BACKGROUND Human endometrium has immense regenerative capacity, growing ~5 mm in 7 days every month. We have previously identified a small population of colony-forming endometrial stromal cells which we hypothesize are mesenchymal stem cells (MSC). The aim of this study was to determine if the co-expression of two perivascular cell markers, CD146 and platelet-derived growth factor-receptor β (PDGF-Rβ), will prospectively isolate endometrial stromal cells which exhibit MSC properties, and determine their location in human endometrium. METHODS Single cell suspensions of human endometrial stromal cells were fluorescence activated cell sorting (FACS) sorted into CD146+PDGF-Rβ+ and CD146−PDGF-Rβ− populations and analysed for colony-forming ability, in vitro differentiation and expression of typical MSC markers. Full thickness human endometrial sections were co-stained for CD146 and PDGF-Rβ. RESULTS FACS stromal CD146+PDGF-Rβ+ stromal cells (1.5% of sorted population) were enriched for colony-forming cells compared with CD146−PDGF-Rβ− cells (7.7 ± 1.7 versus 0.7 ± 0.2% P <0.0001), and also underwent differentiation into adipogenic, osteogenic, myogenic and chondrogenic lineages. They expressed MSC phenotypic surface markers and were located near blood vessels. CONCLUSION This study shows that human endometrium contains a small population of MSC-like cells that may be responsible for its cyclical growth, and may provide a readily available source of MSC for tissue engineering applications.