PCR Analysis of IgH and TCR-γ Gene Rearrangements as a Confirmatory Diagnostic Tool for Lymphoproliferative Disorders

Abstract

This study investigates PCR analysis of immunoglobulin heavy chain (IgH) and T cell receptor (TCR) gene rearrangements on paraffin-embedded tissue sections and bone marrow aspirates of patients suspected to have lymphoproliferative disorders but with inconclusive diagnosis in histopathological examination. 130 samples of patients with inconclusive immunohistochemistry results were evaluated for clonal rearrangement of IgH and TCR genes. Based on histopathology examination, the patients were divided into three groups: the first group without any definite diagnosis of lymphoproliferative disorders (60 cases, 46.2 %), the second group suspected to have a lymphoproliferative disorder but in favor of benign disorders (19 cases, 14.6 %) and the third group suspect to lymphoproliferative disorders but relatively in favor of malignant disorders (51 cases, 39.2 %). After DNA extraction and quality control, semi-nested PCR was performed using consensus primers for amplification of TCR-c and CDR-3 regions of IgH genes. PCR products were analyzed after heteroduplex analysis using polyacrylamide gel electrophoresis, and were subject to silver staining. Totally, in over half of the cases (55.4 %), a monoclonal pattern was found in IgH or TCR-c genes rearrangements. Monoclonal IgH gene rearrangement was detected in 48.1 % of patients, whereas monoclonal TCR-c gene rearrangement was found in 33.6 % of them, which was not statistically significant (P = 0.008). Only in 32 patients (24.6 %) were the results of TCR-c and IgH gene rearrangements consistent with respect to the presence (2.3 %) or absence (22.3 %) of monoclonality. Finally, PCR analysis of TCR-c and IgH gene rearrangements led to definite diagnosis in 105 patients (80.8 %), and only 25 cases (19.2 %) remained inconclusive. Our results emphasize the usefulness of gene rearrangement study in cases without a definite diagnosis in immunohistochemistry studies. Multiple PCR analysis results when combined with patient's clinical course and immunohistochemistry can lead to early diagnosis and subsequent therapy.

The value of detecting immunoglobulin gene rearrangements in the diagnosis of B-cell lymphoma

“…In this study, the BIOMED-2 system was employed to analyze IG gene rearrangements in 244 samples of paraffin embedded lymphoid tissue. Of these, 209 B-cell lymphoma samples were positive for IG gene monoclonal rearrangement, but the 35 lymphadenosis samples did not show IG gene monoclonal rearrangement bands (false negatives and false positives were zero), suggesting that the detection rate was 100%, which is markedly higher than that of other detection systems [ 24 , 25 ] and consistent with Evans's report [ 15 ].…”

Diagnosing Vitreoretinal Lymphomas—An Analysis of the Sensitivity of Existing Tools

“…In addition to its diagnostic value, MyD88 mutation status is more cost effective and has a shorter turnaround time when compared to the established IgH PCR-based assays adapted by pathology laboratories [1]. The PCR-based IgH rearrangement assay requires a larger quantity of cells and generally has a higher limit of detection (10-20% of clonal B cell population) when compared to MyD88 mutation analysis, which can detect 5% or less mutant cells [51,52]. PCR for IgH and TCR gene rearrangements can also rarely yield a false positive result because of nonuniform (skewed) amplification of gene rearrangements.…”