Determination of VH Family Usage in B-Cell Malignancies via the BIOMED-2 IGHPCR Clonality Assay

Abstract

Objectives: To determine whether VH family usage in B-celllymphoproliferative disorders can be deduced from polymerasechain reaction (PCR) product-length information obtainedthrough the BIOMED-2 (Invivoscribe, San Diego, CA) clonalityassay. Methods: We develop an algorithm that uses the sizinginformation of the BIOMED-2 immunoglobulin heavy chain(IGH) clonality assay to deduce VH family usage. PCR withfamily-specific primers on 51 clinical samples containing 54rearranged alleles were used to validate the algorithm. Results: The clonal PCR products in different frameworkreactions contain the same NDN segment (because they arefrom the same allele). Subtracting the size of the framework IIIproduct from the size of the framework I and II products yieldsthe relative position of the framework primer binding sites forthe VH segment used. The VH family can be assigned with theserelative positions because they are VH family specific in theBIOMED-2 assay. The VH family assigned by the algorithm wasconcordant with family-specific PCR results for 49 (96%) of the51 specimens. Conclusions: We have developed an algorithm that can correctlyassign VH family usage when all three BIOMED-2 frameworkreactions produced clonal products. Given the wide adoption ofBIOMED-2 assay, the algorithm can facilitate collection of IGHVH usage data without additional cost to the laboratories.