Inasmuch as human natural killer (NK) cell activity was markedly augmented by a streptococcal immunopotentiator, OK-432, both in vivo and in vitro, the mechanism in which OK-432 augmented human NK cell activity was analyzed. Culture supernatants of nonadherent lymphocytes stimulated with OK-432 significantly augmented NK cell activity. Significant activity of both interferon (IFN) (both alpha- and gamma-types) and interleukin-2 (IL-2) was detected in the culture supernatants from nonadherent lymphocytes. Concomitant treatment of supernatants with anti-IFN-alpha antiserum and pH-2 glycine-HCI buffer or the absorption of supernatants with an IL-2-dependent cell line completely abrogated the NK-augmenting activity, whereas the treatment with either one of these resulted in only partial elimination of the activity. These results indicate that OK-432 stimulates human nonadherent lymphocytes to produce IFN and IL-2 and that both factors are primarily responsible for the NK augmentation by OK-432.