bluuebunny/crossref_metadata_2025 huggingface dataset:
Real-time feedback control of millimeter-wave polarization for LHD
Authors: Felici F. , Goodman T. , Sauter O. , Shimozuma T. , Ito S. , Mizuno Y. , Kubo S. , Mutoh T.
Published: February 2009

Electron cyclotron heating (ECH) is widely used in magnetic fusion devices, and the polarization of the injected millimeter-wave beams plays a crucial role in the propagation and absorption of the beam energy by the plasma. This polarization can be adjusted by grating mirror polarizers placed in the transmission lines which carry the microwaves from the power source to the plasma. In long-pulse devices such as the Large Helical Device (LHD) and ITER, it is desirable to track changes in the plasma and adjust the polarization of the ECH in real time such as to keep the absorption as high as possible and avoid shine-through which may lead to overheating of vessel components. For this purpose a real-time feedback control scheme is envisioned in which a measure of the absorption efficiency can be used to adjust the orientation of the polarizing mirrors toward an optimum. Such a setup has been tested in a low-power test stand as preparation for future implementation in the LHD ECH system. It is shown that a simple search algorithm is efficient and can in principle be used to control either the absorption efficiency or the linear polarization angle.
High power light emitting diode based setup for photobleaching fluorescent impurities
Authors: Herman Tobias K. , Mackowiak Stephan A. , Kaufman Laura J.
Published: February 2009

Single molecule fluorescence experiments, with their associated low signals, require very low background fluorescence in the sample. Even high purity liquids will often possess large numbers of fluorescent impurities that are difficult to completely remove through standard purification techniques such as distillation and recrystallization. We have constructed a simple setup in which such impurities can be photobleached before final sample preparation. The instrument consists of high power light emitting diodes, and it delivers almost 10 W of light to the sample without the heating associated with more conventional light sources or the cost and safety concerns associated with a high power laser.
The effect of stress on the dielectric and tunable properties of barium stannate titanate thin films
Authors: Song Sannian , Zhai Jiwei , Gao Lina , Yao Xi
Published: February 2009

Ba ( Sn 0.15 Ti 0.85 ) O 3 thin films were grown on the LaNiO3 buffered LaAlO3, SrTiO3, MgO, and Al2O3 single crystal substrates, respectively. These substrates provide a systematic change in the stress while maintaining the same film microstructure. The stress in the thin film induces an obvious change in the dielectric behavior. The reduction in the ferroelectric transition temperature with increasing biaxial tensile stress is attributed to the suppression of in-plane polarization due to the small lateral grain size in the films. The in-plane tensile stress in this study reduces the unit cell along electric field in the parallel plate capacitor structure and decreases the tunability.
Modeling edge effects in graphene nanoribbon field-effect transistors with real and mode space methods
Authors: Zhao Pei , Guo Jing
Published: February 2009

A computationally efficient mode space simulation method for atomistic simulation of a graphene nanoribbon field-effect transistor in the ballistic limits is developed. The proposed simulation scheme, which solves the nonequilibrium Green’s function coupled with a three dimensional Poisson equation, is based on the atomistic Hamiltonian in a decoupled mode space. The mode space approach, which only treats a few modes (subbands), significantly reduces the simulation time. Additionally, the edge bond relaxation and the third nearest neighbor effects are also included in the quantum transport solver. Simulation examples show that the mode space approach can significantly decrease the simulation cost by about an order of magnitude, yet the results are still accurate. This article also demonstrates that the effects of the edge bond relaxation and the third nearest neighbor significantly influence the transistor’s performance and are necessary to be included in the modeling.
On the interplay between tungsten and tantalum atoms in Ni-based superalloys: An atom-probe tomographic and first-principles study
Authors: Amouyal Yaron , Mao Zugang , Booth-Morrison Christopher , Seidman David N.
Published: February 2009

The partitioning behavior of W in a multicomponent Ni-based superalloy and in a ternary Ni–Al–W alloy is investigated using atom-probe tomography (APT) and first-principles calculations. APT observations indicate that whereas W partitions preferentially to the γ′(L12)-precipitates in the ternary alloy, its partitioning behavior is reversed in favor of the γ(fcc)-matrix in the multicomponent alloy. First-principles calculations of the substitutional formation energies of W and Ta predict that Ta has a larger driving force for partitioning to the γ′ phase than W. This implies that Ta displaces W from the γ′-precipitates into the γ-matrix in multicomponent alloys.
Frequency modulation atomic force microscopy in ambient environments utilizing robust feedback tuning
Authors: Kilpatrick J. I. , Gannepalli A. , Cleveland J. P. , Jarvis S. P.
Published: February 2009

