Porous silicon (PS) has been prepared using a microwave-assisted hydrofluoric acid (HF) etching method from a silicon wafer pre-implanted with 5 MeV Cu ions. The use of microbeam proton-induced X-ray emission (micro-PIXE) and microbeam Rutherford backscattering techniques reveals for the first time the capability of these techniques for studying the formation of micropores. The porous structures observed from micro-PIXE imaging results are compared to scanning electron microscope images. It was observed that the implanted copper accumulates in the same location as the pores and that at high implanted dose the pores form large-scale patterns of lines and concentric circles. This is the first work demonstrating the use of microwave-assisted HF etching in the formation of PS.

This study examines the nature of highly fragile reaction products that form in low water content supercritical carbon dioxide (scCO2) using a combination of focus ion beam/scanning electron microscopy, confocal Raman spectroscopy, helium ion microscopy (HeIM), and transmission electron microscopy (TEM). HeIM images show these precipitates are fragile rosettes. Using the TEM revealed details on the interfacial structure between the newly formed surface precipitates and the underlying initial solid phases. Detailed microscopy analysis revealed that growth of the precipitates either followed a tip growth mechanism, with precipitates forming directly on the forsterite surface if the initial solid was nonporous (natural forsterite) or growth from the surface of the precipitates, where fluid was conducted through the porous (nanoforsterite) agglomerates to the growth center. Identification of the mechanism of formation of hydrated/hydroxylated magnesium carbonate compound phases is a key factor in unraveling the impact of water recycling on mineral reactivity in low water content scCO2 solutions, which has received a great deal of attention as a result of the potential for CO2 to act as an atmospheric greenhouse gas. Techniques used here to examine these fragile structures are also used to examine a wide range of fragile material surfaces.

The present study investigates the effect of the solidification strategy for AA 6063 alloy on the surface appearance of anodized extrusions. The microstructure of the samples was analyzed using both light optical microscopy and scanning electron microscopy. Results show that if heavy segregation occurs from rapid solidification, coarse Mg2Si particles form, thus reducing the potential for precipitation strengthening by the finer β-Mg2Si developed in the solid state. Differentially-strained regions formed during hot extrusion induce differences in particle size for magnesium silicide (Mg2Si) precipitates. Anodizing generates surface roughness due to Mg2Si particle dissolution and AlFeSi decohesion, which is related to both particle size and deformation. During anodizing, an oxide layer forms on the surface of the extruded products, which can lead to streak formation, usually a subject of rejection due to unacceptable heterogeneous reflectivity.

An analysis of the nucleation mechanism of pyramids formed in (100) silicon in Na2CO3/NaHCO3 solution has been carried out. This texturization process of silicon by means of Na2CO3/NaHCO3 solutions is of special interest because it can be applied to the silicon solar cell industry to texture solar cell surfaces to decrease the front reflection and enhance light trapping in the cells. For this purpose, two microscopy techniques—scanning electron microscopy and atomic force microscopy—have been used to study the different stages of pyramidal nucleation and formation. The different aspects and factors involved in the texturization process require different analysis conditions and microscopy resolution. Tracing the transformation of determined surface areas and structures has been achieved, contributing clarification of the mechanism of pyramid nucleation in Na2CO3/NaHCO3 solutions.

Selective hydrogenation is an important process in petrochemistry to purify feedstock for polymer synthesis. For this process, catalysts containing metallic palladium deposited with an eggshell distribution on porous alumina are usually employed. For this kind of catalyst, the activity is known to be in close relation with the thickness of the palladium crust. As palladium oxide is brown and alumina is white, the palladium distribution in a catalyst bead before the reduction step can be characterized by optical microscopy. We propose an original and automatic procedure of optical image analysis to obtain a fast and robust method to measure the mean crust thickness of a catalyst batch and the corresponding standard deviation. The approach is validated by two different methods. First, we compared the crust thickness with those obtained by electron probe microanalysis. Then, catalytic tests of four samples with varying palladium crust thicknesses were performed and confirmed the expected correlation between activity and crust thickness measured by optical microscopy coupled with image analysis.

A hybrid nanobeam diffraction/imaging method, which combines well-developed diffraction imaging with nanobeam diffraction (NBD) pattern analysis, is described for structural mapping of disordered materials. Spatially resolved crystallographic information is obtained by NBD imaging by collecting NBD patterns at predefined intervals within a field of interest. The resulting dataset of NBD patterns is preprocessed to produce a spectral-imaging-like dataset and is further analyzed via multivariate statistical analysis methods in order to extract the relevant structural components and their distribution within the area of the sample under study without prior knowledge. Additional radial distribution function analysis of either the principal components or averaged data provides real-space maps of short-range order within the field of interest. This technique is demonstrated for two systems, one with multiple amorphous phases and one with multiple phases (amorphous and nanocrystalline) with similar chemistry.

