bluuebunny/crossref_metadata_2025 huggingface dataset:
A scanning electron microscopic study on dissolving process of cholesterol gallstones by chenodeoxycholic acid (CDCA)
Authors: Kanetaka T. , Cho M. , Kawamura S. , Sado T. , Hara K.
Published: June 2020

The authors have investigated the dissolution process of human cholesterol gallstones using a scanning electron microscope(SEM). This study was carried out by comparing control gallstones incubated in beagle bile with gallstones obtained from patients who were treated with chenodeoxycholic acid(CDCA).

The cholesterol gallstones for this study were obtained from 14 patients. Three control patients were treated without CDCA and eleven patients were treated with CDCA 300-600 mg/day for periods ranging from four to twenty five months. It was confirmed through chemical analysis that these gallstones contained more than 80% cholesterol in both the outer surface and the core.

The specimen were obtained from the outer surface and the core of the gallstones. Each specimen was attached to alminum sheet and coated with carbon to 100Å thickness. The SEM observation was made by Hitachi S-550 with 20 kV acceleration voltage and with 60-20, 000X magnification.
Investigations into the mechanism of hormone release by rat steroid-producing cells
Authors: Nussdorfer G. G. , Neri G. , Robba C. , Mazzocchi G. , Bortolussi M.
Published: June 2020

Biochemical evidence suggests that an exocytotic mechanism underlies hormone release in steroid producing cells. However, the demonstration of secretory granules has been so far elusive, probably owing to the fact that in steroid-producing cells there is little or no appreciable intracellular storage of hormonal products. Since numerous reports indicate that microtubules play a permissive role in exocytotic mechanisms, we devised to investigate the effects of vinblastine (an antimicrotubular agent) on rat lutein and adrenocortical cells.

Fifteen adult male rats were divided into 3 experimental groups. Group 1 served as a control; group 2 was given an ip. injection of 25mg/kg of vinblastine (Velban, Lilly) 2 hours before sacrifice; group 3 was treated as group 2, but in addition received an ip. injection of 20IU/kg of ACTH 1 hour before sacrifice. An analogous experiment was performed using fifteen pseudogravid prepubertal rats, except that group 3 received 250 IU of HCG 1 hour before sacrifice. The corticosterone and progesterone concentrations in male and female rats, respectively, were determined by radioimmunological methods, in both the peripheral plasma and the homogenates of the right adrenal or ovary.
Effects of renovascular hypertension on rat adrenal zona glomerulosa
Authors: Mazzocchi G. , Rebuffat P. , Robba C. , Vassanelli P. , Nussdorfer G. G.
Published: June 2020

It is well known that the rat adrenal zona glomerulosa steroidogenic activity is controlled by the renin-angiotensin system. The ultrastructural changes in the rat zona glomerulosa cells induced by renovascular hypertension were described previously, but as far as we are aware no correlated biochemical and morphometric investigations were performed.

Twenty adult male albino rats were divided into 2 experimental groups. One group was subjected to restriction of blood flow to the left kidney by the application of a silver clip about the left renal artery. The other group was sham-operated and served as a control. Renovascular hypertension developed in about 10 days: sistolic blood pressure averaged 165 ± 6. 4 mmHg, whereas it was about 110 ± 3. 8 mmHg in the control animals. The hypertensive and control rats were sacrificed 20 days after the operation. The blood was collected and plasma renin activity was determined by radioimmunological methods. The aldosterone concentration was radioimmunologically assayed both in the plasma and in the homogenate of the left capsular adrenal gland.
Healing in the central nervous system of mammals
Authors: Ashhurst Doreen E. , Berry M.
Published: June 2020

Regeneration of axons in the mammalian CNS appears to be limited to a small group of unmyelinated fibres; the majority of axons are unable to grow across a lesion. Despite much work, the mechanism by which the growth of these axons is inhibited, is not understood. A mature, fully healed lesion is filled by collagenous tissue and bordered by the foot processes of astrocytes. Little is known of the early stages in the healing of a lesion, and since it was considered that the sequence of events in the days immediately following injury might be of special significance in determining whether the regenerating axons cross the lesion, this study was initiated.

Lesions made with a knife were placed stereotactically in the left cerebral hemisphere of 30-day-old Wistar rats of either sex. After periods of 1, 2, 4, 8, 12 and 30 days, the brains were perfused first with 20% and then 5% glutaraldehyde in 0. 1M phosphate buffer, pH 7. 2 containing 0. 05M sucrose.
Towards compositional imaging using auger electron signals
Authors: El Gomati M M , Prutton M , Roberts R H , Barkshire I R , Kenny P G , Greenwood J G
Published: June 2020

Auger electron spectroscopy (AES) is a well established, quantitative surface analytical technique with reasonable accuracy of the order of 1% of an atomic monolayer. When it is combined with a small electron beam diameter, high resolution concentration maps of the surface distribution of elements can be obtained. This has been demonstrated to be a useful and powerful method in surface analysis. However, because of the rather low efficiency of Auger electron production (∼ 10-5 - 10-4 per incident electron) long frame scan times (of the order of hours) have to be employed in the case of multi-element composite samples. The raw images often reflect not only surface elemental distribution but also electron beam fluctuations. In addition, subsurface atomic number variations as well as local surface topography are known to alter the contrast of these images. In order to quantify Auger maps to give concentration distribution of the surface elements, sample and instrumental effects have to be separated.
Compositional mapping using large-angle electron scattering
Authors: Pennycook S. J. , Jesson D. E.
Published: June 2020

