bluuebunny/crossref_metadata_2025 huggingface dataset:
Critical Role of EMPA in Discovery of a New Lunar Mineral
Authors: Snow M G , Vaniman D T
Published: July 2020

Natural rock-forming minerals and many synthetic materials have traditionally been characterized by electron microprobe analysis (EMPA). However, recent advances in technology have created attractive alternatives to EMPA that focus instead on bulk chemistry and high-resolution surface studies. The purpose of this abstract is to demonstrate the continued usefulness of careful electron microprobe analysis as part of a well-coordinated analytical study. To this end, a brief summary is presented of the role the electron microprobe played in the recent discovery of yoshiokaite, a new Ca, Al-silicate mineral and one of only three new minerals discovered on the Moon.[l]

Regolith breccias returned by several Apollo missions were subjected to extensive bulk chemical studies at the University of California, Los A n geles.[2] These bulk chemical studies focused particularly on key trace elements that might identify exotic regolith breccias.
The Effect of MASS in Electron-Solid Interactions and the Mystery of the “Heinrich Kink”
Authors: Donovan JJ , Pingitore NE
Published: July 2020

Is average atomic mass or average atomic number more appropriate to describe backscattered electron (BSE) effects in electron-solid interactions? In electron microprobe literature, either one or both of the terms A and Z may be found utilized in various theoretical models to describe electron backscattering. Physical theory would seem to favor atomic number averaging, because backscattering ultimately represents an electromagnetic interaction between the electron and the nucleus. At this scale, mass and its associated property of gravity, is 10 times less powerful than the electromagnetic force. Nonetheless, some empirical results show a strong relation between atomic mass and electron scattering, thus endorsing mass averaging.

The periodic table presents an opportunity for distinguishing between these two influences: those pairs of elements for which an increase in Z yields a decrease in average A (e.g., Ar-39.95 and K-39.10; Co-58.93 and Ni-58.71; Te-127.60 and 1-126.90) due to natural isotope abundances.
Refining Equipment for High Resolution in-Lens Cryo-Sem
Authors: Apkarian Robert P , Lee Stephen , Keiper Jason
Published: July 2020

Advanced metal coating methods have greatly improved the localization of secondary electrons (SEI) from biological specimens staged in an in-lens field emission SEM. Using these metal coatings, molecular level structural information from cell membranes and isolated bio-molecules have been more accurately recorded from frozen-hydrated samples due to the presence of bound water rather than from chemically fixed and dried samples. Although in-lens cryo-SEM systems have provided images with equivalent resolution to TEM-replicas, several conditions are encountered which require optimizing techniques and hardware.

A drop of hydrated Bakers yeast on a gold Balzers carrier was plunge-frozen in ethane at its melting point. Thermal properties of ethane, such as the large difference between melting and boiling points, and its low viscosity increase the heat transfer coefficient during plunging thereby leading to faster cooling of the sample than with other cryogens. Carriers were transferred to the cryo-workstation of the Oxford CT 3500 system.
Design and Construction of a Quantitative Uniaxial Straining Stage For The Environmental SEM
Authors: Mansfield John F , Thouless Michael D , Stefano Justin A , Holbrook Judd
Published: July 2020

The environmental scanning electron microscope (or Environmental SEM) is a powerful tool in the materials science laboratory. The large sample chamber and ability to operate in a very poor vacuum (>5Torr) makes it an ideal device for a wide variety of in-situ studies. A three point bending stage has been used in the ElectroScan E3 environmental SEM at the University of Michigan for over four years. The stage is extremely flexible and can be used to view samples in both planar and transverse views. It also allows for quantitative recording of the stress and strain exerted on the sample via a load cell and a linear variable differential transducer. The output s of the load cell are acquired into an Apple Macintosh computer via a National Instruments analog-to-digital acquisition card. To extend the capability of EMAL's in-situ tensile testing laboratory, a uniaxial tensile stage has been constructed in collaboration with the Department of Mechanical Engineering and Applied Mechanics.
In Situ Mechanical Testing of Dry and Hydrated Cellular Materials in the Environmental Scanning Electron Microscope (ESEM)
Authors: Stokes DJ , Donald AM
Published: July 2020

We present a method for directly observing and quantifying the mechanical deformation of cellular foods, such as breadcrumb and Chinese water chestnut, with the aim of identifying the key differences in the nature of cell wall failure and crack propagation. These factors are of importance to the food industry, and this work may enhance our understanding of the changes in behaviour of crisp cellular foods when subjected to a moist environment.

Breadcrumb coatings are commonly applied to a core substrate such as meat, poultry, fish and vegetables. After flash-frying the product is often frozen, but during storage there is a tendency for the coating to absorb water from the substrate. The driving force for this process is the large difference in water activity level aw between the two materials. Incorrect reconstitution of the product, for example by microwave cooking, leads to a significant loss of crispness.
Investigation of Medium to High Strain Deformation Microstructures Using an Automated Electron Backscattering Pattern (EBSP) System
Authors: Godfrey A , Krieger Lassen N C , Hughes D A , Juul Jensen D
Published: July 2020

The speed of automated EBSP orientation gathering has now increased to a point where it is possible to examine large areas of deformed material at a resolution close to the length scale of the dislocation substructures involved (typically <1μm for strains of greater than ∊= l). Investigation of such deformed samples invariably gives rise to diffraction images of low quality and to many double patterns. To assist in a critical selection of the data, it is usual to store a measure of the indexing confidence for each point. Here we describe how deformed samples, which provide the appropriate range of pattern qualities, can be used to calibrate acceptance criteria for unsupervised EBSP measurements on deformation microstructures. An application in the use of such criteria is also described.

