We have investigated the specific activation by soluble antigen of rat or mouse long-term T helper cell lines using antigen-presenting cell (APC)-free culture conditions. Some T cell lines specific for avidin or myelin basic protein responded to native antigen in the absence of added APC. Responses in the absence of APC were substantial and specific although, as would be expected, lower than in the presence of APC. Proliferation could not be inhibited by culture with anti-Ia antibodies and the ability of lines to respond to antigen in the absence of APC did not correlate with the endogeneous surface Ia expression of the lines. Furthermore, irradiated T cells were unable to act as presenting cells for lines cells of the same or a different specificity. This suggests that the T cells did not present antigen to each other, and demonstrates, along with other data shown, that activation cannot be attributed to undetected APC remaining in the cultures. Anti-avidin T cell lines differed markedly in their ability to respond to avidin in the absence of added APC.S2, an anti-avidin line of H-2s genotype consistently responded well to avidin seen in the absence of added APC; K2, an H-2k anti-avidin line, responded moderately and B3, and H-2b anti-avidin line, although the most prolific responder in the presence of APC, never responded to antigen in their absence. Z1a, a Lewis rat-derived T cell line specific for myelin basic protein, proliferated well in response to the antigen in the absence of added APC. The present findings demonstrate that some T cells can recognize and respond to native antigens without the mediation of specialized APC.