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Nat Methods
2014 Feb 01;112:156-62. doi: 10.1038/nmeth.2784.
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Single-molecule evaluation of fluorescent protein photoactivation efficiency using an in vivo nanotemplate.
Durisic N, Laparra-Cuervo L, Sandoval-Álvarez A, Borbely JS, Lakadamyali M.
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Photoswitchable fluorescent probes are central to localization-based super-resolution microscopy. Among these probes, fluorescent proteins are appealing because they are genetically encoded. Moreover, the ability to achieve a 1:1 labeling ratio between the fluorescent protein and the protein of interest makes these probes attractive for quantitative single-molecule counting. The percentage of fluorescent protein that is photoactivated into a fluorescently detectable form (i.e., the photoactivation efficiency) plays a crucial part in properly interpreting the quantitative information. It is important to characterize the photoactivation efficiency at the single-molecule level under the conditions used in super-resolution imaging. Here, we used the human glycine receptor expressed in Xenopus oocytes and stepwise photobleaching or single-molecule counting photoactivated localization microcopy (PALM) to determine the photoactivation efficiency of fluorescent proteins mEos2, mEos3.1, mEos3.2, Dendra2, mClavGR2, mMaple, PA-GFP and PA-mCherry. This analysis provides important information that must be considered when using these fluorescent proteins in quantitative super-resolution microscopy.
Adam,
Structural basis of enhanced photoconversion yield in green fluorescent protein-like protein Dendra2.
2009, Pubmed
Adam,
Structural basis of enhanced photoconversion yield in green fluorescent protein-like protein Dendra2.
2009,
Pubmed Annibale,
Quantitative photo activated localization microscopy: unraveling the effects of photoblinking.
2011,
Pubmed Bálint,
Correlative live-cell and superresolution microscopy reveals cargo transport dynamics at microtubule intersections.
2013,
Pubmed Betzig,
Imaging intracellular fluorescent proteins at nanometer resolution.
2006,
Pubmed Brakemann,
A reversibly photoswitchable GFP-like protein with fluorescence excitation decoupled from switching.
2011,
Pubmed Dempsey,
Evaluation of fluorophores for optimal performance in localization-based super-resolution imaging.
2011,
Pubmed Durisic,
Stoichiometry of the human glycine receptor revealed by direct subunit counting.
2012,
Pubmed
,
Xenbase Fernández-Suárez,
Fluorescent probes for super-resolution imaging in living cells.
2008,
Pubmed Goldin,
Maintenance of Xenopus laevis and oocyte injection.
1992,
Pubmed
,
Xenbase Grotjohann,
Diffraction-unlimited all-optical imaging and writing with a photochromic GFP.
2011,
Pubmed Gunzenhäuser,
Quantitative super-resolution imaging reveals protein stoichiometry and nanoscale morphology of assembling HIV-Gag virions.
2012,
Pubmed Gurskaya,
Engineering of a monomeric green-to-red photoactivatable fluorescent protein induced by blue light.
2006,
Pubmed Habuchi,
Reversible single-molecule photoswitching in the GFP-like fluorescent protein Dronpa.
2005,
Pubmed Habuchi,
mKikGR, a monomeric photoswitchable fluorescent protein.
2008,
Pubmed Hess,
Ultra-high resolution imaging by fluorescence photoactivation localization microscopy.
2006,
Pubmed Hoi,
A monomeric photoconvertible fluorescent protein for imaging of dynamic protein localization.
2010,
Pubmed Huang,
Whole-cell 3D STORM reveals interactions between cellular structures with nanometer-scale resolution.
2008,
Pubmed Huang,
Breaking the diffraction barrier: super-resolution imaging of cells.
2010,
Pubmed Kanchanawong,
Nanoscale architecture of integrin-based cell adhesions.
2010,
Pubmed Lakadamyali,
3D multicolor super-resolution imaging offers improved accuracy in neuron tracing.
2012,
Pubmed Lando,
Quantitative single-molecule microscopy reveals that CENP-A(Cnp1) deposition occurs during G2 in fission yeast.
2012,
Pubmed Lee,
Counting single photoactivatable fluorescent molecules by photoactivated localization microscopy (PALM).
2012,
Pubmed Lippincott-Schwartz,
Photoactivatable fluorescent proteins for diffraction-limited and super-resolution imaging.
2009,
Pubmed Lynch,
Native glycine receptor subtypes and their physiological roles.
2009,
Pubmed McEvoy,
mMaple: a photoconvertible fluorescent protein for use in multiple imaging modalities.
2012,
Pubmed McKinney,
A bright and photostable photoconvertible fluorescent protein.
2009,
Pubmed Patterson,
A photoactivatable GFP for selective photolabeling of proteins and cells.
2002,
Pubmed Puchner,
Counting molecules in single organelles with superresolution microscopy allows tracking of the endosome maturation trajectory.
2013,
Pubmed Renz,
Plasticity of the asialoglycoprotein receptor deciphered by ensemble FRET imaging and single-molecule counting PALM imaging.
2012,
Pubmed Rust,
Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM).
2006,
Pubmed Sengupta,
Probing protein heterogeneity in the plasma membrane using PALM and pair correlation analysis.
2011,
Pubmed Sengupta,
Quantifying spatial organization in point-localization superresolution images using pair correlation analysis.
2013,
Pubmed Shroff,
Dual-color superresolution imaging of genetically expressed probes within individual adhesion complexes.
2007,
Pubmed Simonson,
Counting bungarotoxin binding sites of nicotinic acetylcholine receptors in mammalian cells with high signal/noise ratios.
2010,
Pubmed Subach,
Photoactivatable mCherry for high-resolution two-color fluorescence microscopy.
2009,
Pubmed Ulbrich,
Subunit counting in membrane-bound proteins.
2007,
Pubmed
,
Xenbase Wang,
A technical consideration concerning the removal of oocyte vitelline membranes for patch clamp recording.
2004,
Pubmed
,
Xenbase Wiedenmann,
EosFP, a fluorescent marker protein with UV-inducible green-to-red fluorescence conversion.
2004,
Pubmed Xu,
Actin, spectrin, and associated proteins form a periodic cytoskeletal structure in axons.
2013,
Pubmed Zhang,
Rational design of true monomeric and bright photoactivatable fluorescent proteins.
2012,
Pubmed
