Immunoprecipitation of radiolabeled human thyroid cell proteins by serum from patients with autoimmune thyroid disease.
Characterization of the antigen to thyroid-stimulating immunoglobulin (TSI), presumably the TSH receptor, remains elusive. We, therefore, attempted immunoprecipitation of the TSI antigen using cultured human thyroid cells radiolabeled to high specific activity by [35S]methionine (all proteins labeled) or by 125I and lactoperoxidase (primarily cell surface proteins labeled). Both techniques produced similar results. Crude membrane preparations from the labeled cells were solubilized in buffer containing 1% Triton and then incubated with TSI-containing or control serum. Immunoglobulin was precipitated with goat antihuman immunoglobulin and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. About 30 proteins were immunoprecipitated by both normal and control sera. However, only 1 protein (mol wt, approximately 320,000) was preferentially selected by TSI-containing serum (6 of 6 tested). This protein was also immunoprecipitated by the majority of sera from patients with Hashimoto's thyroiditis. The 320,000 mol wt protein constituted only a minute amount of total radiolabeled membrane protein, because it was not apparent on electrophoresis of the membrane extract before immunoprecipitation. It corresponded (in migration) to a human thyroglobulin (Tg) standard. Tg also inhibited immunoprecipitation of the 320,000 mol wt band. The anti-Tg antibody titer correlated significantly with the degree of immunoprecipitation of the 320,000 mol wt band. No specific immunoprecipitation of any protein was observed when radiolabeled guinea pig membrane proteins were used. These data provide insight into the difficulties involved in the identification of the TSH receptor with serum containing TSI.
D000941:Antigens / Q000032:analysis; D001327:Autoimmune Diseases / Q000097:blood / Q000276:immunology*; D011232:Chemical Precipitation; D006111:Graves Disease / Q000097:blood* / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology*; D018828:Immunoglobulins, Thyroid-Stimulating; D007158:Immunologic Techniques; D008565:Membrane Proteins / Q000276:immunology; D013961:Thyroid Gland / Q000276:immunology*
D000941:Antigens; D007074:Immunoglobulin G; D018828:Immunoglobulins, Thyroid-Stimulating; D008565:Membrane Proteins