We have previously described an in vitro sensitization (IVS) procedure which enabled the generation of therapeutic T cells from tumor-bearing mice for adoptive immunotherapy. The procedure involved culture of tumor-draining lymph node (TDLN) cells with irradiated tumor in the presence of interleukin-2 (IL-2). The availability of many recombinant cytokines affords an opportunity to examine their effects on the immune response to tumor. In this study, we investigated the effect of tumor necrosis factor-alpha (TNF alpha) on the generation and function of IVS cells utilized in adoptive immunotherapy of the murine MCA 106 sarcoma. TNF alpha administered iv at nontherapeutic doses was found to enhance the antitumor efficacy mediated by IVS cells plus IL-2 in the treatment of pulmonary metastases. In contrast, TNF alpha administration to mice bearing progressive footpad tumors had inhibitory effects on the sensitization of tumor-reactive cells in TDLN since IVS cells generated from these animals displayed a diminished antitumor effect. This effect appeared to be due to a reduced number of tumor-reactive lymphoid cells in the TDLN since TNF alpha added to IVS cultures did not alter the antitumor efficacy of the resultant IVS effector cells. These findings indicate the divergent effects of TNF alpha on the immune response to tumor and adoptive immunotherapy with IVS cells.