Evaluation of High-Affinity Monoclonal Antibodies\nand Antibody-Drug Conjugates by Homogenous Time-Resolved FRET

The\nrapid growth of therapeutic monoclonal antibodies demands greater\naccessibility to scalable methods of evaluating antigen binding. Homogenous\nTR-FRET is ideal for preliminary screening but has not been reported\nto assay these interactions due to their high-affinity and complex\nsolution-phase kinetics. Here we report the development of a competition\nassay to rank-order the relative affinities of these drugs for a common\nantigen. The assay is compatible with automation, requires no modification\nof the analytes, and measures affinities as low as single-digit picomolar.\nWe further demonstrate applications to inform the development of antibody-drug\nconjugates. The assay may aid discovery and manufacturing of therapeutic\nantibodies as a low-cost, high-throughput alternative to existing\ntechnologies.