Vibratome sectioning for enhanced preservation of the cytoarchitecture of the mammalian organ of Corti.
Vibratome sectioning for enhanced preservation of the cytoarchitecture of the mammalian organ of Corti.
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DOI：
10.3791/2793
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发表时间：
2011-06
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影响因子：
--
通讯作者：
K. Shim
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文献类型：
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作者：
K. Shim
哺乳动物的Corti器官是由机械感觉毛发和非感觉支持细胞组成的高度有序的细胞马赛克（见（1,2））。这种细胞马赛克的可视化通常需要Corti器官的横切面。特别是，非感觉柱和deiter细胞，其细胞核位于毛细胞的基部，如果没有Corti器官的横切面，就无法看到。然而，哺乳动物Corti器官的精细细胞结构，包括柱和deiter细胞的精细细胞质过程，很难通过常规的组织学方法（如石蜡和冷冻切片）保存，这些方法与标准的免疫组织化学染色技术兼容。在这里，我描述了一个简单而强大的程序，包括耳蜗振动体切片，整个坐垫的这些振动体切片的免疫组织化学染色，然后是共聚焦显微镜。该方法已广泛用于多种器官的免疫组织化学分析，包括小鼠肢体芽、斑马鱼肠道、肝脏、胰腺和心脏（见（3-6）所选的例子）。此外，该方法在胚胎和成年小鼠的Corti突变体和对照器官的柱细胞数量成像和定量方面都取得了成功(7)。然而，这种方法目前尚未广泛用于检查哺乳动物的Corti器官。该程序的潜力，既提供增强保存的优良细胞结构的Corti成人器官和允许各种细胞类型的定量描述。
The mammalian organ of Corti is a highly ordered cellular mosaic of mechanosensory hair and nonsensory supporting cells (reviewed in (1,2)).Visualization of this cellular mosaic often requires that the organ of Corti is cross-sectioned. In particular, the nonsensory pillar and Deiters' cells, whose nuclei are located basally with respect to the hair cells, cannot be visualized without cross-sectioning the organ of Corti. However, the delicate cytoarchitecture of the mammalian organ of Corti, including the fine cytoplasmic processes of the pillar and Deiters' cells, is difficult to preserve by routine histological procedures such as paraffin and cryo-sectioning, which are compatible with standard immunohistochemical staining techniques. Here I describe a simple and robust procedure consisting of vibratome sectioning of the cochlea, immunohistochemical staining of these vibratome sections in whole mount, followed by confocal microscopy. This procedure has been used widely for immunhistochemical analysis of multiple organs, including the mouse limb bud, zebrafish gut, liver, pancreas, and heart (see (3-6) for selected examples). In addition, this procedure was sucessful for both imaging and quantitificaton of pillar cell number in mutant and control organs of Corti in both embryos and adult mice (7). This method, however, is currently not widely used to examine the mammalian organ of Corti. The potential for this procedure to both provide enhanced preservation of the fine cytoarchitecture of the adult organ of Corti and allow for quantification of various cell types is described.