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Preparation of Samples for Electron Microscopy01:20

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To be visualized by an electron microscope, either transmission or scanning, biological samples need to be fixed (stabilized) so the electron beam does not destroy them and dried thoroughly (desiccated/dehydrated) so the vacuum does not affect them. Fixation needs to be done as quickly as possible because the sample properties will start changing as soon as it is removed from its natural environment. For example, in a tissue sample, the oxygen levels begin decreasing, causing an altered...
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Assessment of Kidney Function in Mouse Models of Glomerular Disease
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Sample Preparation and Stereological Methods for the Study of Glomerular Ultrastructure Using Electron Microscopy.

Kathryn E White1

  • 1EM Research Services, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne, UK. Kathryn.white@ncl.ac.uk.

Methods in Molecular Biology (Clifton, N.J.)
|November 9, 2019
PubMed
Summary

This chapter details conventional and serial block face scanning electron microscopy techniques for preparing renal tissue. It provides protocols for processing, sectioning, imaging, and quantitative data analysis for kidney research.

Keywords:
Electron microscopyGlomerulusMesangiumPodocyteStereology

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Area of Science:

  • Nephrology
  • Microscopy
  • Biological Sciences

Background:

  • Transmission electron microscopy (TEM) is crucial for ultrastructural analysis of renal tissue.
  • Advances in microscopy necessitate updated protocols for optimal tissue preparation.

Purpose of the Study:

  • To describe conventional methods for preparing renal tissue for TEM.
  • To introduce and detail serial block face scanning electron microscopy (SBF-SEM) for renal tissue.
  • To provide comprehensive protocols for tissue processing, sectioning, imaging, and quantitative analysis.

Main Methods:

  • Detailed protocols for conventional fixation, embedding, and sectioning for TEM.
  • Step-by-step guide for serial block face scanning electron microscopy (SBF-SEM) sample preparation.
  • Methods for image acquisition and analysis using both TEM and SBF-SEM.

Main Results:

  • Established protocols for high-resolution imaging of renal ultrastructure using TEM.
  • Demonstrated the utility of SBF-SEM for generating 3D reconstructions of renal tissue.
  • Provided methods for extracting quantitative data from electron microscopy images.

Conclusions:

  • Both conventional TEM and SBF-SEM are valuable techniques for renal tissue analysis.
  • The described protocols facilitate detailed ultrastructural and quantitative studies of the kidney.
  • This chapter serves as a practical guide for researchers in renal histology and pathology.