r -m i n a t i o n s a r e p r o p o r t i o n a l c o u n t e r s e m p l o y i n g d i s c r i m i n a t i o n t e c h n i q u e s .

T h e r e v e r s e c h a r a c t e r i s t i c , i . e . , g a m m a s e n s i t i v i t y and n e g l i g i b l e n e u t r o n r e s p o n s e , i s s o m e w h a t m o r e e a s i l y a c c o m p l i s h e d . One of t he s i m p l e s t and m o s t e f f e c t i v e d e v i c e s i s an i o n i z a t i o n c h a m b e r h a v i n g a w a l l of a n o n - h y d r o g e n o u s c o n d u c t i n g s u b s t a n c e of low a t o m i c n u m b e r , s u c h a s g r a p h i t e o r a l u m i n i u m , and a g a s s u c h a s c a r b o n d i o x i d e , a r g o n , e t c . A i r h a s a n a p p r e c i a b l e r e s p o n s e t o t h e r m a l n e u t r o n s b e c a u s e of c a p t u r e by n i t r o g e n , bu t in m a n y e x p e r i m e n t s i nvo lv ing c a l i f o r n i u m - 2 5 2 t h e t h e r m a l n e u t r o n c o m p o n e n t shou ld b e m i n o r . If a c h a m b e r of t h i s type i s e m p l o y e d , t o g e t h e r wi th a T E c h a m b e r of e q u a l s h a p e a n d s i m i l a r c o n s t r u c t i o n , d o s e s due to n e u t r o n s and g a m m a r a y s c a n both b e d e t e r m i n e d wi th a n a c c u r a c y a p p r o a c h i n g a f e w p e r c e n t . C o r r e c t i o n s c a n be m a d e f o r the n e u t r o n s e n s i t i v i t y of t h e g a m m a - s e n s i t i v e c h a m b e r , w h i c h f o r a n a l u m i n i u m c h a m b e r f i l l e d wi th a r g o n i s a b o u t 1.5% in the c a s e of c a l i f o r n i u m - 2 5 2 n e u t r o n s .

A l t e r n a t i v e m e t h o d s of a s s e s s i n g g a m m a - r a y d o s e s i n c l u d e e m p l o y m e n t of e n e r g y - i n d e p e n d e n t GM c o u n t e r s and , p r o v i d e d t h a t r i g o r o u s c a l i b r a t i o n s and d e v e l o p m e n t p r o c e d u r e s a r e a v a i l a b l e , a l s o p h o t o g r a p h i c e m u l s i o n s .

C h a p t e r 4

RADIATION QUALITY AND MICRODOSIMETRY

4. 1, L I N E A R E N E R G Y T R A N S F E R ( L E T )

In m a n y b i o l o g i c a l e x p e r i m e n t s c a r r i e d out wi th c a l i f o r n i u m - 2 5 2 , the a b s o r b e d d o s e s due to n e u t r o n s and g a m m a r a d i a t i o n w i l l b e found to b e a p p r o x i m a t e l y equa l . T h i s m e a n s t h a t e a c h of t h e s e r a d i a t i o n s c o n t r i b u t e s a b o u t hal f of a l l t he i o n i z a t i o n s a n d e x c i t a t i o n s t a k i n g p l a c e in the i r r a d i a t e d t i s s u e s . N e v e r t h e l e s s t he b i o l o g i c a l e f f e c t of t he g a m m a r a d i a t i o n i s s o m u c h l e s s t h a n t h a t of t he n e u t r o n s a s t o a s s u m e a l m o s t n e g l i g i b l e p r o -p o r t i o n s , p a r t i c u l a r l y a t t h e low d o s e s and low d o s e - r a t e s m a n d a t e d by w e a k s o u r c e s .

T h e r e a s o n f o r t h i s r e m a r k a b l e d i f f e r e n c e in e f f e c t i v e n e s s i s a d i f f e r e n c e in d i s t r i b u t i o n of a b s o r b e d e n e r g y . A l though t h e e n e r g y of t h e e l e c t r o n s s e t in m o t i o n by t h e g a m m a r a d i a t i o n i s of t he s a m e o r d e r of m a g n i t u d e a s t h a t of t h e p r o t o n s r e c o i l i n g f r o m n e u t r o n s , t he f o r m e r h a v e r a n g e s of t he o r d e r of 1 m m w h i l e the l a t t e r h a v e r a n g e s t h a t a r e r o u g h l y a h u n d r e d t i m e s l e s s . T h i s i s , in t u r n , due t o t he f a c t t ha t t he m u c h h e a v i e r and t h e r e f o r e m u c h s l o w e r p r o t o n s a r e m o r e l i k e l y t o i o n i z e o r e x c i t e a t o m s t h r o u g h wh ich t h e y p a s s . T h e l i n e a r e n e r g y t r a n s f e r ( L E T ) , wh ich i s t he p h y s i c a l m e a s u r e of t he r a t e of e n e r g y l o s s of t h e s e p a r t i c l e s , i s g e n e r a l l y of t h e o r d e r of a f e w t e n t h s keV/fian f o r t he e l e c t r o n s , and of the o r d e r of t e n s of keV/ym f o r t he p r o t o n s . T h e s e v a l u e s a r e m e r e l y t y p i c a l a v e r a g e s (ICRU R e p o r t 16, 1970; R o s s i , 1967, 1968).

D e p e n d i n g on the i n i t i a l e n e r g y of the c h a r g e d p a r t i c l e and d i s t a n c e f r o m i t s o r i g i n , i t s L E T c a n v a r y b y m o r e t h a n a n o r d e r of m a g n i t u d e . B e c a u s e of t he c o n s i d e r a b l e v a r i a b i l i t y of L E T , s p e c t r a l d i s t r i b u t i o n s a r e n e e d e d f o r m e a n i n g f u l r e p r e s e n t a t i o n s . T h e s e u s u a l l y t a k e t h e f o r m of t h e d i s t r i b u t i o n of d o s e in L E T , i. e . d (L) , t he f r a c t i o n of t he d o s e e x p e n d e d p e r u n i t i n t e r v a l of L E T , i s p l o t t e d a g a i n s t L . F i g u r e 5 g i v e s s u c h d i s t r i b u t i o n s f o r 2 5 2 Cf n e u t r o n s a s o b s e r v e d _in f r e e s p a c e and in a p h a n t o m .

T h e m e a n v a l u e of s u c h a d i s t r i b u t i o n , Lp, i s the d o s e a v e r a g e of t he L E T . A n o t h e r a v e r a g e , LT , r e p r e s e n t s t he L E T a v e r a g e d o v e r t r a c k l e n g t h . N e i t h e r a v e r a g e i s of a p p r e c i a b l e s i g n i f i c a n c e in r a d i o b i o l o g y . R a d i a t i o n s t h a t h a v e t he s a m e L D , o r r a d i a t i o n s t h a t h a v e t h e s a m e L T , m a y n e v e r t h e l e s s d i f f e r s u b s t a n t i a l l y in b i o l o g i c a l e f f e c t i v e n e s s .

4 . 2 . L I N E A L E N E R G Y y

W h e n a h igh s p e e d e l e c t r o n i o n i z e s a n a t o m , t he l i b e r a t e d s e c o n d a r y e l e c t r o n s m a y h a v e a w i d e r a n g e of e n e r g i e s , bu t a t y p i c a l a v e r a g e v a l u e i s 100 eV. C o n s e q u e n t l y , if a n e l e c t r o n h a v i n g a n L E T of 0.2 keV/ium t r a v e r s e s a b i o l o g i c a l s t r u c t u r e h a v i n g a d i a m e t e r of 1 jum, i t w i l l on t h e a v e r a g e u n d e r g o two c o l l i s i o n s . H o w e v e r , t h e s e c o l l i s i o n s a r e s u b j e c t t o s t a t i s t i c a l f l u c t u a t i o n s , and t h e r e i s an a p p r e c i a b l e p r o b a b i l i t y t h a t t h e i r n u m b e r i s bo th l a r g e r a n d s m a l l e r t h a n 2, i n c l u d i n g 0. On the o t h e r h a n d , a p r o t o n h a v i n g a n L E T of 100 keV/ i im h a s i n s u f f i c i e n t e n e r g y to t r a v e l a d i s t a n c e of 1 ¡jm; i t m a y t h e r e f o r e no t e v e n t r a v e r s e the r e g i o n .

23

2 4 CHAPTER 10

0.5

0.4

0.3

-J 0 .2

0.1

1 1 1

ft

i i

1 \ PHANTOM

• J X ' r \

' / >> . 1 ! / ' A I R — ^ J r /

/

i i i

\

•

10' I O 2 10*

L E T L (keV / ( im)

FIG. 5. L-d(L) vs. L (L is plotted on a logari thmic scale) for neutrons from 252Cf. The curve marked 'a i r ' was obtained in free air . The curve marked 'phantom' was obtained in a T . E. phantom 7.5 cm from the source. The ordinate is L 'd(L) rather than d(L) because the abscissa is logari thmic. As a result the fraction of the dosé in any LET interval is proportional to the corresponding area under the curve.

FIG. 6. y • d(y) vs. y (y is plotted on a logar i thmic scale) for neutrons and gamma radiation from 252Cf. The curves were obtained inside a phantom for 1-fjm and 2-(jm spherical cavit ies. The ordinate is y d ( y ) rather than d(y) because the abscissa is logar i thmic.

RADIATION QUALITY AND MICRODOSIMETRY 25

FIG. 7. Diagram of spherical proportional counter.

26 CHAPTER 10

A n o t h e r c o n s i d e r a t i o n i s t ha t in t r a v e r s i n g a d e f i n e d v o l u m e , s u c h a s a s p h e r e , e i t h e r p a r t i c l e m a y h a v e v a r y i n g p a t h l e n g t h s w i th in t he v o l u m e .

F o r t h e s e and s i m i l a r r e a s o n s , it i s d i f f i c u l t o r i m p o s s i b l e t o d e t e r -m i n e t he e n e r g y d i s t r i b u t i o n in i r r a d i a t e d m a t t e r on t he b a s i s of L E T s p e c t r a . T h e l i n e a l e n e r g y y i s d e f i n e d a s t he e n e r g y l a i d down by a n i n d i v i d u a l p a r t i c l e ( a n d / o r i t s s e c o n d a r i e s ) in a v o l u m e , d iv ided by t h e m e a n d i a m e t e r of t he v o l u m e . T h e r e g i o n i s u s u a l l y a s s u m e d to h a v e the s h a p e of a s p h e r e , in wh ich c a s e t he m e a n d i a m e t e r i s t w o - t h i r d s of t he s p h e r e d i a m e t e r .

T h e l i n e a l e n e r g y y i s a s t o c h a s t i c q u a n t i t y , wh ich m e a n s t h a t it c a n a s s u m e a v a r i e t y of v a l u e s u n d e r s e e m i n g l y f i x e d c o n d i t i o n s . T h u s , if a p a r t i c l e of L E T L t r a v e r s e s a 1 - ^ m s p h e r e a l o n g i t s m a x i m u m d i a m e t e r , y m a y be e x p e c t e d t o b e | L , bu t c o n s i d e r a b l e d e v i a t i o n s f r o m t h i s v a l u e m a y a l s o be o b s e r v e d , p a r t i c u l a r l y if L i s low. W h e n d i s t r i b u t i o n s of d(y) v e r s u s y a r e g i v e n , it i s n e c e s s a r y not on ly t o s p e c i f y t he s h a p e , bu t a l s o t he d i m e n s i o n s of t he v o l u m e of i n t e r e s t . A s t h e s e b e c o m e p r o g r e s s i v e l y s m a l l e r , t he d(y) d i s t r i b u t i o n b r o a d e n s b e c a u s e of i n c r e a s i n g s t a t i s t i c a l f l u c t u a t i o n s . F i g u r e 6 s h o w s d(y) d i s t r i b u t i o n s a s o b s e r v e d in l - / j m and 2-jum s p h e r e s in a p h a n t o m . H e r e t h e d i f f e r e n c e in s i z e i s not s u f f i c i e n t t o r e s u l t in a p p r e c i a b l e c h a n g e s of t h e d i s t r i b u t i o n .

4. 3. S P E C I F I C E N E R G Y z

In g e n e r a l , t h e d e l i v e r y of a c e r t a i n a b s o r b e d d o s e r e s u l t s in t h e t r a v e r s a l b y s e v e r a l c h a r g e d p a r t i c l e s in s o m e r e g i o n in t he i r r a d i a t e d t i s s u e . S o m e w h a t s i m i l a r l y to t he m a n n e r in wh ich y i s t he s t o c h a s t i c a n a l o g of L E T , t he s p e c i f i c e n e r g y z i s t he a n a l o g of t he a b s o r b e d d o s e . D i s t r i b u t i o n s of z c a n b e c a l c u l a t e d once d i s t r i b u t i o n s in t e r m s of y a r e known. T h e s p e c i f i c e n e r g y i s a q u a n t i t y of c o n s i d e r a b l e i m p o r t a n c e in t h e o r e t i c a l r a d i o b i o l o g y , bu t it w i l l not b e c o n s i d e r e d in any d e t a i l in t h i s p u b l i c a t i o n .

4. 4. T I S S U E - E Q U I V A L E N T P R O P O R T I O N A L C O U N T E R S

In t h e c o n v e n t i o n a l o p e r a t i o n of a t i s s u e - e q u i v a l e n t i o n i z a t i o n c h a m b e r , t he t o t a l c h a r g e p r o d u c e d by a l a r g e n u m b e r of d i r e c t l y i o n i z i n g p a r t i c l e s i s d e t e r m i n e d , t o d e r i v e the a b s o r b e d d o s e in t i s s u e . E v a l u a t i o n s of L E T and of m i c r o d o s i m e t r i c q u a n t i t i e s r e q u i r e t he d e t e r m i n a t i o n of t he e n e r g y i n c r e m e n t s p r o d u c e d by i n d i v i d u a l c h a r g e d p a r t i c l e s . T h i s in t u r n n e c e s s i t a t e s a m p l i f i c a t i o n of t h e m i n u t e p u l s e s p r o d u c e d by s i n g l e p a r t i c l e s , wh ich i s a c c o m p l i s h e d t h r o u g h the u s e of p r o p o r t i o n a l c o u n t e r s and t h e i r a c c e s s o r y e l e c t r o n i c e q u i p m e n t , c o n s i s t i n g of a p r e a m p l i f i e r , a p u l s e a m p l i f i e r and a m u l t i c h a n n e l a n a l y s e r ( R o s s i and R o s e n z w e i g , 1955) . B y u s i n g t h i s t e c h n o l o g y , i t i s p o s s i b l e t o m e a s u r e a s p e c t r u m w h i c h i s in e s s e n c e p r o p o r t i o n a l t o the f r e q u e n c y s p e c t r u m of y. F o r r e a s o n s of s y m m e t r y a n d s i m p l i c i t y , t he c a v i t y in wh ich t h e s e m e a s u r e m e n t s a r e c a r r i e d out i s u s u a l l y s p h e r i c a l , and t h e p r e s s u r e i s a d j u s t e d t o s u c h a v a l u e t h a t un i t d e n s i t y t i s s u e s p h e r e s h a v i n g d i a m e t e r s of t he o r d e r of 1 ijm a r e s i m u l a t e d . F i g u r e 7 i s a d i a g r a m of a s p h e r i c a l p r o p o r t i o n a l c o u n t e r a s u t i l i z e d in m i c r o d o s i m e t r y . M e a s u r e m e n t s wi th s u c h d e v i c e s h a v e b e e n r e p o r t e d f o r c a l i f o r n i u m - 2 5 2 r a d i a t i o n ( D i c e l l o , G r o s s & K r a l j e v i c , 1972).

C h a p t e r 5

BIOLOGICAL T E S T SYSTEMS

5 . 1 . E N D - P O I N T S S C O R E D

At one t i m e o r a n o t h e r a l m o s t e v e r y l i v ing s p e c i e s h a s b e e n i r r a d i a t e d in t h e n a m e of r a d i o b i o l o g y . In g e n e r a l , b i o l o g i c a l s y s t e m s u s e d f o r t h e s t u d y of r a d i a t i o n e f f e c t s m a y be d iv ided in to two c a t e g o r i e s :

(A) S y s t e m s w h i c h in one w a y o r a n o t h e r s c o r e c e l l k i l l i ng , i . e . t h e l o s s of r e p r o d u c t i v e i n t e g r i t y of c e l l s .

(B) S y s t e m s w h i c h invo lve a m u t a t i o n of a g e r m c e l l o r a s o m a t i c c e l l , o r a n e o p l a s t i c t r a n s f o r m a t i o n .

(A) S y s t e m s d e p e n d e n t on c e l l k i l l i ng

5 . 2 . I N - V I T R O C E L L C U L T U R E

One of t h e m o s t p o p u l a r e x p e r i m e n t a l s y s t e m s i s t h e i n - v i t r o c u l t u r e of m a m m a l i a n c e l l s . By t h e m i d - 1 9 5 0 s , i m p r o v e m e n t s in g r o w t h m e d i a and h a n d l i n g t e c h n i q u e s m a d e p o s s i b l e the c u l t u r e of s i n g l e i s o l a t e d c e l l s .

If a s i n g l e c e l l c a n d iv ide m a n y t i m e s and p r o d u c e a c o l o n y v i s i b l e to t he n a k e d e y e , it h a s p r o v i d e d u n e q u i v o c a l p r o o f t h a t i t s r e p r o d u c t i v e i n t e g r i t y i s i n t a c t . A c e l l t h a t i s unab le to f o r m a co lony i s , by d e f i n i t i o n , r e p r o d u c t i v e l y d e a d . T h e l o s s of r e p r o d u c t i v e i n t e g r i t y a s a f u n c t i o n of r a d i a t i o n d o s e i s known a s a d o s e - s u r v i v a l c u r v e . T h i s t e c h n i q u e w a s f i r s t d e s c r i b e d by P u c k and M a r c u s (1956), who u s e d H e L a c e l l s d e r i v e d f r o m a h u m a n c a r c i n o m a of t h e c e r v i x . S u b s e q u e n t l y i t p r o v e d p o s s i b l e to c u l t u r e a w i d e v a r i e t y of c e l l t y p e s , bo th n o r m a l and m a l i g n a n t , o r i g i n a t i n g in s e v e r a l s p e c i e s of m a m m a l s , and to d e t e r m i n e r a d i a t i o n d o s e - s u r v i v a l c u r v e s .

A t y p i c a l e x p e r i m e n t t o d e t e r m i n e a r a d i a t i o n s u r v i v a l c u r v e f o r c e l l s of a n e s t a b l i s h e d c e l l l i n e i s p e r f o r m e d a s f o l l o w s . C e l l s f r o m an a c t i v e l y g r o w i n g s t o c k c u l t u r e a r e p r e p a r e d into a s u s p e n s i o n of s i n g l e c e l l s and c o u n t e d w i t h a h e m a c y t o m e t e r o r e l e c t r o n i c c e l l c o u n t e r , a n d known n u m b e r s a r e s e e d e d in to P e t r i d i s h e s . S o m e d i s h e s a r e r e s e r v e d f o r c o n t r o l s , and o t h e r s e x p o s e d to a r a n g e of g r a d e d d o s e s of r a d i a t i o n .

T h e d i s h e s a r e t h e n i n c u b a t e d a t 37°C f o r a p e r i o d of one to two w e e k s , d e p e n d i n g upon the c e l l c y c l e of t h e p a r t i c u l a r c e l l l i n e , to a l l ow o p p o r t u n i t y f o r c e l l s w h i c h h a v e r e t a i n e d t h e i r r e p r o d u c t i v e i n t e g r i t y t o d iv ide m a n y t i m e s and f o r m v i s i b l e c o l o n i e s . E a c h c o l o n y c o n s i s t s of c e l l s w h i c h a r e a l l t h e p r o g e n y of a s i n g l e a n c e s t o r . A f t e r t h e c u l t u r e s a r e f i x e d and s t a i n e d , a c o u n t i s m a d e of t h e n u m b e r of c o l o n i e s p e r d i s h .

F r o m u n i r r a d i a t e d c o n t r o l d i s h e s , t h e p l a t i n g e f f i c i e n c y ( P . E . ) i s c a l c u l a t e d , d e f i n e d to be :

^ p g _ N u m b e r of c o l o n i e s c o u n t e d ^ ^qq 0 ' ' N u m b e r of c e l l s s e e d e d

27

2 8 CHAPTER 10

FIG.8 . Typical^survival curves for m a m m a l i a n cells exposed to radiat ion. Cel l surviving fract ion, on a logar i thmic scale , is plotted against dose on a l inear sca le . For densely ionizing radiations, such as a - r ays or low-energy neutrons, the dose response is a straight l ine (curve A) characterized by one parameter , the 37°lo dose slope (D 0 ) . For X or y-rays, which are sparsely ionizing, the dose-response relationship has an ini t ia l shoulder before tending to become straight (curve B). In this case, two parameters are needed to specify the curve: D0 and n, or al ternatively D0 and Dq.

T h e P . E . i s t y p i c a l l y 50 to 90%. I d e a l l y t h i s q u a n t i t y s h o u l d be 100%, bu t i t s e l d o m i s f o r a v a r i e t y of r e a s o n s , i n c l u d i n g s u b o p t i m a l g r o w t h m e d i u m , u n c e r t a i n t i e s i n c o u n t i n g t h e o r i g i n a l c e l l c o n c e n t r a t i o n , and t h e t r a u m a of h a n d l i n g and t r y p s i n i z a t i o n .

T h e f r a c t i o n of c e l l s s u r v i v i n g a g i v e n d o s e of r a d i a t i o n i s g i v e n by:

C o l o n i e s c o u n t e d S u r v i v i n g f r a c t i o n = _ .,-, :—, _ ~—rrvrz B C e l l s s e e d e d X P . E . / 1 0 0

T h e n u m b e r of c e l l s s e e d e d p e r d i s h i s a d j u s t e d a c c o r d i n g to t he r a d i a t i o n d o s e , s o t h a t a p p r o x i m a t e l y t h e s a m e n u m b e r of c e l l s s u r v i v e and g r o w up in to c o l o n i e s .

S u r v i v a l d a t a o b t a i n e d by the p r o c e d u r e o u t l i n e d a b o v e a r e u s u a l l y p r e s e n t e d in t h e f o r m s h o w n in F i g . 8, w i t h d o s e on a l i n e a r s c a l e and s u r v i v i n g f r a c t i o n on a l o g a r i t h m i c s c a l e . F o r d e n s e l y i o n i z i n g r a d i a t i o n s , s u c h a s a - r a y s o r l o w - e n e r g y n e u t r o n s , t h e s u r v i v a l d a t a c l o s e l y a p p r o x i -m a t e to a s t r a i g h t l i n e f r o m t h e o r i g i n ( s e e A in F i g . 8). In t h i s c a s e the s u r v i v a l c u r v e i s c o m p l e t e l y d e s c r i b e d by one p a r a m e t e r , D0 o r s l o p e , d e f i n e d to be t h e d o s e r e q u i r e d to r e d u c e t he c e l l p o p u l a t i o n by a f a c t o r of 1 / e , i . e . 0 . 37 .

BIOLOGICAL TEST SYSTEMS 29

F o r s p a r s e l y i o n i z i n g r a d i a t i o n s , the s u r v i v a l c u r v e h a s a b r o a d i n i t i a l s h o u l d e r a t low d o s e s , f o l l o w e d by a p o r t i o n w h i c h b e c o m e s s t r a i g h t o r a l m o s t s t r a i g h t in a s e m i l o g a r i t h m i c p lo t ( s e e c u r v e B in F i g . 8). At l e a s t two p a r a m e t e r s a r e n e c e s s a r y to s p e c i f y a c u r v e of t h i s k ind . F i r s t t he s l o p e , o r D0 , of t he s t r a i g h t p o r t i o n of t h e c u r v e , d e f i n e d a s b e f o r e . S e c o n d t h e e x t r a p o l a t i o n n u m b e r , w h i c h i s found by e x t r a p o l a t i n g t he s t r a i g h t p o r t i o n of t h e s u r v i v a l c u r v e un t i l i t c u t s t he ' s u r v i v i n g f r a c t i o n 1 a x i s .

T h e s i z e of t he e x t r a p o l a t i o n n u m b e r (n in F i g . 8) i s an i n d i c a t i o n of t h e e x t e n t of the s h o u l d e r e x h i b i t e d by the s u r v i v a l c u r v e . A l t e r n a t i v e l y , t he s h o u l d e r m a y be d e s c r i b e d by the ' q u a s i - t h r e s h o l d d o s e ' . T h i s q u a n t i t y (Dq in F i g . 8) i s t h e d o s e a t w h i c h t h e s t r a i g h t p o r t i o n of t h e s u r v i v a l c u r v e , e x t r a p o l a t e d b a c k w a r d s , c u t s t he d o s e a x i s .

S u r v i v a l c u r v e s h a v e b e e n d e t e r m i n e d f o r a g r e a t v a r i e t y of m a m m a l i a n c e l l s i n c u l t u r e , o r i g i n a t i n g i n n o r m a l o r m a l i g n a n t t i s s u e s i n s e v e r a l d i f f e r e n t s p e c i e s of a n i m a l s . W i t h f ew e x c e p t i o n s , t h e DQ f o r a l l c e l l s s t u d i e d w i t h X - r a y s f a l l s i n t h e r a n g e 1 0 0 - 200 r a d , w h i l e t h e e x t r a p o l a t i o n n u m b e r v a r i e s f r o m 1.5 to 10.

5 . 3 . IN-VIVO S U R V I V A L D A T A

T h e f i r s t i n - v i v o s u r v i v a l c u r v e w a s d e s c r i b e d by H e w i t t and W i l s o n i n 1959 . T h e s e w o r k e r s u s e d a l y m p h o c y t i c l e u k e m i a i n C B A m i c e , w h i c h w a s of s p o n t a n e o u s o r i g i n . T h e y p r e p a r e d a s i n g l e c e l l s u s p e n s i o n f r o m t h e i n f i l t r a t e d l i v e r of a n a n i m a l w i t h a d v a n c e d d i s e a s e , and found t h a t t h e t u m o u r cou ld b e t r a n s p l a n t e d by i n j e c t i n g a known n u m b e r of c e l l s in to t h e p e r i t o n e a l c a v i t i e s of r e c i p i e n t m i c e , w h i c h s u b s e q u e n t l y d e v e l o p e d l e u k e m i a . The l e u k e m i a cou ld be t r a n s m i t t e d , on t he a v e r a g e , by t h e i n j e c t i o n of on ly 2 c e l l s . T h i s q u a n t i t y i s known a s t h e T D g 0 . O t h e r d o n o r a n i m a l s w e r e t h e n g i v e n g r a d e d d o s e s of r a d i a t i o n , and t h e c o r r e s p o n d i n g TD 5 0 v a l u e s d e t e r m i n e d . T h e T D 5 0 s f o r t he i r r a d i a t e d a n i m a l s w e r e , of c o u r s e , m u c h b i g g e r t h a n f o r c o n t r o l b e c a u s e m a n y of t h e c e l l s w e r e s t e r i l i z e d by t h e r a d i a t i o n . F o r a g i v e n d o s e , t h e s u r v i v i n g f r a c t i o n i s g i v e n by:

S u r v i v i n g f r a c t i o n = — c o n t r o l TD 5 0 i r r a d i a t e d

T h i s p r o c e d u r e i s known a s t he d i lu t i on a s s a y t e c h n i q u e , and h a s b e c o m e t h e b a s i s f o r o b t a i n i n g c e l l s u r v i v a l c u r v e s f o r m a n y i n - v i v o s y s t e m s , i n c l u d i n g s o l i d t u m o u r s .

A n u m b e r of e x p e r i m e n t a l t r a n s p l a n t a b l e s o l i d t u m o u r s , s o m e of w h i c h a r o s e s p o n t a n e o u s l y in t h e f i r s t p l a c e w h i l e o t h e r s w e r e i n i t i a l l y i n d u c e d w i t h a c h e m i c a l c a r c i n o g e n , c a n be p a s s e d f r o m o n e g e n e r a t i o n of a n i m a l s to t h e n e x t . T u m o u r s f r o m d o n o r a n i m a l s a r e r e m o v e d , chopped up in to s m a l l p i e c e s , and the c e l l s p r e p a r e d in to a s i n g l e c e l l s u s p e n s i o n by t r y p s i n i z a t i o n . V a r i o u s known n u m b e r s of t h e s e c e l l s c a n t h e n be t r a n s -p l a n t e d s u b c u t á n e o us ly in to g r o u p s of r e c i p i e n t a n i m a l s . By i r r a d i a t i n g s o m e of t h e d o n o r a n i m a l s , i t i s p o s s i b l e to c a l c u l a t e t he TD 5 0 v a l u e s f o r c o n t r o l and i r r a d i a t e d a n i m a l s , and t h u s a r r i v e a t t h e f r a c t i o n of c e l l s s u r v i v i n g .

3 0 CHAPTER 10

3 0 0 0 R A D TO MOAT T E S T D O S E D TO C E N T R A L A R E A

O B S E R V E D R E G R O W T H O F S K I N N O D U L E IN

C E N T R A L A R E A

FIG. 9. Technique used to isolate an area of skin for experimental irradiation. A 30-kV X-ray machine is used to irradiate an annulus of skin to a massive dose to produce a 'moat ' of dead cells. An isolated area of intact skin in the centre is then given a test dose of radiation, and observed for the regrowth of skin nodules. (Redrawn from Withers, 1967).

5. 4 . SURVIVAL C U R V E F O R MOUSE SKIN C E L L S

It i s p o s s i b l e to d e t e r m i n e t h e s u r v i v a l c u r v e f o r e p i t h e l i a l c e l l s i n t h e m o u s e by m e a n s of a n i n g e n i o u s t e c h n i q u e d e v i s e d by W i t h e r s (1967) . T h e b a s i s of t he t e c h n i q u e i s i l l u s t r a t e d i n F i g u r e 9. H a i r i s p l u c k e d f r o m a n a r e a on t h e b a c k of t h e m o u s e , and a l o w - v o l t a g e X - r a y m a c h i n e i s u s e d t o i r r a d i a t e a n a n n u l u s of s k i n to a m a s s i v e d o s e of t h e o r d e r of 3000 r a d . T h i s p r o d u c e s a ' m o a t ' of d e a d c e l l s . In t he c e n t r e of t h i s m o a t i s an i s o l a t e d i s l a n d of i n t a c t c e l l s , w h i c h h a v e b e e n p r o t e c t e d d u r i n g the f i r s t e x p o s u r e by m e a n s of a s m a l l m e t a l s p h e r e (in f a c t a b a l l b e a r i n g w a s u s e d ) .

T h i s s m a l l a r e a of i n t a c t s k i n w a s t h e n g i v e n a t e s t d o s e of r a d i a t i o n and s u b s e q u e n t l y o b s e r v e d f o r r e g r o w t h of s k i n . I s o l a t e d n o d u l e s of s k i n r e g r o w i n g w i th in t h i s a r e a s o m e d a y s l a t e r i n d i c a t e d t h a t one o r m o r e s k i n c e l l s had s u r v i v e d . If no c e l l s s u r v i v e d i n t h e t e s t a r e a , h e a l i n g cou ld on ly t a k e p l a c e a t a m u c h l a t e r d a t e by i n f i l t r a t i o n of c e l l s a c r o s s t h e h e a v i l y i r r a d i a t e d m o a t . To o b t a i n a s u r v i v a l c u r v e , t e s t a r e a s of d i f f e r e n t s i z e w e r e i r r a d i a t e d to a r a n g e of d o s e s , and t h e n u m b e r of s k i n n o d u l e s c o u n t e d . T h i s w a s t h e f i r s t i n - v i v o s u r v i v a l c u r v e f o r a n o r m a l t i s s u e .

5 . 5 . SURVIVAL C U R V E F O R MOUSE J E J U N A L C R Y P T C E L L S

W i t h e r s and E l k i n d (1970) d e s c r i b e d a t e c h n i q u e to m e a s u r e t h e s u r v i v a l c h a r a c t e r i s t i c of t h e c r y p t c e l l s in t h e m o u s e j e j u n u m . G r o u p s of adu l t m i c e w e r e g i v e n g r a d e d d o s e s of X - r a y s i n t h e r a n g e 1000 - 2600 r a d t o t h e w h o l e b o d y . Abou t 3 | d a y s a f t e r i r r a d i a t i o n , t h e a n i m a l s w e r e s a c r i f i c e d and s e g m e n t s of t he j e j u n u m r e m o v e d . A f t e r r o u t i n e h i s t o l o g i c a l p r o c e s s i n g , t r a n s v e r s e s e c t i o n s of t h e j e j u n u m w e r e e x a m i n e d , and t h e n u m b e r of v i a b l e c r y p t s s c o r e d .

A c r y p t w a s a s s u m e d t o b e r e g e n e r a t i n g if t h e r e w e r e 10 o r m o r e c e l l s , e a c h w i t h a p r o m i n e n t n u c l e u s and l i t t l e c y t o p l a s m , ly ing c l o s e t o g e t h e r and a p p e a r i n g c r o w d e d . N o n - v i a b l e c r y p t s c o n t a i n e d no c e l l s , o r w e r e s p a r s e l y

FIG. 10. Photograph or a mouse's spleen. The mouse has been supralethally irradiated to steri l ize all the cells of its spleen, before nucleated isologous bone marrow cells from a donor mouse were in jec ted intravenously. Some of these cells are trapped in the spleen and grow into colonies or nodules. (Photograph kindly supplied by Dr. A. Carsten).

BIOLOGICAL TEST SYSTEMS 1192

p o p u l a t e d by e n l a r g e d c e l l s wi th p r o m i n e n t e o s i n o p h i l i c c y t o p l a s m . T h e n u m b e r of r e g e n e r a t i n g c r y p t s in e a c h t r a n s v e r s e s e c t i o n w a s r e c o r d e d , and c o m p a r e d wi th c o n t r o l s , w h i c h on a v e r a g e c o n t a i n 160 c r y p t s p e r c i r c u m f e r e n c e .

F r o m t h e d a t a o b t a i n e d w i t h g r a d e d d o s e s of X - r a y s i t i s p o s s i b l e to c o n s t r u c t a s u r v i v a l c u r v e f o r t h e s t e m c e l l s c o n s t i t u t i n g t h e c r y p t s of t h e j e j u n u m .

5. 6 . THE S P L E E N C O L O N Y ASSAY S Y S T E M

A s y s t e m w a s d e v e l o p e d by M c C u l l o c h and T i l l (1962) to d e t e r m i n e t he s u r v i v a l c u r v e f o r t he c o l o n y - f o r m i n g c e l l s in t he bone m a r r o w of m i c e . R e c i p i e n t a n i m a l s w e r e s u p r a l e t h a l l y i r r a d i a t e d wi th a d o s e of abou t 900 r a d . T h i s d o s e s t e r i l i z e s a l l of t he p o t e n t i a l l y d iv id ing c e l l s in t he a n i m a l ' s s p l e e n . N u c l e a t e d i s o l o g o u s bone m a r r o w c e l l s , t a k e n f r o m a d o n o r a n i m a l , w e r e t h e n i n j e c t e d i n t r a v e n o u s l y i n to t h e r e c i p i e n t a n i m a l s ; s o m e of t h e c e l l s l o d g e d in the s p l e e n , w h e r e t h e y f o r m e d n o d u l e s 10 to 11 d a y s l a t e r . At t h i s t i m e the s p l e e n s w e r e r e m o v e d , and the n o d u l e s c o u n t e d . A p i c t u r e of a s p l e e n wi th n o d u l e s i s s h o w n in F i g . 10. It w a s found t h a t , o n a v e r a g e , 1 0 4 c e l l s had to be i n o c u l a t e d in to a r e c i p i e n t a n i m a l in o r d e r to p r o d u c e a c o l o n y . T h i s low ' p l a t i n g e f f i c i e n c y ' c a n be a t t r i b u t e d to t h e f a c t t h a t m o s t of t h e c e l l s i n t h e n u c l e a t e d bone m a r r o w s u s p e n s i o n a r e f u l l y d i f f e r e n t i a t e d c e l l s , and would n e v e r b e c a p a b l e u n d e r any c i r c u m -s t a n c e s of f o r m i n g a c o l o n y . By i r r a d i a t i n g s o m e of t he d o n o r a n i m a l s to g r a d e d d o s e s i t i s p o s s i b l e to p r o d u c e a s u r v i v a l c u r v e f o r t h e c o l o n y - f o r m i n g c e l l s i n t h e bone m a r r o w .

5 . 7 . S E E D L I N G S O F VICIA F A B A

L o n g b e f o r e t he a d v e n t of c e l l c u l t u r e t e c h n i q u e s , e i t h e r i n v i v o o r in v i t r o , r a d i o b i o l o g i c a l t e s t s y s t e m s w e r e u s e d in w h i c h t h e e n d - p o i n t s

3 2 CHAPTER 10

s c o r e d d e p e n d e d i n one way o r a n o t h e r upon the k i l l i ng of c e l l s . F o r e x a m p l e , a s e a r l y a s 1913 M o t t r a m w a s u s i n g the g r o w t h i n h i b i t i o n of t h e p r i m a r y r o o t of s e e d l i n g s of V i c i a f a b a a s a m e t h o d of s c o r i n g r a d i a t i o n d a m a g e .

D u r i n g the 1930s and 1940s t h i s s i m p l e s e e d l i n g s y s t e m w a s u s e d to e l u c i d a t e m a n y of t he b a s i c c h a r a c t e r i s t i c s of r a d i a t i o n r e s p o n s e . T h e v a r i a t i o n of b i o l o g i c a l e f f e c t i v e n e s s wi th r a d i a t i o n q u a l i t y , t h e o x y g e n e f f e c t , t he d o s e r a t e e f f e c t , a s w e l l a s r e p a i r b e t w e e n s p l i t d o s e s , w e r e a l l i n v e s t i g a t e d wi th V i c i a s e e d l i n g s ( G r a y and Read , 1942) .

W h i l e t h e r e a r e c o m p l i c a t i n g f a c t o r s , s u c h a s m i t o t i c d e l a y and e f f e c t s on d i f f e r e n t i a t i o n , t h e r e d u c e d g r o w t h of an i r r a d i a t e d p l a n t s e e d l i n g i s due , p r i m a r i l y , to t h e k i l l i ng of a p r o p o r t i o n of d iv id ing c e l l s in t h e m e r i s t e m .

5 . 8 . A N I M A L L E T H A L I T Y DUE TO W H O L E - B O D Y IRRADIATION

In t he e a r l y d a y s of r a d i o b i o l o g y , m a n y e x p e r i m e n t s w e r e p e r f o r m e d in w h i c h a n i m a l s w e r e e x p o s e d t o t a l - b o d y to r a d i a t i o n , and the on ly e n d -p o i n t s c o r e d w a s t he d e a t h of t he a n i m a l . In t he c a s e of t he m o u s e , and indeed of any of t h e s m a l l r o d e n t s , two d i s t i n c t m o d e s of d e a t h c a n b e d i s t i n g u i s h e d a s a r e s u l t of t o t a l - b o d y i r r a d i a t i o n (Bond, F l i e d n e r and A r c h a m b e a u , 1965; L a n g h a m , 1967) .

A f t e r d o s e s i n t h e r a n g e of 400 to 800 r a d , a n i m a l s d i e s o m e w e e k s l a t e r a s a r e s u l t of d a m a g e to t h e h a e m a t o p o i e t i c s y s t e m . A d o s e of t h i s o r d e r h a s l i t t l e o r no e f f e c t on t he c i r c u l a t i n g b l o o d c e l l s , w h i c h a r e m a t u r e f u n c t i o n i n g c e l l s i n c a p a b l e of f u r t h e r d i v i s i o n s . T h e r a d i a t i o n k i l l s a p r o -p o r t i o n of t h e p r i m i t i v e s t e m c e l l s i n t he bone m a r r o w , t h e l y m p h n o d e s and t h e s p l e e n . No e f f e c t s of t he r a d i a t i o n a r e a p p a r e n t f o r s e v e r a l w e e k s un t i l t h e c i r c u l a t i n g c e l l s i n t h e b lood r e a c h t h e end of t h e i r u s e f u l l i f e , • d ie , and a r e r e m o v e d . R e p l a c e m e n t b lood c e l l s a r e no t a v a i l a b l e s i n c e t he r a d i a t i o n k i l l e d t h e p r i m i t i v e s t e m c e l l s . C o n s e q u e n t l y , t h e e x p o s e d a n i m a l s u f f e r s f r o m h a e m o r r h a g e s due to a l a c k of p l a t e l e t s , w e a k n e s s b e c a u s e of a n a e m i a ( lack of r e d c e l l s ) , and a t t he s a m e t i m e r e s i s t a n c e to i n f e c t i o n i s a t a m i n i m u m b e c a u s e of t he s p a r s i t y of w h i t e c e l l s . A n i m a l s d ie up to 30 d a y s p o s t - i r r a d i a t i o n . 1

At h i g h e r d o s e s , i n e x c e s s of a 1000 r a d , d e a t h o c c u r s 3 t o 4 d a y s a f t e r i r r a d i a t i o n , due t o a denud ing of t he l in ing of t h e g u t . T h e gut i s a c l e a r e x a m p l e of a s e l f - r e n e w a l t i s s u e . T h e s u r f a c e of the gu t i s c o m p o s e d of a v a s t n u m b e r of v i l l i , f i n g e r - l i k e p r o t r u s i o n s c o m p o s e d of c e l l s w h i c h a r e f u l l y d i f f e r e n t i a t e d , a b l e to p e r f o r m the s p e c i a l f u n c t i o n of a b s o r b i n g n o u r i s h -m e n t f r o m t h e food , but i n c a p a b l e of f u r t h e r d i v i s i o n s . T h e s e f u n c t i o n a l c e l l s a r e c o n t i n u a l l y w o r n a w a y and s l o u g h e d off f r o m t h e t o p s of t h e v i l l i , and m u s t be c o n t i n u o u s l y r e p l a c e d by c e l l s b o r n in t he c r y p t s , w h i c h a r e f a c t o r i e s of a c t i v e l y d iv id ing c e l l s l o c a t e d a t t he b a s e of e a c h v i l l u s . W h e n the gu t i s e x p o s e d to a d o s e of abou t 1000 r a d , m a n y of t h e c r y p t c e l l s a r e k i l l e d , t h u s c u t t i n g off t he s u p p l y of new d i f f e r e n t i a t e d c e l l s . T h e e f f e c t of t he r a d i a t i o n i s no t a p p a r e n t i m m e d i a t e l y , b e c a u s e the d i f f e r e n t i a t e d f u n c t i o n i n g c e l l s of t he v i l l i a r e u n a f f e c t e d by the r a d i a t i o n , and c o n t i n u e t o p e r f o r m t h e i r s p e c i a l i z e d f u n c t i o n s . H o w e v e r , in t he c o u r s e of t i m e , a s t h e v i l l i a r e w o r n away and s l o u g h e d of f , t h e r e a r e no new c e l l r e p l a c e -m e n t s . A s a r e s u l t , t he gu t i s d e n u d e d of v i l l i , and g r o s s h o l e s a p p e a r . D e a t h s o o n f o l l o w s a s a r e s u l t of l o s s of body f l u i d s , and g r o s s i n f e c t i o n s .

