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Electron Microscope Tomography and Single-particle Reconstruction01:07

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Transmission electron microscopy (TEM) can be used to determine the 3D structure of biological samples with the help of techniques such as electron microscope tomography and single-particle reconstruction. While single-particle reconstruction can examine macromolecules and macromolecular complexes in vitro conditions only, tomography permits the study of cell components or small cells in vivo.
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A scanning electron microscope (SEM) is used to study the surface features of a sample by using an electron beam that scans the sample surface in a two-dimensional manner. Typically, areas between ~1 centimeter to 5 micrometers in width can be imaged. SEM can be used to image bacteria, viruses, tissues as well as larger samples like insects. Conventional SEM gives a magnification ranging from 20X to 30,000X and spatial resolution of 50 to 100 nanometers.
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The early pioneers of microscopy opened a window into the invisible world of microorganisms. In 1830, Joseph Jackson Lister created an essentially modern light microscope. The 20th century saw the development of microscopes that leveraged nonvisible light, such as fluorescence microscopy that uses an ultraviolet light source and electron microscopy that uses short-wavelength electron beams. These advances significantly improved magnification, image resolution, and contrast. By comparison, the...
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The wavelengths of visible light ultimately limit the maximum theoretical resolution of images created by light microscopes. Most light microscopes can only magnify 1000X, and a few can magnify up to 1500X. Electrons, like electromagnetic radiation, can behave like waves, but with wavelengths of 0.005 nm, they produce significantly greater resolution up to 0.05 nm as compared to 500 nm for visible light. An electron microscope (EM) can create a sharp image that is magnified up to 2,000,000X.
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Related Experiment Video

Updated: Jan 4, 2026

Cryo-EM and Single-Particle Analysis with Scipion
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Cryo-EM and Single-Particle Analysis with Scipion

Published on: May 29, 2021

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Hybrid Electron Microscopy Normal Mode Analysis with Scipion.

Mohamad Harastani1, Carlos Oscar S Sorzano2, Slavica Jonić1

  • 1Sorbonne Université, UMR CNRS 7590, Muséum National d'Histoire Naturelle, IRD, Institut de Minéralogie, de Physique des Matériaux et de Cosmochimie, IMPMC, Paris, France.

Protein Science : a Publication of the Protein Society
|November 7, 2019
PubMed
Summary
This summary is machine-generated.

A new Hybrid Electron Microscopy Normal Mode Analysis (HEMNMA) software analyzes single-particle images to reveal continuous conformational changes in molecular complexes. This free, open-source tool, ContinuousFlex, is available for Scipion V2.0.

Keywords:
continuous conformational changescryo-electron microscopydynamicsnormal mode analysissingle-particle analysissoftwarestructure

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Area of Science:

  • Structural Biology
  • Biophysics
  • Computational Biology

Background:

  • Analyzing continuous conformational changes in molecular complexes is crucial for understanding their function.
  • Existing methods may struggle when conformational changes dominate orientational variations in single-particle images.
  • Hybrid Electron Microscopy Normal Mode Analysis (HEMNMA) was previously developed to address these challenges.

Purpose of the Study:

  • To present a new, user-friendly, free, and open-source software implementation of HEMNMA.
  • To make HEMNMA analysis accessible as part of the Scipion V2.0 ecosystem.
  • To facilitate the study of continuous conformational variability in molecular complexes from electron microscopy data.

Main Methods:

  • HEMNMA computes normal modes of a reference model (structure or EM map) for a molecular complex.
  • It analyzes single-particle images to determine the full distribution of conformational variability.
  • Simultaneous iterative optimization of particle conformation, orientation, and shift parameters is employed.

Main Results:

  • A new HEMNMA implementation, ContinuousFlex, is now available as a Scipion V2.0 plugin.
  • This software is independent of MATLAB, enhancing its accessibility.
  • The ContinuousFlex plugin is user-friendly, free, and open-source.

Conclusions:

  • The ContinuousFlex plugin provides a powerful and accessible tool for analyzing continuous conformational changes in molecular complexes using electron microscopy.
  • This advancement enables deeper insights into molecular dynamics and function.
  • The open-source nature promotes wider adoption and further development in structural biology research.