荷斯坦奶牛乳腺上皮细胞的分离培养

目的：建立荷斯坦奶牛乳腺上皮细胞的分离培养方法。方法：采用组织块种植法培养奶牛乳腺上皮细胞,利用胰蛋白酶差时消化法分离、纯化上皮细胞。结果：成功培养出奶牛乳腺上皮细胞,显微镜下观察,纯化的乳腺上皮细胞呈典型上皮细胞形态,细胞之间排列紧密,呈鹅卵石铺路样,形态均一,多角形的单层聚集。通过荧光免疫细胞染色方法对细胞骨架蛋白-角蛋白18进行鉴定,呈现阳性反应。乳腺上皮细胞增殖旺盛,经25次以上传代后长势仍然良好。结论：采用组织块种植法结合胰酶差时消化法成功获得纯化的奶牛乳腺上皮细胞。

Isolation and culture of Holstein-bovine mammary epithelial cells

Objective：To establish the methods of isolation and culture of Holstein-bovine mammary epithelial cells.Methods：Tissues explant was employed to culture bovine mammary epithelial cells and trypsin digesting from time interval was used to purify the cells.Results：The mammary gland epithelial cells were cultivating.When observed by microscope,the purified cells showed typical characters of epithelial cells,tightly arranging cell to cell,flagstone-like,homogeneous cell morphology.The tissue specific expression of cytokeratin 18 gene in mammary epithelial cells was detected by immunofluorescent cytochemical staining.Bovine mammary gland epithelial cells proliferated exuberantly when cultured up to passage 25.Conclusion：Purified bovine mammary gland epithelial cells were obtained by using tissues explant culture and trypsin digesting from time interval.