Protein Production, Structural Biology, and Nucleic Acid Sequencing and Analysis

CORE B SUMMARY This Program Project focuses on a group of closely related and clinically important antibody-mediated thrombocytopenias in which β3 integrins (αIIbβ3, αVβ3, and single-chain β3 fragments) and platelet chemokines including platelet factor 4 (PF4) and platelet basic protein isoforms (PBPi) are among the target antigens. Purified antibodies of human and mouse origin, and protein antigens including integrins, PF4, PBPi, and their mutants represent essential materials shared among the three Projects for the proposed structural and functional studies. In addition, next-generation sequencing (NGS) is a powerful tool that will be utilized by Projects 2 and 3 to isolate anti-PF4 and anti-PBPi monoclonal antibodies, and investigate the molecular and cellular mechanisms underlying the ontogeny of B cells that generate harmful antibodies resulting in antibody-mediated thrombocyto- penia and associated pathogenesis. The overall goal of Core B is to provide centralized and standardized large-scale antibody and protein production and authentication services, structure-based data interpretation of antibody-antigen interaction and functional assays, as well as centralized NGS and data analysis to each of the Projects in our Program, which should reduce the time and effort, maximize cost savings, ensure data repro- ducibility, cross-validation, and interpretation in parallel and/or complementary studies.