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Journal Article

Coordinate-targeted fluorescence nanoscopy with multiple off states.

MPS-Authors
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Danzl,  J. G.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons189436

Sidenstein,  S.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons185783

Gregor,  C.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons39413

Urban,  N. T.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons189438

Ilgen,  P.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons15269

Jakobs,  S.
Research Group of Mitochondrial Structure and Dynamics, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons15210

Hell,  S. W.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

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Supplementary Material (public)

2253029_Suppl_1.pdf
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(Supplementary material), 18MB

2253029_Suppl_2.avi
(Supplementary material), 18MB

2253029_Suppl_3.avi
(Supplementary material), 5MB

2253029_Suppl_4.avi
(Supplementary material), 4MB

2253029_Suppl_5.avi
(Supplementary material), 11MB

2253029_Suppl_6.avi
(Supplementary material), 7MB

Citation

Danzl, J. G., Sidenstein, S., Gregor, C., Urban, N. T., Ilgen, P., Jakobs, S., et al. (2016). Coordinate-targeted fluorescence nanoscopy with multiple off states. Nature Photonics, 10(2), 122-128. doi:10.1038/nphoton.2015.266.


Cite as: http://hdl.handle.net/11858/00-001M-0000-0029-C711-4
Abstract
Far-field super-resolution fluorescence microscopy discerns fluorophores residing closer than the diffraction barrier by briefly transferring them in different (typically ON and OFF) states before detection. In coordinate-targeted super-resolution variants, such as stimulated emission depletion (STED) microscopy, this state difference is created by the intensity minima and maxima of an optical pattern, causing all fluorophores to assume the off state, for instance, except at the minima. Although strong spatial confinement of the on state enables high resolution, it also subjects the fluorophores to excess intensities and state cycles at the maxima. Here, we address these issues by driving the fluorophores into a second off state that is inert to the excess light. By using reversibly switchable fluorescent proteins as labels, our approach reduces bleaching and enhances resolution and contrast in live-cell STED microscopy. Using two or more transitions to off states is a useful strategy for augmenting the power of coordinate-targeted super-resolution microscopy.