Extending Microscopic Resolution with Single-Molecule Imaging and Active Control
- 9 June 2012
- journal article
- review article
- Published by Annual Reviews in Annual Review of Biophysics
- Vol. 41 (1), 321-342
- https://doi.org/10.1146/annurev-biophys-050511-102250
Abstract
Superresolution imaging of biological structures provides information beyond the optical diffraction limit. One class of superresolution techniques uses the power of single fluorescent molecules as nanoscale emitters of light combined with emission control, variously described by the acronyms PALM/FPALM/STORM and many others. Even though the acronyms differ and refer mainly to different active-control mechanisms, the underlying fundamental principles behind these “pointillist” superresolution imaging techniques are the same. Circumventing the diffraction limit requires two key steps. The first step (superlocalization) is the detection and localization of spatially separated single molecules. The second step actively controls the emitting molecules to ensure a very low concentration of single emitters such that they do not overlap in any one imaging frame. The final image is reconstructed from time-sequential imaging and superlocalization of the single emitting labels decorating the structure of interest. The statistical, imaging, and active-control strategies for achieving superresolution imaging with single molecules are reviewed.Keywords
This publication has 82 references indexed in Scilit:
- Super-Resolution Imaging of the Nucleoid-Associated Protein HU in Caulobacter crescentusBiophysical Journal, 2011
- Super-resolution Microscopy of Lipid Bilayer PhasesJournal of the American Chemical Society, 2011
- Molecular Orientation Affects Localization Accuracy in Superresolution Far-Field Fluorescence MicroscopyNano Letters, 2010
- Superresolution Imaging of Chemical Synapses in the BrainNeuron, 2010
- Superresolution Imaging of Targeted Proteins in Fixed and Living Cells Using Photoactivatable Organic FluorophoresJournal of the American Chemical Society, 2010
- Molecules and Methods for Super-Resolution ImagingMethods in Enzymology, 2010
- Photoactivatable fluorescent proteins for diffraction-limited and super-resolution imagingTrends in Cell Biology, 2009
- DCDHF Fluorophores for Single‐Molecule Imaging in CellsChemphyschem, 2009
- A Photoactivatable Push−Pull Fluorophore for Single-Molecule Imaging in Live CellsJournal of the American Chemical Society, 2008
- Precise Nanometer Localization Analysis for Individual Fluorescent ProbesBiophysical Journal, 2002