Chromosome analysis for calibration curve

creation for potential retrospective dose estimation

Joint Institute for Nuclear Research

Laboratory of Radiation Biology

Students Gijana S.

Ngoepe PNM.

Seepei LE.

Supervisor Dr. Kutsalo P.

OUTLINE

Aim

Introduction

Methodology

Results

Conclusion

Acknowledgements

AIM

Creating calibration curve for retrospective dose estimation, on

the basis of the frequencies of unstable chromosomal aberrations

induced by the proton beam used in cancer therapy at the

Laboratory of Nuclear Problems at JINR.

INTRODUCTION

Introduction

There have been numerous radiation accidents:

• Hiroshima & Nogasaki, Japan, 1945

• Semipalatinsk, USSR, 1949-1963

• Chernobyl, USSR, 1986

• Tokia-Mura, Japan, 1999

• Byalujstok, Poland, 2001

Application of ionising radiation has increased.

Biological dosimetry allows assessing of dose of radiation by

analysis of damages induced in chromosomes of human peripheral

blood lymphocytes.

Lymphocytes Close correspondence between aberrations induced

in lymphocytes in vivo and in vitro.

Lymphocytes are type of cells found mainly in a DNA

presynthetic stage of a cell cycle.

Lymphocyte

Chromosomes

Chromosomes-

contain genetic

material, which makes

organisms what they

are.

DNA - main cell

target for the action of

ionising radiation.

Cell cycle

Cell cycle - events

that occur from the

completion of one

division until the

next completion of

one division of a cell.

In metaphase, centromeres are clearly visible and

analysis of dicentrics is possible.

(a) S-phase (b) Early G2-phase (c) Late G2-phase

(d) Metaphase (e) Anaphase (f) G1-phase

Chromosomal aberrations Different aberrations that can be observed on metaphase

analysis are dicentrics, centric rings, acentric rings,

chromosome fragments, chromatid breaks, etc.

Centric ring

Acentric

ring

Dic

METHODOLOGY

EQUIPMENT

Lithium

heparin tubes

Proton

irradiator

Incubator

Laminar

air flow

cabinet Pipette Centrifuge Light microscope

Isolation of lymphocytes

• Studies can be done on either whole blood or isolated

peripheral blood lymphocytes

Isolation of lymphocytes Checking the spread of chromosomes before staining

Working under sterile conditions

SCHEME OF EXPERIMENT Standard Metaphase Analysis (unstable aberrations)

Colcemid

47h

Fixation

48-50h

Observation using

light microscope

Culture of

Irradiated

blood

PHA

0h

RESULTS AND DISCUSSION

RESULTS

Results as measured by students

RESULTS

Results as measured by Dr. Kutsalo

CONCLUSION

The number of aberrations is dependent on the radiation dose

The retrospective dose can be determined from the number of

aberrations using the calibration curves that we have created

ACKNOWLEDGEMENTS

Dr. Mkaza

Prof. Lekala

Mr. Malaza

Dr. Kutsalo

National Research Foundation

Department of Science and Technology

Joint Institute for Nuclear Research

Thank you!