Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, DC, USA.
Laboratory for Fluorescence Dynamics, Department of Biomedical Engineering, University of California, Irvine, CA, USA.
Nat Rev Nephrol. 2021 Feb;17(2):128-144. doi: 10.1038/s41581-020-00337-8. Epub 2020 Sep 18.
Fluorescence microscopy, in particular immunofluorescence microscopy, has been used extensively for the assessment of kidney function and pathology for both research and diagnostic purposes. The development of confocal microscopy in the 1950s enabled imaging of live cells and intravital imaging of the kidney; however, confocal microscopy is limited by its maximal spatial resolution and depth. More recent advances in fluorescence microscopy techniques have enabled increasingly detailed assessment of kidney structure and provided extraordinary insights into kidney function. For example, nanoscale precise imaging by rapid beam oscillation (nSPIRO) is a super-resolution microscopy technique that was originally developed for functional imaging of kidney microvilli and enables detection of dynamic physiological events in the kidney. A variety of techniques such as fluorescence recovery after photobleaching (FRAP), fluorescence correlation spectroscopy (FCS) and Förster resonance energy transfer (FRET) enable assessment of interaction between proteins. The emergence of other super-resolution techniques, including super-resolution stimulated emission depletion (STED), photoactivated localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM) and structured illumination microscopy (SIM), has enabled functional imaging of cellular and subcellular organelles at ≤50 nm resolution. The deep imaging via emission recovery (DIVER) detector allows deep, label-free and high-sensitivity imaging of second harmonics, enabling assessment of processes such as fibrosis, whereas fluorescence lifetime imaging microscopy (FLIM) enables assessment of metabolic processes.
荧光显微镜，特别是免疫荧光显微镜，已广泛用于研究和诊断目的的肾功能和病理学评估。20 世纪 50 年代共聚焦显微镜的发展使活细胞成像和肾脏活体成像成为可能；然而，共聚焦显微镜受到其最大空间分辨率和深度的限制。荧光显微镜技术的最新进展使我们能够越来越详细地评估肾脏结构，并为肾脏功能提供了非凡的见解。例如，快速光束振荡的纳米级精确成像（nSPIRO）是一种超分辨率显微镜技术，最初用于肾脏微绒毛的功能成像，能够检测肾脏中的动态生理事件。各种技术，如光漂白后荧光恢复（FRAP）、荧光相关光谱（FCS）和Förster 共振能量转移（FRET），可用于评估蛋白质之间的相互作用。其他超分辨率技术的出现，包括超分辨率受激辐射耗散（STED）、光激活定位显微镜（PALM）、随机光学重建显微镜（STORM）和结构光照明显微镜（SIM），使我们能够以≤50nm 的分辨率对细胞和亚细胞细胞器进行功能成像。通过发射恢复（DIVER）探测器进行的深度成像允许进行深层、无标记和高灵敏度的二次谐波成像，能够评估纤维化等过程，而荧光寿命成像显微镜（FLIM）则能够评估代谢过程。
