Isolation of outer hair cells from the cochlear sensory epithelium in whole-mount preparation using laser capture microdissection
Isolation of outer hair cells from the cochlear sensory epithelium in whole-mount preparation using laser capture microdissection
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DOI：
10.1016/j.jneumeth.2007.01.017
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发表时间：
2007-05-15
影响因子：
3
通讯作者：
Zheng, Jing
中科院分区：
文献类型：
--
作者：
Anderson, Charles T.;Zheng, Jing
外毛细胞（OHC）在哺乳动物耳蜗的频率选择性和信号放大中起重要作用。由于OHC的数量相对较少，并且是耳蜗中的少数细胞，因此分离和分离它们用于cDNA文库创建和蛋白质组学研究等应用是一项具有挑战性的任务。激光捕获显微切割（LCM）旨在从非常薄的组织切片中捕获细胞，它可以从大面积组织中准确分离特定细胞，用于RNA，DNA和蛋白质组学研究。由于耳蜗解剖结构的限制，耳蜗的薄切片包含少量的OHC。因此，我们采用LCM技术从Corti器官全标本中分离OHC，每个标本都含有数百个同时可接近和可收集的OHC。为了比较，我们还使用了更传统的机械剥离。比较了LCM法和传统机械法提取外毛细胞cDNA的质量，并讨论了两种方法的优缺点。类似的方法也可以用于从整装耳蜗制备物中分离大量的内毛细胞和选择的支持细胞。(C)2007 Elsevier B.V.保留所有权利。
Outer hair cells (OHCs) play an important role in frequency selectivity and signal amplification in the mammalian cochlea. Because OHCs are relatively few in number and a minority of the cells in the cochlea, separating and isolating them for applications such as cDNA library creation and proteomic studies is a challenging task. Laser capture microdissection (LCM) is designed to capture cells from very thin tissue sections, it can accurately isolate specific cells from large regions of tissue for RNA, DNA, and proteomic studies. Due to the constraints of cochlear anatomy, thin sections of the cochlea contain small numbers of OHCs. Therefore, we adapted the LCM technique to isolate OHCs from organ of Corti whole-mounts, each of which contain hundreds of OHCs that are simultaneously accessible and collectable. For comparison, we also used a more traditional mechanical dissection. The quality of cDNA derived from the OHCs collected with LCM and with the traditional mechanical method are compared and the merits and limitations of the techniques discussed. A similar approach can also be used to isolate large quantities of inner hair cells and selected supporting cells from the whole-mount cochlear preparation. (C) 2007 Elsevier B.V. All rights reserved.