Core D: Bmem Analysis Core

Abstract The overall goal of this core is to expand and provide centralized analysis of human memory B cells (Bmem) for the research program, to enhance our understanding of humoral responses to influenza vaccination and infection. Specifically, Core D will perform single-cell Nojima cultures to characterize BCR repertoire of human Bmem cells. Nojima culture, developed by members of Core D, is a feeder cell-dependent, B-cell expansion / antibody cloning system to characterize the B-cell repertoire at a single-cell level. Unique features of Nojima cultures include unbiased expansion of single B cells, high cloning efficiency, and ease of sample handling. Combining Nojima cultures with multiplexed antigen-binding assays, we can rapidly determine large numbers of BCR phenotypes (specificity, avidity, cross-reactivity) and genotypes (V(D)J gene sequences) for mouse, human, and non-human primate B cells. Core D activities include 1) isolation of single human Bmem cells from different age groups, 2) generation of clonal IgG antibodies through Nojima cultures, 3) determination of specificity and avidity, and 4) V(D)J gene sequences of the clonal IgG antibodies. Core D will directly interact with all projects and other scientific cores through communications, discussions, and data sharing. The inclusion of this Bmem Analysis Core will provide robust, uniform and comprehensive analysis of the human Bmem repertoire, critical for achieving the overall goals of the Program Project. 1