An automated hologram acquisition system for big-data analysis and for improving the statistical precision of phase analysis has been upgraded with automated particle detection technology. The coordinates of objects in low-magnification images are automatically detected using zero-mean normalized cross-correlation with preselected reference images. In contrast with the conventional scanning acquisitions

AbstractThe lamellar structure of bone, which endows biomechanical rigidity to support the host organism is observed in mammals, including humans. It is therefore essential to develop a quantitative analysis to evaluate the lamellarity of bone, which would especially useful for the pharmacological evaluation of anti-osteoporotic drugs. This study applied a current system for the semi-automatic recognition

We developed a gas environmental heating specimen holder system by applying differential pumping effect to a specimen holder for the in-situ transmission electron microscopy observation and electron energy loss spectroscopy (EELS) analysis of catalytic materials. In the in-situ experiments, using two small orifices and O-rings, the maximum formed gas pressure was ~20 Pa. Also, using a heater membrane

Until now, studies on nail disease have been performed through microscopic diagnosis and microscopic computed tomography (micro-CT). However, these kinds of conventional methods have some limitations. Firstly, the microscopic method is considered the gold standard for medical diagnosis. However, due to the use of fluorescent materials, the sample is damaged and it takes a long time to get results.

The increasing popularity and adoption rate of cryo-electron microscopy is evidenced by a growing number of new microscope installations around the world. The quality and reliability of the instruments improved dramatically in recent years, but site-specific issues or unnoticed problems during installation could undermine productivity. Newcomers to the field may also have limited experience and/or

The stability of β-precipitates in the Zr-1Nb alloy has been studied under Ne ion irradiation of energy 250 keV by in-situ Transmission Electron Microscope (TEM) as a function of irradiation dose. The irradiation was carried out up to ~136 dpa at 573 K. Microstructural investigations have shown that up to ~38 dpa, precipitates showed an increase in size and for irradiation dose higher than 38 dpa,

Microscopy, 2021, 1–5. doi: 10.1093/jmicro/dfaa078

Operando observation using spin-polarized scanning electron microscopy (spin SEM) has been demonstrated by detecting changes in the magnetization in the writing head of a hard disk drive (HDD) during operation. A current-applying system developed for use in the sample stage of a spin SEM enables imaging of the magnetization changes in the writing head of an HDD while the writing head is activated.

Scanning ion conductance microscopy (SICM) is useful for imaging soft and fragile biological samples in liquids because it probes the samples’ surface topography by detecting ion currents under non-contact and force-free conditions. SICM acquires the surface topographical height by detecting the ion current reduction that occurs when an electrolyte-filled glass nanopipette approaches the sample surface

In transmission electron microscope (TEM), both the amplitude and phase of electron beam change when electrons traverse a specimen. The amplitude is easily obtained by the square root of the intensity of a TEM image, while the phase affects defocused images. In order to obtain the phase map and verify the theoretical model of the interaction between electron beam and specimen, a lot of simulations

Methodology for quantitative evaluation of electron radiation damage and calculation of tolerable electron dose was developed to achieve damage-less scanning electron microscope (SEM) observation of beam sensitive polymer film. The radiation damage is typically evaluated with visual impressions of SEM images, however, this method may be unreliable because observer’s subjectivity may affect the results

Correlative light and electron microscopy (CLEM) is an excellent approach for examining the cellular localization of biomolecules. Here, we developed a simple method for CLEM by combining pre-embedding immunohistochemistry with a novel fluorescent probe, namely Fluolid NS Orange, and an embedding resin called ʻDurcupan™ʼ. Specimens were embedded in Durcupan™ or LR White after immunolabeling and post-fixation

Reversibly photoswitchable fluorescent proteins (RSFPs) are a class of fluorescent proteins whose fluorescence can be turned on and off by light irradiation. RSFPs have become essential tools for super-resolution (SR) imaging. Because most SR imaging techniques require high power density illumination, mitigating phototoxicity in cells due to intense light irradiation has been a challenge. Although

