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bioRxiv · 10.1101/170555

RAPID: Real-time image-based autofocus for all wide-field optical microscopy systems

Silvestri, L.·Muellenbroich, M. C.·Costantini, I.·Di Giovanna, A. P.·Sacconi, L.·Pavone, F. S.
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Abstract

Autofocus methods used in biomicroscopy are either based on the search of an optimal focus position - which requires suspending data collection during the optimization process - or on the continuous monitoring of the position of a fiducial plane - which may not coincide with the sample itself. Here, we introduce RAPID (Rapid Autofocus via Pupil-split Image phase Detection), a method for real-time image-based focus stabilization, applicable in all wide-field microscopy systems. We demonstrate that RAPID maintains high image quality in various settings, from in vivo fluorescence imaging to light-sheet microscopy. RAPID provides a universal autofocus solution for automated microscopy, and enables quantitative assays otherwise impossible in a standard microscope, such as 3D tracking of fast-moving organisms.

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Silvestri, L., Muellenbroich, M. C., Costantini, I., Di Giovanna, A. P., Sacconi, L., Pavone, F. S.. 2017-07-31. RAPID: Real-time image-based autofocus for all wide-field optical microscopy systems. https://doi.org/10.1101/170555

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