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Sample records for hybrid enrichment assay

  1. Automated UF6 Cylinder Enrichment Assay: Status of the Hybrid Enrichment Verification Array (HEVA) Project: POTAS Phase II

    Energy Technology Data Exchange (ETDEWEB)

    Jordan, David V.; Orton, Christopher R.; Mace, Emily K.; McDonald, Benjamin S.; Kulisek, Jonathan A.; Smith, Leon E.

    2012-06-01

    Pacific Northwest National Laboratory (PNNL) intends to automate the UF6 cylinder nondestructive assay (NDA) verification currently performed by the International Atomic Energy Agency (IAEA) at enrichment plants. PNNL is proposing the installation of a portal monitor at a key measurement point to positively identify each cylinder, measure its mass and enrichment, store the data along with operator inputs in a secure database, and maintain continuity of knowledge on measured cylinders until inspector arrival. This report summarizes the status of the research and development of an enrichment assay methodology supporting the cylinder verification concept. The enrichment assay approach exploits a hybrid of two passively-detected ionizing-radiation signatures: the traditional enrichment meter signature (186-keV photon peak area) and a non-traditional signature, manifested in the high-energy (3 to 8 MeV) gamma-ray continuum, generated by neutron emission from UF6. PNNL has designed, fabricated, and field-tested several prototype assay sensor packages in an effort to demonstrate proof-of-principle for the hybrid assay approach, quantify the expected assay precision for various categories of cylinder contents, and assess the potential for unsupervised deployment of the technology in a portal-monitor form factor. We refer to recent sensor-package prototypes as the Hybrid Enrichment Verification Array (HEVA). The report provides an overview of the assay signatures and summarizes the results of several HEVA field measurement campaigns on populations of Type 30B UF6 cylinders containing low-enriched uranium (LEU), natural uranium (NU), and depleted uranium (DU). Approaches to performance optimization of the assay technique via radiation transport modeling are briefly described, as are spectroscopic and data-analysis algorithms.

  2. Hybrid Enrichment Assay Methods for a UF6 Cylinder Verification Station: FY10 Progress Report

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Leon E.; Jordan, David V.; Orton, Christopher R.; Misner, Alex C.; Mace, Emily K.

    2010-08-01

    Pacific Northwest National Laboratory (PNNL) is developing the concept of an automated UF6 cylinder verification station that would be located at key measurement points to positively identify each cylinder, measure its mass and enrichment, store the collected data in a secure database, and maintain continuity of knowledge on measured cylinders until the arrival of International Atomic Energy Agency (IAEA) inspectors. At the center of this unattended system is a hybrid enrichment assay technique that combines the traditional enrichment-meter method (based on the 186 keV peak from 235U) with non-traditional neutron-induced high-energy gamma-ray signatures (spawned primarily by 234U alpha emissions and 19F(alpha, neutron) reactions). Previous work by PNNL provided proof-of-principle for the non-traditional signatures to support accurate, full-volume interrogation of the cylinder enrichment, thereby reducing the systematic uncertainties in enrichment assay due to UF6 heterogeneity and providing greater sensitivity to material substitution scenarios. The work described here builds on that preliminary evaluation of the non-traditional signatures, but focuses on a prototype field system utilizing NaI(Tl) and LaBr3(Ce) spectrometers, and enrichment analysis algorithms that integrate the traditional and non-traditional signatures. Results for the assay of Type-30B cylinders ranging from 0.2 to 4.95 wt% 235U, at an AREVA fuel fabrication plant in Richland, WA, are described for the following enrichment analysis methods: 1) traditional enrichment meter signature (186 keV peak) as calculated using a square-wave convolute (SWC) algorithm; 2) non-traditional high-energy gamma-ray signature that provides neutron detection without neutron detectors and 3) hybrid algorithm that merges the traditional and non-traditional signatures. Uncertainties for each method, relative to the declared enrichment for each cylinder, are calculated and compared to the uncertainties from an attended

  3. Hybrid Enrichment Verification Array: Module Characterization Studies

    Energy Technology Data Exchange (ETDEWEB)

    Zalavadia, Mital A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Smith, Leon E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); McDonald, Benjamin S. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Kulisek, Jonathan A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Mace, Emily K. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Deshmukh, Nikhil S. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2016-03-01

    The work presented in this report is focused on the characterization and refinement of the Hybrid Enrichment Verification Array (HEVA) approach, which combines the traditional 186-keV 235U signature with high-energy prompt gamma rays from neutron capture in the detector and surrounding collimator material, to determine the relative enrichment and 235U mass of the cylinder. The design of the HEVA modules (hardware and software) deployed in the current field trial builds on over seven years of study and evolution by PNNL, and consists of a ø3''×3'' NaI(Tl) scintillator coupled to an Osprey digital multi-channel analyzer tube base from Canberra. The core of the HEVA methodology, the high-energy prompt gamma-ray signature, serves as an indirect method for the measurement of total neutron emission from the cylinder. A method for measuring the intrinsic efficiency of this “non-traditional” neutron signature and the results from a benchmark experiment are presented. Also discussed are potential perturbing effects on the non-traditional signature, including short-lived activation of materials in the HEVA module. Modeling and empirical results are presented to demonstrate that such effects are expected to be negligible for the envisioned implementation scenario. In comparison to previous versions, the new design boosts the high-energy prompt gamma-ray signature, provides more flexible and effective collimation, and improves count-rate management via commercially available pulse-processing electronics with a special modification prompted by PNNL.

  4. Assay of the deuterium enrichment of water via acetylene.

    Science.gov (United States)

    Previs, S F; Hazey, J W; Diraison, F; Beylot, M; David, F; Brunengraber, H

    1996-06-01

    A technique is presented for measuring the 2H enrichment of water in biological samples when this enrichment is greater than 0.2%. The sample is reacted with calcium carbide to form acetylene gas, which is determined by gas chromatography electron impact ionization mass spectrometry. Ion-molecule reactions, resulting in proton abstraction, are minimized by lowering the electron ionization energy from the usual 70 eV to 45 eV. This technique is much more rapid and economical than the classical isotope ratio mass spectrometric assay of the enrichment of hydrogen gas derived from reduction of water.

  5. Hybridization Capture Using Short PCR Products Enriches Small Genomes by Capturing Flanking Sequences (CapFlank)

    DEFF Research Database (Denmark)

    Tsangaras, Kyriakos; Wales, Nathan; Sicheritz-Pontén, Thomas;

    2014-01-01

    Solution hybridization capture methods utilize biotinylated oligonucleotides as baits to enrich homologous sequences from next generation sequencing (NGS) libraries. Coupled with NGS, the method generates kilo to gigabases of high confidence consensus targeted sequence. However, in many experimen...

  6. Detection of Campylobacter jejuni and Campylobacter coli in foods by enrichment culture and polymerase chain reaction enzyme-linked immunosorbent assay.

    Science.gov (United States)

    Bolton, F J; Sails, A D; Fox, A J; Wareing, D R A; Greenway, D L A

    2002-05-01

    A polymerase chain reaction (PCR) assay based on a solution hybridization format with colorimetric end-point detection (PCR ELISA) was investigated for the specific detection of Campylobacter jejuni and Campylobacter coli in food samples following enrichment culture. One hundred fifteen samples of raw meat and offal (poultry, porcine, ovine, and bovine), raw shellfish, and artificially contaminated milk were enriched in blood-free Campylobacter Enrichment Broth for 48 h. Enrichment cultures were subcultured to Campylobacter blood-free selective agar plates, and presumptive isolates were identified by phenotypic methods. DNA was extracted from 1-ml aliquots of the enrichment cultures using a rapid extraction method, and the DNA was used as the template in a PCR ELISA. A comparison of the PCR ELISA with the enrichment culture and subculture to selective agar method showed that the results of 112 of the 115 samples tested were in agreement by both methods. Seventy-one of the various food samples were positive in the PCR ELISA, and 70 samples were positive by culture. The PCR ELISA had a sensitivity of 99% and a specificity of 96%, with a positive predictive value of 97% and a negative predictive value of 98%. The PCR ELISA is a rapid, sensitive, and specific method for the detection of C. jejuni and C. coli in foods following enrichment culture and significantly reduces the time required for their detection.

  7. Uncertainty Quantification in Application of the Enrichment Meter Principle for Nondestructive Assay of Special Nuclear Material

    Directory of Open Access Journals (Sweden)

    Tom Burr

    2015-01-01

    Full Text Available Nondestructive assay (NDA of special nuclear material (SNM is used in nonproliferation applications, including identification of SNM at border crossings, and quantifying SNM at safeguarded facilities. No assay method is complete without “error bars,” which provide one widely used way to express confidence in assay results. NDA specialists typically partition total uncertainty into “random” and “systematic” components so that, for example, an error bar can be developed for the SNM mass estimate in one item or for the total SNM mass estimate in multiple items. Uncertainty quantification (UQ for NDA has always been important, but greater rigor is needed and achievable using modern statistical methods. To this end, we describe the extent to which the guideline for expressing uncertainty in measurements (GUM can be used for NDA. Also, we describe possible extensions to the GUM by illustrating UQ challenges in NDA that it does not address, including calibration with errors in predictors, model error, and item-specific biases. A case study is presented using gamma spectra and applying the enrichment meter principle to estimate the 235U mass in an item. The case study illustrates how to update the ASTM international standard test method for application of the enrichment meter principle using gamma spectra.

  8. Quantitative rRNA-targeted solution-based hybridization assay using peptide nucleic acid molecular beacons.

    Science.gov (United States)

    Li, Xu; Morgenroth, Eberhard; Raskin, Lutgarde

    2008-12-01

    The potential of a solution-based hybridization assay using peptide nucleic acid (PNA) molecular beacon (MB) probes to quantify 16S rRNA of specific populations in RNA extracts of environmental samples was evaluated by designing PNA MB probes for the genera Dechloromonas and Dechlorosoma. In a kinetic study with 16S rRNA from pure cultures, the hybridization of PNA MB to target 16S rRNA exhibited a higher final hybridization signal and a lower apparent rate constant than the hybridizations to nontarget 16S rRNAs. A concentration of 10 mM NaCl in the hybridization buffer was found to be optimal for maximizing the difference between final hybridization signals from target and nontarget 16S rRNAs. Hybridization temperatures and formamide concentrations in hybridization buffers were optimized to minimize signals from hybridizations of PNA MB to nontarget 16S rRNAs. The detection limit of the PNA MB hybridization assay was determined to be 1.6 nM of 16S rRNA. To establish proof for the application of PNA MB hybridization assays in complex systems, target 16S rRNA from Dechlorosoma suillum was spiked at different levels to RNA isolated from an environmental (bioreactor) sample, and the PNA MB assay enabled effective quantification of the D. suillum RNA in this complex mixture. For another environmental sample, the quantitative results from the PNA MB hybridization assay were compared with those from clone libraries.

  9. Enriching Student Learning of Astronomy in Online Courses via Hybrid Texts

    Science.gov (United States)

    Montgomery, M.

    2016-01-01

    Hybrid texts such as Horizons: Exploring the Universe, Hybrid (with CengageNOW) and Universe, Hybrid (with CengageNOW) are designed for higher education learning of astronomy in undergraduate online courses. In these hybrid texts, quiz and test bank questions have been revised to minimize easy look-up of answers by students via the Internet and discussion threads have been re-designed to allow for student selection of learning and for personalized learning, for example. By establishing connections between the student and the course content, student learning is enriched, students spend more time learning the material, student copying of answers is minimized, and student social engagement on the subject matter is increased. In this presentation, we discuss how Hybrid texts in Astronomy can increase student learning in online courses.

  10. Detection of human papillomavirus DNA by the hybrid capture assay

    Directory of Open Access Journals (Sweden)

    Carvalho Maria Odete O.

    2003-01-01

    Full Text Available Human Papillomavirus (HPV infection is the main cause of cervical cancers and cervical intraepithelial neoplasias (CIN worldwide. Consequently, it would be useful to evaluate HPV testing to screen for cervical cancer. Recently developed, the second-generation Hybrid Capture (HCA II test is a non-radioactive, relatively rapid, liquid hybridization assay designed to detect 18 HPV types, divided into high and low-risk groups. We evaluated 1055 women for HPV infection with the HCA II test. Five hundred and ten (48.3% of these women had HPV infection; 60 (11.8% had low cancer-risk HPV DNA; 269 (52.7% had high-risk HPV types and 181 (35.5% had both groups. Hence, 450 women (88.2% in this HPV-infected group had at least one high risk HPV type, and were therefore considered to be at high risk for cancer. Among the group with Papanicolaou (Pap test results, the overall prevalence of HPV DNA was 58.4%. Significant differences in HPV infection of the cervix were detected between Pap I (normal smears and Pap IV (carcinomas (p<0.0001. Values of HPV viral load obtained for Pap I and SILs were significantly different, with an upward trend (p<0.0001, suggesting a positive correlation between high viral load values and risk of SIL. Because of the high costs of the HCA II test, its use for routine cervical mass screening cannot be recommended in poor countries. Nevertheless, it is a useful tool when combined with cytology, diagnosing high-risk infections in apparently normal tissues. Use of this technique could help reduce the risk of cancer.

  11. Hybrid Enrichment Verification Array: Module Characterization Studies Version 2

    Energy Technology Data Exchange (ETDEWEB)

    Zalavadia, Mital A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Smith, Leon E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); McDonald, Benjamin S. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Kulisek, Jonathan A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Mace, Emily K. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Deshmukh, Nikhil S. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2015-12-01

    The work presented in this report is focused on the characterization and refinement of the HEVA approach, which combines the traditional 186-keV 235U signature with high-energy prompt gamma rays from neutron capture in the detector and surrounding collimator material, to determine the relative enrichment and 235U mass of the cylinder. The design of the HEVA modules (hardware and software) deployed in the current field trial builds on over seven years of study and evolution by PNNL and consists of a ø3''×3'' NaI(Tl) scintillator coupled to an Osprey digital multi-channel analyzer tube base from Canberra. In comparison to previous versions, the new design boosts the high energy prompt gamma-ray signature, provides more flexible and effective collimation, and improves count-rate management via commercially available pulse-processing electronics with a special modification prompted by PNNL.

  12. PCR-Reverse Blot Hybridization Assay for Screening and Identification of Pathogens in Sepsis

    OpenAIRE

    Choi, Yeonim; Wang, Hye-young; Lee, Gyusang; Park, Soon-Deok; Jeon, Bo-Young; Uh, Young; Kim, Jong Bae; Lee, Hyeyoung

    2013-01-01

    Rapid and accurate identification of the pathogens involved in bloodstream infections is crucial for the prompt initiation of appropriate therapy, as this can decrease morbidity and mortality rates. A PCR-reverse blot hybridization assay for sepsis, the reverse blot hybridization assay (REBA) Sepsis-ID test, was developed; it uses pan-probes to distinguish Gram-positive and -negative bacteria and fungi. In addition, the assay was designed to identify bacteria and fungi using six genus-specifi...

  13. Real-time assays with molecular beacons and other fluorescent nucleic acid hybridization probes.

    Science.gov (United States)

    Marras, Salvatore A E; Tyagi, Sanjay; Kramer, Fred Russell

    2006-01-01

    A number of formats for nucleic acid hybridization have been developed to identify DNA and RNA sequences that are involved in cellular processes and that aid in the diagnosis of genetic and infectious diseases. The introduction of hybridization probes with interactive fluorophore pairs has enabled the development of homogeneous hybridization assays for the direct identification of nucleic acids. A change in the fluorescence of these probes indicates the presence of a target nucleic acid, and there is no need to separate unbound probes from hybridized probes. The advantages of homogeneous hybridization assays are their speed and simplicity. In addition, homogeneous assays can be combined with nucleic acid amplification, enabling the detection of rare target nucleic acids. These assays can be followed in real time, providing quantitative determination of target nucleic acids over a broad range of concentrations.

  14. Multilayer interparticle linking hybrid MOF-199 for noninvasive enrichment and analysis of plant hormone ethylene.

    Science.gov (United States)

    Zhang, Zhuomin; Huang, Yichun; Ding, Weiwei; Li, Gongke

    2014-04-01

    Ethylene, an important plant hormone, is of utmost importance during many developmental processes of plants. However, the efficient enrichment and analysis of trace ethylene still remains a challenge. A simple and mild multilayer interparticle linking strategy was proposed to fabricate a novel hybrid MOF-199 enrichment coating. Strong chemical interparticle linkages throughout the coating improved the durability and reproducibility of hybrid MOF-199 coating dramatically. This coating performed a significant extraction superiority of ethylene over commonly used commercial coatings, attributed to the multiple interactions including "molecular sieving effect", hydrogen bonding, open metal site interaction, and π-π affinity. The hybridization of multiwalled carbon nanotubes (MWCNTs) with MOF-199 further improved the enrichment capability and also acted as a hydrophobic "shield" to prevent the open metal sites of MOF-199 from being occupied by water molecules, which effectively improved the moisture-resistant property of MOF-199/CNTs coating. Finally, this novel enrichment method was successfully applied for the noninvasive analysis of trace ethylene, methanol, and ethanol from fruit samples with relatively high humidity. The low detection limit was 0.016 μg/L for ethylene. It was satisfactory that trace ethylene could be actually detected from fruit samples by this noninvasive method. Good recoveries of spiked grape, wampee, blueberry, and durian husk samples were obtained in the range of 90.0-114%, 79.4-88.6%, 78.5-86.8%, and 85.2-105% with the corresponding relative standard deviations of 4.8-9.8%, 6.9-8.9%, 3.8-8.1%, and 9.3-10.5% (n = 3), respectively.

  15. Sampling Enrichment toward Target Structures Using Hybrid Molecular Dynamics-Monte Carlo Simulations.

    Science.gov (United States)

    Yang, Kecheng; Różycki, Bartosz; Cui, Fengchao; Shi, Ce; Chen, Wenduo; Li, Yunqi

    2016-01-01

    Sampling enrichment toward a target state, an analogue of the improvement of sampling efficiency (SE), is critical in both the refinement of protein structures and the generation of near-native structure ensembles for the exploration of structure-function relationships. We developed a hybrid molecular dynamics (MD)-Monte Carlo (MC) approach to enrich the sampling toward the target structures. In this approach, the higher SE is achieved by perturbing the conventional MD simulations with a MC structure-acceptance judgment, which is based on the coincidence degree of small angle x-ray scattering (SAXS) intensity profiles between the simulation structures and the target structure. We found that the hybrid simulations could significantly improve SE by making the top-ranked models much closer to the target structures both in the secondary and tertiary structures. Specifically, for the 20 mono-residue peptides, when the initial structures had the root-mean-squared deviation (RMSD) from the target structure smaller than 7 Å, the hybrid MD-MC simulations afforded, on average, 0.83 Å and 1.73 Å in RMSD closer to the target than the parallel MD simulations at 310K and 370K, respectively. Meanwhile, the average SE values are also increased by 13.2% and 15.7%. The enrichment of sampling becomes more significant when the target states are gradually detectable in the MD-MC simulations in comparison with the parallel MD simulations, and provide >200% improvement in SE. We also performed a test of the hybrid MD-MC approach in the real protein system, the results showed that the SE for 3 out of 5 real proteins are improved. Overall, this work presents an efficient way of utilizing solution SAXS to improve protein structure prediction and refinement, as well as the generation of near native structures for function annotation.

  16. Small RNA in situ hybridization in Caenorhabditis elegans, combined with RNA-seq, identifies germline-enriched microRNAs.

    Science.gov (United States)

    McEwen, Tamara J; Yao, Qiuming; Yun, Sijung; Lee, Chin-Yung; Bennett, Karen L

    2016-10-15

    Over four hundred different microRNAs (miRNAs) have been identified in the genome of the model organism the nematode Caenorhabditis elegans. As the germline is dedicated to the preservation of each species, and almost half of all the cells in an adult nematode are germline, it is likely that regulatory miRNAs are important for germline development and maintenance. In C. elegans the miR35 family has strong maternal effects, contributing to normal embryogenesis and to adult fecundity. To determine whether any particular miRNAs are greatly enriched in the C. elegans germline we used RNA-seq to compare the miRNA populations in several germline-defective strains of adult C. elegans worms, including glp-4(germline proliferation-4), glh-1(germline helicase-1) and dcr-1(dicer-1). Statistical analyses of RNA-seq comparisons identified 13 miRNAs that are germline-enriched, including seven members of the well-studied miR35 family that were reduced as much as 1000-fold in TaqMan qRT PCR miRNA assays. Along with the miR35s, six others: miR-56 (a member of the miR51 family),-70, -244, -260 , -788 and -4813, none of which previously considered as such, were also identified by RNA-seq as germline-enriched candidates. We went on to develop a successful miRNA in situ hybridization protocol for C. elegans, revealing miR35s specifically concentrate during oogenesis in the pachytene region of the gonad, and persist throughout early embryogenesis, while in adult animals neither let-7 nor miR-228 has a germline-bias.

  17. Scoping study to expedite development of a field deployable and portable instrument for UF6 enrichment assay

    Energy Technology Data Exchange (ETDEWEB)

    Chan, George; Valentine, John D.; Russo, Richard E.

    2017-09-14

    The primary objective of the present study is to identity the most promising, viable technologies that are likely to culminate in an expedited development of the next-generation, field-deployable instrument for providing rapid, accurate, and precise enrichment assay of uranium hexafluoride (UF6). UF6 is typically involved, and is arguably the most important uranium compound, in uranium enrichment processes. As the first line of defense against proliferation, accurate analytical techniques to determine the uranium isotopic distribution in UF6 are critical for materials verification, accounting, and safeguards at enrichment plants. As nuclear fuel cycle technology becomes more prevalent around the world, international nuclear safeguards and interest in UF6 enrichment assay has been growing. At present, laboratory-based mass spectrometry (MS), which offers the highest attainable analytical accuracy and precision, is the technique of choice for the analysis of stable and long-lived isotopes. Currently, the International Atomic Energy Agency (IAEA) monitors the production of enriched UF6 at declared facilities by collecting a small amount (between 1 to 10 g) of gaseous UF6 into a sample bottle, which is then shipped under chain of custody to a central laboratory (IAEA’s Nuclear Materials Analysis Laboratory) for high-precision isotopic assay by MS. The logistics are cumbersome and new shipping regulations are making it more difficult to transport UF6. Furthermore, the analysis is costly, and results are not available for some time after sample collection. Hence, the IAEA is challenged to develop effective safeguards approaches at enrichment plants. In-field isotopic analysis of UF6 has the potential to substantially reduce the time, logistics and expense of sample handling. However, current laboratory-based MS techniques require too much infrastructure and operator expertise for field deployment and operation. As outlined in the IAEA Department of Safeguards Long

  18. Rapid detection of Shigella and enteroinvasive Escherichia coli in produce enrichments by a conventional multiplex PCR assay.

    Science.gov (United States)

    Binet, Rachel; Deer, Deanne M; Uhlfelder, Samantha J

    2014-06-01

    Faster detection of contaminated foods can prevent adulterated foods from being consumed and minimize the risk of an outbreak of foodborne illness. A sensitive molecular detection method is especially important for Shigella because ingestion of as few as 10 of these bacterial pathogens can cause disease. The objectives of this study were to compare the ability of four DNA extraction methods to detect Shigella in six types of produce, post-enrichment, and to evaluate a new and rapid conventional multiplex assay that targets the Shigella ipaH, virB and mxiC virulence genes. This assay can detect less than two Shigella cells in pure culture, even when the pathogen is mixed with background microflora, and it can also differentiate natural Shigella strains from a control strain and eliminate false positive results due to accidental laboratory contamination. The four DNA extraction methods (boiling, PrepMan Ultra [Applied Biosystems], InstaGene Matrix [Bio-Rad], DNeasy Tissue kit [Qiagen]) detected 1.6 × 10(3)Shigella CFU/ml post-enrichment, requiring ∼18 doublings to one cell in 25 g of produce pre-enrichment. Lower sensitivity was obtained, depending on produce type and extraction method. The InstaGene Matrix was the most consistent and sensitive and the multiplex assay accurately detected Shigella in less than 90 min, outperforming, to the best of our knowledge, molecular assays currently in place for this pathogen.

  19. Dorsal horn-enriched genes identified by DNA microarray, in situ hybridization and immunohistochemistry

    Directory of Open Access Journals (Sweden)

    Koblan Kenneth S

    2002-08-01

    Full Text Available Abstract Background Neurons in the dorsal spinal cord play important roles in nociception and pain. These neurons receive input from peripheral sensory neurons and then transmit the signals to the brain, as well as receive and integrate descending control signals from the brain. Many molecules important for pain transmission have been demonstrated to be localized to the dorsal horn of the spinal cord. Further understanding of the molecular interactions and signaling pathways in the dorsal horn neurons will require a better knowledge of the molecular neuroanatomy in the dorsal spinal cord. Results A large scale screening was conducted for genes with enriched expression in the dorsal spinal cord using DNA microarray and quantitative real-time PCR. In addition to genes known to be specifically expressed in the dorsal spinal cord, other neuropeptides, receptors, ion channels, and signaling molecules were also found enriched in the dorsal spinal cord. In situ hybridization and immunohistochemistry revealed the cellular expression of a subset of these genes. The regulation of a subset of the genes was also studied in the spinal nerve ligation (SNL neuropathic pain model. In general, we found that the genes that are enriched in the dorsal spinal cord were not among those found to be up-regulated in the spinal nerve ligation model of neuropathic pain. This study also provides a level of validation of the use of DNA microarrays in conjunction with our novel analysis algorithm (SAFER for the identification of differences in gene expression. Conclusion This study identified molecules that are enriched in the dorsal horn of the spinal cord and provided a molecular neuroanatomy in the spinal cord, which will aid in the understanding of the molecular mechanisms important in nociception and pain.

  20. Microinjection and Fluorescence In Situ Hybridization Assay for Studying mRNA Export in Mammalian Cells.

    Science.gov (United States)

    Wang, Ke; Shi, Min; Cheng, Hong

    2017-01-01

    Microinjection and Fluorescence in situ Hybridization (FISH) assay is a useful method for mRNA export studies, which can overcome the problems of traditional transfection in cells. Here, we describe the method of microinjection and FISH assay applied in investigation of mRNA export. By this method we can estimate the mRNA export kinetics, examining mRNA export in cells with low transfection efficiencies, and observing nuclear export of aberrant RNAs.

  1. Final Report Nucleic Acid System - Hybrid PCR and Multiplex Assay Project Phase 2

    Energy Technology Data Exchange (ETDEWEB)

    Koopman, R P; Langlois, R G; Nasarabadi, S; Benett, W J; Colston, B W; Johnson, D C; Brown, S B; Stratton, P L; Milanovich, F P

    2002-04-17

    This report covers phase 2 (year 2) of the Nucleic Acid System--Hybrid PCR and Multiplex Assay project. The objective of the project is to reduce to practice the detection and identification of biological warfare pathogens by the nucleic acid recognition technique of PCR (polymerase chain reaction) in a multiplex mode using flow cytometry. The Hybrid instrument consists of a flow-through PCR module capable of handling a multiplexed PCR assay, a hybridizing module capable of hybridizing multiplexed PCR amplicons and beads, and a flow cytometer module for bead-based identification, all controlled by a single computer. Multiplex immunoassay using bead-based Luminex flow cytometry is available, allowing rapid screening for many agents. PCR is highly specific and complements and verifies immunoassay. It can also be multiplexed and detection provided using the bead-based Luminex flow cytometer. This approach allows full access to the speed and 100-fold multiplex capability of flow cytometry for rapid screening as well as the accuracy and specificity of PCR. This project has two principal activities: (1) Design, build and test a prototype hybrid PCR/flow cytometer with the basic capabilities for rapid, broad spectrum detection and identification, and (2) Develop and evaluate multiplex flow analysis assay protocols and reagents for the simultaneous detection of PCR products. This project requires not only building operationally functional instrumentation but also developing the chemical assays for detection of priority pathogens. This involves development and evaluation of multiplex flow analysis assay protocols and reagents for the simultaneous detection of PCR products.

  2. Evaluation of a multi-marker immunomagnetic enrichment assay for the quantification of circulating melanoma cells

    Directory of Open Access Journals (Sweden)

    Freeman James B

    2012-09-01

    Full Text Available Abstract Background Circulating melanoma cells (CMCs are thought to be valuable in improving measures of prognosis in melanoma patients and may be a useful marker of residual disease to identify non-metastatic patients requiring adjuvant therapy. We investigated whether immunomagnetic enrichment targeting multiple markers allows more efficient enrichment of CMCs from patient peripheral blood than targeting a single marker. Furthermore, we aimed to determine whether the number of CMCs in patient blood was associated with disease stage. Methods We captured CMCs by targeting the melanoma associated markers MCSP and MCAM as well as the melanoma stem cell markers ABCB5 and CD271, both individually and in combination, by immunomagnetic enrichment. CMCs were enriched and quantified from the peripheral blood of 10 non-metastatic and 13 metastatic melanoma patients. Results Targeting all markers in combination resulted in the enrichment of more CMCs than when any individual marker was targeted (p  Conclusions Our results demonstrated that a combination of markers should be targeted for optimal isolation of CMCs. In addition, there are significantly more CMCs in metastatic patients compared with non-metastatic patients and therefore quantification of CMCs may prove to be a useful marker of disease progression.

  3. Evaluation of a fluorescent DNA hybridization assay for the detection of Neisseria gonorrhoeae.

    Science.gov (United States)

    Cano, R J; Palomares, J C; Torres, M J; Klem, R E

    1992-07-01

    This study evaluates a four-hour fluorescent DNA hybridization assay using both known bacterial isolates and clinical specimens. A biotinylated oligonucleotide probe from a sequence of the plasmid-encoded gene cppB was used. Hybrids were detected by addition of a streptavidin-alkaline phosphatase conjugate, followed by incubation for 30 min in a fluorescent substrate for alkaline phosphatase. The level of detection of the fluorescent assay was 0.1 pg of cryptic plasmid DNA or 200 cfu of the plasmid-containing strain NG 34/85 of Neisseria gonorrhoeae. A total of 119 reference strains of Neisseria gonorrhoeae and other related bacteria were tested for reactivity with the probe. All Neisseria gonorrhoeae strains, including eight plasmid-free strains, hybridized with the probe. Fluorescence ratios were 2.67 for plasmid-free strains and 3.85 for plasmid-containing strains. Of the heterologous microorganisms tested, only one of six strains of Neisseria cinerea gave a fluorescence ratio above the 2.0 cut-off value for positivity with the probe at a cell density of 1 x 10(4) cfu. The probe was also evaluated using clinical specimens from 100 patients attending a clinic for sexually transmitted diseases. The sensitivity of the assay was 100% while the specificity was 97.5%. Positive and negative predictive values were 91.2% and 100%, respectively. The fluorescent DNA hybridization assay for the detection of Neisseria gonorrhoeae described here thus appears to be a highly specific and sensitive assay.

  4. Bonding and bio-properties of hybrid laser/magnetron Cr-enriched DLC layers

    Energy Technology Data Exchange (ETDEWEB)

    Jelinek, Miroslav, E-mail: jelinek@fzu.cz [Institute of Physics, Academy of Sciences of the Czech Republic, v.v.i, Na Slovance 2, 18221 Prague (Czech Republic); Czech Technical University in Prague, Faculty of Biomedical Engineering, nam. Sitna 3105, 27201 Kladno (Czech Republic); Zemek, Josef [Institute of Physics, Academy of Sciences of the Czech Republic, v.v.i, Na Slovance 2, 18221 Prague (Czech Republic); Vandrovcová, Marta; Bačáková, Lucie [Institute of Physiology of the Czech Academy of Sciences of the Czech Republic, v.v.i., Videnska 1083, 14220 Prague 4 (Czech Republic); Kocourek, Tomáš; Remsa, Jan; Písařík, Petr [Institute of Physics, Academy of Sciences of the Czech Republic, v.v.i, Na Slovance 2, 18221 Prague (Czech Republic); Czech Technical University in Prague, Faculty of Biomedical Engineering, nam. Sitna 3105, 27201 Kladno (Czech Republic)

    2016-01-01

    Chromium-enriched diamond-like carbon (DLC) layers were prepared by a hybrid technology using a combination of pulsed laser deposition (PLD) and magnetron sputtering. XRD revealed no chromium peaks, indicating that the layers are mostly amorphous. Carbon (sp{sup 2} and sp{sup 3} bonds) and chromium bonds were determined by XPS from C 1s, O 1s, and Cr 2p photoelectron peaks. Depending on the deposition conditions, the concentration of Cr in DLC layers moved from zero to 10 at.% for as-received sample surfaces, and to about 31 at.% after mild sputter-cleaning by argon ion cluster beam. It should be noted that the most stable Cr{sup 3+} bonding state is in Cr{sub 2}O{sub 3} and Cr(OH){sub 3}, and that there is the toxic Cr{sup 6+} state in CrO{sub 3}. The surface content of hexavalent chromium in the Cr 2p3/2 spectra is rather low, but discernible. The population density of Saos-2 cells was the highest in samples containing higher concentrations of chromium 7.7 and 10 at.%. This means that higher concentrations of chromium supported the cell adhesion and proliferation. In addition, as revealed by a LIVE/DEAD viability/cytotoxicity kit, the cells on all Cr-containing samples maintained high viability (96 to 99%) on days 1 and 3 after seeding. However, this seemingly positive cell behavior could be associated with the risk of dedifferentiation and oncogenic transformation of cells. - Highlights: • DLC and chromium-enriched DLC layers were prepared by hybrid laser–magnetron deposition. • The content of chromium in DLC varied up to 10 at.% (31 at.% after ion beam sputtering). • The surface content of toxic hexavalent chromium in the Cr 2p3/2 spectra is rather low but discernible. • Higher concentrations of chromium supported the cell adhesion and proliferation. • Cells on all Cr-containing samples maintained high viability (96 to 99%).

  5. Assay of protein and peptide adducts of cholesterol ozonolysis products by hydrophobic and click enrichment methods.

    Science.gov (United States)

    Windsor, Katherine; Genaro-Mattos, Thiago C; Miyamoto, Sayuri; Stec, Donald F; Kim, Hye-Young H; Tallman, Keri A; Porter, Ned A

    2014-10-20

    Cholesterol undergoes ozonolysis to afford a variety of oxysterol products, including cholesterol-5,6-epoxide (CholEp) and the isomeric aldehydes secosterol A (seco A) and secosterol B (seco B). These oxysterols display numerous important biological activities, including protein adduction; however, much remains to be learned about the identity of the reactive species and the range of proteins modified by these oxysterols. Here, we synthesized alkynyl derivatives of cholesterol-derived oxysterols and employed a straightforward detection method to establish secosterols A and B as the most protein-reactive of the oxysterols tested. Model adduction studies with an amino acid, peptides, and proteins provide evidence for the potential role of secosterol dehydration products in protein adduction. Hydrophobic separation methods-Folch extraction and solid phase extraction (SPE)-were successfully applied to enrich oxysterol-adducted peptide species, and LC-MS/MS analysis of a model peptide-seco adduct revealed a unique fragmentation pattern (neutral loss of 390 Da) for that species. Coupling a hydrophobic enrichment method with proteomic analysis utilizing characteristic fragmentation patterns facilitates the identification of secosterol-modified peptides and proteins in an adducted protein. More broadly, these improved enrichment methods may give insight into the role of oxysterols and ozone exposure in the pathogenesis of a variety of diseases, including atherosclerosis, Alzheimer's disease, Parkinson's disease, and asthma.

  6. Quantitative data analysis methods for bead-based DNA hybridization assays using generic flow cytometry platforms.

    Science.gov (United States)

    Corrie, S R; Lawrie, G A; Battersby, B J; Ford, K; Rühmann, A; Koehler, K; Sabath, D E; Trau, M

    2008-05-01

    Bead-based assays are in demand for rapid genomic and proteomic assays for both research and clinical purposes. Standard quantitative procedures addressing raw data quality and analysis are required to ensure the data are consistent and reproducible across laboratories independent of flow platform. Quantitative procedures have been introduced spanning raw histogram analysis through to absolute target quantitation. These included models developed to estimate the absolute number of sample molecules bound per bead (Langmuir isotherm), relative quantitative comparisons (two-sided t-tests), and statistical analyses investigating the quality of raw fluorescence data. The absolute target quantitation method revealed a concentration range (below probe saturation) of Cy5-labeled synthetic cytokeratin 19 (K19) RNA of c.a. 1 x 10(4) to 500 x 10(4) molecules/bead, with a binding constant of c.a. 1.6 nM. Raw hybridization frequency histograms were observed to be highly reproducible across 10 triplex assay replicates and only three assay replicates were required to distinguish overlapping peaks representing small sequence mismatches. This study provides a quantitative scheme for determining the absolute target concentration in nucleic acid hybridization reactions and the equilibrium binding constants for individual probe/target pairs. It is envisaged that such studies will form the basis of standard analytical procedures for bead-based cytometry assays to ensure reproducibility in inter- and intra-platform comparisons of data between laboratories. (c) 2008 International Society for Advancement of Cytometry.

  7. Detection of Phaeocystis globosa using sandwich hybridization integrated with nuclease protection assay (NPA-SH)

    Institute of Scientific and Technical Information of China (English)

    ZHEN Yu; MI Tiezhu; YU Zhigang

    2008-01-01

    Phaeocystis globosa Scherffel is one of the common harmful algae species in coastal waters of the southeastern China. In this study, sandwich hybridization integrated with nuclease protection assay (NPA-SH) was used to qualitatively and quantitatively detect P. globosa. Results showed that this method had good applicability and validity in analyzing the samples from laboratory cultures and from fields. The linear regression equation for P. globosa was obtained, and the lowest detection number of cells was 1.8×104 cells. Statistics showed that there was no distinct difference between the results of detecting the microalgae by NPA-SH and traditional microscopy. This technique has good reliability, accuracy, and can give a remarkably high sample processing rate. Sandwich hybridization integrated with nuclease protection assay will provide an efficient alternative to microscopic method for monitoring and investigating the bloom of P. globosa.

  8. Construction of libraries enriched for sequence repeats and jumping clones, and hybridization selection for region-specific markers

    Energy Technology Data Exchange (ETDEWEB)

    Kandpal, R.P.; Kandpal, G.; Weissman, S.M. (Yale Univ. School of Medicine, New Haven, CT (United States))

    1994-01-04

    The authors describe a simple and rapid method for constructing small-insert genomic libraries highly enriched for dimeric, trimeric, and tetrameric nucleotide repeat motifs. The approach involves use of DNA inserts recovered by PCR amplification of a small-insert sonicated genomic phage library or by a single-primer PCR amplification of Mbo I-digested and adaptor-ligated genomic DNA. The genomic DNA inserts are heat denatured and hybridized to a biotinylated oligonucleotde. The biotinylated hybrids are retained on a Vectrex-avidin matrix and eluted specifically. The eluate is PCR amplified and cloned. More than 90% of the clones in a library enriched for (CA)[sub n] microsatellites with this approach contained clones with inserts containing CA repeats. They have also used this protocol for enrichment of (CAG)[sub n] and (AGAT)[sub n] sequence repeats and for Not I jumping clones. They have used the enriched libraries with an adaptation of the cDNA selection method to enrich for repeat motifs encoded in yeast artificial chromosomes.

  9. Edge-enriched, porous carbon-based, high energy density supercapacitors for hybrid electric vehicles.

    Science.gov (United States)

    Kim, Yong Jung; Yang, Cheol-Min; Park, Ki Chul; Kaneko, Katsumi; Kim, Yoong Ahm; Noguchi, Minoru; Fujino, Takeshi; Oyama, Shigeki; Endo, Morinobu

    2012-03-12

    Supercapacitors can store and deliver energy by a simple charge separation, and thus they could be an attractive option to meet transient high energy density in operating fuel cells and in electric and hybrid electric vehicles. To achieve such requirements, intensive studies have been carried out to improve the volumetric capacitance in supercapacitors using various types and forms of carbons including carbon nanotubes and graphenes. However, conventional porous carbons are not suitable for use as electrode material in supercapacitors for such high energy density applications. Here, we show that edge-enriched porous carbons are the best electrode material for high energy density supercapacitors to be used in vehicles as an auxiliary powertrain. Molten potassium hydroxide penetrates well-aligned graphene layers vertically and consequently generates both suitable pores that are easily accessible to the electrolyte and a large fraction of electrochemically active edge sites. We expect that our findings will motivate further research related to energy storage devices and also environmentally friendly electric vehicles. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. A combined enrichment and aptamer pulldown assay for Francisella tularensis detection in food and environmental matrices.

    Directory of Open Access Journals (Sweden)

    Elise A Lamont

    Full Text Available Francisella tularensis, a Gram-negative bacterium and causative agent of tularemia, is categorized as a Class A select agent by the Centers for Disease Control and Prevention due to its ease of dissemination and ability to cause disease. Oropharyngeal and gastrointestinal tularemia may occur due to ingestion of contaminated food and water. Despite the concern to public health, little research is focused on F. tularensis detection in food and environmental matrices. Current diagnostics rely on host responses and amplification of F. tularensis genetic elements via Polymerase Chain Reaction; however, both tools are limited by development of an antibody response and limit of detection, respectively. During our investigation to develop an improved culture medium to aid F. tularensis diagnostics, we found enhanced F. tularensis growth using the spent culture filtrate. Addition of the spent culture filtrate allowed for increased detection of F. tularensis in mixed cultures of food and environmental matrices. Ultraperformance liquid chromatography (UPLC/MS analysis identified several unique chemicals within the spent culture supernatant of which carnosine had a matching m/z ratio. Addition of 0.625 mg/mL of carnosine to conventional F. tularensis medium increased the growth of F. tularensis at low inoculums. In order to further enrich F. tularensis cells, we developed a DNA aptamer cocktail to physically separate F. tularensis from other bacteria present in food and environmental matrices. The combined enrichment steps resulted in a detection range of 1-106 CFU/mL (starting inoculums in both soil and lettuce backgrounds. We propose that the two-step enrichment process may be utilized for easy field diagnostics and subtyping of suspected F. tularensis contamination as well as a tool to aid in basic research of F. tularensis ecology.

  11. Ribosomal protein genes are highly enriched among genes with allele-specific expression in the interspecific F1 hybrid catfish.

    Science.gov (United States)

    Chen, Ailu; Wang, Ruijia; Liu, Shikai; Peatman, Eric; Sun, Luyang; Bao, Lisui; Jiang, Chen; Li, Chao; Li, Yun; Zeng, Qifan; Liu, Zhanjiang

    2016-06-01

    Interspecific hybrids provide a rich source for the analysis of allele-specific expression (ASE). In this work, we analyzed ASE in F1 hybrid catfish using RNA-Seq datasets. While the vast majority of genes were expressed with both alleles, 7-8 % SNPs exhibited significant differences in allele ratios of expression. Of the 66,251 and 177,841 SNPs identified from the datasets of the liver and gill, 5420 (8.2 %) and 13,390 (7.5 %) SNPs were identified as significant ASE-SNPs, respectively. With these SNPs, a total of 1519 and 3075 ASE-genes were identified. Gene Ontology analysis revealed that genes encoding cytoplasmic ribosomal proteins (RP) were highly enriched among ASE genes. Parent-of-origin was determined for 27 and 30 ASE RP genes in the liver and gill, respectively. The results indicated that genes from both channel catfish and blue catfish were involved in ASE. However, each RP gene appeared to be almost exclusively expressed from only one parent, indicating that ribosomes in the hybrid catfish were in the "hybrid" form. Overall representation of RP transcripts among the transcriptome appeared lower in the F1 hybrid catfish than in channel catfish or blue catfish, suggesting that the "hybrid" ribosomes may work more efficiently for translation in the F1 hybrid catfish.

  12. A novel SERRS sandwich-hybridization assay to detect specific DNA target.

    Directory of Open Access Journals (Sweden)

    Cécile Feuillie

    Full Text Available In this study, we have applied Surface Enhanced Resonance Raman Scattering (SERRS technology to the specific detection of DNA. We present an innovative SERRS sandwich-hybridization assay that allows specific DNA detection without any enzymatic amplification, such as is the case with Polymerase Chain Reaction (PCR. In some substrates, such as ancient or processed remains, enzymatic amplification fails due to DNA alteration (degradation, chemical modification or to the presence of inhibitors. Consequently, the development of a non-enzymatic method, allowing specific DNA detection, could avoid long, expensive and inconclusive amplification trials. Here, we report the proof of concept of a SERRS sandwich-hybridization assay that leads to the detection of a specific chamois DNA. This SERRS assay reveals its potential as a non-enzymatic alternative technology to DNA amplification methods (particularly the PCR method with several applications for species detection. As the amount and type of damage highly depend on the preservation conditions, the present SERRS assay would enlarge the range of samples suitable for DNA analysis and ultimately would provide exciting new opportunities for the investigation of ancient DNA in the fields of evolutionary biology and molecular ecology, and of altered DNA in food frauds detection and forensics.

  13. Fluorescence in situ hybridization in combination with the comet assay and micronucleus test in genetic toxicology

    Directory of Open Access Journals (Sweden)

    Hovhannisyan Galina G

    2010-09-01

    Full Text Available Abstract Comet assay and micronucleus (MN test are widely applied in genotoxicity testing and biomonitoring. While comet assay permits to measure direct DNA-strand breaking capacity of a tested agent MN test allows estimating the induced amount of chromosome and/or genome mutations. The potential of these two methods can be enhanced by the combination with fluorescence in situ hybridization (FISH techniques. FISH plus comet assay allows the recognition of targets of DNA damage and repairing directly. FISH combined with MN test is able to characterize the occurrence of different chromosomes in MN and to identify potential chromosomal targets of mutagenic substances. Thus, combination of FISH with the comet assay or MN test proved to be promising techniques for evaluation of the distribution of DNA and chromosome damage in the entire genome of individual cells. FISH technique also permits to study comet and MN formation, necessary for correct application of these methods. This paper reviews the relevant literature on advantages and limitations of Comet-FISH and MN-FISH assays application in genetic toxicology.

  14. Photoelectrochemical competitive DNA hybridization assay using semiconductor quantum dot conjugated oligonucleotides.

    Science.gov (United States)

    Baş, Deniz; Boyaci, Ismail Hakki

    2011-05-01

    A competitive DNA hybridization assay based on the photoelectrochemistry of the semiconductor quantum dot-single stranded DNA conjugates (QD-ssDNA) was developed. Hybridization of QD-ssDNA with the capture probe DNA immobilized on the indium-tin oxide electrodes enables photocurrent generation when the electrochemical cell was illuminated with a light source. Upon the competition between QD-ssDNA and single-stranded target DNA, the photocurrent response decreased with the increase in the target DNA concentration. A linear relationship between the photocurrent and the target DNA concentration was obtained (R(2) = 0.991). The selectivity of system towards the target DNA was also demonstrated using non-complementary sample.

  15. Fabrication of uniform DNA-conjugated hydrogel microparticles via replica molding for facile nucleic acid hybridization assays.

    Science.gov (United States)

    Lewis, Christina L; Choi, Chang-Hyung; Lin, Yan; Lee, Chang-Soo; Yi, Hyunmin

    2010-07-01

    We identify and investigate several critical parameters in the fabrication of single-stranded DNA conjugated poly(ethylene glycol) (PEG) microparticles based on replica molding (RM) for highly uniform and robust nucleic acid hybridization assays. The effects of PEG-diacrylate, probe DNA, and photoinitiator concentrations on the overall fluorescence and target DNA penetration depth upon hybridization are examined. Fluorescence and confocal microscopy results illustrate high conjugation capacity of the probe and target DNA, femtomole sensitivity, and sequence specificity. Combined, these findings demonstrate a significant step toward simple, robust, and scalable procedures to manufacture highly uniform and high-capacity hybridization assay particles in a well-controlled manner by exploiting many advantages that the batch processing-based RM technique offers. We envision that the results presented here may be readily applied to rapid and high-throughput hybridization assays for a wide variety of applications in bioprocess monitoring, food safety, and biological threat detection.

  16. Simultaneous Detection of Three Arboviruses Using a Triplex RT-PCR Enzyme Hybridization Assay

    Institute of Scientific and Technical Information of China (English)

    Dan Dong; Shi-hong Fu; Li-hua Wang; Zhi Lv; Tai-yuan Li; Guo-dong Liang

    2012-01-01

    Arboviruses represent a serious problem to public health and agriculture worldwide.Fast,accurate identification of the viral agents of arbovirus-associated disease is essential for epidemiological surveillance and laboratory investigation.We developed a cost-effective,rapid,and highly sensitive one-step "triplex RT-PCR enzyme hybridization"assay for simultaneous detections of Japanese Encephallitis virus (JEV,Flaviviridae)Getah virus (GETV,Togaviridae),and Tahyna virus (TAHV,Bunyaviridae) using three pairs of primers to amplify three target sequences in one RT-PCR reaction.The analytical sensitivity of this assay was 1 PFU/mL for JEV,10PFU/mL for GETV,and 10 PFU/mL for TAHV.This assay is significantly more rapid and less expensive than the traditional serological detection and single RT-PCR reaction methods.When “triplex RT-PCR enzyme hybridization” was applied to 29 cerebrospinal fluid(CSF)samples that were JEV-positive by normal RT-PCR assay,all samples were strongly positive for JEV,but negative for GETV and TAHV,demonstrating a good sensitivity,specificity,and performance at CSF specimen detection.

  17. Cross-reactivity profiles of hybrid capture II, cobas, and APTIMA human papillomavirus assays

    DEFF Research Database (Denmark)

    Preisler, Sarah Nørgaard; Rebolj, Matejka; Ejegod, Ditte Møller

    2016-01-01

    evaluated to what extent these can be explained by cross-reactivity, i.e. positive test results without evidence of high-risk HPV genotypes. The patterns of cross-reactivity have been thoroughly studied for hybrid capture II (HC2) but not yet for newer HPV assays although the manufacturers claimed...... no or limited frequency of cross-reactivity. In this independent study we evaluated the frequency of cross-reactivity for HC2, cobas, and APTIMA assays.METHODS:Consecutive routine cervical screening samples from 5022 Danish women, including 2859 from women attending primary screening, were tested with the three...... cytology and positive high-risk HPV test results were invited for repeated testing in 18 months.RESULTS:Cross-reactivity to low-risk genotypes was detected in 109 (2.2 %) out of 5022 samples on HC2, 62 (1.2 %) on cobas, and 35 (0.7 %) on APTIMA with only 10 of the samples cross-reacting on all 3 assays...

  18. Visual detection technique for efficient screening and isolation of Salmonella based on a novel enrichment assay using chromatography membrane.

    Science.gov (United States)

    Tang, F; Xiong, Y; Zhang, H; Wu, K; Xiang, Y; Shao, J-B; Ai, H-W; Xiang, Y-P; Zheng, X-L; Lv, J-R; Sun, H; Bao, L-S; Zhang, Z; Hu, H-B; Zhang, J-Y; Chen, L; Lu, J; Liu, W-Y; Mei, H; Ma, Y; Xu, C-F; Fang, A-Y; Gu, M; Xu, C-Y; Chen, Y; Chen, Z; Sun, Z-Y

    2016-03-01

    To detect Salmonella more efficiently and isolate strains more easily, a novel and simple detection method that uses an enrichment assay and two chromogenic reactions on a chromatography membrane was developed. Grade 3 chromatography paper is used as functionalized solid phase support (SPS), which contains specially optimized medium. One reaction for screening is based on the sulfate-reducing capacity of Salmonella. Hydrogen sulfide (H2S) generated by Salmonella reacts with ammonium ferric citrate to produce black colored ferrous sulfide. Another reaction is based on Salmonella C8 esterase that is unique for Enterobacteriaceae except Serratia and interacts with 4-methylumbelliferyl caprylate (MUCAP) to produce fluorescent umbelliferone, which is visible under ultraviolet light. A very low detection limit (10(1) CFU ml(-1)) for Salmonella was achieved on the background of 10(5) CFU ml(-1) Escherichia coli. More importantly, testing with more than 1,000 anal samples indicated that our method has a high positive detection rate and is relatively low cost, compared with the traditional culture-based method. It took only 1 day for the preliminary screening and 2 days to efficiently isolate the Salmonella cells, indicating that the new assay is specific, rapid, and simple for Salmonella detection. In contrast to the traditional culture-based method, this method can be easily used to screen and isolate targeted strains with the naked eye. The results of quantitative and comparative experiments showed that the visual detection technique is an efficient alternative method for the screening of Salmonella spp. in many applications of large-sized samples related to public health surveillance.

  19. A pilot study applying the plant Anchored Hybrid Enrichment method to New World sages (Salvia subgenus Calosphace; Lamiaceae).

    Science.gov (United States)

    Fragoso-Martínez, Itzi; Salazar, Gerardo A; Martínez-Gordillo, Martha; Magallón, Susana; Sánchez-Reyes, Luna; Moriarty Lemmon, Emily; Lemmon, Alan R; Sazatornil, Federico; Granados Mendoza, Carolina

    2017-02-09

    We conducted a pilot study using Anchored Hybrid Enrichment to resolve relationships among a mostly Neotropical sage lineage that may have undergone a recent evolutionary radiation. Conventional markers (ITS, trnL-trnF and trnH-psbA) have not been able to resolve the relationships among species nor within portions of the backbone of the lineage. We sampled 12 representative species of subgenus Calosphace and included one species of Salvia's s.l. closest relative, Lepechinia, as outgroup. Hybrid enrichment and sequencing were successful, yielding 448 alignments of individual loci with an average length of 704bp. The performance of the phylogenomic data in phylogenetic reconstruction was superior to that of conventional markers, increasing both support and resolution. Because the captured loci vary in the amount of net phylogenetic informativeness at different phylogenetic depths, these data are promising in phylogenetic reconstruction of this group and likely other lineages within Lamiales. However, special attention should be placed on the amount of phylogenetic noise that the data could potentially contain. A prior exploration step using phylogenetic informativeness profiles to detect loci with sites with disproportionately high substitution rates (showing "phantom" spikes) and, if required, the ensuing filtering of the problematic data is recommended. In our dataset, filtering resulted in increased support and resolution for the shallow nodes in maximum likelihood phylogenetic trees resulting from concatenated analyses of all the loci. Additionally, it is expected that an increase in sampling (loci and taxa) will aid in resolving weakly supported, short deep internal branches.

  20. Evaluation of a reverse-hybridization StripAssay for the detection of genetic polymorphisms leading to acenocoumarol sensitivity.

    Science.gov (United States)

    Gialeraki, Argyri; Markatos, Christos; Grouzi, Elisabeth; Merkouri, Efrosyni; Travlou, Anthi; Politou, Marianna

    2010-04-01

    Acenocoumarol is mainly catabolized by CYP2C9 isoform of cytochrome P450 (CYP) liver complex and exerts its anticoagulant effect through the inhibition of Vitamin K Epoxide Reductase (VKOR). The most important genetic polymorphisms which lead to an impaired enzymatic activity and therefore predispose to acenocoumarol sensitivity, are considered to be CYP2C9*2 (Arg144Cys), CYP2C9*3 (Ile359Leu) and VKORC1-1639G>A, respectively. In this study we compared the results of the PGXThrombo StripAssay kit (ViennaLab Diagnostics,Vienna, Austria) with direct DNA sequencing and in house Restriction Fragment Length Polymorphisms (RFLP) for the detection of the aforementioned Single Nucleotide Polymorphisms (SNPs). The reverse hybridization StripAssay was found to be equally effective with RFLP and direct DNA sequencing for the detection of CYP2C9*2 and CYP2C9*3 polymorphisms, respectively. The comparison of the RFLP reference method with the reverse hybridization StripAssay for the detection of VKORC1-1639 G>A polymorphism showed that the reverse hybridization StripAsssay might misclassify some A/A homozygotes as heterozygotes. Optimization of the hybridization procedures may eliminate the extra low signal band observed in some samples at the reverse hybridization StripAssay and improve its diagnostic value.

  1. Droplet Digital Enzyme-Linked Oligonucleotide Hybridization Assay for Absolute RNA Quantification

    Science.gov (United States)

    Guan, Weihua; Chen, Liben; Rane, Tushar D.; Wang, Tza-Huei

    2015-09-01

    We present a continuous-flow droplet-based digital Enzyme-Linked Oligonucleotide Hybridization Assay (droplet digital ELOHA) for sensitive detection and absolute quantification of RNA molecules. Droplet digital ELOHA incorporates direct hybridization and single enzyme reaction via the formation of single probe-RNA-probe (enzyme) complex on magnetic beads. It enables RNA detection without reverse transcription and PCR amplification processes. The magnetic beads are subsequently encapsulated into a large number of picoliter-sized droplets with enzyme substrates in a continuous-flow device. This device is capable of generating droplets at high-throughput. It also integrates in-line enzymatic incubation and detection of fluorescent products. Our droplet digital ELOHA is able to accurately quantify (differentiate 40% difference) as few as ~600 RNA molecules in a 1 mL sample (equivalent to 1 aM or lower) without molecular replication. The absolute quantification ability of droplet digital ELOHA is demonstrated with the analysis of clinical Neisseria gonorrhoeae 16S rRNA to show its potential value in real complex samples.

  2. Applying Subtractive Hybridization Technique to Enrich and Amplify Tumor-Specific Transcripts of Esophageal Squamous Cell Carcinoma.

