Two approaches have been developed to induce TSH receptor antibodies in mice with properties resembling those in Graves' disease, the Shimojo model of injecting live fibroblasts coexpressing the TSH receptor and major histocompatibility complex antigen Class II, and TSH receptor-DNA vaccination. Thyroid-stimulating antibodies appear to occur less commonly after DNA vaccination, but there has been no direct comparison of these models. We performed a three-way comparison of 1) AKR/N and 2) BALB/c mice vaccinated with TSH receptor-DNA and 3) AKR/N mice injected with fibroblasts expressing the TSH receptor and the major histocompatibility complex antigen class II of AKR/N mice. TSH receptor-DNA vaccinated mice had low or undetectable levels of TSH receptor antibodies determined by ELISA or flow cytometry. Nonspecific binding precluded comparisons with sera from Shimojo mice by these assays. TSH binding inhibition and thyroid-stimulating antibody were undetectable in TSH receptor-DNA vaccinated mice. In Shimojo mice, TSH binding inhibition was positive in approximately 60%, and thyroid-stimulating antibodies were positive in hyperthyroid animals. Unlike the negative antibody data, splenocytes from TSH receptor-vaccinated (but not Shimojo) mice proliferated and produced the Th1 cytokine interferon-gamma in response to TSH receptor antigen. In conclusion, DNA vaccination is less effective at inducing TSH receptor antibodies than the Shimojo approach, but it permits the future characterization of TSH receptor-specific T cells generated without adjuvant.