Middle-aged women have a lower prevalence and incidence of cardiovascular events compared with men. The mechanisms responsible for this gender-specific difference are unclear. Numeric and functional impairments of bone marrow–derived circulating endothelial progenitor cells (EPCs) are associated with increased cardiovascular and cerebrovascular morbidity and mortality. It is currently unknown whether there are gender-related differences in EPC number and function in middle-aged adults. We tested the hypothesis that EPCs isolated from middle-aged women demonstrate greater colony-forming capacity and migratory activity compared with men of similar age. Peripheral blood samples were collected from 50 sedentary adults, 25 men (59 ± 1 years of age) and 25 women (58 ± 1 years of age). Mononuclear cells were isolated and preplated for 2 days, and nonadherent cells were further cultured for 7 days to determine EPC colony-forming units. Migratory activity of EPCs was determined using a modified Boyden chamber. The number of EPC colony-forming units was significantly higher (∼150%) in samples collected from women (16 ± 3) compared with that collected from men (7 ± 1). In addition, EPC migration (relative fluorescent units) was ∼40% greater in women (729 ± 74) than in men (530 ± 67). In conclusion, these results demonstrate that EPC colony-forming capacity and migratory activity are higher in middle-aged women than in men.