Characterization of Dynamics in Materials Using Time-Resolved Electron Microscopy Incorporating Electron Counting and Electron Correlation Spectroscopy

IntroductionCurrently, the resolution of structural detail in conventional transmission electron microscopy (CTEM) of organic materials is limited by the electron beam damage suffered by the sample. It has recently been shown that the imaging modes now available in scanning transmission electron microscopy (STEM) provide the potential for improvements in resolution over CTEM methods. However, the effect of the sensitivity of an organic sample itself to beam damage has not been compared in STEM versus CTEM. In STEM each element of the sample area is briefly illuminated with a small diameter, high flux, scanning electron beam, whereas in CTEM the entire sample area is continuously illuminated with a large diameter, lower flux, static electron beam. Although it has been shown that the amount of sample damage in CTEM is independent of dose rate(2), the electron flux in STEM may range from 3 to 4 orders of magnitude greater than that in CTEM, with a possible influence on damage rate. Generally, the effects of beam size, high flux, and short dwell time of the scanning electron beam on sensitivity of the sample to damage in STEM are not known.

An elastic protein with a secondary structure distinct from all well-known load-bearing proteins is found in the byssus of the giant clam, Tridacna maxima . The byssus consists of a bundle of hundreds of individual threads, each measuring about about 100 μm in diameter, which exhibit a tendon-like mechanical response. The amino acid composition of Tridacna byssus, however, is unlike tendon collagen, lacking high glycine, proline, and hydroxyproline. Wide-angle X-ray scattering (WAXS) and small-angle X-ray scattering (SAXS) measurements suggest that the constituent nanofibrils of the byssal threads are distinct from known secondary structure motifs previously reported for elastic proteins including the collagen triple-helix, the β-sheet nanocrystalline domains of silks, or the double-stranded coiled-coil regions of intermediate filaments. Instead, X-ray diffraction data indicate a structural organization in which four coiled-coil α-helices form a stable rope-like structure, which then further pack in a pseudohexagonal lattice to form nanofibrils. Amino acid composition analysis shows unusually high concentrations of acidic as well as basic residues, suggesting that the four-helix structure is stabilized by strong ionic interactions between oppositely charged residues in neighboring strands. The composition also suggests additional stabilization by disulfide cross-linking. On a larger scale, scanning and conventional transmission electron microscope (STEM and TEM) observations indicate that the nanofibrils exhibit an alternating periodicity of about 500 nm along the axial direction. A molecular model that combines the mechanical properties with the structural characteristics of the Tridacna byssal threads is proposed.

There are mainly two complementary imaging modes in transmission electron microscopy (TEM): Conventional TEM (CTEM) and scanning TEM (STEM). In the CTEM mode the specimen is illuminated with a plane electron wave, and the direct image formed by the objective lens is recorded in the image plane. STEM is based on scanning the specimen surface with a focused electron beam and collecting scattered electrons with an extended disk or ring-shaped detector. Here we show that combination of CTEM imaging with STEM illumination generally allows extending the point resolution of CTEM imaging beyond the diffraction limit. This new imaging mode improves imaging characteristics, is more robust against chromatic aberration, exhibits direct structural imaging with superior precision, visualizes light elements with excellent contrast, and even allows us to overcome the conventional information limit of a microscope.

Adult channel catfish were exposed to 0 or 15 ppm arsenic as sodium arsenate in synthetic lake water for approximately six months. After sacrifice, livers were removed and examined by conventional transmission electron microscopy, optical microscopy, scanning electron microscopy, and electron probe X-ray microanalysis. Cytoplasmic structures in the nuclei were found to contain highly concentrated deposits of iron. Thirty percent of these structures were also found to contain arsenic at a level above the minimum detectable level for the conditions of the study. The size, shape and relative position of the cytoplasmic structures were the same as the hemosiderin granules observed by conventional transmission electron microscopy. (JMT)

