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Single‐Molecule Localization Microscopy using mCherry
Author(s) -
Winterflood Christian M.,
Ewers Helge
Publication year - 2014
Publication title -
chemphyschem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.016
H-Index - 140
eISSN - 1439-7641
pISSN - 1439-4235
DOI - 10.1002/cphc.201402423
Subject(s) - mcherry , green fluorescent protein , biophysics , fluorescence , fusion protein , super resolution microscopy , chemistry , resolution (logic) , protein subcellular localization prediction , microscopy , biomolecule , fluorescence microscope , nanotechnology , biochemistry , biology , materials science , physics , recombinant dna , computer science , optics , artificial intelligence , gene
We demonstrate the potential of the commonly used red fluorescent protein mCherry for single‐molecule super‐resolution imaging. mCherry can be driven into a light‐induced dark state in the presence of a thiol from which it can recover spontaneously or by irradiation with near UV light. We show imaging of subcellular protein structures such as microtubules and the nuclear pore complex with a resolution below 40 nm. We were able to image the C‐terminus of the nuclear pore protein POM121, which is on the inside of the pore and not readily accessible for external labeling. The photon yield for mCherry is comparable to that of the latest optical highlighter fluorescent proteins. Our findings show that the widely used mCherry red fluorescent protein and the vast number of existing mCherry fusion proteins are readily amenable to super‐resolution imaging. This obviates the need for generating novel protein fusions that may compromise function or the need for external fluorescent labeling.