TEM in materials science—past,present and future

The progress in transmission electron microscopy as applied in materials science is reviewed briefly, from the era of replica techniques in the 1940s and 1950s, through the development of the diffraction contrast technique in the late 1950s and 1960s, to the present day when instrumental resolution is sufficient to obtain structure images of a wide variety of crystalline solids. One of the most important advances has been the development of combined imaging and microanalytical techniques in a single instrument. The versatility of the TEM technique with atomic resolution and microanalytical facilities, the variety of signals and contrast effects available, and its universal application, establishes it as an essential tool in materials science for the foreseeable future.     

Structure and morphology of the in situ composites of a liquid crystalline polymer and polycarbonate

In situ composites were prepared via melt blending of a liquid crystalline polymer (LCP) and polycarbonate using a twin screw extruder. The structure and morphology of these composites were analysed using both transmission electron microscopy (TEM) and scanning electron microscopy. The LCP phases were able to orientate and form in situ submicrometre fibres during the extrusion and post-extrusion drawing. TEM images as well as selected-area diffraction patterns were obtained from the materials. The effects of both composition, i.e. LCP content, and post-extrusion draw-down ratio on the development of the in situ formed LCP fibres were studied in detail. A skin–core morphological differentiation is observed in these materials where well-defined LCP fibres of higher aspect ratios were formed in the skin region. However, a significant amount of unelongated LCP particles were found coexisting with the less well-defined fibres in the core region of the extrudates. This skin–core differentiation was found to be dependent on the composition and the processing conditions, e.g. draw ratio. In this instance, electron microscopy is proven to be a powerful technique not only for direct observation of the formation, dimensions and morphology of the in situ LCP fibres, but also for the qualitative and quantitative characterization of the molecular orientation and crystalline structures in these fibres using selected-area electron diffraction. It is observed that the skin–core differentiation becomes more distinct in the in situ composites containing a higher percentage of LCP but diminishes when the material is processed at higher post-extrusion draw ratio.     

Polyphosphate at the Streptomyces lividans cytoplasmic membrane is enhanced in the presence of the potassium channel KcsA

The distribution of polyphosphate (polyP) within the cytoplasmic membrane of Streptomyces lividans hyphae or protoplasts has been determined at high spatial resolution by elemental mapping using energy‐filtered electron microscopy (EFTEM). The results revealed that polyP was best traceable after its interaction with lead ions followed by their precipitation as lead sulphide. Concomitant studies of the S.lividans wildtype (WT) strain and its co‐embedded mutant ΔK (lacking a functional kcsA gene) were conducted by labelling as the surface matrix of either one was labelled by cationic colloidal thorium dioxide. Within the WT strain, additional polyP was found to accumulate distinctly at the inner face of the cytoplasmic membrane. After removal of the cell wall (within protoplasts), the polyP‐derived lead‐sulphide (PbS) precipitate formed clusters of fibrillar material extending up to 50 nm into the cytoplasm. This feature was absent in the ΔK mutant strain. Together the results revealed that the presence of the KcsA channel and the structured polyP coincide.     

In-situ observation of shape and atomic structure of Xe nanocrystals embedded in aluminium

An imaging technique to determine in situ the shape and atomic structure of nanosized Xe crystals embedded in Al is described using high-resolution transmission electron microscopy (HRTEM). The Xe nanocrystals, with sizes less than 5 nm were prepared by the implantation of 30 keV Xe⁺ into Al at room temperature. The fcc Xe nanocrystals are mesotactic with the Al lattice and have a lattice parameter ≈ 50% larger than that of Al. HRTEM images of the Xe were not clear in [110] zone axis illumination because of the small number of Xe atoms relative to Al atoms in any atom column. An off-axial imaging technique that consists of tilting the specimen several degrees from a zone axis and defocusing to suppress the Al lattice fringes is employed for the 110 projection of the Xe/Al system and the structure of the Xe nanocrystals is successfully imaged. The Xe images clearly represent projections of cuboctahedra with faces parallel to eight Al {111} planes truncated by six {100} planes. The results of multislice image simulations using a three-dimensional atomic model agreed well with the results obtained by the off-axial imaging technique. The usefulness of the technique is demonstrated with observations of crystal defects introduced into the Xe under intense 1000 keV electron irradiation.     

