bluuebunny/crossref_metadata_2025 huggingface dataset:
Genistein Inhibits Cell Proliferation and Induces Mitochondrial and Nuclear Alterations That Indicate Mechanisms Involved in Apoptosis
Authors: Gobert Geoffrey , Curren Ed , Welshons Wade , Sun Qing-yuan , Schatten Heide
Published: July 2020

A highly significant correlation between reduced incidence of breast cancer in Asian countries and consumption of soy suggests that specific components in soy may have anticarcinogen activity. The soy ingredients genistein and daidzein have been found to inhibit induced breast tumors in animal and cell culture models. These isoflavones are known to be both agonists and antagonists of estrogen activity but only genistein is also a potent inhibitor of tyrosine kinases which are the primary intracellular signalling mechanisms associated with the regulation of cell proliferation.

Genistein promotes cell proliferation in breast cancer cells at low concentrations in its function as estrogen agonist but inhibits cell proliferation at high concentrations (30 μM). In order to investigate the underlying mechanisms by which high concentration of genistein inhibit cell proliferation we treated MCF-7 cells with increasing concentrations of genistein and analyzed cells by immunofluorescence and transmission electron microscopy (TEM).
Effects of Co60 Irradiation on Muscle Fiber Types of the Grass Frog
Authors: Cohen Glenn M , Scott Margaret F
Published: July 2020

Striated skeletal muscle has been considered radioresistant because it is highly differentiated and post-mitotic. Striated muscle does, however, respond to irradiation with morphological and biochemical changes after short and long latency periods; vascular and/or neurological impairments might contribute to the delayed responses to irradiation.

The objective of the present study was to determine the susceptibility of three amphibian muscle fiber types to Co60 irradiation. In amphibians, the three major fiber types are 1) large twitch fibers, which contain low levels of mitochondrial enzymes and lipids, but intermediate levels of glycogen; 2) small twitch fibers, which contain high levels of both glycolytic and mitochondrial enzymes (FIG. 1); and tonic fibers, which contain low levels of all three histochemical markers. Thus, the determination of susceptibility of different amphibian fiber types to irradiation might indicate whether the metabolic characteristics of the fibers, rather than morphological or electrical properties, could serve as an early indicator of radiation damage.
Electron Microscopic Evaluation of Vector Quality for Gene Therapy
Authors: Yu Qian-Chun , Ma Xiaokui , Croyle Maria , Qu Guang , Huang James , Gao Guangping , Hughes Joe , Wilson James
Published: July 2020

Successful gene therapy is dependent upon well regulated, long-term expression of transgenes in the target tissue. The development and production of very high quality vectors are one of the most important steps leading to delivery of the desired genes. Although several types of vectors have been used for the delivery of transgene to the target cells of the host, our institute routinely uses recombinant viral vectors including adenovirus and adeno-associated virus (rAAV) for gene therapy studies. To assure high quality of the viral vectors being used for animal experiment as well as clinical trials, a number of quality control protocols, including electron microscopy, was developed and implemented to monitor the overall purity, and density of the vectors.

Transmission electron microscopy (TEM), combined with optimized procedure of negative staining for virus, offers a unique means to generate visual data of the vector preparation; and it has gained increasing application to help assess the quality of vectors.
Mineralization of Collagen in Avian Tendon Examined by Tapping Mode Atomic Force Microscopy
Authors: Siperko L M , Landis W J
Published: July 2020

Type I collagen fibrils provide a structural framework for mineral nucleation, growth, and development during calcification of tissues such as bone, dentin and tendon of many vertebrates. Previous work has described a general mechanistic model of normal collagen- mineral interaction in which nucleation events take place on collagen surfaces and in channels or gaps defined by periodic (64-70 nm) collagen hole and overlap regions. These sites are created in early collagen molecular and macromolecular assembly. The model suggests that, once nucleated, the mineral within channel spaces grows into irregularly shaped platelets of small size and these develop into larger platelets whose detectable crystallographic c-axis direction parallels the long axis direction of the collagen fibrils with which they associate. The fate of mineral nucleated on collagen surfaces is unclear, although it is known that the volume of extracellular tissue space between adjacent collagen fibrils, like that within the fibrils, also becomes mineralized.
Stylet Length of Silverleaf Whitefly Adults and Nymphs And The Mechanism of Stylet Insertion Into the Leaves of Host Plants
Authors: Freeman Thomas P , Buckner James S , Nelson Dennis R
Published: July 2020

Silverleaf whiteflies, Bemisia argentifolii Bellows and Perring, feed on a wide variety of plants and are capable of causing significant economic damage. Host selection may be determined by a wide variety of factors. Adult whiteflies may feed on one leaf and then move to another, however, their feeding and oviposition choices may well determine the success of egg hatch and nymph survival. Current theory suggests that whiteflies feed (Fig. 1) and oviposit on the abaxial surface of leaves due to the closer proximity of phloem tissue. This is believed to be particularly important for nymphs which are stationary during much of their development and have shorter stylets than adults. We examined whitefly stylet penetration into cotton and hibiscus leaves using both light and scanning electron microscopic techniques.

