bluuebunny/crossref_metadata_2025 huggingface dataset:
CIV, An Iridovirus With a Pseudo T=147 Capsid Structure
Authors: Yan X , Olson N H , Bergoin M , Rossmann M G , Baker T S
Published: July 2020

Chilo iridescent virus (CIV), the type species of the genus Iridovirusin the family Iridoviridae,was first isolated from the rice stem borer, Chilo suppressalis,a member of Lepidoptera insect order. ‘2 CIV is being considered as a potential pest control agent because of its wide host range which includes leafhoppers and some arthropods. The virion (∼lxl09 daltons) encapsidates a 209 kbp, dsDNA genome within its fiber-covered, icosahedral shell. The virion is composed of about 26 structural proteins, two (P50 and P'50, 50kDa each) of which are the major capsid proteins. P50 migrates as a monomer in either a reducing or non-reducing SDS-PAGE gel and is exposed to the surface, whereas P'50 exists as a disulfide-bond, stabilized trimer in a non-reducing gel and is only partially exposed. Virions contain 9% lipid present as a lipid bilayer within the protein capsid.
Infection of Tick Cells By Two Nonpathogenic Strains of Rickettsia
Authors: Palmer Ann T , Munderloh Ulrike G , Kurtti Timothy J
Published: July 2020

Ticks harbor several different genera of endosymbiotic bacteria. These obligate intracellular bacteria, most of which appear to be nonpathogenic for the tick, persist in nature by being transmitted transovarially from generation to generation. Those in the genus Rickettsiabelong to the spotted fever group (SFG) of rickettsia which includes both mammalian pathogenic and nonpathogenic species. The species that cause spotted fevers in humans, e.g. Rickettsia rickettsii, are well characterized but those not known to cause disease remain poorly understood. Transmission electron microscopy on SFG rickettsia cultured in tick cell cultures offers a powerful way to examine dynamic host parasite interactions in these apparently nonpathogenic SFG species.

We have successfully isolated two unknown, nonpathogenic rickettsia strains in tick cell lines. T2 was isolated from an Ixodes ricinus tick collected in the English Garden in Munich, Germany and is propagated in an Ixodes scapularis cell line, ISE6.
First Report of a Putative Cyanophage, Mc-1, of Microcoleus Sp.
Authors: Rosowski J R , Shaffer J J , Martin E L , Kokjohn T A , Lee K W
Published: July 2020

A current review (1) lists 21 cyanophages of coccoid and filamentous heterocystic and non-heterocystic blue-green algae (BGA). The capsid diameters of these viruses range from 50-90 nm, and none have been previously reported for Microcoleus,a sheath-producing, filamentous, surface colonizing BGA (Fig. 1), of the order Oscillatoriales, and class Cyanophyceae (= Cyanobacteria). From a freshwater culture of Microcoleussp. we report here a putative cyanophage with a side-to-side hexagonal capsid diameter of 140 nm, based on measurements from thin sections of algal filaments prepared for transmission electron microscopy (TEM).

An earlier report (2) documented the ultrastructure of mucilage secretion, and its role in sheath, trichome and filament formation in an isolate of the BGA Microcoleus sp. grown in soil-water medium. Now we report large cyanophage-like particles in a majority of cells making up the trichomes. These putative hexagonal viral capsids (Figs. 2-5) are mainly clustered in the center or to one side of the nucleoplasm.
Infrared Spectroscopic Micro-Imaging of Macromolecular Leakage In Acute Lung Injury Tissue
Authors: Wang J , Fata P , Yip B , Oppenheimer L , O’Connor R , Stimpson R , Mantsch H H
Published: July 2020

Acute lung injury is defined as a syndrome of inflammation resulting in increased capillary-alveolar permeability that is associated with a constellation of clinical, radiological and physiological abnormalities. Adult respiratory distress syndrome (ARDS), relatively common in the intensive care, is a more specific, severe form of ALI, with diverse causes. Diffuse alveolar damage is the pathognomonic feature of the syndrome. Under the pathophysiologic conditions of increasing capillary membrane permeability, large macromolecules can gain access to the interstitium, resulting in pulmonary interstitial edema, and from there enter the alveolar space in severe cases. Detecting the distribution of macromolecules in lung tissue, therefore, constitutes evidence of the injury. Traditional pathologic examination of biopsy/autopsy materials is the only route to provide pathologic evidence of the capillary-alveolar membrane damage induced lung injury. Although morphometric analyses are more laborious, they provide qualitative assessment of morphological changes that is an important aspect of acute lung injury, see Fig.l.
Comparison of Ultrastructural Morphology of Tissue fixed in Formalin and Prefer™
Authors: Billings Steven D , Goheen Michael P
Published: July 2020

Ultrastructural examination is sometimes necessary for diagnosis of surgical pathology material. Not uncommonly, tissue is not fixed in 3% glutaraldehyde prior to realizing that electron microscopy would be valuable or even necessary to render a diagnosis. It is well known that formalin can be an adequate fixative for ultrastructural evaluation of tissue. Increasingly, non-formalin based fixatives have become more popular with routine surgical pathology practice. Prefer™, a glyoxal-based fixative, has become a popular substitute for formalin in many institutions. We are not aware of any study that has compared ultrastructural morphology of tissue fixed in formalin to tissue fixed in a glyoxal-based fixative.