BIOLOGICAL TEST SYSTEMS 3 3

D e a t h due t o t o t a l - b o d y i r r a d i a t i o n , t h e r e f o r e , i s u l t i m a t e l y a r e s u l t of c e l l k i l l i n g i n a c r i t i c a l s e l f - r e n e w a l t i s s u e , e i t h e r t he h a e m a t o p o i e t i c s y s t e m o r t h e g a s t r o i n t e s t i n a l t r a c t , d e p e n d i n g upon t h e d o s e l e v e l . T h e s e r a t h e r c r u d e e n d - p o i n t s , w h i c h w e r e used in t he e a r l y d a y s of r a d i o b i o l o g y , p r o v i d e d r o u g h e s t i m a t e s of t h e r e l a t i v e b i o l o g i c a l e f f e c t i v e n e s s of n e u t r o n s p r o d u c e d by f i s s i o n c o m p a r e d wi th g a m m a r a y s o r X - r a y s .

(B) S y s t e m s invo lv ing a m u t a t i o n o r t r a n s f o r m a t i o n

5 . 9 . G E N E T I C M U T A T I O N S

R a d i a t i o n c a n p r o d u c e i r r e p a r a b l e d a m a g e in t h e g e n e t i c a p p a r a t u s of c e l l s w h i c h s u r v i v e t h e r a d i a t i o n e x p o s u r e . Such a c h a n g e i s s a i d to be a m u t a t i o n . If a m u t a t i o n o c c u r s in a g e r m c e l l , t he c o n s e q u e n c e s w i l l be f e l t , not by t h e i n d i v i d u a l in w h i c h t h e c h a n g e t a k e s p l a c e , but by f u t u r e g e n e r a t i o n s . A m u t a t i o n in a s o m a t i c c e l l , by c o n t r a s t , m a y be h a r m f u l to t h e i n d i v i d u a l e x p o s e d bu t w i l l no t be p a s s e d on to h i s d e s c e n d a n t s .

U n t i l r e l a t i v e l y r e c e n t y e a r s , t h e on ly a v a i l a b l e d a t a on r a d i a t i o n -i n d u c e d g e n e t i c e f f e c t s c a m e f r o m e x p e r i m e n t s wi th t h e f r u i t f l y D r o s o p h i l a . In t h i s c r e a t u r e it i s p o s s i b l e to s t u d y l e t h a l m u t a t i o n s , o r to s t u d y g e n e m u t a t i o n s w h i c h r e s u l t i n c h a n g e s in t h e a p p e a r a n c e of t h e f r u i t - f l y , s u c h a s m i s - s h a p e d w i n g s , o r a c h a n g e i n e y e c o l o u r . In t h i s s y s t e m , t h e r e i s a l i n e a r r e l a t i o n s h i p b e t w e e n t h e i n c i d e n c e of m u t a t i o n s and r a d i a t i o n d o s e . In add i t i on , t h e n u m b e r of m u t a t i o n s p r o d u c e d by a g i v e n d o s e of X - r a y s i s t h e s a m e , w h e t h e r t h e d o s e i s d e l i v e r e d in a s i n g l e e x p o s u r e , o r s p r e a d ou t o v e r a p e r i o d of t i m e in a s e r i e s of d o s e f r a c t i o n s ( S p e n c e r and S t e r n , 1948) .

In t he y e a r s f o l l owing t h e S e c o n d W o r l d W a r , l a r g e - s c a l e e x p e r i m e n t s h a v e b e e n u n d e r t a k e n to d e t e r m i n e m u t a t i o n r a t e s in t h e m o u s e . S p e c i f i c g e n e m u t a t i o n s m a y b e s c o r e d , w h i c h c a u s e e a s i l y r e c o g n i z e d c h a n g e s in c o a t c o l o u r and obvio.us d e f e c t s of t h e e a r o r t a i l . In t h i s m a m m a l i a n s y s t e m , r a d i a t i o n p r o d u c e d g e n e t i c e f f e c t s f o r m a v e r y c o m p l i c a t e d p i c t u r e . E a c h m u t a t i o n h a s a d i f f e r e n t r a d i o s e n s i t i v i t y , and t h e r e i s an i m p o r t a n t d o s e - r a t e e f f e c t ( R u s s e l l , 1963a ,b ) .

5 . 1 0 . SOMATIC M U T A T I O N S

S o m a t i c m u t a t i o n s m a y be s t u d i e d in a s i m p l e s y s t e m s u c h a s T r a d e s -c a n t i a , by o b s e r v i n g e a s i l y r e c o g n i z e d c o l o u r c h a n g e s in t h e c e l l s of t he s t a m e n h a i r s ( S p a r r o w , U n d e r b r i n k and R o s s i , 1972). One of t he a d v a n t a g e s of t h i s s y s t e m i s t h a t a v a s t n u m b e r of s a m p l e s m a y be s t u d i e d , s o t h a t t he e f f e c t s of v e r y s m a l l d o s e s of r a d i a t i o n m a y be i n v e s t i g a t e d .

5 . 1 1 . N E O P L A S T I C T R A N S F O R M A T I O N S

T h e i n d u c t i o n of t u m o u r s a s a r e s u l t of r a d i a t i o n h a s b e e n o b s e r v e d f r o m the e a r l i e s t u s e s of X - r a y s . T h e m e c h a n i s m by w h i c h r a d i a t i o n c a r c i n o g e n e s i s o c c u r s i s by no m e a n s c l e a r . An a t t r a c t i v e t h e o r y s p e c u l a t e s t h a t c a r c i n o g e n e s i s r e s u l t s f r o m the m u t a t i o n of a n o r m a l t i s s u e c e l l , but t h i s m u s t be on ly p a r t of a m u c h m o r e c o m p l i c a t e d p i c t u r e .

34 CHAPTER 10

S t u d i e s of c a r c i n o g e n e s i s in a n i m a l s h a v e b e e n of two t y p e s . F i r s t , t o t a l - b o d y i r r a d i a t i o n i s c a r r i e d out at low to i n t e r m e d i a t e d o s e s us ing s t r a i n s of a n i m a l s w h i c h a r e p e c u l i a r l y s e n s i t i v e and show a h igh n a t u r a l i n c i d e n c e of t he n e o p l a s m t h a t i s be ing s t u d i e d . S e c o n d , v e r y l a r g e d o s e s of r a d i a t i o n , w h i c h would b e l e t h a l if g i v e n w h o l e - b o d y , a r e a p p l i e d t o l i m i t e d a r e a s of t i s s u e .

Many e x p e r i m e n t s h a v e b e e n p e r f o r m e d to s t u d y t h e i n c i d e n c e of l e u k e m i a in .mice e x p o s e d to w h o l e - b o d y r a d i a t i o n , b e c a u s e i t w a s though t t h a t the r i s k of l e u k e m i a w a s t he m o s t s e r i o u s h a z a r d t h a t h u m a n b e i n g s f a c e d a f t e r i r r a d i a t i o n .

The p r o d u c t i o n of s o l i d t u m o u r s in a n i m a l s i s m o r e d i f f i c u l t to s t u d y . T h e r e a r e n u m e r o u s r e p o r t s i n t he l i t e r a t u r e of t h e i n d u c t i o n of m a m m a r y t u m o u r s by i o n i z i n g r a d i a t i o n s in m i c e , g u i n e a p i g s and r a t s . T h e m o s t e x t e n s i v e s t u d i e s h a v e b e e n c a r r i e d out in t h e S p r a g u e D a w l e y r a t . In th i s s p e c i e s t h e r e i s a h igh n a t u r a l i n c i d e n c e of m a m m a r y n e o p l a s m s l a t e in l i f e , a l t h o u g h s u c h t u m o u r s a r e not f r e q u e n t l y o b s e r v e d d u r i n g the f i r s t 18 m o n t h s . If a d o s e of r a d i a t i o n i s d e l i v e r e d e a r l y i n l i f e , in abou t the 6 th w e e k , a l a r g e n u m b e r of t u m o u r s m a y be c o u n t e d d u r i n g t h e f i r s t y e a r of l i f e w h e n the c o n t r o l b a c k g r o u n d r a t e i s v e r y low ( S h e l l a b a r g e r , C r o n k i t e , Bond and L i p p i n c o t t , 1957) .

C h a p t e r 6

R E L A T I V E BIOLOGICAL E F F E C T I V E N E S S

6 . 1 . D E F I N I T I O N

T h e f o r m a l d e f i n i t i o n of t he r e l a t i v e b i o l o g i c a l e f f e c t i v e n e s s (RBE) i s a s f o l l o w s . T h e R B E of s o m e t e s t r a d i a t i o n r c o m p a r e d wi th t h e s t a n d a r d r a d i a t i o n i s d e f i n e d by t h e r a t i o D s / D r , w h e r e Ds and Dj a r e t h e d o s e s of t h e s t a n d a r d r a d i a t i o n and t h e t e s t r a d i a t i o n , r e s p e c t i v e l y , r e q u i r e d f o r e q u a l b i o l o g i c a l e f f e c t . T h e s t a n d a r d r a d i a t i o n i s u s u a l l y 250 kV X - r a d i a t i o n o r g a m m a r a d i a t i o n .

R B E t u r n s ou t to be a c o m p l i c a t e d q u a n t i t y , s i n c e i t v a r i e s w i t h t he b i o l o g i c a l s y s t e m u s e d , and a l s o w i t h t he d o s e l e v e l , o r l e v e l of b i o l o g i c a l d a m a g e i n t h a t g i v e n s y s t e m . In add i t i on , t h e R B E i s a l s o found to v a r y c o n s i d e r a b l y if t he d o s e - r a t e c h a n g e s o v e r a w i d e r a n g e .

6 . 2 . R B E AS A F U N C T I O N O F DOSE

In t h e e a r l y d a y s of r a d i o b i o l o g y , t he s t u d y of R B E a p p e a r e d to be /

s t r a i g h t f o r w a r d b e c a u s e on ly s i m p l e t e s t s y s t e m s w e r e a v a i l a b l e w h i c h had a s i n g l e u n e q u i v o c a l e n d - p o i n t . To i l l u s t r a t e t h e po in t we wi l l d i s c u s s a s p e c i f i c e x a m p l e . •

S u p p o s e i t i s d e c i d e d to m e a s u r e t h e R B E of f a s t n e u t r o n s c o m p a r e d w i t h g a m m a r a y s , u s i n g a s a b i o l o g i c a l t e s t s y s t e m t h e l e t h a l i t y t f p l a n t s e e d l i n g s . G r o u p s of p l a n t s would b e e x p o s e d to v a r i o u s d o s e s of g a m m a r a y s , w h i l e p a r a l l e l g r o u p s would b e e x p o s e d to g r a d e d n e u t r o n d o s e s . At t h e end of t h e p e r i o d of o b s e r v a t i o n i t wou ld be p o s s i b l e to c a l c u l a t e t h e d o s e of g a m m a r a y s , and t h e n t h e d o s e of n e u t r o n s , w h i c h c a u s e d the d e a t h of hal f t h e p l a n t s i n t h e g r o u p . T h i s q u a n t i t y i s known a s t he LD 5 0 , t he m e a n l e t h a l d o s e . In a t y p i c a l c a s e t he L D g 0 f o r g a m m a r a y s m a y b e 450 r a d w h i l e t he c o r r e s p o n d i n g q u a n t i t y f o r f a s t n e u t r o n s m a y be 300 r a d . T h e R B E of n e u t r o n s r e l a t i v e to g a m m a r a y s i s t h e n s i m p l y t h e r a t i o 4 5 0 / 3 0 0 , i . e . 1 .5 .

T h e s i t u a t i o n b e c o m e s m o r e c o m p l i c a t e d i f , i n s t e a d , a t e s t s y s t e m i s c h o s e n w h e r e i t i s p o s s i b l e t o o b s e r v e t he b i o l o g i c a l r e s p o n s e o v e r a r a n g e of d o s e s . T h i s i s the c a s e w h e n c e l l s u r v i v a l c u r v e s a r e d e t e r m i n e d . W h e n X - r a y s and 1 - M e V n e u t r o n s a r e c o m p a r e d , t h e s u r v i v a l c u r v e s h a v e d i f f e r e n t s h a p e s . The X - r a y s u r v i v a l c u r v e h a s an i n i t i a l s h o u l d e r , wh i l e t he n e u t r o n c u r v e i s an e x p o n e n t i a l f u n c t i o n of d o s e . C o n s e q u e n t l y , t he R B E w i l l d e p e n d upon t h e l e v e l of b i o l o g i c a l d a m a g e (and t h e r e f o r e d o s e ) c h o s e n .

T h i s po in t i s i l l u s t r a t e d in F i g . 11, w h i c h s h o w s s u r v i v a l c u r v e s o b t a i n e d w i t h m a m m a l i a n c e l l s in c u l t u r e e x p o s e d to a r a n g e of d o s e s of , on t h e one hand f a s t n e u t r o n s , on t he o t h e r hand 2 5 0 - k V X - r a y s . T h e R B E m a y b e c a l c u l a t e d f r o m t h e s e s u r v i v a l c u r v e s a s t he r a t i o of d o s e s n e e d e d to p r o d u c e t h e s a m e b i o l o g i c a l e f f e c t . If the e n d - p o i n t c h o s e n f o r c o m p a r i s o n i s a s u r v i v i n g f r a c t i o n of 0. 02, t h e n t h e d o s e of n e u t r o n s n e c e s s a r y i s 400 r a d , w h i l e t h e c o r r e s p o n d i n g d o s e of X - r a y s i s 1200 r a d . T h e R B E i s t h e n t h e

35

3 6 CHAPTER 10

DOSE (rod)

FIG. 11. Typica l survival curves for m a m m a l i a n cells exposed to 1 -MeV fast neutrons and X-rays. The survival curves for the two types of radiat ion have different shapes, and as a consequence the RBE is a function of dose.

10

~1 i i i i j i i i i i i i i_

A C U T E E X P O S U R E S 1-MeV neutrons

_i L 100

N E U T R O N DOSE ( r o d )

- i i I ' ' ' ' 1000

FIG. 12. RBE as a function of neutron dose, calculated from the data in Fig. 11. RBE varies inversely with the square root of the neutron dose excep t at low doses where i t tends towards a l imit ing va lue .

q u o t i e n t , 1 2 0 0 / 4 0 0 , i . e . 3 .0 . If , h o w e v e r , t he c o m p a r i s o n i s m a d e a t t h e s u r v i v i n g f r a c t i o n of 0 .6 , i t i s e v i d e n t t h a t t h e n e u t r o n d o s e r e q u i r e d i s on ly 64 r a d w h i l e t h e c o r r e s p o n d i n g X - r a y i s 300 r a d ; t he r e s u l t a n t R B E i s 3 0 0 / 6 4 , i . e . 4 .7 .

It i s m o s t i n s t r u c t i v e to p l o t the R B E a g a i n s t t h e n e u t r o n d o s e . F o r m a n y b i o l o g i c a l s y s t e m s , a l i n e a r r e l a t i o n s h i p h a s b e e n f o u n d . T h i s i s i l l u s t r a t e d in F i g . 12, O v e r a w ide r a n g e of d o s e s t h e l i ne h a s a s l o p e of m i n u s h a l f , i n d i c a t i n g t h a t R B E i s i n v e r s e l y p r o p o r t i o n a t e t o t h e s q u a r e r o o t of t h e d o s e .

RELATIVE BIOLOGICAL EFFECTIVENESS 3 7

DOSE (rod) 600 800 1000 1200 1400 1600

R A D I U M G A M M A - R A Y S

FIG. 13. At low dose-rate , survival curves for gamma rays become exponential functions of dose, but the slope of the survival curve varies with dose-rate . For neutrons, the slope of the survival curve varies l i t t le with dose-ra te . In this situation, RBE no longer varies with dose, but now varies with dose-rate as shown in Fig. 14.

GAMMA-RAY DOSE-RATE ( r a d / h )

20 30 40 50 60 80 100

FIG. 14. Variation with dose-rate of the RBE of 252Cf neutrons compared with radium gamma rays, calculated from the data of Fig. 13.

6 . 3 . R B E AS A F U N C T I O N O F D O S E - R A T E

A f u r t h e r c o m p l i c a t i o n o c c u r s if t h e r a d i a t i o n i s d e l i v e r e d a t low d o s e -r a t e s , s o t h a t t h e e x p o s u r e t i m e e x t e n d s o v e r m a n y h o u r s o r d a y s . In th i s c a s e t h e s u r v i v a l c u r v e f o r g a m m a r a y s v a r i e s c r i t i c a l l y wi th d o s e - r a t e , w h e r e a s t he s u r v i v a l c u r v e f o r n e u t r o n s v a r i e s v e r y l i t t l e w i t h d o s e - r a t e (F ig . 13). A s a r e s u l t i t i s p o s s i b l e to o b t a i n a who le r a n g e of R B E v a l u e s

38 CHAPTER 10

DOSE ( r o d )

FIG. 15. The changing shape of survival curves with LET. As the LET increases in going from X-rays to fast neutrons the slope of the survival curve gets steeper, while the extrapolation number is reduced.

" 1 — 100

T 1000 0.1

T " 10

L E T ^ t k e V / j i m of tissue)

FIG. 16. Variation with LET of the RBE as determined for m a m m a l i a n cells in culture. Curves 1, 2 and 3 refer to ce l l survival levels of 0. 8, 0 . 1 and 0 . 0 1 . For a given LET, RBE depends on the level of biological damage, and therefore on the dose. (From Barendsen, 1968).

RELATIVE BIOLOGICAL EFFECTIVENESS 3 9

LET^ (keV/pmof unit density tissue)

FIG. 17. OER, as determined with cultured m a m m a l i a n cells, plotted as a function of LET. Closed circles represent data with monoenerget ic charged part icles. The open tr iangle represents 250-kVp X-rays with a track average LET of 1 . 3 k e V / p m . (From Barendsen et a l . , 1966).

d e p e n d i n g upon w h i c h s u r v i v a l c u r v e s a r e c o m p a r e d . T h i s i s i l l u s t r a t e d in F i g . 14. U n d e r t h e s e c i r c u m s t a n c e s , R B E v a l u e s m a k e l i t t l e s e n s e u n l e s s d o s e - r a t e s a r e c h o s e n w h i c h r e s u l t in the s a m e a m o u n t of b i o l o g i c a l d a m a g e in t h e s a m e o v e r a l l e x p o s u r e t i m e .

6 . 4 . R B E AS A F U N C T I O N O F L E T

A s the L E T i n c r e a s e s , t he c h a r a c t e r i s t i c s h a p e of t h e s u r v i v a l c u r v e c h a n g e s . F o r a c u t e g a m m a - r a y e x p o s u r e s , t he c e l l s u r v i v a l c u r v e h a s a l a r g e i n i t i a l s h o u l d e r a t low d o s e s , and t e n d s to b e c o m e s t r a i g h t e r a t h i g h e r d o s e s . F o r a l p h a r a y s , t he s u r v i v a l c u r v e i s an e x p o n e n t i a l f u n c t i o n of d o s e . F o r n e u t r o n s of e n e r g i e s i n e x c e s s of a few MeV, t he s h a p e i s i n t e r -m e d i a t e b e t w e e n t h e s e two e x t r e m e s . In g e n e r a l , a s t h e L E T of t h e r a d i a t i o n i n c r e a s e s , t h e s l o p e o f t h e s u r v i v a l c u r v e g e t s s t e e p e r , and the s i z e of t h e i n i t i a l s h o u l d e r g e t s s m a l l e r . T h i s i s i l l u s t r a t e d in F i g . 15.

U s i n g m a m m a l i a n c e l l s in c u l t u r e , R B E v a l u e s h a v e b e e n d e t e r m i n e d f o r m a n y d i f f e r e n t r a d i a t i o n s , s p a n n i n g a w i d e r a n g e of L E T v a l u e s . In F i g . 16 R B E i s p l o t t e d a s a f u n c t i o n of L E T . A s the L E T i n c r e a s e s , t he R B E i n c r e a s e s s l o w l y a t f i r s t , and t h e n m o r e r a p i d l y a s t he L E T e x c e e d s 10 k e V / / u m . B e t w e e n 10 and 100 k e V / V m t h e R B E i n c r e a s e s r a p i d l y w i t h i n c r e a s i n g L E T , and in f a c t r e a c h e s a m a x i m u m at a r o u n d 100 k e V / j u m . B e y o n d t h i s v a l u e f o r t h e L E T t h e R B E f a l l s t o a l o w e r v a l u e owing to t he p h e n o m e n o n of ' o v e r - k i l l ' .

6 . 5 . O E R AS A F U N C T I O N O F L E T

A m o s t i m p o r t a n t r e l a t i o n s h i p e x i s t s b e t w e e n L E T and the o x y g e n e n h a n c e m e n t r a t i o (OER) . As d i s c u s s e d in C h a p t e r 7, v a r i o u s t y p e s of r a d i a t i o n e x h i b i t q u i t e d i f f e r e n t O E R s . F o r g a m m a r a y s w h i c h a r e s p a r s e l y i o n i z i n g , and h a v e a low L E T , t h e O E R h a s a v a l u e of a b o u t 2 .5 to 3.

4 0 CHAPTER 10

N e u t r o n s of 14 M e V , w h i c h a r e of i n t e r m e d i a t e i o n i z i n g d e n s i t y , show c h a r a c t e r i s t i c a l l y an O E R of a p p r o x i m a t e l y 1 .6 . F o r h i g h - L E T a l p h a p a r t i c l e s , t he O E R t e n d s t o w a r d s un i ty . B a r e n d s e n and h i s c o l l e a g u e s (1966) u s e d m a m m a l i a n c e l l s c u l t u r e d i n v i t r o to i n v e s t i g a t e t h e O E R f o r a w h o l e r a n g e of r a d i a t i o n t y p e s e n c o m p a s s i n g a w i d e s p e c t r u m of L E T v a l u e s . T h e i r r e s u l t s a r e s h o w n in F i g . 17, w h e r e O E R i s p l o t t e d a s a f u n c t i o n of L E T . At low L E T , c o r r e s p o n d i n g to X o r g a m m a r a y s , t he O E R i s b e t w e e n 2.5 and 3; a s t h e L E T i n c r e a s e s , t h e O E R f a l l s s l o w l y a t f i r s t , un t i l t h e L E T e x c e e d s a v a l u e of about 60 k e V / p m , a f t e r wh ich t h e O E R f a l l s r a p i d l y arid r e a c h e s uni ty by t h e t i m e the L E T h a s r e a c h e d a p p r o x i m a t e l y 200 k e V / ^ m .

C h a p t e r 7

E F F E C T OF M O L E C U L A R OXYGEN

7 . 1 . H I S T O R I C A L

A l a r g e n u m b e r of c h e m i c a l and p h a r m a c o l o g i c a l a g e n t s h a v e b e e n d i s c o v e r e d w h i c h m o d i f y t he b i o l o g i c a l a c t i o n of i o n i z i n g r a d i a t i o n s . As it t u r n s ou t , n o t h i n g e x c e e d s t h e d r a m a t i c e f f e c t of m o l e c u l a r oxygen . T h e o x y g e n e f f e c t w a s d i s c o v e r e d in t he 1920s by t h e u s e of r e l a t i v e l y s i m p l e b i o l o g i c a l s y s t e m s . As e a r l y a s 1921 it had b e e n n o t ed by H o l t h u s e n t h a t a s c a r i s e g g s w e r e r e l a t i v e l y r e s i s t a n t to r a d i a t i o n in t he a b s e n c e of oxygen . A c o r r e l a t i o n b e t w e e n r a d i o s e n s i t i v i t y and the p r e s e n c e of oxygen w a s m a d e i n d e p e n d e n t l y by P e t r y in 1923 , f r o m a s t u d y of t he e f f e c t s of r a d i a t i o n on v e g e t a b l e s e e d s . T h e m a g n i t u d e of t he oxygen e f f e c t w a s a l s o f i r s t d e t e r m i n e d wi th a s i m p l e p l an t s y s t e m , n a m e l y t he g r o w t h i nh ib i t i on of s e e d l i n g s of V i c i a f a b a .

7 . 2 . C E L L SURVIVAL C U R V E S IN T H E P R E S E N C E AND A B S E N C E O F OXYGEN

T h e e a r l i e r d i s c o v e r i e s wi th s i m p l e p l a n t s y s t e m s h a v e s i n c e b e e n r e p e a t e d wi th m a m m a l i a n c e l l s in c u l t u r e . S u r v i v a l c u r v e s wi th m a m m a l i a n c e l l s e x p o s e d to g a m m a r a y s in t h e p r e s e n c e o r a b s e n c e of oxygen a r e i l l u s t r a t e d in F i g . 18. T h e s h a p e of t h e s u r v i v a l c u r v e i s t he s a m e u n d e r b o t h a e r a t e d and hypox ic c o n d i t i o n s ; a l l t h a t i s d i f f e r e n t i s t h e m a g n i t u d e of t he d o s e r e q u i r e d to p r o d u c e a g i v e n d e g r e e of b i o l o g i c a l d a m a g e .

T h e e x a m p l e i l l u s t r a t e d i s t y p i c a l of m a n y s u c h e x p e r i m e n t s p u b l i s h e d in t he l i t e r a t u r e ; a t a l l l e v e l s of s u r v i v a l t he d o s e r e q u i r e d u n d e r h y p o x i c c o n d i t i o n s i s a p p r o x i m a t e l y t h r e e t i m e s a s g r e a t a s t he d o s e r e q u i r e d u n d e r a e r a t e d c o n d i t i o n s . T h e r a t i o of hypox ic t o a e r a t e d d o s e s r e q u i r e d t o p r o d u c e t h e s a m e b i o l o g i c a l e f f e c t i s t h e s a m e f o r a l l s u r v i v a l l e v e l s , and b e c a u s e of t h i s , o x y g e n i s s a i d t o b e a d o s e - m o d i f y i n g a g e n t . T h i s r a t i o i s c a l l e d t he o x y g e n e n h a n c e m e n t r a t i o (OER) . T h e OER h a s b e e n d e t e r m i n e d f o r a w ide r a n g e of b i o l o g i c a l s y s t e m s and d i f f e r e n t e n d - p o i n t s , and i t s v a l u e f o r X o r g a m m a r a y s a l m o s t a l w a y s f a l l s w i th in t h e r a n g e 2 .5 to 3.

T h e oxygen e f f e c t f o r o t h e r t y p e s of i o n i z i n g r a d i a t i o n s i s i l l u s t r a t e d in F i g . 19. F o r a l p h a p a r t i c l e s , w h i c h a r e d e n s e l y i o n i z i n g , the s u r v i v a l c u r v e i s a n e x p o n e n t i a l f u n c t i o n of d o s e ; i . e . t he s u r v i v a l c u r v e d o e s no t h a v e a n i n i t i a l s h o u l d e r . In t h i s c a s e , s u r v i v a l e s t i m a t e s m a d e in t he p r e s e n c e o r a b s e n c e of o x y g e n f a l l a l o n g a c o m m o n l i n e . T h e O E R i s u n i t y , o r in o t h e r w o r d s t h e r e i s n o oxygen e f f e c t . F o r r a d i a t i o n s of i n t e r m e d i a t e i o n i z i n g d e n s i t y , s u c h a s n e u t r o n s , t h e oxygen e f f e c t i s v e r y m u c h s m a l l e r t h a n f o r X o r g a m m a r a y s . In the e x a m p l e s h o w n , t he O E R f o r 1 5 - M e V n e u t r o n s is a p p r o x i m a t e l y 1.6 .

T o s u m up , t he o x y g e n e f f e c t i s l a r g e and i m p o r t a n t in the c a s e of s p a r s e l y i o n i z i n g r a d i a t i o n s s u c h a s g a m m a r a y s , it i s a b s e n t f o r d e n s e l y i o n i z i n g r a d i a t i o n s s u c h a s a l p h a p a r t i c l e s , and it h a s a n i n t e r m e d i a t e v a l u e f o r f a s t n e u t r o n s .

4 1

42 CHAPTER 10

1 0 1000

T

D O S E ( r o d ) 2000 3 0 0 0 4 0 0 0

H A M S T E R C E L L S E X P O S E D TO 7 - R A Y S

FIG.18. Survival curves for m a m m a l i a n cells exposed to gamma rays under aerated and hypoxic conditions. The oxygen enhancement rat io (OER), defined to be the rat io of doses without and with oxygen needed to produce the same biological e f fec t , is about 3, and is independent of dose.

7 . 3 . TIMING O F T H E OXYGEN E F F E C T

F o r the oxygen e f f e c t to b e o b s e r v e d , oxygen m u s t b e p r e s e n t while the r a d i a t i o n i s b e i n g a b s o r b e d . T h i s s i m p l e s t a t e m e n t i s g e n e r a l l y thought to be t r u e , a l though m a n y i ngen ious e x p e r i m e n t s have b e e n d e s i g n e d t o e l u c i d a t e the m e c h a n i c s of the oxygen e f f e c t m o r e fu l ly .

One of the e a r l y e x p e r i m e n t s , and the s i m p l e s t of i t s k ind , w a s p e r -f o r m e d by W r i g h t and H o w a r d - F l a n d e r s (1957), who i r r a d i a t e d the t a i l s of young r o d e n t s wi th a s h o r t p u l s e of e l e c t r o n s , and s c o r e d the e f f e c t of the r a d i a t i o n by o b s e r v i n g the ex ten t to which the s u b s e q u e n t g r o w t h of the t a i l was inh ib i t ed . T h e e f f e c t of a g iven d o s e i s r e d u c e d if the t a i l i s m a d e hypoxic by c l a m p i n g off the blood supply to the t a i l b e f o r e i r r a d i a t i o n , s i n c e , in c o m m o n wi th a l l m a m m a l i a n c e l l s , the s e n s i t i v e c a r t i l a g e c e l l s c o n c e r n e d wi th the g r o w t h of the bone a r e p r o t e c t e d f r o m r a d i a t i o n by hypox ia . The s e n s i t i v i t y of the t a i l w a s found to d r o p to a l e v e l c h a r a c t e r i s t i c of hypoxia if the c l a m p w a s app l i ed f o u r s e c o n d s b e f o r e the e x p o s u r e to X - r a y s ; th i s t i m e was p r o b a b l y n e e d e d f o r the t i s s u e s to e x h a u s t t he oxygen t r à p p e d in t h e m a f t e r the c l a m p had b e e n a p p l i e d .

The m o s t r e c e n t e x p e r i m e n t s of th is k ind , involv ing the s h o r t e s t r e s o l v i n g t i m e , have b e e n p e r f o r m e d by M i c h a e l et a l . (1973) at the G r a y L a b o r a t o r y in London . Us ing s h o r t b u r s t s of e l e c t r o n s , and g a s - m i x i n g t e c h n i q u e s b a s e d on an e x p l o s i o n to quickly a d m i t oxygen to the b i o l o g i c a l s y s t e m b e f o r e o r a f t e r r a d i a t i o n , they w e r e a b l e to show tha t r a d i a t i o n s e n s i t i v i t y c h a r a c t e r i s t i c of a e r a t e d cond i t ions could be obta ined in b a c t e r i a l o r m a m m a l i a n c e l l s if the oxygen was a d m i t t e d s e v e r a l m i l l i o n t h s of a s e c o n d a f t e r the r a d i a t i o n e x p o s u r e . T h i s f ind ing is c o n s i s t e n t wi th the no t ion tha t the oxygen e f f e c t i s involved wi th the p r o d u c t i o n of f r e e r a d i c a l s ,

EFFECT OF MOLECULAR OXYGEN 43

X - R A Y S

Hypoxic

0.01

0.001 ~ i — ' — i — ' — r - ' — i — < — i — 1 — i — ' — r 2 0 0 4 0 0 6 0 0 8 0 0 1000 1200 1400 1600 1800 2 0 0 0

Dose (rod)

2 0 0 4 0 0 6 0 0 Dose (rod)

2> 0.1-

; 0.01-

0.001-

» - R A Y S

\ N E

Aerated ¿

lypoxic

\

O E R =1.0

0 100 2 0 0 3 0 0 Dose (rad)

FIG. 19. Survival curves for mammal i an cells under aerated and hypoxic conditions and exposed to X-rays,

fast neutrons and a - p a r t i c l e s . (From Barendsen e t a l . , 1966; and Broerse e t a l . , 1967).

which have a l i f e t i m e of about 1 0 " 5 s , b e c a u s e the oxygen e f f e c t i s o b s e r v e d if oxygen i s admit ted to the b i o l o g i c a l s p e c i m e n during the l i f e t i m e of the f r e e r a d i c a l s .

7 . 4 . OXYGEN CONCENTRATION REQUIRED

A key ques t ion c o n c e r n i n g the oxygen e f f e c t i s the concentra t ion requ ired for the e f f e c t to be o b s e r v e d . Is it a t race of oxygen, or i s it a l a r g e amount of oxygen that i s involved? Many e x p e r i m e n t s have b e e n p e r f o r m e d with b a c t e r i a and m a m m a l i a n c e l l s in a t tempts to a n s w e r this quest ion .

44 CHAPTER 10

DOSE (krad)

FIG. 20. Survival curves for m a m m a l i a n cells X-irradiated in the presence of different concentrations of oxygen. Open circles, a i r ; closed circles 2200 ppm of oxygen or a part ial pressure of 1.7 mm of mercury; open squares, 355 ppm of oxygen; closed squares, 100 ppm of oxygen; open triangles, 10 ppm of oxygen. (From Elkind et a l . , 1963).

T h e r e a r e d i f f e r e n c e s b e t w e e n t h e r e s u l t s o b t a i n e d wi th v a r i o u s t e s t s y s t e m s , bu t t he s i m i l a r i t i e s a r e m u c h m o r e s t r i k i n g and m o r e i m p o r t a n t t h a n the d i f f e r e n c e s .

F i g u r e 20 i s a d r a m a t i c e x a m p l e of wha t h a p p e n s to t he s u r v i v a l c u r v e f o r m a m m a l i a n c e l l s w h e n o x y g e n i s i n t r o d u c e d in to the s y s t e m . C u r v e A i s c h a r a c t e r i s t i c of the r e s p o n s e u n d e r c o n d i t i o n s of e q u i l i b r a t i o n wi th a i r . C u r v e B i s a s u r v i v a l c u r v e f o r i r r a d i a t i o n in a s low a l e v e l of hypox ia a s c a n b e o b t a i n e d u n d e r e x p e r i m e n t a l c o n d i t i o n s , wh ich c o r r e s p o n d s to abou t 10 p p m of oxygen in t he g a s p h a s e . T h e i n t r o d u c t i o n of 100 p p m i s r e a d i l y n o t i c e a b l e b e c a u s e t h e s l o p e of t he s u r v i v a l c u r v e b e c o m e s s t e e p e r . At a c o n c e n t r a t i o n of 2200 p p m , w h i c h i s a b o u t of oxygen , t he s u r v i v a l c u r v e is a p p r o x i m a t e l y ha l f way b e t w e e n t h o s e c h a r a c t e r i s t i c of a e r a t e d and h y p o x i c c o n d i t i o n s .

It h a s b e e n s h o w n in o t h e r e x p e r i m e n t s tha t by t he t i m e a c o n c e n t r a t i o n of oxygen c o r r e s p o n d i n g to 2% h a s b e e n r e a c h e d , the s u r v i v a l curv.e i s v i r t u a l l y i n d i s t i n g u i s h a b l e f r o m t h a t o b t a i n e d u n d e r c o n d i t i o n s of n o r m a l a e r a t i o n . F u r t h e r m o r e , c h a n g i n g f r o m a i r t o 100% oxygen a t h igh p r e s s u r e d o e s no t f u r t h e r a f f e c t t he s l o p e of the s u r v i v a l c u r v e .

It i s e v i d e n t , t h e n , tha t v e r y s m a l l a m o u n t s of o x y g e n a r e s u f f i c i e n t t o p r o d u c e t h e d r a m a t i c and i m p o r t a n t oxygen e f f e c t o b s e r v e d wi th X o r g a m m a r a y s . T h e oxygen t e n s i o n of m o s t n o r m a l t i s s u e s i s s i m i l a r t o tha t of v e n o u s b lood o r l y m p h , w h i c h is in the r a n g e of 20 to 40 m m of m e r c u r y . F r o m a r a d i o b i o l o g i c a l point of v i e w , t h e r e f o r e , n o r m a l t i s s u e s a r e u s u a l l y c o n s i d e r e d to b e w e l l o x y g e n a t e d , a l t h o u g h t h e r e a r e a few e x c e p t i o n s . T h e f a c t t h a t s u c h s m a l l q u a n t i t i e s of o x y g e n m o d i f y b i o l o g i c a l r e s p o n s e i n t r o d u c e s t e c h n i c a l c o m p l i c a t i o n s when e x p e r i m e n t s on t h i s s u b j e c t a r e c a r r i e d out .

EFFECT OF MOLECULAR OXYGEN 45

7. 5. MECHANISM OF THE OXYGEN E F F E C T

T h e r e i s no unanimity a m o n g s t e x p e r t s in the f i e l d c o n c e r n i n g the m e c h a n i s m of the oxygen e f f e c t , although it i s g e n e r a l l y a g r e e d that oxygen a c t s at the l e v e l of the f r e e r a d i c a l s .

The chain of event s f r o m the absorpt ion of radiat ion to the f ina l e x p r e s s i o n of b i o l o g i c a l damage m a y be s u m m a r i z e d a s f o l l o w s . The absorpt ion of radiat ion l e a d s to the product ion of fas t charged p a r t i c l e s ; the charged p a r t i c l e s in p a s s i n g through the b i o l o g i c a l m a t e r i a l produce a n u m b e r of ion p a i r s . T h e s e ion p a i r s have a v e r y shor t l i f e t i m e (of the o r d e r of 10" 1 0 s ) ; f r o m t h e s e ion p a i r s a r e produced f r e e r a d i c a l s that a r e highly r e a c t i v e m o l e c u l e s . They a r e important b e c a u s e the ir l i f e t i m e of 10"5 s i s apprec iab ly l o n g e r than that of the ion p a i r s .

It i s the f r e e r a d i c a l s that break c h e m i c a l bonds and in i t ia te the cha in of e v e n t s which r e s u l t s in the f ina l e x p r e s s i o n of b i o l o g i c a l damage . If oxygen i s p r e s e n t , it r e a c t s with the f r e e r a d i c a l s to produce an organic p e r o x i d e which i s a n o n - r e s t o r a b l e f o r m of the targe t m a t e r i a l ; that i s , it has a l r e a d y r e s u l t e d in a changed c h e m i c a l c o m p o s i t i o n of the m a t e r i a l e x p o s e d to the radiat ion. With no oxygen p r e s e n t , th is r e a c t i o n cannot take p l a c e , and m a n y of the ion ized target m o l e c u l e s m a y r e p a i r or r e c o v e r the abi l i ty to funct ion n o r m a l l y .

C h a p t e r 8

C E L L AGE AND RADIOSENSITIVITY

8.1 . T H E C E L L C Y C L E

M a m m a l i a n c e l l s r e p l i c a t e and i n c r e a s e in n u m b e r by c e l l d i v i s i o n o r m i t o s i s . When a c e l l d i v i d e s , two d a u g h t e r c e l l s a r e p r o d u c e d , e a c h of wh ich c a r r i e s a c h r o m o s o m e c o m p l e m e n t i d e n t i c a l to t h a t of the m o t h e r c e l l . If g r o w i n g c e l l s a r e c o n t i n u o u s l y o b s e r v e d wi th a c o n v e n t i o n a l l igh t m i c r o -s c o p e , t he on ly even t t ha t can b e c l e a r l y d i s t i n g u i s h e d is t he p r o c e s s of m i t o s i s i t s e l f . F o r m o s t of the c e l l c y c l e , the c h r o m o s o m e m a t e r i a l i s d i f f u s e and no t c l e a r l y s e e n , but f o r a s h o r t p e r i o d of t i m e b e f o r e m i t o s i s it c o n d e n s e s in to d i s c r e t e and r e c o g n i z a b l e e n t i t i e s . The c h r o m o s o m e s s e p a r a t e in to two g r o u p s and m o v e to the two p o l e s of the ce l l ; d i v i s i o n t h e n o c c u r s to f o r m two d a u g h t e r c e l l s . T h i s p r o c e s s of m i t o s i s o c c u p i e s a p e r i o d of abou t one h o u r . T h e r e m a i n d e r of the ce l l c y c l e , the s o - c a l l e d i n t e r p h a s e , o c c u p i e s a l l of t he i n t e r m i t o t i c p e r i o d , and wi th a c o n v e n t i o n a l m i c r o s c o p e no e v e n t s of i n t e r e s t c a n b e i d e n t i f i e d d u r i n g t h i s t i m e .

