The resolution-limiting aberrations of round electromagnetic lenses can now be successfully overcome via the use of multipole element “aberration correctors.” The installation and performance of a hexapole-based corrector (CEOS GmbH) integrated on the probe-forming side of a JEOL 2200FS FEG STEM/TEM is described. For the resolution of the microscope not to be severely compromised by its environment, a new, specially designed building at Oak Ridge National Laboratory has been built. The Advanced Microscopy Laboratory was designed with the goal of providing a suitable location for aberration-corrected electron microscopes. Construction methods and performance of the building are discussed in the context of the performance of the microscope. Initial performance of the microscope on relevant specimens and modifications made to eliminate resolution-limiting conditions are also discussed.

We compare experimental fluctuation electron microscopy (FEM) speckle data with electron diffraction simulations for thin amorphous carbon and silicon samples. We find that the experimental speckle intensity variance is generally more than an order of magnitude lower than kinematical scattering theory predicts for spatially coherent illumination.

We hypothesize that decoherence, which randomizes the phase relationship between scattered waves, is responsible for the anomaly. Specifically, displacement decoherence can contribute strongly to speckle suppression, particularly at higher beam energies. Displacement decoherence arises when the local structure is rearranged significantly by interactions with the beam during the exposure. Such motions cause diffraction speckle to twinkle, some of it at observable time scales.

We also find that the continuous random network model of amorphous silicon can explain the experimental variance data if displacement decoherence and multiple scattering is included in the modeling. This may resolve the longstanding discrepancy between X-ray and electron diffraction studies of radial distribution functions, and conclusions reached from previous FEM studies.

Decoherence likely affects all quantitative electron imaging and diffraction studies. It likely contributes to the so-called Stobbs factor, where high-resolution atomic-column image intensities are anomalously lower than predicted by a similar factor to that observed here.

We have recently reported the synthesis of one-dimensional nanobelt structures of ZnO, SnO2, In2O3, CdO, Ga2O3, and PbO2 by evaporating the desired commercial metal oxide powders at high temperatures (Science (2001), 291, 1947). The as-synthesized oxide nanobelts are pure, structurally uniform, single crystalline, and most of them free from dislocations. The beltlike morphology appears to be a unique and common structural characteristic for the family of semiconducting oxides. In the present article, we focus on the twin and stacking fault planar defects found in oxide nanobelts and nanowires although they are rarely observed. Some interesting and unique growth morphologies are presented to illustrate the roles played by surface energy and kinetics in growth. It is shown that the surfaces of the oxide nanobelts are enclosed by the low-index, low-energy crystallographic facets. The growth morphology is largely dominated by the growth kinetics.

Sedimentary carbonate rocks are one of the principal porous structures in natural reservoirs of hydrocarbons such as crude oil and natural gas. Efficient hydrocarbon recovery requires an understanding of the carbonate pore structure, but the nature of sedimentary carbonate rock formation and the toughness of the material make proper analysis difficult. In this study, a novel preparation method was used on a dolomitic carbonate sample, and selected regions were then serially sectioned and imaged by focused ion beam–scanning electron microscopy. The resulting series of images were used to construct detailed three-dimensional representations of the microscopic pore spaces and analyze them quantitatively. We show for the first time the presence of nanometer-scale pores (50–300 nm) inside the solid dolomite matrix. We also show the degree of connectivity of these pores with micron-scale pores (2–5 μm) that were observed to further link with bulk pores outside the matrix.

Arterial physiology relies on a delicate three-dimensional (3D) organization of cells and extracellular matrix, which is remarkably altered by vascular diseases like abdominal aortic aneurysms (AAA). The ability to explore the micro-histology of the aorta wall is important in the study of vascular pathologies and in the development of vascular constitutive models, i.e., mathematical descriptions of biomechanical properties of the wall. The present study reports and validates a fast image processing sequence capable of quantifying collagen fiber organization from histological stains. Powering and re-normalizing the histogram of the classical fast Fourier transformation (FFT) is a key step in the proposed analysis sequence. This modification introduces a powering parameter w, which was calibrated to best fit the reference data obtained using classical FFT and polarized light microscopy (PLM) of stained histological slices of AAA wall samples. The values of w = 3 and 7 give the best correlation (Pearson's correlation coefficient larger than 0.7, R2 about 0.7) with the classical FFT approach and PLM measurements. A fast and operator independent method to identify collagen organization in the arterial wall was developed and validated. This overcomes severe limitations of currently applied methods like PLM to identify collagen organization in the arterial wall.

