bluuebunny/crossref_metadata_2025_split huggingface dataset:
Subcellular Localozation of TFPI in Human Umbilical Vein Endothelial Cells (HUVEC)
Authors: Olsen Randi , Webster Paul , Hansen John Bjarne 
Published: June 2020

Tissue factor pathway inhibitor (TFPI) is a serine protease inhibitor which regulates tissue factor (TF) induced blood coagulation by inhibiting the action of factor VIIa-TF complexes and factor Xa. In plasma, more than 80 % of the TFPI is bound to lipoproteins, 10 -20 % is carrier free, whereas platelets contains a small amount of TFPI. A major pool is probably assosiated with the vascular endothelium (1). This pool may have a crucial regulatory function because the endothelial surface serves as a binding site for factor VII/VIIa, mediating the activation of the extrinsic coagulation system.The normal distribution of TFPI within human umbilical vein endothelial cells (HUVEC) was examined on ultrathin cryosections labeled with specific antibodies and colloidal gold probes (2,3). Label was seen associated with the cell surface and with many intracellular structures. The surface label was patchy, the signal intensity varied between cells and between different parts of the same cell. Intracellular TFPI was found in many compartments including the Golgi complex (fig.l) a finding that was expected as these cells are the main site of TFPI synthesis, and endocytic organelles. Anti TFPI-antibodies were seen over organelles that also contained 3-(2,4-Dinitroanilino)-3'-amino-N-methyldipropylamine (DAMP), a compound that accumulates in organelles with low lumenal pH. DAMP was visualized using anti-DNP antibodies (fig.2). Structures that contained exclusively TFPI or DAMP were also seen, suggesting that TFPI did not enter all endocytic organelles and was not exclusively localized to low pH compartments. The TFPI-antibodies also colocalized with specific markers of endosomes (MPR) (fig.3) and lysosomes (LAMP-1) (fig.4)(4). TFPI colocalized with both, demonstrating the appearence of TFPI also in these endocytic compartments.
Three-dimensional imaging of nucleosomes from transcriptionally active genes using electron spectroscopic imaging
Authors: Czarnota G. J. , Bazett-Jones D. P. , Ottensmeyer F. P. 
Published: June 2020

The three-dimensional structure of the nucleosome was determined using particles purified from transcriptionally active genes in conjunction with electron spectroscopic imaging, and quaternion-assisted angular reconstitution procedures. The results reveal a configuration which is very different from the canonical compact crystallographic structure for this fundamental chromosome subunit, implying a structural disruption of the nucleosome with the activation of gene expression in accord with numerous physico-chemical observations.Previous analyses of nucleosomes purified from transcriptionally quiescent genes have indicated numerous structural states dependent on factors in vitro which modify charge based interactions in nucleoprotein complexes. Nucleosomes from transcriptionally active genes undergo chemical alterations in vivo which similarly modify charge based interactions. In order to investigate the effects of the gene expression associated chemical alterations on nucleosome structure, particles were purified from transcriptionally active genes using mercury affinity chromatography. These nucleosome particles are hyperacetylated with respect to particles from transcriptionally quiescent genes. Here additionally, sulphydryls normally buried within the protein core of the transcriptionally inactive particle are exposed to chemical modifying agents thus facilitating purification as described.
High resolution rotary shadowing of herpes simplex virus helicase proteins
Authors: Moretz R. C. , Crute James J. 
Published: June 2020