    Science.gov (United States)

    Mahmoudian, Reihaneh Alsadat; Abbaszadegan, Mohammad Reza; Gholamin, Mehran

    2017-04-01

    Subtractive hybridization (SH) as an efficient and powerful approach can be applied to isolate differentially expressed transcripts as well as detect of involved mRNAs in various cellular processes, particularly diseases and malignancies. This procedure leads to the enrichment of specific low copy transcripts of tumor cells. Having developed a new approach for SH to isolate tumor specific transcripts, we facilitated discovery of uniquely expressed genes in esophageal squamous cell carcinoma (ESCC). Total RNA was extracted from the fresh tumoral and their adjacent normal tissues, and purified using the Switch Mechanism At the 5' end of Reverse Transcript (SMART) method. Following cDNA synthesis of normal mRNAs using magnetic beads, it was hybridized with tumor mRNAs. To enhance efficiency of subtraction, hybridization was repeated three rounds. Finally, amplification of subtracted tumor-specific transcripts was carried out using in vitro transcription. The subtracted tumoral mRNAs was analyzed quantitatively using real-time PCR for both tumor-specific and housekeeping genes. The subtracted mRNA was confirmed as tumor-specific mRNA pool using RT-PCR and quantitative real-time PCR assessment. The elevated level of tumor-specific transcripts such as MAGE-A4 and CD44 as well as declined copy number of housekeeping genes such as GAPDH, β actin and β2-microglobulin, were confirmed in subtracted tumoral mRNA. The presence of tumor genes was confirmed after the SH procedure. The designed SH method in combination with SMART technique can isolate and amplify high quality tumor-specific transcripts even from small amount of tumor tissues. Removal of common transcripts from the extracted tumoral mRNAs using SH, leads to the enrichment of tumor-specific transcripts. The isolated transcripts are of interest because of their probable roles in ESCC progression and development. In addition, these tumor-specific mRNAs can be applied for future vaccine cancer studies.

  3. Detection of Prorocentrum donghaiense using sandwich hybridization integrated with nuclease protection assay

    Institute of Scientific and Technical Information of China (English)

    CHEN Jie; ZHEN Yu; MI Tiezhu; YU Zhigang

    2009-01-01

    Prorocentrum donghaiense is an important harmful algae bloom (HAB) causing creature in China's seas, and the conventional visual detection can not cope with long-term monitoring and highthroughput sampling projects. An assay for P. donghaiense with sandwich hybridization integrated with nuclease protection assay (NPA-SH) was established. Tests with mixed samples and spiked field ones confirmed its good specificity and sensitivity. The cell number of P. donghaiense correlated well with the optical density, and the regression equation is y=4×10-6x+ 0.694 9, in which x is the cell number, and y is the optical density, with r2=0.953 5. These results show that the NPA-SH method has good feasibility in the detection of P. donghaiense. Results of NPA-SH and microscopy are excellent for each sample. The NPA-SH method was a simple way in quantitative detection of P. donghaiense, and the whole process could be finished in about six hours, which provided a new approach in high-throughput sampling and long-term monitoring of P. donghaiense.

  4. A LightCycler real-time PCR hybridization probe assay for detecting food-borne thermophilic Campylobacter

    DEFF Research Database (Denmark)

    Perelle, S.; Josefsen, Mathilde Hartmann; Hoorfar, Jeffrey

    2004-01-01

    Cycler real-time PCR assay (LC-PCR), which used fluorescent hybridization probes was developed. The test incorporated an internal amplification control co-amplified with the 16S rRNA gene of Campylobacter to monitor potential PCR inhibitors and ensure successful amplifications. The specificity study involving...

  5. Hybrid Capture 2 and cobas human papillomavirus assays perform similarly on SurePath samples from women with abnormalities

    DEFF Research Database (Denmark)

    Fornari, D; Rebolj, M; Bjerregaard, B

    2016-01-01

    OBJECTIVE: In two laboratories (Departments of Pathology, Copenhagen University Hospitals of Herlev and Hvidovre), we compared cobas and Hybrid Capture 2 (HC2) human papillomavirus (HPV) assays using SurePath® samples from women with atypical squamous cells of undetermined significance (ASCUS) at...

  6. Preparation of hydrazine functionalized polymer brushes hybrid magnetic nanoparticles for highly specific enrichment of glycopeptides.

    Science.gov (United States)

    Huang, Guang; Sun, Zhen; Qin, Hongqiang; Zhao, Liang; Xiong, Zhichao; Peng, Xiaojun; Ou, Junjie; Zou, Hanfa

    2014-05-07

    Hydrazide chemistry is a powerful technique in glycopeptides enrichment. However, the low density of the monolayer hydrazine groups on the conventional hydrazine-functionalized magnetic nanoparticles limits the efficiency of glycopeptides enrichment. Herein, a novel magnetic nanoparticle grafted with poly(glycidyl methacrylate) (GMA) brushes was fabricated via reversible addition-fragmentation chain transfer (RAFT) polymerization, and a large amount of hydrazine groups were further introduced to the GMA brushes by ring-opening the epoxy groups with hydrazine hydrate. The resulting magnetic nanoparticles (denoted as Fe3O4@SiO2@GMA-NHNH2) demonstrated the high specificity of capturing glycopeptides from a tryptic digest of the sample comprising a standard non-glycosylated protein bovine serum albumin (BSA) and four standard glycoproteins with a weight ratio of 50 : 1, and the detection limit was as low as 130 fmol. In the analysis of a real complex biological sample, the tryptic digest of hepatocellular carcinoma, 179 glycosites were identified by the Fe3O4@SiO2@GMA-NHNH2 nanoparticles, surpassing that of 68 glycosites by Fe3O4@SiO2-single-NHNH2 (with monolayer hydrazine groups on the surface). It can be expected that the magnetic nanoparticles modified with hydrazine functionalized polymer brushes via RAFT technique will improve the specificity and the binding capacity of glycopeptides from complex samples, and show great potential in the analysis of protein glycosylation in biological samples.

  7. Detection of human papillomavirus in pterygium and conjunctival papilloma by hybrid capture II and PCR assays.

    Science.gov (United States)

    Takamura, Y; Kubo, E; Tsuzuki, S; Akagi, Y

    2008-11-01

    To elucidate the putative role of human papillomavirus (HPV) infection in pterygium and conjunctival papilloma. Hybrid capture II (HC-II) and polymerase chain reaction (PCR) assays were performed to detect HPV in pterygium (42 samples obtained from 40 patients) and conjunctival papilloma (8 samples from 6 patients). The amount of HPV DNA was evaluated by measurement of relative light units (RLUs) on a luminometer. All papilloma samples were positive for HPV DNA by PCR and HC-II. The RLU values for specimens of recurrent and re-recurrent papilloma were markedly higher than those for specimens of primary lesions. HPV was detected by PCR in 2 of 42 (4.8%) beta-globin-positive pterygium specimens, whereas HC-II showed that HPV was negative in all pterygium samples. Our results support the hypothesis that HPV DNA is associated with the pathogenesis of conjunctival papilloma, but not pterygium. RLU measurement by HC-II may serve as a marker for evaluating the activity of HPV in conjunctival tumours.

  8. A hybrid neural network structure for application to nondestructive TRU waste assay

    Energy Technology Data Exchange (ETDEWEB)

    Becker, G. [Idaho National Engineering Lab., Idaho Falls, ID (United States)

    1995-12-31

    The determination of transuranic (TRU) and associated radioactive material quantities entrained in waste forms is a necessary component. of waste characterization. Measurement performance requirements are specified in the National TRU Waste Characterization Program quality assurance plan for which compliance must be demonstrated prior to the transportation and disposition of wastes. With respect to this criterion, the existing TRU nondestructive waste assay (NDA) capability is inadequate for a significant fraction of the US Department of Energy (DOE) complex waste inventory. This is a result of the general application of safeguard-type measurement and calibration schemes to waste form configurations. Incompatibilities between such measurement methods and actual waste form configurations complicate regulation compliance demonstration processes and illustrate the need for an alternate measurement interpretation paradigm. Hence, it appears necessary to supplement or perhaps restructure the perceived solution and approach to the waste NDA problem. The first step is to understand the magnitude of the waste matrix/source attribute space associated with those waste form configurations in inventory and how this creates complexities and unknowns with respect to existing NDA methods. Once defined and/or bounded, a conceptual method must be developed that specifies the necessary tools and the framework in which the tools are used. A promising framework is a hybridized neural network structure. Discussed are some typical complications associated with conventional waste NDA techniques and how improvements can be obtained through the application of neural networks.

  9. Comparison of kDNA PCR-hybridization assay with three PCR methods for canines visceral Leishmaniasis diagnosis

    Energy Technology Data Exchange (ETDEWEB)

    Pilatti, Marcia M.; Andrade, Antero S.R. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil)], e-mail: marciapilatti@yahoo.com.br, e-mail: antero@cdtn.br; Ferreira, Sidney A. [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Parasitologia], e-mail: saninoalmeida@gmail.com

    2009-07-01

    The sensitivity of the kDNA PCR-Hybridization assay, which uses radioactive DNA probes (labeled with {sup 32}P), was compared with three conventional PCR methods used for canine visceral leishmaniasis diagnosis. All PCR methods had two steps: a first amplification followed by hybridization or by a new amplification (nested or semi nested). Two methods (kDNA PCR-Hybridization and kDNA snPCR) used primers addressed to kinetoplast minicircles and the other two methods to the coding (LnPCR) and intergenic noncoding regions (ITS-1 nPCR) of the ribosomal rRNA genes. The comparison was accomplished in two groups of 23 infected dogs using samples collected by the conjunctival swab procedure. In the Group 1 the DNA was extracted from cotton swabs by phenol-chloroform and in Group 2 by boiling. The most efficient PCR methods in the Group 1 were those based on kDNA targets. The kDNA PCR-Hybridization was able to detect parasites in 22/23 dogs (95.6%) and in 40/46 samples (86.9%). The kDNA snPCR was positive for 21/23 dogs (91.3%) and for 40/46 samples (86.9%). The positivities of the kDNA based methods were significantly higher than the positivities verified for the methods based on ribosomal rRNA genes (p<0.05). In the Group 2 the kDNA PCR- Hybridization showed a better performance detecting parasites in 18/23 dogs (78.3%) and in 31/46 samples (67.4%), significantly higher than the other three methods (p<0.05). The higher sensitivity of the minicircle kDNA based assays reported by others was confirmed in this study and kDNA PCR-Hybridization showed the best sensitivity among the assays evaluated. (author)

  10. Investigation of parameters that affect the success rate of microarray-based allele-specific hybridization assays.

    Directory of Open Access Journals (Sweden)

    Lena Poulsen

    Full Text Available BACKGROUND: The development of microarray-based genetic tests for diseases that are caused by known mutations is becoming increasingly important. The key obstacle to developing functional genotyping assays is that such mutations need to be genotyped regardless of their location in genomic regions. These regions include large variations in G+C content, and structural features like hairpins. METHODS/FINDINGS: We describe a rational, stable method for screening and combining assay conditions for the genetic analysis of 42 Phenylketonuria-associated mutations in the phenylalanine hydroxylase gene. The mutations are located in regions with large variations in G+C content (20-75%. Custom-made microarrays with different lengths of complementary probe sequences and spacers were hybridized with pooled PCR products of 12 exons from each of 38 individual patient DNA samples. The arrays were washed with eight buffers with different stringencies in a custom-made microfluidic system. The data were used to assess which parameters play significant roles in assay development. CONCLUSIONS: Several assay development methods found suitable probes and assay conditions for a functional test for all investigated mutation sites. Probe length, probe spacer length, and assay stringency sufficed as variable parameters in the search for a functional multiplex assay. We discuss the optimal assay development methods for several different scenarios.

  11. Prevalence of human papillomavirus infection in the genital tract determined by hybrid capture assay

    Directory of Open Access Journals (Sweden)

    Fernanda N. Carestiato

    Full Text Available Human Papillomavirus (HPV infection is the most prevalent sexually-transmitted virus worldwide. It is known to be the etiological agent of cervical cancer and cervical intraepithelial neoplasia (CIN. Consequently, there is strong motivation to evaluate HPV testing in cervical cancer screening. Recently developed, the second generation of the hybrid capture test (HCA II is a non-radioactive, relatively rapid, hybridization assay, designed to detect 18 HPV types divided into high and low-risk groups. We evaluated 7,314 patients (5,833 women and 1,481 men for HPV infection by HCA II. Among them, 3,008 (41.1% presented HPV infection: 430 (14.2% had HPV DNA of low risk for cancer, 1,631 (54.2% had high risk HPV types and 947 (31.5% had both types. The prevalence in females was 44.9%. The prevalence of HPV DNA in the group for which cytological results were available was slightly higher: 55.3% (1007/1824. Significant differences were detected in the frequency of HPV infection of the cervix between normal cases and those with high-grade squamous-intraepithelial lesions (HSIL(P<0.0001. Among males, the prevalence was 26.2%, composed of 9.1% in Group A, 9.7% in Group B and 7.4% with multiple infections. We observed that male prevalence was lower and that low-risk types were more frequent than in females. HPV viral load was significantly greater in SILs than in normal or inflammatory cases (P<0.0001, suggesting an association between high viral load values and risk of SIL. Because of high costs, the HCA II test cannot be recommended for routine mass screening for cervical infection in poor countries. Nevertheless, it was found to be a useful tool, when combined with cytology, discovering high-risk infections in apparently normal tissues and revealing silent infections that may be responsible for the maintenance of HPV in the general population. These findings point to the need for close and careful management of patients, thereby reducing overtreatment

  12. A pre-breeding screening program for transgenic boars based on fluorescence in situ hybridization assay.

    Science.gov (United States)

    Bou, Gerelchimeg; Sun, Mingju; Lv, Ming; Zhu, Jiang; Li, Hui; Wang, Juan; Li, Lu; Liu, Zhongfeng; Zheng, Zhong; He, Wenteng; Kong, Qingran; Liu, Zhonghua

    2014-08-01

    For efficient transgenic herd expansion, only the transgenic animals that possess the ability to transmit transgene into next generation are considered for breeding. However, for transgenic pig, practically lacking a pre-breeding screening program, time, labor and money is always wasted to maintain non-transgenic pigs, low or null transgenic transmission pigs and the related fruitless gestations. Developing a pre-breeding screening program would make the transgenic herd expansion more economical and efficient. In this technical report, we proposed a three-step pre-breeding screening program for transgenic boars simply through combining the fluorescence in situ hybridization (FISH) assay with the common pre-breeding screening workflow. In the first step of screening, combined with general transgenic phenotype analysis, FISH is used to identify transgenic boars. In the second step of screening, combined with conventional semen test, FISH is used to detect transgenic sperm, thus to identify the individuals producing high quality semen and transgenic sperm. In the third step of screening, FISH is used to assess the in vitro fertilization embryos, thus finally to identify the individuals with the ability to produce transgenic embryos. By this three-step screening, the non-transgenic boars and boars with no ability to produce transgenic sperm or transgenic embryos would be eliminated; therefore only those boars could produce transgenic offspring are maintained and used for breeding and herd expansion. It is the first time a systematic pre-breeding screening program is proposed for transgenic pigs. This program might also be applied in other transgenic large animals, and provide an economical and efficient strategy for herd expansion.

  13. Estrogenic/Antiestrogenic Activities of Polycyclic Aromatic Hydrocarbons and Their Monohydroxylated Derivatives by Yeast Two-Hybrid Assay

    OpenAIRE

    Hayakawa, Kazuichi; Onoda, Yu; Tachikawa, Chihiro; Hosoi, Shinzo; Yoshita, Morio; Chung, Sang Woon; Kizu, Ryoichi; Toriba, Akira; Kameda, Takayuki; Tang, Ning

    2007-01-01

    Estrogenic/antiestrogenic activities of 14 polycyclic aromatic hydrocarbons (PAHs) and 63 monohydroxylated PAHs (OHPAHs) having 2 to 6 rings were evaluated by yeast two-hybrid assay expressing human estrogen receptor α. Relative effective potencies of estrogenic and antiestrogenic activities were calculated as the inverse values of the relative concentration of the test compound that gave the same activities of E2 and 4-hydroxytamoxifen, respectively. PAHs did not show any estrogenic/antiestr...

  14. Antioxidant activity of yoghurt peptides: Part 1-in vitro assays and evaluation in omega-3 enriched milk

    DEFF Research Database (Denmark)

    Farvin, Sabeena; Baron, Caroline; Nina Skall, Nielsen

    2010-01-01

    The aim of the present study was to investigate important factors contributing to the high oxidative stability of fish-oil-enriched yoghurt, with particular emphasis on the possible antioxidative effects of peptides released during yoghurt fermentation. Yoghurt samples were stripped from sugars...

  15. Targeted capture enrichment assay for non-invasive prenatal testing of large and small size sub-chromosomal deletions and duplications

    Science.gov (United States)

    Tsangaras, Kyriakos; Kypri, Elena; Loizides, Charalambos; Ioannides, Marios; Achilleos, Achilleas; Mina, Petros; Keravnou, Anna; Sismani, Carolina; Koumbaris, George; Patsalis, Philippos C.

    2017-01-01

    Noninvasive prenatal testing (NIPT) using whole genome and targeted sequencing has become increasingly accepted for clinical detection of Trisomy 21 and sex chromosome aneuploidies. Few studies have shown that sub-chromosomal deletions or duplications associated with genetic syndromes can also be detected in the fetus noninvasively. There are still limitations on these methodologies such as the detection of variants of unknown clinical significance, high number of false positives, and difficulties to detect small aberrations. We utilized a recently developed targeted sequencing approach for the development of a NIPT assay, for large and small size deletions/duplications, which overcomes these existing limitations. Artificial pregnancies with microdeletion/microduplication syndromes were created by spiking DNA from affected samples into cell free DNA (cfDNA) from non-pregnant samples. Unaffected spiked samples and normal pregnancies were used as controls. Target Capture Sequences (TACS) for seven syndromes were designed and utilized for targeted capture enrichment followed by sequencing. Data was analyzed using a statistical pipeline to identify deletions or duplications on targeted regions. Following the assay development a proof of concept study using 33 normal pregnancies, 21 artificial affected and 17 artificial unaffected pregnancies was carried out to test the sensitivity and specificity of the assay. All 21 abnormal spiked-in samples were correctly classified as subchromosomal aneuploidies while the 33 normal pregnancies or 17 normal spiked-in samples resulted in a false positive result. We have developed an NIPT assay for the detection of sub-chromosomal deletions and duplications using the targeted capture enrichment technology. This assay demonstrates high accuracy, high read depth of the genomic region of interest, and can identify deletions/duplications as small as 0.5 Mb. NIPT of fetal microdeletion/microduplication syndromes can be of enormous benefit

  16. Detection of oligonucleotide hybridization on a single microparticle by time-resolved fluorometry: hybridization assays on polymer particles obtained by direct solid phase assembly of the oligonucleotide probes.

    Science.gov (United States)

    Hakala, H; Heinonen, P; Iitiä, A; Lönnberg, H

    1997-01-01

    Oligodeoxyribonucleotides were assembled by conventional phosphoramidite chemistry on uniformly sized (50 microns) porous glycidyl methacrylate/ethylene dimethacrylate (SINTEF) and compact polystyrene (Dynosphere) particles, the aminoalkyl side chains of which were further derivatized with DMTrO-acetyl groups. The linker was completely resistant toward ammonolytic deprotection of the base moieties. The quality of oligonucleotides was assessed by repeating the synthesis on the same particles derivatized with a cleavable ester linker. The ability of the oligonucleotide-coated particles to bind complementary sequences via hybridization was examined by following the attachment of oligonucleotides bearing a photoluminescent europium(III) chelate to the particles. The fluorescence emission was measured directly on a single particle. The effects of the following factors on the kinetics and efficiency of hybridization were studied: number of particles in a given volume of the assay solution, loading of oligonucleotide on the particle, concentration of the target oligonucleotide in solution, length of the hybridizing sequence, presence of noncomplementary sequences, and ionic strength. The fluorescence signal measured on a single particle after hybridization was observed to be proportional to the concentration of the target oligonucleotide in solution over a concentration range of 5 orders of magnitude.

  17. Uncovering Arabidopsis membrane protein interactome enriched in transporters using mating-based split ubiquitin assays and classification models

    Directory of Open Access Journals (Sweden)

    Jin eChen

    2012-06-01

    Full Text Available High-throughput data are a double-edged sword; for the benefit of large amount of data, there is an associated cost of noise. To increase reliability and scalability of high-throughput protein interaction data generation, we tested the efficacy of classification to enrich potential protein-protein interactions (pPPIs. We applied this method to identify interactions among Arabidopsis membrane proteins enriched in transporters. We validated our method with multiple retests. Classification improved the quality of the ensuing interaction network and was effective in reducing the search space and increasing true positive rate. The final network of 541 interactions among 239 proteins (of which 179 are transporters is the first protein interaction network enriched in membrane transporters reported for any organism. This network has similar topological attributes to other published protein interaction networks. It also extends and fills gaps in currently available biological networks in plants and allows building a number of hypotheses about processes and mechanisms involving signal-transduction and transport systems.

  18. Uncovering Arabidopsis Membrane Protein Interactome Enriched in Transporters Using Mating-Based Split Ubiquitin Assays and Classification Models

    Science.gov (United States)

    Chen, Jin; Lalonde, Sylvie; Obrdlik, Petr; Noorani Vatani, Azam; Parsa, Saman A.; Vilarino, Cristina; Revuelta, Jose Luis; Frommer, Wolf B.; Rhee, Seung Y.

    2012-01-01

    High-throughput data are a double-edged sword; for the benefit of large amount of data, there is an associated cost of noise. To increase reliability and scalability of high-throughput protein interaction data generation, we tested the efficacy of classification to enrich potential protein–protein interactions. We applied this method to identify interactions among Arabidopsis membrane proteins enriched in transporters. We validated our method with multiple retests. Classification improved the quality of the ensuing interaction network and was effective in reducing the search space and increasing true positive rate. The final network of 541 interactions among 239 proteins (of which 179 are transporters) is the first protein interaction network enriched in membrane transporters reported for any organism. This network has similar topological attributes to other published protein interaction networks. It also extends and fills gaps in currently available biological networks in plants and allows building a number of hypotheses about processes and mechanisms involving signal-transduction and transport systems. PMID:22737156

  19. Nucleic Acid Sandwich Hybridization Assay with Quantum Dot-Induced Fluorescence Resonance Energy Transfer for Pathogen Detection

    Directory of Open Access Journals (Sweden)

    Cheng-Chung Chou

    2012-12-01

    Full Text Available This paper reports a nucleic acid sandwich hybridization assay with a quantum dot (QD-induced fluorescence resonance energy transfer (FRET reporter system. Two label-free hemagglutinin H5 sequences (60-mer DNA and 630-nt cDNA fragment of avian influenza viruses were used as the targets in this work. Two oligonucleotides (16 mers and 18 mers that specifically recognize two separate but neighboring regions of the H5 sequences were served as the capturing and reporter probes, respectively. The capturing probe was conjugated to QD655 (donor in a molar ratio of 10:1 (probe-to-QD, and the reporter probe was labeled with Alexa Fluor 660 dye (acceptor during synthesis. The sandwich hybridization assay was done in a 20 μL transparent, adhesive frame-confined microchamber on a disposable, temperature-adjustable indium tin oxide (ITO glass slide. The FRET signal in response to the sandwich hybridization was monitored by a homemade optical sensor comprising a single 400 nm UV light-emitting diode (LED, optical fibers, and a miniature 16-bit spectrophotometer. The target with a concentration ranging from 0.5 nM to 1 μM was successfully correlated with both QD emission decrease at 653 nm and dye emission increase at 690 nm. To sum up, this work is beneficial for developing a portable QD-based nucleic acid sensor for on-site pathogen detection.

  20. Simultaneous analysis of folic acid and pantothenic acid in foods enriched with vitamins by stable isotope dilution assays

    Energy Technology Data Exchange (ETDEWEB)

    Rychlik, Michael

    2003-10-24

    Folic and pantothenic acid were quantified in multivitamin products by stable isotope dilution assays using [{sup 2}H{sub 4}]folic acid and [{sup 13}C{sub 3},{sup 15}N]pantothenic acid as the internal standards. Detection was achieved by liquid chromatography/mass spectrometry which enabled unequivocal determination of the vitamins. Due to the very simple extraction procedure, analysis of the vitamins was completed within 2 h. When analyzing multivitamin sweets, the intra-assay and inter-assay coefficient of variation was 3.2% (n=5) and 3.1% (n=5) for folic acid and 4.5% (n=5) as well as 6.5% (n=7) for pantothenic acid, respectively. Along with the precision data, recovery values of 99.4% for folic acid and 103% for pantothenic acid at addition levels of 6 mg/kg and 600 {mu}g/kg, respectively, to starch products proved the accuracy of the new method. Application of the stable isotope dilution assay to fruit juices, whey products, cereals, sweets, pharmaceuticals, wheat flour and salt fortified with one or both vitamins revealed that for the majority of products the labeled pantothenic acid contents were exceeded by about 30%, whereas for folic acid also significantly lower contents than the label claim were found.

  1. Multilevel correlations in the biological phosphorus removal process: From bacterial enrichment to conductivity-based metabolic batch tests and polyphosphatase assays.

    Science.gov (United States)

    Weissbrodt, David G; Maillard, Julien; Brovelli, Alessandro; Chabrelie, Alexandre; May, Jonathan; Holliger, Christof

    2014-12-01

    Enhanced biological phosphorus removal (EBPR) from wastewater relies on the preferential selection of active polyphosphate-accumulating organisms (PAO) in the underlying bacterial community continuum. Efficient management of the bacterial resource requires understanding of population dynamics as well as availability of bioanalytical methods for rapid and regular assessment of relative abundances of active PAOs and their glycogen-accumulating competitors (GAO). A systems approach was adopted here toward the investigation of multilevel correlations from the EBPR bioprocess to the bacterial community, metabolic, and enzymatic levels. Two anaerobic-aerobic sequencing-batch reactors were operated to enrich activated sludge in PAOs and GAOs affiliating with "Candidati Accumulibacter and Competibacter phosphates", respectively. Bacterial selection was optimized by dynamic control of the organic loading rate and the anaerobic contact time. The distinct core bacteriomes mainly comprised populations related to the classes Betaproteobacteria, Cytophagia, and Chloroflexi in the PAO enrichment and of Gammaproteobacteria, Alphaproteobacteria, Acidobacteria, and Sphingobacteria in the GAO enrichment. An anaerobic metabolic batch test based on electrical conductivity evolution and a polyphosphatase enzymatic assay were developed for rapid and low-cost assessment of the active PAO fraction and dephosphatation potential of activated sludge. Linear correlations were obtained between the PAO fraction, biomass specific rate of conductivity increase under anaerobic conditions, and polyphosphate-hydrolyzing activity of PAO/GAO mixtures. The correlations between PAO/GAO ratios, metabolic activities, and conductivity profiles were confirmed by simulations with a mathematical model developed in the aqueous geochemistry software PHREEQC. © 2014 Wiley Periodicals, Inc.

  2. Sensitive electrochemical determination of miRNAs based on a sandwich assay onto magnetic microcarriers and hybridization chain reaction amplification.

    Science.gov (United States)

    Torrente-Rodríguez, R M; Campuzano, S; Montiel, V Ruiz-Valdepeñas; Montoya, J J; Pingarrón, J M

    2016-12-15

    A novel electrochemical approach for determination of miRNAs involving a sandwich hybridization assay onto streptavidin-magnetic beads (Strep-MBs), hybridization chain reaction (HCR) amplification and amperometric detection at disposable screen-printed carbon electrodes is reported. Using miRNA-21 as the target analyte, a dynamic linear range from 0.2 to 5.0nM with a 60pM (1.5fmol in 25μL) detection limit was obtained. The achieved sensitivity is 24-fold higher than a non-HCR amplification approach involving conventional sandwich type assay onto MBs. Moreover, the whole assay time lasted 1h 45min which is remarkably shorter than other reported methodologies. The methodology exhibited full selectivity against other non-complementary miRNAs as well as an acceptable discrimination between homologous miRNA family members. The applicability of this novel approach was demonstrated by determining mature miRNA-21 in total RNA (RNAt) extracted from tumor cells and human tissues.

  3. Visual search of neuropil-enriched RNAs from brain in situ hybridization data through the image analysis pipeline hippo-ATESC.

    Directory of Open Access Journals (Sweden)

    Roberto Ugolotti

    Full Text Available MOTIVATION: RNA molecules specifically enriched in the neuropil of neuronal cells and in particular in dendritic spines are of great interest for neurobiology in virtue of their involvement in synaptic structure and plasticity. The systematic recognition of such molecules is therefore a very important task. High resolution images of RNA in situ hybridization experiments contained in the Allen Brain Atlas (ABA represent a very rich resource to identify them and have been so far exploited for this task through human-expert analysis. However, software tools that may automatically address the same objective are not very well developed. RESULTS: In this study we describe an automatic method for exploring in situ hybridization data and discover neuropil-enriched RNAs in the mouse hippocampus. We called it Hippo-ATESC (Automatic Texture Extraction from the Hippocampal region using Soft Computing. Bioinformatic validation showed that the Hippo-ATESC is very efficient in the recognition of RNAs which are manually identified by expert curators as neuropil-enriched on the same image series. Moreover, we show that our method can also highlight genes revealed by microdissection-based methods but missed by human visual inspection. We experimentally validated our approach by identifying a non-coding transcript enriched in mouse synaptosomes. The code is freely available on the web at http://ibislab.ce.unipr.it/software/hippo/.

  4. Metal-organic frameworks@graphene hybrid aerogels for solid-phase extraction of non-steroidal anti-inflammatory drugs and selective enrichment of proteins.

    Science.gov (United States)

    Zhang, Xiaoqiong; Liang, Qionglin; Han, Qiang; Wan, Wei; Ding, Mingyu

    2016-06-20

    Graphene aerogel (GA)-supported metal-organic framework (MOF) particles with a three-dimensional (3D) architecture were fabricated for the first time via a facile template-free "sol-cryo" method. The prepared MOFs@graphene hybrid aerogels exhibit a 3D interconnected macroporous framework of graphene sheets with uniform dispersion of MOF particles. We also report the first attempt at using the hybrid aerogels as adsorbents for the solid-phase extraction (SPE) of non-steroidal anti-inflammatory drugs (NSAIDs) and the selective enrichment of proteins. The macroporous skeletons of GA provide both low backpressure and rapid mass transfer in SPE application, thus overcoming the obstacle of high backpressure caused by directly packing submicron or micron sized MOF particles into SPE cartridges. Excellent performances including satisfactory recoveries, high sensitivity and good reproducibility were achieved in the extraction of five NSAIDs. The hybrid aerogels also showed an interesting ability for selective enrichment of ribonuclease A (RNase A) and simultaneous exclusion of cytochrome C (Cyt C) and lysozyme (Lyz), which could be attributed to the electrostatic interactions between proteins and the positively charged coordinatively unsaturated metal sites (CUS) in MIL-101. We believe that this work will promote the application of MOFs in adsorption and separation, and our synthetic strategy could be further extended to develop other graphene-based hybrid aerogels.

  5. Paper-based solid-phase nucleic acid hybridization assay using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Noor, M Omair; Shahmuradyan, Anna; Krull, Ulrich J

    2013-02-05

    A paper-based solid-phase assay is presented for transduction of nucleic acid hybridization using immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) were FRET-paired with Cy3 acceptor. Hybridization of Cy3-labeled oligonucleotide targets provided the proximity required for FRET-sensitized emission from Cy3, which served as an analytical signal. The assay exhibited rapid transduction of nucleic acid hybridization within minutes. Without any amplification steps, the limit of detection of the assay was found to be 300 fmol with the upper limit of the dynamic range at 5 pmol. The implementation of glutathione-coated QDs for the development of nucleic acid hybridization assay integrated on a paper-based platform exhibited excellent resistance to nonspecific adsorption of oligonucleotides and showed no reduction in the performance of the assay in the presence of large quantities of noncomplementary DNA. The selectivity of nucleic acid hybridization was demonstrated by single-nucleotide polymorphism (SNP) detection at a contrast ratio of 19 to 1. The reuse of paper over multiple cycles of hybridization and dehybridization was possible, with less than 20% reduction in the performance of the assay in five cycles. This work provides an important framework for the development of paper-based solid-phase QD-FRET nucleic acid hybridization assays that make use of a ratiometric approach for detection and analysis.

  6. Bacterial Bioburden Decrease in Orthokeratology Lens Storage Cases After Forewarning: Assessment by the DNA Dot Hybridization Assay.

    Science.gov (United States)

    Fang, Po-Chiung; Lo, Jung; Chang, Tsung C; Chien, Chun-Chih; Hsiao, Chang-Chun; Tseng, Shin-Ling; Lai, Yu-Hsuan; Kuo, Ming-Tse

    2017-05-01

    The aim of this study was to measure the changes in the bacterial bioburden in orthokeratology (OK) lens storage cases using the DNA dot hybridization assay (DHA) after forewarning patients about their bacterial contamination severity. Thirty-one OK lens wearers were prospectively enrolled in this study. Dot hybridization assay was used for serial measurements of bacterial bioburden in OK storage cases after lenses had been soaked for approximately 6 hr. After the first assessment, the lens wearers were informed of the extent of case contamination and the possible risk of microbial keratitis (MK), and best practices for lens care and lens case hygiene were reviewed and reinforced. A second assessment by the same DHA method was performed after approximately 6 months. Two universal bacterial probes confirmed a significant decrease in bacterial bioburden at the second assessment (P<0.01 and P<0.001). Genus-specific probes showed significant reductions in Acinetobacter and Klebsiella (P=0.02 and P=0.01), but not in Pseudomonas (P=0.42). Making OK lens wearers aware of the bacterial bioburden in their lens cases resulted in improved quality of case care and reduced bioburden. Our results suggest that a strategy of bioburden assessment with forewarning could be a useful method to decrease the incidence of OK-related MK.

  7. Identifying Gene Regulatory Networks in Arabidopsis by In Silico Prediction, Yeast-1-Hybrid, and Inducible Gene Profiling Assays.

    Science.gov (United States)

    Sparks, Erin E; Benfey, Philip N

    2016-01-01

    A system-wide understanding of gene regulation will provide deep insights into plant development and physiology. In this chapter we describe a threefold approach to identify the gene regulatory networks in Arabidopsis thaliana that function in a specific tissue or biological process. Since no single method is sufficient to establish comprehensive and high-confidence gene regulatory networks, we focus on the integration of three approaches. First, we describe an in silico prediction method of transcription factor-DNA binding, then an in vivo assay of transcription factor-DNA binding by yeast-1-hybrid and lastly the identification of co-expression clusters by transcription factor induction in planta. Each of these methods provides a unique tool to advance our understanding of gene regulation, and together provide a robust model for the generation of gene regulatory networks.

  8. Determination of estrogen receptor {beta}-mediated estrogenic potencies of hydroxylated PCBS by a yeast two-hybrid assay

    Energy Technology Data Exchange (ETDEWEB)

    Kuroki, H.; Kumate, M.; Nakaoka, H.; Yonekura, S. [Daiichi Coll. of Pharmaceutical Sciences, Fukuoka (Japan); Nishikawa, J.; Nishihara, T. [Osaka Univ., Osaka (Japan)

    2004-09-15

    Several environmental phenolic chemicals such as Nonylphenol and Bisphenol A (BPA) have been previously shown to possess estrogenic properties. In the previous paper, we have investigated the estrogenic activity of a series of hydroxylated PCBs (OH-PCBs) by a yeast two-hybrid assay (estrogen receptor{alpha} (ER{alpha}) -TIF2), in which the expression of estrogenic activity is based on the interaction of chemicals with ER{alpha}, and demonstrated that 4'-OH-CB30 and 4'-OH-CB61 are more estrogenic than BPA, one of the environmental estrogens. We have showed that one chlorine substitution adjacent to 4-OH at 3- or 5-position significantly reduces the ER{alpha}-mediated estrogenic activity of 4-OH-PCBs. Thus, 4'-OH-CB25 and 4-OH-CB56 showed a very weak estrogenicity. We have also showed that 4-OH-PCBs with two chlorine substitutions adjacent to 4-OH at 3- and 5-position such as 4'-OH-CB79 (hydroxylated metabolite of CB77) and persistent 4-OH-PCBs retained in human blood (4-OH-CB107, 4-OH-CB146 and 4-OH-CB187) have no ER{alpha}-mediated estrogenic activity. ER is known to have two subtypes, namely ER{alpha} and ER{beta} and it is reported that ligand, some agonist and antagonist have a different binding affinity for ER{alpha} and ER{beta}. However, there is limited information on ER{beta}-mediated endocrine disrupting potency. In this study, we examined the ER{beta}-mediated estrogenic activity of a series of OH-PCBs, including environmentally relevant 4-OH-PCBs by a yeast two-hybrid assay (ER{beta}-TIF2).

  9. Fluorometric polyethyleneglycol-peptide hybrid substrates for quantitative assay of protein disulfide isomerase

    DEFF Research Database (Denmark)

    Christiansen, Camilla; St Hilaire, Phaedria M; Winther, Jakob R.

    2004-01-01

    In eukaryotic cells the enzyme protein disulfide isomerase (PDI) is responsible for the formation and reshuffling of disulfide bonds in secretory proteins. The reaction carried out by PDI involves interaction with a highly complex mixture of polypeptide molecules that are in the process of folding....... This means that PDI activity is typically measured in the context of a globular protein folding pathway. The absence of small, well-defined substrates for the quantitation of both oxidation and reduction reactions constitutes an inherent problem in the analysis of PDI activity. We describe a new type...... of substrate for PDI where two cysteine-containing oligopeptides are connected by an onameric ethylene glycol linker. We term such hybrid compounds PEGtides. The oligopeptides are each marked with a fluorescent aminobenzoic acid and a quenching nitrotyrosine group, respectively. The reversible formation...

  10. Diagnosis of visceral Leishmaniasis in asymptomatic dogs by the KDNA PCR-hybridization assay using noninvasive samples

    Energy Technology Data Exchange (ETDEWEB)

    Leite, Rodrigo Souza; Andrade, Antero Silva Ribeiro de [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil). Lab. de Radiobiologia], e-mail: rleite2005@gmail.com; Ferreira, Sydney de Almeida; Ituassu, Leonardo Trindade; Melo, Maria Norma de [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Centro de Ciencias Biologicas. Dept. de Parasitologia], e-mail: saninoalmeida@gmail.com

    2009-07-01

    The visceral leishmaniasis (VL) in Brazil is caused by Leishmania (Leishmania) chagasi and the asymptomatic dogs may transmit the parasite to sand flies vectors. The VL epidemiological control in Brazil involves the elimination of seropositive dogs, insecticide treatment and systematic treatment of human cases. Therefore, the accurate diagnosis is important in order to avoid the disease transmission or unnecessary culling of dogs. Serological tests are used for screening of dogs. However, these techniques present limitations. The Polymerase Chain Reaction (PCR) is an attractive alternative to the diagnosis in this context; but non-invasive samplings have great importance because they are simpler, painless and less resisted by dog-owners. This study aimed at evaluating conjunctival swab (CS) for canine VL diagnosis. In this methodology a sterile cotton swab is used to sampling the dog conjunctiva in both eyes. Thirty asymptomatic seropositive dogs were used. The samples were analyzed by the kDNA PCR-hybridization procedure in which the PCR products are hybridized with cloned kDNA mini-circles labeled with {sup 32}P[]dCTP. In addition, blood (B) was collected from each animal. L. chagasi was identified in 90% of CS samples and 13,6% of B samples. The high sensitivity obtained with asymptomatic dogs, in which the diagnosis is more difficult due the low number of parasites in the samples, allow concluding that the conjunctival swab associated to the kDNA PCR-hybridization assay provides a valuable alternative tool for the direct diagnosis of canine leishmaniasis. (author)

  11. Cross-reactivity profiles of hybrid capture II, cobas, and APTIMA human papillomavirus assays

    DEFF Research Database (Denmark)

    Preisler, Sarah Nørgaard; Rebolj, Matejka; Ejegod, Ditte Møller

    2016-01-01

    Background High-risk Human Papillomavirus (HPV) testing is replacing cytology in cervical cancer screening as it is more sensitive for preinvasive cervical lesions. However, the bottleneck of HPV testing is the many false positive test results (positive tests without cervical lesions). Here, we...... assays. None of the 35 genotypes was detected in 49 (1.0 %), 162 (3.2 %), and 56 (1.1 %) samples, respectively. In primary screening at age 30 to 65 years (n = 2859), samples of 72 (25 %) out of 289 with high-risk infections on HC2 and 

  12. Implementation of a multiregion hybridization assay to characterize HIV-1 strains detected among injecting drug users in Manipur, India.

    Science.gov (United States)

    Sarkar, Roni; Sengupta, Satarupa; Mullick, Ranajoy; Singh, N Brajachand; Sarkar, Kamalesh; Chakrabarti, Sekhar

    2009-01-01

    We have implemented the latest technology of a multiregion hybridization assay (MHAbce, version 2) for the molecular characterization of HIV-1 among injecting drug users (IDUs) of Manipur, India. This study provides a more detailed analysis on the basis of probes designed from eight different genomic regions of HIV-1, to achieve a clear picture of HIV-1 genomic diversity in Manipur. Out of 30 samples, 15 were found to be of subtype C, 1 of subtype B, 5 with dual-probe reactivity, 8 with multigenomic recombination pattern and 1 sample showed both dual-probe reactivity and multigenomic variations. In contrast, the heteroduplex mobility assay (HMA) with respect to gag and env genes revealed 21 samples to be of subtype C (gag C/env C), 3 samples of subtype B (gag B/env B) and 6 samples of B/C recombinants (gag C/env B). MHAbce illustrates the occurrence of inter- and intragenomic variants and dual infection in an IDU population from India. It also indicates the possibility of the presence of new circulating recombinant forms of HIV-1 strains, which might have been difficult to trace by HMA alone.

  13. The next-generation Hybrid Capture High-Risk HPV DNA assay on a fully automated platform.

    Science.gov (United States)

    Eder, Paul S; Lou, Jianrong; Huff, John; Macioszek, Jerzy

    2009-07-01

    A next-generation diagnostic system has been developed at QIAGEN. The QIAensemble system consists of an analytical subsystem (JE2000) that utilizes a re-engineered Hybrid Capture chemistry (NextGen) to maintain the high level of clinical sensitivity established by the digene High-Risk HPV DNA Test (HC2), while creating improved analytical specificity as shown both in plasmid-based analyses and in processing of clinical specimens. Limit-of-detection and cross-reactivity experiments were performed using plasmid DNA constructs containing multiple high-risk (HR) and low-risk (LR) HPV types. Cervical specimens collected into a novel specimen collection medium, DCM, were used to measure stability of specimens, as well as analytical specificity. Signal carryover, instrument precision, and specimen reproducibility were measured on the prototype JE2000 system using the automated NextGen assay. The Limit of Detection (LOD) is HPV 16 plasmid in the automated assay. No cross-reactivity (signal above cutoff) was detected on the automated system from any of 13 LR types tested at 10(7) copies per assay. Within-plate, plate-to-plate, and day-to-day performance in the prototype system yielded a CV of 20%. No indication of target carryover was found when samples containing up to 10(9) copies/ml of HPV DNA type 16 were processed on the JE2000 instrument. In an agreement study with HC2, 1038 donor cervical specimens were tested in both the manual NextGen assay and HC2 to evaluate agreement between the two tests. After eliminating discrepant specimens that were adjudicated by HR-HPV genotyping, the adjudicated positive agreement was 98.5% (95% CI: 94.6, 99.6). The JE2000 prototype system automates NextGen assay processing, yielding accurate, reproducible, and highly specific results with both plasmid analytical model tests and cervical specimens collected in DCM. The final system will process more than 2000 specimens in an 8-hour shift, with fully continuous loading.

  14. An integrated closed-tube 2-plex PCR amplification and hybridization assay with switchable lanthanide luminescence based spatial detection.

    Science.gov (United States)

    Lahdenperä, Susanne; Spangar, Anni; Lempainen, Anna-Maija; Joki, Laura; Soukka, Tero

    2015-06-21

    Switchable lanthanide luminescence is a binary probe technology that inherently enables a high signal modulation in separation-free detection of DNA targets. A luminescent lanthanide complex is formed only when the two probes hybridize adjacently to their target DNA. We have now further adapted this technology for the first time in the integration of a 2-plex polymerase chain reaction (PCR) amplification and hybridization-based solid-phase detection of the amplification products of the Staphylococcus aureus gyrB gene and an internal amplification control (IAC). The assay was performed in a sealed polypropylene PCR chip containing a flat-bottom reaction chamber with two immobilized capture probe spots. The surface of the reaction chamber was functionalized with NHS-PEG-azide and alkyne-modified capture probes for each amplicon, labeled with a light harvesting antenna ligand, and covalently attached as spots to the azide-modified reaction chamber using a copper(i)-catalyzed azide-alkyne cycloaddition. Asymmetric duplex-PCR was then performed with no template, one template or both templates present and with a europium ion carrier chelate labeled probe for each amplicon in the reaction. After amplification europium fluorescence was measured by scanning the reaction chamber as a 10 × 10 raster with 0.6 mm resolution in time-resolved mode. With this assay we were able to co-amplify and detect the amplification products of the gyrB target from 100, 1000 and 10,000 copies of isolated S. aureus DNA together with the amplification products from the initial 5000 copies of the synthetic IAC template in the same sealed reaction chamber. The addition of 10,000 copies of isolated non-target Escherichia coli DNA in the same reaction with 5000 copies of the synthetic IAC template did not interfere with the amplification or detection of the IAC. The dynamic range of the assay for the synthetic S. aureus gyrB target was three orders of magnitude and the limit of detection of 8 p

  15. Discrimination of clostridium species using a magnetic bead based hybridization assay

    Science.gov (United States)

    Pahlow, Susanne; Seise, Barbara; Pollok, Sibyll; Seyboldt, Christian; Weber, Karina; Popp, Jürgen

    2014-05-01

    Clostridium chauvoei is the causative agent of blackleg, which is an endogenous bacterial infection. Mainly cattle and other ruminants are affected. The symptoms of blackleg are very similar to those of malignant edema, an infection caused by Clostridium septicum. [1, 2] Therefore a reliable differentiation of Clostridium chauvoei from other Clostridium species is required. Traditional microbiological detection methods are time consuming and laborious. Additionally, the unique identification is hindered by the overgrowing tendency of swarming Clostridium septicum colonies when both species are present. [1, 3, 4] Thus, there is a crucial need to improve and simplify the specific detection of Clostridium chauvoei and Clostridium septicum. Here we present an easy and fast Clostridium species discrimination method combining magnetic beads and fluorescence spectroscopy. Functionalized magnetic particles exhibit plentiful advantages, like their simple manipulation in combination with a large binding capacity of biomolecules. A specific region of the pathogenic DNA is amplified and labelled with biotin by polymerase chain reaction (PCR). These PCR products were then immobilized on magnetic beads exploiting the strong biotin-streptavidin interaction. The specific detection of different Clostridium species is achieved by using fluorescence dye labeled probe DNA for the hybridization with the immobilized PCR products. Finally, the samples were investigated by fluorescence spectroscopy. [5

  16. Paper-based solid-phase multiplexed nucleic acid hybridization assay with tunable dynamic range using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Noor, M Omair; Krull, Ulrich J

    2013-08-06

    A multiplexed solid-phase nucleic acid hybridization assay on a paper-based platform is presented using multicolor immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize two types of QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) and red-emitting QDs (rQDs) served as donors with Cy3 and Alexa Fluor 647 (A647) acceptors. The gQD/Cy3 FRET pair served as an internal standard, while the rQD/A647 FRET pair served as a detection channel, combining the control and analytical test zones in one physical location. Hybridization of dye-labeled oligonucleotide targets provided the proximity for FRET sensitized emission from the acceptor dyes, which served as an analytical signal. Hybridization assays in the multicolor format provided a limit of detection of 90 fmol and an upper limit of dynamic range of 3.5 pmol. The use of an array of detection zones was designed to provide improved analytical figures of merit compared to that which could be achieved on one type of array design in terms of relative concentration of multicolor QDs. The hybridization assays showed excellent resistance to nonspecific adsorption of oligonucleotides. Selectivity of the two-plex hybridization assay was demonstrated by single nucleotide polymorphism (SNP) detection at a contrast ratio of 50:1. Additionally, it is shown that the use of preformed QD-probe oligonucleotide conjugates and consideration of the relative number density of the two types of QD-probe conjugates in the two-color assay format is advantageous to maximize assay sensitivity and the upper limit of dynamic range.

  17. On-chip multiplexed solid-phase nucleic acid hybridization assay using spatial profiles of immobilized quantum dots and fluorescence resonance energy transfer

    Energy Technology Data Exchange (ETDEWEB)

    Noor, M. Omair; Tavares, Anthony J.; Krull, Ulrich J., E-mail: ulrich.krull@utoronto.ca

    2013-07-25

    Graphical abstract: -- Highlights: •Solid-phase multiplexed QD-FRET nucleic acid assay in electrokinetic fluidic chip. •Concurrent detection of two oligonucleotides based on channel length coverage. •Selection of “turn-on” and “turn-off” signals from two acceptor dyes and two colors of immobilized QDs, respectively. •No loss in assay sensitivity when implementing multiplexed assay format. -- Abstract: A microfluidic based solid-phase assay for the multiplexed detection of nucleic acid hybridization using quantum dot (QD) mediated fluorescence resonance energy transfer (FRET) is described herein. The glass surface of hybrid glass-polydimethylsiloxane (PDMS) microfluidic channels was chemically modified to assemble the biorecognition interface. Multiplexing was demonstrated using a detection system that was comprised of two colors of immobilized semi-conductor QDs and two different oligonucleotide probe sequences. Green-emitting and red-emitting QDs were paired with Cy3 and Alexa Fluor 647 (A647) labeled oligonucleotides, respectively. The QDs served as energy donors for the transduction of dye labeled oligonucleotide targets. The in-channel assembly of the biorecognition interface and the subsequent introduction of oligonucleotide targets was accomplished within minutes using a combination of electroosmotic flow and electrophoretic force. The concurrent quantification of femtomole quantities of two target sequences was possible by measuring the spatial coverage of FRET sensitized emission along the length of the channel. In previous reports, multiplexed QD-FRET hybridization assays that employed a ratiometric method for quantification had challenges associated with lower analytical sensitivity arising from both donor and acceptor dilution that resulted in reduced energy transfer pathways as compared to single-color hybridization assays. Herein, a spatial method for quantification that is based on in-channel QD-FRET profiles provided higher analytical

  18. A Paper-Based Sandwich Format Hybridization Assay for Unlabeled Nucleic Acid Detection Using Upconversion Nanoparticles as Energy Donors in Luminescence Resonance Energy Transfer

    Directory of Open Access Journals (Sweden)

    Feng Zhou

    2015-09-01

    Full Text Available Bioassays based on cellulose paper substrates are gaining increasing popularity for the development of field portable and low-cost diagnostic applications. Herein, we report a paper-based nucleic acid hybridization assay using immobilized upconversion nanoparticles (UCNPs as donors in luminescence resonance energy transfer (LRET. UCNPs with intense green emission served as donors with Cy3 dye as the acceptor. The avidin functionalized UCNPs were immobilized on cellulose paper and subsequently bioconjugated to biotinylated oligonucleotide probes. Introduction of unlabeled oligonucleotide targets resulted in a formation of probe-target duplexes. A subsequent hybridization of Cy3 labeled reporter with the remaining single stranded portion of target brought the Cy3 dye in close proximity to the UCNPs to trigger a LRET-sensitized emission from the acceptor dye. The hybridization assays provided a limit of detection (LOD of 146.0 fmol and exhibited selectivity for one base pair mismatch discrimination. The assay was functional even in undiluted serum samples. This work embodies important progress in developing DNA hybridization assays on paper. Detection of unlabeled targets is achieved using UCNPs as LRET donors, with minimization of background signal from paper substrates owing to the implementation of low energy near-infrared (NIR excitation.

  19. Multiplexed Nucleic Acid Hybridization Assays Using Single-FRET-Pair Distance-Tuning.

    Science.gov (United States)

    Qiu, Xue; Guo, Jiajia; Jin, Zongwen; Petreto, Alexandra; Medintz, Igor L; Hildebrandt, Niko

    2017-07-01

    Multiplexed photoluminescence (PL) detection plays an important role in chemical and biological sensing. Here, it is shown that time-gated (TG) detection of a single terbium-donor-based Förster resonance energy transfer (FRET) pair can be used to selectively quantify low nanomolar concentrations of multiple DNAs or microRNAs in a single sample. This study demonstrates the applicability of single-TG-FRET-pair multiplexing for molecular (Tb-to-dye) and nanoparticle (Tb-to-quantum-dot) biosensing. Both systems use acceptor-sensitization and donor-quenching for quantifying biomolecular recognition and modification of the donor-acceptor distance for tuning the PL decays. TG intensity detection provides extremely low background noise and a quick and simple one-step assay format. Single-TG-FRET-pair multiplexing can be combined with spectral and spatial resolution, paving the way for biosensing with unprecedented high-order multiplexing capabilities. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Identification and characterization of protein interactions in the mammalian mRNA processing body using a novel two-hybrid assay

    Energy Technology Data Exchange (ETDEWEB)

    Bloch, Donald B., E-mail: bloch@helix.mgh.harvard.edu; Nobre, Rita A.; Bernstein, Gillian A.; Yang, Wei-Hong

    2011-09-10

    Components of the mRNA processing body (P-body) regulate critical steps in mRNA storage, transport, translation and degradation. At the core of the P-body is the decapping complex, which removes the 5' cap from de-adenylated mRNAs and mediates an irreversible step in mRNA degradation. The assembly of P-bodies in Saccharomyces cerevisiae, Arabidopsis thaliana and Drosophila melanogaster has been previously described. Less is known about the assembly of mammalian P-bodies. To investigate the interactions that occur between components of mammalian P-bodies, we developed a fluorescence-based, two-hybrid assay system. The assay depends on the ability of one P-body component, fused to an exogenous nuclear localization sequence (NLS), to recruit other P-body components to the nucleus. The assay was used to investigate interactions between P-body components Ge-1, DCP2, DCP1, EDC3, RAP55, and RCK. The results of this study show that the modified two-hybrid assay can be used to identify protein interactions that occur in a macromolecular complex. The assay can also be used to efficiently detect protein interaction domains. The results provide important insights into mammalian P-body assembly and demonstrate similarities, and critical differences, between P-body assembly in mammalian cells compared with that of other species. -- Research highlights: {yields} A two-hybrid assay was developed to study interactions in macromolecular complexes. {yields} The assay was applied to interactions between components of mRNA P-bodies. {yields} The assay effectively and efficiently identified protein interaction domains. {yields} P-body assembly in mammalian cells differs from that in other species.

  1. Study comparing human papillomavirus (HPV) real-time multiplex PCR and Hybrid Capture II INNO-LiPA v2 HPV genotyping PCR assays

    DEFF Research Database (Denmark)

    Iftner, Thomas; Germ, Liesje; Swoyer, Ryan

    2009-01-01

    Human papillomavirus (HPV) DNA genotyping is an essential test to establish efficacy in HPV vaccine clinical trials and HPV prevalence in natural history studies. A number of HPV DNA genotyping methods have been cited in the literature, but the comparability of the outcomes from the different...... methods has not been well characterized. Clinically, cytology is used to establish possible HPV infection. We evaluated the sensitivity and specificity of HPV multiplex PCR assays compared to those of the testing scheme of the Hybrid Capture II (HCII) assay followed by an HPV PCR/line hybridization assay...... (HCII-LiPA v2). SurePath residual samples were split into two aliquots. One aliquot was subjected to HCII testing followed by DNA extraction and LiPA v2 genotyping. The second aliquot was shipped to a second laboratory, where DNA was extracted and HPV multiplex PCR testing was performed. Comparisons...

  2. Simultaneous detection of several oligonucleotides by time-resolved fluorometry: the use of a mixture of categorized microparticles in a sandwich type mixed-phase hybridization assay.

    Science.gov (United States)

    Hakala, H; Virta, P; Salo, H; Lönnberg, H

    1998-12-15

    Porous, uniformly sized (50 micrometer) glycidyl methacrylate/ethylene dimethacrylate particles (SINTEF) were used as a solid phase to construct a sandwich type hybridization assay that allowed simultaneous detection of up to six oligonucleotides from a single sample. The assay was based on categorization of the particles by two organic prompt fluorophores, viz. fluorescein and dansyl, and quantification of the oligonucleotide hybridization by time-resolved fluorometry. Accordingly, allele-specific oligodeoxyribonucleotide probes were assembled on the particles by conventional phosphoramidite strategy using a non-cleavable linker, and the category defining fluorescein and/or dansyl tagged building blocks were inserted in the 3'-terminal sequence. An oligonucleotide bearing a photoluminescent europium(III) chelate was hybridized to the complementary 3'-terminal sequence of the target oligonucleotide, and the resulting duplex was further hybridized to the particle-bound allele-specific probes via the 5'-terminal sequence of the target. After hybridization each individual particle was subjected to three different fluorescence intensity measurements. The intensity of the prompt fluorescence signals of fluorescein and dansyl defined the particle category, while the europium(III) chelate emission quantified the hybridization. The length of the complementary region between the target oligonucleotide and the particle-bound probe was optimized to achieve maximal selectivity. Furthermore, the kinetics of hybridization and the effect of the concentration of the target oligomer on the efficiency of hybridization were evaluated. By this approach the possible presence of a three base deletion (DeltaF508), point mutation (G542X) and point deletion (1078delT) related to cystic fibrosis could unequivocally be detected from a single sample.

