'Electron tomography' ('ET') is a tomography technique for obtaining detailed 3D structures of sub-cellular macro-molecular objects. Electron tomography is an extension of traditional transmission electron microscopy and uses a transmission electron microscope to collect the data. In the process, a beam of electrons is passed through the sample at incremental degrees of rotation around the center of the target sample. This information is collected and used to assemble a three-dimensional image of the target. Current resolutions of ET systems are in the 5–20 nm range, suitable for examining supra-molecular multi-protein structures, although not the secondary and tertiary structure of an individual protein or polypeptide.
In the field of biology, bright-field transmission electron microscopy (BF-TEM) and high-resolution TEM (HRTEM) are the primary imaging methods for tomography tilt series acquisition. However, there are two issues associated with BF-TEM and HRTEM. First, acquiring an interpretable 3-D tomogram requires that the projected image intensities vary monotonically with material thickness. This condition is difficult to guarantee in BF/HRTEM, where image intensities are dominated by phase-contrast with the potential for multiple contrast reversals with thickness, making it difficult to distinguish voids from high-density inclusions. Second, the contrast transfer function of BF-TEM is essentially a high-pass filter – information at low spatial frequencies is significantly suppressed – resulting in an exaggeration of sharp features. However, the technique of annular dark-field scanning transmission electron microscopy (ADF-STEM) more effectively suppresses phase and diffraction contrast, providing image intensities that vary with the projected mass-thickness of samples up to micrometres thick for materials with low atomic number. ADF-STEM also acts as a low-pass filter, eliminating the edge-enhancing artifacts common in BF/HRTEM. Thus, provided that the features can be resolved, ADF-STEM tomography can yield a reliable reconstruction of the underlying specimen which is extremely important for its application in material science. In 2010, a 3D resolution of 0.5±0.1×0.5±0.1×0.7±0.2 nm was achieved with a single-axis ADF-STEM tomography. Presently, the highest electron tomography resolution is around 2.4 angstrom as demonstrated by UCLA Miao group using a gold nanoparticle. This technique has recently been used to directly visualize the atomic structure of screw dislocations in nanoparticles.
Different tilting methods
The most popular tilting methods are the single-axis and the dual-axis tilting methods. Dual-axis tomography is sometimes referred to as conical tomography as well. By using dual-axis tilting, the elongation effect is reduced by a factor of however, twice as many images need to be taken.
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