Nucleic acid fragmentation on the millisecond timescale using a conventional X-ray rotating anode source: application to protein–DNA footprinting

Nucleic acid fragmentation (footprinting) by ·OH radicals is used often as a tool to probe nucleic acid structure and nucleic acid–protein interactions. This method has proven valuable because it provides structural information with single base pair resolution. Recent developments in the field introduced the ‘synchrotron X-ray footprinting’ method, which uses a high-flux X-ray source to produce single base pair fragmentation of nucleic acid in tens of milliseconds. We developed a complementary method that utilizes X-rays generated from a conventional rotating anode machine in which nucleic acid footprints can be generated by X-ray exposures as short as 100–300 ms. Our theoretical and experimental studies indicate that efficient cleavage of nucleic acids by X-rays depends upon sample preparation, energy of the X-ray source and the beam intensity. In addition, using this experimental set up, we demonstrated the feasibility of conducting X-ray footprinting to produce protein–DNA protection portraits at sub-second timescales.     

Precise Cobb Angle Measurement System Based on Spinal Images Merging Function

Background

Convenient and precise measurement of the Cobb angle using a small size X-ray detector has been required for local clinics.

A field method for the cephalometric x-ray study of skulls in early Nubian cemeteries

The cephalometer, a clinical and research instrument employed by the orthodontic profession to record standardized x-rays of the human skull, has been modified for research work in remote areas where the usual laboratory facilities are not available. This cephalometer was specifically designed to record cephalograms (head radiographs) on the living Nubian population and their extensive skeletal record. The cephalometer consists of a light, portable tripod stand and an aluminum beam to which is attached at one end a Wehmer cephalostat (head holder), and on the other end an isotope radiation source. Hence, the radiation source and the cephalostat were held in the standard relationship of five feet from source to the mid-saggittal plane of the cranium. Ytterbium-169, with a half life of 32.5 days, was used to expose the film. The isotope and the use of the new Experimental Polaroid X-ray Film permitted the x-ray examination of Nubian skulls with no conventional power source or darkroom facilities.     

Evidence for the absence of photoreduction of the metal centers of cytochrome C oxidase by X-irradiation

Samples of X-irradiated cytochrome c oxidase were examined by electron paramagnetic resonance and optical spectroscopy. Both radiation from the Stanford Synchrotron Radiation Laboratory and a conventional X-ray source (W target) were utilized. The X-ray flux from these sources ranges from 10(9) to 10(13) photons/s. No evidence was found for photoreduction of the metal centers in the enzyme by X-ray photons. These results demonstrate that the integrity of cytochrome c oxidase is maintained using the conditions under which X-ray absorption measurements are presently being made.     

Dynamics of accelerated ions in coronal loops and model of a gamma-ray source

The dynamics of ions accelerated to energies of 10–100 MeV/nucleon in an electric field of ～0.01–0.1 V/cm, which has a component directed along the magnetic field of solar coronal loops with a characteristic size of ～100000 km, is considered. The motion of fast ions trapped in a current-carrying magnetic loop that has a magnetic mirror at its base (the mirror trap model) is analyzed. The applicability of the obtained theoretical results to interpret gamma-ray bursts that, according to the data of the RHESSI space observatory, occurred on July 23, 2002 and October 28, 2003, is discussed. In those two bursts, a single and a pair gamma source displaced relative to the hard X-ray sources were localized in the 2.223-MeV neutron-capture line. On the basis of complex analysis of multi-wavelength (X-ray, gamma-ray, and optical) observations and the data on fast solar protons, a new topological model of the source of accelerated particles (of the mirror trap type) and a new scenario of the event that occurred on July 23, 2002 are proposed. Evidence of the possibility of particle acceleration by the electric field in coronal mass-ejection loops during large solar flares is obtained. The simulation results indicate that the gamma-ray source in the excitation lines (4.1–6.7 MeV) should coincide with the region where the accelerated ions interact with the background plasma of the solar atmosphere above the spot of the flare active region.     

