bluuebunny/crossref_metadata_2025_split huggingface dataset:
Time-resolved cryo-electron microscopy using spray mixing
Authors: Berriman John , Unwin Nigel 
Published: June 2020

It is often necessary to investigate the three-dimensional structures of biological molecules, or complexes, in short-lived, transitional states in order to obtain a clear understanding of how they work. A simple method is described to this end for cryo-electron microscopy. The two reactants (e.g. protein complex and ligand) are mixed by the coalescence of spray droplets containing one component with a thin, grid-supported aqueous film containing the other. The transitional structure is then trapped by adjusting the delay between the droplet impact and freezing. As an alternative to the photolytic release of caged compounds, this method does not require special chemicals and avoids any heating or photolysis of the sample. It should be possible to catch reaction times as short as 1 millisecond.The freezing device was a standard guillotine-type plunger in which the grid, mounted on forceps, is allowed to drop by free-fall from a height of several centimeters above the ethane surface.
Silver enhancement of nanogold and undecagold
Authors: Hainfeld James F. , Furuya Frederic R. 
Published: June 2020

Silver enhancement of colloidal gold is commonly used to improve visibility of small gold particles in the electron microscope, or to visualize immunogold labeling in the light or confocal microscope. Silver enhancement can also produce a signal visible by eye in such applications as immunodot blotsor staining of gels or transfers with gold reagents. A recent advance in gold technology has been theuse of gold clusters, which are gold compounds, and can be covalently linked to antibodies or other molecules. Two gold clusters developed for such uses are undecagold (Au11 ), which has a 0.8 nm gold core, and a larger, 1.4 nm gold core cluster, •Nanogold•. Because of their small size, many uses requirefurther enlargement via silver deposition.Here we report quantitative studies of the silver reaction rates for Nanogold and undecagold. These were followed by measuring the UV-visible (UV-vis) spectra at 1 min intervals of solutions of gold clusters mixed with a light insensitive silver enhancer (LI Silver). Solutions were in a 1 cm path length disposable cuvette and spectra were recorded using a Hewlett Packard 8452A diode array spectrophotometer.
Characterization of cryosectioned cell suspensions by STEM
Authors: Shi S. L. , Sun S. Q. , Nash T. E. , Leapman R. D. 
Published: June 2020

To exploit the improved resolution and sensitivity that is now available for biological microanalysis it is crucial to optimize the specimen preparation techniques. This requires rapid freezing and ultrathin cryosectioning to a thickness of around 100 nm. Recently, it has been shown that low-dose scanning transmission electron microscopy (STEM) when applied to such specimens can yield structural detail which is comparable to that from conventional thin plastic sections. After the water is removed from the cryosections by freeze drying in vacuum, the dark-field STEM signal provides a high contrast image of the dry mass distributions of the specimen. In this paper we demonstrate the quality of ultrastructure that is achievable for suspensions of cultured cells which are ideal systems to study since the culture medium can be readily controlled. The method is illustrated by application to the protozoan parasite, Giardia lamblia.G. lamblia trophozoites, cultured in TYI-S-33 growth medium, were centrifuged at low speed to obtain a very high density suspension of cells.
Synaptic connectivity in hamster circumvallate taste buds
Authors: Zubrzycki K. J. 
Published: June 2020

The hamster is an interesting animal model for studying gustation because of its remarkable responsiveness to sweet stimuli. Previous investigators of hamster circumvallate taste have categorized cells into type I, II, and basal cells. The goals of the present study were to determine the ultrastructure of the various cell types and to analyze synaptic contacts in circumvallate taste buds of the hamster.Adult female Syrian hamsters were prepared for electron microscopy and thick Epon sections (2-4μm) were collected on microscope slides. Selected sections were then reembedded for thin sectioning. Thin sections were sliced and mounted onto Formvar-coated slot grids and poststained in uranyl acetate and lead citrate. The sections were examined with a Hitachi H-7000 transmission electron microscope at 100 kV.The circumvallate (=vallate) papilla of the hamster is located in the midline of the posterior portion of the tongue. Type I (dark cells), type II cells (light cells) and basal cells were identified within these taste buds.
Organization of primary afferent and local-circuit synapses in the olfactory glomerulus
Authors: Greer Charles A. , Bartolomei Juan C. , Dembner Jeffrey M. 
Published: June 2020

