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PubMed1998

Image processing based on the combination of high-resolution electron microscopy and electron diffraction.

A method of crystal structure determination by electron crystallographic image processing based on the combination of high-resolution electron microscopy (HREM) and electron diffraction is introduced. It consists of two stages: image deconvolution and resolution enhancement. In the first stage an image taken at an arbitrary defocus condition is transformed into the structure image with the resolution depending on the resolution of the electron microscope. In the second stage the image resolution is enhanced to the diffraction resolution limit by combining the electron diffraction data and using the phase extension technique so that in the final image most unoverlapped atoms can be resolved individually. The experimental diffraction intensities are corrected for approximating to square structure factors. The principle of the image processing and the procedure of diffraction intensity correction are briefly described and the results of applications are illustrated. Since the method is based on the weak phase object approximation (WPOA), the validity of WPOA is discussed by introducing an approximate image contrast theory named pseudo weak phase object approximation (PWPOA) to demonstrate the image contrast change with the crystal thickness for very thin crystals.

Copper↗

PubMed1998

Kinetic modeling of omega-transamination for enzymatic kinetic resolution of alpha-methylbenzylamine.

A kinetic model for omega-transaminase from Bacillus thuringiensis JS64 was developed by using the King-Altman method to simulate the kinetic resolution of alpha-methylbenzylamine (alpha-MBA). Starting from a ping-pong bi-bi mechanism, a complete kinetic model including substrate inhibition only in the reverse reaction (i.e., transamination between acetophenone and L-alanine) was developed. The asymmetric synthesis of (S)-alpha-MBA proved to be difficult due to a much lower maximum reverse reaction rate than the maximum forward reaction rate, thermodynamically exergonic forward reaction (i.e., transamination between (S)-alpha-MBA and pyruvate), and the severe product and substrate inhibition of the reverse reaction. Experimental values for kinetic parameters show that the product inhibition constant of (S)-alpha-MBA is the most important parameter on determining the resolution reaction rate, suggesting that the resolution reaction rate will be very low unless (S)-alpha-MBA strongly inhibits the reverse reaction. Using the kinetic model, the kinetic resolution of alpha-MBA in aqueous buffer was simulated, and the simulation results showed a high degree of consistency with experimental data over a range of reaction conditions. Various simulation results suggest that the crucial bottleneck in the kinetic resolution of alpha-MBA lies mainly in the accumulation of acetophenone in reaction media as the reaction proceeds, whereas L-alanine exerts a little inhibitory effect on the reaction. The model predicts that removing acetophenone produced during the reaction can enhance the reaction rate dramatically. Indeed, the biphasic reaction system is capable of extracting acetophenone from the aqueous phase, showing a much higher reaction rate compared to a monophasic reaction system. The kinetic model was also useful in predicting the properties of other, better enzymes as well as the optimal concentrations of amino acceptor and enzyme in the resolution reaction.

Acetophenones↗

PubMed1998

The influence of the resolution of reciprocal ST segment changes on in-hospital complications of acute myocardial infarction after percutaneous coronary angioplasty.

