小鼠胚胎成纤维细胞的分离与培养

目的 探讨影响小鼠胚胎成纤维细胞（MEF）分离和培养的一些因素,以建立稳定的MEF培养体系。方法 取BALB／c小鼠胚胎分离成纤维细胞,利用体外培养体系,对MEF的生长形态,生长曲线及细胞周期进行观察,并对不同胚胎日龄以及不同胰酶作用时间对MEF分离及培养的影响进行研究。结果 13.5d胎龄鼠胚的MEF分离效果优于10.5d,18.5d胎龄鼠胚;MEF在体外为贴壁生长型细胞,第三代细胞增殖旺盛;在室温条件下,0.25%胰酶消化MEF时间以3－5min为宜。结论 MEF在第3代增殖旺盛,最适宜作胚胎干细胞的饲养层。

Isolation and Culture of Mouse Embryonic Fibroblast

Objective To establish a stable culture system of mouse embryonic fibroblast(MEF). Methods We isolated MEF from the embryos of BALB/c mouse and investigated the morphology, growth curve and cell cycle of MEF in vitro. The effects of the gestational age (days) of mouse embryos and the time of digestion on the isolation and culture of MEF were assessed. Results The mouse embryo of 13.5 d is better than that of 10.5 d and 18.5 d on the isolation of MEF. MEF in vitro is a kind of adherent cell with good ability for proliferation in passage 3. In room temperature, the digestive time of 0.25% trypsin should be 3 5 min. Conclusion MEFs(P3) are quite suited to be a feeder layer of embryonic stem cells.