Frequency modulation atomic force microscopy (FM-AFM) is rapidly evolving as the technique of choice in the pursuit of high resolution imaging of biological samples in ambient environments. The enhanced stability afforded by this dynamic AFM mode combined with quantitative analysis enables the study of complex biological systems, at the nanoscale, in their native physiological environment. The operational bandwidth and accuracy of constant amplitude FM-AFM in low Q environments is heavily dependent on the cantilever dynamics and the performance of the demodulation and feedback loops employed to oscillate the cantilever at its resonant frequency with a constant amplitude. Often researchers use ad hoc feedback gains or instrument default values that can result in an inability to quantify experimental data. Poor choice of gains or exceeding the operational bandwidth can result in imaging artifacts and damage to the tip and/or sample. To alleviate this situation we present here a methodology to determine feedback gains for the amplitude and frequency loops that are specific to the cantilever and its environment, which can serve as a reasonable “first guess,” thus making quantitative FM-AFM in low Q environments more accessible to the nonexpert. This technique is successfully demonstrated for the low Q systems of air (Q∼40) and water (Q∼1). In addition, we present FM-AFM images of MC3T3-E1 preosteoblast cells acquired using the gains calculated by this methodology demonstrating the effectiveness of this technique.
Interferences in photodetachment of a triatomic negative ion
Authors: Afaq A. , Ahmad Iftikhar , Ahmad M. A. , Rashid A. , Tahir B. A. , Hussain Muhammad Tahir
Published: February 2009

The photodetachment of a triatomic negative ion is studied and the detached-electron wave function is obtained as a superposition of coherent waves originating from each atom of the system. The photodetached electron flux is evaluated on a screen placed at a large distance from the system, which displays strong interferences. A simple analytical formula is also obtained for the total photodetachment cross section. The formula approaches one time the cross sections for the one-center and two-center systems in the high photon energy limits. Also it approaches three times the cross section for one-center system in the low photon energy limits.
Determination of MgO/AlN heterojunction band offsets by x-ray photoelectron spectroscopy
Authors: Yang A. L. , Song H. P. , Liu X. L. , Wei H. Y. , Guo Y. , Zheng G. L. , Jiao C. M. , Yang S. Y. , Zhu Q. S. , Wang Z. G.
Published: February 2009

MgO is a promising gate dielectric and surface passivation film for GaN/AlGaN transistors, but little is known of the band offsets in the MgO/AlN system. X-ray photoelectron spectroscopy was used to measure the energy discontinuity in the valence band (ΔEv) of MgO/AlN heterostructures. A value of ΔEv=0.22±0.08 eV was obtained. Given the experimental band gap of 7.83 eV for MgO, a type-I heterojunction with a conduction band offset of ∼1.45 eV is found. The accurate determination of the valence and conduction band offsets is important for use of III-N alloys based electronic devices.
Self-organized periodic domain structure for second harmonic generations in ferroelastic β′-(Sm,Gd)2(MoO4)3 crystal lines on glass surfaces
Authors: Tsukada Yuta , Honma Tsuyoshi , Komatsu Takayuki
Published: February 2009

Ferroelastic β′-(Sm,Gd)2(MoO4)3 (SGMO) crystal lines are patterned on the surface of Sm2O3–Gd2O3–MoO3–B2O3 glasses by continuous wave Yb:YVO4 laser irradiations (wavelength is 1080 nm, power is 1.2 W and scanning speed is 5 μm/s). Crystal lines show self-organized periodic domain structures with periodic refractive index changes. The intensity of second harmonic (SH) waves changes periodically depending on the domain structure. It is proposed from polarized micro-Raman scattering spectra and the azimuthal dependence of SH intensities that the orientation of (MoO4)2− tetrahedra in SGMO crystals changes gradually and periodically along the crystal line growth direction due to the spontaneous strains in SGMO crystals.
Demonstration of diffraction enhancement via Bloch surface waves in a-SiN:H multilayers
Authors: Liscidini Marco , Galli Matteo , Patrini Maddalena , Loo Richard W. , Goh M. Cynthia , Ricciardi Carlo , Giorgis Fabrizio , Sipe J. E.
Published: February 2009

By exploiting the excitation of a Bloch surface wave at a wavelength λ=670 nm, we observe a diffraction enhancement of more than 45 times from a rabbit IgG protein grating printed on a-SiN:H multilayers. Our results demonstrate that the use of surface states in dielectric multilayers is very promising for the realization of the next generation of diffraction-based biosensors.
Auxetic behavior under electrical loads in an induced ferroelectric phase
Authors: Tan X. , Jo W. , Granzow T. , Frederick J. , Aulbach E. , Rödel J.
Published: February 2009

The longitudinal and transverse strains were measured as a function of applied electric fields in a bulk ceramic sample of Pb0.99Nb0.02[(Zr0.57Sn0.43)0.94Ti0.06]0.98O3 at room temperature. Instead of a transverse contraction, a transverse expansion was observed in the electric-field-induced ferroelectric phase after the antiferroelectric-to-ferroelectric phase transition. Therefore, an auxetic behavior was established in monolithic ferroelectric polycrystalline ceramics under electrical loads. The behavior is characterized by a negative strain ratio that is analogous to the Poisson’s ratio. The transverse expansion leads to a large hydrostatic piezoelectric coefficient dh, which suggests new applications of antiferroelectric ceramics in piezoelectric devices.
Silver-clad nitride semiconductor laser diode
Authors: Bour David , Chua Christopher , Yang Zhihong , Teepe Mark , Johnson Noble
Published: February 2009