Herpes viruses cause numerous human diseases. Herpes Simplex viruses (HSV1 and HSV2), varicella zoster virus (VSV),cytomegalovirus (CMV), and Epstein-Barr Virus (EBV) cause conditions ranging from peripheral sores (HSV, VSV), generalized viremia (CMV) and recurrent malaise (EBV). The common targets for HSV are neurons in the CNS, peripheral nerve cells or plexuses of sensory ganglia in highly innervated epithelium (e.g. lips, genitals).Interfering with the mechanisms of infection and virus production depends on the ability to target specific stages of viral reproduction. Helicases are enzymes that are essential for the duplication of viral DNA. The origin binding protein (OBP) helicase (encoded by the DNA replication gene UL9) binds to sequences in DNA at the three replication origins. These origins are points within the HSV chromosome where DNA replication initiates. The protein is a 83kDa polypeptide which exists as a dimer. Although the role of the UL9 helicase in HSV DNA replication has not been exactly determined, by analogy to other eukaryotic origin binding replication proteins, the binding and enzymatic action of the UL9 helicase to the origin sequences may initiate the assembly of a multiprotein replication complex. Helicase-primase (the product of the DNA replication genes UL5. UL8 and UL52) acts to unwind double-stranded DNA at growing replication forks. The helicaseprimase activity is expressed by a 213kDa heterodimer consisting of the 99kDa UL5 and 114kDA UL52 subunits. ICP8 (SSB—UL29 gene product) is a single-stranded binding protein of 122kDa that complexes with OBP (two SSB/OBP dimer). Each of these gene products has been expressed in insect cells with a recombinant baculovirus and purified to near homogeneity. Assays have been established for the screening of compounds that could inhibit the helicase activity of HSV 1 helicases. Analysis of helicase/DNA substrate complexes under different conditions will be facilitated by a better understanding of the structure of the individual proteins.
SEM image sharpness analysis
Authors: Postek M. T. , Vladar A. E. 
Published: June 2020

Fully automated or semi-automated scanning electron microscopes (SEM) are now commonly used in semiconductor production and other forms of manufacturing. The industry requires that an automated instrument must be routinely capable of 5 nm resolution (or better) at 1.0 kV accelerating voltage for the measurement of nominal 0.25-0.35 micrometer semiconductor critical dimensions. Testing and proving that the instrument is performing at this level on a day-by-day basis is an industry need and concern which has been the object of a study at NIST and the fundamentals and results are discussed in this paper.In scanning electron microscopy, two of the most important instrument parameters are the size and shape of the primary electron beam and any image taken in a scanning electron microscope is the result of the sample and electron probe interaction. The low frequency changes in the video signal, collected from the sample, contains information about the larger features and the high frequency changes carry information of finer details. The sharper the image, the larger the number of high frequency components making up that image. Fast Fourier Transform (FFT) analysis of an SEM image can be employed to provide qualitiative and ultimately quantitative information regarding the SEM image quality.
How to get the most of a SEM using the CASINO monte carlo program
Authors: Drouin D. , Hovington P. , Gauvin R. , Beauvais J. 
Published: June 2020

High resolution scanning electron microscopy (SEM) has become more and more popular for several reason. First, the new field emission (FE) source of electron is now able to produce a high current density electron probe even at very low energies. It is possible to perform analysis with spatial resolution comparable to one obtain in conventional transmission electron microscope (TEM). The major inconvenient of TEM analysis are related to sample preparation. Fabrication of electron transparency film is generally a complex and time consuming process.To fully exploit a SEM, microscope users must have an accurate knowledge of the electron matter interaction. In order to help microscope users determine the appropriate microscope conditions, Monte Carlo simulation programs have been developed. Those programs, use random number sequences and complex models to accurately determine the signals recorded in a SEM (X-Ray and Backscattered electron). With the new generation of powerful personal computers, it is now possible to rapidly predict SEM outputs corresponding to the user’s microscope conditions. The CASINO program is a shareware that can be used to fully exploit a SEM.
Investigation of low-loss spectra and near-edge fine structure of polymers by peels
Authors: Heckmann W. 
Published: June 2020

