bluuebunny/crossref_metadata_2025_split huggingface dataset:
Comparison of Focused Ion Beam and Conventional Techniques on Tem Specimen Preparation of Metal-Ceramic Interfaces
Authors: Ramirez de Arellano López A , Chiou W-A , Faber K T 
Published: July 2020

Abstract
The results of TEM analyses of materials are critically dependent on the quality of the sample prepared. Although numerous techniques have been developed in the last two decades, differential thinning of inhomogeneous materials remains a serious problem. Recently, focused ion beam (FIB) technique has been introduced for cross-sectional sample preparation for TEM and SEM.
A novel system for depositing a fine-grain (∼ 200 nm) ceramic coating on a metal surface via a patent pending Small-Particle Plasma Spray (SPPS) technique has been developed at the Basic Industry Research Laboratory of Northwestern University. To understand the properties of the coated surface, the ceramic/metal interface and the microstructure of the ceramic coating must be investigated. This paper presents a comparison of the microstructure of an A12O3 coating on a mild steel substrate prepared using conventional and FEB techniques.
Designing an Electron Microscopy Facility: Step by Step Procedure
Authors: Murphy Judy A 
Published: July 2020

Abstract
Designing a microscopy facility can be completed in a step by step approach, whether it be a completely new facility or an existing one moving into expanded quarters. The procedure is independent of size. There are several major factors that must be taken into consideration. Check lists can be compiled by the microscopist. One should easily be able to start at any point along the way. The time and effort initially put into the design will pay for itself many times over in convenience and efficiency of the resulting laboratory.
Prior to drawing the actual design of the microscopy facility but critical to the design specifics, it is necessary to compile the following information: considerations for determining the size of the facility; general EM lab requirements which affect site determination; activity functional flow diagram; EM room requirements; a list of all the equipment to be placed in the EM facility and exact specifications for each; design features not associated with specific equipment; activities requiring space that are not specific to the equipment; general building requirements not specifically related to microscopy operations; and ergonomic and psychological factors.
Some Simple Quantitative Applications of Vascular Corrosion Casting
Authors: Hossler Dr Fred E 
Published: July 2020

Abstract
The primary use of vascular corrosion casting has been and likely will continue to be for describing the 3-dimensional anatomy of the microvasculature of organs and tissues. In these studies the assumption is made that the corrosion cast faithfully replicates the anatomy, dimensions, and distribution of the vasculature of these tissues and organs. If this assumption is accurate, and it appears to be nearly so in tissues where measurements have been made, it follows that the casts should lend themselves to quantitative measurements. I summarize here some simple quantitative measurements we have made from corrosion casts. For these studies, cannulas were inserted into arteries leading to the organ of interest in anesthetized animals, and the blood was removed by flushing with Ringer solution or buffered saline, at physiological temperatures and pressures. Resin consisting of Mercox (or Mercox diluted with methyl methacrylate monomer) and catalyst was infused via the same cannula until the onset of polymerization.
Cryo-Electron Microscopy and Image Reconstruction of PBCV-1, an ALGAL Virus with T=169, Icosahedral Lattice Symmetry
Authors: Yan X , Olson N H , Van Etten J L , Baker T S 
Published: July 2020

Abstract
Paramecium bursaria Chlorella virus l(PBCV-l), the type species of the Pycodnaviridae family, is a large, plaque-forming virus that replicates in certain unicellular, exsymbiotic, Chlorella-like green algae. The virion (∼lxl09 daltons) has a 330,740 base pair dsDNA genome encapsulated within an icosahedral shell of ∼190nm diameter. Purified virions contain more than 50 different proteins, which account for 65% of the total virion mass and range in size from 10 to more than 200kDa. Vp54, the major capsid protein (40% of total virion protein) is a myristilated glycoprotein but it is not phosphorylated. Three other proteins are located on the virus surface. Virions contain 5-10% lipid, located inside the glycoprotein shell, which is required for virus infectivity. Little is known about the detailed ultrastructure of PBCV-1 virions, though some information has been obtained with sectioned, negatively-stained, and metal-shadowed specimens. We have used cryo-electron microscopy and three-dimensional (3D) image reconstruction methods to examine the native morphology of PBCV-1.
On The Unique Structural Organization of the Saccharomyces Cerevisiae Pyruvate Dehydrogenase Complex
Authors: Stoops James K , Zhou Z Hong , Schroeter John P , Kolodziej Steven J , Cheng R Holland , Baker Timothy S , McCarthy Diane L B , Yazdi Mohammed A , Maeng Cheol-Young , Reed Lester J 
Published: July 2020

