We previously reported on the 86 kDa natural killer target molecule associated with transformation of the oncogene-transfected rat fibroblasts. This molecule may participate in the lethal hit phase of cytotoxicity by natural killer (NK) cells. Originally, this molecule was defined by mAb109, but mAb109 could react only with rat tumor lines. In this report, to determine whether the 86 kDa molecule could be utilized as a natural killer target molecule in mammalian cells, we developed a polyclonal anti-86 kDa antibody (pAb109). Our data indicated that pAb109 preferentially reacted with NK-susceptible lines such as mouse YAC-1, rat W31 and human K562 cells, but reacted only weakly with NK-resistant mouse EL-4, rat fetal fibroblast WFB and human fetal fibroblast HEPM. In a cytotoxicity experiment, pAb109 F(ab')2 fragments could inhibit the cytolysis by NK cells of W31 and K562 cells. However, these fragments did not inhibit the cytotoxicity of non-NK cells such as CD3+, CD4-, CD8- T cell receptor alpha beta- T cells (presumably gamma delta T cells) to W31 cells. Taken together, these data suggest that the cell transformation-associated 86 kDa molecule may be critical in NK cytotoxicity, and a candidate for the NK target structure in mammalian tumor cells.