Visualization of superparamagnetic nanoparticles in vascular tissue using XμCT and histology

In order to increase the dose of antineoplastic agents in the tumor area, the concept of magnetic drug targeting (MDT) has been developed. Magnetic nanoparticles consisting of iron oxide and a biocompatible cover layer suspended in an aqueous solution (ferrofluid) serve as carriers for chemotherapeutics being enriched by an external magnetic field after intra-arterial application in desired body compartments (i.e., tumor). We established an ex vivo model to simulate in vivo conditions in a circulating system consisting of magnetic iron oxide nanoparticles passing an intact bovine artery and being focused by an external magnetic field to study their distribution in the vessel. Micro-computed X-ray tomography (XμCT) and histology can elucidate the arrangement of these particles after application. XμCT-analysis has been performed on arterial sections after MDT in order to determine the distribution of the nanoparticles. These measurements have been carried out with a cone X-ray source and corresponding histological sections were stained with Prussian blue. It could be shown that combining XμCT and histology offers the opportunity for a better understanding of the mechanisms of nanoparticle deposition in the vascular system after MDT.     

Synchrotron radiation SAXS of 11S soya globulin

Small-angle X-ray scattering using the Daresbury synchrotron source has been employed to obtain scattering curves from a 5% solution of the 11S soya globulin. The high intensity of the source allowed exposure times to be reduced by up to 1000 times compared with those for a conventional X-ray generator. Submaxima at higher angles were recorded which have not been reported previously. This improved resolution appears to result from reduced aggregation and/or denaturation of the protein due to the very short exposure times. Such detail in the scattering curve should be of importance for structural modelling of the proteins, particularly in the case of the 11S soya globulin for which intact individual subunits cannot be isolated.     

A variable-energy electron microbeam: a unique modality for targeted low-LET radiation

We have designed and constructed a low-cost, variable-energy low-LET electron microbeam that uses energetic electrons to mimic radiation damage produced by gamma and X rays. The microbeam can access lower regions of the LET spectrum, similar to conventional X-ray or 60Co gamma-ray sources. The device has two operating modes, as a conventional microbeam targeting single cells or subpopulations of cells or as a pseudo broad-beam source allowing for direct comparison with conventional sources. By varying the incident electron energy, the target cells can be selectively exposed to different parts of the energetic electron tracks, including the track ends.     

Stopped-flow solution scattering using synchrotron radiation: Apparatus,data collection and data analysis

We have constructed an experimental system, under remote control, for stopped-flow X-ray scattering using synchrotron radiation. It has been used, in conjunction with an annular detector and its associated electronics, to obtain good scattering curves, with time-slices as short as 200 ms, in a new study of the dissociation of the enzyme complex aspartate transcarbamylase. The data have been analysed by new statistical methods, and they agree well with the results from parallel chemical quench experiments. For studying dissociation reactions, stopped-flow X-ray scattering is a quite practical method, which need not use very much more material than conventional stopped-flow experiments.     

Extracellular synthesis of gold bionanoparticles by <Emphasis Type="Italic">Nocardiopsis</Emphasis> sp. and evaluation of its antimicrobial,antioxidant and cytotoxic activities

Advancement of biological process for the synthesis of bionanoparticles is evolving into a key area of research in nanotechnology. The present study deals with the biosynthesis, characterization of gold bionanoparticles by Nocardiopsis sp. MBRC-48 and evaluation of their antimicrobial, antioxidant and cytotoxic activities. The gold bionanoparticles obtained were characterized by UV–visible spectroscopy, X-ray diffraction analysis, Fourier transform infrared spectroscopy, field emission scanning electron microscopy, energy dispersive X-ray analysis and transmission electron microscopy (TEM). The synthesized gold bionanoparticles were spherical in shape with an average of 11.57 ± 1.24 nm as determined by TEM and dynamic light scattering (DLS) particle size analyzer, respectively. The biosynthesized gold nanoparticles exhibited good antimicrobial activity against pathogenic microorganisms. It showed strong antioxidant activity as well as cytotoxicity against HeLa cervical cancer cell line. The present study demonstrated the potential use of the marine actinobacterial strain of Nocardiopsis sp. MBRC-48 as an important source for gold nanoparticles with improved biomedical applications including antimicrobial, antioxidant as well as cytotoxic agent.     

