Fast, 3D Isotropic Imaging of Whole Mouse Brain Using Multiangle‐Resolved Subvoxel SPIM

The recent integration of light‐sheet microscopy and tissue‐clearing has facilitated an important alternative to conventional histological imaging approaches. However, the in toto cellular mapping of neural circuits throughout an intact mouse brain remains highly challenging, requiring complicated mechanical stitching, and suffering from anisotropic resolution insufficient for high‐quality reconstruction in 3D. Here, the use of a multiangle‐resolved subvoxel selective plane illumination microscope (Mars‐SPIM) is proposed to achieve high‐throughput imaging of whole mouse brain at isotropic cellular resolution. This light‐sheet imaging technique can computationally improve the spatial resolution over six times under a large field of view, eliminating the use of slow tile stitching. Furthermore, it can recover complete structural information of the sample from images subject to thick‐tissue scattering/attenuation. With Mars‐SPIM, a digital atlas of a cleared whole mouse brain (≈7 mm × 9.5 mm × 5 mm) can readily be obtained with an isotropic resolution of ≈2 µm (1 µm voxel) and a short acquisition time of 30 min. It provides an efficient way to implement system‐level cellular analysis, such as the mapping of different neuron populations and tracing of long‐distance neural projections over the entire brain. Mars‐SPIM is thus well suited for high‐throughput cell‐profiling phenotyping of brain and other mammalian organs.