Fourteen microsatellite loci were characterized for the Rufous-throated Antbird (Thamnophilidae: Gymnopithys rufigula), an Amazonian obligate ant-following bird. Eight novel polymorphic microsatellite loci were isolated using next-generation sequencing and six additional loci were cross-amplified using primers previously designed for other bird species. All 14 loci were screened using 40 samples from central Amazon. The number of alleles per locus ranged from 3 to 20, whilst the observed and expected heterozygosities varied from 0.200 to 0.975, and 0.186 to 0.928, respectively. No linkage disequilibrium was found among the loci. Only one locus deviated from Hardy-Weinberg equilibrium. Among the eight novel loci, six were cross-amplified in at least one additional Thamnophilidae species, whilst three loci cross-amplified in two other Dendrocolaptidae species. This set of markers will be useful for future population genetic structure and parentage analysis of Amazonian forest birds.