To clarify the immunopharmacological action of an extract isolated from inflamed skin of rabbits inoculated with vaccinia virus (Neurotropin), its effect on delayed type hypersensitivity (DTH) response to sheep red blood cells (SRBC) in mice was examined. Neurotropin enhanced the DTH response in C57BL/6 mice which were low responders to SRBC, but not in either BALB/c or C3H/He mice (high responders) when administered i.p. for 4 consecutive days prior to sensitization. However, Neurotropin did not affect the formation of plaque-forming cells to SRBC in C57BL/6 mice under the condition where it enhanced the DTH response. We further examined the mechanism by which Neurotropin enhanced the DTH response in C57BL/6 mice by means of cell transfer experiments. Spleen cells from mice administered Neurotropin i.p. for 4 days, but not saline, could enhance the DTH response when transferred i.v. into normal syngeneic mice just before sensitization. However, the treatment of the spleen cells with anti-Thy-1.2 + complement (C) or with anti-Lyt-1.2 + C, but not with anti-Lyt-2.2 + C, abrogated its enhancing effect. The depletion of macrophages from the cells had no effect. On the other hand, the spleen cells from mice administered Neurotropin had no enhancing effect in the effector phase of DTH response, and they showed a helper T cell activity in a DTH helper T cell assay system in which cyclophosphamide-treated mice were used as recipients. These results suggest that Neurotropin enhances the DTH response in low responder mice through the induction of Lyt-1+2- DTH helper T cells.