Two crystals, chosen with orientations that are known to develop a well defined texture spread during deformation, were channel die deformed to a strain of ∊=1.5 (78% reduction).
Esem Study of Film Formation in Latices Polymerised in Presence of Starch
Authors: Stelmashenko N , Donald AM
Published: July 2020

The Environmental Scanning Electron Microscope (ESEM) was used to study film-formation in vinyl latices polymerised in the presence of varying amount and type of starch. An understanding of the effect of native biopolymers on film forming mechanisms in latex has technological and industrial relevance because it can lead to the creation of novel latices with potentially lower cost and improved biosustainability.

Film formation studies were carried out on a range of samples selected by ICI Paints as the most promising candidates for the novel latices. These included vinyl latices polymerised with 2 - 15% of potato and waxy maize starch, taken either in native or modified. Normally, no surfactants were used during latex polymerisation; a few samples were prepared for comparative purposes when polymerisation was carried out both with and without the addition of surfactant. Latex films were spun-cast on glass slides before insertion into the microscope.
Optimisation of the S.E. Signal to Background Ratio in ESEM
Authors: Keller TH , Thiel BL , Donald AM
Published: July 2020

We have performed a theoretical and experimental study of the signal composition in the Environmental SEM (ESEM) with the intention of forming a set of general guidelines for optimising the signal to background ratio. In the ElectroScan ESEM, a gas ionisation cascade is used to amplify the secondary electron signals emanating from the specimen surface. The presence of gas in the chamber also gives rise to a pressure dependent background signal derived from ionisation events between gas molecules and high energy primary beam and backscattered electrons, as well as secondary electrons generated by the probe skirt.

The signal collected by an environmental secondary detector (ESD) (ElectroScan, 1991) or a gaseous secondary detector (GSED) (ElectroScan, 1994) is an amplified signal which is a composite of at least three contributions. These are the amplified currents arising from the ionisation of the gas by high energy electrons from the primary (Ipe) and backscattered electrons (Ihse).
Using Secondary Electron Contrast for Imaging Water-Oil Emulsions in the Environmental SEM (ESEM)
Authors: Stokes DJ , Thiel BL , Donald AM
Published: July 2020

The ESEM is able to image liquids in their natural state, unlike conventional SEM, and offers significantly better resolution than optical microscopy. The ability to image binary liquid mixtures, such as water-oil emulsions, depends upon sufficient contrast for differentiation of the dispersed and continuous phases. We have made observations of such water-oil emulsions in an Electroscan 2010 Environmental SEM, and have duly obtained images with good contrast. Experiments were carried out on emulsions of vegetable oil-in-water, polybutene-in-water and water-in-polybutene, all emulsified with nonionic polysaccharide surfactants. Each sample was imaged at a temperature of approximately 3°C, in water vapour at a pressure of 4 torr, in order to maintain the liquid state.

From the ESEM micrographs in Figures 1, 2 and 3, it is clear that water exhibits a greater secondary electron emission than an unsaturated hydrocarbon.
Biometrology? Finding Biological, Pathological, Diagnostic Meaning From Critical Length-Scale Measurements
Authors: Allen MJ , Elings V B
Published: July 2020

The atomic force microscope (AFM) is now a fairly widely used metrological tool for measuring surfaces with nanometer-scale precision, in 3-D. A number of biologists have noticed that untreated biological cells and molecules deposited onto surfaces can also be analyzed by AFM in air or fluids provided the AFM imaging forces do not greatly excede the compressiblity of the specimen. What has gone largely unnoticed in the biological microscopy community is the fact that AFM provides a wealth of opportunity with regard to 3D, nanometer-scale morphometric and biochemical measurements of biomaterials, cells and molecules. The ability to obtain such precise length-scale measurements on biological structures does open up a whole new branch of biological microscopy- something we have termed, Biometrology. A broad definition of Biometrology is, the science of quantifying biological structures such that the measured quantities give rise to biological or diagnostic information.
Development of Polarization Modulation Near-Field Scanning Optical Microscope and its Application to Mapping Defect-Induced Birefringence in SrTiO3 Bicrystals
Authors: Hsu Julia W P , McDaniel E B , McClain S C
Published: July 2020

Photoelastic measurement is a sensitive optical technique to map strain fields in otherwise isotropic materials. To extend this method to the submicron scale, we combine dynamic polarimetry with nearfield scanning optical microscopy (NSOM) and construct a polarization modulation NSOM (PMNSOM). The 670 nm laser light passes first through a linear polarizer (oriented at 90°), and then through a photoelastic modulator (PEM), and finally through a quarter wave plate. The PEM introduces a sinusoidally time varying phase shift δ0sin(2πft) into the +45° polarization component, where the modulation frequency/is the resonant frequency (50 kHz) of the PEM quartz element. The quarter wave plate (oriented at 0°) transforms this elliptically polarized light into linearly polarized light with its orientation varying sinusoidally at the modulation frequency. This polarized light is then coupled into a single-mode optical fiber leading to the NSOM tip.
Magnetically-Oscillated Probe AFM for Imaging and Stiffness Measurements at the Liquid-Solid Interface
Authors: Lindsay SM , Han Wenhai , Tao Nongjian
Published: July 2020

