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Cryo-EM Single Particle Analysis

Single particle analysis is a group of related computerized image processing techniques used to analyze images from transmission electron microscopy (TEM).

What is Cryo-EM Single Particle Analysis?

Cryo-electron microscopy (cryo-EM) of single-particle specimens is used to determine the structure of proteins and macromolecular complexes without the need for crystals. These methods were developed to improve and extend the information obtainable from TEM images. Individual images of stained or unstained particles are very noisy, and so hard to interpret. Combining several digitised images of similar particles together gives an image with stronger and more easily interpretable features. Using cryo-electron microscopy it has become possible to generate reconstructions with sub-nanometer resolution and near-atomic resolution first in the case of highly symmetric viruses, and now in smaller, asymmetric proteins as well.

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Direct Electron Detectors and Cryo-EM

The remarkable progress in single-particle cryo-EM in the last 10 years has primarily been enabled by the development of direct electron detection (DED) cameras1,2,3. DED cameras have a superior detective quantum efficiency (DQE), a measure of the combined effects of the signal and noise performance of an imaging system4, and the underlying complementary metal-oxide semiconductor (CMOS) technology makes it possible to collect dose-fractionated image stacks, referred to as movies, that allow computational correction of specimen movements5,6 and electron counting for the best results.

Cryo-EM structure of human T-cell leukemia virus type-1 (HTLV-1) intasome

More reliable 3D density maps

Together, these features produce images of unprecedented quality, which, in turn, improves the results of digital image processing. In parallel, the continually increasing computer power allows the use of increasingly sophisticated image processing algorithms, resulting in greatly improved and more reliable 3D density maps.

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