牛乳腺上皮细胞体外分离、培养和鉴定的研究

doi:10.3969/j.issn.1008-0864.2010.04.13

›› 2010, Vol. 12 ›› Issue (4): 68-72.DOI: 10.3969/j.issn.1008-0864.2010.04.13

牛乳腺上皮细胞体外分离、培养和鉴定的研究

李瑞国1,苗朝华2,安晓荣3,陈永福3

(1.中国生物技术发展中心, 北京 100036|2.中国农业科学院生物技术研究所, 北京 100081|3.中国农业大学农业生物技术国家重点实验室, 北京 100094)

收稿日期:2010-06-23 修回日期:2010-07-07 出版日期:2010-08-15 发布日期:2010-07-26
通讯作者: 安晓荣,教授,博士,主要研究方向为动物基因工程。Tel:010-62733463;E-mail: xra@cau.edu.cn
作者简介:李瑞国,副研究员,博士,研究方向为动物生理学。Tel:010-88225162。
基金资助:
国家863计划项目(2002AA206311);国家自然科学基金项目(30871783)资助。

Studies of in Vitro Isolation Culture and Identification of |Bovine Mammary Gland Epithelial Cells

LI Rui-guo1, MIAO Chao-hua2, AN Xiao-rong3, CHEN Yong-fu3

(1.China National Center for Biotechnology Development, Beijing 100036|2.Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081|3.State Key Laboratory for Agro-biotechnology, China Agricultural University, Beijing 100094, China)

Received:2010-06-23 Revised:2010-07-07 Online:2010-08-15 Published:2010-07-26

摘要/Abstract

摘要：

建立可用于基因打靶的乳腺上皮细胞系是乳腺生物反应器研制的重要环节,分别采用胰蛋白酶、胶原酶及其两者的混合消化液分离获得了包括少量成纤维细胞在内的原代牛乳腺上皮细胞,比较了不同消化方法的优缺点。利用Trypsin-EDTA阶段消化法在体外培养牛乳腺上皮细胞,观察了此细胞培养到第5代、10代、15代和20代时的细胞形态特征和生长特性,比较了该细胞在胶原基质和混合型基质包被培养皿中的培养效果。以乳腺腺泡上皮细胞中的细胞特征蛋白角蛋白18为标志,利用角蛋白18单克隆抗体进行了细胞免疫组化鉴定,证明分离培养细胞的上皮特性。

关键词: 牛;乳腺上皮细胞系;体外培养;免疫组化鉴定

Abstract:

Establishment of mammary gland epithelial cell line for gene targeting is an important step for production of mammary gland bioreactor. Primary bovine mammary gland epithelial cell including a few fibroblast cells were isolated by using Trypsin, Collagenase and their combination. The advantages and shortcomings among different isolation methods were compared. Bovine mammary gland epithelial cells were cultured in vitro by using Trypsin-EDTA stage digestion method. Their cell morphology and growth characteristics were observed when the cell was in the 5th, 10th, 15th and 20th generation to compare the growth effects between cells cultured in single collagen coated dish and multifactor coated dish. The cells epithelial characteristic was identified by immunochemical experiment using cytokeratin-18, which is a specific cytokeratin for mammary gland epithelial cell.

Key words: bovine, mammary gland epithelial cell, in vitro, immunochemical identification

中图分类号:

Q954.6-33+2

李瑞国1,苗朝华2,安晓荣3,陈永福3. 牛乳腺上皮细胞体外分离、培养和鉴定的研究[J]. , 2010, 12(4): 68-72.

LI Rui-guo1, MIAO Chao-hua2, AN Xiao-rong3, CHEN Yong-fu3. Studies of in Vitro Isolation Culture and Identification of |Bovine Mammary Gland Epithelial Cells[J]. , 2010, 12(4): 68-72.

牛乳腺上皮细胞体外分离、培养和鉴定的研究

Studies of in Vitro Isolation Culture and Identification of |Bovine Mammary Gland Epithelial Cells

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