bluuebunny/crossref_metadata_2025_split huggingface dataset:
The organization of the microtubule associated protein tau in Alzheimer paired helical filaments
Authors: Ruben George C. , Iqbal Khalid , Grundke-Iqbal Inge , Johnson John E. 
Published: June 2020

Neurofibrillary tangles (NFT) of paired helical filaments (PHF) are the most characteristic brain lesions of Alzheimer disease and their abundance determines the diagnosis. This histopathological hallmark occurs with greater frequency in patients with clinical dementia of the Alzheimer type than in normal individuals of the same age. Therefore, it is important that we establish and understand the arrangement of subunits in PHF. This knowledge, we believe, could be key to understanding the pathogenesis of neurofibrillary degeneration.Intracellular Alzheimer NFT fixed in OsO4or KMnO4in the absence of glutaraldehyde treatment were first observed by transmission electron microscopy (TEM) of thin sections and reported by Kidd. The tangles he observed contained helical pairs of ~10 nm filaments separated center to center by 15 nm with a double helical period of ~160 nm that he named paired helical filaments. Subsequent thin sectioning work with a similar fixation treatment confirmed that the PHF were helical with crossover width, T=12.8±2 nm, and a wide region, W=24±3.1 nm, which occured every L=65−80 nm.
Correlative atomic-force microscopy and high-resolution scanning electron microscopy of proteins attached to platelet surfaces
Authors: Simmons S.R. , Eppell S.J. , Marchant R.E. , Albrecht R.M. 
Published: June 2020

The atomic force microscope (AFM) has provided images at submolecular or atomic scale resolution of biological macromolecules attached to surfaces such as mica, graphite, or synthetic phospholipid membranes. Because the AFM can be operated with the sample in air, vacuum, or immersed in a liquid such as a biological buffer, it has the potential for high resolution imaging of the structure and organization of macromolecules on surfaces of cells in the hydrated or even living state. Realization of this potential would allow observation of molecular processes at the cell surface without the necessity for preparation of the sample for electron microscopy. To date, however, the AFM has yielded images of cell surfaces only at relatively low magnifications, and has not provided the atomic resolution achieved on hard, crystalline surfaces.Previously we have utilized correlative video-enhanced light microscopy, high voltage transmission electron microscopy, and low voltage, high resolution scanning electron microscopy (HRSEM)
Development of high-performance objective lens polepiece for ultrahigh resolution Analytical Electron Microscope
Authors: Fukushima K. , Kaneyama T. , Hosokawa F. , Tsuno H. , Honda T. , Kersker M. 
Published: June 2020

Recently, in the materials science field, the ultrahigh resolution analytical electron microscope (UHRAEM) has become a very important instrument to study extremely fine areas of the specimen. The requirements related to the performance of the UHRAEM are becoming gradually severer. Some basic characteristic features required of an objective lens are as follows, and the practical performance of the UHRAEM should be judged by totally evaluating them.1) Ultrahigh resolution to resolve ultrafine structure by atomic-level observation.2) Nanometer probe analysis to analyse the constituent elements in nm-areas of the specimen.3) Better performance of x-ray detection for EDS analysis, that is, higher take-off angle and larger detection solid angle.4) Higher specimen tilting angle to adjust the specimen orientation.To attain these requirements simultaneously, the objective lens polepiece must have smaller spherical and chromatic aberration coefficients and must keep enough open space around the specimen holder in it.
Demonstration of GAD in purkinje-cell terminals with silver enhanced gold immunocytochemistry
Authors: Gilerovitch Helen G. , Bishop Georgia A. , King James S. , Burry Richard W. 
Published: June 2020

The neurotransmitter gamma-amino butyric acid (GABA) is synthesized in presynaptic terminals by glutamic acid decarboxylase (GAD). Antibodies to GAD have been used to localize presynaptic terminals in the cerebellar cortex that use GABA as a neurotransmitter. In the cerebellum, the Purkinje cell makes synaptic contacts on dendrites and somata of neurons in the cerebellar nuclei. We were interested in developing a technique that combined light microscopic analysis of GAD distribution in Purkinje cell terminals with high quality electron microscopic localization of GAD.The approach taken in this study was to combine high glutaraldehyde/metabisulfite fixation technique with the preembedding N-propyl gallate (NPG) silver enhanced gold procedure. Mice were perfused with buffered 3% glutaraldehyde and 0.4% sodium metabisulfite. Vibratome sections (60 μm) were permeabilized with 0.3% Triton, and incubated with a monoclonal anti-GAD antibody. The sections were incubated in goat-anti-mouse 1 nm Nanogold, and the gold was enhanced with the NPG silver enhancement solution for 10 minutes.
Immunoelectron microscopic localization of actin in developing rat leg muscle
Authors: Fassel Theresa A. , Greaser Marion L. 
Published: June 2020