The AFM is a natural tool for exploring liquid-solid interfaces, but oscillating probe methods are limited by the viscous action of the liquid on the probe. If the probe is driven acoustically, high driving amplitudes are required to obtain a response from the probe, and this can give rise to spurious responses. The problem is solved if the probe is driven directly, using a magnetic film on the tip and a solenoid to provide a driving field. This method, Magnetic A/C mode (MacMode) has provided the highest resolution in both images of biomolecules and in force spectroscopy at the liquid-solid interface.

The interaction force is reduced because the microscope can be operated at low amplitudes of oscillation (0.5 to 5nm) with small changes (0.1 to 0.5nm) in amplitude used for sensing. This often results in greatly improved resolution, presumably because small aperities on the tip are not damaged.
Comparative Surface Study of Atomic Images With Variable Temperature UHF-AFM and STM
Authors: Iwatsuki M , Suzuki K , Kitamura S , Kersker M
Published: July 2020

The AFM technique has made a remarkable progress recently. However, relatively few studies using the UHV-AFM have been reported compared to those using atmospheric AFMs or UHV-STMs. Atomic-resolution images of the clean sample surface in ultrahigh vacuum, which is relatively active and not observable in atmosphere, have been reported only few examples. The intense reaction between the tip of the cantilever and the sample in contact mode imaging, one of the techniques used in atomic imaging in atmosphere, often damages the surface of the sample, making it difficult to acquire atomic-resolution images in UHV.

In order to reduce the strong interaction between the tip and the sample, the noncontact imaging technique for the UHV-AFM (NC-AFM: JAFM-4500XT) has been developed. However, none of the atmospheric NC-AFMs has succeeded in achieving atomic level resolution.
The Surface Morphology and Characterisation of Electronic Properties of Boron Implanted Microwave Plasma CVD Diamond Films by Atomic Force and Scanning Tunneling Microscopies
Authors: Fitzgerald A G , Fan Y , John P , Troupe C E , Wilson J I B
Published: July 2020

The surface morphology and electronic properties of a low energy boron implanted diamond films with shallow doping, prepared by microwave plasma enhanced chemical vapour deposition (CVD), have been characterised by atomic force microscopy (AFM), scanning tunneling microscopy (STM) and scanning tunneling spectroscopy (STS) techniques.

Both AFM and STM images taken at different locations on the films have exhibited similar morphological features on the (100) crystal surfaces. The crystal surfaces are not atomically flat but are composed of many hillocks as shown in Fig 1(a) to 1(c). The majority of values measured from the peaks of hillocks to the valleys are in the range of 2 to 5 nm, and the diameter of these hillocks is in the range of 50 to 250 nanometers. These crystal surface morphological features are believed to be caused in the high energy boron ion implantation process.
Using AFM Phase Lag Data to Indentefy Microconstituents With Varying Values of Elastic Modulus
Authors: Leonard DN , Batchelor AD , Russell PE
Published: July 2020

The atomic force microscope (AFM) has allowed microscopists to observe surface topography of non-conductive samples on the atomic level. One AFM mode that scanned probe microscopists have recently shown an interest in is the phase lag imaging mode. It has already been demonstrated that the resulting phase lag data can be used to identify different densities of microlayered polyethylene. However a quantitative understanding of phase lag imaging has yet to be fully developed. With a better understanding of the phase lag data it may be possible to estimate the modulus or other material properties of a sample from an AFM phase image alone.

The need for better understanding phase lag data is essential to increasing the knowledge of a how a material's microstructure behaves on the nanometer scale. This investigation focused on AFM phase lag data produced while scanning microstructures with large differences in modulus values between the compositional phases.
Analysis of Selected Area Diffraction Patterns With Winjade
Authors: Walck S D , Ruzakowski-Athey P
Published: July 2020

The analysis of Selected Area Diffraction (SAD) patterns that are collected from a single phase material having sufficient crystallites to provide continuous rings is relatively straightforward. However, when this condition is not met and there may be several phases present having rings of a spotty nature, the pattern is complex and can be quite difficult to analyze manually because of the vast number of discrete spots. WinJade from MDI is an X-ray diffraction (XRD) analysis program with an Electron Diffraction Program Module (EDPM) that can be used to aid in the analysis of SAD patterns. The EDPM produces Integrated Circular Density Plots (ICDP), which are one-dimensional intensity profiles plotted as a function of equivalent XRD 20 values or crystal d-spacings. These ICDP's can be overlayed with XRD patterns or with reference lines from the NIST and JCPDS crystalline databases for direct comparisons.
Trace Element Detection by X-Ray Fluorescence
Authors: Madden M C , Wong R , Wherry D C
Published: July 2020

Due primarily to the absence of Bremsstrahlung radiation, fluorescent spectra obtained with x-ray excitation of a sample have a lower background than those obtained with electron excitation. This can be used to detect elements in liquids at very low concentrations under certain conditions. The technique consists of placing a small measured volume of liquid to be analyzed for trace elements on a thin, low atomic number film; evaporating the liquid to dryness; placing it under an x-ray beam; and obtaining a fluorescent spectra. The residue from the evaporated liquid contains any trace elements present, which then produce characteristic peaks in the fluorescent spectrum. These peaks can be used to measure concentrations quantitatively.