Cryo X-ray microscope with flat sample geometry for correlative fluorescence and nanoscale tomographic imaging

X-ray imaging offers a new 3-D view into cells. With its ability to penetrate whole hydrated cells it is ideally suited for pairing fluorescence light microscopy and nanoscale X-ray tomography. In this paper, we describe the X-ray optical set-up and the design of the cryo full-field transmission X-ray microscope (TXM) at the electron storage ring BESSY II. Compared to previous TXM set-ups with zone plate condenser monochromator, the new X-ray optical layout employs an undulator source, a spherical grating monochromator and an elliptically shaped glass capillary mirror as condenser. This set-up improves the spectral resolution by an order of magnitude. Furthermore, the partially coherent object illumination improves the contrast transfer of the microscope compared to incoherent conditions. With the new TXM, cells grown on flat support grids can be tilted perpendicular to the optical axis without any geometrical restrictions by the previously required pinhole for the zone plate monochromator close to the sample plane. We also developed an incorporated fluorescence light microscope which permits to record fluorescence, bright field and DIC images of cryogenic cells inside the TXM. For TXM tomography, imaging with multi-keV X-rays is a straightforward approach to increase the depth of focus. Under these conditions phase contrast imaging is necessary. For soft X-rays with shrinking depth of focus towards 10nm spatial resolution, thin optical sections through a thick specimen might be obtained by deconvolution X-ray microscopy. As alternative 3-D X-ray imaging techniques, the confocal cryo-STXM and the dual beam cryo-FIB/STXM with photoelectron detection are proposed.     

The bioelectric field of the catfish Ictalurus nebulosus.

The variability of the bioelectric field of the electrosensitive catfish, Ictalurus nebulosus, was investigated by recording the potential variation occurring when the fish passed a stationary electrode, and by recording the field of a stationary fish by a 15-electrode array. A good first order approximation of the recorded field of a 20 cm long fish is a dipole dc source with the source and sink about 7 cm apart, carrying a current of about 1 microA in water with a specific resistivity of 3.3 kohm cm. At 5 cm distance from the dipole axis such a source generates an electric potential swing in the order of 50 microV in free space, head negative, tail positive. Superimposed on the basic component are respiration related fluctuations, and fluctuations related to the activity of the alimentary canal, gills, and skin. Novel stimuli, or stressors like investigators approaching the aquarium, evoke sudden increases in field strength. which last about 15 min. Demineralization of the aquarium water causes changes in field strength and reversal of field polarity. The administration of food causes field variations in the vicinity of the anal opening. The bioelectric field shows diurnal fluctuations of 100 microV. The peak is at about 04:00, the dip at 14:00. The fluctuations of the bioelectric field are sufficiently strong and specilic to serve as electrical stimuli to other electrosensitive catfish. It is suggested that the field changes allow a simple form of electrocommunication. i.e. inform conspecifics about some physiological properties of the field source. The cellular mechanisms underlying the fluctuations of the bioelectric field are homeostatic processes mediated by ion pumps and ion channels.     

Dark-Field Imaging: Recent developments and potential clinical applications

This paper describes an X-ray phase contrast imaging technique using analyzer-based optics called X-ray Dark-Field Imaging that has been under development for the past 10 years. We describe the theory behind XDFI, the X-ray optics required for implementing it in practice, and algorithms used for 2D, 2.5D, and 3D image reconstruction. The XDFI optical chain consists of an asymmetrically cut, Bragg-type monochromator-collimator that provides a planar monochromatic X-ray beam, a positioning stage for the specimens, a Laue-case angle analyzer, and one or two cameras to capture the dark and bright field images. We demonstrate the soft-tissue discrimination capabilities of XDFI by reconstructing images with absorption and phase contrast. By using a variety of specimens such as breast tissue with cancer, joints with articular cartilage, ex-vivo human eye specimen, and others, we show that refraction-based contrast derived from XDFI is more effective in characterizing anatomical features, articular pathology, and neoplastic disease than conventional absorption-based images. For example, XDFI of breast tissue can discriminate between the normal and diseased terminal duct lobular unit, and between invasive and in-situ cancer. The final section of this paper is devoted to potential future developments to enable clinical and histo-pathological applications of this technique.     