Adult silverleaf whitefly stylets were found to range from 100 urn to more than 300 um in length.
Light and Electron Microscopy Study of the Metacestode (Coenurus) of Taenia multiceps
Authors: Ramirez-Bon Enrique , S. Daniel Gonzalez
Published: July 2020

Introduction:- The metacestode (larval stage) of Taenia multiceps, it is characterized by its big bladder fluid filled and multiple scoleces. Affect very frequently herbivorous mammals like sheeps, hares and rabbits (1); and occasionally humans (2), causing disorders very similars to that caused by Cysticercus cellulosae (3), the larval stage of Taenia solium. The adult parasite, Taenia multiceps, it is found in the intestine of domestic and wild carnivorous like dogs, wolves and coyotes.

In this paper we describe the morphology at the optical and electron microscope level of coenuri collected from hares.

Material and Methods.- Coenuri collected from hares, were dissected from muscle tissues and fixed in glutaraldehyde 2.5% in 0.1M phosphate buffer pH 7.4, postfixed in 1% osmium tetroxide in the same buffer. Inclusion in medcast resin (Pelco). Sections were stained with uranyl acetate and lead citrate, and observed in a Zeiss EM 109 electron microscope.
Pulmonary Neuroepithelial Bodies in Neonatal Rats Chronically Exposed to Nitric Oxide
Authors: Hung Kuen-Shan , Her Xay K
Published: July 2020

Pulmonary neuroepithelial bodies (NEBs) consist of clusters of innervated endocrine cells localized in human and animal lungs. They are prominent in late fetal and neonatal lungs and their density declines with increased postnatal age. Gas sensing functions of these structures have been suggested by Lauweryns and Cokelaere. Subsequent experiments show that chronic hypoxia leads to an increase in the size or number of NEBs, and prolonged hyperoxia also causes elevation of NEB numbers. Animals exposed to cigarette smoke also had an increased number of immunoreactive NEBs.

Inhaled nitric oxide (NO) can reverse pulmonary vasoconstriction induced by hypoxic breathing or ligation of ductus arteriosus in newborn and young animals. These studies led to clinical use of NO for treatment of pediatric patients with pulmonary hypertension. However, direct effects of inhaled NO on various components of the lung are not clearly understood.
Ultrastructure of the Stomach of Aphanius Dispar (Cyprinodontidae), With Special Reference to Stress from Starvation
Authors: Ba-Omar T A , Victor R
Published: July 2020

The effect of starvation on the mucosa of the Aphanius dispar stomach is investigated. The gastric erosion resulting from stress due to starvation over a period of 24 - 120 hours has been described at the ultrastructural level.

Specimens of A. dispar with a total length (TL) range of 30.4-46.3 mm, weighing from 0.43-0.70 gm were collected from Wadi Al-Khod, Sultanate of Oman (Lat. 23° 30’ N; Long. 58 ° 40'E). Thirty six fish were acclimatized for two weeks and were fed with Tetramin flakes, ad libitum, twice daily. Thirty of these fish were subsequently starved in a chamber separated from the six control fish that were fed normally in the same tank. Six starving fish were randomly sacrificed at 24, 48, 72, 96 and 120 hr intervals for histological studies. At the end of the experiment, the six control fish were also sacrificed. Stomachs were fixed and processed for TEM.
Immunocytochemistrical Studies on Cytoskeletal Change in Cultured Cardiomyocyte of Tilapia
Authors: Tung L C
Published: July 2020

In the present study, the myofibril regeneration in the long-term cultured fish cardiomyocytes was studied with immunocytochemistry.

Adult Tilapia heart was dissociated into a single-cell suspension with collagenase and protease-minced tissue method. The culture medium was Eagle's minimal essential medium (MEM) with Earle's salts, supplemented with 10% fetal calf serum, 1 x nonessential amino acid mixture, 100 IU/ml penicillin G, and 100 μg/ml streptomycin. The cultured cells were grown in a humidified CO2 incubator at 28°Cand in a medium without glutamine for eliminating fibroblast contamination. In the initial 24 h culture, the elongated-shape cells gradually shortened from their both ends and rounded up. Over 5 to 6 days postcultivation, the cells attached to the bottom of the culture flask and began to protrude pseudopodia. The cells could not be subcultured and also proliferated indefinitely. The life span of cells in culture was 30 to 60 days.
The Effect of a 20-Minute Exposure of Colloidal Silver on the Membrane of Human Peripheral Blood Lymphocytes and Raji Cells
Authors: Garvin K R , Hintze C J , Bowden R D , Johnson D A , Tidwell J E , Standing M D , Murray B K , O'Neill KL
Published: July 2020

Apoptosis (programmed cell death) is an important form of physiological cell death displayed under divergent conditions by an enormous variety of tissues. Apoptosis can be characterized by cell shrinkage, membrane blebbing, and DNA fragmentation (1). Data from recent reports have suggested that many types of cancer are accompanied by the loss of apoptotic ability (2). Many manufacturers have hailed colloidal silver as an effective treatment of many diseases including cancer (3). The FDA has recently finalized a rule stating that drug products containing colloidal silver or silver salts are not recognized as safe and effective. They conclude that product labels claiming that scientific evidence suggests otherwise are incorrect (4). In this study we undertook to examine the effects of colloidal silver on both Raji cells, a B lymphocyte lymphoma, (ATCC # CCL-86) and normal, peripheral blood lymphocytes (PBL). We noted a decrease in viability of 90% in both Raji cells and lymphocytes after exposure to colloidal silver.
Structural Details as Clues to Understanding Nacre Formation
Authors: Yao N , Markiewicz D J , Aksay I A
Published: July 2020

Mollusk nacre, the opalescent lining of shells such as abalone, has a fracture toughness about 3000 times greater than that of aragonite (CaCO3), the mineral of which it is almost entirely composed. Since it owes its fracture strength and toughness to its composite organic/inorganic microstructure, details of nacre formation provide clues for materials scientists hoping to mimic nacre's microstructure in fracture-resistant synthetic materials. TEM and SEM provide detailed structural information, which contributes to the understanding of nacre.

