When various synthetic double- or single-stranded DNAs were incubated with spleen cells from mice (BALB/c or CDF1) at 37 degrees for 20 hr, it was found that some of the DNAs augmented NK activity and produced factors in the culture supernatants which showed antiviral activity and activity to render mouse macrophages cytotoxic toward tumor cells. Poly(dG,dC) showed the strongest activities, when incubated with spleen cells from lipopolysaccharide-nonresponsive mice, C3H/HeJ. The activity of the culture supernatant to activate macrophages was completely abolished by a small amount of anti-IFN gamma antibody. On the other hand, the virus-inhibitory activity of the supernatant was mostly neutralized by anti-IFN alpha/beta. When IMC tumor cells (5 x 10(5) cells) were mixed with poly(dG,dC) (100 micrograms) and then inoculated intradermally into CDF1 mice, the tumor did not take, while tumors grew progressively and killed the mice in a control group inoculated with tumor cells alone. Direct cytotoxicity of poly(dG,dC) at a concentration of 1,000 micrograms/ml against IMC cells was not observed in vitro.