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bioRxiv · 10.1101/2024.07.08.602588

Array Detection Enables Large Localization Range for Simple and Robust MINFLUX

Slenders, E.·Patil, S.·Held, M. O.·Zunino, A.·Vicidomini, G.
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Abstract

The MINFLUX concept significantly enhances the spatial resolution of single-molecule localization microscopy (SMLM) by overcoming the limit imposed by the fluorophores photon counts. Typical MINFLUX microscopes localize the target molecule by scanning a zero-intensity focus around the molecule in a circular trajectory, with smaller trajectory diameters yielding lower localization uncertainties for a given number of photons. Since this approach requires the molecule to be within the scanned trajectory, MINFLUX typically relies on a photon-demanding iterative scheme with decreasing trajectory diameters. Although the iterative procedure does not substantially reduce the photon efficiency of MINFLUX, this approach is prone to misplacements of the trajectory and increases the systems complexity. In this work, we introduce ISM-FLUX, a novel implementation of MINFLUX using image-scanning microscopy (ISM) with a single-photon avalanche diode (SPAD) array detector. ISM-FLUX provides precise MINFLUX localization within the trajectory while maintaining conventional photon-limited uncertainty outside it. The robustness of ISM-FLUX localization results in a larger localization range and greatly simplifies the architecture, which may facilitate broader adoption of MIN-FLUX.

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Slenders, E., Patil, S., Held, M. O., Zunino, A., Vicidomini, G.. 2024-07-11. Array Detection Enables Large Localization Range for Simple and Robust MINFLUX. https://doi.org/10.1101/2024.07.08.602588

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