牛奶中乳腺上皮细胞的分离培养及其鉴定.pdfVIP

生物技术通报 ·研究报告· BIOTECHNOLOGY BULLETIN 年第 期 2013 3 牛奶中乳腺上皮细胞的分离培养及鉴定 崔新洁  王婷  刘秉春  陶林  王潇 （内蒙古大学生命科学学院，呼和浩特 010010） 摘 要 ： 旨在建立简便、经济、纯净的牛乳腺上皮细胞的体外培养方法，并对所培养细胞进行鉴定。以新鲜牛奶为材料， 离心收集细胞进行原代培养并传代，通过测定生长曲线、乳腺上皮细胞骨架蛋白-角蛋白8 和18、β-酪蛋白、核型等对所培养细胞 进行鉴定，以确定其为乳腺上皮细胞。离心收集到的细胞原代培养3 d 后形成多个单克隆，细胞呈典型的铺路石状，同时伴随少量 的吞噬细胞，继续培养至单克隆汇合时，细胞表层产生大小不等的乳滴，吞噬细胞大多转移到乳滴中，传代后，得到纯净的乳腺 上皮细胞，吞噬细胞随传代而消失 ；细胞生长曲线呈 “S”型，RT-PCR 法扩增到了乳腺上皮细胞的骨架蛋白8、18 以及β-酪蛋白， 细胞免疫组化检测到了细胞角蛋白8 ，核型为29 对端着丝粒常染色体，1 对近端着丝粒性染色体。通过以上试验证明，乳汁分离 法可以培养纯净的、具有正常分裂增生能力的牛乳腺上皮细胞，建立了乳腺上皮细胞的体外培养模型。 关键词 ： 乳汁 牛乳腺上皮细胞 细胞骨架蛋白 泌乳 ， Isolation Culture and Identification of Bovine Mammary Epithelial Cell Cui Xinjie Wang Ting Liu Bingchun Tao Lin Wang Xiao （College of Lif e Sciences ，Inner Mongolia University ，Hohhot 0 100 10 ） It was to set up a simple, economic and pure method to culture and identify bovine mammary epithelial cell （BMEC ） Abstract:  in vitro, which were collected from centrifuging the fresh milk. Then, the cell growth curve, keratin8, keratin 18, Beta casein and karyotype were identified. After three days, several cells grew together, and the shape looked like typical paving stones. At the same time, there were also a few phagocytes. Continue to culture several days, all cells grew together, and there were some emulsion droplets secreted to the outside of the cells with almost all phagocytes transferring into the emulsion droplets. When the cells