The microstructure, chemical composition and mechanical strength of heterogeneous materials of mantis shrimp (Oratosquilla oratoria) saddle were studied. As the key component of the striking system, the saddle was comprised of two distinct layers including outer layer and inner layer. Outer layer contained blocky microtubules and exhibited compact appearance. The inner layer presented a typical periodic

This paper presents a quick and straightforward method to evaluate image distortion in scanning electron microscopy (SEM) using a certified reference material (CRM) as a test specimen. The CRM has a square dot-array structure, whose dot-pitch has an accredited value. By calculating the distance between each dot of the CRM via image analysis, we can detect the distortion in the image as variations of

Nickel nanocrystals have received much attention for their ferromagnetic properties. The crystal properties are strongly dependent on their facets and therefore detailed study of their morphology, facets and orientation is critical for magnetic applications. In this work, equilibrium crystal shapes of self-assembled nickel nanocrystals on the (111) termination of strontium titanate (SrTiO3) at room

The measurement of the volume of blood cells is important for clinical diagnosis and patient management. While digital holography microscopy (DHM) has been used to obtain such information, previous off-axis setups usually involve a separated reference beam and are thus not very easy to implement. Here, we use the simple in-line Gabor setup without separation of a reference beam to measure the shape

The structure analysis of amorphous materials still leaves much room for improvement. Owing to the lack of translational or rotational symmetry of amorphous materials, it is important to develop a different approach from that used for crystals for the structure analysis of amorphous materials. Here, the angstrom-beam electron diffraction method was used to obtain the local structure information of

The performance of a direct electron detector using silicon-on-insulator (SOI) technology in a low-voltage transmission electron microscope (LVTEM) is evaluated. The modulation transfer function and detective quantum efficiency of the detector are measured under backside illumination. The SOI-type detector is demonstrated to have high sensitivity and high efficiency for the direct detection of low-energy

We have previously indicated that a single injection of alendronate, one of the nitrogen-containing bisphosphonates (NBPs), affects murine hematopoietic processes, such as the shift of erythropoiesis from bone marrow (BM) to spleen, disappearance of BM-resident macrophages, the increase of granulopoiesis in BM and an increase in the number of osteoclasts. NBPs induce apoptosis and the formation of

AbstractIn-liquid frequency modulation atomic force microscopy (FM-AFM) has been used for visualizing subnanometer-scale surface structures of minerals, organic thin films and biological systems. In addition, three-dimensional atomic force microscopy (3D-AFM) has been developed by combining it with a three-dimensional (3D) tip scanning method. This method enabled the visualization of 3D distributions

AbstractUltra-high voltage electron microscopy is useful for research utilizing high-penetration thickness of electron beam, in situ observation, or irradiation effects by the particle characteristics of electrons. In this review, the importance of non-equilibrium materials science research by a combination with irradiation effects and in situ observation is shown, and examples of some research are

AbstractWe have developed an amplitude-division type Mach-Zehnder electron interferometer (MZ-EI). The developed MZ-EI is composed of single crystals corresponding to amplitude-division beam splitters, lenses corresponding to mirrors and an objective aperture. The spacings and azimuth angles of interference fringes can be controlled by single crystal materials and their orientations and by diffraction

AbstractThe dynamics of cerebrospinal fluid (CSF) are essential for maintaining homeostasis in the central nervous system. Despite insufficiently detailed descriptions of their structural and molecular properties for a century, cranial arachnoid granulations (CAGs) on meninges have been thought to participate in draining CSF from the subarachnoid space into the dural sinuses. However, recent studies

AbstractIn this study, a method was developed for examining the distribution pattern of inorganic substances dispersed in hydrogel-filled medical devices. Transmission electron microscopy (TEM) using ultra-thin cryosectioning (owing to hydrogel’s water content) was performed on contact lenses with an iris pattern in which the distribution pattern of inorganic pigments was problematic. We confirmed