Abstract
Dihydrohpoamide acetyl transferase (E2), a catalytic and structural component of a multienzyme complex that catalyzes the oxidative decarboxylation of pyruvate, forms the central core to which the other components are bound. We have utilized protein engineering and 3-D electron microscopy to study the structural organization of the largest multienzyme complex known (Mr ∼ 107). The structures of the truncated 60-mer core (tE2) and complexes of the tE2 associated with a binding protein (BP), and the BP associated with its dihydrohpoamide dehydrogenase (BP'E3) and the intact E2 associated with BP and the pyruvate dehydrogenase (E1) were determined (Figs. 1 and 2). The tE2 core is a pentagonal dodecahedron consisting of 20 cone-shaped trimers interconnected by 30 bridges.
Previous studies have given rise to the generally accepted belief that BP and BP'E3 components are bound on the outside of the E2 scaffold and that E1 is similarly bound to the core in variable positions by flexible tethers.
The Ribosome - Three-Dimensional Structure and Ligand-Binding Studies
Authors: Frank J , Penczek P , Malhotra A , Gabashvili I , Grassucci R , Heagle A , Srivastava S , Burkhardt N , Jünemann R , Nierhaus KH , Agrawal RK 
Published: July 2020

Abstract
To date, cryo-electron microscopy has become the most successful technique for exploring the structure of the ribosome and for studying binding positions of its various ligands, with the resolution slowly extending toward 10 Å. Obstacles in the attempts to improve resolution are the limited stability and coherence of the electron microscope, the statistics of data collection, and the conformational heterogeneity of the specimen. The last factor in this list proved to be the reason why it has been difficult to go past 18-20 Å with many specimens despite the use of state-of-the-art electron microscopes and inclusion of tens of thousand of projections.
A breakthrough has been achieved with a protein synthesis initiation-like complex in which mRNA and fMet-tRNA is bound to the E. coli ribosome. The high occupancy and extraordinary conformational homogeneity of this specimen has enabled us to reach a resolution of 15 Å.
Molecular Architecture of Ribonucleoprotein Vaults
Authors: Stewart P L , Rome L H , Kong L B 
Published: July 2020

Abstract
Vaults are ribonucleoprotein complexes highly conserved between species as diverse as Dictyostelium, rat, and human. Thousands of vault particles exist per cell in various tissues including liver, lung, kidney and spleen. These complexes have been found in the cytoplasm as well as associated with the nuclear membrane. Vaults have a large barrel-like middle section with a protruding cap on each end. These structures have an average mass of 12.9 ± 1 MDa, as measured by STEM, and are composed of 3 proteins, p210, pi92 and pi04, as well as an RNA strand of approximately 90 bases. The major vault protein, pl04, constitutes the majority of the particle mass, existing in 96 copies per structure. The major vault protein was recently recognized as the lung-resistance related protein (LRP), a known multidrug resistance gene that is over-expressed in certain cancer cells, including ovarian and myeloid leukemia cells.
A three-dimensional structure of the vault complex was generated using the QVIEW and IMAGIC-5 software packages (Fig. 1).
Macromolecular Assemblies Designed for Controlled Proteolysis
Authors: Walz J , Koster AJ , Tamura T , Baumeister W 
Published: July 2020

Abstract
Since cellular structures are rebuilt continually, protein degradation is essential for the maintenance of homeostasis. Misfolded proteins ensuing from genetic defects or environmental stress, are prone to aggregation; chaperones and proteases cooperate in minimizing such unproductive interactions. Last, but not least, protein degradation serves to terminate the lifespan of many regulatory proteins at distinct times and thus features as a key regulatory element itself. Proteins destined for degradation must be recognized and selected within the crowded environment of the cell. The stratagem of self-compartmentalization is key to controlling cellular proteolysis (1).
In recent years, a number of multisubunit proteolytic complexes have been described which possess large internal cavities or nano-compartments. This allows them to confine the proteolytic action to their interior; access to these inner compartments is usually restricted to the unfolded proteins. This, in turn, makes it necessary for these proteases to interact - either in a transient or in a permanent manner
Formation of Proteasome-PA700 Complexes Directly Correlates with Activation of Peptidase Activity
Authors: Gogol E , Adams G , Crotchett B , Slaughter C , DeMartino G 
Published: July 2020

