Application of Electron Diffraction to Biological Electron Microscopy

Three methods by which electron diffraction may be applied to problems in electron microscopy are discussed from a fundamental point of view, and experimental applications with biological specimens are demonstrated for each case. It is shown that wide-angle electron diffraction provides valuable information for evaluating specimen damage that can occur either during specimen preparation or while in the electron beam. Dark-field electron microscopy can be used both to enhance the image contrast and to provide highly restricted and therefore highly specific information about the object. Low-angle electron diffraction provides quantitative information about the object structure in the range from 20 A to ~ 1000 A. Lowangle electron diffraction also demonstrates the important role of Fourier contrast with biological specimens, which are usually characterized by structural features with dimensions of 20 A or larger.     

Electron cryotomography

Electron cryotomography is an emerging technique that allows the structures of unique biological objects such as individual macromolecules, viruses, and even small whole cells to be reconstructed in their near-native states in three dimensions (3-D) to an approximate 5-nm resolution. The required instrumentation, sample preparation and limitations, data collection, typical results, and future prospects are summarized briefly.     

Anomalous Electron Transport in One-Dimensional Electron Cyclotron Drift Turbulence

Plasma Physics Reports - The transverse electron current due to the crossed electric and magnetic fields results in the robust instability driven by the electron $${\mathbf{E}} \times...     

Generation of Low-Energy High-Current Electron Beams in Plasma-Anode Electron Guns

This paper is a review of studies on the generation of low-energy high-current electron beams in electron guns with a plasma anode and an explosive-emission cathode. The problems related to the initiation of explosive electron emission under plasma and the formation and transport of high-current electron beams in plasma-filled systems are discussed consecutively. Considerable attention is given to the nonstationary effects that occur in the space charge layers of plasma. Emphasis is also placed on the problem of providing a uniform energy density distribution over the beam cross section, which is of critical importance in using electron beams of this type for surface treatment of materials. Examples of facilities based on low-energy high-current electron beam sources are presented and their applications in materials science and practice are discussed.     

Electron microscopic morphometry

The fundamental concepts of morphometry, the principal correction factors for systematic errors and the basic principles of efficient sampling are outlined for quantitative morphology at the electron microscopic level of resolution. The important usefulness of correlating electron microscopic morphometry with complementary light microscopic morphometry is emphasized.     

Electron microscope tomography

Electron microscope tomography allows three-dimensional reconstruction of ultrastructural objects at the molecular level. The method is general and not limited to symmetric, or regularly ordered structures. Alone, or in combination with immunoelectron microscopy and electron spectroscopic imaging, electron microscope tomography is a powerful technique in cell and molecular biology.     

Damage to Biological Samples Caused by the Electron Beam during Electron Microscopy

A new method has been developed which measures directly the beam damage suffered by biological specimens in the electron microscope. This method involves the use of radioautography to measure specific radioactivity of labeled specimens, either exposed or unexposed to the beam. Using this technique, it has been found that macromolecular samples such as ribosomes and R17 virions are severely damaged during standard electron microscopic operations: from 15 to 40% of the mass of the sample may be lost in a 30 sec exposure to the beam.     

Electron microscopy of cholesterol

Negative staining and platinum-carbon shadowing have been used to prepare electron microscope specimens from aqueous colloidal suspensions of cholesterol microscrystals and from crystalline suspensions in methanol and ethanol. Microcrystals prepared by injection of alcoholic solutions of cholesterol into water exhibit angular conformations of varying regularity which contain a number of parallel cholesterol bilayers. The electron optical images of the cholesterol microcrystals, oriented horizontally and ‘on-edge’, obtained by both negative staining and metal shadowing, are in good agreement. Metal shadowing does, however, reveal greater detail within microcrystal clusters than does negative staining, as well as of the bilayer steps at microcrystal edges. The needle-like crystals (from methanol) and plate-like crystals (from ethanol) present considerable difficulties for the negative staining technique, because of their thickness and the consequent depth of the surrounding negative stain. Small crystals are, nevertheless, shown to possess multiple cholesterol bilayers. Platinum-carbon shadowing of cholesterol crystals taken directly from methanol and ethanol provides more satisfactory images than negative staining. The large depth of focus of the transmission electron microscope enables the stacked cholesterol bilayers to be clearly defined at the edges of crystals. The results obtained are discussed in relation to the physicochemical and biological properties of cholesterol, which underlie the fundamental difficulty encountered when fixing and staining cholesterol for thin sectioning, and also the role of cholesterol insolubility in the formation of gallstones.     

Electron tomography of viruses

Understanding the molecular architectures of enveloped and complex viruses is a challenging frontier in structural biology. In these viruses, the structural and compositional variation from one viral particle to another generally precludes the use of either crystallization or image averaging procedures that have been successfully implemented in the past for highly symmetric viruses. While advances in cryo electron tomography of unstained specimens provide new opportunities for identification and molecular averaging of individual subcomponents such as the surface glycoprotein spikes on purified viruses, electron tomography of stained and plunge-frozen cells is being used to visualize the cellular context of viral entry and replication. Here, we review recent developments in both areas as they relate to our understanding of the biology of heterogeneous and pleiomorphic viruses.     

Electron transfer in biology

Table VII presents a list of the topics I have discussed. Underlying biological electron transfer which employs metal ions overwhelmingly is the intimacy of the interaction between metal ion properties and protein properties. Attacking the problems is attacking a cornerstone of life - bioenergetics. It is appropriate that this is the Heyrovsky Memorial Lecture since he devised the polarograph which is a device for coupling electrolytes (protons) in solution with electrons in metal atoms.     

Electron microscope study ofLeptospira

A leptospira consists of the cytoplasmic cylinder (contained within its own wall) which winds helically anticlockwise around the axistyle; both these components are enclosed by the outer covering membrane enveloping the whole organism.