Odorant molecules are transduced by olfactory receptor cells whose axons join to form the olfactory nerve which distributes across the surface of the olfactory bulb (OB). Axons exit the nerve layer to terminate within the glomerular neuropil of the OB. While there appears a gross topography between the epithelium and OB4, it is clear that extensive topographic reorganization of axons occurs within the olfactory nerve. To better understand the mechanisms that may contribute to the establishment of glomerular-specific fascicles and functional domains within the OB, we have investigated axonal organization within the nerve and the intraglomerular distribution of primary afferent synapses using light, confocal and electron microscopy.Sprague-Dawley rats, 30 to 50 days postnatal, were anesthetized, lightly perfused with 0.9% NaCl and the OBs removed. Crystals of the lipophilic dye, Dil, were inserted into the olfactory nerve layer and the tissue placed in 4% paraformaldehyde at room temperature for 10 - 30 days.
An ultrastructural and biochemical study of the effects of a 5-lipoxygenase inhibitor on peroxisome proliferation in rat liver and kidney
Authors: Tahir S. K. , Gagne G. D. , Miller M.F. 
Published: June 2020

The present study was undertaken to assess the effects of a drug candidate on the morphological and biochemical changes associated with peroxisome proliferation in the liver and kidneys of CD®BR rats. Zileuton, (Abbott-64077), a benzothiophene N-hydroxyurea, is a 5-lipoxygenase inhibitor which has potential therapeutic uses for conditions such as rheumatoid arthritis. To date, several hypolipidemic drugs, certain plasticizers, and a number of unrelated compounds are known to induce peroxisome proliferation in animals. Peroxisomes (also referred to as microbodies) are single membrane-bound organelles found in most tissues, however, their occurrence in hepatocytes is greater than in any other cell type. In the kidneys, peroxisomes are predominantly located in the cortex.The prolonged proliferation of hepatocellular peroxisomes is of concern since chronic proliferation of peroxisomes has been associated with an increase in hepatic tumors in rodents. In general, peroxisome proliferation can be assessed by an increase in peroxisome number and an increase in fatty acid β-oxidation activity.
Automated counting and 3-D morphometry of nuclei in thick tissue slices from laser- scanning confocal microscopy data
Authors: Ancin H. , Roysam B. , Chestnut M.H. , Otte T.E. , Szarowski D.H. , Turner J.N. 
Published: June 2020

This paper presents recent advances in automated three-dimensional (3-D) image analysis methods for cell counting, and quantitative measurement of various nuclear properties in thick (30-120 μm) tissue sections that are imaged by a laser-scanning confocal microscope. The technical advances reported here are: (i) improved 3-D nuclear separation methods for analyzing samples containing large numbers of nuclei per unit volume, and large connected clusters; (ii) methods for adapting the image analysis system to handle a much larger variety of specimens with greater variability in image parameters, such as intensities, nuclear shapes and sizes; and (iii) methods for assisting a user in selecting parameter inputs to the counting system.Improved 3-D nuclear separation was achieved by computing image gradients from each optical slice. Perona and Malik’s algorithm was used to enhance the image gradient at true nuclear boundaries while suppressing the undesirable intra-nuclear image gradients. The result was used to compute a new proximity index for the partitional cluster analysis method.
Fast automated mosaic synthesis method for 2-D/3-D image analysis of specimens much wider than the field of view
Authors: Becker D.E. , Ancin H. , Roysam B. , Turner J.N. 
Published: June 2020

We present an efficient, robust, and widely-applicable technique for computational synthesis of wide-area images from a series of overlapping partial views. The synthesized image is the set union of the areas covered by the partial views, and is called the “mosaic”. One application is the laser-scanning confocal microscopy of specimens that are much wider than the field of view of the microscope. Another is imaging of the retinal periphery using a standard fundus imager. This technique can also be used to combine the results of various forms of image analysis, such as cell counting and neuron tracing, to generate large representations that are equivalent to processing the total mosaic, rather than the individual partial views.The synthesis begins by computing a concise set of landmark points for each partial view. The type of landmarks used can vary greatly depending on the application. For instance, in the retinal imaging application, the vascular branching and crossover points are a natural choice. Likewise, the locations of cells in Figs. 1 and 2 provide a natural set of landmarks for joining these images.
Association of pituitary adenoma with neuronal christoma: A TEM study
Authors: Horvath Eva , Kovacs Kalman , Scheithauer B. W. , Lloyd R. V. , Smyth H. S. 
Published: June 2020

