Abstract: Functional and structural analyses of major histocompatibility complex (MHC) class I molecules of the Aotus genus are necessary to validate it as a solid animal model for biomedical research. We thus isolated, cloned and sequenced exons 2 and 3 from three Aotus species ( A. nancymaae , A. nigriceps and A. vociferans ). We found 24 sequences, which divided into two different groups (Ao‐g1 and Ao‐g2). A further sequence was identified as a processed pseudogene (Aona‐PS2). Both sequence evolution and variability analyses showed that Ao‐g1 and Ao‐g2 display similar characteristics to Catarrhini's classical loci, such as positive selection pressure at the peptide binding region (PBR) high variability and a trans ‐specific evolution pattern.