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Influence of initial electron beam characteristics on monte carlo calculated absorbed dose distributions for linear accelerator electron beams.

The least known parameters in a Monte Carlo simulation of a linear accelerator treatment head are often the properties of the initial electron beam directed onto the exit vacuum window. Several initial beams with different spatial fluence distributions, angular divergences and energy spectra have been transported through the geometry of a scattering foil accelerator. The electron beam characteristics (energy spectrum and angular distribution) at the phantom surface and the subsequent relative absorbed dose distribution in a water phantom were calculated. The dose distribution was found to be insensitive to the geometrical properties of the initial beam. Furthermore, the lateral dose profiles are unaffected by the energy spectrum of the initial beam. The effect on the depth-dose curve is negligible if the initial energy spectrum is symmetric (e.g., Gaussian shaped) and its full width at half maximum (FWHM) is less than approximately 10% of the most probable energy. A larger FWHM will decrease the normalized dose gradient, but will not affect the dose in the build-up region. An asymmetric wedge shaped spectrum with a low-energy extension simultaneously increases the dose in the build-up region and decreases the dose gradient. The relationship between the energy spectral width and the normalized dose gradient is, however, smaller than published analytical expressions indicate. Some well-established energy-range relationships were shown to be accurate for most of the initial beams studied. The energy spectrum at the phantom surface was also derived from a measured depth-dose curve through different methods. The extracted spectrum depends on the beam model and the spectral reconstruction algorithm. Even though the depth-dose curve is fairly independent of initial beam characteristics, a correct description of the low-energy tail of the energy spectrum is important to obtain good agreement between measured and Monte Carlo calculated doses in the build-up region.

Electrons↗

Low-voltage backscattered electron imaging of non-coated biological samples in a low-vacuum environment using a variable-pressure scanning electron microscope with a YAG-detector.

A variable-pressure scanning electron microscope (VP-SEM) equipped with a high-sensitive backscattered electron (BSE) detector of the YAG type was applied to studies of biological tissue samples. The rat kidney and trachea were fixed, dehydrated in ethanol, critical point-dried and examined in the VP-SEM under a specimen chamber pressure of 1 to 150 Pa. The high-resolution surface images of the non-coated specimens were obtained in a low-vacuum (1-20 Pa) environment at an accelerating voltage of 5 kV, while the images at 10-20 kV contained information beneath the surface of the specimens. The observation in the VP-SEM equipped with YAG detector in a low-vacuum (1-20 Pa) environment at low (3-5 kV) accelerating voltages is useful for the three-dimensional analysis of the surface morphology of biological non-coated samples.

Animals↗

In vivo monitoring of the potassium channel KcsA in Streptomyces lividans hyphae using immuno-electron microscopy and energy-filtering transmission electron microscopy.

The previous discovery of the Streptomyces lividans kcsA gene and its overexpression followed by the functional reconstitution of the purified gene product has resulted in new strategies to explore this channel protein in vitro. KcsA has evolved as a general model to investigate the structure/function relationship of ion channel proteins. Using specific antibodies raised against a domain of KcsA lacking membrane-spanning regions, KcsA has now been localized within numerous separated clusters between the outer face of the cytoplasm and the cell envelope in substrate hyphae of the S. lividans wild-type strain but not in a designed chromosomal disruption mutant DeltaK, lacking a functional kcsA gene. Previous findings had revealed that caesium ions led to a block of KcsA channel activity within S. lividans protoplasts fused to giant vesicles. As caesium can be scored by electron energy loss spectroscopy better than potassium, this technique was applied to hyphae that had been briefly exposed to caesium instead of potassium ions. Caesium was found preferentially at the cell envelope. Compared to the DeltaK mutant, the relative level of caesium was approximately 30 % enhanced in the wild-type. This is attributed to the presence of KcsA channels. Additional visualization by electron spectroscopic imaging supported this conclusion. The data presented are believed to represent the first demonstration of in vivo monitoring of KcsA in its original host.

Cesium↗

Monilethrix: an electron microscopic and electron histochemical study.

We have carried our electron microscopic and electron histochemical investigations on affected hair follicles and hair shafts to elucidate further the structural abnormality in monilethrix. The structural abnormalities manifest in the cortex and cuticle of the hair shaft could be traced to the locality of the 'zone of keratinization' of the follicle. Abnormal orientation of the cortical cell proteins, folding of the cuticle cell membrane and varying amounts of endocuticular material were all observed in the region prior to full keratinization. We observed structural abnormalities within all the component layers of the inner root sheath adjacent to the faults within the hair shaft, indicating a possible asynchrony within the monilethrix follicle.

