Rapid 3D isotropic imaging of whole organ with double-ring light-sheet microscopy and self-learning side-lobe elimination.

Bessel-like plane illumination forms a new type of light-sheet microscopy with ultra-long optical sectioning distance that enables rapid 3D imaging of fine cellular structures across an entire large tissue. However, the side-lobe excitation of conventional Bessel light sheets severely impairs the quality of the reconstructed 3D image. Here, we propose a self-supervised deep learning (DL) approach that can completely eliminate the residual side lobes for a double-ring-modulated non-diffraction light-sheet microscope, thereby substantially improving the axial resolution of the 3D image. This lightweight DL model utilizes the own point spread function (PSF) of the microscope as prior information without the need for external high-resolution microscopy data. After a quick training process based on a small number of datasets, the grown-up model can restore sidelobe-free 3D images with near isotropic resolution for diverse samples. Using an advanced double-ring light-sheet microscope in conjunction with this efficient restoration approach, we demonstrate 5-minute rapid imaging of an entire mouse brain with a size of ∼12 mm × 8 mm  ×  6 mm and achieve uniform isotropic resolution of ∼4 µm (1.6-µm voxel) capable of discerning the single neurons and vessels across the whole brain.