Double-Antibody Radioimmunoassay for B16 Melanoma Antibodies 2

Abstract

A double-antlbody radioimmunoassay was developed that quantitated the antibody response to melanoma-associated antigens (MAA). The assay was based on the coprecipitation by anti-immunoglobulin of complexes of radiolabeled MAA with tumor antibodies. MAA were radiolabeled by incubation of melanoma cells in tissue culture with 3H-leucine. Serum of C578L/6J mice, immunized by repeated injections of irradiated B16 melanoma cells, bound 4–8 times as much radiolabeled MAA as did serum of normal mice. The relationship was linear between the amount of MAA bound and the amount of antiserum used. The assay was sufficiently sensitive and reproducible to measure changes in the levels of antibodies to 816 melanoma during its natural growth. Antibody levels declined during the late phase of tumor growth. There were cross-reactions between the serum of mice immunized to mammary adenocarcinoma and MAA and between antimelanoma serum and antigens in syngeneic fibroblasts, which suggests the presence of shared antigenic determinants between these tumors and normal mouse tissue.