Hair↗

Ultrastructural aluminium detection in amphibian tissues by electron spectroscopic imaging and electron energy-loss spectroscopy.

Aluminium causes a variety of toxic effects in living organisms but very little is known about its uptake, pathways and locations of deposition. We have applied electron spectroscopic imaging (ESI) and electron energy-loss spectroscopy (EELS) to locate aluminium at the ultrastructural level in amphibian larvae from acidic ponds. It is found diffusely bound or precipitated in cell organelles. The spatial resolution of aluminium detection is high. The elemental composition of small areas can be demonstrated by EELS. Three different fixation procedures give similar results. The two- and three-window methods at the K- and L-edges are compared.

Aluminum↗

Demonstration of calcium in dermal melanocytes of Xenopus laevis and Poecilia reticulata with electron energy-loss spectroscopy and electron spectroscopic imaging.

The subcellular distribution of calcium in dermal melanocytes of Xenopus laevis and Poecilia reticulata has been analysed. Using two cytochemical methods, phosphate precipitation and a combined oxalate-pyroantimonate technique, electron energy-loss spectroscopy and electron spectroscopic imaging have been applied for elemental analysis. Both precipitation techniques revealed a high calcium content in the melanosomes of both species. Calcium was also located in the vicinity of collagen fibrils and in the plasma membrane.

Animals↗

Transcriptional regulation of the proton translocating NADH dehydrogenase genes (nuoA-N) of Escherichia coli by electron acceptors, electron donors and gene regulators.

The promoter region and transcriptional regulation of the nuoA-N gene locus encoding the proton-translocating NADH:quinone oxidoreductase was analysed. A 560 bp intergenic region upstream of the nuo locus was followed by a gene (designated lrhA for LysR homologue A) coding for a gene regulator similar to those of the LysR family. Disruption of lrhA did not affect growth (respiratory or non-respiratory) or expression of nuo significantly. Transcriptional regulation of nuo by electron acceptors, electron donors and the transcriptional regulators ArcA, FNR, NarL and NarP, and by IHF (integration host factor) was studied with protein and operon fusions containing the promoter region up to base pair -277 ('nuo277') or up to base pair -89 ('nuo899'). The expression of the nuo277-lacZ fusions was subject to ArcA-mediated anaerobic repression and NarL(+ nitrate)-mediated anaerobic activation. FNR and IHF acted as weak repressors under anaerobic conditions. Expression of nuo899-lacZ was stimulated during anaerobic fumarate respiration and aerobically by C4 dicarboxylates. Therefore, expression of nuo is regulated by O2 and nitrate via ArcA, NarL, FNR and IHF at sites within the -277 region, and by other factors including C4 dicarboxylates at a site between -277 and -899. A physiological role for the transcriptional stimulation by O2 and nitrate is suggested.

Aerobiosis↗

Cytochrome c2 is essential for electron transfer to nitrous oxide reductase from physiological substrates in Rhodobacter capsulatus and can act as an electron donor to the reductase in vitro. Correlation with photoinhibition studies.

