Micro-computed tomography with the highly intense, monochromatic X rays produced by the synchrotron is a superior method to nondestructively measure the local absorption in three-dimensional space. Because biological tissues and cells consist mainly of water as the surrounding medium, higher absorbing agents have to be incorporated into the structures of interest. Even without X-ray optics such as refractive lens, one can uncover the stain distribution with the spatial resolution of about 1 μm. Incorporating the stain at selected cell compartments, for example, binding to the RNA/DNA, their density distribution becomes quantified. In this communication, we demonstrate that tomograms obtained at the beamlines BW2 and W2 (HASYLAB at DESY, Hamburg, Germany) and 4S (SLS, Villigen, Switzerland) clearly show that the RNA/DNA-stained HEK 293 cell clusters have a core of high density and a peripheral part of lower density, which correlate with results of optical microscopy. The inner part of the clusters is associated with nonvital cells as the result of insufficient oxygen and nutrition supply. This necrotic part is surrounded by (6 ± 1) layers of vital cells.

We have implemented and tested a new automatic method for the montage synthesis and three-dimensional (3D) reconstruction of large tissue volumes from confocal laser scanning microscopy data (CLSM). This method relies on maximization of the phase correlation between adjacent images. It was tested on a large specimen (a murine heart) that was cut into a number of individual sections with thickness appropriate for CLSM. The sections were scanned horizontally (in-plane) and vertically (perpendicular to the optical planes) to produce “tiles” of a 3D volume. Phase correlation maximization was applied to the montage synthesis of in-plane tiles and 3D alignment of optical slices within a given physical section. The performance of the new method is evaluated.

Performing reflection-mode (backscatter-mode) confocal microscopy on cells growing on reflective substrates gives images that have improved contrast and are more easily interpreted than standard reflection-mode confocal micrographs (Keith et al., 1998). However, a number of factors degrade the quality of images taken with the highest-resolution microscope objectives in this technique. We here describe modifications to reflection-enhanced backscatter confocal microscopy that (partially) overcome these factors. With these modifications of the technique, it is possible to visualize structures the size—and refractility—of individual microtubules in intact cells. Additionally, we demonstrate that this technique, in common with fluorescence techniques such as standing wave widefield fluorescence microscopy and 4-Pi confocal microscopy, offers improved resolution in the Z-direction.

Electron energy loss spectroscopy (EELS), X-ray photoelectron spectroscopy (XPS), and transmission electron microscopy have been used to study iron catalysts for Fischer–Tropsch synthesis. When silica-containing iron oxide precursors are activated in flowing CO, the iron phase segregates into iron carbide crystallites, leaving behind some unreduced iron oxide in an amorphous state coexisting with the silica binder. The iron carbide crystallites are found covered by characteristic amorphous carbonaceous surface layers. These amorphous species are difficult to analyze by traditional catalyst characterization techniques, which lack spatial resolution. Even a surface-sensitive technique such as XPS shows only broad carbon or iron peaks in these catalysts. As we show in this work, EELS allows us to distinguish three different carbonaceous species: reactive amorphous carbon, graphitic carbon, and carbidic carbon in the bulk of the iron carbide particles. The carbidic carbon K edge shows an intense “π*” peak with an edge shift of about 1 eV to higher energy loss compared to that of the π* of amorphous carbon film or graphitic carbon. EELS analysis of the oxygen K edge allows us to distinguish the amorphous unreduced iron phase from the silica binder, indicating these are two separate phases. These results shed light onto the complex phase transformations that accompany the activation of iron catalysts for Fischer–Tropsch synthesis.

A fixed-bed reactor has been designed and constructed for ex situ transmission electron microscopy (TEM) studies of heterogeneous catalysts. The ex situ facility exposes a fully prepared TEM sample on a grid to actual process conditions (e.g., temperature, pressure, gas composition, etc.) by placing the grid at the exit section of a conventional fixed-bed reactor. A unique reactor design allows grid transfer into the electron microscope and back into the reactor again under a controlled (inert) environment, thus allowing time-resolved monitoring of catalyst morphology changes under realistic, well-controlled conditions. This facility stands completely independent of the TEM. Thus, no special TEM modifications are required and long-term ex situ studies do not impact microscope utilization. The utility of the facility is demonstrated via the oxidation of intermediate size (∼20–∼80 nm) supported copper particles.

