Fig. 1

Schematic workflow illustrating the steps involved in the study. a Selection step, and b validation step. Arthritis was induced in collagen-induced arthritis (CIA) mice by immunization with type II collagen (CII). Immunoglobulin G (IgG) was purified by protein G beads from serum of mice presenting with acute arthritis and naïve mice. A phage display library was used to select mimotopes against purified IgG from CIA mice (three cycles of biopanning), with a subtractive step against IgG from naïve mice. IgG-bound phages from CIA mice were competitively eluted by incubation with total proteins extracted from inflamed joints. Total proteins extracted from inflamed joints were also used for anti-M12 antibody recognition in Western blot. Immunoreactivity of each selected mimotope was tested by phage exyme-linked immunosorbent assay (ELISA). After DNA extraction and in silico and in vitro analysis, the most reactive mimotope (M12) was identified as a peptide that mimics a predicted antigenic site of the human carbonic anhydrase III protein. Validation of the M12 synthetic peptide as a possible rheumatoid arthritis (RA) autoantigen was carried out by ELISA assay