To address this question, we took sections of fresh tissue from three different tumors (metastatic melanoma involving a lymph node, squamous cell carcinoma of the lung, and mixed papillary serous and clear cell carcinoma of the ovary) and fixed a portion of each in 3% glutaraldehyde, 10% neutral buffered formalin, and Prefer™.
Skeletal Muscle Ultrastructural Alterations In Alcoholism
Authors: Müller B , Finol HJ , Montes de Oca I , Mayorca. A
Published: July 2020

Two forms of muscular alterations have been described in alcoholic patients, one acute, the so called Hypokalemic alcoholic Myopathy; focal or diffuse pain, swelling, tenderness and weakness of skeletal muscle are the main clinical features, and the chronic one with proximal muscle weakness wich progresses slowly in a period of weeks or months Ultrastructurally myofibril disorganization and necrosis where described.

In this work we present a systematic study of skeletal muscle alterations in four alcoholic patients who attended the Department of Internal Medicine at Caracas University Hospital. The patients were males, ages between 56 and 62 years old, and presented chronic alcoholism with muscle weakness with a more distal distribution and polymyalgia. Biopsies were taken from quadriceps femorismuscle with a percutaneous neddle and processed for routine transmission electron microscopy.

Muscle fibers showed different degrees of atrophy with myofibril disorganization (Fig.l) and disappearence (Fig.2). Myonuclei were hyperchromatic and intermyofibrilar spaces were widened and exhibited abundant lipid droplets.
Kidney Ultrastructural Pathology In Plasmodium Berghei Murine Malaria: Tubular Cell and Peritubular Capdllary Alterations
Authors: Pulido-Méndez M , Finol HJ , Rodríguez-Acosta A , Márquez A , González N , Andrade G , Boada-Sucre A
Published: July 2020

Renal function impairment can be found in severe or complicated P. falciparum malaria. Some studies report 2-5% incidence of acute renal failure associated with high mortality (1). Causes of acute renal damage are not clearly understood but massive hemoglobinuria seems unlikely in the majority of cases and different immunopathogenic mechanisms leading to tubulointerstitial nephritis are currently proposed (2). Ultrastructure of renal lesions in malaria have been performed in humans (3) and in some infections of rodents and monkeys (4). These studies report abnormalities mainly in the glomerular area and references to tubular cell pathologic changes are still limited. This work describes tubular and capillary alterations in P. berghei malaria.

P. berghei parasitized erythrocytes (strain from the Swiss Tropical Medicine Institute) were inoculated intraperitoneally in C57BL/6 male mice. Renal samples were obtained at 13th-14th days post-infection when severe illness was evident and parasitaemia reached 40-50%. Samples were processed by routine techniques for transmission electron microscopy and observed in Hitachi H-500 and H-7100 electron microscopes.
Successful Management of Infectious Pathogens From Microbial Outbreak Specimens: Safety and Identification
Authors: Humphrey Charles D
Published: July 2020

Despite significant advances in molecular biology and immunology, electron microscopy (EM) continues to be useful for morphologic identification of infectious microbes in body fluid and cultured specimens from humans and experimental animals. Morphologic identification using negative stain and/or thin-section EM is often a primary tool in public health investigations, followed by immunologic, molecular, or cultured isolation. Difficulties encountered are usually related to the length of time between specimen (e.g. stool) collection and preparation for EM and/or handling and shipping conditions. Another negative mitigating factor may be multi-lab interventions prior to arrival. Appropriate specimen collection, storage, and handling prior to attempts to identify the agent are important factors for successful identification.

Barrier-control guidelines called “Universal Precautions for Laboratories“ have for many years been recommended by U.S. and international health services organizations. The guidelines are based on the premise that safe work sites result from a combination of engineering controls, management policies, work practices, and, when required, prompt medical intervention.
Vascular Regression In Gill Filters of Tadpoles of Xenopus Laevis Daudin: A Qualitative and Quantitative Sem Study on Vascular Corrosion Casts
Authors: H. Bartel  , B. Minnich  , I. Margreiter  , A. Lametschwandtner
Published: July 2020

During metamorphosis amphibian tadpoles regress gills, tail and other larval structures. Thus gills are a good model to study how a highly complex three-dimensional vascular network regresses and finally disappears. Presently, only the microvascularization of external gills of larvae of Hynobius  dunni(2) and Litoria ewingi(3) is examined. The present study analyzes the spatio-temporal changes which the gill 3D-vascular network undergoes during metamorphosis using SEM of microvascular corrosion castings and a newly developed 3D-morphometry system (4, 5).

