Journal: Frontiers in Physiology
Article Title: Serial intravital 2-photon microscopy and analysis of the kidney using upright microscopes
doi: 10.3389/fphys.2023.1176409
Figure Lengend Snippet: Intravital microscopy setup for upright imaging of the kidney across mm and sub-µm scales with a custom 3D printed animal holder. (A) 3D model and dimensions of a winged abdominal imaging window (wAIW) for serial imaging made from titanium. (B) wAIW slotted into a custom 3D-printed animal holder. The animal holder can be assembled from indicated parts that were printed separately on a consumer-grade 3D printer. (C) Individual steps of wAIW mounting. The wAIW gets slotted into the central plate of the animal holder and is locked into place by a top plate fixated through 2 lateral clips and 2 M6 bolts and nuts. (D) Anesthetized mouse after wAIW implantation showing correct orientation of the wings (facing dorsally and ventrally) to prevent hindlimb hindrance. Note the attached kidney surface to the coverslip. Scale bar = 10 mm. (E) Experimental setup for intravital microscopy of the kidney or other abdominal organs with an upright 2-photon microscope. The anesthetized mouse is mounted to the animal holder and placed on a heating pad on the microscope stage. (F) Maximum intensity projection of an intravital 2-photon microscopy 3D atlas scan acquired in a Cdh5-Confetti mouse kidney illustrates the potential of intravital microscopy to image mm-scale tissue regions. Scale bar = 500 µm. (G) High-resolution view (4096 × 4096 px, 124 nm/px) of a Cdh5-Confetti mouse provides paired morphological and functional readouts. Endothelial cells expressing the Brainbow2.1 reporter system can be identified through 10 different color combinations . A Setau-665-BSA dye (gray) labels the lumen of peritubular capillaries (ǂ) and apical albumin reabsorption in renal proximal tubules (arrowheads). Scale bar = 100 µm. (H) Zoomed-in inset of Setau-665-BSA detection channel (displayed in grey) demonstrates blood plasma labeling in peritubular capillaries (ǂ).
Article Snippet: Intravital imaging data were acquired on three different upright 2-photon microscopes: a Bruker Ultima IV (Bruker, United States), a Bruker Ultima Investigator Plus (Bruker, United States) and an Olympus FVMPE-RS (Olympus, Japan).
Techniques: Intravital Microscopy, Imaging, Microscopy, Functional Assay, Expressing, Labeling
