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
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Samples are rapidly frozen in a thin layer of vitreous ice on specialized grids
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Vitrification preserves the sample in a near-native state without ice crystal formation
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Typical sample thickness is 10-100 nm, ideally matching the maximum particle dimension
Imaging Conditions
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Imaging is performed at cryogenic temperatures, typically around -188°C using liquid ethane as a coolant
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Low-electron dose conditions are used to minimize radiation damage to sensitive biological samples
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Accelerating voltages of 200-300 kV enable greater sample penetration
Key Advantages
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Does not require sample crystallization, unlike X-ray crystallography
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Can image a wide range of biological structures, from individual proteins to whole cells
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Capable of achieving near-atomic resolution, with some structures resolved to 1.5 Å or better
Data Collection and Processing
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Multiple 2D projection images are collected of randomly oriented particles
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Advanced computational methods combine these 2D images to reconstruct 3D structures
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Large datasets of hundreds of thousands of particle images may be processed
Applications
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Determination of high-resolution structures of proteins, viruses, and macromolecular complexes
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Visualization of dynamic biological processes through image series
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Study of structures difficult or impossible to crystallize, like membrane proteins
Sub-techniques. Cryo-EM encompasses several related methods, including:
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Single-particle analysis
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Cryo-electron tomography
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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.