This technique has been around for some time, but with detection limits at the parts per million (ppm) level. Although there are many definitions of detection limit, the most commonly used one is from Burton:
High Resolution X-ray Microscopy of Frozen Hydrated Samples
Authors: Schneider G
Published: July 2020

X-ray microscopy is a rapidly developing field stimulated by the development of brilliant X-ray sources and high resolution X-ray lenses. It provides higher resolution than optical microscopy and higher penetration power than electron microscopy. Therefore, X-ray microscopy allows high resolution imaging of thick hydrated samples. The two dominating processes determining the contrast in X-ray microscopy are photoelectric absorption and phase shift. For this reason X-ray microscopy can be performed in amplitude and phase contrast. The Göttingen X-ray microscope at the BESSY electron storage ring in Berlin is operating in both contrast modes and is used for different application fields, for example in biology, biophysics, medicine, colloid chemistry, and soil sciences.

Especially biological objects are sensitive to ionizing radiation. Theoretical investigations show that X-ray images of frozen-hydrated specimen can be obtained without radiation induced artifacts. Therefore, an object stage for cryogenic specimen was developed and implemented on the Gottingen transmission X-ray microscope (TXM) at the electron storage ring BESSY.
High Spatial Resolution Soft X-Ray Microscopy and Microanalysis of Thick and Hydrated Materials
Authors: Meyer-Ilse W , Brown J T , Magowan C , Yeung J , Kurtis K E , Monteiro P , Tonner B P , Nealson K , Lelièvre S , Larabell C
Published: July 2020

The Center for X-ray Optics (CXRO) built and operates a high-resolution soft x-ray microscope (XM-1) at the Advanced Light Source in Berkeley. We report on the use of this instrument in a variety of scientific fields, including biology, civil engineering and environmental sciences.

The microscope is a conventional (full field) x-ray microscope, which uses zone plate lenses to provide high resolution transmission images. The optical setup is similar to the Göttingen x-ray microscope, operated at the BESSY synchrotron radiation facility in Berlin, Germany. A condenser zone plate, fabricated by the Göttingen group, is illuminating the sample and an objective zone plate, fabricated by Erik Anderson (CXRO), is forming an enlarged image on an x-ray CCD camera. While the optical path of the microscope is in vacuum, the sample is at atmospheric pressure, flushed by helium. The spatial resolution of our microscope is 43 nm, measured as the distance from 10%-90% intensity in the image of a knife-edge.
Imaging, Spectroscopy and Tomography of Frozen Hydrated Specimens With the Cryo Scanning Transmission X-Ray Microscope at The NSLS
Authors: Maser J , Jacobsen C , Wang Y , Osanna A , Winn B , Kirz J
Published: July 2020

With the steady improvement of x-ray optics with high resolution and efficiency, and continued development or adaptation of different imaging and measuring techniques, soft x-ray microscopy has emerged as a powerful method to image and analyze fully hydrated specimens of several micrometer thickness at sub-optical resolution (for a recent overview, see ref. 1). We report on experiments performed with the cryo scanning transmission x-ray microscope (cryo-STXM), which has recently come into operation at the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory.

Cryo-STXM uses x-rays with energies between the absorption edge of Carbon (E = 284 eV) and Oxygen (E = 543 eV) from the soft x-ray undulator at the NSLS. Fully hydrated specimens such as eucaryotic cells in water or ice layers of up to 10 micrometer thickness can be imaged without any additional need for contrast enhancing techniques.
Applications of Laboratory-Based X-Ray Microfluorescence Analysis in Archaeometry
Authors: Pingitore NE , Leach JD , Iglesias A , Sampson CG , Carmichael DL , Peterson JA
Published: July 2020

X-ray microfluorescence (XRMF) is spatially resolved x-ray fluorescence (XRF) analysis of elemental composition. Formerly available only at synchrotron sources, recent development of commercial instruments has brought a tamer version of this technique into the size and price range of the academic laboratory. Our Kevex Omicron system features a primary beam diameter as small as 30 μm, with an energy dispersive spectrometer collecting the x-ray spectrum from Na to U. The micro-positioning stage and optical microscope permit selective spatially resolved analysis on the surfaces of objects as large as a desktop dictionary. The large sample stage, functional spatial resolution, and non-destructive analysis present significant opportunities for elemental analysis of a variety of archaeological materials. In particular, objects which for aesthetic, religious, regulatory, or economic reasons cannot be subjected to any alteration in sample preparation may still be analyzed by XRMF. Our laboratory has been conducting a number of investigations which illustrate the potential impact of XRMF technology on archaeometry.
X-Ray Radiation Damage of Polymers in a Scanning Transmission X-Ray Microscope
Authors: Coffey T , Ade H , Urquhart S , Smith AP
Published: July 2020