The association of a pituitary adenoma with nervous tissue consisting of neuron-like cells and neuropil is a rare abnormality. In the majority of cases, the pituitary tumor is a chromophobic adenoma, accompanied by acromegaly. Histology reveals widely variable proportions of endocrine and nervous tissue in alternating or intermingled patterns. The lesion is perceived as a composite one consisting of two histogenetically distinct parts. It has been suggested that the neuronal component, morphologically similar to secretory neurons of the hypothalamus, may initiate adenoma formation by releasing stimulatory substances. Immunoreactivity for growth hormone releasing hormone (GRH) in the neuronal component of some cases supported this view, whereas other findings such as consistent lack of growth hormone (GH) cell hyperplasia in the lesions called for alternative explanation.Fifteen tumors consisting of a pituitary adenoma and a neuronal component have been collected over a 20 yr. period. Acromegaly was present in 11 patients, was equivocal in one, and absent in 3.
Morphology of Muerto Canyon virus, causative agent of hantavirus pulmonary syndrome
Authors: Goldsmith C. S. , Humphrey C. D. , Elliott L. H. , Zaki S. R. 
Published: June 2020

Hantavirus pulmonary syndrome (HPS) is a newly described illness which was first recognized in May, 1993, in the southwestern US. The disease is characterized by fever, myalgias, headache, and cough,with rapid development of fatal respiratory distress. The causative agent, Muerto Canyon virus (MCV), was recently isolated at CDC from a deer mouse, Peromyscus maniculatus, the most common vector for this virus. In order to fully understand this newly discovered virus, we have characterized the isolate by thin section and negative stain TEM, by SEM, and by pre- and post-embedded immunolabeling of thin sections.MCV is a hantavirus, a member of the family Bunyaviridae. In thin sections of infected Vero E6 culture cells, the extracellular particles are spherical to oval, with fuzzy surface projections along the envelope (figs. 1,2). The interior of the virus contains strands of nucleocapsid. Particles are similar in morphology to Hantann virus, the causative agent of Korean hemorrhagic fever. The negative staining of purified particles (fig. 3) also revealed typical hantavirus morphology of a spherical particle with grid-like surface. Examination by SEM showed scattered particles along the cell surface and among the filopodia.
Identification of the pulmonary syndrome hantavirus by direct and colloidal gold immune Electron Microscopy
Authors: Humphrey C. D. , Goldsmith C.S. , Elliott L. , Zaki S.R. 
Published: June 2020

An outbreak of unexplained acute pulmonary syndrome with high fatality was recognized in the spring of 1993 in the southwestern United States. The cause of the illness was quickly identified serologically and genetically as a hantavirus and the disease was named hantavirus pulmonary syndrome (HPS). Recently, the virus was isolated from deer mice which had been trapped near the homes of HPS patients, and cultivated in Vero E6 cells. We identified the cultivated virus by negative-stain direct and colloidal gold immune electron microscopy (EM).Virus was extracted, clarified, and concentrated from unfixed and 0.25% glutaraldehyde fixed supernatant fluids of infected Vero E6 cells by a procedure described previously. Concentrated virus suspensions tested by direct EM were applied to glow-discharge treated formvar-carbon filmed grids, blotted, and stained with 0.5% uranyl acetate (UA) or with 2% phosphotungstic acid (PTA) pH 6.5. Virus suspensions for immune colloidal gold identification were adsorbed similarly to filmed grids but incubated for 1 hr on drops of 1:50 diluted monoclonal antibody to Prospect Hill virus nucleoprotein or with 1:50 diluted sera from HPS virus infected deer mice.
Acetonitrile is better than ethanol as a dehydrating agent for cells prepared for SEM
Authors: Holshek J. G. , Akins R. E. 
Published: June 2020

