ClearScope: Sub-Cellular Light-Sheet Imaging of Intact, Cleared Tissue
Fay MG, Lang PJ, Denu DS, O’Connor NJ, Haydock B, Blaisdell J, Roussel N, Wilson A, Aronson SM, Angstman PJ, Gong C, Butola T, Devinsky O, Basu J, Tomer R, Glaser JR. ClearScope: a fully integrated light sheet theta microscope for sub-cellular resolution imaging without lateral size constraints. bioRxiv [Preprint]. 2024:2024.08.15.608141. doi: 10.1101/2024.08.15.608141.
Background: Three-dimensional (3D) imaging of cleared, intact brain tissue is essential for mapping neural connectivity and investigating structural changes in neurological and neuropsychiatric disorders. Light-sheet fluorescence microscopy (LSFM) offers rapid, high-resolution imaging but is limited by lateral (XY) specimen size constraints due to its orthogonal optical design.
Hypothesis: This study tested the hypothesis that a fully integrated light sheet theta microscopy (LSTM) system – ClearScope – can achieve sub-cellular resolution imaging of intact, cleared specimens without lateral size limitations, while providing an accessible and streamlined workflow.
Methods: ClearScope employs dual oblique illumination and a perpendicular detection path, enabling deep tissue imaging regardless of lateral dimensions. It integrates an automated detection objective changer, adaptive refractive index mismatch correction, intelligent tissue scanning workflows and comprehensive pre- and post-processing tools. Performance was evaluated using fluorescent microspheres and biological samples, including whole mouse brains and human cortex.
Results: ClearScope achieves lateral resolution of ~0.33 µm using a 20x detection objective and axial resolutions of 5.4–6.7 µm across the field of view. Imaging intact mouse brains and human tissue yielded high-quality, minimally distorted 3D images suitable for digital neuron tracing, vascular reconstruction and stereologic analysis. The system reliably handles large samples and varied refractive indices with intuitive user interfaces.
https://pubmed.ncbi.nlm.nih.gov/39229056/
Conclusions: ClearScope advances volumetric imaging by enabling sub-cellular resolution without lateral size constraints. Its integrated, user-friendly platform expands access to whole-organ imaging in neuroscience, systems biology and related disciplines.
