Localization-Based Super-Resolution Imaging of Cellular Structures

  • Pakorn Kanchanawong
  • Clare M. Waterman
Part of the Methods in Molecular Biology book series (MIMB, volume 1046)


Fluorescence microscopy allows direct visualization of fluorescently tagged proteins within cells. However, the spatial resolution of conventional fluorescence microscopes is limited by diffraction to ~250 nm, prompting the development of super-resolution microscopy which offers resolution approaching the scale of single proteins, i.e., ~20 nm. Here, we describe protocols for single molecule localization-based super-resolution imaging, using focal adhesion proteins as an example and employing either photoswitchable fluorophores or photoactivatable fluorescent proteins. These protocols should also be easily adaptable to imaging a broad array of macromolecular assemblies in cells whose components can be fluorescently tagged and assemble into high density structures.

Key words

Super-resolution microscopy Focal adhesions Localization microscopy TIRF PALM Single molecules Photoswitchable fluorophores Photoactivatable fluorescent proteins 



PK is supported by the Singapore National Research Foundation under the NRF Fellowship (NRFF-2011-04). CMW is supported by the Division of Intramural Research, National Heart, Lung, and Blood Institute, National Institutes of Health. We thank Harald Hess and Gleb Shtengel (Howard Hughes Medical Institute, Janelia Farm Research Campus), and Michael Davidson (The Florida State University) for advice, equipment, reagents, and collaboration related to this work.


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Copyright information

© Springer Science+Business Media, LLC 2013

Authors and Affiliations

  • Pakorn Kanchanawong
    • 1
  • Clare M. Waterman
    • 2
  1. 1.Department of Bioengineering, Mechanobiology InstituteNational University of SingaporeSingaporeSingapore
  2. 2.Cell Biology and Physiology CenterNational Heart Lung and Blood Institute, National Institutes of HealthBethesdaUSA

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