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Updated: May 11, 2025

Serial Block-Face Scanning Electron Microscopy SBEM for the Study of Dendritic Spines
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New Approaches Based on Serial-Block Face Electron Microscopy to Investigate the Peripheral Nervous System.

Vitalijs Borisovs1,2, Mario Bossi1, Laura Matino3

  • 1Experimental Neurology Unit, School of Medicine and Surgery, Università di Milano-Bicocca, Monza, Italy.

Journal of the Peripheral Nervous System : JPNS
|April 19, 2025
PubMed
Summary

Automated uranyl-free staining offers a competitive alternative for serial block face scanning electron microscopy (SBF-SEM) of peripheral nerves and dorsal root ganglia (DRG). This method streamlines 3D ultrastructural analysis, making complex imaging more accessible.

Keywords:
dorsal root gangliaperipheral nervescanning electron microscopyserial block face

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Area of Science:

  • Electron Microscopy
  • Neuroscience
  • Biotechnology

Background:

  • Serial block face scanning electron microscopy (SBF-SEM) provides high-resolution 3D imaging but faces challenges in sample preparation and reconstruction.
  • Peripheral nerves and dorsal root ganglia (DRG) present unique complexities for ultrastructural analysis.

Purpose of the Study:

  • To evaluate and optimize sample preparation techniques for SBF-SEM in peripheral nerve and DRG specimens.
  • To compare manual gold standard staining with automated transmission electron microscopy (TEM) and uranyl-free en bloc preparation methods.
  • To assess the feasibility of automated 3D reconstruction for these challenging tissues.

Main Methods:

  • Three distinct SBF-SEM sample preparation techniques were tested: manual high molecular weight staining, automated standard TEM preparation, and automated uranyl-free en bloc preparation.
  • Specimens from mouse caudal nerve and DRG were utilized.
  • Image analysis and 3D rendering were performed using integrated software solutions.

Main Results:

  • The manual high molecular weight staining method showed superior results overall.
  • Automated standard TEM and uranyl-free methods yielded high-quality images for caudal nerve specimens.
  • Automated 3D rendering was successful for all tested samples.
  • Uranyl-free preparation significantly improved contrast in DRG specimens, overcoming challenges with standard TEM preparation.

Conclusions:

  • Automated uranyl-free staining is a viable alternative to traditional methods for SBF-SEM, offering competitive results with some limitations.
  • High-quality SBF-SEM imaging of peripheral nerves is achievable using standard TEM preparation, enabling analysis of previously embedded samples.
  • These advancements facilitate more accessible ultrastructural analysis of neural tissues.