Two different approaches were used to identify new microsatellite polymorphisms among captive Bolivian squirrel monkeys (Saimiri boliviensis). In the first case, PCR primers for published human microsatellite loci were screened using genomic DNA from squirrel monkeys. Six polymorphic loci were identified using DNA samples from 19 unrelated individuals. The average heterozygosity among these six loci is 0.73. In the second set of experiments, a DNA library was created from Saimiri genomic DNA, and clones were selected from that library by screening with probes containing di-, tri-, and tetranucleotide repeats. Six novel microsatellites were identified this way, with an average heterozygosity of 0.59. Primer pairs for these six cloned microsatellites were also screened using a series of DNA samples from ten other platyrrhine species to assess the potential utility of these loci in other taxa. This study provides 12 new DNA polymorphisms that will be useful for various studies of this genus and demonstrates that both approaches can be used to develop new DNA polymorphisms in platyrrhine species.