The Detection of Antigens in Immune Complexes

Abstract

A procedure is described for the purification of soluble immune complexes (IC) from biological fluids and for the detection of antibody-bound antigen. A model IC, prepared with various amounts of human serum albumin (HSA) and constant amounts of anti-HSA, gave a 44% recovery after gel filtration on Sephacryl S-300, affinity chromatography on protein A-Sepharose, and concentration of eluates therefrom before analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was calculated that 20 ng of antibody-bound HSA should be detectable when polyacrylamide gels are subjected to a direct radioimmunoassay involving the use of rabbit anti-HSA, 125I-protein A and autoradiography, thus defining the sensitivity of the procedure for detecting IC-bound antigen in a model system. The procedure has direct relevance to the examination of IC of unknown composition present in sera of individuals with various diseases.