An abalone shell was fractured to expose the stacked-plate morphology for imaging in a Philips XL30 FEG-SEM. Also, an ion-milled, cross sectional sample was prepared for imaging in a Philips CM200 FEG-TEM. While SEM shows tablets of aragonite with a highly uniform thickness of 0.5 μm, stacked in interdigitated columns (Fig. 1), cross-sectional TEM reveals organic layers, ∼10 nm thick, which separate the aragonite lamella (Fig. 2), consistent with previous studies.
Low-Loss EELS Spectral Fingerprints of Lipids and Protein
Authors: Aitouchen A , Shi S , Libera M , Misra M
Published: July 2020

Because of their amphiphilic nature, lipids tend to self-assemble into membranes and other topologically complex structures that can serve a number of functions in both natural and synthetic environments. The structure of lipid assemblies often varies on length scales of 2-100 nm, and morphological studies of such structures often require electron-optical methods. Image contrast in an electron microscope is usually generated by large defocus for unstained samples or by positive/negative staining methodologies. We currently are developing alternate approaches to generate image contrast based on spatially resolved Electron Energy Loss Spectroscopy (EELS) [1]. This has allowed us to distinguish between different lipid species as well as between lipids and protein. Low-loss spectra taken from cholesterol, ceramide and protein (BSA) show that these materials have characteristic spectroscopic fingerprints due to both π and σ valence-electron excitations that are sufficiently different to distinguish between them.
Quantitative Analysis of Metal Sequestering in Different Cell Components of Gloeocapsa alpicola in the Overplus Phase
Authors: Goldberg JJ , Jensen TE
Published: July 2020

The elemental composition of polyphosphate bodies (PPB's) and other components of the cell have previously been determined by using energy dispersive X-ray spectrometers(EDX). In this present study we perform a quantitative analysis of normally grown cells and cells that were grown in the overplus phenomenon.

Cells of Gloeocapsa alpicola were grown in modified Fitzgerald's media and harvested after a logarithmic growth phase of 14 days. The overplus cells were grown, as has been previously described. The cells were exposed to 20ppm of Al, Cd, Cu, Mn, Ni, Pb, and Zn, as well as a control, for one hour. The cells were then air dried on formvar coated grids or fixed and embedded in EPON according to Luft's procedure.

For Xn-ay analysis cells were first located using the TEM mode, and then the microscope was switched to the STEM mode. Analyses of cell components were carried out using the spot mode (75Kv).
Gene Expression Profiling Following Instillation of Diesel Exhaust Particles in Rat Lung: A First Study
Authors: Reynolds Lucy , BéruBé Kelly , Jones Timothy , Richards Roy
Published: July 2020

Epidemiological studies conducted first in the USA and later in the UK, suggest that a relationship exists between increased cardio-respiratory hospital admissions, morbidity and mortality rates and increases in PM10 concentrations. In urban environments, ultrafine diesel exhaust particles (DEP), accounts for 20-80 % by mass of the airborne PM10 arising from vehicular activities. In previous work, we used well characterised DEP as a surrogate for PM10 and examined its bioreactivity in vivo by assessing lung permeability, inflammation and epithelial cell markers in lavage fluid. Delivery of a single instillate of l mg DEP into the rat lung was not found to cause progressive damage but did produce a transient change in lung permeability. In the experiment described here, we instilled two different doses (control [NaCl], 0.25 and 1.25 mg) of DEP into the rat lung and assessed the responses using the methods described above with the addition of a new technique known as gene expression profiling.
On the Peak to Background Ratio of X-Rays emitted from Rough Surfaces
Authors: Gauvin Raynald , Lifshin Eric
Published: July 2020

The classical schemes to convert the x-ray intensity into concentration, using the ZAF or the ϕ (ρz ) methods, are valid for specimens having homogeneous composition and flat surfaces. Quantitative schemes have also been developed for x-ray microanalysis of multi-layered specimens. More recently, a quantitative method has been proposed for the microanalysis of spherical inclusions embedded in a matrix as well as Monte Carlo simulations of x-ray emission from porous materials.

