Novel In Vitro Booster Vaccination to Rapidly Generate Antigen-specific Human Monoclonal Antibodies

Nandin IS, et al., 214(9):2811, J Exp Med. , 2017

Identification of novel pathogen antigens and production of human therapeutic antibodies are important in the continued development of vaccines to prevent infections disease. In this study a novel in vitro approach using human B-Cells induced to produce antigen specific antibodies is described. Antigens were linked to streptavidin coated beads along with a TLR ligand, CpG, known to induce B-Cell proliferation and differentiation into plasma cells. It is hypothesized that B-cell receptor specific internalization of CpG will lead to activation and proliferation of antigen specific B-Cells within a population. Initially a common B-Cell receptor kappa chain was targeted with a particle coated with anti-ƙ light chain antibody and CpG to allow optimization of B-Cell activation. B-Cell activation was determined by expression of CDs by flow cytometry, phosphorylation of key intracellular signalling proteins and RNA-seq analysis of stimulated B-Cells. Subsequently, the ability of the system to induce proliferation and differentiation of B-Cell subsets for tetanus toxoid, influenza haemaglutinin subtypes H1N1, H5N1, H7N9 and gp120 was tested. Plasma cells were generated and produced antibodies shown to be antigen specific by ELISA. The affinity of the generated antibodies for the antigen was determined using SPR or BLI (Octet) technologies. The Octet data was generated using streptavidin biosensors hydrated in 1X kinetics buffer for at least 10min and loaded with 12.3 μM antigen. Association of antibody from 0.094 to 60nM in 2-fold dilutions and subsequent dissociation in 1X kinetics buffer was used to generate sensorgrams for analysis of kinetics parameter ka, kd and KD.

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