High-angle electron scattering represents a relatively small fraction of the total elastic scattering, typically 10%, but is still orders of magnitude larger than the intensity of secondary excitations such as X rays or core loss inelastically scattered electrons. It is possible, therefore, to form an image from thin samples at high resolution with acceptable signal to noise ratio in a reasonable collection time. The STEM high-angle annular detector introduced by Howie detects only the scattering which occurs very close to the atom sites so that the scattering cross section a approaches the Z2 dependence of unscreened Rutherford scattering. Furthermore, in all but the very thinnest crystals the detected signal is dominated by thermal diffuse scattering, which is generated incoherently by each atom in proportion to the electron intensity close to its site. Due to the large angular range of the detector, dynamical effects in the outgoing thermal diffuse scattering (Kikuchi lines) are averaged out and have no effect on the total intensity detected.
Comparison of luminal surface observations of the healing response of an experimental small-diameter woven Dacron and expanded polytetrafluoroethylene arterial prosthesis
Authors: Wolf Michael F. , Coscio Michael R. , Bianco Richard W.
Published: June 2020

In surgical vascular reconstruction, the principal synthetic prosthesis materials are DacronTM (woven or knitted polyethyleneterephthalate [PET] fibers) and expanded polytetrafluoroethylene (ePTFE). Following implantation, the markedly different microstructure and material chemistry of each synthetic blood conduit imparts a characteristic healing response. The ideal intimal healing response however is the rapid development of a stable nonthrombogenic autologous surface, preferably lined with functional confluent endothelial cells. Since small diameter prostheses (less than 6 mm I.D.) made of either material are considered to perform inadequately in peripheral vascular and coronary artery reconstructions, novel materials and composites are the focus of much investigation. In this report, scanning electron microscopy (SEM) is used to follow and compare the luminal surface healing response of a 4 mm I.D. ePTFE and a novel woven microfiber Dacron™ prosthesis coated with a glow-discharge plasma tetrafluoroethylene (PET+pTFE).
Microbiologically induced corrosion of stainless steel by Desulfovibrio vulgaris: An scanning electron microscope study
Authors: Garcia Roger , Li Fang , Hendrickson Lester
Published: June 2020

The corrosion of mild steel by sulfate reducing bacteria has been studied quite extensively. However, with the replacement of mild steels with stainless steel in many of these applications numerous sightings of corroding stainless steel have been made as well. Initially, the cathodic depolarization theory was widely accepted as the mechanism for both. The essential part of this theory involves the removal of hydrogen from the metal surface. Hydrogenase in Desulfovibrio allows utilization of elemental hydrogen from the cathode of the corrosion cell. This causes the reduction of sulfate whereby the biological cell gets its energy via a respiration process. Finally, the oxygen from the sulfate becomes available to the cathode and hence corrosion is enhanced. Without this reducing action the cathode would become polarized thereby decreasing the EMF and lowering the corrosion rate. Among other proposed mechanisms are differential aeration cells and corrosive products produced by the bacteria.
Undecagold labeling of tRNA
Authors: Hainfeld James F. , Alford Kyra M. , Sprinzl Mathias , Mandiyan Valsan , Tumminia Santa J. , Boublik Miloslav
Published: June 2020

The undecagold (Au11) cluster was used to covalently label tRNA molecules at two specific ribonucleotides, one at position 75, and one at position 32 near the anticodon loop. Two different Au11 derivatives were used, one with a monomaleimide and one with a monoiodacetamide to effect efficient reactions.

The first tRNA labeled was yeast tRNAphe which had a 2-thiocytidine (s2C) enzymatically introduced at position 75. This was found to react with the iodoacetamide-Aun derivative (Fig. 1) but not the maleimide-Aun (Fig. 2). Reaction conditions were 37° for 16 hours. Addition of dimethylformamide (DMF) up to 70% made no improvement in the labeling yield. A high resolution scanning transmission electron micrograph (STEM) taken using the darkfield elastically scattered electrons is shown in Fig. 3.
Processing of mRNA and proteins at the neuromuscular synapse
Authors: Bursztajn S. , Berman S. A.
Published: June 2020

Muscle cells are multinucleated and each nucleus has the capacity of expressing many genes. Both in vivo and in vitro the AChR clusters appear to be associated with clusters of nuclei. To answer the question whether each nucleus in a multinucleated myotube is equally active in expressing the AChR message and how this may be regulated by motor neuron we have carried out in situ hybridization with radiolabeled probes to the AChR α subunit RNA. We have found that muscle cells do not express the mRNA at equal levels. The response we have observed is not due to the death, or the metabolic inactivity of some nuclei. Cells hybridized with a probe for U1 small nuclear RNA, show grains in every nucleus with approximately the same grain distribution. To better understand how the motor neuron may regulate the AChR gene expression we have combined in situ hybridization with immunocytochemistry. We radiolabeled the AChR intron/exon probes with two (35S) thiodeoxynucleoside triphosphates (dNYPs).
High-pressure freezing of cells in suspension for low-voltage scanning electron microscopy (LVSEM)
Authors: Malecki Marek , Pawley James , Ris Hans
Published: June 2020

