目的探讨脐带的间充质干细胞分化为成骨细胞的影响因素和免疫组织化学鉴定。方法收集306例冻存的健康胎儿脐带,采用华尔通胶组织块贴壁法从脐带组织中分离间充质干细胞,利用荧光显微镜观察原代细胞的细胞形态。脐带间充质干细胞的免疫表型和细胞周期采用免疫组织化学检测。复苏冻存的脐带间充质干细胞,并传代培养至10代。对第10代的脐带间充质干细胞进行成骨诱导,其成骨能力分别通过钙结节和骨涎蛋白的免疫荧光检测以及茜素红染色检测来确定。结果免疫组织化学结果显示,培养的细胞高表达间充质干细胞的表面标志物CD73、CD90和CD105,但是不表达造血细胞的表面标志物CD34和CD45。复苏后细胞的存活率是90%。细胞周期显示,第10代的细胞有80%处于G0/G1期,20%处于S+G2/M期。成骨细胞刺激因子诱导干细胞的骨涎蛋白染色呈阳性,并形成矿化的钙结节。结论冻存的脐带间充质干细胞在成骨细胞刺激因子下能被诱导分化为成骨细胞,具有高度自我增殖和多向分化能力。 Objective To investigate the factors that affect the differentiation of umbilical cord mesenchymal stem cells into osteoblasts and identify by immunohistochemistry. Methods Umbilical cord blood samples were collected from 306 frozen fetuses. Mesenchymal stem cells (MSCs) were isolated from umbilical cord tissue by Walton gum tissue adhesion method. The morphology of primary cells was observed by fluorescence microscopy. The immunophenotype and cell cycle of umbilical cord mesenchymal stem cells were detected by immunohistochemistry. Recovery of cryopreserved umbilical cord mesenchymal stem cells, and subculture to 10 generations. The 10th generation of umbilical cord mesenchymal stem cells were induced to osteogenesis, and their osteogenic capacity was determined by immunofluorescent detection of calcium nodules and bone sialoprotein, and alizarin red staining. Results Immunohistochemical results showed that the cultured cells highly expressed the mesenchymal stem cell surface markers CD73, CD90 and CD105 but did not express the hematopoietic cell surface markers CD34 and CD45. The survival rate of cells after resuscitation is 90%. The cell cycle showed that 80% of the cells in passage 10 were in G0 / G1 phase and 20% in S + G2 / M phase. Osteoblast stimulating factor-induced bone sialoprotein staining of stem cells was positive and formed mineralized calcium nodules. Conclusion The cryopreserved umbilical cord mesenchymal stem cells can be induced to differentiate into osteoblasts under osteoblast stimulating factor, with a high degree of self-proliferation and multi-directional differentiation.