Peripheral blood lymphocytes of patients with autoimmune thyroid diseases were studied using monoclonal antibodies reacting with cell surface antigens of activated T-cells (Ia+T), as well as their helper-inducer (Ia+TH/I) and suppressor-cytotoxic (Ia+TS/C) subsets, using two-color dye labeling and dual laser activated cell sorter analyses. Compared to normal subjects, hyperthyroid Graves' disease patients had significantly higher percent Ia+T values in association with an increase in percent Ia+TH/I as well as a reduction in percent Ia+TS/C; whereas patients with hypothyroid Hashimoto's thyroiditis as well as those with postpartum thyroiditis studied in the hyperthyroid phase also had a significant but lesser increase in percent Ia+T-cells, but their percent Ia+TH/I subset was significantly decreased, whereas the percent Ia+TS/C subset was increased; and patients with toxic nodular goiter or factitious hyperthyroidism (nonimmunogenic causes of hyperthyroidism) had a significant increase in percent Ia+T-cells without a significant difference in their Ia+T subsets or their ratios in comparison to controls. These studies demonstrated the feasibility of detecting Ia+T-cells and their subset characteristics using two-color dye labeling and dual laser flow microfluorocytometric methodology. In both the active and treated phases of Graves' disease, Hashimoto's thyroiditis, and postpartum thyroiditis, the percent Ia+T-cells was increased compared to normal subjects, with the highest values occurring in hyperthyroid Graves' disease. Furthermore, patients with hyperthyroid Graves' disease had the opposite changes in percent Ia+TH/I and Ia+TS/C subsets as compared to patients with either untreated hypothyroid Hashimoto's disease or the hyperthyroid phase of postpartum thyroiditis, suggesting that the pathogenic mechanisms involved in Hashimoto's disease and the destructive hyperthyroidism of painless thyroiditis are similar, and that they are both distinctly different from that of hyperthyroid Graves' disease.