Aotus monkeys received 4 doses of Plasmodium falciparum EBA-175 region II vaccine as plasmid DNA (Dv-Dv) or recombinant protein in adjuvant (Pv-Pv) or as 3 doses of DNA and 1 dose of protein (Dv-Pv). After 3 doses, antibody titers were ∼104 in DNA-immunized monkeys and 106 in protein-immunized monkeys. A fourth dose did not significantly boost antibody responses in the Dv-Dv only or Pv-Pv only groups, but titers were boosted to ∼106 in monkeys in the Dv-Pv group. Four weeks after the last immunization, the animals were challenged with 104 P. falciparum-parasitized erythrocytes. Peak levels of parasitemia were lower in the 16 monkeys that received region II-containing plasmids or proteins than in the 16 controls (geometric mean: 194, 178 and 410, 110 parasites/μL, respectively; P = .013, Student's t test). Three of 4 monkeys in the Dv-Pv group did not require treatment. These data demonstrate that immunization with EBA-175 region II induces a significant antiparasite effect in vivo.