High-fidelity tissue super-resolution imaging achieved with confocal2 spinning-disk image scanning microscopy

作者：Qianxi Liang, Wei Ren, Boya Jin, Liang Qiao, Xichuan Ge, Yunzhe Fu, Xiaoqi Lu, Meiqi Li, Peng Xi · 发表于：Light Science & Applications · 年份：2025 · DOI：10.1038/s41377-025-01930-x · 被引用次数：11 · 研究领域：Advanced Fluorescence Microscopy Techniques、Optical Coherence Tomography Applications、Digital Holography and Microscopy

Abstract Super-resolution imaging has revolutionized our ability to visualize biological structures at subcellular scales. However, deep-tissue super-resolution imaging remains constrained by background interference, which leads to limited depth penetration and compromised imaging fidelity. To overcome these challenges, we propose a novel imaging system, confocal² spinning-disk image scanning microscopy (C 2 SD-ISM). It integrates a spinning-disk (SD) confocal microscope, which physically eliminates out-of-focus signals, forming the first confocal level. A digital micromirror device (DMD) is employed for sparse multifocal illumination, combined with a dynamic pinhole array pixel reassignment (DPA-PR) algorithm for ISM super-resolution reconstruction, forming the second confocal level. The dual confocal configuration enhances system resolution, while effectively mitigating scattering background interference. Compared to computational out-of-focus signal removal, SD preserves the original intensity distribution as the penetration depth increases, achieving an imaging depth of up to 180 μm. Additionally, the DPA-PR algorithm effectively corrects Stokes shifts, optical aberrations, and other non-ideal conditions, achieving a lateral resolution of 144 nm and an axial resolution of 351 nm, and a linear correlation of up to 92% between the original confocal and the reconstructed image, thereby enabling high-fidelity super-resolution imaging. Moreover, the system’s programmable illum...