Advances in fluorescence microscopy techniques to study kidney function

• Published in: Nature reviews. Nephrology Vol. 17; no. 2; pp. 128 - 144
• Main Authors: Ranjit, Suman, Lanzanò, Luca, Libby, Andrew E., Gratton, Enrico, Levi, Moshe
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 01-02-2021; Nature Publishing Group
• Subjects: 631/1647/245; 631/80; 692/4022/272; Humans; Kidney - diagnostic imaging; Kidney - metabolism; Kidney - physiopathology; Kidney Diseases - diagnostic imaging; Medicine; Medicine & Public Health; Microscopes; Microscopy; Microscopy, Fluorescence; Nephrology; Review Article
• ISSN: 1759-5061; 1759-507X
• DOI: 10.1038/s41581-020-00337-8

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.
This Review provides an overview of fluorescence-based microscopy techniques that have been used to study molecular processes within the kidney. The authors describe how the development of cutting-edge technologies has enabled high spatiotemporal resolution of molecular interactions and processes, and how these approaches have aided our understanding of kidney dynamics.
Key points
Microscopic imaging has revolutionized our understanding of kidney structure and physiology.
Kidney physiology is determined by dynamic processes and, although understanding of kidney structures at super-resolution is needed to understand kidney function, knowledge of kidney structure per se cannot explain kidney function.
Only a few super-resolution microscopy techniques, including super-resolution stimulated emission depletion (STED) microscopy and nanoscale precise imaging by rapid beam oscillation (nSPIRO), can simultaneously provide high-resolution structural information and insights into kidney dynamics.
Imaging of individual cells in tissues requires methods that are capable of imaging deep into the tissue, such as two-photon excitation and autofluorescence methods, imaged using methodologies such as the deep imaging via emission recovery (DIVER) detector; single photon excitation using dyes and immunofluorescence approaches cannot penetrate deep into live tissue.