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Sub-Diffraction-Limit Imaging with Stochastic Optical Reconstruction Microscopy

  • Mark Bates
  • Bo Huang
  • Michael J. Rust
  • Graham T. Dempsey
  • Wenqin Wang
  • Xiaowei Zhuang
Chapter
Part of the Springer Series in Chemical Physics book series (CHEMICAL, volume 96)

Summary

Light microscopy is a widely used imaging method in biomedical research. However, the resolution of conventional optical microcopy is limited by the diffraction of light, making structures smaller than 200 nm difficult to resolve. To overcome this limit, we have developed a new form of fluorescence microscopy - Stochastic Optical Reconstruction Microscopy (STORM). STORM makes use of single-molecule imaging methods and photo-switchable fluorescent probes to temporally separate the otherwise spatially overlapping images of individual molecules. An STORM image is acquired over a number of imaging cycles, and in each cycle only a subset of the fluorescent labels is switched on such that each of the active fluorophores is optically resolvable from the rest. This allows the position of these fluorophores to be determined with nanometer accuracy. Over the course of many such cycles, the positions of numerous fluorophores are determined and used to construct a super-resolution image. Using this method, we have demonstrated multi-color, three-dimensional (3D) imaging of biomolecules and cells with ∼ 20 nm lateral and ∼ 50 nm axial resolutions. In principle, the resolution of this technique can reach the molecular scale.

Keywords

Localization Accuracy Centroid Position Superresolution Imaging Imaging Cycle Stochastic Optical Reconstruction Microscopy 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Notes

Acknowledgements

This work is supported by in part by the NIH (to X.Z.). X.Z. is a Howard Hughes Medical Institute Investigator.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2010

Authors and Affiliations

  • Mark Bates
    • 1
  • Bo Huang
    • 2
    • 3
  • Michael J. Rust
    • 4
  • Graham T. Dempsey
    • 5
  • Wenqin Wang
    • 4
  • Xiaowei Zhuang
    • 6
  1. 1.School of Engineering and Applied SciencesCambridgeUSA
  2. 2.Department of Chemistry and Chemical BiologyCornell UniversityIthacaUSA
  3. 3.Howard Hughes Medical Institute Harvard UniversityCambridgeUSA
  4. 4.Department of PhysicsHarvard UniversityCambridgeUSA
  5. 5.Program in BiophysicsHarvard UniversityCambridgeUSA
  6. 6.Department of Chemistry and Chemical BiologyHoward Hughes Medical Institute Harvard UniversityCambridgeUSA

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