An edge-emitting laser diode is described in which the epitaxial upper cladding layer is replaced with a metal contact selected for its optical as well as electrical properties. The structure is demonstrated with an In0.1Ga0.9N multiple-quantum-well laser diode operating at 412 nm with silver rather than the typical p-type AlGaN for the upper cladding. Silver is effective as a cladding layer because of its very low index of refraction at the lasing wavelength, so that the guided optical mode exhibits minimal penetration into the silver. Eliminating the epitaxial upper cladding layer reduces thermal degradation of the InGaN multiple-quantum-well active layer.
Domain wall anisotropic magnetoresistance in planar nanowires
Authors: Bogart L. K. , Atkinson D.
Published: February 2009

The anisotropic magnetoresistance (AMR) of individual magnetic domain walls in planar nanowires has been systematically investigated as a function of the micromagnetic wall structure, width, and thickness of the nanowire. Experimentally derived thickness dependence of both the thin film resistivity and the anisotropic magnetoresistance were incorporated into the calculations. We found that the AMR value can be used to identify the wall structure and that the wall resistance is sensitive to wire width for widths less than 300 nm. Furthermore, in comparison with the detailed analysis here, a previous simplified domain wall model significantly underestimates domain wall resistance in narrower wires.
Lifetime recovery in ultrahighly titanium-doped silicon for the implementation of an intermediate band material
Authors: Antolín E. , Martí A. , Olea J. , Pastor D. , González-Díaz G. , Mártil I. , Luque A.
Published: February 2009

The doping of conventional semiconductors with deep level (DL) centers has been proposed to synthesize intermediate band materials. A recent fundamental study of the nonradiative recombination (NRR) mechanisms predicts the suppression of the NRR for ultrahigh DL dilutions as a result of the delocalization of the impurity electron wave functions. Carrier lifetime measurements on Si wafers doped with Ti in the 1020–1021 cm−3 concentration range show an increase in the lifetime, in agreement with the NRR suppression predicted and contrary to the classic understanding of DL action.
Time-dependent electrical double layer with blocking electrode
Authors: Han Z. J. , Morrow R. , Tay B. K. , McKenzie D.
Published: February 2009

This paper deals with the experimental observation of time-dependent electrical double layer (EDL) in electrolyte. A potential-distance diagram is used to fully understand different stages in the formation of EDL. The influence of the thickness of the blocking layer and the ionic strength to the formation of EDL is discussed based on the equivalent circuit. With this simple method, it is found that in addition to Debye screening length, the frequency has to be considered if an alternating electric field is used to control the movement of charged biomolecules inside EDL.
Dual-wavelength neodymium-doped yttrium aluminum garnet laser with chromium-doped yttrium aluminum garnet as frequency selector
Authors: Yu Haohai , Zhang Huaijin , Wang Zhengping , Wang Jiyang , Yu Yonggui , Zhang Xingyu , Lan Ruijun , Jiang Minhua
Published: February 2009

We report on a potential source for the generation of terahertz radiation using the well-known neodymium-doped yttrium aluminum garnet as the laser material, and chromium-doped yttrium aluminum garnet (Cr:YAG) as the frequency selector and saturable absorber. Continuous-wave output power as high as 5.28 W at 1052 nm was achieved. With Cr:YAG, the laser has a dual-wavelength output at 1052 and 1064 nm. The maximum average output power, pulse repetition rate, and pulse energy are 3.75 W, 41.2 kHz, and 90.1 μJ, respectively. We also found that the intensity ratio of the two oscillating modes can be tuned by adjusting the pump power.
Iridescent large-area ZrO2 photonic crystals using butterfly as templates
Authors: Chen Yu , Gu Jiajun , Zhu Shenmin , Fan Tongxiang , Zhang Di , Guo Qixin
Published: February 2009

Intact ZrO2 (with refractive index of 2.12) replica, which is large in size (about 3×4 cm2), has been synthesized by using natural butterfly wings as templates. Microstructure characters of original butterfly wing scales are maintained faithfully in this biomorphic ZrO2. All replicas can reflect iridescent visible lights, which can even be observed by naked eyes. Optical microscope investigations indicate that colors reflected by one single scale are different from those done by the overlapped two or even more scales. Colors are not only determined by materials’ refractive index, observation angle, and the structure of every single scale, but also by its piled number and modes. With the increase in the number of piled scales, the color is not simply redshifted or blueshifted, which is the most direct and powerful evidence for structural colors.
The role of aluminum oxide buffer layer in organic spin-valves performance
Authors: Zhan Y. Q. , Liu X. J. , Carlegrim E. , Li F. H. , Bergenti I. , Graziosi P. , Dediu V. , Fahlman M.
Published: February 2009