AbstractPancreatic islet microcirculation, consisting of pancreatic islet microvascular endothelial cells (IMECs) and pericytes (IMPCs), provides crucial support for the physiological function of pancreatic islet. Emerging evidence suggests that pancreatic islet microcirculation is impaired in type 1 diabetes mellitus (T1DM). Here, we investigated the potential ultrastructural protective effects of

AbstractSherds of ancient ceramics potteries, dating back to the seventh century BC and excavated from southern Italy, were analyzed in terms of microstructure and porosity (SEM), elemental composition (EDX), chemical bonding and mineralogical components (FTIR) in order to establish correlation with firing temperature, firing condition and provenance. Si/Al ratio from EDX analysis showed three major

AbstractHere, we describe a method for tracking intracellular processing of small interfering RNA (siRNA) containing complexes using automated microscopy controls and image acquisition to minimize user effort and time. This technique uses fluorescence colocalization to monitor dual-labeled fluorescent siRNAs delivered by silica nanoparticles in different intracellular locations, including the early/late

AbstractWe have developed a new certified reference material (CRM) for image sharpness evaluation and magnification calibration for scanning electron microscopy (SEM). Designed to be suitable for the image sharpness evaluation by the derivative method, the CRM has nanoscale tungsten dot-array structure fabricated on silicon substrate, which gives steep contrast transition from the dot to the substrate

AbstractUnroofing, which is the mechanical shearing of a cell to expose the cytoplasmic surface of the cell membrane, is a unique preparation method that allows membrane cytoskeletons to be observed by cryo-electron microscopy, atomic force microscopy, freeze-etching electron microscopy and other methods. Ultrasound and adhesion have been known to mechanically unroof cells. In this study, unroofing

A phase imaging method that measures the phase shift existing at a dislocation’s core is described. The method uses the interference of two symmetrically diffracted beams on the optic axis by means of an electron biprism. Each diffracted beam carries half the phase of the dislocation core. When combined, the entire phase shift of the dislocation core is obtained.

Multiciliogenesis is a cascading process for generating hundreds of motile cilia in single cells. In vertebrates, this process has been investigated in the ependyma of brain ventricles and the ciliated epithelia of the airway and oviduct. Although the early steps to amplify centrioles have been characterized in molecular detail, subsequent steps to establish multicilia have been relatively overlooked

AbstractCorrelative light and electron microscopy (CLEM) methods combined with live imaging can be applied to understand the dynamics of organelles. Although recent advances in cell biology and light microscopy have helped in visualizing the details of organelle activities, observing their ultrastructure or organization of surrounding microenvironments is a challenging task. Therefore, CLEM, which

We have been using sandwich freezing of living yeast and bacteria followed by freeze-substitution for observing close-to-native ultrastructure of cells. Recently, sandwich freezing of glutaraldehyde-fixed cultured cells and human tissues have been found to give excellent preservation of ultrastructure of cells and tissues. These studies, however, have been conducted using a handmade sandwich freezing

In this paper, we review the current state of phase plate imaging in a transmission electron microscope. We focus especially on the hole-free phase plate design, also referred to as the Volta phase plate. We discuss the implementation, operating principles and applications of phase plate imaging. We provide an imaging theory that accounts for inelastic scattering in both the sample and in the hole-free

Escherichia coli chaperonin GroEL, which is a large cylindrical protein complex comprising two heptameric rings with cavities of 4.5 nm each in the center, assists in intracellular protein folding with the aid of GroES and adenosine triphosphate (ATP). Here, we investigated the possibility that GroEL can also encapsulate metal nanoparticles (NPs) up to ∼5 nm in diameter into the cavities with the aid

