Three-dimensional visualization of dislocations by serial sectioning of SEM-ECCI images was performed on a creep-deformed nickel-based heat-resistant alloy. By performing serial tomograms of ECCI images, a volume of approximately 5 μm × 5 μm × 0.3 μm volume was successfully reconstructed. By incorporating the crystal orientation information obtained by the SEM-EBSD method, we confirmed that the three-dimensional arrangement of dislocations, such as slip planes and cross-slip, is accurately reflected in the three-dimensional volume. Thus, serial sectioning of ECCI images enables us to observe and analyze the bulk dislocation microstructure in three dimensions in the micrometer scale range.

Focused ion beam systems have been widely used for TEM specimen preparation in the field of research development and industry. However, FIB damaged layers are formed on the specimen surfaces during the FIB milling with high energy Ga ion beam. To remove the FIB damaged layers, low energy Ga or Ar ion beams are applied to the TEM specimens after the FIB milling, which are extremely effective for semiconductor and ceramics materials. Such the low energy Ga and Ar ion beam were applied to thin platinum foils, which were thinned by high energy Ga ion beam, to investigate the damaged layer formed on the surfaces of the platinum foils. In addition, results of 3D reconstruction of EuBa₂Cu₃O_{7 – y} superconductive layer with BaHfO₃ rods using FIB-SEM system were reported. Following the results, problems and future developments of the 3D reconstruction technique using the FIB-SEM system are described.

Xe plasma FIB offers an extremely high etching rate at least 50 times higher than common liquid metal ion source based Ga FIB. In recent years, plasma FIB-SEM systems become more important and popular in many application fields, such as failure analysis in the semiconductor industry, large-volume 3D tomography with EDS and EBSD analysis for metals, ceramics, rock samples, and various materials in the material science field. Some materials and samples exhibit poor cross section quality with some FIB induced artifacts (terrace/rippling, curtaining). Such artifacts are more pronounced on plasma FIB instruments due to their high probe current. Suppression methods for such artifacts become more important to utilize plasma FIB-SEM systems. However, there are few reports in domestic journals describing characteristics of these artifacts from the viewpoint of high-current milling. Therefore, this article introduces FIB induced artifacts from the viewpoint of the forming process and some techniques for high current artifact-free milling by plasma FIB.

In the field of electron microscopy, automation has gained attention, emerging as a key topic in conferences. Automation levels vary by context. The paper categorizes required automation styles in semiconductor fabrication lines, academia, and analysis centers. After that, it focuses on high level automation of TEM analysis in semiconductor fabrication lines. This automation requires high success rates using AI-based high robustness solutions. Workflow-wise, managing sample flow and data with high traceability is essential. It includes standardizing Lamella Carrier (LC) and LC Container (LCC) with unique ID codes of SEMI standards.

Convergent-beam electron diffraction (CBED) using nanoelectron probes is used for symmetry determination of local areas and quantitative crystal structure analysis (QCBED) based on the dynamical diffraction theory. The development of fast and sensitive two-dimensional pixelated electron detectors has led to the rapid development of 4D-STEM, which combines the CBED method with scanning transmission electron microscopy, allowing the analysis of non-uniform structures such as interfaces. Structural analyses of twin boundaries of CaTiO3 and BaTiO3 using 4D-STEM and an approach of dynamical diffraction calculations toward quantitative structural analysis using 4D-STEM are presented.

In situ hybridization (ISH) is a robust technique employed to detect specific RNA molecules within cells. While conventional ISH methods utilizing hapten-labeled probes are effective in detecting multiple RNA targets, the detection process remains complex and required longer time. To address this, we present a novel application of fluorescence resonance energy transfer (FRET)-based molecular beacon (MB) probes for ISH. MCF-7 cells and C57BL/6J mouse uterus were subjected to ISH experiments. MB probes targeting ERα mRNA and 28S rRNA were labeled with Cy3/BHQ-2 and 6-FAM/DABCYL, respectively. Conventional probes were labeled with digoxigenin. In MCF-7 cells, 28S rRNA exhibited distinct signals in the nucleolus and cytoplasm of all cells, while ERα mRNA signals were observed in some nucleoli. In the uterus, complementary MB probes successfully detected 28S rRNA, whereas negative control slides exhibited no signals. Furthermore, 28S rRNA was detected in all cells, whereas ERα mRNA was predominantly detected in the epithelium. Notably, fluorescence intensity of 28S rRNA significantly decreased when subjected to 1 or 2 base-mismatched sequences, indicating highly specific target RNA detection. In summary, FRET-based MB probes offer valuable advantages in ISH, including expedited hybridization kinetics, high sensitivity and specificity.

Serial semi-thin sections are useful for sample preparation for array tomography and CLEM observation, but they have the problem of frequent wrinkling when attached to substrates such as glass slides and silicon wafers by pulling up a diagonally placed substrate or by dropping water levels. In the case of ultra-thin sections, this method rarely causes wrinkling, but in the case of semi-thin sections, wrinkling is thought to occur due to insufficient stretching of the sections. In this research, we developed a new device that transfers a large number of serial semi-thin sections floating on water from a diamond knife with a large boat while retaining the water necessary for stretching onto a hot plate at once, and heats and extends the sections while keeping them in place. A glass slide with a water-repellent frame is placed on the device, and placed horizontally on a knife boat and sunk in water. After sectioning, the floating sections in the frame are lifted vertically by the device and can be moved directly onto the hotplate. This method enables to obtain a wide range of sequential tissue images without wrinkles and a detailed three-dimensional analysis of the distribution of intracellular organelles and cellular structures.

Quantum electron microscopy is an emerging field of study at the boundary between electron microscopy and quantum information science. While quantum technologies such as quantum computing have increasingly been discussed recently, it is generally hard to find truly useful applications in the near future, as we explain in the main text. We argue that quantum electron microscopy could be an exception to this rule and it would give us tangible quantum advantage. Specifically, we expect to obtain an increased amount of information from radiation-sensitive specimens, that are often important in fields such as structural biology and soft materials science.