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Related Experiment Video

Updated: Nov 14, 2025

Correlative Light Electron Microscopy CLEM for Tracking and Imaging Viral Protein Associated Structures in Cryo-immobilized Cells
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HPM live μ for a full CLEM workflow.

Xavier Heiligenstein1, Marit de Beer2, Jérôme Heiligenstein1

  • 1CryoCapCell, Kremlin-Bicêtre, France.

Methods in Cell Biology
|March 12, 2021
PubMed
Summary
This summary is machine-generated.

Correlative microscopy (CM) integrates imaging techniques for biological research. This study introduces a high-pressure freezing system coupled to a light microscope for improved cryo-arrest in correlative light and electron microscopy (CLEM).

Keywords:
Bio-imagingCorrelative Light and Electron MicroscopyCryo-electron microscopyCryo-light microscopyElectron microscopyHigh-pressure freezingLive cell imagingVitrificationVolume electron microscopy

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Area of Science:

  • Biotechnology
  • Microscopy
  • Cell Biology

Background:

  • Multimodal imaging, particularly Correlative Light and Electron Microscopy (CLEM), revolutionizes biological and biomedical material analysis across length scales.
  • Traditional electron microscopy (EM) sample preparation involves chemical fixation, which can introduce artifacts; rapid cryogenic fixation (vitrification) preserves ultrastructure and protein fluorescence.
  • Bridging live imaging and cryo-arrest is crucial for capturing dynamic biological events in CLEM.

Purpose of the Study:

  • To develop and optimize a high-pressure freezing (HPF) system directly coupled to a light microscope for minimizing the time between live imaging and cryo-arrest.
  • To enhance sample preservation and capture biological events in CLEM workflows.
  • To explore the potential of modified HPF technology for imaging 3D samples and ensuring homogeneous deep vitrification.

Main Methods:

  • Development of a high-pressure freezing (HPF) system directly integrated with a light microscope.
  • Optimization of sample preservation and timing for live imaging to cryo-arrest transition.
  • Modification of HPF environmental parameters for improved vitrification of 2D and 3D samples.

Main Results:

  • Successful integration of HPF with light microscopy, enabling rapid cryo-arrest post-live imaging.
  • Demonstrated optimization of sample preservation and reduced time lag for capturing biological events.
  • Revisited HPF technology shows potential for homogeneous deep vitrification in 3D samples.

Conclusions:

  • The developed HPF system significantly enhances CLEM protocols by preserving cellular ultrastructure and fluorescence.
  • This integrated approach facilitates multi-step, multi-modal imaging for dynamic biological studies.
  • The optimized HPF system holds promise for advanced CLEM applications, particularly for correlating live imaging with electron microscopy.