The merozoite surface protein-1 (MSP-1) is a leading candidate for a vaccine targeted at the erythrocytic stages of plasmodial parasite development. Recently, there has been increasing interest in this polypeptide, particularly in the carboxyl-terminal EGF-like domains. We have previously shown that this region from Plasmodium yoelii, when expressed in native configuration, could immunize mice against an otherwise lethal challenge infection. In this model system, protection appears to be predominantly mediated by antibodies. In all rodent immunization studies to date, however, the immunogen has contained both of the postulated EGF-like domains. We report here on the efficacy of immunization with the individual EGF-like domains from P. yoelii in elicitation of a protective host response. Although all animals developed some level of antibody in response to the various immunogens, only those animals immunized with both EGF-like domains produced antibodies which could recognize the native MSP-1 molecule. Antibodies generated against the individual EGF-like domains did cross-react with the double EGF-like domain structure, suggesting that the immunogens had retained elements of native configuration. In addition, only those animals which generated antibodies capable of recognizing native MSP-1 showed any level of protection from challenge infection. These results suggest that determinants unique to the double EGF-like domain structure may be necessary for the generation of antibodies specific for the native configuration of MSP-1 and that these antibodies may play a significant role in protection.