Quantitative real-time PCR method for detection of B-lymphocyte monoclonality by comparison of kappa and lambda immunoglobulin light chain expression.
BACKGROUND
An abnormal IgLkappa:IgLlambda ratio has long been used as a clinical criterion for non-Hodgkin B-cell lymphomas. As a first step toward a quantitative real-time PCR-based multimarker diagnostic analysis of lymphomas, we have developed a method for determination of IgLkappa:IgLlambda ratio in clinical samples.


METHODS
Light-up probe-based real-time PCR was used to quantify IgLkappa and IgLlambda cDNA from 32 clinical samples. The samples were also investigated by routine immunohistochemical analysis and flow cytometry.


RESULTS
Of 32 suspected non-Hodgkin lymphoma samples analyzed, 28 were correctly assigned from real-time PCR measurements assuming invariant PCR efficiencies in the biological samples. Four samples were false negatives. One was a T-cell lymphoma, one was a diffuse large B-cell lymphoma, and one was reanalyzed and found lymphoma-positive by in situ calibration, which takes into account sample-specific PCR inhibition. Twelve of the samples were fine-needle aspirates, and these were all correctly assigned.


CONCLUSIONS
This work is a first step toward analyzing clinical samples by quantitative light-up probe-based real-time PCR. Quantitative real-time PCR appears suitable for high-throughput testing of cancers by measuring expression of tumor markers in fine-needle aspirates.
Ståhlberg
Anders
A
Department of Molecular Biotechnology, Chalmers University of Technology, 405 30 Gothenburg, Sweden.
Aman
Pierre
P
Ridell
Börje
B
Mostad
Petter
P
Kubista
Mikael
M
D000465:Algorithms; D001402:B-Lymphocytes / Q000737:chemistry*; D006801:Humans; D007145:Immunoglobulin kappa-Chains / Q000032:analysis* / Q000235:genetics; D007146:Immunoglobulin lambda-Chains / Q000032:analysis* / Q000235:genetics; D008198:Lymph Nodes / Q000737:chemistry / Q000473:pathology; D016393:Lymphoma, B-Cell / Q000737:chemistry / Q000175:diagnosis / Q000473:pathology; D008954:Models, Biological; D016133:Polymerase Chain Reaction
D007145:Immunoglobulin kappa-Chains; D007146:Immunoglobulin lambda-Chains