High-Resolution, Large Imaging Volume, and Multi-View Single Objective Light-Sheet Microscopy

Recent developments in Oblique Plane Microscopy (OPM) have shown that it can achieve high spatio-temporal resolution. Here we describe a single objective light-sheet microscope based on oblique plane illumination that achieves: (i) large field of view and high-resolution imaging via a custom remote focusing objective; (ii) fast volumetric imaging by means of light-sheet stabilised stage scanning – a novel scanning modality that extends the imaging volume without compromising imaging speed nor quality; (iii) multi-view imaging by alternating the orientation of light-sheet illumination and detection to improve the image quality on large samples; (iv) simpler design and ergonomics by remote placement of coverslips to allow inverted imaging, enabling imaging across scales in a high-throughput format. Overall, we achieved a resolution of 450 nm laterally and 2 μm axially and a field of view of 3000 μm × 800 μm × 300 μm. We demonstrate the speed, field of view, resolution and versatility of our novel instrument by imaging various systems, including zebrafish whole brain activity, Drosophila egg chamber development, and zebrafish development – up to nine embryos simultaneously.

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