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

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

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)

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

Electron 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. High-resolution

To visualize the fine structure of compacted DNA in Synechococcus elongatus PCC 7942, which appears at a specific time in the regular light/dark cycle prior to cell division, Chrom EM (Ou et al., 2017) with some modifications was applied. After staining DNA with DRAQ5, the cells were fixed and irradiated by red laser in the presence of DAB, and subsequently fixed with OsO4. A system with He-Ne laser

Ptychography 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 microscopy

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 two-dimensional holograms. The third-order tensor, which is decomposed into a core tensor and three factor matrices, is approximated as a lower-rank

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

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

This study was aimed to verify the cellular interplay between vascular endothelial cells and surrounding cells in the chondro-osseous junction of murine tibiae. Many CD31-positive endothelial cells accompanied with Dolichos Biflorus Agglutinin lectin-positive septoclasts invaded into the hypertrophic zone of the tibial epiphyseal cartilage. MMP9 immunoreactive cytoplasmic processes of vascular endothelial

This study aimed to reproduce the characteristics of original adenocarcinoma tumors in vitro. The pancreatic cell line, SUIT-58, derived from a moderately differentiated adenocarcinoma of metastatic pancreatic cancer was used. The cells have a sheet structure in conventional cell culture without forming glands or exhibiting mucin production in the lumen. First, the necessity of scaffolds to create

Since the majority of a tendon’s dry weight is collagen fibers, tendon healing consists mainly of collagen repair and observing three-dimensional networks of collagen fibers with scanning electron microscopy (SEM) is optimal for investigating this process. In this report, a cell-maceration/SEM method was used to investigate extrasynovial- tendon (unwrapped tendon in synovial tissue such as the tendon

We investigate the effect that recording off-axis electron holograms on pixelated detectors, such as charge-coupled devices (CCD) and direct-detection devices (DDD), can have on measured amplitudes and phases. Theory will be developed for the case of perfectly uniform interference fringes illuminating an imperfect detector with gain variations and pixel displacements. We will show that both these types

Accuracy of electron density distribution analysis using large-angle convergent-beam electron diffraction (LACBED) patterns is evaluated for different convergence angles. An orbital ordered state of FeCr2O4 is used as an example of the analysis. Ideal orbital-ordered and non-ordered states are simulated by using orbital scattering factors. LACBED patterns calculated for the orbital-ordered state were

This paper reviews the basics of electron holography as an introduction of the holography part of this special issue in Microscopy. We discuss the general principle of holography and interferometry regarding measurements and analyses of phase distributions, first using the optical holography. Next, we discuss physical phenomena peculiar to electron waves that cannot be realized by light waves and principles

This review examines methods of magnetic flux density measurements from the narrow grain boundary (GB) regions, the thickness of which is of the order of nanometers, produced in Nd-Fe-B-based sintered magnets. Despite of the complex crystallographic microstructure and the significant stray magnetic field of the sintered magnet, recent progress in electron holography allowed for determination of the

The aquatic carnivorous plant Aldrovanda vesiculosa L. is critically endangered worldwide; its peculiar lifestyle raises many questions and poses problems both intriguing on their own and relevant to conservation. While establishing a culture system for its propagation and restoring its natural habitat in Hozoji pond in Saitama, Japan, we conducted ultrastructural observations to examine the various

The extended Rayleigh resolution measure was introduced to give a generalized resolution measure that can be readily applied to imaging and resolving particles that have finite size. Here, we make a detailed analysis of the influence of the particle size on this resolution measure. We apply this to scanning electron microscopy, under simple assumption of a Gaussian electron beam intensity distribution

Materials characterization is indispensable for materials development. In particular, spectroscopy provides atomic configuration, chemical bonding and vibrational information, which are crucial for understanding the mechanism underlying the functions of a material. Despite its importance, the interpretation of spectra using human-driven methods, such as manual comparison of experimental spectra with

We try to improve the limit of the phase estimation of the interference fringe at low electron dose levels in electron holography by a noise reduction method. In this paper, we focus on unsupervised approaches to apply it to electron beam-sensitive and unknown samples and describe an overview of denoising methods used widely in image processing, such as wiener filter, total variation denoising, nonlocal

The combination of scanning transmission electron microscopy (STEM) with analytical instruments has become one of the most indispensable analytical tools in materials science. A set of microscopic image/spectral intensities collected from many sampling points in a region of interest, in which multiple physical/chemical components may be spatially and spectrally entangled, could be expected to be a

