Identifying and sorting live vaccine-specific B cells from peripheral blood and bone marrow using spectral enhanced flow cytometry 3898

Abstract Description Detection of vaccine-specific B cells and antibody secreting cells (ASC) are important to inform HIV vaccine strategies. Herein, we describe the development of a flow cytometry-based assay using cell surface mounted Ig-traps and fluorescently labelled antigen-specific probes to detect HIV vaccine-specific B cells. Moreover, this technique is suitable for downstream characterization with transcriptomics and functional cell culture-based assays. Broader applications include the study of infections, cancer and allergy. To refine assay parameters, peripheral blood (PB) and bone marrow (BM) samples were isolated from non-human primates (NHPs) that received a bivalent group M HIV mRNA vaccine with or without a boost of A244+B.63521 adjuvanted with ALFQ. Briefly, Ig-traps are bound to the cell surface and allowed to naturally secrete antibodies which are trapped and bound to the cell surface. This allows previously un-probable ASC populations to become probe-able without fixation/permeabilization. These molecular constructs are used in conjunction with our NHP flow cytometry panel and analyzed with a spectrally enabled cytometer to characterize and sort ASCs. Our preliminary results show that this technique successfully identified and sorted HIV-specific ASCs in NHP samples. This refined technique will be used to detect ASCs in preclinical and clinical vaccine and therapeutic trials to further inform future vaccination strategies and capture novel bnAbs Topic Categories Technological Innovations in Immunology (TECH)