This study was undertaken to determine an immunologically active regimen for the administration of recombinant gamma-interferon (rIFN-gamma). The patient population included patients with completely resected melanoma, stage I (Clark's level IV or V) or stage II. All patients exhibited no evidence of disease (NED) at the time of the study. Patients received rIFN-gamma by intramuscular (IM) injection daily for 15 days at 0.0001 mg/m2, 0.001 mg/m2, 0.01 mg/m2, 0.1 mg/m2 (ten patients/group), or 0.25 mg/m2 (five patients). Interferon (IFN) was well tolerated, with non-dose-limiting constitutional symptoms occurring in the majority of patients at 0.1 mg/m2 and 0.25 mg/m2. All five patients receiving 0.25 mg/m2 developed elevated transaminase levels, which led to a discontinuation of therapy in one patient. Immunological activity was assessed by serial measurements of natural killer (NK) cell activity, hydrogen peroxide production by monocytes, and changes in expression of Fc receptors and human leukocyte class II antigen (HLA-DR) on monocytes. These changes were determined at baseline (X2), six to seven time points during rIFN-gamma therapy, and two times after the last dose of rIFN-gamma. No changes were observed at the two lowest doses. At the 0.01 mg/m2 dose, all parameters were elevated but not as consistently nor to the same levels as seen following administration of 0.1 mg/m2. At 0.25 mg/m2, H2O2 production was enhanced, but unlike at 0.1 mg/m2, it declined during the last few days of IFN therapy. Subcutaneous (SC) administration was compared with IM administration using the 0.1 mg/m2 dose. SC administration resulted in enhanced H2O2 production and Fc receptor expression by monocytes. More consistent elevations in peroxide generation and higher levels of Fc receptor expression were seen following SC administration. No significant difference was found between the two routes of administration. A comparison of two schedules, daily and three times weekly, suggested that monocyte activation may return to normal 72 hours after IFN administration. Of the doses tested, 0.1 mg/m2 administered daily appeared to be the most effective biological response modifier (BRM) regimen, and because of ease of administration, we favor the SC route.