A SMART method for isolating monoclonal antibodies from individual rhesus macaque memory B cells.

ABSTRACT: Characterizing antigen-specific B cells is a critical component of vaccine and infectious disease studies in rhesus macaques (RMs). However, it is challenging to capture immunoglobulin variable (IgV) genes from individual RM B cells using 5' multiplex (MTPX) primers in nested PCR reactions. In particular, the diversity within RM IgV gene leader sequences necessitates large 5' MTPX primer sets to amplify IgV genes, decreasing PCR efficiency. To address this problem, we developed a switching mechanism at the 5' ends of the RNA transcript (SMART)-based method for amplifying IgV genes from single RM B cells to capture Ig heavy and light chain pairs. We demonstrate this technique by isolating simian immunodeficiency virus (SIV) envelope-specific antibodies from single-sorted RM memory B cells.

SUBMITTER: Weinfurter JT 

PROVIDER: S-EPMC10842827 | biostudies-literature | 2024 Feb

REPOSITORIES: biostudies-literature