In order to delineate the important antigenic determinants, human thyroiditis and rabbit antihuman thyroglobulin sera have been tested against native and modified thyroglobulins. Reduction or trypsinization of thyroglobulin produced a substantial diminution of its reactivity with rabbit antisera and almost abolished its reaction with thyroiditis sera. Thyroglobulin transiently exposed to 8 M urea retained the ability to precipitate with rabbit antibodies but did not precipitate with human antibodies. It was possible to demonstrate, however, that urea-treated thyroglobulin formed soluble complexes with human antibodies. No new antigenic sites for the human serum were produced by any of the modifications. The soluble complexes formed between rabbit and human antibodies and native thyroglobulin have been compared by sucrose gradient ultracentrifugation. The complexes formed with human antibodies were more homogeneous than those formed with rabbit antibodies at a given antibody-antigen ratio. It is concluded that human antithyroglobulin antibodies possess a more restricted range than rabbit antibodies. All of the antigenic determinants active in human autoimmunization are exposed on native thyroglobulin, and the sites with the highest affinities are destroyed by modification of the secondary and tertiary structure of the molecule.