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bioRxiv · 10.1101/2020.08.27.266999

3D Interferometric Lattice Light-Sheet Imaging

Cao, B.·Wang, G.·Li, J.·Pertsinidis, A.
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Abstract

Understanding cellular structure and function requires live-cell imaging with high spatio-temporal resolution and high detection sensitivity. Direct visualization of molecular processes using single-molecule/super-resolution techniques has thus been transformative. However, extracting the highest-resolution 4D information possible from weak and dynamic fluorescence signals in live cells remains challenging. For example, some of the highest spatial resolution methods, e.g. interferometric (4Pi) approaches1-6 can be slow, require high peak excitation intensities that accelerate photobleaching or suffer from increased out-of-focus background. Selective-plane illumination (SPIM)7-12 reduces background, but most implementations typically feature modest spatial, especially axial, resolution. Here we develop 3D interferometric lattice light-sheet (3D-iLLS) imaging, a technique that overcomes many of these limitations. 3D-iLLS provides, by virtue of SPIM, low light levels and photobleaching, while providing increased background suppression and significantly improved volumetric imaging/sectioning capabilities through 4Pi interferometry. We demonstrate 3D-iLLS with axial resolution and single-particle localization precision down to <100nm (FWHM) and <10nm (1{sigma}) respectively. 3D-iLLS paves the way for a fuller elucidation of sub-cellular phenomena by enhanced 4D resolution and SNR live imaging.

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Cao, B., Wang, G., Li, J., Pertsinidis, A.. 2020-08-28. 3D Interferometric Lattice Light-Sheet Imaging. https://doi.org/10.1101/2020.08.27.266999

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