Spleen cells from adult thymectomized mice (ATX) were assayed in a syngeneic graft vs host (GVH) model based upon enlargement of the draining popliteal lymph node following syngeneic cell inoculation into the hind footpad. Spleen cells from ATX mice have been found to induce a significantly higher increase in the weight of the regional lymph node than that induced by the injection of normal spleen cells. Irradiated spleen cells from ATX donors did not cause a similar increase, suggesting either that proliferation of the transferred cells was required at some stage of the reaction or that autoreactive cells are radiosensitive. Autoreactive cells were found in the spleen of mice 2 to 3 months after the thymectomy but were never found in the lymph nodes of such animals or in the thymus of intact mice. They are not phagocytic adherent cells and are not retained on nylon wool columns, which suggests that they belong to the T-cell lineage. Autoreactivity is lost when spleen cells from ATX donors are depleted of autologous rosette-forming cells (A-RFC) by centrifugation on a Ficoll-Hypaque gradient after rosette formation. Autoreactive spleen lymphocytes might belong to the population of A-RFC previously characterized as a population of immature T cells.