We studied 120 patients (M:F 105:15, mean age 57.5 +/- 10.1 years) with acute myocardial infarction (MI) successfully treated with percutaneous coronary angioplasty (PTCA) to analyze the influence of the resolution of the ST segment elevation and depression after intervention to 1 month composite endpoints of reinfarction or reocclusion, development of congestive heart failure (CF) and death. Sum of preintervention and postintervention ST segment elevation and depression and the rate of resolution of these ST segment elevations and depressions were recorded for every patient. A total of 17 (14.2%) composite endpoint events (events group) were recorded (7 reocclusion or recurrent MI, 9 CF, and 1 death). On univariate analysis, events group patients were older (53.3 +/- 9.9 vs. 58.8 +/- 9.1 years, P = 0.032), had lesser resolution of ST segment elevations (85 +/- 24% vs. 44 +/- 55%, P = 0.017) and depression (72 +/- 26% vs. 52 +/- 30%, P = 0.009), had greater preintervention ST segment elevation (17.49 +/- 12.95 mm vs. 28.38 +/- 20.41 mm, P = 0.045), had lower ejection fraction (59.3 +/- 10.2% vs. 43.6 +/- 9.4%, P < 0.001), and had more frequent multivessel disease (71% vs. 47%, P = 0.048) compared to the nonevents group. Time from angina to reperfusion, residual stenosis, sex, infarct location and infarct-related vessel distribution were similar. On multivariate analysis (logistic regression with backward likelihood ratio) only older age (P = 0.0752), lesser rate of resolution of ST segment depression (P = 0.0262) and lower ejection fractions (P = 0.0014) were retained as predictors of the composite endpoints. Relative risk conferred by less than 50% resolution of ST segment depressions for composite endpoints were 3.78 (95% CI 1.63-8.73). We conclude that the lack of resolution of the sum of reciprocal ST segment depressions identifies a subgroup of acute MI patients with greater morbidity after primary PTCA.

Angioplasty, Balloon, Coronary↗

PubMedDate not supplied

High-resolution echo-planar fMRI of human visual cortex at 3.0 tesla.

Known specialized properties of the human visual cortex have been used to investigate the role of spatial resolution on fMRI using blood oxygenation level dependent (BOLD) echo-planar MRI at 3.0 tesla. The magnitude of BOLD signal changes has been examined at low (3.1 x 3.1 x 3.0 mm3) and high (0.8 x 1.6 x 3.0 mm3) resolution using both gradient-echo and spin-echo EPI. Paradigms were designed to activate primary visual cortex (V1/V2) and more specialized areas associated with detection of color (V4) and motion (V5). Sensitivity of activation maps increased at higher resolution despite the decreased total signal intensity at the smaller voxel size, presumably from reduced partial volume averaging. The greater microvascular selectivity of high-resolution spin-echo imaging enabled distinct activation patterns sensitive to motion to be detected in V1/V2 that were not apparent with gradient-echo imaging. The spatial resolution at 3.0 tesla was constrained by the size of physiological head motion relative to the voxel dimensions rather than SNR or the hemodynamic response of BOLD contrast. The higher spatial resolution at 3.0 tesla with more selective spin-echo EPI can further refine functional mapping within the cerebral cortex.

Brain Mapping↗

PubMed2000

Correlation of high-resolution breast MR imaging with histopathology; validation of a technique.

A high-resolution three-dimensional surface gradient coil set was used to obtain magnetic resonance (MR) images of breast specimens, using a gradient-echo pulse sequence (TR/TE 1000/8 msec, flip angle 75 degrees), with 117 micrometer in-plane resolution and 1 mm slice thickness. Breast tissues were obtained from one autopsy and three surgical specimens. High-resolution breast MR images and histopathology sections (7 micrometer thickness) were acquired in the same anatomical plane. Radiographs were acquired of the sliced specimens (approximately 5 mm thick) so that images from all three methods could be correlated. It was found that in vitro high-resolution breast MRI correlated well with low-resolution microscopic histology, demonstrating normal anatomy (lobules, ducts, connective tissue strands, blood vessels) and pathology (tumor content, margins, and presence of microcalcifications) of the breast more clearly than conventional pre-gadolinium breast MRI. High-resolution breast MRI may improve specificity, when added to a conventional breast MRI protocol.

Biopsy↗

PubMed2002

Liquid chromatographic resolution of N-acyl-alpha-amino acids as their anilide derivatives on a chiral stationary phase based on (S)-leucine.

