AU cell-surface antigen of human malignant melanoma: Solubilization and partial characterization

Abstract

AU antigen is defined by reactions of sera from patient AU with cell-surface antigens of cultured autologous melanoma cells (SK-MEL-28). Past studies established that no available cell type other than AU melanoma expressed AU antigen. By use of antibody inhibition tests for antigen detection, limited papain digestion of AU melanoma cells was found to result in the solubilization of AU antigen along with β(2)-microglobulin (β2m) and HLA allogeneic and xenogeneic specificities. Comparable papain treatment of other melanoma and non-melanoma cell lines solubilized β2m and HLA, but did not result in the release of antigen with AU reactivity. Maximum yield of AU antigen from AU melanoma cells was obtained after very short (5-15 min) digestion times in contrast to the more prolonged proteolysis required for maximum HLA and β2m release. AU antigen was not immunoprecipitated by rabbit antiserum against β2m or HLA under conditions leading to partial or complete removal of β2m of HLA. At least a proportion of the molecules with AU determinants appear to be glycoproteins, as indicated by specific affinity for Lens culinaris hemagglutinin (LcH). After affinity chromatography on LcH-agarose, the specific activity of AU antigen was increased 50-fold. As determined by gel filtration chromatography, AU antigen has a molecular weight in the range of 20,000-50,000.