Generating a panel of clinically relevant monoclonal antibody to monitor conjugation conditions for pneumococcal polysaccharide vaccines 9261

Abstract Description The pneumococcal conjugate vaccine has been available for more than two decades. However, the impact of conjugation-induced structure changes in pneumococcal polysaccharides on clinical immunogenicity is not clearly understood. Under-conjugation may lead to weakened T-cell-dependent immune response, while over-conjugation or harsh conjugation conditions can alter critical polysaccharide epitopes, impairing the antibody response. We propose generating clinically relevant monoclonal antibodies (mAbs) with confirmed neutralizing ability by isolating polysaccharide-specific B-cells from human subjects to optimize the conjugation process. First, vaccinated donors were screened by using Multiplex Opsonophagocytic killing assay (MOPA) to identify those with high titers against three target serotypes. Single memory B cells were then sorted for in vitro culture using flow cytometry, and supernatants were tested for the presence of polysaccharide- and/or carrier-protein-specific antibodies. IgG genes from positive clones were then PCR amplified and sequenced. Recombinant IgGs were made to verify reactivity and functionality in ELISA and MOPA. Epitopes of these mAbs will be identified using glycan microarrays and/or nuclear magnetic resonance (NMR) spectroscopy. Finally, this mAb panel will be used to detect potential changes in the binding affinity of pneumococcal polysaccharides conjugated under different conditions/extensions, aiming to identify the best conjugation conditions. Funding Sources Supported by Merck & Co., Inc Topic Categories Vaccines and Immunotherapy (VAC)