Human mesenchymal stem cells (hMSCs) have shown great potential for therapeutic purposes. However, the low frequencies of hMSCs in the body and difficulties in expanding their numbers in vitro have limited their clinical use. In order to develop an alternative strategy for the expansion of hMSCs in vitro, we coated tissue culture polystyrene with keratins extracted from human hair and studied the behavior of cells from 2 donors on these surfaces. The coating resulted in a homogeneous distribution of nanosized keratin globules possessing significant hydrophilicity. Results from cell attachment assays demonstrated that keratin-coated surfaces were able to moderate donor-to-donor variability when compared with noncoated tissue culture polystyrene. STRO-1 expression was either sustained or enhanced on hMSCs cultured on keratin-coated surfaces. This translated into significant increases in the colony-forming efficiencies of both hMSC populations, when the cells were serially passaged. Human hair keratins are abundant and might constitute a feasible replacement for other biomaterials that are of animal origin. In addition, our results suggest that hair keratins may be effective in moderating the microenvironment sufficiently to enrich hMSCs with high colony-forming efficiency ex vivo, for clinical applications.