Abstract
The eukaryotic 20S proteasome (700 kDa) serves as the core complex for the breakdown of most cytosolic proteins by the ubiquitin-dependent pathway. This cylinder-shaped proteolytic assembly (17 nm long, 11 nm diameter) consists of four stacked heptameric rings that help to form a central cavity where degradation occurs. The catalytic sites are located on the cavity surface of the two middle (3 rings while the outer a rings serve as docking sites for various regulatory complexes.
PA700 (700 kDa, 20 protein subunits) is the major regulatory complex of the proteasome, forming a “cap” that attaches to either or both proteasome ends. The resulting superassembly structurally resembles the larger, more active 26S proteasome and exhibits both a higher intrinsic peptidase activity as well as a wider range of substrate specificity, including ubiquitin-tagged proteins. A second regulatory complex, the modulator (300 kDa), further stimulates the effects of PA700 at subsaturating concentrations of the activator with no detectable effect on the proteasome in the absence of PA700.
Proteolytic Control of Bacteriophage HK97 Capsid Maturation.
Authors: Duda Robert L , Conway James F , Cheng Naiqian , Steven Alasdair C , Hendrix Roger W 
Published: July 2020

Abstract
HK97 is a tailed temperate bacteriophage of E. coli that builds an icosahedral capsid using steps that include regulated assembly, proteolysis, radical conformational changes and the formation of novel covalent bonds (Fig. 1). This pathway is being exploited as a model system to explore how the formation of multiprotein complexes can be regulated by each of these mechanisms. We have identified and purified at least four intermediates (Prohead I, Prohead II, Head I and Head II) and examined them by cryo-electron microscopy and three dimensional reconstruction procedures (Fig. 2). Comparison of particle reconstructions at resolution of about 25 - 30 A have lead to major insights into the causes and purposes of the regulated changes that we have also characterized biochemically and genetically.
Prohead I consists of 420 copies of the 42 kDa gp5 capsid protein arranged as 72 blister-shaped morphological capsomers in a thick walled hollow T=7 icosahedral particle with a diameter of -470 Å.
Dual-Labeled Probes for Fluorescence and Electron Microscopy
Authors: Powell Richard D , Halsey Carol M R , Gutierrez Edmund , Hainfeld James F , Furuya Frederic R 
Published: July 2020

Abstract
Fluorescein and the 1.4 nm Nanogold cluster have been separately attached to polyclonal antibody Fab’ fragments to generate combined fluorescent and gold secondary antibody probes (1, 2), and in comparisons with Nanogold, colloidal gold and fluorescein-labeled secondary antibodies, neither label was found to be significantly compromised by the presence of the other. The proximity to a gold label with which a fluorophore retains sufficient fluorescence emission intensity to be useful is limited by non-radiative fluorescence resonance energy transfer (FRET) (3). This is a function of the degree of overlap of the fluorophore emission spectrum with the metal particle absorbtion spectrum, and of the separation of the metal particle and the fluorophore (4); for an effective dual-labeled probe, separation should be greater than the Forster distance (3), at which 50 % of excited-state decay occurs by FRET.
Correlative Instrumental Neutron Activation Analysis, LM, And TEM to Track in Vivo Distribution of Colloidal Gold Spheres in BALB/C Mice
Authors: Hillyer JF , Albrecht RM 
Published: July 2020

Abstract
Colloidal gold is routinely used conjugated to ligands or antibodies as a label for the detection of cell structures by transmission (TEM) and scanning (SEM) electron microscopy. Silver enhancement, a procedure that amplifies the size of gold spheres, is used for light microscopic (LM) observation of colloidal gold. These techniques are valuable in localization studies but it is difficult to quantitate the level of labeling in larger volumes of tissue solely by microscopic means.
Instrumental Neutron Activation Analysis (INAA) can be used to provide a quantitative index of gold accumulation at the organ and tissue levels. Given that gold is not naturally found in the body in any substantial amount and that colloidal gold and ligand conjugates can be prepared to yield uniform bead sizes, the amount of label can be determined at the organ and tissue levels in whole animal systems.
Properties and Applications of EGFP, Enhanced Color Variants of GFP, and Unstable Derivatives of GFP
Authors: Kitts PA , Li X , Piston DW , Chervenak R , Kain SR 
Published: July 2020