The electronic structures of the 8-hydroxyquinoline-aluminum (Alq3)/Al2O3/Co interfaces were studied by photoelectron spectroscopy. A strong interface dipole was observed, which leads to a reduction in the electron injection barrier. The x-ray photoelectron spectroscopy spectra further indicate that the Al2O3 buffer layer prevents the chemical interaction between Alq3 molecules and Co atoms. X-ray magnetic circular dichroism results demonstrate that a Co layer deposited on an Al2O3 buffered Alq3 layer shows better magnetic ordering in the interface region than directly deposited Co, which suggests a better performance of spin valves with such a buffer layer. This is consistent with the recent results from [Dediu et al., Phys. Rev. B 78, 115203 (2008)].
Frequently Asked Questions
Authors: Grabher Gernot , Stark David
Published: February 2009

What's this got to do with social geography?
Cognition, Context, and Multimethod Approaches to Economic Decision Making
Authors: Strauss Kendra
Published: February 2009

Research in behavioural economics, as in economic geography, seeks to address fundamental questions about cognition, the status of human rationality, and social-cum-spatial structure in economic decision making. Critiques of the ‘strong’ model of economic rationality that underpins models of behaviour in orthodox economics by those working under the rubric of the former, such as Thaler and Sunstein, are gaining the attention of politicians and policy makers. Yet the ‘limits to rationality’ thesis nevertheless lacks a credible appreciation of the diversity of social life and the significance of context in framing behaviour. Economic geography, while taking seriously the issues of the coconstitution of culture and economy and the relational nature of economic decision making, has moved away from behaviouralism at a time when critical approaches are arguably more salient than ever. I put forward the argument for an engagement with behavioural economic approaches to decision making while critically examining the need for multimethod approach that can accommodate quantitative, statistical, and experimental approaches and qualitative work that takes seriously social identity and attributes including gender, age, social class, and aspirations. The challenge of theorising the social context in which choices are made is discussed with reference to Bourdieu's notion of habitus, and three exemplars are used to illustrate the strengths and weaknesses of different methodological approaches. Herbert Simon's metaphor, which conceptualises the decision-making moment as a pair of scissors, is suggested as a way of thinking about the intersection between cognition and the decision-making environment.
Power in Numbers: A Call for Analytical Generosity toward New Political Strategies
Authors: Hamilton Trina
Published: February 2009

This paper is a methodological and epistemological reflection on the power of numbers to contribute to the debate over the potential and limitations of market politics as a regulatory force in the global economy. Informal regulatory networks, including transnational corporate campaigns, form a new sphere of politics which leap-frogs the state and targets corporations directly concerning their social and environmental impacts. I describe my statistical analysis of corporate campaigns targeting US multinationals to argue in favor of analytical generosity when evaluating these new political forms. I argue that a reflexive and critical quantification can provide new insights into stakeholder power and contemporary political processes.
Adapting Standards of Care Under Extreme Conditions
Authors: Gebbie Kristine M. , Peterson Cheryl A. , Subbarao Italo , White Kathleen M.
Published: February 2009

The filing of criminal charges against a group of clinicians in New Orleans for failure to meet expected standards of care following the hurricanes of 2005 made the growing concern among health professionals about care provided during extreme emergencies or disasters all too real. Questions about what may lead to censure, penalties from licensing boards, or lawsuits have come from nurses, physicians, and many other licensed health professionals. A panel convened by the American Nurses Association that included representatives of medical, public health, hospital, and government agencies considered the ethical, professional, and practical aspects of meeting standards of care in such circumstances. Clinicians are reminded that in emergencies, it is only the circumstances that change (perhaps radically); neither the individual’s professional competency nor the basic professional standard of care is different. In making prioritized decisions under such circumstances, the individual’s ethical framework is utilitarian, and there are 3 areas for action, even when some routine tasks are set aside: maintain worker and patient safety; maintain airway, breathing, and circulation; and establish or maintain infection control. Policy recommendations such as state legislation for the adoption of comprehensive immunity for volunteer health care workers, and the establishment of a medical review panel as arbitration board are also suggested. The resulting white paper summarizes the issues and provides guidance to individual professionals, institutions in which they work, and emergency planners. (Disaster Med Public Health Preparedness. 2009;3:111–116)
Chromosome Conformation Capture
Authors: Cope Nathan F. , Fraser Peter
Published: February 2009

INTRODUCTION

Chromosome conformation capture (3C) is a technique used to detect the spatial organization of chromosomal DNA in fixed cells. DNA sequences in spatial proximity in the nucleus or engaged in physical interactions (such as those between genes and regulatory elements) can be assessed quantitatively to provide a measure that potentially reflects their frequency of association and/or their proximity. 3C can be used to study long-range interactions between DNA sequences on the same chromosome (intrachromosomal) or between different chromosomes (interchromosomal). Briefly, chromatin fragments in proximity are fixed with formaldehyde, followed by digestion with restriction enzymes, nuclear lysis, dilution of the cross-linked complexes, and intramolecular ligation. The cross-links are subsequently removed and the DNA is purified. Polymerase chain reaction (PCR) is then used to amplify DNA fragments containing novel ligation junctions using primers specific for a pair of DNA sequences under investigation. Here we describe a detailed 3C protocol and experimental controls that can be applied to investigate the nuclear juxtaposition of any two genomic regions, in cis or trans.
DNA Extraction from Formalin-Fixed, Paraffin-Embedded Tissue
Authors: Tang Weining , David Freedom B. , Wilson Malania M. , Barwick Benjamin G. , Leyland-Jones Brian R. , Bouzyk Mark M.
Published: February 2009