Vascular casts of gills of tadpoles of the South African Clawed Toad, Xenopus laevis Daudin of stages 57 to 62 (6) were analyzed by SEM. Stereopaired images (tilting angle: 6 degrees) grabbed with a slow scan frame grabber (Orion 4.27) from the SEMs photo-display were imported into a 3Dmorphometry system (6) to measure vessel diameters, interbranching distances, branching angles and branching orders of filter row venules. This study reports the changes in the branching orders only.
Physiological Aspects of Vascular Corrosion Casting
Authors: Olson KR
Published: July 2020

Vascular corrosion replicas provide unique three-dimensional models of both macro- and microcirculatory angio-architecture. In most replication procedures, blood is initially flushed from the vasculature with a low viscosity, non-clotting medium, such as saline, prior to infusion with the unpolymerized resin. While this removes cellular elements from the vessels and prevents coagulation, the chemical characteristics of the saline and the mode in which it is infused can potentially affect vascular smooth muscle. This can be disadvantageous if subsequent distribution of the resin does not faithfully mimic normal perfusion pathways, or if the vascular lumen is either narrowed due to vasospasm or distended due to excessive intravascular pressure. Alternatively, the composition of the saline can be modified to enhance the replication process. This may be done merely to ensure complete filling of the vasculature or to evaluate anatomical foci instrumental in physiological and pharmacological regulation of perfusion distribution.
Exact Detection of Arterial and Portal Branches of the Terminal Afferent Vessels in the Rat Liver By Means of Corrosion Casts and Edx
Authors: Spornitz Udo M , Bartuskova Irena , Morson Gianni
Published: July 2020

The physiological necessity of the dual afferent blood supply of the liver and the morphological features of the macrocirculation are well understood. The microcirculation, however, still poses a number of unsolved questions. In particular the nature of the terminal junctions of the arterial and portal blood stream with the sinusoids has so far not been satisfactorily elucidated (1-2). Scanning electron microscopy alone or in combination with corrosion casts cannot solve the problem, because it remains difficult to tell the different vessels apart. Under favorable circumstances the terminal portal vein is the easiest to be distinguished with corrosion casts. This is due to the fact that its relatively wide branches surround the classical liver lobules along the major parts of their circumference. The diameter of the terminal hepatic artery is in its initial segments not as wide as the initial segments of the terminal portal vein.
Angioarchitecture of Palatine Mucosa of Young Rabbits : A Sem Study of Vascular Corrosion Casts
Authors: Kronka Marcia C , Watanabe Ii-Sei
Published: July 2020

The angioarchitecture of the hard palate mucosae of young rabbits was investigated by using microvascular casting of low density resin (Mercox®). The injected specimens were examined by scanning electron microscope (SEM). The palatine mucosa and the adjacent gingiva have an important function during the mastication, vocalization and swallowing. The purpose of this paper is to analyse the structure of the micro vasculature of the palatine mucosa.

The animals were anesthetized and perfused with approximately 500ml of prewarmed (37°C) heparinized saline. Then, 30-40ml of Mercox® CL2-B, prepared conforming the manufacterer's intructions, were carefully injected via aorta artery under manual pressure. After the complete resin polymerization, specimens were dissected and macerated with 10% NaOH for several days at room temperature. The vascular casts were rinsed in distelled water, dried, mounted in stubs, coated with gold and examined in JEOL 6100 scanning electron microscope.
Triggered Morphology Generation in a Biosynthetic Model Spider Dragline Silk Protein
Authors: Valluzzi R , Szela S , Kirschner D , Kaplan D
Published: July 2020

Recombinant DNA techniques were used to prepare a protein modeled after the consensus sequence of Nephila clavipesspider dragline silk, incorporating methionine residues to serve as redox “triggers”. In addition a water-soluble 27 residue peptide model of the dragline silk consensus amorphous sequence, representing a single amorphous block in the protein sequence, was prepared and characterized to gain additional insight into the behavior of the amorphous phase. X-ray diffraction, electron diffraction, transmission electron microscopy (TEM), and Fourier transform infrared spectroscopy (FTIR) were used to characterize the ability of the recombinant protein to form (β-sheet crystals and the effect of the oxidation state of the redox trigger on crystallinity and noncrystalline order in the sample. The formation of intractable β-sheet crystallites is a major cause of insolubility in proteins that can form this type of secondary structure. Changes in crystallinity were observed when triggered/reduced (insoluble) and untriggered/oxidized (soluble) protein samples were compared.
Strategies For Immunodissection Of The Connective Tissue Matrix And Basement Membranes
Authors: Keene Douglas R , Tufa Sara F
Published: July 2020

Connective tissue matrices are quite diversified and include that composing skin, tendon, bone, cartilage, cornea and many others. The main component of the connective tissue matrix is collagen, composing approximately 70% of the dry weight of the human body. More members of the collagen family are discovered each year, with over twenty types described to date. Many of these collagens are tissue specific. In addition to the collagens, the connective tissue matrix is also the residence of epithelial and endothelial basement membranes and many other molecules including a variety of proteoglycans and elastic fiber components. The cells within the matrix are highly differentiated and task specific.

Using immunocytochemical technique applied at the level of the electron microscope, we have focused our resources to understand the structure and functional role of these matrix molecules.
Live Tissue Video-Microscopy Of Different Vascular Beds In The Eye: A Microcosmosm Of Untouched Microcirculation
Authors: Becker Matthias D , Martin Tammy M , Planck Stephen R , Rosenbaum J T
Published: July 2020

The eye offers the unique possibility of direct visualization of microvessels in readily accessible tissues. Due to the compact anatomy of the eye, different microvascular beds of different embryological origin can be examined in albino mice without traumatic tissue preparation, Epifluorescence microscopy in combination with different fluorescent dyes has been applied. To observe individual cells in vessels, rhodamine 6G can be used to stain leukocytes and platelets, and acridine orange stains nuclei of leukocytes and endothelial cells. Fluorescein-isothiocyanate (FITC)-dextran labels plasma.

