Poly I:C treatment can inhibit the ability of macrophages (M phi) to induce antigen-specific T-cell proliferation. This study investigated whether this inhibition is the result of suppressor or cytotoxic activity. Pretreatment of M phi with indomethacin in vivo, in vitro or both failed to reverse the inhibition of T-cell proliferation induced by poly I:C-treated, keyhole limpet haemocyanin (KLH)-pulsed M phi, suggesting that prostaglandin production does not mediate the inhibition of T-cell proliferation. The transfer of supernatants from cultures containing poly I:C-treated, KLH-pulsed M phi to cultures containing saline-treated, KLH-pulsed M phi and T cells did not inhibit T-cell proliferation, suggesting that the inhibition of T-cell proliferation by poly I:C is not mediated by the production of soluble suppressor factors. As addition of poly I:C-treated, KLH-pulsed M phi to cultures containing saline-treated, KLH-pulsed M phi did not significantly inhibit KLH-specific T-cell proliferation, the inhibition of T-cell proliferation is also not mediated by direct cell contact or short-range soluble suppressor factors. In addition, poly I:C-treated, KLH-pulsed M phi did not induce cytolysis of syngeneic T cells. These results indicate that cytotoxic or suppressor effector functions of M phi are not involved in the mechanism by which poly I:C inhibits M phi-induced, antigen-specific T-cell proliferation.