The energy filtered imaging technique has so far been carried out in a scanning transmission electron microscope (STEM) fitted with a sector type energy analyzer. The STEM has a disadvantage of low beam parallelity because it uses a convergent beam, while the conventional transmission electron microscope (CTEM) allows good phase contrast and diffraction contrast to be obtained because of the high parallelity of the beam, and allows also high resolution images to be obtained. The technique to obtain energy filtered CTEM images has thus far been carried out by a Castaing-Henry type filter or an Ω type filter. However, these filters have the disadvantage of lower energy resolution than conventional sector type energy analyzer at the present time. This paper reports energy filtered CTEM images of MgO smoke, obtained using a new scanning CTEM image technique and a high energy resolution sector type energy analyzer which can resolve bulk and surface plasmon energy.

High density lipoproteins of d = 1.12 to 1.21 g/ml from human serum (HDL3) were studied by electron microscopy with both negative staining and freeze-etching techniques. For the negatively stained specimens, a modified conventional transmission electron microscope as well as a scanning transmission electron microscope were used. The freeze-etched specimens were examined by a conventional transmission electron microscope. The diameter of HDL3 was found to be 105 +/- 4 A by freeze-etching and 94 +/- 6 A by negative staining. The surface of the HDL3 particles exhibited about 12 discrete domains, 28 +/- 3 A (freeze-etched) and 28 +/- 4 A (negatively stained) in diameter, of undefined chemical composition. Moreover, the freeze-etched specimens revealed an inner core 40 +/- 2 A in diameter, corresponding to estimated values reported previously. All information is consistent with the HDL3 model proposed by B. W. Shen, F. J. Kézdy, and A. M. Scanu [(1977) Proc. Natl. Acad. Sci. USA 74, 837-841], with additional evidence for well-defined surface substructure. The consistency of the images obtained with the various electron microscopy techniques and the marked change in the appearance of the surface in the HDL3 preparations that were digested by phospholipase A2 (EC 3.1.1.4) support the validity of the interpretation.

In many fields of research in science, engineering, and medicine, electron microscopy as a method for directly imaging submicroscopic structures has become increasingly important in recent decades. Electron microscopy (EM) includes several different techniques: conventional transmission electron microscopy (TEM), high-resolution electron microscopy (HREM), highvoltage electron microscopy (HVEM), scanning electron microscopy (SEM), analytical electron microscopy (AEM), emission electron microscopy (EEM), and others. In the past the central aim of using electron microscopy was structure determination, but recently it has been of growing importance for also investigating different processes, i.e., changes in materials by interaction with several influential factors (e.g., heat, electric or mechanic fields, mechanical loading). Of particular interest is the study of the micromechanical processes of deformation and fracture. Therefore, electron microscopy is a very powerful tool for materials science. The present review reports on some capabilities and limitation of the application of electron microscopy to solid polymers. In Section II the techniques of electron microscopy are briefly reviewed, followed by a section that describes the main methods of specimen preparation of solid polymeric materials. In Section IV the results of several applications of electron microscopy are discussed to reveal the morphology as well as the micromechanical deformation processes of several polymeric materials.

The effects of parenteral injection of aluminum, indium, gadolinium, or terbium in rats have been previously studied in several organs such as the liver, the kidneys, etc., but never in mammary glands. In this work, we have attempted to study the subcellular localization of these elements after their intraperitoneal administration. Their subsequent effects in the lactating mammary gland cells have also been studied. Our results using conventional transmission electron microscopy have shown that the lysosomes of the mammary glandular epithelial cells are the intracellular site of accumulation of the studied elements. Our results have also show intracellular deteriorations such as an expanded ergastoplasm and altered mitochondria after intraperitoneal injection of aluminum and indium.