A chiral stationary phase (CSP 1) derived from N-(3,5-dinitrobenzoyl)leucine N-phenyl N-alkylamide was used for the liquid chromatographic resolution of anilide derivatives of N-acyl-alpha-amino acids and the chromatographic resolution results were compared with those from four other commercial CSPs. The chromatographic resolution results showed that CSP 1 was most effective among five CSPs used in this study. The chiral recognition mechanism exerted by CSP 1 for the resolution of anilide derivatives of N-acyl-alpha-amino acids is proposed to involve a face-to-face pi-pi interaction and two hydrogen bonding interactions between the CSP and the analytes from the chromatographic resolution behaviors of slightly modified anilide derivatives of N-acyl-alpha-amino acids. The chiral recognition mechanism proposed is quite similar to that advanced previously for the resolution of N-(3,5-methoxybenzoyl)-alpha-amino acids on CSP 1, even though the interaction sites of the two types of analytes were totally different from each other. The apparent similarity of the two chiral recognition mechanisms was assumed to stem from the identical interaction modes of the two types of analytes with the CSP. In addition, the dependence of the enantioselectivity of anilide derivatives of N-acyl-alpha-amino acids on the length of the alkyl tail of the N-acyl group of analytes was rationalized to stem from the intercalation of the N-acyl group of the (R)-enantiomer of analytes between the tethers of the CSP.

Journal Article↗

PubMed2001

Solvent-free optical resolution of N-methylamphetamine by distillation after partial diastereoisomeric salt formation.

Solvent-free optical resolution of N-methylamphetamine was developed by distillation after partial diastereoisomeric salt formation. From the 18 chiral acids tested by this method, five provide by this method resolution: O,O'-dibenzoyltartaric acid, O,O'-di-p-toluoyltartaric acid, 6-methoxy-alpha-methyl-2-naphthaleneacetic acid (Naproxen), the cis-permetrinic acid, and the 2-phenoxypropionic acid. Among them the O,O'-dibenzoyltartaric acid in water-free form provided the more effective resolution. The efficiency of this resolution S = 0.74 is in the range of the industrial-scale resolutions and not worse than the efficiency achieved by optical resolution via fractional crystallization.

Journal Article↗

PubMed2001

Theoretical versus experimental resolution in optical microscopy.

The aim of this article is to compare experimental resolution under different conditions with theoretical resolution predicted using electromagnetic diffraction theory. Imaging properties of fluorescent beads of three different diameters (0.1 microm, 0.2 microm, and 0.5 microm) as well as imaging properties of four different fluorescence-stained DNA targets (ABL gene, BCR gene, centromere 6, and centromere 17) are studied. It is shown how the dependence of the resolution on object size varies with wavelength (520 nm versus 580 nm), type of microscopy (wide-field, confocal using Nipkow disk, confocal laser scanning) and basic image processing steps (median and gaussian filters). Furthermore, specimen influence on the resolution was studied (the influence of embedding medium, coverglass thickness, and depth below the coverglass). Both lateral and axial resolutions are presented. The results clearly show that real objects are far from being points and that experimental resolution is often much worse than the theoretical one. Although the article concentrates on fluorescence imaging using high NA objectives, similar dependence can also be expected for other optical arrangements.

DNA↗

PubMed2003

The impact of spatial resolution and respiratory motion on MR imaging of atherosclerotic plaque.

PURPOSE: To examine the impact of spatial resolution and respiratory motion on the ability to accurately measure atherosclerotic plaque burden and to visually identify atherosclerotic plaque composition. MATERIALS AND METHODS: Numerical simulations of the Bloch equations and vessel wall phantom studies were performed for different spatial resolutions by incrementally increasing the field of view. In addition, respiratory motion was simulated based on a measured physiologic breathing pattern. RESULTS: While a spatial resolution of > or = 6 pixels across the wall does not result in significant errors, a resolution of < or = 4 pixels across the wall leads to an overestimation of > 20%. Using a double-inversion T2-weighted turbo spin echo sequence, a resolution of 1 pixel across equally thick tissue layers (fibrous cap, lipid, smooth muscle) and a respiratory motion correction precision (gating window) of three times the thickness of the tissue layer allow for characterization of the different coronary wall components. CONCLUSIONS: We found that measurements in low-resolution black blood images tend to overestimate vessel wall area and underestimate lumen area.