Studies were conducted on the arrangement of α-actin in muscle cells in Sprague-Dawley rats at 1, 3, 5, 7, 30 and over 100 days of age. The α-actin isoform was distinguished from the nonmuscle β- and τ- isoforms by using the α-specific monoclonal antibody HUC1-1. The antibody was used in a two stage post-embedding procedure with colloidal gold conjugated to goat anti-mouse IgG.Muscle samples were placed in a relaxing solution (100 mM KCl, 10mM Imidazole, 10mM MgCl2, 2mM EGTA, 4mM ATP, 1mM NiCl2, 1mM PMSF, 100 ¼M leupeptin, 50% glycerol) for 1 hr on ice or overnight at -20°C. Samples were transferred to a skinning solution (100mM KCl, 10mM KH2PO4, 5mM MgCl2, ImM EGTA, 1% glucose, 0.2% Triton X-100, 0.02% NaN3) for 1 hr on ice and then into 3.7% formaldehyde in rigor buffer (RB - 75mM KCl, 5mM K2HPO4, 2mM MgCl2, 2mM EGTA, 2mM NaN3) for 1 hr for fixation.
Ultrastructural localization of antigenic sites in pancreatic tissue using in situ (metal-mirror) cryofixation
Authors: Lightfoot Fred G. 
Published: June 2020

There have been previously reported methods using in situ cryofixation via metal mirror freezing to study the ultrastructural characteristics of lung tissue while in its native state (inflated), as well as the elemental composition in subcellular compartments of the proximal tubule in whole (unexcised) tissue. Many studies have been done to determine the most appropriate way to conserve both antigenicity and structural integrity of various tissues. In most of these protocols there are steps which are likely to reduce or destroy the tissue antigenicity. In this report, an application using in situ cryofixation of pancreatic tissue for optimal ultrastructure and antigenicity will be described.Male rats were anesthetized with sodium pentobarbital (50mg/kg). The abdomen was opened and the pancreas was carefully elevated, using forceps, being careful to maintain integrity of the mesentery of the duodenum and pancreas along with their blood supply. The tissue was rapidly frozen using the PS1000 cryofixation unit (Delaware Diamond Knives, Wilmington, DE), a pneumatically operated metal mirror instrument precooled to nitrogen temperature.
Ultrastructural application of the patsc-GMS staining in various kidney diseases
Authors: Namimatsu Shigeki , Nakamura Shin-ichi , Nakagami Yoshizo , Oguro Tatsuo , Asano Goro 
Published: June 2020

We have previously reported a new staining method, PATSC-GMS, for transmission electron microscopy. In the present study, we evaluated the stainability and usefulness of the PATSC-GMS method in tissues from renal diseases.Fifty human kidney biopsies comprising normal (n=2), membranous glomerulonephritis (n=10), Alport's syndrome (n=l), IgA nephropathy (n=4), and other cases (n=33) were studied. Samples were pre-fixed in 2.5% glutaraldehyde, post-fixed in 1% osmium tetroxide, and embedded in Epok-812 after dehydration. Ultrathin sections were made and placed on nickel grid mesh. The sections of the tissues were Pre-incubated in ammonia water(25%) for 10minutes, oxidized in 1% periodic acid for 20 minutes, reacted in 0.1% thiosemicarbazid (TSC) for lminute, post-incubated in ammonia water(25%) for 5minutes and stained with gelatin methenamine silver(GMS) solution: (50ml of 3% methenamine, 5ml of 2% gelatin, 5ml of 5% silver nitrate, 40ml of distilled water, and 5-6ml of 5% sodium tetraborate) was performed for 50C 50minutes in the oven.
Localization of atrial natriuretic peptide in normal human atrial tissue by immunoelectron microscopy
Authors: Wei Chi-Ming , Hukee Margaret , McGregor Christopher G.A. , Burnett John C. 
Published: June 2020

