The Sensitivity of Existing TCRγ PCR Primer Sets Is Sufficient To Eliminate TCRβ Southern Blot Analysis for Clonality Assessment.
Amir Behdad, Eric Olsen, Wayne Grody, Joshua Deignan. Cedars-Sinai Medical Center, Los Angeles, CA; University of California, Los Angeles
Background: Clonality assessment is an important tool in the diagnosis of suspected lymphoproliferative disorders. Southern blot (SB) analysis is currently considered the gold standard, and is commonly either used alone or in conjunction in cases which are found to be negative by PCR. However, there are newer PCR primer sets which may have the potential to completely eliminate Southern Blot analysis for the detection of clonality in blood and bone marrow specimens.
Design: We evaluated existing records from 120 patients, including 33 bone marrow and 87 peripheral blood specimens, submitted to our laboratory between 2008 and 2010. All samples were studied using the Invivoscribe (Non-BIOMED-2) multiplex PCR reagents for the immunoglobulin heavy chain (IGH) and T-cell receptor-γ (TCRG). Any case with negative PCR results (n=63) was further evaluated by SB (either IGH or T-cell receptor-β (TCRB)). The results from both PCR and SB were then compared with the clinical diagnosis.
Results: PCR results for IGH and TCRG were available for 62 and 96 cases, respectively. The PCR results in 85% of the IGH cases and 45% of the TCRG cases were negative, and SB was performed only for those cases. The results of SB and PCR were concordant in all cases where T-cell clonality was assessed. B-cell clonality was missed only in three neoplastic cases with B-cell lineage, while T-cell clonality (by PCR) was positive for all of these three cases. B-cell clonality by SB was checked for only one of the cases and was positive. The overall analytical sensitivity was 100% for TCRG PCR and was 75% for IGH PCR. In 86% of all cases, the molecular findings correlated with the histopathological diagnosis, which is consistent with other previously published findings. Interestingly, TCR clonality by PCR was also detected in the blood of 15 patients with no definitive histopathological diagnosis of a lymphocytic malignancy.
Conclusions: TCRB Southern blot analysis did not enable the detection of any TCR rearrangements which were not already found using our existing PCR primer sets, and TCRG PCR may add additional sensitivity to cases which are negative for IGH PCR for the clonal assessment of malignant B-cell proliferations. IGH SB should continue to be used for all negative IGH PCR cases, though alternate IGH primer sets exist (ex. BIOMED-2) which may increase the analytical sensitivity of PCR for B-cell malignancies and enable the elimination of IGH SB.
Tuesday, March 1, 2011 1:00 PM
Poster Session IV # 167, Tuesday Afternoon