Since the description of Peromyscus leucopus easti Paradiso, 1960, scant work has been directed toward examining the evolutionary relationships of P. leucopus in the mid-Atlantic region. We used morphometric and molecular analyses to examine the phylogenetic relationships of 3 subspecies known from the eastern United States (P. l. easti, P. l. leucopus, and P. l. noveboracensis); additional specimens from North and South Dakota (P. l. aridulus) were incorporated in the molecular analysis. Morphometric analyses revealed that large mice are found in New England and the Appalachian Mountains as far south as west-central North Carolina and that intermediate-sized mice inhabit the Cumberland Plateau and the lowlands from Virginia southward at least to Alabama. These groups represent P. l. noveboracensis and P. l. leucopus, respectively. The smallest specimens, which are referable to P. l. easti, were found in the vicinity of Princess Anne County, Virginia. The largest mice were found on the southern half of Hatteras Island, North Carolina. Examination of D-loop sequence data revealed that there is no apparent restriction of gene flow among P. leucopus on the mainland of the eastern United States, including those from Princess Anne County, Virginia. However, mice from the southern end of Hatteras Island, North Carolina, were as distinct from specimens from the eastern United States as were mice from North and South Dakota. The population of white-footed mice from Hatteras Island, North Carolina, represents an undescribed subspecies.