A single layer of LaAlO3 with a nominal thickness of one unit cell, which is sandwiched between a SrTiO3 substrate and a SrTiO3 capping layer, is quantitatively investigated by high-resolution transmission electron microscopy. By the use of an aberration-corrected electron microscope and by employing sophisticated numerical image simulation procedures, significant progress is made in two aspects. First, the structural as well as the chemical features of the interface are determined simultaneously on an atomic scale from the same specimen area. Second, the evaluation of the structural and chemical data is carried out in a fully quantitative way on the basis of the absolute image contrast, which has not been achieved so far in materials science investigations using high-resolution electron microscopy. Considering the strong influence of even subtle structural details on the electronic properties of interfaces in oxide materials, a fully quantitative interface analysis, which makes positional data available with picometer precision together with the related chemical information, can contribute to a better understanding of the functionality of such interfaces.

Imaging of latex particles, especially those with low glass transition temperature (Tg) has been a challenging issue. Different sample preparation methods for characterization of the morphology of a poly(n-butyl acrylate)/polystyrene two-phase latex are discussed and compared in this study. A method via hydroxyethyl cellulose embedding combined with ruthenium tetraoxide (RuO4) staining for scanning transmission electron microscope (STEM) observation is developed. By using this method, the spherical shape of latex particles can be maintained without deformation. The degree of incorporation of RuO4 into latex particles and cellulose matrix is different, which makes latex particles readily identifiable from cellulose matrix under STEM. A series of latexes with different structures such as copolymer latex and organic-inorganic hybrid latex were also successfully investigated by this method. The results indicate this specimen preparation method can be applied to study the morphology of a wide range of latex systems.

This in vitro study evaluated the effect of light curing unit (LCU) type, mouthwashes, and soft drink on chemical degradation of a nanofilled resin composite. Samples (80) were divided into eight groups: halogen LCU, HS—saliva (control); HPT—Pepsi Twist®; HLC—Listerine®; HCP—Colgate Plax®; LED LCU, LS—saliva (control); LPT—Pepsi Twist®; LLC—Listerine®; LCP—Colgate Plax®. The degree of conversion analysis and the measure of the peak area at 2,930 cm−1 (organic matrix) of resin composite were done by Fourier-transform Raman spectroscopy (baseline, after 7 and 14 days). The data were subjected to multifactor analysis of variance (ANOVA) at a 95% confidence followed by Tukey's HSD post-hoc test. The DC ranged from 58.0% (Halogen) to 59.3% (LED) without significance. Differences in the peak area between LCUs were found after 7 days of storage in S and PT. A marked increase in the peak intensity of HLC and LLC groups was found. The soft-start light-activation may influence the chemical degradation of organic matrix in resin composite. Ethanol contained in Listerine® Cool Mint mouthwash had the most significant degradation effect. Raman spectroscopy is shown to be a useful tool to investigate resin composite degradation.

Dragonfly eye beads are considered to be the earliest types of glass objects in China, and in the past have been considered as evidence of culture interaction or trade between West and East Asia. In this article, synchrotron radiation microcomputed tomography and μ-probe energy dispersive X-ray fluorescence were used to determine the chemical composition, microstructure, and manufacturing technology of four dragonfly eye beads, excavated from a Chu tomb at the Shenmingpu site, Henan Province, China, dated stylistically to the Middle and Late Warring State Period (475 bc–221 bc). First, a nondestructive method was used to differentiate the material types including faience (glazed quartz), frit, glazed pottery (clay ceramic), and glass. Three beads were identified as faience and one bead as glazed pottery. The glaze recipe includes quartz, saltpeter, plant ash, and various copper, and is classified as belonging to the K2O-CaO-SiO2 glass system, which indicates that these beads were not imported from the West. Based on computed tomography slices, the manufacturing technology of the faience eye beads appears to include the use of an inner core, molding technology, and the direct application glazing method. These manufacturing features are consistent with the techniques used in China during this same time period for bronze mold-casting, proto-porcelain, and glass.

