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bioRxiv · 10.1101/2020.04.06.027037

Fast multi-directional DSLM for confocaldetection without striping artifacts

Ricci, P.·Sancataldo, G.·Gavryusev, V.·Franceschini, A.·Muellenbroich, C.·Silvestri, L.·Pavone, F. S.
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Abstract

In recent years light-sheet fluorescence microscopy (LSFM) has become a cornerstone technology for neuroscience, improving quality and capabilities of 3D imaging. By selectively illuminating a single plane, it provides intrinsic optical sectioning and fast image recording, while minimizing out of focus fluorescence background, sample photo-damage and photo-bleaching. However, images acquired with LSFM are often affected by light absorption or scattering effects, leading to un-even illumination and striping artifacts. In this work we present an optical solution to this problem, via fast multi-directional illumination of the sample, based on an acousto-optical deflector (AOD). We demonstrate that this pivoting system is compatible with confocal detection in digital scanned laser light-sheet fluorescence microscopy (DSLM) by using a pivoted elliptical-Gaussian beam. We tested its performance by acquiring signals emitted by specific fluorophores in several mouse brain areas, comparing the pivoting beam illumination and a traditional static one, measuring the point spread function response and quantifying the striping reduction. We observed real-time shadow suppression, while preserving the advantages of confocal detection for image contrast.

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Ricci, P., Sancataldo, G., Gavryusev, V., Franceschini, A., Muellenbroich, C., Silvestri, L., Pavone, F. S.. 2020-04-06. Fast multi-directional DSLM for confocaldetection without striping artifacts. https://doi.org/10.1101/2020.04.06.027037

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