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CRYO-ELECTRON MICROSCOPY

Cryo-electron microscopy (cryo-EM) is a powerful imaging technique in structural biology. Here are the key features of this technique:

Sample Preparation

  • Samples are rapidly frozen in a thin layer of vitreous ice on specialized grids

  • Vitrification preserves the sample in a near-native state without ice crystal formation

  • Typical sample thickness is 10-100 nm, ideally matching the maximum particle dimension
     

Imaging Conditions

  • Imaging is performed at cryogenic temperatures, typically around -188°C using liquid ethane as a coolant

  • Low-electron dose conditions are used to minimize radiation damage to sensitive biological samples

  • Accelerating voltages of 200-300 kV enable greater sample penetration
     

Key Advantages

  • Does not require sample crystallization, unlike X-ray crystallography

  • Can image a wide range of biological structures, from individual proteins to whole cells

  • Capable of achieving near-atomic resolution, with some structures resolved to 1.5 Å or better

Data Collection and Processing

  • Multiple 2D projection images are collected of randomly oriented particles

  • Advanced computational methods combine these 2D images to reconstruct 3D structures

  • Large datasets of hundreds of thousands of particle images may be processed
     

Applications

  • Determination of high-resolution structures of proteins, viruses, and macromolecular complexes

  • Visualization of dynamic biological processes through image series

  • Study of structures difficult or impossible to crystallize, like membrane proteins
     

Sub-techniques. Cryo-EM encompasses several related methods, including:

  • Single-particle analysis

  • Cryo-electron tomography

  • Cryo-correlative light and electron microscopy (cryo-CLEM)

Cryo-EM has revolutionized structural biology by enabling the visualization of biological structures in their native state at unprecedented resolutions, earning its developers the 2017 Nobel Prize in Chemistry.

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