bluuebunny/crossref_metadata_2025_split huggingface dataset:
The effect of temperature on high-resolution TEM image contrast
Authors: Geipel T. , Mader W. , Pirouz P. 
Published: June 2020

Temperature affects both elastic and inelastic scattering of electrons in a crystal. The Debye-Waller factor, B, describes the influence of temperature on the elastic scattering of electrons, whereas the imaginary part of the (complex) atomic form factor, fc = fr + ifi, describes the influence of temperature on the inelastic scattering of electrons (i.e. absorption). In HRTEM simulations, two possible ways to include absorption are: (i) an approximate method in which absorption is described by a phenomenological constant, μ, i.e. fi; - μfr, with the real part of the atomic form factor, fr, obtained from Hartree-Fock calculations, (ii) a more accurate method in which the absorptive components, fi of the atomic form factor are explicitly calculated. In this contribution, the inclusion of both the Debye-Waller factor and absorption on HRTEM images of a (Oll)-oriented GaAs crystal are presented (using the EMS software.Fig. 1 shows the the amplitudes and phases of the dominant 111 beams as a function of the specimen thickness, t, for the cases when μ = 0 (i.e. no absorption, solid line) and μ = 0.1 (with absorption, dashed line).
Acquisition and analysis of fluorescence ratio images using a multispectral microanalytical imaging program
Authors: Freudenrich Craig C. , Boyle Robert T. , Kopf David A. , Ingram Peter , LeFurgey Ann 
Published: June 2020

Any study regarding the influence of physiological, pathological, or pharmacological/toxicological factors on cellular ion homeostasis requires knowledge of multiple parameters including cytoplasmic ion concentrations, ion fluxes, and subcellular elemental distribution so that an accurate assessment of the state of ion homeostasis can be obtained. These multiple parameters are often obtained by using different types of microscopy and microanalysis. For example, cytoplasmic ion concentrations are measured by fluorescence digital imaging microscopy using various ion indicators, while cellular and subcellular element contents are determined by electron probe X-ray microanalysis (EPXMA) digital imaging. For correlating information from various microscopies, it would be advantageous to have one software package and image format that operates on the same computer platform. Here, we describe the modification of an existing software (ImagNSpect) developed for EPXMA digital imaging for use in analyzing fluorescence digital images.Fluorescence images (490 and 440 nm excitation, 540 nm emission) of BCECF-loaded cultured rat hepatocytes (Fig. 1a, b) were obtained on an inverted fluorescence microscope (Carl Zeiss, Inc., Thorn wood, NY) equipped with a rotating excitation filter wheel, a CCD camera (Photometries, Ltd., Tucson, AZ), a controller box, and a Macintosh Quadra 800 computer (Apple Computer Corp., Cupertino, CA).
Light and Electron Microscopic localization of inducible nitric oxide synthase in the heart of rats with myocarditis
Authors: Nishikawa T. , Ishiyama S. , Takeda K. , Shimojo T. , Hiroe M. , Kasajima T. 
Published: June 2020

Nitric oxide synthase (NOS) is an enzyme involved in the synthesis of nitric oxide (NO). The inducible form of NOS (iNOS) is induced in the presence of cytokines and produces massive amounts of NO which may be harmful to living tissue. In acute myocarditis, a serious disease which can be fatal, extensive myocardial cell damage associated with massive inflammatory cell infiltration is usually observed and large amounts of cytokines are released from inflammatory cells. However, the mechanism of the heart tissue injury is still unclear. In this study, we investigated the role of iNOS in the myocardial tissue injury in acute myocarditis.The heart tissue specimens were taken from Lewis rats with experimentally induced autoimmune myocarditis. An immunohistochemical study was performed using iNOS antibody raised in New Zealand white rabbits immunized with synthetic peptides (17 amino acids) corresponding to the C-terminal of macrophage iNOS. The specificity of the antibody was confirmed by Western blot analysis. Immunoelectron microscopic studies were carried out by the gold-labeled IgG method using ultrathin sections of LR white resin-embedded material.
The molecular structure of thiol-monolayers on gold imaged with an Atomic Force Microscope
Authors: Jaschke M. , Wolf H. , Schönherr H , Ringsdorf H. , Bamberg E. , Butt H.-J. 
Published: June 2020

