Some comments on models for field enhancement.

To estimate the apex field-enhancement factor gamma(a)associated with a pointed protrusion on a flat planar surface, the simple physical models of a 'floating sphere at emitter-plane potential' and a 'hemisphere on a post' are often discussed. The corresponding mathematical expressions have the form: gamma(a)=m+h/rho, where rho is the sphere or hemisphere radius, h is its 'height above the emitter plane', and m is a constant variously taken as 0, 2 or 3. Recent numerical simulations for the 'hemisphere on a post' model, reported elsewhere by two of us (CJE and GV) and by Kokkaris, Modinos and Xanthakis, have shown that all of these simple formulae significantly overpredict gamma(a) if h/rho is large. This article first reexamines the basis of these simple formulae and confirms that they are less secure than is sometimes thought. The formulae reported elsewhere as fits to the numerical results are then quoted and compared with the simple formulae, and with the known exact analytical result for the 'hemi-ellipsoid on a plane' model. Discrepancies can be rationalised. Some general conclusions are drawn.

Field Emission and Electron Microscopy.

An overview and new results are presented of the investigations carried out in the last 5 years on nano-sized tips by means of electron microscopy, an electron optical bench, and computation. Tungsten and, in particular, carbon nano-tips prepared by carbon contamination in a scanning electron microscope, were studied for applications as field-emission electron sources. Several features of their use are described and the results concerning the determination of some of their basic properties are reported.

Lipid oxidation deletes the nanodomain organization of artificial membranes.

Nanoscopic domains with different crystal structures have been induced in closed artificial membranes and have been directly imaged by atomic force microscopy at a spatial resolution better than 0.3 nm. These observations provide experimental evidence to the hydrophobic mismatching theory of lateral phase separation phenomena. Under oxidant conditions, the lipid-lipid assembly reorganizes into a new steady-state structure with disappearance of specific nanodomains. This finding may contribute to understanding the mechanism of peroxidative damage to membrane properties. In fact, alterations of specific modes of molecular conformation and packing may lead to perturbation of specific properties.

Hyaluronic acid by atomic force microscopy.

Hyaluronic acid (HA) of different molecular weights has been examined by atomic force microscopy (AFM) in air. This technique allows 3-D surface images of soft samples without any pretreatment, such as shadowing or staining. In the present study we examined the supermolecular organization of HA chains when deposited on mica and graphite, to better understand the interchain and intrachain interactions of HA molecules in solution. The concentration of the solution deposited varied from 0.001 to 1 mg/ml. On both substrates, and independent of the concentration, high-molecular-mass HA formed networks in which molecules ran parallel for hundreds of nanometers, giving rise to flat sheets and tubular structures that separate and rejoin into similar neighboring aggregates. Accurate measurements of the thickness of the thinnest sheets were consistent with a monolayer of HA molecules, 0.3 nm thick, strongly indicating lateral aggregation forces between chains as well as rather strong hydrophilic interactions between mica and HA. The results agree with an existing model of HA tertiary structure in solution in which the network is stabilized by both hydrophilic and hydrophobic interactions. Our images support this model and indicate that hydrophobic interactions between chains may exert a pivotal role in aqueous solution.

Study of elastic fiber organization by scanning force microscopy.

Elastic fibers of beef ligamentum nuchae were observed by atomic force microscopy and data compared with those obtained by conventional and freeze-fracture electron microscopy. Fresh isolated elastin fibers as well as thin sections of ligament fragments, which were fixed and embedded either in relaxed or in stretched conditions, were analysed. The results confirm that, at least in beef ligamentum nuchae, elastic fibers consist of beaded filaments which can be oriented by stretching in the direction of the force applied. Moreover, atomic force microscopy revealed that these beaded filaments are laterally connected by periodical bridges which become more pronounced upon stretching. The data clearly show that elastin molecules are organized in a rather ordered array, at least at the super-molecular level, and a depiction of the elastin organization in beef ligamentum nuchae is attempted.

Atomic force microscopy observations of acyl chains in phospholipids.

The potential use of atomic force microscopy (AFM) to image the mode of assembly and to measure the corresponding lattice parameters of model systems consisting of ordered aggregates of cardiolipin molecules has been investigated. An unprecedented resolution of about 0.2 nm has been achieved on suitably prepared specimens. This enables the orientational order and the positional correlations of the individual molecules in the lattice to be defined, and submolecular details, such as the acyl chains and the polar groups, to be imaged. The structural parameters derived from AFM have been compared with those obtained by transmission electron diffraction of the same specimen and found to be in excellent agreement. AFM turns out to be a powerful and probably a unique tool to reveal local phase variations in systems, such as biological membranes, that have non-homogeneous composition and organization.

Control of specimen orientation and environment.

Application of electron microscopy in a wide variety of fields of investigation has placed ever-expanding demands on the various components of the instrument. In situ specimen manipulation is one such demand and can often be critical to the success of an experiment. Control of specimen orientation is the most common manipulation, but control of a variety of other physical and chemical parameters may also be important. Temperature, gaseous and/or liquid environment, and mechanical operations are examples. Control and variation of these parameters in a small device (occupying a few cm3) operated in a strong magnetic field inside a vacuum system is often a considerable challenge. This must also be done at extreme stability: at least as good as the resolution limit of the microscope. Optimization of stage performance is too often sacrificed for optical performance or vice versa. The next generation of objective lenses and specimen stages are being designed in concert: an approach which should lead to an improved in situ laboratory, observed with optimum optics.

A method for the improvement of the visibility of transmission electron microscope images.

A method is presented of improving the visibility of transmission electron microscope images in any situation in which a high resolution in only one chosen direction is of interest. The technique is based on the use of slot shaped objective apertures. Such apertures are of reduced area relative to a circular aperture giving the same all round resolution. The background intensity due to inelastically scattered electrons is thus reduced. The aperture device developed is described, while the value of the method is demonstrated by its application to the observation of dislocations. Further possible applications are indicated.