Atrial natriuretic peptide (ANP) is a 28-amino acid peptide which is secreted from atrial myocytes and possesses natriuretic, vasoactive and renin inhibiting actions. In chronic congestive heart failure (CHF), plasma concentrations of ANP are significantly increased in the circulation secondary to enhanced atrial and ventricular synthesis. In a variety of non-human species, ANP has been detected by light and electron microscopy. However, the localization of ANP in normal human cardiac tissue remains unclear. Therefore, the current studies were designed to determine the localization of ANP in normal human atrial tissue.Cardiac tissue of three normal human right atria were obtained from the donor heart undergoing primary cardiac transplantation. Atrial tissue was fixed for 24 hours at 4°C in 4% formaldehyde and 1% glutaraldehyde in 0.1 M phosphate buffer. These specimens were rinsed in phosphate buffer, dehydrated in a series of ethyl alcohols, embedded in Quetol 651, and polymerized at 42°C for 48 hours.
Cytochemical localization of free-radical-derived oxidants in human coronary atherectomy plaques
Authors: Ann Ellis E. , Grant Maria B. , Wargovich Thomas J. 
Published: June 2020

Atherosclerosis is a multifactorial disease which leads to occlusion of vessels and potentially fatal intravascular thrombosis. Biochemical and immunocytochemical studies of oxidized low density lipoprotein (LDL) and treatment with antioxidants have demonstrated a role for oxidation in antherogenesis in animal models. The enzyme NADH-oxidase produces superoxide radicals (O2·-) and the free radical derived oxidant, hydrogen peroxide (H2O2). We used the cerium NADH-oxidase cytochemical localization technique to identify sites of free radical derived oxidation in coronary artery atherosclerotic lesions removed from patients undergoing coronary atherectomy.Atherectomy plaques were fixed in cold, sodium cacodylate buffered (pH 7.4) 5% acrolein within 5 min after removal from patients. Specimens were washed in buffer and processed as previously described for cytochemical localization of NADH-oxidase. Cross sections of the plaques were examined and photographed, without post staining, at 75 kV in the transmission electron microscope.NADH-oxidase activity, shown by cerium perhydroxide precipitate, was confined to the extracellular matrix and foam cells of the fatty streak.
Serial thin sectioning for pre- and post-embedding immunohistochemistry: Achieving consistent and reliable results for a 3-D reconstruction
Authors: Milroy Antonia M. 
Published: June 2020

In recent years many new techniques and instruments for 3-Dimensional visualization of electron microscopic images have become available. Higher accelerating voltage through thicker sections, photographed at a tilt for stereo viewing, or the use of confocal microscopy, help to analyze biological material without the necessity of serial sectioning. However, when determining the presence of neurotransmitter receptors or biochemical substances present within the nervous system, the need for good serial sectioning (Fig. 1+2) remains. The advent of computer assisted reconstruction and the possibility of feeding information from the specimen viewing chamber directly into a computer via a camera mounted on the electron microscope column, facilitates serial analysis. Detailed information observed at the subcellular level is more precise and extensive and the complexities of interactions within the nervous system can be further elucidated.We emphasize that serial ultra thin sectioning can be performed routinely and consistently in multiple user electron microscopy laboratories. Initial tissue fixation and embedding must be of high quality.
Ultrastructural study of basement membrane re-formation after intimal denudation
Authors: Yokoyama Munehiro , Ishiwata Toshiyuki , Asano Goro 
Published: June 2020

The basement membrane (BM) underneath the endothelial cell(EC) layer plays important roles in the maintenance of cell polarity and the regulation of cell migration, adhesion, proliferation and differentiation. The model of denudation of the aortic intima with a balloon catheter is useful in research into interactions among cell components and extra-cellular matrix in the regenerative process of ECs in atherogenesis. In this study, re-formation of BM was examined during re-endothelialization after intimal denudation, using transmission electron microscopy.Male Wistar rats, weighing about 400 g were used and endothelial denudation of the thoracic aorta was performed with a vascular balloon catheter (1.5 Fr., DOW CORNING, CORP). The catheter was inserted into the thoracic aorta via the left common carotid artery. After inflation of the balloon with saline solution, the catheter was passed through the length of the aorta about 2.5 cm three times. The animals were sacrificed at 15 min, 3, 7, 14, 21 and 28 days after endothelial denudation and the thoracic aortas were removed.
Adherence of enterotoxigenicKlebsiella pneumoniaeto hela cells and expression of type 3 fimbriae
Authors: Miliotis M.D. , Tall B.D. , Gray R.T. , Hall R.H. 
Published: June 2020

