Splenic T and B lymphocyte and null cell populations were analyzed in NZB mice as autoimmune disease developed during the first year of life. In the B lymphocytes, a progressive shift occurred from the surface IgD bright subset to the surface IgM bright subset. There was a slight increase in the ratio of Ly-2- to Ly-2+ T cells. Splenomegaly was not detected until after 40 wk of age and was primarily due to an increase in the number of null cells. This change was accompanied by an increase in the size, as determined by narrow-angle forward light scatter, of the Ly-2+ but not the Ly-2- T cells, an elevation of IgG-containing plasma cells, and the appearance of anti-erythrocyte autoantibody. The splenic B cell subset distribution and the enlargement of the Ly-2+ T cells were reflected in the peripheral blood, whereas the T cell subset ratio was not. The B cell subset alteration did not correlate with any of the other changes observed. Statistical associations were found between the ratios of T cell subsets, the enlargement of the Ly-2+ T cells, and the increased number of null cells, suggesting a linkage among those late changes that immediately precede the development of overt autoimmune disease.