  3. USE OF FLUORESCENCE IN SITU HYBRIDIZATION ASSAY ON URINE SEDIMENT CELLS TO DIAGNOSE URINARY BLADDER CANCER AND ITS RECURRENCESY

    Directory of Open Access Journals (Sweden)

    I. E. Vorobtsova

    2014-07-01

    Full Text Available Fluorescence in situ hybridization (FISH assay was used to detect tumor cells in the urine sediment of patients diagnosed as having urinary bladder cancer (UBC. For this, the investigators applied a fluorescence DNA probe kit (UroVysion that could reveal the cytogenetic abnormalities characteristic for UBC, such as hyperploidy for chromosomes 3, 7, and 17 and deletion of the 9p21 locus, in the cast-off cells. Twenty-eight patients with the primary diagnosis of UBC, 12 with its suspected recurrence, 3 subjects without UBC were examined. The findings were compared with cystoscopic data after urine samples were taken. The sensitivity of the UroVysion test totaled 78.5 ± 9.7 % for all stages of primary cancer (pT1-pT4, 87.5 ± 11.6 % for its early stage (рТ1, and 100 % for UBC recurrences. Hyperploidy was a predominant type of cytogenetic abnormalities in the cast-off tumor cells. Among the abnormal cells, the types of hyperploidy (tri-, tetrasomy were most common for chromosome 3 and less for chromosome 7. Thus, the UroVysion test is a noninvasive highly sensitive tool that may be used in clinical practice to improve the diagnosis of UBC, to detect recurrences, and to monitor the efficiency of treatment.

  4. USE OF FLUORESCENCE IN SITU HYBRIDIZATION ASSAY ON URINE SEDIMENT CELLS TO DIAGNOSE URINARY BLADDER CANCER AND ITS RECURRENCESY

    Directory of Open Access Journals (Sweden)

    I. E. Vorobtsova

    2011-01-01

    Full Text Available Fluorescence in situ hybridization (FISH assay was used to detect tumor cells in the urine sediment of patients diagnosed as having urinary bladder cancer (UBC. For this, the investigators applied a fluorescence DNA probe kit (UroVysion that could reveal the cytogenetic abnormalities characteristic for UBC, such as hyperploidy for chromosomes 3, 7, and 17 and deletion of the 9p21 locus, in the cast-off cells. Twenty-eight patients with the primary diagnosis of UBC, 12 with its suspected recurrence, 3 subjects without UBC were examined. The findings were compared with cystoscopic data after urine samples were taken. The sensitivity of the UroVysion test totaled 78.5 ± 9.7 % for all stages of primary cancer (pT1-pT4, 87.5 ± 11.6 % for its early stage (рТ1, and 100 % for UBC recurrences. Hyperploidy was a predominant type of cytogenetic abnormalities in the cast-off tumor cells. Among the abnormal cells, the types of hyperploidy (tri-, tetrasomy were most common for chromosome 3 and less for chromosome 7. Thus, the UroVysion test is a noninvasive highly sensitive tool that may be used in clinical practice to improve the diagnosis of UBC, to detect recurrences, and to monitor the efficiency of treatment.

  5. A hybrid stochastic-deterministic computational model accurately describes spatial dynamics and virus diffusion in HIV-1 growth competition assay.

    Science.gov (United States)

    Immonen, Taina; Gibson, Richard; Leitner, Thomas; Miller, Melanie A; Arts, Eric J; Somersalo, Erkki; Calvetti, Daniela

    2012-11-01

    We present a new hybrid stochastic-deterministic, spatially distributed computational model to simulate growth competition assays on a relatively immobile monolayer of peripheral blood mononuclear cells (PBMCs), commonly used for determining ex vivo fitness of human immunodeficiency virus type-1 (HIV-1). The novel features of our approach include incorporation of viral diffusion through a deterministic diffusion model while simulating cellular dynamics via a stochastic Markov chain model. The model accounts for multiple infections of target cells, CD4-downregulation, and the delay between the infection of a cell and the production of new virus particles. The minimum threshold level of infection induced by a virus inoculum is determined via a series of dilution experiments, and is used to determine the probability of infection of a susceptible cell as a function of local virus density. We illustrate how this model can be used for estimating the distribution of cells infected by either a single virus type or two competing viruses. Our model captures experimentally observed variation in the fitness difference between two virus strains, and suggests a way to minimize variation and dual infection in experiments.

  6. Interfacial chemistry and the design of solid-phase nucleic acid hybridization assays using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    Science.gov (United States)

    Algar, W Russ; Krull, Ulrich J

    2011-01-01

    The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling the regeneration and reuse of solid-phase QD-FRET hybridization assays. FRET-sensitized emission from acceptor dyes associated with hybridization events at immobilized QD donors provides the analytical signal in these assays. The minimization of active sensing area reduces background from QD donor PL and allows the resolution of smaller amounts of acceptor emission, thus lowering the LOD. The association of multiple acceptor dyes with each hybridization event can enhance FRET efficiency, thereby improving sensitivity. Many previous studies have used interfacial protein layers to generate selectivity; however, transient destabilization of these layers is shown to prevent efficient regeneration. To this end, we report a protein-free interfacial chemistry and demonstrate the specific detection of as little as 2 pmol of target, as well as an improved capacity for regeneration.

  7. A hybrid enrichment process combining conjugated polymer extraction and silica gel adsorption for high purity semiconducting single-walled carbon nanotubes (SWCNT)

    Science.gov (United States)

    Ding, Jianfu; Li, Zhao; Lefebvre, Jacques; Cheng, Fuyong; Dunford, Jeffrey L.; Malenfant, Patrick R. L.; Humes, Jefford; Kroeger, Jens

    2015-09-01

    A novel purification process for the enrichment of sc-SWCNTs that combines selective conjugated polymer extraction (CPE) with selective adsorption using silica gel, termed hybrid-CPE (h-CPE), has been developed, providing a high purity sc-SWCNT material with a significant improvement in process efficiency and yield. Using the h-CPE protocol, a greater than 5 fold improvement in yield can be obtained compared to traditional CPE while obtaining sc-SWCNT with a purity >99.9% as assessed by absorption spectroscopy and Raman mapping. Thin film transistor devices using the h-CPE derived sc-SWCNTs as the semiconductor possess mobility values ranging from 10-30 cm2 V-1 s-1 and current ON/OFF ratio of 104-105 for channel lengths between 2.5 and 20 μm.A novel purification process for the enrichment of sc-SWCNTs that combines selective conjugated polymer extraction (CPE) with selective adsorption using silica gel, termed hybrid-CPE (h-CPE), has been developed, providing a high purity sc-SWCNT material with a significant improvement in process efficiency and yield. Using the h-CPE protocol, a greater than 5 fold improvement in yield can be obtained compared to traditional CPE while obtaining sc-SWCNT with a purity >99.9% as assessed by absorption spectroscopy and Raman mapping. Thin film transistor devices using the h-CPE derived sc-SWCNTs as the semiconductor possess mobility values ranging from 10-30 cm2 V-1 s-1 and current ON/OFF ratio of 104-105 for channel lengths between 2.5 and 20 μm. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr04851f

  8. Comparative proteomic analysis of a membrane-enriched fraction from flag leaves reveals responses to chemical hybridization agent SQ-1 in wheat

    Directory of Open Access Journals (Sweden)

    Qilu eSong

    2015-08-01

    Full Text Available The induction of wheat male fertile lines by using the chemical hybridizing agent SQ-1 (CHA-SQ-1 is an effective approach in the utilization of heterosis; however, the molecular basis of male fertility remains unknown. Wheat flag leaves are the initial receptors of CHA-SQ-1 and their membrane structure plays a vital role in response to CHA-SQ-1 stress. To investigate the response of wheat flag leaves to CHA-SQ-1 stress, we compared their quantitative proteomic profiles in the absence and presence of CHA-SQ-1. Our results indicated that wheat flag leaves suffered oxidative stress during CHA-SQ-1 treatments. Leaf O2-, H2O2, and malonaldehyde levels were significantly increased within 10 h after CHA-SQ-1 treatment, while the activities of major antioxidant enzymes such as superoxide dismutase, catalase, and guaiacol peroxidase were significantly reduced. Proteome profiles of membrane-enriched fraction showed a change in the abundance of a battery of membrane proteins involved in multiple biological processes. These variable proteins mainly impaired photosynthesis, ATP synthesis protein mechanisms and were involved in the response to stress. These results provide an explanation of the relationships between membrane proteomes and anther abortion and the practical application of CHA for hybrid breeding.

  9. New Prototype Safeguards Technology Offers Improved Confidence and Automation for Uranium Enrichment Facilities

    Energy Technology Data Exchange (ETDEWEB)

    Brim, Cornelia P.

    2013-04-01

    An important requirement for the international safeguards community is the ability to determine the enrichment level of uranium in gas centrifuge enrichment plants and nuclear fuel fabrication facilities. This is essential to ensure that countries with nuclear nonproliferation commitments, such as States Party to the Nuclear Nonproliferation Treaty, are adhering to their obligations. However, current technologies to verify the uranium enrichment level in gas centrifuge enrichment plants or nuclear fuel fabrication facilities are technically challenging and resource-intensive. NNSA’s Office of Nonproliferation and International Security (NIS) supports the development, testing, and evaluation of future systems that will strengthen and sustain U.S. safeguards and security capabilities—in this case, by automating the monitoring of uranium enrichment in the entire inventory of a fuel fabrication facility. One such system is HEVA—hybrid enrichment verification array. This prototype was developed to provide an automated, nondestructive assay verification technology for uranium hexafluoride (UF6) cylinders at enrichment plants.

  10. A paper-based resonance energy transfer nucleic acid hybridization assay using upconversion nanoparticles as donors and quantum dots as acceptors

    Energy Technology Data Exchange (ETDEWEB)

    Doughan, Samer; Uddayasankar, Uvaraj; Krull, Ulrich J., E-mail: ulrich.krull@utoronto.ca

    2015-06-09

    Highlights: • Covalent immobilization of upconversion nanoparticles on paper. • LRET-based label free DNA detection using quantum dots as acceptors. • Use of polyethylene glycol to eliminate non-specific adsorption of quantum dots. • Improved analytical performance compared to analogous assays. - Abstract: Monodisperse aqueous upconverting nanoparticles (UCNPs) were covalently immobilized on aldehyde modified cellulose paper via reduction amination to develop a luminescence resonance energy transfer (LRET)-based nucleic acid hybridization assay. This first account of covalent immobilization of UCNPs on paper for a bioassay reports an optically responsive method that is sensitive, reproducible and robust. The immobilized UCNPs were decorated with oligonucleotide probes to capture HPRT1 housekeeping gene fragments, which in turn brought reporter conjugated quantum dots (QDs) in close proximity to the UCNPs for LRET. This sandwich assay could detect unlabeled oligonucleotide target, and had a limit of detection of 13 fmol and a dynamic range spanning nearly 3 orders of magnitude. The use of QDs, which are excellent LRET acceptors, demonstrated improved sensitivity, limit of detection, dynamic range and selectivity compared to similar assays that have used molecular fluorophores as acceptors. The selectivity of the assay was attributed to the decoration of the QDs with polyethylene glycol to eliminate non-specific adsorption. The kinetics of hybridization were determined to be diffusion limited and full signal development occurred within 3 min.

  11. Detection of Streptococcus suis by in situ hybridization, indirect immunofluorescence, and peroxidase-antiperoxidase assays in formalin-fixed, paraffin-embedded tissue sections from pigs

    DEFF Research Database (Denmark)

    Boye, Mette; Feenstra, Anne Avlund; Tegtmeier, Conny

    2000-01-01

    methods, an indirect immunofluorescence assay and a peroxidase-antiperoxidase method, using polyclonal antibodies also were developed. The specificity of the oligonucleotide probe was examined by whole-cell and dot-blot hybridization against reference strains of the 35 serotypes of S. suis and other...... closely related streptococci and other bacteria commonly isolated from pigs. The probe was specific for S, suis serotypes 1-31. The specificity of the polyclonal antibodies, which has previously been evaluated for use in diagnostic bacteriology for typing of serotype 2, was further evaluated...... in experimentally infected murine tissue with pure culture of different serotypes of S. suis, related streptococci, and other bacteria commonly found in pigs. The polyclonal antibodies against S. suis serotype 2 cross-reacted with serotypes 1 and 1/2 in these assays. The in situ hybridization...

  12. Resonance Energy Transfer-Based Nucleic Acid Hybridization Assays on Paper-Based Platforms Using Emissive Nanoparticles as Donors.

    Science.gov (United States)

    Doughan, Samer; Noor, M Omair; Han, Yi; Krull, Ulrich J

    2017-01-01

    Quantum dots (QDs) and upconverting nanoparticles (UCNPs) are luminescent nanoparticles (NPs) commonly used in bioassays and biosensors as resonance energy transfer (RET) donors. The narrow and tunable emissions of both QDs and UCNPs make them versatile RET donors that can be paired with a wide range of acceptors. Ratiometric signal processing that compares donor and acceptor emission in RET-based transduction offers improved precision, as it accounts for fluctuations in the absolute photoluminescence (PL) intensities of the donor and acceptor that can result from experimental and instrumental variations. Immobilizing NPs on a solid support avoids problems such as those that can arise with their aggregation in solution, and allows for facile layer-by-layer assembly of the interfacial chemistry. Paper is an attractive solid support for the development of point-of-care diagnostic assays given its ubiquity, low-cost, and intrinsic fluid transport by capillary action. Integration of nanomaterials with paper-based analytical devices (PADs) provides avenues to augment the analytical performance of PADs, given the unique optoelectronic properties of nanomaterials. Herein, we describe methodology for the development of PADs using QDs and UCNPs as RET donors for optical transduction of nucleic acid hybridization. Immobilization of green-emitting QDs (gQDs) on imidazole functionalized cellulose paper is described for use as RET donors with Cy3 molecular dye as acceptors for the detection of SMN1 gene fragment. We also describe the covalent immobilization of blue-emitting UCNPs on aldehyde modified cellulose paper for use as RET donors with orange-emitting QDs (oQDs) as acceptors for the detection of HPRT1 gene fragment. The data described herein is acquired using an epifluorescence microscope, and can also be collected using technology such as a typical electronic camera.

  13. Heterogeneous oligonucleotide-hybridization assay based on hot electron-induced electrochemiluminescence of a rhodamine label at oxide-coated aluminum and silicon electrodes

    Energy Technology Data Exchange (ETDEWEB)

    Spehar-Deleze, Anna-Maria [Laboratory of Sensors, Actuators and Microsystems, Institute of Microtechnology, University of Neuchatel, Rue Jaquet-Droz 1, CH-2007 Neuchatel (Switzerland) and Laboratory of Inorganic and Analytical Chemistry, Helsinki University of Technology, Kemistintie 1, FIN-02015 HUT (Finland)]. E-mail: anna-maria.spehar@unine.ch; Suomi, Johanna [Laboratory of Inorganic and Analytical Chemistry, Helsinki University of Technology, Kemistintie 1, FIN-02015 HUT (Finland); Jiang Qinghong [Laboratory of Inorganic and Analytical Chemistry, Helsinki University of Technology, Kemistintie 1, FIN-02015 HUT (Finland); Rooij, Nico de [Laboratory of Sensors, Actuators and Microsystems, Institute of Microtechnology, University of Neuchatel, Rue Jaquet-Droz 1, CH-2007 Neuchatel (Switzerland); Koudelka-Hep, Milena [Laboratory of Sensors, Actuators and Microsystems, Institute of Microtechnology, University of Neuchatel, Rue Jaquet-Droz 1, CH-2007 Neuchatel (Switzerland); Kulmala, Sakari [Laboratory of Inorganic and Analytical Chemistry, Helsinki University of Technology, Kemistintie 1, FIN-02015 HUT (Finland)

    2006-07-28

    This paper describes a heterogeneous oligonucleotide-hybridization assay based on hot electron-induced electrochemiluminescence (HECL) of a rhodamine label. Thin oxide-film coated aluminum and silicon electrodes were modified with an aminosilane layer and derivatized with short, 15-mer oligonucleotides via diisothiocyanate coupling. Target oligonucleotides were conjugated with tetramethylrhodamine (TAMRA) dye at their amino modified 5' end and hybridization was detected using HECL of TAMRA. Preliminary results indicate sensitivity down to picomolar level and low nonspecific adsorption. The sensitivity was better on oxide-coated silicon compared to oxide-coated aluminum electrodes and two-base pair mismatched hybrids were successfully discriminated. The experimental results presented here might be useful for the design of disposable electrochemiluminescent DNA biosensors.

  14. Interfacial Chemistry and the Design of Solid-Phase Nucleic Acid Hybridization Assays Using Immobilized Quantum Dots as Donors in Fluorescence Resonance Energy Transfer

    OpenAIRE

    Krull, Ulrich J.; W. Russ Algar

    2011-01-01

    The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling th...

  15. Study comparing human papillomavirus (HPV) real-time multiplex PCR and Hybrid Capture II INNO-LiPA v2 HPV genotyping PCR assays.

    Science.gov (United States)

    Iftner, Thomas; Germ, Liesje; Swoyer, Ryan; Kjaer, Susanne Kruger; Breugelmans, J Gabrielle; Munk, Christian; Stubenrauch, Frank; Antonello, Joseph; Bryan, Janine T; Taddeo, Frank J

    2009-07-01

    Human papillomavirus (HPV) DNA genotyping is an essential test to establish efficacy in HPV vaccine clinical trials and HPV prevalence in natural history studies. A number of HPV DNA genotyping methods have been cited in the literature, but the comparability of the outcomes from the different methods has not been well characterized. Clinically, cytology is used to establish possible HPV infection. We evaluated the sensitivity and specificity of HPV multiplex PCR assays compared to those of the testing scheme of the Hybrid Capture II (HCII) assay followed by an HPV PCR/line hybridization assay (HCII-LiPA v2). SurePath residual samples were split into two aliquots. One aliquot was subjected to HCII testing followed by DNA extraction and LiPA v2 genotyping. The second aliquot was shipped to a second laboratory, where DNA was extracted and HPV multiplex PCR testing was performed. Comparisons were evaluated for 15 HPV types common in both assays. A slightly higher proportion of samples tested positive by the HPV multiplex PCR than by the HCII-LiPA v2 assay. The sensitivities of the multiplex PCR assay relative to those of the HCII-LiPA v2 assay for HPV types 6, 11, 16, and 18, for example, were 0.806, 0.646, 0.920, and 0.860, respectively; the specificities were 0.986, 0.998, 0.960, and 0.986, respectively. The overall comparability of detection of the 15 HPV types was quite high. Analyses of DNA genotype testing compared to cytology results demonstrated a significant discordance between cytology-negative (normal) and HPV DNA-positive results. This demonstrates the challenges of cytological diagnosis and the possibility that a significant number of HPV-infected cells may appear cytologically normal.

  16. Genomic Signatures of North American Soybean Improvement Inform Diversity Enrichment Strategies and Clarify the Impact of Hybridization

    Directory of Open Access Journals (Sweden)

    Justin N. Vaughn

    2016-09-01

    Full Text Available Crop improvement represents a long-running experiment in artificial selection on a complex trait, namely yield. How such selection relates to natural populations is unclear, but the analysis of domesticated populations could offer insights into the relative role of selection, drift, and recombination in all species facing major shifts in selective regimes. Because of the extreme autogamy exhibited by soybean (Glycine max, many “immortalized” genotypes of elite varieties spanning the last century have been preserved and characterized using ∼50,000 single nucleotide polymorphic (SNP markers. Also due to autogamy, the history of North American soybean breeding can be roughly divided into pre- and posthybridization eras, allowing for direct interrogation of the role of recombination in improvement and selection. Here, we report on genome-wide characterization of the structure and history of North American soybean populations and the signature of selection in these populations. Supporting previous work, we find that maturity defines population structure. Though the diversity of North American ancestors is comparable to available landraces, prehybridization line selections resulted in a clonal structure that dominated early breeding and explains many of the reductions in diversity found in the initial generations of soybean hybridization. The rate of allele frequency change does not deviate sharply from neutral expectation, yet some regions bare hallmarks of strong selection, suggesting a highly variable range of selection strengths biased toward weak effects. We also discuss the importance of haplotypes as units of analysis when complex traits fall under novel selection regimes.

  17. Genomic Signatures of North American Soybean Improvement Inform Diversity Enrichment Strategies and Clarify the Impact of Hybridization.

    Science.gov (United States)

    Vaughn, Justin N; Li, Zenglu

    2016-09-08

    Crop improvement represents a long-running experiment in artificial selection on a complex trait, namely yield. How such selection relates to natural populations is unclear, but the analysis of domesticated populations could offer insights into the relative role of selection, drift, and recombination in all species facing major shifts in selective regimes. Because of the extreme autogamy exhibited by soybean (Glycine max), many "immortalized" genotypes of elite varieties spanning the last century have been preserved and characterized using ∼50,000 single nucleotide polymorphic (SNP) markers. Also due to autogamy, the history of North American soybean breeding can be roughly divided into pre- and posthybridization eras, allowing for direct interrogation of the role of recombination in improvement and selection. Here, we report on genome-wide characterization of the structure and history of North American soybean populations and the signature of selection in these populations. Supporting previous work, we find that maturity defines population structure. Though the diversity of North American ancestors is comparable to available landraces, prehybridization line selections resulted in a clonal structure that dominated early breeding and explains many of the reductions in diversity found in the initial generations of soybean hybridization. The rate of allele frequency change does not deviate sharply from neutral expectation, yet some regions bare hallmarks of strong selection, suggesting a highly variable range of selection strengths biased toward weak effects. We also discuss the importance of haplotypes as units of analysis when complex traits fall under novel selection regimes.

  18. Fluorescence in situ hybridization-flow cytometry-cell sorting-based method for separation and enrichment of type I and type II methanotroph populations.

    Science.gov (United States)

    Kalyuzhnaya, Marina G; Zabinsky, Rebecca; Bowerman, Sarah; Baker, David R; Lidstrom, Mary E; Chistoserdova, Ludmila

    2006-06-01

    A fluorescence in situ hybridization-flow cytometry (FISH/FC)-based method was optimized using artificial mixtures of pure cultures of methanotrophic bacteria. Traditional oligonucleotide probes targeting 16S rRNAs of type I (MG84/705 probe) and type II (MA450 probe) methanotrophs were labeled with fluorescein or Alexa fluor and used for FISH, followed by fluorescence-activated FC analysis and cell sorting (FACS). The method resulted in efficient separation of target cells (type I or type II methanotrophs) from the artificial mixtures. The method was then applied for detection and enrichment of type I and type II methanotroph populations from a natural sample, Lake Washington sediment. Cells were extracted from the sediment, fixed, and subjected to FISH/FC/FACS. The resulting subpopulations were analyzed by reverse transcriptase PCR surveys of 16S rRNA, pmoA (encoding a subunit of particulate methane monooxygenase), and fae (encoding formaldehyde-activating enzyme) genes. The functional gene analysis indicated specific separation of the type I and type II methanotroph populations. 16S rRNA gene analysis revealed that type I methanotrophs comprised 59% of the subpopulation separated using the type I-specific probe and that type II methanotrophs comprised 47.5% of the subpopulation separated using the type II-specific probe. Our data indicate that the FISH/FC/FACS protocol described can provide significant enrichment of microbial populations of interest from complex natural communities and that these can be used for genetic tests. We further tested the possibility of direct whole-genome amplification (WGA) from limited numbers of sorted cells, using artificial mixtures of microbes whose genome sequences are known. We demonstrated that efficient WGA can be achieved using 10(4) or more cells separated by 16S rRNA-specific FISH/FC/FACS, while fewer cells resulted in less specific WGA.

  19. Detection of oligonucleotide hybridization on a single microparticle by time-resolved fluorometry: quantitation and optimization of a sandwich type assay.

    Science.gov (United States)

    Hakala, H; Mäki, E; Lönnberg, H

    1998-01-01

    Uniformly sized (50 micro m) porous glycidyl methacrylate/ethylene dimethacrylate particles (SINTEF) were used as the solid phase in a sandwich type mixed-phase hybridization assay based on time-resolved fluorescence detection on a single particle. These particles were coated with oligodeoxyribonucleotide probes by conventional phosphoramidite chain assembly. An oligodeoxyribonucleotide bearing a photoluminescent europium(III) chelate, ¿2,2',2",2"'-¿¿4'-¿4"'-[(4, 6-dichloro-1,3,5-triazin-2-yl)amino]phenyl¿-2,2':6',2"-terpyrid ine-6, 6"-diyl¿bis(methylenenitrilo)¿tetrakis(acetato)¿eur opi um(III), was hybridized to a complementary sequence of the target oligonucleotide, and the resulting duplex was further hybridized to the particle-bound probes. The latter binding was quantified by time-resolved measurement of the emission signal of a single particle. Kinetics of hybridization and the effect of the concentration of the target oligomer and the fluorescently tagged probe on the efficiency of hybridization were studied. The intensity of the emission signal was linearly related to the concentration of the target oligomer over a range of 5 orders of magnitude. The length of the complementary region between the target oligomer and the particle-bound probe was varied, and the effect of point mutations and deletions on the hybridization efficiency was determined in each case. The maximal selectivity was observed with 10-16-base pair complementary sequences, the optimal length depending on the oligonucleotide loading on the particle. Discrimination between the complete matches and point mismatches was unequivocal, a single point mutation and/or deletion decreasing the efficiency of hybridization by more than 2 orders of magnitude.

  20. Hybrid monolith sorbent of polypyrrole-coated graphene oxide incorporated into a polyvinyl alcohol cryogel for extraction and enrichment of sulfonamides from water samples.

    Science.gov (United States)

    Chullasat, Kochaporn; Nurerk, Piyaluk; Kanatharana, Proespichaya; Kueseng, Pamornrat; Sukchuay, Thanyaporn; Bunkoed, Opas

    2017-04-08

    A hybrid monolith sorbent of polypyrrole-coated graphene oxide embedded in polyvinyl alcohol cryogel was prepared and used as an effective solid phase extraction sorbent for the determination of trace sulfonamides. The large surface areas with many adsorption sites of polypyrrole and graphene oxide facilitated the high adsorption of sulfonamides via hydrogen bonding, π-π and hydrophobic interactions. The high porosity of the polyvinyl alcohol cryogel helped to reduce the back pressure that occurs in a conventional packed solid phase extraction cartridge. The effecting parameters on the extraction efficiency including the type of sorbent, the polymerization time, desorption conditions, the sample pH, the sample volume, the sample flow rate, and ionic strength were investigated and optimized. Under the optimum conditions, the developed method provided a wide linear range from 0.20 to 100.0 μg L(-1) for sulfadiazine, sulfathiazole and sulfamerazine; and from 0.10 to 100 μg L(-1) for sulfamethazine, sulfamonomethoxine and sulfadimethoxine. The limits of detection were 0.20 μg L(-1) for sulfadiazine, sulfathiazole and sulfamerazine; and 0.10 μg L(-1) for sulfamethazine, sulfamonomethoxine and sulfadimethoxine. The developed hybrid monolith polypyrrole-coated graphene oxide embedded in the polyvinyl alcohol cryogel sorbent provided good recoveries in the range of 85.5-99.0% with RSDs of less than 5.0%. The sorbent offered a good reproducibility, was robust and can be reused at least 10 times. It was successfully applied for the extraction and enrichment of sulfonamides from normal and supplemented water samples. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Investigation of Parameters that Affect the Success Rate of Microarray-Based Allele-Specific Hybridization Assays

    DEFF Research Database (Denmark)

    Poulsen, Lena; Søe, Martin Jensen; Moller, Lisbeth Birk

    2011-01-01

    Background: The development of microarray-based genetic tests for diseases that are caused by known mutations is becoming increasingly important. The key obstacle to developing functional genotyping assays is that such mutations need to be genotyped regardless of their location in genomic regions...... buffers with different stringencies in a custom-made microfluidic system. The data were used to assess which parameters play significant roles in assay development. Conclusions: Several assay development methods found suitable probes and assay conditions for a functional test for all investigated mutation...... sites. Probe length, probe spacer length, and assay stringency sufficed as variable parameters in the search for a functional multiplex assay. We discuss the optimal assay development methods for several different scenarios....

  2. Development of a Rapid Reverse Blot Hybridization Assay for Detection of Clinically Relevant Antibiotic Resistance Genes in Blood Cultures Testing Positive for Gram-Negative Bacteria.

    Science.gov (United States)

    Wang, Hye-Young; Yoo, Gilsung; Kim, Juwon; Uh, Young; Song, Wonkeun; Kim, Jong Bae; Lee, Hyeyoung

    2017-01-01

    Rapid and accurate identification of the causative pathogens of bloodstream infections is crucial for the prompt initiation of appropriate antimicrobial therapy to decrease the related morbidity and mortality rates. The aim of this study was to evaluate the performance of a newly developed PCR-reverse blot hybridization assay (REBA) for the rapid detection of Gram-negative bacteria (GNB) and their extended-spectrum β-lactamase (ESBL), AmpC β-lactamase, and carbapenemase resistance genes directly from the blood culture bottles. The REBA-EAC (ESBL, AmpC β-lactamase, carbapenemase) assay was performed on 327 isolates that were confirmed to have an ESBL producer phenotype, 200 positive blood culture (PBCs) specimens, and 200 negative blood culture specimens. The concordance rate between the results of REBA-EAC assay and ESBL phenotypic test was 94.2%. The sensitivity, specificity, positive predictive value, and negative predictive value of the REBA-EAC assay for GNB identification in blood culture specimens were 100% (95% CI 0.938-1.000, P < 0.001), 100% (95% CI 0.986-1.000, P < 0.001), 100% (95% CI 0.938-1.000, P < 0.001), and 100% (95% CI 0.986-1.000, P < 0.001), respectively. All 17 EAC-producing GNB isolates from the 73 PBCs were detected by the REBA-EAC assay. The REBA-EAC assay allowed easy differentiation between EAC and non-EAC genes in all isolates. Moreover, the REBA-EAC assay was a rapid and reliable method for identifying GNB and their β-lactamase resistance genes in PBCs. Thus, this assay may provide essential information for accelerating therapeutic decisions to achieve earlier appropriate antibiotic treatment during the acute phase of bloodstream infection.

  3. Performance of the Aptima High-Risk Human Papillomavirus mRNA Assay in a Referral Population in Comparison with Hybrid Capture 2 and Cytology▿

    Science.gov (United States)

    Clad, Andreas; Reuschenbach, Miriam; Weinschenk, Johanna; Grote, Ruth; Rahmsdorf, Janina; Freudenberg, Nikolaus

    2011-01-01

    This study compared the Aptima human papillomavirus (HPV) (AHPV; Gen-Probe Incorporated) assay, which detects E6/E7 mRNA from 14 high-risk types, the Hybrid Capture 2 HPV DNA (HC2; Qiagen Incorporated) test, and repeat cytology for their ability to detect high-grade cervical lesions (cervical intraepithelial neoplasia grade 2+ [CIN2+]) in women referred to colposcopy due to an abnormal Papanicolaou (Pap) smear. A total of 424 clinical specimens, stored in liquid-based cytology (LBC) vials at room temperature for up to 3 years, were tested by repeat cytology, the AHPV assay, and the HC2 test. Assay results were compared to each other and to histology results. The overall agreement between the AHPV assay and the HC2 test was 88.4%. The sensitivity (specificity) of cytology, the HC2 test, and the AHPV assay for the detection of CIN2+ was 84.9% (66.3%), 91.3% (61.0%), and 91.7% (75.0%) and for the detection of CIN3+ was 93.9% (54.4%), 95.7% (46.0%), and 98.2% (56.3%), respectively. Of the disease-positive specimens containing high-risk HPV (HR HPV) DNA as determined by Linear Array (Roche Diagnostics), the AHPV assay missed 3 CIN2 and 1 microfocal CIN3 specimen, while the HC2 test missed 6 CIN2, 4 CIN3, and 1 cervical carcinoma specimen. The AHPV assay had a sensitivity similar to but a specificity significantly higher (P < 0.0001) than the HC2 test for the detection of CIN2+. The AHPV assay was significantly more sensitive (P = 0.0041) and significantly more specific (P = 0.0163) than cytology for the detection of disease (CIN2+). PMID:21191046

  4. Performance of the Aptima high-risk human papillomavirus mRNA assay in a referral population in comparison with Hybrid Capture 2 and cytology.

    Science.gov (United States)

    Clad, Andreas; Reuschenbach, Miriam; Weinschenk, Johanna; Grote, Ruth; Rahmsdorf, Janina; Freudenberg, Nikolaus

    2011-03-01

    This study compared the Aptima human papillomavirus (HPV) (AHPV; Gen-Probe Incorporated) assay, which detects E6/E7 mRNA from 14 high-risk types, the Hybrid Capture 2 HPV DNA (HC2; Qiagen Incorporated) test, and repeat cytology for their ability to detect high-grade cervical lesions (cervical intraepithelial neoplasia grade 2+ [CIN2+]) in women referred to colposcopy due to an abnormal Papanicolaou (Pap) smear. A total of 424 clinical specimens, stored in liquid-based cytology (LBC) vials at room temperature for up to 3 years, were tested by repeat cytology, the AHPV assay, and the HC2 test. Assay results were compared to each other and to histology results. The overall agreement between the AHPV assay and the HC2 test was 88.4%. The sensitivity (specificity) of cytology, the HC2 test, and the AHPV assay for the detection of CIN2+ was 84.9% (66.3%), 91.3% (61.0%), and 91.7% (75.0%) and for the detection of CIN3+ was 93.9% (54.4%), 95.7% (46.0%), and 98.2% (56.3%), respectively. Of the disease-positive specimens containing high-risk HPV (HR HPV) DNA as determined by Linear Array (Roche Diagnostics), the AHPV assay missed 3 CIN2 and 1 microfocal CIN3 specimen, while the HC2 test missed 6 CIN2, 4 CIN3, and 1 cervical carcinoma specimen. The AHPV assay had a sensitivity similar to but a specificity significantly higher (P < 0.0001) than the HC2 test for the detection of CIN2+. The AHPV assay was significantly more sensitive (P = 0.0041) and significantly more specific (P = 0.0163) than cytology for the detection of disease (CIN2+).

  5. Development of a Rapid Reverse Blot Hybridization Assay for Detection of Clinically Relevant Antibiotic Resistance Genes in Blood Cultures Testing Positive for Gram-Negative Bacteria

    Science.gov (United States)

    Wang, Hye-young; Yoo, Gilsung; Kim, Juwon; Uh, Young; Song, Wonkeun; Kim, Jong Bae; Lee, Hyeyoung

    2017-01-01

    Rapid and accurate identification of the causative pathogens of bloodstream infections is crucial for the prompt initiation of appropriate antimicrobial therapy to decrease the related morbidity and mortality rates. The aim of this study was to evaluate the performance of a newly developed PCR-reverse blot hybridization assay (REBA) for the rapid detection of Gram-negative bacteria (GNB) and their extended-spectrum β-lactamase (ESBL), AmpC β-lactamase, and carbapenemase resistance genes directly from the blood culture bottles. The REBA-EAC (ESBL, AmpC β-lactamase, carbapenemase) assay was performed on 327 isolates that were confirmed to have an ESBL producer phenotype, 200 positive blood culture (PBCs) specimens, and 200 negative blood culture specimens. The concordance rate between the results of REBA-EAC assay and ESBL phenotypic test was 94.2%. The sensitivity, specificity, positive predictive value, and negative predictive value of the REBA-EAC assay for GNB identification in blood culture specimens were 100% (95% CI 0.938–1.000, P < 0.001), 100% (95% CI 0.986–1.000, P < 0.001), 100% (95% CI 0.938–1.000, P < 0.001), and 100% (95% CI 0.986–1.000, P < 0.001), respectively. All 17 EAC-producing GNB isolates from the 73 PBCs were detected by the REBA-EAC assay. The REBA-EAC assay allowed easy differentiation between EAC and non-EAC genes in all isolates. Moreover, the REBA-EAC assay was a rapid and reliable method for identifying GNB and their β-lactamase resistance genes in PBCs. Thus, this assay may provide essential information for accelerating therapeutic decisions to achieve earlier appropriate antibiotic treatment during the acute phase of bloodstream infection. PMID:28232823

  6. Multiplex Real-Time PCR Assays for Screening of Shiga Toxin 1 and 2 Genes, Including All Known Subtypes, and Escherichia coli O26-, O111-, and O157-Specific Genes in Beef and Sprout Enrichment Cultures.

    Science.gov (United States)

    Harada, Tetsuya; Iguchi, Atsushi; Iyoda, Sunao; Seto, Kazuko; Taguchi, Masumi; Kumeda, Yuko

    2015-10-01

    Shiga toxin family members have recently been classified using a new nomenclature into three Stx1 subtypes (Stx1a, Stx1c, and Stx1d) and seven Stx2 subtypes (Stx2a, Stx2b, Stx2c, Stx2d, Stx2e, Stx2f, and Stx2g). To develop screening methods for Stx genes, including all of these subtype genes, and Escherichia coli O26-, O111-, and O157-specific genes in laboratory investigations of Shiga toxin-producing E. coli (STEC) foodborne cases, we developed multiplex real-time PCR assays and evaluated their specificity and quantitative accuracy using STEC and non-STEC isolates, recombinant plasmids, and food enrichment cultures and by performing STEC spiking experiments with beef and sprout enrichment cultures. In addition, we evaluated the relationship between the recovery rates of the target strains by direct plating and immunomagnetic separation and the cycle threshold (CT) values of the real-time PCR assays for the Stx subtypes and STEC O26, O111, and O157 serogroups. All three stx1- and seven stx2-subtype genes were detected by real-time PCR with high sensitivity and specificity, and the quantitative accuracy of this assay was confirmed using control plasmids and STEC spiking experiments. The results of the STEC spiking experiments suggest that it is not routinely possible to isolate STEC from enrichment cultures with real-time PCR CT values greater than 30 by direct plating on MacConkey agar, although highly selective media and immunomagnetic beads were able to isolate the inoculated strains from the enrichment cultures. These data suggest that CT values obtained from the highly quantitative real-time PCR assays developed in this study provide useful information to develop effective isolation strategies for STEC from food samples. The real-time PCR assays developed here are expected to aid in investigations of infections or outbreaks caused by STEC harboring any of the stx-subtype genes in the new Stx nomenclature, as well as STEC O26, O111, and O157.

  7. A sandwich-hybridization assay for simultaneous determination of HIV and tuberculosis DNA targets based on signal amplification by quantum dots-PowerVision™ polymer coding nanotracers.

    Science.gov (United States)

    Yan, Zhongdan; Gan, Ning; Zhang, Huairong; Wang, De; Qiao, Li; Cao, Yuting; Li, Tianhua; Hu, Futao

    2015-09-15

    A novel sandwich-hybridization assay for simultaneous electrochemical detection of multiple DNA targets related to human immune deficiency virus (HIV) and tuberculosis (TB) was developed based on the different quantum dots-PowerVision(TM) polymer nanotracers. The polymer nanotracers were respectively fabricated by immobilizing SH-labeled oligonucleotides (s-HIV or s-TB), which can partially hybrid with virus DNA (HIV or TB), on gold nanoparticles (Au NPs) and then modified with PowerVision(TM) (PV) polymer-encapsulated quantum dots (CdS or PbS) as signal tags. PV is a dendrimer enzyme linked polymer, which can immobilize abundant QDs to amplify the stripping voltammetry signals from the metal ions (Pb or Cd). The capture probes were prepared through the immobilization of SH-labeled oligonucleotides, which can complementary with HIV and TB DNA, on the magnetic Fe3O4@Au (GMPs) beads. After sandwich-hybridization, the polymer nanotracers together with HIV and TB DNA targets were simultaneously introduced onto the surface of GMPs. Then the two encoding metal ions (Cd(2+) and Pb(2+)) were used to differentiate two viruses DNA due to the different subsequent anodic stripping voltammetric peaks at -0.84 V (Cd) and -0.61 V (Pb). Because of the excellent signal amplification of the polymer nanotracers and the great specificity of DNA targets, this assay could detect targets DNA as low as 0.2 femtomolar and exhibited excellent selectivity with the dynamitic range from 0.5 fM to 500 pM. Those results demonstrated that this electrochemical coding assay has great potential in applications for screening more viruses DNA while changing the probes.

  8. Rapid identification of Salmonella serovars in feces by specific detection of virulence genes, invA and spvC, by an enrichment broth culture-multiplex PCR combination assay.

    Science.gov (United States)

    Chiu, C H; Ou, J T

    1996-10-01

    In order to make a rapid and definite diagnosis of Salmonella enteritis in children, an enrichment broth culture-multiplex PCR combination assay was devised to identify Salmonella serovars directly from fecal samples. Two pairs of oligonucleotide primers were prepared according to the sequences of the chromosomal invA and plasmid spvC genes. PCR with these two primers would produce either one amplicon (from the invA gene) or two amplicons (from the invA and spvC genes), depending on whether or not the Salmonella bacteria contained a virulence plasmid. The fecal sample was diluted 10- to 20-fold into gram-negative enrichment broth and incubated to eliminate inhibitory compounds and also to allow selective enrichment of the bacteria. One or two amplicons were obtained, the expected result if Salmonella bacteria were present. The detection limit of this PCR was about 200 bacteria per reaction mixture. The primers were specific, as no amplification products were obtained with 18 species and 22 isolates of non-Salmonella bacteria tested which could be present in the feces or cause contamination. In contrast, when 23 commonly seen Salmonella serovars (38 isolates) were tested, all were shown to carry the invA gene and seven concomitantly harbored the spvC gene of the virulence plasmid. This assay was applied to the diagnosis of Salmonella enteritis in 57 children who were suffering from mucoid and/or bloody diarrhea. Of the 57 children, 38 were PCR positive and 22 were culture positive. There were two culture-positive samples that were not detected by PCR. Thus, this PCR assay showed an efficiency of 95% (38 of 40), which is much higher than the 60% (24 of 40) by culture alone. Not only is this method more sensitive, rapid, and efficient but it will cause only an incremental increase in the cost of stool processing, since enrichment cultivation of fecal samples from diarrheal patients using gram-negative enrichment broth is a routine practice for identification in many

  9. A binary functional substrate for enrichment and ultrasensitive SERS spectroscopic detection of folic acid using graphene oxide/Ag nanoparticle hybrids.

    Science.gov (United States)

    Ren, Wen; Fang, Youxing; Wang, Erkang

    2011-08-23

    Herein graphene oxide/Ag nanoparticle hybrids (GO/PDDA/AgNPs) were fabricated according to a self-assembly procedure. Using the obtained GO/PDDA/AgNPs as SERS substrates, an ultrasensitive and label-free detection of folic acid in water and serum was demonstrated based on the inherent SERS spectra of folic acid. The modified graphene oxide exhibited strong enrichment of folic acid due to the electrostatic interaction, and the self-assembled Ag nanoparticles greatly enhanced the SERS spectra of folic acid, both of which led to an ultrahigh sensitivity. Therefore, although the SERS enhancement of p-ATP on GO/PDDA/AgNPs was weaker than that on Ag nanoparticles, the SERS signals of folic acid on GO/PDDA/AgNPs were much stronger than that on Ag nanoparticles. To improve the detection, the concentration of GO/PDDA/AgNPs was optimized to reduce background of the graphene oxide. The SERS spectra of the folic acid showed that the minimum detected concentration of folic acid in water was as low as 9 nM with a linear response range from 9 to 180 nM. To estimate the feasibility of the detection method based on GO/PDDA/AgNPs for the practical applications, diluted serum containing different concentrations of folic acid was taken as real samples. It was established that the sensitivity and the linear range for the folic acid in serum were comparable to that in water. This ultrasensitive and label-free SERS detection of folic acid based on GO/PDDA/AgNPs offers great potential for practical applications of medicine and biotechnology.

  10. Comparative study of ProEx C immunocytochemistry and UroVysion fluorescent in-situ hybridization assays on urine cytology specimens

    Directory of Open Access Journals (Sweden)

    Sue Chang

    2015-01-01

    Full Text Available Background: Detection of urothelial carcinoma (UC by urine cytology can be challenging. Recently, ProEx C has been studied as a marker to improve detection of UC. ProEx C is an assay targeting expression of topoisomerase IIa and minichromosome maintenance protein-2 and is currently utilized to assist in diagnoses of the gynecological specimens. In this study, we compared the utility of ProEx C and UroVysion in urine specimens. Materials and Methods: Twenty-seven urine specimens with UroVysion assay analysis and surgical biopsy follow-up were selected. The smears were stained with ProEx C. ProEx C and UroVysion assay results were separated into two categories based on surgical biopsy follow-up (benign or neoplastic. Surgical biopsy diagnoses were used as the gold standard for comparative evaluation of the two assays. The surgical follow-up was 9 benign, 2 low grade, and 16 high grade UCs. Results: The sensitivity was 88.9% for ProEx C and 55.6% for UroVysion, while the specificity was 77.8% for ProEx C and 44.4% for UroVysion. Positive predictive value was 88.9% for ProEx C and 66.7% for UroVysion. Negative predictive value was 77.8% and 33.3% for ProEx C and UroVysion, respectively. Using the two-tailed paired t-test, P value of 0.033 was obtained when ProEx C stain was compared with the UroVysion assay. Conclusion: ProEx C immunocytochemistry has a more favorable performance than fluorescent in-situ hybridization with a significant difference between the two assays using paired two-tail t-test (P = 0.0033.

  11. Simultaneous Detection of CDC Category "A" DNA and RNA Bioterrorism Agents by Use of Multiplex PCR & RT-PCR Enzyme Hybridization Assays

    Directory of Open Access Journals (Sweden)

    Kelly J. Henrickson

    2009-10-01

    Full Text Available Assays to simultaneously detect multiple potential agents of bioterrorism are limited. Two multiplex PCR and RT-PCR enzyme hybridization assays (mPCR-EHA, mRT-PCR-EHA were developed to simultaneously detect many of the CDC category “A” bioterrorism agents. The “Bio T” DNA assay was developed to detect: Variola major (VM, Bacillus anthracis (BA, Yersinia pestis (YP, Francisella tularensis (FT and Varicella zoster virus (VZV. The “Bio T” RNA assay (mRT-PCR-EHA was developed to detect: Ebola virus (Ebola, Lassa fever virus (Lassa, Rift Valley fever (RVF, Hantavirus Sin Nombre species (HSN and dengue virus (serotypes 1-4. Sensitivity and specificity of the 2 assays were tested by using genomic DNA, recombinant plasmid positive controls, RNA transcripts controls, surrogate (spiked clinical samples and common respiratory pathogens. The analytical sensitivity (limit of detection (LOD of the DNA asssay for genomic DNA was 1×100~1×102 copies/mL for BA, FT and YP. The LOD for VZV whole organism was 1×10-2 TCID50/mL. The LOD for recombinant controls ranged from 1×102~1×103copies/mL for BA, FT, YP and VM. The RNA assay demonstrated LOD for RNA transcript controls of 1×104~1×106 copies/mL without extraction and 1×105~1×106 copies/mL with extraction for Ebola, RVF, Lassa and HSN. The LOD for dengue whole organisms was ~1×10-4 dilution for dengue 1 and 2, 1×104 LD50/mL and 1×102 LD50/mL for dengue 3 and 4. The LOD without extraction for recombinant plasmid DNA controls was ~1×103 copies/mL (1.5 input copies/reaction for Ebola, RVF, Lassa and HSN. No cross-reactivity of primers and probes used in both assays was detected with common respiratory pathogens or between targeted analytes. Clinical sensitivity was estimated using 264 surrogate clinical samples tested with the BioT DNA assay and 549 samples tested with the BioT RNA assay. The clinical specificity is 99.6% and 99.8% for BioT DNA assay and BioT RNA assay, respectively. The

  12. Quantitative bioanalysis of antibody-conjugated payload in monkey plasma using a hybrid immuno-capture LC-MS/MS approach: Assay development, validation, and a case study.

    Science.gov (United States)

    Liu, Ang; Kozhich, Alexander; Passmore, David; Gu, Huidong; Wong, Richard; Zambito, Frank; Rangan, Vangipuram S; Myler, Heather; Aubry, Anne-Françoise; Arnold, Mark E; Wang, Jian

    2015-10-01

    Antibody drug conjugates (ADCs) are complex molecules composed of two pharmacologically distinct components, the cytotoxic payload and the antibody. The measurement of the payload molecules that are attached to the antibody in vivo is important for the evaluation of the safety and efficacy of ADCs, and can also provide distinct information compared to the antibody-related analytes. However, analyzing the antibody-conjugated payload is challenging and in some cases may not be feasible. The in vivo change in drug antibody ratio (DAR), due to deconjugation, biotransformation or other clearance phenomena, generates unique and additional challenges for ADC analysis in biological samples. Here, we report a novel hybrid approach with immuno-capture of the ADC, payload cleavage by specific enzyme, and LC-MS/MS of the cleaved payload to quantitatively measure the concentration of payload molecules still attached to the antibody via linker in plasma. The ADC reference material used for the calibration curve is not likely to be identical to the ADC measured in study samples due to the change in DAR distribution over the PK time course. The assay clearly demonstrated that there was no bias in the measurement of antibody-conjugated payload for ADC with varying DAR, which thus allowed accurate quantification even when the DAR distribution dynamically changes in vivo. This hybrid assay was fully validated based on a combination of requirements for both chromatographic and ligand binding methods, and was successfully applied to support a GLP safety study in monkeys.

  13. Quantitative analysis of low-abundance serological proteins with peptide affinity-based enrichment and pseudo-multiple reaction monitoring by hybrid quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Kim, Kwang Hoe; Ahn, Yeong Hee; Ji, Eun Sun; Lee, Ju Yeon; Kim, Jin Young; An, Hyun Joo; Yoo, Jong Shin

    2015-07-02

    Multiple reaction monitoring (MRM) is commonly used for the quantitative analysis of proteins during mass pectrometry (MS), and has excellent specificity and sensitivity for an analyte in a complex sample. In this study, a pseudo-MRM method for the quantitative analysis of low-abundance serological proteins was developed using hybrid quadrupole time-of-flight (hybrid Q-TOF) MS and peptide affinity-based enrichment. First, a pseudo-MRM-based analysis using hybrid Q-TOF MS was performed for synthetic peptides selected as targets and spiked into tryptic digests of human serum. By integrating multiple transition signals corresponding to fragment ions in the full scan MS/MS spectrum of a precursor ion of the target peptide, a pseudo-MRM MS analysis of the target peptide showed an increased signal-to-noise (S/N) ratio and sensitivity, as well as an improved reproducibility. The pseudo-MRM method was then used for the quantitative analysis of the tryptic peptides of two low-abundance serological proteins, tissue inhibitor of metalloproteinase 1 (TIMP1) and tissue-type protein tyrosine phosphatase kappa (PTPκ), which were prepared with peptide affinity-based enrichment from human serum. Finally, this method was used to detect femtomolar amounts of target peptides derived from TIMP1 and PTPκ, with good coefficients of variation (CV 2.7% and 9.8%, respectively), using a few microliters of human serum from colorectal cancer patients. The results suggest that pseudo-MRM using hybrid Q-TOF MS, combined with peptide affinity-based enrichment, could become a promising alternative for the quantitative analysis of low-abundance target proteins of interest in complex serum samples that avoids protein depletion.

  14. Comparison of analytical and clinical performance of CLART HPV2 genotyping assay to Linear Array and Hybrid Capture 2

    DEFF Research Database (Denmark)

    Ejegod, Ditte Møller; Rebolj, Matejka; Bonde, Jesper

    2015-01-01

    prone to inter-observer variability. The reading of test results on the CLART HPV2 genotyping assay is, on the other hand, automated. The aim of our study was to directly compare the detection of HPV genotypes and high-grade cervical intraepithelial neoplasia (CIN) by CLART, Linear Array (LA...

  15. Fluorescence in situ hybridization assay detects upper urinary tract transitional cell carcinoma in patients with asymptomatic hematuria and negative urine cytology.

    Science.gov (United States)

    Huang, W T; Li, L Y; Pang, J; Ruan, X X; Sun, Q P; Yang, W J; Gao, X

    2012-01-01

    We evaluated the performance of a multiprobe FISH (fluorescence in situ hybridization) assay for noninvasive detection of upper urinary tract transitional cell carcinoma (UUT-TCC) in patients with asymptomatic hematuria and negative urine cytology. Voided urine samples from 285 patients with asymptomatic hematuria and negative urine cytology were prospectively analyzed by FISH technique. FISH assays were performed to detect chromosomal changes frequently associated with TCC, including aneuploidy of chromosomes 3, 7 and 17, and loss of the 9p21 locus. Eleven (3.9%) had a positive FISH result. Of the 11 patients, nine (81.8%) were found to have a TCC of the upper urinary tract, while no patients with negative FISH findings were found to have UUT-TCC. In this selected cohort, the sensitivity and specificity of FISH for the detection of UUT-TCC was 100% and 99.3%, respectively. Our preliminary data suggest that the clinical utility of FISH assay of chromosomes 3, 7, 9, and 17 as a noninvasive ancillary test for the diagnosis of UUT-TCC in a selected patient population with asymptomatic hematuria and negative urine cytology and by significant high sensitivity and specificity may be a reliable diagnostic approach for early detection of UUT-TCC patients. Further larger prospective and multicenter trials are needed to confirm our results.

  16. Development and Evaluation of a Rapid, Simple, and Sensitive Immunochromatographic Assay To Detect Thermostable Direct Hemolysin Produced by Vibrio parahaemolyticus in Enrichment Cultures of Stool Specimens

    OpenAIRE

    Kawatsu, Kentaro; Ishibashi, Masanori; Tsukamoto,Teizo

    2006-01-01

    Thermostable direct hemolysin (TDH) is considered to be a major virulence factor in Vibrio parahaemolyticus, and most cases of V. parahaemolyticus diarrhea in humans are caused by tdh gene-positive strains. In the present study, we developed an immunochromatographic assay to detect TDH (TDH-ICA) and evaluated the utility of TDH-ICA for the diagnosis of V. parahaemolyticus diarrhea. TDH-ICA allowed the detection of 0.2 ng/ml of TDH within 10 min. Fecal homogenates were spiked with various numb...

  17. Assessment of biological and colony hybridization assays for detection of the aerobactin system in Escherichia coli from urinary tract infections.

    Science.gov (United States)

    Orskov, I; Williams, P H; Svanborg Edén, C; Orskov, F

    1989-01-01

    A total of 466 E. coli strains from urinary tract infections (UTI) were screened for the presence and expression of the aerobactin system by a colony hybridization test and a bioassay. A probe carrying part of the genes for aerobactin synthesis was used. A total of 43.1% (201) of the strains were positive in the probe test and undoubtedly positive in the bioassay. When doubtfully positive bioassays were included, this figure rose to 49.8% (232). An additional 4.9% (23) of the strains were positive in the colony hybridization test only while 44% (205) of the strains were negative in both tests. Doubtfully positive bioassays were probably due either to a false positive reaction or to a weak expression of the aerobactin system. 01:K1:H- strains were characteristically probe positive and doubtfully positive in the bioassay. The incidence of isolates positive by both methods or by only one of them was significantly higher among isolates from cases of pyelonephritis (Py) than among those from asymptomatic bacteriuria (ABU) and normal feces (FN) (P less than 0.01).

  18. Detection of plum pox virus by enzyme-linked immunosorbent assay in some apricot and peach varieties and hybrids in Romania.

    Science.gov (United States)

    Toma, S; Isac, M; Balan, V; Ivascu, A

    1998-09-01

    Plum pox virus (PPV) is a potyvirus widely spread in many species of the Prunus genus such as plum, apricot, peach, sweet cherry and others. This potyvirus causes great damage to stone fruit trees in Romania and other European countries as Hungary, Italy, Czech Republic, France, Spain, Greece, Turkey, and Slovak Republic. The Research Station for Fruit Tree Growing Baneasa in Bucharest has realized many studies on the epidemiology and spread of PPV and also on the disease symptomatology and detection possibilities. The control of sharka disease by sanitary selection measures requires corresponding detection techniques. The aim of this study was to determine the presence or absence of PPV in some apricot and peach varieties and hybrids in 1995-1997 by the enzyme-linked immunosorbent assay (ELISA) and to verify if some of our biological materials evaluated as symptom-free under field conditions for many years are also virus-free and can be considered healthy.

  19. A reliable solid phase microextraction-gas chromatography-triple quadrupole mass spectrometry method for the assay of selenomethionine and selenomethylselenocysteine in aqueous extracts: difference between selenized and not-enriched selenium potatoes.

    Science.gov (United States)

    Gionfriddo, Emanuela; Naccarato, Attilio; Sindona, Giovanni; Tagarelli, Antonio

    2012-10-17

    A new analytical approach is exploited in the assay of selenium speciation in selenized and not selenium enriched potatoes based on the widely available solid-phase microextraction (SPME) coupled to-GC-triple quadrupole mass spectrometry (SPME-GC-QqQ MS) method. The assay of selenomethionine (SeMet) and selenomethylselenocysteine (SeMeSeCys) in potatoes here reported provides clues to the effectiveness of SPME technique combined with gas chromatography-tandem mass spectrometry, which could be of general use. For the exploitation of the GC method, the selected analytes were converted into their N(O,S)-alkoxycarbonyl alkyl esters derivatives by direct treatment with alkyl chloroformate in aqueous extracts. The performance of five SPME fibers and three chloroformates were tested in univariate mode and the best results were obtained using the divinylbenzene/carboxen/polydimethylsiloxane fiber and propylchloroformate. The variables affecting the efficiency of SPME analysis were optimized by the multivariate approach of design of experiment (DoE) and, in particular, a central composite design (CCD) was applied. Tandem mass spectrometry in selected reaction monitoring (SRM) has allowed the elimination of matrix interferences, providing reconstructed chromatograms with well-resolved peaks and the achievement of very satisfactory detection and quantification limits. Both precision and recovery of the proposed protocol tested at concentration of 8 and 40 μg kg(-1) (dry matter), offered values ranging from 82.3 to 116.3% and from 8.5 to 13.1% for recovery and precision, respectively. The application of the method to commercial samples of selenized and not selenium enriched potatoes proved that the Se fertilization increases significantly the concentration of these bioavailable selenoamino acids.

