Quick and reliable method for retina dissociation and separation of rod photoreceptor perikarya from adult mice
Quick and reliable method for retina dissociation and separation of rod photoreceptor perikarya from adult mice
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DOI：
10.1016/j.mex.2015.01.002
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发表时间：
2015-01-01
期刊：
影响因子：
1.9
通讯作者：
Solovei, Irina
中科院分区：
文献类型：
--
作者：
Feodorova, Yana;Koch, Mirja;Solovei, Irina
一个纯的和丰富的人口成年杆perikarya可以利用在不同的研究有关的核功能，如基因表达分析，旨在阐明细胞类型和疾病之间的关系[1]。分选是基于特定的细胞表面标记或荧光标记的报告蛋白。在这里，我们描述了一种简单可靠的方法，分离视杆细胞胞体，而不使用染色程序或转基因小鼠。然而，这种方法仅限于从成年小鼠视网膜中分选视杆细胞。成熟的视杆细胞具有倒置的核结构，这是由这些核的光学功能决定的[2]。细胞核中心异染色质的高背向散射可以作为通过前向散射和侧向散射进行FAC-sorting的选择标准。木瓜蛋白酶分离系统分离视网膜的方法（Wothington Biochemical Corporation）进行了优化。设计了一种易于遵循的分步视网膜解离方案。仅基于细胞核中异染色质的高反向散射，通过前向和侧向散射进行分类。(C)2015作者由爱思唯尔公司出版
A pure and abundant population of adult rod perikarya can be exploited in different studies concerning nuclear functions such as gene expression analyses which aim at elucidating the relationship between cell type and disease [1]. Sorting is based either on specific cell-surface markers or fluorescently labeled reporter proteins. Here, we describe a simple and reliable method for separation of rod photoreceptor perikarya without the use of staining procedures or transgenic mice. This method is limited, however, to sorting rod photoreceptors from adult mouse retina. Mature rods possess an inverted nuclear architecture which is determined by the optical functions of these nuclei [2]. The high backscatter of heterochromatin in the core of the nucleus can be used as a selection criterion for FAC-sorting by forward and sideward scatter.The procedure for retina dissociation using the Papain Dissociation System (Wothington Biochemical Corporation) was optimized.An easy to follow step-by-step protocol for retina dissociation was devised.Rod perikarya were FAC- sorted by forward and sideward scatter based solely on the high backscatter of heterochromatin in their nuclei. (C) 2015 The Authors. Published by Elsevier B.V.