Breaking the Diffraction Barrier: Super-Resolution Imaging of Cells

Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158, USA.
Cell (Impact Factor: 32.24). 12/2010; 143(7):1047-58. DOI: 10.1016/j.cell.2010.12.002
Source: PubMed


Anyone who has used a light microscope has wished that its resolution could be a little better. Now, after centuries of gradual improvements, fluorescence microscopy has made a quantum leap in its resolving power due, in large part, to advancements over the past several years in a new area of research called super-resolution fluorescence microscopy. In this Primer, we explain the principles of various super-resolution approaches, such as STED, (S)SIM, and STORM/(F)PALM. Then, we describe recent applications of super-resolution microscopy in cells, which demonstrate how these approaches are beginning to provide new insights into cell biology, microbiology, and neurobiology.

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    • "However, the requirement of sample embedding for high-quality serial sectioning poses extra challenges for super-resolution imaging. Since STORM imaging relies on switching and localization of individual fluorophores to reconstruct super-resolution images (Huang et al., 2010; Rust et al., 2006), the resolution of a STORM image depends not only on the localization precision of individual fluorophores determined by their photon output but also on the localization density determined by the labeling density. Achieving optimal STORM resolution thus requires the labeling and embedding conditions to simultaneously retain optimal fluorophore properties and high-density labeling in resin-embedded samples. "
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    • "Among the methods of acquiring super-resolution fluorescence images [1] [2], structured illumination microscopy (SIM) offers a relatively modest, twofold resolution improvement over widefield microscopy [3]. However, as SIM uses only a relatively small number of widefield images to capture the information required to improve resolution, it is in principle more suitable for live sample imaging; SIM offers the advantages of fast acquisition over a large area and weaker irradiation of the sample compared to alternative techniques such as stimulated emission depletion [4] and single-molecule localisation [5] [6] [7], and it is compatible with all fluorophores used in widefield and confocal imaging. "
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