  20. Screening of agonistic activities against four nuclear receptors in wastewater treatment plants in Japan using a yeast two-hybrid assay

    Institute of Scientific and Technical Information of China (English)

    Daisuke Inoue; Koki Nakama; Kazuko Sawada; Taro Watanabe; Hisae Matsui; Kazunari Sei; Tsuyoshi Nakanishi; Michihiko Ike

    2011-01-01

    To assess the potential endocrine disruptive effects through multiple nuclear receptors (NRs), especially non-steroidal NRs, in municipal wastewater, we examined the agonistic activities on four NRs (estrogen receptor c, thyroid hormone receptor α, retinoic acid receptor α and retinoid X receptor α) of untreated and treated wastewater from municipal wastewater treatment plants (WWTPs) in Japan using a yeast two-hybrid assay.Investigation of the infiuent and effluent of seven WWTPs revealed that agonistic activities against steroidal and non-steroidal NRs were always detected in the infiuents and partially remained in the effluents.Further investigation of four WWTPs employing conventional activated sludge, pseudo-anoxic-oxic, anoxic-oxic and anaerobic-anoxic-oxic processes revealed that the ability to reduce the agonistic activity against each of the four NRs varies depending on the treatment process.These results indicated that municipal wastewater in Japan commonly contains endocrine disrupting chemicals that exert agonistic activities on steroidal and non-steroidal NRs, and that some of these chemicals are released into the natural aquatic environment.Although the results obtained in yeast assays suggested that measured levels of non-steroidal NR agonists in the effluent of WWTPs were not likely to cause any biological effect, further study is required to assess their possible risks in detail.

  1. Data on regulation of the gene for the adipocyte-enriched micropeptide Adig/Smaf1 by qPCR analysis and luciferase reporter assay

    Directory of Open Access Journals (Sweden)

    Gang Ren

    2016-12-01

    Full Text Available This article describes qPCR analysis for the Adig/Smaf1 gene in multiple in vitro adipocyte differentiation models including white and brown adipogenesis, cell lines and primary cultures. The article also contains qPCR data for transcript levels of Adig/Smaf1 in a wide panel of murine tissues. Expression of Adig/Smaf1 transcript in white and brown adipose tissue in fasted and refed mice is reported and also data for Adig/Smaf1 transcript expression in genetically obese ob/ob mice. Data on the effects of siRNA-mediated knockdown of Srebp1c on Adig/Smaf1 transcript levels in 3T3-L1 adipocytes are shown. Luciferase reporter assays provide data for regulation of an ~ 2 kb fragment of the 5′ flanking region of Adig/Smaf1 gene by PPARγ/RXRα. This data is related to a research article describing Adig/Smaf1 protein expression, “Expression, regulation and functional assessment of the 80 amino acid Small Adipocyte Factor 1 (Smaf1 protein in adipocytes” (G. Ren, P. Eskandari, S. Wang, C.M. Smas, 2016 [1].

  2. A sensitive and selective LC-MS/MS analysis coupled with an online sample enrichment technique for H295R steroidogenesis assay and its application in the investigation of the effect of sildenafil on steroidogenesis.

    Science.gov (United States)

    Kang, Soyoung; Park, Sol; Kim, Mi Jie; Oh, Seung Min; Chung, Kyu Hyuck; Lee, Sooyeun

    2013-11-01

    An in vitro steroidogenesis assay using H295R human adenocarcinoma cells is a useful tool for the fast identification of compounds that affect the production of testosterone and 17β-estradiol. Selective and sensitive hormone measurement by liquid chromatography-tandem mass spectrometry (LC-MS/MS) can make this assay more reliable. Therefore, in the present study, a sensitive and selective method for the quantification of testosterone and 17β-estradiol in the H295R steroidogenesis assay was developed and fully validated using LC-MS/MS coupled with an online sample enrichment technique. To prove its usefulness, the method developed was applied to investigate the effect of sildenafil on steroidogenesis. Cell medium samples were diluted and prepared using solid-phase extraction. The samples were prepared on ice and were not kept for more than 30 min to prevent degradation of hormones. The extracts were dried, reconstituted, filtered, and analyzed by LC-MS/MS with polarity switching electrospray ionization. The validation results for selectivity, matrix effect, recovery, linearity, precision, and accuracy were satisfactory. The limits of detection for testosterone and 17β-estradiol were 5 and 10 pg/mL, respectively, and the limit of quantification for both testosterone and 17β-estradiol was 10 pg/mL, which was in accordance with the OECD guideline. No degradation was observed under the storage conditions for 7 and 14 days at -80 °C as well as after three freeze-thaw cycles, whereas 17β-estradiol was degraded after 1 h on ice during sample processing. The method developed was successfully used for the investigation of the effect of sildenafil on steroidogenesis. This method can be very useful for the initial selection of drugs with androgenic and/or estrogenic effects for specific purposes, e.g., in the selection of drugs that are used to reverse the effects of chemical castration.

  3. Development of in situ hybridization and PCR assays for the detection of Enterocytozoon hepatopenaei (EHP), a microsporidian parasite infecting penaeid shrimp.

    Science.gov (United States)

    Tang, Kathy F J; Pantoja, Carlos R; Redman, Rita M; Han, Jee Eun; Tran, Loc H; Lightner, Donald V

    2015-09-01

    A microsporidian parasite, Enterocytozoon hepatopenaei (abbreviated as EHP), is an emerging pathogen for penaeid shrimp. EHP has been found in several shrimp farming countries in Asia including Vietnam, Thailand, Malaysia, Indonesia and China, and is reported to be associated with growth retardation in farmed shrimp. We examined the histological features from infected shrimp collected from Vietnam and Brunei, these include the presence of basophilic inclusions in the hepatopancreas tubule epithelial cells, in which EHP is found at various developmental stages, ranging from plasmodia to mature spores. By a PCR targeting the 18S rRNA gene, a 1.1kb 18S rRNA gene fragment of EHP was amplified, and this sequence showed a 100% identity to EHP found in Thailand and China. This fragment was cloned and labeled with digoxigenin-11-dUTP, and in situ hybridized to tissue sections of infected Penaeus vannamei (from Vietnam) and P. stylirostris (Brunei). The results of in situ hybridization were specific, the probe only reacted to the EHP within the cytoplasmic inclusions, not to a Pleistophora-like microsporidium that is associated with cotton shrimp disease. Subsequently, we developed a PCR assay from this 18S rRNA gene region, this PCR is shown to be specific to EHP, did not react to 2 other parasitic pathogens, an amoeba and the cotton shrimp disease microsporidium, nor to genomic DNA of various crustaceans including polychaetes, squids, crabs and krill. EHP was detected, through PCR, in hepatopancreatic tissue, feces and water sampled from infected shrimp tanks, and in some samples of Artemia biomass. Copyright © 2015 Elsevier Inc. All rights reserved.

  4. Locked Nucleic Acid-Based In Situ Hybridization Reveals miR-7a as a Hypothalamus-Enriched MicroRNA with a Distinct Expression Pattern

    DEFF Research Database (Denmark)

    Herzer, S; Silahtaroglu, A; Meister, B

    2012-01-01

    present in the hypothalamus, miR-7a, was the only miRNA found to be enriched in the hypothalamus, with low or no expression in other parts of the central nervous system (CNS). Within the hypothalamus, strong miR-7a expression was distinct and restricted to some hypothalamic nuclei and adjacent areas. mi...

  5. Evaluation of the Punch-it™ NA-Sample kit for detecting microbial DNA in blood culture bottles using PCR-reverse blot hybridization assay.

    Science.gov (United States)

    Kim, Jungho; Wang, Hye-Young; Kim, Seoyong; Park, Soon Deok; Yu, Kwangmin; Kim, Hyo Youl; Uh, Young; Lee, Hyeyoung

    2016-09-01

    DNA extraction efficiency affects the success of PCR-based method applications. The Punch-it™ NA-Sample kit for extracting DNA by using paper chromatography is technically easy to use and requires just two reagents and only 10min to complete. The Punch-it™ NA-Sample kit could be offered as a rapid, accurate, and convenient method for extracting bacterial and fungal DNA from blood culture bottles. We compared the efficiencies of the commercial kit (Punch-it™ NA-Sample kit) and an in-house conventional boiling method with Chelex-100 resin for DNA extraction from blood culture bottles. The efficiency of the two DNA extraction methods was assessed by PCR-reverse blot hybridization assay (PCR-REBA, REBA Sepsis-ID) for detecting Gram positive (GP) bacteria, Gram negative (GN) bacteria, and Candida species with 196 positive and 200 negative blood culture bottles. The detection limits of the two DNA extraction methods were 10(3)CFU/mL for GP bacteria, 10(3)CFU/mL for GN bacteria, and 10(4)CFU/mL for Candida. The sensitivity and specificity of the Punch-it™ NA-Sample kit by REBA Sepsis-ID were 95.4% (187/196) and 100% (200/200), respectively. The overall agreement of the two DNA extraction methods was 98.9% (392/396). Three of four samples showing discrepant results between the two extraction methods were more accurately matched up with the Punch-it™ NA-Sample kit based on conventional culture methods. The results indicated that the Punch-it™ NA-Sample kit extracted bacterial and fungal DNA in blood culture bottles and allowed extracted DNA to be used in molecular assay. Copyright © 2016. Published by Elsevier B.V.

  6. Bioinspired near-infrared-excited sensing platform for in vitro antioxidant capacity assay based on upconversion nanoparticles and a dopamine-melanin hybrid system.

    Science.gov (United States)

    Wang, Dong; Chen, Chuan; Ke, Xuebin; Kang, Ning; Shen, Yuqing; Liu, Yongliang; Zhou, Xi; Wang, Hongjun; Chen, Changqing; Ren, Lei

    2015-02-11

    A novel core-shell structure based on upconversion fluorescent nanoparticles (UCNPs) and dopamine-melanin has been developed for evaluation of the antioxidant capacity of biological fluids. In this approach, dopamine-melanin nanoshells facilely formed on the surface of UCNPs act as ultraefficient quenchers for upconversion fluorescence, contributing to a photoinduced electron-transfer mechanism. This spontaneous oxidative polymerization of the dopamine-induced quenching effect could be effectively prevented by the presence of various antioxidants (typically biothiols, ascorbic acid (Vitamin C), and Trolox). The chemical response of the UCNPs@dopamine-melanin hybrid system exhibited great selectivity and sensitivity toward antioxidants relative to other compounds at 100-fold higher concentration. A satisfactory correlation was established between the ratio of the "anti-quenching" fluorescence intensity and the concentration of antioxidants. Besides the response of the upconversion fluorescence signal, a specific evaluation process for antioxidants could be visualized by the color change from colorless to dark gray accompanied by the spontaneous oxidation of dopamine. The near-infrared (NIR)-excited UCNP-based antioxidant capacity assay platform was further used to evaluate the antioxidant capacity of cell extracts and human plasma, and satisfactory sensitivity, repeatability, and recovery rate were obtained. This approach features easy preparation, fluorescence/visual dual mode detection, high specificity to antioxidants, and enhanced sensitivity with NIR excitation, showing great potential for screening and quantitative evaluation of antioxidants in biological systems.

  7. Detection of high-risk subtypes of human papillomavirus in cervical swabs: routine use of the Digene Hybrid Capture assay and polymerase chain reaction analysis.

    LENUS (Irish Health Repository)

    Brennan, M M

    2012-02-03

    Human papillomaviruses (HPVs) are major causative agents in the pathogenesis of cervical cancer, and more than twenty types are associated with its development. With the introduction of liquid-based preparation systems, it is envisaged that large-scale HPV testing will be established in the near future. Preliminary studies demonstrate the accessibility of these samples for DNA testing using both the Digene Hybrid Capture assay (DHCA) and polymerase chain reaction (PCR) techniques. This study aims to assess the validity and sensitivity of the DHCA system to detect high-risk HPV DNA, using two sets of HPV consensus primers (Gp5+\\/Gp6+ and MY09\\/MY11) in tandem with routine assessment of cervical smear and biopsy samples. Results indicate that the combination of DHCA and PCR detects more high-grade lesions than does the DHCA alone. DHCA-negative cases were categorised by subsequent PCR amplification into low-grade HPV-negative (12\\/16) cervical lesions and high-grade HPV-positive (7\\/9) cervical lesions. Gp5+\\/Gp6+ primers were less sensitive in detecting HPV-positive samples than was the MY09\\/MY11 primer set. These results support the use of high-risk HPV testing by DHCA, with subsequent analysis of DHCA-negative samples by PCR using the MY09\\/MY11 primers.

  8. Diagnosis of anaplastic lymphoma kinase rearrangement in cytological samples through a fluorescence in situ hybridization-based assay: Cytological smears versus cell blocks.

    Science.gov (United States)

    Zito Marino, Federica; Rossi, Giulio; Brunelli, Matteo; Malzone, Maria Gabriella; Liguori, Giuseppina; Bogina, Giuseppe; Morabito, Alessandro; Rocco, Gaetano; Franco, Renato; Botti, Gerardo

    2017-05-01

    Anaplastic lymphoma kinase (ALK) status analysis of lung cytological specimens should be successfully encouraged in routine practice because biopsy specimens are not always available. To date, the US Food and Drug Administration has approved both fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) as diagnostic tests for identifying ALK-positive patients eligible for treatment with crizotinib. Although ALK IHC is an optimal diagnostic tool, FISH becomes mandatory in equivocal cases. ALK FISH of paraffin-embedded tissue material is still the gold standard, whereas the cytological specimen assay has not yet been completely standardized. Many controversial data have been reported on the adequacy of cytology cell blocks (CBs) versus conventional smears for FISH testing. This review discusses some critical issues related to ALK FISH of cytological samples, including the triaging of collected specimens to optimize the material, the use of CBs versus conventional smears, and alternative methods for an ALK rearrangement diagnosis. Conventional smears have the advantages of an immediate evaluation, no probe tissue-related artifactual loss, no fixation-related alterations, and usually sufficient material for an analytic preparation. On the other hand, CBs have several advantages, including the appropriate conservation of the tissue architecture, an absence of problems related to cell overlapping, and the ability to evaluate neoplastic cells in a dark field. Cancer Cytopathol 2017;125:303-312. © 2017 American Cancer Society. © 2017 American Cancer Society.

  9. Facile synthesis of boronate-decorated polyethyleneimine-grafted hybrid magnetic nanoparticles for the highly selective enrichment of modified nucleosides and ribosylated metabolites.

    Science.gov (United States)

    Li, Hua; Shan, Yuanhong; Qiao, Lizhen; Dou, Abo; Shi, Xianzhe; Xu, Guowang

    2013-12-03

    Ribosylated metabolites, especially modified nucleosides, have been extensively evaluated as cancer-related biomarkers. Boronate adsorbents are considered to be promising materials for extracting them from complex matrices. However, the enrichment of ribosylated metabolites in low abundance is still a challenge due to the limited capacity and selectivity of the existing boronate adsorbents. In this study, a novel type of magnetic nanoparticles named Fe3O4@SiO2@PEI-FPBA was synthesized by grafting polyethyleneimine (PEI) onto the surface of Fe3O4@SiO2 before modification by boronate groups. The high density of the amino groups on the PEI chains supplied a large number of binding sites for boronate groups. Thus, the adsorption capacity (1.34 ± 0.024 mg/g) of the nanoparticles, which is 6- to 7-fold higher than that of analogous materials, was greatly improved. The unreacted secondary amines and tertiary amines of the PEI enhanced the aqueous solubility of the nanoparticles, which could efficiently reduce nonspecific adsorption. The nanoparticles were able to capture 1,2 cis-diol nucleosides from 1000-fold interferences. Moreover, the flexible chains of PEI were favorable for effective enrichment and quick equilibration (nanoparticles. Among them, 43 were identified to be nucleosides and other ribosylated metabolites. Nine low abundance modified nucleosides were detected for the first time. In conclusion, Fe3O4@SiO2@PEI-FPBA is an attractive candidate material for the highly selective enrichment of 1,2-cis-diol compounds.

  10. Study on separation and enrichment of rhodium in silver-containing materials by lead fire assay%铅试金分离富集含银物料中铑的研究

    Institute of Scientific and Technical Information of China (English)

    林海山; 唐维学; 戴凤英; 刘天平

    2013-01-01

    报道了一个改进的铅试金分离富集-ICP-AES测定含银物料中铑的方法.用钯粉或钯银粉混合物作灰吹保护剂,分别研究了钯与银的质量比、合粒中钯银与铑的质量比对分离富集合银物料中铑的影响.结果表明:当钯量:银量≥2:1时,铑在灰吹过程的损失可忽略;当钯银量:铑量≥40∶1时合粒中铑在常压下可以完全溶解;当银量:钯量≥1∶10时,合粒可以用HNO3(1+3)快速溶解.研究了钯量对电感耦合等离子体发射光谱法测定铑的影响.方法回收率98.2% ~ 100.5%,对实际样品进行分析,测定值与推荐值一致,相对标准偏差小于4.0%(n=5).%An improvement was made on the separation and enrichment of rhodium by lead fire assay with ICP-AES method. Using palladium powder or a mixture of palladium silver as protective reagent for cupellation, the effects of various testing factors, such as the mass ratio of palladium to silver and the mass ratio of palladium-silver to rhodium, have been studied in detail when rhodium was enriched. The results indicated that loss of rhodium could be ignored for cupellation process at Rh; Pd ≥ 2; 1, rhodium of kernel could be completely dissolved at atmospheric pressure and Pd-Ag:Rh≥40:1, kernel of precious metals could be dissolved with HNO3 (1 +3) at Ag:Pd≥l:10. The effects of palladium on the determination of rhodium with inductively coupled plasma atomic emission spectrometry had been studied. The recoveries of silver in samples ranged from 98. 2% to 100. 5%. Found values were consistent with recommended values, and the relative standard derivations (RSDs) were less than 4. 0 % ( n =5) in the real samples.

  11. A novel asymmetric-loop molecular beacon-based two-phase hybridization assay for accurate and high-throughput detection of multiple drug resistance-conferring point mutations in Mycobacterium tuberculosis.

    Science.gov (United States)

    Chen, Qinghai; Wu, Nan; Xie, Meng; Zhang, Bo; Chen, Ming; Li, Jianjun; Zhuo, Lisha; Kuang, Hong; Fu, Weiling

    2012-04-01

    The accurate and high-throughput detection of drug resistance-related multiple point mutations remains a challenge. Although the combination of molecular beacons with bio-immobilization technology, such as microarray, is promising, its application is difficult due to the ineffective immobilization of molecular beacons on the chip surface. Here, we propose a novel asymmetric-loop molecular beacon in which the loop consists of 2 parts. One is complementary to a target, while the other is complementary to an oligonucleotide probe immobilized on the chip surface. With this novel probe, a two-phase hybridization assay can be used for simultaneously detecting multiple point mutations. This assay will have advantages, such as easy probe availability, multiplex detection, low background, and high-efficiency hybridization, and may provide a new avenue for the immobilization of molecular beacons and high-throughput detection of point mutations.

  12. Label-Free and Enzyme-Free Homogeneous Electrochemical Biosensing Strategy Based on Hybridization Chain Reaction: A Facile, Sensitive, and Highly Specific MicroRNA Assay.

    Science.gov (United States)

    Hou, Ting; Li, Wei; Liu, Xiaojuan; Li, Feng

    2015-11-17

    Homogenous electrochemical biosensing strategies have attracted substantial attention, because of their advantages of being immobilization-free and having rapid response and improved recognition efficiency, compared to heterogeneous biosensors; however, the high cost of labeling and the strict reaction conditions of tool enzymes associated with current homogeneous electrochemical methods limit their potential applications. To address these issues, herein we reported, for the first time, a simple label-free and enzyme-free homogeneous electrochemical strategy based on hybridization chain reaction (HCR) for sensitive and highly specific detection of microRNA (miRNA). The target miRNA triggers the HCR of two species of metastable DNA hairpin probes, resulting in the formation of multiple G-quadruplex-incorporated long duplex DNA chains. Thus, with the electrochemical indicator Methylene Blue (MB) selectively intercalated into the duplex DNA chain and the multiple G-quadruplexes, a significant electrochemical signal drop is observed, which is dependent on the concentration of the target miRNA. Thus, using this "signal-off" mode, a simple, label-free and enzyme-free homogeneous electrochemical strategy for sensitive miRNA assay is readily realized. This strategy also exhibits excellent selectivity to distinguish even single-base mismatched miRNA. Furthermore, this method also exhibits additional advantages of simplicity and low cost, since both expensive labeling and sophisticated probe immobilization processes are avoided. Therefore, the as-proposed label-free and enzyme-free homogeneous electrochemical strategy may become an alternative method for simple, sensitive, and selective miRNA detection, and it has great potential to be applied in miRNA-related clinical diagnostics and biochemical research.

  13. Locked Nucleic Acid-Based In Situ Hybridization Reveals miR-7a as a Hypothalamus-Enriched MicroRNA with a Distinct Expression Pattern

    DEFF Research Database (Denmark)

    Herzer, S; Silahtaroglu, A; Meister, B

    2012-01-01

    MicroRNAs (miRNAs) are short (22 nucleotides) non-coding ribonucleic acid (RNA) molecules that post-transcriptionally repress expression of protein-coding genes by binding to 3'-untranslated regions of the target mRNAs. In order to identify miRNAs selectively expressed within the hypothalamus...... present in the hypothalamus, miR-7a, was the only miRNA found to be enriched in the hypothalamus, with low or no expression in other parts of the central nervous system (CNS). Within the hypothalamus, strong miR-7a expression was distinct and restricted to some hypothalamic nuclei and adjacent areas. mi......R-7a expression was particularly prominent in the subfornical organ, suprachiasmatic, paraventricular, periventricular, supraoptic, dorsomedial and arcuate nuclei. Identical expression patterns for miR-7a was seen in mouse and rat hypothalamus. By combining LNA-FISH with immunohistochemistry...

  14. The human interactome knowledge base (hint-kb): An integrative human protein interaction database enriched with predicted protein–protein interaction scores using a novel hybrid technique

    KAUST Repository

    Theofilatos, Konstantinos A.

    2013-07-12

    Proteins are the functional components of many cellular processes and the identification of their physical protein–protein interactions (PPIs) is an area of mature academic research. Various databases have been developed containing information about experimentally and computationally detected human PPIs as well as their corresponding annotation data. However, these databases contain many false positive interactions, are partial and only a few of them incorporate data from various sources. To overcome these limitations, we have developed HINT-KB (http://biotools.ceid.upatras.gr/hint-kb/), a knowledge base that integrates data from various sources, provides a user-friendly interface for their retrieval, cal-culatesasetoffeaturesofinterest and computesaconfidence score for every candidate protein interaction. This confidence score is essential for filtering the false positive interactions which are present in existing databases, predicting new protein interactions and measuring the frequency of each true protein interaction. For this reason, a novel machine learning hybrid methodology, called (Evolutionary Kalman Mathematical Modelling—EvoKalMaModel), was used to achieve an accurate and interpretable scoring methodology. The experimental results indicated that the proposed scoring scheme outperforms existing computational methods for the prediction of PPIs.

  15. Comparison of peptide nucleic acid fluorescence in situ hybridization assays with culture-based matrix-assisted laser desorption/ionization-time of flight mass spectrometry for the identification of bacteria and yeasts from blood cultures and cerebrospinal fluid cultures.

    Science.gov (United States)

    Calderaro, A; Martinelli, M; Motta, F; Larini, S; Arcangeletti, M C; Medici, M C; Chezzi, C; De Conto, F

    2014-08-01

    Peptide nucleic acid fluorescence in situ hybridization (PNA FISH) is a molecular diagnostic tool for the rapid detection of pathogens directly from liquid media. The aim of this study was to prospectively evaluate PNA FISH assays in comparison with culture-based matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) identification, as a reference method, for both blood and cerebrospinal fluid (CSF) cultures, during a 1-year investigation. On the basis of the Gram stain microscopy results, four different PNA FISH commercially available assays were used ('Staphylococcus aureus/CNS', 'Enterococcus faecalis/OE', 'GNR Traffic Light' and 'Yeasts Traffic Light' PNA FISH assays, AdvanDx). The four PNA FISH assays were applied to 956 positive blood cultures (921 for bacteria and 35 for yeasts) and 11 CSF cultures. Among the 921 blood samples positive for bacteria, PNA FISH gave concordant results with MALDI-TOF MS in 908/921 (98.64%) samples, showing an agreement of 99.4% in the case of monomicrobial infections. As regards yeasts, the PNA FISH assay showed a 100% agreement with the result obtained by MALDI-TOF MS. When PNA FISH assays were tested on the 11 CSF cultures, the results agreed with the reference method in all cases (100%). PNA FISH assays provided species identification at least one work-day before the MALDI-TOF MS culture-based identification. PNA FISH assays showed an excellent efficacy in the prompt identification of main pathogens, yielding a significant reduction in reporting time and leading to more appropriate patient management and therapy in cases of sepsis and severe infections.

  16. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas

    2013-01-21

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling the macroscopic observation. Such tests are most often used to explore antibody-antigen reactions. Agglutination has been used for protein assays using a biotin/streptavidin system as well as a hybridization based assay. The agglutination systems are prone to selftermination of the linking analyte, prone to active site saturation and loss of agglomeration at high analyte concentrations. We investigated the molecular target/ligand interaction, explaining the common agglutination problems related to analyte self-termination, linkage of the analyte to the same bead instead of different microbeads. We classified the agglutination process into three kinds of assays: a two- component assay, a three-component assay and a stepped three- component assay. Although we compared these three kinds of assays for recognizing DNA and protein molecules, the assay can be used for virtually any molecule, including ions and metabolites. In total, the optimized assay permits detecting analytes with high sensitivity in a short time, 5 min, at room temperature. Such a system is appropriate for POC testing.

  17. Detection of Human Papillomavirus DNA in Cervical Samples: Analysis of the New PGMY-PCR Compared To the Hybrid Capture II and MY-PCR Assays and a Two-Step Nested PCR Assay

    OpenAIRE

    Giovannelli, Lucia; Lama, Anna; Capra, Giuseppina; Giordano, Viviana; Aricò, Pietro; Ammatuna, Pietro

    2004-01-01

    The PGMY-PCR for human papillomavirus (HPV) was evaluated, in parallel with nested PCR (nPCR), in samples with noted Hybrid Capture II (HCII) and MY-PCR results. PGMY-PCR detected HPV DNA in 2.5% of HCII-negative-MY-PCR-negative samples and in 71.7% of HCII-positive-MY-PCR-negative samples; also, it detected the MY-PCR-negative-nPCR-negative types HPV-42, HPV-44, HPV-51, HPV-87, and HPV-89.

  18. Experimental conditions improving in-solution target enrichment for ancient DNA

    DEFF Research Database (Denmark)

    Cruz-Dávalos, Diana I.; Llamas, Bastien; Gaunitz, Charleen

    2017-01-01

    High-throughput sequencing has dramatically fostered ancient DNA research in recent years. Shotgun sequencing, however, does not necessarily appear as the best-suited approach due to the extensive contamination of samples with exogenous environmental microbial DNA. DNA capture-enrichment methods...... or the whole genome. We found that varying quantities of the starting probes had only moderate effect on capture outcomes. Starting DNA, probe tiling, the hybridization temperature and the proportion of endogenous DNA all affected the assay, however. Additionally, probe features such as their GC content...... extracted from degraded and ancient remains....

  19. Comparison of multiplex real-time PCR and PCR-reverse blot hybridization assay for the direct and rapid detection of bacteria and antibiotic resistance determinants in positive culture bottles.

    Science.gov (United States)

    Wang, Hye-Young; Kim, Seoyong; Kim, Jungho; Park, Soon Deok; Kim, Hyo Youl; Uh, Young; Lee, Hyeyoung

    2016-09-01

    The aim of this study was to evaluate the performance of a commercially available multiplex real-time PCR assay and a PCR-reverse blot hybridization assay (PCR-REBA) for the rapid detection of bacteria and identification of antibiotic resistance genes directly from blood culture bottles and to compare the results of these molecular assays with conventional culture methods. The molecular diagnostic methods were used to evaluate 593 blood culture bottles from patients with bloodstream infections. The detection positivity of multiplex real-time PCR assay for Gram-positive bacteria, Gram-negative bacteria and Candida spp. was equivalent to PCR-REBA as 99.6 %, 99.1 % and 100 %, respectively. Using conventional bacterial cultures as the gold standard, the sensitivity, specificity, positive predictive value and negative predictive value of these two molecular methods were 99.5 % [95 % confidence interval (CI), 0.980-1.000; PReal-methicillin-resistant Staphylococcusaureus multiplex real-time PCR assay targeting the mecA gene to detect methicillin resistance was lower than that of the PCR-REBA method, detecting an overall positivity of 98.4 % (n=182; 95 % CI, 0.964-1.000; P<0.009) and 99.5 % (n=184; 95 % CI, 0.985-1.000; P<0.0001), respectively. The entire two methods take about 3 h, while results from culture can take up to 48-72 h. Therefore, the use of these two molecular methods was rapid and reliable for the characterization of causative pathogens in bloodstream infections.

  20. A highly sensitive, multiplex broad-spectrum PCR-DNA-enzyme immunoassay and reverse hybridization assay for rapid detection and identification of Chlamydia trachomatis serovars.

    NARCIS (Netherlands)

    Quint, K.D.; Doorn, L.J. van; Kleter, B.; Koning, M.N. de; Munckhof, H.A. van den; Morre, S.A.; Harmsel, B. ter; Weiderpass, E.; Harbers, G.; Melchers, W.J.G.; Quint, W.G.V.

    2007-01-01

    Chlamydia trachomatis (Ct) comprises distinct serogroups and serovars. The present study evaluates a novel Ct amplification, detection, and genotyping method (Ct-DT assay). The Ct-DT amplification step is a multiplex broad-spectrum PCR for the cryptic plasmid and the VD2-region of ompl. The Ct-DT de

  1. Use of fluorescence in situ hybridization in combination with the comet assay in DNA damage and repa%彗星荧光原位杂交技术在检测特定基因损伤与修复中的应用

    Institute of Scientific and Technical Information of China (English)

    张炳珍; 王志萍

    2011-01-01

    @@ 彗星荧光原位杂交技术(fluorescence in situ hybridization in combination with the Comet assay,Comet-FISH)是彗星试验(Comet assay,又称单细胞凝胶电泳)和荧光原位杂交(fluorescence in situ hybridization,FISH)技术的结合,是一种在单细胞水平上检测特定基因损伤和修复的有效方法[1].

  2. Enrichment and Preservation of Architectural Knowledge

    DEFF Research Database (Denmark)

    Beetz, Jakob; Blümel, Ina; Dietze, Stefan

    2016-01-01

    approaches of semi-automatically deriving building models from point cloud data sets acquired from laser scans and the integration and overlay of such representations with explicit Building Information Models (BIM). We introduce novel ways for the further semantic enrichment of such hybrid building models...

  3. Comparison of three targeted enrichment strategies on the SOLiD sequencing platform.

    Directory of Open Access Journals (Sweden)

    Dale J Hedges

    Full Text Available Despite the ever-increasing throughput and steadily decreasing cost of next generation sequencing (NGS, whole genome sequencing of humans is still not a viable option for the majority of genetics laboratories. This is particularly true in the case of complex disease studies, where large sample sets are often required to achieve adequate statistical power. To fully leverage the potential of NGS technology on large sample sets, several methods have been developed to selectively enrich for regions of interest. Enrichment reduces both monetary and computational costs compared to whole genome sequencing, while allowing researchers to take advantage of NGS throughput. Several targeted enrichment approaches are currently available, including molecular inversion probe ligation sequencing (MIPS, oligonucleotide hybridization based approaches, and PCR-based strategies. To assess how these methods performed when used in conjunction with the ABI SOLID3+, we investigated three enrichment techniques: Nimblegen oligonucleotide hybridization array-based capture; Agilent SureSelect oligonucleotide hybridization solution-based capture; and Raindance Technologies' multiplexed PCR-based approach. Target regions were selected from exons and evolutionarily conserved areas throughout the human genome. Probe and primer pair design was carried out for all three methods using their respective informatics pipelines. In all, approximately 0.8 Mb of target space was identical for all 3 methods. SOLiD sequencing results were analyzed for several metrics, including consistency of coverage depth across samples, on-target versus off-target efficiency, allelic bias, and genotype concordance with array-based genotyping data. Agilent SureSelect exhibited superior on-target efficiency and correlation of read depths across samples. Nimblegen performance was similar at read depths at 20× and below. Both Raindance and Nimblegen SeqCap exhibited tighter distributions of read depth

  4. Doing more with less: fluorescence in situ hybridization and gene sequencing assays can be reliably performed on archival stained tumor tissue sections.

    Science.gov (United States)

    Pelosi, Giuseppe; Perrone, Federica; Tamborini, Elena; Fabbri, Alessandra; Testi, Maria Adele; Busico, Adele; Settanni, Giulio; Picciani, Benedetta; Bovio, Enrica; Sonzogni, Angelica; Valeri, Barbara; Garassino, Marina; De Braud, Filippo; Pastorino, Ugo

    2016-04-01

    Little is known about molecular testing on tumor tissue retrieved from stained sections, for which there may be a clinical need. We retrospectively analyzed 112 sections from 56 tumor patients using either fluorescence in situ hybridization (FISH) with different probes (19 sections from 17 patients) or Sanger or targeted next generation sequencing for detection of BRAF, EGFR, KRAS, C-KIT, and TP53 mutations (93 sections from 39 patients). Tumor tissue sections had been stained by hematoxylin and eosin (H&E) (42 sections) or by immunohistochemistry for cytoplasmic or nuclear/nuclear-cytoplasmic markers (70 sections) with a peroxidase (P-IHC, with 3,3'-diaminobenzidine as chromogen) or alkaline phosphatase label (AP-IHC, with Warp Red™ as chromogen). For FISH analysis, the concordance rate between the original diagnosis and that obtained on H&E- or P-IHC-stained tissue sections (AP-IHC was not on record for this set of patients) was 95% (18 out of 19 tumor sections). Only one tumor sample, diffusely positive for MLH1, did not yield any nuclear hybridization signal. For sequencing analysis, the concordance rate was 100% on negative P-IHC and positive AP-IHC-stained sections, regardless of the subcellular localization of the reaction product. Mutations were detected in only 52% of cases expressing nuclear/nuclear-cytoplasmic markers, regardless of the sequencing technology used (p = 0.0002). In conclusion, stained sections may be a valuable resource for FISH or sequencing analysis, but on cases expressing nuclear markers sequencing results need to be interpreted cautiously.

  5. Rover waste assay system

    Energy Technology Data Exchange (ETDEWEB)

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J. [Idaho National Engineering Lab., Idaho Falls, ID (United States)

    1997-11-01

    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched {sup 235}U contaminated piping, tank sections, and debris from the Rover nuclear rocket fuel processing facility at the Idaho Chemical Processing Plant. A scanning system translates a NaI(Tl) detector/collimator system over the structural components where both relative and calibrated measurements for {sup 137}Cs are made. Uranium-235 concentrations are in operation and is sufficiently automated that most functions are performed by the computer system. These functions include system calibration, problem identification, collimator control, data analysis, and reporting. Calibration of the system was done through a combination of measurements on calibration standards and benchmarked modeling. A description of the system is presented along with the methods and uncertainties associated with the calibration and analysis of the system for components from the Rover facility. 4 refs., 2 figs., 4 tabs.

  6. Comparison of multiplex PCR hybridization-based and singleplex real-time PCR-based assays for detection of low prevalence pathogens in spiked samples.

    Science.gov (United States)

    Hockman, Donna; Dong, Ming; Zheng, Hong; Kumar, Sanjai; Huff, Matthew D; Grigorenko, Elena; Beanan, Maureen; Duncan, Robert

    2017-01-01

    Molecular diagnostic devices are increasingly finding utility in clinical laboratories. Demonstration of the effectiveness of these devices is dependent upon comparing results from clinical samples tested with the new device to an alternative testing method. The preparation of mock clinical specimens will be necessary for the validation of molecular diagnostic devices when a sufficient number of clinical specimens is unobtainable. Examples include rare pathogens, some of which are pathogens posing a biological weapon threat. Here we describe standardized steps for developers to follow for the culture and quantification of three organisms used to spike human whole blood to create mock specimens. The three organisms chosen for this study were the Live Vaccine Strain (LVS) of Francisella tularensis, surrogate for a potential biothreat pathogen, Escherichia coli, a representative Gram-negative bacterium and Babesia microti (Franca) Reichenow Peabody strain, representing a protozoan parasite. Mock specimens were prepared with blood from both healthy donors and donors with nonspecific symptoms including fever, malaise, and flu-like symptoms. There was no significant difference in detection results between the two groups for any pathogen. Testing of the mock samples was compared on two platforms, Target Enriched Multiplex-PCR (TEM-PCR™) and singleplex real-time PCR (RT-PCR). Results were reproducible on both platforms. The reproducibility demonstrated by obtaining the same results between two testing methods and between healthy and symptomatic mock specimens, indicates the standardized methods described for creating the mock specimens are valid and effective for evaluating diagnostic devices.

  7. A new method for quantitative analysis of the T cell receptor V region repertoires in healthy common marmosets by microplate hybridization assay.

    Science.gov (United States)

    Kitaura, Kazutaka; Fujii, Yoshiki; Matsutani, Takaji; Shirai, Kenji; Suzuki, Satsuki; Takasaki, Tomohiko; Shimada, Shin; Kametani, Yoshie; Shiina, Takashi; Takabayashi, Shuji; Katoh, Hideki; Ogasawara, Kouetsu; Kurane, Ichiro; Suzuki, Ryuji

    2012-10-31

    The common marmoset, Callithrix jacchus, is one of the smallest primates and is increasingly used for an experimental nonhuman primate model in many research fields. Analysis of T cell receptor (TCR) repertoires is a powerful tool to investigate T cell immunity in terms of antigen specificity and variability of TCR expression. However, monoclonal antibodies specific for many TCR Vα or Vβ chains have not been created. We have recently identified a large number of TCRα chain variable (TRAV) and TCRβ chain variable (TRBV) sequences from a cDNA library of common marmosets. The purpose of this study is to develop a new method for analysis of TCR repertoires in the common marmoset using this sequence information. This method is based on a microplate hybridization technique using 32 TRAV-specific and 32 TRBV-specific oligoprobes following an adaptor-ligation PCR. This enables the easy quantitation of the respective TRAV and TRBV expression levels. No cross-hybridization among specific-oligoprobes and very low variances in repeated measures of the same samples was found, demonstrating high specificity and reproducibility. Furthermore, this method was validated by an antihuman Vβ23 antibody which specifically bound to marmoset Vβ23. Using this method, we analyzed TCR repertoires from various tissue samples (PBMCs, spleen, lymph node and thymus) and isolated T cell subpopulations (CD4+CD8+, CD4+CD8− and CD4−CD8+) from the thymus of 10 common marmosets. Neither tissue-specific nor T cell subpopulation-specific differences was found in TRAV and TRBV repertoires. These results suggest that, unlike mice, TCR repertoires in the common marmoset are not affected by endogenous superantigens and are conserved among individuals, among tissues, and among T cell subpopulations. Thus, TCR repertoire analysis with high specificity and reproducibility is a very useful technique, with the potential to replace flow cytometric analysis using a panel of TRV-specific antibodies, many

  8. A direct assay of carboxyl-containing small molecules by SALDI-MS on a AgNP/rGO-based nanoporous hybrid film.

    Science.gov (United States)

    Hong, Min; Xu, Lidan; Wang, Fangli; Geng, Zhirong; Li, Haibo; Wang, Huaisheng; Li, Chen-Zhong

    2016-04-25

    Silver nanoparticles (AgNPs) and reduced graphene oxide (rGO) hybrid nanoporous structures fabricated by the layer-by-layer (LBL) electrostatic self-assembly have been applied as a simple platform for the rapid analysis of carboxyl-containing small molecules by surface-assisted laser desorption/ionization (D/I) mass spectrometry (SALDI-MS). By the simple one-step deposition of analytes onto the (AgNP/rGO)9 multilayer film, the MS measurements of various carboxyl-containing small molecules (including amino acids, fatty acids and organic dicarboxylic acids) can be done. In contrast to other energy transfer materials relative to AgNPs, the signal interferences of a Ag cluster (Agn(+) or Agn(-)) and a C cluster (Cn(+) or Cn(-)) have been effectively reduced or eliminated. The effects of various factors, such as the pore structure and composition of the substrates, on the efficiency of D/I have been investigated by comparing with the (AgNP)9 LBL nanoporous structure, (AgNP/rGO)9/(SiO2NP)6 LBL multilayer film and AgNP/prGO nanocomposites.

  9. Prediction of human cell radiosensitivity: Comparison of clonogenic assay with chromosome aberrations scored using premature chromosome condensation with fluorescence in situ hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Sasai, K.; Evans, J.W.; Kovacs, M.S. [Stanford Univ. School of Medicine, CA (United States)] [and others

    1994-12-01

    The purpose of the present investigation was to determine whether chromosome aberrations scored by premature chromosome condensation (PCC) and fluorescence in situ hybridization (FISH) can predict the radiosensitivity of human cell lines, thereby providing a possible means of assessing the in situ radiosensitivity of normal tissues and the radiocurability of individual human cancers. We used four cells lines of different radiosensitivity: normal human fibroblasts (AG1522), ataxia-telangiectasia fibroblasts (AT2052), a human fibrosarcoma cell line (HT1080), and a human melanoma cell line (melanoma 903). These were irradiated in plateau phase with a range of doses and assessed both for clonogenic cell survival and for aberrations in a single chromosome (number 4) immediately after, and 24 h after irradiation. The initial number of breaks in chromosome 4 was proportional to irradiation dose and was identical for all the different human cell lines, irrespective of radiosensitivity. On the other hand, the number of chromosome 4 breaks remaining 24 h after irradiation reflected the radiosensitivity of the cells such that the relationship between residual chromosome aberrations and cell survival was the same for the different cell lines. These results suggest that the scoring of chromosome aberrations in interphase using FISH with PCC holds considerable promise for predicting the radiosensitivity of normal and tumor tissues in situ. 28 refs., 4 figs.

  10. The epidemiology of tick-borne haemoparasites as determined by the reverse line blot hybridization assay in an intensively studied cohort of calves in western Kenya.

    Science.gov (United States)

    Njiiri, Nyawira E; Bronsvoort, B Mark deC; Collins, Nicola E; Steyn, Helena C; Troskie, Milana; Vorster, Ilse; Thumbi, S M; Sibeko, Kgomotso P; Jennings, Amy; van Wyk, Ilana Conradie; Mbole-Kariuki, Mary; Kiara, Henry; Poole, E Jane; Hanotte, Olivier; Coetzer, Koos; Oosthuizen, Marinda C; Woolhouse, Mark; Toye, Philip

    2015-05-30

    The development of sensitive surveillance technologies using PCR-based detection of microbial DNA, such as the reverse line blot assay, can facilitate the gathering of epidemiological information on tick-borne diseases, which continue to hamper the productivity of livestock in many parts of Africa and elsewhere. We have employed a reverse line blot assay to detect the prevalence of tick-borne parasites in an intensively studied cohort of indigenous calves in western Kenya. The calves were recruited close to birth and monitored for the presence of infectious disease for up to 51 weeks. The final visit samples from 453 calves which survived for the study period were analyzed by RLB. The results indicated high prevalences of Theileria mutans (71.6%), T. velifera (62.8%), Anaplasma sp. Omatjenne (42.7%), A. bovis (39.9%), Theileria sp. (sable) (32.7%), T. parva (12.9%) and T. taurotragi (8.5%), with minor occurrences of eight other haemoparasites. The unexpectedly low prevalence of the pathogenic species Ehrlichia ruminantium was confirmed by a species-specific PCR targeting the pCS20 gene region. Coinfection analyses of the seven most prevalent haemoparasites indicated that they were present as coinfections in over 90% of the cases. The analyses revealed significant associations between several of the Theileria parasites, in particular T. velifera with Theileria sp. sable and T. mutans, and T. parva with T. taurotragi. There was very little coinfection of the two most common Anaplasma species, although they were commonly detected as coinfections with the Theileria parasites. The comparison of reverse line blot and serological results for four haemoparasites (T. parva, T. mutans, A. marginale and B. bigemina) indicated that, except for the mostly benign T. mutans, indigenous cattle seem capable of clearing infections of the three other, pathogenic parasites to below detectable levels. Although the study site was located across four agroecological zones, there was

  11. Unravelling the Bacterial Vaginosis-Associated Biofilm: A Multiplex Gardnerella vaginalis and Atopobium vaginae Fluorescence In Situ Hybridization Assay Using Peptide Nucleic Acid Probes.

    Directory of Open Access Journals (Sweden)

    Liselotte Hardy

    Full Text Available Bacterial vaginosis (BV, a condition defined by increased vaginal discharge without significant inflammation, is characterized by a change in the bacterial composition of the vagina. Lactobacillus spp., associated with a healthy vaginal microbiome, are outnumbered by BV-associated organisms. These bacteria could form a polymicrobial biofilm which allows them to persist in spite of antibiotic treatment. In this study, we examined the presence of Gardnerella vaginalis and Atopobium vaginae in vaginal biofilms using Peptide Nucleic Acid (PNA probes targeting these bacteria. For this purpose, we developed three new PNA probes for A. vaginae. The most specific A. vaginae probe, AtoITM1, was selected and then used in an assay with two existing probes, Gard162 and BacUni-1, to evaluate multiplex FISH on clinical samples. Using quantitative polymerase chain reaction (qPCR as the gold standard, we demonstrated a sensitivity of 66.7% (95% confidence interval: 54.5% - 77.1% and a specificity of 89.4% (95% confidence interval: 76.1% - 96% of the new AtoITM1 probe. FISH enabled us to show the presence of a polymicrobial biofilm in bacterial vaginosis, in which Atopobium vaginae is part of a Gardnerella vaginalis-dominated biofilm. We showed that the presence of this biofilm is associated with high bacterial loads of A. vaginae and G. vaginalis.

  12. Unravelling the Bacterial Vaginosis-Associated Biofilm: A Multiplex Gardnerella vaginalis and Atopobium vaginae Fluorescence In Situ Hybridization Assay Using Peptide Nucleic Acid Probes.

    Science.gov (United States)

    Hardy, Liselotte; Jespers, Vicky; Dahchour, Nassira; Mwambarangwe, Lambert; Musengamana, Viateur; Vaneechoutte, Mario; Crucitti, Tania

    2015-01-01

    Bacterial vaginosis (BV), a condition defined by increased vaginal discharge without significant inflammation, is characterized by a change in the bacterial composition of the vagina. Lactobacillus spp., associated with a healthy vaginal microbiome, are outnumbered by BV-associated organisms. These bacteria could form a polymicrobial biofilm which allows them to persist in spite of antibiotic treatment. In this study, we examined the presence of Gardnerella vaginalis and Atopobium vaginae in vaginal biofilms using Peptide Nucleic Acid (PNA) probes targeting these bacteria. For this purpose, we developed three new PNA probes for A. vaginae. The most specific A. vaginae probe, AtoITM1, was selected and then used in an assay with two existing probes, Gard162 and BacUni-1, to evaluate multiplex FISH on clinical samples. Using quantitative polymerase chain reaction (qPCR) as the gold standard, we demonstrated a sensitivity of 66.7% (95% confidence interval: 54.5% - 77.1%) and a specificity of 89.4% (95% confidence interval: 76.1% - 96%) of the new AtoITM1 probe. FISH enabled us to show the presence of a polymicrobial biofilm in bacterial vaginosis, in which Atopobium vaginae is part of a Gardnerella vaginalis-dominated biofilm. We showed that the presence of this biofilm is associated with high bacterial loads of A. vaginae and G. vaginalis.

  13. PD-L1 Expression by Two Complementary Diagnostic Assays and mRNA In Situ Hybridization in Small Cell Lung Cancer

    Science.gov (United States)

    Yu, Hui; Batenchuk, Cory; Badzio, Andrzej; Boyle, Theresa A.; Czapiewski, Piotr; Chan, Daniel C.; Lu, Xian; Gao, Dexiang; Ellison, Kim; Kowalewski, Ashley A.; Rivard, Christopher J.; Dziadziuszko, Rafal; Zhou, Caicun; Hussein, Maen; Richards, Donald; Wilks, Sharon; Monte, Marc; Edenfield, William; Goldschmidt, Jerome; Page, Ray; Ulrich, Brian; Waterhouse, David; Close, Sandra; Jassem, Jacek; Kulig, Kimary; Hirsch, Fred R.

    2017-01-01

    Introduction Therapeutic antibodies to immune checkpoints show promising results. Programmed death-ligand 1 (PD-L1), an immune checkpoint ligand, blocks the cancer immunity cycle by binding the PD-L1 receptor (programmed death 1). We investigated PD-L1 protein expression and messenger RNA (mRNA) levels in SCLC. Methods PD-L1 protein expression and mRNA levels were determined by immunohistochemistry (IHC) with SP142 and Dako 28-8 PD-L1 antibodies and in situ hybridization in primary tumor tissue microarrays in both tumor cells and tumor-infiltrating immune cells (TIICs) obtained from a limited-disease SCLC cohort of 98 patients. An additional cohort of 96 tumor specimens from patients with extensive-disease SCLC was assessed for PD-L1 protein expression in tumor cells with Dako 28-8 antibody only. Results The overall prevalence of PD-L1 protein expression in tumor cells was 16.5%. In the limited-disease cohort, the prevalences of PD-L1 protein expression in tumor cells with SP142 and Dako 28-8 were 14.7% and 19.4% (tumor proportion score cutoff ≥1%) and PD-L1 mRNA ISH expression was positive in 15.5% of tumor samples. Increased PD-L1 protein/mRNA expression was associated with the presence of more TIICs (p < 0.05). The extensive-disease cohort demonstrated a 14.9% positivity of PD-L1 protein expression in tumor cells with Dako 28-8 antibody. Conclusions A subset of SCLCs is characterized by positive PD-L1 and/or mRNA expression in tumor cells. Higher PD-L1 and mRNA expression correlate with more infiltration of TIICs. The prevalence of PD-L1 in SCLC is lower than that published for NSCLC. The predictive role of PD-L1 expression in SCLC treatment remains to be established. PMID:27639678

  14. Enriched MU-Calculi Module Checking

    CERN Document Server

    Ferrante, Alessandro; Parente, Mimmo

    2008-01-01

    The model checking problem for open systems has been intensively studied in the literature, for both finite-state (module checking) and infinite-state (pushdown module checking) systems, with respect to Ctl and Ctl*. In this paper, we further investigate this problem with respect to the Mu-calculus enriched with nominals and graded modalities (hybrid graded Mu-calculus), in both the finite-state and infinite-state settings. Using an automata-theoretic approach, we show that hybrid graded Mu-calculus module checking is solvable in exponential time, while hybrid graded Mu-calculus pushdown module checking is solvable in double-exponential time. These results are also tight since they match the known lower bounds for Ctl. We also investigate the module checking problem with respect to the hybrid graded Mu-calculus enriched with inverse programs (Fully enriched Mu-calculus): by showing a reduction from the domino problem, we show its undecidability. We conclude with a short overview of the model checking problem ...

  15. Disagreement between Human Papillomavirus Assays

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Preisler, Sarah; Ejegod, Ditte Møller

    2014-01-01

    assays. Positive agreement between the assays was measured as the conditional probability that the results of all compared assays were positive given that at least one assay returned a positive result. Of all 5,064 samples, 1,679 (33.2%) tested positive on at least one of the assays. Among these, 41......We aimed to determine the disagreement in primary cervical screening between four human papillomavirus assays: Hybrid Capture 2, cobas, CLART, and APTIMA. Material from 5,064 SurePath samples of women participating in routine cervical screening in Copenhagen, Denmark, was tested with the four......% tested positive on all four. Agreement was lower in women aged ≥ 30 years (30%, vs. 49% at samples (29%, vs. 38% in follow-up samples), and in women with concurrent normal cytology (22%, vs. 68% with abnormal cytology). Among primary screening samples from women aged 30...

  16. Enzyme assays.

    Science.gov (United States)

    Reymond, Jean-Louis; Fluxà, Viviana S; Maillard, Noélie

    2009-01-07

    Enzyme assays are analytical tools to visualize enzyme activities. In recent years a large variety of enzyme assays have been developed to assist the discovery and optimization of industrial enzymes, in particular for "white biotechnology" where selective enzymes are used with great success for economically viable, mild and environmentally benign production processes. The present article highlights the aspects of fluorogenic and chromogenic substrates, sensors, and enzyme fingerprinting, which are our particular areas of interest.

  17. Simultaneous identification of Aspergillus and Mucorales species by reverse line blot (RLB) hybridization assay%应用反向线点杂交技术鉴定临床常见曲霉属和毛霉目真菌

    Institute of Scientific and Technical Information of China (English)

    赵作涛; 李丽丽; 王晓阳; 万喆; 陈伟; 李若瑜

    2011-01-01

    Objective To Rapidly differentiate Aspergillus and Mucorales species by reverse line blot hybridization (RLB). Methods Totally 98 isolates including five Aspergillus strains ( Aspergillus fumigatus, Aspergillus flavus,Aspergillus niger, Aspergillus terreus and Aspergillus nidulans ) and seven Mucorales species ( Mucor heimalis ,Mucor racemosus ,Mucor cercinelloidea,Rhizopus arrhizus,Rhizopus microsporus,Rhizomucor pusillus and Absidia corymbifera ) were obtained from Research Center for Medical Mycology and Mycoses, Peking University. ITS1 and ITS4 fungal universal primers were chosen for PCR amplification, and the amplified products were used for reverse line blot hybridization with 12 fungal specie-specific probes. RLB data were compared with those by traditional fungal morphology and ITS sequencing methods. Results RLB showed high sensitivity and specificity , with 100% correct identification percentage of all the isolates and no cross hybridization between the species-specific probes. Eight negative control stains ( Candida albicans ,Fusarium solani ,Scedosporium apiospermum ,Penicillium marneffei ,Exophiala verrucosa,Aspergillus clavatus,Aspergillus japonicus and Cunninghamella elegans ) also showed negative by RLB. The analytical sensitivity of RLB was 1. 8 x 10-3 ng/μL by 10-fold serial dilution of Aspergillus fumigatus genomic DNA. Conclusions The RLB assay provides a rapid and reliable option for laboratory diagnosis and identification of Aspergillus and Mucorales species.%目的 应用反向线点杂交技术(reverse line blot hybridization,RLB)快速鉴定临床常见的曲霉属和毛霉目真菌.方法 收集我院真菌和真菌病研究中心保存的5种曲霉菌(烟曲霉、黄曲霉、黑曲霉、土曲霉、构巢曲霉)和7种毛霉目真菌(冻土毛霉菌、总状毛霉菌、卷枝毛霉菌、少根根霉、小孢根霉、微小根毛霉、伞状犁头霉),共计98株菌株.利用真菌通用引物ITS1和ITS4对菌株进行PCR扩增,用12个

  18. Profile of World Uranium Enrichment Programs-2009

    Energy Technology Data Exchange (ETDEWEB)

    Laughter, Mark D [ORNL

    2009-04-01

    demonstrated commercially. In the early 1980s, six countries developing gas centrifuge technology (United States, United Kingdom, Germany, the Netherlands, Japan, and Australia) along with the International Atomic Energy Agency and the European Atomic Energy Community began developing effective safeguards techniques for GCEPs. This effort was known as the Hexapartite Safeguards Project (HSP). The HSP had the goal of maximizing safeguards effectiveness while minimizing the cost to the operator and inspectorate, and adopted several recommendations, such as the acceptance of limited-frequency unannounced access inspections in cascade halls, and the use of nondestructive assay measurements and tamper-indicating seals. While only the HSP participants initially committed to implementing all the measures of the approach, it has been used as a model for the safeguards applied to GCEPs in additional states. Uranium enrichment capacity has continued to expand on all fronts in the last few years. GCEP capacity is expanding in anticipation of the eventual shutdown of the less-efficient GDPs, the termination of the U.S.-Russia HEU blend-down program slated for 2013, and the possible resurgence of nuclear reactor construction as part of an expected 'Nuclear Renaissance'. Overall, a clear trend in the world profile of uranium enrichment plant operation is the continued movement towards multinational projects driven by commercial and economic interests. Along this vein, the safeguards community is continuing to develop new safeguards techniques and technologies that are not overly burdensome to enrichment plant operators while delivering more effective and efficient results. This report provides a snapshot overview of world enrichment capacity in 2009, including profiles of the uranium enrichment programs of individual states. It is a revision of a 2007 report on the same topic; significant changes in world enrichment programs between the previous and current reports are emphasized

  19. Synthesis of core-shell hydrophilic polymer-silica hybrid material and its application in N-glycan enrichment%新型核壳型亲水聚合物-硅胶杂化填料的合成及其在蛋白质N-糖链富集中的应用

    Institute of Scientific and Technical Information of China (English)

    白海红; 范超; 沈丙权; 田芳; 邓玉林; 潘一廷; 秦伟捷; 钱小红

    2015-01-01

    蛋白质的 N-糖基化修饰与多种重要的生理、病理进程密切相关,是多种重大疾病诊断标志物研究的热点。由于糖蛋白本身多是低丰度表达的蛋白质,且糖链结构具有高度微不均一性,这使得蛋白质糖基化修饰的分析具有一定的挑战。本研究利用表面引发-原子转移自由基聚合( SI-ATRP )法,以带有双键的氨基葡萄糖为单体(GMAG),成功制备了新型核壳型亲水聚合物-硅胶杂化填料(pGMAG-SiO2)。由于在硅胶表面引入致密的亲水聚合物层,该填料不仅保持了硅胶良好的机械强度,而且显著提高了其亲水性,因此非常适合作为亲水填料用于蛋白质的N-糖链富集。以麦芽七糖和鸡卵清蛋白的N-糖链为研究对象,考察了该填料对N-糖链的富集效果,并将该杂化填料成功用于人血浆中糖蛋白 N-糖链的富集检测,共鉴定了47种糖型。以上结果表明,pGMAG-SiO2填料对N-糖链具有较高的亲和性,可以用于 N-糖链的高覆盖率鉴定。%Protein N-glycosylation is one of the most important post-translational modifications closely correlated with many important biological and pathological processes. The structural alterations of N-linked glycans in glycoproteins are always associated with many diseases,such as diabetes,heart failure and malignant tumors. Therefore,it is very important to establish sen-sitive methods for high-throughput N-glycan profiling. However,the low abundance of the N-glycoproteins and the heterogeneity of the N-glycans make it a challenge to analyse the pro-tein glycosylation sensitively. In this work,we had synthesized core-shell hydrophilic polymer-silica hybrid materials(pGMAG-SiO2)for the efficient enrichment of protein N-glycans. Firstly, pGMAG-SiO2 was prepared by in situ growth of glucose polymer on the surface of silica microp-articles using surface-initiated atom transfer radical polymerization( SI

  20. Dynamical investigation of macromolecular hybridization bioassays

    CERN Document Server

    Bittner, R; Wixforth, A

    2002-01-01

    A novel sensoric technique for the dynamical in situ investigation of a hybridization bio assay is presented, which utilizes a metal bead labeling method. Therein, hybridization results in an increased metal coverage on parts of a substrate, where it takes place. Our sensing principle relies on the measurement of the radio frequency impedance of the hybridization spots. We propose several examples for sensor devices.