Arteriosclerosis↗

PubMed2001

Critical role of spatial resolution in dynamic contrast-enhanced breast MRI.

The spatial resolution of three-dimensional (3D) gradient-echo T1-weighted images, from 40 women with 25 malignant and 23 benign lesions, was purposely degraded to determine the role of spatial resolution in recording, analysis, and diagnosis of dynamic contrast-enhanced breast MRI. Images were recorded and analyzed at pixel resolution according to the 3TP method (Degani et al., Nat Med 1997;3:780-782). Reduction in spatial resolution degraded the appearance of foci with fast wash-in and fast washout dynamics. This resulted in an increase in false-negative diagnoses. The sensitivity for differentiating between malignant and benign lesions, using threshold criteria defined by the 3TP analysis, of 76% decreased to 60% and 24% for a 2- and 4-fold reduction in spatial resolution, respectively, without affecting significantly the high specificity (96-100%). In order to minimize false-negative diagnoses of contrast-enhanced breast MRI and maintain high specificity, it is essential to record and analyze the dynamic behavior at high spatial resolution. J. Magn. Reson. Imaging 2001;13:862-867.

Biopsy↗

PubMed2003

High-resolution 3D proton spectroscopic imaging of the human brain at 3 T: SNR issues and application for anatomy-matched voxel sizes.

In a systematic study on the interdependence of linewidth, signal-to-noise ratio (SNR), and spatial resolution in 3D proton spectroscopic imaging ((1)H-SI) at 3 T, we demonstrate reduced linewidths with increased spatial resolution due to reduced magnetic inhomogeneity within the brain. High-precision quantitative data (0.75-0.094 cm(3)) were obtained for all resolutions, enabling the creation of metabolic maps that display details such as the ventricles, sulci, and gyri. High-resolution (1)H-SI allows differences in metabolic ratios to be estimated for anatomically defined regions in gray (GM) and white matter (WM). Seven distinct regions in a healthy brain were anatomically segmented and their metabolic ratios were compared quantitatively. Data from a tumor patient are also presented to demonstrate potential clinical applications. Because of the high resolution, the metabolite ratios could be determined for distinct pathologic regions within the tumor and its surroundings. The method was additionally applied to a patient with patchy Pelizaeus Merzbacher disease (PMD), and compared to single-voxel spectroscopy performed in the same session. High-resolution SI data were demonstrated in our study to allow the direct matching of anatomic and metabolic images. This may enhance the clinical value of (1)H-SI.

Adult↗

PubMed2003

SNR improvement for multiinjection time-resolved high-resolution CE-MRA of the peripheral vasculature.

Peripheral MR angiography (MRA) should ideally provide images over a large field of view with high spatial resolution and adequate temporal resolution to accommodate differences in regional filling times. Image subtraction is usually used to remove background signals. In examination protocols involving multiple injections at multiple sites, previously injected contrast present in the mask image provides a substantial decrease in the subtraction image signal. Bolus chase methods avoid this problem but provide limited time for acquisition of high-resolution images at each station. We present here a technique applied to peripheral angiography that provides high spatial and temporal resolution while maintaining high SNR in multiple injection examinations. Undersampled projection imaging was used to increase spatial resolution relative to a previously reported technique using a Cartesian acquisition technique. Late acquisition of high spatial frequencies and temporal matched-filtering were used to increase spatial resolution and SNR, respectively. Temporal correlation analysis was applied to permit multistation examinations without mask subtraction, thus providing an additional gain in SNR relative to multistation subtraction methods. Quantitative analysis is provided to evaluate the signal and noise behavior in the matched-filtering process due to multiinjection and mask subtraction.

Computer Simulation↗

PubMed1993

Resolution of synthetic chi structures by the FLP site-specific recombinase.

