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

Eric Waltari,Peter S Kim,Aaron Mcgeever,Natalia Friedland,Krista M Mccutcheon

doi:10.3389/fimmu.2019.01452

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 peripheral blood mononuclear cells (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. Using in vitro polyclonal stimulation, we 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.

Highlights

One of public health’s most cost-effective interventions to prevent and reduce the disease burden of infectious disease is vaccination
Examples of typical B cell receptor (BCR) amplicon products used for next generation sequencing (NGS), which looked similar between peripheral blood mononuclear cells (PBMCs) and stimulated PBMC, are shown in Supplementary Image S1
We provide methods to allow for a deep analysis of the clonal diversity, persistence and hierarchy of antibodies in the memory repertoire from PBMCs

Summary

Introduction

One of public health’s most cost-effective interventions to prevent and reduce the disease burden of infectious disease is vaccination. Memory B cells survive in a functionally quiescent state in tissues and can be found at low frequency in PBMCs after a pathogen is eliminated, re-activating and differentiating into antibody secreting plasmablasts within a week of a secondary infection [4,5,6]. Evaluating both types of adaptive B-cell memory is challenged by the anatomical inaccessibility and rarity of these cells. A variety of serological assays for polyclonal antibody measurements have been developed [7] and, more recently, the applications of generation sequencing (NGS) and single cell sequencing technologies have offered high resolution

Methods

Results

Conclusion