We use the Scanning Transmission X-ray Microscope (STXM) at the National Synchrotron Light Source (NSLS) (1) to acquire images and spectra of polymers. To interpret data correctly, the effects X-ray radiation has on polymers must be understood. We have therefore started to characterize radiation damage in a variety of carbonyl containing polymers in two ways. First, we want to ascertain the critical dose for mass loss and the critical dose for the carbonyl in a variety of polymers and relate the critical dose to the polymer structure. (The critical dose is the radiation dose at which the optical density of the material is decreased by 1/e of its original value.) We also want to understand the damage mechanism. STXM acquires images and Near Edge X-ray Absorption Fine Structure (NEXAFS) spectra by using X-ray photons to excite inner shell electrons to unoccupied valence orbitals or to the continuum.
Characterizing Inhomogeneous Deformation Substructure in Large Single Crystals by Microscopy and XRD - A Synergystic Analysis
Authors: Kingman P W
Published: July 2020

Characterization of deformed crystals containing extensive inhomogeneities and localized lattice reorientations presents a formidable challenge, particularly when the available sample volume is limited and/or irregular in configuration. The crystals analyzed in this work were originally high-symmetry single crystal tungsten rods, 6.9 mm in diameter and 102 mm long, which had been ballistically launched against steel blocks. At ordnance velocities (1.5 km/sec), the rods evert as they penetrate the block, resulting in a cylindrical cavity partially lined with a hollow tube of tungsten, with a slug resting at the bottom. These tungsten residuals contained many localizations, fractures, and assemblages of discrete sub-entities, typically 100-200 microns in size, within a macroscopic flow pattern extending over many millimeters. The measured cavity depths displayed a strong crystallographic symmetry dependence. Because the post-event samples, which were either blocks containing polished axial sections through the cavity or loose ejecta, were both massive and frangible, area-resolved X-ray diffraction, utilizing a fine-beam Laue technique with a precision specimen translator to permit correlation with micrographs, was used to map and characterize the lattice sub-structure.
X-Ray Microscopy Developments at the ESRF
Authors: Kaulich B , Oestreich S , Barrett R , Susini J
Published: July 2020

The ESRF source properties offer the required coherence and brilliance to extend X-ray microscopy techniques, which have been extensively used in the soft X-ray region, to higher energies. This will enable new investigations such as the study of thicker specimens compared to the soft X-ray region, access to absorption edges of elements of major interest in the biological and materials sciences, in particular from Potassium to Chromium, the use of X-ray fluorescence for trace element mapping with high spatial resolution and the application of phase contrast.

Two microscopes are under development at the ESRF to cover a wide range of biological and material sciences applications:

A scanning transmission X-ray microscope (STXM) working in the photon energy range of 0.5-8keV. Whilst in its simplest form this might consist of taking multiple images of a single sample region at different incident energies, an interesting extension is to perform highly spatially resolved XAS scans on small regions of the sample.
High Resolution Transmission Electron Microscopy at Zero Cs
Authors: O'Keefe Michael A
Published: July 2020

Now that correctors for objective lens spherical aberration are becoming feasible, questions have been raised about the usefulness of high-resolution transmission electron microscopy at zero Cs and the possible difficulties of interpretation of such images. In general, high resolution TEM images are interpreted either by comparison with simulations from model structures or by contrast transfer functions (CTFs) to determine the weight (and sense) of spatial contributions to images from corresponding diffracted beams. At zero Cs, HREM image simulations will work, but a projected charge density theory should be used (instead of CTF theory) to interpret images. Both theories use approximations; CTF theory relies on kinematic scattering and PCD theory on zero Cs and limited defocus.
First Application of a Spherical-Aberration Corrected Microscope Material Science
Authors: Kabius B , Urban K , Haider M , Uhlemann S , Schwan E , Rose H
Published: July 2020

One of the most challenging tasks for high-resolution electron microscopy (HREM) is the atomistic investigation of defects and interfaces in thin films of semiconductors and ceramics. In particular, the application of these materials in electronic devices requires the understanding of the atomistic structure responsible for electronic and optical properties. The imaging of these structures requires a point resolution down to 0.1 nm. For example the smallest atomic spacing of GaAs in the (110) projection is 0.14 nm. According to Scherzer the point resolution of the TEM is proportional to spherical-aberration coefficient Cs1/4 and to the wave length λ3/4. Commercial medium-voltage microscopes up to 400 kV offer only a point resolution of 0.16 nm due to the high spherical-aberration constant Cs of the electromagnetic objective lens. Decreasing the wave length λ by increasing the electron energy improves the point resolution, but has the drawback of severe radiation damage and high costs.
Modification Of a Conventional TEM (CTEM) for Lorentz Microscopy
Authors: Wall M , Bajt S , Cerjan C
Published: July 2020

We have modified a CTEM in order to perform Lorentz microscopy experiments at high magnification and resolution on magnetic materials. The modification consists of the adaptation of a second side entry goniometer (SEG) to the CTEM (JEOL 200CX STEM) column above the objective lens, in a region of the column where the measured residual magnetic fields are < 0.5 Gauss with the objective lens in a fully excited state (see Figure 1).