The FLP site-specific recombinase is encoded by the two micron circle, an endogenous plasmid of Saccharomyces cerevisiae. FLP-mediated recombination in vitro proceeds via a short-lived Holliday (chi) intermediate. We have made a synthetic chi structure containing two FLP recognition target (FRT) sequences in order to investigate resolution by purified FLP protein. We found that incubation of this model substrate with FLP generated two pairs of linear products in equal quantities. Thus, resolution was equally likely to occur in either direction. Alteration of FLP binding sites, so as to inhibit binding, affected the direction of resolution; cleavage was reduced adjacent to the altered binding site. The overall efficiency of resolution increased when one FLP binding site was mutated. In investigating the series of mutated chi structures we found that resolution requires only two intact FLP binding sites. However, the non-specific protein-DNA interaction of additional FLP molecules may also be required. Thus, resolution is more tolerant of the loss of FLP binding sites than is the complete recombination reaction.

Base Sequence↗

PubMed1999

Shope fibroma virus DNA topoisomerase catalyses holliday junction resolution and hairpin formation in vitro.

The telomeres of poxviral chromosomes comprise covalently closed hairpin structures bearing mismatched bases. These hairpins are formed as concatemeric replication intermediates and are processed into mature, unit-length genomes. The structural transitions and enzymes involved in telomere resolution are poorly understood. Here we show that the type I topoisomerase of Shope fibroma virus (SFV) can promote a recombination reaction which converts cloned SFV replication intermediates into hairpin-ended molecules resembling mature poxviral telomeres. Recombinant SFV topoisomerase linearised a palindromic plasmid bearing 1.5 kb of DNA encoding the SFV concatemer junction, at a site near the centre of inverted-repeat symmetry. Most of these linear reaction products bore hairpin tips as judged by denaturing gel electrophoresis. The resolution reaction required palindromic SFV DNA sequences and was inhibited by compounds which block branch migration (MgCl2) or poxviral topoisomerases. The resolution reaction was also slow, needed substantial quantities of topoisomerase, and required that the palindrome be extruded in a cruciform configuration. DNA cleavage experiments identified a pair of suitably oriented topoisomerase recognition sites, 90 bases from the centre of the cloned SFV terminal inverted repeat, which may mark the resolution site. These data suggest a resolution scheme in which branch migration of a Holliday junction through a site occupied by covalently bound topoisomerase molecules, could lead to telomere resolution.

DNA Topoisomerases, Type I↗

PubMedDate not supplied

Low-resolution density maps from atomic models: how stepping "back" can be a step "forward".

Atomic-resolution structures have had a tremendous impact on modern biological science. Much useful information also has been gleaned by merging and correlating atomic-resolution structural details with lower-resolution (15-40 A), three-dimensional (3D) reconstructions computed from images recorded with cryo-transmission electron microscopy (cryoTEM) procedures. One way to merge these structures involves reducing the resolution of an atomic model to a level comparable to a cryoTEM reconstruction. A low-resolution density map can be derived from an atomic-resolution structure by retrieving a set of atomic coordinates editing the coordinate file, computing structure factors from the model coordinates, and computing the inverse Fourier transform of the structure factors. This method is a useful tool for structural studies primarily in combination with 3D cryoTEM reconstructions. It has been used to assess the quality of 3D reconstructions, to determine corrections for the phase-contrast transfer function of the transmission electron microscope, to calibrate the dimensions and handedness of 3D reconstructions, to produce difference maps, to model features in macromolecules or macromolecular complexes, and to generate models to initiate model-based determination of particle orientation and origin parameters for 3D reconstruction.

Algorithms↗

PubMed2000

Low- and high-resolution mapping of DNA damage at specific sites.