Thiols spontaneously form monolayers on gold. An important question is which factors determine the structure of the thiols. The two extreme positions are: (a) the structure of the underlying gold determines the packing of the thiol molecules; (b) the geometry of the thiol molecules and the interaction between thiols determine their two-dimensional structure.To help answer this question two series of thiol molecules on gold (111) were imaged by atomic force microscopy. In order to reduce the force between tip and sample, and to prevent distortion of the thiol monolayer, all images were taken in ethanol at forces between 0.5 nN and 3 nN.The first series consisted of several azobenzene-thiols:Although these molecules possess a terminal alkyl chain, their lattice is different from the well known hexagonal (√3 × √3) R 30° lattice (with respect to the underlying gold (111) lattice) of alkane-thiols. Instead, we found two stable rectangular lattices which could both be described by lattice constants of 0.61 nm and 0.79 nm, and an angle of 89°. These structures were also found on polycrystalline gold. Also, varying the length of the alkyl chain (molecules (i) and (ii)) or using a disulfide instead of a thiol (molecules (ii) and (iii)) did not change the lattice constants. Thus we conclude that the lattice for azobenzenethiols is end-group dominated.
Measuring forces between biological macromolecules with the Atomic Force Microscope: characterization and applications
Authors: Lee Gil U , Chrisey Linda , Colton Richard J. 
Published: June 2020

Structure and function in biological macromolecular systems such as proteins and polynucleotides are based on intermolecular interactions that are short ranged and chemically specific. Our knowledge of these molecular interactions results from indirect physical and thermodynamic measurements such as x-ray crystallography, light scattering and nuclear magnetic resonance spectroscopy. Direct measurement of molecular interaction forces requires that the state of a system be monitored with near atomic resolution while an independent force is applied to the system of 10−12 to 10−9 Newton magnitude. The atomic force microscope (AFM) has recently been applied to the study of single molecular interactions. The microfabricated cantilever of the AFM, a force transducer of small yet variable stiffness and high resonance frequency, produces a transducer of 10−15 N/Hz1/2 force sensitivities and 0.01 nm position accuracy.This presentation describes the AFM measurement of the molecular interaction forces in the model ligand-receptor system streptavidin-biotin and between complementary strands of DNA.
Tomographic reconstruction of mitochondria
Authors: Mannella C.A. , Marko M. , Buttle K. , Penczek P. , Leith A. , Frank J. 
Published: June 2020

During oxidative phosphorylation, free energy of substrate oxidation is used to generate electrochemical gradients across the mitochondrial inner membrane. Full understanding of this process requires knowing the pathways for internal diffusion of ions and metabolites inside the organelle. This, in turn, requires detailed information about the organization of the membranes that compartmentalize the mitochondrion and the distribution of transport proteins (proton pumps, ion channels, metabolite carriers) on these membranes.We have undertaken the study of the compartmentation of rat-liver mitochondria (conventionally fixed and plastic-embedded) using electron microscopic tomography. Reconstructions of isolated mitochondria have been computed using projection images collected on the Albany high-voltage electron microscope from sections (0.5-1-μm thick) tilted around one axis over +/− 70° at 2° increments. A problem encountered with this approach is the directional loss of resolution due to the “missing wedge” of information in Fourier space in the direction parallel to the tilt axis. To overcome this problem, we have developed an alignment algorithm that allows projections collected over two or more tilting directions to be used in the same modified back-projection calculation.
Practical electron tomography
Authors: Woodcock C.L. 
Published: June 2020

Despite the potential of the technique, electron tomography has yet to be widely used by biologists. This is in part related to the rather daunting list of equipment and expertise that are required. Thanks to continuing advances in theory and instrumentation, tomography is now more feasible for the non-specialist. One barrier that has essentially disappeared is the expense of computational resources. In view of this progress, it is time to give more attention to practical issues that need to be considered when embarking on a tomographic project. The following recommendations and comments are derived from experience gained during two long-term collaborative projects.Tomographic reconstruction results in a three dimensional description of an individual EM specimen, most commonly a section, and is therefore applicable to problems in which ultrastructural details within the thickness of the specimen are obscured in single micrographs. Information that can be recovered using tomography includes the 3D shape of particles, and the arrangement and dispostion of overlapping fibrous and membranous structures.
Detection and mapping of interactions between chromatin and the nuclear envelope in drosophila embryos
Authors: Marshall W.F. , Dernburg A.F. , Harmon B. , Sedat J.W. 
Published: June 2020