S ince c e l l d i v i s i o n i s a c y c l i c a l p h e n o m e n o n , r e p e a t e d in e a c h g e n e r a t i o n of the c e l l s , it i s u s u a l to r e p r e s e n t it a s a c i r c l e as shown in F i g . 21. The t o t a l c i r c u m f e r e n c e of the c i r c l e r e p r e s e n t s the m i t o t i c c y c l e t i m e f o r the c e l l s (T c) ; t he p e r i o d of m i t o s i s i s r e p r e s e n t e d by M.

8 .2 . A U T O R A D I O G R A P H Y

H o w a r d and P e l e (1951, 1953) w e r e t he f i r s t to f u r t h e r s u b d i v i d e the m i t o t i c c y c l e by t he u s e of r a d i o a c t i v e l a b e l l e d DNA p r e c u r s o r s . The t e c h -n i q u e of a u t o r a d i o g r a p h y , e m p l o y e d in t h e s e e x p e r i m e n t s , i s c a r r i e d out a s f o l l o w s .

The g r o w i n g c e l l s a r e fed a p r e c u r s o r u s e d in the bu i ld ing of DNA m o l e c u l e s , c o m m o n l y t h y m i d i n e . T h e t h y m i d i n e is l a b e l l e d wi th a b e t a e m i t t i n g i s o t o p e ; in the e a r l y w o r k of H o w a r d a n d P e l e , the r a d i o a c t i v e l a b e l u s e d w a s p h o s p h o r u s ( 3 2 P) but n o w a d a y s t r i t i u m (SH) i s a l m o s t e x c l u s i v e l y u s e d , b e c a u s e of t he s h o r t e r r a n g e of b e t a r a y s . T h i s r a d i o a c t i v e l a b e l l e d DNA p r e c u r s o r i s m a d e a v a i l a b l e t o t h e g r o w i n g popu l a t i on of c e l l s f o r a s h o r t p e r i o d of t i m e , a n d is i n c o r p o r a t e d in to the DNA of t h o s e c e l l s w h i c h a t the t i m e a r e a c t i v e l y s y n t h e s i z i n g DNA; c e l l s which a r e not m a k i n g DNA w i l l not t a k e u p the r a d i o a c t i v e l a b e l .

The c e l l p r e p a r a t i o n is t h e n f ixed and s t a i n e d , s o tha t the c e l l s a r e r e a d i l y v i s i b l e , and c o v e r e d wi th a th in l a y e r of n u c l e a r e m u l s i o n . B e t a r a y s f r o m the i n c o r p o r a t e d t r i t i a t e d t h y m i d i n e l e a v e the c e l l s a n d , in p a s s i n g t h r o u g h the c o v e r i n g l a y e r of n u c l e a r e m u l s i o n , p r o d u c e t r a c k s w h i c h c a n b e s e e n when s u b s e q u e n t l y the e m u l s i o n is d e v e l o p e d . When the p r e p a r a t i o n i s v i e w e d t h r o u g h a m i c r o s c o p e , c e l l s a r e s e e n to h a v e a l a r g e n u m b e r of b l a c k d o t s o v e r the n u c l e u s r e s u l t i n g f r o m the b e t a r a y s t h a t w e r e e m i t t e d . T h e s e a r e c e l l s t h a t h a v e i n c o r p o r a t e d t he i s o t o p e , b e c a u s e they w e r e s y n t h e s i z i n g DNA d u r i n g the l a b e l l i n g p e r i o d . O t h e r c e l l s do not h a v e any

4 7

4 8 CHAPTER 10

M ( Mitosis)

S ( D N A synthetic phase) FIG.21. The principal events in the mitot ic cycle for act ively growing cells a r e : mitosis (M) and the period of DNA synthesis (S). Gj and G2 are 'gaps* of apparent inact ivi ty .

g r a i n s located over the ir n u c l e i , and this i s in terpre ted to m e a n that the c e l l s w e r e not a c t i v e l y s y n t h e s i z i n g DNA at the t i m e the r a d i o a c t i v e l a b e l was m a d e a v a i l a b l e to them.

8.3. THE PHASES OF THE C E L L CYCLE

U s i n g this t echnique , it can be shown that c e l l s incorporate the rad io -ac t ive labe l only during a c e r t a i n w e l l de f ined f rac t ion of the c e l l c y c l e . T h e r e i s an i n t e r v a l b e t w e e n m i t o s i s and DNA s y n t h e s i s in which no l a b e l i s incorporated; th i s i n t e r v a l , or 'gap', i s known a s the Gj gap. A f t e r DNA s y n t h e s i s h a s b e e n c o m p l e t e d , there i s a s e c o n d in terva l , or gap, b e f o r e m i t o s i s which has b e e n t e r m e d G2.

A l l p r o l i f e r a t i n g m a m m a l i a n c e l l s , whe ther in cu l ture or growing n o r m a l l y in a t i s s u e , have a m i t o t i c c y c l e which can be divided into t h e s e const i tuent par t s ; m i t o s i s , M, f o l l o w e d by Gj, a per iod of DNA s y n t h e s i s , S, and G2, a f t e r which m i t o s i s o c c u r s again. The l engths of t h e s e v a r i o u s parts of the m i t o t i c c y c l e v a r y a c c o r d i n g to the par t i cu lar c e l l s that a r e s tudied. The tota l c e l l c y c l e m a y v a r y f r o m about 10 hours , which i s the s h o r t e s t known c e l l c y c l e in m a m m a l i a n c e l l s , to many hundreds of h o u r s , which i s c o m m o n l y s e e n in s o m e plant m a t e r i a l s and s o m e n o r m a l t i s s u e s in m a m m a l i a n s y s t e m s . Most of this s p r e a d in c y c l e t i m e s b e t w e e n d i f f erent c e l l s i s a r e s u l t of a d r a m a t i c v a r i a t i o n in the length of the Gj per iod . The r e m a i n i n g c o m p o n e n t s of the c e l l c y c l e (namely M, S and Gz) vary c o m p a r a -t ive ly l i t t le b e t w e e n d i f f erent c e l l s in d i f f erent c i r c u m s t a n c e s .

CELL AGE AND RADI0SENSITIV1TY 49

1.0

0.5

0.2

< 0 .1 >

œ 0.05 U

UJ 0.02

I LÜ 0 . 0 1

I 0.005

0.002

0 . 0 0 1

0.0005, 0 200 400 600 800 1000 1200 1400

DOSE (rad)

FIG.22. Cell survival curves for Chinese hamster cells at various stages in the cell cycle . For cells in mitosis, the survival curve is steep and has no shoulder. For cells in late S, the curve is less steep and has a big shoulder. G t and early S are intermediate in sensitivity. (From Sinclair, 1968).

8.4. SYNCHRONOUS C E L L C U L T U R E S

T h e r a d i o s e n s i t i v i t y of c e l l s v a r i e s wi th t h e i r pos i t i on o r age in the m i t o t i c c y c l e . A s tudy of t h i s p h e n o m e n o n w a s m a d e p o s s i b l e by the d e v e l o p -m e n t of t e c h n i q u e s to p r o d u c e s y n c h r o n o u s l y d iv id ing c e l l p o p u l a t i o n s , in which at a g iven t i m e a l l of t he c e l l s occupy the s a m e p h a s e of t he c e l l c y c l e .

Two t e c h n i q u e s h a v e b e e n u s e d to p r o d u c e s y n c h r o n o u s l y d iv id ing c e l l p o p u l a t i o n s . T h e f i r s t of t h e s e i s known a s the ' m i t o t i c h a r v e s t ' t e c h n i q u e , d e s c r i b e d by T e r a s i m a and T o l m a c h (1963) and s u i t a b l e only f o r c e l l s which g row in a m o n o l a y e r a t t a c h e d to the s u r f a c e of a g r o w t h v e s s e l . T h e t e c h -nique exp lo i t s the f ac t tha t when c e l l s a r e c l o s e t o m i t o s i s , they round up and b e c o m e l o o s e l y a t t a c h e d to the s u r f a c e . If at th i s s t a g e the g r o w t h m e d i u m o v e r the c e l l s i s s h a k e n , m i t o t i c c e l l s b e c o m e d e t a c h e d f r o m the s u r f a c e and f loa t in the m e d i u m . If t h e s e c e l l s a r e r e s e e d e d into new P e t r i d i s h e s , the popu la t i on c o n s i s t s a l m o s t e n t i r e l y of m i t o t i c c e l l s . When t h e s e c e l l c u l t u r e s a r e i ncuba t ed at 37°C, the c e l l s m o v e t o g e t h e r s y n c h r o n o u s l y t h r o u g h the c e l l c y c l e . By d e l i v e r i n g a d o s e of r a d i a t i o n a t v a r i o u s t i m e s a f t e r the i n i t i a l h a r v e s t i n g of the m i t o t i c c e l l s , the r a d i o s e n s i t i v i t y a s a func t i on of c e l l age m a y b e s t u d i e d .

A n o t h e r way to ob ta in s y n c h r o n o u s l y d iv id ing c e l l s i s to u s e a d r u g which i n t e r f e r e s wi th the p r o g r e s s of c e l l s wi th in n o r m a l m i t o t i c c y c l e s . A n u m b e r of d i f f e r e n t s u b s t a n c e s can b e u s e d , t he m o s t p o p u l a r of which i s h y d r o x y u r e a .

5 0 CHAPTER 10

0.2 - A 1 1 1 1

CHINESE HAMSTER CELLS i i

0. r \ ~z

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0.01 1 1-

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0.002 I f s f Gl ^ s ^ G» ^Hm 0.001 !_.. L 1 1 ! 1

TIME (hours)

FIG. 23. Forms of age response for (A) cells with a short G, such as Chinese hamster cells, and (B) cells with a long G t such as HeLa cel ls . (From Sinclair, 1969).

T h e a d v a n t a g e of t h i s m e t h o d i s tha t it m a y be a p p l i e d t o s o m e t i s s u e s y s t e m s a s w e l l a s to c e l l s g r o w n in c u l t u r e . When h y d r o x y u r e a is added to a popu-l a t i o n of d i v i d i n g c e l l s it h a s two e f f e c t s ; in the f i r s t p l a c e c e l l s in the S p h a s e t a k e up the d r u g and a r e k i l l ed . S e c o n d , the d r u g i m p o s e s a b lock a t t h e end of t h e Gj p e r i o d , and c e l l s t ha t o c c u p y the G 2 , M and G j c o m p a r t -m e n t s p r o g r e s s t h r o u g h the c e l l c y c l e and a c c u m u l a t e a t t h i s b lock . By the end of a t r e a t m e n t wi th the d r u g l a s t i n g m a n y h o u r s , a l l of the v i a b l e c e l l s in t he p o p u l a t i o n w i l l b e s i t u a t e d in a n a r r o w window a t the end of Gj , p o i s e d r e a d y to e n t e r S. If t he d r u g i s t h e n r e m o v e d f r o m t h e p o p u l a t i o n , t h i s s y n c h r o n i z e d c o h o r t of c e l l s w i l l p r o c e e d on t h r o u g h t h e ce l l c y c l e in a s y n c h r o n i z e d f a s h i o n .

8 .5 . X - IRRADIAT ION O F SYNCHRONOUSLY DIVIDING C E L L S

U s i n g C h i n e s e h a m s t e r c e l l s g r o w n in c u l t u r e , S i n c l a i r (1968) d e t e r m i n e d s u r v i v a l c u r v e s at a n u m b e r , of d i s c r e t e p o i n t s d u r i n g the c e l l c y c l e . The r e s u l t s a r e shown in F i g . 22. S u r v i v a l c u r v e s a r e shown f o r m i t o t i c c e l l s (M),

CELL AGE AND RAD10SENSIT1VITY 5.1

f o r c e l l s in G 1 ; a n d f o r c e l l s in t he e a r l y and l a t e p a r t s of DNA s y n t h e t i c p h a s e (S). It i s a t o n c e ev iden t t ha t t h e m o s t s e n s i t i v e c e l l s a r e t h o s e in m i t o s i s ; t hey exh ib i t a s u r v i v a l c u r v e w h i c h i s qu i t e s t e e p a n d h a s n o i n i t i a l s h o u l d e r . At t he o t h e r e x t r e m e of s e n s i t i v i t y , t he s u r v i v a l c u r v e f o r c e l l s in t h e l a t t e r p a r t of t he S - p h a s e exh ib i t a s u r v i v a l c u r v e w h i c h i s l e s s s t e e p t h a n t h a t f o r c e l l s in M , and is c h a r a c t e r i z e d by a v e r y b r o a d i n i t i a l s h o u l d e r . T h e o t h e r p h a s e s of t he c y c l e , n a m e l y Gj a n d t h e e a r l y p a r t of S, a r e i n t e r -m e d i a t e in s e n s i t i v i t y b e t w e e n t h e s e two e x t r e m e s .

8 .6 . A G E R E S P O N S E F U N C T I O N F O R H E L A AND H A M S T E R C E L L S

T h e two c e l l l i n e s m o s t c o m m o n l y u s e d in r a d i o b i o l o g i c a l r e s e a r c h a r e H ë L a c e l l s , d e r i v e d o r i g i n a l l y f r o m a b i o p s y of a h u m a n c a r c i n o m a of t h e c e r v i x , and h a m s t e r c e l l s d e r i v e d f r o m one o r o t h e r of the r e g e n e r a t i n g t i s s u e s of t h e C h i n e s e h a m s t e r . T h e m o s t i m p o r t a n t d i f f e r e n c e b e t w e e n H e L a and h a m s t e r c e l l s i s the l e n g t h of the m i t o t i c c y c l e , w h i c h f o r t h e two l i n e s i s a p p r o x i m a t e l y 24 and 10 h o u r s , r e s p e c t i v e l y . M o s t of t h i s d i f f e r e n c e c a n b e a t t r i b u t e d to a m a r k e d v a r i a t i o n in t h e l e n g t h of t he G j p h a s e . In t h e c a s e of C h i n e s e h a m s t e r c e l l s , G j o c c u p i e s l i t t l e m o r e t h a n a n h o u r , and i n d e e d in s o m e h a m s t e r l i n e s it c a n b a r e l y b e d i s t i n g u i s h e d a t a l l . By c o n t r a s t H e L a c e l l s , in c o m m o n wi th m o s t c e l l s t ha t h a v e a l o n g e r c y c l e , h a v e a G j p h a s e w h i c h o c c u p i e s a s i g n i f i c a n t p o r t i o n of the t o t a l c e l l c y c l e .

F i g u r e 23 i s a c o m p a r i s o n of t he s h a p e of the a g e r e s p o n s e f o r H e L a c e l l s w h i c h r e p r e s e n t c e l l s w i t h i n an a p p r e c i a b l e Gj , and h a m s t e r c e l l s , wh ich a r e r e p r e s e n t a t i v e of c e l l s h a v i n g v e r y s h o r t G1# When the t i m e s c a l e i s a d j u s t e d s o t h a t t he S p e r i o d h a s a c o m p a r a b l e l eng th f o r b o t h c e l l l i n e s , t he g e n e r a l p a t t e r n of c y c l i c v a r i a t i o n i s v e r y s i m i l a r , the only i m p o r t a n t d i f f e r e n c e b e i n g the e x t r a s t r u c t u r e which is e v i d e n t d u r i n g the G j p h a s e in t he c a s e of H e L a c e l l s . M a n y i n v e s t i g a t o r s h a v e p e r f o r m e d c o m p a r a b l e e x p e r i m e n t s wi th a v a r i e t y of s u b - l i n e s of h a m s t e r and H e L a c e l l s , and h a v e o b t a i n e d e s s e n t i a l l y s i m i l a r r e s u l t s . T h e m a i n c h a r a c -t e r i s t i c s of t he v a r i a t i o n of r a d i o s e n s i t i v i t y wi th c e l l a g e in t he m i t o t i c c y c l e m a y b e s u m m a r i z e d a s f o l l o w s : (a) C e l l s a t o r c l o s e to m i t o s i s a r e t h e m o s t s e n s i t i v e . (b) C e l l s in t he l a t t e r p a r t of t he S - p h a s e a r e u s u a l l y t he m o s t r e s i s t a n t . (c) F o r t h o s e c e l l l i n e s tha t h a v e a n a p p r e c i a b l e Ga p e r i o d , a s e c o n d

r e s i s t a n t p h a s e i s e v i d e n t e a r l y in G j , f o l l o w e d by a s e n s i t i v e p e r i o d t o w a r d s t h e end of G^.

(d) G 2 i s u s u a l l y s e n s i t i v e , p e r h a p s a s s e n s i t i v e a s M.

C h a p t e r 9

R E P A I R OF S U B L E T H A L DAMAGE AND THE D O S E - R A T E E F F E C T

9. 1. S H A P E O F T H E SURVIVAL C U R V E AND S U B L E T H A L D A M A G E

The s u r v i v a l c u r v e f o r m a m m a l i a n c e l l s e x p o s e d to X - r a y s h a s a c h a r a c t e r i s t i c s h a p e . When s u r v i v i n g f r a c t i o n i s p l o t t e d a g a i n s t d o s e on a s e m i l o g a r i t h m i c p lo t t h e r e i s a b r o a d i n i t i a l s h o u l d e r , f o l l o w e d by a s t e e p e r and s t r a i g h t e r p o r t i o n . T h i s t h r e s h o l d t y p e of r e s p o n s e i m p l i e s t h a t d a m a g e m u s t b e a c c u m u l a t e d f o r a l e t h a l e f f e c t to b e c o m e e v i d e n t . C e l l s w h i c h h a v e a c c u m u l a t e d l e s s t h a n the c r i t i c a l a m o u n t of d a m a g e n e c e s s a r y to r e s u l t in l e t h a l i t y a r e s a i d to be ' s u b l e t h a l l y ' d a m a g e d ; g i v e n t i m e , s u c h c e l l s m a y be ab l e to r e p a i r , and c o m p l e t e l y r e c o v e r f r o m , t h i s t y p e of d a m a g e .

9 . 2 . S P L I T - D O S E E X P E R I M E N T S

E l k i n d and Su t ton (1959, 1960) m a d e an e x t e n s i v e s t u d y of t h e r e p a i r of s u b l e t h a l d a m a g e by c o m p a r i n g t h e e f f e c t of a s i n g l e d o s e of r a d i a t i o n wi th t h e s a m e t o t a l d o s e d iv ided in to two e q u a l f r a c t i o n s , s p a c e d by v a r i o u s t i m e i n t e r v a l s . T h e r e s u l t s of a s p l i t d o s e e x p e r i m e n t a r e s h o w n in F i g . 24. A s t h e t i m e i n t e r v a l b e t w e e n the two h a l v e s of t h e d o s e i s i n c r e a s e d , t he s u r v i v i n g f r a c t i o n i n c r e a s e s u n t i l a p l a t e a u i s r e a c h e d a f t e r a t i m e i n t e r v a l of abou t 2 h o u r s . In t he c a s e of C h i n e s e h a m s t e r c e l l s , t h e r e i s a s u b s e q u e n t d ip in t h e r e c o v e r y c u r v e a t about 6 h o u r s ; t h i s i s a r e s u l t of t h e age r e s p o n s e func t ion .

When an a s y n c h r o n o u s p o p u l a t i o n of c e l l s i s e x p o s e d to a r e l a t i v e l y l a r g e d o s e of r a d i a t i o n , c e l l s in t he m o r e s e n s i t i v e p h a s e s of t h e c y c l e (M and G 2 ) a r e r e d u c e d to a m u c h l o w e r s u r v i v i n g f r a c t i o n t h a n S - p h a s e c e l l s , w h i c h a r e r e s i s t a n t . T h e p o p u l a t i o n of c e l l s w h i c h s u r v i v e s t h e f i r s t of a p a i r of s p l i t d o s e s , t h e r e f o r e , t e n d s to be c o m p o s e d p r i n c i p a l l y of S - p h a s e c e l l s . When the t i m e i n t e r v a l b e t w e e n s p l i t d o s e s i s 4 t o 6 h o u r s , t h o s e c e l l s w h i c h w e r e in t h e S - p h a s e at the t i m e of t h e f i r s t d o s e f r a c t i o n h a v e m o v e d a r o u n d and a r e now in a s e n s i t i v e p h a s e of t h e c e l l c y c l e (M o r G 2 ) by t h e t i m e t h e s e c o n d d o s e f r a c t i o n i s d e l i v e r e d .

The p a t t e r n of r e p a i r c h a r a c t e r i s t i c a l l y s e e n b e t w e e n s p l i t d o s e s i s , t h e r e f o r e , a c o m b i n a t i o n of two p r o c e s s e s w h i c h t a k e p l a c e s i m u l t a n e o u s l y . F i r s t , t h e r e i s t h e p r o m p t r e p a i r of s u b l e t h a l r a d i a t i o n d a m a g e , and s e c o n d , t h e r e i s p r o g r e s s i o n of c e l l s t h r o u g h the ce l l c y c l e d u r i n g t h e i n t e r v a l b e t w e e n the s p l i t d o s e s .

9. 3. R E P A I R AND SURVIVAL C U R V E S H A P E

F o r c e l l s in c u l t u r e t h e r e i s u s u a l l y a good c o r r e l a t i o n b e t w e e n the e x t e n t of r e p a i r of s u b l e t h a l d a m a g e , and t h e m a g n i t u d e of t he s h o u l d e r of the s u r v i v a l c u r v e . T h i s i s to be e x p e c t e d s i n c e bo th a r e m a n i f e s t a t i o n s

53

5 4 CHAPTER 10

0 1 b 1 I 1 I 1 I 1 I 1 I

/ Z O

707 + 8 0 4 rod /

O < Ca

^ 0 . 0 1

z > > cc 3 V)

Incubation at 3 7 ° C between doses

0 . 0 0 1 I i I i I i I i I i I i 0 2 4 6 8 10 12 HOURS B E T W E E N T W O DOSES

FIG.24. Survival of Chinese hamster cells following two dose fractions separated by various t ime intervals. The rise in surviving fraction in the first few hours is due to the prompt repair of sub-lethal d a m a g e . The subsequent dip in the curve is a result of survivoK from the first dose fraction moving around to a more sensitive phase of the cel l cyc le . (Redrawn from Elkind and Sutton, 1960).

of the s a m e b a s i c p h e n o m e n o n . C e l l l i n e s w h i c h a r e c h a r a c t e r i z e d by a s u r v i v a l c u r v e wi th a b r o a d s h o u l d e r , e x h i b i t a l a r g e a m o u n t of r e p a i r of s u b l e t h a l d a m a g e b e t w e e n s p l i t d o s e s ; c o n v e r s e l y , c e l l l i n e s wi th a s m a l l s h o u l d e r to t h e i r s u r v i v a l c u r v e s s h o w a l i m i t e d ab i l i t y t o r e p a i r s u b l e t h a l d a m a g e .

By m e a n s of s i m i l a r s p l i t - d o s e e x p e r i m e n t s , r e p a i r of s u b l e t h a l r a d i a t i o n d a m a g e h a s b e e n d e m o n s t r a t e d in j u s t abou t e v e r y b i o l o g i c a l t e s t s y s t e m f o r w h i c h a q u a n t i t a t i v e e n d - p o i n t i s a v a i l a b l e . T h i s i n c l u d e s c e l l s c u l t u r e d in v ivo , t h e r e s p o n s e of i n t a c t n o r m a l s k i n in t h e m o u s e , l e t h a l e f f e c t s of w h o l e body r a d i a t i o n in a n i m a l s , t h e T C D 5 0 of e x p e r i m e n t a l t u m o u r s in a n i m a l s , 1 t h e s u r v i v a l of c r y p t c e l l s in t h e m o u s e j e j u n u m , a s w e l l a s a v a r i e t y of p l a n t s y s t e m s .

9 . 4 . D O S E - H A T E E F F E C T

In t he c o u r s e of e x p e r i m e n t s in r a d i o b i o l o g y an e x t r e m e l y wide r a n g e of r a d i a t i o n d o s e - r a t e s h a s b e e n e m p l o y e d , e x t e n d i n g f r o m a few r a d s p e r d a y up to t h o u s a n d s of r a d s in a f r a c t i o n of a s e c o n d . O v e r t h e r a n g e of d o s e - r a t e s f r o m 10 r a d / h up to 1000 r a d / m i n , s t u d i e s h a v e b e e n u n d e r t a k e n w i t h a n u m b e r of b i o l o g i c a l s y s t e m s , but t h e m o s t c o m p l e t e d a t a a r e a v a i l a b l e f o r m a m m a l i a n c e l l s in c u l t u r e . T h i s d r a m a t i c d o s e - r a t e e f f e c t i s a d i r e c t c o n s e q u e n c e of t h e r e p a i r of s u b l e t h a l d a m a g e .

C o n t i n u o u s low d o s e - r a t e i r r a d i a t i o n m a y be c o n s i d e r e d to be e q u i v a l e n t to the a p p l i c a t i o n of an i n f i n i t e n u m b e r of s m a l l f r a c t i o n s . F i g u r e 25 i l l u s t r a t e s an i d e a l i z e d e x p e r i m e n t in w h i c h e a c h d o s e i s d e l i v e r e d , no t in

1 The tumour can be transmitted on the average by the implantat ion of a quantity of cells known as the TCD 5 0 . This is analogous to the TD50 explained in Section 5 . 3 .

REPAIR OF SUBLETHAL DAMAGE AND THE DOSE-RATE EFFECT 55

0 . 1

z O ( -o < o :

O J O I o z > > cc o

CO

1 0 . 0 0 1

0.0001 FIG.25. Ideal ized exper iment . Curve A is the survival curve for a single acu te exposure to X-rays. Curve F is obtained if each dose is given as a series of small fractions of size Ü! , with full repair between dose fractions. Multiple small fractions approximate to a continuous low dose-rate exposure. (From Elkind and Whitmore, 1 9 6 1 ) .

D O S E •

a s i n g l e e x p o s u r e , but in a n u m b e r of equa l f r a c t i o n s of s i z e D, wi th a t i m e i n t e r v a l b e t w e e n the f r a c t i o n s s u f f i c i e n t for r e p a i r of s u b l e t h a l d a m a g e to be c o m p l e t e . T h e s h o u l d e r of the c u r v e i s r e p e a t e d wi th e a c h f r a c t i o n ; if on ly s i n g l e s u r v i v a l p o i n t s w e r e d e t e r m i n e d , c o r r e s p o n d i n g to e q u a l d o s e i n c r e m e n t s , the o v e r a l l s u r v i v a l c u r v e that would be o b s e r v e d i s s h o w n by the b r o k e n l i n e , l a b e l l e d F .

R e a s o n i n g a long t h e s e l i n e s , L a j t h a and O l i v e r (1961) s e t out to p r e d i c t the s u r v i v a l c u r v e s that would be e x p e c t e d f o r v a r i o u s l o w d o s e - r a t e t r e a t -m e n t s . T h e i r m o d e l w a s b a s e d on the c h a r a c t e r i s t i c s of the s u r v i v a l c u r v e f o r a c u t e e x p o s u r e s , and a s s u m e s that r e p a i r of s u b l e t h a l d a m a g e t a k e s p l a c e d u r i n g the l o w d o s e - r a t e e x p o s u r e s . T h e i r p r e d i c t i o n s a r e s h o w n in F i g . 26; a s the d o s e - r a t e i s l o w e r e d the s u r v i v a l c u r v e b e c o m e s p r o -g r e s s i v e l y l e s s s t e e p , w h i l e at the s a m e t i m e the e x t r a p o l a t i o n n u m b e r t e n d s t o w a r d s uni ty .

56 CHAPTER 10

100 2 0 0 5 0 0 4 0 0 5 0 0 6 0 0 7 0 0 8 0 0

FIG.26. Predicted survival curves for cells exposed to X o r y - r a y s at various low dose-rates. (From Lajtha and Oliver, 1961).

DOSE ( r a d ) 0 2 0 0 4 0 0 6 0 0 8 0 0 1000

FIG.27. Survival curves of HeLa cells exposed to gamma rays over a range of dose-rates. (From Hall and Bedford, 1964b).

REPAIR OF SUBLETHAL DAMAGE AND THE DOSE-RATE EFFECT 5 7

1 0 '

10° 2 1 . 5 r a d / h

o o I 2 0 r a d / m ¡ n « 5 3 . 3 r a d / h

\ 1 0 " »

5 0 0 1 0 0 0 1500 2 0 0 0 2 5 0 0

DOSE ( r a d )

FIG.28. Survival curves of Chinese hamster cells exposed to 6 0 Co gamma rays at different dose-rates.

(Unpublished data due to J .S . Bedford).

9. 5. D O S E - R A T E E F F E C T F O R C E L L S IN C U L T U R E

F i g u r e 27 i s an e x a m p l e of t he d o s e - r a t e e f f e c t f o r H e L a c e l l s . F i g u r e 28 s h o w s d o s e - r a t e d a t a f o r C h i n e s e h a m s t e r c e l l s w h i c h , b e c a u s e t h e y h a v e a m u c h l a r g e r s h o u l d e r in t h e i r a c u t e X - r a y s u r v i v a l c u r v e , a r e m o r e s e n s i t i v e t o c h a n g e s in d o s e - r a t e . F o r m o s t m a m m a l i a n c e l l s y s t e m s , the p r i n c i p a l d o s e - r a t e e f f e c t m a y b e o b s e r v e d b e t w e e n 100 and 1 r a d / m i n . The d o s e - r a t e e f f e c t in t h i s r a n g e i s due to t h e r e p a i r of s u b l e t h a l d a m a g e , w h i c h t a k e s p l a c e d u r i n g the r a d i a t i o n e x p o s u r e .

A s t he d o s e - r a t e i s r e d u c e d m o r e and m o r e , s u b l e t h a l d a m a g e m a y b e r e p a i r e d b e c a u s e t h e a c c u m u l a t i o n of r a d i a t i o n i n j u r y i s s p r e a d out o v e r a l o n g e r p e r i o d of t i m e . C o n s e q u e n t l y , t he c e l l k i l l i n g p o t e n t i a l of a g iven d o s e of r a d i a t i o n d e c r e a s e s w i t h t i m e . P r o v i d e d tha t c e l l s a r e no t a l l o w e d to d iv ide t h e r e i s l i t t l e f u r t h e r d o s e - r a t e e f f e c t be low 1 r a d / m i n , e s s e n t i a l l y a l l s u b l e t h a l d a m a g e i s r e p a i r e d d u r i n g the e x p o s u r e , and t h e r e s i d u a l c e l l k i l l i n g e f f e c t i s due to n o n - r e p a r a b l e i n j u r y . If a b i o l o g i c a l s y s t e m i s u s e d in w h i c h c e l l s a r e a b l e to d iv ide , t h e p i c t u r e i s c o m p l i c a t e d d u r i n g a l o n g r a d i a t i o n e x p o s u r e a t a v e r y low d o s e - r a t e . U n d e r t h e s e c o n d i t i o n s , t he i n - v i t r o c e l l c u l t u r e t e c h n i q u e d o e s not p r o v i d e a r e l e v a n t a n s w e r , b e c a u s e t h e r e i s an a d d i t i o n a l d o s e - r a t e e f f e c t r e s u l t i n g f r o m c e l l p r o l i f e r a t i o n . The po in t a t w h i c h c e l l d i v i s i o n i s s t o p p e d by a low d o s e - r a t e i r r a d i a t i o n i s a f u n c t i o n of t h e d o s e p e r c e l l c y c l e .

9. 6. VERY L O W D O S E - R A T E S

T h e e f f e c t of v e r y low d o s e - r a t e s h a s b e e n i n v e s t i g a t e d by m e a s u r i n g the g r o w t h r a t e of c u l t u r e d m a m m a l i a n c e l l s , and by o b s e r v i n g r e n e w a l t i s s u e s of s m a l l a n i m a l s . N i a s and L a j t h a (1964) s t u d i e d t h e g r o w t h of

5 8 CHAPTER 10

H e L a c e l l s e x p o s e d c o n t i n u o u s l y to b e t a r a y s f r o m t r i t i a t e d w a t e r a t d o s e -r a t e s of 30 to 300 r a d / d a y . A d o s e - r a t e of 30 r a d / d a y had a b a r e l y d e t e c t -a b l e e f f e c t o v e r t he 1 1 - d a y p e r i o d of t h e e x p e r i m e n t .

C o u r t e n a y (1965) s h o w e d t h a t a s u s p e n s i o n c u l t u r e of m o u s e l y m p h o m a c e l l s , i r r a d i a t e d c o n t i n u o u s l y a t a d o s e - r a t e of 72 r a d / d a y , g r e w e x p o n e n t i a l l y , a l t h o u g h m o r e s l o w l y t h a n n o r m a l , f o r a p e r i o d of o v e r 10 m o n t h s . It w a s l a t e r r e p o r t e d t h a t c u l t u r e s of t h e s e c e l l s had s u r v i v e d c o n t i n u o u s i r r a d i a t i o n a t 120 r a d / d a y f o r 2 y e a r s , d u r i n g w h i c h t i m e r a ' d i o r e s i s t a n t m u t a n t s a p p e a r e d in t he e x p o s e d c u l t u r e s .

A c e l l p o p u l a t i o n c a n m a i n t a i n a s t e a d y s t a t e u n d e r c o n t i n u o u s i r r a d i a t i o n in a n u m b e r of t he r e n e w a l t i s s u e s in s m a l l a n i m a l s , a s l ong a s the p r o -d u c t i o n of n e w c e l l s by d i v i s i o n i s a b l e to m o r e t h a n b a l a n c e t h e d e a t h -r a t e of c e l l s due to i r r a d i a t i o n . T h e d o s e - r a t e t h a t c a n b e t o l e r a t e d v a r i e s w i t h t h e s p e c i e s and t y p e of t i s s u e c o n c e r n e d .

T h e t e s t e s a p p e a r to b e the m o s t s e n s i t i v e t i s s u e s t u d i e s ; r e p r o d u c t i o n c a n be m a i n t a i n e d in m a l e r a t s and m i c e f o r 10 g e n e r a t i o n s o r m o r e u n d e r c o n t i n u o u s e x p o s u r e a t a d o s e - r a t e of 2 r a d / d a y , but a s l i g h t i n c r e a s e in d o s e - r a t e above t h i s l e v e l r e s u l t s in a d e p l e t i o n of t h e t e s t e s c e l l p o p u l a -t ion . At t h e o t h e r e x t r e m e , t he s m a l l i n t e s t i n e in t h e r a t h a s b e e n shown to m a i n t a i n ce l l d i v i s i o n and a s t e a d y - s t a t e c e l l p o p u l a t i o n when c o n t i n u o u s l y i r r a d i a t e d a t a d o s e - r a t e a s h i g h a s 400 r a d / d a y . T h e b l o o d - f o r m i n g t i s s u e s a r e i n t e r m e d i a t e b e t w e e n t h e s e two e x t r e m e s ; r e d c e l l p r o d u c t i o n in t he r a t c a n b e m a i n t a i n e d a t c l o s e t o n o r m a l l e v e l s f o r m o n t h s whi le b e i n g e x p o s e d to 50 r a d / d a y .

L a m e r t o n and C o u r t e n a y (1968) d r e w a t t e n t i o n to t h r e e p r i n c i p a l f a c t o r s w h i c h can d e t e r m i n e t he r e s p o n s e of r e n e w a l t i s s u e s to c o n t i n u o u s i r r a d i a -t i o n . F i r s t , t h e c e l l u l a r s e n s i t i v i t y of t he s t e m c e l l s i n v o l v e d . S e c o n d , t h e d u r a t i o n of t h e c e l l c y c l e , s i n c e a s m a l l e r d o s e p e r c e l l c y c l e w i l l b e a c c u m u l a t e d in c e l l s wh ich t u r n o v e r a t a r a p i d r a t e . And t h i r d , t h e a b i l i t y of s o m e t i s s u e s to a d a p t to t h e new t r a u m a of c o n t i n u o u s i r r a d i a t i o n .

9. 7. R E P A I R AND T H E D O S E - R A T E E F F E C T WITH HIGH L E T IRRADIATION

F o r a g iven b i o l o g i c a l t e s t s y s t e m , t he m a g n i t u d e of t h e s h o u l d e r on t he s u r v i v a l c u r v e , and t h e r e f o r e t h e a m o u n t of s u b l e t h a l d a m a g e r e p a i r i n d i c a t e d by a s p l i t - d o s e e x p e r i m e n t , v a r i e s wi th t he type of r a d i a t i o n u s e d . A s a l r e a d y s t a t e d , t h e s h o u l d e r of t h e s u r v i v a l c u r v e i s g r e a t e s t f o r X - r a y s , i s c o m p l e t e l y a b s e n t f o r d e n s e l y i o n i z i n g r a d i a t i o n s s u c h a s a l p h a p a r t i c l e s , and h a s an i n t e r m e d i a t e v a l u e f o r f a s t n e u t r o n s . S p l i t -d o s e e x p e r i m e n t s i n d i c a t e r e p a i r of s u b l e t h a l d a m a g e w h i c h v a r i e s in m u c h t h e s a m e way . F o r X - r a y s , a g r e a t d e a l of d o s e s p a r i n g r e s u l t s f r o m d i v i d i n g t h e d o s e in to a n u m b e r of f r a c t i o n s . F o r a l p h a p a r t i c l e s t h e b i o l o g i c a l e f f e c t of a d o s e d i v i d e d in to f r a c t i o n s i s n e a r l y t h e s a m e a s t h a t of t h e s a m e t o t a l d o s e g iven in a s i n g l e f r a c t i o n , p r o v i d e d t h a t t he t i m e o v e r w h i c h t h e f r a c t i o n a t e d r e g i m e i s s p r e a d i s not l ong e n o u g h f o r a p p r e c i a b l e c e l l d i v i s i o n t o t a k e p l a c e . F o r 1 5 - M e V n e u t r o n s , t he r e s u l t s o b t a i n e d a r e i n t e r m e d i a t e b e t w e e n t h e s e two e x t r e m e s , a s would b e e x p e c t e d f r o m t h e s h a p e of t he s u r v i v a l c u r v e ; t h e r e i s a s m a l l a m o u n t of r e p a i r b e t w e e n s p l i t d o s e s . F o r l o w e r e n e r g y n e u t r o n s , s u c h a s f i s s i o n s p e c t r u m n e u t r o n s , t h e r e i s l i t t l e o r no r e p a i r b e t w e e n s p l i t d o s e s , s i n c e t he s u r v i v a l c u r v e i s e s s e n t i a l l y an e x p o n e n t i a l f u n c t i o n of d o s e .

C h a p t e r 10

L A T E E F F E C T S

10. 1. RADIATION HAZARDS

The n e e d f o r c a u t i o n in t he u s e of i o n i z i n g r a d i a t i o n s , w h i c h w i l l be d i s c u s s e d in C h a p t e r 11, i s due to t he l a t e e f f e c t s t h a t h a v e b e e n o b s e r v e d a t l o n g t i m e s a f t e r e x p o s u r e to r a d i a t i o n . L o n g - t e r m , l a t e , o r d e l a y e d e f f e c t s a r e t hough t to b e due to i r r e p a r a b l e d a m a g e to t he g e n e t i c m a t e r i a l of c e l l s wh ich s u r v i v e t he r a d i a t i o n e x p o s u r e . T h e s e e f f e c t s c a n b e p r o -duced by l o w e r d o s e s , and by l o w e r d o s e - r a t e s , t han i s the c a s e f o r a c u t e c e l l k i l l i n g e f f e c t s , w h i c h a r e u s u a l l y t he e n d - p o i n t of s h o r t - t e r m e x p e r i -m e n t s . L a t e e f f e c t s w i l l be d i s c u s s e d u n d e r f o u r s e p a r a t e h e a d i n g s : (a) e f f e c t s on t he e m b r y o and foe tus , (b) n o n - s p e c i f i c l i f e - s h o r t e n i n g , (c) c a r c i n o g e n e s i s , and (d) g e n e t i c e f f e c t s .

10. 2. RADIATION E F F E C T S ON T H E E M B R Y O AND F O E T U S

M o d e r a t e d o s e s of i o n i z i n g r a d i a t i o n h a v e b e e n s h o w n to p r o d u c e c a t a s t r o p h i c e f f e c t s on t h e d e v e l o p i n g e m b r y o o r f o e t u s , b o t h in t e r m s of g r o s s l e t h a l i t y , and in t he p r o d u c t i o n of a n o m a l i e s . M o s t e x p e r i m e n t a l d a t a in t h i s f i e l d h a v e b e e n o b t a i n e d wi th t h e m o u s e , bu t i t i s p r o b a b l y j u s t i f i e d to a s s u m e tha t t he m a j o r e f f e c t s s e e n in t he m o u s e w h e n r a d i a t i o n i s d e l i v e r e d a t s p e c i f i c s t a g e s of d e v e l o p m e n t would a l s o o c c u r in t h e h u m a n at t he e q u i v a l e n t s t a g e s .

R u s s e l l and R u s s e l l (1954) d i v i d e d up t h e t o t a l d e v e l o p m e n t p e r i o d in u t e r o in to t h r e e s t a g e s : (a) p r e - i m p l a n t a t i o n , wh ich e x t e n d s f r o m f e r t i l i z a t i o n to the t i m e when the e m b r y o a t t a c h e s to the w a l l of t he u t e r u s , o c c u p i e s d a y s 0 to 5j in t he m o u s e , wh ich c o r r e s p o n d s to t h e f i r s t 9 d a y s p o s t - c o n c e p t i o n in t h e h u m a n . D u r i n g t h i s p e r i o d t he p r i n c i p a l r e s u l t of e x p o s u r e t o r a d i a t i o n i s a h igh i n c i d e n c e of p r e n a t a l d e a t h s , e x p r e s s e d p r i m a r i l y a s a d e c r e a s e in l i t t e r s i z e .

O r g a n o g e n e s i s , t he p e r i o d d u r i n g wh ich the m a j o r o r g a n s a r e d e v e l o p i n g , e x t e n d s f r o m day 5 j to 13^ in t h e m o u s e , wh ich c o r r e s p o n d s to t he p e r i o d f r o m the 9th day to t h e 6th week in the h u m a n be ing .

