Thiele Jan Christoph, Jungblut Marvin, Helmerich Dominic A, Tsukanov Roman, Chizhik Anna, Chizhik Alexey I, Schnermann Martin J, Sauer Markus, Nevskyi Oleksii, Enderlein Jörg
Third Institute of Physics-Biophysics, Georg August University, 37077 Göttingen, Germany.
Department of Biotechnology and Biophysics, Biocenter, University of Würzburg, Am Hubland, 97074 Würzburg, Germany.
Sci Adv. 2022 Jun 10;8(23):eabo2506. doi: 10.1126/sciadv.abo2506. Epub 2022 Jun 8.
Over the past two decades, super-resolution microscopy has seen a tremendous development in speed and resolution, but for most of its methods, there exists a remarkable gap between lateral and axial resolution, which is by a factor of 2 to 3 worse. One recently developed method to close this gap is metal-induced energy transfer (MIET) imaging, which achieves an axial resolution down to nanometers. It exploits the distance-dependent quenching of fluorescence when a fluorescent molecule is brought close to a metal surface. In the present manuscript, we combine the extreme axial resolution of MIET imaging with the extraordinary lateral resolution of single-molecule localization microscopy, in particular with direct stochastic optical reconstruction microscopy (STORM). This combination allows us to achieve isotropic three-dimensional super-resolution imaging of subcellular structures. Moreover, we used spectral demixing for implementing dual-color MIET-STORM that allows us to image and colocalize, in three dimensions, two different cellular structures simultaneously.
在过去二十年中，超分辨率显微镜在速度和分辨率方面取得了巨大发展，但对于其大多数方法而言，横向分辨率和轴向分辨率之间存在显著差距，轴向分辨率要差2至3倍。最近开发的一种缩小这一差距的方法是金属诱导能量转移（MIET）成像，它可实现低至纳米级的轴向分辨率。当荧光分子靠近金属表面时，它利用了与距离相关的荧光猝灭。在本论文中，我们将MIET成像的极高轴向分辨率与单分子定位显微镜（尤其是直接随机光学重建显微镜（STORM））的非凡横向分辨率相结合。这种结合使我们能够实现亚细胞结构的各向同性三维超分辨率成像。此外，我们使用光谱解混来实现双色MIET-STORM，这使我们能够在三维空间中同时对两种不同的细胞结构进行成像和共定位。
