Combining 3D single molecule localization strategies for reproducible bioimaging

Clément Cabriel, Nicolas Bourg, Pierre Jouchet, Guillaume Dupuis, Christophe Leterrier, Aurélie Baron, Marie-Ange Badet-Denisot, Boris Vauzeilles, Emmanuel Fort, and Sandrine Lévêque-Fort

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Nov 27, 2018
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Received Date: 16th November 18

We developed a 3D localization-based super-resolution technique providing a slowly varying localization precision over a 1 μm range with precisions down to 15 nm. The axial localization is performed through a combination of point spread function (PSF) shaping and supercritical angle fluorescence (SAF), which yields absolute axial information. Using a dual-view scheme, the axial detection is decoupled from the lateral detection and optimized independently to provide a weakly anisotropic 3D resolution over the imaging range. This method can be readily implemented on most homemade PSF shaping setups and provides drift-free, tilt-insensitive and achromatic results. Its insensitivity to these unavoidable experimental biases is especially adapted for multicolor 3D super-resolution microscopy, as we demonstrate by imaging cell cytoskeleton, living bacteria membranes and axon periodic submembrane scaffolds. We further illustrate the interest of the technique for biological multicolor imaging over a several-μm range by direct merging of multiple acquisitions at different depths.

Read in full at bioRxiv.

This is an abstract of a preprint hosted on an independent third party site. It has not been peer reviewed but is currently under consideration at Nature Communications.

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