基于抗体的荧光激活细胞分选法分离哺乳动物卵原干细胞
Isolation of Mammalian Oogonial Stem Cells by Antibody-Based Fluorescence-Activated Cell Sorting.
作者信息
Navaroli Deanna M, Tilly Jonathan L, Woods Dori C
机构信息
Laboratory of Aging and Infertility Research, Northeastern University, 360 Huntington Avenue, Boston, MA, 02115, USA.
Department of Biology, Northeastern University, Boston, MA, 02115, USA.
出版信息
Methods Mol Biol. 2016;1457:253-68. doi: 10.1007/978-1-4939-3795-0_19.
The ability to isolate and subsequently culture mitotically active female germ cells from adult ovaries, referred to as either oogonial stem cells (OSCs) or adult female germline stem cells (aFGSCs), has provided a robust system to study female germ cell development under multiple experimental conditions, and in many species. Flow cytometry or fluorescence-activated cell sorting (FACS) is an integral part of many isolation and characterization protocols. Here, we provide methodological details for antibody-based flow cytometric isolation of OSCs using antibodies specific for external epitopes of the proteins Ddx4 or Ifitm3, alone or in combination with the use of fluorescent reporter mice. Beginning with sample preparation, we provide point-by-point instructions to guide researchers on how to isolate OSCs using flow cytometry.
从成年卵巢中分离并随后培养有丝分裂活跃的雌性生殖细胞的能力，这些细胞被称为卵原干细胞（OSCs）或成年雌性生殖系干细胞（aFGSCs），为在多种实验条件下以及许多物种中研究雌性生殖细胞发育提供了一个强大的系统。流式细胞术或荧光激活细胞分选（FACS）是许多分离和表征方案中不可或缺的一部分。在这里，我们提供了基于抗体的流式细胞术分离OSCs的方法细节，使用针对蛋白质Ddx4或Ifitm3外部表位的特异性抗体，单独使用或与荧光报告小鼠结合使用。从样品制备开始，我们提供逐点说明，以指导研究人员如何使用流式细胞术分离OSCs。
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