Cryo-Electron Microscopy

Abstract

Cryo-electron microscopy (cryo-EM) includes three technical methods, (1) rapid freezing for vitreous ice-embedding, (2) observation of frozen hydrated specimens, and (3) image processing for three-dimensional structural analysis. The three-dimensional structural analysis can be performed in three different ways. Electron crystallography can decipher the structure of membrane proteins at the highest resolution (atomic level). Single particle analysis now allows the determiration of the structure of highly purified proteins and complexes (non-crystalline biomolecules) in solution at the near-atomic level. Electron tomography can reveal the three-dimensional structure of an ultrathin-section of the cells and/or tissues. The resolution of the structure obtained by electron tomography is not very high (nm level), however it is possible to reveal the individual structures of biomolecular assemblies or cellular organelles, in close-to-native condition in the cell. A technical development for cryo-EM should be the introduction of a new CMOS camera for the direct detection of electrons. Using such camera, a short movie (usually 2-3 seconds), comprising numerous images with a few hundred milliseconds exposure each can be recorded. Such a movie has a demonstrated value, as it can compensate for the specimen motion induced by irradiation of electron beam. Improvements in image processing algorithms and computer programs are also essential for achieving three-dimensional structures at a better resolution.