The development and morphology of the immature phases of Pachycrepoideus vindemmiae (Rondani, 1875) (Hymenoptera, Pteromalidae) are described from a laboratory rearing culture maintained on Ceratitis capitata (Wiedemann, 1824) (Diptera, Tephritidae) using microscopic techniques, including light and scanning electron microscopy. The surface of the chorion of the egg is granulated, and the micropyle occurs at the anterior end. The labrum of the first instar larva does not have sensilla, and the second to fourth instar larvae have setae on the head. The mature larva is characterized by the position and number of the integumental differentations (sensilla and setae). On completion of larval development, an adecticous and exarate pupa is produced. As for the adult, the mandibles of the pupae are toothed. Five larval instars are recorded, based on statistical analyses of the sizes of the larval mandibles in combination with characters such as the number of exuviae and excretion of the meconium. Developmental time from egg to adult emergence was ∼18–20 days for males and ∼21–23 days for females at 21–26°C, 55–85 relative humidity, and a 16L:8D photoperiod. The results show that the eggs and different larval instars of this parasitoid can be unambiguously identified only by scanning electron microscope.

Electron microscopy of enclosed liquid samples requires the thinnest possible membranes as enclosing windows as well as nanoscale liquid sample thickness to achieve the best possible resolution. Today liquid sample systems for transmission electron microscopy (TEM) are typically made from two sandwiched microchips with thin membranes. We report on a new microfabricated chip system based on a monolithic design that enables membrane geometry on the scale of a few micrometers. The design is intended to reduce membrane deflection when the system is under pressure, a microfluidic channel for improved flow geometry, and a better space angle for auxiliary detectors such as energy-dispersive X-ray spectroscopy. We explain the system design and fabrication and show the first successful TEM images of liquid samples in the chips.

The measurement of absolute crystal polarity is crucial to understanding the structural properties of many planar defects in compound semiconductors. Grain boundaries, including twin boundaries, in the sphalerite lattice are uniquely characterized by the crystallographic misorientation of individual grains and the direction of the crystal polarity in domains adjoining the grain boundary. To evaluate crystal polarity in gallium phosphide (GaP), asymmetrical interference contrast in convergent-beam electron-diffraction (CBED) patterns was used to ascertain the nature and direction of polar bonds. The direction of the asymmetry in the electron diffraction reflections was correlated with the crystal polarity of a sample with known polarity. The CBED technique was applied to determine the polar orientation of grains adjoining Σ = 3 coherent and lateral twin boundaries in polycrystalline GaP.

The recent advent of a novel design of in situ heating technology for electron microscopes has permitted unprecedented control of elevated temperature studies of catalytic materials, particularly when coupled with the sub-Ångström imaging performance of a modern aberration-corrected scanning transmission electron microscope (STEM). Using micro-electro-mechanical-systems (MEMS)-based Aduro™ heating chips from Protochips, Inc. (Raleigh, NC, USA) allows nearly instantaneous heating and cooling of catalyst powders, avoiding effects of temperature ramping as experienced with standard heating stages. The heating technology also provides stable operation limited only by the inherent drift in the microscope stage, thus allowing full image resolution to be achieved even at elevated temperatures. The present study details the use of both the high X-Y spatial resolution in both dark-field and simultaneous bright-field imaging, along with the high resolution in Z (depth sectioning) provided by the large probe incidence semiangle in the aberration-corrected instrument, to characterize the evolution of microstructure in a commercial Au/Fe2O3 water-gas shift catalyst during elevated temperature treatment. The phenomenon of Au diffusion to the surface of hematite support particles to form discrete crystalline Au nanoparticles in the 1–2 nm size range, after a prior leaching treatment to remove surface Au species has been characterized.

Atom probe tomography (APT) is a powerful technique to characterize buried three-dimensional nanostructures in a variety of materials. Accurate characterization of those nanometer-scale clusters and precipitates is of great scientific significance to understand the structure–property relationships and the microstructural evolution. The current widely used cluster analysis method, a variant of the density-based spatial clustering of applications with noise algorithm, can only accurately extract clusters of the same atomic density, neglecting several experimental realities, such as density variations within and between clusters and the nonuniformity of the atomic density in the APT reconstruction itself (e.g., crystallographic poles and other field evaporation artifacts). This clustering method relies heavily on multiple input parameters, but ideal selection of those parameters is challenging and oftentimes ambiguous. In this study, we utilize a well-known cluster analysis algorithm, called ordering points to identify the clustering structures, and an automatic cluster extraction algorithm to analyze clusters of varying atomic density in APT data. This approach requires only one free parameter, and other inputs can be estimated or bounded based on physical parameters, such as the lattice parameter and solute concentration. The effectiveness of this method is demonstrated by application to several small-scale model datasets and a real APT dataset obtained from an oxide-dispersion strengthened ferritic alloy specimen.

