21世纪初诞生的超分辨光学成像技术凭借纳米级空间分辨率、低损制样等优点,迅速发展成为生命科学研究中不可或缺的技术手段。其中单分子定位超分辨成像(SMLM)技术更是由于其成像原理易懂、空间分辨率极高等特点,一直受到科研工作者的青睐,不断取得重要的技术和应用进展。首先回顾了SMLM的工作原理,讨论了其光路搭建、图像重建、漂移校正等关键技术问题。介绍了两类代表性SMLM技术。列举了多种多色SMLM方法,并分析了各自的优缺点。介绍了SMLM成像参数的改进研究,包括横/纵分辨率的提高、成像视野和深度的改善。介绍了SMLM和深度学习,SMLM和电镜等成像手段结合的关联成像研究进展。讨论了SMLM数据提取与分析方法。最后列举了SMLM在细胞生物学中的重要应用,并展望了SMLM未来的发展方向。期望该综述能为SMLM工作者提供有益的启发和参考,推进SMLM在生命科学研究中的深入应用。

Super-resolution microscopy invented at the beginning of the 21 ^st century has rapidly become a indispensable method in life science research owing to their nanoscale spatial resolution, low damage of sample preparation, and so on. Among a variety of super-resolution imaging techniques, single-molecule localization super-resolution microscopy (SMLM) with straightforward principle and outstanding spatial resolution gains more and more attention from researchers, thereby continuously making significant progress on the techniques and applications. Firstly, this paper reviewed the principle of SMLM and discussed some technical problems including the optical path building, the image reconstruction, and the drift correction. Two types of representative SMLMs were indroduced and discussed. Then, diversified multi-color SMLMs were introduced and their advantages and disadvantages were analyzed. The improvement of imaging parameters including the lateral/axial spatial resolution, imaging field, and imaging depth of SMLM was subsequently discussed. The research progress of correlated imaging of the SMLM combined with deep learning and SMLM combined with the electron microscope was further introduced. Moreover, the extraction and analysis methods of SMLM data were discussed. Finally, some important applications of SMLM in cell biology were listed and the development prospects of SMLM were discussed. We hope the present review could be a useful reference for the SMLM users and provides novel insights for them, thus promoting the in-depth applications of SMLM in life science research.