Isolation, Culture and Cryopreservation of Human Bone Marrow – Derived Mesenchymal Stem Cells

Human bone marrow derived mesenchymal stem cells (BMMSCs), a source of important autologous stem cells without ethical debate, have been used in both experimental and clinical treatment. Therefore, this research aims to isolate, culture and cryopreserve BMMSCs. Mesenchymal stem cells (MSCs) were preserved in each cryovial by slow freezing – rapid thawing method. Post-cryopreserved MSCs was analyzed the viability and the stemness by their expression of some specific markers, differentiation potential and proliferation ability. The results of this research showed it is easy to isolate the BMMSCs by culturing the mononuclear cells derived from human bone marrow in DMEM/F12 plus 15% FBS, 10 ng/ml FGF and 1% antibiotic–mycotic; the viability rates of thawed MSCs were 72.95+/-6.14%. Cryopreserved MSCs maintained the stemness after 1 year storage in nitrogen liquid. They expressed CD13, CD44, CD90, CD166 markers and were negative for CD14, CD34, CD45 and HLA-DR markers in similar to fresh MSCs. In addition, they also owned and maintained adipogenic differentiation potential. So that BMMSCs can be in vitro cultured, proliferated and preserved and used for long periods of time in regenerative medicine.