With the specimen positioned above the objective lens, it is necessary to increase the excitation of the lens by approximately 20% in order to bring the image into focus. This combination of a specimen positioned above the object pole piece, and the higher objective lens excitation, affects several changes in the optics. First, if a specimen is positioned within the pole piece gap and the objective lens is turned off for Lorentz microscopy purposes, the highest typical magnification achievable is approximately 1,000 X and the resolution is not much better than 0.1μm.
A New 200kv Energy Filter Field Emission Transmission Electron Microscope
Authors: Kaneyama T , Tsuno K , Honda T , Kersker M , Tsuda K , Terauchi M , Tanaka M
Published: July 2020

In the field of biological and materials sciences, the importance of energy filter transmission electron microscope(EF-TEM) is increasing. Because it is a powerful instrument for contrast enhancement and obtaining elemental mapping images. We have developed a 200kV EF-TEM equipped with a fieldemission gun and in-column spectrometer. The new EF-TEM JEM-2010FEF inherits the performance in high resolution imaging and analysis from field emission TEM. The outer view is shown in Fig.l.

Figure 2 shows the lens configuration of JEM-2010FEF. An in-column Q-type spectrometer is introduced within the imaging lens system. It was designed to have image distortion less than 1% and dispersion power 1.2p.m/eV for 200keV electrons. There is no need of compensating procedure of distortion. Imaging lens system consists of two objective lenses, three intermediate lenses and three projector lenses. The 8-stage imaging lens system enables wide range of imaging modes equal to conventional TEMs; energy spectroscopic image of magnification from ×200 to × 1,500,000, energy spectroscopic diffraction of camera length from 200mm to 2,000mm.
The CM120-Biofiiter: Digital Imaging Of Frozen Hydratedspecimens and Elemental Mapping
Authors: Lücken U , de Jong AF , Kundmann M , Chemy D , Leapman R
Published: July 2020

A new system for energy-filtered electron microscopy (EFTEM) has been developed, adapted for the study of life science specimens. Special attention has been given to integration, ease of use and the typical problems encountered while investigating unstained and cryo-specimens. Low-dose, cryocapabilities and a high-contrast objective lens have been combined with an imaging energy filter, digital image recording with phosphor scintillator and advanced image-processing capabilities. The CM120-BioFilter is an integration of the CM120-BioTWIN or the CM120TWIN with the Gatan postcolumn imaging filter GIF100. The BioTWIN objective lens (Cs=6.2 mm, f=5.9 mm) is optimized for high-contrast imaging. The TWIN lens(Cs=2mm,f=2.8mm) is optimized for high resolution in structural biology. The GIF 100 is a computer-controlled, second-order corrected energy filter with a cooled multiscan CCD for digital image recording. Improvement of the contrast can be achieved if inelastically scattered electrons are removed.
Quantitative Visible-Light and Electron Phase Microscopy
Authors: Barty A , Nugent KA , Paganin D , Roberts A
Published: July 2020

At the University of Melbourne we have been pursuing an ongoing program of investigation into the recovery of phase information from intensity measurements [1-9]. In this paper we consider the implications of this work in optical and electron microscopy.

Many objects of interest to biologists are phase objects which means that light is slowed and refracted in the object but not absorbed. Techniques such as phase-contrast microscopy and Nomarski differential interference contrast (DIC) microscopy are traditionally used to render the phase structure visible but do not directly map the phase distribution and are unable to produce quantitative data. These techniques also entangle the phase and amplitude information and so have limitations where both phase and amplitude information is required. In this paper, we demonstrate quantitative non-interferometric recovery of microscopic phase structure using incoherent illumination of the type commonly found in optical microscopes, and show that our results correlate with structure observed using Nomarski DIC techniques.
Isotopic Measurements of Inorganic Material by Time-Of-Flight SIMS
Authors: Fahey A J
Published: July 2020

Isotopic measurements via Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) have generally not been considered as practical because of the low duty cycle at which ToF-SIMS instruments operate and the corresponding low data rate. The recent discovery of pre-solar material in meteorites has shown that large variations in isotopic ratios (several orders of magnitude for some elements) exist in small (∼1 μm), refractory meteoritic grains. These grains are ideal candidates for ToF-SIMS, which consumes little sample material, compared to dynamic, magneticsector SIMS. ToF-SIMS also allows for parallel detection of all species present in the sample; thus, multiple isotopic systems can be studied in one measurement. As a prerequisite to studying the isotopic composition of meteoritic materials, preliminary determinations of ratios for a number of elements have been made on materials of known isotopic composition. This allows us to investigate problems that may be unique to ToF-SIMS for the measurement of isotopic ratios.
Visualizing Neuronal Sturcture by Combining GFP-Transfection With 2-Photon Microscopy In Living Brain Slices
Authors: Horch H L Wilson , Katz L C
Published: July 2020

Research in our lab is aimed at examining how signaling molecules affect the differentiation of neurons in the developing cortex. Most morphological studies rely on single time point images of cells within a population. It is obviously advantageous to examine changes in individual living cells over time. By combining 2-photon imaging with co-transfection of genes for green fluorescent protein (GFP) and neurotrophins in living neurons, we can asses the effects of these on individual living cells.