The ultrastructure of cells suspended in physiological fluids or cell culture media can only be studied if the living processes are stopped while the cells remain in suspension. Attachment of living cells to carrier surfaces to facilitate further processing for electron microscopy produces a rapid reorganization of cell structure eradicating most traces of the structures present when the cells were in suspension. The structure of cells in suspension can be immobilized by either chemical fixation or, much faster, by rapid freezing (cryo-immobilization). The fixation speed is particularly important in studies of cell surface reorganization over time. High pressure freezing provides conditions where specimens up to 500μm thick can be frozen in milliseconds without ice crystal damage. This volume is sufficient for cells to remain in suspension until frozen. However, special procedures are needed to assure that the unattached cells are not lost during subsequent processing for LVSEM or HVEM using freeze-substitution or freeze drying. We recently developed such a procedure.
Morphological effects of sulfur mustard on a human skin equivalent
Authors: Petrali J.P. , Oglesby S.B. , Justus T.A.
Published: June 2020

We have previously reported morphological correlates of sulfur mustard (HD) toxicity in several model systems: the human skin grafted athymic nude mouse; the hairless guinea pig; and human cells in culture. We are now describing HD effects in a human skin equivalent, TESTSKIN®, and comparing these effects with those already reported for animal models and cells in culture. The human skin equivalent (HSE) is used here as an organotypic in vitro model system to bridge the knowledge gap between HD effects in monotypic cells in culture and animal in vivo effects. Additionally, HSE allowed study of HD toxicity which circumvented the concern of using human biopsied tissue.
Ultrastructural evaluation of the effects induced by a macrolide antibiotic (LY281389) on L6 muscle cells in vitro
Authors: Horn J. W. , White S. L. , Laska D. A. , Buening M. K. , Novilla M. N. , Jensen C. B. , Hoover D. M.
Published: June 2020

Toxic doses of LY281389 (9-N(n-propyl)erythromycylamine), a cationic amphophilic macrolide antibiotic, induce a generalized cytoplasmic vacuolar change in tissues of rats and dogs. The present in vitro study using L6 cells (a line derived from neoplastic skeletal muscle) was done to evaluate sequentially early cytologic changes.

The L6 cells seeded on tissue culture plates were allowed to grow to a uniform density. The culture medium was then replaced with control medium or medium containing 0.25 mg/ml LY281389 for exposures of 0.5, 2, 6, 12, 24, or 48 hours. The L6 cells were then fixed for 1-hour in modified Karnovsky's solution (pH 7.2), rinsed with 0.1 M sodium cacodylate buffer (pH 7.2) and postfixed for an additional hour in 2% osmium tetroxide (pH 7.2). Following a second buffer rinse, the L6 cells were dehydrated in graded ethanol solutions. The cells on each plate were embedded with epoxy resin (EPON 812), selected areas were scribed out and re-embedded for orientation. Ultrathin sections (silver to gold color interference range) were mounted on copper 200 mesh grids, counterstained, and examined using a Philips EM410LS electron microscope at 60kV.
Localization of the cystic fibrosis gene product in a transfected Sf9 insect cell line
Authors: Ackerley C.A. , Kartner N. , Riordan J.R. , Phillips M.J.
Published: June 2020

The cystic fibrosis gene product (CFTR) is thought to be either directly or indirectly involved with Cl- conductance in epithelial cells in which it is produced. Utilizing the Baculovirus expression vector system, CFTR has been expressed in Sf9 insect cells which in turn display a regulated Cl- conductance employing the same techniques as those used to demonstrate Cl- conductance in epithelial cells. This was not observed in mock-infected Bgalactosidase producing or non-infected Sf9 cells.

Two cell lines, one infected with CFTR expressing recombinant baculovirus and the other infected with B-galactosidase expressing recombinant baculovirus were fixed in 2% paraformaldehyde and 0.001% glutaraldehyde in 0.1 M HEPES buffer, PH7.2 for 2 hours. They were then rinsed several times with buffer, infiltrated with 2% agarose and infused with 2.3 M sucrose for at least 24 hours. The samples were then frozen by rapid immersion in liquid nitrogen cooled FREON 22 and ultrathin cryosections cut with a cryoultramicrotome and immunogold labelled with purified monclonal antibodies against CFTR (1) using the method described by Griffiths.
Elemental identification by x-ray images of pathologic tissue
Authors: Mumaw Virgil R.
Published: June 2020

Elemental localization in biological specimens, using x-ray microanalysis, can best be demonstrated with x-ray images. X-ray images provide useful detail about elemental localization as well as relative quantitative information. The comparison of different elements using color overlay images graphically demonstrates interrelationships in ultrathin sections. Pathology specimens with unknown crystal formations were studied using x-ray imaging.

