A simple procedure for isolation of Bufo arenarum C3 complement fraction and preparation of antiserum

Because of the need for antibodies in our studies involving the third component of complement in Bufo arenarum, we performed a simple procedure to purify C3 from B. arenarum serum to use as antigen in the preparation of the antiserum. The strategy was based on the well-known ability of C3 to bind to zymosan (Zy), a yeast cell wall extract comprised of polysaccharides. The Zy-bound fraction showed cross reactivity with a commercial antibody to human C3 as well as a similar electrophoretic profile (SDS-PAGE) to C3 from other species. The Zy-C3 complex resulting from binding Zy to B. arenarum serum was injected into rabbits and the antiserum against this C3-like fraction was purified by protein A-Sepharose chromatography. The purified C3 antibody was found to be suitable for immunochemical studies.