Expansion Microscopy of Ciliary Proteins.
Expansion Microscopy of Ciliary Proteins.
复制标题
纤毛蛋白的放大显微镜。
DOI：
10.1007/978-1-0716-3507-0_4
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发表时间：
2024
期刊：
影响因子：
--
通讯作者：
Shi,Xiaoyu
中科院分区：
文献类型：
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作者：
Park,Sohyeon;Shi,Xiaoyu
标记保留扩展显微镜(LR-EXM)是一种样品制备技术，它将细胞或组织嵌入到可膨胀的水凝胶中，并对样品进行膨胀，使其能够达到任何传统荧光显微镜的高分辨率。聚合过程中的荧光损失和蛋白质变性一直是标准膨胀显微镜的主要局限性。为了最大限度地减少荧光损失，LR-EXM使用了三官能团锚，这种锚可以在聚合和变性过程中存活下来，然后在扩张后引入荧光团。通过使用LR-EXM，人们可以在分子尺度上研究初级纤毛的结构，与以前介绍的扩张显微镜方法相比，具有更高的信噪比。在这一章中，我们描述了一个详细的程序，展示了LR-exm是如何用于研究睫状蛋白的。
Label-retention expansion microscopy (LR-ExM) is a sample preparation technique, which embeds the cells or tissues in a swellable hydrogel and expands the sample so that one can achieve a high resolution with any conventional fluorescence microscopes. Fluorescence loss during polymerization and protein denaturation have been a major limitation of standard expansion microscopy. To minimize fluorescence loss, LR-ExM uses trifunctional anchors, which can survive from polymerization and denaturation, and then introduce fluorophores after expansion. By using LR-ExM, one can study the structure of primary cilia at molecular-scale resolution with a much higher signal-to-noise ratio, compared with previously introduced expansion microscopy methods. In this chapter, we describe a detailed procedure showing how LR-ExM is used to study ciliary proteins.