Minutes-timescale 3D isotropic imaging of entire organs at subcellular resolution by content-aware compressed-sensing light-sheet microscopy

Fang, Chunyu; Chu, Tingting; Yu, Tingting; Huang, Yujie; Li, Yusha; Wan, Peng; Feng, Wenyang; Wang, Xuechun; Mei, Wei; Zhu, Dan; Fei, Peng

Instant 3D imaging of entire organs and organisms at cellular resolution is a recurring challenge in life science. Here we report on a computational light-sheet microscopy able to achieve minute-timescale mapping of entire macro-scale organs at high spatial resolution, thereby overcoming the throughput limit of current 3D microscopy implementations. Through combining a dual-side confocally-scanned Bessel light-sheet illumination which provides thinner-and-wider optical sectioning of deep tissues, with a content-aware compressed sensing (CACS) computation pipeline which further improves the contrast and resolution based on a single acquisition, our method yields 3D images with high, isotropic spatial resolution and rapid acquisition improved by two-orders of magnitude. We demonstrate the imaging of whole brain (∼400 mm3), entire gastrocnemius and tibialis muscles (∼200 mm3) of mouse at subcellular resolution (0.5-μm isovoxel) and ultra-high throughput of 5∼10 minutes per sample. Various system-level cellular analyses, such as mapping cell populations at different brain sub-regions, tracing long-distance projection neurons over the entire brain, and calculating neuromuscular junction occupancy across whole muscle, were also readily enabled by our method.