In 1986, an outbreak of diarrheal disease in Texas was attributed to contaminated iced tea in whichKlebsiella pneumoniaewas the predominant flora. Although the ten strains tested were of different serotypes, they all produced a heat labile enterotoxin. Some of these enterotoxigenic strains (ETKP) also exhibited mannose-resistant hemagglutinins, which were later found to be type 3 MR/K fimbriae. In this study we extended our observations and investigated fimbrial expression, as well as HeLa cell adherence behavior, using negative staining, immunogold labeling, and scanning electron microscopy (SEM).ETKP strains were grown on colonization factor antigen agar overnight at 37° C for expression of fimbriae. The presence of fimbriae and immunogold labeling of the cells was determined as described by Wolfeet al.Antiserum was prepared against purified fimbriae in rabbits and dilutions of primary and secondary antibodies (goat anti-rabbit IgG conjugated with 10 nm gold particles) in phosphate buffered saline, pH 7.2, containing 1 % bovine serum albumin and 0.5% Tween 20 were used to label cells.
Bacterial adherence and biofilm behavior ofVibrio vulnificus
Authors: Tall B. D. , Gray R. T. , Shah D. B. 
Published: June 2020

Vibrio vulnificus, an opportunistic human pathogen, is found as member of the normal microflora of shellfish and other seafoods, many of which are eaten raw. Though usually not harmful,V. vulnificusis responsible for causing fulminating septicemia in immunocompromised individuals. In previous light microscopic studies, we showed data suggesting that isogenic unencapsulated phase variants were more adherent to HeLa cells than were counterpart encapsulated phase variants. In this study, we extended our observations by comparing phase variant capsular morphology stained with Alcian blue (AB) and Ruthenium red (RR), and investigated the dynamics of biofilm formation by these organisms to glass coverslips (CS) using quantitative plate counts and scanning electron microscopy (SEM).To characterize the morphology of capsules expressed by these organisms, we stained cells grown on trypticase soy agar containing 1 % NaCl (TSA/NaCl) with AB and then prepared them for electron microscopy (EM) according to the method described by Hendleyet al.
Thickness measurement in quantitative Electron Microscopy
Authors: Heng Yew M. 
Published: June 2020

Quantitative electron microscopy generally implies the measurement of elemental content and/or structures in a given specimen or the 3D reconstruction of a structure. When a feature in a specimenis being quantified in a transmission electron microscope (TEM), the specimen thickness has to bedetermined for at least two reasons. Firstly, thickness is a required parameter in the calculation, for example, of volume and secondly, it is required for the calibration of instrumental conditions ortheoretical parameters such that correction for thickness effects due to conditions such as radiation damage, x-ray absorption or multiple scattering of electrons can be performed. However, it is a common knowledge that specimen thickness is difficult to be determined accurately. The difficulty is mainly due to the nature of TEM specimens. There is no universal method which can be used.Therefore, in order to select a few appropriate methods, it is necessary that a wide range of methods be reviewed and critically assessed.Numerous methods are available for the measurement of the thickness of a specimen to be analysed in a TEM (Table 1).
Data acquisition and processing for automated electron microscopy
Authors: Krivanek O.L. , de Ruijter W.J. , Meyer C.E. , Leber M.L. , Wilbrink J. 
Published: June 2020

