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JMD 2005, Vol. 7, No. 2
Copyright © 2005 American Society for Investigative Pathology & Association for Molecular Pathology

B-Cell Clonality Determination Using an Immunoglobulin {kappa} Light Chain Polymerase Chain Reaction Method

Reetesh K. Pai*, Artemis E. Chakerian*, John M. Binder{dagger}, Mitual Amin{dagger} and David S. Viswanatha*{ddagger}

From the Department of Pathology, * Experimental Pathology Laboratory, University of New Mexico; Tricore Reference Laboratories, {ddagger} Albuquerque, New Mexico; and the Department of Pathology, {dagger} William Beaumont Hospital, Troy, Michigan

To augment the detection of clonality in B-cell malignancies, we designed a consensus primer {kappa} light chain gene (Ig{kappa}) polymerase chain reaction (PCR) assay in combination with a consensus primer immunoglobulin heavy chain gene (IgH) PCR assay. Its efficacy was then evaluated in a series of 86 paraffin tissue samples comprising neoplastic and reactive lymphoproliferations. Analysis after PCR was accomplished by 10% native polyacrylamide gel electrophoresis after heteroduplex pretreatment of PCR products and by a post-PCR chip-based capillary electrophoresis analytic method. Overall, 49 of 68 (72%) of mature B-cell neoplasms yielded discrete Ig{kappa} gel bands within the predicted size range with no clonotypic Ig{kappa} products observed among reactive lymphoid or T-cell proliferations. The application of Ig{kappa} PCR improved overall sensitivity from 81% with IgH PCR alone to 90% with combined Ig{kappa}/IgH PCR, with this effect being most notable in germinal center-related lymphomas. Sequencing of positive Ig{kappa} rearrangements revealed that most rearrangements involved members of the V{kappa}1 (40%) and V{kappa}2 (34%) gene families along with J{kappa}1 (26%), J{kappa}2 (23%), and J{kappa}4 (51%) gene segments. Involvement of V{kappa} pseudogenes was identified in 24% of cases with V{kappa}-KDE rearrangements. Our results demonstrate the efficacy of Ig{kappa} PCR in improving the detection rate of clonality in B-cell neoplasms and further introduce a novel post-PCR chip-based capillary electrophoresis analytic method for rapid PCR fragment size evaluation.




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