Compositional imaging with electron energy loss spectroscopy (EELS) can be performed in both the energy-filtering transmission electron microscope (EFTEM) and in the scanning transmission electron microscope (STEM). Quantitative elemental distributions are obtained from core-edges produced by inner-shell excitations, although more detailed information about chemical bonding and electronic structure is also available from the fine structure associated with valence electron excitations. The fixed-beam EFTEM can provide data from large numbers of pixels very rapidly and offers an advantage for analysis of extended specimen regions containing relatively high atomic concentrations. Acquisition of entire spectra at each pixel in the field-emission STEM (spectrum-imaging technique) provides improved flexibility and accuracy despite the longer recording times. Spectrum-imaging allows post facto data processing with parameters that can be varied after acquisition is completed. In suitably thin specimens, EELS compositional mapping can provide a sensitivity of a few atoms for certain elements.

Dynamic charging is the condition in which the magnitude of the charge on the surface of a non-conductor, such as a polymer, under electron irradiation appears to change with the incident current density. Measurements confirm that this effect does occur and show that the magnitude of the effect depends on the relationship between the incident beam energy and the energy (E2) at which the sample has a total electron yield of unity. The effect arises from changes in the leakage resistance of the sample, and two possible explanations for this behavior are suggested.

Ethanol (ethyl alcohol) has long been a standard reagent used in preparing tissues for light and electron microscopy. After fixation, tissues are usually dehydrated with ethanol before being embedded in paraffin or plastic. In this study we show that the ethanol-infiltrated tissue can be frozen and sectioned directly, without embedding. When tissue impregnated with ethanol is cooled below about -117¡C with liquid nitrogen, the ethanol solidifies without appreciable crystallization. The frozen tissue can then be sectioned in a commercial cryoultramicrotome that is set at -155 to -170°C to produce semithin frozen sections (0.5-3 μm thick) for light microscopy, or ultrathin frozen sections (50-100 nm thick) for electron microscopy. Sections are picked up and mounted on glass slides or EM grids by means that are in current use for ice ultrathin frozen sectioning. Since there is no apparent freezing damage, the morphology in these ethanol frozen sections of unembedded tissue appears generally quite good, often resembling that obtained by conventional EM techniques. Examples are provided that illustrate the use of this material for immunocytochemistry at the light and electron microscope levels.

We present a novel combined atomic force microscope (AFM) and confocal laser scanning microscope (CLSM). The sample is supported from the side by a boom from a piezo tube scanner, allowing top and bottom access to the sample. The sample is scanned above an inverted microscope objective with a fixed optical path for fluorescent CLSM imaging. An AFM positioned directly above the sample simultaneously measures surface topography. The piezo tube scanner is angled, placing its center of scan curvature directly above the microscope objective. This geometry allows flat scans up to 300 μm on a side. Because the sample is scanned, the AFM and CLSM images are acquired in direct registration.

Four leaf preparation techniques (air drying, tetramethylsilane air drying, critical point drying, and freeze substitution) used in scanning electron microscopy (SEM) were evaluated with respect to the degree of cellular distortion they produce in stomatal guard cells of leaves of Dactylis glomerata and Elymus canadensis. Surface morphological distortion and cuticle disruption in the air-dried and tetramethylsilane air-dried leaves, and cuticle disruption within the critical point-dried tissue made it difficult to obtain measurements.The freeze-substituted tissue experienced little cuticle disturbance, and the cellular morphology appeared normal. The length of the guard cells did not significantly differ between the air-dried, tetramethylsilane air-dried, critical point-dried, or freeze-substituted samples. Widths did significantly vary, with the freeze-substituted tissue having lower values than tissues treated with the other treatments. Freeze substitution methodology produced SEM images that appear to be less distorted and allow easy and precise measurement.

An improved, jet electropolishing technique for the production of germanium TEM specimens has been developed. The use of slightly modified, commercially available equipment permits rapid, reliable thinning of foils exhibiting large electron transparent regions. The use of automatic termination of polishing produces cleaner foil surfaces than incremental final polishing did previously.