The microstructural and morphological properties of the as-prepared tin (Sn) coating on ferrous substrates affects the lifetime as well as the applicability of the coating itself. In order to improve our knowledge of this system, strips of steel St-37 were coated with Sn in the form of gas-atomized powder, with an average diameter of 110±10 μm, using plasma spraying. The plasma gas was an Ar-He mixture and the current intensity was 650 A. The as-casted coating was characterized with scanning electron microscopy (SEM), X-ray diffraction (XRD) and conventional transmission electron microscopy (CTEM). It was deduced that the coating is characterized by high integrity, although certain pores were detected far from the Fe-Sn interface. However they refer to less than 1% of the total volume. Nevertheless the examination with SEM and XRD proved that the coating is mainly composed of pure Sn. However, interdiffusion takes place between the two phases (Fe-Sn) at their interface as deduced by CTEM. As a result, certain Fe-Sn phases are formed, which seem to be scattered in the coating close to the interface in the form of small inclusions.

The installation of an imaging energy filter extends the imaging and analytical capabilities of a conventional transmission electron microscope (CTEM) considerably. We report here on experiments which demonstrate the versatility of such a device. The arrangement of our system is shown in Fig. 1. The experiments performed pertain to:The improvement of contrast in thick specimens by exclusion of inelastically scattered electrons. The spatial resolution of the electron microscope can be preserved by the correction of second-order aberrations with additional electromagnetic hexapoles.The determination of the concentration of light elements with EELS at a spatial resolution better than 10 nm. The chemical structure of ceramic material (SI3N4and SiC with additions) could thus be elucidated. A study on the extended fine structure in the energy loss spectra of the oxygen and magnesium K edges in MgO (Fig. 2) did confirm the theoretical prediction for that material.

Characterization of deformation localization in cold-rolled metastable β-Ti–Nb–Ta–Zr alloy

Crystalline structures of carbon complexes in amorphous carbon films

It is well known that the combination of a field emission gun (FEG) and a conventional transmission electron microscope (CTEM) is extremely important for nanometer area analysis in analytical electron microscopy. However, the smaller illumination angle and reduced energy spread of FEG than those of a conventional electron gun (W hair pin filament or LaB6) give a slowly damping envelop function in phase contrast transfer function (PCTF). Thus the FEG ensures application not only to analytical microscopy but also to high resolution electron microscopy to improve the information limit.In a high voltage electron microscope (above 200 kV), high-speed vacuum pumps have to be provided below the acceleration tube to get an ultra high vacuum (UHV) around the field emission tip located at the top of the acceleration tube. However, this method is not always the best way to provide UHV because of the poor vacuum conductance caused by the electrodes inside the acceleration tube.

Multicolored gene reporters for light microscopy are indispensable for biomedical research, but equivalent genetic tools for electron microscopy (EM) are still rare despite the increasing importance of nanometer resolution for reverse engineering of molecular machinery and reliable mapping of cellular circuits. We here introduce the fully genetic encapsulin/cargo system of Quasibacillus thermotolerans (Qt), which in combination with the recently characterized encapsulin system from Myxococcus xanthus (Mx) enables multiplexed gene reporter imaging via conventional transmission electron microscopy (TEM) in mammalian cells. Cryo-electron reconstructions revealed that the Qt encapsulin shell self-assembles to nanospheres with T = 4 icosahedral symmetry and a diameter of ∼43 nm harboring two putative pore regions at the 5-fold and 3-fold axes. We also found that upon heterologous expression in mammalian cells, the native cargo is autotargeted to the inner surface of the shell and exhibits ferroxidase activity leading to efficient intraluminal iron biomineralization, which enhances cellular TEM contrast. We furthermore demonstrate that the two differently sized encapsulins of Qt and Mx do not intermix and can be robustly differentiated by conventional TEM via a deep learning classifier to enable automated multiplexed EM gene reporter imaging.

We report our experience with a prototype combined light and electron microscope (the LEM 2000) with particular reference to its application to routine surgical histopathology. We found its major advantages over conventional transmission electron microscopies were due to the large grid size (7 mm diameter), low magnification capacity (x 250), and the built-in microprocessor for recording areas of interest. These features combine to reduce sampling errors and greatly facilitate orientation and relocation of fields of diagnostic importance.