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Martina Zaninotto
In the early 2000s an assay that measured
serum immunoglobulin-free light chains
(FLC) was developed
….these novel polyclonal antibodies
reacted with only those epitopes that were
hidden when bound to heavy chain but
available when not associated with heavy
chain
…the serum FLC assay in combination with
serum PEL and serum IFE is sufficient to
screen for pathological monoclonal plasma
Proliferative disorders other than AL…
-20
-15
-10
-5
0
5
10
15
20
CV
%
Mean = 18.19 mg/L
mean CV% =7.05 2008-2009
2014
2014
2012
What are the requirements of anti-FLC antibodies?
What are the desirable performance goals?
Why is there a lack of harmonisation of patient results?
What are the optimal sample dilution procedures?
How do laboratories manage non harmonised measurement?
What reference intervals should be used?
……………………?
Biochimica Clinica, 2013
Specificity of selected monoclonal antibodies for FLC
moAb Kappa moAb Kappa
moAb lambda moAb lambda
● FLC kappa
○ IgG1 kappa
● FLC lambda
○ IgG1 lambda
te Velthius H et al 2011
No
cross-reaction with intact
immunoglobulins
Monoclonal antibody
based-method
Policlonal antibody based-
method (Robson E et al, 2007)
(te Velthius H et al, 2011)
ETF Policlonal A Monoclonal A
K light chains MM
IgA K MM
IgA K MM
IgA K MM
Tate J et al, 2013
Pretorius CJ et al 2012
Controlli Freelite N latex
Low High Low High
media, mg/L
Kappa
CV %
15.92 29.35 14.00 34.87
5.8 9.7 7.5 11.9
media, mg/L
Lambda
CV %
28.22 60.38 11.92 32.20
8.5 9.6 3.6 4.8
Braga et al 2012
Hansen CT et al, 2014
CVi 4.3%, CVa 7.3% RCV 24% CVi 7.0%, CVa 4.5% RCV 23%
Is the assays comparable in
EQA schemes?
kappa
Distribution n ALTM mg/L SD mg/L CV%
053 31 5.43 2.83 52.20
054 30 6.71 5.22 77.8
055 34 3.45 2.51 72.8
056 48 3914.41 2729.16 69.7
061 Beckman 4 23574.0 10228 43.4
061 Behring 20 57.1 29.2 51.1
063 Beckman 7 4965.14 1325.2 26.7
063 Behring 22 4464.55 1045.7 23.4
(2009)
Vercammen M, 2011
2013
FLC Kappa
23.000 mg/L
FLC lambda
57.000 mg/L
pre-reaction signal
main-reaction signal
threshold value
for pre-reaction
te Velthius H, 2011
Vercammen M, 2011
Jacobs JFM, 2011
Jacobs JFM, 2011
Kappa
mg/L
Lambda
mg/L
k/l Analizzatore
Altinier S et al
CCLM, 2010
4.52 –22.33 4.84 – 21.88 0.15 – 3.35 BNII, nefelometro
Freelite method
Katzmann JA et al
Clin Chem, 2002
3.3–19.4 5.7 – 26.3 0.26 – 1.65 BNII, nefelometro
Freelite method
Pattenden RJ
Ann Clin Biochem,
2007
4.1–41 11.6 – 45.7 0.3 – 1.3 BNII, nefelometro
Freelite method
Beetham R et al
Ann Clin Biochem
2007
9.4-18.7 20.1 – 34.7 0.21 – 1.40 Image, nefelometro
Freelite method
Pattenden RJ
Ann Clin Biochem
2007
6.2–35.3 2.7 – 20.6 0.7 – 3.4 AU400 turbidimetro
Freelite method
Te Velthius H et al
CCLM , 2011
6.7-22.4 8.3-27.0 0.31 – 1.56 BN Systems
N Latex method
Tate J et al, 2013
0
20
40
60
80
100
120
140
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
L BS
L siemens
Cate
ne le
gg
ere
la
mb
da
,
mg
/L
Freelite
N_Latex
Autotrapianto
2007
Sospensione
Terapia 2009
POEMS + amiloidosi cardica
Monitoraggio 2007-2011
0
5
10
15
20
25
30
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
27
22
Ca
ten
e le
gg
ere
la
mb
da
,
mg
/L
POEMS + amiloidosi cardica
Monitoraggio 2007-2011
0,00
0,50
1,00
1,50
2,00
2,50
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
Ka
pp
a/l
am
bd
a r
ati
o
POEMS + amiloidosi cardiaca
Monitoraggio 2007-2011
Freelite
N-Latex
0
100
200
300
400
500
600
700
1 2 3 4 5 6 7
K BS
K siemens
Mieloma micromolecolare K
Monitoraggio 2007-2010
Ca
ten
e le
gg
ere
ka
pp
a,
mg
/L
Autotrapianto
2007
Freelite
N Latex
0
20
40
60
80
100
120
140
160
1 2 3 4 5 6
0
20
40
60
80
100
120
1 2 3 4 5 6 7
Mieloma micromolecolare K
Monitoraggio 2007-2010
Kap
pa/L
am
bd
a r
ati
o
0
10
20
30
40
50
60
1 2 3 4 5 6
09-2009
01-2010
05-2010
Palladini G et al
Mollee P et al, 2013
Performance characteristics strongly affect the
clinical value of the test and
laboratory specialists have a duty to promote
the continuous improvement of analytical
quality specifications,
as well as
to recommend the appropriate clinical
interpretation/utilization of results obtained
with currently available assays...
M.Plebani, 2005
Right test
Best quality
Sensible request for good clinical
reasons
Expert interpretation by people who
are well trained
Continual evaluation of practice
Dr. Joanna Sheldon
Protein Reference Unit
St. George’s Hospital
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