F a r f e w e r p r e n a t a l d e a t h s a r e s e e n a s a c o n s e q u e n c e of i r r a d i a t i o n d u r i n g o r g a n o g e n e s i s , bu t i n s t e a d a g r e a t v a r i e t y of c o n g e n i t a l a n o m a l i e s of a s t r u c t u r a l n a t u r e m a y a p p e a r . D o s e s a s low a s 18 R of X - r a y s a r e qui te s u f f i c i e n t to p r o d u c e an a p p r e c i a b l e n u m b e r of a n o m a l i e s in s m a l l r o d e n t s . The p r o d u c t i o n of a s p e c i f i c d e f e c t i s a s s o c i a t e d wi th a d e f i n i t e t i m e d u r i n g t h i s p e r i o d of o r g a n o g e n e s i s , u s u a l l y the t i m e of t h e f i r s t m o r p h o l o g i c a l e v i d e n c e of d i f f e r e n t i a t i o n in the o r g a n , o r p o r t i o n of t he o r g a n , i nvo lved (Rugh 1962).

The f o e t a l p e r i o d e x t e n d s f r o m about day 14 o n w a r d s in t h e m o u s e , and c o r r e s p o n d s t o 6 w e e k s o n w a r d in the h u m a n . I r r a d i a t i o n d u r i n g t h i s p e r i o d l e a d s to f a r f e w e r o b v i o u s s t r u c t u r a l a n o m a l i e s . N e v e r t h e l e s s , f u n c t i o n a l d i s a b i l i t i e s wh ich a r e m u c h m o r e d i f f i c u l t to i d e n t i f y can o c c u r a s a r e s u l t of i r r a d i a t i o n d u r i n g t h i s p e r i o d . It h a s b e e n s u g g e s t e d t h a t g r o w t h d i s o r d e r s , and p o s s i b l y a l o w e r i n g of s u b s e q u e n t i n t e l l i g e n c e ,

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6 0 CHAPTER 10

r e s u l t f r o m e x p o s u r e to r a d i a t i o n a t t h i s t i m e . Much h i g h e r d o s e s of i r r a d i a t i o n a r e r e q u i r e d to k i l l e m b r y o s d u r i n g t h i s p e r i o d t h a n a t e a r l i e r s t a g e s of d e v e l o p m e n t .

B e c a u s e of t h e s e c o n s i d e r a t i o n s , it i s i m p o r t a n t to e n s u r e t h a t p r e g n a n t w o m e n a r e no t i r r a d i a t e d , p a r t i c u l a r l y d u r i n g t h e e a r l y w e e k s f o l l o w i n g c o n c e p t i o n . S i n c e p r e g n a n c y c a n b e u n d e t e c t e d d u r i n g t h e s e e a r l y t i m e s when t h e f o e t u s i s m o s t v u l n e r a b l e , a l l w o m e n of c h i l d - b e a r i n g age s h o u l d be a s s u m e d to b e p r e g n a n t a t a l l t i m e s , and r a d i a t i o n l e v e l s kep t to a m i n i m u m . If d i a g n o s t i c X - r a y s a r e n e e d e d f o r m e d i c a l r e a s o n s , t hey shou ld b e t a k e n d u r i n g the 10 d a y s f o l l o w i n g the o n s e t of a m e n s t r u a l p e r i o d to avo id a c c i d e n t a l i r r a d i a t i o n of a c o n c e p t u s .

10. 3. N O N - S P E C I F I C L I F E - S H O R T E N I N G

T h e r e a r e m a n y r e p o r t s in t h e l i t e r a t u r e t h a t e x p o s u r e t o i o n i z i n g r a d i a t i o n s s h o r t e n s t he l i f e s p a n of s m a l l a n i m a l s . A f t e r a d o s e of r a d i a t i o n i n s u f f i c i e n t to c a u s e e a r l y l e t h a l i t y , a n i m a l s a p p e a r to r e c o v e r c o m p l e t e l y ; b lood c o u n t s r e t u r n t o n o r m a l , g a s t r o i n t e s t i n a l s y m p t o m s d i s a p p e a r , and w e i g h t r e t u r n s a l m o s t to n o r m a l . N e v e r t h e l e s s , t h e s e a n i m a l s d ie s o o n e r t h a n c o n t r o l s . R o t b l a t and L i n d o p (1961) found a l i n e a r r e l a t i o n s h i p b e t w e e n l i f e s h o r t e n i n g and r a d i a t i o n d o s e . F o r e a c h 100 r a d of X - r a y s d e l i v e r e d a s an a c u t e e x p o s u r e , t he l i f e e x p e c t a n c y of t h e m i c e w a s r e d u c e d by about 5%.

I r r a d i a t e d s u r v i v o r s b e h a v e j u s t l i k e o l d e r u n i r r a d i a t e d a n i m a l s . In o t h e r w o r d s , t h e y a p p e a r to l o s e p a r t of t h e i r y o u t h o r e a r l y l i f e . No n e w o r d i f f e r e n t d i s e a s e s h a v e b e e n o b s e r v e d a s l a t e e f f e c t s in i r r a d i a t e d a n i m a l s . M o s t r e p o r t s in t he l i t e r a t u r e c l a i m t h a t a l l c a u s e s of d e a t h a r e a c c e l e r a t e d by r a d i a t i o n , and f o r t h i s r e a s o n t h e t e r m s ' n o n - s p e c i f i c l i f e - s h o r t e n i n g ' and ' r a d i a t i o n a g i n g ' h a v e b e e n i n t r o d u c e d . H o w e v e r , t h e r e i s i n c r e a s i n g e v i d e n c e f r o m m o r e r e c e n t s t u d i e s of h u m a n and a n i m a l p o p u l a t i o n s , t h a t in t he low to i n t e r m e d i a t e d o s e r a n g e , l i f e - s h o r t e n i n g can be a c c o u n t e d f o r e n t i r e l y by the i n d u c t i o n of n e o p l a s m s .

T h e on ly e v i d e n c e in the h u m a n b e i n g f o r a l i f e - s h o r t e n i n g e f f e c t of r a d i a t i o n c o m e s f r o m the s tudy of A m e r i c a n r a d i o l o g i s t s ; t h i s i n d i c a t e d tha t t he l i f e - s p a n of t h o s e who d i ed d u r i n g t h e p e r i o d 1945-54 w a s s h o r t e r t h a n t h a t of o t h e r g r o u p s of m e d i c a l p r a c t i t i o n e r s . The a n a l y s i s of t h e s e da t a h a s b e e n s e v e r e l y c r i t i c i z e d , s o t he c o n c l u s i o n s a r e in doub t . In add i t ion , s i m i l a r s t u d i e s of B r i t i s h r a d i o l o g i s t s o v e r t h e p e r i o d f r o m 1897 to 1957 f a i l e d t o s h o w any c o m p a r a b l e s h o r t e n i n g of l i f e , a l t h o u g h t h e s e r a d i o l o g i s t s i n c l u d e d m a n y who w e r e e x p o s e d in t h e e a r l y d a y s w h e n r a d i a t i o n s a f e t y s t a n d a r d s w e r e v e r y l a x .

1 0 . 4 . RADIATION C A R C I N O G E N E S I S

T h e p r o d u c t i o n of n e o p l a s m s by r a d i a t i o n h a s b e e n o b s e r v e d f r o m the e a r l y u s e s of X - r a y s . R a d i a t i o n c a r c i n o g e n e s i s h a s b e e n s t u d i e d e x t e n s i v e l y in s m a l l a n i m a l s , and a c h a r a c t e r i s t i c r e l a t i o n s h i p b e t w e e n i n c i d e n c e and the X - r a y d o s e i s s h o w n in F i g . 29. As t he d o s e i s i n c r e a s e d t h e i n c i d e n c e of m a l i g n a n c y a l s o i n c r e a s e s , u p to a m a x i m u m wh ich u s u a l l y o c c u r s a t about 300 r a d . F o r h i g h e r d o s e s t h e i n c i d e n c e d r o p s a g a i n .

LATE EFFECTS 6 1

40-

30-

1 20-

t /

/ / o

\

(0-

50 tOO 150 300 X-RAY DOSE (rad)

400 450

FIG. 29. Incidence of myeloid leukemia in RF m a l e mice exposed to whole-body X-i r radia t ion. (From Upton, 1961).

T o o b t a i n m e a n i n g f u l r e s u l t s w i th a r e a s o n a b l e n u m b e r of a n i m a l s , it i s c o m m o n p r a c t i c e to u s e a s t r a i n of a n i m a l s w h i c h show a h igh n a t u r a l i n c i d e n c e of t h e d i s e a s e . The m o s t c o m m o n e x a m p l e s a r e l e u k e m i a in s p e c i f i c s t r a i n s of m i c e , o r m a m m a r y c a r c i n o m a in a p a r t i c u l a r t y p e of r a t . W h i l e t h e c h o i c e of s u c h s y s t e m s m a k e s t he s t u d i e s m u c h e a s i e r to p e r f o r m , the r e s u l t s a r e l e s s u s e f u l t h a n would be t h e c a s e if t h e e x p e r i m e n t s cou ld be p e r f o r m e d wi th a n i m a l s t h a t do not show a p a r t i c u l a r s u s c e p t i b i l i t y to the t u m o u r s t u d i e d .

T h e r e a r e s o m a n y i n s t a n c e s on r e c o r d of m a l i g n a n t n e o p l a s m s b e i n g p r o d u c e d in h u m a n b e i n g s a s a r e s u l t of i r r a d i a t i o n , t h a t it i s no t n e c e s s a r y to d e p e n d e x c l u s i v e l y upon a n i m a l d a t a in t h i s f i e ld .

Sk in c a n c e r w a s o f t e n o b s e r v e d in e a r l y w o r k e r s wi th X - r a y s who g a v e l i t t l e c o n s i d e r a t i o n to r a d i a t i o n p r o t e c t i o n . L u n g c a n c e r o c c u r s f r e q u e n t l y a m o n g s t t h e p i t c h b l e n d e m i n e r s in Saxony , and t h e u r a n i u m m i n e r s in C o l o r a d o , who a s a r e s u l t of t h e i r o c c u p a t i o n i n h a l e q u a n t i t i e s of r a d i o -a c t i v e g a s .

Bone t u m o u r s h a v e b e e n o b s e r v e d in t he r a d i u m d i a l p r i n t e r s , who w e r e m o s t l y young g i r l s e m p l o y e d in f a c t o r i e s p r o d u c i n g l u m i n o u s c l o c k s and w a t c h e s . It w a s c o m m o n p r a c t i c e f o r t h e s e p e o p l e to l i c k t h e i r b r u s h in to s h a p e w i t h t h e i r t o n g u e a f t e r it had b e e n d ipped in to the p a i n t c o n t a i n i n g r a d i u m . O v e r a p e r i o d of y e a r s , a p p r e c i a b l e q u a n t i t i e s of r a d i o a c t i v e m a t e r i a l w e r e i n g e s t e d , and m o s t of the w o r k e r s who did not die of a n a e m i a , l a t e r d i ed of bone t u m o u r s . .

T h e c o n t r a s t m e d i u m t h o r o t r a s t i s r a d i o a c t i v e , and p a t i e n t s in w h o m i t w a s u s e d h a v e an e l e v a t e d i n c i d e n c e of c a n c e r of t h e l i v e r .

T h e s u r v i v o r s of t h e a t o m b o m b e x p l o s i o n s in H i r o s h i m a and N a g a s a k i c o m p r i s e t h e l a r g e s t g r o u p of i r r a d i a t e d h u m a n b e i n g s , and c l e a r l y s h o w an e l e v a t e d i n c i d e n c e of l e u k e m i a and of o t h e r t y p e s of c a n c e r a s w e l l .

In t h e U n i t e d K i n g d o m , d u r i n g t h e l a t e 1930s and e a r l y 1940s , m a n y t h o u s a n d s of p e o p l e w e r e t r e a t e d wi th X - r a y s f o r t he r e l i e f of p a i n a s s o c i a t e d

6 2 CHAPTER 10

wi th a n k y l o s i n g s p o n d y l i t i s . T h e s e p a t i e n t s w e r e s u b s e q u e n t l y s h o w n to h a v e an i n c r e a s e d i n c i d e n c e of l e u k e m i a .

T h e r e i s s o m e e v i d e n c e , a l t h o u g h not v e r y c o n v i n c i n g , t h a t A m e r i c a n r a d i o l o g i s t s s h o w an i n c r e a s e d i n c i d e n c e of l e u k e m i a c o m p a r e d wi th t he g e n e r a l p o p u l a t i o n , bu t t he t o t a l n u m b e r of c a s e s i s v e r y s m a l l , and the s t a t i s t i c a l v a l i d i t y i s in doubt . C h i l d r e n who w e r e t r e a t e d m a n y y e a r s ago f o r an e n l a r g e d t h y m u s g land c e r t a i n l y show an e l e v a t e d i n c i d e n c e of c a n c e r of t h e t h y r o i d .

T h e r e can b e no doubt , t h e n , t h a t r a d i a t i o n r e c e i v e d by h u m a n b e i n g s r e s u l t s in an i n c r e a s e in n e o p l a s t i c d i s e a s e . The J a p a n e s e s u r v i v o r s and the B r i t i s h s p o n d y l i t i c p a t i e n t s a r e t he on ly g r o u p s l a r g e enough f o r any q u a n t i t a t i v e a s s e s s m e n t to b e m a d e .

Whi l e i t i s l e u k e m i a t h a t i s a s s o c i a t e d wi th t h e a t o m b o m b in t h e m i n d s of m o s t p e o p l e , i t i s b e c o m i n g i n c r e a s i n g l y a p p a r e n t a s t he y e a r s p a s s t h a t i t m a y no t b e t he m o s t i m p o r t a n t m a l i g n a n c y a s s o c i a t e d wi th r a d i a t i o n . A s t h e J a p a n e s e s u r v i v o r s a r e s t u d i e d f o r a l o n g e r p e r i o d of t i m e , l ung c a n c e r , b r e a s t c a n c e r , and p a r t i c u l a r l y t h y r o i d c a n c e r , a l l a p p e a r m o r e f r e q u e n t l y t h a n in t h e J a p a n e s e p o p u l a t i o n a s a w h o l e . T h e s e t y p e s of c a n c e r t a k e m u c h l o n g e r to d e v e l o p ; w h e r e a s t he i n c i d e n c e of r a d i a t i o n -i n d u c e d l e u k e m i a r e a c h e s a p e a k 5 to 7 y e a r s a f t e r i r r a d i a t i o n and d e c l i n e s by 15 to 20 y e a r s , o t h e r t y p e s of c a n c e r s a r e on ly now a p p e a r i n g , n e a r l y 30 y e a r s a f t e r t h e e x p l o s i o n of t h e b o m b . In t he l ong t e r m it a p p e a r s l i k e l y t h a t a t l e a s t 5, if no t 10, t i m e s a s m a n y - p e o p l e w i l l d ie of o t h e r t y p e s of c a n c e r c a u s e d by t h e b o m b , a s h a v e d i ed of l e u k e m i a .

T h e q u a n t i t a t i v e d a t a ' w h i c h e x i s t r e l a t i n g t h e n u m b e r of c a s e s of n e o p l a s m s to t he d o s e r e c e i v e d , a l l r e f e r to r e l a t i v e l y l a r g e d o s e s of r a d i a t i o n c o n s i s t i n g of m o r e t h a n 50 r a d of a m i x t u r e of n e u t r o n s and g a m m a r a y s . R i s k e s t i m a t e s f o r the n u m b e r of c a n c e r s p r o d u c e d by low d o s e s of r a d i a t i o n , s u c h a s t h o s e r e c e i v e d o c c u p a t i o n a l l y o r in t h e c o u r s e of m e d i c a l d i a g n o s t i c X - r a y s , c a n only b e m a d e on t h e b a s i s of a s e r i e s of t e n u o u s a s s u m p t i o n s . It i s no t t he p u r p o s e of t h i s book to d i s c u s s s u c h r i s k e s t i m a t e s , d e t a i l s of wh ich c a n b e found e l s e w h e r e . In t h e p a s t , g e n e t i c e f f e c t s w e r e c o n s i d e r e d to be t h e on ly i m p o r t a n t c o n s e q u e n c e s of low d o s e s of r a d i a t i o n , but t h e r e i s now r e a s o n to b e l i e v e t h a t s o m a t i c e f f e c t s m a y be e q u a l l y i m p o r t a n t .

10. 5. G E N E T I C E F F E C T S

A c o n s i d e r a b l e d e g r e e of u n c e r t a i n t y m u s t be a c c e p t e d in e s t i m a t i n g the g e n e t i c h a z a r d s of r a d i a t i o n e x p o s u r e in m a n . T h e l a r g e s t g r o u p of h u m a n b e i n g s e x p o s e d to r a d i a t i o n a r e t he J a p a n e s e s u r v i v o r s of H i r o s h i m a and N a g a s a k i , and y e t u p to t h e p r e s e n t t i m e no i n c r e a s e h a s b e e n o b s e r v e d in t he i n c i d e n c e of p r e n a t a l o r n e o n a t a l d e a t h s , o r in t he f r e q u e n c y of m a l f o r m a t i o n s . H o w e v e r , by g e n e t i c s t a n d a r d s , t he n u m b e r of p e o p l e e x p o s e d w a s s m a l l , and i n s u f f i c i e n t t i m e h a s e l a p s e d f o r r e c e s s i v e m u t a t i o n s to b e e x p r e s s e d . C o n s e q u e n t l y , d a t a f r o m the i r r a d i a t i o n of s m a l l m a m m a l s i s a l l t h a t i s a v a i l a b l e .

In t h e y e a r s f o l l o w i n g the Second W o r l d W a r , l a r g e - s c a l e e x p e r i m e n t s w e r e u n d e r t a k e n to d e t e r m i n e t h e m u t a t i o n r a t e s in t he m o u s e f o l l o w i n g i r r a d i a t i o n ( R u s s e l l 1963a, b) . S p e c i f i c g e n e m u t a t i o n s w e r e s c o r e d , w h i c h

LATE EFFECTS 6 3

i n c l u d e d a c h a n g e in coa t c o l o u r and o b v i o u s d e f e c t s of t he e a r o r t a i l . T h e r e s u l t s of t h e s e e x p e r i m e n t s t u r n e d ou t to be v e r y c o m p l i c a t e d .

In g e n e r a l , t h e m a l e i s m u c h m o r e s e n s i t i v e t h a n t h e f e m a l e a s f a r a s t he g e n e t i c e f f e c t s of a g iven d o s e of r a d i a t i o n a r e c o n c e r n e d . T h e r e i s e v i d e n c e of an i m p o r t a n t d o s e - r a t e e f f e c t ; a r e d u c t i o n in d o s e - r a t e r e s u l t e d in a s i g n i f i c a n t and i m p o r t a n t r e d u c t i o n in t h e a m o u n t of g e n e t i c d a m a g e p r o d u c e d by a g iven d o s e of r a d i a t i o n . A t i m e i n t e r v a l b e t w e e n i r r a d i a t i o n and c o n c e p t i o n i s known to r e d u c e t he n u m b e r of m u t a t i o n s p r o d u c e d , w h e t h e r m a l e o r f e m a l e i s e x p o s e d ( R u s s e l l 1965).

E x t r a p o l a t i n g t he m o u s e d a t a t o m a n l e a d s to t he c o n c l u s i o n t h a t t h e doub l ing d o s e e q u i v a l e n t , 2 t h a t i s t he d o s e r e q u i r e d to doub le t h e s p o n t a n e o u s m u t a t i o n r a t e , p r o b a b l y l i e s b e t w e e n 10 and 200 r e m . When h u m a n b e i n g s r e c e i v e r a d i a t i o n , w h e t h e r a s m e m b e r s of t h e g e n e r a l p u b l i c o r in t he c o u r s e of t h e i r o c c u p a t i o n , c o n d i t i o n s a r e s u c h t h a t m u t a t i o n r a t e s would be e x p e c t e d to b e low. E i t h e r t h e d o s e - r a t e i s low and c o n t i n u o u s a s in o c c u p a t i o n e x p o s u r e o r , w h e r e h igh d o s e - r a t e s a r e i n v o l v e d , t he t o t a l d o s e p e r e x p o s u r e i s s m a l l . On t h e r a r e o c c a s i o n w h e n a l a r g e d o s e i s a b s o r b e d in a r a d i a t i o n a c c i d e n t , a s i g n i f i c a n t p r o p o r t i o n of t he d e l e t e r i o u s g e n e t i c e f f e c t s can b e a v o i d e d if c o n c e p t i o n i s d e f e r r e d f o r s o m e m o n t h s .

* See the defini t ion of dose-equivalent H in Section 1 1 . 1 .

C h a p t e r 11

RADIATION P R O T E C T I O N

1 1 . 1 . P E R M I S S I B L E DOSES

It s h o u l d be a s s u m e d t h a t any e x p o s u r e to i o n i z i n g r a d i a t i o n c a r r i e s s o m e r i s k of d e l e t e r i o u s e f f e c t s and tha t , t h e r e f o r e , u n n e c e s s a r y e x p o s u r e s shou ld b e a v o i d e d . It s hou ld a l s o b e r e a l i z e d t h a t n e u t r o n s r e p r e s e n t an e s p e c i a l h a z a r d . T h e f o l l o w i n g i m p o r t a n t p o i n t s a r e c o n d e n s e d f r o m ICRU p u b l i c a t i o n 9 (1965) and t h e N C R P r e p o r t s 38 and 39 (1971).

T h e p r i n c i p a l q u a n t i t y of i m p o r t a n c e in r a d i a t i o n p r o t e c t i o n i s t he d o s e e q u i v a l e n t H, m e a s u r e d in r e m , wh ich i s t he p r o d u c t of a b s o r b e d d o s e D and t h e q u a l i t y f a c t o r Q. T h u s

H = QD

Q d e p e n d s on t h e L E T of t h e c h a r g e d p a r t i c l e s t h a t p r o d u c e D. It i s I . 0 f o r g a m m a r a y s and 8. 5 f o r n e u t r o n s f r o m 2 5 2 C f .

T h e n e u t r o n t i s s u e k e r m a in f r e e a i r a t 1 m f r o m a 1 (jg 2 5 2 C f s o u r c e i s a b o u t 0. 2 m r a d / h . The m a x i m u m v a l u e of D n e a r t h e s u r f a c e of a h u m a n body a t t h a t d i s t a n c e i s abou t 0. 28 m r a d / h ( the l a r g e r v a l u e i s d u e t o b a c k -s c a t t e r i n g by t h e body) . A c c o r d i n g to t he a b o v e e q u a t i o n t he d o s e e q u i v a l e n t at t he body s u r f a c e i s abou t 2 . 4 m r e m / h . The d o s e e q u i v a l e n t d u e to g a m m a r a d i a t i o n i s abou t 0. 15 i n r e m / h .

No e x p e r i m e n t e r o l d e r t h a n 18 y e a r s s h o u l d r e c e i v e , in any w e e k , a d o s e e q u i v a l e n t t h a t i s m o r e t h a n 100 m r e m . No m e m b e r of t h e g e n e r a l popu l a t i on , and no i n d i v i d u a l y o u n g e r t h a n 18 y e a r s , s h o u l d r e c e i v e m o r e t h a n 10 m r e m / w e e k .

I I . 2. R E Q U I R E M E N T S F O R A RADIATION S A F E T Y P R O G R A M

T h e f o l l o w i n g a r e the m i n i m u m r e q u i r e m e n t s f o r an a d e q u a t e r a d i a t i o n s a f e t y p r o g r a m .

(1) An i n d i v i d u a l p e r s o n a l l y r e s p o n s i b l e f o r s o u r c e s m u s t b e d e s i g n a t e d b e f o r e t he r e c e i p t of s o u r c e s . He m u s t k e e p a n i n v e n t o r y w h i c h e n a b l e s h i m to a c c o u n t f o r a l l s o u r c e s a t a l l t i m e s , m u s t e n s u r e t h a t s o u r c e s a r e h a n d l e d p r o p e r l y and s a f e l y , and m u s t b e r e s p o n s i b l e f o r t he p e r f o r m a n c e of l e a k t e s t s ( s e e be low) .

(2) E f f o r t s shou ld b e m a d e to e n s u r e t h a t s o u r c e s do in f a c t h a v e t he s t a t e d 2 5 2Cf c o n t e n t . T h i s r e q u i r e s m e a s u r e m e n t s of t i s s u e k e r m a in f r e e a i r w h i c h m a y e i t h e r be t h e t o t a l t i s s u e k e r m a o r m e r e l y t h a t due to g a m m a r a d i a t i o n ( s ee be low) .

(3) T h e r e m u s t be a d e q u a t e s o u r c e s e c u r i t y . T h i s r e q u i r e s t h a t when no t in a c t u a l u s e s o u r c e s a r e e i t h e r l o c k e d in a c o n t a i n e r o r t h a t t h e c o n t a i n e r i s in a l o c k e d r o o m w i t h a c c e s s c o n t r o l l e d by the r e s p o n s i b l e i n d i v i d u a l .

6 5

6 6 CHAPTER 10

(4) The c o n t a i n e r ( cask ) in wh ich s o u r c e s a r e s t o r e d s h o u l d h a v e a d e q u a t e s h i e l d i n g . T h e b a s i c c r i t e r i o n i s t h a t s o u r c e s be s t o r e d in s u c h a w a y t h a t no i n d i v i d u a l r e c e i v e s a d o s e e q u i v a l e n t in e x c e s s of 100 m r e m / w e e k . T h e s h i e l d i n g r e q u i r e d to a c c o m p l i s h t h i s d e p e n d s on l o c a t i o n and o c c u p a n c y of t h e e n v i r o n s of t h e c a s k . H o w e v e r , a s a m i n i m u m the s h i e l d i n g s h o u l d be s u c h t h a t the d o s e e q u i v a l e n t r a t e a t i t s s u r f a c e no t e x c e e d 200 m r e m / h and t h a t a t a d i s t a n c e of 1 m f r o m t h e c a s k to d o s e e q u i v a l e n t r a t e b e l e s s t h a n 10 m r e m / h .

(5) The c a s k and the r o o m in w h i c h it i s n o r m a l l y kep t s h o u l d c a r r y w a r n i n g s i g n s . If a t a l l p o s s i b l e s o u r c e s sho u l d b e p r o v i d e d wi th l a b e l s bu t t h e y shou ld in any c a s e a l w a y s b e s t o r e d in l a b e l l e d c o n t a i n e r s t h a t c l e a r l y i d e n t i f y t h e n a t u r e and s t r e n g t h of s o u r c e s .

(6) S o u r c e s s h o u l d n e v e r b e t o u c h e d b y h a n d and the u s e of t o n g s i s e s s e n t i a l . T h e t o n g s s h o u l d be s o c o n s t r u c t e d a s to m i n i m i z e t he p o s s i b i l i t y of m e c h a n i c a l d a m a g e to t he s o u r c e .

(7) S o u r c e s shou ld be l e a k - t e s t e d a t i n t e r v a l s no t e x c e e d i n g 6 m o n t h s . In l e a k t e s t i n g , t h e s o u r c e shou ld e i t h e r be w iped a g a i n s t f i l t e r p a p e r (when h e l d by tongs ) o r a l t e r n a t i v e l y t h e s o u r c e s h o u l d be i m m e r s e d in e t h y l a l c o h o l f o l l o w e d by e v a p o r a t i o n of t h e s o l v e n t . T h e f i l t e r p a p e r , o r t h e r e s i d u e a f t e r e v a p o r a t i o n , s h o u l d be c o u n t e d f o r a l p h a a c t i v i t y . Any l e a k i n g s o u r c e s h o u l d i m m e d i a t e l y be p l a c e d in an a i r t i gh t c o n t a i n e r and r e t u r n e d to t h e m a n u f a c t u r e r .

(8) Many of t h e i n s t r u m e n t s u t i l i z e d in p h y s i c s e x p e r i m e n t s h a v e a s e n s i t i v i t y c o m p a r a b l e t o o r e v e n l e s s t h a n t h a t c o r r e s p o n d i n g to d o s e e q u i v a l e n t r a t e s n e a r p e r m i s s i b l e l e v e l s . In e x p e r i m e n t s r e q u i r i n g a p p r e c i a b l e p e r i o d s of t i m e it m a y t h e r e f o r e o f t e n b e n e c e s s a r y to p r o v i d e a r e m o t e r e a d o u t d e v i c e r a t h e r t h a n h a v i n g the e x p e r i m e n t e r r e m a i n in t he i m m e d i a t e v i c i n i t y of t h e d e t e c t o r . In t h i s c o n n e c t i o n s p e c i a l c o n s i d e r a -t i on shou ld be g i v e n to m i n i m i z i n g e x p o s u r e of t h e e y e s .

11. 3. SHIELDING R E Q U I R E M E N T S F O R C A L I F O R N I U M - 2 5 2

W a t e r - e x t e n d e d p o l y e s t e r s (WEP) o r p o l y e t h y l e n e a r e p r o b a b l y t he b e s t s h i e l d i n g m a t e r i a l s f o r 2 5 2 Cf s o u r c e s . P a r a f f i n h a s v i r t u a l l y t h e s a m e a t t e n u a t i o n c h a r a c t e r i s t i c s , and a l t hough it i s a f i r e h a z a r d , it i s f r e q u e n t l y e m p l o y e d e s p e c i a l l y when e n c l o s e d in m e t a l l i c c o n t a i n e r s . W a t e r , w h i c h i s t he s i m p l e s t n e u t r o n s h i e l d i n g m a t e r i a l a v a i l a b l e , i s no t r e c o m m e n d e d f o r s t o r a g e c o n t a i n e r s b e c a u s e of e v a p o r a t i o n o r l e a k a g e . H o w e v e r , w a t e r -f i l l e d b u c k e t s m a y s e r v e a s t e m p o r a r y s h i e l d s p r o v i d e d tha t t he s o u r c e i s r e t a i n e d in a c l o s e d m e t a l t ube t e r m i n a t i n g n e a r t h e c e n t r e of t h e b u c k e t w h i c h p r e v e n t s c o n t a c t b e t w e e n t h e s o u r c e and t h e w a t e r .

In c a l c u l a t i n g r e q u i r e d s h i e l d i n g t h i c k n e s s e s t he fo l l owing a p p r o x i m a -t i o n s 3 c a n be u s e d :

3 None of these approximations are entirely correct . Although they should result in adequately conserva-t ive radiation protection design they should not be employed for other purposes.

RADIATION PROTECTION 67

1. The c a l i f o r n i u m - 2 5 2 i s c o n s i d e r e d a s o u r c e of n e u t r o n s on ly and p r o d u c e s a d o s e e q u i v a l e n t r a t e of 2. 5 m r e m / h a t a d i s t a n c e of 1 m p e r jug of 2 5 2 C f .

2. T h e i n v e r s e s q u a r e l a w a p p l i e s . 3. T h e half v a l u e l a y e r (HVL) of W E P , p o l y e t h y l e n e , p a r a f f i n o r w a t e r

i s 5 c m .

A s an e x a m p l e , c o n s i d e r a s o u r c e of 25 y.g w h i c h i s to b e p l a c e d in a c o n t a i n e r w h i c h i s 2 m r e m o v e d f r o m an a r e a w h e r e p e r s o n n e l s p e n d 50 h o u r s p e r week . Wi th a m a x i m u m p e r m i s s i b l e d o s e e q u i v a l e n t of 100 m r e m / w e e k t h e m a x i m u m p e r m i s s i b l e d o s e e q u i v a l e n t r a t e i s 2 m r e m • h " 1 . W i t h 2. 5 m r e m • h"1 p e r pg a t 1 m t h e d o s e e q u i v a l e n t r a t e d u e to t h e u n s h i e l d e d s o u r c e i s , a t 1 m , 2. 5X 25 = 62. 5 m r e m • h"1 and at 2 m i t i s 62. 5 / 4 = 15. 8 m r e m • h" 1 . T h i s i s abou t e igh t t i m e s a s h i g h a s t h e l i m i t i n g d o s e r a t e . C o n s e q u e n t l y 3 HVL, o r 15 c m , of s h i e l d i n g m a t e r i a l (hav ing a h y d r o g e n c o n t e n t of m o r e t h a n 10%) i s r e q u i r e d . T h u s t he c a s k m u s t h a v e a d i a m e t e r and a h e i g h t of abou t 30 c m if t he s m a l l c e n t r a l t u b e in w h i c h the s o u r c e i s s t o r e d h a s n e g l i g i b l e d i a m e t e r .

P a r t 2

LABORATORY MANUAL

C h a p t e r 12

G E N E R A L INFORMATION F O R LABORATORY E X P E R I M E N T S

1 2 . 1 . A PRE-REQUISITE FOR A L L E X P E R I M E N T E R S

P e r s o n s p e r f o r m i n g e x p e r i m e n t s with 252Cf should be f a m i l i a r with the contents of Chapter 11 c o n c e r n i n g rad ia t ion p r o t e c t i o n regu la t ion and p r o c e d u r e s . Ideal ly a c o u r s e of in s t ruc t ion should have b e e n comple ted which inc ludes a l l ' the m a t e r i a l included in the in s t ruc t iona l s y l l a b u s .

1 2 . 2 . DETERMINATIONS OF DOSE FOR E X P E R I M E N T S WITH CALIFORNIUM-252

Although a t tempts at independent d e t e r m i n a t i o n s a re not d i s couraged , it i s l ike ly that in the major i ty of i n s t a n c e s the m o s t a c c u r a t e v a l u e s f o r g a m m a radiat ion, as w e l l as neutrons , of t i s s u e k e r m a in f r e e a ir or of the absorbed dose in phantoms wi l l be obtained by mult ip ly ing the appropriate data in Appendix I by the m a s s of a s o u r c e (in^ug). The v a l u e s of the m a s s , as g i v e n by the manufac turer , should be a c c u r a t e to 2 or 3%. T h e s e must of c o u r s e be c o r r e c t e d f o r decay in the fo l lowing way .

P r o c u r e a s h e e t of s e m i l o g a r i t h m i c graph paper . The l o g a r i t h m i c ordinate should be labe l l ed ' C o r r e c t i o n f a c t o r ' and the l i n e a r a b s c i s s a ' T i m e in y e a r s ' . Draw a s tra ight l ine def ined by 0 t i m e and 1.0 c o r r e c t i o n f a c t o r and 2 . 6 5 y e a r t i m e and 0 . 5 c o r r e c t i o n f a c t o r . U s e th i s chart to d e t e r m i n e the d e c a y c o r r e c t i o n for any s o u r c e of 2 5 2 Cf.

The m a s s of 252Cf in the s o u r c e may , h o w e v e r , be unknown or u n c e r t a i n . In th i s connec t ion it should be r e a l i z e d that s o u r c e s of unequal s t rength can have the s a m e e x t e r n a l appearance and that it i s t h e r e f o r e p o s s i b l e that b e c a u s e of i n t e r c h a n g e s the wrong m a s s i s attr ibuted to s o u r c e s . Thus if there i s no r e l i a b l e c e r t i f i c a t i o n of s o u r c e s t rength ava i lab le , the e x p e r i -m e n t e r m u s t d e t e r m i n e it on his own and such m e a s u r e m e n t s should in any c a s e be c a r r i e d out as part of the laboratory e x e r c i s e s . Various methods for a c c o m p l i s h i n g th i s are g i v e n in l a t e r chapters .

12. 3. VARIATIONS FROM THE MANUAL

The e x p e r i m e n t s d e s c r i b e d in the c h a p t e r s which fo l low have b e e n s e l e c t e d for d idact ic v a l u e . The d e s c r i p t i o n s a re b e l i e v e d to be adequate to guide the student towards the c o r r e c t a p p r o a c h e s . V a r i a n c e s f r o m the indicated p r o c e d u r e s , as w e l l as fur ther s t u d i e s , may provide valuable addit ional e x p e r i e n c e provided that they are c a r r i e d out under the s u p e r -v i s i o n of an i n s t r u c t o r who i s s u f f i c i e n t l y e x p e r i e n c e d to f o r e s e e p o s s i b l e c o m p l i c a t i o n s . B e c a u s e of the v a r i e t y of i n s t r u m e n t s and techniques that may be employed , r e s u l t s are o f t en unpredic table and are t h e r e f o r e not s p e c i f i e d in deta i l .

7 1

7 2 CHAPTER 10

1 2 . 4 . E Q U I P M E N T AVAILABILITY

A l l the e x p e r i m e n t s d e s c r i b e d below r e q u i r e s p e c i a l i z e d equipment , which in the c a s e of the m o r e advanced s tud ie s outl ined in Chapter 15 may be di f f icul t or i m p o s s i b l e to cons truc t l o c a l l y . In this c a s e purchase may be n e c e s s a r y . Informat ion on c o m m e r c i a l s u p p l i e r s of such equipment i s ava i lab le f r o m the D o s i m e t r y Sect ion of the Internat ional A t o m i c E n e r g y A g e n c y .

C h a p t e r 13

E X P E R I M E N T S WITH A GM COUNTER

1 3 . 1 . COUNTER R E S P O N S E TO NEUTRONS

Any p r o p e r l y o p e r a t i n g G e i g e r - M ü l l e r (GM) c o u n t e r i s m u c h m o r e s e n s i t i v e to g a m m a than to neutron r a d i a t i o n if s e n s i t i v i t y i s r e l a t e d to t i s s u e k e r m a . H e n c e GM c o u n t e r e x p e r i m e n t s c a n be c o n s i d e r e d to be un-a f f e c t e d by the p r e s e n c e of n e u t r o n s ( e x c e p t f o r capture g a m m a r a d i a t i o n w h i c h m a y b e c o m e i m p o r t a n t w h e n neutrons a r e t h e r m a l i z e d - s e e b e l o w ) .

1 3 . 2 . ELIMINATION OF B E T A RADIATION

Suspend a 2 5 2Cf s o u r c e at l e a s t 1 m f r o m the ground. P l a c e the GM c o u n t e r at such a d i s t a n c e that the count ing ra te i s w e l l w i t h i n the a c c e p t a b l e l i m i t s ( n e g l i g i b l e p i l e - u p ) . P l a c e i n c r e a s i n g t h i c k n e s s of a l u m i n i u m (Al) f o i l around the s o u r c e . The f o i l should be in the f o r m of c o n c e n t r i c t u b e s hav ing a w a l l t h i c k n e s s 4 of l e s s than 0 . 1 m m . D e t e r m i n e (and plot) the count ing r a t e as a func t ion of w a l l t h i c k n e s s . Note the t h i c k n e s s r e q u i r e d beyond w h i c h t h e r e i s no m a r k e d change of count ing r a t e . At th i s point v i r t u a l l y a l l the ß-radiation i s a b s o r b e d . Conduct a l l e x p e r i m e n t s in C h a p t e r s 13 and 14 wi th the s o u r c e s h i e l d e d by the t h i c k n e s s of .A 1 d e t e r m i n e d by th i s e x p e r i m e n t .

Note: If the s o u r c e c a p s u l a t i o n or the w a l l of the GM c o u n t e r i s th ick no ß-radiation m a y be d e t e c t a b l e .

1 3 . 3 . INVERSE SQUARE LAW

T h i s e x p e r i m e n t should be conducted in a l a r g e r o o m or o u t - o f - d o o r s . P l a c e the 2 5 2Cf s o u r c e n e a r the c e n t r e of the r o o m h a l f w a y b e t w e e n

f l o o r und c e i l i n g . P l a c e the GM c o u n t e r at the s a m e he ight , and at a d i s t a n c e d. D e t e r m i n e the count ing ra te v e r s u s d, and plot th i s on l oga -r i t h m i c graph p a p e r . (On l o g a r i t h m i c paper both a x e s a r e l o g a r i t h m i c . ) O v e r at l e a s t part of the m e a s u r e m e n t range the l ine should have a s l o p e of - 2 . L e s s e r s l o p e s m a y be o b s e r v e d at v e r y shor t or v e r y long d i s t a n c e s .

The d e c r e a s e d s l o p e at s h o r t e r d i s t a n c e i s quite p o s s i b l y due to p i l e - u p ( i . e . a p u l s e rate w h i c h i s too rapid to b e . r e s o l v e d by the c o u n t e r or i t s c i r c u i t ) . T h i s m a y be v e r i f i e d by s h i e l d i n g the s o u r c e by a p p r o x i m a t e l y 1 inch of lead and noting the r e s u l t a n t d e c r e a s e at c l o s e a s w e l l a s at i n t e r m e d i a t e d i s t a n c e s . If p i l e - u p i s invo lved , the p e r c e n t a g e d e c r e a s e in count ing r a t e should be l e s s at the c l o s e r d i s t a n c e .

F l a t t e n i n g of the count ing ra te v e r s u s d i s t a n c e c u r v e at l a r g e d i s t a n c e s m a y w e l l be due to s c a t t e r i n g . V e r i f y th i s b y r e p e a t i n g the e x p e r i m e n t with both s o u r c e and counter at s m a l l e r d i s t a n c e s f r o m the ground, f ina l ly d e t e r m i n i n g

4 An alternative method is to roll up a long ribbon of foil around a peg that has a slightly larger diameter than the source. Partially withdraw the peg so that the source may be placed in the resulting cylindrical cavity. If the free end of the ribbon is weighted, rotation of the peg will gradually reduce the absorber thickness.

7 3

7 4 CHAPTER 10

the r e l a t i o n with both s o u r c e and counter p laced on the f l oor of the r o o m (or the ground if o u t - o f - d o o r s ) . Attempt to i n c r e a s e the e f f e c t s t i l l fur ther by r e p e a t i n g the e x p e r i m e n t with s o u r c e and counter p laced at the ins ide c o r n e r b e t w e e n the f l oor and the m a s o n r y w a l l . Max imum s c a t t e r i n g would be o b s e r v e d if both s o u r c e and counter w e r e m o v e d r e l a t i v e to e a c h o ther in a c y l i n d r i c a l hole ins ide a m a s s i v e block of m a t t e r . Th i s g e o m e t r y can be a c h i e v e d if in a v e r t i c a l a r r a n g e m e n t the s o u r c e i s p laced at the end of a c l o s e d tube i m m e r s e d cen tra l l y in a l a r g e tank of w a t e r with the counter l o w e r e d to v a r i o u s depths ins ide the tube.

