Point by Point: An Introductory Guide to Sample Preparation for Single-Molecule, Super-Resolution Fluorescence Microscopy.
Point by Point: An Introductory Guide to Sample Preparation for Single-Molecule, Super-Resolution Fluorescence Microscopy.
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DOI：
10.1002/9780470559277.ch140241
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发表时间：
2015-06-01
影响因子：
--
通讯作者：
Vaughan, Joshua C
中科院分区：
文献类型：
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作者：
Halpern, Aaron R;Howard, Marco D;Vaughan, Joshua C
单分子、基于定位的超分辨率显微镜能够在经典的250 nm衍射极限下揭示详细的亚细胞结构和蛋白质分布，但有效地利用这项技术需要仔细的样品准备、数据采集和数据分析的组合，这可能会让新手研究人员望而却步。在该方案中，提供了关于制备用于超分辨率显微镜的坚固的参考样品的详细说明，包括细胞骨架(微管)、膜结合的细胞器(过氧化物体)和支架蛋白(网状蛋白)。这些样品也构成了生物研究人员感兴趣的具有代表性的成像目标的子集，并突出了样品制备方面的差异和相似之处。
Single-molecule, localization-based, super resolution microscopy is able to reveal detailed subcellular structures and protein distributions below the classical 250-nm diffraction limit of light, but utilizing this technique effectively requires a combination of careful sample preparation, data acquisition, and data analysis, which can be daunting to novice researchers. In this protocol, detailed instructions on preparation of robust reference samples for super-resolution microscopy, including the cytoskeleton (microtubules), membrane-bound organelles (peroxisomes), and scaffold proteins (clathrin), are provided. These samples also constitute a representative subset of imaging targets of interest to biological researchers and highlight the differences and similarities in sample preparation.