Formalin fixed tissue from autopsy and surgical pathology were examined. For this study lung, lymph nodes and skin were used. Some specimens were removed from paraffin blocks and deparaffinized prior to processing for electron microscopy. Tissues were dehydrated and embedded in PolyBed epoxy resin (Polysciences, Warrington, PA). No osmium postfixation was used. Thin unstained sections were coated with carbon using a vacuum evaporator. The sections were examined and analyzed using a Philips EM 400 TEM/STEM microscope equipped with an EDAX PV9900 microanalyzer with the EDAX imaging package. X-ray images were collected using dwell times varying from 10 msec to 1 sec. Longer dwell times resulted in collection times as long as 14 hrs.
Antigen presentation between cultured cerebral smooth muscle and endothelial cells: Confocal laser scanning microscopy study
Authors: Ghoneim Shams , Fabry Zsuzsa , Keiner Judy , Hart Michael
Published: June 2020

We have previously demonstrated with transmission and scanning electron microscopy that cultured endothelia and smooth muscle cells derived from cerebral microvessels can maintain their in vivo close relationship. This close association of the two cell population has functional implications which could explain humoral transmission of substances, the exchange of metabolites or play a significant role in autoimmune responses.

It is generally accepted that the expression of antigen on the surface of a given cell is necessary for antigen presentation by that cell type. The role of antigen presentation and expression on brain cells could be significant in diseases such as multiple sclerosis (MS) and experimental allergic encephalitis (EAE).
IVEM and SEM images of whole cells cultured on EHS matrix (Matrigel)
Authors: Smith Nancy R. , Benson Stephen , Larabell Carolyn
Published: June 2020

To investigate the interactions of cells with the extracellular matrix (ECM), rat lung fibroblasts (RLF) were cultured on an ECM extracted from EHS sarcoma to serve as an in vitro morphogenic model system. Surface features of RLF/EHS matrix interactions were viewed using the scanning electron microscope (SEM) and the intermediate voltage electron microscope (IVEM).

For SEM, RLF were plated on EHS coated 13 mm glass coverslips at a density of 1 X 104/cm2. Coverslips were coated by spreading 25 μl of chilled EHS matrix (15 mg/ml) on the coverslip and incubating at 37° for 60 min. For IVEM, formvar coated gold grids were submerged in EHS matrix prior to polymerization. Controls consisted of cells grown on glass or gold grids minus ECM.
Surface structure of the photosystem II complex
Authors: Miller Kenneth R. , Jacob Jules S.
Published: June 2020

The Photosystem II (PS-II) complex is organized around a photosynthetic reaction center (RC) embedded in the photosynthetic membrane. PS-II traps the energy of sunlight and uses it drive highenergy electron transport across the photosynthetic membrane. PS-II is closely associated with a group of proteins known as the oxygen-evolving complex (OEC), which are bound to the inner surface of the photosynthetic membrane. This complex splits water to yield electrons that are passed to the RC, releasing molecular oxygen. We have used freeze-etch electron microscopy to study 2-dimensional crystals of the PS-II complex obtained from a photosynthetic mutant of barley (viridiszb63) kindly provided by Dr. David Simpson of the Carlsberg Institute of Copenhagen (Simpson & von Wettstein, 1980). The photosynthetic membranes of these mutant plants lack photosystem I, and consequently contain extensive crystalline membrane regions enriched in PS-II. These plants are an excellent source of PS-II sheetlike crystals, obtainable without the use of detergents or chemical modification: Figure 1, prepared by quick-freezing, deep-etching, and rotary shadowing, illustrates the appearance of these sheetlike crystals.
Maternal plastids in alfalfa egg cells: A qualitative and quantitative study
Authors: Zhu T. , Mogensen H. L. , Smith S. E.
Published: June 2020

Genetic studies have shown that alfalfa (Medicago sativa L.) has a predominance of paternal plastid inheritance. Results from cytological studies, which have quantified plastids and plastid nucleoids within generative and sperm cells of alfalfa, demonstrate that male germ cell composition does not adequately explain known differences in patterns of plastid inheritance among specific genotypes. To determine the behavior and fate of female plastids, we are currently studying unfertilized eggs, zygotes and early embryos. Here we report preliminary results on the cytological composition of alfalfa egg cells in genotype, a relatively strong female plastid transmitter, and genotype CUFB, a relatively weak female.
Ultrastructure of Plasmopara halstedii containing virus-like particles
Authors: Freeman Thomas , Gulya Thomas
Published: June 2020

Mycoviruses have been found in more than 100 species of fungi and it is estimated that they may occur in as many as 500 species (Buck, 1986; Hollings, 1978; and Lemke, 1981). Mycoviruses range in size from small, isometric particles (25 nm in diameter) to long, flexuous rods exceeding 2000 nm in length, with the majority being isometric particles ranging in size from 25-50 nm (Buck, 1986). Mycoviruses hae been reported in all groups of fungi, but have been least frequently observed in the Oomycetes.

