The majority of neotropical primates (NP) are endangered, however the impact on the health of these animals caused by parasitic infections, such as Plasmodium sp. infection is not well understood, particularly the immune response. This study aims to evaluate the humoral immune response of wild and captive NP naturally exposed to Plasmodium spp infection. More than one hundred serum samples of NP, belonging to all Platyrrhini families, from different municipalities of Brazilian Atlantic Forest region were used in immunoenzymatic assay for the detection of IgG antibodies against Plasmodium antigens: (i) peptides of circumsporozoite protein, as a marker antigen of infection exposition; and (ii) three antigen markers of the establishment of blood infection (DBPII, AMA-1 and EBP). Eighty-five percent of NP had antibodies against exposure marker, and 76% against at least one of the establishment of blood infection markers. Levels of antibodies were higher in free-living than in captive monkeys from the same area. A high percentage of adults compared to non-adults were seropositive and showed higher antibody levels. Our findings suggested that NP from all families were somehow exposed to malaria infection, but only in some of them the erythrocytic infection was well-established. NP from Atelidae and Pitheciidae were the most responsive. This study will help the identification of potential reservoirs for malaria and the understanding of the course of immune response during natural Plasmodium infection in neotropical primates.