Interactions between chromatin and nuclear envelope (NE) have been implicated in chromatin condensation, gene regulation, nuclear reassembly, and organization of chromosomes within the nucleus. To further investigate the physiological role played by such interactions, it will be necessary to determine which loci specifically interact with the nuclear envelope. This will not only facilitate identification of the molecular determinants of this interaction, but will also allow manipulation of the pattern of chromatin-NE interactions to probe possible functions. We have developed a microscopic approach to detect and map chromatin-NE interactions inside intact cells.Fluorescence in situ hybridization (FISH) is used to localize specific chromosomal regions within the nucleus of Drosophila embryos and anti-lamin immunofluorescence is used to detect the nuclear envelope. Widefield deconvolution microscopy is then used to obtain a three-dimensional image of the sample (Fig. 1). The nuclear surface is represented by a surface-harmonic expansion (Fig 2). A statistical test for association of the FISH spot with the surface is then performed.
Near-field microscopy of membrane domains
Authors: Hwang J. , Betzig E. , Edidin M. 
Published: June 2020

Results from several different methods for probing the lateral organization of cell surface membranes indicate that these membranes are patchy, divided into domains. The data suggest that on average these domains are 0.1-1 μm across and that they persist for 10’s to 1000’s of seconds. At least some domains in this size range, when labeled by fluorescent proteins or lipids ought to be detectable by conventional, far-field, fluorescence microscopy. However, though some images are consistent with a domain structure for membranes, most far-field images of fluorescent cell surfaces lack the detail necessary to define domains.We have used near-field scanning optical microscopy, NSOM, of fluorescent-labeled cells to visualize membrane patchiness on the nanometer scale. This method yields images with resolutions of 50 nm or less. In our near-field microscope the labeled sample is illuminated by a optical fiber probe, with an aperture of 50-80nm. The probe is scanned over the cell surface at a distance of ˜ 10 nm from the surface. Only surface fluorescence is excited by the scanned probe.
Confocal microscopy of single living cells
Authors: Lemasters John J. 
Published: June 2020

The advent of laser scanning confocal microscopy solves the dilemma of studying thick specimens with optical microscopy by creating optical slices less than 1 μm in thickness. Increasingly, confocal microscopy is an essential analytical tool for studying the structure and physiology of living cells. Because confocal microscopy collects light from only a fraction of the specimen volume, greater illumination is required. Consequently, photodamage and photobleaching are greater considerations, especially for study for living cells where repeated measurements over time are desired. To minimize photodamage, laser intensity should be attenuated by 100-1000 fold, photomultiplier circuits should be operated at highest sensitivity, and stable fluorophores should be used. When these conditions are met, literally hundreds of high resolution confocal images can be obtained from single cells loaded with parameter sensitive fluorophores.The number of parameter-specific fluorophores useful for observing single living cells by confocal microscopy is large and increasing. By labeling with calcein and collecting serial images, the volume, shape and surface topography of single living cells are reconstructed with results rivaling scanning electron micrographs.
Surgical repair of long bone defects or trauma by guided tissue engineering with composite implants of porous bioactive glass-ceramic (A-WGC) particles bound by a plaster/CaO/P2O5cement contained by a resorbable vicryl™ mesh
Authors: Hanker J.S. , Hanker L.C. , Giammara B.L. 
Published: June 2020

