Soluble cytotoxic factors from mouse spleen cells have been shown to selectively lyse NK sensitive target cells. Lysis of target cells is assessed by trypan blue uptake or 51Cr--release assay in a 16-48 hour assay. The possible role of such natural killer cytotoxic factors (NKCF) in the mechanism of natural killer cell-mediated cytotoxicity (NKCMC) has been examined. Several lines of evidence are presented which indicate that there exists a strong correlation between lysis by NKCF and lysis in NKCMC. For instance, (1) NKCF are generated following stimulation of mouse spleen cells with NK sensitive targets; (2) Lysis of NKCF is selective for NK sensitive targets and is species specific; (3) Mice with poor NK activity, such as Bg/Bg mice, produce poor NKCF: (4) There is concomittant inhibition of NKCMC and NKCF activities by blocking RAT serum; and (5) Several known characteristics of the mechanism of NKCMC are shown to be shared in the NKCF system. Based on these findings, we propose a model for NKCMC in which lysis by NK effector cells is the result of multiple steps, namely target binding to an NK effector cell, activation of the lytic mechanism, and involvement of NKCF to mediate lysis. Accordingly, for targets to be NK sensitive, they ought to be able to interact and bind with NK effectors, activate the NK cells, bind NKCF, and be sensitive to the NKCF lytic activity.