Ion channeling contrast (iCC) and electron channeling contrast (eCC) are caused by variation in signal resulting from changes in the angle of the incident beam and the crystal lattice with respect to the target. iCC is directly influenced by the incident ion range in crystalline materials. The ion range is larger for low-index crystal orientated grains, resulting in the emission of fewer secondary electrons at the surface yielding a lower signal. Ions are stopped closer to the surface for off-axis grains, resulting in the emission of many secondary electrons yielding a higher signal. Conversely, backscattered electrons (BSEs) are the primary contribution to eCC. BSEs are diffracted or channeled to form an electron channeling pattern (ECP). The BSE emission of the ECP peaks when the electron beam is normal to the surface of an on-axis grain, and therefore a bright signal is observed. Thus, iCC and eCC images yield inverse contrast behavior for on-axis oriented grains. Since there is a critical angle associated with particle channeling, accurately determining grain boundary locations require the acquisition of multiple images obtained at different tilt conditions.

Fluorescence resonance energy transfer (FRET) microscopy is a powerful technique to quantify dynamic protein-protein interactions in live cells. Total internal reflection fluorescence (TIRF) microscopy can selectively excite molecules within about 150 nm of the glass-cell interface. Recently, these two approaches were combined to enable high-resolution FRET imaging on the adherent surface of living cells. Here, we show that interference fringing of the coherent laser excitation used in TIRF creates lateral heterogeneities that impair quantitative TIRF-FRET measurements. We overcome this limitation by using a two-dimensional scan head to rotate laser beams for donor and acceptor excitation around the back focal plane of a high numerical aperture objective. By setting different radii for the circles traced out by each laser in the back focal plane, the penetration depth was corrected for different wavelengths. These modifications quell spatial variations in illumination and permit calibration for quantitative TIRF-FRET microscopy. The capability of TIRF-FRET was demonstrated by imaging assembled cyan and yellow fluorescent protein–tagged HIV-Gag molecules in single virions on the surfaces of living cells. These interactions are shown to be distinct from crowding of HIV-Gag in lipid rafts.

We used hybrid detectors (HyDs) to monitor the trajectories and interactions of promyelocytic leukemia (GFP-PML) nuclear bodies (NBs) and mCherry-53BP1-positive DNA lesions. 53BP1 protein accumulates in NBs that occur spontaneously in the genome or in γ-irradiation-induced foci. When we induced local DNA damage by ultraviolet irradiation, we also observed accumulation of 53BP1 proteins into discrete bodies, instead of the expected dispersed pattern. In comparison with photomultiplier tubes, which are used for standard analysis by confocal laser scanning microscopy, HyDs significantly eliminated photobleaching of GFP and mCherry fluorochromes during image acquisition. The low laser intensities used for HyD-based confocal analysis enabled us to observe NBs for the longer time periods, necessary for studies of the trajectories and interactions of PML and 53BP1 NBs. To further characterize protein interactions, we used resonance scanning and a novel bioinformatics approach to register and analyze the movements of individual PML and 53BP1 NBs. The combination of improved HyD-based confocal microscopy with a tailored bioinformatics approach enabled us to reveal damage-specific properties of PML and 53BP1 NBs.

The aim of this study was to analyze whether the mesenchymal stromal cells (MSCs) isolated from palatal tissue grafts harvested in order to cover gingival recessions have the basic characteristics of stem cells. The palatal tissue cells were processed using a special culture medium that stimulated the development of only undifferentiated cellular lines. Cells at passage 4 were evaluated by flow cytometry to examine the expression of specific surface markers and were tested for multilineage differentiation capacity. These cells collected at passage 4 were also investigated for the capacity to cluster into embryoid body aggregates. Palatal MSCs displayed positive staining for the mesenchymal markers CD29, CD73, CD105, CD 49e, and CD44, but did not express hematopoietic markers CD34/45. The palatal MSCs successfully differentiated into osteogenic, adipogenic, and chondrogenic lineages. When seeded in special conditions, palatal MSCs propagated into unattached spheres resembling embryoid body aggregates consisting both of differentiated and undifferentiated cells as revealed at the ultrastructural evaluation. It is concluded that the isolated palatal MSCs fulfilled the basic criteria defining the stem cells. This new source of stem cells characterized here for the first time opens new perspectives on possible applications in basic research and in regenerative medicine.

Defining the ultrastructure of endocytic sites and localization of endocytic proteins in Saccharomyces cerevisiae by immunoelectron microscopy is central in understanding the mechanisms of membrane deformation and scission during endocytosis. We show that an improved sample preparation protocol based on high-pressure freezing, freeze substitution, and low-temperature embedding allows us to maintain the cellular fine structure and to immunolabel green fluorescent protein–tagged endocytic proteins or actin in the same sections. Using this technique we analyzed the stepwise deformation of endocytic membranes and immunolocalized the endocytic proteins Abp1p, Sla1p, Rvs167p, and actin, and were able to draw a clear ultrastructural distinction between endocytic sites and eisosomes by immunolocalizing Pil1p. In addition to defining the geometry and the fine structure of budding yeast endocytic sites, we observed associated actin filaments forming a cage-like meshwork around the endocytic membrane.