Measurement of DNA damage and repair at the nucleotide level in intact cells has provided compelling evidence for the molecular details of these events as they occur in intact organisms. Furthermore, these measurements give the most accurate picture of the rates of repair in different structural domains of DNA in chromatin. In this report, we describe two methods currently used in our laboratories to map DNA lesions at (or near) nucleotide resolution in yeast cells. The low-resolution method couples damage-specific strand breaks in DNA with indirect end-labeling to measure DNA lesions over a span of 1.5 to 2 kb of DNA sequence. The resolution of this method is limited by the resolution of DNA length measurements on alkaline agarose gels (about +/-20 bp on average). The high-resolution method uses streptavidin magnetic beads and special biotinylated oligonucleotides to facilitate end-labeling of DNA fragments specifically cleaved at damage sites. The latter method maps DNA damage sites at nucleotide resolution over a shorter distance (<500 bp), and is constrained to the length of DNA resolvable on DNA sequencing gels. These methods are used in tandem for answering questions regarding DNA damage and repair in different chromatin domains and states of gene expression.

Biotinylation↗

PubMed1993

The influence of image resolution on the positron emission tomographic measurement of caudate glucose consumption.

The aim of this study was to investigate the influence of image resolution on (a) relative and absolute values of caudate glucose consumption (rCMRGlc) determined by positron emission tomography (PET), and (b) the detection of significant differences in these metabolic values between groups of subjects. For this purpose, raw data of cerebral accumulation of fluorine-18 fluorodeoxyglucose (FDG) obtained in 11 normal subjects and in nine patients with unilateral thalamic infarction were reconstructed using filtered backprojection with four different cut-off frequencies (CFs), yielding images with a transaxial resolution of 5.7, 7.1, 8.9 and 11 mm (full-width at half-maximum; FWHM). Absolute values of caudate rCMRGlc decreased significantly by more than 30% over the range of image resolutions studied. Bilateral ratios of caudate rCMRGlc were insensitive to variations in image resolution. Levels of significance assessing the differences in mean metabolic values between patients and controls were all below 0.01. They were, however, slightly better at image resolutions of 7.1 and 8.9 mm than at a resolution of 5.7 mm. These data indicate (a) that relative values of rCMRGlc are better suited to compare quantitative results from different PET cameras than are absolute values, and (b) that the CF used for the filtered back-projection exerts a small but not negligible influence on levels of significance assessing differences in metabolic values between groups of subjects.

Adult↗

PubMed1995

Influence of transcription and replication on the in situ resolution of immunoglobulin heavy-chain constant region genes: an interphase cytogenetics analysis.

An interphase cytogenetics analysis was performed to investigate whether replication and transcription could influence in situ resolution of immunoglobulin (Ig) heavy chain constant region genes. A plasmid probe recognizing five C gamma segments separated by known linear DNA distances was hybridized in situ and visualized by digital fluorescence microscopy. In interphase nuclei from phytohemagglutinin (PHA)-stimulated lymphocytes, the gamma genes were resolved as one to three signals per allele in the majority of nuclei, whereas in a minority, complex patterns of several signals per allele could be observed. The latter were restricted to nuclei in an early stage of the S phase, as assessed by hybridization experiments performed in cells grown in the presence of bromodeoxyuridine. To investigate whether the in situ resolution of the C gamma segments could vary as a function of the transcription activity of the locus, the C gamma probe was subsequently hybridized to nuclei from a mature B cell line (JVM-2), which produces gamma transcripts as shown by in situ RNA hybridization experiments. Primary human fibroblasts were further used as representative of a non-lymphoid cell type with transcriptionally inactive Ig genes. When Gl nuclei from the three cell types were compared in terms of the in situ resolution of the C gamma locus, JVM-2 cells were found to include the highest percentage of higher resolution patterns (three to five signals per allele in 28% of nuclei), fibroblasts the lowest (three signals per allele, 2%), while PHA-stimulated lymphocytes occupied an intermediate position between the other two cell types (three or four signals per allele, 15%). The data show that the in situ resolution of Ig C gamma genes varies throughout the cell cycle and is influenced by the transcriptional activity of the locus. The variability of the resolution patterns observed appears to reflect different levels of chromatin packaging, which in turn are likely to influence the probe accessibility to its target. These observations are relevant for the interpretation of data from interphase cytogenetics analysis of independent, but closely spaced, DNA segments.

Cells, Cultured↗