Large field-of-view and isotropic light sheet microscopy with aberration-free tunable foci

Light-sheet microscopy stands out as a powerful tool in biological imaging due to its exceptional performance in fluorescence imaging. However, achieving both high sectioning performance and a vast field of view (FOV) poses a fundamental challenge in conventional light-sheet microscopy. The light-sheet thickness is typically constrained to 1 μm for a wide FOV, potentially compromising resolution. To address this limitation, we introduce an axial scanning light-sheet microscopy (ASLM) technique integrated with aberration-free tunable foci to enable high-NA excitation while maintaining a generous FOV. The proposed scheme successfully achieves isotropic resolution of 280 nm in a three-dimensional imaging system, encompassing a FOV of 80 × 80 μm and an impressive imaging speed of 60 ms per frame. These remarkable characteristics underscore the immense potential of ASLM for high- spatiotemporal resolution imaging.