Transmission electron microscopy has changed from a purely imaging method to an analytical method. This has been facilitated particularly by equipping electron microscopes with energy filters and with parallel electron energy loss spectrometers (PEELS). Because of their relatively high energy resolution (1 to 2 eV) they provide information not only on the elements present but also on the type of bonds between the molecular groups. Polymers are radiation sensitive and the molecular bonds change as the spectrum is being recorded. This can be observed with PEEL spectrometers that are able to record spectra with high sensitivity and in rapid succession.A PEEL spectrum can be divided into a low loss range and an inner shell loss range of higher energy. The low loss spectra of polymers always show a broad peak at about 22 eV and a further peak at 7 eV, if aromatic groups are present, as is the case with PS (Fig. 1). In the course of exposure, the intensity of this peak decreases, a sign that the benzene ring is destroyed by the radiation (Fig. 2).
Extraction and identification of fillers and pigments from pyrolyzed rubber and tire samples
Authors: Sadhukhan P. , Zimmerman J. B. 
Published: June 2020

Rubber stocks, specially tires, are composed of natural rubber and synthetic polymers and also of several compounding ingredients, such as carbon black, silica, zinc oxide etc. These are generally mixed and vulcanized with additional curing agents, mainly organic in nature, to achieve certain “designing properties” including wear, traction, rolling resistance and handling of tires. Considerable importance is, therefore, attached both by the manufacturers and their competitors to be able to extract, identify and characterize various types of fillers and pigments. Several analytical procedures have been in use to extract, preferentially, these fillers and pigments and subsequently identify and characterize them under a transmission electron microscope.Rubber stocks and tire sections are subjected to heat under nitrogen atmosphere to 550°C for one hour and then cooled under nitrogen to remove polymers, leaving behind carbon black, silica and zinc oxide and 650°C to eliminate carbon blacks, leaving only silica and zinc oxide.
A shear stabilized biaxial texture in a lamellar block copolymer
Authors: Polis D. L. , Pinheiro B. S. , Lakis R. E. , Winey K.I. 
Published: June 2020

Block copolymers spontaneously self-assemble into a variety of morphologies. Recent studies have produced a biaxial texture in poly(styrene-b-ethylene propylene), SEP, diblock copolymers by applying oscillatory shear. This biaxial texture consists of “parallel” lamellae (normal to lamellae aligned perpendicular to shearing surfaces) and “transverse” lamellae (normal to lamellae aligned parallel to shearing direction) according to small-angle X-ray scattering, SAXS. The present study has determined how these two populations of lamellae are arranged and how they relax upon quiescent annealing by examining the superstructure via FE-SEM.Sample preparation for the JEOL 6300 FE-SEM was designed to preserve a sample's orientation with respect to the shearing direction and provide a large viewing area (.1 - .3 mm2) relative to what is typical with cryo-ultramicrotomed sections for TEM. Bulk samples were cut and mounted on a specimen stub with one of the 3 orthogonal directions of the shearing geometry oriented perpendicular to the stub surface.
TEM study of a biofilm on copper corrosion
Authors: Chiou W. A. , Kohyama N. , Little B. , Wagner P. , Meshii M. 
Published: June 2020

The corrosion of copper and copper alloys in a marine environment is of great concern because of their widespread use in heat exchangers and steam condensers in which natural seawater is the coolant. It has become increasingly evident that microorganisms play an important role in the corrosion of a number of metals and alloys under a variety of environments. For the past 15 years the use of SEM has proven to be useful in studying biofilms and spatial relationships between bacteria and localized corrosion of metals. Little information, however, has been obtained using TEM capitalizing on its higher spacial resolution and the transmission observation of interfaces. The research presented herein is the first step of this new approach in studying the corrosion with biological influence in pure copper.Commercially produced copper (Cu, 99%) foils of approximately 120 μm thick exposed to a copper-tolerant marine bacterium, Oceanospirillum, and an abiotic culture medium were subsampled (1 cm × 1 cm) for this study along with unexposed control samples.
Fracture investigation of Fe-Zn alloy coating on steel sheets after deformation
Authors: Lin Yuli , Hou Wen-Shen 
Published: June 2020

