NK cell activity of four human premature infants between 28 and 33 wk of gestation and eleven human fetuses at 9 to 22 wk of gestation was tested against the K-562 cell line in a 4-hr 51Cr-release assay. Cord blood lymphocytes from premature infants expressed well-developed NK capacity, although the level of cytotoxicity was lower than that of full-term newborns or adults. Cells prepared from fetal liver displayed cytotoxicity against K-562 targets in four out of eleven fetuses, whereas cells from fetal bone marrow, spleen, and thymus expressed only marginal or negative anti-K-562 killing. NK cell activity in the fetal liver was observed as early as at 9 wk of gestation. The functional NK capacity of premature infants and fetuses was augmented in vitro by IFN-alpha treatment. Fetal cells without spontaneous NK activity did not develop cytotoxicity against K-562 target cells in the presence of IFN-alpha. The present results corroborate the concept of the intrauterine development of human NK cell activity.