Biophysical considerations for development of antibody-based therapeutics

Abstract

Development of therapeutic monoclonal antibodies (mAbs) requires rigorous measurements of the kinetic and thermodynamic binding properties of antibody-antigen complexes for drug candidate optimization and the design of clinical dosing strategies. For measuring the dissociation equilibrium constants of mAbs binding reversibly to antigens, two premier technologies are commonly used: Biacore surface plasmon resonance (SPR) and the solution-based kinetic exclusion assay (KinExA). This chapter details the correct experimental design, the proper use of the instrumentation, optimal data processing, instrument limitations and potential sources of artifacts, as well as a rigorous comparison between SPR and KinExA approaches. Biacore applications for high-throughput kinetic screening and epitope binning are briefly presented. Additionally, the use of cell-based affinity assays using fluorescence activated cell sorting and KinExA is discussed for instances where purified antigens outside a cell membrane lose their native structure and/or functionality.