Image processing is one of the most important applications of recent machine learning (ML) technologies. Convolutional neural networks (CNNs), a popular deep learning-based ML architecture, have been developed for image processing applications. However, the application of ML to microscopic images is limited as microscopic images are often 3D/4D, that is, the image sizes can be very large, and the images

Significant inventions about electron microscopes have been attained in the last two decades. One of the most important inventions is of the aberration-correction techniques, which continue to spread over research laboratories in the world. Thanks to the picometer-sized electron probe produced by the aberration correction, for example, the precision in crystal-structure analysis and/or chemical mapping

An automated acquisition system for collecting a large number of electron holograms, to improve the statistical precision of phase analysis, was developed. A technique for shifting the electron beam in combination with stage movement allows data to be acquired over a wide area of a TEM-specimen grid. Undesired drift in the hologram position, which may occur during the hologram acquisition, can be corrected

In this review, we focus on the applications of machine learning methods for analyzing image data acquired in imaging flow cytometry technologies. We propose that the analysis approaches can be categorized into two groups based on the type of data, raw imaging signals or features explicitly extracted from images, being analyzed by a trained model. We hope that this categorization is helpful for understanding

Single-molecule imaging analysis has been applied to study the dynamics and kinetics of molecular behaviors and interactions in living cells. In spite of its high potential as a technique to investigate the molecular mechanisms of cellular phenomena, single-molecule imaging analysis has not been extended to a large scale of molecules in cells due to the low measurement throughput as well as required

An ionic liquid (IL) is a salt consisting of only cations and anions, which exists in the liquid state at room temperature. Interestingly ILs combine various favorable physicochemical properties, such as negligible vapor pressure, flame resistance, relatively high ionic conductivity, wide electrochemical window, etc. To take advantage of two specific features of ILs, viz. their nonvolatile and antistatic

Dendritic spines are small protrusions that receive most of the excitatory inputs to the pyramidal neurons in the neocortex and the hippocampus. Excitatory neural circuits in the neocortex and hippocampus are important for experience-dependent changes in brain functions, including postnatal sensory refinement and memory formation. Several lines of evidence indicate that synaptic efficacy is correlated

The temporal resolution in scanning transmission electron microscopy (STEM) is limited by the scanning system of an electron probe, leading to only a few frames per second (fps) at most in the current microscopes. To push the boundary of atomic-resolution STEM imaging into dynamic observations, an unprecedentedly faster scanning system combined with fast electron detection systems should be a prerequisite

Stacking faults are easily formed in silicon carbide (SiC) crystals, and this is also the case for SiC nanowires. The stacking faults exercise influences on SiC’s properties, therefore it is important to understand their formation mechanism and to control their formation for applications of SiC and its nanowires. In this study, we propose a method for investigating stacking faults’ formation mechanism

Lensless Fourier transform holography has been developed. By treating Bragg diffraction waves as object waves and a transmitted spherical wave as a reference wave, these two waves are interfered and recorded as holograms away from the reciprocal plane. In this method, reconstruction of holograms requires only one Fourier transform. Application of this method to analyze vortex beams worked well and

Investigation of solid electrolyte interphases (SEIs) on negative electrode surfaces is essential to improve the stable charge-discharge performance of rechargeable lithium-air batteries (Li-O2 batteries). In this study, a direct investigation of SEI films is conducted using analytical transmission electron microscopy (TEM). A thin Cu specimen is prefabricated for TEM observation and is utilised as

Various protein-labeling methods based on the specific interactions between genetically encoded tags and synthetic probes have been proposed to complement fluorescent protein-based labeling. In particular, labeling methods based on enzyme reactions have been intensively developed by taking advantage of the highly specific interactions between enzymes and their substrates. In this approach, the peptides

Transmission electron microscopy (TEM) and scanning transmission electron microscopy (STEM) enable the visualization of three-dimensional (3D) microstructures ranging from atomic to micrometer scales using 3D reconstruction techniques based on computed tomography algorithms. This 3D microscopy method is called electron tomography (ET) and has been utilized in the fields of materials science and engineering

Image contrast between carbon nanotubes (CNTs) and polytetrafluoroethylene (PTFE) in a CNT/PTFE composite film, which is difficult to obtain by conventional backscattered electron (BSE) imaging, was optimized to better elucidate the distribution of CNT in the film. Ultra-low landing energy condition (0.3 keV in this study) was used to prevent specimen damage due to electron beam irradiation. Signal

The theoretical conditions for small-angle inelastic scattering where the incident electron can effectively be treated as a particle moving in a uniform potential is examined. The motivation for this work is the recent development of a multislice method that combines plasmon energy losses with elastic scattering using Monte Carlo methods. Since plasmon excitation is delocalized, it was assumed that