Versatile high-speed volumetric imaging from microscopic to macroscopic scale by self-adaptive oblique plane microscopy

Abstract There is an increasing need for large-scale high-speed volumetric recording in complex multi-cellular model systems to define dynamic processes. Oblique plane microscopy (OPM) provides a solution that features oblique illumination, rapid optical scanning, and remote focusing to achieve real-time 4D microscopy. OPM implements light sheet imaging via a single primary objective lens, making the entire space below the objective accessible for large specimens, such as living mouse brain. Yet it is challenging to adopt OPM beyond a microscopic scale (i.e. size < 1mm), limiting its broad applications. Here we present a self-adaptive OPM that leverages Abbe’s sine condition to unlock its flexibility across a range of field-of-views (FOVs) (up to 8 mm2) and resolutions (down to 2.2 µm3). This versatility enables brain-wide single neuron volumetric calcium imaging in behaving larval zebrafish (1×0.4 mm2 FOV at 5 Hz) and capillary blood cell tracking in living mouse brain (>3×3 mm2 FOV) with a sweeping 0.32 mm wide volume section at 100 Hz. In optically cleared mouse brain, the flexibility allows a screen-and-zoom capability by sequentially imaging the whole brain at low-and-high magnifications to locate and resolve subcellular structures such as dendritic tress and spines. By offering a switchable imaging resolution, volume, and speed, the self-adaptive OPM achieves a versatile platform for studying a wide range of multi-cellular model system, whether in vivo or fixed and optically cleared. Competing Interest Statement The authors have declared no competing interest. Footnotes This version has improved resolution in the PDF to correct some illegible text in the figures.