Experimental evaluation of depth-of-interaction correction in a small-animal positron emission tomography scanner

Human and small-animal positron emission tomography (PET) scanners with cylindrical geometry and conventional detectors exhibit a progressive reduction in radial spatial resolution with increasing radial distance from the geometric axis of the scanner. This "depth-of-interaction" (DOI) effect is sufficiently deleterious that many laboratories have devised novel schemes to reduce the magnitude of this effect and thereby yield PET images of greater quantitative accuracy. Here we examine experimentally the effects of a particular DOI correction method (dual-scintillator phoswich detectors with pulse shape discrimination) implemented in a small-animal PET scanner by comparing the same phantom and same mouse images with and without DOI correction. The results suggest that even this relatively coarse, two-level estimate of radial gamma ray interaction position significantly reduces the DOI parallax error. This study also confirms two less appreciated advantages of DOI correction: a reduction in radial distortion and radial source displacement as a source is moved toward the edge of the field of view and a resolution improvement detectable in the central field of view likely owing to improved spatial sampling.     

X-ray Dose Reduction through Adaptive Exposure in Fluoroscopic Imaging

X-ray fluoroscopy is widely used for image guidance during cardiac intervention. However, radiation dose in these procedures can be high, and this is a significant concern, particularly in pediatric applications. Pediatrics procedures are in general much more complex than those performed on adults and thus are on average four to eight times longer¹. Furthermore, children can undergo up to 10 fluoroscopic procedures by the age of 10, and have been shown to have a three-fold higher risk of developing fatal cancer throughout their life than the general population^2,3.We have shown that radiation dose can be significantly reduced in adult cardiac procedures by using our scanning beam digital x-ray (SBDX) system⁴-- a fluoroscopic imaging system that employs an inverse imaging geometry^5,6 (Figure 1, Movie 1 and Figure 2). Instead of a single focal spot and an extended detector as used in conventional systems, our approach utilizes an extended X-ray source with multiple focal spots focused on a small detector. Our X-ray source consists of a scanning electron beam sequentially illuminating up to 9,000 focal spot positions. Each focal spot projects a small portion of the imaging volume onto the detector. In contrast to a conventional system where the final image is directly projected onto the detector, the SBDX uses a dedicated algorithm to reconstruct the final image from the 9,000 detector images.For pediatric applications, dose savings with the SBDX system are expected to be smaller than in adult procedures. However, the SBDX system allows for additional dose savings by implementing an electronic adaptive exposure technique. Key to this method is the multi-beam scanning technique of the SBDX system: rather than exposing every part of the image with the same radiation dose, we can dynamically vary the exposure depending on the opacity of the region exposed. Therefore, we can significantly reduce exposure in radiolucent areas and maintain exposure in more opaque regions. In our current implementation, the adaptive exposure requires user interaction (Figure 3). However, in the future, the adaptive exposure will be real time and fully automatic.We have performed experiments with an anthropomorphic phantom and compared measured radiation dose with and without adaptive exposure using a dose area product (DAP) meter. In the experiment presented here, we find a dose reduction of 30%.     

Spectroscopic imaging at compact inverse Compton X-ray sources

While K-edge subtraction (KES) imaging is a commonly applied technique at synchrotron sources, the application of this imaging method in clinical imaging is limited although results have shown its superiority to conventional clinical subtraction imaging. Over the past decades, compact synchrotron X-ray sources, based on inverse Compton scattering, have been developed to fill the gap between conventional X-ray tubes and synchrotron facilities. These so called inverse Compton sources (ICSs) provide a tunable, quasi-monochromatic X-ray beam in a laboratory setting with reduced spatial and financial requirements. This allows for the transfer of imaging techniques that have been limited to synchrotrons until now, like KES imaging, into a laboratory environment. This review article presents the first studies that have successfully performed KES at ICSs. These have shown that KES provides improved image quality in comparison to conventional X-ray imaging. The results indicate that medical imaging could benefit from monochromatic imaging and KES techniques. Currently, the clinical application of KES is limited by the low K-edge energy of available iodine contrast agents. However, several ICSs are under development or already in commissioning which will provide monochromatic X-ray beams with higher X-ray energies and will enable KES using high-Z elements as contrast media. With these developments, KES at an ICS has the ability to become an important tool in pre-clinical research and potentially advancing existing clinical imaging techniques.     