Individual neurons in brain slices from ferret visual cortex were transfected with GFP using particle mediated gene-transfer, or Biolistics. Using the Helios Gene Gun (Bio-Rad), DNA-coated gold particles are blasted into living brain slices. Neurons are transfected when a gold particle lands in the nucleus. Within 24 to 36 hours individual cells are well-filled with GFP allowing visualization of dendrites, spines, and axons.
Crista Junctions of Mitochondria Visualized by Electron Tomography
Authors: Perkins G A , Song J Y , Tarsa L , McCarty C , Deerinck T J , Ellisman M H , Frey T G
Published: July 2020

Electron microscope tomography is a useful method for deriving three-dimensional (3-D) structures from tilt series. We used an intermediate, high-voltage electron microscope operated at 400 kV to study the 3-D structure of selectively stained or labeled mitochondria from diverse sources. We have examined the structure of neuronal mitochondria and mitochondria from brown adipose tissue (BAT) employing either single-axis or double-axis techniques over a tilt range of ±60° with 1° or 2° angular increments (Fig. 1). Although BAT mitochondria contains only lamellar cristae with no tubular cristae as we have observed in neuronal mitochondria, we found that the inner membrane of BAT mitochondria invaginates to form cristae only through narrow, tubular openings or crista junctions (Fig. 2).

Recent interest in the possibility of structural artifacts in chemically fixed mitochondria imaged in situ motivated us to perform electron tomography on cryofixed brain and brown fat tissue.
Electron Microscopic Tomography of Cellular Organelles: Chemical Fixation Vs. Cryo-Substitution of Rat-Liver Mitochondria
Authors: Mannella CA , Buttle K , Tessitore K , Rath BK , Hsieh C , D'Arcangelis D , Marko M
Published: July 2020

Electron microscopic tomography is proving to be a valuable tool for investigating the 3D structure and organization of cellular organelles. Important progress is being made in the application of the technique to frozen-hydrated material, but it is likely that success with thick specimens will be limited by the low contrast and beam sensitivity of naked biological material. Thus, optimizing procedures for fixing, embedding, staining, and selectively labelling cells for 3D electron microscopy remains a priority.

Tomography of chemically fixed and plastic-embedded rat-liver tissue and isolated mitochondria has shown that the cristae (the invaginations of the mitochondrial inner membrane) are pleiomorphic and connected to each other and to the surface of the inner membrane by tubular regions 30-40 nm in diameter. This basic design feature has important implications for the microcompartmentation of ions and molecules within this organelle.
3d Structure Of Periodic Cubic-Phase Inner Membranes In Mitochondria of Chaos Carolinensis
Authors: Deng Y , Mieczkowski M , Marko M , Buttle K , Rath BK , Mannella CA
Published: July 2020

The mitochondrial inner membrane contains the machinery of oxidative phosphorylation. This membrane has invaginations called cristae which vary widely in shape between organisms and between tissues in the same organism. Electron microscopic tomography indicates that, despite this pleiomorphism, there is a common design feature, namely, the cristal membranes connect to each other and to the periphery of the inner membrane by tubular regions 30-40 nm in diameter. This finding has important implications for the internal diffusion of ions, metabolites and macromolecules within mitochondria.

In some types of mitochondria, the cristae exhibit periodicity.1 In the case of the amoeba Chaos carolinensis, detailed analysis and modeling of thin-section images of mitochondria in starved cells indicate that the highly curved cristae correspond to periodic cubic surfaces. We are undertaking electron microscopic tomographic and crystallographic approaches to more thoroughly characterize these membrane phases and, in particular, establish the continuity of the internal compartments which they define.
The Centrosome: Three-Dimensional Structure of a Cell Organelle at 40A Resolution Obtained by Electrontomography
Authors: Ruiz T , Radermacher M , Rath B , Rieder C , Bornens M
Published: July 2020

The organization of the microtubule network of eukariotic cells is controlled by the centrosome. This organelle which duplicates once during the cell cycle comprises two centrioles and a dense filamentous material called pericentriolar material. The centrioles are cylinders of ≈ 0.25 μm in diameter and ≈ 0.5μm in length and are composed of 9 microtubule triplets at the proximal end and 9 doublets at the distal end. For the last 50 years the centrosome and the centrioles in particular have been the object of many optical and electron microscopy studies. But, in spite of their highly symmetric organization, their fine structure is still not well understood. The reconstruction presented here is a substantial step forward towards a more detailed understanding of their structure.

Centrosomes were isolated from a human lymphoblastoma cell line (1). 10 nm gold particles were applied to formvar-carbon coated grids to provide markers for the alignment procedure (2,3). Centrosomes were centrifuged onto the grids and negatively stained.
Using Animation to Enhance 3D Visualization: A Strategy for a Production Environment
Authors: Dougherty M T , Chiu W
Published: July 2020

The use of animation can significantly enhance the visualization of three-dimensional (3D) structures. It can present a focused train of thought, or it can be used to systematically scan through previously unfathomable quantities of data to examine for unknown features and consistencies. In order to establish a modern animation facility requires a variety of technical, psychological and artistic skills, in addition to advanced computer graphics and related equipment. 3D graphics combined with animation has proven to be a very effective tool in storytelling. The dynamic visual image is uniquely suited for thought experiments, simulations and traversing vast quantities of data otherwise incomprehensible to the human mind. Sometimes animation is the only method that allows a clear presentation of complex empirical or theoretical information in many dimensions. Scientific visualization is by its nature an exploratory process. And frequently animations are iteratively refined and polished to enhance comprehensibility for the researchers and their peers.
Atomic Model of Tubulin by Electron Crystallography
Authors: Nogales Eva , Downing Kenneth H
Published: July 2020