Wedge polishing was used to prepare one-dimensional Si n-p junction and Si p-channel metal-oxide-silicon field effect transistor (pMOSFET) samples for precise and quantitative electrostatic potential analysis using off-axis electron holography. To avoid artifacts associated with ion milling, cloth polishing with 0.02-μm colloidal silica suspension was used for final thinning. Uniform thickness and no significant charging were observed by electron holography analysis for samples prepared entirely by this method. The effect of sample thickness was investigated and the minimum thickness for reliable results was found to be ∼160 nm. Below this thickness, measured phase changes were smaller than expected. For the pMOSFET sample, quantitative analysis of two-dimensional electrostatic potential distribution showed that the metallurgical gate length (separation between two extension junctions) was ∼54 nm, whereas the actual gate length was measured to be ∼70 nm by conventional transmission electron microscopy. Thus, source and drain junction encroachment under the gate was 16 nm.

Solid particles in snow deposits, sampled in mining and Pb-processing area of Žerjav, Slovenia, have been investigated using scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM/EDS). Identified particles were classified as geogenic–anthropogenic, anthropogenic, and secondary weathering products. Geogenic–anthropogenic particles were represented by scarce Zn- and Pb-bearing ore minerals, originating from mine waste deposit. The most important anthropogenic metal-bearing particles in snow were Pb-, Sb- and Sn-bearing oxides and sulphides. The morphology of these particles showed that they formed at temperatures above their melting points. They were most abundant in snow sampled closest to the Pb-processing plant and least abundant in snow taken farthest from the plant, thus indicating that Pb processing was their predominant source between the last snowfall and the time of sampling. SEM/EDS analysis showed that Sb and Sn contents in these anthropogenic phases were higher and more variable than in natural Pb-bearing ore minerals. The most important secondary weathering products were Pb- and Zn-containing Fe-oxy-hydroxides whose elemental composition and morphology indicated that they mostly resulted from oxidation of metal-bearing sulphides emitted from the Pb-processing plant. This study demonstrated the importance of single particle analysis using SEM/EDS for differentiation between various sources of metals in the environment.

Abstract: Quantitative X-ray mapping in the analytical electron microscope (AEM) could improve the statistics of grain-boundary segregation measurements if high spatial resolution can be maintained at lower magnifications (<500 kX). Typically, only about 10 boundaries are analyzed because of the difficulty of conventional AEM measurements; however, a low-magnification quantitative X-ray map could contain twice this number of boundaries in a single field of view. Microscope conditions and mapping parameters have been explored for operation at ∼250 kX, under a variety of conditions to illustrate the trade-offs between various characteristics, such as analytical resolution, counting statistics, magnification, and acquisition time. From these data, it is possible to extrapolate to maps generated under different conditions and estimate their limitations with respect to these characteristics. A simple model has been developed to describe the behavior of inclined grain boundaries that can be used to estimate the detectability of segregant as a function of boundary tilt. Using quantitative X-ray maps, grain boundary Cu coverage has been measured from 55 boundaries in Al–4 wt.% Cu with minimal user effort. For fine-grained thin films, mapping is substantially more efficient than other methods of data acquisition and may be used to measure segregation at large numbers of boundaries.

Extended abstract of a paper presented at Microscopy and Microanalysis 2008 in Albuquerque, New Mexico, USA, August 3 – August 7, 2008

Fiber alignment plays a critical role in the structure and function of cells and tissues. While fiber alignment quantification is important to experimental analysis and several different methods for quantifying fiber alignment exist, many studies focus on qualitative rather than quantitative analysis perhaps due to the complexity of current fiber alignment methods. Speed and sensitivity were compared in edge detection and fast Fourier transform (FFT) for measuring actin fiber alignment in cells exposed to shear stress. While edge detection using matrix multiplication was consistently more sensitive than FFT, image processing time was significantly longer. However, when MATLAB functions were used to implement edge detection, MATLAB's efficient element-by-element calculations and fast filtering techniques reduced computation cost 100 times compared to the matrix multiplication edge detection method. The new computation time was comparable to the FFT method, and MATLAB edge detection produced well-distributed fiber angle distributions that statistically distinguished aligned and unaligned fibers in half as many sample images. When the FFT sensitivity was improved by dividing images into smaller subsections, processing time grew larger than the time required for MATLAB edge detection. Implementation of edge detection in MATLAB is simpler, faster, and more sensitive than FFT for fiber alignment quantification.