Iris, limbus, conjunctival and choroidal vessels under normal and inflammatory conditions (endotoxin-induced uveitis) were examined in BALB/c mice and Lewis rats. Digital video technology can be applied to intravital microscopy with great benefit: video quality remains on a high level and the amount of time for the quantification procedure and the need of expensive software is reduced compared to classical video recording.
Astrosclereids In Needles Of Old-Growth Douglas-Fir Trees
Authors: Apple M , Soeldner A , Hamill R , Tiekotfer K
Published: July 2020

Old-growth Douglas-fir trees in the Pacific Northwest are venerable giants that often live for 500 years and reach heights of over 75 meters. Their needles are relatively ephemeral and small but have the important role of interacting with the atmosphere in order to transpire and photosynthesize. Within the photosynthetic mesophyll tissue of Douglas-fir needles, there are large, non-living cells with lignified secondary cell walls that are known as astrosclereids. Apparent channels in the secondary wall may provide a route for exchange or transport of materials between the astrosclereid lumen and mesophyll cells or the vascular cylinder. Astrosclereids may be involvev d in storage of secondary metabolites such as tannin and may develop in response to fungi, mistletoe, or other pathogens. More knowledge is needed about the development, structure and function of astrosclereids.

Needles were collected from sapling and old-growth Douglas-fir, Pseudotsuga menziesii, (Mirb.) Franco, trees at the Wind River Canopy Crane in Carson, Washington and from three sites in the Cascade Mountains of Oregon in 1997 and 1998.
Visualization Of The Transcription Factor Agl15 In Plant Embryos: A Case Of Regulation Via Cellular Localization
Authors: Fernandez DE , Perry SE , Lehti MD
Published: July 2020

AGL15 (AGAMOUS-like 15) was originally identified as a sequence preferentially expressed in developing plant embryos by comparing PCR-generated ‘differential displays’ of embryo and non-embryo RNA populations. The sequence is expressed at low levels and encodes a member of the MADS domain family (named for the first four members to be identified: MCM1, AGAMOUS, DEFICIENS, and SERUM RESPONSE FACTOR) of regulatory factors. Members of the MADS domain family play pivotal roles in critical developmental events in a wide variety of organisms: in specification of mating type in yeast (MCM1), in responses to the environment in human cells (SRF), in control of mesoderm differentiation in Drosophila and muscle differentiation in humans (MEF2), and in determination of meristem and organ identity during flower development in plants (AGAMOUS, DEFICIENS/APETALA3, GLOBOSA/PISTILLATA, etc.). AGL15 is the only MADS domain factor identified to date that is expressed in a highly preferential manner in plant embryos.
Morphological And Immunocytochemical Analysis Of Polyhydroxybuterate Distribution In Transgenic Plants
Authors: Colburn SM , Biest N , Hao M , Houmiel K , Mitsky T , Padgette S , Reiser S , Slater S , Tran M , Valentine H , Gruys K
Published: July 2020

Poly-β-hydroxybuterate (PHB), a biodegradable polyester, is accumulated as a storage compound in many species of bacteria. It belongs to a class of polymers called polyhydroxyalkanoates (PHAs). PHA's were originally identified in 1923.However these polymers first came to the attention of industry in the early 1980's when they were recognized as having thermoplastic properties.Monsanto currently markets a form of PHA produced via fermentation under the trade name Biopol™. It is used to make biodegradable plastic products such as credit cards, bottles and disposable drinking cups. However fermentation is an expensive route to production. The use of crop plants genetically engineered to produce PHA's could provide a less expensive source of the plastic and therefore a more attractive alternative to traditional non-biodegradable petrochemical derived plastics. PHB production in plants was first demonstrated in 1992 in transgenic Arabidopsis thaliana by Chris Somerville and coworkers. They showed that PHB production was tolerated by the plant if it was targeted to the chloroplasts.
Chloroplast Ultrastructure And Absorption Properties Of The Alga Phaeocystis Antarctica Karsten: A Qualitative Study Using Electron Tomography
Authors: Moisan Tiffany A , Ellisman Mark , Sosinsky Gina
Published: July 2020

Understanding the light-harvesting properties of algae and higher plants are a fundamental topic in photosynthesis research. Much oceanographic research has focused on characterizing the in vivo chlorophyll-specific absorption coefficient, a*ph (λ) in phytoplankton because it serves as an input variable for bio-optical modeling of photosynthesis using remote sensing instrumentation such as moorings, drifters, and satellites. Values of a*ph (λ) vary spectrally and the magnitude depends on accessory pigments, photo-protective pigments, and pigment packaging effects. Several studies have shown that the contribution of cellular characteristics to a*ph(λ) varies with growth conditions including temperature, light, and nutrients. It has been shown that a*ph (λ) values in Phaeocystis vary predictably at 4°C over light intensities under light limitation. Phaeocystis demonstrated significant pigment package effects that depended on single cell diameter and thylakoid membrane stacking. Using thick sections obtained from fixed and embedded cultures of colonial P. antarctica, we calculated tomographic reconstructions of individual chloroplasts under light-limiting conditions for net photosynthesis in order to gain an understanding of the continuity of thylakoid membranes and understand the spatial relationship between the pyrenoid, the starch containing organelle, and thylakoid membranes.
Colocalization Of Markers Of The Endocytic And Biosynthetic Pathways In Multivesicular Bodies Of Murine Alveolar Type Ii Epithelial Cells
Authors: Na Cheng-Lun , Weaver Timothy E
Published: July 2020

