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bioRxiv · 10.1101/2021.01.31.429063

Consequences of the Nyquist-Shannon sampling criterion in Mesoscopic Multiphoton Microscopy to avail full-field sub-micron resolution resolvability

Borah, B. J.·Lee, J.-C.·Chi, H.-H.·Hsiao, Y.-T.·Yen, C.-T.·Sun, C.-K.
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Abstract

With a limited effective voxel rate, to date, each laser-scanning mesoscopic multiphoton microscope (MPM), despite securing an ultra-large field of view (FOV) and an ultra-high optical resolution simultaneously, experiences a fundamental issue with digitization; i.e., inability to satisfy the Nyquist-Shannon sampling criterion to resolve the optics-limited sub-micron resolution over the whole FOV. Such a system either neglects the criterion degrading the digital resolution to twice the pixel size, or significantly reduces the imaging area and/or the imaging speed to respect the digitization. Here we introduce a Nyquist figure of merit parameter to assess this issue, further to comprehend a maximum aliasing-free FOV and a cross-over excitation wavelength for a laser scanning MPM system. Based on our findings we demonstrate an ultra-high voxel rate acquisition in a custom-built mesoscopic MPM system to exceed the Nyquist-rate for a >3800 FOV-resolution ratio while not compromising the imaging speed as well as the photon-budget.

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Borah, B. J., Lee, J.-C., Chi, H.-H., Hsiao, Y.-T., Yen, C.-T., Sun, C.-K.. 2021-02-02. Consequences of the Nyquist-Shannon sampling criterion in Mesoscopic Multiphoton Microscopy to avail full-field sub-micron resolution resolvability. https://doi.org/10.1101/2021.01.31.429063

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