Dynamic superresolution imaging of endogenous proteins on living cells at ultra-high density.

@article{Giannone2010DynamicSI,
  title={Dynamic superresolution imaging of endogenous proteins on living cells at ultra-high density.},
  author={Gr{\'e}gory Giannone and Eric Hosy and Florian Levet and Audrey Constals and Katrin Schulze and Alexander I. Sobolevsky and Michael P. Rosconi and Eric Gouaux and Robert Tamp{\'e} and Daniel Choquet and Laurent Cognet},
  journal={Biophysical journal},
  year={2010},
  volume={99 4},
  pages={
          1303-10
        }
}
Versatile superresolution imaging methods, able to give dynamic information of endogenous molecules at high density, are still lacking in biological science. Here, superresolved images and diffusion maps of membrane proteins are obtained on living cells. The method consists of recording thousands of single-molecule trajectories that appear sequentially on a cell surface upon continuously labeling molecules of interest. It allows studying any molecules that can be labeled with fluorescent… Expand
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References

SHOWING 1-10 OF 47 REFERENCES
Repetitive reversible labeling of proteins at polyhistidine sequences for single-molecule imaging in live cells.
TLDR
A novel, generic labeling strategy that offers free choice of fluorophores while being minimally perturbing is demonstrated by labeling engineered polyhistidine sequences of the serotonin-gated 5-HT(3) receptor on the surface of live mammalian cells. Expand
Imaging Intracellular Fluorescent Proteins at Nanometer Resolution
TLDR
This work introduced a method for optically imaging intracellular proteins at nanometer spatial resolution and used this method to image specific target proteins in thin sections of lysosomes and mitochondria and in fixed whole cells to image retroviral protein Gag at the plasma membrane. Expand
Imaging of single molecule diffusion.
TLDR
Methodology for visualization of the motion of individual fluorescent molecules is applied to imaging of the diffusional path of single molecules in a phospholipid membrane by usingospholipids carrying one rhodamine dye molecule. Expand
Live-cell photoactivated localization microscopy of nanoscale adhesion dynamics
TLDR
By allowing observation of a wide variety of nanoscale dynamics, live-cell PALM provides insights into molecular assembly during the initiation, maturation and dissolution of cellular processes. Expand
Dynamic multiple-target tracing to probe spatiotemporal cartography of cell membranes
TLDR
An analytical single-particle tracking method and tool, multiple-target tracing (MTT), that takes advantage of the high spatial resolution provided by single-fluorophore sensitivity to generate dynamic maps at high densities of tracked particles, thereby providing global representation of molecular dynamics in cell membranes. Expand
Autofluorescent proteins in single-molecule research: applications to live cell imaging microscopy.
TLDR
The yellow-fluorescent protein mutant eYFP is superior compared to all the fluorescent proteins for single-molecule studies in vivo and is found to dynamically track the entity to which it is anchored when bound to membranes of live cells. Expand
Subdiffraction-resolution fluorescence imaging of proteins in the mitochondrial inner membrane with photoswitchable fluorophores.
TLDR
Subdiffraction-resolution fluorescence imaging of intracellular F(0)F(1)-ATP synthase and cytochrome c oxidase in the inner membrane of mitochondria is demonstrated and it is demonstrated how quantitative data, i.e. the protein distribution in the membrane, can be derived and compared. Expand
Superresolution microscopy on the basis of engineered dark states.
TLDR
It is demonstrated that superresolution imaging can be carried out with practically every single-molecule compatible, synthetic fluorophore by controlling their emission properties by increasing the duration of the off-states and creating radical ion states of severalfold longer lifetimes. Expand
Subdiffraction-resolution fluorescence microscopy of myosin-actin motility.
TLDR
A novel approach for video-like subdiffraction microscopy based on rapid and reversible photoswitching of commercially available organic carbocyanine fluorophores is introduced and in vitro studies on the motility of fluorophore-labeled actin filaments along myosin II are demonstrated. Expand
High-density mapping of single-molecule trajectories with photoactivated localization microscopy
TLDR
This work created spatially resolved maps of single-molecule motions by imaging the membrane proteins Gag and VSVG, and obtained several orders of magnitude more trajectories per cell than traditional single-particle tracking enables. Expand
...
1
2
3
4
5
...