High-resolution epitope mapping of commercial antibodies to ANCA antigens by yeast surface display

Epitope mapping provides critical insight into antibody-antigen interactions. Epitope mapping of autoantibodies from patients with autoimmune diseases can help elucidate disease immunogenesis and guide the development of antigen-specific therapies. Similarly, epitope mapping of commercial antibodies targeting known autoantigens enables the use of those antibodies to test specific hypotheses. Anti-Neutrophil Cytoplasmic Autoantibody (ANCA) vasculitis results from the formation of autoantibodies to multiple autoantigens, including myeloperoxidase (MPO), proteinase-3 (PR3), plasminogen (PLG), and peroxidasin (PXDN). To perform high-resolution epitope mapping of commercial antibodies to these autoantigens, we developed a novel yeast surface display library based on a series of >5000 overlapping peptides derived from their protein sequences. Using both FACS and magnetic bead isolation of reactive yeast, we screened 19 commercially available antibodies to the ANCA autoantigens. This approach to epitope mapping resulted in highly specific, fine epitope mapping, down to single amino acid resolution in many cases. Our study also identified cross-reactivity between some commercial antibodies to MPO and PXDN, which suggests that patients with apparent autoantibodies to both proteins may be the result of cross-reactivity. Together, our data validate yeast surface display using maximally overlapping peptides as an excellent approach to linear epitope mapping.