Introduction

Formalin-fixed, paraffin-embedded (FFPE) tissue is one of the most widely practiced methods for clinical sample preservation and archiving. It is estimated that, worldwide, over a billion tissue samples, most of them FFPE, are being stored in numerous hospitals, tissue banks, and research laboratories. These archived samples could potentially provide a wealth of information in retrospective molecular studies of diseased tissues. While standard for histopathology and microscopic investigation (e.g., hematoxylin and eosin [H&E] staining, immunohistochemistry [IHC], and tissue microarray [TMA]), FFPE samples pose a major challenge for molecular pathologists, because nucleic acids are heavily modified and trapped by extensive protein-nucleic acid and protein-protein cross linking. Historically, FFPE samples were not considered to be a viable source for molecular analyses. Recently, however, it has been discovered that with appropriate protease digestion, it is possible to release microgram amounts of DNA and RNA from FFPE samples. The purified nucleic acids, although highly fragmented, are suitable for a variety of downstream genomic and gene expression analyses, such as polymerase chain reaction (PCR), quantitative reverse transcription PCR (qRT-PCR), microarray, array comparative genomic hybridization (CGH), microRNA, and methylation profiling. Several commercial kits are currently available for FFPE extraction. The protocol reported here is adapted from the Ambion RecoverAll Total Nucleic Acid Isolation Kit, but includes several modifications. Although our protocol focuses on DNA isolation, the RecoverAll Kit can also be utilized to recover RNA, including microRNA.
Virus-Induced Gene Silencing as a Tool for Delivery of dsRNA into Plants: Figure 1.
Authors: Padmanabhan Meenu , Dinesh-Kumar Savithramma P.
Published: February 2009

INTRODUCTION

The inherent RNA silencing mechanism in plants has been effectively manipulated as a tool for the targeted down-regulation of genes. Numerous methods have been employed to initiate this homology-based RNA degradation process, but all rely on the activity of double-stranded RNAs (dsRNAs) corresponding to the gene of interest. Virus-induced gene silencing (VIGS) has gained acceptance as the tool of choice for transient induction of silencing. It involves creation of engineered viruses carrying sequences corresponding to the host gene to be silenced. Infection leads to synthesis of viral dsRNA, an intermediate step in viral replication. This activates the anti-viral RNA silencing pathway, resulting in down-regulation of the host gene transcript. While several VIGS vectors have been developed, the Tobacco Rattle Virus (TRV) provides the most robust results in terms of efficiency, ease of application, and absence of disease symptoms. Engineered TRV vectors carrying host-derived segments are transformed into Agrobacterium tumefaciens, which is then introduced into the plant. This protocol outlines a simple procedure for introducing the TRV-based binary vectors pTRV1 and pTRV2 into solanaceous plants such as Nicotiana benthamiana and Solanum lycopersicon (tomato), as well as Arabidopsis.
Somatic Hybridization in the Moss Physcomitrella patens Using PEG-Induced Protoplast Fusion
Authors: Cove David J. , Perroud Pierre-François , Charron Audra J. , McDaniel Stuart F. , Khandelwal Abha , Quatrano Ralph S.
Published: February 2009

INTRODUCTION

As an alternative to sexual crossing, protoplasts from two strains of moss (Physcomitrella patens) can be hybridized using polyethylene glycol (PEG). Although the efficiency is low, it requires no sophisticated apparatus. Hybrids are readily obtained using complementary auxotrophic mutants or strains with transgenic antibiotic resistance markers. It is now routine to obtain hybrids using transgenic strains that are hygromycin- or G418-resistant by selecting hybrids on medium containing both antibiotics.
Chemical and UV Mutagenesis of Spores and Protonemal Tissue from the Moss Physcomitrella patens
Authors: Cove David J. , Perroud Pierre-François , Charron Audra J. , McDaniel Stuart F. , Khandelwal Abha , Quatrano Ralph S.
Published: February 2009

INTRODUCTION

This protocol describes how to mutagenize spores and protonemal tissue from moss (Physcomitrella patens) using chemicals or ultraviolet (UV) light. Spores are mutagenized using the alkylating agents N-methyl-N′-nitro-N-nitrosoguanidine (NTG) and ethyl methanesulfonate (EMS), and protonemal tissue is mutagenized with NTG and UV light. Compared to alkylating agents, UV is less effective as a mutagen, but it may be advantageous because it is less hazardous and may not lead to clustered lesions.
Transformation of the Moss Physcomitrella patens Using Direct DNA Uptake by Protoplasts
Authors: Cove David J. , Perroud Pierre-François , Charron Audra J. , McDaniel Stuart F. , Khandelwal Abha , Quatrano Ralph S.
Published: February 2009