For the case of specimens having a non-planar surface, a quantitative method based on the peak to background ratio, using photons of the same energy, has been proposed1. However, this method has some pitfalls. First, this method is based on the assumption that the peak to background ratio is independent of the specimen roughness which is not strictly correct because the ionization cross sections and the bremstrallung cross sections are not the same. Therefore, the shapes of the ϕ (p z ) curves are not the same for characteristic and continuum photon of the same energy resulting in different absorption corrections. The result is that the peak to background ratio will vary with beam position on a rough surface.
Electron microprobe analysis under conditions of non-normal Electron beam incidence
Authors: Lifshin E , Gauvin R
Published: July 2020

From its inception, electron microprobe analysis was almost exclusively done under conditions of normal electron probe incidence. The radial symmetry of this geometry greatly simplified the development of quantitative equations, and these equations where further refined based on large amounts of data also collected at normal incidence. However, as x-ray detection systems where added to scanning electron microscopes (SEMs), samples were often viewed under conditions of non-normal incidence and attempts were made to modify the various correction procedures to give acceptable quantitative results. Little justification for these methods has ever been published and so the current study was undertaken to compare theoretically calculated x-ray emission from a well characterized sample, in this case NiAl (.685 wt. % Ni,.315 wt. %A1) with experimentally measured results collected as a function of tilt angle. The theoretical calculation where done using a Monte Carlo (MC) program developed by Gauvin and Lifshin.
Improving the Analtical Accuracy in the Analysis of Particles by Employing Low Voltage Analysis:
Authors: Small JA , Armstrong JT
Published: July 2020

The energy of the electron beam, in conventional electron probe microanalysis, is generally in the range of 15-25 keV which provides the necessary overvoltage to excite efficiently the K and L x-ray lines for elements with atomic numbers in the range of about 5-83. One of the primary microanalytical methods for obtaining compositional information on particles is X-ray analysis in the electron probe and these same voltage criteria have been applied to the procedures developed for this purpose. The main difference in analytical procedures for bulk samples and particles is that corrections have to be applied to the particle k-ratios or calculated compositions to compensate for: 1) the penetration or scattering of electrons out of the particle volume and 2) variations in the absorption due to particle geometry of x-rays less than about 3 keV. In general, particle corrections improve the accuracy and reduce the relative uncertainty estimates from several tens of percent for uncorrected data to about 10% for corrected data.
A New Parametrization of Mott Scattering Cross Section for Monte Carlo Simulation
Authors: Demers Hendrix , Gauvin Raynald
Published: July 2020

In the recent years the Monte-Carlo simulation has been used successfully to exploit and understand fully the capabilities of electron microscopes. In this paper, we propose a new parametrization of Mott scattering cross-section for the calculation of the total cross-section as well as the polar angle of collision. This parametrization gives better results than Rutherford cross-section for Monte Carlo simulation at low beam energy without the numerous data files needed to use the exact Mott cross-section.

The calculation of elastic scattering cross-section can be performed with the Rutherford equation using the screening parameter, δ, the energy of the incident electron, and the electron wavelength, differential cross-section is given by:
Development of a Spectral Database for Testing Quantitative Electron Probe Microanalysis of Rough, Bulk Samples
Authors: Newbury Dale E
Published: July 2020

The vast majority of applications of electron probe x-ray microanalysis takes place on specimens which deviate significantly from the ideal configuration. Classic quantitative x-ray microanalysis makes the tacit assumption that the only reason the unknown differs in emitted x-ray intensity from standards is that there is a difference in composition between them. While this seems trivial, it forces the analyst to eliminate a major non-compositional source of possible intensity differences, namely the geometric effects associated with surface roughness. An important early paper by Yakowitz demonstrated that deviations from an ideal flat surface due to surface topography could seriously degrade the accuracy of analysis. Yakowitz interrupted the grinding and polishing procedure on pure elements and alloys at various stages and measured the variance in the x-ray intensity as the probe was scanned across the surface as compared to the predicted distribution based on counting statistics.
Tackling the Complexities of Analyzing Phases in Metallurgical SLAGS
Authors: Livi KJT , Farthing DJ , Veblen LA , Wing BA
Published: July 2020

In many areas of the world slags pose potential environmental threats to their surroundings. Heavy metals, toxic and radioactive elements contained in some slags may leach into drinking water and contaminate agricultural areas. Leaching experiments can estimate short-term alteration rates, but such tests cannot reproduce long-term weathering processes. Such studies are particularly important in the case of U- and Th-bearing slags which may remain hazardous for 103 104 years. These radioactive slags are of concern to decisions regarding the termination of a nuclear materials license and the subsequent allowances for unrestricted public use of the smelting sites and slag piles. To thoroughly evaluate leaching from slags, one must understand the distribution of elements among the slag phases and the stability of each phase. Questions regarding the distribution of elements among multiple phases are best answered by detailed electron microprobe (EMP) studies.

Slags are often mineralogically and compositionally complex, and may have complicated exposure histories.
EBSD in FESEM and LVSEM
Authors: Camus Patrick
Published: July 2020

Background

Electron backscattered diffraction (EBSD) is a very powerful analysis technique to understand the crystallography of a sample in the SEM. Typically, a high beam current is needed to obtain good results because of inefficient collection of the patterns. In W-filament SEM's, the high beam current necessitates a large spot size and requisite low image resolution. The large beam current can also be a source of problems for insulating samples or sample mounts by causing charging and the resulting image drift.