  1. Profile of World Uranium Enrichment Programs - 2007

    Energy Technology Data Exchange (ETDEWEB)

    Laughter, Mark D [ORNL

    2007-11-01

    of the future, but has yet to be demonstrated commercially. In the early 1980s, six countries developing gas centrifuge technology (United States, United Kingdom, Germany, the Netherlands, Japan, and Australia) along with the International Atomic Energy Agency (IAEA) and the European Atomic Energy Community (EURATOM) began developing effective safeguards techniques for GCEPs. This effort was known as the Hexapartite Safeguards Project (HSP). The HSP had the goal of maximizing safeguards effectiveness while minimizing the cost to the operator and inspectorate, and adopted several recommendations, such as the acceptance of limited-frequency unannounced access (LFUA) inspections in cascade halls, and the use of nondestructive assay (NDA) measurements and tamper-indicating seals. While only the HSP participants initially committed to implementing all the measures of the approach, it has been used as a model for the safeguards applied to GCEPs in additional states. This report provides a snapshot overview of world enrichment capacity in 2007, including profiles of the uranium enrichment programs of individual states. It is based on open-source information, which is dependent on unclassified sources and may therefore not reflect the most recent developments. In addition, it briefly describes some of the safeguards techniques being used at various enrichment plants, including implementation of HSP recommendations.

  2. Profile of World Uranium Enrichment Programs - 2007

    Energy Technology Data Exchange (ETDEWEB)

    Laughter, Mark D [ORNL

    2007-11-01

    of the future, but has yet to be demonstrated commercially. In the early 1980s, six countries developing gas centrifuge technology (United States, United Kingdom, Germany, the Netherlands, Japan, and Australia) along with the International Atomic Energy Agency (IAEA) and the European Atomic Energy Community (EURATOM) began developing effective safeguards techniques for GCEPs. This effort was known as the Hexapartite Safeguards Project (HSP). The HSP had the goal of maximizing safeguards effectiveness while minimizing the cost to the operator and inspectorate, and adopted several recommendations, such as the acceptance of limited-frequency unannounced access (LFUA) inspections in cascade halls, and the use of nondestructive assay (NDA) measurements and tamper-indicating seals. While only the HSP participants initially committed to implementing all the measures of the approach, it has been used as a model for the safeguards applied to GCEPs in additional states. This report provides a snapshot overview of world enrichment capacity in 2007, including profiles of the uranium enrichment programs of individual states. It is based on open-source information, which is dependent on unclassified sources and may therefore not reflect the most recent developments. In addition, it briefly describes some of the safeguards techniques being used at various enrichment plants, including implementation of HSP recommendations.

  3. Comparing the Cervista HPV HR test and Hybrid Capture 2 assay in a Dutch screening population: improved specificity of the Cervista HPV HR test by changing the cut-off.

    Directory of Open Access Journals (Sweden)

    Aniek Boers

    Full Text Available The diagnostic performance of the widely-used Cervista HPV HR test was compared to the Hybrid Capture 2 (HC2 test in a Dutch population-based cervical cancer screening program. In 900 scrapings of women with normal cytomorphology, specificity was 90% (95%CI: 87.84-91.87 for the Cervista HPV HR test and 96% (95%CI: 94.76-97.37 for the HC2 test with 93% agreement between both tests (κ = 0.5, p<0.001. The sensitivity for CIN2+ using 65 scrapings of women with histological-confirmed CIN2+ was 91% (95%CI: 80.97-96.51 for the Cervista HPV HR test and 92% (95%CI: 82.94-97.43 for the HC2 test with 95% agreement between both tests (κ = 0.7, p<0.001. Fifty-seven of 60 HC2 negative/Cervista positive cases tested HPV-negative with PCR-based HPV assays; of these cases 56% were defined as Cervista triple-positive with FOZ values in all 3 mixes higher than the second cut-off of 1.93 (as set by manufacturer. By setting this cut-off at 5.0, specificity improved significantly without affecting sensitivity. External validation of this new cut-off at 5.0 in triple-positive scrapings of women selected from the SHENCCASTII database revealed that 22/24 histological normal cases now tested HPV-negative in the Cervista HPV HR test, while CIN2+ lesions remained HPV-positive. The intra-laboratory reproducibility of the Cervista HPV HR test (n = 510 showed a concordance of 92% and 93% for cut-off 1.93 and 5.0 (κ = 0.83 and κ = 0.84, p<0.001 and inter-laboratory agreement of the Cervista HPV HR test was 90% and 93% for cut-off 1.93 and 5.0 (κ = 0.80 and κ = 0.85, p<0.001. In conclusion, the specificity of the Cervista HPV HR test could be improved significantly by increasing the second cut-off from 1.93 to 5.0, without affecting the sensitivity of the test in a population-based screening setting.

  4. Angiogenesis Assays.

    Science.gov (United States)

    Nambiar, Dhanya K; Kujur, Praveen K; Singh, Rana P

    2016-01-01

    Neoangiogenesis constitutes one of the first steps of tumor progression beyond a critical size of tumor growth, which supplies a dormant mass of cancerous cells with the required nutrient supply and gaseous exchange through blood vessels essentially needed for their sustained and aggressive growth. In order to understand any biological process, it becomes imperative that we use models, which could mimic the actual biological system as closely as possible. Hence, finding the most appropriate model is always a vital part of any experimental design. Angiogenesis research has also been much affected due to lack of simple, reliable, and relevant models which could be easily quantitated. The angiogenesis models have been used extensively for studying the action of various molecules for agonist or antagonistic behaviour and associated mechanisms. Here, we have described two protocols or models which have been popularly utilized for studying angiogenic parameters. Rat aortic ring assay tends to bridge the gap between in vitro and in vivo models. The chorioallantoic membrane (CAM) assay is one of the most utilized in vivo model system for angiogenesis-related studies. The CAM is highly vascularized tissue of the avian embryo and serves as a good model to study the effects of various test compounds on neoangiogenesis.

  5. Experimental conditions improving in-solution target enrichment for ancient DNA.

    Science.gov (United States)

    Cruz-Dávalos, Diana I; Llamas, Bastien; Gaunitz, Charleen; Fages, Antoine; Gamba, Cristina; Soubrier, Julien; Librado, Pablo; Seguin-Orlando, Andaine; Pruvost, Mélanie; Alfarhan, Ahmed H; Alquraishi, Saleh A; Al-Rasheid, Khaled A S; Scheu, Amelie; Beneke, Norbert; Ludwig, Arne; Cooper, Alan; Willerslev, Eske; Orlando, Ludovic

    2016-08-27

    High-throughput sequencing has dramatically fostered ancient DNA research in recent years. Shotgun sequencing, however, does not necessarily appear as the best-suited approach due to the extensive contamination of samples with exogenous environmental microbial DNA. DNA capture-enrichment methods represent cost-effective alternatives that increase the sequencing focus on the endogenous fraction, whether it is from mitochondrial or nuclear genomes, or parts thereof. Here, we explored experimental parameters that could impact the efficacy of MYbaits in-solution capture assays of ~5000 nuclear loci or the whole genome. We found that varying quantities of the starting probes had only moderate effect on capture outcomes. Starting DNA, probe tiling, the hybridization temperature and the proportion of endogenous DNA all affected the assay, however. Additionally, probe features such as their GC content, number of CpG dinucleotides, sequence complexity and entropy and self-annealing properties need to be carefully addressed during the design stage of the capture assay. The experimental conditions and probe molecular features identified in this study will improve the recovery of genetic information extracted from degraded and ancient remains.

  6. Uranium Conversion & Enrichment

    Energy Technology Data Exchange (ETDEWEB)

    Karpius, Peter Joseph [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2017-02-06

    The isotopes of uranium that are found in nature, and hence in ‘fresh’ Yellowcake’, are not in relative proportions that are suitable for power or weapons applications. The goal of conversion then is to transform the U3O8 yellowcake into UF6. Conversion and enrichment of uranium is usually required to obtain material with enough 235U to be usable as fuel in a reactor or weapon. The cost, size, and complexity of practical conversion and enrichment facilities aid in nonproliferation by design.

  7. Enrichment of high ammonia tolerant methanogenic culture

    DEFF Research Database (Denmark)

    Fotidis, Ioannis; Karakashev, Dimitar Borisov; Proietti, Nicolas

    plants. The methods used today to counteract ammonia inhibition are slow and costexpensive. A new biological approach to avoid or counteract ammonia inhibition by using ammonia tolerant methanogens, could provide a sustainable solution for cost-effective digestion of abundant ammonia-rich wastes. The aim...... of the current study was to isolate and identify methanogenic cultures tolerant to high ammonia concentrations. A mixed methanogenic population was stepwise exposed to ammonia concentrations (1 to 9.26 g NH4+-N L-1) during an enrichment process with successive batch cultivations. The methanogenic population...... was derived from a full scale biogas reactor (Hashøj, Denmark), fed with 75% animal manure and 25% food industries organic waste. Basal anaerobic medium was used for the enrichment along with sodium acetate (1 g HAc L-1) as a carbon source. Fluorescence insitu hybridization (FISH) was used to determine...

  8. Enriching the Catalog

    Science.gov (United States)

    Tennant, Roy

    2004-01-01

    After decades of costly and time-consuming effort, nearly all libraries have completed the retrospective conversion of their card catalogs to electronic form. However, bibliographic systems still are really not much more than card catalogs on wheels. Enriched content that Amazon.com takes for granted--such as digitized tables of contents, cover…

  9. Hybrid Baryons

    CERN Document Server

    Page, P R

    2003-01-01

    We review the status of hybrid baryons. The only known way to study hybrids rigorously is via excited adiabatic potentials. Hybrids can be modelled by both the bag and flux-tube models. The low-lying hybrid baryon is N 1/2^+ with a mass of 1.5-1.8 GeV. Hybrid baryons can be produced in the glue-rich processes of diffractive gamma N and pi N production, Psi decays and p pbar annihilation.

  10. ICP-MS Determination of Platinum Metals in Black Shale Enriched by Tin Fire Assay%锡试金富集-电感耦合等离子体质谱法测定黑色页岩中的铂族元素

    Institute of Scientific and Technical Information of China (English)

    王君玉; 孙自军; 袁润蕾; 王盈; 李杰; 王敏捷

    2013-01-01

    黑色页岩中的铂族元素经锡试金富集后,所得贵金属沉淀物用盐酸-硝酸(3+1)混合酸溶解,用电感耦合等离子体质谱法测定样品中的铂族元素.以镥为内标元素,用轻质氧化镁-高纯碳化硅(2+1)作覆盖剂,以防止锇和钌的挥发损失.铂族元素在一定的质量浓度范围内与其光谱强度呈线性关系,检出限(3σ)在0.018~0.20 ng· g-1之间.方法的加标回收率在80.0%~110%之间,相对标准偏差(n=11)在5.9%~14%之间.%Platinum metals in black shale were enriched by the method of tin fire assay.The precious metal precipitate obtained was dissolved in mixed acid of HCl-HNC3 (3 + 1),and contents of platinum metals in the solution were determined by ICP-MS.Lutetium was used as internal standard; and a mixture of light MgO and high purity SiC (2 + 1) was used as covering reagent to avoid the volatilization of Os and Ru.Linear relationships between values of spectral intensity and mass concentration of the platinum metals were kept in definite ranges.Detection limits (3σ) found were in the range of 0.018-0.20 ng · g-1.Tests for recovery were made by standard addition method,giving values of recovery in the range of 80.0%-110% with RSD's (n=11) in the range of 5.9%-14%.

  11. A comparison of two real-time polymerase chain reaction assays using hybridization probes targeting either 16S ribosomal RNA or a subsurface lipoprotein gene for detecting leptospires in canine urine.

    Science.gov (United States)

    Gentilini, Fabio; Zanoni, Renato Giulio; Zambon, Elisa; Turba, Maria Elena

    2015-11-01

    Leptospires are excreted in the urine of infected animals, and the prompt detection of leptospiral DNA using polymerase chain reaction (PCR) is increasingly being used. However, contradictory data has emerged concerning the diagnostic accuracy of the most popular PCR assays that target either the 16S ribosomal RNA (rrs) or the subsurface lipoprotein (LipL32) genes. In order to clarify the effect of the gene target, a novel hydrolysis probe-based, quantitative real-time PCR (qPCR) assay targeting the LipL32 gene was developed, validated, and then compared directly to the previously described rrs hydrolysis probe-based qPCR using a convenience collection of canine urine samples. The novel LipL32 qPCR assay was linear from 5.9 × 10(6) to 59 genome equivalents per reaction. Both the LipL32 and the rrs qPCR assays showed a limit of detection of 10 target copies per reaction indicating an approximately equivalent analytical sensitivity. Both assays amplified all 20 pathogenic leptospiral strains tested but did not amplify a representative collection of bacteria commonly found in voided canine urine. When the field samples were assayed, 1 and 5 out of 184 samples yielded an amplification signal in the LipL32 and rrs assays, respectively. Nevertheless, when the limit of detection was considered as the cutoff for interpreting findings, the 4 discordant cases were judged as negative. In conclusion, our study confirmed that both LipL32 and rrs are suitable targets for qPCR for the detection of leptospiral DNA in canine urine. However, the rrs target requires the mandatory use of a cutoff value in order to correctly interpret spurious amplifications.

  12. Profile of World Uranium Enrichment Programs-2009

    Energy Technology Data Exchange (ETDEWEB)

    Laughter, Mark D [ORNL

    2009-04-01

    demonstrated commercially. In the early 1980s, six countries developing gas centrifuge technology (United States, United Kingdom, Germany, the Netherlands, Japan, and Australia) along with the International Atomic Energy Agency and the European Atomic Energy Community began developing effective safeguards techniques for GCEPs. This effort was known as the Hexapartite Safeguards Project (HSP). The HSP had the goal of maximizing safeguards effectiveness while minimizing the cost to the operator and inspectorate, and adopted several recommendations, such as the acceptance of limited-frequency unannounced access inspections in cascade halls, and the use of nondestructive assay measurements and tamper-indicating seals. While only the HSP participants initially committed to implementing all the measures of the approach, it has been used as a model for the safeguards applied to GCEPs in additional states. Uranium enrichment capacity has continued to expand on all fronts in the last few years. GCEP capacity is expanding in anticipation of the eventual shutdown of the less-efficient GDPs, the termination of the U.S.-Russia HEU blend-down program slated for 2013, and the possible resurgence of nuclear reactor construction as part of an expected 'Nuclear Renaissance'. Overall, a clear trend in the world profile of uranium enrichment plant operation is the continued movement towards multinational projects driven by commercial and economic interests. Along this vein, the safeguards community is continuing to develop new safeguards techniques and technologies that are not overly burdensome to enrichment plant operators while delivering more effective and efficient results. This report provides a snapshot overview of world enrichment capacity in 2009, including profiles of the uranium enrichment programs of individual states. It is a revision of a 2007 report on the same topic; significant changes in world enrichment programs between the previous and current reports are emphasized

  13. Hybrid vehicles

    Energy Technology Data Exchange (ETDEWEB)

    West, J.G.W. [Electrical Machines (United Kingdom)

    1997-07-01

    The reasons for adopting hybrid vehicles result mainly from the lack of adequate range from electric vehicles at an acceptable cost. Hybrids can offer significant improvements in emissions and fuel economy. Series and parallel hybrids are compared. A combination of series and parallel operation would be the ideal. This can be obtained using a planetary gearbox as a power split device allowing a small generator to transfer power to the propulsion motor giving the effect of a CVT. It allows the engine to run at semi-constant speed giving better fuel economy and reduced emissions. Hybrid car developments are described that show the wide range of possible hybrid systems. (author)

  14. Synchronizability Analysis of Harmonious Unification Hybrid Preferential Model

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    The harmonious unification hybrid preferential model uses the dr ratio to adjust the proportion of deterministic preferential attachment and random preferential attachment, enriched the only deterministic preferential network model,

  15. Methods for Determining the Statistical Significance of Enrichment or Depletion of Gene Ontology Classifications under Weighted Membership

    Directory of Open Access Journals (Sweden)

    Ernesto eIacucci

    2012-02-01

    Full Text Available High-throughput molecular biology studies, such as microarray assays of gene expression, two-hybrid experiments for detecting protein interactions, or ChIP-Seq experiments for transcription factor binding, often result in an interesting set of genes—say, genes that are co-expressed or bound by the same factor. One way of understanding the biological meaning of such a set is to consider what processes or functions, as defined in an ontology, are over-represented (enriched or under-represented (depleted among genes in the set. Usually, the significance of enrichment or depletion scores is based on simple statistical models and on the membership of genes in different classifications. We consider the more general problem of computing p-values for arbitrary integer additive statistics, or weighted membership functions. Such membership functions can be used to represent, for example, prior knowledge on the role of certain genes or classifications, differential importance of different classifications or genes to the experimenter, hierarchical relationships between classifications, or different degrees of interestingness or evidence for specific genes. We describe a generic dynamic programming algorithm that can compute exact p-values for arbitrary integer additive statistics. We also describe several optimizations for important special cases, which can provide orders-of-magnitude speed up in the computations. We apply our methods to datasets describing oxidative phosphorylation and parturition and compare p-values based on computations of several different statistics for measuring enrichment. We find major differences between p-values resulting from these statistics, and that some statistics recover gold standard annotations of the data better than others. Our work establishes a theoretical and algorithmic basis for far richer notions of enrichment or depletion of gene sets with respect to gene ontologies than has previously been available.

  16. Exosome enrichment of human serum using multiple cycles of centrifugation.

    Science.gov (United States)

    Kim, Jeongkwon; Tan, Zhijing; Lubman, David M

    2015-09-01

    In this work, we compared the use of repeated cycles of centrifugation at conventional speeds for enrichment of exosomes from human serum compared to the use of ultracentrifugation (UC). After removal of cells and cell debris, a speed of 110 000 × g or 40 000 × g was used for the UC or centrifugation enrichment process, respectively. The enriched exosomes were analyzed using the bicinchoninic acid assay, 1D gel separation, transmission electron microscopy, Western blotting, and high-resolution LC-MS/MS analysis. It was found that a five-cycle repetition of UC or centrifugation is necessary for successful removal of nonexosomal proteins in the enrichment of exosomes from human serum. More significantly, 5× centrifugation enrichment was found to provide similar or better performance than 5× UC enrichment in terms of enriched exosome protein amount, Western blot band intensity for detection of CD-63, and numbers of identified exosome-related proteins and cluster of differentiation (CD) proteins. A total of 478 proteins were identified in the LC-MS/MS analyses of exosome proteins obtained from 5× UCs and 5× centrifugations including many important CD membrane proteins. The presence of previously reported exosome-related proteins including key exosome protein markers demonstrates the utility of this method for analysis of proteins in human serum.

  17. Thermal breeder fuel enrichment zoning

    Science.gov (United States)

    Capossela, Harry J.; Dwyer, Joseph R.; Luce, Robert G.; McCoy, Daniel F.; Merriman, Floyd C.

    1992-01-01

    A method and apparatus for improving the performance of a thermal breeder reactor having regions of higher than average moderator concentration are disclosed. The fuel modules of the reactor core contain at least two different types of fuel elements, a high enrichment fuel element and a low enrichment fuel element. The two types of fuel elements are arranged in the fuel module with the low enrichment fuel elements located between the high moderator regions and the high enrichment fuel elements. Preferably, shim rods made of a fertile material are provided in selective regions for controlling the reactivity of the reactor by movement of the shim rods into and out of the reactor core. The moderation of neutrons adjacent the high enrichment fuel elements is preferably minimized as by reducing the spacing of the high enrichment fuel elements and/or using a moderator having a reduced moderating effect.

  18. ENRICHMENT OF OBO ONTOLOGIES

    Science.gov (United States)

    Bada, Michael; Hunter, Lawrence

    2006-01-01

    This paper describes a frame-based integration of the three GO subontologies, the Chemicals of Biological Interest ontology (ChEBI), and the Cell Type Ontology (CTO) in which relationships between elements of the ontologies are modeled in a way that better captures the relational semantics between biological concepts represented by the terms, rather than between the terms themselves, than previous frame-based efforts. We also describe a methodology for creating suggested enriching assertions of the form (subject, relationship, object) by identifying patterns in GO terms, mapping these patterns and subpatterns to relationships, matching concepts to these patterns and subpatterns, and integrating these assertions into the ontologies. Using this methodology, a large number of reliable assertions linking previously unlinked OBO terms using a wide variety of specific, hierarchically arranged relationships were created: A predicted assertion was made for 62% of GO terms that matched one of 31 patterns, and 97% of these predicted assertions were assessed to be valid; a further 429 assertions (corresponding to 6% of the matching terms) were manually created, resulting in an initial set of 4,497 assertions. Furthermore, this methodology programmatically integrates assertions into a base ontology such that each assertion is fully consistent with respect to higher (i.e., more general) relevant class and slot levels. Such an integration is absent from previous compositional efforts, and we argue its necessity for the creation of coherent biological ontologies when linking previously unlinked terms. PMID:17011833

  19. Earthquake forecast enrichment scores

    Directory of Open Access Journals (Sweden)

    Christine Smyth

    2012-03-01

    Full Text Available The Collaboratory for the Study of Earthquake Predictability (CSEP is a global project aimed at testing earthquake forecast models in a fair environment. Various metrics are currently used to evaluate the submitted forecasts. However, the CSEP still lacks easily understandable metrics with which to rank the universal performance of the forecast models. In this research, we modify a well-known and respected metric from another statistical field, bioinformatics, to make it suitable for evaluating earthquake forecasts, such as those submitted to the CSEP initiative. The metric, originally called a gene-set enrichment score, is based on a Kolmogorov-Smirnov statistic. Our modified metric assesses if, over a certain time period, the forecast values at locations where earthquakes have occurred are significantly increased compared to the values for all locations where earthquakes did not occur. Permutation testing allows for a significance value to be placed upon the score. Unlike the metrics currently employed by the CSEP, the score places no assumption on the distribution of earthquake occurrence nor requires an arbitrary reference forecast. In this research, we apply the modified metric to simulated data and real forecast data to show it is a powerful and robust technique, capable of ranking competing earthquake forecasts.

  20. City model enrichment

    Science.gov (United States)

    Smart, Philip D.; Quinn, Jonathan A.; Jones, Christopher B.

    The combination of mobile communication technology with location and orientation aware digital cameras has introduced increasing interest in the exploitation of 3D city models for applications such as augmented reality and automated image captioning. The effectiveness of such applications is, at present, severely limited by the often poor quality of semantic annotation of the 3D models. In this paper, we show how freely available sources of georeferenced Web 2.0 information can be used for automated enrichment of 3D city models. Point referenced names of prominent buildings and landmarks mined from Wikipedia articles and from the OpenStreetMaps digital map and Geonames gazetteer have been matched to the 2D ground plan geometry of a 3D city model. In order to address the ambiguities that arise in the associations between these sources and the city model, we present procedures to merge potentially related buildings and implement fuzzy matching between reference points and building polygons. An experimental evaluation demonstrates the effectiveness of the presented methods.

  1. Conceptual Ideas for New Nondestructive UF6 Cylinder Assay Techniques

    Energy Technology Data Exchange (ETDEWEB)

    Miller, Karen A. [Los Alamos National Laboratory

    2012-05-02

    Nondestructive assay (NDA) measurements of uranium cylinders play an important role in helping the International Atomic Energy Agency (IAEA) safeguard uranium enrichment plants. Traditionally, these measurements have consisted of a scale or load cell to determine the mass of UF{sub 6} in the cylinder combined with a gamma-ray measurement of the 186 keV peak from {sup 235}U to determine enrichment. More recently, Los Alamos National Laboratory (LANL) and Pacific Northwest National Laboratory (PNNL) have developed systems that exploit the passive neutron signal from UF{sub 6} to determine uranium mass and/or enrichment. These include the Uranium Cylinder Assay System (UCAS), the Passive Neutron Enrichment Meter (PNEM), and the Hybrid Enrichment Verification Array (HEVA). The purpose of this report is to provide the IAEA with new ideas on technologies that may or may not be under active development but could be useful for UF{sub 6} cylinder assay. To begin, we have included two feasibility studies of active interrogation techniques. There is a long history of active interrogation in the field of nuclear safeguards, especially for uranium assay. Both of the active techniques provide a direct measure of {sup 235}U content. The first is an active neutron method based on the existing PNEM design that uses a correlated {sup 252}Cf interrogation source. This technique shows great promise for UF{sub 6} cylinder assay and is based on advanced technology that could be implemented in the field in the near term. The second active technique is nuclear resonance fluorescence (NRF). In the NRF technique, a bremsstrahlung photon beam could be used to illuminate the cylinder, and high-resolution gamma-ray detectors would detect the characteristic de-excitation photons. The results of the feasibility study show that under certain measurement geometries, NRF is impractical for UF6 cylinder assay, but the 'grazing transmission' and 'secant transmission' geometries

  2. Functionalized periodic mesoporous organosilicas for enhanced and selective peptide enrichment.

    Science.gov (United States)

    Wan, Jingjing; Qian, Kun; Zhang, Jun; Liu, Fang; Wang, Yunhua; Yang, Pengyuan; Liu, Baohong; Yu, Chengzhong

    2010-05-18

    The analysis of peptides by the mass spectrometry (MS) technique is important in modern life science. The enrichment of peptides can increase the detection efficiency and is sometimes indispensable for collecting the information on proteins with low-abundance. Herein, we first report that functionalized periodic mesoporous organosilica (PMO) materials have a superior peptide enrichment property. It is demonstrated that the PMO materials with an organo-bridged (-CH(2)-) hybrid wall composition display a highly enhanced peptide enrichment ability compared to the pure silica material (SBA-15) with similar mesostructured parameters and morphology. More importantly, by surface modification of PMO with amino groups (denoted NH(2)-PMO), PMO and NH(2)-PMO with opposite charged surfaces (-25.2 and +39.0 mV, respectively) show selective affinities for positively and negatively charged peptides, respectively. By directly adding PMO, NH(2)-PMO as well as pure silica materials to the peptides solution with a low concentration (1-2 fmol/microL), 36 and 28 peptides can be detected from the BSA digestion in the presence of PMO and NH(2)-PMO, respectively, while only 6 and 4 are monitored in the case of SBA-15 enrichment and from solution without enrichment, respectively. Moreover, 69.4% (25 of 36) of enriched peptides by PMO have pI > or = 6 and 80% (21 of 28) of enriched peptides by NH(2)-PMO possess pI PMO and PMO enrichment together, 51 peptides can be identified with a MOWSE score of 333. It is also noted that similar conclusions can also be obtained from the peptides solution originated from other proteins. This might be an important contribution to the understanding of the interaction between peptides and porous hosts, and the proposed method is promising for the development of both material science and biotechnology.

  3. Ensaio de potência da alfaepoetina: Comparação de camundongos Swiss Webster, NIH, C57BL/6, BALB/c com o híbrido B6D2F1 /Potency assay of epoetin alpha: Comparison of Swiss Webster, NIH, C57BL/6, BALB/c mice with the hybrid B6D2F1

    Directory of Open Access Journals (Sweden)

    Igor Barbosa da Silva

    2013-08-01

    Full Text Available Neste estudo comparamos os resultados de ensaios de potência da alfaepoetina (EPOhr realizados com camundongos de diferentes colônias e linhagens (Swiss Webster, NIH, C57BL/6 e BALB/c com aqueles de ensaios conduzidos com o híbrido B6D2F1, o úni-co camundongo recomendado pela Farmacopeia Europeia (FE. Fêmeas de diferentes colônias e linhagens, pesando 16-18 gramas, receberam uma única dose de EPOhr por via subcutânea (30, 90 ou 270 UI/animal, 0,2 mL/camundongo. As potências biológi-cas de apresentações de 4.000 UI/mL da EPOhr foram avaliadas utilizando um material de referência de trabalho de alfaepoetina (3.773 UI/mL anteriormente testado junto ao padrão de referência internacional BRP (European Pharmacopeia Biological Refe-rence Preparation. Os resultados indicaram que camundongos das colônias e linhagens examinadas atingiram critérios estatísticos (FE para um ensaio válido de potência da eritropoietina e, portanto, podem ser considerados como alternativas ao uso do híbrido B6D2F1. Os ensaios com camundongos BALB/c, entretanto, foram os que produziram re-sultados mais semelhantes aos obtidos com os híbridos B6D2F1, em relação à contagem média de reticulócitos em resposta a 30, 90 e 270 UI/camundongo, e aos coefi cientes angulares (inclinação e lineares (intersecção da curva dose-resposta (curvas paralelas praticamente superpostas. --------------------------------------------------------------------------- In this study we compared the outcomes of epoetin alpha (rhEPO potency assays per-formed with Swiss Webster, NIH, C57BL/6 and BALB/c mice with those of the assay conducted with the B6D2F1 hybrid, the only mice recommended by the European Phar-macopoeia (EP. Female mice from different breeding stocks and strains, weighing 16-18 g, received a single subcutaneous injection of (30, 90 or 270 IU per mouse, 0.2 mL per mouse of rhEPO. Biological potencies 4000 IU/mL rhEPO pharmaceutical forms from di-fferent batches

  4. Microsatellite DNA capture from enriched libraries.

    Science.gov (United States)

    Gonzalez, Elena G; Zardoya, Rafael

    2013-01-01

    Microsatellites are DNA sequences of tandem repeats of one to six nucleotides, which are highly polymorphic, and thus the molecular markers of choice in many kinship, population genetic, and conservation studies. There have been significant technical improvements since the early methods for microsatellite isolation were developed, and today the most common procedures take advantage of the hybrid capture methods of enriched-targeted microsatellite DNA. Furthermore, recent advents in sequencing technologies (i.e., next-generation sequencing, NGS) have fostered the mining of microsatellite markers in non-model organisms, affording a cost-effective way of obtaining a large amount of sequence data potentially useful for loci characterization. The rapid improvements of NGS platforms together with the increase in available microsatellite information open new avenues to the understanding of the evolutionary forces that shape genetic structuring in wild populations. Here, we provide detailed methodological procedures for microsatellite isolation based on the screening of GT microsatellite-enriched libraries, either by cloning and Sanger sequencing of positive clones or by direct NGS. Guides for designing new species-specific primers and basic genotyping are also given.

  5. Hydrogen-enriched fuels

    Energy Technology Data Exchange (ETDEWEB)

    Roser, R. [NRG Technologies, Inc., Reno, NV (United States)

    1998-08-01

    NRG Technologies, Inc. is attempting to develop hardware and infrastructure that will allow mixtures of hydrogen and conventional fuels to become viable alternatives to conventional fuels alone. This commercialization can be successful if the authors are able to achieve exhaust emission levels of less than 0.03 g/kw-hr NOx and CO; and 0.15 g/kw-hr NMHC at full engine power without the use of exhaust catalysts. The major barriers to achieving these goals are that the lean burn regimes required to meet exhaust emissions goals reduce engine output substantially and tend to exhibit higher-than-normal total hydrocarbon emissions. Also, hydrogen addition to conventional fuels increases fuel cost, and reduces both vehicle range and engine output power. Maintaining low emissions during transient driving cycles has not been demonstrated. A three year test plan has been developed to perform the investigations into the issues described above. During this initial year of funding research has progressed in the following areas: (a) a cost effective single-cylinder research platform was constructed; (b) exhaust gas speciation was performed to characterize the nature of hydrocarbon emissions from hydrogen-enriched natural gas fuels; (c) three H{sub 2}/CH{sub 4} fuel compositions were analyzed using spark timing and equivalence ratio sweeping procedures and finally; (d) a full size pick-up truck platform was converted to run on HCNG fuels. The testing performed in year one of the three year plan represents a baseline from which to assess options for overcoming the stated barriers to success.

  6. CLEAN: CLustering Enrichment ANalysis

    Directory of Open Access Journals (Sweden)

    Medvedovic Mario

    2009-07-01

    Full Text Available Abstract Background Integration of biological knowledge encoded in various lists of functionally related genes has become one of the most important aspects of analyzing genome-wide functional genomics data. In the context of cluster analysis, functional coherence of clusters established through such analyses have been used to identify biologically meaningful clusters, compare clustering algorithms and identify biological pathways associated with the biological process under investigation. Results We developed a computational framework for analytically and visually integrating knowledge-based functional categories with the cluster analysis of genomics data. The framework is based on the simple, conceptually appealing, and biologically interpretable gene-specific functional coherence score (CLEAN score. The score is derived by correlating the clustering structure as a whole with functional categories of interest. We directly demonstrate that integrating biological knowledge in this way improves the reproducibility of conclusions derived from cluster analysis. The CLEAN score differentiates between the levels of functional coherence for genes within the same cluster based on their membership in enriched functional categories. We show that this aspect results in higher reproducibility across independent datasets and produces more informative genes for distinguishing different sample types than the scores based on the traditional cluster-wide analysis. We also demonstrate the utility of the CLEAN framework in comparing clusterings produced by different algorithms. CLEAN was implemented as an add-on R package and can be downloaded at http://Clusteranalysis.org. The package integrates routines for calculating gene specific functional coherence scores and the open source interactive Java-based viewer Functional TreeView (FTreeView. Conclusion Our results indicate that using the gene-specific functional coherence score improves the reproducibility of the

  7. Hybrid Metaheuristics

    CERN Document Server

    2013-01-01

    The main goal of this book is to provide a state of the art of hybrid metaheuristics. The book provides a complete background that enables readers to design and implement hybrid metaheuristics to solve complex optimization problems (continuous/discrete, mono-objective/multi-objective, optimization under uncertainty) in a diverse range of application domains. Readers learn to solve large scale problems quickly and efficiently combining metaheuristics with complementary metaheuristics, mathematical programming, constraint programming and machine learning. Numerous real-world examples of problems and solutions demonstrate how hybrid metaheuristics are applied in such fields as networks, logistics and transportation, bio-medical, engineering design, scheduling.

  8. Hybrid intermediaries

    OpenAIRE

    Cetorelli, Nicola

    2014-01-01

    I introduce the concept of hybrid intermediaries: financial conglomerates that control a multiplicity of entity types active in the "assembly line" process of modern financial intermediation, a system that has become known as shadow banking. The complex bank holding companies of today are the best example of hybrid intermediaries, but I argue that financial firms from the "nonbank" space can just as easily evolve into conglomerates with similar organizational structure, thus acquiring the cap...

  9. Hybrid composites

    CSIR Research Space (South Africa)

    Jacob John, Maya

    2009-04-01

    Full Text Available effect was observed for the elongation at break of the hybrid composites. The impact strength of the hybrid composites increased with the addition of glass fibres. The tensile and impact properties of thermoplastic natural rubber reinforced short... panels made from conventional structural materials. Figure 3 illustrates the performance of cellular biocomposite panels against conventional systems used for building and residential construction, namely a pre- cast pre-stressed hollow core concrete...

  10. The APTIMA HPV assay versus the Hybrid Capture 2 test in triage of women with ASC-US or LSIL cervical cytology: a meta-analysis of the diagnostic accuracy.

    Science.gov (United States)

    Arbyn, Marc; Roelens, Jolien; Cuschieri, Kate; Cuzick, Jack; Szarewski, Ann; Ratnam, Sam; Reuschenbach, Miriam; Belinson, Suzanne; Belinson, Jerome L; Monsonego, Joseph

    2013-01-01

    Testing for DNA of 13 high-risk HPV types with the Hybrid Capture 2 (HC2) test has consistently been shown to perform better in triage of women with cervical cytology results showing atypical squamous cells of undetermined significance (ASC-US) but often not in triage of low-grade squamous intraepithelial lesions (LSIL) detected in cervical cancer screening. In a meta-analysis, we compared the accuracy of the APTIMA HPV test, which identifies RNA of 14 high-risk HPV types, to HC2 for the triage of women with ASC-US or LSIL. Literature search-targeted studies where the accuracy of APTIMA HPV and HC2 for detection of underlying CIN2/3+ was assessed concomitantly including verification of all cases of ASC-US and LSIL. HSROC (Hierarchical Summary ROC) curve regression was used to compute the pooled absolute and relative sensitivity and specificity. Eight studies, comprising 1,839 ASC-US and 1,887 LSIL cases, were retrieved. The pooled sensitivity and specificity of APTIMA to triage ASC-US to detect underlying CIN3 or worse was 96.2% (95% CI = 91.7-98.3%) and 54.9% (95% CI = 43.5-65.9%), respectively. APTIMA and HC2 showed similar pooled sensitivity; however, the specificity of the former was significantly higher (ratio: 1.19; 95% CI = 1.08-1.31 for CIN2+). The pooled sensitivity and specificity of APTIMA to triage LSIL were 96.7% (95% CI = 91.4-98.9%) and 38.7% (95% CI = 30.5-47.6%) for CIN3+. APTIMA was as sensitive as HC2 but more specific (ratio: 1.35; 95% CI = 1.11-1.66). Results were similar for detection of CIN2 or worse. In both triage of ASC-US and LSIL, APTIMA is as sensitive but more specific than HC2 for detecting cervical precancer.

  11. Evaluación de una técnica de hibridación reversa para identificación rápida de micobacterias en Chile Evaluation of a reverse hybridization assay for the identification of Mycobacterium in Chile

    Directory of Open Access Journals (Sweden)

    Tamara Leiva C

    2012-03-01

    Full Text Available Objetivos: Se evalúa una técnica para la identificación de micobacterias basada en la nueva tecnología de hibridación en tiras con sondas (Line Probe Assays-LiPAs. Métodos: Se analizaron 74 cepas, correspondientes a 23 especies y/o complejos, preclasificadas mediante pruebas bioquímicas tradicionales, sondas genéticas y PRA (análisis de patrones de restricción, identificables y no identi-ficables a nivel de especie por el kit utilizado. El kit evaluado, GenoType CM (Hain Lifescience, Nehren, Alemania, permite la identificación genética molecular de 14 de las especies micobacterianas más comunes, mediante una PCR múltiple e hibridación reversa del producto en tiras con sondas de regiones genéticas de ARNr de 23S. Resultados: Con la utilización de este ensayo para identificación de Micobacterias se obtuvo 94,0% (CI95% 84,4-98,0 de sensibilidad y 88,0% (CI95% 46,7-99,3 de especificidad totales. Conclusiones: Se concluye que GenoType CM constituye una herramienta adecuada para la identificación de micobacterias, rápida, sensible, operativa en las actuales condiciones de trabajo del Laboratorio de Referencia Nacional de Micobacterias en Chile y que podría constituir un avance para el acortamiento en los tiempos que demora el proceso, lo que implica una mejor oportunidad de aplicación de tratamiento sólo en los casos en que éste sea requerido.Objective: Identification for Mycobacterium assay based in the new technology of reverse hybridization DNA probe assay was evaluated (Line Probe Assays-LiPAs. Methods: 74 strains belonging to 23 mycobacterial species or complex classified previously by classical biochemical methods, genetic probes and PRA (patterns of restriction analysis, with and without specific pattern expected to be identified at specie level were analysed.The utilized test, GenoType CM (Hain Lifescience, Nehren, Alemania, is able of identifying 14 of the most common mycobacterial species after a multiplex PCR

  12. Defining the needs for gas centrifuge enrichment plants advanced safeguards

    Energy Technology Data Exchange (ETDEWEB)

    Boyer, Brian David [Los Alamos National Laboratory; Erpenbeck, Heather H [Los Alamos National Laboratory; Miller, Karen A [Los Alamos National Laboratory; Swinhoe, Martyn T [Los Alamos National Laboratory; Ianakiev, Kiril [Los Alamos National Laboratory; Marlow, Johnna B [Los Alamos National Laboratory

    2010-04-05

    Current safeguards approaches used by the International Atomic Energy Agency (IAEA) at gas centrifuge enrichment plants (GCEPs) need enhancement in order to verify declared low-enriched (LEU) production, detect undeclared LEU production and detect highly enriched uranium (HEU) production with adequate detection probability using nondestructive assay (NDA) techniques. At present inspectors use attended systems, systems needing the presence of an inspector for operation, during inspections to verify the mass and {sup 235}U enrichment of declared UF{sub 6} containers used in the process of enrichment at GCEPs. In verifying declared LEU production, the inspectors also take samples for off-site destructive assay (DA) which provide accurate data, with 0.1% to 0.5% measurement uncertainty, on the enrichment of the UF{sub 6} feed, tails, and product. However, taking samples of UF{sub 6} for off-site analysis is a much more labor and resource intensive exercise for the operator and inspector. Furthermore, the operator must ship the samples off-site to the IAEA laboratory which delays the timeliness of results and interruptions to the continuity of knowledge (CofK) of the samples during their storage and transit. This paper contains an analysis of possible improvements in unattended and attended NDA systems such as process monitoring and possible on-site analysis of DA samples that could reduce the uncertainty of the inspector's measurements and provide more effective and efficient IAEA GCEPs safeguards. We also introduce examples advanced safeguards systems that could be assembled for unattended operation.

  13. Initial report on characterization of excess highly enriched uranium

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1996-07-01

    DOE`s Office of Fissile Materials Disposition assigned to this Y-12 division the task of preparing a report on the 174.4 metric tons of excess highly enriched U. Characterization included identification by category, gathering existing data (assay), defining the likely needed processing steps for prepping for transfer to a blending site, and developing a range of preliminary cost estimates for those steps. Focus is on making commercial reactor fuel as a final disposition path.

  14. Identification and characterization of Argonaute gene family and meiosis-enriched Argonaute during sporogenesis in maize

    Institute of Scientific and Technical Information of China (English)

    Zuxin Zhang

    2014-01-01

    Argonaute (AGO) proteins play a key role in regulation of gene expression through smal RNA‐directed RNA cleavage and translational repression, and are essential for multiple developmental processes. In the present study, 17 AGO genes of maize (Zea mays L., ZmAGOs) were identified using a Hidden Markov Model and validated by rapid amplifica-tion of cDNA ends assay. Subsequently, quantitative PCR revealed that expressions of these genes were higher in reproductive than in vegetative tissues. AGOs presented five temporal and spatial expression patterns, which were likely modulated by DNA methylation, 50‐untranslated exons and microRNA‐mediated feedback loops. Intriguingly, ZmAGO18b was highly expressed in tassels during meiosis. Furthermore, in situ hybridization and immunofluorescence showed that ZmA- GO18b was enriched in the tapetum and germ cel s in meiotic anthers. We hypothesized that ZmAGOs are highly expressed in reproductive tissues, and that ZmAGO18b is a tapetum and germ cel‐specific member of the AGO family in maize.

  15. Biomimetic fiber assembled gradient hydrogel to engineer glycosaminoglycan enriched and mineralized cartilage: An in vitro study.

    Science.gov (United States)

    Mohan, Neethu; Wilson, Jijo; Joseph, Dexy; Vaikkath, Dhanesh; Nair, Prabha D

    2015-12-01

    The study investigated the potential of electrospun fiber assembled hydrogel, with physical gradients of chondroitin sulfate (CS) and sol-gel-derived bioactive glass (BG), to engineer hyaline and mineralized cartilage in a single 3D system. Electrospun poly(caprolactone) (PCL) fibers incorporated with 0.1% w/w of CS (CSL) and 0.5% w/w of CS (CSH), 2.4% w/w of BG (BGL) and 12.5% w/w of BG (BGH) were fabricated. The CS showed a sustained release up to 3 days from CSL and 14 days from CSH fibers. Chondrocytes secreted hyaline like matrix with higher sulfated glycosaminoglycans (sGAG), collagen type II and aggrecan on CSL and CSH fibers. Mineralization was observed on BGL and BGH fibers when incubated in simulated body fluid for 14 days. Chondrocytes cultured on these fibers secreted a mineralized matrix that consisted of sGAG, hypertrophic proteins, collagen type X, and osteocalcin. The CS and BG incorporated PCL fiber mats were assembled in an agarose-gelatin hydrogel to generate a 3D hybrid scaffold. The signals in the fibers diffused and generated continuous opposing gradients of CS (chondrogenic signal) and BG (mineralization) in the hydrogel. The chondrocytes were encapsulated in hybrid scaffolds; live dead assay at 48 h showed viable cells. Cells maintained their phenotype and secreted specific extracellular matrix (ECM) in response to signals within the hydrogel. Continuous opposing gradients of sGAG enriched and mineralized ECM were observed surrounding each cell clusters on gradient hydrogel after 14 days of culture in response to the physical gradients of raw materials CS and BG. A construct with gradient mineralization might accelerate integration to subchondral bone during in vivo regeneration.

  16. Predictive Assay For Cancer Targets

    Energy Technology Data Exchange (ETDEWEB)

    Suess, A; Nguyen, C; Sorensen, K; Montgomery, J; Souza, B; Kulp, K; Dugan, L; Christian, A

    2005-09-19

    Early detection of cancer is a key element in successful treatment of the disease. Understanding the particular type of cancer involved, its origins and probable course, is also important. PhIP (2-amino-1-methyl-6 phenylimidazo [4,5-b]pyridine), a heterocyclic amine produced during the cooking of meat at elevated temperatures, has been shown to induce mammary cancer in female, Sprague-Dawley rats. Tumors induced by PhIP have been shown to contain discreet cytogenetic signature patterns of gains and losses using comparative genomic hybridization (CGH). To determine if a protein signature exists for these tumors, we are analyzing expression levels of the protein products of the above-mentioned tumors in combination with a new bulk protein subtractive assay. This assay produces a panel of antibodies against proteins that are either on or off in the tumor. Hybridization of the antibody panel onto a 2-D gel of tumor or control protein will allow for identification of a distinct protein signature in the tumor. Analysis of several gene databases has identified a number of rat homologs of human cancer genes located in these regions of gain and loss. These genes include the oncogenes c-MYK, ERBB2/NEU, THRA and tumor suppressor genes EGR1 and HDAC3. The listed genes have been shown to be estrogen-responsive, suggesting a possible link between delivery of bio-activated PhIP to the cell nucleus via estrogen receptors and gene-specific PhIP-induced DNA damage, leading to cell transformation. All three tumors showed similar silver staining patterns compared to each other, while they all were different than the control tissue. Subsequent screening of these genes against those from tumors know to be caused by other agents may produce a protein signature unique to PhIP, which can be used as a diagnostic to augment optical and radiation-based detection schemes.

  17. Interaction of the heterotrimeric G protein alpha subunit SSG-1 of Sporothrix schenckii with proteins related to stress response and fungal pathogenicity using a yeast two-hybrid assay

    Directory of Open Access Journals (Sweden)

    González-Méndez Ricardo

    2010-12-01

    Full Text Available Abstract Background Important biological processes require selective and orderly protein-protein interactions at every level of the signalling cascades. G proteins are a family of heterotrimeric GTPases that effect eukaryotic signal transduction through the coupling of cell surface receptors to cytoplasmic effector proteins. They have been associated with growth and pathogenicity in many fungi through gene knock-out studies. In Sporothrix schenckii, a pathogenic, dimorphic fungus, we previously identified a pertussis sensitive G alpha subunit, SSG-1. In this work we inquire into its interactions with other proteins. Results Using the yeast two-hybrid technique, we identified protein-protein interactions between SSG-1 and other important cellular proteins. The interactions were corroborated using co-immuneprecipitation. Using these techniques we identified a Fe/Mn superoxide dismutase (SOD, a glyceraldehyde-3-P dehydrogenase (GAPDH and two ion transport proteins, a siderophore-iron transporter belonging to the Major Facilitator Superfamily (MFS and a divalent-cation transporter of the Nramp (natural resistance-associated macrophage protein family as interacting with SSG-1. The cDNA's encoding these proteins were sequenced and bioinformatic macromolecular sequence analyses were used for the correct classification and functional assignment. Conclusions This study constitutes the first report of the interaction of a fungal G alpha inhibitory subunit with SOD, GAPDH, and two metal ion transporters. The identification of such important proteins as partners of a G alpha subunit in this fungus suggests possible mechanisms through which this G protein can affect pathogenicity and survival under conditions of environmental stress or inside the human host. The two ion transporters identified in this work are the first to be reported in S. schenckii and the first time they are identified as interacting with fungal G protein alpha subunits. The association

  18. LA PROTEÍNA PTHB DE Xanthomonas axonopodis pv. Manihotis ES AUTOACTIVA EN ENSAYOS DE DOBLE HÍBRIDO The PthB Protein from Xanthomonas axonopodis pv. Manihotis is an Autoactive in Yeast Two-Hybrid Assays

    Directory of Open Access Journals (Sweden)

    JULIANA GIL

    the yeast two hybrid expression vector pLAW10, generating a fusion protein with the Binding Domain (BD of the transcription factor GAL4. In this work, PthB was cloned in a translational fusion with Gal4-BD (DNA Binding Domain. After transforming this construct into a yeast strain, autoactivation of the reporter genes was observed, even at the highest concentrations of 3-AT. The deletion of the first, second or both NLS and the AAD did not eliminate the ability of autoactivation of PthB. These results show the impossibility of using PthB to screen a cassava cDNA library to identify the proteins interacting with PthB.

  19. Fluorescence in situ hybridization and aminocytes karyotyping for 1809 aminotic cells assay%1809份羊水细胞荧光原位杂交技术与核型检测结果分析

    Institute of Scientific and Technical Information of China (English)

    于丽; 付杰; 马京梅; 李昶; 潘虹; 杨慧霞

    2014-01-01

    目的:探讨荧光原位杂交( FISH)技术联合羊水细胞培养在检测胎儿染色体异常的临床应用价值。方法回顾性总结2012年7月1日至2013年12月31日在北京大学第一医院就诊的1809例孕17~38周进行FISH产前诊断妊娠妇女的详细临床资料及随访信息。羊水间期细胞选用13、18、21、X、Y特异性探针检测,同时进行常规羊水细胞培养核型分析,将两者结果进行对照分析。结果1809份羊水间期细胞FISH检测均成功,其中检出正常核型1767例,非整倍体39例、三倍体1例、嵌合体2例,与常规细胞染色体核型分析结果一致,另外34例结构异常、5例嵌合体及12例正常变异FISH技术未能检出。结论 FISH技术检测胎儿常见染色体数目异常具有快速简便、准确率高等优点,FISH技术联合羊水细胞培养能够更好的服务于产前诊断。%Objective To analyze the clinical application of fluorescence in situ hybridization ( FISH) and karyotype analysis in prenatal diagnosis of chromosomal abnormalities .Methods The prenatal diagnosis of chromosome aneuploidies by FISH analysis of chromosome-specific probes (chromosome 13,18, 21,X,Y) in interphase amniocytes of 1 809 pregnant women of 17-38 weeks of gestational age was used , comparisons with the karyotyping results was done simultaneously.All the 1 809 cases came from Peking University First Hospital from July 1,2012 to December 31,2013, and the relevant clinical data and birth follow-up information were collected.Results All the 1 809 cases had been successfully examined by FISH , including 1 767 normal cases and 42 cases of numerical abnormality (39 cases of aneuploid, 1 case of triploid and 2 cases of mosaicism),which were consistent with the karyotyping analysis .What′s more,34 cases of chromosomal structural abnormalities , 5 cases of chimera and 12 cases of normal variant were failed to detected by FISH.Conclusion With the advantages of high

  20. Colorimetric protein assay techniques.

    Science.gov (United States)

    Sapan, C V; Lundblad, R L; Price, N C

    1999-04-01

    There has been an increase in the number of colorimetric assay techniques for the determination of protein concentration over the past 20 years. This has resulted in a perceived increase in sensitivity and accuracy with the advent of new techniques. The present review considers these advances with emphasis on the potential use of such technologies in the assay of biopharmaceuticals. The techniques reviewed include Coomassie Blue G-250 dye binding (the Bradford assay), the Lowry assay, the bicinchoninic acid assay and the biuret assay. It is shown that each assay has advantages and disadvantages relative to sensitivity, ease of performance, acceptance in the literature, accuracy and reproducibility/coefficient of variation/laboratory-to-laboratory variation. A comparison of the use of several assays with the same sample population is presented. It is suggested that the most critical issue in the use of a chromogenic protein assay for the characterization of a biopharmaceutical is the selection of a standard for the calibration of the assay; it is crucial that the standard be representative of the sample. If it is not possible to match the standard with the sample from the perspective of protein composition, then it is preferable to use an assay that is not sensitive to the composition of the protein such as a micro-Kjeldahl technique, quantitative amino acid analysis or the biuret assay. In a complex mixture it might be inappropriate to focus on a general method of protein determination and much more informative to use specific methods relating to the protein(s) of particular interest, using either specific assays or antibody-based methods. The key point is that whatever method is adopted as the 'gold standard' for a given protein, this method needs to be used routinely for calibration.

  1. Suppression subtractive hybridization.

    Science.gov (United States)

    Ghorbel, Mohamed T; Murphy, David

    2011-01-01

    Comparing two RNA populations that differ from the effects of a single independent variable, such as a drug treatment or a specific genetic defect, can establish differences in the abundance of specific transcripts that vary in a population dependent manner. There are different methods for identifying differentially expressed genes. These methods include microarray, Serial Analysis of Gene Expression (SAGE), and quantitative Reverse-Transcriptase Polymerase Chain Reaction (qRT-PCR). Herein, the protocol describes an easy and cost-effective alternative that does not require prior knowledge of the transcriptomes under examination. It is specifically relevant when low levels of RNA starting material are available. This protocol describes the use of Switching Mechanism At RNA Termini Polymerase Chain Reaction (SMART-PCR) to amplify cDNA from small amounts of RNA. The amplified cDNA populations under comparison are then subjected to Suppression Subtractive Hybridization (SSH-PCR). SSH-PCR is a technique that couples subtractive hybridization with suppression PCR to selectively amplify fragments of differentially expressed genes. The resulting products are cDNA populations enriched for significantly overrepresented transcripts in either of the two input RNAs. These cDNA populations can then be cloned to generate subtracted cDNA library. Microarrays made with clones from the subtracted forward and reverse cDNA libraries are then screened for differentially expressed genes using targets generated from tester and driver total RNAs.

  2. Absolute nuclear material assay

    Science.gov (United States)

    Prasad, Manoj K [Pleasanton, CA; Snyderman, Neal J [Berkeley, CA; Rowland, Mark S [Alamo, CA

    2012-05-15

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  3. Enrichment of light hydrocarbon mixture

    Science.gov (United States)

    Yang; Dali; Devlin, David; Barbero, Robert S.; Carrera, Martin E.; Colling, Craig W.

    2010-08-10

    Light hydrocarbon enrichment is accomplished using a vertically oriented distillation column having a plurality of vertically oriented, nonselective micro/mesoporous hollow fibers. Vapor having, for example, both propylene and propane is sent upward through the distillation column in between the hollow fibers. Vapor exits neat the top of the column and is condensed to form a liquid phase that is directed back downward through the lumen of the hollow fibers. As vapor continues to ascend and liquid continues to countercurrently descend, the liquid at the bottom of the column becomes enriched in a higher boiling point, light hydrocarbon (propane, for example) and the vapor at the top becomes enriched in a lower boiling point light hydrocarbon (propylene, for example). The hollow fiber becomes wetted with liquid during the process.

  4. 31 CFR 540.316 - Uranium enrichment.

    Science.gov (United States)

    2010-07-01

    ... 31 Money and Finance: Treasury 3 2010-07-01 2010-07-01 false Uranium enrichment. 540.316 Section... FOREIGN ASSETS CONTROL, DEPARTMENT OF THE TREASURY HIGHLY ENRICHED URANIUM (HEU) AGREEMENT ASSETS CONTROL REGULATIONS General Definitions § 540.316 Uranium enrichment. The term uranium enrichment means the process...

  5. HistoFlex-a microfluidic device providing uniform flow conditions enabling highly sensitive, reproducible and quantitative in situ hybridizations

    DEFF Research Database (Denmark)

    Søe, Martin Jensen; Okkels, Fridolin; Sabourin, David;

    2011-01-01

    slides of spotted DNA microarrays when applying probe concentrations generally used in in situ hybridization (ISH) assays. The HistoFlex's novel ability in online monitoring of an in situ hybridization assay was demonstrated using direct fluorescent detection of hybridization to 18S rRNA. Tissue sections...... were not visually damaged during assaying, which enabled adapting a complete ISH assay for detection of microRNAs (miRNA). The effects of flow based incubations on hybridization, antibody incubation and Tyramide Signal Amplification (TSA) steps were investigated upon adapting the ISH assay...

