The overlap of external morphometric measurements between Peromyscus leucopus (White-footed Mouse) and P. gossypinus (Cotton Mouse) makes species determination challenging. Peromyscus were live-trapped at Poinsett State Park, Sumter County, SC, and identified using field and laboratory techniques. Our measurements for hind-foot length, total length, and tail length overlapped with published values for both species. Body mass, as measured in the field, was a good criterion that identified our animals as Cotton Mice (n = 29). This identification was confirmed by digital radiography, microsatellite DNA markers, and glucose phosphate isomerase analysis. Our results indicated that a combination of field and laboratory techniques is a valuable approach for positively identifying morphologically similar species.