David Unnersjö-Jess1,2, Amer Ramdedovic1,2, Martin Höhne1,2, Linus Butt1, Felix C Koehler1,2, Roman-Ulrich Müller1,2, Peter F Hoyer3, Hans Blom4, Bernhard Schermer1,2, Thomas Benzing1,2. 1. Department II of Internal Medicine and Center for Molecular Medicine Cologne (CMMC), University of Cologne, Cologne, Germany. 2. Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany. 3. Pediatric Nephrology, Pediatrics II, University of Duisburg-Essen, Essen, Germany. 4. Science for Life Laboratory, Department of Applied Physics, Royal Institute of Technology, Solna, Sweden.
Abstract
Background: Diseases of the glomeruli, the renal filtration units, are a leading cause of progressive kidney disease. Assessment of the ultrastructure of podocytes at the glomerular filtration barrier is essential for diagnosing diverse disease entities, providing insight into the disease pathogenesis, and monitoring treatment responses. Methods: Here we apply previously published sample preparation methods together with stimulated emission depletion and confocal microscopy for resolving nanoscale podocyte substructure. The protocols are modified and optimized in order to be applied to formalin-fixed paraffin-embedded (FFPE) samples. Results: We successfully modified our protocols to allow for deep three-dimensional stimulated emission depletion and confocal imaging of FFPE kidney tissue with similar staining and image quality compared with our previous approaches. We further show that quantitative analysis can be applied to extract morphometrics from healthy and diseased samples from both mice and humans. Conclusions: The results from this study could increase the feasibility of implementing optical kidney imaging protocols in clinical routines because FFPE is the gold-standard method for storage of patient samples.
Background: Diseases of the glomeruli, the renal filtration units, are a leading cause of progressive kidney disease. Assessment of the ultrastructure of podocytes at the glomerular filtration barrier is essential for diagnosing diverse disease entities, providing insight into the disease pathogenesis, and monitoring treatment responses. Methods: Here we apply previously published sample preparation methods together with stimulated emission depletion and confocal microscopy for resolving nanoscale podocyte substructure. The protocols are modified and optimized in order to be applied to formalin-fixed paraffin-embedded (FFPE) samples. Results: We successfully modified our protocols to allow for deep three-dimensional stimulated emission depletion and confocal imaging of FFPE kidney tissue with similar staining and image quality compared with our previous approaches. We further show that quantitative analysis can be applied to extract morphometrics from healthy and diseased samples from both mice and humans. Conclusions: The results from this study could increase the feasibility of implementing optical kidney imaging protocols in clinical routines because FFPE is the gold-standard method for storage of patient samples.
Authors: David Unnersjö-Jess; Lena Scott; Sonia Zambrano Sevilla; Jaakko Patrakka; Hans Blom; Hjalmar Brismar Journal: Kidney Int Date: 2017-12-11 Impact factor: 10.612
Authors: Bernhard Schermer; Thomas Benzing; Linus Butt; David Unnersjö-Jess; Martin Höhne; Aurelie Edwards; Julia Binz-Lotter; Dervla Reilly; Robert Hahnfeldt; Vera Ziegler; Katharina Fremter; Markus M Rinschen; Martin Helmstädter; Lena K Ebert; Hayo Castrop; Matthias J Hackl; Gerd Walz; Paul T Brinkkoetter; Max C Liebau; Kálmán Tory; Peter F Hoyer; Bodo B Beck; Hjalmar Brismar; Hans Blom Journal: Nat Metab Date: 2020-05-11
Authors: Octavian Bucur; Feifei Fu; Mike Calderon; Geetha H Mylvaganam; Ngoc L Ly; Jimmy Day; Simon Watkin; Bruce D Walker; Edward S Boyden; Yongxin Zhao Journal: Nat Protoc Date: 2020-04-01 Impact factor: 13.491
Authors: Peter Ilgen; Stefan Stoldt; Lena-Christin Conradi; Christian Andreas Wurm; Josef Rüschoff; B Michael Ghadimi; Torsten Liersch; Stefan Jakobs Journal: PLoS One Date: 2014-07-15 Impact factor: 3.240