1 3 . 4 . GAMMA-RAY TISSUE KERMA IN F R E E AIR

Cal ibrate the GM counter with a nucl ide emi t t ing hard g a m m a r a y s with a known e x p o s u r e rate constant . The e x p o s u r e rate constant i s the e x p o s u r e rate ( e . g . in R-h"1) f r o m 1 Ci of the nucl ide at a d i s tance of 1 m . F o r radium this constant i s 0 . 8 2 5 R - m 2 . Ci"1-h"l (or mR • m2- mCi"1-h"1) for an unshie lded s o u r c e in equ i l ibr ium with i t s daughter products . 5 En-c a p s u l a t i o n of the s o u r c e r e q i r e s a c o r r e c t i o n of about 6.5% per m i l l i m e t r e of P t - I r a l loy . T h i s ca l ibra t ion e s t a b l i s h e s the re la t ion between e x p o s u r e X and counts NQ. It may be a s s u m e d that the n u m e r i c a l va lue of the t i s s u e k e r m a i s 0 . 9 6 t i m e s that of the e x p o s u r e in r o e n t g e n s .

Thus if the counter i s e x p o s e d to g a m m a radiat ion f r o m another s o u r c e and Ns counts are r e g i s t e r e d the t i s s u e k e r m a i s

Ky = 0 . 9 6 rad r o e n t g e n

X r o e n t g e n s count N s I counts

Ut i l i z ing th i s ca l ibra t ion d e t e r m i n e the m a s s of 2 5 2Cf in a s o u r c e , u s i n g the i n f o r m a t i o n on g a m m a - r a y t i s s u e k e r m a rate in f r e e a ir as g iven in Appendix I.

Th i s ca l ibra t ion i s of l imi t ed v a l u e . One p o s s i b l e s o u r c e of e r r o r i s e n e r g y dependence of the counter which depends m a r k e d l y on the c o n s t r u c t i o n and c o m p o s i t i o n of the counter . If p o s s i b l e , th i s might be inves t iga ted u t i l i z ing nuc l ides emi t t ing g a m m a r a y s of v a r i o u s e n e r g i e s . Another fac tor which may be of i m p o r t a n c e i s uncerta inty r e g a r d i n g the t h i c k n e s s of encap-su la t ion of the 252Cf s o u r c e , as w e l l as that of the ca l ibrat ing s o u r c e . However , it i s unl ike ly that the e r r o r i n c u r r e d i s m o r e than 20%, and th i s kind of m e a s u r e m e n t should be adequate for e s t a b l i s h i n g the s o u r c e s trength for heal th p h y s i c s p u r p o s e s .

As w i l l be shown in Chapter 14, i on iza t ion c h a m b e r s are m o r e sui table for d e t e r m i n a t i o n of the s t rength of c a l i f o r n i u m s o u r c e s .

1 3 . 5 . ATTENUATION E X P E R I M E N T S

The d e t e r m i n a t i o n of absorpt ion c o e f f i c i e n t s of v a r i o u s m a t e r i a l s i s b e s t c a r r i e d out under condi t ions w h e r e the s o u r c e i s c o l l i m a t e d s o as to produce a narrow beam which i s b a r e l y l a r g e r than the d e t e c t o r . A second

5 Strictly speaking this number applies per gram rather than per curie of radium. However the d i f ference involved can be neglec ted .

EXPERIMENTS WITH A GM COUNTER 75

c o l l i m a t o r def ining e s s e n t i a l l y the s a m e beam i s p laced i m m e d i a t e l y in front of the d e t e c t o r and v a r i o u s t h i c k n e s s e s of the m a t e r i a l to be in-v e s t i g a t e d are p laced b e t w e e n the two c o l l i m a t o r s . The f i r s t c o l l i m a t o r e n s u r e s that as l i t t le m a t e r i a l as prac t i cab le i s i rrad ia ted , thus reduc ing the product ion of s e c o n d a r y rad ia t ions . The c o l l i m a t o r in front of the de tec tor ac t s p r i m a r i l y as a sh i e ld for such s e c o n d a r y rad ia t ions .

In the c a s e of 2 5 2 Cf, ra ther m a s s i v e c o l l i m a t o r s would be requ ired and the e x p e r i m e n t would not be par t i cu lar ly i n s t r u c t i v e b e c a u s e of the wide range of e n e r g i e s of the g a m m a e m i s s i o n s . If e x p e r i m e n t s a long t h e s e l i n e s are n e v e r t h e l e s s contemplated , it would be s i m p l e s t (but not bes t ) to a c c o m p l i s h the r e q u i r e d c o l l i m a t i o n by i n s e r t i n g both the s o u r c e and the GM counter into t h i c k - w a l l e d l ead tubes (wal l t h i c k n e s s of the o r d e r of 1 c m or m o r e ) .

T h e r e i s , h o w e v e r , an e x p e r i m e n t which i s i n s t r u c t i v e and y i e l d s in-f o r m a t i o n needed for l a t e r e x p e r i m e n t s . Unlike the attenuat ion e x p e r i m e n t d e s c r i b e d above , which i s a narrow beam e x p e r i m e n t , th i s i s a sh ie ld ing study which i s c a r r i e d out under broad b e a m cond i t ions .

Although the opt imum v e r s i o n of this e x p e r i m e n t i s one in which con-c e n t r i c c y l i n d r i c a l s h i e l d s are ut i l i zed , a s i m p l e r and quite adequate modi -f i c a t i o n i s one in which f lat s h e e t s are u s e d . T h e s e s h e e t s should be about 30 c m square , about \ cm thick in the c a s e of lead and about 2 c m thick in the c a s e of para f f in . To avoid damage to the s o u r c e a r e c e s s should be made in the c e n t r e of one of the s h e e t s into which the s o u r c e c a n be p laced and held with tape . The s o u r c e and the counter are p laced about a m e t r e apart and the counting rate d e t e r m i n e d as a funct ion of a b s o r b e r t h i c k n e s s . Th i s e x p e r i m e n t i s c a r r i e d out with a b s o r b e r s applied to the front only as w e l l as with m a t e r i a l applied in the back. It w i l l v e r y l ike ly be found that p l a c e m e n t of p a r a f f i n behind the s o u r c e (as s e e n f r o m the counter) w i l l in fact i n c r e a s e the counting r a t e . Th i s i s due to neutron capture by hydrogen and the e m i s s i o n of a 2. 2 - M e V g a m m a ray . A f t e r t h e s e s tud ie s are c o m p l e t e , two th in -wa l l ed cans ( such as f lat o i l cans) are f i l l ed with w a t e r and.the attenuation obtained i s c o m p a r e d with that of an equal t h i c k n e s s of paraf f in . The d i f f e r e n c e should be s l i g h t .

An i n c r e a s i n g concentra t ion of boron in the w a t e r cans should be produced by gradual ly adding s o m e boron-conta in ing sa l t (e . g. borax) . T h i s should r e s u l t in a gradual d e c r e a s e of the counting rate with a l e v e l l i n g out at boron c o n c e n t r a t i o n s of the o r d e r of 10 g / l i t r e . Note should be taken of the fact that boron when p r e s e n t in e i t h e r tank r e d u c e s the count r a t e .

If a l i th ium ra ther than a boron salt i s u s e d , the reduct ion in count rate should be e v e n g r e a t e r b e c a u s e the (n, a) r e a c t i o n with 6 Li , unlike that with MB , r e s u l t s in neg l ig ib l e s e c o n d a r y g a m m a e m i s s i o n (the exc i t ed ''Li nuc leus produced in the boron r e a c t i o n e m i t s a 0 . 4 8 - M e V g a m m a photon). How-e v e r , l i thium s a l t s m a y not be read i ly procurable and in addition, s o m e of the c o m m e r c i a l l y avai lable l i thium i s depleted in 6 Li .

C h a p t e r 14

E X P E R I M E N T S WITH LARGE IONIZATION CHAMBERS

14. 1. INSTRUMENTATION REQUIREMENTS

The e x p e r i m e n t s d e s c r i b e d in th i s chapter r e q u i r e two c h a m b e r s , e a c h having a v o l u m e of the o r d e r of 1 l i t r e or m o r e . The ion izat ion current in t h e s e c h a m b e r s m u s t be m e a s u r e d with an appropr iate e l e c t r o m e t e r . C a r e should be e x e r c i s e d to avoid p a r a s i t i c ionizat ion , i. e. ion izat ion o c c u r r i n g in gas v o l u m e s o ther than those of the ion iza t ion c h a m b e r s .

Under o p t i m u m condi t ions the ion izat ion c h a m b e r s have a shape s i m i l a r to that shown in F i g . 1 with one chamber be ing m a d e of t i s s u e - e q u i v a l e n t p l a s t i c n o r m a l l y f i l l ed with a t i s s u e - e q u i v a l e n t gas m i x t u r e , and the other chamber c o n s t r u c t e d of a lumin ium and f i l l ed with argon. However , for the p u r p o s e s of the e x p e r i m e n t s d i s c u s s e d h e r e s i m p l e r t y p e s would s u f f i c e . C h a m b e r s of the type shown in F i g . 30 are r e l a t i v e l y e a s y to c o n s t r u c t and they can be m a d e s u f f i c i e n t l y g a s - t i g h t by applying s i m p l e s e a l i n g compounds (par t i cu lar ly v a r i o u s k inds of wax). The m a j o r c r i t i c a l component of such a c h a m b e r i s the insu la tor , which can be c o n s t r u c t e d of Luc i te , p o l y s t y r e n e , po lye thy lene and v a r i o u s ha logenated d e r i v a t i v e s of p o l y -e thy lene , such as t e t ra f luoropo lye thy l ene or v a r i o u s ch lor inated v e r s i o n s . I n s u l a t o r s m u s t be kept s c r u p u l o u s l y c lean fo l l owing a thorough wash ing with soap so lu t ion and r e p e a t e d r i n s i n g by d i s t i l l e d w a t e r , and they should n e v e r be touched by hand on c r i t i c a l s u r f a c e s (which in the d e s i g n shown in F i g . 30 are the curved s u r f a c e s ) . One s u c h c h a m b e r should be c o n -s t ruc ted of a lumin ium and the other of a hydrogenous p l a s t i c . The b e s t s e l e c t i o n for th i s i s t i s s u e - e q u i v a l e n t p la s t i c which in addit ion to having the c o r r e c t hydrogen content i s a l s o conducting. If th is i s not ava i lab le other hydrogenous p l a s t i c s , s u c h a s po lye thy lene , p o l y s t y r e n e of methy l m e t h a c r y l a t e (Luci te or P e r s p e x ) can be u s e d . It w i l l be n e c e s s a r y in this c a s e to paint the ins ide s u r f a c e s of such a c h a m b e r with a thin l a y e r of co l lo ida l graphite (Aquadag) which should be v i g o r o u s l y rubbed with paper to i m p r o v e conduct iv i ty . Al l s u r f a c e s m u s t be e l e c t r i c a l l y in terconnec ted and contact m u s t be e s t a b l i s h e d to the outs ide , p r e f e r a b l y by m e a n s of a thin pin which p e n e t r a t e s one of the p l a s t i c w a l l s and i s s e c u r e d by wax.

It should be p o s s i b l e to f i l l at l e a s t the hydrogenous c h a m b e r with a v a r i e t y of g a s e s , and it i s v e r y d e s i r a b l e that the c h a m b e r s be s trong enough to p e r m i t evacuat ion . If the la t ter r e q u i r e m e n t i s not met , the gas a t m o s p h e r e in the c h a m b e r can s t i l l be m o d i f i e d by r e p e a t e d flushing, but cer ta in e x p e r i m e n t s d e s c r i b e d be low wi l l not be p o s s i b l e .

In al l i on izat ion c h a m b e r m e a s u r e m e n t s note should be taken of the t e m p e r a t u r e T and the p r e s s u r e P , and r e a d i n g s should be c o r r e c t e d to s tandard t e m p e r a t u r e and p r e s s u r e by mul t ip l i ca t ion of the fac tor

7 60 T 273 P

Here T i s the abso lute t e m p e r a t u r e in d e g r e e s Ke lv in and P i s the p r e s s u r e in torr .

7 7

7 8 CHAPTER 10

» 2 - .

C - .

Aluminium

Insu lo tor t

A lumin ium or

Hydrogenous Plast ic

FIG.30. Diagram of a s imple ionization chamber suitable for experiments with cal i fornium-252. The chamber, envelope is made of a luminium and in order to withstand evacuat ion the end plates (Px and P2 ) should be about 7. 5 mm thick and the cylindrical portion T should be about 3 mm thick. The cr i t ical insulators (Tx and T 2 ) and the collect ing electrode C are secured by a single screw B, which also serves as conductor. The liner L made of thin 1 mm) mater ia l is a luminium for the neutron insensitive chamber and T. E. plastic or another hydrogenous plastic for the T. E. or hydrogenous chamber . It is supported by rings (Rj and Rj) which may be made of any insulator. Contact is established through the insulated connector V. Gases can be admitted or removed through tubes G1 and G 2 .

The chamber should be sealed with wax that melts at moderate tempera ture 50°C). Especial care should be taken when the wax is applied to chambers (W) cut in insulator I 2 . The wax should not form a bridge across the h igh- impedance insulation.

If L is not made from e lec t r ica l ly conducting mater ia l its inside surfaces must be coated with a thin layer of conducting graphite (Aquadag) and contact must be established to V. This may be accomplished by a continuous coating of the inside and the outside of the pieces (2 plates plus cylinder) that constitute L.

The e lec t rometer is connected between the electrostat ic shield (S) and B. The collecting potential is applied between V and the chamber envelope.

EXPERIMENTS WITH LARGE IONIZATION CHAMBERS 7 9

The saturat ion c h a r a c t e r i s t i c s of the c h a m b e r should be d e t e r m i n e d by o b s e r v i n g the ion izat ion current I a s a funct ion of the applied c o l l e c t i n g vo l tage V. The c o r r e c t va lue of the ionizat ion current should be obtained by ex trapo la t ion of a plot in which l / l i s plotted aga ins t 1 /V. Saturat ion wi l l r e q u i r e h igher v o l t a g e s with neutrons as c o m p a r e d with -y-radiation; it a l s o w i l l r e q u i r e higher v o l t a g e s at higher gas p r e s s u r e s .

T h e s e m e a s u r e m e n t s should be made with both p o l a r i t i e s of the c o l l e c t -ing vo l tage . If s u c h m e a s u r e m e n t s are p e r f o r m e d , it should be kept in mind that, i m m e d i a t e l y a f ter r e v e r s a l of the c o l l e c t i n g potent ial , d i s p l a c e -ment c u r r e n t s m a y o c c u r in i n s u l a t o r s b e c a u s e of changes in po lar iza t ion . T h e s e should be o b s e r v e d in the absence of rad ia t ion s o u r c e s .

14. 2. NEUTRON INSENSITIVE CHAMBER

The a l u m i n i u m c h a m b e r u s e d in these e x p e r i m e n t s i s b e s t f i l l e d with argon but CO2 i s a c c e p t a b l e a s we l l . Air , which i s of c o u r s e the e a s i e s t gas to1 u s e , i s s o m e w h a t i n f e r i o r b e c a u s e of the t h e r m a l neutron r e s p o n s e of n i t rogen (the 1 4N(n, p) 1 4C r e a c t i o n which y i e l d s a proton of about 0. 7 - M e V energy) . This r e s u l t s in a no t i ceab le neutron s e n s i t i v i t y when dea l ing with heav i ly m o d e r a t e d 2 5 2Cf s o u r c e s .

E x p e r i m e n t s to be p e r f o r m e d with this chamber a r e e s s e n t i a l l y the s a m e as t h o s e d e s c r i b e d in S e c t i o n s 13 .4 and 13. 5. The chamber should be ca l ibrated by a s o u r c e e m i t t i n g g a m m a radia t ion which i s at l e a s t predominant ly m o r e e n e r g e t i c than about 0. 5 MeV. Beyond th i s e n e r g y and the e n e r g y of the m o s t e n e r g e t i c g a m m a radia t ion e m i t t e d by the ca l i forn ium s o u r c e the c h a m b e r r e s p o n s e should be substant ia l ly e n e r g y -independent. The c o r r e c t i o n fac tor 0. 96 should again be applied.

B e c a u s e of the e n e r g y independence of the c h a m b e r this d e t e r m i n a t i o n of the m a s s of the 2 5 2Cf inner s o u r c e wi l l be m o r e r e l i a b l e and the only r e m a i n i n g uncerta inty m a y be that re la t ing to s o u r c e encapsulat ion . H o w e v e r , it i s quite un l ike ly that the combined e r r o r s r e l a t i n g to encapsula t ion of the 252Cf and ca l ibrat ion s o u r c e s e x c e e d 20% and the de terminat ion of the 2 5 2Cf m a s s should in m a n y c a s e s be accurate to ± 10%.

The r e s u l t s obtained in the attenuation e x p e r i m e n t , when c a r r i e d out with the ion izat ion c h a m b e r , should be substant ia l ly the s a m e as those o b s e r v e d with the GM counter .

14. 3. E X P E R I M E N T S WITH THE HYDROGENOUS CHAMBER

The g a m m a - r a y s e n s i t i v i t y of th i s chamber i s again e s t a b l i s h e d by ca l ibrat ion as in the c a s e of the non-hydrogenous chamber and the GM counter , and the s a m e c o r r e c t i o n f a c t o r i s applied.

If the c h a m b e r i s t i s s u e equivalent (a r e q u i r e m e n t which for the p u r p o s e s of t h e s e e x p e r i m e n t s i s adequate ly m e t if both wal l and gas contain 10% hydrogen by m a s s ) the neutron s e n s i t i v i t y of the c h a m b e r w i l l be equal to the g a m m a s e n s i t i v i t y except for a W c o r r e c t i o n : the W for neutron s e c o n d a r i e s i s a p p r o x i m a t e l y 4% l a r g e r than that for the s e c o n d a r y e l e c t r o n s produced by g a m m a r a y s . This c o r r e c t i o n i s in the oppos i te d i r e c t i o n f r o m that for the r e l a t i o n between e x p o s u r e and t i s s u e k e r m a for g a m m a radiat ion and as a r e s u l t the read ing o f the c h a m b e r i s within

80 CHAPTER 18

about 1% the s a m e when it is exposed to x r o e n t g e n s of g a m m a r a y s or x r a d s of neu t rons . However , the c o r r e c t i o n f ac to r 0. 96 m u s t be appl ied to the g a m m a component of the r ad i a t ion . If RA1 and R T E r e p r e s e n t the r e a d i n g of the a lumin ium and T. E. c h a m b e r s in mu l t i p l e s of the r e a d i n g o b s e r v e d p e r roen tgen of h a r d g a m m a rad ia t ion , and if t h e s e c h a m b e r s a r e exposed in a mixed f ie ld of g a m m a r a y s in n e u t r o n s , t he i r r e a d i n g s wil l thus be as fol lows:

R A 1 = 1 . 0 4 K y

R T E = 1 . 0 4 K y + K N

w h e r e Ky and K N a r e the t i s s u e k e r m a in r a d s due to g a m m a r a y s and n e u t r o n s r e s p e c t i v e l y .

It fo l lows tha t

K y = 0 . 9 6 RA1

K N ~ R T E " R A I

Cal ib ra t ion of a 252 C F s o u r c e with a p a i r of ioniza t ion c h a m b e r s us ing

the above equat ions i s s u p e r i o r to the me thods ment ioned thus f a r . In p a r t i c u l a r K N i s v i r t ua l l y una f fec ted by s o u r c e encapsu la t ion because the P t — Ir al loy has negl ig ib le neu t ron a t tenuat ion in the t h i c k n e s s e s employed. If the hydrogenous c h a m b e r has a hydrogen content o the r than 10%, i t s neu t ron sens i t i v i ty m a y , to a good approx imat ion , be c o n s i d e r e d to be p r o p o r t i o n a l to i ts r e l a t i v e hydrogen content . Thus if the c h a m b e r wall i s polyethylene i ts compos i t ion i s b a s i c a l l y (CH2) , c o r r e s p o n d i n g to a hydrogen content of 14. 3%. If such a c h a m b e r i s u s e d with an a lumin ium c h a m b e r

a s s u m i n g that the c h a m b e r i s homogeneous , i . e. is f i l led with a gas that a l so has the compos i t ion CH2 (such as e thylene) .

On the o the r hand, the a tomic compos i t ion of p o l y s t y r e n e is (CH), c o r r e s p o n d i n g to a hydrogen content of 7. 7%. In th is c a s e

K n = ( R C H - R a i > / ° - 7 7

and a su i tab le gas i s ace ty lene . The i m p o r t a n c e of c h a m b e r homogenei ty can be d e m o n s t r a t e d in an

i n s t r u c t i v e e x p e r i m e n t that should be c a r r i e d out if the hydrogenous c h a m b e r has su f f i c i en t m e c h a n i c a l s t r e n g t h to wi ths tand evacuat ion . As s ta ted in Sect ion 3. 3 t h e r e is exac t compensa t ion be tween p ro tons o r ig ina t ing in the wal l and expending p a r t o r a l l of t h e i r ene rgy in the gas (wall contr ibut ion) and p ro tons tha t o r ig ina te in the gas (gas contr ibut ion) . Th i s compensa t ion does not occur if the gas contains (on a p e r c e n t by weight bas i s ) m o r e o r l e s s hydrogen than does the wal l . F o r c h a m b e r s of the d imens ions u n d e r d i s cus s ion , and f o r 252Cf n e u t r o n s , the gas cont r ibut ion is quite impor t an t . Consequent ly , if the gas h a s the wrong hydrogen content an e r r o r i s i n c u r r e d tha t i n c r e a s e s a s the gas p r e s s u r e i s i n c r e a s e d .

EXPERIMENTS WITH LARGE IONIZATION CHAMBERS 8 1

In the e x p e r i m e n t des igned to d e m o n s t r a t e th is e f fec t the c a l i f o r n i u m s o u r c e should be sh ie lded by about 2 in. of lead in o r d e r to r e d u c e the g a m m a cont r ibut ion . It should be p laced c lose to the c h a m b e r to e n s u r e subs t an t i a l r e a d i n g s . If the r e a d i n g of a homogeneous c h a m b e r is plot ted v e r s u s gas p r e s s u r e , a s t r a i g h t l ine should r e s u l t . However , if the c h a m b e r i s f i l led with C 0 2 o r with a i r , a curved l ine should r e s u l t in which the s lope d e c r e a s e s with i n c r e a s i n g p r e s s u r e of the gas . C o n v e r s e l y , if one f i l l s the c h a m b e r with a gas having a v e r y high hydrogen content (e. g. me thane which has a 25% hydrogen content) the r e v e r s e c u r v a t u r e wil l be o b s e r v e d . It should be noted that the s lope of the cu rve n e a r the o r ig in i s p r o p o r t i o n a l to the p roduc t SW (see Sec t ions 2. 6 and 3. 3).

C h a p t e r 15

ADVANCED E X P E R I M E N T S IN DOSIMETRY AND MICRODOSIMETRY

15. 1. GENERAL REMARKS

The e x p e r i m e n t s d e s c r i b e d in th i s s ec t ion r e q u i r e soph i s t i ca t ed dev ices . Only expe r i enced i n v e s t i g a t o r s should a t t empt to c o n s t r u c t them. F a m i l i a r i t y with the l i t e r a t u r e r e f e r e n c e s i s e s s e n t i a l and only the ma in f e a t u r e s of the e x p e r i m e n t a l p r o c e d u r e wil l be given.

15. 2. DOSE DISTRIBUTION IN A TISSUE-EQUIVALENT MEDIUM

B e c a u s e of the s t r o n g a t tenuat ion of the r ad i a t i on (pa r t i cu l a r l y that of neu t rons ) dose m e a s u r e m e n t s in t i s sue -equ iva l en t m a t e r i a l m u s t be c a r r i e d out within d i s t a n c e s that a r e of the o r d e r of c e n t i m e t r e s . This r e q u i r e s the e m p l o y m e n t of a v e r y s m a l l c h a m b e r of the type shown in F ig . 4. When us ing mul t ip l i ca t ion c h a m b e r s , it i s e s s e n t i a l that t h e r e be a s t eady flow of gas at an a c c u r a t e l y con t ro l led p r e s s u r e and that the a c c e l e r a t i n g vo l tage , which is of the o r d e r of 1000 V, be s t ab i l i zed to 0. 1% o r b e t t e r . Mechan ica l m e a n s m u s t be p rov ided to move e i t he r the c h a m b e r o r the c a l i f o r n i u m s o u r c e with pos i t ioning a c c u r a c y of the o r d e r of m i l l i m e t r e s .

If th i s a p p a r a t u s is ava i lab le , i sodose c u r v e s for both neu t rons and g a m m a r a d i a t i o n s can be d e t e r m i n e d .

15. 3. MICRODOSIMETRY AND THE DETERMINATION O F L E T DISTRIBUTIONS

T h e s e e x p e r i m e n t s r e q u i r e a s p h e r i c a l t i s s u e - e q u i v a l e n t p ropo r t i ona l counter with p r e - a m p l i f i e r , a m p l i f i e r and m u l t i - c h a n n e l a n a l y s e r as wel l as a w e l l - r e g u l a t e d gas flow s y s t e m . In addit ion s o m e kind of c a l i b r a t i ng s o u r c e m u s t be p rov ided . The s i m p l e s t i s a co l l ima ted s o u r c e of a - r a d i a t i o n .

In the s p e c t r u m o b s e r v e d , p u l s e s due to e l e c t r o n s and p ro tons a r e well s e p a r a t e d and a s m a l l cont r ibut ion f r o m h e a v i e r r e c o i l i n g nuc le i should be v i s ib le as wel l . F i g u r e 6 i s a s e m i l o g a r i t h m i c r e p r e s e n t a t i o n of the l o g a r i t h m of l i nea l ene rgy v e r s u s the p roduc t of y • d(y). The l a t t e r is obta ined by mul t ip l i ca t ion of the f r equency d i s t r ibu t ion f(y) by y 2 . B e c a u s e of d i f f e r e n c e s in s t a t i s t i c a l f luc tua t ion and f in i te r a n g e s the s p e c t r a depend somewha t on the magni tude of the s t r u c t u r e that is s imu la t ed .

Ut i l iz ing a p r o c e d u r e d e s c r i b e d in Attix et al. (1968), one m a y d e t e r -mine the L E T d i s t r ibu t ion of dose fo r the high L E T por t ion of the c a l i f o r n i u m s p e c t r u m . Th i s i s given in F ig . 5.

83

C h a p t e r 16

E X P E R I M E N T S WITH TRADE SC ANTIA

16. 1. THE SYSTEM DESCRIBED

T r a d e s c a n t i a pa ludosa was f i r s t u sed by Sax b e f o r e the Second World War to s tudy induced c h r o m o s o m e a b e r r a t i o n s , and s ince that t i m e n u m e r o u s r ad iob io log ica l e x p e r i m e n t e r s have ut i l ized s p e c i e s o r v a r i e t i e s of T r a d e s c a n t i a . Many t i s s u e s o r o r g a n s of th is plant may be u s e d e f f i c ien t ly in v a r i o u s e x p e r i m e n t a l p r o c e d u r e s , including g r o s s l e tha l i ty , changes in weight or' he ight of the plant , inhibit ion of f lower produc t ion , r educ t i on in the n u m b e r of s t a m e n h a i r s , and c h r o m o s o m e b r e a k a g e . In many ways the m o s t u s e f u l end-poin t , however , i s the s c o r i n g of a b e r r a t i o n s which r e s u l t in changes of co lour in the s t a m e n h a i r s .

T r a d e s c a n t i a clone 02 is a diploid plant (2n = 12), h e t e r o z y g o u s f o r f lower co lour , which is thought to be a r e s u l t of a n a t u r a l c r o s s be tween blue and pink f l o w e r e d f o r m s or spec i e s . It i s a p e r e n n i a l h e r b a c e o u s plant with n a r r o w t a p e r i n g l e a v e s , not unlike s o m e of the m e d i u m s i z e g r a s s e s , and it r e a c h e s a height of about 2 f t (see F i g . 31).

Techn iques for the s u c c e s s f u l and ef f ic ien t r ad iob io log ica l u s e of the s t a m e n h a i r s y s t e m have been evolved dur ing the pas t two decades . I ts f i r s t ex tens ive u s e as a t e s t s y s t e m fo r muta t ion induction was in the Biosa te l l i t e p r o g r a m m e . It ha s subsequent ly been used for R B E e x p e r i -m e n t s with m o n o e n e r g e t i c n e u t r o n s , and m o s t r e c e n t l y for e x p e r i m e n t s involving h igh-energy n i t rogen ion b e a m s (Sparrow, Unde rb r ink and Ross i , 1972; Underb r ink , S cha i r er and Spa r row, 1973).

The s t a m e n h a i r s cons i s t of long f i l a m e n t s of cont iguous s ingle ce l l s , in which ioniz ing r a d i a t i o n can induce s e v e r a l t ypes of m u t a t i o n s or a b e r r a t i o n s (see F ig . 32). Some a b e r r a t i o n s , such a s the change f r o m the wild type blue colour to pink in clone 02, a r e c o n s i d e r e d to be t r u e soma t i c muta t ions in the c l a s s i c a l s e n s e , s ince the plant i s h e t e r o z y g o u s f o r blue o r pink f lower colour .

The s t a m e n h a i r s y s t e m has a n u m b e r of ou ts tanding advan tages for r ad iob io log ica l work:

(a) S tamen h a i r s a p p e a r to be p roduced l a r g e l y by r e p e a t e d d iv is ions of the t e r m i n a l or s u b t e r m i n a l ce l l s , with only a few i n t e r s t i t i a l d iv i s ions . At c e r t a i n deve lopmenta l s t a g e s , they appea r to have e s s e n t i a l l y a s ing le o r two cel l m e r i s t e m which f a v o u r s the study of l o s s of r e p r o d u c t i v e in tegr i ty a f t e r i r r a d i a t i o n .

(b) Individual i n f l o r e s c e n c e s contain a compac t c l u s t e r of s m a l l f l ower buds so that u n i f o r m i r r a d i a t i o n is p o s s i b l e even with s m a l l r ad i a t i on b e a m s .

(c) L a r g e ce l l popula t ions with we l l -de f ined ce l l l i neages can be ana lysed with r e g a r d to the locat ion of spec i f i c k inds of a b e r r a t i o n s .

85

FIG.31. Mature stock plant of Tradescantia c lone 02, showing its characteristic growth habit. (Picture by courtesy of A. Underbrink).

EXPERIMENTS WITH TRADESCANTIA 8 7

HAIRLESS

FIG.32. Typical stamen with associated stamen hairs as they appear when properly prepared for scoring, (Picture by courtesy of A . Underbrink).

EXPERIMENTS WITH TRADESCANTIA 8 9

(d) The handl ing of the m a t e r i a l and the s c o r i n g of muta t ions i s r e a d i l y l e a r n e d and eas i ly p e r f o r m e d . In addit ion a r e l a t i v e l y s h o r t e x p e r i m e n t a l pe r iod i s r e q u i r e d .

(e) T r a d e s c a n t i a clone 02 has a high r a d i o s e n s i t i v i t y . A s igni f ican t y ie ld of mu ta t ions r e s u l t s f r o m s ingle acute doses as low as 10 m r a d of 430-keV n e u t r o n s and 250 m r a d of X - r a y s .

(f ) Genet ic va r i ab i l i t y i s m i n i m a l s ince the p lan ts a r e usua l ly p r o p a -gated asexua l ly by cut t ings .

The me thods u s e d for the cul t ivat ion of the p lan t s , and t he i r p r e p a r a t i o n fo r e x p e r i m e n t s , ha s been d e s c r i b e d in g r e a t de ta i l by Unde rb r ink , S c h a i r e r and S p a r r o w (1974).

16. 2. LOW DOSE-RATE E X P E R I M E N T WITH CALIFORNIUM-252 AND/OR RADIUM USING TRADESCANTIA

F o r a typ ica l e x p e r i m e n t , app rox ima te ly 50 cut t ings m u s t be p r e p a r e d . T h e s e should p r e f e r a b l y be roo ted for a low d o s e - r a t e e x p e r i m e n t , s i n c e long e x p o s u r e t i m e s a r e needed. A l t e rna t ive ly , the cut t ings may be m a i n t a i n e d in Hoagland ' s so lu t ion dur ing the pe r iod of i r r a d i a t i o n . Rooted and un roo t ed cut t ings a r e i l l u s t r a t e d in F ig . 33.

A s m a l l i r r a d i a t i o n f i x t u r e m u s t be c o n s t r u c t e d of a p l a s t i c m a t e r i a l such as Luci te . About 10 i n f l o r e s c e n c e s can be a c c o m m o d a t e d in a s p a c e 2. 5 cm by 2. 5 cm by 1. 25 cm. A volume of t h e s e d imens ions can be i r r a d i a t e d u n i f o r m l y by m e a n s of two p lana r a r r a y s of r ad ioac t i ve s o u r c e s , p a r a l l e l to one another and spaced by 1. 25 cm. Such an a r r a n g e m e n t can be p roduced with as few a s 10 r a d i o a c t i v e s o u r c e s . One f ix tu re should be c o n s t r u c t e d for u s e with c a l i f o r n i u m - 2 5 2 and an iden t i ca l s e t - u p des igned f o r u s e with a g a m m a e m i t t e r such a s r a d i u m or coba l t -60 .

Approx ima te ly 10 i n f l o r e s c e n c e s a r e i r r a d i a t e d with c a l i f o r n i u m - 2 5 2 , Ein equal n u m b e r i r r a d i a t e d with g a m m a r a y s , and a s i m i l a r n u m b e r r e s e r v e d fo r c o n t r o l s . In any one given e x p e r i m e n t , a l im i t ed n u m b e r of exposu re t i m e s should be used , s ince it i s adv isab le to u s e as many s a m p l e s as pos s ib l e f o r each dose . A r a n g e of to ta l doses can be obtained by m e a n s of r e p l i c a t e e x p e r i m e n t s . F o r c a l i f o r n i u m - 2 5 2 rad ia t ion , doses within the r a n g e 1 to 50 r a d give s c o r e a b l e n u m b e r s of muta t ions , while fo r the p a r a l l e l g a m m a - r a y e x p e r i m e n t s doses of 10 to 100 r a d a r e a p p r o p r i a t e .

A f t e r the i r r a d i a t i o n e x p o s u r e , the cut t ings a r e ma in ta ined in Hoagland ' s solut ion gently a e r a t e d in a growth c h a m b e r under c a r e f u l l y con t ro l l ed condi t ions . F r o m the 7th to the 18th day p o s t - i r r a d i a t i o n , each f l ower that b l o o m s i s p icked, and the s t a m e n h a i r s a r e r e m o v e d and mounted in p a r a f f i n oi l , so tha t they may be examined with a d i s s e c t i n g m i c r o s c o p e . The de ta i l ed m e t h o d s have been d e s c r i b e d by Underb r ink , S c h a i r e r and Spa r row (1974).

F o r each r ad ia t ion dose , the t o t a l n u m b e r of mutant events , divided by the to ta l n u m b e r of s t a m e n h a i r s s c o r e d dur ing the p o s t - i r r a d i a t i o n pe r iod , i s ave raged . The mutan t event f r e q u e n c y o b s e r v e d in con t ro l s , which should be v e r y low indeed, is s u b t r a c t e d f r o m the va lues obta ined in i r r a d i a t e d s a m p l e s . The data a r e then p r e s e n t e d on a double l o g a r i t h m i c plot, with pink mutan t even t s p e r h a i r plot ted aga ins t the dose in r a d s .

9 0 CHAPTER 16

FIG.34. Neutron and X-ray dose-response curves for p ink-mutan t events in s tamen hairs of Tradescantia c lone 02. The points represent average values obtained by dividing the total number of mutant events by the total number of s tamen hairs scored from day 11 to day 15 af ter irradiation, when the mutat ion frequency is highest. The horizontal l ine represents the mutant -event frequency in control experiments . (Taken from Sparrow, Underbrink and Rossi, 1972).

F r o m t h e s e data it wil l be p o s s i b l e to ca lcu la te the r e l a t i v e b iological e f f e c t i v e n e s s fo r th i s s o m a t i c mu ta t ion of c a l i f o r n i u m - 2 5 2 rad ia t ion c o m p a r e d with r a d i u m g a m m a r a y s at low d o s e - r a t e .

Data for T r a d e s c a n t i a exposed to n e u t r o n s and g a m m a r a y s at low d o s e - r a t e have not as yet a p p e a r e d in the l i t e r a t u r e . F i g u r e 34 i l l u s t r a t e s r e s u l t s that have been obta ined fo r acute e x p o s u r e s to 430-keV neu t rons and 250-kV X - r a y s . In t h e s e e x p e r i m e n t s a v e r y wide r a n g e of doses was u sed , extending ove r a r a n g e that wil l not be p r a c t i c a b l e at low dose r a t e s . However , F ig . 34 gives s o m e indicat ion of the r ep roduc ib i l i t y of the r e s u l t s that can be obta ined with th is b io logica l s y s t e m , and the way in which they should be p r e s e n t e d .

C h a p t e r 17

E X P E R I M E N T S WITH ROOT M E R I S T E M S

1 7 . 1 . THE CHOICE OF THE SYSTEM

The e f f ec t of ionizing r ad i a t i on on seed l ings of Vicia faba was f i r s t r e p o r t e d by M o t t r a m (1913). J u s t be fo re the Second World W a r , Gray and Read began a deta i led inves t iga t ion of the g r o s s e f f e c t s shown by r o o t s a f t e r i r r a d i a t i o n , namely r educ t i on or c e s s a t i o n of g rowth . Using these c r i t e r i a to a s s e s s r ad i a t i on damage , they p e r f o r m e d t h e i r c l a s s i c a l expe r i -m e n t s to d e m o n s t r a t e the inf luence of oxygen on the r e s p o n s e of l iving ce l l s to ioniz ing r a d i a t i o n . They a l so inves t iga ted the r e l a t i ve e f f i c iency of d i f f e ren t types of r ad i a t i on in producing b io logica l damage (Gray und Read, 1942 a - d ; Gray and Scholes 1951; Read, 1952 a, b; Read, 1959.)

Fol lowing the work of t h e s e p ionee r s , a n u m b e r of r ad iob io log i s t s have s ince chosen the bean roo t as a t e s t ob j ec t . Obse rva t i on of g r o s s damage has been pa r a l l e l ed by a s tudy of the h is to logy and cytology of the root t ip, so that the abil i ty of r ad ia t ion to p roduce mi to t i c delay and c h r o m o s o m e a b e r r a t i o n s has a l so been inves t iga ted in de t a i l . T h e r e a r e a n u m b e r of r e a s o n s why so many people have e l ec t ed to work with th is b io logica l m a t e r i a l .

(a) The c h e a p n e s s of the m a t e r i a l and equipment needed, t o g e t h e r with the s impl i c i ty of the t ech iques , a r e obvious a t t r a c t i o n s .

(b) Without the co l l abora t ion of o ther w o r k e r s , bean r o o t s can be used by phys i c i s t s who have no f o r m a l t r a in ing in biology.

(c) The m e r i s t e m a t i c , d i f f e ren t i a t ing and ful ly d i f f e r en t i a t i ng ce l l s a r e s e p a r a t e d into d i f f e r en t zones in the roo t , mak ing it pos s ib l e f o r each to be i r r a d i a t e d s e p a r a t e l y .

(d) Although the r ad ia t ion biology of the bean root has a f a sc ina t ion of i t s own, the r e a l i n t e r e s t l ies in the fac t that the g e n e r a l r e p o n s e c lose ly p a r a l l e l s that of o ther s i m p l e o r g a n i s m s and of human t i s s u e s .

(e) The t echn iques used allow a suf f ic ien t n u m b e r of individual seed l ings ( s e v e r a l hundreds) to be used in each e x p e r i m e n t to r e d u c e r andom e r r o r s to an acceptab le leve l .

(f) Any s u f f e r i n g o r d i s c o m f o r t to e x p e r i m e n t a l a n i m a l s , so of ten n e c e s s a r y in i r r a d i a t i o n e x p e r i m e n t s , is avoided.

(g) It i s f e a s ib l e to i r r a d i a t e bean roo t s u n d e r condi t ions that a r e v i r tua l ly i m p o s s i b l e with e x p e r i m e n t a l an ima l s ; f o r example , high p r e s s u r e s of v a r i o u s g a s e s of the o r d e r of 100 atm may be used , o r pro longed pe r i ods of hypoxia may be imposed upon bean roo t s , which would not be t o l e r a t e d by whole a n i m a l s o r even m a m m a l i a n ce l l s in c u l t u r e .

(h) The ce l l s of the root m e r i s t e m s conta in only 6 p a i r s of r e l a t ive ly l a r g e c h r o m o s o m e s , which a r e v e r y convenient fo r cy to logica l s tudy.

91

9 2 CHAPTER 18

FIG. 35. Pattern of root growth of Vicia seedlings exposed to radiation. There is l i t t le , if any, reduction in growth for the first day, but subsequently the dai ly growth-rate falls to a min imum 5 to 7 days post- irradiat ion. If the dose has not been too large, the growth-rate later recovers. The two most commonly used scores of radiation damage are: (a) growth in 10 days, i . e . the area under the curve up to the 10th day expressed as a fraction of controls; (b) the minimum growth-rate , likewise expressed as a fraction of controls.

E x p e r i m e n t s involving the rad iobio logy of Vicia faba fa l l into t h r e e d is t inct g r o u p s . F i r s t , the o b s e r v a t i o n of the g r o s s e f f e c t s of r ad ia t ion on root g rowth . The reduc t ion o r c e s s a t i o n of root growth has been used as an ind ica to r to c o m p a r e the b iological e f f e c t s of d i f f e r en t types of r ad ia t ion o r t r e a t m e n t r e g i m e s . Second, the m i c r o s c o p i c study of the n o r m a l i r r a d i a t e d root t ip at the c e l l u l a r l eve l . F r o m t h e s e s tud ies an a t tempt has been made to inves t iga te the m e c h a n i s m of r ad ia t ion damage , and the p r i m a r y even t s which r e s u l t in the growth inhibi t ion of roo t growth have been o b s e r v e d . Th i rd , the study of ap ica l m e r i s t e m s , including that of the b r o a d bean, u s ing ionizing r ad i a t i on to d i s t u r b the n o r m a l p a t t e r n of behav iour , and by doing so shed light on the s t r u c t u r e and m e t a b o l i s m of the apex .

