It has recently been shown that tumor necrosis factor (TNF) induces increased Ia antigen expression on a malignant murine macrophage cell line, and that TNF is synergistic with gamma interferon (IFN) in inducing Ia expression. This finding raises the possibility that TNF serves as a non-interferon macrophage activating factor in vivo. Since it is known that TNF has different effects on malignant and benign cells, we chose to evaluate the effects of recombinant TNF on primary cultures of murine peritoneal macrophages (MP). Neither human nor murine TNF increased the proportion of MP which expressed Ia antigen, and TNF actually partially prevented the IFN-induced increase in Ia. However, culture with TNF activated MP for increased hydrogen peroxide production in response to phorbol myristate acetate (PMA) and antagonized the IFN-induced decrease in the proportion of the MP bearing receptors for the Fc region of IgG2b. TNF and IFN were additive in increasing peroxide production. Both human and murine TNF had the same effects on MP, and MP from C3H/HeN (endotoxin sensitive) and C3H/HeJ (endotoxin resistant) mice responded comparably to TNF and IFN. Our results support the hypothesis that TNF has some macrophage-activating activity, but its effects are selective and not identical with those of IFN.