It has been shown, by imaging gold (200) planes, that it is possible to achieve better than 0.20-nm structural resolution in cryo-transmission electron microscopy (cryo-TEM). This has been done using commercially available cryo equipment and using a 300-kV field emission gun (FEG) TEM. The images of 15-nm gold particles embedded in amorphous frozen water clearly show the (111) planes (separated by 0.235 nm) in gold. Fourier transform demonstrates the presence of (200) planes in the image, proving a resolution of better than 0.20 nm. The experimental results are supported by image simulations using the multislice method. These simulations suggest that it should be possible to achieve the same resolution even in smaller particles and particles of lighter elements. The crucial experimental problem to overcome is keeping the thickness of the amorphous film low and to work at low electron dose conditions.

A combination of transmission electron microscopy (TEM) and in situ tensile testing in an environmental scanning electron microscopy (ESEM) was used to evaluate the static bulk and dynamic surface morphologies of medical polyurethanes. TEM results showed phase-separated hard segment and soft segment structures. Surface morphology as a function of strain was studied using ESEM in conjunction with a tensometer.

Ultramicrotomy, the technique of cutting nanometers-thin slices of material using a diamond knife, was applied to prepare transmission electron microscope (TEM) specimens of nanoporous poly(methylsilsesquioxane) (PMSSQ) thin films. This technique was compared to focused ion beam (FIB) cross-section preparation to address possible artifacts resulting from deformation of nanoporous microstructure during the sample preparation. It was found that ultramicrotomy is a successful TEM specimen preparation method for nanoporous PMSSQ thin films when combined with low-energy ion milling as a final step. A thick, sacrificial carbon coating was identified as a method of reducing defects from the FIB process which included film shrinkage and pore deformation.

Amorphous 1–2-nm-wide intergranular films in ceramics dictate many of their properties. The detailed investigation of structure and chemistry of these films pushes the limits of today's transmission electron microscopy. We report on the reconstruction of the one-dimensional potential profile across the film from an experimentally acquired tilt series of energy-filtered electron diffraction patterns. Along with the potential profile, the specimen thickness, film orientation with respect to the grain lattice and specimen surface, and the absolute specimen orientation with respect to the laboratory frame of reference are retrieved.

Scanning electron microscopy (SEM) is widely used in the science of materials and different parameters were developed to characterize the surface roughness. In a previous work, we studied the surface topography with fractal dimension at low scale and two parameters at high scale by using the variogram, that is, variance vs. step log–log graph, of a SEM image. Those studies were carried out with the FERImage program, previously developed by us. To verify the previously accepted hypothesis by working with only an image, it is indispensable to have reliable three-dimensional (3D) surface data. In this work, a new program (EZEImage) to characterize 3D surface topography in SEM has been developed. It uses fast cross correlation and dynamic programming to obtain reliable dense height maps in a few seconds which can be displayed as an image where each gray level represents a height value. This image can be used for the FERImage program or any other software to obtain surface topography characteristics. EZEImage also generates anaglyph images as well as characterizes 3D surface topography by means of a parameter set to describe amplitude properties and three functional indices for characterizing bearing and fluid properties.

Scanning electron microscopy (SEM) is widely used in surface studies and continuous efforts are carried out in the search of estimators of different surface characteristics. By using the variogram, we developed two of these estimators that were used to characterize the surface roughness from the SEM image texture. One of the estimators is related to the crossover between fractal region at low scale and the periodic region at high scale, whereas the other estimator characterizes the periodic region. In this work, a full study of these estimators and the fractal dimension in two dimensions (2D) and three dimensions (3D) was carried out for emery papers. We show that the obtained fractal dimension with only one image is good enough to characterize the roughness surface because its behavior is similar to those obtained with 3D height data. We show also that the estimator that indicates the crossover is related to the minimum cell size in 2D and to the average particle size in 3D. The other estimator has different values for the three studied emery papers in 2D but it does not have a clear meaning, and these values are similar for those studied samples in 3D. Nevertheless, it indicates the formation tendency of compound cells. The fractal dimension values from the variogram and from an area versus step log–log graph were studied with 3D data. Both methods yield different values corresponding to different information from the samples.

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