The Use of Immunoglobulin Gene Rearrangement Polymerase Chain Reaction Assays for Detection of B-Cell Clonality for Plasma Cell Neoplasms Using Novel PCR PrimersLloyd Hutchinson*, Paul J Lee, Bruce A Woda, Ediz F Cosar, Mandi-Lee Caporelli BS and Xiuling Meng
Department of Pathology, University of Massachusetts Memorial Health Care, USA
- *Corresponding Author:
- Lloyd Hutchinson
Department of Pathology
University of Massachusetts Memorial Health Care
Innovation Dr., biotech, 3 Worcester, MA 01605, USA
Tel: +1 (413)545 0111
E-mail: [email protected]
Received Date: June 14, 2016; Accepted Date: October 15, 2016; Published Date: October 18, 2016
Citation: Hutchinson L, Lee PJ, Woda BA, Cosar EF, Caporelli M, et al. (2016) The Use of Immunoglobulin Gene Rearrangement Polymerase Chain Reaction Assays for Detection of B-Cell Clonality for Plasma Cell Neoplasms Using Novel PCR Primers. J Mol Biomark Diagn 7:302. doi: 10.4172/2155-9929.1000302
Copyright: © 2016 Hutchinson L, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
We have designed a standardized protocol with multiplex-primer sets capable of detecting majority IG kappa (IGK) and IG lambda (IGL) light chain rearrangements in plasma cell neoplasms (PCN). Thirty primers were combined in three multiplexed PCR reactions to target IGK, KDE and IGL rearrangements. Variable region (V) primers were designed to prevent “primer dimers”, provide matching melting temperatures (Tm), minimize amplicon size, and optimize sequencing time. Amplicons were subjected to capillary gel electrophoresis for analysis. In a discovery series, we tested 37 plasma cells neoplasms PCN (28 PCNs at diagnosis and 9 PCNs post-treatment). The assay investigated an additional 52 prospective PCN cases in the validation series. Results were compared to bone marrow morphology, immunohistochemical (IHC), flow cytometry data, and standard IGH FRIII gene rearrangement assay. In the discovery series, the following sensitivities/specificities were obtained for mature B-cell neoplasms: IGH FRIII: 29.7%/100%, IGK: 80.4%/100%, KDE: 25.0%/100%, and IGL: 35.1%/96.8%. The combination of IGH FRIII, IGK, and KDE detected 83.8% (31/37) vs 67.3% (35/52) in the discovery vs validation series, respectively, for the PCN population. Interestingly, 21.2% (11/52) of the validation samples positive by IG clonality, were negative by IHC and flow cytometry. In IHC/flow cytometry positive cases with a PCN representing a tumor burden of >50%, 10% to 50%, 1% to 10%, 0% to 1% of cells, the combined sensitivity of the IG clonality assay was 100% (20/20), 72% (23/32), 53% (10/19) and 20% (1/5) respectively. This IGK/IGL clonality assay has good sensitivity at diagnosis.