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      Fluorescence nanoscopy in cell biology.

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          Abstract

          Fluorescence nanoscopy uniquely combines minimally invasive optical access to the internal nanoscale structure and dynamics of cells and tissues with molecular detection specificity. While the basic physical principles of 'super-resolution' imaging were discovered in the 1990s, with initial experimental demonstrations following in 2000, the broad application of super-resolution imaging to address cell-biological questions has only more recently emerged. Nanoscopy approaches have begun to facilitate discoveries in cell biology and to add new knowledge. One current direction for method improvement is the ambition to quantitatively account for each molecule under investigation and assess true molecular colocalization patterns via multi-colour analyses. In pursuing this goal, the labelling of individual molecules to enable their visualization has emerged as a central challenge. Extending nanoscale imaging into (sliced) tissue and whole-animal contexts is a further goal. In this Review we describe the successes to date and discuss current obstacles and possibilities for further development.

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          Most cited references167

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          Three-dimensional super-resolution imaging by stochastic optical reconstruction microscopy.

          Recent advances in far-field fluorescence microscopy have led to substantial improvements in image resolution, achieving a near-molecular resolution of 20 to 30 nanometers in the two lateral dimensions. Three-dimensional (3D) nanoscale-resolution imaging, however, remains a challenge. We demonstrated 3D stochastic optical reconstruction microscopy (STORM) by using optical astigmatism to determine both axial and lateral positions of individual fluorophores with nanometer accuracy. Iterative, stochastic activation of photoswitchable probes enables high-precision 3D localization of each probe, and thus the construction of a 3D image, without scanning the sample. Using this approach, we achieved an image resolution of 20 to 30 nanometers in the lateral dimensions and 50 to 60 nanometers in the axial dimension. This development allowed us to resolve the 3D morphology of nanoscopic cellular structures.
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            Far-field optical nanoscopy.

            In 1873, Ernst Abbe discovered what was to become a well-known paradigm: the inability of a lens-based optical microscope to discern details that are closer together than half of the wavelength of light. However, for its most popular imaging mode, fluorescence microscopy, the diffraction barrier is crumbling. Here, I discuss the physical concepts that have pushed fluorescence microscopy to the nanoscale, once the prerogative of electron and scanning probe microscopes. Initial applications indicate that emergent far-field optical nanoscopy will have a strong impact in the life sciences and in other areas benefiting from nanoscale visualization.
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              Beiträge zur Theorie des Mikroskops und der mikroskopischen Wahrnehmung

              E. Abbe (1873)
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                Author and article information

                Journal
                Nat. Rev. Mol. Cell Biol.
                Nature reviews. Molecular cell biology
                Springer Science and Business Media LLC
                1471-0080
                1471-0072
                Nov 2017
                : 18
                : 11
                Affiliations
                [1 ] Max Planck Institute for Biophysical Chemistry, Department of NanoBiophotonics, Am Fassberg 11, 37077 Göttingen, Germany.
                [2 ] Max Planck Institute for Medical Research, Department of Optical Nanoscopy, Jahnstrasse 29, 69120 Heidelberg, Germany.
                [3 ] German Cancer Research Center (DKFZ), BioQuant, Im Neuenheimer Feld 267, 69120 Heidelberg, Germany.
                [4 ] University of Göttingen Medical Faculty, Department of Neurology, Robert-Koch-Strasse 40, 37075 Göttingen, Germany.
                Article
                nrm.2017.71
                10.1038/nrm.2017.71
                28875992
                0a19fd4e-eb25-4545-b511-bc9ea8328d9c
                History

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