INTRODUCTION

This protocol describes how to transform moss (Physcomitrella patens) protoplasts using polyethylene glycol (PEG)-mediated DNA uptake. The transformation rates for direct uptake by protoplasts of DNA with and without genomic sequence (a targeting construct) are typically 10−5 and 10−3, respectively. (These are the frequencies of stable transformants among regenerants surviving the transformation procedure.)
Transformation of Moss Physcomitrella patens Gametophytes Using a Biolistic Projectile Delivery System
Authors: Cove David J. , Perroud Pierre-François , Charron Audra J. , McDaniel Stuart F. , Khandelwal Abha , Quatrano Ralph S.
Published: February 2009

INTRODUCTION

This method makes use of biolistic bombardment to propel gold particles coated with DNA into protonemal tissue of the moss Physcomitrella patens. It is especially suitable for transient gene expression studies, but it can be used to obtain stable transformants.
Isolation of Single Choanoflagellate Cells from Field Samples and Establishment of Clonal Cultures
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. This protocol describes how to enrich freshly collected field samples and isolate single choanoflagellate cells. Two methods are commonly used: isolation by micropipetting single cells and isolation by dilution. The two methods are complementary and each has its advantages and disadvantages. The micropipette technique requires considerable experience. However, provided a single cell has been isolated, any culture arising from it will be clonal. The dilution technique does not require the same level of skill, but single-cell isolation is not ensured. For the latter technique, repeating the isolation procedure using higher dilutions is advised to ensure that only one cell is the basis of an established culture.
Long-Term Frozen Storage of Choanoflagellate Cultures
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. Choanoflagellates can be stored under stable conditions by freezing them with liquid nitrogen, as described here. To prevent cross-contamination among vials stored in the liquid phase of a liquid nitrogen freezer, the use of CryoFlex tubing to fully seal the vials is encouraged.
Visualizing the Subcellular Localization of Actin, β-Tubulin, and DNA in Monosiga brevicollis
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. They occur as spherical to ovoid cells exhibiting morphological similarities to the choanocytes (feeding cells) of sponges. Specifically, they are characterized by a single anterior flagellum surrounded by a funnel-shaped collar composed of 30 or more actin-based tentacles or microvilli. These structures can be visualized by staining for actin and β-tubulin. In this protocol, Monosiga brevicollis cells are fixed in dilute formaldehyde, attached to coverslips, and then exposed to antibodies against β-tubulin and an actin-specific stain; a simple DNA stain highlights the nucleus. The central challenge of this protocol is to treat the cells gently so that they retain their ultrastructure and remain attached to the coverslips during processing.
Preparation of Total RNA from Monosiga brevicollis and Other Choanoflagellates
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. The preparation of RNA is an important prerequisite for characterizing the gene expression profile of choanoflagellates under different conditions. Although most standard protocols (e.g., those using TRIzol) are sufficient for many purposes, the approach described here minimizes RNA degradation and provides a high-quality template for downstream protocols, including real-time polymerase chain reaction.
Rapid Preparation of Genomic DNA from Monosiga brevicollis and Other Choanoflagellates
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. This protocol describes a straightforward but crude isolation strategy for genomic DNA that has been effective on all tested choanoflagellate species. The resulting genomic DNA provides a suitable template for a variety of polymerase chain reaction (PCR) techniques, but is too impure for procedures that require very high-quality DNA such as Southern blotting.
Preparation of High-Molecular-Weight Genomic DNA from Monosiga brevicollis and Other Choanoflagellates
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. This protocol describes a simple genomic DNA isolation strategy that should be effective for almost all choanoflagellate species and will provide genomic templates suitable for polymerase chain reactions and Southern blot analysis.
Separation of Choanoflagellate and Bacterial Genomic DNA
Authors: King Nicole , Young Susan L. , Abedin Monika , Carr Martin , Leadbeater Barry S.C.
Published: February 2009

INTRODUCTION

Choanoflagellates are heterotrophic nanoflagellates: small, colorless protozoa that are present in marine and freshwater environments as well as in hydrated soils. Because they are the closest living relatives of the metazoa, the study of their cell biology and genomes promises to provide new insights into metazoan ancestry and origins. However, the preparation of genomic DNA from cultures of Monosiga brevicollis or other choanoflagellates can be complicated by the presence of abundant bacterial DNA. Indeed, bacterial DNA can represent as much as 90% of the total yield from a standard genomic DNA preparation. This protocol describes the use of a CsCl gradient to separate choanoflagellate genomic DNA from the DNA of its bacterial prey. This strategy works only when the G+C content of the choanoflagellate genomic DNA is significantly different from that of its bacterial food. The final DNA sample from this protocol is expected to be 90%-95% enriched for choanoflagellate genomic DNA and to provide an adequate template for genome sequencing projects.
The OP9-DL1 System: Generation of T-Lymphocytes from Embryonic or Hematopoietic Stem Cells In Vitro
Authors: Holmes Roxanne , Zúñiga-Pflücker Juan Carlos
Published: February 2009