1. Addition of nitrous oxide to a periplasmic fraction released from Rhodobacter capsulatus strains MT1131, N22DNAR+ or AD2 caused oxidation of c-type cytochrome, as judged by the decrease in absorbance at 550 nm. The periplasmic fraction catalysed reduction of nitrous oxide in the presence of either isoascorbate plus phenazine ethosulphate or reduced methyl viologen. The rates with these two electron donors were similar and were comparable to the activity observed with a quantity of cells equivalent to those from which the periplasm sample had been derived. Activity in the periplasm could not be observed with ascorbate plus 2,3,5,6-tetramethyl-p-phenylenediamine although this reductant was effective with intact cells treated with myxothiazol to block the activity of the cytochrome-bc1 complex. 2. Cells of R. capsulatus MTG4/S4, a mutant from which the gene for cytochrome c2 has been specifically deleted, did not catalyse detectable rates of nitrous-oxide reduction. A nitrous-oxide reductase activity was present, as shown by activity of both cells and a periplasmic fraction with isoascorbate plus phenazine ethosulphate as reductant. The rates in cells and the periplasmic fraction were similar to those observed in the corresponding wild-type strain (MT1131). In contrast to wild-type cells, 2,3,5,6-tetramethyl-p-phenylenediamine and N,N,N',N'-tetramethyl-p-phenylenediamine [Ph(NMe2)2] were ineffective as mediators of electrons from isoascorbate. Visible absorption spectra showed that no detectable cytochromes in either the periplasm or intact cells of the MTG4/S4 mutant were oxidised by nitrous oxide. 3. Purified ferroycytochrome c2 from R. capsulatus was oxidised by nitrous oxide in the presence of periplasm from R. capsulatus MTG4/S4. The rate of oxidation was proportional to the amount of periplasm added, but was considerably lower than the rate of nitrous-oxide reduction observed with the same periplasmic fraction when either ascorbate plus phenazine ethosulphate or reduced methyl viologen were used as substrates. The oxidation of cytochrome c2 was inhibited by acetylene and by low concentrations of NaCl. 4. Oxidation of ferrocytochrome c2 by nitrous oxide was observed when the purified cytochrome was mixed with a preparation of nitrous-oxide reductase. However, oxidation of ferrocytochrome c' by nitrous oxide was not observed in the presence of the reductase. The observations with the mutant MTG4/S4 suggest that cytochrome c2 is the only periplasmic cytochrome involved in nitrous-oxide reduction. 5. Nitrous-oxide-dependent oxidation of a c-type cytochrome was observed in a periplasmic fraction from Paracoccus denitrificans, provided the fraction was first reduced.(ABSTRACT TRUNCATED AT 400 WORDS)

Cytochrome c Group↗

Chemical mechanism of the Gram stain and synthesis of a new electron-opaque marker for electron microscopy which replaces the iodine mordant of the stain.

Crystal violet (hexamethyl-para-rosaniline chloride) interacts with aqueous KI-I2 during the Gram stain via a simple metathetical anion exchange to produce a chemical precipitate. There is an apparent 1:1 stoichiometry between anion (I-) and cation (hexamethyl-para-rosaniline+) during the reaction and, since the small chloride anion is replaced by the bulkier iodide, the complex formed becomes insoluble in water. It is this same precipitate which forms in the cellular substance of bacteria (both gram-positive and gram-negative types) and which initiates the Gram reaction. Potassium trichloro(eta 2-ethylene)-platinum(II), as an electronopaque marker for electron microscopy, was chemically synthesized, and it produced an anion in aqueous solution which was compatible with crystal violet for the Gram stain. It interacted with crystal violet in a similar manner as iodide to produce an insoluble complex which was chemically and physically analogous to the dye-iodide precipitate. This platinum anion therefore allows the Gram staining mechanism to be followed by electron microscopy.

Bacillus subtilis↗

Electron transfer flavoprotein from Methylophilus methylotrophus: properties, comparison with other electron transfer flavoproteins, and regulation of expression by carbon source.

When grown on methylated amines as a carbon source, Methylophilus methylotrophus synthesizes an electron transfer flavoprotein (ETF) which is the natural electron acceptor of trimethylamine dehydrogenase. It is composed of two dissimilar subunits of 38,000 and 42,000 daltons and 1 mol of flavin adenine dinucleotide. It was reduced by trimethylamine dehydrogenase to a stable anionic semiquinone form, which could not be converted, either enzymatically or chemically, to the fully reduced dihydroquinone. This ETF exhibited spectral properties which were nearly identical to ETFs from bacterium W3A1, Paracoccus denitrificans, and pig liver mitochondria. M. methylotrophus ETF cross-reacted immunologically and enzymatically with the ETF of bacterium W3A1 but not with the other two ETFs. In M. methylotrophus and bacterium W3A1, ETF and trimethylamine dehydrogenase were each expressed during growth on trimethylamine and were each absent during growth on methanol.

Animals↗

Comparison of direct electron microscopy, immune electron microscopy, and rotavirus enzyme-linked immunosorbent assay for detection of gastroenteritis viruses in children.

