Cao Shanbo, Wang Fang, Liu Lin
The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, China.
Methods Mol Biol. 2013;1074:111-23. doi: 10.1007/978-1-62703-628-3_9.
Isolation and culture of primary embryonic stem (ES) cell colonies are the first critical step towards establishment of stable ES cell lines. Here we introduce a novel method designated as "Separate and Seed" that contributes remarkably to efficient derivation of bovine primary ES cell colonies from blastocysts. The bovine ES cell colonies can self-renew to passage 10 with the growth factors bFGF and BIO. The bovine ES cells exhibit morphology typical of ES cells and express pluripotent molecular markers including Oct4, Nanog, SSEA1, SSEA4, and alkaline phosphatase (AP). These pluripotent markers may be used for the characterization of authentic bovine ES cell lines. Although continued efforts are required for improving long-term culture of bovine ES cells, this novel "Separate and Seed" method plus the growth factors bFGF and BIO provides an initial effective step that may eventually lead to the derivation of authentic bovine ES cells.
原代胚胎干细胞（ES）集落的分离和培养是建立稳定ES细胞系的首要关键步骤。在此，我们介绍一种名为“分离与接种”的新方法，该方法对从囊胚高效获得牛原代ES细胞集落有显著贡献。牛ES细胞集落在添加生长因子bFGF和BIO的情况下可自我更新至第10代。牛ES细胞呈现出典型的ES细胞形态，并表达包括Oct4、Nanog、SSEA1、SSEA4和碱性磷酸酶（AP）在内的多能性分子标志物。这些多能性标志物可用于鉴定真正的牛ES细胞系。尽管仍需持续努力来改善牛ES细胞的长期培养，但这种新的“分离与接种”方法加上生长因子bFGF和BIO提供了一个初步的有效步骤，最终可能会成功获得真正的牛ES细胞。