Automated electron microscopy promises to perform many tasks better and faster than a human operator. It should also allow the operator to concentrate on the larger picture without having to worry about countless details that can be best handled by a computer. It requires three essential components: 1) data acquisition system that provides the computer with high-quality data on line, 2) computer and software able to analyze the incoming data in real time, and 3) control links that enable the computer to adjust the important microscope parameters.An optimized system architecture is shown schematically in Fig. 1. The microscope is equipped with various microprocessors that control its hardware, and provide data processing abilities devoted to different types of signals (e.g., X-ray spectra). These microprocessors use a standardized communication protocol to communicate over a standard network (such as AppleTalk or Ethernet) with a “master computer”, which provides the user interface, as well as the computing power necessary to handle the most demanding tasks.
HVEM tomography: A new tool for the cell biologist
Authors: Frank Joachim 
Published: June 2020

The technique of three-dimensional imaging with the transmission electron microscope is a new tool ready to be used by the cell biologist. By itself, the transmission microscope is not capable of producing 3D images, but it allows collection of a tilt series of projections in a wide angular range. From such a series, a 3D image can be reconstructed using established mathematical principles. The first reconstructions of this kind were obtained by Walter Hoppe’s group in 1974, but the full potential of this approach was not realized until the mid-80’s when computers with sufficient speed and memory became available.Especially within the past three years, there has been a rapid development in the areas of instrumentation, computation, and visualization. New tilt stages meet the demands of high accuracy and wide angular coverage. Direct image readout systems and on-line control of the microscope make it possible to design automated data collection systems.
STEM measurement of subcellular water distributions
Authors: Leapman R.D. , Sun S.Q. , Shi S-L. , Buchanan R.A. , Andrews S.B. 
Published: June 2020

Recent advances in rapid-freezing and cryosectioning techniques coupled with use of the quantitative signals available in the scanning transmission electron microscope (STEM) can provide us with new methods for determining the water distributions of subcellular compartments. The water content is an important physiological quantity that reflects how fluid and electrolytes are regulated in the cell; it is also required to convert dry weight concentrations of ions obtained from x-ray microanalysis into the more relevant molar ionic concentrations. Here we compare the information about water concentrations from both elastic (annular dark-field) and inelastic (electron energy loss) scattering measurements.In order to utilize the elastic signal it is first necessary to increase contrast by removing the water from the cryosection. After dehydration the tissue can be digitally imaged under low-dose conditions, in the same way that STEM mass mapping of macromolecules is performed. The resulting pixel intensities are then converted into dry mass fractions by using an internal standard, e.g., the mean intensity of the whole image may be taken as representative of the bulk water content of the tissue.
Energy-filtered reflection electron microscopy and reflection high-energy electron diffraction on Zeiss 912 TEM
Authors: LIU JINGYUE 
Published: June 2020

In reflection electron microscopy (REM) and reflection high energy electron diffraction (RHEED) the average path length of the elastically scattered electrons in the crystal ranges from 10 -100 nm and a significant portion of the electrons in the RHEED pattern spots used for imaging is inelastically scattered. The excitations of surface plasmons, bulk plasmons and valence electrons involves energy losses of 10 ∽30 eV. Thus the image contrast and resolution in REM are degraded due to chromatic aberration of the objective lens. The use of energy filters in a TEM should offer significant improvement in resolution and contrast of REM images. We present here some new results on the investigation of resolution limit and contrast mechanisms in energy filtered REM images.The experiments were performed on a Zeiss 912 TEM fitted with an Omega magnetic imaging energy filter. Digital RHEED patterns and REM images were acquired into 1024 pixels by 1024 pixels via a Gatan 679 CCD camera fitted to the microscope.
Compositional biological mapping with x-rays in the electron microscope
Authors: LeFurgey Ann , Kopf D.A. , Ingram Peter 
Published: June 2020

New ideas in microprobe analysis over the last decade have refuted the notion that electron probe xray imaging is limited to multispectral data as qualitative maps. Electron microscopy is still the most widely used technique for high resolution imaging of biological materials. The acquisition and processing of digital quantitative elemental images of one to several cells in an electron microscope is now accomplished with advanced, commonly available microcomputers. A system developed in our laboratories uses a graphics-based microcomputer to acquire spectrally filtered xray data that is fitted to standards, to display the data and images on-line and in real time, and to correct the multispectral image with regard to specimen drift. Statistics in the data acquisition is the limiting factor in both the image resolution and the ability to automatically recognize regions which need longer acquisition times.Except in routine clinical or diagnostic situations, specimen preparation requires the tissues or cells to be rapidly frozen in a viable state in order to preserve their inherent elemental and anatomical integrity.