Steel coating technology has made great progress in recent years to meet the demand for corrosion-resistant steel. Fe-Zn coated steel sheet is one of those which has been developed with increasing production in response to the brisk demand in the automobile, construction and household electrical appliance industriesThe adhesion of coating to substrate is a critical property of any coating system for mechanical integrity and environmental protection of the substrate. The investigation of fracture on the coating layer and/or on any interfaces after deformation, specially the initiation of failure provides fundamental acknowledge to understand major factors contributing to the adhesion of coatings. This paper presents the results of fracture on Fe-Zn alloy coatings on steel sheets after deformation.
Stabilizing the nanostructure in ball-milled iron alloys through the addition of oxide precipitates
Authors: Doğan C.P. , Rawers J.C. 
Published: June 2020

High-energy ball milling has proved to be an effective means of producing nanostructured metal alloys from elemental powders. Interest has focussed on this processing method because of its potential to form metastable compositions with a combination of properties not attainable by more conventional processing techniques. Consolidation of the resulting powders into commercially-viable solids has proven difficult, however. Because of the metastable nature of the compositions formed and the extremely high boundary area that makes up the nanostructure, these materials are very susceptible to exposure to elevated temperatures. Since most conventional consolidation techniques involve high temperatures, the nanostructure within the powders is typically lost during this processing step. Thus, a reliable means of enhancing the thermal stability of these materials, allowing economical consolidation, is clearly needed.One possible method to enhance thermal stability is to introduce a homogeneous distribution of stable second phase particles that will pin the grain boundaries, allowing retention of the nanostructure at temperatures high enough for effective consolidation.
Imaging focused x-rays from glass capillaries
Authors: Steel Eric B. , Jach Terrence 
Published: June 2020

Recent developments of glass capillaries allow x-ray probes to be focused down to micrometer sizes. These probes show promise for microanalysis applications, if they can reliably deliver high intensity, well formed x-ray beams to a specimen. We have developed an optical bench for viewing and evaluating the focused x-rays emitted from these capillaries. The optical bench allows pulse counting, direct viewing of the x-ray probe spatial distributions, and energy pass band by using exchangeable detectors. Our results are used to determine the probe forming capabilities of the lenses and to indicate problems and directions for improving the manufacturing of the x-ray optics.The x-ray optical bench is shown in a schematic diagram in figure A. It consists of a conventional CuKα x-ray diffraction tube joined to an optical table. An optical rail extends across them. A shielded box with a hinged side covers the source and the optical rail. The x-ray optic and exchangeable detector system are located on computer controlled stages to allow fine adjustments in all necessary translation and tilt directions. This optical bench permits measurements to be devised, performed, and changed with a minimum of difficulty. The optical bench allows for maximum flexibility in sample positioning, detector positioning, source size, and source collimation. The quantities that we evaluate in each optical element include focal spot size, spot uniformity, total spot flux, spot aberrations, diffuse scattered radiation, and depth of field or convergence.
Imaging Deep Optical Sections with Two-Photon Excitation Fluorescence Microscopy
Authors: White J.G. , Centonze V.F. , Wokosin D.L. 
Published: June 2020

The technique of optical sectioning allows the visualization of a succession of images of parallel planes within a thick specimen with little or no out-of-focus interference. Ultimately, a limit is reached on the depth to which optical sections can be obtained from a given sample. This limit, up to the working distance of the objective, is largely determined by the degree of light scattering encountered by the incident excitation beam as well as the returning emission signal.Confocal imaging was one of the first optical sectioning techniques applied to fluorescence imaging. Two-photon excitation imaging is a recently developed alternative optical sectioning technique for fluorescence imaging where an excitation wavelength of around twice the excitation peak of the fluorophore is used in a laser-scanning microscope. This excitation wavelength produces very little fluorophore excitation in the bulk of the sample, but when the incident photons are confined in space and time sufficient two-photon absorption events can take place to obtain rapid imaging of fluorophores. With high peak powers—obtained with a sub-picosecond pulsed laser source focused by a lens—sufficient photon density can be obtained for easily detectable two-photon events. Thus fluorophore excitation occurs as two photons are absorbed essentially simultaneously, which act effectively as a single photon of twice the energy (half the wavelength). Two-photon events have a quadratic dependence on intensity, and, therefore, decrease rapidly away from the focal volume of the lens. In a raster scanning system, fluorophore excitation is confined to the optical section being viewed as fluorophore away from the lens focal volume is not excited by the long-wavelength illumination.
Analysis of Cellular Organelles and Macromolecular Assemblies by Electron Energy Loss Spectroscopy
Authors: Leapman R.D. , Andrews S.B. 
Published: June 2020

