The mechanism of the cytostatic and cytocidal activities of TNF was studied in human tumour cells. BT-20 breast and ME-180 cervical cancer cells were significantly growth-inhibited by TNF, but other cells were not. When protein synthesis was inhibited by cycloheximide, however, TNF was cytotoxic for all cells except BT-20 cells. This suggests that different mechanisms are responsible for the cytostatic and cytocidal activities of TNF. The sensitivity of different cell lines could not be correlated with the number or affinity of TNF receptors. Some protease inhibitors completely protected human and murine cells from TNF cytotoxicity. Inhibitors of chymotrypsin-like proteases were more effective than inhibitors of trypsin-like proteases. Reversible and irreversible inhibitors (such as alkylating compounds) were both protective. The cells were best protected when pretreated with inhibitors before the addition of TNF. When the protease inhibitors were removed the cells gradually lost their resistance to TNF cytotoxicity. The inhibitors did not interfere with the functioning of TNF-receptor complexes, since SK-MEL-109 melanoma cells treated with a protease inhibitor synthesized TNF-induced proteins. These findings suggest that a protease is involved in the cytocidal activity of TNF.