  6. Nematode Indicators of Organic Enrichment

    NARCIS (Netherlands)

    Ferris, H.; Bongers, A.M.T.

    2006-01-01

    The organisms of the soil food web, dependent on resources from plants or on amendment from other sources, respond characteristically to enrichment of their environment by organic matter. Primary consumers of the incoming substrate, including bacteria, fungi, plant-feeding nematodes, annelids, and

  7. Nematode Indicators of Organic Enrichment

    NARCIS (Netherlands)

    Ferris, H.; Bongers, A.M.T.

    2006-01-01

    The organisms of the soil food web, dependent on resources from plants or on amendment from other sources, respond characteristically to enrichment of their environment by organic matter. Primary consumers of the incoming substrate, including bacteria, fungi, plant-feeding nematodes, annelids, and s

  8. Fast facility spent-fuel and waste assay instrument. [Fluorinel Dissolution and Fuel Storage (FAST) Facility

    Energy Technology Data Exchange (ETDEWEB)

    Eccleston, G.W.; Johnson, S.S.; Menlove, H.O.; Van Lyssel, T.; Black, D.; Carlson, B.; Decker, L.; Echo, M.W.

    1983-01-01

    A delayed-neutron assay instrument was installed in the Fluorinel Dissolution and Fuel Storage Facility at Idaho National Engineering Laboratory. The dual-assay instrument is designed to measure both spent fuel and waste solids that are produced from fuel processing. A set of waste standards, fabricated by Los Alamos using uranium supplied by Exxon Nuclear Idaho Company, was used to calibrate the small-sample assay region of the instrument. Performance testing was completed before installation of the instrument to determine the effects of uranium enrichment, hydrogenous materials, and neutron poisons on assays. The unit was designed to measure high-enriched uranium samples in the presence of large neutron backgrounds. Measurements indicate that the system can assay low-enriched uranium samples with moderate backgrounds if calibrated with proper standards.

  9. Clinical Application and Evaluation of Polymerase Chain Reaction-Mic rowell Plate Hybridization Assay in the Detection of Mycobacterium tuberculosis%聚合酶链反应-微孔板杂交法检测结核杆菌的临床应用及其评价

    Institute of Scientific and Technical Information of China (English)

    杨正林; 傅明德; 杜琼; 杨明清; 乔国蓉; 唐蓉

    2001-01-01

    目的 对聚合酶链反应(PCR)-微孔板杂交法检测结核杆菌的临床适用性和可行性进行评价。方法 收集1130份临床标本,分别用抗酸染色、培养、PCR电泳及PCR微孔板杂交法进行结核杆菌检测,并结合临床诊断和疗效观察对实验结果进行分析。结果 100份临床证实无结核病的体液标本经抗酸染色和PCR电泳,分别有1例和2例假阳性,但经培养和PCR微孔板杂交法未查出阳性;其余1030份高度怀疑含有结核杆菌的体液标本的检测结果,以PCR微孔板杂交阳性检出例数最高(481/1030),其次为PCR电泳(406/1030)、培养(365/1030)以及抗酸染色(256/1030);χ2检验结果显示,PCR微孔板杂交的结核杆菌阳性检出例数与其它三种方法比较均呈显著或极显著性差异(P<0.0083及P<0.0017)。结论 PCR-微孔板杂交方法是特异、灵敏、准确、快速的结核杆菌检测方法,具有推广应用价值。%Objective To evaluate the clinical application an d feasibilityof polymerase chain reaction-microwell plate hybridization assay in the detection of M ycobacterium tuberculosis. Methods 1130 specimens with strong s uspicion for myc obacterium tuberculosis were collected from the hospitals and were detected by fast bacilli stain,culture,PCR-electrophoresis and PCR-microwell plate hybridi za tion respectively. The laboratory results were analyzed in combination with the symptoms and signs of patients and the observations on treatment.Also detected were 100 samples from the clinically evidenced non-tuberculosis patients. Results In the 100 samples collected from the patients without tuber culosis, the PCR- h ybridization method and culture method did not detect Mycobacterium tuberculosis , but the fast bacilli stain method and PCR-electrophoresis method brought out o ne and two false-positive results respectively. The results of testing the 1030 clinical samples which probably contained

  10. Hybrid microelectronic technology

    Science.gov (United States)

    Moran, P.

    Various areas of hybrid microelectronic technology are discussed. The topics addressed include: basic thick film processing, thick film pastes and substrates, add-on components and attachment methods, thin film processing, and design of thick film hybrid circuits. Also considered are: packaging hybrid circuits, automating the production of hybrid circuits, application of hybrid techniques, customer's view of hybrid technology, and quality control and assurance in hybrid circuit production.

  11. Lump corrections for radioactive waste assay.

    Science.gov (United States)

    Miller, T J

    2009-09-01

    Previous studies have shown that automated radioactive waste assay techniques, such as segmented gamma scanner (SGS) and automated qualitative and quantitative (AQ2), have severely underestimated fissile material due to either the malfunction or absence of appropriate lump correction routines. This paper examines the application of manual techniques, such as Monte Carlo N particle (MCNP) and spectral non-destructive assay platform (SNAP) software, to lump corrections in plutonium (Pu), enriched uranium (EU) and depleted uranium (DU) waste streams. Excellent results have been obtained when comparing MCNP with SNAP and applying the SNAP lump correction routine to a range of simulated and typical wastes containing various Pu and EU lump sizes. It has been concluded that the need for lump corrections was relatively rare and usually apparent from abnormal gamma ray peak area ratios, since most AWE waste streams are only lightly shielded.

  12. Genomic Prediction of Barley Hybrid Performance

    Directory of Open Access Journals (Sweden)

    Norman Philipp

    2016-07-01

    Full Text Available Hybrid breeding in barley ( L. offers great opportunities to accelerate the rate of genetic improvement and to boost yield stability. A crucial requirement consists of the efficient selection of superior hybrid combinations. We used comprehensive phenotypic and genomic data from a commercial breeding program with the goal of examining the potential to predict the hybrid performances. The phenotypic data were comprised of replicated grain yield trials for 385 two-way and 408 three-way hybrids evaluated in up to 47 environments. The parental lines were genotyped using a 3k single nucleotide polymorphism (SNP array based on an Illumina Infinium assay. We implemented ridge regression best linear unbiased prediction modeling for additive and dominance effects and evaluated the prediction ability using five-fold cross validations. The prediction ability of hybrid performances based on general combining ability (GCA effects was moderate, amounting to 0.56 and 0.48 for two- and three-way hybrids, respectively. The potential of GCA-based hybrid prediction requires that both parental components have been evaluated in a hybrid background. This is not necessary for genomic prediction for which we also observed moderate cross-validated prediction abilities of 0.51 and 0.58 for two- and three-way hybrids, respectively. This exemplifies the potential of genomic prediction in hybrid barley. Interestingly, prediction ability using the two-way hybrids as training population and the three-way hybrids as test population or vice versa was low, presumably, because of the different genetic makeup of the parental source populations. Consequently, further research is needed to optimize genomic prediction approaches combining different source populations in barley.

  13. Cell viability assays: introduction.

    Science.gov (United States)

    Stoddart, Martin J

    2011-01-01

    The measurement of cell viability plays a fundamental role in all forms of cell culture. Sometimes it is the main purpose of the experiment, such as in toxicity assays. Alternatively, cell viability can be used to -correlate cell behaviour to cell number, providing a more accurate picture of, for example, anabolic -activity. There are wide arrays of cell viability methods which range from the most routine trypan blue dye exclusion assay to highly complex analysis of individual cells, such as using RAMAN microscopy. The cost, speed, and complexity of equipment required will all play a role in determining the assay used. This chapter aims to provide an overview of many of the assays available today.

  14. Tube-Forming Assays.

    Science.gov (United States)

    Brown, Ryan M; Meah, Christopher J; Heath, Victoria L; Styles, Iain B; Bicknell, Roy

    2016-01-01

    Angiogenesis involves the generation of new blood vessels from the existing vasculature and is dependent on many growth factors and signaling events. In vivo angiogenesis is dynamic and complex, meaning assays are commonly utilized to explore specific targets for research into this area. Tube-forming assays offer an excellent overview of the molecular processes in angiogenesis. The Matrigel tube forming assay is a simple-to-implement but powerful tool for identifying biomolecules involved in angiogenesis. A detailed experimental protocol on the implementation of the assay is described in conjunction with an in-depth review of methods that can be applied to the analysis of the tube formation. In addition, an ImageJ plug-in is presented which allows automatic quantification of tube images reducing analysis times while removing user bias and subjectivity.

  15. Transgenic Animal Mutation Assays

    Institute of Scientific and Technical Information of China (English)

    Tao Chen; Ph.D.D.A.B.T.

    2005-01-01

    @@ The novel transgenic mouse and rat mutation assays have provided a tool for analyzing in vivo mutation in any tissue, thus permitting the direct comparison of cancer incidence with mutant frequency.

  16. Assays for thrombopoietin

    Energy Technology Data Exchange (ETDEWEB)

    McDonald, T.P.

    1977-01-01

    In summary, thrombopoietin levels have been determined indirectly by measuring thrombocytopoiesis in assay animals (platelet counting, measurement of isotope incorporation into newly formed platelets, changes in platelet sizes, or alterations in number and size of megakaryocytes) and by use of an immunoassay. Although much work remains, it seems clear at the present time that isotopic uptake into platelets of specially prepared assay mice (rebound-thrombocytosis) is superior to the other techniques now available for the measurement of thrombopoietin. However, the ideal assay for TSF which is specific, rapid, and inexpensive is yet to be developed. An immunoassay is in the development stage, but will require additional work before it can be utilized for the routine assay of TSF.

  17. Detection of 11q23/MLL gene rearrangement in hematological malignancies with fluorescence in situ hybridization assay%间期荧光原位杂交技术检测恶性血液病的11q23/MLL基因重排

    Institute of Scientific and Technical Information of China (English)

    彭智; 冯文莉; 肖志坚; 周迎春; 刘光平; 刘基铎

    2008-01-01

    目的 分析伴有11q23/MLL基因重排的恶性血液病的细胞遗传学特点,探讨荧光原位杂交技术( FISH)在诊断及鉴定恶性血液病11q23/MLL基因重排中的价值.方法 用间期FISH分析11q23/MLL基因易位细胞的30例恶性血液病患者的核型特征, 用MLL双色分离探针绿色标记在 (5′MLL, 光谱绿) 和 (3′MLL, 光谱桔红).结果 应用常规细胞遗传学及间期FISH分析白血病患者30例,结果显示11q23+/MLL+患者9例(30.0%),11q23-/MLL+患者4例(13.3%),11q23+/MLL-患者2例(6.7%),11q23-/MLL-患者15例,检测到部分病例染色体核型分析与间期FISH方法检测11q23异常与MLL基因重排不一致.结论 FISH在检测11q23/MLL基因重排方面与传统的常规细胞遗传学相比具有检出率高的优势,能更有效、直观地分析恶性血液病的染色体异常,对于恶性血液病的诊断以及异常染色体的检出具有广泛的应用前景.%Objective To analyze the cytogenetical characteristic of 11q23/MLL gene rearrangement in hematological malignancies and to investigate the value of fluorescence in situ hybridization (FISH) assay in the diagnosis and appraisal of 11q23/MLL gene rearrangement.Methods FISH assay was performed to analyze the karyotypic characteristic of 11q23/MLL gene rearranged cells in 30 patients with hematological malignancies. The dual color probe was adopted. 5′MLL was labeled with spectrum green and 3′MLL labeled with spectrum orange.Results The incidence of 11q23+/MLL+ in acute leukemia (AL) patients was 30.0% (10 out of 30 cases) and 11q23-/MLL+ was 13.3% (4 cases) , and 11q23+/MLL- was 6.7% (2 cases) and the karyotype of 15 cases was normal. For some patients, different results were obtained by using conentional cytogenetical analysis and interphased FISH assay for detecting 11q23/MLL gene rearrangements.Conclusion FISH assay has greater advantage over cytogenetical study in the analysis of 11q23/MLL abnormality. It is also a promising tool in

  18. New Rapid Spore Assay

    Science.gov (United States)

    Kminek, Gerhard; Conley, Catharine

    2012-07-01

    The presentation will detail approved Planetary Protection specifications for the Rapid Spore Assay for spacecraft components and subsystems. Outlined will be the research and studies on which the specifications were based. The research, funded by ESA and NASA/JPL, was conducted over a period of two years and was followed by limited cleanroom studies to assess the feasibility of this assay during spacecraft assembly.

  19. Enrichment of lanthanides in aragonite

    Institute of Scientific and Technical Information of China (English)

    瞿成利; 路波; 刘刚

    2009-01-01

    Using the constant addition technique,the coprecipitation of lanthanum,gadolinium,and lutetium with aragonite in seawater was experimentally investigated at 25 ℃.Their concentrations in aragonite overgrowths were determined by inductive coupled plasma mass spectrometer.All these lanthanides were strongly enriched in aragonite overgrowths.The amount of lanthanum,gadolinium,and lutetium incorporated into aragonite accounted for 57%-99%,50%-89%,and 40%-91% of their initial total amount,respectively.With the in...

  20. Enrichment and aggression in primates.

    Science.gov (United States)

    Honess, P E; Marin, C M

    2006-01-01

    There is considerable evidence that primates housed under impoverished conditions develop behavioural abnormalities, including, in the most extreme example, self-harming behaviour. This has implications for all contexts in which primates are maintained in captivity from laboratories to zoos since by compromising the animals' psychological well-being and allowing them to develop behavioural abnormalities their value as appropriate educational and research models is diminished. This review examines the extensive body of literature documenting attempts to improve living conditions with a view to correcting behavioural abnormalities and housing primates in such a way that they are encouraged to exhibit a more natural range and proportion of behaviours, including less self-directed and social aggression. The results of housing, feeding, physical, sensory and social enrichment efforts are examined with specific focus on their effect on aggressive behaviour and variation in their use and efficacy. It is concluded that while inappropriate or poorly distributed enrichment may encourage aggressive competition, enrichment that is species, sex, age and background appropriate can dramatically reduce aggression, can eliminate abnormal behaviour and substantially improve the welfare of primates maintained in captivity.

  1. Fluorescence Hybridization Assay Based On Chitosan-Linked Softarrays

    Science.gov (United States)

    2003-07-01

    was incubated in the wells to reduce the Schiff base resulting from the reaction of aldehyde and amine groups. After this reaction, the yellowish...color representative of a Schiff base disappeared and the background fluorescence signal dropped to the initial ~8 to 12 fluorescence intensity (FI

  2. Yeast Protein–Protein Interaction Assays and Screens

    NARCIS (Netherlands)

    Folter, de S.; Immink, G.H.

    2011-01-01

    Most transcription factors fulfill their role in protein complexes. As a consequence, information about their interaction capacity sheds light on a protein’s function and the molecular mechanism underlying this activity. The yeast two-hybrid GAL4 (Y2H) assay is a powerful method to unravel and ident

  3. Comparison of a PCR serotyping assay, Check&Trace assay for Salmonella, and Luminex Salmonella serotyping assay for the characterization of Salmonella enterica identified from fresh and naturally contaminated cilantro.

    Science.gov (United States)

    Jean-Gilles Beaubrun, J; Ewing, L; Jarvis, K; Dudley, K; Grim, C; Gopinath, G; Flamer, M-L; Auguste, W; Jayaram, A; Elmore, J; Lamont, M; McGrath, T; Hanes, D E

    2014-09-01

    Salmonella enterica isolated from fresh cilantro samples collected through the USDA/AMS Microbiological Data Program (MDP) were used to compare a PCR serotyping assay against the Check&Trace assay and the Luminex (BioPlex) Salmonella serotyping assay. The study was conducted to evaluate the effectiveness of the three methods for serotyping Salmonella from both enrichment broth cultures and pure Salmonella cultures. In this investigation, Salmonella spp. serotyping was conducted using 24 h enrichment broth cultures and pure Salmonella cultures from cilantro samples, with the PCR serotyping assay. Conversely, the Check&Trace and Luminex for Salmonella assays required pure cultures for Salmonella serotyping. The cilantro samples contained S. enterica serovar Montevideo, Newport, Saintpaul, and Tennessee, identified by the PCR serotyping assay and Check&Trace for Salmonella, but the Luminex assay only identified two of the four serotypes of the cilantro samples. The anticipated impact from this study is that the PCR serotyping assay provides a time- and cost-effective means for screening, identifying and serotyping Salmonella using DNA extracted from 24 h enrichment cilantro samples.

  4. Network-based functional enrichment

    Directory of Open Access Journals (Sweden)

    Poirel Christopher L

    2011-11-01

    Full Text Available Abstract Background Many methods have been developed to infer and reason about molecular interaction networks. These approaches often yield networks with hundreds or thousands of nodes and up to an order of magnitude more edges. It is often desirable to summarize the biological information in such networks. A very common approach is to use gene function enrichment analysis for this task. A major drawback of this method is that it ignores information about the edges in the network being analyzed, i.e., it treats the network simply as a set of genes. In this paper, we introduce a novel method for functional enrichment that explicitly takes network interactions into account. Results Our approach naturally generalizes Fisher’s exact test, a gene set-based technique. Given a function of interest, we compute the subgraph of the network induced by genes annotated to this function. We use the sequence of sizes of the connected components of this sub-network to estimate its connectivity. We estimate the statistical significance of the connectivity empirically by a permutation test. We present three applications of our method: i determine which functions are enriched in a given network, ii given a network and an interesting sub-network of genes within that network, determine which functions are enriched in the sub-network, and iii given two networks, determine the functions for which the connectivity improves when we merge the second network into the first. Through these applications, we show that our approach is a natural alternative to network clustering algorithms. Conclusions We presented a novel approach to functional enrichment that takes into account the pairwise relationships among genes annotated by a particular function. Each of the three applications discovers highly relevant functions. We used our methods to study biological data from three different organisms. Our results demonstrate the wide applicability of our methods. Our algorithms are

  5. Ancient hybridization and genomic stabilization in a swordtail fish.

    Science.gov (United States)

    Schumer, Molly; Cui, Rongfeng; Powell, Daniel L; Rosenthal, Gil G; Andolfatto, Peter

    2016-06-01

    A rapidly increasing body of work is revealing that the genomes of distinct species often exhibit hybrid ancestry, presumably due to postspeciation hybridization between closely related species. Despite the growing number of documented cases, we still know relatively little about how genomes evolve and stabilize following hybridization, and to what extent hybridization is functionally relevant. Here, we examine the case of Xiphophorus nezahualcoyotl, a teleost fish whose genome exhibits significant hybrid ancestry. We show that hybridization was relatively ancient and is unlikely to be ongoing. Strikingly, the genome of X. nezahualcoyotl has largely stabilized following hybridization, distinguishing it from examples such as human-Neanderthal hybridization. Hybridization-derived regions are remarkably distinct from other regions of the genome, tending to be enriched in genomic regions with reduced constraint. These results suggest that selection has played a role in removing hybrid ancestry from certain functionally important regions. Combined with findings in other systems, our results raise many questions about the process of genomic stabilization and the role of selection in shaping patterns of hybrid ancestry in the genome. © 2016 John Wiley & Sons Ltd.

  6. NEAT: an efficient network enrichment analysis test

    OpenAIRE

    Signorelli, Mirko; Vinciotti, Veronica; Wit, Ernst C

    2016-01-01

    Background Network enrichment analysis is a powerful method, which allows to integrate gene enrichment analysis with the information on relationships between genes that is provided by gene networks. Existing tests for network enrichment analysis deal only with undirected networks, they can be computationally slow and are based on normality assumptions. Results We propose NEAT, a test for network enrichment analysis. The test is based on the hypergeometric distribution, which naturally arises ...

  7. Hybrid Gear

    Science.gov (United States)

    Handschuh, Robert F. (Inventor); Roberts, Gary D. (Inventor)

    2016-01-01

    A hybrid gear consisting of metallic outer rim with gear teeth and metallic hub in combination with a composite lay up between the shaft interface (hub) and gear tooth rim is described. The composite lay-up lightens the gear member while having similar torque carrying capability and it attenuates the impact loading driven noise/vibration that is typical in gear systems. The gear has the same operational capability with respect to shaft speed, torque, and temperature as an all-metallic gear as used in aerospace gear design.

  8. Hybrid Qualifications

    DEFF Research Database (Denmark)

    has turned out as a major focus of European education and training policies and certainly is a crucial principle underlying the European Qualifications Framework (EQF). In this context, «hybrid qualifications» (HQ) may be seen as an interesting approach to tackle these challenges as they serve «two...... masters», i.e. by producing skills for the labour market and enabling individuals to progress more or less directly to higher education. The specific focus of this book is placed on conditions, structures and processes which help to combine VET with qualifications leading into higher education...

  9. Comparison of three human papillomavirus DNA assays and one mRNA assay in women with abnormal cytology

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Lynge, Elsebeth; Ejegod, Ditte;

    2014-01-01

    OBJECTIVE: To compare the clinical characteristics of four human papillomavirus (HPV) assays: hybrid capture 2 (HC2), cobas, CLART, and APTIMA in Danish women with abnormal cytology. METHODS: SurePath samples from 367 consecutive women from Copenhagen, with atypical squamous cells of undetermined...

  10. Enriching Music and Language Arts Experiences

    Science.gov (United States)

    Flohr, John W.

    2006-01-01

    The article focuses on enriching music and language arts experiences of students. Music can enrich literature and language arts, poetry, theater arts, transitions, science, and math, as well as help meet special learner needs. A well-understood example of enrichment is the alphabet song. A music or classroom teacher using the alphabet song helps…

  11. How Did the IGM Become Enriched?

    CERN Document Server

    Aguirre, A; Aguirre, Anthony; Schaye, Joop

    2006-01-01

    The enrichment of the intergalactic medium with heavy elements is a process that lies at the nexus of poorly-understood aspects of physical cosmology. We review current understanding of the processes that may remove metals from galaxies, the basic predictions of these models, the key observational constraints on enrichment, and how intergalactic enrichment may be used to test cosmological simulations.

  12. Next-generation hybridization and introgression

    Science.gov (United States)

    Twyford, A D; Ennos, R A

    2012-01-01

    Hybridization has a major role in evolution—from the introgression of important phenotypic traits between species, to the creation of new species through hybrid speciation. Molecular studies of hybridization aim to understand the class of hybrids and the frequency of introgression, detect the signature of ancient hybridization, and understand the behaviour of introgressed loci in their new genomic background. This often involves a large investment in the design and application of molecular markers, leading to a compromise between the depth and breadth of genomic data. New techniques designed to assay a large sub-section of the genome, in association with next-generation sequencing (NGS) technologies, will allow genome-wide hybridization and introgression studies in organisms with no prior sequence data. These detailed genotypic data will unite the breadth of sampling of loci characteristic of population genetics with the depth of sequence information associated with molecular phylogenetics. In this review, we assess the theoretical and methodological constraints that limit our understanding of natural hybridization, and promote the use of NGS for detecting hybridization and introgression between non-model organisms. We also make recommendations for the ways in which emerging techniques, such as pooled barcoded amplicon sequencing and restriction site-associated DNA tags, should be used to overcome current limitations, and enhance our understanding of this evolutionary significant process. PMID:21897439

  13. Validation of Folate-Enriched Eggs as a Functional Food for Improving Folate Intake in Consumers.

    Science.gov (United States)

    Altic, Leslie; McNulty, Helene; Hoey, Leane; McAnena, Liadhan; Pentieva, Kristina

    2016-11-30

    Functional foods enriched with folate may be beneficial as a means of optimizing folate status in consumers. We recently developed novel eggs enriched with folate through folic acid supplementation of the hen's feed, but their potential to influence consumer folate status is unknown because the natural folate forms incorporated into the eggs may not necessarily be retained during storage and cooking. This study aimed to determine the stability of natural folates in folate-enriched eggs under typical conditions of storage and cooking. Total folate was determined by microbiological assay following tri-enzyme treatment in folate-enriched eggs and un-enriched (barn and free-range) on the day they were laid, after storage (up to 27 days) and after using four typical cooking methods (boiling, poaching, frying, scrambling) for different durations. On the day of laying, the folate content of enriched eggs was found to be significantly higher than that of un-enriched barn or free-range eggs (mean ± SD; 123.2 ± 12.4 vs. 41.2 ± 2.8 vs. 65.6 ± 18.5 µg/100 g; p functional foods with enriched folate content. Further studies will confirm their effectiveness in optimizing the biomarker folate status of consumers.

  14. Clinical validation of an ultra high-throughput spiral microfluidics for the detection and enrichment of viable circulating tumor cells.

    Directory of Open Access Journals (Sweden)

    Bee Luan Khoo

    Full Text Available Circulating tumor cells (CTCs are cancer cells that can be isolated via liquid biopsy from blood and can be phenotypically and genetically characterized to provide critical information for guiding cancer treatment. Current analysis of CTCs is hindered by the throughput, selectivity and specificity of devices or assays used in CTC detection and isolation.Here, we enriched and characterized putative CTCs from blood samples of patients with both advanced stage metastatic breast and lung cancers using a novel multiplexed spiral microfluidic chip. This system detected putative CTCs under high sensitivity (100%, n = 56 (Breast cancer samples: 12-1275 CTCs/ml; Lung cancer samples: 10-1535 CTCs/ml rapidly from clinically relevant blood volumes (7.5 ml under 5 min. Blood samples were completely separated into plasma, CTCs and PBMCs components and each fraction were characterized with immunophenotyping (Pan-cytokeratin/CD45, CD44/CD24, EpCAM, fluorescence in-situ hybridization (FISH (EML4-ALK or targeted somatic mutation analysis. We used an ultra-sensitive mass spectrometry based system to highlight the presence of an EGFR-activating mutation in both isolated CTCs and plasma cell-free DNA (cf-DNA, and demonstrate concordance with the original tumor-biopsy samples.We have clinically validated our multiplexed microfluidic chip for the ultra high-throughput, low-cost and label-free enrichment of CTCs. Retrieved cells were unlabeled and viable, enabling potential propagation and real-time downstream analysis using next generation sequencing (NGS or proteomic analysis.

  15. Against vaccine assay secrecy.

    Science.gov (United States)

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M

    2015-01-01

    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors.

  16. Against vaccine assay secrecy

    Science.gov (United States)

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M

    2015-01-01

    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors. PMID:25826194

  17. Enrichment planting without soil treatment

    Energy Technology Data Exchange (ETDEWEB)

    Hagner, Mats

    1998-12-31

    Where enrichment planting had been carried out with either of the two species Picea abies and Pinus contorta, the survival of the planted seedlings was at least as good as after planting in a normal clear cut area treated with soil scarification. This was in spite of the fact that the seedlings were placed shallow in the humus layer without any soil treatment. However, they were sheltered from insects by treatment before planting. Where enrichment planting was carried out with Pinus sylvestris the survival in dense forest was poor, but in open forest the survival was good. The growth of planted seedlings was enhanced by traditional clearing and soil treatment. However, this was for Pinus sylvestris not enough to compensate for the loss of time, 1-2 years, caused by arrangement of soil scarification. The growth of seedlings planted under crown cover was directly related to basal area of retained trees. However, the variation in height growth among individual seedlings was very big, which meant that some seedlings grow well also under a fairly dense forest cover. The pioneer species Pinus sylvestris reacted more strongly to basal area of retained trees than did the shade tolerant species Picea abies. Enrichment planting seems to be a necessary tool for preserving volume productivity, at places where fairly intensive harvest of mature trees has been carried out in stands of ordinary forest type in central Sweden. If double seedlings, with one Picea abies and one Pinus sylvestris, are used, the probability for long term establishment is enhanced 13 refs, 20 figs, 4 tabs

  18. Intuitionistic hybrid logic

    DEFF Research Database (Denmark)

    Braüner, Torben

    2011-01-01

    Intuitionistic hybrid logic is hybrid modal logic over an intuitionistic logic basis instead of a classical logical basis. In this short paper we introduce intuitionistic hybrid logic and we give a survey of work in the area.......Intuitionistic hybrid logic is hybrid modal logic over an intuitionistic logic basis instead of a classical logical basis. In this short paper we introduce intuitionistic hybrid logic and we give a survey of work in the area....

  19. Continuity Controlled Hybrid Automata

    OpenAIRE

    Bergstra, J. A.; Middelburg, C.A.

    2004-01-01

    We investigate the connections between the process algebra for hybrid systems of Bergstra and Middelburg and the formalism of hybrid automata of Henzinger et al. We give interpretations of hybrid automata in the process algebra for hybrid systems and compare them with the standard interpretation of hybrid automata as timed transition systems. We also relate the synchronized product operator on hybrid automata to the parallel composition operator of the process algebra. It turns out that the f...

  20. Boron enrichment in martian clay.

    Directory of Open Access Journals (Sweden)

    James D Stephenson

    Full Text Available We have detected a concentration of boron in martian clay far in excess of that in any previously reported extra-terrestrial object. This enrichment indicates that the chemistry necessary for the formation of ribose, a key component of RNA, could have existed on Mars since the formation of early clay deposits, contemporary to the emergence of life on Earth. Given the greater similarity of Earth and Mars early in their geological history, and the extensive disruption of Earth's earliest mineralogy by plate tectonics, we suggest that the conditions for prebiotic ribose synthesis may be better understood by further Mars exploration.

  1. Boron Enrichment in Martian Clay

    Science.gov (United States)

    Nagashima, Kazuhide; Freeland, Stephen J.

    2013-01-01

    We have detected a concentration of boron in martian clay far in excess of that in any previously reported extra-terrestrial object. This enrichment indicates that the chemistry necessary for the formation of ribose, a key component of RNA, could have existed on Mars since the formation of early clay deposits, contemporary to the emergence of life on Earth. Given the greater similarity of Earth and Mars early in their geological history, and the extensive disruption of Earth's earliest mineralogy by plate tectonics, we suggest that the conditions for prebiotic ribose synthesis may be better understood by further Mars exploration. PMID:23762242

  2. Digital doorway: enriching your mind

    CSIR Research Space (South Africa)

    Cambridge

    2008-11-01

    Full Text Available stream_source_info Cambridge_P_2008.pdf.txt stream_content_type text/plain stream_size 4576 Content-Encoding UTF-8 stream_name Cambridge_P_2008.pdf.txt Content-Type text/plain; charset=UTF-8 Digital Doorway: Enriching... your mind G CAMBRIDGE Meraka Institute, CSIR, PO Box 395, Pretoria, 0001 gcambrid@csir.co.za The Digital Doorway is a joint project between the South African Department of Science and Technology (DST) and the Meraka Institute of the CSIR...

  3. CTL ELISPOT assay.

    Science.gov (United States)

    Ranieri, Elena; Popescu, Iulia; Gigante, Margherita

    2014-01-01

    Enzyme-linked immune absorbent spot (Elispot) is a quantitative method for measuring relevant parameters of T cell activation. The sensitivity of Elispot allows the detection of low-frequency antigen-specific T cells that secrete cytokines and effector molecules, such as granzyme B and perforin. Cytotoxic T cell (CTL) studies have taken advantage with this high-throughput technology by providing insights into quantity and immune kinetics. Accuracy, sensitivity, reproducibility, and robustness of Elispot resulted in a wide range of applications in research as well as in diagnostic field. Actually, CTL monitoring by Elispot is a gold standard for the evaluation of antigen-specific T cell immunity in clinical trials and vaccine candidates where the ability to detect rare antigen-specific T cells is of relevance for immune diagnostic. The most utilized Elispot assay is the interferon-gamma (IFN-γ) test, a marker for CD8(+) CTL activation, but Elispot can also be used to distinguish different subsets of activated T cells by using other cytokines such as T-helper (Th) 1-type cells (characterized by the production of IFN-γ, IL-2, IL-6, IL-12, IL-21, and TNF-α), Th2 (producing cytokines like IL-4, IL-5, IL-10, and IL-13), and Th17 (IL-17) cells. The reliability of Elispot-generated data, by the evaluation of T cell frequency recognizing individual antigen/peptide, is the core of this method currently applied widely to investigate specific immune responses in cancer, infections, allergies, and autoimmune diseases. The Elispot assay is competing with other methods measuring single-cell cytokine production, e.g., intracellular cytokine by FACS or Miltenyi cytokine secretion assay. Other types of lymphocyte frequency and function assays include limiting dilution assay (LDA), cytotoxic T cell assay (CTL), and tetramer staining. Compared with respect to sensitivity the Elispot assay is outranking other methods to define frequency of antigen-specific lymphocytes. The method

  4. Assays for calcitonin receptors

    Energy Technology Data Exchange (ETDEWEB)

    Teitelbaum, A.P.; Nissenson, R.A.; Arnaud, C.D.

    1985-01-01

    The assays for calcitonin receptors described focus on their use in the study of the well-established target organs for calcitonin, bone and kidney. The radioligand used in virtually all calcitonin binding studies is /sup 125/I-labelled salmon calcitonin. The lack of methionine residues in this peptide permits the use of chloramine-T for the iodination reaction. Binding assays are described for intact bone, skeletal plasma membranes, renal plasma membranes, and primary kidney cell cultures of rats. Studies on calcitonin metabolism in laboratory animals and regulation of calcitonin receptors are reviewed.

  5. Development and Validation of a HPV-32 Specific PCR Assay

    Directory of Open Access Journals (Sweden)

    Leigh Janet

    2009-06-01

    Full Text Available Abstract Background Human Papillomavirus-32 (HPV-32 has traditionally been associated with focal-epithelial-hyperplasia (FEH. It is also present in 58% of oral warts of HIV-positive individuals whose prevalence is increasing. Current methods for the detection of HPV-32 are labor-intensive and insensitive so the goal of this work was to develop a highly sensitive and easy to use specific polymerase chain reaction (PCR assay. Materials and methods An HPV-32 L1 specific PCR assay was developed and optimized. The sensitivity and specificity was compared to previous assays utilized for detection (PGMY and MY09/11 PCR with dot blot hybridization using cloned HPV-32 L1, the closely related HPV-42 L1 as well as clinical samples (oral swabs and fluids from 89 HIV-positive subjects. Results The HPV-32 specific PCR assay showed improved sensitivity to 5 copies of HPV-32 as compared to the PGMY PCR, MY09/11 PCR and dot blot which had a limit of detection of approximately 3,000 copies. Using the HPV-32 dot blot hybridization assay as the gold standard, the HPV-32 specific PCR assay has a sensitivity of 95.8% and 88.9% by sample and subject, respectively, and specificity was 87.8% and 58.8% by sample and subject, respectively. The low sensitivity is due to the HPV-32 specific PCR assays ability to detect more HPV-32 positive samples and may be the new gold standard. Conclusion Due to the ease, sensitivity, and specificity the HPV-32 specific PCR assay is superior to previous assays and is ideal for detection of HPV-32 in large cohorts. This assay provides an excellent tool to study the natural history of HPV-32 infection and the development of oral warts.

  6. Environmental enrichment for primates in laboratories

    Science.gov (United States)

    Buchanan-Smith, H. M.

    2010-06-01

    Environmental enrichment is a critical component of Refinement, one of the 3Rs underlying humane experimentation on animals. In this paper I discuss why primates housed in laboratories, which often have constraints of space and study protocols, are a special case for enrichment. I outline a framework for categorising the different types of enrichment, using the marmoset as a case study, and summarise the methods used to determine what animals want/prefer. I briefly review the arguments that enrichment does not negatively affect experimental outcomes. Finally I focus on complexity and novelty, choice and control, the underlying features of enrichment that makes it successful, and how combined with a thorough understanding of natural history we can put effective enrichment into practice in laboratories. Throughout the paper I emphasise the need to evaluate enrichment to ensure it is having the desired effect.

  7. Hybridized tetraquarks

    Directory of Open Access Journals (Sweden)

    A. Esposito

    2016-07-01

    Full Text Available We propose a new interpretation of the neutral and charged X,Z exotic hadron resonances. Hybridized-tetraquarks are neither purely compact tetraquark states nor bound or loosely bound molecules but rather a manifestation of the interplay between the two. While meson molecules need a negative or zero binding energy, its counterpart for h-tetraquarks is required to be positive. The formation mechanism of this new class of hadrons is inspired by that of Feshbach metastable states in atomic physics. The recent claim of an exotic resonance in the Bs0π± channel by the D0 Collaboration and the negative result presented subsequently by the LHCb Collaboration are understood in this scheme, together with a considerable portion of available data on X,Z particles. Considerations on a state with the same quantum numbers as the X(5568 are also made.

  8. Hybridized Tetraquarks

    CERN Document Server

    Esposito, A.; Polosa, A.D.

    2016-01-01

    We propose a new interpretation of the neutral and charged X, Z exotic hadron resonances. Hybridized-tetraquarks are neither purely compact tetraquark states nor bound or loosely bound molecules. The latter would require a negative or zero binding energy whose counterpart in h-tetraquarks is a positive quantity. The formation mechanism of this new class of hadrons is inspired by that of Feshbach metastable states in atomic physics. The recent claim of an exotic resonance in the Bs pi+- channel by the D0 collaboration and the negative result presented subsequently by the LHCb collaboration are understood in this scheme, together with a considerable portion of available data on X, Z particles. Considerations on a state with the same quantum numbers as the X(5568) are also made.

  9. Preventive effects of selenium-enriched spiruline (SESP) on radiation pneumonitis.

    Science.gov (United States)

    Bai, Y; Wang, D; Cui, X; Yang, Z; Zhu, M; Zhang, Z; Xia, G; Gong, Y

    1998-01-01

    We studied the early effects of selenium-enriched spiruline (SESP) on radiation-induced pulmonary fibrosis using histopathology, cDNA-mRNA dot-blot hybridization, and a biochemical technique for quantifying the hydroxyproline content. We found marked lung injury in the irradiated group, whereas only slight hyperemia, hemorrhage, exudation, and thickness of the interalveolar wall in the lung in the selenium-treated group. The hydroxyproline (hyp) content in the lungs and the synthesis of type III collagen mRNA decreased in the SESP-treated group. Selenium-enriched spiruline may have a protective effect on the development of radiation pneumonitis.

  10. New oligosaccharyltransferase assay method.

    Science.gov (United States)

    Kohda, Daisuke; Yamada, Masaki; Igura, Mayumi; Kamishikiryo, Jun; Maenaka, Katsumi

    2007-11-01

    We developed a new in vitro assay for oligosaccharyltransferase (OST), which catalyzes the transfer of preassembled oligosaccharides on lipid carriers onto asparagine residues in polypeptide chains. The asparagine residues reside in the sequon, Asn-X-Thr/Ser, where X can be any amino acid residue except Pro. We demonstrate the potency of our assay using the OST from yeast. In our method, polyacrylamide gel electrophoresis is used to separate the glycopeptide products from the peptide substrates. The substrate peptide is fluorescently labeled and the formation of glycopeptides is analyzed by fluorescence gel imaging. Two in vitro OST assay methods are now widely used, but both the methods depend on previous knowledge of the oligosaccharide moiety: One method uses lectin binding as the separation mechanism and the other method uses biosynthetically or chemoenzymatically synthesized lipid-linked oligosaccharides as donors. N-linked protein glycosylation is found in all three domains of life, but little is known about the N-glycosylation in Archaea. Thus, our new assay, which does not require a priori knowledge of the oligosaccharides, will be useful in such cases. Indeed, we have detected the OST activity in the membrane fraction from a hyperthermophilic archaeon, Pyrococcus furiosus.

  11. Hyaluronic Acid Assays

    DEFF Research Database (Denmark)

    Itenov, Theis S; Kirkby, Nikolai S; Bestle, Morten H

    2015-01-01

    BACKGROUD: Hyaluronic acid (HA) is proposed as a marker of functional liver capacity. The aim of the present study was to compare a new turbidimetric assay for measuring HA with the current standard method. METHODS: HA was measured by a particle-enhanced turbidimetric immunoassay (PETIA) and enzyme...

  12. Instrument for assaying radiation

    Energy Technology Data Exchange (ETDEWEB)

    Coleman, Jody Rustyn; Farfan, Eduardo B.

    2016-03-22

    An instrument for assaying radiation includes a flat panel detector having a first side opposed to a second side. A collimated aperture covers at least a portion of the first side of the flat panel detector. At least one of a display screen or a radiation shield may cover at least a portion of the second side of the flat panel detector.

  13. Detection of Campylobacter jejuni and Campylobacter coli in Environmental Waters by PCR Enzyme-Linked Immunosorbent Assay

    OpenAIRE

    Sails, A. D.; Bolton, F. J.; Fox, A. J.; Wareing, D. R. A.; Greenway, D. L. A.

    2002-01-01

    A PCR enzyme-linked immunosorbent assay (ELISA) assay was applied to the detection of Campylobacter jejuni and Campylobacter coli in environmental water samples after enrichment culture. Bacterial cells were concentrated from 69 environmental water samples by using filtration, and the filtrates were cultured in Campylobacter blood-free broth. After enrichment culture, DNA was extracted from the samples by using a rapid-boiling method, and the DNA extracts were used as a template in a PCR ELIS...

  14. HistoFlex--a microfluidic device providing uniform flow conditions enabling highly sensitive, reproducible and quantitative in situ hybridizations.

    Science.gov (United States)

    Søe, Martin Jensen; Okkels, Fridolin; Sabourin, David; Alberti, Massimo; Holmstrøm, Kim; Dufva, Martin

    2011-11-21

    A microfluidic device (the HistoFlex) designed to perform and monitor molecular biological assays under dynamic flow conditions on microscope slide-substrates, with special emphasis on analyzing histological tissue sections, is presented. Microscope slides were reversibly sealed onto a cast polydimethylsiloxane (PDMS) insert, patterned with distribution channels and reaction chambers. Topology optimization was used to design reaction chambers with uniform flow conditions. The HistoFlex provided uniform hybridization conditions, across the reaction chamber, as determined by hybridization to microscope slides of spotted DNA microarrays when applying probe concentrations generally used in in situ hybridization (ISH) assays. The HistoFlex's novel ability in online monitoring of an in situ hybridization assay was demonstrated using direct fluorescent detection of hybridization to 18S rRNA. Tissue sections were not visually damaged during assaying, which enabled adapting a complete ISH assay for detection of microRNAs (miRNA). The effects of flow based incubations on hybridization, antibody incubation and Tyramide Signal Amplification (TSA) steps were investigated upon adapting the ISH assay for performing in the HistoFlex. The hybridization step was significantly enhanced using flow based incubations due to improved hybridization efficiency. The HistoFlex device enabled a fast miRNA ISH assay (3 hours) which provided higher hybridization signal intensity compared to using conventional techniques (5 h 40 min). We further demonstrate that the improved hybridization efficiency using the HistoFlex permits more complex assays e.g. those comprising sequential hybridization and detection of two miRNAs to be performed with significantly increased sensitivity. The HistoFlex provides a new histological analysis platform that will allow multiple and sequential assays to be performed under their individual optimum assay conditions. Images can subsequently be recorded either in

  15. Continuity Controlled Hybrid Automata

    NARCIS (Netherlands)

    Bergstra, J.A.; Middelburg, C.A.

    2004-01-01

    We investigate the connections between the process algebra for hybrid systems of Bergstra and Middelburg and the formalism of hybrid automata of Henzinger et al. We give interpretations of hybrid automata in the process algebra for hybrid systems and compare them with the standard interpretation of

  16. Continuity controlled Hybrid Automata

    NARCIS (Netherlands)

    Bergstra, J.A.; Middelburg, C.A.

    2008-01-01

    We investigate the connections between the process algebra for hybrid systems of Bergstra and Middelburg and the formalism of hybrid automata of Henzinger et al. We give interpretations of hybrid automata in the process algebra for hybrid systems and compare them with the standard interpretation of

  17. The validation and clinical implementation of BRCAplus: a comprehensive high-risk breast cancer diagnostic assay.

    Directory of Open Access Journals (Sweden)

    Hansook Kim Chong

    Full Text Available Breast cancer is the most commonly diagnosed cancer in women, with 10% of disease attributed to hereditary factors. Although BRCA1 and BRCA2 account for a high percentage of hereditary cases, there are more than 25 susceptibility genes that differentially impact the risk for breast cancer. Traditionally, germline testing for breast cancer was performed by Sanger dideoxy terminator sequencing in a reflexive manner, beginning with BRCA1 and BRCA2. The introduction of next-generation sequencing (NGS has enabled the simultaneous testing of all genes implicated in breast cancer resulting in diagnostic labs offering large, comprehensive gene panels. However, some physicians prefer to only test for those genes in which established surveillance and treatment protocol exists. The NGS based BRCAplus test utilizes a custom tiled PCR based target enrichment design and bioinformatics pipeline coupled with array comparative genomic hybridization (aCGH to identify mutations in the six high-risk genes: BRCA1, BRCA2, PTEN, TP53, CDH1, and STK11. Validation of the assay with 250 previously characterized samples resulted in 100% detection of 3,025 known variants and analytical specificity of 99.99%. Analysis of the clinical performance of the first 3,000 BRCAplus samples referred for testing revealed an average coverage greater than 9,000X per target base pair resulting in excellent specificity and the sensitivity to detect low level mosaicism and allele-drop out. The unique design of the assay enabled the detection of pathogenic mutations missed by previous testing. With the abundance of NGS diagnostic tests being released, it is essential that clinicians understand the advantages and limitations of different test designs.

  18. In vitro antioxidant activities of antioxidant-enriched toothpastes.

    Science.gov (United States)

    Battino, M; Ferreiro, M S; Armeni, T; Politi, A; Bompadre, S; Massoli, A; Bullon, P

    2005-03-01

    Several forms of periodontal diseases (PD) are often associated with modified phagocytosing leukocytes and contemporary free radical production. Host antioxidant defenses could benefit from toothpastes used as adjuncts to counteract plaque-associated bacteria. The aim of the present study was to determine possible antioxidant activity (AA) of 12 differently antioxidant-enriched toothpastes, regardless of their efficacy as antimicrobial agents. Toothpastes were enriched alternatively with sodium ascorbyl phosphate, alpha-tocopherol acetate, pycnogenol, allantoin and methyl salycilate or a mixture of these. AA was tested in a cell-free system with a ABTS-decolorization assay improved by means of a flow injection analysis device. Comet assay, using NCTC 2544 keratinocytes, was performed to test if it was possible to identify any protection against in vitro DNA fragmentation provoked by a challenge with H(2)O(2) in cultures pre-incubated with toothpaste extracts. Only toothpastes containing sodium ascorbyl phosphate displayed clear AA with I(50) values ranging between 50 and 80 mg of toothpaste/ml water. COMET analysis of cells challenged with H(2)O(2) in presence of toothpaste extracts revealed a limited protection exerted by sodium ascorbyl phosphate. The results described herein indicate that toothpastes containing sodium ascorbyl phosphate possess AA. All the data were obtained in systems in vitro and the demonstration of in vivo AA is desirable. These findings could be useful in the treatment and maintenance of some forms of PD and should be considered when arranging new toothpaste formulations.

  19. The corneal pocket assay.

    Science.gov (United States)

    Ziche, Marina; Morbidelli, Lucia

    2015-01-01

    The cornea in most species is physiologically avascular, and thus this assay allows the measurement of newly formed vessels. The continuous monitoring of neovascular growth in the same animal allows the evaluation of drugs acting as suppressors or stimulators of angiogenesis. Under anesthesia a micropocket is produced in the cornea thickness and the angiogenesis stimulus (tumor tissue, cell suspension, growth factor) is placed into the pocket in order to induce vascular outgrowth from the limbal capillaries. Neovascular development and progression can be modified by the presence of locally released or applied inhibitory factors or by systemic treatments. In this chapter the experimental details of the avascular cornea assay, the technical challenges, and advantages and disadvantages in different species are discussed. Protocols for local drug treatment and tissue sampling for histology and pharmacokinetic profile are reported.

  20. Kinetic Tetrazolium Microtiter Assay

    Science.gov (United States)

    Pierson, Duane L.; Stowe, Raymond; Koenig, David

    1993-01-01

    Kinetic tetrazolium microtiter assay (KTMA) involves use of tetrazolium salts and Triton X-100 (or equivalent), nontoxic, in vitro color developer solubilizing colored metabolite formazan without injuring or killing metabolizing cells. Provides for continuous measurement of metabolism and makes possible to determine rate of action of antimicrobial agent in real time as well as determines effective inhibitory concentrations. Used to monitor growth after addition of stimulatory compounds. Provides for kinetic determination of efficacy of biocide, greatly increasing reliability and precision of results. Also used to determine relative effectiveness of antimicrobial agent as function of time. Capability of generating results on day of test extremely important in treatment of water and waste, disinfection of hospital rooms, and in pharmaceutical, agricultural, and food-processing industries. Assay also used in many aspects of cell biology.

  1. Hybride textuelle Strukturen und hybride textuelle Einheiten. Ein Beitrag zur Theorie der Wörterbuchform

    Directory of Open Access Journals (Sweden)

    Herbert Ernst Wiegand

    2011-10-01

    hierarchical structures are element-heterogeneous whilst the structurecarrying set of all hierarchical pure structures are element-homogeneous. In the structural diagrams of hierarchical hybrid article structures, the nodes for those text segments that establish the hybrid status of the structures are connected with the nodes for the text constituents either by means of arrows for the text-architectonic upward/downward relations, so that the structural graphs are architectonically enriched, or by means of specially marked edges that connect the nodes for the non-functional text segments and those for the internally-expanded functional item additions with the nodes for the text constituents. To each type of hierarchical pure article structure belong various types of hybrid article structures. The same applies to hierarchical pure item structures. Only a selection from the typology of hybrid article and item structures are discussed as well as a small selection of hybrid textual units that display determining features of two text segment classes (cf. the keywords.

    Keywords: ELEMENT-HETEROGENEOUS STRUCTURE-CARRYING SET, FUNCTIONAL ITEM ADDITION, FUNCTIONAL-POSITIONAL SEGMENTATION, HIERARCHICAL ARCHITECTONICALLY ENRICHED ARTICLE MICROSTRUCTURE, HIERARCHICAL HYBRID ARTICLE MICROSTRUCTURE, HIERARCHICAL HYBRID DEEP DOUBLE GLOSS-CONDITIONED ITEM STRUCTURE, HIERARCHICAL HYBRID EXHAUSTIVE ITEMS STRUCTURE, HIERARCHICAL HYBRID GLOSS-CONDITIONED ITEM STRUCTURE, HIERARCHICAL HYBRID ITEMS CONSTITUENT STRUCTURE WITH GLOSS-CONDITIONED PARTIAL STRUCTURE, HIERARCHICAL HYBRID ITEMS STRUCTURE, HIERARCHICAL HYBRID MINIMISED GLOSSCONDITIONED ITEM STRUCTURE, HIERARCHICAL HYBRID SHALLOW DOUBLE GLOSSCONDITIONED ITEM STRUCTURE, HIERARCHICAL HYBRIDARTICLE CONSTITUENT STRUCTURE, HIERARCHICAL HYBRIDTEXT CONSTITUENT STRUCTURE, HIERARCHICAL PURE TEXT CONSTITUENT STRUCTURE, HYBRID CROSS-REFERENCE MARKER, ITEM RELATION, NON-FUNCTIONAL-POSITIONAL SEGMENTATION, ORDERING RELATION, SEGMENTATIVE ISOLATION

  2. Stabilization of concentration fluctuations in mixed membranes by hybrid lipids

    Science.gov (United States)

    Palmieri, Benoit; Safran, Samuel

    2012-02-01

    Finite-size domains have been observed at the surface of cells. These lipids ``rafts'' are stable nanodomains enriched in saturated lipids and cholesterol. While line tension favors macrodomains, one explanation for raft stabilization suggests that the membrane composition is tuned close to a spinodal temperature. From this point of view, rafts are long-lived concentration fluctuations in the mixed phase. We propose a ternary mixture model for the cell membrane that includes hybrid lipids which have one saturated and one unsaturated hydrocarbon chain. Finite amount of hybrid lipids reduces the packing incompatibility at the saturated/unsaturated lipid interface and stabilizes the concentration fluctuations. Hybrid-Hybrid interactions are included in the model and further increase the life-time of the rafts and decrease their length-scales. Moreover, the hybrid has extra orientational degrees of freedom that may lead to modulated phases.

  3. B cell helper assays.

    Science.gov (United States)

    Abrignani, Sergio; Tonti, Elena; Casorati, Giulia; Dellabona, Paolo

    2009-01-01

    Activation, proliferation and differentiation of naïve B lymphocytes into memory B cells and plasma cells requires engagement of the B cell receptor (BCR) coupled to T-cell help (1, 2). T cells deliver help in cognate fashion when they are activated upon recognition of specific MHC-peptide complexes presented by B cells. T cells can also deliver help in a non-cognate or bystander fashion, when they do not find specific MHC-peptide complexes on B cells and are activated by alternative mechanisms. T-cell dependent activation of B cells can be studied in vitro by experimental models called "B cell helper assays" that are based on the co-culture of B cells with activated T cells. These assays allow to decipher the molecular bases for productive T-dependent B cell responses. We show here examples of B cell helper assays in vitro, which can be reproduced with any subset of T lymphocytes that displays the appropriate helper signals.

  4. The characteristics of the spindle fiber attachments(SFAs) enriched fraction from mouse nuclei

    Institute of Scientific and Technical Information of China (English)

    NIZUMEI; NINGYI; 等

    1993-01-01

    The characteristics of the particulate mouse centromere enriched fraction from isolated nuclei obtained in our laboratory were investigated by indirect immunofluorescence,test of the activity of microtubule organizing center(MTOC),SDS-PAGE,and fluorescence in situ hybridization.Most of the particles of the fraction are complexes of DNA and kinetochore proteins and show MTOC activity.The DNA isolated from the fraction can hybridize with DNA in the regions of the primary constrictions of all chromosomes of ascions of the primary constrictions of all chromosomes of ascites cells.The kinetochore proteins isolated from the fraction are mainly those with molecular weight of 55 KD and 59 KD.Results suggested that the fraction obtained is a centromere enriched nuclear fraction as indicated in our previous report.

  5. COMPARISON OF SAMPLE PREPARATION METHODS FOR CHIP-CHIP ASSAYS

    OpenAIRE

    O'Geen, Henriette; Nicolet, Charles M.; Blahnik, Kim; Green, Roland; Farnham, Peggy J.

    2006-01-01

    A single ChIP sample does not provide enough DNA for hybridization to a genomic tiling array. A commonly used technique for amplifying the DNA obtained from ChIP assays is linker-mediated PCR (LMPCR). However, using this amplification method, we could not identify Oct4 binding sites on genomic tiling arrays representing 1% of the human genome (ENCODE arrays). In contrast, hybridization of a pool of 10 ChIP samples to the arrays produced reproducible binding patterns and low background signals...

  6. Reversal of Cocaine Addiction by Environmental Enrichment

    National Research Council Canada - National Science Library

    Marcello Solinas; Claudia Chauvet; Nathalie Thiriet; Rana El Rawas; Mohamed Jaber

    2008-01-01

    .... On the other hand, positive conditions, such as environmental enrichment, can reduce the reinforcing effects of psychostimulants and may provide protection against the development of drug addiction...

  7. Environmental enrichment in farm, zoo, companion and experimental animals

    Directory of Open Access Journals (Sweden)

    Vučinić Marijana

    2009-01-01

    Full Text Available The paper deals with environmental enrichment for domestic animals at farms, animals in zoos, experimental animals and pet animals. Also, the paper defines and describes different strategies of environmental enrichment. Environmental enrichment is a simple and effective mean of prevention of boredom, behavioral disorders as well as an effective mean of improving animal welfare in farm, zoo, companion and experimental animals. Different items and materials may be used for environmental enrichment. They need to be evaluated for use by taking into account the following: the species of an animal, its needs, habits and capabilities, the type of an enrichment device, the device's ability to stimulate the animal's interest and the safety of the device. Enrichment programmes should always include two forms of enrichment: behavioral enrichment and environmental enrichment. Enrichment comes in many forms such as structural or physical enrichment, sensory enrichment (auditory and olfactory stimulation, dietary enrichment, manipulatable enrichment and social enrichment.

  8. Filter characteristics influencing circulating tumor cell enrichment from whole blood.

    Directory of Open Access Journals (Sweden)

    Frank A W Coumans

    Full Text Available A variety of filters assays have been described to enrich circulating tumor cells (CTC based on differences in physical characteristics of blood cells and CTC. In this study we evaluate different filter types to derive the properties of the ideal filter for CTC enrichment. Between 0.1 and 10 mL of whole blood spiked with cells from tumor cell lines were passed through silicon nitride microsieves, polymer track-etched filters and metal TEM grids with various pore sizes. The recovery and size of 9 different culture cell lines was determined and compared to the size of EpCAM+CK+CD45-DNA+ CTC from patients with metastatic breast, colorectal and prostate cancer. The 8 µm track-etched filter and the 5 µm microsieve had the best performance on MDA-231, PC3-9 and SKBR-3 cells, enriching >80% of cells from whole blood. TEM grids had poor recovery of ∼25%. Median diameter of cell lines ranged from 10.9-19.0 µm, compared to 13.1, 10.7, and 11.0 µm for breast, prostate and colorectal CTC, respectively. The 11.4 µm COLO-320 cell line had the lowest recovery of 17%. The ideal filter for CTC enrichment is constructed of a stiff, flat material, is inert to blood cells, has at least 100,000 regularly spaced 5 µm pores for 1 ml of blood with a ≤10% porosity. While cell size is an important factor in determining recovery, other factors must be involved as well. To evaluate a filtration procedure, cell lines with a median size of 11-13 µm should be used to challenge the system.

