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In vitro reconstitution of B cell receptor–antigen interactions to evaluate potential vaccine candidates
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In vitro reconstitution of B cell receptor–antigen interactions to evaluate potential vaccine candidates
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Journal Article
In vitro reconstitution of B cell receptor–antigen interactions to evaluate potential vaccine candidates
2016
Overview
This protocol describes an immunogen evaluation pipeline containing two main components that enable vaccine candidates to be rank-ordered.
Predicting immune responses before vaccination is challenging because of the complexity of the governing parameters. Nevertheless, recent work has shown that B cell receptor (BCR)-antigen engagement
in vitro
can prove a powerful means of informing the design of antibody-based vaccines. We have developed this principle into a two-phased immunogen evaluation pipeline to rank-order vaccine candidates. In phase 1, recombinant antigens are screened for reactivity to the germline precursors that produce the antibody responses of interest. To both mimic the architecture of initial antigen engagement and facilitate rapid immunogen screening, these antibodies are expressed as membrane-anchored IgM (mIgM) in 293F indicator cells. In phase 2, the binding hits are multimerized by nanoparticle or proteoliposome display, and they are evaluated for BCR triggering in an engineered B cell line displaying the IgM sequences of interest. Key developments that complement existing methodology in this area include the following: (i) introduction of a high-throughput screening step before evaluation of more time-intensive BCR-triggering analyses; (ii) generalizable multivalent antigen-display platforms needed for BCR activation; and (iii) engineered use of a human B cell line that does not display endogenous antibody, but only ectopically expressed BCR sequences of interest. Through this pipeline, the capacity to initiate favorable antibody responses is evaluated. The entire protocol can be completed within 2.5 months.
Publisher
Nature Publishing Group UK,Nature Publishing Group
Subject