Immunocytochemistry studies have established that multivesicular bodies (MVBs) in alveolar type II epithelial cells are important for processing pulmonary surfactant protein precursors (proSP; Voorhout et al., 1992 and 1993). Localization of pulmonary surfactant protein A (SP-A) gold and cationic ferritin to the MVBs demonstrated that MVBs also serve as endosomal/lysosomal compartments for recycling SPs (Williams et al., 1984, Ryan et al., 1989). However, these studies did not elucidate whether the biosynthesis of pulmonary SPs and endocytosis occur in the same MVBs. In this study, we used primary murine alveolar type II epithelial cell cultures as a model to determine if the biosynthetic and endocytic compartments colocalized in MVBs. The endocytic compartments of MVBs were identified using BSA gold conjugates, while the biosynthetic pathway was detected with antibodies directed against proSPs.

Alveolar type II epithelial cells isolated from 6 weeks old C57B/6 mice were grown on Transwell cell inserts in the presence of 6 nm BSA gold conjugates (BSAG6) adjusted to final concentration of OD520 5.0 in DMEM, 25 mM HEPES, and 10% fetal bovine serum, pH 7.3, at 37 °C for 20 hours.
Ultrastructure of Candida Albicans And Saccharomyces Cerevisiae After Treatment With Sch 56301 (An Aureobasidin)
Authors: Dovey-Hartman BJ , Cramer C , Greene J , Shaw KJ , Hare RS , DiDomenico B
Published: July 2020

Aureobasidins are cyclic depsipeptide natural products that have been shown to be fungicidal against a wide variety of fungi. We have studied the effect of an aureobasidin-like compound, SCH 56301 on the morphology and cytology of yeast cells. For both, Candida albicans and Saccharomyces cerevisiae, the normal cell cycle is disrupted by sub-lethal concentrations of the compound. In Saccharomyces, cells accumulate at the G2/M border of the cell cycle, DNA replication is essentially complete and a short nuclear spindle is visible; however, nuclear migration and spindle elongation do not occur and consequently a small round daughter cell remains attached to the mother cell. Changes in cell wall morphology are visible at old bud scars remaining on mother cells. Under the conditions of preparation of the specimens, there may have been insufficient cross-linking during treatment with SCH 56301 so that intracellular pressure appears to be pushing on a weakened cell wall at these points.
Centrosome Structure And Function Is Altered By Experimental Manipulations With Formamide: Implications For Abnormal Cell Divisions During Cancer
Authors: Schatten Heide , Hueser Christopher N , Chakrabarti Amitabha
Published: July 2020

The formation of abnormal mitosis associated with cancer has been intriguing for many decades. While microtubules had been the focus of previous studies, recent research has focused on centrosomes, microtubule organizing centers which organize the mitotic apparatus during cell division. During normal mitosis centrosomes form two poles but in cancer, centrosomes can form three, four, or more poles, and organize tripolar, quadripolar, and multipolar mitoses, respectively. This has severe consequences for genomic stability because chromosomes are separated unequally to three, four, or more poles. This can result in aneuploidy and gene amplifications with multiple defects in cellular regulation. It can result in malignancy that is accompanied by cell cycle imbalances and abnormal cell proliferation. While radiation and chemical agents are known to damage DNA and can lead to cell cycle abnormalities, the damage of centrosome structure leading to abnormal mitosis deserves also consideration.
Mapping Phosphorus In Macromolecular Assemblies At Near Single Atom Sensitivity By Stem-Eels
Authors: Leapman RD , Rizzo NW
Published: July 2020

In principle, electron energy loss spectroscopy (EELS) in the scanning transmission electron microscope (STEM) should be able to detect single atoms of phosphorus in biological macromolecules. Such a capability would provide a direct method for measuring the degree of phosphorylation or nucleotide binding which are important in determining the functional states of protein assemblies. Although it has already been possible to map heavy elements, e.g., uranium, at near single atom sensitivity, this has not yet been achievable for lighter atoms like phosphorus. One difficulty in imaging light atoms in a biological specimen is that there is insufficient contrast to visualize the atoms directly so, in a stationary probe analysis, it is not known where to place the probe. Identifying single phosphorus atoms therefore necessarily involves compositional mapping in which EELS information is collected pixel by pixel, a technique known as spectrum-imaging. Recently, improvements have been made in the acquisition of such images, especially the ability to correct for specimen drift in a robust fashion.
Rapid Procedure For Detecting Scrapie-Associated Fibrils In Chronic Wasting Disease Of Elk And Mule Deer
Authors: Hearne CE , Clapper JL , DeVries KL , Williams ES
Published: July 2020

Chronic wasting disease (CWD) is a transmissible spongiform encephalopathy (TSE) of mule deer and elk. Affected animals are characterized clinically by a combination of abnormal behavior and gradual loss of body condition. The disease is invariably fatal. Diagnosis of CWD is made by histologic examination of the central nervous system and microscopic lesions of CWD are typical of the TSEs. Brains from CWD-suspect elk and mule deer can be examined by transmission electron microscopy (TEM) for the presence of scrapie-associated fibrils (SAF). Western blot methods identify abnormal prion protein (PrPres) which accumulates in the brains of animals with TSE, including CWD.