An approximate 10% suspension in water of the first available stool sample from 411 infants and young children with acute gastroenteritis was examined by electron microscopy (EM) after 2 min of negative staining. This procedure enabled the detection of 88% of the 199 rotavirus infections, all of the 22 adenovirus infections, and 47% of the 15 approximately 27-nm virus infections ultimately detected by a combination of techniques, including immune electron microscopy (IEM) and rotavirus enzyme-linked immunosorbent assay (ELISA). Of the 204 infections detected by direct EM of stools, 76% were detected within 2 min of viewing, and 94% were detected within 6 min of viewing. Type 1 and type 2 rotavirus particles were visualized with approximately equal efficiency, although type 2 rotavirus infections were more common. Rectal swab preparations were clearly inferior to stool preparations for the detection of virus infection by direct EM. IEM examination was required for efficient visualization of viruses in rectal swab specimens. ELISA was the most sensitive method for the detection of rotaviruses; with this method, all infections in which rotavirus particles were visualized by EM or IEM were detected. However, 73% of the 1,834 specimens which were presumptively positive for rotavirus by conventional indirect ELISA proved to be falsely positive on the basis of EM, IEM, blocking ELISA, confirmatory ELISA, or a combination of these methods. False-positive rotavirus ELISA reactions apparently were eliminated when fecal specimens were tested in a modified confirmatory ELISA with a lower dilution of rotavirus-negative (pre-immunization) than rotavirus-positive (post-immunization) capture antibody from the same animal.

Acute Disease↗

Choroidal vasculature changes in spontaneously hypertensive rats - transmission electron microscopy and scanning electron microscopy with casts.

PURPOSE: The purpose of the present study was to elucidate the specific morphological changes of the choroidal vasculature in long-term hypertension. METHODS: The choroidal vessels of spontaneously hypertensive (SHR) rats were examined with corrosion casts/scanning electron microscope (SEM) and the transmission electron microscope (TEM). RESULTS: In 18-month-old SHRs, corrosion casts and SEM showed tortuosity, caliber irregularity and generalized narrowing of the choroidal arteries. The draining venules were prominently decreased in number. The ampullae of the vortex veins were narrow, but arteriovenous anastomoses, crossing defects and obstruction were not seen. The choriocapillaris was elongated and engorged. TEM of the choroidal vessels showed hypertrophy of smooth muscle cells and irregular thickening of the basement membrane and narrow lumen of the choriocapillaris. The RPE and Bruch's membrane were intact. CONCLUSION: The choroidal vascular bed showed extensive morphological changes. This study revealed that hypertensive choroidopathy has specific features such as tortuosity, caliber irregularity, generalized narrowing of arteries, decreased number of draining venules and elongated and engorged choriocapillaris. These findings indicate that hypertensive choroidopathy is as important as retinopathy.

Animals↗

Simultaneous visualization of myeloperoxidase reactivity and immunogold labeling by backscattered electron imaging with the scanning electron microscope.

We describe a method to label circulating human granulocytes with an immunogold marker and then incubate them for demonstration of myeloperoxidase activity. The cells were observed in the backscattered electron imaging (BEI) mode of the scanning electron microscope. This permits simultaneous visualization of cell surface morphology, the immunological surface marker, and the cytochemical reactivity of each individual cell. Enhanced identification of the various cell types and more precise characterization of cell surface features in the different steps of leukocyte differentiation are expected to result from application of this technique.

Gold↗

Electronic structure of brain: structure-activity relationships between electronic structure and neurotransmitters based on molecular hardness concept.

In order to understand the relation between the electronic structure of neurotransmitters and the brain, a model of the brain based on absolute hardness (eta) and absolute electronegativity (chi) is described. It was found that the coordinate r(chi, eta) of electronic structures of neurotransmitters obtained using the parameters eta and chi can be graphically classified into three groups: catecholamine type (group I), gamma-aminobutanoic acid (GABA) type (group II), and acetylcholine (ACh) type (group III) in the eta-chi diagram. The results suggest that the brainstem and neocortex in the brain are chemically soft and hard, respectively, because they show that the myelinated nerve is chemically soft and the unmyelinated nerve is chemically hard. If one calculates the r(chi, eta) to understand which group a drug belongs to, one can predict the target receptors of the drug from the eta-chi diagram. Using eta-chi maps, one is then able to design medications like antidepressants, tranquilizers, and ACh agonists.

Brain↗

Improved CNDO/S calculation of electronic spectra of organic compounds. I. New CNDO/S calculation by using an improved method of one-center electron repulsion integral.

