Evaluation of the efficacy of 3 nonviable blood-stage antigens-schizont Ag, merozoite Ag, and "French press" Ag-of Plasmodium knowlesi, emulsified in Freund's complete adjuvant (FCA), was carried out in a study involving 32 rhesus monkeys. After 2 immunizations, administered 6 weeks apart, monkeys were challenged with a variant of P. knowlesi different from that used for immunization. All 8 control monkeys that received either PBS or FCA developed a severe parasitaemia and died of the infection within 12 days after parasite challenge. In the group that received the freeze-thawed schizont Ag, 5 of the 8 monkeys died at the same time as the controls, 1 died a day later, and 2 survived the infection (maximum parasitaemia: 0.7%; 3.2%). In the freeze-thawed merozoite Ag group, 1 monkey died at the same time as the controls, 5 monkeys died between 14 and 17 days after challenge, and 2 monkeys survived the infection (maximum parasitaemia: <0.01%; 2.7%). In the lyophilized French press Ag group, 2 monkeys died at the same time as the controls, 2 died between 12 and 15 days after challenge, and 4 survived the infection (maximum parasitaemia: 1.2%; 0.4%; 0.9%; 0.07%).Immunized monkeys failed to gain weight during the period of immunization and abscess formation at injection sites was comparable in monkeys of the 3 Ag groups. Reticulocytosis and anaemia developed in all monkeys that survived longer than 12 days after challenge, even in monkeys with very low-grade levels of parasitaemia.Various tests were performed to assess humoral or cell-mediated immune responses during the period of immunization and after challenge. No clear-cut relationship could be established between the results of any of these tests and the survival of immunized monkeys after parasite challenge. Some of the results, however, suggest that there might be some relationship between cell-mediated responses and the survival of immunized monkeys beyond the survival time of control monkeys.