目的探讨人骨髓间充质干细胞(hMSCs)体外分离培养、鉴定和标记的方法以及生物学特性。方法无菌条件下穿刺人髂后上嵴抽取骨髓,采用直接贴壁法分离纯化hMSCs,体外扩增,用免疫细胞化学法及流式细胞仪分别对培养的hMSCs进行鉴定,同时检测第3代和同代溴脱氧尿嘧啶(5bromo2’deoxyuridine,BrdU)标记的hMSCs细胞周期。结果体外培养的原代hMSCs在培养24小时内可见少量贴壁细胞,7天达到融合,免疫细胞化学示CD44和CD105表达阳性,CD34表达阴性,流式细胞仪示CD44阳性率为89.64%,CD34为0.11%。大部分hMSCs核抗BrdU染色阳性,阳性率达90%。细胞周期显示第3代和同代BrdU标记的hMSCs约有90%的细胞处于G0/G1期,第10代为80%。结论hMSCs在体外很容易分离培养和扩增,但随着传代次数的增加,hMSCs变得宽大扁平且增殖速度减慢,出现衰老现象;用BrdU标记hMSCs对其细胞周期无明显影响,可作为标记细胞的一种有效手段。 OBJECTIVE: To study the isolation, cultivation, identification and labeling of human bone marrow mesenchymal stem cells (hMSCs) in vitro and their biological characteristics. Methods The bone marrow was isolated from the posterior crista of human iliac under sterile conditions. The hMSCs were isolated and purified by direct adherent method. The hMSCs were expanded in vitro. The cultured hMSCs were identified by immunocytochemistry and flow cytometry. And bromodeoxyuridine (BrdU) labeled hMSCs. Results Primary hMSCs cultured in vitro showed a small amount of adherent cells within 24 hours after culturing, and fused for 7 days. Immunocytochemistry showed positive expression of CD44 and CD105, negative expression of CD34. The positive rate of CD44 in flow cytometry was 89.64% 0.11%. The majority of hMSCs were positive for BrdU staining, with a positive rate of 90%. The cell cycle showed that about 90% of the cells in the third and the generation BrdU-labeled hMSCs were in the G0 / G1 phase, and the tenth generation was 80%. Conclusions hMSCs can be easily isolated and cultured in vitro. However, hMSCs become flat and flat and the proliferation rate slows down with the passage of time. Senescence phenomenon is observed in hMSCs. The hMSCs labeled with BrdU have no effect on the cell cycle and can be used as markers An effective means of cells.