Conventional SAF purification procedures for TEM examination of CWD-suspect brain tissue require both high speed and ultracentrifugation steps followed by Proteinase K enzyme treatment. Modifications used in this experiment include early Proteinase K, or Proteinase K and Collagenase treatment prior to high speed centrifugation and the elimination of the ultracentrifugation step.
Glucose-6-Sulfate Is A Negative Stain/Sustain
Authors: Massover W H
Published: July 2020

It is increasingly suspected that negative staining should be capable of imaging protein structure at much higher resolution levels than are usually achieved. The heavy metal salts traditionally used as negative stains give structural information only to 20-25Å (2.0-2.5nm); this limit probably is caused by several defective properties of these compounds. If new reagents without these defects can be identified, then higher resolution details should be imaged. The finding that some light atom salts can be used successfully for negative staining greatly increases the range of candidate compounds available for evaluation.

A covalently modified monosaccharide, glucose-6-phosphate, recently was shown to function as an unconventional negative stain. This natural organic reagent directly combines the desirable structurepreserving actions of glucose with added scattering from phosphorus and potassium atoms. It is not known if sugar derivatives containing other light atoms in place of phosphorus can serve as a negative stain.
Age-Related Changes In Acetylcholinesterase Activity In The Efferent System Of The Cochlea Of C57bl/6 Mice
Authors: Cohen Glenn M , Shah Bela A
Published: July 2020

C57BL/6 mice, along with other mouse genotypes, have served as models of human presbycusis. C57BL/6 mice suffer from progressively severe hearing impairments, attributable primarily to losses of both hair cells (sensory receptors) and spiral ganglion cells (afferent neurons). Their age-related hair cell degeneration in the cochlea begins in the basal turn and progresses to the apex.

Auditory nerves also undergo age-related changes. The mammalian cochlear nerve is mixed, being composed largely of afferent (sensory) fibers and a small population of efferent fibers. Because efferent nerves are primarily cholinergic, we used acetylcholinesterase (AChE) as a marker enzyme for determining both age-related changes in AChE activity of the efferent fibers and also for tracing degenerative patterns in efferent pathways within the organ of Corti.

Twelve C57BL/6 mice of the following ages were used: four young (23 days), four middle-aged (13 months), and four old (22.5 months) mice.
Utility Of Secondary Guard Hairs In Animal Hair Identification
Authors: Yates B C
Published: July 2020

Variability among hairs of humans is dependent on body source and ethnicity. Variability in the hairs of non-human mammals is also dependent on somatic source, but may exhibit a wide range of characteristics among hairs from any given site. Humans do not have secondary guard hairs as defined below, but forensic scientists involved in trace analysis must recognize a different suite of variability that exists when examining animal hairs. Hairs other than primary guard hairs from the dorsum, which are often the only representative hairs illustrated in animal hair atlases, may be present as trace evidence in a forensic case in which the origin of the hair is unknown. An overview of the different grades of animal hair that could be present in a forensic case or natural history inquiry places the secondary, tertiary, etc. guard hairs in a continuum that is based on relative diameters.
Typical Application In A Metallurgical Failure Analysis Laboratory
Authors: P.E. Bruce LWong
Published: July 2020

Metallurgical failure analysis is generally straight forward in many cases. However, there are times when it is difficult to really discern the fracture characteristics due to surface degradation, improper handling, or subsequent mechanical damage from improper planning.

Dr. Alan Tettleman, a founder of Failure Analysis and Associates, in San Jose, CA, once told me you can not properly conduct any product failure analysis without metallography, and especially without scanning electron microscopy. In many ways, I agree with him. There are obvious times, however, when various physical properties and deformations are apparent. Those are generally when a consultant is not required. As in your field, there are many references that assist the usage and implementation of the scanning electron microscope and energy dispersive x-ray analysis system. Many of the references introduce the procedure and some of the cautionary steps that should be taken in preparation of samples and in the interpretation of the fracture.
Insights Into The Past: The Use Of Microscopy In The Conservation Of Cultural Materials.
Authors: Claire Dean J
Published: July 2020

Probably an example of every type of material ever manipulated and utilized by man through out history will eventually find its way to a conservator for treatment and preservation. The term “art conservator”, commonly used to describe those that work to protect and preserve the physical remains of mankind's invention and creativity, is a misnomer. We are better described as conservators of cultural materials, as our work involves the preservation of a huge range of materials and artifacts, many of which would not qualify as “art” under its most accepted definition. From the sublime (a Leonardo da Vinci cartoon) to the ridiculous (a 19th century saloon singer's dress made entirely from rattlesnake skins), from the ancient (paleolithic cave paintings) to the new (a pair of Nike® Air Jordans), if they are in a collection somewhere, they are candidates for examination by a conservator.