To reduce the spot size with adequate beam currents for imaging, field-emission (FE) guns are used. For most FESEM's, the beam current is markedly reduced from that produced by W-filament instruments. Even though the image resolution is better, the EBSD performance tends to suffer at low beam currents for standard hardware configurations. To regain the performance of a comparable W-filament system, the EBSD hardware should be changed for the low beam current operation. This paper will address the hardware considerations for low beam current applications.
Reduced Unit Cell Determination From Unindexed EBSD Patterns
Authors: Michael J R , Goehner R P
Published: July 2020

Electron backscatter diffraction (EBSD) is a technique that can provide identification of unknown crystalline phases while exploiting the excellent imaging capabilities of the scanning electron microscope (SEM). Phase identification using EBSD has now progressed to the point that it is commercially available. Phase identification in the SEM requires high quality EBSD patterns that can only be collected using either film or charge coupled device (CCD)-based cameras. High quality EBSD patterns obtained in this manner show many diffraction features that are useful in the determination of the unit cell of the sample.’ This paper will discuss the features in the EBSD patterns and the procedure used to determine the reduced unit cell of the sample.

One of the major advantages of EBSD over electron diffraction in the transmission electron microscope is the remarkable field of view that is routinely attained. The large angular view of the diffraction pattern permits many zone axes and their associated symmetries to be viewed in a single pattern or at most a few patterns.
Spatial Distribution of Crystallographic Orientational Variants in Yag-Al2O3 Directionally Solidified Eutectic Composite
Authors: Frazer Colleen S , Dickey Elizabth C
Published: July 2020

YAG-A12O3 directionally solidified eutectic composites show great promise in applications requiring high temperature strength and creep resistance.’ The orientation relationships between YAG and Al2O3 have been established within the composite by use of SEM Electron Backscatter Diffraction (EBSD) and x-ray diffraction (XRD). The orientation relationships of were determined. Two related variants of A12O3 were discovered in the sample. EBSD was employed to determine the spatial distribution of the variants within the sample.

The microstructure of the eutectic composite, shown in Figure 1, reflects a “chinese script” pattern, neither lamellar nor fibrous. The directionally solidified eutectic fibers, approximately 2.5 mm in diameter, were cut into disks normal to the fiber axis by a diamond saw and polished by with diamond-imbedded paper to reduce chances of phase-preferential wear. Samples used for EBSD were ion milled for 30 minutes at low power and incident angle to improve the patterns.
Identification of cu6sn5 and cu10sn3 Phases in Admixed Dispersal Phase Dental Amalgam by High Resolution Trasmission Electron Microscopy Combined with Convergent Beam Electron Diffraction.
Authors: Ramon Caballero , Amado Quintero , Oswaldo Hilders
Published: July 2020

Creep process in dental amalgams has been a subject of study. The performance of dental amalgams depends on the creep resistance of different products offered in the market. Creep process occurs at mouth temperatures which are slightly lower than Gamma 1 matrix phase (Ag2Hg3) melting point (310°K). One successful attempt to improve the creep resistance has been the addition of a dispersed phase (particles of eutectic Ag-Cu phase) in conventional amalgams. By this method, a new phase (η- Cu6Sn5) has been produced which inhibits the detrimental gamma 2 (Sn8Hg) formation. It has been proposed that the presence of finely distributed particles of η-Cu6Sn5 in the matrix lowers the creep rate. This behavior has been attributed to the pinning effect of these fine particles on the dislocations produced during the creep process (1). The presence of η-Cu6Sn5 phase finely dispersed in the matrix has been explained by the initial formation of reaction rings around the eutectic Ag-Cu spherical particles.
Apoptosis During the Knee Joint Development of Mouse
Authors: Hwang D H , Lim H S , Kim Y W
Published: July 2020

Apoptosis has been known as a significant element in the process of embryonic morphogenesis (1). Many studies on the role of apoptosis during development have been done using several organ systems, such as limb bud (2), nervous system(3), and inner ear (4). Recently, cell death with many morphological characteristics of apoptosis was reported in the developmental study of chick knee joint (5). This study was designed to observe the appearance of apoptotic cells during the joint cavity formation of mouse knee joint by transmission electron microscopy and TUNEL staining. The fetus were collected from l5thday(15D) to 19th day(l 9D) of pregnancy.

We observed many blood vessels and mesenchymal cells at 15D between the cartilages of femur and tibia. Mesenchymal cells had condensed heterochromatin and dilated rough endoplasmic reticulum (rER). At 16D, the joint clefts were first formed. Apoptotic cells, with characteristic nuclear fragmentation and condensation, appeared between the joint cleft and the capillaries.
Antioxidants Stimulate Germinal Vesicle Breakdown, But Inhibit Chromosome Formation And Spindle Assembly In Porcine Oocytes
Authors: Bailey GW , McKernan S , Price RL , Walck SD , Charest P-M , Gauvin R , Sun Qing-Yuan , Prather Randall S , Schatten Heide
Published: July 2020

