# ClearScope: A Fully Integrated Light-Sheet Theta Microscope for Sub-Micron-Resolution Imaging Without Lateral Size Constraints

## TL;DR

ClearScope is a new microscope that can image large brain samples in high detail without size limits, helping study brain structure and disease.

## Contribution

A fully integrated and user-friendly light-sheet theta microscope system that enables sub-micron-resolution imaging of large biological specimens without lateral size constraints.

## Key findings

- ClearScope enables subcellular-resolution imaging of large cleared brain specimens.
- The system supports seamless workflow from image acquisition to quantitative analysis.
- Demonstrated performance includes high-resolution 3D imaging of mouse and human brain samples.

## Abstract

Three-dimensional (3D) ex vivo imaging of cleared tissue from intact brains from animal models, human brain surgical specimens, and large postmortem human and non-human primate brain specimens is essential for understanding physiological neural connectivity and pathological alterations underlying neurological and neuropsychiatric disorders. Contemporary light-sheet microscopy enables rapid, high-resolution imaging of large, cleared samples but is limited by the orthogonal arrangement of illumination and detection optics, which constrains specimen size. Light-sheet theta microscopy (LSTM) overcomes this limitation by employing two oblique illumination paths while maintaining a perpendicular detection geometry. Here, we report the development of a next-generation, fully integrated and user-friendly LSTM system that enables uniform subcellular-resolution imaging (with subcellular resolution determined by the lateral performance of the system) throughout large specimens without constraining lateral (XY) dimensions. The system provides a seamless workflow encompassing image acquisition, data storage, pre- and post-processing, enhancement and quantitative analysis. Performance is demonstrated by high-resolution 3D imaging of intact mouse brains and human brain samples, including complete downstream analyses such as digital neuron tracing, vascular reconstruction and design-based stereological analysis. This enhanced and accessible LSTM implementation enables rapid quantitative mapping of molecular and cellular features in very large biological specimens.