Objective: Adult stem cells such as mesenchymal stem cells (MSCs) are multipotent. Because of their low immunogenicity MSCs significantly can be used in cell therapy, regenerative medicine and clinical applications. MSCs have been used in the transplant of many organs such as liver, kidneys and heart and also are able to treat many disorders. However, isolation of MSCs from adult tissues such as bone marrow and adipose tissues requires invasive procedures and the availability of an appropriate donor. The amount of MSCs that can be gained from a single donor is limited and the capability of these cells for long-term proliferation is quite poor. ESCs can differentiate into MSCs in certain culture conditions. ESC–derived MSCs could provide unlimited MSCs source for several clinical treatments for instance, mesodermal tissues repair and even for improvement of human ESC-derived HSCs engraftment. Moreover, in a study while treated with IFN-g, ESC–derived MSCs expressed lower HLA-DR than BM-derived MSCs. Materials and Methods: In this research, we derived hMSCs from human ESCs. hESCs were cultured on MEFs that were inactivated by mitomycin C, after that hESCs were passaged mechanically using glass beads and formed embryoid bodies in petridishes (EBs), EBs were then spread into well plates using different medias (hESCs media, DMEM with 10% FBS, DMEM with 20% FBS), later cells were passaged into flasks and finally MSCs were characterized by immunolocalisation using confocal microscopy. Results: As a result, the ESC–derived MSCs adhered to the flask and had a spindle-shaped that resembled BM-derived MSCs morphology. Also ESC–derived MSCs were positive for CD44, CD105, CD29 markers the same as BM-derived MSCs. Furthermore MSCs were negative for CD31 and CD45 markers as expected. Conclusion: Our ESC–derived MSCs had two of the three criteria to be defined as MSCs; they adhered to plastic in standard culture conditions. Also MSCs expressed specific cell surface antigens and lack certain other antigens. However the MSCs should in addition have the ability to differentiate into osteocytes, adipocytes and chondrocytes that needs further experiments to be done. In conclusion, large amount of MSCs can be derived from clinically compliant hESCs. However, additional in vivo studies are required to direct differentiate MSCs into specific tissue types for clinical use. Also gain more knowledge of how MSCs reduce immune responses after transplantation is necessary.