Isolation and Culture of Human Bone Marrow Mesenchymal Stem Cells

The specimens were collected from The First Affiliated Hospital of Zhengzhou University, which was approved by the ethics committee of the hospital, and the informed consent was obtained and signed. 10 ml of human bone marrow blood was added to Ficoll solution at a proportion of 1:1 in the lower layer of the centrifuge tube, and then, 10 ml of bone marrow blood was slowly added to the upper layer. The centrifugal speed was 2,000 rpm with up and down of 1 for 20 min. After centrifugation, the centrifuge tube was slowly removed. At this time, the liquid was stratified, and the mononuclear cell layer was carefully drawn into a new centrifuge tube. Then, 10 ml of PBS was added and centrifuged at 1,500 rpm for 10 min. One milliliter of culture medium (containing 1% penicillin Streptomycin Solution and 10% Australian fetal bovine serum within DMEM medium) was added to the suspension and inoculated into a 10 cm culture dish. The culture medium was changed every 3 days after inoculation. When the cells reached 80% healing, they were subcultured.