Electron microscopy study of different stages of oxidation of Ti-47Al-2Nb-2Cr-0.15B and Ti-45Al-15Nb at 900°C

The initial stage of oxidation of Ti-47Al-2Nb-2Cr-0.15B and Ti-45Al-15Nb alloys was studied. Studies reveal that the X and NbCr₂ phases will form in advance in the transition layer of the 2Nb2Cr alloy compared with the 15Nb alloy. The adherence between the nitride layer and underling layer is good. However, an obvious boundary exists between the mixed layer and the inner layer. The recrystallizing of the base alloy leads to an increase of the volume fraction of grain boundaries that can increase the oxidation rate at the initial oxidation stage. An Nb-based compound forms in the transition layer, which can prevent the formation of X phase.     

Advantages of indium–tin oxide-coated glass slides in correlative scanning electron microscopy applications of uncoated cultured cells

A method of direct visualization by correlative scanning electron microscopy (SEM) and fluorescence light microscopy of cell structures of tissue cultured cells grown on conductive glass slides is described. We show that by growing cells on indium–tin oxide (ITO)-coated glass slides, secondary electron (SE) and backscatter electron (BSE) images of uncoated cells can be obtained in high-vacuum SEM without charging artefacts. Interestingly, we observed that BSE imaging is influenced by both accelerating voltage and ITO coating thickness. By combining SE and BSE imaging with fluorescence light microscopy imaging, we were able to reveal detailed features of actin cytoskeletal and mitochondrial structures in mouse embryonic fibroblasts. We propose that the application of ITO glass as a substrate for cell culture can easily be extended and offers new opportunities for correlative light and electron microscopy studies of adherently growing cells.     

TC4钛合金锯齿形切屑绝热剪切带的微观组织和显微硬度变化

使用金相显微镜、透射电子显微镜观察了TC4钛合金在不同切削速度下形成的锯齿形切屑的绝热剪切带的微观组织形貌,测量了绝热剪切带的显微硬度。结果表明,在较低切削速度下,绝热剪切带为形变带;在较高切削速度下,绝热剪切带为由细小等轴晶粒和局部发生了β相转变为α″相的马氏体相组成的转变带;随着切削速度的进一步提高,在绝热剪切带中观察到了非晶化这一新现象。无论绝热剪切带是形变带还是转变带,其显微硬度都随切削速度的增大而增大。根据绝热剪切带微观组织和显微硬度的变化规律,绝热剪切带显微硬度的强化可分为三个阶段:形变强化、细晶和马氏体相变强化、非晶强化。     

Scanning electron microscopy of nonconductive specimens at critical energies in a cathode lens system

A method for scanning electron microscopy imaging of nonconductive specimens, based on measurement and utilisation of a critical energy, is described in detail together with examples of its application. The critical energy, at which the total electron yield curve crosses the unit level, is estimated on the basis of measurement of the image signal development from the beginning of irradiation. This approach, concentrated onto the detected signal as the only quantity crucial for the given purpose of acquiring a noncharged micrograph, evades consequences of any changes in an irradiated specimen that influence the total electron yield curve and possibly also the critical energy value. Implementation of the automated method, realised using a cathode lens-equipped scanning electron microsope (SEM), enables one to establish a mean rate of charging over the field of view and its dependence on the electron landing energy. This dependence enables one to determine the energy of a minimum damage of the image of the given field of view. Factors influencing reliability and applicability of the method are discussed and examples of noncharged micrographs of specimens from both life and material science fields are presented.     

Advantages of stereo imaging of metallic surfaces with low voltage backscattered electrons in a field emission scanning electron microscope

High emission current backscattered electron (HC-BSE) stereo imaging at low accelerating voltages (≤ 5 keV) using a field emission scanning electron microscope was used to display surface structure detail. Samples of titanium with high degrees of surface roughness, for potential medical implant applications, were imaged using the HC-BSE technique at two stage tilts of + 3° and − 3° out of the initial position. A digital stereo image was produced and qualitative height, depth and orientation information on the surface structures was observed. HC-BSE and secondary electron (SE) images were collected over a range of accelerating voltages. The low voltage SE and HC-BSE stereo images exhibited enhanced surface detail and contrast in comparison to high voltage (> 10 keV) BSE or SE stereo images. The low voltage HC-BSE stereo images displayed similar surface detail to the low voltage SE images, although they showed more contrast and directional sensitivity on surface structures. At or below 5 keV, only structures a very short distance into the metallic surface were observed. At higher accelerating voltages a greater appearance of depth could be seen but there was less information on the fine surface detail and its angular orientation. The combined technique of HC-BSE imaging and stereo imaging should be useful for detailed studies on material surfaces and for biological samples with greater contrast and directional sensitivity than can be obtained with current SE or BSE detection modes.     

