Functional Enrichment and Analysis of Antigen-Specific Memory B Cell Antibody Repertoires in PBMCs

Abstract

Phenotypic screening of antigen-specific antibodies in human blood is a common diagnostic test for infectious agents and a correlate of protection after vaccination. In addition to long-lived antibody secreting plasma cells residing in the bone marrow, memory B cells are a latent source of antigen-experienced, long-term immunity that can be found at low frequencies in circulating PBMCs. Assessing the genotype, clonal frequency, quality, and function of antibodies resulting from an individual's persistent memory B cell repertoire can help inform the success or failure of immune protection. We have applied ELISPOT cell culture methods to functionally expand the memory repertoire from PBMCs and clonally map monoclonal antibodies from this population. We show that combining deep sequencing of stimulated memory B cell repertoires with retrieving single antigen-specific cells is a promising approach in evaluating the latent, functional B cell memory in PBMCs.

Functional Enrichment and Analysis of Antigen-Specific Memory B Cell Antibody Repertoires in PBMCs

“…This manuscript has been released as a Pre-Print at bioRxiv at https://www.biorxiv.org/content/10.1101/557769v1 (28). We thank Payton Anders Weidenbacher of Stanford University for providing the hemagglutinin and MEDI8852 proteins used for single cell antibody FACS and characterization, and thank Gerry Meixiong for assisting with the Reflow pipeline steps.…”

Analysis of Antigen‐Specific Human Memory B Cell Populations Based on In Vitro Polyclonal Stimulation

“…This can be addressed using MPAb‐purified IgG, for example, in assays that measure the functionality of antiviral Abs (Tesini et al., 2019). In vitro MBC stimulation also has value as a step to facilitate cloning of MBC‐expressed Ab genes through enrichment of MBCs and increasing transcript levels (Rajan et al., 2018; Waltari, McGeever, Friedland, Kim, & McCutcheon, 2019).…”