What transmission electron microscopes can visualize now and in the future.
Our review concentrates on the progress made in high-resolution transmission electron microscopy (TEM) in the past decade. This includes significant improvements in sample preparation by quick-freezing aimed at preserving the specimen in a close-to-native state in the high vacuum of the microscope. Following advances in cold stage and TEM vacuum technology systems, the observation of native, frozen hydrated specimens has become a widely used approach. It fostered the development of computer guided, fully automated low-dose data acquisition systems allowing matched pairs of images and diffraction patterns to be recorded for electron crystallography, and the collection of entire tilt-series for electron tomography. To achieve optimal information transfer to atomic resolution, field emission electron guns combined with acceleration voltages of 200-300 kV are now routinely used. The outcome of these advances is illustrated by the atomic structure of mammalian aquaporin-O and by the pore-forming bacterial cytotoxin ClyA resolved to 12 A. Further, the Yersinia injectisome needle, a bacterial pseudopilus and the binding of phalloidin to muscle actin filaments were chosen to document the advantage of the high contrast offered by dedicated scanning transmission electron microscopy (STEM) and/or the STEM's ability to measure the mass of protein complexes and directly link this to their shape. Continued progress emerging from leading research laboratories and microscope manufacturers will eventually enable us to determine the proteome of a single cell by electron tomography, and to more routinely solve the atomic structure of membrane proteins by electron crystallography.
Müller
Shirley A
SA
Maurice E. Müller Institute for Structural Biology, Biozentrum, University of Basel, Klingelbergstr. 70, CH-4056 Basel, Switzerland. shirley.mueller@unibas.ch
Aebi
Ueli
U
Engel
Andreas
A
D003205:Computing Methodologies; D005544:Forecasting; D005615:Freezing; D049673:History, 20th Century; D049674:History, 21st Century; D046911:Macromolecular Substances / Q000737:chemistry; D046529:Microscopy, Electron, Transmission / Q000266:history / Q000379:methods* / Q000639:trends; D011506:Proteins / Q000737:chemistry; D013048:Specimen Handling / Q000379:methods
D046911:Macromolecular Substances; D011506:Proteins