Recent advances in electron energy loss spectroscopy (EELS) have significantly extended the range of applications for biological microanalysis. For example, EELS can now detect physiological concentrations of the important element calcium in rapidly frozen cells with a sensitivity greater than that achievable by energy-dispersive x-ray spectroscopy (EDXS). It can also detect small numbers of phosphorus atoms bound to macromolecular assemblies, and measure water distributions in frozen hydrated tissue. Here we discuss some of these developments in the context of detection limits and mapping techniques in the scanning transmission electron microscope (STEM) and energy-filtering transmission electron microscope (EFTEM).The useful information about elemental composition in EELS of biological specimens generally resides in weak core-edge signals corresponding to atomic concentrations in the 10−5−10−3(1–100 mmol/kg dry weight) range. For example, the Ca L2,3signal/background ratio is typically only 10−3and it is necessary to measure differences in signal that are only 104 of the background. Changes in low-loss fine structure corresponding to varying chemical composition are also very subtle; for example, detection of a 3% change in water content requires reliable measurement of a 0.1 eV shift in the low-loss intensity maximum. To extract such information requires efficient parallel detection of the energy loss spectrum and a high-brightness source to provide a sufficient number of incident electrons. The dedicated STEM is particularly well-suited for analyzing low concentrations of biological elements. If desired, the probe current can be reduced into the picoampere range for low-dose, high-resolution imaging prior to elemental analysis. The STEM’S field-emission source can then be used to deliver a current approaching 10 nA into a ~10 nm diameter probe. High electron flux conditions are ideal for spectrum-imaging applications where adequate counting statistics must be achieved within a limited pixel dwell time. The cold field-emission source of the STEM has the additional advantage of providing electrons with a narrow energy spread of <0.5 eV which is important in fine structure studies.
Calcium Quantitation with A Parallel EELS/Cooled CCD System
Authors: Ho Ruoya , Shao Zhifeng , Somlyo Andrew P. 
Published: June 2020

The value of electron energy loss spectroscopy (EELS) for analyzing low concentrations of phosphorus and calcium, in biological cryo-sections, has been well documented. With currently available techniques, the sensitivities of EELS for phosphorus and calcium can be three to ten fold higher than those of electron probe microanalysis (EPMA).Our experiments were performed with an EELS system including a Philips CM200 electron microscope equipped with a field emission gun and a STEM unit, and a GATAN 666 PEELS. We replaced the original fiber optically coupled photodiode array on the PEELS with a cooled CCD camera with lens coupling for better performance. The microscope is operated at 200 keV, allowing us to use relatively thick cryo-sections (λi = 1700 Å for carbon at 200 keV). We also used the shortest camera length in the STEM unit for achieving higher collection angle. According to our experience, the 2 mm aperture on the PEELS provides the best compromise of energy resolution and collection angle. Under these conditions, the collection semi-angle of this system is about 7 mrad and the energy resolution (full width at half maximum without specimen) is about 2.2~2.4 eV. The main limitations on using a larger collection angle are the 90 mm column between the viewing screen of the microscope and the entrance aperture of the PEELS and the poor energy resolution obtained with larger PEELS aperture.