Sunflower (Helianthus annuus) seedlings were inoculated with a North Dakota isolate of race 2 Plasmopara halstedii, the causal agent of sunflower downy mildew. Systemic disease symptoms developed within 14 days and samples were prepared for transmission electron microscopy using standard techniques.
Elemental composition of polyphosphate bodies in aluminum-treated Pisolithus tinctorius: Comparison of chemical and freeze-substitution methods
Authors: McQuattie C. J. , Klomparens K. L.
Published: June 2020

The toxic effects of aluminum (Al) ions on plants growing in acidic soils may be ameliorated by mycorrhizal (M) fungi, which form symbiotic associations with roots of many forest trees. Pisolithus tinctorius, a M fungus found in many acid soils, is tolerant of high levels of Al. It is not known, however, if Al is excluded from the hyphae or compartmentalized within specific organelles or areas of the hyphae. Many M fungi have been shown to accumulate divalent cations, especially Ca, with P in polyphosphate (polyP) bodies. Whether Al ions might also be localized in polyP bodies is unclear. Previously, most ion localization studies in M fungi have been performed using conventional chemical fixation (CF) methods. However, ion movement due to aqueous fixation has been demonstrated. The inhibition of ion movement during fixation is a superior feature of specimen preparation by rapid freezing followed by freeze substitution (FS). The purpose of this study was to determine by energy dispersive X-ray microanalysis (EDS) of ultrathin sections the elemental composition of polyP bodies and other hyphal areas in P. tinctorius grown at different Al levels and prepared by CF or FS methods.
An ultrastructural study of the epicuticle of Ascaris suum
Authors: Ramnani D. M. , Cain G. D.
Published: June 2020

Ascaris suum, an intestinal parasite of swine, has a complex body wall, consisting of nine distinct layers when observed by transmission electron microscopy. The outermost layer of the cortex is a trilaminar region, the epicuticle, that is similar in appearance to plasma membrane. Parasitic nematodes interact with their hosts through this layer. The structure of the epicuticle is being studied with transmission and scanning electron microscopes.

Parasites were obtained from an abattoir at Waterloo, Iowa, and maintained in Harpur's saline medium for up to two days. Cuticles were separated by microdissection and washed in 0.1M phosphate buffer, pH 7.2. Trump's universal fixative (4% Formaldehyde: 1% Glutaraldehyde in phosphate buffer, pH 7.2 ) was used for 1 h at room temperature for primary fixation. Surface charge of epicuticle was studied by incubating cuticle with cationized ferritin (Polysciences) at a concentration of lmg/ml in 0.1M phosphate buffer, pH 7.2, for 30 minutes at room temperature.
X-ray microanalysis and microbial colonization of leaves in Lake Wallenpaupack in eastern Pennsylvania
Authors: Majumdar S.K. , Daehler C.C. , Baker M.A.
Published: June 2020

The microbial decomposition of leaf litter provides a major source of organic carbon and other nutrients and is important in ensuring the stability of lake ecosystems. Lake leaf litter can also act as a sink for immobilization of nutrients and elements. This investigation utilizes the scanning electron microscope and the energy-dispersive x-ray microanalysis technique to study the effects of agricultural or septic tank leakage pollutants on microbial colonization and elemental accumulation in leaves submerged in a lake in eastern Pennsylvania.

White oak and red maple leaves were placed in mesh bags in the littoral zone of three cove sites at lake Wallenpaupack, a 13 mile-long, 5,700 acre lake located in the Pocono Mountains in eastern Pennsylvania. Cove 1 was surrounded by a wooded area and received water input from a wetland preserve. Cove 2 was suspected of contamination by septic tank leakage. Cove 3 received water input from a tributary passing through a heavily farmed drainage basin.
Cytodex dextran beads as a substrate for observing the effects of selenium on bacterial morphology
Authors: Regnier G. , Smith N. , Gonzalez R. , Dixon B.
Published: June 2020

Selenium, through the process of bioconcentration, can accumulate to reach toxic levels in the tissues of birds, fish and other wildlife. To date, little research has focused on the effect os selenium on bacteria and in particular, on the nitrifying bacteria which provide a considerable beneficial impact on the environment. Preliminary studies in our laboratory, have indicated that selenium can disrupt the nitrogen cycle by inhibiting the conversion of nitrite to nitrate by Nitrobacter spp. The effect of selenium on Nitrobacter is as of yet unknown. This break in the normal cycle allows nitrite, which is highly toxic in low levels to aquatic animals, to accumulate in the environment. This study was undertaken to investigate the effects of selenium assimilation, by nitrifying bacteria, on the nitrogen cycle and any resultant morphological changes in cell structure.
Artifacts in microscopy: Photography and interpretation
Authors: Wergin William P. , Pooley Christopher D.
Published: June 2020

For the biologist, artifacts can be defined as any manifestations that result from mechanical manipulations or chemical reagents and are not representative of authentic structural relationships. In electron microscopy, living biological tissues are generally sampled, fixed, frozen, dehydrated, embedded, critical point dried, sectioned, mounted, stained, and/or coated - mechanical and chemical manipulations that undoubtedly produce artifacts. However, potential problems do not end once the specimen is in the microscope, because numerous artifacts, which we will classify as photographic and interpretive, are also associated with recording images, processing films, printing negatives, and understanding the final print. Artifacts associated with these four procedures will be addressed in this discussion.
A comparison of CeB6 and LaB6 thermionic cathodes
Authors: Christman L. A. , Schwind G. A. , Swanson L. W.
Published: June 2020

In recent years lanthanum hexaboride (LaB6) has proven to be an excellent thermionic cathode material, and has become the standard for most electron beam applications. The reasons that LaB6 is superior to tungsten as a material for thermionic cathodes are that it has a low work function and low volatility at high temperature. It is also relatively insensitive to most contaminants (or can at least be thermally cleaned of most contaminants).