We describe here real-time, in situ observations of the formation of nanosize germanium (Ge) islands on silicon (Si). The deposition of Ge onto electron-transparent Si(100) takes place in a UHV transmission electron microscope that has been modified to allow chemical vapor deposition to be carried out in the polepiece. We recorded the growth process at video rate and were therefore able to follow the evolution of individual islands. As the islands grew, we observed a coarsening process similar to classical Ostwald ripening, but which leads at certain times to a bimodal distribution of island sizes. We show that this phenomenon can be understood using a model in which a conventional coarsening process is modified by a transition between two different island shapes.

Electron microprobe analysis of U, Th, and Pb in naturally occurring minerals can indicate their age. Where the Pb is entirely due to the radioactive decay of U and Th, the time since mineral formation or equilibration can be calculated. Uraninite (UO2), monazite (REE PO4), zircon (ZrSiO4), and xenotime (YPO4) have been used, the latter containing U and/or Th in minor proportions. Any stable U- or Th-bearing phase can be considered. Careful analysis is required with attention to interferences, background measurement, detection limits, and Pb-free sample preparation. Extended counting times (600 s) at a probe current >200 nA are recommended. Ages can be determined from uraninite older than 2 Ma for a Pb detection limit of 0.02% and up to 700−1,000 Ma, after which Pb can be lost from the structure. The youngest monazite ages permitted by the Pb detection limit are 50–100 Ma and ages greater than 3,000 Ma have been determined. The method does not provide the detail of isotopic methods, but results can be obtained more readily. Examples show dating of cheralite ((Ca,Ce)(Th,Ce)(PO4)2), a rock containing primary and secondary UO2, and a suite of detrital uraninite grains that formed a part of a mineral exploration program.

The charge-coupled devices used in electron microscopy are coated with a scintillating crystal that gives rise to a severe modulation transfer function (MTF). Exact knowledge of the MTF is imperative for a good correspondence between image simulation and experiment. We present a practical method to measure the MTF above the Nyquist frequency from the beam blocker's shadow image. The image processing has been fully automated and the program is made public. The method is successfully tested on three cameras with various beam blocker shapes.

Theories have been put forward on the etiology of sialoliths; however, a comprehensive understanding of their growth mechanisms is lacking. In an attempt to fill this gap, the current study has evaluated the internal architecture and growth patterns of a set of 30 independent specimens of sialoliths characterized at different scales by computed microtomography and electron microscopy. Tomography reconstructions showed cores in most of the sialoliths. The cores were surrounded by concentric or irregular patterns with variable degrees of mineralization. Regardless of the patterns, at finer scales the sialoliths consisted of banded and globular structures. The distribution of precipitates in the banded structures is compatible with a Liesegang–Ostwald phenomenon. On the other hand, the globular structures appear to arise from surface tension effects and to develop self-similar features as a result of a viscous fingering process. Electron diffraction patterns demonstrated that Ca- and P-based electrolytes crystallize in a structure close to that of hydroxyapatite. The organic matter contained sulfur with apparent origin from sulfated components of secretory material. These results cast new light on the mechanisms involved in the formation of sialoliths.

The molecular distribution in nanocolloids of poly(dimethyl siloxane) (PDMS) and an organic copolymer (methyl acrylate co-methyl methacrylate co-vinyl acetate) preserved in a frozen aqueous solution was investigated using cryovalence electron energy-loss spectroscopy (EELS) coupled with a scanning transmission electron microscope. Low energy-loss spectra depend upon valence electron structure, and we show that they are substantially different for the PDMS, the copolymer, and the vitrified water studied here. Combining a high efficiency detection system and the use of high-signal low-loss spectra in EELS, we achieved a spatial resolution of 8 nm without serious beam-induced specimen damage in this radiation-sensitive soft-materials system. To obtain quantitative phase maps of silicone and copolymer composition within individual nanoparticles, spectrum datasets were processed via multiple least squares fitting. Quantitative line profiles from the resulting compositional maps indicate that the PDMS lobe of biphasic nanoparticles contained a significant amount of the copolymer and a diffuse interface was formed. Since the nanoparticle synthesis involves polymerization of acrylate monomer dissolved in PDMS nanoparticle precursors, these results suggest that the evolution of the nanocolloid morphology during synthesis is kinetically frozen as the acrylate copolymer achieves some critical molecular weight.