The structure of tubulin, the major protein component of microtubules, has been obtained by electron crystallography of zinc-induced 2-D crystals. The atomic model of the tubulin dimer, built into a 3.7 Å density map, shows that α- and β-tubulin have basically identical structures. Each monomer is very compact and is formed by two interacting beta sheets surrounded by helices. The structure can be divided into three sequential domains that are functionally distinctive. The N-terminal domain forms a Rossmann fold that binds the nucleotide. Connected to the nucleotide-binding domain by the core helix is a second domain that in β-tubulin contains the binding site of taxol. The C-terminal region is formed by two long helices that constitute most of the interactive, outside surface of the microtubule. The nucleotide-binding and second domains are common with FtsZ, a bacterial homologue of tubulin essential for cell division. The bacterial protein, which lacks the C-terminal helices of tubulin, has generally shorter loops and contains an extra helix at the N-terminus.
Crystallographic Reconstruction of the Acrosomal Process from Limulus Polyphemus Sperm
Authors: Sherman M B , Jakana J , Sun S , Matsudaira P , Chiu W , Schmid M F
Published: July 2020

The acrosomal process is an intracellular quasi-crystalline organelle in the head of the sperm of the horseshoe crab Limulus polyphemus. It consists of 100 - 130 actin-scruin filaments packed together in a pseudo-hexagonal lattice2 and is up to 60 (im long with a diameter of 0.1 μm. Scruin-scruin interactions are responsible for cross-linking the actin filaments together in the bundle. The goal of the current study is to reveal interfilament interactions in the bundle. We have taken tilt series images in the electron microscope3 to reconstruct its three-dimensional structure without assuming helical symmetry.

The acrosomal process was purified as described.34 Bundles were embedded in vitreous ice over holes on a holey carbon film on copper grids. The specimen was kept at -167°C in a JEOL 4000EX electron microscope operating at 400 kV. Straight 6-10 (im long bundles were found using a TV-rate CCD camera in defocused diffraction mode.
Structure of a Recombinant Gap Junction Channel at 7Å Resolution
Authors: Unger VM , Entrikin D W , Guan X , Kumar NM , Gilula NB , Yeager M
Published: July 2020

Gap junction membrane channels mediate the electrical and metabolic coupling between cells. The channels are formed by the end-to-end docking of two hemichannels (connexons), each of which is formed by a hexameric cluster of protein subunits (connexins). The principal gap junction protein in the heart, α1 connexin (also designated Cx43), mediates action potential propagation between cells in order to synchronize cardiac contraction.

We recently utilized electron cryo-microscopy and image analysis to examine frozen-hydrated two-dimensional (2D) crystals of a recombinant, truncated α1- connexin (α1Cx263T). The projection map at 7Å resolution revealed that each 30kD connexin subunit has a transmembrane α-helix that lines the aqueous pore and a second α-helix in close contact with the membrane lipids [Nature Struct. Biol. 4: 39-43 (1997)]. The distribution of densities allowed us to propose a model in which the two apposing connexons that form the channel are staggered by ∼30°. Furthermore, apparent non-crystallographic twofold axes predicted that the two apposing connexons adopt identical conformations.
Electron Crystallography of a Prokaryotic Potassium Channel
Authors: Sui HX , Li HL , Ghanshani S , Lee S , Walian PJ , Wu CL , Chandy KG , Jap BK
Published: July 2020

Potassium channels are ubiquitous ion channel proteins which play a crucial role in a broad spectrum of important cell functions. KcsA is a potassium channel found in the bacterium, Streptomyces lividan, and is believed to have only two transmembrane helices.

The KcsA protein has been cloned, overexpressed and purified to homogeneity, and reconstituted with phospholipids to form two-dimensional (2-D) crystals. Crystal samples were embedded in trehalose and examined in a cryo-holder (-170 °C) using a JEOL 4000EX electron microscope operated at 400kV in low dose mode. Images were recorded in spot scan-mode at a magnification of 60,000X, then digitized on a Perkin Elmer PDS flat bed microdensitometer and processed by using the MRC program suite. Processed images were brought to a common phase origin, and the amplitudes and phases from individual images were vectorially combined.
User Access to Lorentz Microscopy at the NCEM, LBNL, Berkeley.
Authors: Verbist K , Nelson C , Krishnan K
Published: July 2020

A standard Philips CM200FEG electron microscope, without the special Lorentz lens, has been optimized for Lorentz imaging. The necessary field-free sample region is obtained by switching off the objective lens in the free lens mode. The limited range of magnification is compensated for by a post-column Gatan image filter (GIF) which magnifies by a factor of _ 20. Fresnel imaging is performed by defocusing with the diffraction lens. The use of low angle diffraction, in combination with the apertures located at the selected area aperture plane, allow Foucault imaging. The TEM analog of differential phase contrast (DPC) imaging has been implemented. This method makes it possible to obtain quantitave induction maps of the in-plane magnetization. TEM DPC is based on a series of Foucault images, recorded with different incremental beam tilts, which are processed to yield images equivalent to the quadrant signals obtained by the STEM DPC technique.