Correlative light and electron microscopy (CLEM) has been in use for several years, however it has remained a costly method with difficult sample preparation. Here, we report a series of technical improvements developed for precise and cost-effective correlative light and scanning electron microscopy (SEM) and focused ion beam (FIB)/SEM microscopy of single cells, as well as large tissue sections. Customized coordinate systems for both slides and coverslips were established for thin and ultra-thin embedding of a wide range of biological specimens. Immobilization of biological samples was examined with a variety of adhesives. For histological sections, a filter system for flat embedding was developed. We validated ultra-thin embedding on laser marked slides for efficient, high-resolution CLEM. Target cells can be re-located within minutes in SEM without protracted searching and correlative investigations were reduced to a minimum of preparation steps, while still reaching highest resolution. The FIB/SEM milling procedure is facilitated and significantly accelerated as: (i) milling a ramp becomes needless, (ii) significant re-deposition of milled material does not occur; and (iii) charging effects are markedly reduced. By optimizing all technical parameters FIB/SEM stacks with 2 nm iso-voxels were achieved over thousands of sections, in a wide range of biological samples.

Atomic-size vortex beams have great potential in probing the magnetic moment of materials at atomic scales. However, the limited depth of field of vortex beams constrains the probing depth in which the helical phase front is preserved. On the other hand, electron channeling in crystals can counteract beam divergence and extend the vortex beam without disrupting its topological charge. Specifically, in this article, we report that atomic vortex beams with topological charge ±1 can be coupled to the 2p columnar bound states and propagate for more than 50 nm without being dispersed and losing its helical phase front. We give numerical solutions to the 2p columnar orbitals and tabulate the characteristic size of the 2p states of two typical elements, Co and Dy, for various incident beam energies and various atomic densities. The tabulated numbers allow estimates of the optimal convergence angle for maximal coupling to 2p columnar orbital. We have also developed analytic formulae for beam energy, convergence angle, and hologram-dependent scaling for various characteristic sizes. These length scales are useful for the design of pitch-fork apertures and operations of microscopes in the vortex-beam imaging mode.

Specialized intermediate filaments (IFs) have critical importance for the clearness and uncommon transparency of vertebrate lens fiber cells, although the physical mechanisms involved are poorly understood. Recently, an unusual low-scattering light transport was also described in retinal Müller cells. Exploring the function of IFs in Müller cells, we have studied the morphology and distribution pattern of IFs and other cytoskeletal filaments inside the Müller cell main processes in the foveolar part of the avian (pied flycatcher) retina. We found that some IFs surrounded by globular nanoparticles (that we suggest are crystallines) are present in almost every part of the Müller cells that span the retina, including the microvilli. Unlike IFs implicated in the mechanical architecture of the cell, these IFs are not connected to any specific cellular membranes. Instead, they are organized into bundles, passing inside the cell from the endfeet to the photoreceptor, following the geometry of the processes, and repeatedly circumventing numerous obstacles. We believe that the presently reported data effectively confirm that the model of nanooptical channels built of the IFs may provide a viable explanation of Müller cell transparency.

We review our studies of the initial oxidation stages of Cu(001) thin films as investigated by in situ ultra-high vacuum transmission electron microscopy. We present our observations of surface reconstruction and the nucleation to coalescence of copper oxide during in situ oxidation in O2. We have proposed a semi-quantitative model, where oxygen surface diffusion is the dominant mechanism of the initial oxidation stages of Cu. We have also investigated the effect of water vapor on copper oxidation. We have observed that the presence of water vapor in the oxidizing atmosphere retards the rate of Cu oxidation and Cu2O is reduced when exposed directly to steam.

