Epitope mapping is essential for the understanding how an antibody works. Given millions of antibodies short of epitope information, there is an urgent need for high-throughput epitope mapping. Here we combined a commercial phage display of random peptide library of 109 diversity with next generation sequencing to develop Antibody binding epitope Mapping (AbMap) technology. Over two hundred antibodies were analyzed in a single test and epitopes were determined for >50% of them. Strikingly, the antibodies were able to recognize different proteins from multiple species with similar epitopes. We successfully identified the epitopes of 14 anti-PD-1 antibodies, including Sintilimab (i.e., L128, A129, and P130), and confirmed that the binding epitopes of Nivolumab and Sintilimab are very close to the binding interface of PD-1 and PD-L1. The predicted conformational epitopes of Pembrolizumab and Nivolumab are consistent with their antibody-antigen co-crystal structures. AbMap is the first technology enables high-throughput epitope mapping.

Antibody binding epitope Mapping (AbMap) of hundred antibodies in a single run