Human peripheral blood lymphocytes (PBL), obtained from patients with a variety of malignancies, when incubated in vitro with phytohemagglutinin (PHA), lysed fresh autologous tumors during a short-term 51Cr-release assay. These PHA-activated killer (PAK) cells produced maximum lysis of tumor cells by 4 to 8 hr of co-cultivation. PHA incubation induced the generation of cells lytic for autologous and allogeneic tumors but not autologous or allogeneic PBL. Cold target inhibition studies demonstrated that autologous and allogeneic tumors of various histologic types all shared the determinants recognized by PAK cells. Some adherent cells were necessary for generation of these PAK, but higher numbers were suppressive. Augmentation of tumor cell lysis was seen when adherent cells were partially removed before PHA activation. The PAK effector cell was OKT3+, OKT8+. The precursor cell of the PAK was separated from natural killer (NK) cells on Percoll gradients and was generated from thoracic duct lymphocytes, a population devoid of NK cells. Furthermore, the phenotype of the majority of the precursor cells was OKT3+, OKM1-. Activation by PHA for 2 days was found to be an efficient and convenient method for generating lymphoid cells lytic for fresh autologous human tumor. The biologic and possible therapeutic role of these cells is currently being investigated.