Immunohistochemical studies using immunized Guinea pig sera with features of anti-human thyroid, eye and skeletal antibody and Graves' sera.

Type 2 5' deiodinase enzyme was observed in both thyroid and eye muscle tissues, highlighting its possible role as a common antigen in thyroid-associated ophthalmopathy. Sera of 105 Graves' patients and 40 controls, and immunized guinea pig sera against TCSS peptide, showing homology to the amino acid sequence from 132 to 152 of type 2 5' deiodinase, were investigated to demonstrate the binding effects to human thyroid, eye and skeletal muscle tissues. Twenty-two Graves' patients were positive for anti-TCSS peptide antibodies, of whom 18 cases had ophthalmopathy. The levels of anti-TCSS peptide antibodies were higher not only in Graves' patients with (P<0.0001) and without (P<0.036) eye symptoms compared to controls but also the difference was significant between patients with and without ophthalmopathy (P<0.049). In Western blot, immunized sera showed binding reactions to the supernatant fractions of human thyroid, eye and skeletal muscle tissues at the range of 29 kDa. Patient sera with Graves' ophthalmopathy resulted in positive reactions directed to membrane areas in thyroid follicular cells, and to fibers in eye and skeletal muscles using immunohistochemical method, while no positive staining was present after adding control sera. The binding features of immunized guinea pig sera exhibited similar staining in all human tissues but could be blocked with Graves' sera. Our results suggest that type 2 5' deiodinase enzyme protein could play a role as an antigen in Graves' disease. Immunized guinea pig sera against TCSS peptide exhibited similar binding reactions and stainings to human thyroid, eye and skeletal muscle tissues as patient sera with Graves' ophthalmopathy.