Sample Preparation for STED Microscopy
Sample Preparation for STED Microscopy
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DOI：
10.1007/978-1-60761-404-3_11
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发表时间：
2010-01-01
期刊：
影响因子：
--
通讯作者：
Jakobs, Stefan
中科院分区：
文献类型：
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作者：
Wurm, Christian A.;Neumann, Daniel;Jakobs, Stefan
自从十九世纪末发现衍射势垒以来，人们普遍认为，使用远场光学显微镜不可能解析比光波长的一半更精细的结构细节。 STED 显微镜的出现表明，至少对于荧光成像来说，这些限制是可以克服的。由于 STED 显微镜是一种远场技术，原则上可以使用与传统共焦显微镜相同的样品制备方法。然而，分辨率的提高需要额外的预防措施，以确保样本的结构保存。我们提出了强大的协议来生成 STED 显微镜测试样本。这些珠子样品和免疫标记哺乳动物细胞的方案可用作起点，以适应现有的标记策略以满足亚衍射分辨率显微镜的要求。
Since the discovery of the diffraction barrier in the late nineteenth Century, it has been commonly accepted that with far-field optical microscopy it is not possible to resolve structural details considerably finer than half the wavelength of light. The emergence of STED microscopy showed that, at least for fluorescence imaging, these limits can be overcome. Since STED microscopy is a far-field technique, in principle, the same sample preparation as for conventional confocal microscopy may be utilized. The increased resolution, however, requires additional precautions to ensure the structural preservation of the specimen. We present robust protocols to generate test samples for STED microscopy. These protocols for bead samples and immunolabeled mammalian cells may be used as starting points to adapt existing labeling strategies for the requirements of sub-diffraction resolution microscopy.