Exploiting the tunability of stimulated emission depletion microscopy for super-resolution imaging of nuclear structures

Nature Communications
Maria J SarmentoLuca Lanzanò

Abstract

Imaging of nuclear structures within intact eukaryotic nuclei is imperative to understand the effect of chromatin folding on genome function. Recent developments of super-resolution fluorescence microscopy techniques combine high specificity, sensitivity, and less-invasive sample preparation procedures with the sub-diffraction spatial resolution required to image chromatin at the nanoscale. Here, we present a method to enhance the spatial resolution of a stimulated-emission depletion (STED) microscope based only on the modulation of the STED intensity during the acquisition of a STED image. This modulation induces spatially encoded variations of the fluorescence emission that can be visualized in the phasor plot and used to improve and quantify the effective spatial resolution of the STED image. We show that the method can be used to remove direct excitation by the STED beam and perform dual color imaging. We apply this method to the visualization of transcription and replication foci within intact nuclei of eukaryotic cells.

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Citations

Apr 14, 2019·Journal of Biophotonics·Luca PescePaolo Bianchini
Apr 17, 2021·Nature Methods·Lorenzo ScipioniEnrico Gratton
May 7, 2021·Annual Review of Biophysics·Leonel MalacridaEnrico Gratton
Jul 8, 2021·Molecular Systems Biology·Liangqi Xie, Zhe Liu
Jul 6, 2021·Biophysical Journal·Aymeric Le GratietAlberto Diaspro
Mar 14, 2021·Microscopy Research and Technique·Zhaoyang WuPeng Xi

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Methods Mentioned

BETA
electron microscopy
fluorescence microscopy
light diffraction
fluorescence correlation spectroscopy
Fluorescence
super-resolution microscopy

Software Mentioned

STED
ImageJ
STORM
SPLIT
MATLAB

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