Freshly obtained normal lymphoid cells kill certain tumor target cells in vitro. Using peripheral blood lymphocytes (PBLs) and the human tumor target cell line BT-20, we have defined a tumor necrosis factor (TNF)-dependent, cell-mediated cytotoxic mechanism that is homologous to the murine natural cytotoxic (NC) cell activity. Human NC cell activity was detected in freshly isolated PBLs and was augmented by short in vitro pulses with recombinant human interleukin-2 but not with recombinant human alfa interferon. Monoclonal anti-TNF antibodies inhibited the killing of the target cells. The independence of interferon and the mediation of killing by TNF distinguish human NC cell activity from natural killer and lymphokine-activated killer cell activities.