INTRODUCTION

Differentiation of mouse embryonic stem cells (ESCs) or hematopoietic stem cells (HSCs) from fetal liver or bone marrow into T-lymphocytes can be achieved in vitro with the support of OP9-DL1 cells, a bone-marrow-derived stromal cell line that ectopically expresses the Notch ligand, Delta-like 1 (Dll1). This approach provides a simple, versatile, and efficient culture system that allows for the commitment, differentiation, and proliferation of T-lineage cells from different sources of stem cells. This article contains a series of protocols, the first of which describes the establishment, maintenance, and storage of OP9 and OP9-DL1 cells. Subsequent protocols detail how to co-culture the OP9 and OP9-DL1 cells with either ESCs or HSCs from fetal liver or bone marrow, leading to in vitro differentiation of the stem cells into lymphocytes.
Patient safety incidents associated with airway devices in critical care: a review of reports to the UK National Patient Safety Agency
Authors: Thomas A. N. , McGrath B. A.
Published: February 2009

We used key words and letter sequences to identify airway‐associated patient safety incidents submitted to the UK National Patient Safety Agency from critical care units in England and Wales. We identified 1085 such airway incidents submitted in the two years from October 2005 to September 2007. Three hundred and twelve incidents (28.8%) involved neonates or babies. Of the total 1085 incidents, 200 (18.4%) were associated with tracheal intubation, 53 (4.9%) with tracheostomy and 893 (82.3%) were post‐procedure problems. One hundred and ten incidents (10.1%) were associated with more than temporary harm. Eighty‐eight intubation incidents were associated with equipment problems. Partial displacement of tubes resulted in more than temporary harm to the patient more frequently than complete tube displacement (15.7% vs 3.8%). Capnography was not described in any cases of displacement or blockage of tracheal or tracheostomy tubes. Recommendations concerning minimum standards for capnography, availability and checking of equipment and tracheostomy placement are made.
Influence of food intake on the bioavailability and efficacy of oral calcitonin
Authors: Karsdal Morten A. , Byrjalsen Inger , Azria Möise , Arnold Michel , Choi Les , Riis Bente J. , Christiansen Claus
Published: February 2009

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• No orally bioavailable peptides are currently approved by the US Food and Drug Administration or the European Medicines Agency.

• Attempts have been made to develop an oral formulation of salmon calcitonin (sCT).

• However, the effect of food intake on the bioavailability and efficacy of oral peptide formulations has not been systematically investigated.

WHAT THIS STUDY ADDS

• We provide evidence that oral doses of sCT may be optimally received in the fasting state or prior to meal intake.

• Fasting or preprandial dosing led to improved bioavailability and effect on pharmacodynamic biomarkers of efficacy.

• The results may aid other researchers involved in the formulation of oral peptides.

AIMS To investigate the influence of food intake on the bioavailability and pharmacodynamic effects of salmon calcitonin (sCT).

METHODS A single‐blind, randomized, partly placebo‐controlled study was conducted in 36 healthy postmenopausal female volunteers aged 62–74 years. The influence of food intake on oral dosing with 0.8 mg of sCT at 22.00 h was evaluated for a (i) predose meal at 18.00 h, (ii) predose meal at 20.00 h, (iii) predose meal at 21.00 h, (iv) postdose meal at 22.10 h, (v) no meal, and (vi) meal at 20.00 h and placebo at 22.00 h. Study biomarkers were plasma sCT levels and changes in the bone resorption marker CTX‐I (C‐terminal telopeptide of collagen type I).

RESULTS The predose meal at 18.00 and 21.00 h significantly decreased relative oral bioavailability of sCT to 26% [95% confidence interval (CI) 0.09, 0.73 and 0.09, 0.75, P= 0.009 and P= 0.01]. The meal consumed 10 min after dosing decreased the oral bioavailability of sCT to 59% (95% CI 0.21, 1.68), although nonsignificant (P= 0.48). This decreased bioavailability led to lower relative suppression of serum CTX‐I, with an AUC of the 4‐h efficacy response of −91%–×–hours for those receiving a meal at 18.00 h, compared with −238%–×–hours for fasting subjects. The Dunnett‐adjusted difference between these two treatment sequences was 147%–×–hours (95% CI 68, 225) (P= 0.0003). The AUC was comparable among fasting subjects and those consuming a meal 10 min after dosing.

CONCLUSIONS Postprandial dosing may limit the bioavailability of orally administered sCT. Maximal benefit can be achieved by dosing at least 10 min prior to meal time.
Metformin increases plasma ghrelin in Type 2 diabetes
Authors: Doogue Matthew P. , Begg Evan J. , Moore M. Peter , Lunt Helen , Pemberton Chris J. , Zhang Mei
Published: February 2009

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• Metformin, unlike the other major antihyperglycaemic drugs, is not associated with weight gain.