Abstract
The green fluorescent protein (GFP) has great potential as a tool for biologists because it can be used as an in vivo real time reporter of protein localization and gene expression in a variety of experimental systems. Wild type GFP, however, has several undesirable properties including low brightness, a significant lag in the development of fluorescence, complex photoisomerization, inefficient protein folding at 37°C, and poor expression in several species. To improve upon these qualities, we have combined an ultra-bright variant of GFP, GFPmutl, with a synthetic gene sequence containing codons preferentially found in highly expressed human proteins. The combination of improved fluorescence intensity and higher expression levels yields an enhanced variant (EGFP) (Table 1) that greatly increases the sensitivity of this reporter.
The natural green emission of GFP can conveniently be monitored by optics designed to detect fluorescein. There are, however, many potential applications for GFP that require additional emission colors.
GFP Fusions for Fluorescence Detection of Ca2+ and Ca2+-Calmodulin in Living Cells
Authors: Persechini Anthony 
Published: July 2020

Abstract
We have previously described fluorescent indicators for Ca2+ (FIP-CAs) and (Ca2+)4-calmodulin (FIP-CBs) whose responses are based on a ligand-dependent decrease in fluorescence energy transfer (FRET) between GFP variants. The indicators for (Ca2+)4-calmodulin contain calmodulin-binding domains, those for Ca2+ also contain an integral calmodulin (CaM) domain. We have developed new versions of these indicators constructed with enhanced blue- and red-shifted GFPs suitable for stable and transient expression in mammalian cells, and have begun to use them to investigate the relationships between the free intracellular concentrations of Ca2+ ([Ca2+]i) and (Ca2+)4-CaM ([(Ca2+)4-CaM]i). When the blue-shifted fluorophore is excited at 380 nm these constructs exhibit an emission peak at 505 nm due to FRET to the red-shifted fluorophore.
We have made FIP-CBs with dissociation constants for (Ca2+)4-CaM of 0.5 nM, 20 nM, 300 nM and &gt; 20 μM by introducing R →Q substitutions in the CaM-binding sequence, and have stablyexpressed them in HEK-293 cells (Fig. 1).
Molecular pathology - Implications for Diagnosis and Pathogenesis of Infectious Diseases.
Authors: Zaki SR 
Published: July 2020

Abstract
Molecular pathology represents a distinctive new discipline in medicine with many potential applications that have been heretofore untapped. This discipline is introducing important changes in the concepts of disease causation and in the classification of diseases. Transfer of molecular genetic technology and knowledge to the pathology laboratory is giving the pathologist new and powerful tools to diagnose, classify, and monitor disease. Methodologies such as in situ hybridization (ISH), immunohistochemistry (IHC), and the polymerase chain reaction (PCR) will continue to allow more systematic and accurate classification of human diseases. IHC and ISH allow the identification and localization of cells containing specific antigens or nucleic acid sequences. The main advantage of these methods is their ability to localize targets and define their morphologic features in various tissues, allowing correlations with histopathologic changes.
Ultrastructural Characterization of Virus-Like Particles Produced by a Recombinant Baculovirus Expression Vector
Authors: Wasserman A J , Wang H , Clark G , Megill J R , Durham SK 
Published: July 2020

Abstract
Analysis of Hepatitis C virus (HCV) assembly and packaging, and the study of virus-cell interactions, have been impeded by the lack of a robust cell culture system and a viable small animal model. Recently, an alternative approach utilizing recombinant gene technology has been implemented. We describe a system in which specific HCV genes are cloned into a baculovirus to form a recombinant baculovirus expression vector that produces HCV proteins that self-assemble into enveloped virus-like particles (VLP). Four HCV structural genes were cloned into the baculovirus expression vector under the control of the polyhedrin promoter, and were then recombined with baculovirus DNA. A baculovirus with no HCV insert served as a negative control (wild-type). Ovary cells (Spodoptera frugiperda) were infected with either the recombinant or wild type baculovirus. Five-days post-infection, a group of cells were harvested, fixed in McDowell-Trumps solution, and routinely prepared for transmission electron microscopy (TEM).
Centrosome Proliferation in the Human Androgen-Responsive LNCaP and the Androgen-Independent DU145 Prostate Cancer Cell Lines
Authors: Schatten Heide , Ripple Maureen , Balczon Ron , Taylor Meghan , Crosser Michael 
Published: July 2020

