Successful immunotherapy of early S.C. or i.p. (816) melanoma in syngeneic C57BL/6 (86) mice was achieved with S.C. peri-lesional injections (for S.C. tumors) o r i.p. injections (for i.p. tumors) of recombinant human interleukin 2 (rlL-2) and recombinant murine interferon-gamma (rlFN-7). Over a 28day period, rlL-2 and rlFN-y were injected 14 times. Results with this combination were additive with S.C. tumors and synergistic with i.p. tumors. Treatment with 6,250 U-25,000 U of rlL-2 and 2 pg of rlFN-y began 1-3 days after S.C. inoculation of melanoma. On day 50, 87% (72183) of mice thus treated were completely free of tumor. None of the 78 control mice (tumor + buffer) survived. Of mice receiving either rlL-2 o r rlFN-y alone, 59% (47179) and 53% (44/83), respectively. were tumor-free. 1.p. tumors were also eradicated by i.p. injections of rlL-2 (50,000 U) with rlFN-y (5,10, and I5 pg) as judged by absence of tumor in 81 O h (21/26) o f mice autopsied between days 45 and 65. N o control mice survived, and only 17% (2/12) and 2Ooh (6/30) given either rlL-2 o r rlFN-y separately (i.p.) were tumor-free. Doses of rlFN-7 from 1-4 pg were more beneficial in eliminating I-day S.C. melanomas than were higher doses. and local S. C. treatment was far superior t o distant or systemic treatment. Non-adherent peritoneal or splenic cells from mice bearing 6-day-old i.p. melanomas and treated with one or both lymphokines on days 3 and 4 were used in cytotoxicity assays in virro. Significant cytotoxicity against cultured melanoma cells was displayed by cells harvested from lymphokine-treated mice, but there was no evidence of the synergism observed with combination treatment of i.p. tumors in vivo. rlFN-y inhibited proliferation of melanoma cells in virro. whereas rlL-2 stirnulated proliferation at 1,000 U/ml. Plating efficiency was increased by a t least 30% by culture with 100 U or 1,000 U of rlL-Uml and both concentrations neutralized the inhibitory effect of 0.05 ng/ml of IFN-y, but not of 0.5 o r 5.0 ng/ml.