Culture and Identification of Mesenchymal Stem Cells Derived from Umbilical Cord Blood

Objective: To establish a method of purification and culturing of mesenchymal stem cells(MSCs) derived from umbilical cord blood and to investigate their biological characteristics.Methods: MSCs were obtained from umbilical cord blood of term deliveries after the mothers signed a donation form.The culture bottles were pretreated coating with self plasma.After 72-96 hours of primary culture,one half of the medium was renewed and non-adherent cells were removed.The medium was renewed by half in every 3-5 days until MSCs reached 80%-90% fusion,then they were subcultured with a propotion of 1∶2.After the third passage,the cells were analyzed with flow cytometry.The following monocloned antibodies as CD29,CD34,CD44 and CD45 were used.The growth cycle and morphological changes of MSCs were monitored by phase contrast microscopy.Results: A cell line derived from umbilical cord blood was acquired which possessed morphological and immunophenotypical characteristics similar to those of MSCs isolated from bone marrow.The potential of proliferation remained unchanging over five passages when the cells were cultured and maintained at low differentiation.By the flow cytometric analysis,CD29 and CD44 were expressed on MSCs while no expression of CD34 and CD45 were found.Conclusion: The presence of MSCs in umbilical cord blood of term infants has been demonstrated,and MSCs can be purified and cultured in vitro for the further studies of the stem cells.