While the b road bean root , Vicia faba , has been the m o s t exhaus t ive ly studied roo t m e r i s t e m , s e r i e s of e x p e r i m e n t s have been p e r f o r m e d a l so with b a r l e y ( E b e r t and B a r b e r , 1961) and a l s o with c o r n seed , Z e a m a y s (Her ing and Shepstone, 19 73). The p r e f e r e n c e shown f o r Vicia faba is due p a r t l y to the s m a l l n u m b e r of d i s t inc t ive c h r o m o s o m e s which th is spec i e s con ta ins , but mos t ly it i s a qu i rk of h i s t o ry r e su l t i ng f r o m the fact that much of the e a r l y r ad iob io log ica l work was p e r f o r m e d in Nor th-w e s t e r n E u r o p e , w h e r e broad beans a r e a c o m m o n vege tab le . *

It i s c e r t a i n that in a lmos t e v e r y count ry of the wor ld t h e r e a r e o t h e r seed l ings , of which the au tho r s a r e u n a w a r e , that would be equal ly su i tab le for r ad iob io log ica l inves t iga t ions of the type to be d e s c r i b e d in th i s c h a p t e r .

EXPERIMENTS WITH ROOT MERISTEMS 93

R e a d e r s of th i s manual , l iving in d i f f e ren t c o u n t r i e s of the wor ld , a r e u r g e d to seek seed l ing s y s t e m s which a r e indigenous to t h e i r p a r t of the wor ld , to extend the r ange of r ad iob io log ica l e x p e r i m e n t s with botanica l s y s t e m s .

1 7 . 2 . GROSS E F F E C T S ON ROOT GROWTH

When seed l ings of Vicia faba a r e exposed to a dose of X - r a y s in the r a n g e of 50 to 250 r ad , the growth r a t e of the p r i m a r y root fol lows a c h a r a c t e r i s t i c p a t t e r n ( see F i g . 35). F o r the f i r s t day the growth r a t e is v i r t u a l l y unaf fec ted , but t h e r e a f t e r f a l l s p r o g r e s s i v e l y un t i l a min imum is r e a c h e d s o m e 4 to 6 days l a t e r , to be followed by a p r o g r e s s i v e r e c o v e r y . The extent to which the g r o w t h - r a t e is d e c r e a s e d , a l so the t ime t aken fo r r e c o v e r y , depends upon the s i ze of the d o s e . F o r the dose range a l r e a d y spec i f i ed , r e c o v e r y is comple te by 10 to 15 days a f t e r t r e a t m e n t , by which t i m e the i r r a d i a t e d r o o t s a r e growing as quickly a s u n i r r a d i a t e d con t ro l r o o t s of the s a m e age . A l a r g e r dose may cause a p r o g r e s s i v e r e d u c t i o n of the growth r a t e to z e r o , and the roo t which t u r n s b rown may not grow again . Th is wil l be the fa te of s o m e r o o t s exposed to doses in e x c e s s of 250 r ad , and to a lmos t a l l of those exposed to 600 r a d . All types of ioniz ing r a d i a t i o n produce the s a m e g r o s s e f f ec t s a s those d e s c r i b e d fo r X - r a y s , but the quant i ta t ive r e l a t i o n s h i p with dose i s v e r y d i f f e r e n t .

Individual r o o t s v a r y c o n s i d e r a b l y , in both growth r a t e and r e s p o n s e to r ad ia t ion , and consequent ly mos t w o r k e r s have e lec ted to expose g roups of be tween 9 and 20 roo t s to e a c h dose l eve l . The r e s u l t s may then be handled by s t a t i s t i c a l me thods . In g e n e r a l t h r e e p a r a m e t e r s have been used by v a r i o u s w o r k e r s to s c o r e the e f f e c t s of r a d i a t i o n on root g rowth .

(a) The ' m e a n le tha l d o s e ' — defined to be the dose which r e s u l t s in c e s s a t i o n of growth f o r 4 o r m o r e days by half of the roo t s of the group (Gray and Read, 1942a; Spalding, Langham and Ander son , 1956, 1958).

(b) The ' m i n i m u m g r o w t h - r a t e ' of the r o o t s , r e a c h e d 6 to 8 days a f t e r i r r a d i a t i o n , e x p r e s s e d as a f r a c t i o n of con t ro l r o o t s of the s a m e length (Gray and Scholes , 1951). Th i s quant i ty is m a r k e d Gm i n in F i g . 35.

(c) The ' g r o w t h in 10 d a y s ' — defined to be the mean i n c r e m e n t in length of the i r r a d i a t e d roo t s in the 10 days fol lowing i r r a d i a t i o n , e x p r e s s e d as a f r a c t i o n of con t ro l r o o t s in the s a m e pe r iod (Read, 1952a; Nea ry , 195 7; Hall , 1961; Hall and La j tha 1963). It i s , in e f f ec t , the a r e a unde r the cu rve up to the 10th day in F i g . 35.

Us ing one o r ano ther of t h e s e p a r a m e t e r s to s c o r e r ad ia t ion damage , a n u m b e r of p r o b l e m s may be inves t iga ted with Vicia s eed l ings .

17. 3. STRUCTURE OF THE ROOT

The s t r u c t u r e of the p r i m a r y root of the b road bean, s t a r t i n g f r o m the t ip, i s roughly as fo l lows . The f i r s t o n e - t h i r d of a m i l l i m e t r e c o n s i s t s of the root cap, the ce l l s of which a r e r e l a t i ve ly i n e r t . The m e r i s t e m occupies the next t h r e e m i l l i m e t r e s of the roo t , and in th i s r eg ion ce l l s a r e ac t ive ly dividing and growing. The next c e n t i m e t r e o r so is t e r m e d the e longat ing zone . Cel l s in th is r eg ion make no m o r e d iv is ions , but

94 CHAPTER 18

* -Neutron beam

t

Gas inlet

-- O-r ing

- G a s outlet

FIG. 36. Diagram of a fixture suitable for irradiation of Vicia seedlings with high energy neutrons. The fixture is filled with growth medium, and bubbled with air or pure nitrogen according to the plan of the experiment. Before reaching the seedlings the neutron beam passes through an appropriate thickness t of tissue-equivalent plastic.

only d i f f e r en t i a t e and e longa te . Ce l l s f r o m the dividing zone pas s into the e longat ing zone and by t h e i r e longat ion p roduce mos t of the o b s e r v e d lengthening of the roo t . Th i s i s a br ief s u m m a r y of the gene ra l l y held p i c tu r e of the roo t t ip (Gray and Scholes , 1951; Read, 1959).

The r educ t ion of roo t growth fol lowing i r r a d i a t i o n i s due en t i r e ly to e f f e c t s on the m e r i s t e m , s ince e x p o s u r e of e i t h e r the m a t u r e sec t ion of the roo t o r the e longat ing zone does not a f fec t root growth provided the m e r i s t e m i tse l f i s sh ie lded (Gray and Scholes , 1951; G r a y and Boag unpubl ished , c i t . Read, 1959.)

1 7 . 4 . CULTURE O F THE SEEDLINGS

Most e x p e r i m e n t s with Vicia fab a have been c a r r i e d out with Sutton' s P r o l i f i c Long Pod b road beans obtained f r o m Sutton and Son, Reading, Eng land . Using s e e d s f r o m th i s s o u r c e e n s u r e s that the m a t e r i a l i s gene t i ca l ly u n i f o r m , and it h a s been the e x p e r i e n c e of many w o r k e r s in d i f f e r en t p a r t s of the wor ld that the r ad iob io log ica l r e s u l t s obtained a r e r e m a r k a b l y c o n s i s t e n t . However , it is obviously not n e c e s s a r y to u s e any p a r t i c u l a r s t r a i n . The bas i c t ech iques have been d e s c r i b e d in de ta i l by Read (1959) and Hal l (1961).

F o r each e x p e r i m e n t , a p p r o x i m a t e l y 500 s e e d s a r e soaked in running t ap w a t e r , the t e m p e r a t u r e of which should be main ta ined c lose to 19°C; v a r i a t i o n s of a few d e g r e e s on e i t h e r s ide wil l r e s u l t in d i f f e r e n c e s in the t i m e r e q u i r e d f o r ge rmina t i on , but wi l l not u l t ima te ly a f fec t the r e s u l t s . A f t e r about 3 days, those that have g e r m i n a t e d , a s judged by the a p p e a r a n c e of a r a d i c a l , a r e planted in moi s t v e r m i c u l i t e o r moi s t sand . The s e e d s a r e

EXPERIMENTS WITH ROOT MERISTEMS 95

allowed to grow in th i s way f o r 4 o r 5 days, be ing kept moi s t dur ing th i s p e r i o d . By the end of th i s growing per iod , the m a j o r i t y should develop a p r i m a r y roo t be tween 6 and 12 cm long, and t h e s e a r e r e m o v e d f r o m the v e r m i c u l i t e , washed in w a t e r , and the t e s t a and p lumule r e m o v e d . E a c h of the s eed l i ngs i s m a r k e d with a n u m b e r on one of the co ty ledons and a r e f e r e n c e point on the hypocotyl . The length f r o m th i s r e f e r e n c e point to the t ip of the growing root i s then m e a s u r e d and r e c o r d e d , and the seed l ings a r e t r a n s f e r r e d to the growing tank .

The m a i n cu l t u r e tank should have a s i ze of a p p r o x i m a t e l y 24 in. by 12 in. and 12 in . deep . In s o m e p a r t s of the wor ld a l l that i s needed f o r growing the seed l ings is the loca l t ap wa te r , while in o the r r eg ions the c o m m e r c i a l supply of w a t e r is so f r e e of t r a c e e l e m e n t s tha t it wi l l not suppor t the growth of s e e d l i n g s . In th i s c a s e it is p r e f e r a b l e to u s e d is t i l led w a t e r to which i s added a c a r e f u l l y con t ro l led m i x t u r e of t r a c e e l e m e n t s such as that sugges t ed by Hall, Ross i , K e l l e r e r , Goodman and Mar ino (19 73).

The t e m p e r a t u r e of the w a t e r in the cu l t u r e tank should be ma in ta ined at 19 ± | ' C s ince the growth r a t e v a r i e s c r i t i c a l l y with t e m p e r a t u r e . Roots die at t e m p e r a t u r e s above 30°C.

The beans a r e placed on a Luci te lid of the tank, with t h e i r r o o t s p a s s i n g downward th rough ho les into the w a t e r . The s e l ec t ed and m a r k e d r o o t s f o r any given e x p e r i m e n t a r e then divided into a n u m b e r of g roups , c a r e f u l l y matched so that each g roup conta ins the s a m e s p e c t r u m of roo t l eng ths . The i r r a d i a t i o n i s c a r r i e d out one o r two days a f t e r the s o r t i n g into g r o u p s . A f t e r i r r a d i a t i o n , the roo t s a r e al lowed to r e m a i n in the m a i n cu l t u r e tank f o r a pe r iod of 10 days, except f o r a br ie f i n t e r v a l each day when the l a t e r a l r o o t s a r e r e m o v e d , and the length of each p r i m a r y roo t i s r e m e a s u r e d .

17 .5 . METHODS O F IRRADIATION

F o r i r r a d i a t i o n , s eed ings a r e held in a f i x t u r e f a b r i c a t e d of Luc i te , s i m i l a r to the des ign d e s c r i b e d by Read (1959), which c o n s t r a i n s a g roup of s eed l ings such that the s ens i t i ve roo t t ips come t o g e t h e r into a s m a l l v o l u m e . The f i x tu re may have an elongated r e c t a n g u l a r c r o s s - s e c t i o n , which r e p r e s e n t s the op t imum des ign f o r expos ing seed l ings to a beam of X - r a y s o r n e u t r o n s . F i g . 36 shows the des ign o f a f i x t u r e u s e d fo r h i g h - e n e r g y n e u t r o n s . A l t e rna t ive ly the f i x tu re may be c i r c u l a r in s ec t i ons , f a b r i c a t e d f r o m a length of Luci te tubing. Th is des ign may be m o r e convenient f o r i r r a d i a t i n g a g roup of s eed l ings with a r i ng of r ad ioac t i ve s o u r c e s spaced a round the f i x t u r e and c o n c e n t r i c with i t .

In e i t h e r c a s e the s ens i t i ve po r t ions of the r o o t s which need to be i r r a d i a t e d , namely the t e r m i n a l half c e n t i m e t r e of each roo t , a r e g a t h e r e d t oge the r into a s m a l l vo lume so that they m a y be exposed to a u n i f o r m r a d i a t i o n f i e l d . The j ig i s . f i l l ed with cu l tu re medium or t ap w a t e r a s the c a s e may be; it i s v e r y i m p o r t a n t to e n s u r e that it i s bubbled v igorous ly with a i r , b e c a u s e roo t m e r i s t e m s u s e a g r ea t dea l of oxygen and the w a t e r in the f i x t u r e i s quickly deoxygena ted .

When it i s r e q u i r e d to i r r a d i a t e the seed l ings u n d e r condi t ions of hypoxia, the f i x tu re should be c losed with a lid f i t t ing onto an O - r i n g , and bubbled v igo rous ly with h igh -pu r i t y n i t rogen f o r about half an hour b e f o r e i r r a d i a t i o n , to scavenge a l l t r a c e s of oxygen f r o m the f i x t u r e . Th is technique has been d e s c r i b e d by Hall and Cavanagh (1967).

96 CHAPTER 18

1 7 . 6 . SCORING RADIATION DAMAGE

The length of the p r i m a r y roo t of each seed l ing i s m e a s u r e d immed ia t e ly b e f o r e i r r a d i a t i o n , and r e m e a s u r e d at the s a m e t i m e daily up to and including the 10th day. The growth i n c r e m e n t in 10 days f o r each individual root i s s imply the d i f f e r e n c e in length be tween the f i r s t and the las t m e a s u r e m e n t . The m e a n 10-day i n c r e m e n t fo r each i r r a d i a t e d group, e x p r e s s e d a s a f r a c t i o n of the c o r r e s p o n d i n g quant i ty fo r u n i r r a d i a t e d con t ro l s , is , as s t a t ed in Sect ion 17 .2 , usua l ly t e r m e d the ' g r o w t h in 10 days ' and has been used fo r many y e a r s as a m e a s u r e of r a d i a t i o n d a m a g e .

E a c h seedl ing i s an independent ent i ty , and if 9 to 14 a r e used f o r each group, with double that n u m b e r r e s e r v e d f o r con t ro l , the r e s u l t s a r e r ead i ly amenab le to e l e m e n t a r y but r i g o r o u s s t a t i s t i c a l ana lys i s (Finney 1952). The daily m e a s u r e m e n t s a r e used to detec t any oddi t ies o r anomal i e s in roo t g rowth that a r e not a s soc i a t ed with the r a d i a t i o n . F o r example , the growth r a t e a f t e r r ad ia t ion does not f a l l dur ing the f i r s t day, but begins to f a l l s ign i f i can t ly t h e r e a f t e r . Roots that f a i l to grow in the f i r s t 24 to 48 h o u r s have p robab ly been damaged by handling dur ing the i r r a d i a t i o n p r o c e d u r e s , and should be excluded f r o m the f ina l c a l cu l a t i ons . Al te rna t ive ly t h e s e addi t ional data may be used to ca l cu la t e the p a r a m e t e r ca l led ' m i n i m u m growth r a t e ' ( see Sect ion 17 .2) .

1 7 . 7 . LOW T E M P E R A T U R E FOR PROLONGED EXPOSURES

At the n o r m a l cu l tu re t e m p e r a t u r e of about 19°C, the ce l l cyc le t i m e of the dividing c e l l s in the m e r i s t e m a t i c r eg ion i s c lose to 24 h o u r s . If ava i lab le d o s e - r a t e s a r e so low that adequate to ta l doses cannot be accu-mula ted in l e s s than 24 h o u r s , then the i n t e r p r e t a t i o n of r e s u l t s b e c o m e s di f f icul t because the p i c tu r e i s compl ica ted by ce l l d ivis ion dur ing the r ad i a t i on e x p o s u r e . Th i s point was d i s c u s s e d in the d o s e - r a t e c h a p t e r of the sy l l abus . At such low d o s e - r a t e s , the d o s e - r a t e e f fec t is due to ce l l p r o l i f e r a t i o n m o r e than to the r e p a i r of sub le tha l d a m a g e .

With plant s eed l ings th is p r o b l e m can be c i r c u m v e n t e d if the t e m p e r a t u r e of the w a t e r in the cu l tu re tank and in the i r r a d i a t i o n f ix tu re i s lowered to about 4°C, when ce l l d iv is ion in the m e r i s t e m , and the m a c r o s c o p i c growth of the roo t , is s topped . Th i s technique was or ig ina l ly d e s c r i b e d by Nea ry (1957) and has been ex tens ive ly used by Hall and L a j t h a (1963), and Hall and Bedford (1964a). The lowered t e m p e r a t u r e does no p e r m a n e n t damage to the r o o t s , and when they a r e subsequent ly r e t u r n e d to t h e i r n o r m a l growing t e m p e r a t u r e of 19°C, the g r o w t h - r a t e r e s u m e s a n o r m a l p a t t e r n . F u r t h e r m o r e , when a dose of r a d i a t i o n is de l ive red to the roo t s at the lower t e m p e r a t u r e , the r a d i a t i o n damage is ' s t o r e d ' . When the roo t s a r e sub-sequent ly rev ived at a l a t e r date to 19°C, the r e s u l t a n t p a t t e r n of growth i s iden t i ca l to that i m m e d i a t e l y a f t e r the s a m e dose de l i ve red to roo t s at 19"C.

Although no ce l l d iv is ion t a k e s p lace in r o o t s main ta ined at th i s low t e m p e r a t u r e , r e c o v e r y of sub le tha l damage a f t e r acute e x p o s u r e s of X - r a y s does t ake place , though m o r e slowly than at 19°C. Us ing th is s y s t e m , t h e r e -f o r e , the e f fec t of v e r y low d o s e - r a t e s involving e x p o s u r e s of s e v e r a l weeks may be inves t iga ted , without the compl ica t ions of ce l l d iv is ion continuing dur ing the i r r a d i a t i o n .

EXPERIMENTS WITH ROOT MERISTEMS 97

1.0

0.9 CALIFORNIUM AT !6rad/hour 0

RADIUM AT 46.9 rad/hour O.E.R. = 2.02

g 0.6

Z 0.5 X

Q Q3

I— 0.4

QL O

Q2

OB

0-1

0 20 30 4 0 50 60 80 100 200 4 0 0 6 0 0 DOSE (rod)

FIG. 37. Data for Vicia seedlings exposed a t low dose-ra te to gamma rays from radium and to the mixed neutron and gamma-ray emission from cal i fornium-252. The score of radiation damage , the 'growth in 10 days1, is plotted on a l inear scale against dose on a logar i thmic scale. The data may then be analysed by parallel straight l ine statistical techniques. (From Hall and Fairchild, 1972).

1 7 . 8 . S P E C I F I C DESIGN O F A LOW D O S E - R A T E E X P E R I M E N T WITH 252Cf AND/OR WITH A GAMMA E M I T T E R SUCH AS RADIUM

T h e f i r s t c r i t i c a l s t e p in the p lann ing of the e x p e r i m e n t i s the d e s i g n and f a b r i c a t i o n of an i r r a d i a t i o n f i x t u r e . T h i s i s p a r t i c u l a r l y t r u e when the a v a i l a b i l i t y of r a d i o a c t i v e s o u r c e s i s l i m i t e d . The m o s t conven ien t d e s i g n i s to e n c l o s e the r o o t s in a L u c i t e tube wi th a d i a m e t e r of one o r two c e n t i m e t r e s , so tha t the roo t t i p s a r e conf ined in a s m a l l v o l u m e . T h e r a d i o a c t i v e s o u r c e s m a y then be d i s t r i b u t e d a r o u n d the tube , c o n c e n t r i c with i t , and at t he l e v e l of the r o o t t i p s . In t h i s way a m a x i m u m dose r a t e c an be ob ta ined , wi th a m i n i m u m n u m b e r of s o u r c e s , to a s many roo t t i p s a s p o s s i b l e .

W h e n t h e p lant s e e d l i n g s a r e c r o w d e d t o g e t h e r in t h i s way, v i g o r o u s a e r a t i o n i s p a r t i c u l a r l y i m p o r t a n t to e n s u r e tha t oxygen i s not dep le t ed in the t i s s u e of the m e r i s t e m s . To ob ta in a f u l l d o s e r e s p o n s e c u r v e f o r s e e d l i n g s exposed to the m i x e d n e u t r o n and g a m m a e m i s s i o n f r o m c a l i f o r n i u m - 2 5 2 , a dose r a n g e f r o m about 10 t o 60 r a d (total d o s e , n e u t r o n s p lus g a m m a r a y s ) n e e d s to be c o v e r e d . If t h i s t o t a l d o s e c a n be ach ieved in a p e r i o d of l e s s t h a n 24 h o u r s , c o r r e s p o n d i n g to a d o s e - r a t e of about 2\, r a d / h o u r , t h e n the e x p e r i m e n t c an be p e r f o r m e d at r o o m t e m p e r a t u r e .

If t he a v a i l a b l e d o s e - r a t e s a r e l o w e r t h a n t h i s , t h e n the t e m p e r a t u r e should be l o w e r e d to 4 ° C du r ing the r a d i a t i o n e x p o s u r e to a r r e s t c e l l d i v i s i o n . F o r a p a r a l l e l e x p e r i m e n t with a p u r e g a m m a e m i t t e r , e . g . r a d i u m , the r a n g e of d o s e s r e q u i r e d v a r i e s wi th the d o s e - r a t e . At about 50 r a d / h o u r a dose r a n g e f r o m 100 to 400 r a d m u s t be c o v e r e d .

98 CHAPTER 18

DOSE ( r o d )

FIG. 38. The z52Cf da ta from Fig. 37, replotted so that "growth in 10 days ' is on a logar i thmic scale and dose on a l inear scale.

Graded d o s e s a r e given to g roups of s eed l ings by r e m o v i n g s o m e of the seed l ings a f t e r v a r i o u s t r e a t m e n t t i m e s . It i s e s s e n t i a l to e n s u r e that the c o n t r o l s a r e t r e a t e d in exac t ly the s a m e way, p a r t i c u l a r l y in the case of lowered t e m p e r a t u r e s o r unconvent ional g a s s i n g t echn iques which may modify the subsequen t g rowth p a t t e r n . E x p e r i m e n t s with Vicia seed l ings exposed to c a l i f o r n i u m - 2 5 2 and rad ium have been d e s c r i b e d by Hall and F a i r c h i l d (1970).

PLOTTING THE DATA

The da ta may be plot ted in two ways . Unti l r e c e n t y e a r s it was con-vent iona l to plot the ' g r o w t h in 10 days ' on a l i n e a r s c a l e aga ins t the dose on a l o g a r i t h m i c s c a l e . The advantage of th i s plot i s that t h e r e i s a l i nea r r e l a t i o n s h i p be tween t h e s e quan t i t i e s plotted in th is way, at l e a s t f o r va lues of the ' g rowth in 10 d a y s ' be tween 0 . 8 and 0 . 2 . Th i s is p a r t i c u l a r l y va luable f o r e x p e r i m e n t s to d e t e r m i n e the oxygen enhancement r a t i o , s ince the dose r e s p o n s e r e l a t i o n s h i p s fo r oxygen and n i t rogen on th is type of plot a r e p a r a l l e l , and the va lue f o r the mean OER, t o g e t h e r with i t s conf idence l imi t s , can be ca lcu la ted us ing s t andard s t a t i s t i c a l methods f o r a p a r a l l e l l ine a s s a y .

In m o r e r e c e n t y e a r s , with the common u s e of m a m m a l i a n ce l l s u r v i v a l c u r v e s , it ha s b e c o m e m o r e u s u a l to plot the ' growth in 10 d a y s ' on a l o g a r i t h m i c s c a l e , aga ins t dose on a l i nea r s c a l e . Dose r e s p o n s e r e l a t ion -sh ips a r e then obtained which a r e ve ry s i m i l a r in shape to those obtained

EXPERIMENTS WITH ROOT MERISTEMS 99

f o r m a m m a l i a n c e l l s . F o r acute e x p o s u r e s of X - r a y s , the dose r e s p o n s e c u r v e has a b road in i t ia l shou lde r , fol lowed by a s t r a igh t po r t ion at h igher d o s e s . At low d o s e - r a t e the shou lder d i s a p p e a r s and the cu rve b e c o m e s a s t r a i g h t l ine, indica t ing that the r e s p o n s e is an exponent ia l funct ion of dose .

Data f r o m Hall and F a i r c h i l d (19 70) have been rep lo t t ed in both ways in F i g s 3 7 and 38.

C h a p t e r 18

MAMMALIAN C E L L S IN C U L T U R E

18. 1. CHOICE O F SYSTEM

M a m m a l i a n ce l l s cu l tu red in v i t r o have been used ex tens ive ly in r ad iob io log ica l e x p e r i m e n t s s i n c e the deve lopment of the technique by P u c k and M a r c u s in 1956. The use of th i s s y s t e m involves a subs t an t i a l cap i t a l i n v e s t m e n t in equipment , the ava i lab i l i ty of soph i s t i ca t ed t echn iques and a c c e s s to suppl ies of t i s s u e c u l t u r e m e d i u m and s e r u m . However , once it ha s been e s t ab l i shed , it i s r e l a t i v e l y s i m p l e to o p e r a t e .

The popu la r i ty of ce l l cu l t u r e t echn iques i s not h a r d to unde r s t and . Since the ce l l l ines used a r e m a m m a l i a n in o r ig in , and f r equen t ly human, i t i s usual ly fe l t t ha t da ta obtained a r e r e l e v a n t to the human c l in ica l s i tua t ion . Cel l cu l t u r e t echn iques a r e f lexib le and can be adapted to a wide r a n g e of e x p e r i m e n t s . C o m p a r e d with an ima l e x p e r i m e n t s , a l a r g e r n u m b e r of s a m p l e s can be used to p rov ide s t a t i s t i c a l va l id i ty to data , at a f r a c t i o n of the cos t involved with in -v ivo t echn iques . When a n s w e r s can be obta ined f r o m i n - v i t r o c u l t u r e s , the s u f f e r i n g involved in a n i m a l e x p e r i m e n t s i s a l so avoided.

18 .2 . C E L L CULTURE TECHNIQUES

Techn iques f o r handling m a m m a l i a n ce l l c u l t u r e s in v i t r o have been d e s c r i b e d in g r e a t de ta i l (Ham and Puck , 1962). With m o d e r n t echn iques of t i s s u e cu l t u r e it i s p o s s i b l e to t ake p i e c e s f r o m a t u m o u r , o r f r o m m a n y n o r m a l r e g e n e r a t i v e t i s s u e s of m a m m a l s , gr ind the s p e c i m e n into s m a l l p i e c e s , s e p a r a t e the individual ce l l s by the use of t r y p s i n which d i s s o l v e s and loosens the ce l l m e m b r a n e , and in th i s way to p r e p a r e a su spens ion of s ing le c e l l s . If t h e s e c e l l s a r e s eeded into a cu l t u r e f l a s k o r P e t r i d ish , cove red with an a p p r o p r i a t e g rowth m e d i u m and main ta ined at 37°C under a s e p t i c condi t ions , they a t tach to the s u r f a c e , grow and divide .

In p r a c t i c e , m o s t f r e s h expian ts p r o d u c e d in th i s way only grow wel l fo r a few weeks , and subsequen t ly s top growing and d ie . A s m a l l p r o -po r t i on of them, p robab ly t h o s e which undergo a mu ta t ion of some s o r t , b e c o m e e s t ab l i shed and cont inue to grow f o r many y e a r s . E v e r y few days the cu l t u r e m u s t be ' f a r m e d 1 . The ce l l s a r e r e m o v e d f r o m the s u r f a c e with t r y p s i n , m o s t of the ce l l s a r e d i s c a r d e d , and the cu l t u r e v e s s e l i s r e s e e d e d with a s m a l l n u m b e r of c e l l s which quickly grow and r epopu la t e the cu l t u r e f l a s k . T h e s e a r e the s o - c a l l e d e s t ab l i shed ce l l l i nes , which have been used ex tens ive ly in e x p e r i m e n t a l c e l l u l a r rad iob io logy .

The e s t ab l i shed ce l l l i nes a r e h a r d y and r e s u l t in r e p r o d u c i b l e r e s u l t s ; they a r e m u c h s i m p l e r to w o r k with than f r e s h exp ian t s . The four m o s t commonly used e s t ab l i shed ce l l l i nes a r e as fo l lows, al though many o t h e r s have been employed .

(a) H e L a ce l l s , de r ived o r ig ina l ly f r o m a human c e r v i c a l c a r c i n o m a . (b) H a m s t e r ce l l l ines de r ived f r o m the lung o r ova ry of a newborn

Ch inese h a m s t e r . (c) Kidney ce l l s of human o r i g i n (Tx c e l l s ) . (d) Mouse L c e l l s , o r ig ina t ing f r o m m o u s e connect ive t i s s u e .

1 0 1

1 0 2 CHAPTER 18

18 .3 . REPRODUCTIVE INTEGRITY

F o r p r o l i f e r a t i n g c e l l s , such a s haema topo ie t i c s t e m ce l l s , c ryp t c e l l s in the gut, o r c e l l s growing in cu l tu re , l o s s of the capac i ty to su s t a in p r o l i f e r a t i o n , i . e . l o s s of r ep roduc t i ve in t eg r i ty , is an a p p r o p r i a t e def in i t ion of ce l l dea th . Th is i s s o m e t i m e s ca l led r e p r o d u c t i v e dea th . The def in i t ion r e f l e c t s a n a r r o w view of r ad i a t i on biology. A ce l l m a y s t i l l be phys ica l ly p r e s e n t and apparen t ly in tac t , i t m a y be able to m a k e p r o t e i n s o r syn thes i ze DNA, it m a y even be able to s t r u g g l e th rough one o r two m i t o s e s , but if i t has los t the capac i ty to divide indef in i te ly , and p r o d u c e a l a r g e n u m b e r of p rogeny , i t i s by def in i t ion dead . Th i s def in i t ion i s r e l e v a n t to the r a d i o -biology of whole a n i m a l s , p lan t s and t h e i r t i s s u e s . It has p a r t i c u l a r r e l e -vance to the r a d i o t h e r a p y of t u m o u r s .

The capabi l i ty of a s ingle ce l l to grow into a l a r g e colony, which can e a s i l y be s e e n with the naked eye, is a convenient proof that i t has r e t a i n e d i ts r e p r o d u c t i v e i n t e g i r t y . The l o s s of th i s abi l i ty as a funct ion of the dose of r a d i a t i o n i s d e s c r i b e d by the dose s u r v i v a l c u r v e s . The f i r s t such s u r v i v a l c u r v e f o r m a m m a l i a n ce l l s was obtained fo r ce l l s g rown in v i t r o , i . e . cu l tu red a r t i f i c i a l l y in P e t r i d i s h e s (Puck and M a r c u s , 1956).

18. 4. SURVIVAL CURVE FOR ATTACHED MAMMALIAN CELLS EXPOSED AT LOW DOSE-RATE TO RADIUM GAMMA RAYS OR THE MIXED NEUTRON AND GAMMA-RAY EMISSION FROM 252Cf

In th i s type of e x p e r i m e n t m a m m a l i a n ce l l s a r e i r r a d i a t e d , while s t i l l s ing le c e l l s o r m i c r o - c o l o n i e s , a t t ached to the s u r f a c e of a cu l t u r e v e s s e l , and subsequen t ly incubated to t e s t f o r the abi l i ty of the ce l l s to p roduce m a c r o s c o p i c colonies as proof of t h e i r s u r v i v a l . It is n e c e s s a r y to a r r a n g e the r a d i u m a n d / o r the c a l i f o r n i u m - 2 5 2 s o u r c e s in an a r r a y such that they un i fo rmly i r r a d i a t e c e l l s a t tached to a n u m b e r of P e t r i d i s h e s . Th i s wi l l not be pos s ib l e un less an a p p r e c i a b l e n u m b e r of r ad ioac t ive s o u r c e s of s u b s t a n t i a l content a r e ava i l ab l e . It i s p r e f e r a b l e to use p l a s t i c P e t r i d i s h e s to avoid d o s i m e t r y p r o b l e m s a s s o c i a t e d with the abso rp t ion of neu t rons in g l a s s . The r ad ioac t ive s o u r c e s m a y be a r r a n g e d in a p l a n a r a r r a y , following a modi f i ed P a t e r s o n and P a r k e r a r r a n g e m e n t (Meredi th , 1967). The P e t r i d i s h e s may be held in a p lane p a r a l l e l to the r ad ioac t i ve s o u r c e s , so that they a r e i r r a d i a t e d at a un i fo rm d o s e - r a t e .

F o r each e x p e r i m e n t , c e l l s f r o m an ac t ive ly growing p a r t l y conf luent s tock bot t le a r e r e m o v e d by t ryps in i za t i on , suspended in fu l l growth m e d i u m , and the ce l l concen t r a t i on i s counted with a h a e m a c y t o m e t e r o r C o u l t e r e l e c t r o n i c ce l l c o u n t e r . A f t e r an a p p r o p r i a t e di lut ion, v a r i o u s n u m b e r s of c e l l s a r e s eeded into s m a l l p l a s t i c P e t r i d i s h e s . F o r 3 5 - m m F a l c o n p l a s t i c P e t r i d i s h e s , no m o r e than about 50 c e l l s c an be allowed to grow_ into co lon ies , o t h e r w i s e they cannot be d i s t inguished and counted. D i s h e s tha t a r e exposed to doses of r a d i a t i o n r e q u i r e m o r e ce l l s to be seeded , so tha t about 50 s u r v i v o r s g row up into co lon ies . The exact n u m b e r in a g iven e x p e r i m e n t m u s t be found by t r i a l and e r r o r .

A f t e r the c e l l s a r e s eeded into the d i shes , they a r e incubated overn igh t at 37°C to allow the c e l l s to b e c o m e f i r m l y a t tached to the s u r f a c e of the t i s s u e c u l t u r e d i s h e s . A f t e r overn igh t a t t achmen t , the i r r a d i a t i o n s a r e c a r r i e d out at r o o m t e m p e r a t u r e , to s top ce l l s dividing dur ing the c o u r s e

MAMMALIAN CELLS IN CULTURE 103

DOSE ( rad ) 100 2 0 0

Experiment 99 Californium-252 -

3 0 0

0 . 0 4

FIG.39. Survival est imates for Chinese hamster cells exposed to ca l i fornium-252 radiat ion at dose-rates from 1 .14 to 4. 68 rad/hour . The doses on the abscissa are the total due to neutrons and gamma rays. The l ine through the data has a 37% dose slope (D0) of 93 rad. (From Hall , 1972).

DOSE (rod )

FIG. 40. Survival estimates for Chinese hamster cells exposed to radium gamma rays at dose-rates from 11.7 to 5 2 . 5 rad /hour . The lines were fi t ted to the data by the method of least squares. (From Hall , 1972).

1 0 4 CHAPTER 18

of the e x p e r i m e n t s . Consequent ly the i r r a d i a t i o n fac i l i ty is se t up at r o o m t e m p e r a t u r e , but of c o u r s e a humid i f ied a t m o s p h e r e of 5% C 0 2 in a i r m u s t be p r e s e r v e d to m a i n t a i n the c o r r e c t pH of the t i s s u e cu l t u r e m e d i u m . Ce l l s a r e exposed for v a r i o u s p e r i o d s of t i m e , with a p r a c t i c a l l imi ta t ion of about 24 hou r s , which i s a l l the m a m m a l i a n ce l l s a t t ached to P e t r i d i shes wil l s u r v i v e at r o o m t e m p e r a t u r e without a l o s s of p la t ing e f f i c i ency . It i s i m p o r t a n t to e n s u r e tha t con t ro l c u l t u r e s a r e used, exposed fo r exac t ly the s a m e length of t i m e to iden t i ca l condi t ions , except tha t they do not r e c e i v e any r ad i a t i on .

At the end of al l of the i r r a d i a t i o n s , the ce l l c u l t u r e s a r e r e t u r n e d to the incuba to r and al lowed to g row up fo r a p e r i o d of s o m e days until co lonies a r e l a r g e enough to be counted e a s i l y . F o r Ch inese h a m s t e r ce l l s , which have a ce l l cyc le t i m e of about 100 h o u r s , the growth incubat ion p e r i o d m u s t be 1 week; f o r H e L a ce l l s , which have a ce l l cycle t i m e of 2 0 - 24 h o u r s , 2 weeks incubat ion i s n e c e s s a r y . At the end of th is p e r i o d of incubat ion, the ce l l s a r e f ixed and s ta ined and the n u m b e r of v i s ib le colonies p e r d i sh counted . F o r th is p u r p o s e it i s convenient to p r o j e c t the image of the p la te onto a l a r g e s c r e e n so that the n u m b e r of colonies p e r d ish can be counted a c c u r a t e l y , and examined c lose ly .

The f r a c t i o n of c e l l s su rv iv ing each r ad i a t i on dose m a y be ca lcu la ted f r o m a knowledge of the n u m b e r of co lon ies on the i r r a d i a t e d p l a t e s c o m -p a r e d with the c o n t r o l p l a t e s . The e x p e r i m e n t m a y be r e p e a t e d f o r v a r i o u s d o s e - r a t e s of c a l i f o r n i u m - 2 5 2 and a l so of r a d i u m . Data obtained in typ ica l e x p e r i m e n t s c a r r i e d out by th i s p r o c e d u r e a r e shown in F i g s 39 and 40. F o r c a l i f o r n i u m - 2 5 2 the s lope of the s u r v i v a l cu rve does not v a r y much with d o s e - r a t e , while f o r r a d i u m the s lope of the s u r v i v a l cu rve i s a s t r o n g funct ion of the d o s e - r a t e used . T h e s e p a r t i c u l a r e x p e r i m e n t s w e r e c a r r i e d out with Ch inese h a m s t e r c e l l s , but they could obviously be r e p e a t e d with any of the e s t ab l i shed ce l l l i n e s .

18. 5. SURVIVAL CURVES AT LOW DOSE-RATE FOR RADIUM AND/OR 2 5 2Cf USING MAMMALIAN C E L L S IRRADIATED IN SUSPENSION

W h e r e a l im i t ed n u m b e r of r ad ioac t i ve s o u r c e s a r e avai lable , it i s o f ten m o r e p r a c t i c a l to obtain an adequate d o s e - r a t e and convenient g e o m e t r y , if the m a m m a l i a n c e l l s a r e i r r a d i a t e d in su spens ion in s m a l l v o l u m e s r a t h e r than a t tached to the s u r f a c e of P e t r i d i s h e s . A convenient technique f o r e x p e r i m e n t s of t h i s s o r t has been d e s c r i b e d by Hall and L e h n e r t (1973) and by Hal l , Ro iz in -Towle , and Colvet t (1974). S i m i l a r t echn iques have been used by D j o r d j e v i c et a l . (1973) and Nias et a l . (1973).

F o r each e x p e r i m e n t c e l l s f r o m an ac t ive ly growing pa r t ly conf luent cu l t u r e f l a s k a r e h a r v e s t e d by t ryps in i za t i on , washed in m e d i u m to r e m o v e e x c e s s t r y p s i n , and p r e p a r e d into a su spens ion cons i s t ing of app rox ima te ly 15 000 ce l l s p e r m i l l i l i t r e . O n e - t h i r d of a m i l l i l i t r e of th i s suspens ion i s p ipe t ted into each of a l a r g e n u m b e r of s m a l l p l a s t i c c o n t a i n e r s . In the p a p e r s r e f e r r e d to, t h e s e p l a s t i c c o n t a i n e r s w e r e f a b r i c a t e d f r o m 1 - m l F a l c o n d i sposab le p ipe t t e s , which w e r e cut into 5 - c m lengths and sea l ed at the top and the bo t tom by the appl ica t ion of hea t . T h e s e p ipe t t e s have an e x t e r n a l d i a m e t e r of 4 m m and an i n t e r n a l d i a m e t e r of 2 m m . A f t e r the c e l l s have been s ea l ed ins ide t h e s e c o n t a i n e r s , gent le cen t r i fuga t ion c a u s e s the ce l l s to s e t t l e out into a v e r y s m a l l vo lume in the bot tom of

MAMMALIAN CELLS IN CULTURE 1 0 5

1.0

D O S E ( r o d )

0 200 400 600 800 1000 1200 1400 S f c ç L L . i — i — i — i — i i — i — i — i i i r ~

.04 A FIG.41. Survival curves for Chinese hamster cells irradiated at low dose-rate with gamma rays from radium, or the mixed neutron and gamma- ray emission from cal i fornium-252. Irradiations were performed with high ce l l concentrations (hypoxic) and low cel l concentrations (aerated). (From Hall , Roizin-Towle and Colvett , 1974).

each . T h e s e c o n t a i n e r s m a y then be p laced in a s m a l l c i r c l e a round a r ad ioac t i ve s o u r c e of c a l i f o r n i u m - 2 5 2 o r of r a d i u m , in o r d e r to obtain the r e q u i r e d d o s e - r a t e . To e n s u r e un i fo rmi ty of dose , the v i a l s should be t u rned s e v e r a l t i m e s dur ing e x p o s u r e .

The p lan of the e x p e r i m e n t i s a s fo l lows . A l a r g e n u m b e r of ce l l s a m p l e s (30 o r 40 can e a s i l y be handled) a r e seeded with about 5000 c e l l s . Some a r e r e s e r v e d f o r con t ro l s and o t h e r s exposed to a r a n g e of g r aded d o s e s . At l e a s t fou r r e p l i c a t e v i a l s should be exposed to each dose . The g r a d e d d o s e s a r e obtained by p lac ing them f o r d i f f e r e n t l eng ths of t i m e at t he f ixed d i s t ance f r o m the r ad ioac t ive s o u r c e s . I r r a d i a t i o n s a r e c a r r i e d out at r o o m t e m p e r a t u r e , and it is found by e x p e r i e n c e tha t ce l l s in s u s -pens ion , p rov ided t h e i r pH is not d i s tu rbed , wil l s u r v i v e f o r at l e a s t 24 h o u r s .