Mammalian oocytes are arrested at the diplotene stage of the first meiotic division. Release of oocytes from their follicles induces meiotic resumption characterized by germinal vesicle breakdown (GVBD), followed by the chromosome formation and metaphase I spindle organization and finally the extrusion the first polar body. Recently it was shown that cellpermeant antioxidants significantly inhibit spontaneous resumption of meiosis in mouse oocytes, which may indicate a role of oxygen radicals in oocyte maturation. The regulation of mouse oocyte meiosis resumption is different from that of large domestic animals in that GVBD is independent of Ca2+ and protein synthesis. The present study investigated the influence of two cell-permeant antioxidants, 2(3)-ter-butyl-4-hydroxyanisole (BHA) and nordihydroguaiaretic acid (NDGA), on porcine oocyte meiosis resumption, chromatin behavior and spindle assembly. Our findings revealed a different role of antioxidants in porcine oocyte meiosis resumption than in mouse oocyte maturation.
Cortical Granule Exocytosis and Fertilization Coat Elevation is Reduced in Aging Sea Urchin Eggs
Authors: Bailey GW , McKernan S , Price RL , Walck SD , Charest P-M , Gauvin R , Chakrabarti Amitabha , Schatten Heide
Published: July 2020

Cortical granules are specialized Golgi-derived membrane-bound secretory granules that are located beneath the plasma membrane in unfertilized sea urchin eggs. Upon fertilization cortical granules discharge in a reaction induced by calcium and release their contents between the plasma membrane and a thin vitelline layer that lines the plasma membrane. Microvilli at the plasma membrane elongate incorporting cortical granule membranes during elongation. The vitelline layer elevates and becomes the egg's fertilization coat that hardens and serves as physical block to polyspermy. While we do not understand the precise mechanisms that participate in cortical granule discharge it is believed that actin plays a role in this process. Because actin and calcium metabolism is affected in aging cells we investigated if cortical granule secretion is affected in aging sea urchin eggs.

Lytechinus pictus eggs were obtained by intracoelomic injection of 0.5M KCI to release the eggs into sea water at 23°C.
Toward Measuring In-Vitro Single Molecule Force Fields Using a Scanning Force Microscope
Authors: Eppell S J , Todd B A , Zypman F R
Published: July 2020

The interaction of surfaces with their surroundings can be described as arising from force fields generated by molecules at the surface of interest. This statement is true for electrostatic interactions of proteins on cell membranes as well as steric interactions of macromolecules on polymer surfaces. To better understand these systems and ultimately to design superior materials, it is necessary to make direct measurements of these near-surface molecular scale force fields. Scanning force microscopy (SFM) holds some promise in the endeavor to make surface force measurements in a full three dimensional submolecularly resolved space. In-vitro measurements present a problem in that the force fields tend to be very short ranged. This necessitates placing the SFM probe very near the surface in order to measure the fields. The ubiquitous van der Waals attraction tends to induce an instability in the SFM instrument under these conditions leading to a non-equilibrium state. We have carefully examined this state of affairs and found that it is essential to use a substantially more sophisticated method to analyze SFM data than is commonly performed.
Dynamic Debonding Force Measurements of Ligand (RGD)-Platelet Receptor (GP Ilb/IIIa) System under Physiological Conditions
Authors: Lee ImShik , Marchant Roger E
Published: July 2020

ABSTRACT

Binding formation between a peptide sequence (GSSSGRGDSPA) which contains the cell adhesion sequence –RGD-found in fibrinogen, vWF, fibrinonectin, and vitronectin and human platelet intergrin GP Ilb/IIIa plays an important role in thrombus formation. Using atomic force microscopy (AFM), we visualized the detailed structures of membrane and submembrane of the cell, and measured the interaction forces between a peptide modified cantilever probe tip and platelet surface from pN to nN levels under physiological buffer. Direct measurements of the debonding force for the RGD ligand - GP Ilb/IIIa system are presented. To eliminate the possible measurement of the hgand-receptor pair, or of the ligand and AFM tip, following desorption of the ligand; the cantilever tip surface was modified with covalent coupling chemistry rather than physical absorption. Our results showed that the single molecular rupturing force was 93.3 ± 10.41 pN with considerable chain extension in the receptor. The rupturing forces showed a logarithmic dependence of the rate of loading
Imaging Cell Surface Macromolecular Distribution by Mapping Intermolecular Interactions with an Atomic Force Microscope
Authors: Bhatia R , Lin H , Quist A , Primbs G , Desai N , Lai R
Published: July 2020

An atomic force microscope (AFM) can image real-time intermolecular interactions between complementary macromolecules, such as receptor-ligand and antigen-antibody with nanometer resolution in hydrated state. Recent developments in AFM imaging allow mapping of such interaction forces over a large surface area such that local as well as global distribution of complementary biomolecules could be ascertained simultaneously. Using the force-volume maps and AFM probe conjugated with antibody, we have mapped the reorganization of specific receptors on the cell surface as well as the resultant changes in cellular micromechanical properties, such as elasticity and cytoskeletal reorganization of the cell (Figure 1).

In present study, we have mapped vascular endothelial growth factor receptor (VEGF-R) in the plasma membrane of cultured endothelial cells using anti-VEGFR - antibody conjugated to AFM probe. VEGF induced changes in cytoskeleton reorganization in endothelial cells were observed by real-time AFM imaging.
Microscopic Analysis of Porous Biodegradable Scaffolds for Tissue Engineering
Authors: Buevich F , Pulapura S , Kohn J
Published: July 2020

Introduction: There is considerable interest in the use of three-dimensional porous scaffolds for tissue regeneration. The presence of an interconnected framework of pores with large surface area facilitates the formation of extracellular matrix and permits cellular ingrowth into implanted structures. For scaffolds to be useful for tissue regeneration, they must maintain good dimensional stability during the lifetime of the implant. While the initial scaffold architecture is often well characterized, a systematic study of the influence of incubation on the scaffold architecture is critical to ensure that the scaffolds retain their interconnected network of pores during their useful lifetime. Herein, we report on the evaluation of the architecture of polyarylate scaffolds and their stability under in vitro conditions using scanning electron microscopy (SEM).