The most important consideration in cathode performance is its volatility at the operating temperature required for achieving the required current density. The work function for clean LaB6 is 2.6 eV compared with 4.5 eV for tungsten, thus it is possible to operate the former cathode at a considerably reduced temperature compared with tungsten. The volatility is greatly reduced, not only because of the lower operating temperature, but also because LaB6 has a relatively high heat of vaporization considering its low value of work function.
STM studies of crystal growth
Authors: Lagally M. G.
Published: June 2020

It has been recognized since the earliest days of crystal growth that kinetic processes of all Kinds control the nature of the growth. As the technology of crystal growth has become ever more refined, with the advent of such atomistic processes as molecular beam epitaxy, chemical vapor deposition, sputter deposition, and plasma enhanced techniques for the creation of “crystals” as little as one or a few atomic layers thick, multilayer structures, and novel materials combinations, the need to understand the mechanisms controlling the growth process is becoming more critical. Unfortunately, available techniques have not lent themselves well to obtaining a truly microscopic picture of such processes. Because of its atomic resolution on the one hand, and the achievable wide field of view on the other (of the order of micrometers) scanning tunneling microscopy (STM) gives us this opportunity. In this talk, we briefly review the types of growth kinetics measurements that can be made using STM. The use of STM for studies of kinetics is one of the more recent applications of what is itself still a very young field.
Chromosome structure and dynamics as revealed by 3-D and 4-D imaging
Authors: Swedlow Jason R. , Hiraoka Yasushi , Paddy Michael R. , Sedat John W. , Agard David A.
Published: June 2020

Cellular DNA is packaged into chromosomes through complex processes whereby the DNA becomes compacted up to 10,000-fold by assembly into higher-order structures. In spite of many decades of intense study, the details of this process are not known. In addition, there is little information describing the dynamics of these higher-order structures during DNA replication, transcription, and mitosis. The goal of our laboratory is to use state-of-the-art 3-D imaging methods to understand how DNA is arranged into chromosomes and how chromosomes are spatially and temporally organized in the nucleus.

Towards this end, we have developed a fluorescence microscope workstation optimized for collecting three-dimensional multi-wavelength data from fixed and living specimens. The multiplewavelength capability of this system allows us to simultaneously examine the spatial relationships of different chromosomal components in 3-D. Fixed samples can be labelled with up to three different fluorophore-conjugated antibodies and a fluorescent DNA binding dye, DAPI. Three-dimensional multi-wavelength data collection proceeds by sequentially recording images of each color, changing the focus, and then repeating the process as many times as necessary.
Confocal light microscopy and electrophysiology of neurons in culture
Authors: Turner J. N. , Szarowski D. H. , Fejtl M. , Ayrapetyan S. N. , Carpenter D. O.
Published: June 2020

Cell volume changes and regulation are thought to be important in the physiology and pathology of neurons. Osmotic challenges alter a number of physiologic parameters including electrical properties, chemosensitivity, and Na pump activity. Thus, a good method for correlating images of the living cell's surface with electrophysiologic measures is needed, and we are exploring the use of confocal light microscopy for imaging neurons in culture.

Isolated neurons from ganglia of Aplvsia California were plated on polylysine coated coverslips in L15 media with 20% hemolymph at room temperature. Cells stained with “Dil” were imaged with a Bio—Rad MRC—600 and an Olympus BH—2. Images were displayed using Bio—Rad's maximum projection procedure, or Vital Images’ Voxel View software run on an IBM RISC 6000 Powerstation 320. Electrophysiologic properties were assessed after imaging.
Instrumentation and methodology for electron crystallography
Authors: Downing Kenneth H.
Published: June 2020

The determination of three-dimensional structures of macromolecules at atomic resolution remains one oμf the great promises of electron microscopy. Instrumental problems which had to be overcome in order to achieve single-atom resolution have long been recognized and to a large extent overcome. It is interesting to note that most of the instrumental developments were already incorporated in a microscope under construction over 20 years ago. However, the application of techniques to circumvent limitations imposed by the sensitivity of organic specimen to radiation damage are still developing. Electron crystallography, which takes advantage of the ability to obtain useful information from images taken at very low exposure, is close to realizing the potential. The state of electron crystallography has advanced rapidly during the last few years. The structure of one protein, bacteriorhodopsin, has been determined from EM data, and several other structures are advanced to the point of fitting the peptide chain to a high-resolution density map.
mRNA localization by Electron microscopicin situhybridization
Authors: Pollock J. A. , Martone M. , Deerinck T. , Ellisman M. H.
Published: June 2020

Localization of specific proteins in cells by both light and electron microscopy has been facilitate by the availability of antibodies that recognize unique features of these proteins. High resolution localization studies conducted over the last 25 years have allowed biologists to study the synthesis, translocation and ultimate functional sites for many important classes of proteins. Recently, recombinant DNA techniques in molecular biology have allowed the production of specific probes for localization of nucleic acids by “in situ”hybridization. The availability of these probes potentially opens a new set of questions to experimental investigation regarding the subcellular distribution of specific DNA's and RNA's. Nucleic acids have a much lower “copy number” per cell than a typical protein, ranging from one copy to perhaps several thousand. Therefore, sensitive, high resolution techniques are required. There are several reasons why Intermediate Voltage Electron Microscopy (IVEM) and High Voltage Electron Microscopy (HVEM) are most useful for localization of nucleic acidsin situ.
Selective staining of neurons for thick-section intermediate and High-Voltage EM studies
Authors: Ellisman M. H. , Wilson C. J.
Published: June 2020