A f t e r comple t ion of al l the i r r a d i a t i o n e x p o s u r e s , each ce l l con ta ine r i s t r e a t e d in t h e fol lowing way . A v o r t e x m i x e r m a y be used to r e s u s p e n d the ce l l s in the con t a ine r , a f t e r which it i s opened and the conten ts a r e mixed thoroughly by v igo rous p ipet t ing with a P a s t e u r p ipe t t e . V a r i o u s f r a c t i o n s of the r e s u l t i n g ce l l su spens ion then can be r e p l a t e d into P e t r i d i s h e s containing f r e s h growth m e d i u m , and al lowed to incuba te at 37°C until co lonies a r e l a r g e enough to be r ead i ly counted. A f t e r being f ixed and s t a ined the n u m b e r of co lon ies p e r d ish i s counted, p r e f e r a b l y by a p r o j e c t i o n t echn ique . The f r a c t i o n of ce l l s su rv iv ing any p a r t i c u l a r r a d i -at ion dose i s ca l cu la t ed f r o m a c o m p a r i s o n of i r r a d i a t e d and con t ro l d i s h e s . Surv iva l c u r v e s obtained by th i s technique a r e shown in F i g . 41, t aken f r o m Hal l , R o i z i n - T o w l e and Colve t t (1974). In th i s p a r t i c u l a r e x p e r i m e n t , s o m e ce l l c o n t a i n e r s w e r e s eeded with ce l l s at a g r e a t e r concen t r a t i on to c a u s e t h e m to use up oxygen and b e c o m e hypoxic . In th i s way the oxygen enhance -m e n t r a t i o , a s wel l a s the r e l a t i v e b io logica l e f f e c t i v e n e s s , could be

106 CHAPTER 18

CHINESE H A M S T E R CELLS C A L I F O R N I U M - 2 5 2

FIG. 42. Family of curves charting the total number of viable cells in a hamster cell-population exposed to a range of dose-rates from californium-252. (From Hall, Rossi and Roizin, 1971).

compu ted . This t echn ique fo r p roduc ing hypoxia is only m a r g i n a l l y s u c -c e s s f u l , but is the b e s t ava i lab le at p r e s e n t in which c e l l s a r e enc losed in p l a s t i c f o r i r r a d i a t i o n with n e u t r o n s .

The e x p e r i m e n t s should be r e p e a t e d s e v e r a l t i m e s f o r a g a m m a -emi t t ing s o u r c e , such a s r a d i u m , and f o r v a r i o u s d o s e - r a t e s of c a l i f o r n i u m - 252.

18. 6. GROWTH-RATE EXPERIMENTS WITH MAMMALIAN CELLS IRRADIATED AT LOW DOSE-RATE WITH RADIUM AND/OR 2 5 2Cf

The p u r p o s e of t h i s e x p e r i m e n t is not to count the p r o p o r t i o n of ce l l s which a r e able to g row up into v i s ib l e co lon ies , cons i s t ing of ce l l s which a r e a l l the d e s c e n d a n t s of a c o m m o n a n c e s t o r , but i n s t ead to o b s e r v e the e f f ec t of low d o s e - r a t e i r r a d i a t i o n on the s i z e of a growing populat ion of c e l l s .

An a r r a y of r a d i u m o r c a l i f o r n i u m - 2 5 2 s o u r c e s m u s t be a s s e m b l e d to p r o v i d e a un i fo rm d o s e - r a t e o v e r a n u m b e r of s m a l l p l a s t i c P e t r i d i s h e s . As in Sec t ion 18 .4 a P a t e r s o n and P a r k e r d i s t r ibu t ion m a y be used, with the P e t r i d i shes loca ted in a p lane p a r a l l e l with the r ad ioac t ive s o u r c e s .

MAMMALIAN CELLS IN CULTURE 107

C H I N E S E H A M S T E R C E L L S R A D I U M

FIG. 43. Family of curves charting the to ta l number of viable cells in a hamster cel l population exposed to a range of dose-rates of radium g a m m a rays. (From Hall, Rossi and Roizin, 1971).

The r a d i a t i o n fac i l i ty f o r th i s e x p e r i m e n t m u s t be conta ined in a 37°C i n c u b a t o r .

F o r th is e x p e r i m e n t a s low-growing ce l l l ine i s p r e f e r a b l e , such a s f o r example HeLa c e l l s with a doubling t i m e of app rox ima te ly 24 h o u r s . F o r each e x p e r i m e n t a n u m b e r of ce l l s ( e . g . 1000) is s eeded into each of a l a r g e n u m b e r of s m a l l p l a s t i c P e t r i d i s h e s . The d i shes a r e incubated overn igh t to allow the ce l l s to b e c o m e f i r m l y a t t ached . Some of the d i s h e s a r e r e s e r v e d f o r c o n t r o l s , and the r e m a i n d e r scheduled f o r i r r a d i a t i o n . A r a n g e of f o u r o r f ive r a d i a t i o n d o s e - r a t e s should be used .

At dai ly i n t e r v a l s one d ish f r o m the con t ro l s e r i e s and one f r o m each r a d i a t i o n d o s e - r a t e a r e r e m o v e d f r o m the incuba to r , and the n u m b e r of ce l l s p e r d ish counted . Th is i s accompl i shed by r e m o v i n g the ce l l s f r o m the d ish by t r y p s i n i z a t i o n , and counting the ce l l concen t r a t i on in the r e s u l t i n g s u s p e n s i o n by m e a n s of a Cou l t e r e l ec t ron i c ce l l c o u n t e r . This count wi l l give the to ta l n u m b e r of ce l l s contained in the d ish , both v iable and non-v i ab l e . As an a l t e r n a t i v e to counting the ce l l s in a ce l l coun te r , a l iquots of the s u s p e n s i o n could be r e - s e e d e d into new P e t r i d i shes containing f r e s h growth m e d i u m , incubated f o r an a p p r o p r i a t e pe r iod of t i m e , and a n u m b e r of co lon ies counted . Th i s would give the n u m b e r of v iab le c e l l s . Th i s should be r e p e a t e d dai ly , counting one con t ro l and one i r r a d i a t e d d ish f o r a pe r iod of s e v e r a l w e e k s .

108 CHAPTER 18

2 20 I 1 1 1 I I I

Neutron \ " 0 . 4 7

Days o f " exposure

o 3 CVJ m c \ j 3 J LJL

2 3 5 7 10 2 5 2 C f DOSE-RATE (rad/h)

FIG. 44. By comparison and interpolation from Figs 42 and 43, it is possible to select dose-rates of radium and californium-252 which result in the same cell survival fraction in the same radiation treatment t ime. The ratio of these dose-rates is the relative biological effectiveness of californium-252 radiation compared with radium gamma rays. The RBE is seen to be a function of dose-rate. (From Hall, Rossi and Roizin, 1971).

The populat ion s i z e in the u n i r r a d i a t e d d i s h e s wi l l i n c r e a s e exponent ia l ly , while the ce l l populat ion (at l e a s t the v iab le ce l l populat ion) in the i r r a d i a t e d d i s h e s wi l l i n c r e a s e at a much s l o w e r r a t e . E x a m p l e s of the p a t t e r n of g rowth obtained in e x p e r i m e n t s with th i s s o r t , t aken f r o m Hal l , R o s s i and Roiz in (1971), a r e shown in F i g s 42 and 43. T h e s e e x p e r i m e n t s w e r e c a r r i e d out with Ch inese h a m s t e r c e l l s , which have a s h o r t ce l l cyc le . If c e l l s with a l onge r ce l l cyc le could be employed, l ower d o s e - r a t e s of r a d i u m a n d / o r c a l i f o r n i u m - 2 5 2 would be a c c e p t a b l e .

F r o m th i s type of e x p e r i m e n t it i s p o s s i b l e to ca lcu la te the RBE of c a l i f o r n i u m - 2 5 2 c o m p a r e d with r a d i u m o v e r a r a n g e of dose r a t e s w h e r e the end-po in t is the ce l l populat ion s i ze under cont inuous i r r a d i a t i o n . RBE ca lcu la t ions of t h i s type a r e i l l u s t r a t e d in F ig . 44. The r a t i o of d o s e - r a t e s of r a d i u m and c a l i f o r n i u m - 2 5 2 which p roduce the s a m e ce l l surv iv ing f r a c t i o n in the s a m e t r e a t m e n t t i m e m a y be def ined a s the r e l a t i v e b io logica l e f f ec t i ve -n e s s of c a l i f o r n i u m - 2 5 2 r ad i a t i on c o m p a r e d with r a d i u m g a m m a r a y s . By c o m p a r i n g F i g s 42 and 43, and by in te rpo la t ing be tween the l imi ted n u m b e r of d o s e - r a t e s ava i lab le ( fac i l i ta ted by a rep lo t t ing of the data , whe re s u r -viving f r a c t i o n i s p lot ted v e r s u s d o s e - r a t e ) a r a n g e of R B E s may be ca lcu la ted fo r d i f f e r e n t d o s e - r a t e s . I t is at once evident that RBE is a s t rong funct ion of d o s e - r a t e .

C h a p t e r 19

RESPONSE OF MOUSE J E J U N A L C R Y P T C E L L S

19 .1 . LIMITATIONS O F THE SYSTEM

E x p e r i m e n t s wi th th i s s y s t e m can only be p e r f o r m e d if a n u m b e r of c a l i f o r n i u m - 2 5 2 s o u r c e s of r e l a t i v e l y l a r g e r ad ioac t i ve content a r e ava i l ab le . R e f e r e n c e should be m a d e to the p a p e r by W i t h e r s , O l ive r and Glenn (1971). The b a s i c b io logica l s y s t e m was desc r ibed in de ta i l by W i t h e r s and Elkind (1970).

19. 2. DESIGN O F THE E X P E R I M E N T

An i r r a d i a t i o n f i x t u r e should be cons t ruc t ed cons i s t ing of a l a r g e polye thylene phantom, which has a cy l ind r i ca l cavi ty f o r m e d along i t s c e n t r a l ax i s , jus t l a r g e enough to contain one m o u s e (approx imate ly 7 c m long with a d i a m e t e r of 2. 2 cm) . The s o u r c e s should be loca ted to f o r m a c y l i n d r i c a l a r r a y p a r a l l e l to the axis of the cavi ty into which the m o u s e i s to be p l aced , and a t a l eve l so tha t the abdomina l r e g i o n of the m o u s e

HOURS OF EXPOSURE 12 16 20 24

FIG. 45. Dose-survival curves for crypt cells as a function of duration of exposure to z52Cf and radium at the dose-rates specified. (From Withers, Oliver and Glenn, 1971).

109

110 CHAPTER 18

i s i r r a d i a t e d at an e s s e n t i a l l y homogeneous d o s e - r a t e . The d i a m e t e r of a c i r c l e around which the r ad ioac t i ve s o u r c e s wi l l be loca ted is obviously a funct ion of t h e i r r ad ioac t ive content . One phan tom may be cons t ruc t ed to house c a l i f o r n i u m - 2 5 2 s o u r c e s , whi le a s i m i l a r phantom may house a g a m m a - e m i t t i n g i so tope such as r a d i u m o r coba l t -60 .

The l imi t ing f a c t o r in the des ign of t h e s e e x p e r i m e n t s i s the fac t that the m o u s e cannot be cons t r a ined within such a r ad i a t i on holder f o r pe r i ods in e x c e s s of 24 h o u r s . To obtain countable n u m b e r s of r e g e n e r a t i n g c r y p t s , doses m u s t be in the r a n g e of 2000 to 4000 r a d f o r low d o s e - r a t e g a m m a r a y s , and 400 to 700 r a d fo r c a l i f o r n i u m - 2 5 2 r ad i a t i on . In p r a c t i c e , t h e r e f o r e , c a l i f o r n i u m - 2 5 2 d o s e - r a t e s cannot be l o w e r than about 20 r a d / hour , and r a d i u m d o s e - r a t e s cannot be lower than about 150 r a d / h o u r .

O v e r a pe r iod of t ime , r e p l i c a t e a n i m a l s m u s t be exposed to g r aded doses of c a l i f o r n i u m - 2 5 2 a n d / o r r a d i u m g a m m a r a y s , by exposing t hem fo r v a r i o u s p e r i o d s of t i m e in the r ad i a t i on f i x t u r e . A f t e r i r r a d i a t i o n , s ec t i ons of the j e junum m u s t be p r e p a r e d and the surv iv ing c r y p t s s c o r e d . The a v e r a g e n u m b e r of surv iv ing c r y p t s p e r c i r c u m f e r e n c e i s then plot ted as a funct ion of dose . In th is way the e f f e c t s of c a l i f o r n i u m - 2 5 2 and of r a d i u m m a y be c o m p a r e d .

F i g . 45 shows the data obtained by W i t h e r s , O l ive r and Glenn (1971) using th is b iological t e s t s y s t e m . Data such as t h e s e a r e of p a r t i c u l a r va lue because they involve the r e s p o n s e of a n o r m a l t i s s u e .

C h a p t e r 20

RADIATION-INDUCED MAMMARY TUMOURS IN THE RAT

20. 1. THE SYSTEM DESCRIBED

T u m o u r induct ion i s a v e r y impor t an t l a te e f f ec t of r ad i a t i on . It is a c a u s e of g r a v e c o n c e r n when human populat ions r e c e i v e s m a l l doses a s a consequence of occupa t iona l o r m e d i c a l e x p o s u r e . As such it has been ex tens ive ly s tudied in a n i m a l s . The two m o s t commonly used an ima l s y s t e m s a r e the p roduc t ion of l eukemia in m i c e , and the induction of m a m m a r y n e o p l a s m s in the Sprague Dawley r a t .

T h e r e is a high n a t u r a l inc idence of m a m m a r y n e o p l a s m s in f e m a l e Sprague Dawley r a t s once they exceed 18 months of age . However , if the a n i m a l s a r e i r r a d i a t e d at about 6 weeks of age , and subsequent ly o b s e r v e d fo r 11 mon ths , a l a r g e n u m b e r of r ad i a t i on - induced t u m o u r s can be o b s e r v e d while e s s e n t i a l l y none a r e s e e n in con t ro l l ed an imal s . -

It i s commonly r e p o r t e d (She l l aba rge r , Cronk i te , Bond and Lippincot t , 1957), tha t m o r e than one t u m o u r m a y be induced in a g iven a n i m a l . Con-sequent ly i t is c o m m o n p r a c t i c e to exc i se t u m o u r s when they r e a c h a con-venien t s i z e of about 1 cm, so that o t h e r t u m o u r s m a y be s c o r e d in the s a m e a n i m a l if they a r i s e at a l a t e r da te . Of c o u r s e , if a second t u m o u r o c c u r s in the s a m e s i t e , it i s p robab ly a r e c u r r e n c e and i s not s c o r e d as an independent en t i ty .

A f t e r a sub le tha l to ta l body dose of rad ia t ion , t u m o u r s begin to appea r 2 o r 3 mon ths a f t e r i r r a d i a t i o n , and addi t ional t u m o u r s a r e found throughout the 11-month pe r iod of o b s e r v a t i o n . A typica l r e s u l t i s shown in F ig . 46 f o r v a r i o u s doses of X and g a m m a r a y s .

The t u m o u r s that a r e o b s e r v e d a r e of d i f f e r ing h i s to log ica l types , and show v a r i o u s d e g r e e s of ma l ignancy . A d e n o f i b r o m a s a r e the m o s t common , followed by a d e n o c a r c i n o m a s ; f i b r o a d e n o m a s and f i b r o s a r c o m a s a r e much l e s s c o m m o n .

20. 2. TUMOUR INDUCTION BY HIGH L E T RADIATIONS

Neu t rons of v a r i o u s e n e r g i e s , including f i s s i o n n e u t r o n s and 430-keV m o n o e n e r g e t i c n e u t r o n s , have been shown to p r o d u c e m a m m a r y t u m o u r s in Sp rague Dawley r a t s with high e f f i c i ency . A dose of only a few r á d s is su f f i c i en t to p roduce a s c o r e a b l e n u m b e r of t u m o u r s . The r e l a t i v e bio-log ica l e f f e c t i v e n e s s of n e u t r o n s c o m p a r e d with g a m m a r a y s in the low dose r a n g e i s v e r y high indeed, and m a y even a p p r o a c h a va lue of 100.

E x p e r i m e n t s with the mixed neu t ron and g a m m a - r a y e m i s s i o n f r o m c a l i f o r n i u m - 2 5 2 is compl i ca t ed by the r e l a t ive ly few s o u r c e s that a r e usual ly ava i lab le to any given e x p e r i m e n t e r . As a r e s u l t only a l im i t ed vo lume of b r e a s t t i s s u e can be i r r a d i a t e d , P r e l i m i n a r y e x p e r i m e n t s that have been p e r f o r m e d by v a r i o u s w o r k e r s , exposing Sprague Dawley r a t s to c a l i f o r n i u m - 2 5 2 s o u r c e s , ind ica te that f e w e r t u m o u r s a r e p roduced

1 1 1

1 1 2 CHAPTER 18

I 2 3 4 5 6 7 8 9 » ! MONTHS AFTER

IRRADIATION 200 R y-RAY

I 2 3 4 5 6 7 8 9 1 0 1 1 MONTHS AFTER

IRRADIATION 400 R y-RAY

I 2 3 4 5 6 7891011" MONTHS AFTER

IRRADIATION 400 R X-RAY

FIG.46. Illustrating the rate of appearance of tumours after irradiation with various doses of X and y-rays. The numbers in brackets refer to the number of animals per group. Clear areas refer to rats with one tumour only, stippled areas to rats with two tumours, diagonal lines to rats with three tumours, cross-hatched areas to rats with four tumours, and solid areas to rats with f ive tumours. (From Shellabarger, Cronkite, Bond and Lippincott, 1957).

than migh t be expec ted f r o m the ava i lab le da ta in the l i t e r a t u r e pe r t a in ing to b e a m s of neu t rons g e n e r a t e d by a c c e l e r a t o r s . Th i s ques t ion i s s t i l l in s o m e doubt, but i t is p robab ly a vo lume e f f e c t . When c a l i f o r n i u m - 2 5 2 s o u r c e s a r e used, only a r e l a t i v e l y s m a l l p r o p o r t i o n of b r e a s t t i s s u e i s i r r a d i a t e d with n e u t r o n s , c o m p a r e d with the whole-body exposu re to e x t e r n a l s o u r c e s of g a m m a r a y s o r n e u t r o n s . Th i s is the p robab le explanat ion, but s o m e whole-body s y s t e m a t i c e f f ec t cannot be ru led out .

20. 3. SPECIFIC DESIGN O F AN E X P E R I M E N T

A c a l i f o r n i u m - 2 5 2 s o u r c e , held in a s m a l l p l a s t i c con ta ine r , is taped onto a r a t in such a way t h a t ' b r e a s t t i s s u e i s held at a d i s t ance of be tween j c m and 1 c m f r o m the r ad ioac t i ve s o u r c e . C a r e f u l e s t i m a t e s should be m a d e of the vo lume of b r e a s t t i s s u e tha t i s exposed to v a r i o u s doses of r a d i a t i o n . The c a l i f o r n i u m - 2 5 2 s o u r c e is a t t ached , in tu rn , to a n u m b e r of a n i m a l s and l e f t in p l ace f o r p e r i o d s va ry ing f r o m 6 to 24 h o u r s . An a p p r o p r i a t e n u m b e r of con t ro l a n i m a l s mus t , of c o u r s e , be used, which a r e sub jec t ed to s i m i l a r handling and t r a u m a .

The a n i m a l s a r e r a n d o m i z e d and exposed when they a r e app rox ima te ly 6 weeks of age . A f t e r the i r r a d i a t i o n s , which m a y take s e v e r a l weeks to

RADIATION-INDUCED MAMMARY TUMOURS IN THE RAT 1 1 3

comple t e s ince a n u m b e r of a n i m a l s m u s t be exposed , the r a t s a r e o b s e r v e d at f r e q u e n t and r e g u l a r i n t e r v a l s fo r a pe r iod of 11 m o n t h s . T u m o u r s that a r i s e a r e c a r e f u l l y exc i sed and cut into s ec t i ons so that they can be c l a s s i f i e d h i s to log ica l ly . By r emov ing t u m o u r s when they a r e s m a l l , a n i m a l s may r e m a i n in the e x p e r i m e n t so that subsequen t t u m o u r s in the s a m e a n i m a l m a y be s c o r e d .

At the end of the e x p e r i m e n t a g r a p h i s p lot ted of the ave rage n u m b e r of t u m o u r s p e r a n i m a l a s a funct ion of dose . The e x p e r i m e n t m a y be r e p e a t e d using a g a m m a - r a y emi t t ing s o u r c e , such as r a d i u m or coba l t -60 .

Da ta r e l a t i ng to the induct ion of t u m o u r s by low d o s e - r a t e i r r a d i a t i o n , and the r e l a t i v e b io logica l e f f i c i ency of f i s s i o n neu t rons c o m p a r e d with g a m m a r a y s , would c o m p r i s e v e r y va luab le and p e r t i n e n t i n f o r m a t i o n . L i t t l e i n f o r m a t i o n on th i s sub jec t i s ava i lab le at the p r e s e n t t i m e .

In the few p r e l i m i n a r y e x p e r i m e n t s that have been r e p o r t e d in the l i t e r a t u r e f o r the p roduc t ion of m a m m a r y t u m o u r s in Sp rague Dawley r a t s a f t e r c a l i f o r n i u m - 2 5 2 i r r a d i a t i o n (Vogel, 1972), no t u m o u r s w e r e o b s e r v e d in a n i m a l s that w e r e exposed to 5 h o u r s with a 1 - f jC i s o u r c e of c a l i f o r n i u m -252 taped s e c u r e l y o v e r the m a m m a r y gland of the r a t . Overn igh t r ad i a t i on , amount ing to 16 to 18 h o u r s , was n e c e s s a r y b e f o r e t u m o u r s w e r e o b s e r v e d in a n i m a l s i r r a d i a t e d in th i s way .

F u r t h e r da ta f r o m Vogel (1973) showed that of 173 r a t s exposed whole -body to f i s s i o n s p e c t r u m n e u t r o n s with d o s e s in the r a n g e 2 to 20 r ad , 7% of the a n i m a l s developed m à m m a r y t u m o u r s by 200 days , and 25% by 350 days p o s t - i r r a d i a t i o n . In the con t ro l populat ion of r a t s spontaneous m a m m a r y n e o p l a s m s w e r e o b s e r v e d in l e s s than 3% of the a n i m a l s during the f i r s t y e a r of l i f e .

Appendix I

PHYSICAL C H A R A C T E R S OF CALIFORNIUM-252

Atomic number 98

Atomic weight 252

Half- l ife for alpha emissions 2 .73 a

Half- l i fe for spontaneous fission 85.5 a

Half- l ife 2 .65 a

Fraction of decays by alpha emission 97<7o

Fraction of decays by spontaneous fission 3°lo

Average number of neutrons per spontaneous fission 3. 8

Decay heat (51<ï> from fission, 4970 from a decay) 39 (JW/(Jg

Specific activity 530 ßCi/ßg

Neutrons

Neutron emission rate 2 . 3 5 x l o V 1 - | J g n

Tissue kerma rate in free ait at 1 m 0.20 mrad-h"1- Mg"'

Surface dose-rate in large tissue phantom at 1 m 0 .28 m rad • h"1 • f ig '1

Neutron spectrum See Table II

Quality Factor (Q) at surface of large phantom ~ 8- 5

TABLE II. NEUTRON FLUENCE RATE AT 1 m FROM A 1 ng 252Cf SOURCE

Energy Interval

MeV cm"2 -s"1

0 - 0 . 5 2 . 2

0 . 5 - 1.0 2 . 9

1 . 0 - 2 . 0 6 . 1

2 . 0 - 3 . 0 3.7

3 . 0 - 4 . 0 2 .2

4 . 0 - 5 . 0 1 .3

5 . 0 - 6 . 0 0 .45

6 . 0 - 7 . 0 0.32

7 . 0 - 8.0 0 . 1

8 . 0 - 10.0 0 .08

1 0 . 0 - 13.0 0.02

0 - 1 3 . 0 19.4

1 1 5

1 1 6 APPENDIX II

TABLE III. GAMMA F L U E N C E RATE AT 1 m FROM A 1 ßg 252Cf SOURCE

Energy Interval MeV

photons-cm" 2 -s" 1

0 - 0 . 5 37

0 . 5 - 1 . 0 45

1 . 0 - 1 .5 14

1 . 5 - 2 . 0 6 . 1

2 . 0 - 2 . 5 1 . 8

2 . 5 - 3 . 0 0 . 8 8

3 . 0 - 3 . 5 0 . 4 5

3 . 5 - 4 . 0 0 . 2 4

4 . 0 - 4 . 5 0 . 1 4

4 . 5 - 5 . 0 0 .065

5 . 0 - 5 . 5 0 . 0 4

5 . 5 - 6 . 0 0 . 0 1

6 . 0 - 6 . 5 0 . 0 1

0 - 6 . 5 105.7

TABLE IV. NEUTRON ABSORBED DOSE DISTRIBUTION IN r a d - Mg"1'!!"1 FOR A 252Cf APPLICATOR TUBE OF 1.5 c m ACTIVE LENGTH AND 0.7 m m WALL THICKNESS (Af te r Colvet t et al., 1972)

0 0 0 068 0 113 0 220 0 578 (OXXXJOOOOOOOOO 0 631 0 238 0 121 0 072 0 5 0 070 0 116 0 226 0 565 2 51 6 49 2 56 0 572 0 226 0 117 0 071 X 0 0 069 0 111 0 203 0 454 1 224 2 05 1 225 0 459 0 205 0 110 0 067 1 5 0 064 0 100 0 175 0 348 0 679 0 948 0 686 0 346 0 177 0 101 0 064 2 0 0 0S9 0 089 0 148 0 257 0 420 0 527 0 423 0 257 0 148 0 090 0 059 2 5 0 0S4 0 079 0 124 0 189 0 278 0 327 0 280 0 192 0 122 0 079 0 054 3 0 0 0468 0 069 0 100 0 143 0 193 0 219 0 195 0 145 0 100 0 068 0 0489 3 S 0 060 0 081 0 110 0 140 0 154 0 141 0 112 0 082 0 059 4 0 0 051 0 067 0 087 0 105 0 114 0 106 0 088 0 068 0 051 4 s 0 0443 0 056 0 069 0 081 0 086 0 082 0 070 0 057 0 0450 S 0 0 0472 0 057 0 064 0 068 0 065 0 057 0 0481 S s 0 .0471 0 052 0 055 0 053 0 0477 t 0 0 0437 0 0453 0 0440

-5 - 4 - 3 - 2 -1 0 1 2 3 4 5

AXIAL DISPLACEMENT (cm)

PHYSICAL CHARACTERISTICS 1 1 7

TABLE V. GAMMA ABSORBED DOSE DISTRIBUTION IN r a d - M g ' ^ h " 1

FOR A 252Cf APPLICATOR TUBE OF 1.5 cm ACTIVE LENGTH AND 0.7 m m WALL THICKNESS (Af te r Colvet t et al. , 1972)

B o .

0 0 0 0291 0 0426 0 072 0 164 (OXXXXXXXXXXX) 0 206 0 089 0 053 0 0362 B o . 0 5 0 0298 0 0438 0 074 0 180 0 902 2 61 0 939 0 213 0 094 0 056 0 0377

H Z w

w U <

1 0 0 0310 0 0478 0 085 0 185 0 513 0 845 0 504 0 191 0 093 0 056 0 0384 H Z w

w U <

1 5 0 0342 0 052 0 085 0 160 0 309 0 409 0 298 0 161 0 088 0 055 0 0384

H Z w

w U <

2 0 0 0358 0 052 0 080 0 132 0 202 0 242 0 197 0 129 0 081 0 053 0 0378

H Z w

w U < 2 5 0 0355 0 0500 0 073 0 108 0 14 3 0 162 0 140 0 103 0 073 0 0 5 1 0 0366 HJ Cl, CO 3 0 0 0346 0 0469 0 065 0 087 0 108 0 118 0 106 0 084 0 063 0 047 2 0 0350

Q 3 5 0 0436 0 058 0 072 0 085 0 090 0 083 0 070 0 055 0 0434 w w 4 0 0 0400 0 051 0 061 0 069 0 072 0 068 0 059 0 0486 0 0392 0 u > CO

4 5 0 0366 0 0443 0 052 0 057 0 060 0 057 0 050 0 0427 0 0353 0 u > CO 5 0 0 0390 0 044 5 0 0487 0 050 0 0481 0 0435 0 0377

1 S 5 0 0387 0 0418 0 0429 0 0413 0 0378

H 6 0 0 0361 0 0370 0 0358

- 5 - 4 - 3 - 2 - 1 0 1 2 3 4 5

AXIAL DISPLACEMENT (cm)

Gamma radiation (Almost exclusively due to fission products in encapsulated sources. Approximate near-equil ibrium values days or weeks af ter encapsulat ion).

Photon emission rate 1 - 3 x 10 7 - s" 1 - j ig" 1

Tissue kerma rate in free air a t 1 m 0 . 1 1 - 0 .14 mrad-h" 1 - fig"1

(The upper l imit is for a source with negligible wall thickness. The lower figure is for a source encapsulated in 0 . 7 m m plat inum-irr idium a l loy . Values for other wall thicknesses larger than about 0 . 2 m m can be approximated by correcting the lower figure by about 1.5% for each 0 . 1 m m of Pt-Ir above or below 0 .7 m m . )

G a m m a radiation spectrum See Table III

Beta radiation

Encapsulated ! 5 zCf sources emi t beta radiation which can penetrate ma t t e r through thicknesses up to about 1 g - c m " z . The importance of this radiation depends greatly on encapsulat ion. With an af ter loading ce l l having a 0 .25 c m Pt-Ir wall and a GM counter having a 0 . 3 g - c m " 2 wall the observed counting rates due to beta and gamma radiation may be approximately equa l . If the wall thickness of the source is 0 . 7 m m Pt-Ir the beta ray counting rate is reduced by about a factor of 5. In all experiments the 2 5 2 Cf sources should be employed with a shield of a luminium to effect ively e l imina te beta radiat ion. The required thickness should be determined exper imental ly (see Chapter 13).

Depth dose distributions in infinite t issue-equivalent phantom

Tables IV and V show the distribution of absorbed neutron and gamma ray dose around applicator tubes of 1 . 5 c m active length and 0 . 7 m m Pt-Ir walls .

Corresponding data for other sources can be obtained by applying encapsulation corrections for g a m m a rays as given above and by uti l izing procedures described by Colvett e t a l . (1972). In most cases these corrections will amount to less than lO1?».

Appendix II

D E S C R I P T I O N OF CALIFORNIUM-252 SOURCES AND T H E P R O T O T Y P E USE AND STORAGE F A C I L I T Y

by I . A . L e r c h and J . Ha ide r

The ava i l ab i l i ty of s m a l l encapsu la ted s o u r c e s of c a l i f o r n i u m - 2 5 2 fo r u s e in educat ion, r ad i a t i on t h e r a p y and indus t ry , ha s r e s u l t e d in a s ign i f i can t s imp l i f i c a t i on of a v a r i e t y of t a s k s . Many of the e x p e r i m e n t s and s tud ie s e n u m e r a t e d in th is book would r e q u i r e a c c e s s to complex and expens ive f ac i l i t i e s in the absence of su i t ab le s o u r c e s .

The p u r p o s e of th is appendix i s to d e s c r i b e b r i e f l y the 252Cf s o u r c e s lent by the USÁEC to the IAEA for l imi ted d i s t r ibu t ion by the D o s i m e t r y Sect ion of the Divis ion of L i f e S c i e n c e s . In addit ion, a s i m p l e and inexpens ive pro to type fac i l i ty i s d e s c r i b e d in de ta i l which i s adequate fo r both the s t o r a g e and e x p e r i m e n t a l u s e of a p p r o x i m a t e l y 10 jag equivalents of the r ad ionuc l ide . It was the obl igat ion of the Agency to give guidance to po ten t i a l u s e r s on the f a b r i c a t i o n of a u s e and s t o r a g e fac i l i ty which would be inexpens ive and r e l a t i v e l y e a s y to build yet s a f e in u s e .

1. IAEA 252Cf SOURCES

1 . 1 . Af t e r load ing ce l l (ALC) and s h o r t a f t e r l oad ing ce l l (SALC)

Both ce l l s , the a f t e r l o a d i n g ce l l (ALC) and the s h o r t a f t e r load ing ce l l (SALC), a r e p r e p a r e d in the s a m e way. The c a l i f o r n i u m - 2 5 2 is e l e c t r o -depos i ted as Cf(OH)3 f r o m a n i t r a t e so lu t ion onto a p la t inum w i r e . The w i r e i s hea ted in a i r to conver t the Cf(OH)3 to CÎ203, a m o r e s t ab le f o r m . The w i r e i s doubly encapsu la ted in P t - 1 0 wt% Ir capsu le s and sea led by fus ion welding. Both ALC and SALC s o u r c e s m e e t ' s p e c i a l f o r m 1

s p e c i f i c a t i o n s . The s o u r c e d i m e n s i o n s , nomina l s o u r c e s t r eng th , and u n i f o r m i t y of 252Cf d i s t r i bu t ion a r e l i s t ed in Tab le VI.

1 . 2 . Radionucl ide content

T h e s o u r c e s wil l conta in r e p u r i f i e d 252Cf. An ana lys i s of the c a l i f o r n i u m i so tope content depos i ted in s o m e

r e c e n t l y p r e p a r e d s o u r c e s i s given in Tab le VII. The data have been taken f r o m " D e s c r i p t i o n of Cf -252 S o u r c e s " , Rev . 2 / 8 / 7 1 , U . S . Atomic E n e r g y C o m m i s s i o n document .

1 . 3 . Cons t ruc t i on

D e s i g n de ta i l s of the s o u r c e c a p s u l e s a r e given in F i g s 47 and 48. The w i r e i s i n s e r t e d into a c lose- f i t t ing P t - 1 0 wt% Ir ce l l which i s

s ea l ed with a P t - I r a l loy plug by a fus ion welding p r o c e s s . The cel l i s

119

1 2 0 APPENDIX II

TABLE VI. DETAILS OF ALC AND SALC Data obtained f r o m Church i l l Hospi ta l , Headington, Oxford, UK

ALC SALC

Active length 30 .0 ± 0 . 8 m m 15 .0 ± 0. 8 m m

External length 33 .0 ± 1 . 0 m m 18.0 ± 1 . 0 m m

External d iameter 0 . 9 5 ± 0 .10 m m 0 . 9 5 ± 0 .10 m m

Mean source strength 1 .90 ng ± 5°/o 1 . 0 1 (ig ± 5<fo (as of 1 May 1972) (as of 1 May 1972)

Source uniformity ± 15<7o ± 15%

TABLE VII. ANALYSIS OF CALIFORNIUM

Isotope At. ft

249 0 . 9

250 11 .2

251 3 . 1

252 84 .2

253 0 . 5

254 0 . 0 4 99 .94

enc losed in a capsu le and c losed with a second P t - I r plug for the a f t e r l o a d i n g c e l l s . The a f t e r l o a d i n g ce l l s a r e sea l ed in c apsu l e s by a fus ion welding p r o c e s s .

A combina t ion X- and au to r ad iog raph of an ALC s o u r c e i s given in F i g s 49a (en la rged view) and 49b (actual s i z e ) .

Addi t ional de t a i l s conce rn ing m e c h a n i c a l p ro to type t es t ing , qual i ty con t ro l and r e s u l t s of r ad i a t i on t e s t ing a r e ava i lab le f r o m the C a l i f o r n i u m - 2 5 2 I n f o r m a t i o n C e n t r e , Savannah Rive r L a b o r a t o r y , Aiken, South C a r o l i n a 29801, USA.

2. P R O T O T Y P E USE AND STORAGE FACILITY

The l e a s t expens ive and m o s t widely ava i lab le r e l a t i v e l y e f f i c ien t f a s t -neu t ron sh ie ld ing m a t e r i a l i s w a t e r . Based upon th is a s sumpt ion , a s i m p l e des ign fo r a u s e and s t o r a g e fac i l i ty capable of a ccommoda t ing a p p r o x i m a t e l y 10 ßg equiva len ts of 252Cf was developed. Addi t ional des ign c r i t e r i a w e r e e s t ab l i shed which governed the f inal f o r m of the p ro to type fac i l i ty . The e l e m e n t s of the des ign c r i t e r i a a r e :

S impl ic i ty

A c c e s s to the i r r a d i a t i o n c h a m b e r m u s t be accompl i shed without the d a n g e r of e x c e s s i v e e x p o s u r e

SOURCES AND FACILITY 1 2 1

Smal l cos t

The f ac i l i t y m u s t be f lex ib le enough to employ as few as 4 o r as m a n y a s 12 s o u r c e s

Loading o r r e m o v a l of s o u r c e s m u s t be s i m p l e and s a f e

The i r r a d i a t i o n c h a m b e r m u s t be in the f o r m of a cy l inde r and have d i m e n s i o n s of a p p r o x i m a t e l y 10 cm height and 10 c m d i a m e t e r and su f f i c i en t sh ie ld ing m u s t be provided for s t o r i n g the f ac i l i ty in a con t ro l led a r e a .

Consp icuous ly absen t f r o m th i s l i s t is a s t r i n g e n t r e q u i r e m e n t that the i r r a d i a t i o n c h a m b e r have a u n i f o r m neut ron and g a m m a f ie ld . The l imi t ed n u m b e r of s o u r c e s and the n a t u r e of a fac i l i ty des igned fo r d idac t ic p u r p o s e s did not appea r to w a r r a n t such a s e v e r e condit ion.

With t h e s e g e n e r a l cons ide r a t i ons e s t ab l i shed , it was dec ided to b a s e the c o n s t r u c t i o n of the f ac i l i ty on an oil d r u m (200 l i t r e o r 55 gal lon type) .

2 . 1 . D e s i g n and cons t ruc t i on de ta i l s

F i g s 50a - i i l l u s t r a t e the cons t ruc t ion de ta i l s fo r the p ro to type u s e and s t o r a g e fac i l i ty . The m a j o r components of the con ta ine r a r e the 2 0 0 - l i t r e oil d r u m , the s t e e l c e n t r a l i r r a d i a t i o n column, the f ixed and movab le p l a s t i c sh i e lds and the p l a s t i c s o u r c e a s s e m b l y . Po lyv iny l ch lo r ide (PVC) was exc lus ive ly u s e d in the cons t ruc t ion of the p ro to type fac i l i ty al though polyethylene o r o the r equivalent p l a s t i c s a r e e n t i r e l y su i t ab l e . P e r s p e c t i v e views of the fac i l i ty as cons t ruc t ed by the IAEA a r e given in F i g s 50a -d .

2 . 2 . Shielding

Shielding e s t i m a t e s w e r e m a d e with the C a l i f o r n i u m - 2 5 2 Shielding Guide by Stoddard and Hoo tman (1971). By a s s u m i n g a s i m p l i f i e d g e o m e t r y and ignor ing the s u r p l u s sh ie ld ing provided by the c e n t r a l i r r a d i a t i o n co lumn and s t e e l wal l of the con ta ine r , a dose equivalent r a t e of l e s s than 70 m r e m / h at the s u r f a c e of the midd le of the con ta ine r and l e s s than 9 m r e m / h on the top s u r f a c e a r e obta ined . Thus , i t i s c l e a r f r o m t h e s e r a t h e r c r u d e l y c o n s e r v a t i v e e s t i m a t e s that the f ac i l i ty m e e t s the s a f e t y c o n s t r a i n t s imposed in C h a p t e r 11. The e l e m e n t s of t h e s e sh ie ld ing e s t i m a t e s m a y be s u m m a r i z e d a s fo l lows:

(a) A lO-jug 252Cf point s o u r c e i s sh ie lded by 23.75 cm of w a t e r at the con ta ine r mid l ine and 38.5 cm of wa te r at the top s u r f a c e . Thus the g e o m e t r i c d i s t r ibu t ion of the s o u r c e s i s i gno red .

(b) The addi t ional sh ie ld ing of the s t ee l conta inment v e s s e l and c e n t r a l co lumn and the PVC s o u r c e h o l d e r s is i g n o r e d .

(c) The m o r e e f f i c ien t a t tenuat ion of the neu t ron flux provided by the PVC fixed and moving shie ld a s s e m b l i e s i s ignored by a s s u m i n g a homogeneous w a t e r sh ie ld .

Howeve r , sh ie ld ing m u s t be provided which p r e v e n t s dose equiva len ts in e x c e s s of 100 m r e m / w e e k to r ad ia t ion w o r k e r s . Th i s s a f e t y r e q u i r e m e n t

1 2 2 APPENDIX II

TABLE VIII. ESTIMATED AND MEASURED DOSE EQUIVALENT RATES

Estimated dose equivalent rate

(mrem/h)

At midl ine surface of the container

Tota l neutron component 31 .0

Tota l gamma component 1 5 . 3

Total dose equivalent 4 6 . 3

At top surface of the container

Tota l neutron component 2 . 2

Tota l gamma component 3 . 1

Tota l dose equivalent 5 . 3

At 1 met re from middle surface of container in air

To ta l neutron component 1 . 0 3

Total g a m m a component 0 . 5 1

To ta l dose equivalent 1 .54

Measured dose equivalent rate

(mrem/h)

4 . 5

4 . 5

9

0 . 7 5 with movable shields in down position and 1 . 5 with movable shields in up position

1 with movable shields in down position and 1 . 1 with movable shields in up position

1 .7 with movable shields in down position and 2 . 6 with movable shields in up position

0 . 7

0 . 8

1 . 5

i s de r ived f r o m IAEA and ICRP r e c o m m e n d a t i o n s but m a y be m o r e o r l e s s s t r i n g e n t in o ther coun t r i e s in a c c o r d a n c e with loca l l eg i s l a t ion . Thus , the ac tua l sh ie ld ing r e q u i r e m e n t s wi l l depend not on the dose equivalent at the s u r f a c e of t he con ta ine r , but the dose equivalent r ece ived by the r a d i a t i o n w o r k e r p r o x i m a l to the fac i l i ty . T h i s dose equivalent wil l be dependent upon the dose equivalent r a t e at s o m e d i s t ance f r o m the con ta ine r and the amount of the to ta l work ing t i m e spent by the u s e r in the v ic in i ty of the con t a ine r . Most i m p o r t a n t : sh ie ld ing e s t i m a t e s m u s t be ve r i f i ed by ac tua l m e a s u r e m e n t with p r e c i s e l y ca l i b r a t ed r ad i a t i on p ro tec t ion s u r v e y d o s i m e t e r s .