Acetonitrile has been shown to be a better dehydrating agent than ethanol for whole tissue prepared for TEM studies. In this paper we show that it is also a better choice for dehydration of cell cultures prepared for SEM studies. The procedures used for the isolation and culture of cardiac cells in the present study have been outlined in detail by Schroedl and Hartzell (1983) and Freerksen, et al. (1984). Cells were cultured in suspension using HARV-Bioreactors (Synthecon, Inc., Friendswood, TX) for 24 hours to allow attachment and then fed each subsequent 48 hours. After a total of six days in suspension culture, cell/bead clusters were collected and washed with PBS.Primary fixation was done at 4°C, all other steps were performed at room temperature. The buffer used in all cases was 0.1M cacodylate, pH 7.3. The cell/bead cultures were processed according to the following protocol. All of the beads were placed in a 1.5 ml microfuge tube and fixed in 2% buffered glutaraldehyde overnight followed by two 5 min washes in buffer, postfixation in 1% buffered OsO4 for 90 min and three washes in buffer. One half of the beads were then placed in another microfuge tube and dehydrated in acetonitrile, while the remaining beads were dehydrated in ethanol.
Morphological comparison of Pavo cristatus and Pavo cristatus ALB feather barbules
Authors: McKinney M. , Gardner J. S. 
Published: June 2020

It was hypothesized that a morphological difference existed between the feather barbules of the colorful, iridescent peafowl and those of the albino peafowl.Previously molted plume feathers from the Indian (Pavo cristatus) and Albino (Pavo cristatus alb) peafowl were collected from the Tracy Aviary in Salt Lake City, Utah. Segments were selected from the eye region of feathers of both species and they were examined directly with the light microscope (LM). Similar segments were prepared for study with the scanning electron microscope (SEM). Portions of barbs possessing barbules were embedded in Spurr’s resin and sectioned for SEM and transmission electron microscope (TEM) evaluation.With the light microscope it was observed that only the barbules exhibited brilliant colors. The barbs and rachis did not. By changing the position of the light source, the shade and intensity were altered, but not the color.SEM images were collected at low magnification (Figs. 1 and 2). Segmentation of the barbules was observed in both Pavo cristatus and Pavo cristatus alb.
Image-coupling methods in CCD cameras for Electron Microscopy
Authors: Mooney P. E. , Krivanek O. L. 
Published: June 2020

It is well established that the charge-coupled device (CCD) is the detector of choice in imaging applications requiring sensitivity, dynamic range, linearity and low geometric distortion. It has also been shown that in the electron microscope, indirect coupling of the image by a scintillator and transfer optic is required to prevent damage to the CCD and to allow for sufficient dynamic range. The question then follows how best to design the coupling to achieve the image quality required for digital imaging in electron microscopy.We have characterized slow-scan CCD cameras with three representative optical couplings (Figure 1):1:1 fiber-optically coupled camera with a large-pixel CCD (TK1024) and both single-crystal and powder scintillators for 100-400 kV applications requiring good sensitivity,1:1 tandem lens-coupled camera with a large-pixel CCD (TK1024) and a powder scintillator mounted on an ultra-thin Al foil for high voltage applications, and3:1 reduction macro lens-coupled camera with a fast, small-pixel CCD (Kodak MegaPlus) and thin scintillator mounted on a glass prism for applications requiring fast read-out, but not high sensitivity.In this abstract we compare the three coupling methods to each other, and also to a TV-rate fiber-optically
coupled CCD camera.
Direct on-line imaging and quantification of 8-12 bit digital image information for microscopy
Authors: Peters Klaus-Ruediger 
Published: June 2020