Differential-phase-contrast scanning transmission electron microscopy (DPC STEM) is a technique to directly visualize local electromagnetic field distribution inside materials and devices at very high spatial resolution. Owing to the recent progress in the development of high-speed segmented and pixelated detectors, DPC STEM now constitutes one of the major imaging modes in modern aberration-corrected

AbstractG-protein-coupled receptors (GPCRs) comprise the largest and most diverse cell surface receptor family, with more than 800 known GPCRs identified in the human genome. Binding of an extracellular cue to a GPCR results in intracellular G protein activation, after which a sequence of events, can be amplified and optimized by selective binding partners and downstream effectors in spatially discrete

AbstractDifferential phase contrast (DPC) imaging in scanning transmission electron microscopy is a technique to visualize electromagnetic field distribution inside specimens at high spatial resolution. However, diffraction contrast strongly hampers electromagnetic contrast in DPC images especially in polycrystalline samples. In this paper, we develop an imaging technique to effectively suppress diffraction

AbstractElectron staining is generally performed prior to observing organic materials via transmission electron microscopy (TEM) to enhance image contrast. However, electron staining can deteriorate organic materials. Here, we demonstrate electrostatic potential imaging of organic materials via differential phase contrast (DPC) scanning transmission electron microscopy (STEM) without electron staining

AbstractA scanning electron microscope transition edge sensor has been developed to analyze the minor or trace constituents contained in a bulk sample and small particles on the sample under a low accelerating voltage (typically <3 keV). The low accelerating voltage enables to improve the spatial analysis resolution because the primary electron diffusion length is limited around the sample surface

AbstractConventional imaging for three-dimensional (3D) ultra-architectural analysis of collagen fibers and fibroblasts is time-consuming and requires numerous ultrathin sections to search the target area. Currently, no method allows 3D ultra-architectural analysis of predetermined areas including spatial relationships between collagen fibers and fibroblasts in vitro. Herein, we developed a new method

AbstractY-branched or side-by-side-branched carbon microtubes with metal filler material were fabricated, and material transport in the branched microtubes with Joule heating was investigated using in situ scanning electron microscopy with micro-electrode probes. When a voltage and electric current were applied, the material enclosed in the microtubes moved from its original position. The movement

AbstractThe advent of high-flux, high-brilliance synchrotron radiation (SR) has prompted the development of high-resolution X-ray imaging techniques such as full-field microscopy, holography, coherent diffraction imaging and ptychography. These techniques have strong potential to establish non-destructive three- and four-dimensional nano-imaging when combined with computed tomography (CT), called nano-tomography

AbstractSome species of lactic acid bacteria used for the production of natural cheese produce exopolysaccharides (EPS). Electron microscopy is useful for analyzing the microstructure of EPS produced by lactic acid bacteria. However, pretreatments used to observe the microstructure of EPS by electron microscopy, such as dehydration and resin embedding, can result in EPS flowing out easily from the

AbstractHigh refractive index organic solvents are commonly used as an imaging medium in tissue clearing approaches. While effective, such solvents provide serious concerns for the safety of users and the equipment, especially in a central microscopy unit. To overcome these concerns, we have developed a large and reusable imaging chamber compatible with the universal mounting frame AK (PeCon GmbH)

AbstractIn electron diffractive imaging, the phase image of a sample is reconstructed from its diffraction intensity through iterative calculations. The principle of this method is based on the Fourier transform relation between the real-space wave field transmitted by the sample and its Fraunhofer diffraction wave field. Since Gerchberg’s experimental work in 1972, various advancements have been achieved

Important breakthroughs in far-field imaging techniques have been made since the first demonstrations of stimulated emission depletion (STED) microscopy. To date, the most straightforward and widespread deployment of STED microscopy has used continuous wave (CW) laser beams for both the excitation and depletion of fluorescence emission. A major drawback of the CW STED imaging technique has been photobleaching