Despite the variety of materials and artifacts facing us, professional conservators use common approaches to the care of collections and we share the same arsenal of tools.
Mapping Organism Expression Levels at Cellular Resolution in Developing Drosophila
Authors: Knowles David W , Biggin Mark D , Richards Stephen , Sudar Damir
Published: July 2020

Sequence specific transcription factors are the predominant regulators of animal gene expression controlling nearly all biological processes. We are developing novel quantitative optical imaging techniques to map gene expression levels at cellular and sub-cellular resolution within an entire organism. Pregastrula Drosophila embryos have been chosen because these embryos allow high resolution 3D optical imaging since they comprise a single layer of dividing cells surrounding a yolk sac. in addition, the transcription network controlling gene expression is well characterized in early Drosophila embryos[1], and is being further dissected by a multi-laboratory collaboration, the Berkeley Collaboration in Drosophila Genomics, which encompasses this work.

Embryos at different stages of development are labeled for total DNA and specific gene products using different fluorophors and imaged in 3D with confocal microscopy (Figure 1). Intensity-based segmentation of the total DNA image[2] produces a nuclear mask which defines the nuclear boundaries, their location and the number of cells within the embryo (Figure 2). Presently, dilation of the nuclear volumes into their nearest-neighbours[3] is used to estimate the boundary of the cell (Figure 3) and superposition of these images produces a morphological mask defining each cell and its nucleus.
Enzyme-Dependant Fluorescence Recovery After Photobleaching (ED-FRAP): Application to Imaging Dehydrogenase Activity in Living Single Cells
Authors: Combs Christian A , Balaban Robert S
Published: July 2020

Fluorescent recovery from photobleaching coupled with confocal microscopy was explored as a potential high-resolution method of imaging the distribution of enzyme activity in single living cardiac myocytes without relying on steady state measurements of fluorescence. On a fundamental level, much remains to be determined regarding how local conditions within a cell affect metabolism. Many studies have suggested that energy metabolism in muscle cells cannot be accurately described assuming a homogenous system of enzymatic reactions [1,2]. The autofluorescence of NADH has been used in many studies as a quantitative assay of mitochondrial energy metabolism [3,4], but studies of steady state fluorescence cannot distinguish between changes in energy production or utilization.

In this study conditions were created where the fluorescent recovery of a probe would be solely dependent on cellular enzymatic activity (Enzyme Dependent Fluorescence Recovery after Photobleaching (ED-FRAP)). Experiments examining the inherent fluorescence of NADH (351nm excitation, 450 nm emission) were conducted on small droplets (less than 325 μm diameter) containing NADH alone, in droplets containing an enzyme system capable of synthesis of NADH (Figure 1A) and on isolated rabbit cardiac myocytes (Figure 1D). Photobleaching of the entire cell or droplet eliminated diffusion or bulk transport of NADH from non-bleached regions. Droplets containing NADH alone did not recover, while droplets containing enzyme were shown to recover exponentially (Figure 1B) with a rate constant of fluorescent recovery (kf) that was proportional to enzyme concentration (Figure 1C).
Unfolding Single RNA Molecules with Optical Tweezers
Authors: Bustamante Carlos , Liphardt Jan , Onoa Bibiana , Smith Steven B , Collin Delphine , Tinoco Ignacio
Published: July 2020

RNA molecules must fold into specific three-dimensional shapes to perform their structural and catalytic functions. Unlike proteins, RNAs secondary structural features are usually stable enough to form by themselves in solution. The reason is that in RNA, the stabilization energy gained from the formation of secondary structure is substantially larger than the energies involved in tertiary interactions. As a result, the formation of tertiary interactions is expected to alter only slightly the pre-existing secondary structural contacts. Moreover, secondary structure prediction is robust and can be made without taking into consideration tertiary folding. However, bulk studies of the energetics and kinetics of their secondary and tertiary folding are often frustrated by the presence of multiple species and multiple folding pathways in solution. These problems are circumvented in single-molecule studies in which the folding/unfolding trajectories of the individual molecules can be followed. The T. thermophila group I intron ribozyme is organized into several domains whose mechanical unfolding can be investigated independently, and whose tertiary contacts are stabilized by numerous Mg++ ions.

We have begun characterization of the ribozyme by analysis of the P5abc domain because:
Probing Single Molecules in Single Living Cells
Authors: Byassee Tyler A , Chan Warren C W , Nie Shuming
Published: July 2020

Direct observation of single molecules and single molecular events inside living cells could dramatically improve our understanding of basic cellular processes (e.g., signal transduction and gene transcription) as well as improving our knowledge on the intracellular transport and fate of therapeutic agents (e.g., antisense RNA and gene therapy vectors). However, a key remaining question is whether single-molecule methodologies could be developed to study complex molecular processes in living cells. in contrast to clean and well-controlled conditions in-vitro, the intracellular environment contains a broad collection of biological macromolecules and fluorescent materials such as porphyrins and flavins. This complex environment is known to produce intense background fluorescence, commonly known as autofluorescence. Thus, a major concern is that this intracellular background could overwhelm the relatively weak signals arising from single molecules.