One of the biggest problems associated with the repair of trauma in long bones is their requirement to withstand greater mechanical stress than any other bones of the body. After the success we achieved with composite hydroxylapatite (HA)/plaster(PP) in craniofacial bone repair in approximately 200 animals (cats and rats) and 300 humans in our laboratory, clinics and surgeries, it was felt that an area which could perhaps benefit from our experience in bone repair might be orthopedic surgery. When first informed of our desire to apply our biomaterials interest and experience to long bone trauma patients, others expected little success with these biomaterials that were so successful in craniofacial surgery.We felt, however, that success might be achieved for long bone repair with plaster of Paris (CaSO4·½H2O) and/or calcium phosphate. The feeling of my laboratory received some support from Larry Hench's 1988 article. In this article he pointed out that bioactive ceramics such as hydroxylapatite (HA) possibly could bond to bone.
Immunocytochemical double staining with the ELF method
Authors: Baskin D. G. , Vathanaprida U C. , Stahl W. L. 
Published: June 2020

The reagent 2-(5'-chloro-2'-phosphoryloxyphenyl)-6-chloro-4-[3H]-quinazolinone has recently become commercially available as the ELF (enzyme-labeled-fluorescence) phosphatase substrate. The ELF method is basically an alkaline phosphatase enzyme immunostaining procedure in which the reaction product is fluorescent instead of chromogenic. The ELF substrate produces a photostable, bright yellow-green fluorescence (500-580 nm) with low background. The use of the ELF substrate as a green label may have advantages over FITC in double-labeling immunocytochemical staining procedures that employ Cy3, which frequently shows residual orange-red fluorescence when viewed with conventional FITC filter sets.To evaluate the suitability of ELF for dual immunofluorescence with Cy3-labeled second antisera, we used paraffin sections of rat pancreas fixed in 4% paraformaldehyde. Primary antibodies were an anti-insulin monoclonal antibody (1:50) (Biogenex) and anti-glucagon polyclonal antibody (1:1K)2, in Tris-HCL, 1% BSA, 0.1% TX-100, pH 7.4. The insulin antibody was applied overnight at 6 °C, followed by goat anti-mouse IgGCy3 (Jackson Immunochemical Research) for 60 min. at room temperature (RT).
Immunoelectron microscopy of the connective tissue matrix following cryotechnical processing
Authors: Keene D.R. , McDonald K. , Ridgway C.C. 
Published: June 2020

Our understanding of the microarchitecture of connective tissues is based primarily on micrographs taken of tissues stabilized by immersion fixation. Considerable extraction of tissue components, particularly small matrix molecules, occurs even during fixation in glutaraldehyde. An alternative to immersion fixation is the technique of cryostabilization whereby tissue components are instantaneously immobilized and surrounded by non-crystaline (amorphous) or microcrystaline ice. The ice is replaced at low temperature (-100 to -85°C) by acetone containing fixative, and the tissue finally embedded in epoxy. A comparison of micrographs taken from tissue embedded following immersion fixation and those taken following cryotechnical processing reveals striking differences in the structure of cells, basement membranes, and matrix (Fig. 1a and 2a).Our recent objective has been to utilize sections from samples which have been stabilized by high pressure freezing for immunocytochemical labeling experiments with the expectation that such studies would result in a more precise localization of matrix components. The results to date suggest that many of the difficulties normally experienced in section surface labeling technique are still a problem: Antigens may be affected by the chemistry of freeze-substitution and choice of embedding media.
Effects of the pseudomonad phytotoxin coronatine on tomato leaf structure and ultrastructure
Authors: Palmer D.A. , Bender C.L. 
Published: June 2020

Coronatine is a non-host-specific phytotoxin produced by several members of the Pseudomonas syringae group of pathovars. The toxin acts as a virulence factor in P. syringae pv. tomato, allowing the organism to multiply to a higher population density and develop larger lesions than mutant strains unable to produce the toxin. The most prominent symptom observed in leaf tissue treated with coronatine is an intense spreading chlorosis; this has been attributed to a loss of chlorophylls a and b in tobacco. Coronatine's effects on membrane integrity and cell ultrastructure have not been previously investigated. The present study describes changes in tomato leaves in response to treatment with purified coronatine, infection by a coronatine-producing strain of P. syringae pv. tomato, and infection by a cor" mutant.In contrast to H2O-treated tissue, coronatine-treated tissue showed a diffuse chlorosis extending approximately 5 mm from the inoculation site. Leaf thickness, cell number, and cell dimensions were similar for both healthy and coronatine-treated, chlorotic tissue; however, the epidermal cell walls were consistently thicker in coronatine-treated leaves (Figs, la and lb).
Metallothionein expression during wool follicle development
Authors: Nishimura N. , Nishimura H. , Tohyama C. , Cam G.R. , Adelson D.L. 
Published: June 2020