Methods are needed that enable the visualization of selectively stained neurons, neuronal processes and intracellular structures which provide higher resolution than can be obtained with the light microscope. Procedures have been developed using thick sections that allow direct visualization of the three-dimensional structure of the neuron and the visualization of intracellular organelles. Using these methods the geometry of neurons may be accurately measured and this information may then be used to test theoretical predictions about the way in which electrical signals of synaptic origin are processed by the cells. Intermediate and high voltage electron microscopes (HVEM) are well suited to this application because of their high resolution and ability to form images of thick sections. Use of these instruments requires development of selective stains that can produce diffuse cytoplasmic staining of specific cells, cell populations or organelle systems on the basis of their functional properties, or organelle systems.
Quantitative chemical mapping: Spatial resolution
Authors: Ourmazd A. , Baumann F.H. , Bode M. , Kim Y.
Published: June 2020

Quantitative Chemical Mapping is an electron microscopic technique capable of revealing compositional variations in crystalline materials. It combines chemical lattice imaging which maps the sample composition, with vector pattern recognition, which quantifies the local information content of the image to measure the local sample composition. Here we briefly address the spatial resolution of this technique, assuming complete familiarity with its theoretical underpinnings.

In chemical imaging, we are concerned with the way that a compositional inhomogeneity is imaged under conditions appropriate for chemical sensitivity, and how the pattern recognition algorithm extracts information from a chemical lattice image. The problem can be formulated as follows. Given a “chemical impulse” of a specific shape, such as a column of Al atoms imbedded in GaAs (approximating a δ-function), an abrupt interface (a θ-function), or a diffuse interface (e.g., with an error function profile), what is the shape of the impulse on the analyzed chemical image? Or, alternatively, what region of the sample contributes to the information content of an image unit cell? By reciprocity, these two formulations are equivalent.
Resolution limits in annular dark-field STEM
Authors: Loane Russell F. , Xu Peirong , Silcox John
Published: June 2020

Annular dark field scanning transmission electron microscopy (ADF STEM) is capable of resolving atomic structures with Z contrast. Better than 2 Å resolution at 100 kV has been demonstrated with a Cs of 1.3 mm and 0.7 mm. The images are apparently simple to interpret and change little with thickness or defocus. However, much of the reported work is qualitative, or semi-quantitative at best, so that the customarily adopted incoherent imaging model is not well established and a clear understanding of the limits (e.g., detectable AZ at a given ratio of spacing to resolution) is lacking. We present an exploration of such limits for the simple specimen, (100) InP, using quantitative ADF STEM and image simulation.
High-voltage and high-resolution TEM in materials research: Penetration and Resolution Issues
Authors: Fisher Robert M.
Published: June 2020

Transmission electron microscopy has produced most of what is known about the microstructures of inorganic materials. Although some internal structures can be seen on etched surfaces using replicas or the SEM, the lack of electron diffraction data eliminates the availability of crystallographic information so that the identity and habit plane of precipitates or characteristics of lattice defects cannot be determined. However, the old extraction replica technique has come back into vogue for analytical microscopy because matrix effects in X-ray or ELS spectra can be eliminated. Extraction replicas can also be used for HREM studies of small constituents in multi-phase materials when standard specimen thinning techniques are inapplicable.
Using high-vacuum evaporation to obtain high-resolution low-temperature images of freeze-fractured membranes from yeast
Authors: Wergin William P. , Erbe Eric F.
Published: June 2020

In previous low temperature studies of fractured yeast membranes that were examined in a conventional SEM, Wergin and Erbe demonstrated that resolution could be enhanced by substituting conventional sputter coating of gold with high vacuum evaporation of platinum/carbon. Although this procedure did not clearly resolve membrane particles, the frozen sample could be removed from the SEM and the Pt/C “coat” or replica could be recovered for further examination. Using instruments with greater resolution such as a TEM, as well as a field emission (FE) SEM, to examine the replica did reveal the membrane particles on the replica; therefore the authors concluded that failure to resolve these membrane structures in a conventional SEM probably resulted from limitations of the instrument rather than the coating procedure. Recently, we had the opportunity to interface a FESEM with a cryosystem; this combination was used to determine if samples shadowed in a high vacuum evaporator could be transferred to a cold stage in a FESEM to resolve the macromolecular structure of yeast plasma membranes.
High-resolution SE-I imaging of biological membranes using a Schottky field-emission in-lens SEM
Authors: Apkarian R.P.
Published: June 2020

Previous notes have described efforts to routinely collect quality SE-I image contrasts of biologically significant particulate membrane features (1-10 nm) in the context of whole cells and tissue fragments. Utilizing SEMs equipped with in-lens specimen stages and field emission sources (Schottky and cold cathode) operated at 15-30 kV and in conjunction with specimens coated with 1 nm Cr films (Z=24), nanometer resolution of biological samples may be attained. This note describes the definition of optimal electron source conditions for the high resolution imaging of cell membrane features μ 10 nm. The Schottky field emitter equipped ISI DS-130F SEM, in-house at the Yerkes Research Facility, was operated at 5-20 kV accelerating voltages (A.V.) and at 4 or 4.8 kV extraction voltages (E.V.). Although we have published images of soft and hard biological sections and isolated cells containing 1-10 nm particle contrasts by operating the SEM at 25-30 kV A.V., we have maintained 4 kV extraction voltage and not attempted using 4.8 kV above 25 kV A.V.
Microscopy studies of powdery mildew on summer squash
Authors: Gardner John S. , Hess W. M.
Published: June 2020

