小鼠胚胎干细胞的分离培养与鉴定

ISOLATION, CULTURE AND IDENTIFICATION OF MURINE EMBRYONIC STEM CELLS OF C57BL/ 6 MICE

目的:培养、分离和鉴定C57BL/6小鼠胚胎干细胞(embryonicstemcell,ESC)。方法:收集小鼠3·5d的囊胚进行培养,用小鼠胚胎成纤维细胞作为饲养层,形成的ESC样集落,经两次亚克隆法分离培养成ESC;进行相差显微镜观察,采用阶段特异性胚胎抗原-1(stage-spesificembryonicantigen,SSEA-1)对培养的ESC进行免疫组化染色。结果:获得了稳定的ESC集落,传至第12代。ESC呈集落样生长,SSEA-1免疫组化染色强阳性。结论:两次亚克隆法获得的细胞具有ESC主要的生物学特性。

Objective： To establish the embryonic stem cell （ESC） lines from C57BL/ 6 species mouse by using the fibroblast cell feeder layers. Methods： The blastocysts of 3. 5 d from C57BL/ 6 species mouse were collected and cultured on the fibroblast cell feeder layers. The ES cell-like colonies were sub-clonally cultured twice and the cells were observed with phase contrast microscope and identified by SSEA-1 immunostaining. Result： The stable ESC clones were obtained and were passaged to the 12th generation. ESC was colony-like, and SSEA-1 immunostaining is positive. Conclusion： The cells obtained through twice sub-clonally culturing still maintain the main characteristics of ESC.