The NM-gamma CNDO/S program previously developed by our group was modified by the introduction of a new one-center electron repulsion integral gamma(AA)(new) approximation, namely, the gamma(AA)(new)-CNDO/S method. The value of this gamma(AA)(new) was evaluated according to the product values of the coefficient C with the gamma(AA) value proposed in our previous paper. This method using a new gamma(AA) was also found to improve the two-center electron repulsion integral gamma(AB) value with respect to the chemical softness proposed by Nishimoto and co-workers, together with the difference between HOMO and LUMO orbital energies. The results calculated by the present improved gamma(AA)(new)-CNDO/S method demonstrated that not only the calculated absorption maxima wavelengths and ionization potentials, but also the order and the assignment of orbitals coincided very well with those based on the results of experiments investigating a variety of polyenes, cyanynes, and polycyclic aromatic hydrocarbons.

Algorithms↗

Calcium measurements with electron probe X-ray and electron energy loss analysis.

This paper presents a broad survey of the rationale for electron probe X-ray microanalysis (EPXMA) and the various methods for obtaining qualitative and quantitative information on the distribution and amount of elements, particularly calcium, in cryopreserved cells and tissues. Essential in an introductory consideration of microanalysis in biological cryosections is the physical basis for the instrumentation, fundamentals of X-ray spectrometry, and various analytical modes such as static probing and X-ray imaging. Some common artifacts are beam damage and contamination. Inherent pitfalls of energy dispersive X-ray systems include Si escape peaks, doublets, background, and detector calibration shifts. Quantitative calcium analysis of thin cryosections is carried out in real time using a multiple least squares fitting program on filtered X-ray spectra and normalizing the calcium peak to a portion of the continuum. Recent work includes the development of an X-ray imaging system where quantitative data can be retrieved off-line. The minimum detectable concentration of calcium in biological cryosections is approximately 300 mumole kg dry weight with a spatial resolution of approximately 100 A. The application of electron energy loss (EELS) techniques to the detection of calcium offers the potential for greater sensitivity and spatial resolution in measurement and imaging. Determination of mass thickness with EELS can facilitate accurate calculation of wet weight concentrations from frozen hydrated and freeze-dried specimens. Calcium has multiple effects on cell metabolism, membrane transport and permeability and, thus, on overall cell physiology or pathophysiology. Cells can be rapidly frozen for EPXMA during basal or altered functional conditions to delineate the location and amount of calcium within cells and the changes in location and concentration of cations or anions accompanying calcium redistribution. Recent experiments in our laboratory document that EPXMA in combination with other biochemical and electrophysiological techniques can be used to study, for example, sodium and calcium compartmentation in cultured cardiac cells. Such analyses can also be used to clarify the role of calcium in anoxic renal cell injury and to evaluate proposed ionic defects in cells of individuals with cystic fibrosis.

Animals↗

Osteoblast activity at the dental implant-bone interface: transmission electron microscopic and high voltage electron microscopic observations.

The purpose of this report is to present transmission electron microscopic and high voltage transmission electron microscopic (HVEM) observations of a longitudinal investigation examining the activities of osteoblasts and associated tissues apposing titanium and alumina oxide ceramic endosteal dental implants. The HVEM permitted 3-dimensional stereologic observations. All observations were obtained from undecalcified interfacial tissues from this in vivo experimental dog model using commercially available implants placed into the mandible. Two similar implants were placed in both sides of the mandible, with implants in 12 of the 18 dogs supporting fixed bridges for either 6 or 12 months. From the study, we observed that a mineralized matrix exists in direct apposition to the implant. Since bone does not interface the entire length of the implant, other interfacial zones were found to exist which consisted of unmineralized tissues. In such zones, we observed that osteoblasts were routinely found directly at the implant interface to the mandibular bone. These interfacial tissues included unmineralized collagen fibers, proteinaceous material, a finely fibrillar matrix, and the osteoblasts. This study has reinforced the concept that the oral tissue-dental implant interface is a dynamic zone consisting of remodeling activities of the osseous cells and extracellular matrices.

Alveolar Process↗

High resolution electron backscatter diffraction with a field emission gun scanning electron microscope.

A scanning electron microscope with a thermal field emission gun (FEGSEM) is found to offer significant improvements in electron backscatter diffraction performance over a conventional W-filament scanning microscope. The spatial resolution is improved by a factor of approximately 3 in the FEGSEM and is optimized at probe currents of 50-300 nA and at 10-15 keV. The angular accuracy is optimized at probe currents above approximately 150 nA and at 30 keV.

Microscopy, Electron, Scanning↗