AbstractAlthough the possibility of locating single atom in three dimensions using the scanning transmission electron microscope (STEM) has been discussed with the advent of aberration correction technology, it is still a big challenge. In this report we have developed deconvolution routines based on maximum entropy method (MEM) and Richardson–Lucy algorithm (RLA), which are applicable to the STEM-annular

Phase-shifting electron holography (PS-EH) is an interference transmission electron microscopy technique that accurately visualizes potential distributions in functional materials, such as semiconductors. In this paper, we briefly introduce the features of the PS-EH that overcome some of the issues facing the conventional EH based on Fourier transformation. Then, we present a high-precision PS-EH technique

In this study, we conducted a quantitative evaluation of dislocation density by scanning electron microscopy electron channeling contrast imaging for α grains of a Ti-6Al-4V alloy deformed at room temperature. The depth of visibility of dislocations is experimentally measured as 140 to 160 nm by a serial sectioning observation. This result is compared with the theoretical value and applied to evaluate

Senile lentigo or age spots are hyperpigmented macules of skin that commonly develop following long-term exposure to ultraviolet radiation. This condition is caused by accumulation of large numbers of melanosomes (melanin granules) produced by melanocytes within neighboring keratinocytes. However, there is still no consensus regarding the melanosome transfer mechanism in senile lentigo. To date, most

AbstractElectron holography was invented for correcting aberrations of the lenses of electron microscopes. It was used to observe the atomic arrangements in crystals after decades of research. Then it was combined with a hardware aberration corrector to enable high-resolution and high-precision analysis. Its applications were further extended to magnetic observations with sub-nanometer resolution.

To visualize the fine structure of compacted DNA of Synechococcus elongatus PCC 7942, which appears at a specific time in the regular light/dark cycle prior to cell division, ChromEM with some modifications was applied. After staining DNA with DRAQ5, the cells were fixed and irradiated by red laser in the presence of 3,3ʹ-diaminobenzidine and subsequently fixed with OsO4. A system with He–Ne laser

In this study, a noise-reduction technique for series low-dose electron holograms using tensor decomposition is demonstrated through simulation. We treated an entire dataset of the series holograms with Poisson noise as a third-order tensor, which is a stack of 2D holograms. The third-order tensor, which is decomposed into a core tensor and three factor matrices, is approximated as a lower-rank tensor

AbstractWe have designed and evaluated a cryo-electron microscopy (cryo-EM) system for higher-resolution single particle analysis and high-precision electron 3D crystallography. The system comprises a JEOL CRYO ARM 300 electron microscope—the first machine of this model—and a direct detection device camera, a scintillator-coupled camera, GPU clusters connected with a camera control computer and software

AbstractA phase retrieval technique based on a transport of intensity equation (TIE) is one of the defocus series reconstruction techniques in microscopy. Since it does not require any dedicated devices like a biprism, and only three defocus images are enough to retrieve phase information, it has been applied to observe magnetic fields, magnetic domains, electrostatic potentials and strains. It is

Microstructural changes during the martensitic transformation from face-centred cubic (FCC) to body-centred cubic (BCC) in an Fe-31Ni alloy were observed by scanning electron microscopy (SEM) with a newly developed Peltier stage available at temperatures to −75°C. Electron channelling contrast imaging (ECCI) was utilized for the in situ observation during cooling. Electron backscatter diffraction analysis

AbstractPtychography is a coherent diffractive imaging technique that can determine how an electron wave is transmitted through an object by probing it in many small overlapping regions and processing the diffraction data obtained at each point. The resulting electron transmission model describes both phase and amplitude changes to the electron wave. Ptychography has been adopted in transmission electron

We describe small-angle electron diffraction (SmAED) and Lorentz microscopy using a conventional transmission electron microscope. In SmAED, electron diffraction patterns with a wide-angular range on the order of 1 × 10−2 rad to 1 × 10−7 rad can be obtained. It is demonstrated that magnetic information of nanoscale magnetic microstructures can be obtained by Fresnel imaging, Foucault imaging and SmAED