Ultrastructural answer of bone marrow erythroid series and of red blood cells (RBCs) in Wistar rats to bee venom (BV) was analyzed by transmission and scanning electron microscopy, and corroborated with hematological data. A 5-day and a 30-day treatment with daily doses of 700 μg BV/kg and an acute-lethal treatment with a single dose of 62 mg BV/kg were performed. The 5-day treatment resulted in a reduced cellularity of the bone marrow, with necrosed proerythroblasts, polymorphous erythroblasts, and reticulocytes with cytoplasmic extensions, and a lower number of larger RBCs, with poikilocytosis (acanthocytosis) and anisocytosis, and reduced concentrations of hemoglobin. After the 30-day treatment, the bone marrow architecture was restored, but polymorphous erythroblasts and reticulocytes with thin extensions could still be observed, while the RBCs in higher number were smaller, many with abnormal shapes, especially acanthocytes. The acute treatment produced a partial depopulation of the bone marrow and ultrastructural changes of erythroblasts including abnormal mitochondrial cristae. The RBCs in lower number were bigger and crenated, with reduced concentrations of hemoglobin. Overall, BV was able to promote stress erythropoiesis in a time- and dose-related manner, mitochondrial cristae modification being a critical factor involved in the toxicity of the BV high doses.

Most blood vessels contain elastin that provides the vessels with the resilience and flexibility necessary to control hemodynamics. Pathophysiological hemodynamic changes affect the remodeling of elastic components, but little is known about their structural properties. The present study was designed to elucidate, in detail, the three-dimensional (3D) architecture of delicate elastic fibers in small vessels, and to reveal their architectural pattern in a rat model. The fine vascular elastic components were observed by a newly developed scanning electron microscopy technique using a formic acid digestion with vascular casts. This method successfully visualized the 3D architecture of elastic fibers in small blood vessels, even arterioles and venules. The subendothelial elastic fibers in such small vessels assemble into a sheet of meshwork running longitudinally, while larger vessels have a higher density of mesh and thicker mesh fibers. The quantitative analysis revealed that arterioles had a wider range of mesh density than venules; the ratio of density to vessel size was higher than that in venules. The new method was useful for evaluating the subendothelial elastic fibers of small vessels and for demonstrating differences in the architecture of different types of vessels.

Studies of human bloodstains on nonbiological materials have been previously carried out using a high-vacuum scanning electron microscope (HV-SEM) in secondary-electron mode without any sample treatment. To assess whether biological substrates can affect the morphology of human erythrocytes in bloodstains, three fragments of different biological material (bone, shell, and wood) were smeared with peripheral human blood. Afterward, the bloodstains were directly examined in secondary-electron mode by an HV-SEM following a procedure initially standardized to be used in uncoated human bloodstains on stone. The obtained results suggest that HV-SEM is suitable for examining untreated bloodstains on biological substrate and that the morphology of erythrocytes in human bloodstains is not affected by the biological nature of the substrate. A cautionary issue regarding bloodstains on nondehydrated biological substrates is that the waiting time required for initiating the HV-SEM examination is by far higher than when using inorganic bloodstain substrates.

Ferruginous bodies observed in lungs of patients affected by mesothelioma, asbestosis, and pulmonary carcinoma are important to relate the illness to exposure, environmental or occupational, to asbestos. Identification of the inorganic phase constituting the core of the ferruginous bodies, formed around asbestos but also around phases different from asbestos, is essential for legal purposes. Environmental scanning electron microscopy/energy dispersive spectroscopy was used to identify the fibrous mineral phase in the core of ferruginous bodies observed directly in thin sections of tissue, without digestion of the biological matrix. Spectra were taken with sequential analyses along a line crossing the core of the ferruginous bodies. By comparing the spectra taken near to and far from the core, the chemical elements that make up the core could be identified.

We propose a nonscanning three-dimensional (3D) optical microscope based on reflectivity-height transformation in applications of biological and transparent plate measurements. The reflectivity of a prism can be transformed to the surface height of the specimen based on geometrical optics and the principle of internal reflection. Thus, the pattern of reflectivity is representative of the surface profile. Using charge-coupled device cameras to obtain the two-dimensional image patterns and combining with its reflectivity pattern, the 3D profile can be generated. The lateral resolution is determined by the diffraction limit, and the vertical resolution is better than several nanometers according to the incident angle and polarization used.