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【目的】探索可靠的小鼠心脏干细胞（ cardiac stem cells ，CSCs）分离方法，并对体外培养的 CSCs进行表型鉴定。【方法】选用 SPF 级3～4周龄 ICR 小鼠，无菌条件下处理心脏组织，胶原酶 A 消化，40μm 筛网过滤，制备单细胞悬液，利用免疫磁珠分选法结合抗干细胞抗原‐1（stem cell antigen‐1，Sca‐1）的磁珠抗体来分离 Sca‐1阳性干细胞，并进一步行免疫荧光、流式细胞及基因表达等鉴定。【结果】利用磁珠分选法结合特异的抗 Sca‐1的磁珠抗体，成功地从小鼠心脏组织中分离出 Sca‐1阳性细胞群。这些细胞具有小、圆、亮等特征，能在体外培养扩增，传代后仍绝大部分表达 Sca‐1抗原，但基本不表达内皮系或造血细胞系的特异抗原 c‐kit 、Flk‐1、CD31、CD34、CD45等。这些 Sca‐1阳性细胞表达多能干细胞特征的 Nanog 、TERT 基因，心脏胚层特异的 ISL‐1、TBX5基因及心肌特异性转录因子 GATA4、Nkx2．5、MEF2C ，但不表达成熟心肌的标志基因α‐M HC 。【结论】利用磁珠分选法结合抗特异干细胞表面标志物 Sca‐1的磁珠抗体能在短期内获得高纯度的Sca‐1阳性 CSCs ；小鼠 Sca‐1阳性 CSCs 能在体外培养增殖，有助于 CSCs 相关的组织工程或分子机制的实验研究。“,”Objective]To establish a reliable method of isolating cardiac stem cells (CSCs) from murine heart and perform further phenotypic identification after in vitro culturing .[Methods] The ICR mice aged 3 ~4 weeks were selected .Their hearts were processed in aseptic conditions ,digested in collagen A and filtered through a 40 μl strainer to obtain single‐cell suspension ultimately .Then Sca‐1 positive (Sca‐1 + ) stem cells were isolated and sorted from single‐cell suspension by immunomagnetic cell sorting (MACS) and microbead conjugated anti‐mouse Sca‐1 antibody .Then the cells were cultured in vitro for further identification of cell surface antigens and gene expression .[Results] Sca‐1 + cells were successfully harvested with a high purity from murine hearts .These Sca‐1 + cells could proliferate gradually in attachment culture .They were charac‐terized by small size ,round or oval shape and high brightness .Most still expressed Sca‐1 antigen and rarely expressed c‐kit ,Flk‐1 ,CD31 ,CD34 and CD45 .There were also the expressions of Nanog ,TERT ,ISL‐1 , TBX5 ,GATA4 ,Nkx2 .5 and MEF2C ,but not α‐M HC .So their phenotypes expressed biomarkers of mutipo‐tential stem cells and cardiac precursors , but those of endothelial , hematopoietic cells or cardiomyocytes .[Conclusion] MACS with microbead conjugated anti‐mouse Sca‐1 antibody may isolate Sca‐1 + stem cells from murine hearts .And these Sca‐1 + cells are confirmed as Sca‐1 + CSCs .Further in vitro culturing and propaga‐tion of these cells shall facilitate future studies on cardiac tissue engineering and molecular mechanisms of car ‐diac cells .