The polymers used in this study were selected from a library of degradable polyarylates. This library is the first reported combinatorial library of biodegradable condensation polymers.
Non-invasive video imaging for interrogating pharmaceutical crystallization processes
Authors: Wilkinson M J , Jennings K H , Hardy M
Published: July 2020

Crystallization is fundamental to much drug manufacture. Consequently a great deal of attention is focused on trying to attain a reproducible, robust process with predictable outcomes.

Crystallization may be interrogated by various means including laser light scattering, infra-red spectroscopy, calorimetery, x-ray diffraction, solid state NMR analysis and optical and electron microscopy. Although laser backscattering is applicable to real time, in-situ studies it detects chord length and hence it is most accurate when all particles are spherical. This is clearly is not so with drug crystals. Crystal analysis by microscopy is usually conducted on samples removed from the reactor. While this approach has value it may lead to post-sampling changes and is not capable of fully characterising the real time dynamics of particle morphology. In-situ epifluorescence microscopy of cells within a bioreactor has been reported. However, this required insertion of a quartz window into the reactor wall which limited the observation position to one fixed location, which would be less appropriate for batch crystallization reactors.
The Role of Microscopy in Preclinical Safety Assessment
Authors: Dovey-Hartman Barbara J
Published: July 2020

Microscopy plays a vital role in assessing the safety of New Chemical Entities (NCE) in the pre-clinical phase of drug development. Light microscopy (LM), transmission electron microscopy (TEM) and scanning electron microscopy (SEM) are used at the Schering-Plough Research Institute (SPRI) for evaluation of NCE. To support regulatory submissions, NCE are routinely tested in rodents in short-term studies such as one-month toxicity studies, and in longterm studies such as oncogenicity studies that may last 24 months. At the completion of a study, the animals are necropsied and the required tissues collected and stored in fixative. The tissues for LM are processed to slides and stained with Hematoxylin and Eosin (H&E). The information derived from the examination of these tissues by LM becomes an essential part of the pathology report. The LM examination of these tissues usually yields the information needed to either progress a NCE or otherwise deter or halt development.
“In-Situ” TEM Studies and “Real-Time” Digital Imaging
Authors: Alani R , Pan M
Published: July 2020

“In-situ” TEM studies is currently going through resurgence for materials microcharacterization. One important aspect of these studies has always been the image recording process. Traditionally, the following analog image recording techniques have been used: i) “static” images taken by photographic films and ii) viewing the images with a TV camera and recording them onto a videotape [1]. On the other hand, with the advent of CCD cameras and its rapid evolution for digital “static” imaging in TEM [2-3], “real-time” digital imaging for “in-situ” experimentation has been in high demand. In this work, we report the utilization of a new CCD camera for both “real-time” and “static” image acquisitions. The performance of the camera is shown for heating/cooling “in-situ” TEM experiments performed on an aluminum alloy using a double tilt hot stage.

We have utilized a newly designed CCD camera that addresses imaging needs for “in-situ” applications.
New Frame-Transfer Wide-Angle Slow-Scan CCD Camera Allows Recording of Distortion-Free Images for Digital Montages
Authors: Hiller SA , Probst W , Seybold V , Zellmann E , Kabius B , Trondle A
Published: July 2020

Modern SSCs combine low noise with high sensitivity and provide digital images with high dynamic and excellent linearity. Since these images are immediately available in a PC, SSCs have become a non-dispensable tool for on-line image processing and software driven automated TEM tuning [1]. Many experiments performed nowadays in the area of high resolution, low dose, holographic reconstruction, and EFTEM [2] would not be possible without the digital input coming from SSCs.

Despite their excellent characteristics SSCs provide a restricted number of individual image points in respect to film. This turned out to be the most prominent disadvantage of SSCs especially for applications where highly resolved large specimen areas need to be captured. Tab. 1 shows a typical example for a biological application where an image detail of 4 nm (plasma membrane) needs to be resolved. Depending on the film material used, this resolution is matched more or less by a TEM magnification of ∼8,000x.
Position Normalization as a Tool to Extract Compositional and Microstructural Profiles from Backscatter and Secondary Electron Images
Authors: Lee P J , Larbalestier D C
Published: July 2020

Position normalization is a method of manipulating data from any image source so that the effective information to noise ratio is dramatically increased. In this paper we present two very different examples of the application of position normalization used in the study of high current density superconductors. In the first example we use FESEM fractography to analyze grain boundary density and grain shape in micron-diameter filaments and then normalize the data obtained from individual grains to their positions with respect to the filament-matrix interface. In this way we extract the change of grain structure with position in apparently inhomogeneous microstructure. In the second example we analyze the backscattered electron intensity, BEI, from the cross-section of a superconductor filament with respect to its proximity to phase boundaries. In this way we extract small trends in the mean atomic number with a spatial resolution of better than 100 nm.
Effective Extinction Distances in Zone Axis Silicon
Authors: Yu Z , Vanfleet R R , Silcox J
Published: July 2020

Many scientific questions encountered in electron microscopy require quantitative deductions from the observations. Comparisons of experimental observations with simulations are however, still relatively rare since measurements of intensity are normally difficult. In this paper we discuss the use of experimental observations of the effective extinction distances for zone axis silicon using a convergent beam STEM mode for comparison with a number of simulations. On the experimental side, the measurements were made with a STEM that provides accurate intensity measurements directly with a digital image recording system. Two theoretical schemes widely used in electron microscopy simulations, multislice simulation and Bloch-wave calculation, were employed for the simulations. In each case, both a TEM case and a STEM case were calculated for comparison.

