组织块再移法分离培养奶山羊乳腺上皮细胞

Study on tissue re-shift method of isolation and culture of goat mammary epithelial cells

通过比较组织块贴壁培养法、组织块再移法、胰蛋白酶消化法、胶原酶Ⅱ消化法和胶原酶消化配合组织块贴壁法分离、培养、纯化山羊乳腺上皮细胞,绘制细胞生长曲线并计算细胞群体倍增时间,研究乳腺上皮细胞培养效果。结果显示,利用胶原酶消化配合组织块贴壁法不能获得正常生长的乳腺上皮细胞;利用组织块贴壁培养法、组织块再移法和胶原酶Ⅱ消化法获得大量的乳腺上皮细胞,所得到的上皮细胞生长曲线呈典型的"S"型,符合细胞生长的一般规律。组织块再移法不仅可快速获得大量纯化的乳腺上皮细胞,而且所得到的细胞经传代后,细胞群体倍增时间最短、增殖能力最强,表明,该法是获得山羊乳腺上皮细胞最适宜的方法。

In this study,in order to obtain a method to culture mammary epithelial cells with high purity in vitro efficiently,5 methods which include adherent tissue culture,tissue re-shift method,trypsin digestion,collagenase Ⅱ digestion and the combination of collagenase digestion with tissue adherence were applied and compared.Goat mammary epithelial cells were isolated,cultured and purified in 5 methods and finally the cell growth curves were made and the cell population doubling time was calculated.The results indicated that the last method was barely efficient whereas adherent tissue culture,tissue re-shift method and collagenase Ⅱdigestion showed high efficiency to get large amount of mammary epithelial cells and the cell growth curve showed a typical ＂S＂ type.It was notable that tissue re-shift method showed high efficiency to get a large number of purified mammary epithelial cells,short cell population doubling time,strong proliferation after cells were obtained after passage,thus suggesting it is an appropriate method to obtain goat mammary epithelial cells.