Agrowastes are produced worldwide in huge quantities and they contain interesting elements for producing inorganic cementing binders, especially silicon. Conversion of agrowastes into ash is an interesting way of yielding raw material used in the manufacture of low-CO2 binders. Silica-rich ashes are preferred for preparing inorganic binders. Sugarcane leaves (Saccharum officinarum, SL) and bamboo leaves (Bambusa vulgaris, BvL and Bambusa gigantea, BgL), and their corresponding ashes (SLA, BvLA, and BgLA), were chosen as case studies. These samples were analyzed by means of optical microscopy, Cryo-scanning electron microscopy (SEM), SEM, and field emission scanning electron microscopy. Spodograms were obtained for BvLA and BgLA, which have high proportions of silicon, but no spodogram was obtained for SLA because of the low silicon content. Different types of phytoliths (specific cells, reservoirs of silica in plants) in the studied leaves were observed. These phytoliths maintained their form after calcination at temperatures in the 350–850°C range. Owing to the chemical composition of these ashes, they are of interest for use in cements and concrete because of their possible pozzolanic reactivity. However, the presence of significant amounts of K and Cl in the prepared ashes implies a limitation of their applications.

This article reviews different methods used to perform quantitative X-ray microanalysis in the electron microscope and also demonstrates the urgency of measuring the fundamental parameters of X-ray generation for the development of accurate standardless quantitative methods. Using ratios of characteristic lines acquired on the same X-ray spectrum, it is shown that the Cliff and Lorimer KA-B factor can be used in a general correction method that is appropriate for all types of specimens and electron microscopes, providing that appropriate corrections are made for X-ray absorption, fluorescence, and indirect generation. Since the fundamental parameters appear in the KA-B factor, only the ratio of the ionization cross sections needs to be known, not their absolute values. In this regard, the measurement of ratios of the KA-B factor (or intensities at different beam energies of the same material with no change of beam spreading in the material) permits the validation for the best models to compute the ratio of ionization cross sections. It is shown, using this method, that the nonrelativistic Bethe equation, to compute ionization cross section, is very close to the equation of E. Casnati et al. (J Phys B15, 155–167, 1982) and also to the equations proposed by D. Bote and F. Salvat (Phys Rev A77, 042701, 2008) for the computation of the ratio of ionization cross sections. The method is extended to show that it could be used to determine the values of the Coster-Kronig transitions factors, an important fundamental parameter for the generation of L and M lines that is mostly known with poor accuracy. The detector efficiency can be measured with specimens where their intensities were measured with an energy dispersive spectrometer detector, the efficiency of which has been measured in an X-ray synchrotron (M. Alvisi et al., Microsc Microanal12, 406–415, 2006). The spatial resolution should always be computed when performing quantitative X-ray microanalysis and the equations of R. Gauvin (Microsc Microanal13(5), 354–357, 2007) for bulk materials and the one presented in this article for thin films should be used. The effects of X-rays generated by fast secondary electrons and by Auger electrons are reviewed, and their effect can be detrimental for the spatial resolution of materials involving low-energy X-ray lines, in certain specific conditions. Finally, quantitative X-ray microanalysis of heterogeneous materials is briefly reviewed.

Magnetotactic bacteria from aquatic environments were analyzed with the electron spectroscopic imaging technique. Rod-shaped bacteria and cocci were present in most of the samples observed. Magnetotactic multicellular aggregates were also observed at some of the sampling sites. The use of electron spectroscopic imaging allowed the observation of magnetosomes inside magnetotactic microorganisms with exceptional clarity. The number, size, and morphology of magnetosomes, as well as their ultrastructural spatial disposition inside the bacterial cell, could be directly observed and associated with the disposition of flagella of the respective cells.This allowed us to examine the structural relationships between magnetosomes and flagella, which are important components in the mechanisms of magnetotaxis. In disrupted magnetotactic multicellular aggregates, connections between cells were also visualized. We believe this technique will be useful in studying not only magnetotactic bacteria but also other uncultured microorganisms from natural environments.