We demonstrate that fluorescence detection of single molecules can be achieved by tightly focusing a laser beam into a living cell (see Figure 1). The observed background fluorescence is indeed higher than that in-vitro (e.g., pure biological buffer), but this background is continuous and stable, and does not significantly interfere with the measurement of single-molecule photon bursts. Specifically, we report single-molecule results on three types of extrinsic fluorescent molecules in cultured human HeLa cells (a cervical cancer cell line).
Study of Ultrastructure and Permeability of Brain Cortex Vascular Endothelium in Rats under Rhythmic Cooling
Authors: A.S. Kaprelyants , L.M. Marchenko , R.K. Migunova
Published: July 2020

Cooling significantly affects cellular ultrastructure [1], as well as structural and functional state of histohematic barrier [2]. The aim of the given work was to study ultrastructural mechanisms of the permeability of brain cortex vascular endothelium under the effect of rhythmic hypothermia. Experiments were conducted on 180-220 g white males rats. Cooling of caudal part of animals was conducted in an automatic regimen by an ice water with 0.1 Hz frequency during 40 and 80 min. Experimental animals were narcotized by intraperitoneal injection of thiopenthal sodium and sodium hydroxybutyrate mixture in respect of 30mg/kg, 100 mg/kg of mass, correspondingly. Material from parietal lobe of brain cortex for electron-microscopic investigations was taken immediately after the effect finishing. Rectal temperature, which made 36-35C, was measured during the experiment. For electron microscopic investigations the samples were fixed by immerging into 4% glutar aldehyde on a phosphate buffer and were additionally fixed by 1% osmium fixative on a phosphate buffer. After dehydration in ethanol, samples were placed into the mixture of epoxy resins Epon-Araldit (Fluka, Sweden). Colloid carbon was used as a tracer for the estimation of permeability of brain capillary endothelium. with this aim 3-5 ml of 10% colloid carbon solution, prepared on 0,85% NaCl solution, were injected into a tail vein of the animals immediately after their decapitation. Preliminary estimation of the material quality was done on semi-fine tissue sections after the staining by toluidine blue.
Increased DNA Damage and Decreased H-MYH Protein Expression in Human Myocardium with End-Stage Congestive Heart Failure
Authors: Lin R , Dong Y , Conte JV , Lu-Chang A-L , Wei C
Published: July 2020

The reactive oxygen species (ROS) such as superoxide radical (O2), hydrogen peroxide (H2O2), and hydroxyl radical (OH), plays a critical role in the pathogenesis of hypertension, atherosclerosis, ischemia-reperfusion injury, stroke, myocardial infarction, and congestive heart failure. A recent study demonstrated that the frequency of mutation in a reporter gene increased in cortical DNA after forebrain ischemia-reperfusion. Eight DNA lesions that are characteristic of DNA damage mediated by free radicals were detected. Among the different oxidative-damage DNA products, 8-oXo-7,8-dihydrodeoxyguanine (8-oxoG) is the most stable and deleterious adduct. Recent studies demonstrated that human MutY homologue (hMYH) protein plays important role in repairing oxidative damage. to date, there is no information regarding the role of hMYH expression in human myocardium with congestive heart failure (CHF). Therefore, the current study designed to investigate the levels of hMYH expression and 8-oxoG in myocardium in the absence or presence of end-stage congestive heart failure. The hypothesis of this study is that increasing DNA damage such as 8-oxoG generation is associated with a reduction of hMYH expression in human myocardium with congestive heart failure.
Observation of the Internal Structure of a Biological Sample using a Fluoresence Type 3-Dimensional Internal Structure Microscope
Authors: YOKOTA Hideo , NAKAMURA Sakiko , MAKTNOUCHI Akitake , HIGUCHI Toshiro
Published: July 2020

We developed a new type 3-Dimensional Internal Structure Microscope (3D-ISM) for the observation of internal structures of samples. The internal structure of a sample is obtained by cutting it into thin slices and observing the cutting side continuously while cutting. The position of the camera, as well as the sample position are fixed. Therefore there is no shift between the sections, and this system can obtain a true color image of the sample, which is a high resolution and a high-quality three-dimensional image compared with X-ray CT and MRI. After repeatedly slicing a sample, the digital image data from the sectional views is transferred to a computer, where 3-dimensional images of the internal structure of the sample are reconstructed. Using this system we analyzed many biological organisms. Just now, we are developing the system which builds in a fluorescent microscope. in this paper, We therefore observed fluorescent-stained samples by a fluorescence microscope installed in the 3D-ISM instrument are shown.

This article presents the outline of the device and the principle of the observation. This system, consisting of a slicing device, reflecting optical microscope, a CCD camera, a laser video recorder, a PC-based control system and an image processing workstation. FIG. 1 shows a schematic view of the cross secsional machine. Before slicing, the sample is embedded into either paraffin or OCT compound. Subsequently, the sample is positioned into a cross-sectioning machine for the slicing operation, which is performed using a cutting blade. The maximum size of the sample which can be observed with this device is ϕ 8x10mm. The thickness of the slices can be adjusted mechanically between 0.5-10 μ m. The cutting speed can range from 0.5 to 4 slices/ s. Each cross-section is viewed by employing a epi-fluoresence microscope and a CCD camera. The object lens used for the observation can use the x2-80. The resulting image is recorded on a laser video disc. After the slicing operation, the 3D sample image is reconstructed on the basis of volume rendering by using a Workstation. These observations can be completely done by full automation.