小鼠胚胎成纤维细胞分离培养研究

Isolation and Culture Research on Mouse Embryonic Fibroblasts

为探索实验室影响小鼠胚胎成纤维细胞（MEF）分离和培养的因素,取昆明白小鼠胚胎分离成纤维细胞,利用体外培养体系,对不同胎龄、不同血清浓度、不同胰蛋白酶浓度和作用时间对MEF分离及培养的影响进行研究,观察MEF的生长形态及其生物学特性。结果表明,在实验室条件下,12.5-14.5 d胎龄的小鼠胎儿MEF分离效果最好;最佳培养基为DMEM添加15%小牛血清;第3代细胞增殖最旺盛;室温条件下,0.125%胰蛋白酶消化MEF时间以8-10 min为宜。在培养ES细胞时,应选择第3-5代的MEF作为饲养层细胞。研究获得了可在实验室分离制备MEF的方法,为实验室进一步分离培养ES细胞奠定基础。

To research the factors which influence the isolation and culture of mouse fibroblast in our lab, the mouse primary embryonic fibroblasts were isolated from mouse fetus of Kunming white at different gestational ages. According to the influence of different serum and trypsin concentration and the digestion time on MEF, the growth behavior and the characteristics of MEF were studied. Results showed that the optimal gestational age for isolation of mouse embryonic fibroblasts was from 12.5 to 14.5 day. At the room temperature, the optimal time for digesting the mouse fetus tissue was 8 - 12 min with digestion solution containing 0. 125% of trypsin. The mouse embryonic fibroblasts were successfully isolated and cultured, which layed the foundation for further isolation and culture of ES ceells.