Abstract
Fundamental understanding of large-scale dynamic connectivity within a living organism requires volumetric imaging over a large field of view (FOV) at biologically relevant speed and resolution. However, most microscopy methods make trade-offs between FOV and depth resolution, making it challenging to observe highly dynamic processes at cellular resolution in 3D across mesoscopic scales (e.g., whole zebrafish larva). To overcome this limitation, we have developed mesoscopic oblique plane microscopy (Meso-OPM) with a diffractive light sheet. By augmenting the illumination angle of the light sheet with a transmission grating, the axial resolution was improved ~6-fold over existing methods and ~2-fold beyond the diffraction limitation of the primary objective lens. We demonstrated an unprecedented FOV up to 5.4 × 3.3 mm with resolution of 2.5× 3 × 6 μm, allowing volumetric imaging of 3D cellular structures with a single scan. Applying Meso-OPM for in vivo imaging of zebrafish larvae, we report here the first in toto whole body volumetric recordings of neuronal activity at 2 Hz volume rate and the first example of whole body volumetric recordings of blood flow dynamics at 5 Hz with 3D cellular resolution.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
1)We correct Eq.(1) in the main text; 2)The legend of Fig. 1 is updated so that panel d only illustrates the comparison of FOV and axial resolution. 3)The angle of the light sheet in the water is corrected. 4)The mismatching of the back aperture of OL1 and OL2 is now explained. 5)The axial FOV of our Meso-OPM is now clarified to be 0.33 mm.
