Detection of clonality in follicular lymphoma using formalin-fixed, paraffin-embedded tissue samples and BIOMED-2 immunoglobulin primers

Abstract

Aims: The BIOMED-2 multiplex PCR protocol is a commonly used procedure for assessing B cell clonality in lymphoma diagnostics. Follicular lymphoma poses a special challenge for PCR-based analyses because of high prevalence of somatic hypermutations in the rearranged immunoglobulin (IG) domains. This study aimed to evaluate the BIOMED-2 protocol performance in detection of B cell clonality in follicular lymphoma using formalin-fixed, paraffin-embedded (FFPE) tissue. Methods: FFPE samples from 118 patients diagnosed with follicular lymphoma in the period 1998-2008 were used in the study. Clonality of IG heavy (IGH) and light chains (IGK, IGL) was assessed using a PCR procedure that was optimised for FFPE tissue. Results: The highest clonal detection rates were 67.8% with the IGH Vo-FR2-Jo assay and 66.1% with the IGK Vκ-Jκ assay. Clonality was detected in 94.9% of all FFPE follicular lymphoma samples when all assays were combined. FFPE samples stored for 1-5 years did not perform significantly differently from those stored for 6-11 years. Interobserver agreement of clonality was tested for all analyses. The lowest score (Cohen's κ value = 0.56) was observed for the IGK Vκ-Jκ clonality assay. Conclusions: An improved PCR protocol for detection of clonality in FFPE samples using BIOMED-2 IG primers is presented. For best performance, a combination of IGH and IGK analyses is recommended.