The goal of this article is first to review the charging effects occurring when an insulating material is subjected to electron irradiation in a scanning electron microscope (SEM) and next their consequences from both scanning electron microscopy and electron probe microanalysis (EPMA) points of view. When bare insulators are observed, the so-called pseudo mirror effect leads to an anomalous contrast and also to an erroneous surface potential, VS, measurement when a Duane–Hunt limit (DHL) method is used. An alternative possibility is to use an electron toroidal spectrometer (ETS), specially adapted to a SEM, which directly gives the VS value. In the case of a bulk specimen coated with a grounded layer, although the layer prevents external effects of the trapped charge, the electric field beneath the coating is reinforced and leads to loss of ionizations that reduces the number of generated X-ray photons. To take into account both effects mentioned above, whether the studied insulator is coated or not, a method is proposed to deduce the trapped charge inside the insulator and the corresponding internal or external electric field.

The role of centrosomes in stem cell division has recently been highlighted and further ascribes important functions to centrosomes in stem cell maintenance, cellular differentiation, and development. Advanced cell and molecular studies coupled with immunofluorescence, electron microscopy, and live cell imaging of specific centrosome proteins have contributed greatly to our knowledge of centrosome composition, structure, and dynamics and have uncovered new insights into mechanisms of centrosome functions in asymmetric cell division. The establishment of asymmetry and differential positioning of mother and daughter centrosomes during stem cell mitosis is important for allowing one cell to maintain stem cell characteristics while the sibling cell undergoes differentiation. Another key role for centrosomes has been revealed in primary cilia of embryonic stem cells that play significant roles in cellular signaling and are therefore critically important for stem cell decisions. Studies of signaling through primary cilia may contribute important information that may aid in the production of specific cells that are suitable for tissue repair and regeneration in the field of regenerative medicine.

Extended abstract of a paper presented at Microscopy and Microanalysis 2013 in Indianapolis, Indiana, USA, August 4 – August 8, 2013.

Extended abstract of a paper presented at Microscopy and Microanalysis 2006 in Chicago, Illinois, USA, July 30 – August 3, 2006

The electron channeling contrast imaging technique was used to investigate the microstructure of copper coatings fabricated by cold gas dynamic spray. The high velocity impact characteristics for cold spray led to the formation of many substructures, such as high density dislocation walls, dislocation cells, deformation twins, and ultrafine equiaxed subgrains/grains. A schematic model is proposed to explain structure refinement of Cu during cold spray, where an emphasis is placed on the role of dislocation configurations and twinning.

Various practical issues affecting atom probe tomography (APT) analysis of III-nitride semiconductors have been studied as part of an investigation using a c-plane InAlN/GaN heterostructure. Specimen preparation was undertaken using a focused ion beam microscope with a mono-isotopic Ga source. This enabled the unambiguous observation of implantation damage induced by sample preparation. In the reconstructed InAlN layer Ga implantation was demonstrated for the standard “clean-up” voltage (5 kV), but this was significantly reduced by using a lower voltage (e.g., 1 kV). The characteristics of APT data from the desorption maps to the mass spectra and measured chemical compositions were examined within the GaN buffer layer underlying the InAlN layer in both pulsed laser and pulsed voltage modes. The measured Ga content increased monotonically with increasing laser pulse energy and voltage pulse fraction within the examined ranges. The best results were obtained at very low laser energy, with the Ga content close to the expected stoichiometric value for GaN and the associated desorption map showing a clear crystallographic pole structure.

Extended abstract of a paper presented at Microscopy and Microanalysis 2005 in Honolulu, Hawaii, USA, July 31--August 4, 2005

Energy dispersive X-ray microanalysis of micron-sized inclusions in steel is of considerable industrial importance. Measured spectra and Monte Carlo simulations show a significant effect of the steel matrix on analysis of CaO-Al2O3-MgO inclusions: the steel matrix filters the softer (Al and Mg) characteristic X-rays, increasing the relative height of the Ca peak. Bulk matrix correction methods would not result in correct inclusion compositions, but operating at a lower acceleration voltage shifts the effect to smaller inclusion sizes.