X-ray diffraction photographs of chicken gizzard G-actin.DNase I complex crystals taken with synchrotron radiation

X-ray diffraction photographs of a chicken gizzard G-actin.DNase I complex crystal have been recorded using the synchrotron radiation beam emitted by the Synchrotron Radiation Source at Daresbury and the Photon Factory at Tsukuba. The resolution limit was extended to 2.4 A and the exposure time was reduced approximately by a factor of 10, when data recorded at the Photon Factory, were compared with those recorded with a conventional rotating-anode source. Using a newly designed Weissenberg camera equipped with a multi-layer line screen, the diffraction data in a 36 degrees oscillation range were recorded on a single film up to 3.5 A resolution.     

Crystallization of a type I 3-dehydroquinase from Salmonella typhi.

Crystals have been grown of a type I 3-dehydroquinase from both Escherichia coli and Salmonella typhi. However, only those from S. typhi diffract to a resolution of 2.3 A on a conventional X-ray source and are suitable for structure determination. The space group has been determined as P2(1)2(1)2 with unit cell dimensions a = 48.01 A, b = 114.29 A, c = 42.87 A. There is one subunit in the asymmetric unit.     

Influence of asynchronous actuation of the Angara-5-1 facility modules on the penetration of the magnetic field inside a wire array

The influence of asynchronous actuation of the Angara-5-1 facility modules on the implosion symmetry of the wire array plasma in the Z-pinch mode is studied. It is shown that an increase in the r.m.s. jitter in the start times of the modules leads to an appreciable azimuthal asymmetry of magnetic field penetration inside the wire array and, as a consequence, to an increase in the duration of the soft X-ray pulse, a decrease in its power, and a shift of the Z-pinch with respect to the array axis. Necessary conditions for axisymmetric pinch implosion are determined. Experimental data on the magnitude and azimuthal distribution of the current (magnetic field) inside the wire array were obtained from magnetic probe measurements. The position of the Z-pinch with respect to the wire array axis was determined from two-dimensional X-ray images and radial optical streak images.     

Study of the implosion characteristics of quasi-spherical wire arrays on the Angara-5-1 facility at currents of up to 4 MA

Results are presented from experimental studies of the spatial distribution of the density of matter in the central part of the discharge gap and the formation of the temporal profile of the X-ray power in the course of implosion of quasi-spherical wire arrays at discharge currents of up to 4 MA. The spatial distribution of the X-ray intensity in the central part of the discharge gap and the temporal profile of the X-ray power are used as implosion characteristics of quasi-spherical wire arrays. The quasi-spherical arrays were formed by the radial stretching of unstrained wires of initially cylindrical and conical wire arrays under the action of the electrostatic field. The temporal profile of the output X-ray pulse in the photon energy range of 0.1–1 keV is shown to depend on both the geometrical parameters of the quasi-spherical array and the longitudinal distribution of its mass. It is found that a 40% increase in the wire mass due to deposition of an additional mass in the equatorial region of a quasi-spherical array leads to a 15% increase in the average current radius of the pinch and a 30% decrease in the X-ray yield. Experiments with quasi-spherical arrays made of kapron fibers with deposited Al and Bi conducting layers were also carried out. It is demonstrated that application of such arrays makes it possible to control the profile and duration of the generated X-ray pulse by varying the mass, material, and location of the deposited layer. It is found that deposition of an additional mass in the form of a thin Bi stripe on tungsten wires near the cathode end of the array allows one to mitigate the influence of the cathode zipper effect on the pinch compression and formation of the X-ray pulse in tungsten arrays.