Expression of mouse embryonic epitope TEC-2 on human carcinoma-derived cell lines and characterization of its glycoprotein carriers.
Monoclonal antibody TEC-02, raised against mouse embryonal carcinoma cells, has been shown to react with murine preimplantation embryos and with a very limited number of adult mouse tissues. The target epitope, TEC-2, is a carbohydrate carried in mouse embryonal carcinoma cells by large glycoprotein-bound glycan. We report here the expression of TEC-2 epitope on human carcinoma-derived cell lines, HeLa and HS, and the properties of its carbohydrate carriers. Immunolabeling of Nonidet P-40 lysates of HeLa cells separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that TEC-2 antigens are polydispersed glycoconjugates of high molecular weight (mostly above 100,000). TEC-2 antigens detected by the two-site sandwich assay, in which the antigen is immobilized and detected with the same antibody, had a slightly higher molecular weight than those detected by the solid-phase assay. This suggests heterogeneity in the number of TEC-2 epitopes per carrier molecule. When the cells were lysed by Triton X-114 and the detergent and aqueous phases were separated by warming and centrifugation, most of the TEC-2 antigenic activity was found in the aqueous phase. TEC-2 antigens isolated by indirect precipitation from [3H]galactose-labeled HeLa cells were degraded by extensive pronase digestion or mild alkaline treatment to glycopeptides or oligosaccharides of low molecular weight. Thus, TEC-2 epitope in human HeLa cells is carried by carbohydrates of only several monosaccharide units. TEC-02 antibody was also found to bind to Tamm-Horsfall glycoprotein isolated from human urine and its binding was enhanced by desialylation. Combined data indicate that TEC-02 antibody recognizes the GalNAc beta 1----4Gal beta 1----4 structure which may be carried on different types of molecule, according to the site of their synthesis.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D000951:Antigens, Neoplasm / Q000032:analysis*; D002352:Carrier Proteins / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D054278:Embryonal Carcinoma Stem Cells; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis*; D005455:Fluorescent Antibody Technique; D006001:Glycoconjugates / Q000032:analysis / Q000276:immunology; D006023:Glycoproteins / Q000032:analysis*; D006367:HeLa Cells; D006801:Humans; D007118:Immunoassay; D051379:Mice; D008970:Molecular Weight; D009088:Mucoproteins / Q000276:immunology; D014411:Neoplastic Stem Cells / Q000276:immunology*; D014407:Tumor Cells, Cultured; D058949:Uromodulin
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D002352:Carrier Proteins; D000939:Epitopes; D006001:Glycoconjugates; D006023:Glycoproteins; D009088:Mucoproteins; C548612:UMOD protein, human; C548613:Umod protein, mouse; D058949:Uromodulin