Cleared tissue dual-view oblique plane microscopy.

Dvinskikh L., Sparks H., Rowe-Brown L., Hong W., Ennis D., Sarnataro R., Carreno D., Tissot FS., Lo Celso C., Riglar DT., Cunnea P., McNeish IA., Dunsby C.

We present a dual-view oblique plane microscope (dOPM) for imaging thick optically cleared tissue samples using a silicone immersion primary objective. The custom-designed remote refocusing relay utilizes stock optics to achieve remote refocusing in refractive index-matched samples. The spatial resolution of the system was characterized using a series of fluorescent bead phantoms with refractive indices ranging from 1.4-1.5, with the point spread function full width at half maximum measuring ∼0.5 µm laterally and ∼1 µm axially for a refractive index-matched bead sample, with minimal degradation over a > 250 µm axial range. We characterize how the remote refocusing performance at sample refractive indices up to n = 1.5 can be partially compensated for using adjustment of the correction collar on the primary objective. We apply the system to imaging a range of biological samples with varied refractive indices. Combined with tiled acquisition, image stitching, and multi-view image fusion, the microscope enables multicolour imaging of mm-wide and ∼250 µm thick optically cleared mouse ovarian cancer, bone marrow, and colon tissue samples, and a whole Drosophila melanogaster fruit fly brain. The system offers a platform for fast and high-resolution, multicolour volumetric imaging across spatial scales, integrated on a commercially available microscope frame.

DOI

10.1364/BOE.572592

Type

Journal article

Publication Date

2025-09-01T00:00:00+00:00

Volume

16

Pages

3725 - 3739

Total pages

14

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