This study investigated metallothionein(MT) expression in proliferating or differentiating cells of developing wool follicles. Dividing cells in foetal sheep skin were identified by incorporated BrdU, which was injected into the jugular vein of a ewe before sacrifice. MT gene expression was assayed using a digoxigenin-labelled sheep MT-1b riboprobe as described previously. Immunohistochemical staining for MT protein and BrdU was carried out using rabbit-anti rat MT-1 serum and anti-BrdU, as described elsewhere. Foetal sheep skin was obtained from 60 to 130 days of gestation, fixed in HistoChoice or Bouin's and embedded in paraffin.At 71 days of gestation, MT-1b mRNA was detected in the epidermis(EP), primarily in the spinous cells (Fig. la). At 90 days, MT-1b mRNA was concentrated at the tip of the follicle rudiment(F), but not in the presumptive dermal papilla(DP) (Fig. 1b). At 112 days, a symmetric distribution of MT-1b mRNA was observed in the follicle bulb matrix (Fig. 1c). However, very little MT-1b mRNA was detected in the matrix cell layers adjacent to DP. At 130 days, strong MT-lb gene expression was localized to the cytoplasm of cells in the follicle bulb(B) and keratogenous zone(K)(Fig. Id).
Substructurel features of LLC-PK1porcine kidney cells grown on Anocell filter supports
Authors: Mia A.J. , Johnson S. , Franklin J. , Johnson J.F. , Puttaparthi K. , Oakford L.X. , Yorio T. 
Published: June 2020

LLC-PK1cells, derived from porcine kidney cortex, have been used for a variety of studies including sodium-coupled hexose transport, vasopressin/calcitonin sensitive adenylate cyclase(1) and vasopressin V1and V2receptor-mediated endocytosis involving clathrin coated pits and vesicles(2,3,4,5). Recently, we reported on the morphological profile of these cells using the techniques of scanning electron microscopy (SEM). SEM studies of confluent LLC-PK1cells exhibited surface morphology similar to intact tissue. These cells also responded to ADH (antidiuretic hormone, vasopressin) or MZ (mezerein), a non-phorbol activator of protein kinase C (PKC) with a propagation of numerous microvilli over the apical plasma membrane typical of ADH responsive tissue. The current study evaluates the cytomorphological features of cultured LLC-PK1cells following hormone addition.LLC-PK1cells were grown on Anocell filter supports using DMEM media in a 5% CO2/air environment in an incubator at 37°C. Upon confluency, cultured cells were washed in buffer and then treated with 100 mU/ml ADH or MZ10-6M for 15 min. Control and stimulated tissues were fixed in 2% glutaraldehyde in PIPES with a postfixation in 1% osmium tetroxide for 1 hr for either SEM or TEM.
FT-IR microspectroscopic analysis of diseased white matter brain tissues
Authors: Le Vine Steven M. , Wetzel David L. 
Published: June 2020

In situ FT-IR microspectroscopy has allowed spatially resolved interrogation of different parts of brain tissue. In previous work the spectrrscopic features of normal barin tissue were characterized. The white matter, gray matter and basal ganglia were mapped from appropriate peak area measurements from spectra obtained in a grid pattern. Bands prevalent in white matter were mostly associated with the lipid. These included 2927 and 1469 cm-1 due to CH2 as well as carbonyl at 1740 cm-1. Also 1235 and 1085 cm-1 due to phospholipid and galactocerebroside, respectively (Figs 1and2). Localized chemical changes in the white matter as a result of white matter diseases have been studied. This involved the documentation of localized chemical evidence of demyelination in shiverer mice in which the spectra of white matter lacked the marked contrast between it and gray matter exhibited in the white matter of normal mice (Fig. 3).The twitcher mouse, a model of Krabbe’s desease, was also studied. The purpose in this case was to look for a localized build-up of psychosine in the white matter caused by deficiencies in the enzyme responsible for its breakdown under normal conditions.