Abstract
Cancer is a disease characterized by uncontrolled cell divisions in which the molecular controls for cytoskeletal regulation are bypassed. Cell division is governed by centrosomes, microtubule-organizing cell organelles which are crucial for the organization of the mitotic apparatus during mitosis and cell division. Because centrosome abnormalities are observed in the most common human cancers, we used immunofluorescence and transmission electron microscopy to determine centrosome organization in the human androgenresponsive prostate cancer cell line LNCaP and the androgen-independent prostate cancer cell line DU145. During interphase, centrosomes are located in close vicinity to the outer nuclear membrane, duplicate during S-phase, and become separated to the mitotic poles during the transition from interphase to mitosis. Centrosome regulation is based on a number of different factors which are only partly understood. Hormones play a role during developmental regulation of prostates which might trigger the activation of centrosome proteins and consequent cell divisions in order to ensure tissue growth.
Effects of Nitric Oxide on Monocrotaline Induced Right Ventricular Hypertrophy and Lung Mast Cell Hyperplasia in Rats
Authors: Hung Kuen-Shan , Duncan William H 
Published: July 2020

Abstract
Monocrotaline (M) induces pulmonary hypertension (PH) characterized by increased pulmonary arterial pressure, thickened pulmonary arterial walls, and right ventricular hypertrophy (RVH). In addition, numerous mast cells (MC) have been localized in M treated rats and these cells may have a role in causing PH and other lung injury because of their content of serotonin, which is a vasoconstrictor. Nitric oxide (NO) is a vasodilator that has been used in treatments of human patients and experimental animals with PH. The purpose of this study is to determine if NO inhalation can prevent or reverse RVH or MC accumulation induced by M.
Adult male Sprague-Dawley rats were divided into 5 groups (C, M3, MN3, M5, MN5). Group C served as controls without treatments. Groups M3 and M5 received one single subcutaneous injection of M (80mg/kg BW) and were sacrificed at 3 weeks (M3) or 5 weeks (M5) after injection.
Ultrastructural Diagnosis of a Pheochromocytoma at Autopsy
Authors: Siew S , Newton B 
Published: July 2020

Abstract
Pheochromocytoma is a rare tumor found in approximately 2 individuals per million of the population. In 90% of cases, it arises from the chromaffin cells of the adrenal medulla. These cells secrete catecholamines: epinephrine and norepinephrine which have a sympathomimetic action. The classical clinical presentation is paroxysms of extreme elevation of systemic blood pressure. Although pheochromocytoma accounts for only 0.1 % of cases of systemic hypertension, it is important to make the diagnosis, as the sudden release of large quantities of catecholamines into the circulation may prove fatal, as happened in our patient.
We present a case of a 25 year old woman, who collapsed suddenly, while she was feeding her 8 month old baby. She went into ventricular fibrillation and was defibrillated 3 times. There was no response to vigorous resuscitative measures.
Past Medical History: No definitive findings. Vague history of palpitations and an “abnormal heart beat”, which had been considered to be benign.
Quantitation of Peroxisome Proliferation using Silver Enhanced Immunogold Labeling on Glycol Methacrylate (GMA) Sections
Authors: Gagne GD , Illi AH , Hickman D , Fagerland JA 
Published: July 2020

Abstract
Assessment of peroxisome proliferation is important in drug safety studies, since peroxisome proliferation often precedes tumor development in rodents. Proliferation can be measured morphometrically on sections by immunostaining for the peroxisomal enzyme catalase. Previous studies have used tissue embedded in acrylic resins such as LR White, which requires tissue sizes of 1-2 mm. Glycol methacrylate (GMA) is an acrylic resin that preserves antigenicity in much larger pieces of tissue. In this study we applied immunogold-silver staining (IGSS) to GMA sections for the quantitation of peroxisome proliferation in rat livers.
Sprague-Dawley rats were given clofibrate, a known peroxisome proliferator, for 14 days. Livers were removed, fixed in neutral buffered formalin for 4 hours, and embedded in GMA. Sections (3 μm thick) were incubated with antibody to catalase followed by protein A-gold (PAG), and the gold label was amplified using a commercial silver enhancement kit. Tissue sections were counterstained with propidium iodide (PI).