Abstract Light sheet microscopy is a powerful technique for high-speed 3D imaging of subcellular dynamics and large biological specimens. How-ever, it often generates datasets ranging from hundreds of gigabytes to petabytes in size for a single experiment. Conventional com-putational tools process such images far slower than the time to acquire them and often fail outright due to memory limitations. To address these challenges, we present LLSM5DTools, a scalable soft-ware solution for efficient petabyte-scale light sheet image processing. This software incorporates a suite of commonly used processing tools that are memory and performance-optimized. Notable advancements include rapid image readers and writers, fast and memory-efficient geometric transformations, high-performance Richardson-Lucy decon-volution, and scalable Zarr-based stitching. These features outperform state-of-the-art methods by over one order of magnitude, enabling the processing of petabyte-scale image data at the full teravoxel rates of modern imaging cameras. The software opens new avenues for biological discoveries through large-scale imaging experiments.

Image processing tools for petabyte-scale light sheet microscopy data