Techniques to Study Antigen-Specific B Cell Responses

Abstract

Antibodies against foreign antigens are a critical component of the overall immune response and can facilitate pathogen clearance during a primary infection and also protect against subsequent infections. Dysregulation of the antibody response can lead to an autoimmune disease, malignancy, or enhanced infection. Since the experimental delineation of a distinct B cell lineage in 1965, various methods have been developed to understand antigen-specific B cell responses in the context of autoimmune diseases, primary immunodeficiencies, infection, and vaccination. In this review, we summarize the established techniques and discuss new and emerging technologies for probing the B cell response in vitro and in vivo by taking advantage of the specificity of B cell receptor (BCR)-associated and secreted antibodies. These include ELISPOT, flow cytometry, mass cytometry, and fluorescence microscopy to identify and/or isolate primary antigen-specific B cells. We also present our approach to identify rare antigen-specific B cells using magnetic enrichment followed by flow cytometry. Once these cells are isolated, in vitro proliferation assays and adoptive transfer experiments in mice can be used to further characterize antigen-specific B cell activation, function, and fate. Transgenic mouse models of B cells targeting model antigens and of B cell signaling have also significantly advanced our understanding of antigen-specific B cell responses in vivo .

Broadly neutralizing human antibodies against dengue virus identified by single B cell transcriptomics

“…Recent advances in monoclonal antibody isolation and characterization (Boonyaratanakornkit and Taylor, 2019; Corti and Lanzavecchia, 2014) have accelerated the identification of bNAbs, including those against flaviviruses. Examples include antibodies d488 (Li et al, 2019) and m366 (Hu et al, 2019), which were cloned from B cells of rhesus macaques receiving an experimental DENV vaccine and from healthy flavivirus-naive humans, respectively, and antibody DM25-3, which was isolated from a mouse immunized with a mature form of DENV2 virus-like particles (Shen et al, 2018).…”

Regulatory B cells in transplantation: roadmaps to clinic

“…In a humanized mouse model of alloimmune‐mediated injury of human skin grafts, the infusion of alloantigen‐specific Tregs resulted in a significant reduction in dermal tissue injury when compared to polyclonal Tregs [69,70]. Antigen‐specific B cells have a low precursor frequency and are difficult to isolate and expand in high yields [71]. The advent of chimeric antigen receptors (CAR) is revolutionizing T‐cell therapy, and could facilitate development of an antigen‐specific IL‐10 + Breg therapy [72].…”

Screening of Antigen-Specific Antibody-Secreting Cells

“…To achieve direct measurements of secreted mAbs by flow cytometry, the secreted protein must be confined to the cell surface or in the vicinity of the secreting cell, where it can be fluorescently labelled. Several secretion capture assays based on an affinity matrix around the secreting cell have been developed: encapsulating cells in a medium that retains the secreted products or by modifying the cell surface with a capture probe (Boonyaratanakornkit and Taylor 2019;Carroll and Al-Rubeai 2004). These methods also allow for the selection of ASCs with high mAb secretory activity against specific target antigens at a single-cell level.…”