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Journal Article

STED nanoscopy with time-gated detection: Theoretical and experimental aspects.

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

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Schönle,  A.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

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Ta,  H.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

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Han,  K. Y.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

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Moneron,  G.
Department of NanoBiophotonics, MPI for biophysical chemistry, Max Planck Society;

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

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

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1602392.pdf
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Citation

Vicidomini, G., Schönle, A., Ta, H., Han, K. Y., Moneron, G., Eggeling, C., et al. (2013). STED nanoscopy with time-gated detection: Theoretical and experimental aspects. PLoS One, 8(1): e54421. doi:10.1371/journal.pone.0054421.


Cite as: https://hdl.handle.net/11858/00-001M-0000-000E-7743-A
Abstract
In a stimulated emission depletion (STED) microscope the region in which fluorescence markers can emit spontaneously shrinks with continued STED beam action after a singular excitation event. This fact has been recently used to substantially improve the effective spatial resolution in STED nanoscopy using time-gated detection, pulsed excitation and continuous wave (CW) STED beams. We present a theoretical framework and experimental data that characterize the time evolution of the effective point-spread-function of a STED microscope and illustrate the physical basis, the benefits, and the limitations of time-gated detection both for CW and pulsed STED lasers. While gating hardly improves the effective resolution in the all-pulsed modality, in the CW-STED modality gating strongly suppresses low spatial frequencies in the image. Gated CW-STED nanoscopy is in essence limited (only) by the reduction of the signal that is associated with gating. Time-gated detection also reduces/suppresses the influence of local variations of the fluorescence lifetime on STED microscopy resolution.