• Ghrelin is an appetite‐stimulating hormone whose concentrations vary in relation to food, obesity and diabetes control.

• Reports are conflicting about how metformin affects ghrelin concentrations, and this study was aimed at resolving this issue in patients with Type 2 diabetes.

WHAT THIS STUDY ADDS

• In this study an increase in ghrelin concentrations was seen in response to metformin treatment in patients with Type 2 diabetes.

• This effect was opposite to what might be expected if the effect of metformin on weight control was mediated via suppression of ghrelin.

• It is likely that the ghrelin response was secondary to improved glycaemic control.

• Meal time changes in appetite and satiety did not correlate with changes in ghrelin, which suggests ghrelin may not be important in meal initiation.

AIMS Metformin treatment of Type 2 diabetes is not usually associated with weight gain, and may assist with weight reduction. Plasma ghrelin concentrations are inversely associated with obesity and food intake. Metformin might therefore affect ghrelin concentrations, although previous studies have shown variable results in this regard. The primary aim of this study was to determine the effect of metformin on plasma ghrelin, appetite and satiety in patients with Type 2 diabetes.

METHODS Eighteen patients with Type 2 diabetes were studied before and after 6 weeks of metformin treatment, which was titrated to 1 g b.d. On the study days patients were fed standard meals of 390 kcal at 08.00 and 12.30 h, plasma samples were collected at 15‐ and 30‐min intervals, and appetite and satiety were measured on visual analogue scales. Changes in the area under the concentration–time curves (AUCs) of plasma ghrelin, insulin, glucose, appetite and satiety were assessed and examined for correlations with metformin AUCs. Changes in fasting adiponectin and leptin were also measured.

RESULTS Treatment with metformin increased the mean AUC (07.30–16.30 h) of plasma ghrelin by 24% (P= 0.003), while decreasing those of glucose by 19% (P < 0.001) and insulin by 19% (P= 0.001). No changes were detected in hunger and satiety, or in fasting adiponectin or leptin concentrations. There were no clear correlations between metformin plasma concentrations (AUC) and changes in plasma glucose, insulin or ghrelin.

CONCLUSIONS Treatment of Type 2 diabetes with metformin was associated with increased plasma ghrelin concentrations, without associated changes in hunger and satiety.
Pharmacokinetics and pharmacodynamics of LC15‐0444, a novel dipeptidyl peptidase IV inhibitor, after multiple dosing in healthy volunteers
Authors: Lim Kyoung Soo , Cho Joo‐Youn , Kim Bo‐Hyung , Kim Jung‐Ryul , Kim Hwa‐Sook , Kim Dong‐Kyu , Kim Sung‐Ho , Yim Hyeon Joo , Lee Sung‐Hack , Shin Sang‐Goo , Jang In‐Jin , Yu Kyung‐Sang
Published: February 2009

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• The importance of efficient drug development using biomarkers has been increasingly emphasized, from preclinical studies to clinical trials.

• However, as yet few ‘validated’ or ‘qualified’ biomarkers are used in early‐stage drug development in terms of clinical pharmacology and disease pathophysiology.

WHAT THIS STUDY ADDS

• This first‐time‐in‐human study provides evidence of the pharmacological activity of LC15‐0444 in humans, by using dipeptidyl peptidase IV activity and active glucagon‐like peptide‐1 concentrations.

• LC15‐0444 possesses pharmacokinetic and pharmacodynamic characteristics that support a once‐daily dosing regimen.

AIMS LC15‐0444 is a selective and competitive inhibitor of dipeptidyl peptidase (DPP) IV with potential for the treatment of Type 2 diabetes. The aim was to investigate the pharmacokinetic (PK) and pharmacodynamic (PD) profiles after multiple oral ascending doses of LC15‐0444 in healthy male subjects.

METHODS A dose block‐randomized, double‐blind, placebo‐controlled, parallel group study was performed in three groups with 10 subjects (eight for active drug; two for placebo) per group; each group received 200, 400 or 600 mg of LC15‐0444 once daily for 10 days. Blood and urine samples were collected up to 24 h after the first dosing and up to 72 h after the last dosing.

RESULTS The LC15‐0444 concentration–time profiles exhibited characteristics of multicompartment disposition. No dose‐ or time‐dependent change in PK parameters was observed. Mean elimination half‐life was in a range 16.6–20.1 h in the dose groups. Mean renal clearance and fraction of unchanged drug excreted in urine was 18.6–21.9 and 0.40–0.48 l h−1, respectively. In the steady state, mean accumulation ratios by dose groups were between 1.22 and 1.31. More than 80% inhibition of DPP IV activity from baseline was sustained for >24 h in all dose groups.

CONCLUSIONS This study provides evidence of the pharmacological activity of LC15‐0444 in humans. LC15‐0444 possesses PK and PD characteristics that support a once‐daily dosing regimen.