Detection and isolation of antigen-specific immune complexes from sera of melanoma patients

Detection of antigen-specific circulating immune complexes (IC) in sera from cancer patients provides an approach for defining tumor antigens that induce a host immune response and could prove useful for purposes of immunoprognosis. In the present study, a sandwich ELISA was developed to detect antigen-specific IC in cancer patients, utilizing a murine monoclonal antibody designated MAb JSI. The MAb was produced by cell fusion using standard hybridoma technology following immunization with a partially purified fetal antigen that had been isolated from the spent culture medium of a melanoma cell line. The partially purified antigen appeared to be broadly expressed on melanomas, sarcomas, and carcinomas. The MAb was mass-produced in pristine-primed mice. MAb JSI reacted with the cultured melanoma cell M14 but not the autologous lymphoblastoid cell L14. Following purification by ammonium sulfate precipitation, the MAb was immobilized on polystyrene plates and utilized to capture antigen-specific immune complexes from sera of melanoma patients which were detected with anti-human globulins. The second antibody in the sandwich was shown to be endogenous human IgG. Antigen-specific immune complexes were present in melanoma patients but only infrequently in sera from normal individuals and patients with active autoimmune disease. Antigen-specific immune complexes detected in melanoma sera were isolated by affinity chromatography utilizing the MAb JSI. The assay described in this report is simple and reproducible, without plate effect (p = 0.97) or time effect (p = 0.34) and could provide a useful new approach to examine the role of antigen-specific circulating immune complex analysis in cancer patients.