Cathodoluminescence imaging—the study of yttrium aluminates distribution in aluminum nitride

Cathodoluminescence imaging is a technique which may combine a high-resolution spectroscopic method with a high-spatial resolution electron imaging technique to characterize chemical identity as well as physical size, shape, and spatial distribution of the materials which have cathodoluminescence. Cathodoluminescence imaging has been used to characterize yttrium aluminates in aluminum nitride ceramics. By digitally combining cathodoluminescence and backscat-tering electron images, it is possible not only to distinguish between different yttrium aluminates, but also to determine the size, shape, and spatial distribution of these phases. This information provides a route to understanding the local residual oxygen concentration before sintering, the amount of sintering aid to be added, and possible sintering schedule. Furthermore, this information may be related to other properties of aluminum nitride such as residual oxygen concentration and thermal conductivity.     

Early mineralization of matrix vesicles in the epiphyseal growth plate

Matrix vesicles (MVs) induce the primary mineralization in collagen-rich hard tissues such as bone, mineralizing cartilage and dentine. Calcium and phosphate ions accumulate at the inner MV membrane. This accumulation takes place in association with phospholipids alone and/or in association with Annexin V, which displays Ca ion channel activity when inserted in membranes; consequently, Annexin V may be involved in Ca uptake by matrix vesicles. The first crystal nuclei are formed at these macromolecules of the MV inner membrane. They grow to stable nanometre-sized particles, dots, which coalesce to form chains of dots along the macromolecules of the MV inner membrane. At the same time, or shortly afterwards, chains of these Ca phosphate dots also develop inside the MVs. The measured centre-to-centre distances between these dots represent approximately the distances between the nucleating sites, called active sites, along the MV matrix molecules. The mineralization does not stop at the MV membrane but expands continuously into the extravesicular region in radial directions to form nodules. These radiating Ca phosphate chains, which coalesce to form needles, are composed of such primary dots, which have developed at the nucleating sites of the corresponding macromolecules.     

A new analytical method for characterising the bonding environment at rough interfaces in high-k gate stacks using electron energy loss spectroscopy

Determining the bonding environment at a rough interface, using for example the near-edge fine structure in electron energy loss spectroscopy (EELS), is problematic since the measurement contains information from the interface and surrounding matrix phase. Here we present a novel analytical method for determining the interfacial EELS difference spectrum (with respect to the matrix phase) from a rough interface of unknown geometry, which, unlike multiple linear least squares (MLLS) fitting, does not require the use of reference spectra from suitable standards. The method is based on analysing a series of EELS spectra with variable interface to matrix volume fraction and, as an example, is applied to a TiN/poly-Si interface containing oxygen in a HfO₂-based, high-k dielectric gate stack. A silicon oxynitride layer was detected at the interface consistent with previous results based on MLLS fitting.     

Peculiarities of diamonds formed in alkaline carbonate-carbon melts at pressures of 8–10 GPA: Scanning electron microscopy and cathodoluminescence data

Diamond crystallization with both spontaneous and seeded nucleation was realized in the system Na₂Mg(CO₃)₂-K₂Mg(CO₃)₂-C (graphite) at 8–10 GPa and 1700–1800°C. The crystallization products were transparent colorless diamond single crystals of octahedral habit up to 100–150 μm in size. Scanning electron microscopy (SEM) showed that the growing diamond material was precipitated on both octahedral {111} and cubic { 100 } faces of synthetic and natural diamond seed crystals by layers of octahedral orientation, much like the growth of the natural diamond. The physicochemical conditions for diamond crystallization are interpreted as a crystal growth from carbon solutions in alkaline-carbonate melts. Color cathodoluminescence scanning electron microscopy (CCL-SEM) and cathodoluminescence (CL) spectroscopy studies suggested specific peculiarities of the synthetic “carbonate-carbon”(CC) diamonds resembling natural crystals in comparison with diamonds produced by metal-carbon (MC) synthesis. The main feature of the CC product is the lack, for both spontaneous and seed stimulated diamonds, of surface color cathodoluminescence as in the case for natural diamonds with lower concentrations of nitrogen impurity (type II). The CL spectra of the CC diamonds showed the simultaneous luminescent three-band system - H3, 575 nm, and a weak blue A-band - the H3 band structure of which resembles that of natural diamonds of type IIa.     