In microscopy a digital pixel-accurate intensity processing (PAIP) technology for differential contrast enhancement became available for precise extraction and display of image information. Utilizing PC-based massively parallel processing at 1,600 million instructions per second (MIPS), the PAIP image enhancement can be performed on/off-line in near-real-time for objective image evaluation of 8-12 bit data.We used a PC-based workstation (PiXision PiX4: Visionary Systems Inc., New Haven, CT) on-line in combination with a JSM-890 FSEM (JEOL, Peabody, MA) as well as off-line with imported digital micrograph data and analyzed digital image contrast characteristics. The system provided automated intensity processing of any data independent of image content, size and depth. The images were first smoothed through averaging within a defined significant intensity range (IRs). Each pixel was provided with a variable automatically adjusting local averaging mask, generating a processing characteristic of non-linear point processing. Then, since the spacial processing was pixel-accurate (no spacial distortions of maintained intensity structures), all the contrast information eliminated by the smoothing could be recovered through subtraction of the smoothed from the original or previously smoothed data, followed by linear scaling to 8 bit.
pH-dependent polymorphism of assemblies of cytokine hMIP-lβ
Authors: Cheng N. , Gronenborn A. M. , Clore C. M. , Steven A. C. 
Published: June 2020

Human macrophage inflammatory protein 1β (hMIP-1β), also known as Act-2, is a small protein that belongs to an extensive class of chemotactic cytokines. The high resolution solution structure of hMIP-1β has recently been determined by multidimensional NMR spectroscopy, revealing a dimeric molecule of 2 x 8 kDa. The fold of the monomer is very similar to that of the related cytokine, interleukin-8, which was solved previously. However, the mode of association whereby two monomers form a dimer is quite different. To maintain a sufficiently high concentration of soluble protein for NMR studies, hMIP-1β was dissolved at a pH of 2.5. During solubility trials, it was noticed that the hMIP-1β-containing solutions became birefringent at higher pHs, suggesting the formation of ordered light-scattering structures. We have investigated this phenomenon by negative staining electron microscopy.hMIP-1β was obtained from an expression vector, extracted from inclusion bodies, and purified as described. Protein solutions at 0.7 mg/ml, 100 mM NaCl, 20mM Na citrate, pH2.3, were dialyzed against 20mM ionic strength buffers at pH 4.5, 5.0 and 5.5 (sodium citrate), and pH 6.0 and 7.0 (sodium phosphate) at 4°C. Samples were withdrawn at various time-points, applied to glow-discharged carbon films, stained briefly (5s) with uranyl acetate, and examined in a Philips EM400RT microscope.
Synthesis of mesoscopic structures by co-assembly
Authors: McGehee M. D. , Gruner S. M. , Yao N. , Chun C. M. , Navrotsky A. , Aksay I. A. 
Published: June 2020

Synthesis of materials with nanoscale (1-100 nm) organization is of utmost importance in various applications. Recently, scientists at Mobil described a surfactant-based procedure which yields mesoporous silicate/aluminosilicate materials with pores up to 10 nm in size and lamellar, hexagonal, and bicontinuous cubic structures. Two models were proposed to explain the formation of these materials. The first model assumes that silica polymerizes around a water-surfactant liquid crystal phase while the second model contends that the polymerization of the silica determines the ordering of the surfactants in a co-assembly fashion. Our time-resolved x-ray diffraction studies show that transitions between structures occur during synthesis, supporting the co-assembly model.The procedure for synthesizing cubic surfactant-silicates was as follows: 10.41 g of TEOS (tetraethoxy silane, Alpha Chemicals) was added to 35.86 g of Arquad (29% by weight C16TMAC1 in water, Akzo Chemicals), 25 ml of 1M NaOH, and 5.34 mL of deionized water. This mixture was stirred for 30 min with a magnetic stir bar, causing the TEOS to disperse with the surfactant solution and hydrolyze.
Rotating specimen stage for angle-indexed videotape microscopy of automotive water-pump seals and ring-shaped specimens
Authors: Laurion Tristan A. , Dolson Richard L. 
Published: June 2020

Videocameras and electronic image storage devices have given new vigor and potential to optical microscopy and parts inspection, as well as a powerful means of recording microscopic motion in live-time biological studies. Video and electronic techniques permit fast exploratory examinations simultaneous with extensive archiving, and allow the option of deferring hard copy printing to the time when reports may be needed. These capabilities aid in improving the quality of scanning electron microscopy (SEM) and other analyses that may follow and have great value in communicating with the customer.Our modular system was conceived to permit the imaging of the entire surface of ring-type objects as well to certify that features later characterized in more detail by SEM have been well chosen. The system uses the Nikon SMZ-U optical microscope equipped with a current technology video camera used in combination with a commercial frame grabber device, a dye transfer printer, and VHS videotape recorder.