S u p p l e m e n t a r y sh ie ld ing m a y be n e c e s s a r y for the p ro t ec t i on of w o r k e r s sub jec t to pro longed phys i ca l contac t with the c o n t a i n e r . In th is eventual i ty , s i n c e roughly 5% of the to ta l dose equivalent i s due to c ap tu r e g a m m a r a y s at the midd le s u r f a c e of the con ta ine r and 30% of the to ta l at the top s u r f a c e , the addit ion of b o r o n to a concen t r a t i on of a p p r o x i m a t e l y 3 m g / c m 3

in the w a t e r shie ld wi l l r e d u c e th i s s o u r c e of g a m m a r ad i a t i on by a p p r o x i m a t e l y 1 / 3 . However , the to ta l g a m m a component due to both

SOURCES AND FACILITY 1 2 3

p r o m p t and cap tu re r e a c t i o n s wil l account for a p p r o x i m a t e l y 40% of the dose equivalent at the midd le s u r f a c e and 80% at the top s u r f a c e . The addi t ion of lead sh ie ld ing r e q u i s i t e to ach ieve a s a t i s f a c t o r y r educ t ion of the dose equivalent r a t e would not pose a s e r i o u s d i f f icu l ty . F o r example , roughly 3.5 cm of lead would r e d u c e the to ta l g a m m a dose equivalent r a t e by a f a c t o r of 1 /10 .

To t e s t the a c c u r a c y of th i s e s t i m a t e of sh ie ld ing e f f i cacy , the p ro to type con ta ine r was loaded with 12 s o u r c e s containing 7. 6 Mg of 262Cf (± 5% s t a n d a r d devia t ion of the mean ) . Shielding was prov ided by a p p r o x i m a t e l y 200 l i t r e s of wa t e r into which bo ron acid (borax) was d i s so lved in su f f i c i en t quant i ty to ach ieve a concen t r a t ion of 5% boron by weight . Tab le VIII conta ins a c o m p a r i s o n of the e s t i m a t e d and m e a s u r e d dose equivalent r a t e s .

Thus , the method fo r e s t ima t ing the dose equivalent r a t e is c o n s e r v a t i v e and p r o v i d e s an adequate m a r g i n for s a f e ty p rov ided that c a r e i s e x e r c i s e d in the des ign and f a b r i c a t i o n of the con ta ine r .

A note of caution m u s t be sounded in echo of Chap t e r 11 concern ing w a t e r as sh ie ld ing m a t e r i a l . P r o v i s i o n s m u s t be m a d e to cont inuously mon i to r the leve l of the w a t e r in the con ta ine r . A m e a n s fo r checking the w a t e r content i s m a d e by the addi t ion of a cove red a c c e s s hole in the top s u r f a c e . Since the con ta ine r cover i s t ight ly f i t ted , however , the l o s s of w a t e r due to evapora t ion wi l l be s m a l l but checks should be p e r f o r m e d weekly .

In the adoption of a given des ign for a u s e and s t o r a g e fac i l i ty , the r e s p o n s i b l e s c i en t i s t m u s t be c a r e f u l to f a m i l i a r i z e h imse l f with the r e c o m m e n d a t i o n s of the ICRP, the IAEA and the laws of h i s own count ry govern ing the s a f e handl ing of r ad ionuc l i de s . In a l l c a s e s , the IAEA s u p p o r t s the s t r i c t e s t and m o s t c o n s e r v a t i v e r egu la t i ons govern ing r a d i a -t ion p ro tec t ion p r o c e d u r e s compa t ib le with the u s e of r ad ionuc l ides for t r a i n i n g .

2 . 3 . U s e of the fac i l i ty

The conta iner was a r b i t r a r i l y des igned to include a 1 0 - c m o. d. c e n t r a l s t e e l co lumn on which the s o u r c e h o l d e r s a r e mounted and within which the movab le shield a s s e m b l y and i r r a d i a t i o n c h a m b e r a r e contained. In the c losed posi t ion, the c e n t r a l co lumn is sh ie lded by the movable PVC plug above and below the i r r a d i a t i o n c h a m b e r . In the extended pos i t ion ( see F i g s 51a and 51b), t he uppe r and lower movab le p lugs and the i r r a d i a t i o n c h a m b e r s l ide as a s ing le unit unt i l the lower movable plug and the f ixed bot tom plug r e s p e c t i v e l y ac t as the uppe r and lower column s h i e l d s .

In the event of it becoming n e c e s s a r y to i n t roduce e l ec t ron i c components into the i r r a d i a t i o n c h a m b e r , l e a d s m a y be put th rough the uppe r movab le plug if c a r e is t aken to ma in t a in the in t eg r i ty of the sh ie ld ing .

As noted in the p r e v i o u s s e c t i o n s , the f ac i l i ty i s des igned to a c c o m m o -da te a m a x i m u m of 12 ALC o r SALC s o u r c e s . However , 4 o r 6 s o u r c e s m a y a l so be employed equ id i s tan t ly about t he c e n t r a l co lumn. F u r t h e r , the s o u r c e s m a y be mounted e i t h e r p r o x i m a l to the c e n t r a l s t e e l co lumn o r r e v e r s e d for the p u r p o s e of conducting inves t iga t ions in the w a t e r sh i e ld . The s o u r c e h o l d e r s a r e fash ioned fo r e a s y r e m o v a l in the event of t h e i r be ing employed outs ide the c o n t a i n e r .

1 2 4 APPENDIX II

A S S E M B L Y S H E A T H

SHEATH

CELL-

1.271 in.

0.038 in.

L0.025 in.

1.221 in.

0.026 in.

0.006 in. WALL-*!

0.02 S in.

-0.006 in.WALL

C A P S U L E I 0.020 in.

0.016 in.-

0.004 in. WAIL •

1.181 in.

1.221 in.

~ r 0.020 in.

0.004 in. - WALL

- 0 024 in. -0.038 in.

F I G . 4 7 . A f t e r l o a d i n g c e l l , m o d e l ALC.

A S S E M B L Y

SHEATH

0.681 in.

0.038 in.

S H E A T H 0.025 in.

0.631 in.

0.681 in.

0.026 in.-

0.006 in.WALL1

C A P S U L E 0.020 in.

0.025 in. 0.016 in.-

0.004 in. WALL -

- 0.006 in.WALL

- 0.038 in.

F I G . 4 8 . A f t e r l o a d i n g c e l l , m o d e l SALC.

0.631 in.

0.059 in.

0.020 in.

0.004 in. - WALL

0.024 in.

SOURCES AND FACILITY 1 2 5

FIG.49. ALC sources, (a) Enlargement of combination X-ray and autoradiograph illustrating the distribution of activity relative to the total dimensions of the needle (radiograph courtesy of Dr. Madvanath, Bhabha Atomic Research Centre, Bombay, India), (b) Actual size contact print of (a).

1 2 6 APPENDIX II

FIG.50a. Inserting a radiation dosimeter into the irradiation chamber of the container. Approxi-mate ly 10 |jg of 2 K Cf are employed in the faci l i ty which contains 200 litres of borated water to serve as shielding. Movable plastic shields permit access to the irradi-ation chamber without danger to the workers handling the fac i l i ty .

FIG.50b. A t issue-equivalent plastic condenser ionization chamber is shown in p lace in the irradiation chamber . Above and below the chamber are movable Polyvinylchloride plugs which shield the radiation workers during the lowering and. withdrawal of the assembly into and out of the container .

(b )

SOURCES AND FACILITY 1 2 7

FIG.50c. The top of the container has been removed to illustrate the emplacement of the array of radiation sources around the central irradiation column. The sources, in the form of small needles, are inserted into plastic holders which are manipulated with long plastic rods. Twelve such sources are placed around the circumference of the central column after the container has been fi l led with borated water (to provide visual clarity, the water is not in the tank).

FIG.50d. This detail photograph illustrates the construction of the source holder and the manipulating rod which is easily disassembled. There is no radiation source in the holder in this photograph.

1 2 8 APPENDIX II

TOP VIEW OF STEEL TANK (WITHOUT COVERING PLATE AND WITHOUT TRANSPORT CARRIAGE)

FIG.50e . Overall drawing of the prototype Californium Use and Storage Facili ty illustrating the p lacement of the de ta i l drawings Figs 50f - i .

SOURCES AND FACILITY 1 2 9

FIG. 50f. Detai l drawing of the top portion movable upper shielding cylinder, the central steel column and the a luminium support ring.

1 3 0 SOURCES AND FACILITY 130

UPPER SHIELDING CYLINDER

(TROVIDUR- PVC)

SOURCE HOLDER WITH

PVC HANDLE IN PLACE ON

PVC RING-

DETAIL CS

CROSS SECTION : E - F

FIG.50g. Deta i l drawing of the central portion of the irradiation chamber in the closed position, the source holders in position in the source holder ring and the support structures which affix the upper and lower movable cylinders.

SOURCES AND FACILITY 1 3 1

DETAIL O

SOURCE HOLDER

CROSS SECTION: C - D

100*

WATERTIGHT WELDED SE AI

DETAIL 0

STAINLESS STEEL TUBE

FIXED BOTTOM SHIELDING CYLINDER

IPVCI

WOODEN SPACER

•AI PLATE

F I G . 50h . D e t a i l d rawings of the source c a r r i e r r ing and the b o t t o m of the c o n t a i n e r and p l a t f o r m a s s e m b l y .

1 3 2 APPENDIX II

DETAIL GO

I i i

I I

y

b) SAMPLE FOR ALC NEEDLES

a) SAMPLE FOR SftLC NEEDLES

FIG. 50i. Detai l drawings of source holders for both ALC and SALC sources.

R E F E R E N C E S

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BARENDSEN, G. W., (1968), in: Current Topics in Radiation Research, Vol. 4 (EBERT, M . , HOWARD, A . , Eds), Amsterdam, North Holland Publishing Co. , 293-356.

BARENDSEN, G. W., KOOT, C . J . , KERSEN, G.R. , BEWLEY, D . K . , FIELD, S .B. , PARNELL, C. J. (1966), The effect of oxygen in impairment of the proliferative capacity of human cells in culture by ionizing radiations of different LET, Int. J. Radiat. Biol. U , 317-327.

BENGTSSON, L . G . , GOODMAN, L . I . , ROSSI, H.H. (1969), Gas multiplication in air-fi l led ionization chambers, Health Phys. 16, 756-758.

BOND, V.P. , FLIEDNER, T . M . , ARCHAMBEAU, J. O. (1965), Mammalian Radiation Lethality. A Disturbance in Cellular Kinetics, Academic Press, New York.

BROERSE, J. J . , BARENDSEN, G. W., KERSEN, G.R. (1967), Survival of cultured human cells after irradiation with fast neutrons of different energies in hypoxic and oxygenated conditions. Int. J. Radiat. Biol. 13, 559

COLVETT, R .D. , ROSSI, H . H . , KRISHNASWAMY, V. (1972), Dose distribution around a californium-252needle, Phys. Med. Biol. JL7, 356-364.

COURTENAY, V. D. (1965), The response to continuous irradiation of the mouse lymphoma L51784 grown in vitro, Int. J. Radiat. Biol. 9, 581-592.

DICELLO, J . F . , GROSS, W. , KRALJEVIC, U. (1972), Radiation quality of californium-252, Phys. Med. Biol. 17, 345-355.

DJORDJEVIC, B., ANDERSON, L. L. , KIM, S.H. (1973), Oxygen enhancement ratios in HeLa cells irradiated with californium and radium sources, Radiology 107 , 429-434.

EBERT, M . , BARBER, D.A. (1961), The effects of X-rays on the extension growth of barley roots, Int. J. Radiat. Biol. 3, 587-593.

ELKIND, M . M . , SUTTON, H. (1959), (1967), X-ray damage and recovery in mammalian cells in culture, Nature (London), 184, 1293-1295.

ELKIND, M.M. , SUTTON, H. (1960), "Radiation response of mammalian cells grown in culture. 1. Repair of X-ray damage in surviving Chinese hamster cells ." Radiat. Res. 13, 556-593.

ELKIND, M . M . , SWAIN, R. W., ALESCIO, T . , SUTTON, H . , MOSES, W. B. (1963), in: Cellular Radiation Biology, Baltimore, Williams and Wilkins Publishing Co . , 442-461.

ELKIND, M . M . , WH1TMORE, G.F. (1967), The Radiobiology of Cultured Mammalian Cells, Gordon and Breach, New York.

FINNEY, D.J. (1952), Statistical Method in Biological Assay, Charles Griffin, London.

GOODMAN, L.J. (1969), A modified tissue equivalent liquid. Health Phys. 16, 763.

GRAY, L. H . , READ, J. (1942a), The effect of ionizing radiations on the broad bean root. Part 1. General notes, Brit. J. Radiol. 15, 11-16.

GRAY, L. H. , READ, J. (1942b), The effect of ionizing radiations on the broad bean root. Part n . The lethal action gamma radiation, Brit. J. Radiol. 15, 39-42.

GRAY, L . H . , READ, J. (1942c), The effect of ionizing radiations on the broad bean root. Part III. The lethal action of neutron radiation, Brit. J. Radiol. 15, 72-76.

GRAY, L. H . , READ, J. (1942d), The effect of ionizing radiations on the broad bean root. Part IV. The lethal action of alpha radiations. Brit. J. Radiol. 15, 320-336.

GRAY, L .H. , SCHOLES, M.E. (1951), The effect of ionizing radiations on the broad bean root. Part VIII. Growth rate studies and histological analysis, Brit. J. Radiol. 24 , 82, 176 , 228 , 285 , 348.

HALL, E.J. (1961), The relatice biological efficiency of X-rays generated at 250 kVp and gamma radiation from a cobalt 60 therapy unit, Brit. J. Radiol. 34, 313.

133

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HALL, E.J. ((1972), A comparison of radium and cal i fornium-252 using cultured m a m m a l i a n cells, Radiology 102, 173-179.

HALL, E . J . , BEDFORD, J. S. (1964a), A comparison of the effects of acute and protracted gamma irradiation on the growth of seedlings of Vicia faba, Int. J. Radiat. Biol. 8, 467.

HALL, E.J. , BEDFORD, J. S. (1964b), Dose-rate , its e f f ec t on the survival of HeLa cells irradiated with gamma rays, Radiat. Res. 22, 305-315.

HALL, E . J . , CAVANAGH, J. (1967), The oxygen e f fec t for acute and protracted radiation exposures measured with seedlings of Vicia faba , Brit. J. Radiol. 40, 128.

HALL, E. J . , FAIRCH1LD, R. G. (1979), Radiobiological measurements with californium 252, Brit. J. Radiol. 43, 263-266.

í HALL, E. J . , LAJTHA, L. G. (1963), The recovery of Vicia faba mer is tem cells from X-radia t ion, Radiat. Res. 20, 187-194.

HALL, E . J . , LEHNERT, S. (1973), The biophysical properties of 3.9 GeV nitrogen ions. IV. OER and RBE determinat ions using cultured m a m m a l i a n cells, Radiat. Res. 55, 431-436.

HALL, E. J. , ROIZIN-TOWLE, L.A. , COLVETT, R. D. (1974), RBE and OER determinat ions for radium and ca l i fornium-252, Radiology 110, 699-704.

HALL, E . J . , ROSSI, H . H . , KELLERER, A . M . , GOODMAN, L. , MARINO, S. (1973), Radiobiological studies with monoenerge t ic neutrons, Radiat. Res. 54, 431-433.

HALL, E . J . , ROSSI, H . H . , ROIZIN, L. A. (1971), Low dose-ra te irradiation of m a m m a l i a n cells with radium and ca l i fornium-252, Radiology 99, 445-451.

HAM, R. G . , Puck, T . T. (1962), "Quanti tat ive colonial growth of isolated m a m m a l i a n cells, " in: Methods in Enzymology, Vol. V (COLSWICK,S. P., KAPLAN, N. O . , Eds), Academic Press, New York.

HERING, E .R . , SHEPSTONE, B.J. (1972), The ef fec t of irradiation of 'Vicia f aba ' roots with 1.5 mHz ultrasound. Brit. J. Radiol. 45, 786-787.

HEWITT, H. B. , WILSON, C. W. (1959), A survival curve for m a m m a l i a n cells irradiated in vivo, Nature, (London)183, 1060-1061.

HINE, G. J . , BROWNELL, G.L. (1956), Radiation Dosimetry, 1st Edn, Academic Press, New York.

HOLTHUSEN, H. (1921), Beiträge zur Biologie der Strahleneinwirkung. Untersuchungen an Askarideneiern, Pfluegers Arch. f . d . ges. Physiol. 187, 1 -24 .

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HOWARD, A . , PELC, S. R. (1953), Synthesis of desoxyribonucleic acid in normal and irradiated cells and its relation to chormosome breakage, Heredity Supplement , pp. 261-273.

ICRP Publication 9 (1965), Recommendations of the Internat ional Commission on Radiological Protection, Int. C o m m . on Radiol. Prot . , Pergamon Press, New York.

ICRU Report 10b (1964), Physical Aspects of Irradiation, Int. Comm. on Rad. Units and M e a s . , National Bureau of Standards Handbook 85.

ICRU Report 13 (1969), Neutron Fluence, Neutron Spectra and Karma, Int. Comm. on Rad. Units and M e a s . , Washington, D. C.

ICRU Report 14 (1969), Radiation Dosimetry: X-Rays and G a m m a Rays with Max imum Photon Energies between 0.6 and 50 MeV, Int . C o m m . on Rad. Units and M e a s . , Washington, D . C .

ICRU Report 16 (1970), Linear Energy Transfer, Int. Comm. on Rad. Units and M e a s . , Washington, D . C .

ICRU Report 19 (1971), Radiation Quanti t ies and Units, Int. C o m m . on Rad. Units and M e a s . , Washington, D . C .

ICRU Report 20 (1971), Radiation Protection Instrumentation and its Applicat ion, Int. Comm. on Rad. Units and M e a s . , Washington, D. C.

JOHNS, H. E. (1964), The. Physics of Radiology, Charles C. Thomas, Springfield, Illinois.

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LAJTHA, L. G . , OLIVER, R. (1961), Some radiobiological considerations in radiotherapy, Brit. J. Radiol. 34, 252-257.

LAMERTON, L. F . , COURTENAY, V. D. (1968), "The steady state under continuous irradiat ion, " in: Dose-rate in M a m m a l i a n Radiation Biology (United States Atomic Energy Commission, Division of Technical Information, Conference 680410), pp. 3 .1 -3 .12 .

LANGHAM, W. H . , Ed. (1967), Radiobiological Factors in Manned Space Flight. Report of the Space Radiation Study Panel of the Life Sciences Commi t t ee , Nat . Acad. Sei. - Nat . Res. Council Publ. No. 1487.

McCULLOCH, E. A . , TILL, J. E. (1962), The sensitivity of cells from normal mouse bone marrow to g a m m a radiation in vitro and in vivo, Radiat. Res. 16, 822-832.

MEREDITH, W.J. (1967), Radium Dosage: The Manchester System, compiled from art icles by Ralston Paterson (and others) edited by W. J. Meredi th. 2nd Edn, Williams and Wilkins, Balt imore, Md.

MICHAEL, B . D . , ADAMS, G. E . , HEWITT, H. B. , JONES, W. B. G . , WATTS, M. E. (1973), The t ime scale of the oxygen e f fec t in irradiated bac ter ia : a sub-mill isecond fast moving study, Radiat. Res. 55, 553.

MOTTRAM, J. C. (1913), On the act ion of beta and g a m m a rays of radium on the cell in d i f ferent states of nuclear division, Rep. Cancer Labs. Middx. Hosp. , 30, 98.

NCRP Report 38 (1971), Protection Against Neutron Radiation, Nat. Council on Rad. Prot, and Meas. , Washington, D. C.

NCRP Report 39 (1971), Basic Radiation Protection Cri ter ia , Nat . Council on Rad. Prot, and M e a s . , Washington, D. C.

NEARY, G. J. (1957), Dependence on oxygen and tempera ture of the sensitivity of broad bean roots to g a m m a radiation, Nature (London) 180, 248.

NIAS, A. H.W. , HOWARD, A . , GREEN, D . , MAJOR, D. (1973), "The response of Chinese hamster (ovary) cells to protracted irradiation from 2 s C f and 6 t o , Brit. J. Radiol. 46, 991-995.

NIAS, A.H.W. , LAJTHA, L. G. (1964). Continuous irradiation with tritiated water of m a m m a l i a n cells in a monolayer , Nature (London) 202, 613-614.

PETRY, E. (1923), Zur Kenntnis der Bedingungen der biologischen Wirkung der Röntgenstrahlen, in . Wirkung von Oxydationsmitteln auf Empfindlichkeit , Biochem. Z . , 135, 353.

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READ, J. (1952a), The e f fec t of ionizing radiations on the broad bean root. Part X. The dependence of the X-ray sensitivity on dissolved oxygen, Brit. J. Radiol. 25, 89-160.

READ, J. (1952b), Mode of addition of X-ray doses given with d i f ferent oxygen concentrations, Brit. J. Radiol. 25, 336-338.

READ, J. (1959), The Radiation Biology of Vicia f aba , Blackwell Scient i f ic Publications, Oxford.

ROSSI, H .H . (1967), "Energy distribution in the absorption of r ad ia t ion , " in: Advances in Biological and Medica l Physics, Vol. 11, Academic Press, New York, 27-85.

ROSSI, H .H . (1968), "Microscopic energy distribution in irradiated mat te r , " in: Radiation Dosimetry, 2nd Edn, Vol. I . , Academic Press,, New York, 43-92 .

ROSSI, H . H . , COLVETT, R. D. (1972), An ionizat ion chamber with high spatial resolution, Phys. Med. Biol. 17, 227-231.

ROSSI, H . H . , FAILLA, G. (1956), Tissue-equivalent ionizat ion chambers, Nucleonics 14, 32-37.

ROSSI, H. H . , ROSENZWEIG, W. (1955), A device for the measurement of dose as a function of specif ic ionizat ion, Radiology 64, 404-410.

ROTBLAT, J . , LIN DOP, P. (1961), Long term effects of a single whole-body exposure of m i c e to ionizing radiations, Proc. R. Soc. (London) 154, 332.

RUGH, R. (1962), Low levels of X-irradiat ion and the early m a m m a l i a n embryo. Am. J. Roentgenol . , Radium T h e r . , Nucl. Med. 87, 559-566.

RUSSELL, W.L. (1963a), Genet ic hazards of radiat ion, Proc. Am. Phil. Soc. 107, 11-17.

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RUSSELL, W.L. (1963b), "The effects of radiation dose-ra te and fract ionat ion on mutation in m i c e , " in: Repair of Radiation Damage,(SOBELS, F . H . , Ed. ) Pergamon Press, New York, 205-217.

RUSSELL, W.L. (1965), Effect of the interval between irradiations and conception on mutation frequency in f ema le m i c e , Proc. Nat. Acad. Sei. 54, 1552-1556.

RUSSELL, L. B. , RUSSELL, W. L. (1954), An analysis of the changing radiation response of the developing mouse embryo, J. Cel l . Comp. Physiol. Suppl. 1, 43, 103-149.

SHELLABARGER, C. J . , CRONKITE, E. P . , BOND, V. P. , LIPPINCOTT, S .W. (1957), The occurrence of m a m m a r y tumors in the rat af ter sublethal whole-body irradiation, Radiat. Res. 6, 501-512.

SINCLAIR, W. K. (1968), Cyc l ic X-ray responses in m a m m a l i a n cells in vitro, Radiat. Res. 33, 620-643.

SINCLAIR, W.K. (1969), " T i m e dose and fract ionat ion in radiobiology as applied to radiotherapy, " in: Proceedings of the Carmel Conference on T i m e and Dose Relationships in Radiation Biology as Applied to Radiotherapy, BNL Report 50203 (C-57), pp. 97-107.

SPALDING, I . F . , LANGHAM, W . , ANDERSON, E . C . (1956), The re la t ive biological effect iveness of t r i t ium beta radiation with the broad bean root (Vicia f a b a ) as a test system, Radiat. Res. 4 , 221-227.

SPALDING, I . F . , LANGHAM, W . , ANDERSON, E . C . (1958), Relative biological effectiveness of alpha radiation with the broad bean root (Vicia faba) as a test system, Radiat. Res. 8, 322-328.

SPARROW, A . H . , UNDERBRINK, A. G . , ROSSI, H .H . (1972), Mutations induced in Tradescantia by small doses of X-rays and neutrons: Analyses of dose-response curves, Science 176, 916-918.

SPENCER, W. P. STERN, C. (1948), Experiments to test the validity of the l inear R-dose mutat ion frequency relat ion in Drosophila a t low dosage, Genetics 33, 43-74 .

STODDARD, D . H . , HOOTMAN, H.E. (1971), Cal i fornium-252 Shielding Guide, AEC Research and Development Report, DP-1246.

TERAS1MA, R. , TOLMACH, L. (1963), X-ray sensitivity and DNA synthesis in synchronous populations of HeLa cells, Science 140,490-492.

UNDERBRINK, A. G . , SCHAIRER, L . A . , SPARROW, A .H . (1973), The biophysical properties of 3.9 GeV nitrogen ions, Radiat. Res. 55, 437-446.

UNDERBRINK, A. G . , SCHAIRER, L. A . , SPARROW, A .H . (1974), "Tradescantia stamen hairs: a radiobio-logical test system appl icable to chemica l mutagenesis," in : Chemica l Mutagens: Principles and Methods of their Detect ion, Vol. 3 (HOLLAENDER, A. , Ed.) Plenum Press, New York.

UPTON, A . C . (1961), The dose-response relation in radiat ion-induced cancer , Cancer Res. 21, 717-729.

VOGEL, H .H . (1972), Mammary carcinogenesis induced by fission neutrons, Radiat. Res. 51, 494.

VOGEL, H .H . (1973), Neutron induced rat m a m m a r y neoplasms, Radiat. Res. 55, 532.

WITHERS, H.R. (1967), The dose-survival relationship for irradiation of epi the l ia l cells of mouse skin, Brit. J. Radiol. 40, 187-194.

WITHERS, H. R . , ELKIND, M . M . (1970), Microcolony survival assay for cells of mouse intestinal mucosa exposed to radiation, Int . J. Radiat. Biol. 17- 261-267.

WRIGHT, E. A . , HOWARD-FLANDERS, P. (1957), The inf luence of oxygen on the radiosensitivity of m a m m a l i a n tissues, Acta Radiol. 48, 26-32.

WITHERS, H . R . , OLIVER, G. D . , GLENN, D. W. (1971), Response of mouse je junal crypt cells to low dose rate irradiation with cal ifornium neutrons or radium gamma-rays , Radiat. Res. 48, 484-494.

BIBLIOGRAPHY

GENERAL RADIOBIOLOGY TEXTS ANDREWS, J. R. (1968), The Radiobiology of Human Cancer Radiotherapy, W. B. Saunders, Philadelphia.

BACQ, Z . M . , ALEXANDER, P. (1961), Fundamentals of Radiobiology, 2nd Edn, Pergamon Press, New York.

BOND, V. P . , FLIEDNER, T . M . , ARCHAMBEAU, J. O. (1965), Mammalian Radiation Lethality: A Disturbance in Cellular Kinetics, Academic Press, New York.

CASARET, A. P. (1968), Radiation Biology, Prentice-Hall, Englewood Cliffs, N.Y.

COGGLE, J. E. (1971), Biological Effects of Radiation, Wykeham Publications (Science Series No. 34), London.

DALRYMPLE, G . V . , GAULDEN, M. E., KOLLMORGEN, G . M . , VOGEL, H . H . , Eds. (1973). Medical Radiation Biology, W. B. Saunders, Philadelphia.

HALL, E.J. (1973), Radiobiology for the Radiologist, Harper and Row, New York.

LANGHAM, N. H. , Ed. (1967), Radiobiological Factors in Manned Space Flight, Report of the Space Radiation Study Panel of the Life Sciences Committee, Nat. Acad. Sei . , Nat. Res. Council, Publ. No. 1487.

LEA, DEA (1956), Actions of Radiations on Living Cells, 2nd Edn, Cambridge University Press, Cambridge, UK.

LOUTIT. J. F. (1962), Irradiation of Mice and Men, University of Chicago Press, Chicago.

PIZZARELLO, D. J . , WITCOFSKI, R.L. (1967), Basic Radiation Biology, Lea & Febinger, Philadelphia.

SPEAR, F.G. (1953), Radiations and Living Cells, John Wiley, New York.

GENERAL PHYSICS TEXTS

ATTIX, F . H . , ROESCH, W.C. , TOCHILIN, E. (1969), Radiation Dosimetry, Vols 1,2 and 3, Academic Press, New York.

GOODWIN, P . N . , QUIMBY, E . H . , MORGAN, R. H. (1970), Physical Foundations of Radiology, Harperand Row, New York.

JOHNS, H .E . , CUNNINGHAM, J. R. (1969), The Physics of Radiology. Charles C. Thomas, Springfield, 111.

REED, C. W. , Ed. (1970), Radiation Dosimetry, Proceedings of the International School of Physics, Academic Press, New York.

PUBLICATIONS CONCERNING CALIFORNIUM

ANDERSON, L. L. (1973), Status of dosimetry for californium-252 medical neutron sources, Phys. Med. Biol. 18, 779-799.

ATKINS, H. L., FAIRCHILD, R. G. , ROBERTSON, J. S. (1970), Comparison of irradiation by Cf and radium on skin of swine, Radiology 96, 161-165.

ATKINS, H. L., FAIRCHILD, R. G. , ROBERTSON, J. S. (1972), Dose-rate effects on RBE of californium and radium radiations of pig skin, Radiology 103, 439-444.

BERRY, R. J. (1971), Hypoxic protection against fast neutrons of different energies — a review. Europ. J. Cancer T, 145-152.

BERRY, R.J.,OLIVER, G . D . , GEORGE, D.L.,WERNER, J . , BRENNAN, J .T . (1973), Survival of murine leukemia cells exposed to californium-252 neutrons under conditions simulating implantation of these sources into tissue in radiotherapy, Brit. J. Radiol. 46, 996-1004.

BUSHONG, S . C . , PRASOD, N . , BRINEY, S . A . , OLIVER, G. D. (1970), Radiocytogenic determination of oxygen enhancement ratio of californium-252, Radiology 96, 167-170.

137

138 BIBLIOGRAPHY

BDSHONG, S . C . . PRASOD, N . , BRINEY, S . A . , OLIVER, G. D. (1973), Radiocytogenetic studies with ca l i forn ium-252, Int. J. Radiat. Biol. 23, 105-112.

CASTRO, J. R. , OLIVER, G. D. , WITHERS, R . , ALMOND, P. R. (1973), Experience with cal i fornium-252 in c l in ical radiotherapy, Am. J. Roentgenol. , Radium Ther. , Nucl. Med . 117, 182-194.

COLVETT, R . D . , ROSSI, H . H . , KRISHNASWAMY, V. (1972), Dose distribution around a cal i fornium-252 needle , Phys. Med . Biol. 17, 356-364.

DEAN, F . K . , BUSHONG, S . C . , PRASOD, N . , BRINEY, S. (1972), The relat ive biological effectiveness of ca l i forn ium-252 on acu te m a m m a l i a n le thal i ty , Radiology 104, 699-703.

DJORDJEVIC, B., ANDERSON, L. L . , KIM, S. H. (1973), Oxygen enhancement ratios in HeLa cells irradiated with cal i fornium and radium sources, Radiology 107, 429-434.

DREW, R . M . , FAIRCHILD, R. G . , ATKINS, H. L. (1972), The oxygen enhancement ratio as measured with HeLa cells and protracted irradiation from 2s2Cf and 13fcs, Radiology 104, 409-413.

FAIRCHILD, R . G . , DREW, R . M . , ATKINS, H. L. (1969), Relative biological e f fec t of !52Cf radiation on HeLa cells in culture. Radiology 93, 143-147.

FAIRCHILD, R. G . , DREW, R . M . , ATKINS, H.L. (1970), Dose-rate effects for various dose-rates of 2 B Cf radiation on HeLa cells in culture, Radiology 96, 171-174.

FAIRCHILD, R . G . , DREW, R . M . , ATKINS, H.L. (1970), The oxygen enhancement ratio for protracted irradiation with Radiology 96, 661-665.

FIELDS, P. R. , et a l . (1956), Transplutonium elements in thermonuclear test debris, Phys. Rev. 102, 180.

HALL, E.J. (1972), A comparison of radium and ca l i fornium-252 using cultured m a m m a l i a n cells, Radiology 102, 173-179.

HALL, E. J. (1972), A determinat ion of the oxygen enhancement ratio of 252Cf using cultured m a m m a l i a n cells, Brit. J. Rádiol. 45, 284-288.

HALL, E.J. (1972), Radiation dose- ra te : a factor of importance in radiobiology and radiotherapy, Brit. J. Radiol. 45, 81-97 .

HALL, E. J., FAIRCHILD, R. G. (1970), Radiobiological measurements with cal i fornium-252, Brit. J. Radiol. 43, 263-266.

HALL, E . J . , ROIZIN-TOWLE, L. A. , COLVETT, R.D. (1974), RBE and OER determinat ions for radium and ca l i fornium-252, Radiology 110, 699-704.

HALL, E . J . , ROSSI, H . H . , ROIZIN, L .A. (1971), Low dose-ra te irradiation of m a m m a l i a n cells with radium and ca l i forn ium-252, Radiology 49 , 445-451.

KRISHNASWAMY, V. (1972), Calculated depth dose tables for cal i fornium-252 sources in tissue, Phys. Med. Biol. 17, 56-63 .

NIAS, A. H. W . , HOWARD, A . , GREENE, D . , MAJOR, D. (1973), The response of Chinese hamster (ovary) cells to protracted irradiation from cal i fornium-252 and cobal t -60 , Brit. J. Radiol. 46 , 991-995.

SCHLEA, C . S . , STODDARD, D . H . (1965), Californium isotopes proposed for intracavitary and intersti t ial radiation therapy with neutrons, Nature (London) 206, 1058.

THOMPSON, S. G. , STREET, K . , CHIORSO, A . , SEABORG, G. T. (1950), The new e lement cal ifornium (Atomic Number 98), Phys. Rev. 80, 790.

WITHERS, H . R . , OLIVER, G. D . , GLENN, D .W. (1971), Response of mouse je junal crypt cells to low dose-rate irradiation with cal ifornium neutrons or radium g a m m a rays, Radiat. Res. 48, 484-494.

ADDITIONAL SAFETY R E F E R E N C E S FOR APPENDIX II

JAEGER, R . G . , BLIZARD, E. P. , CHILTON, A . B . , GROTENHUIS, M. , HÖNIG, A . , JAEGER, T. A. , EISENLOHR, H .H . (1968 and 1970), Engineering Compendium on Radiation Shielding, Vols I and III, Springet' Verlag, Berlin, Heidelberg and New York.

BIBLIOGRAPHY 1 3 9

IAEA Proceedings Series (Vienna)

Neutron Monitoring for Radiation Protection Purposes (Proc. Symp. Vienna, 1972) 2 Vols, IAEA, Vienna (1973).

IAEA Safety Series (Vienna)

No. 1, Safe Handling of Radionuclides, 1973 Edn (1973) No. 6, Regulations for the Safe Transport of Radioactive Materials , 1973 Revised Edn (1973) No. 8, The Use of Film Badges for Personnel Monitoring (1962) No. 9, Basic Safety Standards for Radiation Protection, 1967 Edn (1967) No. 13, The Provision of Radiological Protection Services (1965) No. 25, Medica l Supervision of Radiation Workers (1968) No. 30, Manual on Safety Aspects of the Design and Equipment of Hot Laboratories (1969) NO. 38, Radiation Protection Procedures (1973).

CONVERSION TABLE: FACTORS FOR CONVERTING UNITS TO SI SYSTEM EQUIVALENTS*

SI base units are the metre (m), kilogram (kg), second (s), ampere (A), kelvin (K), candela (cd) and mole (mol). [For further information, see International Standards ISO 1000 (1973), and ISO 31/0 (1974) and its several parts]

Multiply by to obtain

Mass

pound mass (avoirdupois) 1 Ibm 4.536 X 10"1 kg ounce mass (avoirdupois) 1 ozm = 2.835 X 101

S ton (long) (= 2240 Ibm) 1 ton 1.016 X 103 kg ton (short) (= 2000 Ibm) 1 short ton = 9.072 X 102 kg tonne (= metric ton) 1 t 1.00 X 103 kg

Length

statute mile 1 mile = 1.609 X 10° km yard 1 yd 9.144 X 10"' m foot 1 f t 3.048 X 10"1 m inch 1 in 2.54 X 10"2 m mil (= 10~3 in) 1 mil 2.54 X 10"2 mm

Area

hectare 1 ha 1.00 X 104 m 2

(statute mile)2 1 mile2 2.590 X 10° km2

acre 1 acre = 4.047 X 103 m2

yard2 1 yd2 8.361 X 10"1 m 2

foot2 1 f t 2 9.290 X 10"2 m 2

inch2 1 in2 6.452 X 102 mm2

Volume

yard3 1 yd3 7.646 X 10"1 m3

foot3 1 f t 3 2.832 X 10"2 m3

inch3 1 in3 1.639 X 104 mm3

gallon (Brit, or Imp.) 1 gal (Brit) = 4.546 X 10"3 m3

gallon (US liquid) 1 gal (US) = 3.785 X 10"3 m3

litre 1 I 1.00 X 10"3 m3

Force

dyne 1 dyn = 1.00 X 10"5 N kilogram force 1 kgf 9.807 X 10° N poundal 1 pdl 1.383 X 10"1 N pound force (avoirdupois) 1 Ibf 4.448 X 10° N ounce force (avoirdupois) 1 ozf 2.780 X 10"' N

Power

British thermal unit/second 1 Btu/s 1.054 X 103 W calorie/second 1 cal/s 4.184 X 10° W foot-pound force/second 1 f t l b f / s = 1.356 X 10° W horsepower (electric) 1 hp 7.46 X 102 W horsepower (metric) (= ps) 1 ps 7.355 X 102 W horsepower (550 f t l b f / s ) 1 hp 7.457 X 102 W

* Factors are given exactly or to a maximum of 4 significant figures

Multiply by to obtain

Density

pound mass/inch3

pound mass/foot3

Energy

British thermal unit calorie electron-volt erg foot-pound force kilowatt-hour

Pressure

newtons/metre2

atmosphere*» bar centimetres of mercury (0°C) dyne/centimetre2

feet of water (4°C) inches of mercury (0°C) inches of water (4°C) kilogram force/centimetre2

pound force/foot2

pound force/inch2 (= psi)<> torr (0°C) (= mmHg)

Velocity, acceleration

inch/second foot/second (= fps) foot/minute

mile/hour (= mph)

knot free fall, standard (= g) foot/second2

Temperature, thermal conductivity, energy/area, time

Fahrenheit, degrees —32 Rankine 1 Btu-in/ft2-s- °F 1 Btu/ft-s- °F 1 cal/cm s °C 1 Btu/ft2-s 1 cal/cm2-min

Miscellaneous

foot3 /second foot3/minute rad roentgen curie

1 lbm/in3 = 2.768 X 104 kg/m3

1 lbm/ft3 = 1.602 X 10' kg/m3

1 Btu = 1.054 X 103

1 cal = 4.184 X 10° J 1 eV ~ 1.602 X 10"19 J 1 erg = 1.00 X 10"2 J 1 f t - lb f = 1.356 X 10° J 1 kW-h = 3.60 X 106 J

1 N/m2 = 1.00 Pa 1 atm = 1.013 X 10s Pa 1 bar = 1.00 X 10s Pa 1 cmHg = 1.333 X 103 Pa 1 dyn/cm2 = 1.00 X 10"' Pa 1 f tH jO = 2.989 X 103 Pa 1 inHg = 3.386 X 103 Pa 1 inH20 = 2.491 X 102 Pa 1 kgf/cm2 = 9.807 X 104 Pa 1 I bf/ f t2 = 4.788 X 10' Pa 1 lbf/in2 = 6.895 X 103 Pa 1 torr = • 1.333 X 102 Pa

1 in/s = 2.54 X 101 mm/s 1 ft/s = 3.048 X 1CT> m/s 1 ft/mi n = 5.08 X 10"3 m/s

Í4.470 X 10"' m/s 1 mile/h \1.609 X 10° km/h 1 knot = 1.852 X 10° km/h

= 9.807 X 10° m/s2

1 ft/s2 = 3.048 X 10"' m/s2

° F - 3 2 l 5 i ° C

1 9 1 K = 5.189 X 102 W/m-K = 6.226 X 10' W/m-K = 4.184 X 102 W/m-K = 1.135 X 104 W/m2

= 6.973 X 102 W/m2

1 ft3 /s = 2.832 X 10~2 m3/s 1 f t3 /min = 4.719 X 10"4 m3/s

rad = 1.00 X 10"2 J/kg R = 2.580 X 1 0 " C/kg Ci = 3.70 X 1010 disintegration/s

aatm abs: atmospheres absolute; atm (g): atmospheres gauge.

' I b f / i n 2 (g) (= psig): gauge pressure; lbf/in2 abs (= psia); absolute pressure.

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SUBJECT GROUP: I Life Sciences/Dosimetry