A high-resolution electron microscopic study of defects in sodium β′″-alumina

A high-resolution electron-microscopic study of sodium β′″-alumina, a polytype of the more-widely-studied sodium β- and β″-alumina, has been undertaken using the 600 kV instrument at Cambridge University. Images revealed the loss of sodium-containing planes, which had caused crystals to collapse and shear into defect layers. A model for the structure of these defects is proposed, based on the use of computed images and by comparison with high-resolution images of silver β″-alumina.     

Probe three-dimensional structure of soft organized surfactants at solid–liquid interface with an AFM in MAC Mode and contact mode

Atomic force microscopy provides a unique direct-visualization tool to study the three-dimensional structure of adsorbed surfactants on solid surfaces. Ionic surfactant molecules spontaneously adsorbed onto hydrophilic surfaces from aqueous solution above the critical micelle concentration (cmc) have been imaged using an atomic force microscope in magnetic ac mode (MAC Mode) and contact mode. It was found that the soft organized surfactants were highly compressible and therefore showed a wide range of corrugations depending on imaging forces. When using gentle MAC Mode, corrugations of the organized surfactants around half of the estimated height of the proposed surfactant aggregate cylinders on mica have been stably observed. Traditional contact mode operating in the pre-contact double-layer electrostatic interaction region, however, showed significantly reduced height of the organized molecules.     

Intermediate aggregates resulting in the interaction of bile salt with liposomes studied by transmission electron microscopy and light scattering techniques

The interaction of sodium cholate (SC) with phosphatidylcholine liposomes was studied by means of transmission electron microscopy (TEM), changes in the mean particle size (quasielastic light scattering, QELS) and in the static light scattering (SLS) of the system during liposome solubilization. A good correlation was found between the TEM diameter of particles and the mean hydrodynamic diameter (HD) determined by QELS. The intermediate aggregates resulting in this interaction were dependent on the SC concentration in the system. Thus, an initial vesicle growth occurred when the SC concentration in the system was 13.79 mol%. Additional SC amounts (41.17 mol% SC) led to the formation of the largest vesicles (HD 410 nm). Increasing SC amounts led to a slight fall in the vesicle diameter and in the SLS of the system. Thus, for 47.08 mol% SC, TEM images still showed the presence of vesicles albeit with traces of smaller structures and signs of vesicle fusion. When SC concentration exceeded 48 mol% an abrupt decrease in SLS occurred, the size curve starting to show a bimodal distribution. Thus, for 50 mol% SC a sharp distribution curve appeared at 52 nm indicating the formation of small particles and TEM images showed clear signs of vesicle disintegration with formation of tubular structures. The subsequent self organization of these tubular structures (54 mol% SC) led to the formation of open multilayered structures in coexistence with small particles. A gradual increase in the number of these small particles (mixed micelles) led to the complete solubilization of liposomes.     

Quantitative and qualitative characterization of aquatic iron oxyhydroxide particles by EF-TEM

Electron energy-loss spectroscopy (EELS) and elemental imaging under the energy-filtered transmission electron microscope are powerful tools for the characterization of iron-rich particles present in natural waters. Features present in EEL spectra (Fe-M_2,3 Fe-L_2,3 and O-K ionization edges) of goethite (α-FeOOH) have been studied with an energy filter operated at 80 keV to determine optimal quantification and elemental imaging of Fe-rich natural aquatic particles in the 30–200 nm range of thickness. For quantitative aims, the Fe-M_2,3 ionization edge cannot be used easily, but the Fe-L_2,3 edge provides more accurate results owing to a better background extrapolation. The partial cross-section of the Fe(III) M shell has been determined for iron oxide. The use of two-windows (jump-ratio) and three-windows (background stripping) imaging methods is discussed in relation to the specimen thickness.