Contact & Support

3D STED Nanoscopy

The podocyte cell layer in the glomerular filtration barrier is a key player in kidney diseases. Alterations in the architecture of the podocytes and the basement membrane are linked to the onset of renal failure. Improve your workflow in kidney research with 3D STED Nanoscopy.

Standard electron microscopy techniques based on elaborate protocols identify glomerular diseases based on two-dimensional information. The combination of optical clearing and 3D STED Nanoscopy (Stimulated Emission Depletion) with the Leica TCS SP8 STED 3X opens a new avenue for imaging the kidney in unprecedented three-dimensional detail in light microscopy.

Top image: Cleared kidney sample, revealing the podocyte celllayer; magnified podocytes by STED and electron microscopy (EM). Cleared kidney sample courtesy: David Unnersjö-Jess, KTH, Stockholm, Sweden. Scanning EM image of kidney podocytes courtesy: Center for Microscopy and Image Analysis, Univ. of Zurich, Dr. Urs Ziegler. Scale bar is 1 µm.

Contact us for STED imaging solutions!

Please contact us for personal expert advice on our microscopy solutions for STED imaging.

Visualize the ultrastructure of the glomerular filtration barrier by deep 3D Nanoscopy

Even deeper than 150 µm within the tissue, the ultrastructural features of the specimen come to light powered by the new STED WHITE glycerol objective lens. This is achieved not only at 23°C, but also under live cell imaging conditions at 37°C with a comfortable working distance of 300 µm.

Forget the hassle of physical sectioning. Enjoy spectral freedom and keep up with the growing palette of fluorescent biomarkers.

Be at the forefront of clinical research.

3D Nanoscopy Workflow for Cleared Kidney Samples

Nanoscopy with the TCS SP8 STED system equipped with the HC PL APO 93X/1.30 GLYC motCORR – STED WHITE objective enables deep insights into the ultrastructure of healthy and diseased tissue.

In contrast to alternative methods, such as electron microscopy, STED nanosocopy delivers 3D data, and can display several proteins simultaneously.

Order protocol

1. Sample Preparation

The very first step is to dissect the kidney and stabilize its structure and biomolecular content with the help of a hydrogel. Therefore it is  incubated at 4°C in an acrylamid based gel to  allow gel penetration into the tissue.  For polymerization, the temperature is shifted to 37°C.

2. Sectioning

After stabilization in a hydrogel the kidney is cut in slices of up to 500 µm thickness using a vibratome. 

3. Clearing

Optical clearing helps to make thick biological samples transparent and therefore accessible to deep imaging. Moreover, clearing increases antibody penetration depth and staining quality. For a decent clearing, the kidney sections are transferred into a clearing solution and incubated at 50°C for several days.

4. Fluorescence Immunolabeling

Standard immunolabelling protocols deliver excellent signal to noise ratio in cleared samples. The 1st as well as the 2nd antibody are incubated for a whole day to allow a penetration into the thick sample.

5. Mounting

Light scattering occurs due to the mismatch of Refravtive Indices (RI) in the penetrating media. Thus, in the mounting step the RI of the sample and the mounting medium have to be considered. The RI of fixed tissues is around 1.45. Whereas e.g. water has an RI of 1.33, a Fructose solution matches the RI of fixed tissue. Therefore, samples are mounted in fructose to match the refractive index, allowing imaging with high penetration depth.

6. Nanoscopy

Image the ultrastructure of the glomerular filtration barrier at high resolution and at great depth. With the HC PL APO 93X/1.30 GLYC motCORR – STED WHITE Objective mounted on a TCS SP8 STED 3X microscope, nanoscopy up to 100 µm deep is possible. 

7. Visualization

Display your 3D STED results with the LAS X 3 D visualization tool – easy and straight forward.

Further Reading

Blom et al., Chem. Rev., 2017. Stimulated Emission Depletion Microscopy.

Randles et al., Sci. Rep., 2016. Three-dimensional electron microscopy reveals the evolution of glomerular barrier injury.

Unnersjö-Jess et al., Kidney International, 2015: Super-resolution stimulated emission depletion imaging of slit diaphragm proteins in optically cleared kidney tissue

Additional fields of research

  • Neuroscience
  • Pathology
  • Renal diseases
  • Glomerulopathies
  • Clinical research
  • Pathology research
  • Pharmaceutical research

News & Related articles

11. October 2017 White Blood Cell Recruitment to Bacterial Infection

Zebrafish In Vivo Study Using Leica Fluorescence Microscopes

Tile scan image of Mouse brain tissue section with multicolor immunofluorescent labeling

23. May 2017 Find the Missing Links in Advanced Live Cell Research - Leica launches DMi8 S live cell imaging solution

Wetzlar, Germany – Leica introduces the DMi8 S, a new complete solution designed to give researchers the ultimate tool for fast, versatile microscopic imaging of living cells allowing scientists to find, observe and interact with living cells like...

11. April 2017 Full Spectral Freedom for Multi-color Deep Tissue In Vivo Imaging

Leica Microsystems launches SP8 DIVE – The World's First Tunable Deep Imaging Solution

01. November 2016 Explore life’s true nature with super-resolution and nanoscopy

With HyVolution 2 and the Leica TCS SP8 STED ONE, Leica Microsystems paves the way to access imaging beyond the diffraction limit

21. May 2015 GSD Around the World – A Travel Diary on Facebook

Tamara Straube and Peter Laskey will take you on a journey around the world: Together with a team of specialists, they visited 10 countries, organized 11 workshops for GSD super-resolution microscopy, imaging and sample preparation, and met more than...

13. March 2015 EMBL Advanced Course – Intensive Super-Resolution Microscopy Course for Localization Microscopy and STED

From 20th to 25th July 2015 the EMBL (European Molecular Biology Laboratory), Heidelberg will be holding a six-day intensive course on the subject of super-resolution microscopy in cooperation with Leica Microsystems.

19. January 2015 New Leica TCS SP8 at Bates College: „Second of its kind in Maine, new microscope is a game-changer“

According to Bates College, the confocal offers new imaging capabilities and supports research in biology, neuroscience, nanotechnology and photophysics.

03. December 2014 Free Infographic: Which Super-Resolution Microscopy Method is Right for You?

This illustrated guide to super-resolution microscopy helps you compare technologies to best fulfill your individual research needs.

The Leica TCS SP8 STED 3X was chosen among The Scientist Top 10 Innovations 2014

01. December 2014 The Scientist Top 10 Innovations Award for Leica TCS SP8 STED 3X

Leica Microsystems’ STED Super-Resolution Microscope Again Awarded for its Innovative Technology

Leica HyD SMD – the universal detector for SMD and imaging.

19. November 2014 Universal Hybrid Detector for Single Molecule Detection and Imaging at SfN and ASCB

Leica HyD SMD - the Optimal Detector for Precise and Reliable SMD data