High-speed mosaic imaging using scanner-synchronized stage position sampling

SIGNIFICANCE: Two-photon and confocal microscopy can obtain high frame rates; however, mosaic imaging of large tissue specimens remains time-consuming and inefficient, with higher imaging rates leading to a larger fraction of time wasted translating between imaging locations. Strip scanning obtains faster mosaic imaging rates by translating a specimen at constant velocity through a line scanner at the expense of more complex stitching and geometric distortion due to the difficulty of translating at completely constant velocity. AIM: We aim to develop an approach to mosaic imaging that can obtain higher accuracy and faster imaging rates while reducing computational complexity. APPROACH: We introduce an approach based on scanner-synchronous position sampling that enables subwavelength accurate imaging of specimens moving at a nonuniform velocity, eliminating distortion. RESULTS: We demonstrate that this approach increases mosaic imaging rates while reducing computational complexity, retaining high SNR, and retaining geometric accuracy. CONCLUSIONS: Scanner synchronous strip scanning enables accurate, high-speed mosaic imaging of large specimens by reducing acquisition and processing time.