  9. Polymerase chain reaction assay for avian polyomavirus.

    Science.gov (United States)

    Phalen, D N; Wilson, V G; Graham, D L

    1991-05-01

    A polymerase chain reaction assay was developed for detection of budgerigar fledgling disease virus (BFDV). The assay used a single set of primers complementary to sequences located in the putative coding region for the BFDV VP1 gene. The observed amplification product had the expected size of 550 bp and was confirmed to derive from BFDV DNA by its restriction digestion pattern. This assay was specific for BFDV and highly sensitive, being able to detect as few as 20 copies of the virus. By using the polymerase chain reaction, BFDV was detected in adult, nestling, and embryo budgerigar (Melopsitticus undulatus) tissue DNAs and in sera from adult and nestling budgerigars. These results suggest the possibility of persistent infections in adult birds and lend further support to previously described evidence of possible in ovo transmission. BFDV was also detected in chicken embryo fibroblast cell cultures and chicken eggs inoculated with the virus. A 550-bp product with identical restriction enzyme sites was amplified from a suspected polyomavirus isolated from a peach-faced lovebird (Agapornis pesonata) and from tissue DNA from a Hahn's macaw (Ara nobilis) and a sun conure (Aratinga solstitialis) with histological lesions suggestive of polyomavirus infection. These fragments also hybridized with a BFDV-derived probe, proving that they were derived from a polyomavirus very similar, if not identical, to BFDV.

  10. Quinone-Enriched Gold Nanoparticles in Bioelectrochemistry and Charge Storage

    DEFF Research Database (Denmark)

    Wagner, Michal; Qvortrup, Katrine; Tanner, David Ackland

    We are aiming at the development of a general electrochemical platform suitable for multifarious investigations of biogenic materials. The new platform can be attained by grafting organic/metal hybrids with quinone functionality. In-house made and customized anthraquinone derivatives (with linking...... groups ranging from thiophenes to thiols) were chosen as model compounds for linking together bulk quantities of gold nanoparticles and providing desired functionality. Anthraquinone thiophene and thiol derivatives were synthetized via Sonogashira coupling-type reactions. The synthetic pathways...... for merging gold nanoparticles with resultant anthraquinones include one-pot microwave assisted synthesis or after-mixing of separately prepared gold nanoparticles with selected compounds. The quinone-enriched gold nanoparticles can be transferred onto different electrode surfaces, thus enabling facile...

  11. Nondestructive Assay Options for Spent Fuel Encapsulation

    Energy Technology Data Exchange (ETDEWEB)

    Tobin, Stephen J. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Jansson, Peter [Uppsala Univ. (Sweden)

    2014-10-02

    This report describes the role that nondestructive assay (NDA) techniques and systems of NDA techniques may have in the context of an encapsulation and deep geological repository. The potential NDA needs of an encapsulation and repository facility include safeguards, heat content, and criticality. Some discussion of the facility needs is given, with the majority of the report concentrating on the capability and characteristics of individual NDA instruments and techniques currently available or under development. Particular emphasis is given to how the NDA techniques can be used to determine the heat production of an assembly, as well as meet the dual safeguards needs of 1) determining the declared parameters of initial enrichment, burn-up, and cooling time and 2) detecting defects (total, partial, and bias). The report concludes with the recommendation of three integrated systems that might meet the combined NDA needs of the encapsulation/repository facility.

  12. Application of a double-enrichment procedure for microsatellite isolation and the use of tailed primers for high throughput genotyping

    Directory of Open Access Journals (Sweden)

    Fábio Mendonça Diniz

    2007-03-01

    Full Text Available The number of microsatellite loci and their allelic diversity contribute to increase accuracy and informativity of genetic estimates, however, the isolation of microsatellite loci is not only laborious but also quite expensive. We used (GATAn and (GACAn tetranucleotide probes and single- and double-enrichment hybridization to construct and screen a genomic library with an increased proportion of DNA fragments containing repeat motifs. Repeats were found using both types of hybridization but the double-enrichment procedure recovered sequences of which 100% contained (GATAn and (GACAn motifs. Microsatellite loci primers were then designed with an M13R-tail or CAG-tag to produce scorable PCR products with minimal stutter. The approach used in this study suggests that double-enrichment is a worthwhile strategy when isolating repeat motifs from eukaryotic genomes. Moreover, the use of tailed microsatellite primers provides increased resolution for compound microsatellite loci, with a significant decrease in costs.

  13. Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay.

    Science.gov (United States)

    Black, W C; Gorrochotegui-Escalante, N; Duteau, N M

    2006-03-01

    Most single nucleotide polymorphism (SNP) detection requires expensive equipment and reagents. The oligonucleotide ligation assay (OLA) is an inexpensive SNP assay that detects ligation between a biotinylated "allele-specific detector" and a 3' fluorescein-labeled "reporter" oligonucleotide. No ligation occurs unless the 3' detector nucleotide is complementary to the SNP nucleotide. The original OLA used chemical denaturation and neutralization. Heated OLA (HOLA) instead uses a thermal stable ligase and cycles of denaturing and hybridization for ligation and SNP detection. The cost per genotype is approximately US$1.25 with two-allele SNPs or approximately US$1.75 with three-allele SNPs. We illustrate the development of HOLA for SNP detection in the Early Trypsin and Abundant Trypsin loci in the mosquito Aedes aegypti (L.) and at the a-glycerophosphate dehydrogenase locus in the mosquito Anopheles gambiae s.s.

  14. Growth cone collapse assay.

    Science.gov (United States)

    Cook, Geoffrey M W; Jareonsettasin, Prem; Keynes, Roger J

    2014-01-01

    The growth cone collapse assay has proved invaluable in detecting and purifying axonal repellents. Glycoproteins/proteins present in detergent extracts of biological tissues are incorporated into liposomes, added to growth cones in culture and changes in morphology are then assessed. Alternatively purified or recombinant molecules in aqueous solution may be added directly to the cultures. In both cases after a defined period of time (up to 1 h), the cultures are fixed and then assessed by inverted phase contrast microscopy for the percentage of growth cones showing a collapsed profile with loss of flattened morphology, filopodia, and lamellipodia.

  15. FLUIDICS DEVICE FOR ASSAY

    DEFF Research Database (Denmark)

    2007-01-01

    The present invention relates to a device for use in performing assays on standard laboratory solid supports whereon chemical entities are attached. The invention furthermore relates to the use of such a device and a kit comprising such a device. The device according to the present invention is a......, when operatively connected, one or more chambers (21) comprising the chemical entities (41), the inlet(s) (5) and outlet(s) (6) and chambers (21) being in fluid connection. The device further comprise means for providing differing chemical conditions in each chamber (21)....

  16. Radon assay for SNO+

    Energy Technology Data Exchange (ETDEWEB)

    Rumleskie, Janet [Laurentian University, Greater Sudbury, Ontario (Canada)

    2015-12-31

    The SNO+ experiment will study neutrinos while located 6,800 feet below the surface of the earth at SNOLAB. Though shielded from surface backgrounds, emanation of radon radioisotopes from the surrounding rock leads to back-grounds. The characteristic decay of radon and its daughters allows for an alpha detection technique to count the amount of Rn-222 atoms collected. Traps can collect Rn-222 from various positions and materials, including an assay skid that will collect Rn-222 from the organic liquid scintillator used to detect interactions within SNO+.

  17. Oleuropein-Enriched Olive Leaf Extract Affects Calcium Dynamics and Impairs Viability of Malignant Mesothelioma Cells

    Directory of Open Access Journals (Sweden)

    Carla Marchetti

    2015-01-01

    Full Text Available Malignant mesothelioma is a poor prognosis cancer in urgent need of alternative therapies. Oleuropein, the major phenolic of olive tree (Olea europaea L., is believed to have therapeutic potentials for various diseases, including tumors. We obtained an oleuropein-enriched fraction, consisting of 60% w/w oleuropein, from olive leaves, and assessed its effects on intracellular Ca2+ and cell viability in mesothelioma cells. Effects of the oleuropein-enriched fraction on Ca2+ dynamics and cell viability were studied in the REN mesothelioma cell line, using fura-2 microspectrofluorimetry and MTT assay, respectively. Fura-2-loaded cells, transiently exposed to the oleuropein-enriched fraction, showed dose-dependent transient elevations of cytosolic Ca2+ concentration (Ca2+i. Application of standard oleuropein and hydroxytyrosol, and of the inhibitor of low-voltage T-type Ca2+ channels NNC-55-0396, suggested that the effect is mainly due to oleuropein acting through its hydroxytyrosol moiety on T-type Ca2+ channels. The oleuropein-enriched fraction and standard oleuropein displayed a significant antiproliferative effect, as measured on REN cells by MTT cell viability assay, with IC50 of 22 μg/mL oleuropein. Data suggest that our oleuropein-enriched fraction from olive leaf extract could have pharmacological application in malignant mesothelioma anticancer therapy, possibly by targeting T-type Ca2+ channels and thereby dysregulating intracellular Ca2+ dynamics.

  18. Student science enrichment training program

    Energy Technology Data Exchange (ETDEWEB)

    Sandhu, S.S.

    1994-08-01

    This is a report on the Student Science Enrichment Training Program, with special emphasis on chemical and computer science fields. The residential summer session was held at the campus of Claflin College, Orangeburg, SC, for six weeks during 1993 summer, to run concomitantly with the college`s summer school. Fifty participants selected for this program, included high school sophomores, juniors and seniors. The students came from rural South Carolina and adjoining states which, presently, have limited science and computer science facilities. The program focused on high ability minority students, with high potential for science engineering and mathematical careers. The major objective was to increase the pool of well qualified college entering minority students who would elect to go into science, engineering and mathematical careers. The Division of Natural Sciences and Mathematics and engineering at Claflin College received major benefits from this program as it helped them to expand the Departments of Chemistry, Engineering, Mathematics and Computer Science as a result of additional enrollment. It also established an expanded pool of well qualified minority science and mathematics graduates, which were recruited by the federal agencies and private corporations, visiting Claflin College Campus. Department of Energy`s relationship with Claflin College increased the public awareness of energy related job opportunities in the public and private sectors.

  19. Evaluation of a combined macrophyte-epiphyte bioassay for assessing nutrient enrichment in the Portneuf River, Idaho, USA.

    Science.gov (United States)

    Ray, Andrew M; Mebane, Christopher A; Raben, Flint; Irvine, Kathryn M; Marcarelli, Amy M

    2014-07-01

    We describe and evaluate a laboratory bioassay that uses Lemna minor L. and attached epiphytes to characterize the status of ambient and nutrient-enriched water from the Portneuf River, Idaho. Specifically, we measured morphological (number of fronds, longest surface axis, and root length) and population-level (number of plants and dry mass) responses of L. minor and community-level (ash-free dry mass [AFDM] and chlorophyll a [Chl a]) responses of epiphytes to nutrient enrichment. Overall, measures of macrophyte biomass and abundance increased with increasing concentrations of dissolved phosphorus (P) and responded more predictably to nutrient enrichment than morphological measures. Epiphyte AFDM and Chl a were also greatest in P-enriched water; enrichments of N alone produced no measurable epiphytic response. The epiphyte biomass response did not directly mirror macrophyte biomass responses, illustrating the value of a combined macrophyte-epiphyte assay to more fully evaluate nutrient management strategies. Finally, the most P-enriched waters not only supported greater standing stocks of macrophyte and epiphytes but also had significantly higher water column dissolved oxygen and dissolved organic carbon concentrations and a lower pH. Advantages of this macrophyte-epiphyte bioassay over more traditional single-species assays include the use of a more realistic level of biological organization, a relatively short assay schedule (~10 days), and the inclusion of multiple biological response and water-quality measures.

  20. Evaluation of a combined macrophyte–epiphyte bioassay for assessing nutrient enrichment in the Portneuf River, Idaho, USA

    Science.gov (United States)

    Ray, Andrew M.; Mebane, Christopher A.; Raben, Flint; Irvine, Kathryn M.; Marcarelli, Amy M.

    2014-01-01

    We describe and evaluate a laboratory bioassay that uses Lemna minor L. and attached epiphytes to characterize the status of ambient and nutrient-enriched water from the Portneuf River, Idaho. Specifically, we measured morphological (number of fronds, longest surface axis, and root length) and population-level (number of plants and dry mass) responses of L. minor and community-level (ash-free dry mass [AFDM] and chlorophyll a [Chl a]) responses of epiphytes to nutrient enrichment. Overall, measures of macrophyte biomass and abundance increased with increasing concentrations of dissolved phosphorus (P) and responded more predictably to nutrient enrichment than morphological measures. Epiphyte AFDM and Chl a were also greatest in P-enriched water; enrichments of N alone produced no measurable epiphytic response. The epiphyte biomass response did not directly mirror macrophyte biomass responses, illustrating the value of a combined macrophyte–epiphyte assay to more fully evaluate nutrient management strategies. Finally, the most P-enriched waters not only supported greater standing stocks of macrophyte and epiphytes but also had significantly higher water column dissolved oxygen and dissolved organic carbon concentrations and a lower pH. Advantages of this macrophyte–epiphyte bioassay over more traditional single-species assays include the use of a more realistic level of biological organization, a relatively short assay schedule (~10 days), and the inclusion of multiple biological response and water-quality measures.

  1. Enrichment-ELISA for Detection of Salmonella typhi From Food and Water Samples

    Institute of Scientific and Technical Information of China (English)

    S.KUMAR; K.BALAKRISHNA; HV.BATRA

    2008-01-01

    Objective Development of monoclonal antibody based sandwich enzyme linked immunosorbant assay(sELISA)for rapid detection of Salmonella enterica serovar typhi(S.typhi) from food and water samples and optimization of enrichment procedures for use with the developed sELISA to increase the detection Sensitivity of the assay. Methods Spleen cells from BALB/c mice immunized with flagellin(H=d)antigen of S.typhi were fused with Sp2/0 myeloma cells.The hybridoma cell line specific to H=d antigen was established.characterized and ascites raised against one of these clones.The hypefimmune serum to flagellin antigen was raised in New Zealand White rabbits.An sELISA was developed using polyclonalantibody as capture and monoelonal antibody as detection antibody.To design the efficient culture sUrategies for use with the sELISA.different pre-enrichment and enrichment brothswere evaluated.The mediaincluded buffered peptonewater(BPW)and brain heart infusion broth for pre-enrichment and selenite F broth and Rappaport-Vassiliadis broth as enrichment broths.The developed sELISA with preceding enrichment step in BPW(Enrichment-ELISA)was evaluated in various food samples artificially inoculated with S. typhi bacteria.Various food(30)and water(35)samples collected from field were also tested by Enrichment-ELISA and culture method. Results Out of four specific clones to H=d antigen,one clone(#2/56.IgG2a isotype)was usedin sELISA.The sELISA had the detectionlimit of 104-105 cfu of S.typhi.Of the various broths used with sELISA,BPW was found to yield maximum ELISA values.Enrichment-ELISA,when tested in artificially inoculated food samples,generally,could detect 102S.typhi cfu/mL within 10 h from variousfood rinses(meat,vegetable)and milk samples.After overnight enrichment in BPW,as less as 2 bacteria per 10 mL of milk,meat rinse.and chicken rinse could be detected.Only one of the field samples(water)gave false positive result by Enrichment-ELISA.Conclusion In comparison to culture

  2. Inoculation Stress Hypothesis of Environmental Enrichment

    Science.gov (United States)

    Crofton, Elizabeth J.; Zhang, Yafang; Green, Thomas A.

    2014-01-01

    One hallmark of psychiatric conditions is the vast continuum of individual differences in susceptibility vs. resilience resulting from the interaction of genetic and environmental factors. The environmental enrichment paradigm is an animal model that is useful for studying a range of psychiatric conditions, including protective phenotypes in addiction and depression models. The major question is how environmental enrichment, a non-drug and non-surgical manipulation, can produce such robust individual differences in such a wide range of behaviors. This paper draws from a variety of published sources to outline a coherent hypothesis of inoculation stress as a factor producing the protective enrichment phenotypes. The basic tenet suggests that chronic mild stress from living in a complex environment and interacting non-aggressively with conspecifics can inoculate enriched rats against subsequent stressors and/or drugs of abuse. This paper reviews the enrichment phenotypes, mulls the fundamental nature of environmental enrichment vs. isolation, discusses the most appropriate control for environmental enrichment, and challenges the idea that cortisol/corticosterone equals stress. The intent of the inoculation stress hypothesis of environmental enrichment is to provide a scaffold with which to build testable hypotheses for the elucidation of the molecular mechanisms underlying these protective phenotypes and thus provide new therapeutic targets to treat psychiatric/neurological conditions. PMID:25449533

  3. Enrichment Monitor for 235U Fuel Tubes

    Energy Technology Data Exchange (ETDEWEB)

    Winn, W.G.

    2001-08-22

    This report describes the performance of this prototype y-monitor of 235 Uranium enrichment. In this proposed method y-rates associated with 235U and 232U are correlated with enrichment. Instrumentation for appraising fuel tubes with this method has been assembled and tested.

  4. 21 CFR 137.350 - Enriched rice.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 2 2010-04-01 2010-04-01 false Enriched rice. 137.350 Section 137.350 Food and... Related Products § 137.350 Enriched rice. (a) The foods for which definitions and standards of identity are prescribed by this section are forms of milled rice (except rice coated with talc and glucose...

  5. A Component Analysis of Marriage Enrichment.

    Science.gov (United States)

    Buston, Beverley G.; And Others

    Although marriage enrichment programs have been shown to be effective for many couples, a multidimensional approach to assessment is needed in investigating these groups. The components of information and social support in successful marriage enrichment programs were compared in a completely crossed 2 x 2 factorial design with repeated measures.…

  6. Use of laminar flow patterning for miniaturised biochemical assays

    DEFF Research Database (Denmark)

    Regenberg, Birgitte; Krühne, Ulrich; Beyer, M.

    2004-01-01

    Laminar flow in microfluidic chambers was used to construct low (one dimensional) density arrays suitable for miniaturized biochemical assays. By varying the ratio of flows of two guiding streams flanking a sample stream, precise focusing and positioning of the latter was achieved, and reactive...... species carried in the sample stream were deposited on functionalized chip surfaces as discrete 50 mm wide lanes. Using different model systems we have confirmed the method's suitability for qualitative screening and quantification tasks in receptor-ligand assays, recording biotin......-streptavidin interactions, DNA-hybridization and DNA-triplex formation. The system is simple, fast, reproducible, flexible, and has small sample requirements....

  7. Enriching an effect calculus with linear types

    DEFF Research Database (Denmark)

    Egger, Jeff; Møgelberg, Rasmus Ejlers; Simpson, Alex

    2009-01-01

    We define an ``enriched effect calculus'' by conservatively extending  a type theory for computational effects with primitives from linear logic. By doing so, we obtain a generalisation of linear type theory, intended as a formalism for expressing linear aspects of effects. As a worked example, we...... formulate  linearly-used continuations in the enriched effect calculus. These are captured by a fundamental translation of the enriched effect calculus into itself, which extends existing call-by-value and call-by-name linearly-used CPS translations. We show that our translation is involutive. Full...... completeness results for the various linearly-used CPS translations  follow. Our main results, the conservativity of enriching the effect calculus with linear primitives, and the involution property of the fundamental translation, are proved using a category-theoretic semantics for the enriched effect calculus...

  8. RAS - Screens & Assays - Drug Discovery

    Science.gov (United States)

    The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

  9. Solar enriched methane production: Assessment of plant potentialities and applications

    Directory of Open Access Journals (Sweden)

    Vincenzo Piemonte

    2010-04-01

    Full Text Available The crucial environmental issue due to fossil fuel use in our society and industries and more and more perceived by the communities is stimulating the development of innovative technologies with the scope of reducing GHGs and pollutants emissions, improving plants efficiency and exploiting renewable energy sources. The idea proposed in the present work links this context: a novel hybrid plant for the production of a mixture of methane and hydrogen (20%vol, called enriched-methane, from a steam reforming reactor whose heat duty is supplied by a concentrating solar power (CSP plant by means of a molten salt stream is here conceived, modelled and assessed. The enriched-methane mixture can be applied in methane internal combustion engines (ICE reducing CO, CO2, unburned emissions and improving engine efficiency. Moreover, the residual sensible heat of solar-heated molten salt stream can be used to generate medium-pressure steam and to produce electricity by a steam-turbine. Therefore, the plant proposed is co-generative, producing both hydrogen and electricity from a solar source. The behaviour of methane steam reforming reactor is simulated by means of a 2D mathematical model and the design of a cogenerative solar plant is proposed, evaluating its potentialities in terms of MWh of electricity produced and number of vehicles fed by enriched-methane. A single CSP module (surface requirement = 1.5 hectares coupled with a 4-tubes-and-shell shaped reactor is able to produce 686 tons/year of hydrogen, equivalent to 3.430 tons/year of 20%vol H2-CH4 mixture and 3.097 MWh/year of clean electricity.

  10. Detection of Cronobacter sakazakii in powdered infant formula using an immunoliposome-based immunomagnetic concentration and separation assay

    Science.gov (United States)

    Shukla, Shruti; Lee, Gibaek; Song, Xinjie; Park, Jung Hyun; Cho, Hyunjeong; Lee, Eun Ju; Kim, Myunghee

    2016-01-01

    This study aimed to optimize the applicability of an immunoliposome-based immunomagnetic concentration and separation assay to facilitate rapid detection of Cronobacter sakazakii in powdered infant formula (PIF). To determine the detection limit, specificity, and pre-enrichment incubation time (0, 4, 6, and 8 h), assay tests were performed with different cell numbers of C. sakazakii (2 × 100 and 2 × 101 CFU/ml) inoculated in 10 g of PIF. The assay was able to detect as few as 2 cells of C. sakazakii/10 g of PIF sample after 6 h of pre-enrichment incubation with an assay time of 2 h 30 min. The assay was assessed for cross-reactivity with other bacterial strains and exhibited strong specificity to C. sakazakii. Moreover, the assay method was applied to the detection of C. sakazakii in PIF without pre-enrichment steps, and the results were compared with INC-ELISA and RT-PCR. The developed method was able to detect C. sakazakii in spiked PIF without pre-enrichment, whereas INC-ELISA failed to detect C. sakazakii. In addition, when compared with the results obtained with RT-PCR, our developed assay required lesser detection time. The developed assay was also not susceptible to any effect of the food matrix or background contaminant microflora. PMID:27721500

  11. A combined enrichment and aptamer pulldown assay for francisella tularensis detection in food and environmental matrices

    NARCIS (Netherlands)

    Lamont, Elise A.; Wang, Ping; Enomoto, Shinichiro; Borewicz, Klaudyna; Abdallah, Ahmed; Isaacson, Richard E.; Sreevatsan, Srinand; Kourentzi, Katerina

    2014-01-01

    Francisella tularensis, a Gram-negative bacterium and causative agent of tularemia, is categorized as a Class A select agent by the Centers for Disease Control and Prevention due to its ease of dissemination and ability to cause disease. Oropharyngeal and gastrointestinal tularemia may occur due

  12. Deuterium enrichment of interstellar dusts

    Science.gov (United States)

    Das, Ankan; Chakrabarti, Sandip Kumar; Majumdar, Liton; Sahu, Dipen

    2016-07-01

    High abundance of some abundant and simple interstellar species could be explained by considering the chemistry that occurs on interstellar dusts. Because of its simplicity, the rate equation method is widely used to study the surface chemistry. However, because the recombination efficiency for the formation of any surface species is highly dependent on various physical and chemical parameters, the Monte Carlo method is best suited for addressing the randomness of the processes. We carry out Monte-Carlo simulation to study deuterium enrichment of interstellar grain mantle under various physical conditions. Based on the physical properties, various types of clouds are considered. We find that in diffuse cloud regions, very strong radiation fields persists and hardly a few layers of surface species are formed. In translucent cloud regions with a moderate radiation field, significant number of layers would be produced and surface coverage is mainly dominated by photo-dissociation products such as, C, CH_3, CH_2D, OH and OD. In the intermediate dense cloud regions (having number density of total hydrogen nuclei in all forms ˜2 × 10^4 cm^{-3}), water and methanol along with their deuterated derivatives are efficiently formed. For much higher density regions (˜10^6 cm^{-3}), water and methanol productions are suppressed but surface coverage of CO, CO_2, O_2, O_3 are dramatically increased. We find a very high degree of fractionation of water and methanol. Observational results support a high fractionation of methanol but surprisingly water fractionation is found to be low. This is in contradiction with our model results indicating alternative routes for de-fractionation of water.

  13. Solution-based targeted genomic enrichment for precious DNA samples

    Directory of Open Access Journals (Sweden)

    Shearer Aiden

    2012-05-01

    Full Text Available Abstract Background Solution-based targeted genomic enrichment (TGE protocols permit selective sequencing of genomic regions of interest on a massively parallel scale. These protocols could be improved by: 1 modifying or eliminating time consuming steps; 2 increasing yield to reduce input DNA and excessive PCR cycling; and 3 enhancing reproducible. Results We developed a solution-based TGE method for downstream Illumina sequencing in a non-automated workflow, adding standard Illumina barcode indexes during the post-hybridization amplification to allow for sample pooling prior to sequencing. The method utilizes Agilent SureSelect baits, primers and hybridization reagents for the capture, off-the-shelf reagents for the library preparation steps, and adaptor oligonucleotides for Illumina paired-end sequencing purchased directly from an oligonucleotide manufacturing company. Conclusions This solution-based TGE method for Illumina sequencing is optimized for small- or medium-sized laboratories and addresses the weaknesses of standard protocols by reducing the amount of input DNA required, increasing capture yield, optimizing efficiency, and improving reproducibility.

  14. Bacterial assays for recombinagens.

    Science.gov (United States)

    Hoffmann, G R

    1992-12-01

    Two principal strategies have been used for studying recombinagenic effects of chemicals and radiation in bacteria: (1) measurement of homologous recombination involving defined alleles in a partially diploid strain, and (2) measurement of the formation and loss of genetic duplications in the bacterial chromosome. In the former category, most methods involve one allele in the bacterial chromosome and another in a plasmid, but it is also possible to detect recombination between two chromosomal alleles or between two extrachromosomal alleles. This review summarizes methods that use each of these approaches for detecting recombination and tabulates data on agents that have been found to be recombinagenic in bacteria. The assays are discussed with respect to their effectiveness in testing for recombinagens and their potential for elucidating mechanisms underlying recombinagenic effects.

  15. From hybrid swarms to swarms of hybrids

    Science.gov (United States)

    The introgression of modern humans (Homo sapiens) with Neanderthals 40,000 YBP after a half-million years of separation, may have led to the best example of a hybrid swarm on earth. Modern trade and transportation in support of the human hybrids has continued to introduce additional species, genotyp...

  16. The Hybrid Museum: Hybrid Economies of Meaning

    DEFF Research Database (Denmark)

    Vestergaard, Vitus

    2013-01-01

    this article shows that there are two different museum mindsets where the second mindset leans towards participatory practices. It is shown how a museum can support a hybrid economy of meaning that builds on both a user generated economy of meaning and an institutional economy of meaning and adds value to both....... Such a museum is referred to as a hybrid museum....

  17. Hydraulic Hybrid Vehicles

    Science.gov (United States)

    EPA and the United Parcel Service (UPS) have developed a hydraulic hybrid delivery vehicle to explore and demonstrate the environmental benefits of the hydraulic hybrid for urban pick-up and delivery fleets.

  18. Hybrid Management in Hospitals

    DEFF Research Database (Denmark)

    Byrkjeflot, Haldor; Jespersen, Peter Kragh

    2010-01-01

    Artiklen indeholder et litteraturbaseret studium af ledelsesformer i sygehuse, hvor sundhedsfaglig ledelse og generel ledelse mikses til hybride ledelsesformer......Artiklen indeholder et litteraturbaseret studium af ledelsesformer i sygehuse, hvor sundhedsfaglig ledelse og generel ledelse mikses til hybride ledelsesformer...

  19. PCR-反向点杂交基因分型与实时荧光定量PCR检测人乳头瘤病毒的研究%Use of a PCR-based reverse blot hybridization assay for subtyping and real-time quantitative PCR to detect human papilloma virus

    Institute of Scientific and Technical Information of China (English)

    向华国; 曾锦婷; 何婉意; 黎国

    2012-01-01

    Objective To evaluate the significance of a PCR-based reverse blot hybridization (PCR-RDB) assay and realtime quantitative PCR for detecting human papilloma virus in female outpatients. Methods A total of 121 female outpatients were checked for 23 HFV DNA types by PCR-RDB and 13 high-risk HPV genotypes by real-time quantitative PCR. Results According to PCR-RDB, 28.10% of the women(34/121) tested positive while 16. 53%(20/121) tested positive according to real-time quantitative PCR. HPV was detected more often with PCR-RDB than with real-time quantitative PCR (P<0.05). The concordance rate for the two techniques was 93. 39%(113/121). Conclusion PCR-RDB can be used to screen for HPV infection while real-time quantitative PCR facilitates evaluation of the effectiveness of treatment and the prognosis for cervical carcinoma. Combining the two should increase the specificity and sensitivity of HPV detection.%目的 评价PCR-反向点杂交基因分型与实时荧光定量PCR在检测人乳头瘤病毒(HPV)的意义.方法 同时采用PCR-反向点杂交基因分型和实时荧光定量PCR对121例女性官颈脱离细胞标本进行HPV检测.其中PCR-反向点杂交基因分型能检测23种HPV亚型,实时荧光定量PCR定量检测常见的13种高危HPV亚型.结果 PCR-反向点杂交基因分型检测HPV的阳性率为28.10%(34/121),实时荧光定量PCR检测HPV的阳性率为16.53%(20/121),差异有统计学意义(P<0.05);二者检测的符合率为93.39%(113/121).结论 PCR-反向杂交基因分型适用于HPV感染的筛查,而实时荧光定量PCR适用于HPV感染相关疾病的疗效与预后的判断.PCR-反向杂交基因分型与实时荧光定量PCR联合检测可提高HPV检测的特异性和敏感度,对于生殖道HPV感染以及子宫颈癌的早期发现、预防和治疗具有重要意义.

  20. Hybrid male sterility is caused by mitochondrial DNA deletion.

    Science.gov (United States)

    Hayashida, Kenji; Kohno, Shigeru

    2009-07-01

    Although it is known that the hybrid male mouse is sterile just like any other animal's heterogametic sex, the reason why only the male germ cells are impaired has yet to be discovered. TdT-mediated dUTP nick end labeling assay using a confocal fluorescence microscope and DNA fragmentation assay of hybrid testis indicated destruction of the mitochondrial DNA (mtDNA) rather than the nuclear DNA. Previously we reported that maternal mtDNA inheritance is through selective sperm mtDNA elimination based on the sperm factor and two egg factors, and expression of these three factors was recognized in the hybrid testis. It was thereby assumed that mtDNA destruction caused by the expression of maternal mtDNA inheritance system in male germ cells is implicated in the hybrid male sterility of mice.

  1. Preparation and characterization of a laboratory scale selenomethionine-enriched bread. Selenium bioaccessibility.

    Science.gov (United States)

    Sánchez-Martínez, María; Pérez-Corona, Teresa; Caímara, Carmen; Madrid, Yolanda

    2015-01-14

    This study focuses on the preparation at lab scale of selenomethionine-enriched white and wholemeal bread. Selenium was supplemented either by adding selenite directly to the dough or by using lab-made selenium-enriched yeast. The best results were obtained when using fresh selenium-enriched yeast. The optimum incubation time for selenomethionine-enriched yeast preparation, while keeping formation of selenium byproducts to a minimum, was 96 h. Selenium content measured by isotope dilution analysis (IDA)-ICP-MS in Se-white and Se-wholemeal bread was 1.28 ± 0.02 μg g–1 and 1.16 ± 0.02 μg g–1 (expressed as mean ± SE, 3 replicates), respectively. HPLC postcolumn IDA-ICP-MS measurements revealed that selenomethionine was the main Se species found in Se-enriched bread, which accounted for ca. 80% of total selenium. In vitro gastrointestinal digestion assay provided selenium bioaccessibility values of 100 ± 3% and 40 ± 1% for white and wholemeal Se-enriched bread, respectively, being selenomethionine the main bioaccessible Se species in white bread, while in wholemeal bread this compound was undetectable.

  2. Development of a phenol-enriched olive oil with phenolic compounds from olive cake.

    Science.gov (United States)

    Suárez, Manuel; Romero, Maria-Paz; Motilva, Maria-José

    2010-10-13

    The recent information regarding the healthy properties of virgin olive oil phenols and the interest in increasing the value of byproducts from the oil extraction processs support the standardized development of phenol-enriched olive oil. Accordingly, the aim of this research work was to evaluate strategies for the development of a virgin olive oil enriched with phenolic compounds obtained from olive cake to increase phenolic ingestion without the drawback of a higher calorie intake. For this proposal, different combinations of phenolic extracts were evaluated at a range of concentrations to obtain the best prototype of enriched olive oil. To study the functionality of the phenol enrichments, the total phenolic content and the oxidative stability were determined by the Folin-Ciocalteu and Rancimat tests, respectively. In addition, the phenolic composition and antioxidant capacity (ORAC assay) of the oils were studied. Finally, the stability and potential bioaccesibility of the phenolic fraction of the enriched oils were tested by an in vitro gastrointestinal digestion model. Results of the study showed different strategies to select the best prototype of enriched olive oil, taking into consideration not only their phenolic content but also other important factors such as the feasibility of implementing the preparation process in the food industry.

  3. Lab-on-a-Chip Multiplex Assays.

    Science.gov (United States)

    Peter, Harald; Wienke, Julia; Bier, Frank F

    2017-01-01

    Lab-on-a-chip multiplex assays allow a rapid identification of multiple parameters in an automated manner. Here we describe a lab-based preparation followed by a rapid and fully automated DNA microarray hybridization and readout in less than 10 min using the Fraunhofer in vitro diagnostics (ivD) platform to enable rapid identification of bacterial species and detection of antibiotic resistance. The use of DNA microarrays allows a fast adaptation of new biomarkers enabling the identification of different genes as well as single-nucleotide-polymorphisms (SNPs) within these genes. In this protocol we describe a DNA microarray developed for identification of Staphylococcus aureus and the mecA resistance gene.

  4. Specific detection and confirmation of Campylobacter jejuni by DNA hybridization and PCR.

    Science.gov (United States)

    Ng, L K; Kingombe, C I; Yan, W; Taylor, D E; Hiratsuka, K; Malik, N; Garcia, M M

    1997-11-01

    Conventional detection and confirmation methods for Campylobacter jejuni are lengthy and tedious. A rapid hybridization protocol in which a 1,475-bp chromogen-labelled DNA probe (pDT1720) and Campylobacter strains filtered and grown on 0.22-micron-pore-size hydrophobic grid membrane filters (HGMFs) are used was developed. Among the environmental and clinical isolates of C. jejuni, Campylobacter coli, Campylobacter jejuni subsp. doylei, Campylobacter lari, and Arcobacter nitrofigilis and a panel of 310 unrelated bacterial strains tested, only C. jejuni and C. jejuni subsp. doylei isolates hybridized with the probe under stringent conditions. The specificity of the probe was confirmed when the protocol was applied to spiked skim milk and chicken rinse samples. Based on the nucleotide sequence of pDT1720, a pair of oligonucleotide primers was designed for PCR amplification of DNA from Campylobacter spp. and other food pathogens grown overnight in selective Mueller-Hinton broth with cefoperazone and growth supplements. All C. jejuni strains tested, including DNase-producing strains and C. jejuni subsp. doylei, produced a specific 402-bp amplicon, as confirmed by restriction and Southern blot analysis. The detection range of the assay was as low as 3 CFU per PCR to as high as 10(5) CFU per PCR for pure cultures. Overnight enrichment of chicken rinse samples spiked initially with as little as approximately 10 CFU/ml produced amplicons after the PCR. No amplicon was detected with any of the other bacterial strains tested or from the chicken background microflora. Since C. jejuni is responsible for 99% of Campylobacter contamination in poultry, PCR and HGMF hybridization were performed on naturally contaminated chicken rinse samples, and the results were compared with the results of conventional cultural isolation on Preston agar. All samples confirmed to be culture positive for C. jejuni were also identified by DNA hybridization and PCR amplification, thus confirming that

  5. Resin Catalyst Hybrids

    Institute of Scientific and Technical Information of China (English)

    S. Asaoka

    2005-01-01

    @@ 1Introduction: What are resin catalyst hybrids? There are typically two types of resin catalyst. One is acidic resin which representative is polystyrene sulfonic acid. The other is basic resin which is availed as metal complex support. The objective items of this study on resin catalyst are consisting of pellet hybrid, equilibrium hybrid and function hybrid of acid and base,as shown in Fig. 1[1-5].

  6. Mesoscale hybrid calibration artifact

    Science.gov (United States)

    Tran, Hy D.; Claudet, Andre A.; Oliver, Andrew D.

    2010-09-07

    A mesoscale calibration artifact, also called a hybrid artifact, suitable for hybrid dimensional measurement and the method for make the artifact. The hybrid artifact has structural characteristics that make it suitable for dimensional measurement in both vision-based systems and touch-probe-based systems. The hybrid artifact employs the intersection of bulk-micromachined planes to fabricate edges that are sharp to the nanometer level and intersecting planes with crystal-lattice-defined angles.

  7. Realizing the Hybrid Library.

    Science.gov (United States)

    Pinfield, Stephen; Eaton, Jonathan; Edwards, Catherine; Russell, Rosemary; Wissenburg, Astrid; Wynne, Peter

    1998-01-01

    Outlines five projects currently funded by the United Kingdom's Electronic Libraries Program (eLib): HyLiFe (Hybrid Library of the Future), MALIBU (MAnaging the hybrid Library for the Benefit of Users), HeadLine (Hybrid Electronic Access and Delivery in the Library Networked Environment), ATHENS (authentication scheme), and BUILDER (Birmingham…

  8. Homoploid hybrid expectations

    Science.gov (United States)

    Homoploid hybrid speciation occurs when a stable, fertile, and reproductively isolated lineage results from hybridization between two distinct species without a change in ploidy level. Reproductive isolation between a homoploid hybrid species and its parents is generally attained via chromosomal re...

  9. Hybrid armature projectile

    Science.gov (United States)

    Hawke, Ronald S.; Asay, James R.; Hall, Clint A.; Konrad, Carl H.; Sauve, Gerald L.; Shahinpoor, Mohsen; Susoeff, Allan R.

    1993-01-01

    A projectile for a railgun that uses a hybrid armature and provides a seed block around part of the outer surface of the projectile to seed the hybrid plasma brush. In addition, the hybrid armature is continuously vaporized to replenish plasma in a plasma armature to provide a tandem armature and provides a unique ridge and groove to reduce plasama blowby.

  10. Intraply Hybrid Composite Design

    Science.gov (United States)

    Chamis, C. C.; Sinclair, J. H.

    1986-01-01

    Several theoretical approaches combined in program. Intraply hybrid composites investigated theoretically and experimentally at Lewis Research Center. Theories developed during investigations and corroborated by attendant experiments used to develop computer program identified as INHYD (Intraply Hybrid Composite Design). INHYD includes several composites micromechanics theories, intraply hybrid composite theories, and integrated hygrothermomechanical theory. Equations from theories used by program as appropriate for user's specific applications.

  11. Hybrid quantum information processing

    Energy Technology Data Exchange (ETDEWEB)

    Furusawa, Akira [Department of Applied Physics, School of Engineering, The University of Tokyo (Japan)

    2014-12-04

    I will briefly explain the definition and advantage of hybrid quantum information processing, which is hybridization of qubit and continuous-variable technologies. The final goal would be realization of universal gate sets both for qubit and continuous-variable quantum information processing with the hybrid technologies. For that purpose, qubit teleportation with a continuousvariable teleporter is one of the most important ingredients.

  12. Validation of Folate-Enriched Eggs as a Functional Food for Improving Folate Intake in Consumers

    Directory of Open Access Journals (Sweden)

    Leslie Altic

    2016-11-01

    Full Text Available Functional foods enriched with folate may be beneficial as a means of optimizing folate status in consumers. We recently developed novel eggs enriched with folate through folic acid supplementation of the hen’s feed, but their potential to influence consumer folate status is unknown because the natural folate forms incorporated into the eggs may not necessarily be retained during storage and cooking. This study aimed to determine the stability of natural folates in folate-enriched eggs under typical conditions of storage and cooking. Total folate was determined by microbiological assay following tri-enzyme treatment in folate-enriched eggs and un-enriched (barn and free-range on the day they were laid, after storage (up to 27 days and after using four typical cooking methods (boiling, poaching, frying, scrambling for different durations. On the day of laying, the folate content of enriched eggs was found to be significantly higher than that of un-enriched barn or free-range eggs (mean ± SD; 123.2 ± 12.4 vs. 41.2 ± 2.8 vs. 65.6 ± 18.5 µg/100 g; p < 0.001. Storage at refrigerator and room temperature for periods up to the Best Before date resulted in no significant losses to the folate content of folate-enriched eggs. Furthermore, folate in enriched eggs remained stable when cooked by four typical methods for periods up to the maximum cooking time (e.g., 135 ± 22.5, 133.9 ± 23.0 and 132.5 ± 35.1; p = 0.73, for raw, scrambled for 50 s and scrambled for 2 min, respectively. Thus, natural folates in folate-enriched eggs remain highly stable with little or no losses following storage and cooking. These findings are important because they demonstrate the feasibility of introducing folate-enriched eggs into the diet of consumers as functional foods with enriched folate content. Further studies will confirm their effectiveness in optimizing the biomarker folate status of consumers.

  13. Interactive fluorophore and quencher pairs for labeling fluorescent nucleic acid hybridization probes.

    Science.gov (United States)

    Marras, Salvatore A E

    2008-03-01

    The use of fluorescent nucleic acid hybridization probes that generate a fluorescence signal only when they bind to their target enables real-time monitoring of nucleic acid amplification assays. Real-time nucleic acid amplification assays markedly improves the ability to obtain qualitative and quantitative results. Furthermore, these assays can be carried out in sealed tubes, eliminating carryover contamination. Fluorescent nucleic acid hybridization probes are available in a wide range of different fluorophore and quencher pairs. Multiple hybridization probes, each designed for the detection of a different nucleic acid sequence and each labeled with a differently colored fluorophore, can be added to the same nucleic acid amplification reaction, enabling the development of high-throughput multiplex assays. In order to develop robust, highly sensitive and specific real-time nucleic acid amplification assays it is important to carefully select the fluorophore and quencher labels of hybridization probes. Selection criteria are based on the type of hybridization probe used in the assay, the number of targets to be detected, and the type of apparatus available to perform the assay. This article provides an overview of different aspects of choosing appropriate labels for the different types of fluorescent hybridization probes used with different types of spectrofluorometric thermal cyclers currently available.

  14. A Targeted Enrichment Strategy for Massively Parallel Sequencing of Angiosperm Plastid Genomes

    Directory of Open Access Journals (Sweden)

    Gregory W. Stull

    2013-02-01

    Full Text Available Premise of the study: We explored a targeted enrichment strategy to facilitate rapid and low-cost next-generation sequencing (NGS of numerous complete plastid genomes from across the phylogenetic breadth of angiosperms. Methods and Results: A custom RNA probe set including the complete sequences of 22 previously sequenced eudicot plastomes was designed to facilitate hybridization-based targeted enrichment of eudicot plastid genomes. Using this probe set and an Agilent SureSelect targeted enrichment kit, we conducted an enrichment experiment including 24 angiosperms (22 eudicots, two monocots, which were subsequently sequenced on a single lane of the Illumina GAIIx with single-end, 100-bp reads. This approach yielded nearly complete to complete plastid genomes with exceptionally high coverage (mean coverage: 717×, even for the two monocots. Conclusions: Our enrichment experiment was highly successful even though many aspects of the capture process employed were suboptimal. Hence, significant improvements to this methodology are feasible. With this general approach and probe set, it should be possible to sequence more than 300 essentially complete plastid genomes in a single Illumina GAIIx lane (achieving 50× mean coverage. However, given the complications of pooling numerous samples for multiplex sequencing and the limited number of barcodes (e.g., 96 available in commercial kits, we recommend 96 samples as a current practical maximum for multiplex plastome sequencing. This high-throughput approach should facilitate large-scale plastid genome sequencing at any level of phylogenetic diversity in angiosperms.

  15. Highly sensitive detection of human IgG using a novel bio-barcode assay combined with DNA chip technology

    Science.gov (United States)

    Liu, Zhenbao; Zhou, Bo; Wang, Haiqing; Lu, Feng; Liu, Tianjun; Song, Cunxian; Leng, Xigang

    2013-09-01

    A simple and ultrasensitive detection of human IgG based on signal amplification using a novel bio-barcode assay and DNA chip technology was developed. The sensing platform was a sandwich system made up of antibody-modified magnetic microparticles (Ab-MMPs)/human IgG/Cy3-labeled single-stranded DNA and antibody-modified gold nanoparticles (Cy3-ssDNA-Ab-AuNPs). The MMPs (2.5 μm in diameter) modified with mouse anti-human IgG monoclonal-antibodies could capture human IgG and further be separated and enriched via a magnetic field. The AuNPs (13 nm in diameter) conjugated with goat anti-human IgG polyclonal-antibodies and Cy3-ssDNA could further combine with the human IgG/Ab-MMP complex. The Cy3-ssDNA on AuNPs was then released by TCEP to hybridize with the DNA chip, thus generating a detectable signal by the fluorescence intensity of Cy3. In order to improve detection sensitivity, a three-level cascaded signal amplification was developed: (1) The MMP enrichment as the first-level; (2) Large quantities of Cy3-ssDNA on AuNPs as the second-level; (3) The Cy3-ssDNA conjugate with DNA chip as the third-level. The highly sensitive technique showed an increased response of the fluorescence intensity to the increased concentration of human IgG through a detection range from 1 pg mL-1 to 10 ng mL-1. This sensing technique could not only improve the detection sensitivity for the low concentration of human IgG but also present a robust and efficient signal amplification model. The detection method has good stability, specificity, and reproducibility and could be applied in the detection of human IgG in the real samples.

  16. Highly sensitive detection of human IgG using a novel bio-barcode assay combined with DNA chip technology

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhenbao [Central South University, School of Pharmaceutical Sciences (China); Zhou, Bo, E-mail: zhoubo1771@163.com [The Affiliated Zhongda Hospital of Southeast University, Department of Gerontology (China); Wang, Haiqing; Lu, Feng; Liu, Tianjun; Song, Cunxian; Leng, Xigang, E-mail: lengxigyky@163.com [Institute of Biomedical Engineering, Chinese Academy of Medical Sciences and Peking Union Medical College (China)

    2013-09-15

    A simple and ultrasensitive detection of human IgG based on signal amplification using a novel bio-barcode assay and DNA chip technology was developed. The sensing platform was a sandwich system made up of antibody-modified magnetic microparticles (Ab-MMPs)/human IgG/Cy3-labeled single-stranded DNA and antibody-modified gold nanoparticles (Cy3-ssDNA-Ab-AuNPs). The MMPs (2.5 {mu}m in diameter) modified with mouse anti-human IgG monoclonal-antibodies could capture human IgG and further be separated and enriched via a magnetic field. The AuNPs (13 nm in diameter) conjugated with goat anti-human IgG polyclonal-antibodies and Cy3-ssDNA could further combine with the human IgG/Ab-MMP complex. The Cy3-ssDNA on AuNPs was then released by TCEP to hybridize with the DNA chip, thus generating a detectable signal by the fluorescence intensity of Cy3. In order to improve detection sensitivity, a three-level cascaded signal amplification was developed: (1) The MMP enrichment as the first-level; (2) Large quantities of Cy3-ssDNA on AuNPs as the second-level; (3) The Cy3-ssDNA conjugate with DNA chip as the third-level. The highly sensitive technique showed an increased response of the fluorescence intensity to the increased concentration of human IgG through a detection range from 1 pg mL{sup -1} to 10 ng mL{sup -1}. This sensing technique could not only improve the detection sensitivity for the low concentration of human IgG but also present a robust and efficient signal amplification model. The detection method has good stability, specificity, and reproducibility and could be applied in the detection of human IgG in the real samples.

  17. Concordant testing results between various Human Papillomavirus assays in primary cervical cancer screening

    DEFF Research Database (Denmark)

    de Thurah, Lena; Bonde, Jesper; Hoa Lam, Janni Uyen

    2017-01-01

    OBJECTIVES: Human Papillomavirus (HPV) assays are increasingly used for primary cervical screening and HPV vaccination effect monitoring. We undertook a systematic literature review to determine the concordance in positive test results (i.e., detection of HPV infections) between Hybrid Capture 2...... (HC2) and other assays. METHODS: We searched PubMed, Embase and Scopus for studies of primary screening with HC2 and ≥one more assay, with cross-tabulated testing results for the assays. Two authors applied inclusion criteria and three authors extracted data from included studies. For each inter...

  18. The hydrogen hybrid option

    Energy Technology Data Exchange (ETDEWEB)

    Smith, J.R.

    1993-10-15

    The energy efficiency of various piston engine options for series hybrid automobiles are compared with conventional, battery powered electric, and proton exchange membrane (PEM) fuel cell hybrid automobiles. Gasoline, compressed natural gas (CNG), and hydrogen are considered for these hybrids. The engine and fuel comparisons are done on a basis of equal vehicle weight, drag, and rolling resistance. The relative emissions of these various fueled vehicle options are also presented. It is concluded that a highly optimized, hydrogen fueled, piston engine, series electric hybrid automobile will have efficiency comparable to a similar fuel cell hybrid automobile and will have fewer total emissions than the battery powered vehicle, even without a catalyst.

  19. 77 FR 13367 - General Electric-Hitachi Global Laser Enrichment, LLC, Proposed Laser-Based Uranium Enrichment...

    Science.gov (United States)

    2012-03-06

    ... COMMISSION General Electric-Hitachi Global Laser Enrichment, LLC, Proposed Laser-Based Uranium Enrichment...- Hitachi Global Laser Enrichment, LLC (GLE) Uranium Enrichment Facility. On June 26, 2009, GLE submitted a... uranium enrichment facility (the ``proposed action''). The GLE proposes to locate the facility on...

  20. Chromogenic in situ hybridization: a multicenter study comparing silver in situ hybridization with FISH.

    Science.gov (United States)

    Bartlett, J M S; Campbell, Fiona M; Ibrahim, Merdol; Wencyk, Peter; Ellis, Ian; Kay, Elaine; Connolly, Yvonne; O'Grady, Anthony; Di Palma, Silvana; Starczynski, Jane; Morgan, John M; Jasani, Bharat; Miller, Keith

    2009-10-01

    Our purposes were to perform a robust assessment of a new HER2 chromogenic in situ hybridization test and report on concordance of silver in situ hybridization (SISH) data with fluorescence in situ hybridization (FISH) data and on intraobserver and interlaboratory scoring consistency. HER2 results were scored from 45 breast cancers in 7 laboratories using the Ventana (Tucson, AZ) INFORM HER-2 SISH assay and in 1 central laboratory using a standard FISH assay. Overall, 94.8% of cases were successfully analyzed by SISH across the 6 participating laboratories that reported data. Concordance for diagnosis of HER2 amplification by SISH compared with FISH was high (96.0% overall). Intraobserver variability (8.0%) and intersite variability (12.66%) of absolute HER2/chromosome 17 ratios appear to be tightly controlled across all 6 participating laboratories. The Ventana INFORM HER-2 SISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national guidelines for performance of diagnostic tests.

  1. Herbicide resistance screening assay.

    Science.gov (United States)

    Peterson, Joan M

    2009-01-01

    Herbicide resistance screening is a method that can be used not only to determine presence of the enzyme, phosphinothricin acetyltransferase, encoded by either the Bar or the Pat gene in transgenic maize, but also to assess the inheritance ratio of those genes in a segregating population. Herbicide screening can also be used to study linkage of a transgene of interest that was cotransformed with the herbicide resistance marker gene. By combining the herbicide screen assay with a PCR-based screen of leaf tissue DNA for the presence of both the Bar or the Pat gene marker and a cotransformed transgene of interest from the same seedling tissue and maintaining that seedling identity, the researcher can identify linkage or the possible breakdown in linkage of the marker gene and the transgene of interest. Further, the occurrence of "DNA silencing" can be evaluated if an individual seedling that was susceptible to the applied herbicide nonetheless gave PCR data that indicated presence of the gene responsible for herbicide resistance. Similarly, "DNA silencing" of the gene of interest may be investigated if the seeds can be screened and scored for that phenotypic trait in a nondestructive manner prior to planting.

  2. PROTEIN ENRICHMENT OF SPENT SORGHUM RESIDUE USING ...

    African Journals Online (AJOL)

    BSN

    The yeast isolat~s used were Saccharomyces cerevisiae (dry yeast), Candido. A I isolated from ... Key words: S.cerevisiae, Candida sp., sorghum, protein enrichment, animal feed. .... Identifying bacteria using metabolic characteristics Ln:.

  3. Curriculum enrichment through indigenous Zulu games | Roux |

    African Journals Online (AJOL)

    Curriculum enrichment through indigenous Zulu games. ... Abstract. In the South African context, education has emerged from missionary, colonial and. Afrikaner ... qualitative data (focus group discussions, interviews, video and audio tape

  4. Management's Ecstasy and Disparity Over Job Enrichment

    Science.gov (United States)

    King, Albert S.

    1976-01-01

    A case study analyzing job enrichment schemes and manager expectations of increased productivity is presented. It was found that it was the managers' expectations of increased productivity, not the reorganization of work, that led to higher productivity. (EC)

  5. Hybridization and extinction.

    Science.gov (United States)

    Todesco, Marco; Pascual, Mariana A; Owens, Gregory L; Ostevik, Katherine L; Moyers, Brook T; Hübner, Sariel; Heredia, Sylvia M; Hahn, Min A; Caseys, Celine; Bock, Dan G; Rieseberg, Loren H

    2016-08-01

    Hybridization may drive rare taxa to extinction through genetic swamping, where the rare form is replaced by hybrids, or by demographic swamping, where population growth rates are reduced due to the wasteful production of maladaptive hybrids. Conversely, hybridization may rescue the viability of small, inbred populations. Understanding the factors that contribute to destructive versus constructive outcomes of hybridization is key to managing conservation concerns. Here, we survey the literature for studies of hybridization and extinction to identify the ecological, evolutionary, and genetic factors that critically affect extinction risk through hybridization. We find that while extinction risk is highly situation dependent, genetic swamping is much more frequent than demographic swamping. In addition, human involvement is associated with increased risk and high reproductive isolation with reduced risk. Although climate change is predicted to increase the risk of hybridization-induced extinction, we find little empirical support for this prediction. Similarly, theoretical and experimental studies imply that genetic rescue through hybridization may be equally or more probable than demographic swamping, but our literature survey failed to support this claim. We conclude that halting the introduction of hybridization-prone exotics and restoring mature and diverse habitats that are resistant to hybrid establishment should be management priorities.

  6. Spoof Plasmon Hybridization

    CERN Document Server

    Zhang, Jingjing; Luo, Yu; Shen, Xiaopeng; Maier, Stefan A; Cui, Tie Jun

    2016-01-01

    Plasmon hybridization between closely spaced nanoparticles yields new hybrid modes not found in individual constituents, allowing for the engineering of resonance properties and field enhancement capabilities of metallic nanostructure. Experimental verifications of plasmon hybridization have been thus far mostly limited to optical frequencies, as metals cannot support surface plasmons at longer wavelengths. Here, we introduce the concept of 'spoof plasmon hybridization' in highly conductive metal structures and investigate experimentally the interaction of localized surface plasmon resonances (LSPR) in adjacent metal disks corrugated with subwavelength spiral patterns. We show that the hybridization results in the splitting of spoof plasmon modes into bonding and antibonding resonances analogous to molecular orbital rule and plasmonic hybridization in optical spectrum. These hybrid modes can be manipulated to produce enormous field enhancements (larger than 5000) by tuning the separation between disks or alte...