Powdery mildews are characterized by the appearance of spots or patches of a white to grayish, powdery, mildewy growth on plant tissues, entire leaves or other organs. Ervsiphe cichoracearum, the powdery mildew of cucurbits is among the most serious parasites, and the most common. The conidia are formed similar to the process described for Ervsiphe graminis by Cole and Samson. Theconidial chains mature basipetally from a short, conidiophore mother-cell at the base of the fertile hypha which arises holoblastically from the conidiophore. During early development it probably elongates by polar-tip growth like a vegetative hypha. A septum forms just above the conidiophore apex. Additional septa develop in acropetal succession. However, the conidia of E. cichoracearum are more doliform than condia from E. graminis. The purpose of these investigations was to use scanning electron microscopy (SEM) to demonstrate the nature of hyphal growth and conidial formation of E. cichoracearum on field-grown squash leaves.
Computer recognition of symmetry in sad diffraction patterns
Authors: Jackson A. G. , Park J. , LeClair S.
Published: June 2020

Applications of artificial intelligence to transmission electron microscopy are being studied in the context of a system that can learn to recognize images generated in transmission electron microscopy (TEM), provide interpretation of the imaging conditions, and provide assistance to the user in reaching conclusions about the experiment. Several computing programs exist which allow the TEM analyst to generate diffraction patterns, stereograms, lattices and structures for any inorganic crystal structure. These programs, however, do not act on their own. They require a knowledgeable user to strongly interact with them in order to generate the needed information.

The first step beyond such a computation program is an expert system that has in it the collected observations and methodologies of an expert TEM analyst. The user must still interact strongly with the system via questions presented to the user or presented by the user. If a question is asked that is beyond the stored knowledge in the system, no action occurs. Although such a system would be of value to any TEM user, many experiments are conducted on materials that are new or for which information is not available.
A new Electron Microscopy tomography: Least squares pseudoimage reconstruction technique
Authors: Zhang Nengyu
Published: June 2020

The least square pseudoimage (LSP) Reconstruction technique is based on matrix inversion. Due to the absence of information in electron microscopy, usually in an angle range from 60° to 90°, the matrix is degraded. The degraded matrix is used to compute two-dimensional (2D) projections of the three-dimensional (3D) object along the Z-axis (direction of the electron beam) and tilted around the Y-axis (tilt axis). Applying the pseudoinverse of the degraded matrix to electron micrographs, which are 2D projections of the 3D object, gives the pseudoimage.

Since all of the slices which are perpendicular to the Y axis are related to the same degraded matrix, the 3D reconstruction problem can be simplified as series 2D reconstruction problems. All of the 2D pseudoimages are formed by the same pseudoinverse of a degraded matrix. In this case, however, the degraded matrix projects a 2D object onto an ID projection along the Z axis. Each slice is computed from a single line in all of the electron micrographs.
3-D variance of weighted back-projection reconstruction and its application to the detection of 3-D particle conformational changes
Authors: Liu Weiping
Published: June 2020

To overcome the radiation damage in electron microscopy, the method of 3-D reconstruction from single exposure, random conical tilting series has been developed for single particles with one preferred orientation on the grid. Due to the high noise components in the projections which propagate to the 3-D reconstruction, there is the question about the reproducibility of individual features in the reconstruction. This question cannot be answered by a resolution estimation, because resolution describes the average reliability of the features across the 3-D reconstruction. Also when two independent reconstructions of related particles (e.g. labeled vs. unlabeled) are compared, the following questions are raised: could they have arisen from the same structure? where are the feature differences located? how reliable are the conclusions? These questions can be answered by estimating the variances of the 3-D reconstructions. Since for the random conical method, different projections correspond to different particles on the grid, the conformational differences of these particles can contribute a substantial and recurrent portion to the noise components of their projections. A 3-D variance map should also reveal the 3-D locations of such conformational changes.
Zero-loss omega-filtered REM and RHEED on the new Zeiss 912
Authors: Spence J.C.H. , Mayer J.
Published: June 2020

The Zeiss 912 is a new fully digital, side-entry, 120 Kv TEM/STEM instrument for materials science, fitted with an omega magnetic imaging energy filter. Pumping is by turbopump and ion pump. The magnetic imaging filter allows energy-filtered images or diffraction patterns to be recorded without scanning using efficient parallel (area) detection. The energy loss intensity distribution may also be displayed on the screen, and recorded by scanning it over the PMT supplied. If a CCD camera is fitted and suitable new software developed, “parallel ELS” recording results. For large fields of view, filtered images can be recorded much more efficiently than by Scanning Reflection Electron Microscopy, and the large background of inelastic scattering removed. We have therefore evaluated the 912 for REM and RHEED applications. Causes of streaking and resonance in RHEED patterns are being studied, and a more quantitative analysis of CBRED patterns may be possible. Dark field band-gap REM imaging of surface states may also be possible.