The multislice simulations were carried out using codes available from Kirkland. For the TEM case with plane waves at normal incidence on the sample surface, the unscattered (0,0) exit beam gives the Bright Field (BF) intensity.
In-Line Holography and Phase Amplification for Quantitative Phasecoherent Foucault Imaging
Authors: Volkov VV , Zhu Y
Published: July 2020

For magnetic flux mapping in magnetic materials it is important to use a simple, reliable and quantitative method operating with resolution down to few nanometers scale. The traditional off-axes electron holography can reach, in principle, such a resolution. However, the time-consuming phase reconstruction procedure limits its applications to real-time dynamic studies of magnetic domains behavior under varied external field. A novel recently developed in-line holographic approach, based on magnetic interferograms generated by phase-coherent Foucault (PCF)-imaging, complies with most of these demands and is suitable for dynamic studies of domain structure. For instance, it was successfully applied for induction mapping of closure domains in soft Fe-films [1]. It was found, however, that the resolution of PCF-imaging was lower than that of off-axis electron holography, since it is limited by the observed fringe spacing [2].

We report that the real resolution of PCF-imaging can be improved significantly with help of a novel phase amplification method.
Probability of Seeing (001) Cross-Fringes in a Random Cubic Nanocrystal Image
Authors: Qin W , Fraundorf P
Published: July 2020

As a crystal gets smaller, diffraction spots stay visible for larger deviations from the Bragg condition. The upper limit of such deviations is connected to the threshold for getting lattice fringes in TEM images. This in turn allows one to quantify the probability of seeing cross lattice fringes along a certain zone axis. In this abstract we examine a simple semi-empirical model for the probability of detecting (001) zone cross-fringes of a spherical crystal of cubic lattice.

The upper limit for the deviation of crystal orientation from the exact Bragg condition, without losing cross fringes down a given zone, is expressed as the maximum half-angle θ1 between the zone and the electron beam. The solid angle σ subtended by a cone with this half-angle is proportional to the probability px that a randomly-oriented crystal will show the cross-fringes associated with that zone. A schematic, illustrating the principle used to calculate the probability of seeing cross-fringes, is given in Figure 1.
Morphology and Orientation Relationships of Ge Precipitates in Ag-Ge Alloys
Authors: Luo CP , Dahmen U
Published: July 2020

Ag-Ge forms a simple binary eutectic with only two solid phases - a Ag-rich fee phase and a Ge-rich diamond-cubic phase, both with very limited solid solubility for the other element. Because both phases are cubic and there are no intermediate compounds, precipitation of Ge in an Al matrix might be expected to be crystallographically and morphologically simple. However, previous investigations of similar alloys (Al-Ge and Al-Si) have shown that precipitation in such apparently simple eutectic systems can be surprisingly complex [1,2]. For example, it was found that Ge precipitation in Al-Ge alloys depends sensitively on the availability of lattice vacancies and that at least five different orientation relationships and even more types of morphologies can form during aging after a quench. In the course of that work it was discovered that twinning is a key factor in the development of precipitate morphologies. To understand the role of twinning in precipitation this study investigates the characteristics of precipitation in Al-Ge alloys.
Atomic Structure of Y2O3:Eu/LaA1O3 Interfaces
Authors: Gao H-J , Fan XD , Kumar D , Cho KG , Singh RK , Pennycook SJ
Published: July 2020

There has been intense interest in preparing single crystalline yttrium oxide (Y2O3) thin films for applications in ultra-large scale integration (ULSI) gate insulators, ULSI capacitors, and, by addition of suitable dopant species, for electroluminescent devices. Y2O3 has a C-type rare-earth sesquioxide structure, closely related to the fluorite structure with a cell parameter a= 1.060 nm and space group Th (Ia3). LaAIO3 (LAO) is a rhombohedral structure with lattice parameters a = 0.378 nm, θ ≤ 90.5°. The lattice mismatch with the <110> direction of the YO is therefore less than 0.8%, and so we would anticipate epitaxial growth of single crystalline YO thin films on the LAO (001) substrate to be feasible. Eu activated YO thin films were deposited by laser ablation on (001) LAO substrates. TEM bright field images and electron diffraction patterns were recorded in a Philips EM-400 electron microscope operated at lOOkV. Z-contrast imaging was conducted in a VG HB603 STEM at 300kV. In this presentation, we will report the epitaxial growth of YO thin films doped with ∼ wt 4% Eu on a LaAlO3 substrate, and the atomic structure of the interface.