Expansion Microscopy: Protocols for Imaging Proteins and RNA in Cells and Tissues.
Expansion Microscopy: Protocols for Imaging Proteins and RNA in Cells and Tissues.
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DOI：
10.1002/cpcb.56
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发表时间：
2018-09-01
影响因子：
--
通讯作者：
Boyden, Edward S
中科院分区：
文献类型：
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作者：
Asano, Shoh M;Gao, Ruixuan;Boyden, Edward S
膨胀显微镜 (ExM) 是一项最近开发的技术，可通过在成像前对样本进行各向同性物理膨胀，在传统的衍射极限显微镜上对保存的细胞和组织进行纳米级分辨率成像。在 ExM 中，样本中的生物分子和/或荧光标记与均匀合成在整个样本中的致密可膨胀聚合物基质相连，当浸入水中时，该基质会发生 3 维膨胀 4.5 倍线性。自从我们的第一份报告以来，针对蛋白质、RNA 和其他生物分子可视化而优化的 ExM 版本已经出现。在这里，我们描述了使用典型生物实验室中的化学品和硬件进行蛋白质（蛋白质保留 ExM 或 proExM）以及 RNA（扩增荧光原位杂交或 ExFISH）分析的最佳实践、分步 ExM 方案。此外，还提供了处理和安装扩展样品以及使用共焦和光片显微镜对其进行成像的详细协议。 © 2018 John Wiley & Sons, Inc. 版权所有
Expansion microscopy (ExM) is a recently developed technique that enables nanoscale-resolution imaging of preserved cells and tissues on conventional diffraction-limited microscopes via isotropic physical expansion of the specimens before imaging. In ExM, biomolecules and/or fluorescent labels in the specimen are linked to a dense, expandable polymer matrix synthesized evenly throughout the specimen, which undergoes 3-dimensional expansion by 4.5 fold linearly when immersed in water. Since our first report, versions of ExM optimized for visualization of proteins, RNA, and other biomolecules have emerged. Here we describe best-practice, step-by-step ExM protocols for performing analysis of proteins (protein retention ExM, or proExM) as well as RNAs (expansion fluorescence in situ hybridization, or ExFISH), using chemicals and hardware found in a typical biology lab. Furthermore, a detailed protocol for handling and mounting expanded samples and for imaging them with confocal and light-sheet microscopes is provided. © 2018 by John Wiley & Sons, Inc.