In this study, we examined the activity of recombinant interferon (IFN)-gamma against Plasmodium berghei exoerythrocytic forms (EEF) grown in vitro within the highly differentiated human hepatoma cell line HEPG2. We assayed the effect of IFN-gamma on parasite growth by DNA hybridization using a P. berghei specific DNA probe. The specific activity of IFN-gamma against EEF is very high, and depends upon the time of lymphokine addition. When IFN-gamma is added to HEPG2 cells containing intracellular EEF, 6 hr after sporozoite invasion, parasite DNA replication is inhibited by approximately 75% at 10(3) U/ml and 50% at 1 U/ml. This treatment can either abolish or greatly reduce the infectivity of EEF for mice. When added earlier, 3 hr after completion of sporozoite invasion, IFN-gamma inhibits parasite replication to an even greater degree. The highest levels of inhibition were obtained when IFN-gamma was added 6 hr prior to sporozoite invasion (100% inhibition at 10(2) U/ml, approximately 55% inhibition at 0.1 U/ml, and 17% inhibition at 0.001 U/ml). We found that HEPG2 cells express approximately 44,000 surface receptors for IFN-gamma. These data are consistent with the view that IFN-gamma exerts its antimalarial activity by binding to surface receptors on hepatocytes and inducing intracellular changes unfavorable for parasite development. Tryptophan starvation does not appear to be involved in this process. These findings also support the idea that IFN-gamma, released from immune T cells upon encountering sporozoite antigen, may be an important effector mechanism in sterile immunity to sporozoite challenge.