Radiation induced clustering affects the mechanical properties, that is the ductile to brittle transition temperature (DBTT), of reactor pressure vessel (RPV) steel of nuclear power plants. The combination of low Cu and high Ni used in some RPV welds is known to further enhance the DBTT shift during long time operation. In this study, RPV weld samples containing 0.04 at% Cu and 1.6 at% Ni were irradiated to 2.0 and 6.4×1023 n/m2 in the Halden test reactor. Atom probe tomography (APT) was applied to study clustering of Ni, Mn, Si, and Cu. As the clusters are in the nanometer-range, APT is a very suitable technique for this type of study. From APT analyses information about size distribution, number density, and composition of the clusters can be obtained. However, the quantification of these attributes is not trivial. The maximum separation method (MSM) has been used to characterize the clusters and a detailed study about the influence of the choice of MSM cluster parameters, primarily on the cluster number density, has been undertaken.

The local electrode atom probe (LEAP) has become the primary instrument used for atom probe tomography measurements. Recent advances in detector and laser design, together with updated hit detection algorithms, have been incorporated into the latest LEAP 5000 instrument, but the implications of these changes on measurements, particularly the size and chemistry of small clusters and elemental segregations, have not been explored. In this study, we compare data sets from a variety of materials with small-scale chemical heterogeneity using both a LEAP 3000 instrument with 37% detector efficiency and a 532-nm green laser and a new LEAP 5000 instrument with a manufacturer estimated increase to 52% detector efficiency, and a 355-nm ultraviolet laser. In general, it was found that the number of atoms within small clusters or surface segregation increased in the LEAP 5000, as would be expected by the reported increase in detector efficiency from the LEAP 3000 architecture, but subtle differences in chemistry were observed which are attributed to changes in the way multiple hit detection is calculated using the LEAP 5000.

Reservosomes are endocytic compartments found in the posterior region of epimastigotes of Trypanosoma cruzi. In the differentiation from trypomastigotes to epimastigotes (reverse metacyclogenesis in vitro), one has the rare opportunity of following the biogenesis of an endocytic compartment. Metacyclic trypomastigotes incubated in LIT medium highly enriched with fetal calf serum differentiated directly to epimastigotes. In recently differentiated epimastigotes, acidic organelles were found in round compartments spread along the cell body, whereas in control epimastigotes they were found in reservosomes located in the posterior region. Ultrastructural analysis of intermediate forms showed that the cytostome and reservosomes appeared before differentiation to epimastigotes was completed. Many polymorphic reservosomes, with or without lipid inclusions, were observed from the anterior portion of the cell body, in close relationship with the Golgi complex, to the posterior region. Endocytic tracers were observed in the cytostome, flagellar pocket, vesicles, and newly formed reservosomes. Cruzipain, the main protease of T. cruzi, was localized in newly formed reservosomes and in vesicles budding from the trans-Golgi network that seem to fuse with reservosomes. Ingested gold-labeled albumin and cruzipain colocalized in recently formed reservosomes. Endocytosis and immunocytochemical analysis suggested that the endocytic and the secretory pathways may contribute to reservosome formation.

The resolution achieved in low-dose electron microscopy of biological macromolecules is significantly worse than what can be obtained on the same microscopes with more robust specimens. When two-dimensional crystals are used, it is also apparent that the high-resolution image contrast is much less than what it could be if the images were perfect. Because specimen charging is one factor that might limit the contrast and resolution achieved with biological specimens, we have investigated the use of holey support films that have been coated with a metallic film before depositing specimens onto a thin carbon film that is suspended over the holes. Monolayer crystals of paraffin (C44H90) are used as a test specimen for this work because of the relative ease in imaging Bragg spacings at ∼0.4 nm resolution, the relative ease of measuring the contrast in these images, and the similar degree of radiation sensitivity of these crystals when compared to biological macromolecules. A metallic coating on the surrounding support film does, indeed, produce a significant improvement in the high-resolution contrast for a small fraction of the images. The majority of images show little obvious improvement, however, and even the coated area of the support film continues to show a significant amount of beam-induced movement under low-dose conditions. The fact that the contrast in the best images can be as much as 25%–35% of what it would be in a perfect image is nevertheless encouraging, demonstrating that it should be possible, in principle, to achieve the same performance for every image. Routine data collection of this quality would make it possible to determine the structure of large, macromolecular complexes without the need to grow crystals of these difficult specimen materials.