Mesoscopic oblique plane microscopy with a diffractive light-sheet for large-scale 4D cellular resolution imaging

Area of Science:

• Biomedical Imaging
• Microscopy Technology
• Neuroscience

Background:

• Observing dynamic biological processes in 3D across mesoscopic scales requires high-resolution, large-field-of-view imaging.
• Existing microscopy techniques often face trade-offs between field of view and axial resolution, limiting the study of rapid, whole-organism phenomena.

Purpose of the Study:

• To develop a novel microscopy technique, mesoscopic oblique plane microscopy (Meso-OPM), to overcome current FOV and resolution limitations.
• To achieve high-speed, 3D volumetric imaging of biological structures at cellular resolution across mesoscopic scales.

Main Methods:

• Development of Meso-OPM utilizing a diffractive light sheet with an augmented illumination angle via a transmission grating.
• Demonstration of enhanced axial resolution, approximately sixfold improvement over existing methods.
• Achieved a large field of view (5.4 mm × 3.3 mm) with high resolution (2.5 μm × 3 μm × 6 μm) for single-scan volumetric imaging.

Main Results:

• Meso-OPM achieved approximately sixfold improvement in axial resolution.
• Demonstrated volumetric imaging of 3D cellular structures within a large FOV (5.4 mm × 3.3 mm).
• Successfully performed in vivo whole-body volumetric recordings of neuronal activity at 2 Hz and blood flow dynamics at 5 Hz in zebrafish larvae with 3D cellular resolution.

Conclusions:

• Meso-OPM provides a powerful tool for large-scale, high-speed, 3D dynamic imaging in biological systems.
• This technique enables unprecedented insights into whole-organism physiological processes at cellular resolution.
• Meso-OPM overcomes critical limitations in current microscopy, advancing the study of complex biological systems.