  7. Extreme Environments Facilitate Hybrid Superiority – The Story of a Successful Daphnia galeata × longispina Hybrid Clone

    Science.gov (United States)

    Griebel, Johanna; Gießler, Sabine; Poxleitner, Monika; Navas Faria, Amanda; Yin, Mingbo; Wolinska, Justyna

    2015-01-01

    Hybridization within the animal kingdom has long been underestimated. Hybrids have often been considered less fit than their parental species. In the present study, we observed that the Daphnia community of a small lake was dominated by a single D. galeata × D. longispina hybrid clone, during two consecutive years. Notably, in artificial community set-ups consisting of several clones representing parental species and other hybrids, this hybrid clone took over within about ten generations. Neither the fitness assay conducted under different temperatures, or under crowded and non-crowded environments, nor the carrying capacity test revealed any outstanding life history parameters of this hybrid clone. However, under simulated winter conditions (i.e. low temperature, food and light), the hybrid clone eventually showed a higher survival probability and higher fecundity compared to parental species. Hybrid superiority in cold-adapted traits leading to an advantage of overwintering as parthenogenetic lineages might consequently explain the establishment of successful hybrids in natural communities of the D. longispina complex. In extreme cases, like the one reported here, a superior hybrid genotype might be the only clone alive after cold winters. Overall, superiority traits, such as enhanced overwintering here, might explain hybrid dominance in nature, especially in extreme and rapidly changing environments. Although any favoured gene complex in cyclic parthenogens could be frozen in successful clones independent of hybridization, we did not find similarly successful clones among parental species. We conclude that the emergence of the observed trait is linked to the production of novel recombined hybrid genotypes. PMID:26448651

  8. Marine Fish Hybridization

    KAUST Repository

    He, Song

    2017-04-01

    Natural hybridization is reproduction (without artificial influence) between two or more species/populations which are distinguishable from each other by heritable characters. Natural hybridizations among marine fishes were highly underappreciated due to limited research effort; it seems that this phenomenon occurs more often than is commonly recognized. As hybridization plays an important role in biodiversity processes in the marine environment, detecting hybridization events and investigating hybridization is important to understand and protect biodiversity. The first chapter sets the framework for this disseration study. The Cohesion Species Concept was selected as the working definition of a species for this study as it can handle marine fish hybridization events. The concept does not require restrictive species boundaries. A general history and background of natural hybridization in marine fishes is reviewed during in chapter as well. Four marine fish hybridization cases were examed and documented in Chapters 2 to 5. In each case study, at least one diagnostic nuclear marker, screened from among ~14 candidate markers, was found to discriminate the putative hybridizing parent species. To further investigate genetic evidence to support the hybrid status for each hybrid offspring in each case, haploweb analysis on diagnostic markers (nuclear and/or mitochondrial) and the DAPC/PCA analysis on microsatellite data were used. By combining the genetic evidences, morphological traits, and ecological observations together, the potential reasons that triggered each hybridization events and the potential genetic/ecology effects could be discussed. In the last chapter, sequences from 82 pairs of hybridizing parents species (for which COI barcoding sequences were available either on GenBank or in our lab) were collected. By comparing the COI fragment p-distance between each hybridizing parent species, some general questions about marine fish hybridization were discussed: Is

  9. Rapid electrochemiluminescence assays of polymerase chain reaction products.

    Science.gov (United States)

    Kenten, J H; Casadei, J; Link, J; Lupold, S; Willey, J; Powell, M; Rees, A; Massey, R

    1991-09-01

    We demonstrate the first use of an electrochemiluminescent (ECL) label, [4-(N-succimidyloxycarbonylpropyl)-4'-methyl-2,2'- bipyridine]ruthenium(II) dihexafluorophosphate (Origen label; IGEN Inc.), in DNA probe assays. This label allows rapid (less than 25 min) quantification and detection of polymerase chain reaction (PCR)-amplified products from oncogenes, viruses, and cloned genes. For the PCR, we used labeled oligonucleotide primers complementary to human papiloma virus and the Ha-ras oncogene. These samples were followed by ECL analysis or hybridization with specific, Origen-labeled oligonucleotide probes. These studies demonstrate the speed, specificity, and effectiveness of the new ECL labels, compared with 32P, for nucleic acid probe applications. We describe formats involving conventional methodologies and a new format that requires no wash step, allowing simple and rapid sample analysis. These rapid assays also reduce PCR contamination, by requiring less sample handling. Improvements in ECL detectability are currently under investigation for use in DNA probe assays without amplification.

  10. A pre-enrichment step is essential for detection of Campylobacter sp. in turbid pond water.

    Science.gov (United States)

    Abulreesh, H H; Paget, T A; Goulder, R

    2014-06-01

    This work aimed to detect Campylobacter species from naturally contaminated turbid pond water by PCR. A total of 16 water samples were collected from a turbid village pond. Four methods of DNA extraction were applied to centrifuge pellets from eight 100 ml pond water samples prior to attempted detection of Campylobacter by PCR without an enrichment step. These methods were (1) Tris-HCl and sodium dodecyl sulfate followed by phenol:chloroform:isoamylalcohol extraction followed by treatment with DNA clean up kit, (2) proteinase K, (3) Chelex® 100, and (4) boiling. The other eight pond water samples (10 ml and 100 ml) were filtered and filters were incubated overnight in Preston enrichment broth. The centrifuge pellets obtained from enrichment cultures were treated by proteinase K for DNA extraction. Primers CF03 and CF04 for the flagellin genes (flaA and flaB) of Campylobacter jejuni and Campylobacter coli were used for amplifying the extracted DNA. The DNA extracted from eight-100 ml pond water samples that were not subject to selective enrichment was never amplified with primers CF03 and CF04, hence Campylobacter was not detected. In contrast, the DNA that was from samples that were subjected to a selective enrichment step in Preston broth prior to PCR assay always gave amplified bands of 340-380 bp, therefore the presence of Campylobacter was confirmed. Detection of campylobacters from naturally contaminated, turbid, environmental water may not be feasible by direct PCR assay because of low numbers and the presence of high concentration of humic matter and other PCR inhibitors. The enrichment of water samples in selective broth, however, facilitated PCR detection of Campylobacter probably by increasing cell number and by diluting PCR inhibitors.

  11. Anaerobic degradation of propane and butane by sulfate-reducing bacteria enriched from marine hydrocarbon cold seeps.

    Science.gov (United States)

    Jaekel, Ulrike; Musat, Niculina; Adam, Birgit; Kuypers, Marcel; Grundmann, Olav; Musat, Florin

    2013-05-01

    The short-chain, non-methane hydrocarbons propane and butane can contribute significantly to the carbon and sulfur cycles in marine environments affected by oil or natural gas seepage. In the present study, we enriched and identified novel propane and butane-degrading sulfate reducers from marine oil and gas cold seeps in the Gulf of Mexico and Hydrate Ridge. The enrichment cultures obtained were able to degrade simultaneously propane and butane, but not other gaseous alkanes. They were cold-adapted, showing highest sulfate-reduction rates between 16 and 20 °C. Analysis of 16S rRNA gene libraries, followed by whole-cell hybridizations with sequence-specific oligonucleotide probes showed that each enrichment culture was dominated by a unique phylotype affiliated with the Desulfosarcina-Desulfococcus cluster within the Deltaproteobacteria. These phylotypes formed a distinct phylogenetic cluster of propane and butane degraders, including sequences from environments associated with hydrocarbon seeps. Incubations with (13)C-labeled substrates, hybridizations with sequence-specific probes and nanoSIMS analyses showed that cells of the dominant phylotypes were the first to become enriched in (13)C, demonstrating that they were directly involved in hydrocarbon degradation. Furthermore, using the nanoSIMS data, carbon assimilation rates were calculated for the dominant cells in each enrichment culture.

  12. Detection of Human Picornaviruses by Multiplex Reverse Transcription-PCR and Liquid Hybridization

    OpenAIRE

    Jokela, Pia; Joki-Korpela, Päivi; Maaronen, Marita; Glumoff, Virpi; Hyypiä, Timo

    2005-01-01

    A qualitative multiplex reverse transcription (RT)-PCR and liquid hybridization assay for the detection of human enteroviruses, rhinoviruses, parechoviruses, and Aichi virus was developed. Furthermore, a separate assay for the recognition of hepatitis A virus was established to complement the test pattern so that all human picornaviruses were covered. The amplicons, which represented the 5′ untranslated regions of the viral RNA genomes, were identified in liquid hybridization reactions with g...

  13. Evaluation of a novel PCR-based assay for detection and identification of Chlamydia trachomatis serovars in cervical specimens.

    NARCIS (Netherlands)

    Quint, K.D.; Porras, C.; Safaeian, M.; Gonzalez, P.; Hildesheim, A.; Quint, W.G.V.; Doorn, L.J. van; Silva, S.; Melchers, W.J.G.; Schiffman, M.; Rodriguez, A.C.; Wacholder, S.; Freer, E.; Cortes, B.; Herrero, R.

    2007-01-01

    The aims of this study were to compare a novel PCR-based Chlamydia trachomatis detection and genotyping (Ct-DT) assay with the FDA-approved, commercially available C. trachomatis detection Hybrid Capture 2 (HC2) assay and to investigate the C. trachomatis serovar distribution among young women in a

  14. Practical assay issues with the PERT/PBRT assay: a highly sensitive reverse transcriptase assay.

    Science.gov (United States)

    Chang, A; Dusing, S

    2006-01-01

    Product safety testing for retroviruses can be achieved by a panel of screening assays, including electron microscopy, viral gene specific PCRs, virus propagation, and detection of reverse transciptase activity. The application of PCR-based reverse transcriptase assays (PERT) that are approximately a million-fold more sensitive than conventional nucleotide incorporation assays in the testing of biologicals is described. Use of PERT assays can be applied to three areas: (i) screening for adventitious retrovirus contamination; (ii) detecting and quantifying endogenous viral particle load and (iii) monitoring levels of infectious retrovirus generation in cell lines that contain endogenous retroviruses.

  15. Hybrid weakness in a rice interspecific hybrid is nitrogen-dependent, and accompanied by changes in gene expression at both total transcript level and parental allele partitioning

    Science.gov (United States)

    Lin, Xiuyun; Wang, Jie; Yu, Jiamiao; Sun, Yue; Miao, Yiling; Li, Qiuping; Sanguinet, Karen A.; Liu, Bao

    2017-01-01

    Background Hybrid weakness, a phenomenon opposite to heterosis, refers to inferior growth and development in a hybrid relative to its pure-line parents. Little attention has been paid to the phenomenological or mechanistic aspect of hybrid weakness, probably due to its rare occurrence. Methodology/Principal findings Here, using a set of interspecific triploid F1 hybrids between Oryza sativa, ssp. japonica (genome AA) and a tetraploid wild rice species, O. alta (genome, CCDD), we investigated the phenotypic and physiological differences between the F1 hybrids and their parents under normal and nitrogen-limiting conditions. We quantified the expression levels of 21 key genes involved in three important pathways pertinent to the assayed phenotypic and physiological traits by real-time qRT-PCR. Further, we assayed expression partitioning of parental alleles for eight genes in the F1 hybrids relative to the in silico “hybrids” (parental cDNA mixture) under both normal and N-limiting conditions by using locus-specific cDNA pyrosequencing. Conclusions/Significance We report that the F1 hybrids showed weakness in several phenotypic traits at the final seedling-stage compared with their corresponding mid-parent values (MPVs). Nine of the 21 studied genes showed contrasted expression levels between hybrids and parents (MPVs) under normal vs. N-limiting conditions. Interestingly, under N-limiting conditions, the overtly enhanced partitioning of maternal allele expression in the hybrids for eight assayed genes echo their attenuated hybrid weakness in phenotypes, an observation further bolstered by more resemblance of hybrids to the maternal parent under N-limiting conditions compared to normal conditions in a suite of measured physiological traits. Our observations suggest that both overall expression level and differential partitioning of parental alleles of critical genes contribute to condition-specific hybrid weakness. PMID:28248994

  16. Henkin and Hybrid Logic

    DEFF Research Database (Denmark)

    Blackburn, Patrick Rowan; Huertas, Antonia; Manzano, Maria;

    2014-01-01

    Leon Henkin was not a modal logician, but there is a branch of modal logic that has been deeply influenced by his work. That branch is hybrid logic, a family of logics that extend orthodox modal logic with special proposition symbols (called nominals) that name worlds. This paper explains why...... Henkin’s techniques are so important in hybrid logic. We do so by proving a completeness result for a hybrid type theory called HTT, probably the strongest hybrid logic that has yet been explored. Our completeness result builds on earlier work with a system called BHTT, or basic hybrid type theory...... is due to the first-order perspective, which lies at the heart of Henin’s best known work and hybrid logic....

  17. Detection of Salmonella invA gene in shrimp enrichment culture by polymerase chain reaction.

    Science.gov (United States)

    Upadhyay, Bishnu Prasad; Utrarachkij, Fuangfa; Thongshoob, Jarinee; Mahakunkijcharoen, Yuvadee; Wongchinda, Niracha; Suthienkul, Orasa; Khusmith, Srisin

    2010-03-01

    Contamination of seafood with salmonellae is a major public health concern. Detection of Salmonella by standard culture methods is time consuming. In this study, an enrichment culture step prior to polymerase chain reaction (PCR) was applied to detect 284 bp fragment of Salmonella invA in comparison with the conventional culture method in 100 shrimp samples collected from four different shrimp farms and fresh food markets around Bangkok. Samples were pre-enriched in non-selective lactose broth (LB) and selective tetrathionate broth (TTB). PCR detection limit was 10 pg and 10(4) cfu/ml of viable salmonellae with 100% specificity. PCR assay detected 19 different Salmonella serovars belonging to 8 serogroups (B, C1, C2-C3, D1, E1, E4 and K) commonly found in clinical and environmental samples in Thailand. The detection rate of PCR following TTB enrichment (24%) was higher than conventional culture method (19%). PCR following TTB, but not in LB enrichment allowed salmonella detection with 84% sensitivity, 90% specificity and 89% accuracy. Shrimp samples collected from fresh food markets had higher levels of contaminated salmonellae than those from shrimp farms. The results indicated that incorporation of an enrichment step prior to PCR has the potential to be applied for detection of naturally contaminated salmonellae in food, environment and clinical samples.

  18. Molecular and Kinetic Characterization of Planktonic Nitrospira spp. Selectively Enriched from Activated Sludge.

    Science.gov (United States)

    Park, Mee-Rye; Park, Hongkeun; Chandran, Kartik

    2017-03-07

    Nitrospira spp. are chemolithoautotrophic nitrite-oxidizing bacteria (NOB), which are ubiquitous in natural and engineered environments. However, there exist few independent biokinetic studies on Nitrospira spp., likely because their isolation and selective enrichment from environmental consortia such as activated sludge can be challenging. Herein, planktonic Nitrospira spp. cultures closely related to Candidatus Nitrospira defluvii (Nitrospira lineage I) were successfully enriched from activated sludge in a sequencing batch reactor by maintaining sustained limiting extant nitrite and dissolved oxygen concentrations. Morphologically, the enrichment consisted largely of planktonic cells with an average characteristic diameter of 1.3 ± 0.6 μm. On the basis of respirometric assays, estimated maximum specific growth rate (μmax), nitrite half saturation coefficient (KS), oxygen half saturation coefficient (KO), and biomass yield coefficient (Y) of the enriched cultures were 0.69 ± 0.10 d(-1), 0.52 ± 0.14 mg-N/L, 0.33 ± 0.14 mg-O2/L, and 0.14 ± 0.02 mg-COD/mg-N, respectively. These parameters collectively reflect not just higher affinities of this enrichment for nitrite and oxygen, respectively, but also a higher biomass yield and energy transfer efficiency relative to Nitrobacter spp. Used in combination, these kinetic and thermodynamic parameters can help toward the development and application of energy-efficient biological nutrient removal processes through effective Nitrospira out-selection.

  19. Gas centrifuge enrichment plants inspection frequency and remote monitoring issues for advanced safeguards implementation

    Energy Technology Data Exchange (ETDEWEB)

    Boyer, Brian David [Los Alamos National Laboratory; Erpenbeck, Heather H [Los Alamos National Laboratory; Miller, Karen A [Los Alamos National Laboratory; Ianakiev, Kiril D [Los Alamos National Laboratory; Reimold, Benjamin A [Los Alamos National Laboratory; Ward, Steven L [Los Alamos National Laboratory; Howell, John [GLASGOW UNIV.

    2010-09-13

    Current safeguards approaches used by the IAEA at gas centrifuge enrichment plants (GCEPs) need enhancement in order to verify declared low enriched uranium (LEU) production, detect undeclared LEU production and detect high enriched uranium (BEU) production with adequate probability using non destructive assay (NDA) techniques. At present inspectors use attended systems, systems needing the presence of an inspector for operation, during inspections to verify the mass and {sup 235}U enrichment of declared cylinders of uranium hexafluoride that are used in the process of enrichment at GCEPs. This paper contains an analysis of how possible improvements in unattended and attended NDA systems including process monitoring and possible on-site destructive analysis (DA) of samples could reduce the uncertainty of the inspector's measurements providing more effective and efficient IAEA GCEPs safeguards. We have also studied a few advanced safeguards systems that could be assembled for unattended operation and the level of performance needed from these systems to provide more effective safeguards. The analysis also considers how short notice random inspections, unannounced inspections (UIs), and the concept of information-driven inspections can affect probability of detection of the diversion of nuclear material when coupled to new GCEPs safeguards regimes augmented with unattended systems. We also explore the effects of system failures and operator tampering on meeting safeguards goals for quantity and timeliness and the measures needed to recover from such failures and anomalies.

  20. Multiplex target enrichment using DNA indexing for ultra-high throughput SNP detection.

    LENUS (Irish Health Repository)

    Kenny, Elaine M

    2011-02-01

    Screening large numbers of target regions in multiple DNA samples for sequence variation is an important application of next-generation sequencing but an efficient method to enrich the samples in parallel has yet to be reported. We describe an advanced method that combines DNA samples using indexes or barcodes prior to target enrichment to facilitate this type of experiment. Sequencing libraries for multiple individual DNA samples, each incorporating a unique 6-bp index, are combined in equal quantities, enriched using a single in-solution target enrichment assay and sequenced in a single reaction. Sequence reads are parsed based on the index, allowing sequence analysis of individual samples. We show that the use of indexed samples does not impact on the efficiency of the enrichment reaction. For three- and nine-indexed HapMap DNA samples, the method was found to be highly accurate for SNP identification. Even with sequence coverage as low as 8x, 99% of sequence SNP calls were concordant with known genotypes. Within a single experiment, this method can sequence the exonic regions of hundreds of genes in tens of samples for sequence and structural variation using as little as 1 μg of input DNA per sample.

  1. Methylation profiling using methylated DNA immunoprecipitation and tiling array hybridization.

    Science.gov (United States)

    Cheung, Hoi-Hung; Lee, Tin-Lap; Rennert, Owen M; Chan, Wai-Yee

    2012-01-01

    DNA methylation is an important epigenetic modification that regulates development and plays a role in the pathophysiology of many diseases. It is dynamically changed during germline development. Methylated DNA immunoprecipitation (MeDIP) is an efficient, cost-effective method for locus-specific and genome-wide analysis. Methylated DNA fragments are enriched by a 5-methylcytidine-recognizing antibody, therefore allowing the analysis of both CpG and non-CpG methylation. The enriched DNA fragments can be amplified and hybridized to tiling arrays covering CpG islands, promoters, or the entire genome. Comparison of different methylomes permits the discovery of differentially methylated regions that might be important in disease- or tissue-specific expression. Here, we describe an established MeDIP protocol and tiling array hybridization method for profiling methylation of testicular germ cells.

  2. GLANET: genomic loci annotation and enrichment tool.

    Science.gov (United States)

    Otlu, Burçak; Firtina, Can; Keles, Sündüz; Tastan, Oznur

    2017-09-15

    Genomic studies identify genomic loci representing genetic variations, transcription factor (TF) occupancy, or histone modification through next generation sequencing (NGS) technologies. Interpreting these loci requires evaluating them with known genomic and epigenomic annotations. We present GLANET as a comprehensive annotation and enrichment analysis tool which implements a sampling-based enrichment test that accounts for GC content and/or mappability biases, jointly or separately. GLANET annotates and performs enrichment analysis on these loci with a rich library. We introduce and perform novel data-driven computational experiments for assessing the power and Type-I error of its enrichment procedure which show that GLANET has attained high statistical power and well-controlled Type-I error rate. As a key feature, users can easily extend its library with new gene sets and genomic intervals. Other key features include assessment of impact of single nucleotide variants (SNPs) on TF binding sites and regulation based pathway enrichment analysis. GLANET can be run using its GUI or on command line. GLANET's source code is available at https://github.com/burcakotlu/GLANET . Tutorials are provided at https://glanet.readthedocs.org . burcak@ceng.metu.edu.tr or oznur.tastan@cs.bilkent.edu.tr. Supplementary data are available at Bioinformatics online.

  3. Enrichment of Acinetobacter spp. from food samples.

    Science.gov (United States)

    Carvalheira, Ana; Ferreira, Vânia; Silva, Joana; Teixeira, Paula

    2016-05-01

    Relatively little is known about the role of foods in the chain of transmission of acinetobacters and the occurrence of different Acinetobacter spp. in foods. Currently, there is no standard procedure to recover acinetobacters from food in order to gain insight into the food-related ecology and epidemiology of acinetobacters. This study aimed to assess whether enrichment in Dijkshoorn enrichment medium followed by plating in CHROMagar™ Acinetobacter medium is a useful method for the isolation of Acinetobacter spp. from foods. Recovery of six Acinetobacter species from food spiked with these organisms was compared for two selective enrichment media (Baumann's enrichment and Dijkshoorn's enrichment). Significantly (p Acinetobacter was applied to detect Acinetobacter spp. in different foods. Fourteen different presumptive acinetobacters were recovered and assumed to represent nine different strains on the basis of REP-PCR typing. Eight of these strains were identified by rpoB gene analysis as belonging to the species Acinetobacter johnsonii, Acinetobacter calcoaceticus, Acinetobacter guillouiae and Acinetobacter gandensis. It was not possible to identify the species level of one strain which may suggests that it represents a distinct species.

  4. BSA Hybrid Synthesized Polymer

    Institute of Scientific and Technical Information of China (English)

    Zong Bin LIU; Xiao Pei DENG; Chang Sheng ZHAO

    2006-01-01

    Bovine serum albumin (BSA), a naturally occurring biopolymer, was regarded as a polymeric material to graft to an acrylic acid (AA)-N-vinyl pyrrolidone (NVP) copolymer to form a biomacromolecular hybrid polymer. The hybrid polymer can be blended with polyethersulfone (PES) to increase the hydrophilicity of the PES membrane, which suggested that the hybrid polymer might have a wide application in the modification of biomaterials.

  5. Hybrid Action Systems

    DEFF Research Database (Denmark)

    Ronkko, Mauno; Ravn, Anders P.

    1997-01-01

    a differential action, which allows differential equations as primitive actions. The extension allows us to model hybrid systems with both continuous and discrete behaviour. The main result of this paper is an extension of such a hybrid action system with parallel composition. The extension does not change...... the original meaning of the parallel composition, and therefore also the ordinary action systems can be composed in parallel with the hybrid action systems....

  6. HYBRID VEHICLE CONTROL SYSTEM

    Directory of Open Access Journals (Sweden)

    V. Dvadnenko

    2016-06-01

    Full Text Available The hybrid vehicle control system includes a start–stop system for an internal combustion engine. The system works in a hybrid mode and normal vehicle operation. To simplify the start–stop system, there were user new possibilities of a hybrid car, which appeared after the conversion. Results of the circuit design of the proposed system of basic blocks are analyzed.

  7. Nanoscale Organic Hybrid Electrolytes

    KAUST Repository

    Nugent, Jennifer L.

    2010-08-20

    Nanoscale organic hybrid electrolytes are composed of organic-inorganic hybrid nanostructures, each with a metal oxide or metallic nanoparticle core densely grafted with an ion-conducting polyethylene glycol corona - doped with lithium salt. These materials form novel solvent-free hybrid electrolytes that are particle-rich, soft glasses at room temperature; yet manifest high ionic conductivity and good electrochemical stability above 5V. © 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Hybrid radiator cooling system

    Science.gov (United States)

    France, David M.; Smith, David S.; Yu, Wenhua; Routbort, Jules L.

    2016-03-15

    A method and hybrid radiator-cooling apparatus for implementing enhanced radiator-cooling are provided. The hybrid radiator-cooling apparatus includes an air-side finned surface for air cooling; an elongated vertically extending surface extending outwardly from the air-side finned surface on a downstream air-side of the hybrid radiator; and a water supply for selectively providing evaporative cooling with water flow by gravity on the elongated vertically extending surface.

  9. From Antenna to Assay

    Science.gov (United States)

    Moore, Evan G.; Samuel, Amanda P. S.; Raymond, Kenneth N.

    2009-01-01

    Conspectus Ligand-sensitized, luminescent lanthanide(III) complexes are of considerable importance because their unique photophysical properties (microsecond to millisecond lifetimes, characteristic and narrow emission bands, and large Stokes shifts) make them well suited as labels in fluorescence-based bioassays. The long-lived emission of lanthanide(III) cations can be temporally resolved from scattered light and background fluorescence to vastly enhance measurement sensitivity. One challenge in this field is the design of sensitizing ligands that provide highly emissive complexes with sufficient stability and aqueous solubility for practical applications. In this Account, we give an overview of some of the general properties of the trivalent lanthanides and follow with a summary of advances made in our laboratory in the development of highly luminescent Tb(III) and Eu(III) complexes for applications in biotechnology. A focus of our research has been the optimization of these compounds as potential commercial agents for use in Homogeneous Time-Resolved Fluorescence (HTRF) technology. Our approach involves developing high-stability octadentate Tb(III) and Eu(III) complexes that rely on all-oxygen donor atoms and using multi-chromophore chelates to increase molar absorptivity; earlier examples utilized a single pendant chromophore (that is, a single “antenna”). Ligands based on 2-hydroxyisophthalamide (IAM) provide exceptionally emissive Tb(III) complexes with quantum yield values up to ∼60% that are stable at the nanomolar concentrations required for commercial assays. Through synthetic modification of the IAM chromophore and time-dependent density functional theory (TD-DFT) calculations, we have developed a method to predict absorption and emission properties of these chromophores as a tool to guide ligand design. Additionally, we have investigated chiral IAM ligands that yield Tb(III) complexes possessing both high quantum yield values and strong

  10. Hybrid Unifying Variable Supernetwork Model

    Institute of Scientific and Technical Information of China (English)

    LIU; Qiang; FANG; Jin-qing; LI; Yong

    2015-01-01

    In order to compare new phenomenon of topology change,evolution,hybrid ratio and network characteristics of unified hybrid network theoretical model with unified hybrid supernetwork model,this paper constructed unified hybrid variable supernetwork model(HUVSM).The first layer introduces a hybrid ratio dr,the

  11. Large Unifying Hybrid Supernetwork Model

    Institute of Scientific and Technical Information of China (English)

    LIU; Qiang; FANG; Jin-qing; LI; Yong

    2015-01-01

    For depicting multi-hybrid process,large unifying hybrid network model(so called LUHNM)has two sub-hybrid ratios except dr.They are deterministic hybrid ratio(so called fd)and random hybrid ratio(so called gr),respectively.

  12. Exome Capture with Heterologous Enrichment in Pig (Sus scrofa).

    Science.gov (United States)

    Guiatti, Denis; Pomari, Elena; Radovic, Slobodanka; Spadotto, Alessandro; Stefanon, Bruno

    2015-01-01

    The discovery of new protein-coding DNA variants related to carcass traits is very important for the Italian pig industry, which requires heavy pigs with higher thickness of subcutaneous fat for Protected Designation of Origin (PDO) productions. Exome capture techniques offer the opportunity to focus on the regions of DNA potentially related to the gene and protein expression. In this research a human commercial target enrichment kit was used to evaluate its performances for pig exome capture and for the identification of DNA variants suitable for comparative analysis. Two pools of 30 pigs each, crosses of Italian Duroc X Large White (DU) and Commercial hybrid X Large White (HY), were used and NGS libraries were prepared with the SureSelectXT Target Enrichment System for Illumina Paired-End Sequencing Library (Agilent). A total of 140.2 M and 162.5 M of raw reads were generated for DU and HY, respectively. Average coverage of all the exonic regions for Sus scrofa (ENSEMBL Sus_scrofa.Sscrofa10.2.73.gtf) was 89.33X for DU and 97.56X for HY; and 35% of aligned bases uniquely mapped to off-target regions. Comparison of sequencing data with the Sscrofa10.2 reference genome, after applying hard filtering criteria, revealed a total of 232,530 single nucleotide variants (SNVs) of which 20.6% mapped in exonic regions and 49.5% within intronic regions. The comparison of allele frequencies of 213 randomly selected SNVs from exome sequencing and the same SNVs analyzed with a Sequenom MassARRAY® system confirms that this "human-on-pig" approach offers new potentiality for the identification of DNA variants in protein-coding genes.

  13. Bacterial Community Profiling of H2/CO2 or Formate-Utilizing Acetogens Enriched from Diverse Ecosystems

    Science.gov (United States)

    Han, R.; Zhang, L.; Fu, B.; Liu, H.

    2014-12-01

    Synthetic gases are usually generated from either cellulosic agricultural waste combustion or industrial release and could be subsequently transformed into acetate, ethanol, and/or butyrate by homoacetogenic bacteria, which commonly possess reductive acetyl-CoA synthesis pathway. Homoacetogen-based syngas fermentation technology provides an alternative solution to link greenhouse gas emission control and cellulosic solid waste treatment with biofuels production. The objective of our current project is to hunt for homoacetogens with capabilities of highly efficiently converting syngases to chemical solvents. In this study, we evaluated homoacetogens population dynamics during enrichments and pinpointed dominant homoacetogens representing diverse ecosystems enriched by different substrates. We enriched homoacetogens from four different samples including waste activate sludge, freshwater sediment, anaerobic methanogenic sludge, and cow manure using H2/CO2 (4:1) or formate as substrate for homoacetogen enrichment. Along with the formyltetrahydrofolate synthetase (FTHFS) gene (fhs gene)-specific real time qPCR assay and Terminal Restriction Fragment Length Polymorphism (T-RFLP) analysis, 16S rRNA based 454 high-throughput pyrosequencing was applied to reveal the population dynamic and community structure during enrichment from different origins. Enrichment of homoacetogenic populations coincided with accumulations of short chain fatty acids such as acetate and butyrate. 454 high-throughput pyrosequencing revealed Firmicutes and Spirochaetes populations became dominant while the overall microbial diversity decreased after enrichment. The most abundant sequences among the four origins belonged to the following phyla: Firmicutes, Spirochaetes, Proteobacteria, and Bacteroidetes, accounting for 62.1%-99.1% of the total reads. The major putative homoacetogenic species enriched on H2/CO2 or formate belonged to Clostridium spp., Acetobacterium spp., Acetoanaerobium spp

  14. Hybrid Rocket Technology

    National Research Council Canada - National Science Library

    Sankaran Venugopal; K K Rajesh; V Ramanujachari

    2011-01-01

    With their unique operational characteristics, hybrid rockets can potentially provide safer, lower-cost avenues for spacecraft and missiles than the current solid propellant and liquid propellant systems...

  15. Hybrid FOSS Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Armstrong researchers are continuing their efforts to further develop FOSS technologies. A hybrid FOSS technique (HyFOSS) employs conventional continuous grating...

  16. Microbial identification by immunohybridization assay of artificial RNA labels

    Science.gov (United States)

    Kourentzi, Katerina D.; Fox, George E.; Willson, Richard C.

    2002-01-01

    Ribosomal RNA (rRNA) and engineered stable artificial RNAs (aRNAs) are frequently used to monitor bacteria in complex ecosystems. In this work, we describe a solid-phase immunocapture hybridization assay that can be used with low molecular weight RNA targets. A biotinylated DNA probe is efficiently hybridized in solution with the target RNA, and the DNA-RNA hybrids are captured on streptavidin-coated plates and quantified using a DNA-RNA heteroduplex-specific antibody conjugated to alkaline phosphatase. The assay was shown to be specific for both 5S rRNA and low molecular weight (LMW) artificial RNAs and highly sensitive, allowing detection of as little as 5.2 ng (0.15 pmol) in the case of 5S rRNA. Target RNAs were readily detected even in the presence of excess nontarget RNA. Detection using DNA probes as small as 17 bases targeting a repetitive artificial RNA sequence in an engineered RNA was more efficient than the detection of a unique sequence.

  17. Uranium enrichment management review: summary of analysis

    Energy Technology Data Exchange (ETDEWEB)

    1981-01-01

    In May 1980, the Assistant Secretary for Resource Applications within the Department of Energy requested that a group of experienced business executives be assembled to review the operation, financing, and management of the uranium enrichment enterprise as a basis for advising the Secretary of Energy. After extensive investigation, analysis, and discussion, the review group presented its findings and recommendations in a report on December 2, 1980. The following pages contain background material on which that final report was based. This report is arranged in chapters that parallel those of the uranium enrichment management review final report - chapters that contain summaries of the review group's discussion and analyses in six areas: management of operations and construction; long-range planning; marketing of enrichment services; financial management; research and development; and general management. Further information, in-depth analysis, and discussion of suggested alternative management practices are provided in five appendices.

  18. Nutrient enrichment increases mortality of mangroves.

    Directory of Open Access Journals (Sweden)

    Catherine E Lovelock

    Full Text Available Nutrient enrichment of the coastal zone places intense pressure on marine communities. Previous studies have shown that growth of intertidal mangrove forests is accelerated with enhanced nutrient availability. However, nutrient enrichment favours growth of shoots relative to roots, thus enhancing growth rates but increasing vulnerability to environmental stresses that adversely affect plant water relations. Two such stresses are high salinity and low humidity, both of which require greater investment in roots to meet the demands for water by the shoots. Here we present data from a global network of sites that documents enhanced mortality of mangroves with experimental nutrient enrichment at sites where high sediment salinity was coincident with low rainfall and low humidity. Thus the benefits of increased mangrove growth in response to coastal eutrophication is offset by the costs of decreased resilience due to mortality during drought, with mortality increasing with soil water salinity along climatic gradients.

  19. Nutrient Enrichment Increases Mortality of Mangroves

    Science.gov (United States)

    Lovelock, Catherine E.; Ball, Marilyn C.; Martin, Katherine C.; C. Feller, Ilka

    2009-01-01

    Nutrient enrichment of the coastal zone places intense pressure on marine communities. Previous studies have shown that growth of intertidal mangrove forests is accelerated with enhanced nutrient availability. However, nutrient enrichment favours growth of shoots relative to roots, thus enhancing growth rates but increasing vulnerability to environmental stresses that adversely affect plant water relations. Two such stresses are high salinity and low humidity, both of which require greater investment in roots to meet the demands for water by the shoots. Here we present data from a global network of sites that documents enhanced mortality of mangroves with experimental nutrient enrichment at sites where high sediment salinity was coincident with low rainfall and low humidity. Thus the benefits of increased mangrove growth in response to coastal eutrophication is offset by the costs of decreased resilience due to mortality during drought, with mortality increasing with soil water salinity along climatic gradients. PMID:19440554

  20. From hybrid swarms to swarms of hybrids

    Science.gov (United States)

    Stohlgren, Thomas J.; Szalanski, Allen L; Gaskin, John F.; Young, Nicholas E.; West, Amanda; Jarnevich, Catherine S.; Tripodi, Amber

    2015-01-01

    Science has shown that the introgression or hybridization of modern humans (Homo sapiens) with Neanderthals up to 40,000 YBP may have led to the swarm of modern humans on earth. However, there is little doubt that modern trade and transportation in support of the humans has continued to introduce additional species, genotypes, and hybrids to every country on the globe. We assessed the utility of species distributions modeling of genotypes to assess the risk of current and future invaders. We evaluated 93 locations of the genus Tamarix for which genetic data were available. Maxent models of habitat suitability showed that the hybrid, T. ramosissima x T. chinensis, was slightly greater than the parent taxa (AUCs > 0.83). General linear models of Africanized honey bees, a hybrid cross of Tanzanian Apis mellifera scutellata and a variety of European honey bee including A. m. ligustica, showed that the Africanized bees (AUC = 0.81) may be displacing European honey bees (AUC > 0.76) over large areas of the southwestern U.S. More important, Maxent modeling of sub-populations (A1 and A26 mitotypes based on mDNA) could be accurately modeled (AUC > 0.9), and they responded differently to environmental drivers. This suggests that rapid evolutionary change may be underway in the Africanized bees, allowing the bees to spread into new areas and extending their total range. Protecting native species and ecosystems may benefit from risk maps of harmful invasive species, hybrids, and genotypes.

  1. Rare-earth elements enrichment of Pacific seafloor sediments: the view from volcanic islands of Polynesia

    Science.gov (United States)

    Melleton, Jérémie; Tuduri, Johann; Pourret, Olivier; Bailly, Laurent; Gisbert, Thierry

    2014-05-01

    Rare-earth elements (REEs) are key metals for «green» technologies such as energy saving lamps or permanent magnets used in, e.g., wind turbines, hard disk drives, portable phone or electric or hybrid vehicles. Since several years, world demand for these metals is therefore drastically increasing. The quasi-monopolistic position of China, which produces around 95 % of global REEs production, generates risks for the industries that depend on a secure supply of REEs. In response, countries are developing and diversifying their supply sources, with new mining projects located outside China and efforts in the area of REEs recycling. Most of these projects focus on deposits related to carbonatites and alkaline-peralkaline magmatism, which are generally enriched in light REEs (LREEs) compared to the heavy REEs (HREEs)-enriched deposits of the ion-adsorption types, located in southern China. However, a recent study revealed new valuable resources corresponding to seafloor sediments located in the south-eastern and north-central Pacific. The deep-sea mud described by these authors show a higher HREE/LREE ratio than ion-adsorption deposits, a feature which significantly increases their economic interest. The authors suggest mid-ocean ridge hydrothermal activity as an explanation to this anomalous enrichment. However, several contributions have documented considerable REEs enrichment in basalts and peridotitic xenoliths from French Polynesia. Several arguments have been exposed in favour of a supergene origin, with a short migration, suggesting that REEs were collected from weathered basalts. The Tahaa volcanic island (Sous-le-Vent Island, Society Archipelago, French Polynesia) is the first location where such enrichment has been described. New petrographic and mineralogical investigations confirm a supergene mobilization of this abnormal occurrence. REE-bearing minerals (mainly phosphates of the rhabdophane group) are primarily located within basalt vesicles but also in

  2. Validation of Flow Cytometry and Magnetic Bead-Based Methods to Enrich CNS Single Cell Suspensions for Quiescent Microglia.

    Science.gov (United States)

    Volden, T A; Reyelts, C D; Hoke, T A; Arikkath, J; Bonasera, S J

    2015-12-01

    Microglia are resident mononuclear phagocytes within the CNS parenchyma that intimately interact with neurons and astrocytes to remodel synapses and extracellular matrix. We briefly review studies elucidating the molecular pathways that underlie microglial surveillance, activation, chemotaxis, and phagocytosis; we additionally place these studies in a clinical context. We describe and validate an inexpensive and simple approach to obtain enriched single cell suspensions of quiescent parenchymal and perivascular microglia from the mouse cerebellum and hypothalamus. Following preparation of regional CNS single cell suspensions, we remove myelin debris, and then perform two serial enrichment steps for cells expressing surface CD11b. Myelin depletion and CD11b enrichment are both accomplished using antigen-specific magnetic beads in an automated cell separation system. Flow cytometry of the resultant suspensions shows a significant enrichment for CD11b(+)/CD45(+) cells (perivascular microglia) and CD11b(+)/CD45(-) cells (parenchymal microglia) compared to starting suspensions. Of note, cells from these enriched suspensions minimally express Aif1 (aka Iba1), suggesting that the enrichment process does not evoke significant microglial activation. However, these cells readily respond to a functional challenge (LPS) with significant changes in the expression of molecules specifically associated with microglia. We conclude that methods employing a combination of magnetic-bead based sorting and flow cytometry produce suspensions highly enriched for microglia that are appropriate for a variety of molecular and cellular assays.

  3. 21 CFR 136.115 - Enriched bread, rolls, and buns.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 2 2010-04-01 2010-04-01 false Enriched bread, rolls, and buns. 136.115 Section... § 136.115 Enriched bread, rolls, and buns. (a) Each of the foods enriched bread, enriched rolls, and... label statement of ingredients prescribed for bread, rolls or buns by § 136.110, except that: (1) Each...

  4. An Improved Protocol for Quantification of Freshwater Actinobacteria by Fluorescence In Situ Hybridization

    Science.gov (United States)

    Sekar, Raju; Pernthaler, Annelie; Pernthaler, Jakob; Warnecke, Falk; Posch, Thomas; Amann, Rudolf

    2003-01-01

    We tested a previously described protocol for fluorescence in situ hybridization of marine bacterioplankton with horseradish peroxidase-labeled rRNA-targeted oligonucleotide probes and catalyzed reporter deposition (CARD-FISH) in plankton samples from different lakes. The fraction of Bacteria detected by CARD-FISH was significantly lower than after FISH with fluorescently monolabeled probes. In particular, the abundances of aquatic Actinobacteria were significantly underestimated. We thus developed a combined fixation and permeabilization protocol for CARD-FISH of freshwater samples. Enzymatic pretreatment of fixed cells was optimized for the controlled digestion of gram-positive cell walls without causing overall cell loss. Incubations with high concentrations of lysozyme (10 mg ml−1) followed by achromopeptidase (60 U ml−1) successfully permeabilized cell walls of Actinobacteria for subsequent CARD-FISH both in enrichment cultures and environmental samples. Between 72 and >99% (mean, 86%) of all Bacteria could be visualized with the improved assay in surface waters of four lakes. For freshwater samples, our method is thus superior to the CARD-FISH protocol for marine Bacteria (mean, 55%) and to FISH with directly fluorochrome labeled probes (mean, 67%). Actinobacterial abundances in the studied systems, as detected by the optimized protocol, ranged from 32 to >55% (mean, 45%). Our findings confirm that members of this lineage are among the numerically most important Bacteria of freshwater picoplankton. PMID:12732568

  5. Epithelial cells as alternative human biomatrices for comet assay

    Directory of Open Access Journals (Sweden)

    Emilio eRojas

    2014-11-01

    Full Text Available The comet assay is a valuable experimental tool aimed at mapping DNA damage in human cells in vivo for environmental and occupational monitoring, as well as for therapeutic purposes, such as storage prior to transplant, during tissue engineering, and in experimental ex vivo assays. Furthermore, due to its great versatility, the comet assay allows to explore the use of alternative cell types to assess DNA damage, such as epithelial cells. Epithelial cells, as specialized components of many organs, have the potential to serve as biomatrices that can be used to evaluate genotoxicity and may also serve as early effect biomarkers. Furthermore, 80% of solid cancers are of epithelial origin, which points to the importance of studying DNA damage in these tissues. Indeed, studies including comet assay in epithelial cells have either clear clinical applications (lens and corneal epithelial cells or examine genotoxicity within human biomonitoring and in vitro studies. We here review improvements in determining DNA damage using the comet assay by employing lens, corneal, tear duct, buccal, and nasal epithelial cells. For some of these tissues invasive sampling procedures are needed. Desquamated epithelial cells must be obtained and dissociated prior to examination using the comet assay, and such procedures may induce varying amounts of DNA damage. Buccal epithelial cells require lysis enriched with proteinase K to obtain free nucleosomes.Over a thirty year period, the comet assay in epithelial cells has been litlle employed, however its use indicates that it could be an extraordinary tool not only for risk assessment, but also for diagnosis, prognosis of treatments and diseases.

  6. Transporter assays and assay ontologies: useful tools for drug discovery.

    Science.gov (United States)

    Zdrazil, Barbara; Chichester, Christine; Zander Balderud, Linda; Engkvist, Ola; Gaulton, Anna; Overington, John P

    2014-06-01

    Transport proteins represent an eminent class of drug targets and ADMET (absorption, distribution, metabolism, excretion, toxicity) associated genes. There exists a large number of distinct activity assays for transport proteins, depending on not only the measurement needed (e.g. transport activity, strength of ligand–protein interaction), but also due to heterogeneous assay setups used by different research groups. Efforts to systematically organize this (divergent) bioassay data have large potential impact in Public-Private partnership and conventional commercial drug discovery. In this short review, we highlight some of the frequently used high-throughput assays for transport proteins, and we discuss emerging assay ontologies and their application to this field. Focusing on human P-glycoprotein (Multidrug resistance protein 1; gene name: ABCB1, MDR1), we exemplify how annotation of bioassay data per target class could improve and add to existing ontologies, and we propose to include an additional layer of metadata supporting data fusion across different bioassays.

  7. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.

    2016-01-01

    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...... charged metal ions such as Fe3+, Ga3+, Al3+, Zr4+, and Ti4+ has made it possible to enrich phosphorylated peptides from peptide samples. However, the selectivity of most of the metal ions is limited, when working with highly complex samples, e.g., whole-cell extracts, resulting in contamination from...

  8. Optimization of STR locus enrichment for STR profiling of fragmented DNA.

    Science.gov (United States)

    Ham, Seon-Kyu; Kim, Se-Yong; Ahn, Jang-Won; Seo, Bo Young; Woo, Kwang-Man; Choi, Cheol Yong; Lee, Seung-Hwan

    2014-11-01

    DNA degradation is a major obstacle in gaining an accurate profile with standard DNA typing technology. Although alternative genotyping strategies such as mini-STRs and SNPs have proven to be more successful in profiling degraded DNA, these approaches also have limitations. Here, we show that locus enrichment by hybridization of degraded genomic DNA with an STR locus-specific biotinylated oligonucleotide is a powerful approach to overcome problems in STR typing of highly degraded DNA. An experimental investigation of factors affecting the efficiency of this method indicates that the choice of primer and molar ratio of primers to genomic DNA are critical factors in improving enrichment of the STR locus before genotyping with multiplex kits. In addition, we find that indirect capture rather than direct capture with magnetic beads yields better enrichment efficiency for STR locus enrichments. Using these strategies, we demonstrate an improvement in STR typing of DNA from cultured cells damaged by exposure to sunlight or UV. We suggest that this approach could be applied to highly degraded forensic samples alone or in combination with mini-STRs.

  9. Enrichment of putative stem cells from adipose tissue using dielectrophoretic field-flow fractionation

    Science.gov (United States)

    Vykoukal, Jody; Vykoukal, Daynene M.; Freyberg, Susanne; Alt, Eckhard U.; Gascoyne, Peter R. C.

    2009-01-01

    We have applied the microfluidic cell separation method of dielectrophoretic field-flow fractionation (DEP-FFF) to the enrichment of a putative stem cell population from an enzyme-digested adipose tissue derived cell suspension. A DEP-FFF separator device was constructed using a novel microfluidic-microelectronic hybrid flex-circuit fabrication approach that is scaleable and anticipates future low-cost volume manufacturing. We report the separation of a nucleated cell fraction from cell debris and the bulk of the erythrocyte population, with the relatively rare (<2% starting concentration) NG2-positive cell population (pericytes and/or putative progenitor cells) being enriched up to 14-fold. This work demonstrates a potential clinical application for DEP-FFF and further establishes the utility of the method for achieving label-free fractionation of cell subpopulations. PMID:18651083

  10. 77 FR 14838 - General Electric-Hitachi Global Laser Enrichment LLC, Commercial Laser-Based Uranium Enrichment...

    Science.gov (United States)

    2012-03-13

    ... COMMISSION General Electric-Hitachi Global Laser Enrichment LLC, Commercial Laser-Based Uranium Enrichment... applicant) to authorize construction of a laser-based uranium enrichment facility and possession and use of...-based uranium enrichment facility. Revisions to the application were submitted on March 23, 2010;...

  11. Cardiac hybrid imaging

    Energy Technology Data Exchange (ETDEWEB)

    Gaemperli, Oliver [University Hospital Zurich, Cardiac Imaging, Zurich (Switzerland); University Hospital Zurich, Nuclear Cardiology, Cardiovascular Center, Zurich (Switzerland); Kaufmann, Philipp A. [University Hospital Zurich, Cardiac Imaging, Zurich (Switzerland); Alkadhi, Hatem [University Hospital Zurich, Institute of Diagnostic and Interventional Radiology, Zurich (Switzerland)

    2014-05-15

    Hybrid cardiac single photon emission computed tomography (SPECT)/CT imaging allows combined assessment of anatomical and functional aspects of cardiac disease. In coronary artery disease (CAD), hybrid SPECT/CT imaging allows detection of coronary artery stenosis and myocardial perfusion abnormalities. The clinical value of hybrid imaging has been documented in several subsets of patients. In selected groups of patients, hybrid imaging improves the diagnostic accuracy to detect CAD compared to the single imaging techniques. Additionally, this approach facilitates functional interrogation of coronary stenoses and guidance with regard to revascularization procedures. Moreover, the anatomical information obtained from CT coronary angiography or coronary artery calcium scores (CACS) adds prognostic information over perfusion data from SPECT. The use of cardiac hybrid imaging has been favoured by the dissemination of dedicated hybrid systems and the release of dedicated image fusion software, which allow simple patient throughput for hybrid SPECT/CT studies. Further technological improvements such as more efficient detector technology to allow for low-radiation protocols, ultra-fast image acquisition and improved low-noise image reconstruction algorithms will be instrumental to further promote hybrid SPECT/CT in research and clinical practice. (orig.)

  12. Hybrid intelligent engineering systems

    CERN Document Server

    Jain, L C; Adelaide, Australia University of

    1997-01-01

    This book on hybrid intelligent engineering systems is unique, in the sense that it presents the integration of expert systems, neural networks, fuzzy systems, genetic algorithms, and chaos engineering. It shows that these new techniques enhance the capabilities of one another. A number of hybrid systems for solving engineering problems are presented.

  13. A Hybrid Imagination

    DEFF Research Database (Denmark)

    Jamison, Andrew; Christensen, Steen Hyldgaard; Botin, Lars

    contexts, or sites, for mixing scientific knowledge and technical skills from different fields and social domains into new combinations, thus fostering what the authors term a “hybrid imagination”. Such a hybrid imagination is especially important today, as a way to counter the competitive and commercial...

  14. Hybrid trajectory spaces

    NARCIS (Netherlands)

    Collins, P.J.

    2005-01-01

    In this paper, we present a general framework for describing and studying hybrid systems. We represent the trajectories of the system as functions on a hybrid time domain, and the system itself by its trajectory space, which is the set of all possible trajectories. The trajectory space is given a na

  15. Editorial: Hybrid Systems

    DEFF Research Database (Denmark)

    Olderog, Ernst-Rüdiger; Ravn, Anders Peter

    2007-01-01

    An introduction to three papers in a special issue on Hybrid Systems. These paper were first presented at an IFIP WG 2.2 meeting in Skagen 2005.......An introduction to three papers in a special issue on Hybrid Systems. These paper were first presented at an IFIP WG 2.2 meeting in Skagen 2005....

  16. Hybrid reactors. [Fuel cycle

    Energy Technology Data Exchange (ETDEWEB)

    Moir, R.W.

    1980-09-09

    The rationale for hybrid fusion-fission reactors is the production of fissile fuel for fission reactors. A new class of reactor, the fission-suppressed hybrid promises unusually good safety features as well as the ability to support 25 light-water reactors of the same nuclear power rating, or even more high-conversion-ratio reactors such as the heavy-water type. One 4000-MW nuclear hybrid can produce 7200 kg of /sup 233/U per year. To obtain good economics, injector efficiency times plasma gain (eta/sub i/Q) should be greater than 2, the wall load should be greater than 1 MW.m/sup -2/, and the hybrid should cost less than 6 times the cost of a light-water reactor. Introduction rates for the fission-suppressed hybrid are usually rapid.

  17. Hybrid propulsion technology program

    Science.gov (United States)

    1990-01-01

    Technology was identified which will enable application of hybrid propulsion to manned and unmanned space launch vehicles. Two design concepts are proposed. The first is a hybrid propulsion system using the classical method of regression (classical hybrid) resulting from the flow of oxidizer across a fuel grain surface. The second system uses a self-sustaining gas generator (gas generator hybrid) to produce a fuel rich exhaust that was mixed with oxidizer in a separate combustor. Both systems offer cost and reliability improvement over the existing solid rocket booster and proposed liquid boosters. The designs were evaluated using life cycle cost and reliability. The program consisted of: (1) identification and evaluation of candidate oxidizers and fuels; (2) preliminary evaluation of booster design concepts; (3) preparation of a detailed point design including life cycle costs and reliability analyses; (4) identification of those hybrid specific technologies needing improvement; and (5) preperation of a technology acquisition plan and large scale demonstration plan.

  18. Telomeres in ICF syndrome cells are vulnerable to DNA damage due to elevated DNA:RNA hybrids

    Science.gov (United States)

    Sagie, Shira; Toubiana, Shir; Hartono, Stella R.; Katzir, Hagar; Tzur-Gilat, Aya; Havazelet, Shany; Francastel, Claire; Velasco, Guillaume; Chédin, Frédéric; Selig, Sara

    2017-01-01

    DNA:RNA hybrids, nucleic acid structures with diverse physiological functions, can disrupt genome integrity when dysregulated. Human telomeres were shown to form hybrids with the lncRNA TERRA, yet the formation and distribution of these hybrids among telomeres, their regulation and their cellular effects remain elusive. Here we predict and confirm in several human cell types that DNA:RNA hybrids form at many subtelomeric and telomeric regions. We demonstrate that ICF syndrome cells, which exhibit short telomeres and elevated TERRA levels, are enriched for hybrids at telomeric regions throughout the cell cycle. Telomeric hybrids are associated with high levels of DNA damage at chromosome ends in ICF cells, which are significantly reduced with overexpression of RNase H1. Our findings suggest that abnormally high TERRA levels in ICF syndrome lead to accumulation of telomeric hybrids that, in turn, can result in telomeric dysfunction. PMID:28117327

  19. Development of microsatellite markers from an enriched genomic library of pumpkin (Cucurbita moschata L.

    Directory of Open Access Journals (Sweden)

    Nuchjaree Watcharawongpaiboon

    2007-09-01

    Full Text Available Development of microsatellite markers in Pumpkin (Cucurbita moschata L. was performed using the biotin-streptavidin enrichment procedure. One hundred and thirty three clones were randomly selected. After sequence analysis of 31 randomly picked positive colonies, 100% of the colonies were found to contain microsatellite sequences, and 9 primer pairs were designed. Five of the primers tested could amplify pumpkins DNA and can be used for genetic purity testing of the commercial hybrids. This paper reports the first isolation and utilization of microsatellite markers in pumpkin.

  20. The comet assay – from toy to tool

    Directory of Open Access Journals (Sweden)

    Guenter Speit

    2015-04-01

    Full Text Available The comet assay is nowadays the most common method for measuring DNA damage and repair in single cells. It is based on the microelectrophoretic study published by Ostling and Johanson (1984 and was developed by Singh and coworkers (1988 to a versatile technique for quantitation of low levels of DNA damage in individual cells. This alkaline version still is the basis for the triumphant success of the comet assay in basic research into mechanisms of DNA damage and DNA repair, genotoxicity testing, ecotoxicology and human biomonitoring. Important technical improvements (e.g., the use of precoated slides, introduction of image analysis, high throughput methods, automated scoring systems made the assay more robust and more efficient. Modifications of the standard protocol provide more specific information on the type and biological significance of the damage studied. The introduction of lesion-specific endonucleases allowed the characterization of oxidative base damage, alkylation damage and UV-induced pyrimidine dimers. The combination with fluorescence in situ hybridization (FISH made it possible to identify DNA of particular chromosome regions and measure effects of damage and repair in particular genes. The comet assay is being increasingly used in genotoxicity testing. In particular, the in vivo comet assay has become a component of some genotoxicity test strategies and generally accepted test protocols have evolved over the years. A large international collaborative trial sponsored by the Japanese Center for the Validation of Alternative Methods (JaCVAM was recently completed and an OECD test guideline was approved. The comet assay is widely used in human biomonitoring to measure DNA damage as a marker of exposure to genotoxic agents or to investigate genoprotective effects. However, there are still problems in comparing results from different laboratories and there is need for reducing inter-laboratory variation and identification of standard

  1. Functional properties of soy-enriched tapioca

    African Journals Online (AJOL)

    my mord

    Enrichment of tapioca with soy flour to enhance its nutritional value is feasible however; increase in the ..... entrapment of oil and the binding of fat to the polar chains of proteins ..... Metri AC, Bion FM, Oliveira SRP, Lopes SML (2003). Cassava ...

  2. 21 CFR 137.165 - Enriched flour.

    Science.gov (United States)

    2010-04-01

    ... riboflavin, 24 milligrams of niacin, 0.7 milligrams of folic acid, and 20 milligrams of iron. (b) It may... calcium or wheat germ is excluded in calculating ash content. (f) All ingredients from which the food is... 21 Food and Drugs 2 2010-04-01 2010-04-01 false Enriched flour. 137.165 Section 137.165 Food...

  3. Enrichment Education: Key to NCLEX Success.

    Science.gov (United States)

    Wolahan, Caryle G. Hussey; Wieczorek, Rita Reis

    1991-01-01

    A concomitant decrease in test scores and major change in the composition of the student body was dealt with by the College of Nursing at the State University of New York through the Nursing Education Success Plan. The plan addresses student preparation for the National Council Licensure Exam (NCLEX), curriculum revision, remediation/enrichment,…

  4. Implementing the Schoolwide Enrichment Model in Brazil

    Science.gov (United States)

    de Souza Fleith, Denise; Soriano de Alencar, Eunice M. L.

    2010-01-01

    The Schoolwide Enrichment Model (SEM) has been one of the most widely used models in the education of the gifted in Brazil. It has inspired the political and pedagogical project of the Centers of Activities of High Abilities/Giftedness recently implemented in 27 Brazilian states by the Ministry of Education. In this article, our experience in…

  5. The enrichment history of cosmic metals

    CERN Document Server

    Wiersma, Robert P C; Vecchia, Claudio Dalla; Booth, C M; Theuns, Tom; Aguirre, Anthony

    2010-01-01

    We use a suite of cosmological, hydrodynamical simulations to investigate the chemical enrichment history of the Universe. Specifically, we trace the origin of the metals back in time to investigate when various gas phases were enriched and by what halo masses. We find that the age of the metals decreases strongly with the density of the gas in which they end up. At least half of the metals that reside in the diffuse intergalactic medium (IGM) at redshift zero (two) were ejected from galaxies above redshift two (three). The mass of the haloes that last contained the metals increases rapidly with the gas density. More than half of the mass in intergalactic metals was ejected by haloes with total masses less than 1e11 solar masses and stellar masses less than 1e9 solar masses. The range of halo masses that contributes to the enrichment is wider for the hotter part of the IGM. By combining the `when' and `by what' aspects of the enrichment history, we show that metals residing in lower density gas were typically...

  6. Recommendations based on semantically enriched museum collections

    NARCIS (Netherlands)

    Wang, Y.; Stash, N.; Aroyo, L.; Gorgels, P.; Rutledge, L.; Schreiber, G.

    2008-01-01

    This article presents the CHIP demonstrator1 for providing personalized access to digital museum collections. It consists of three main components: Art Recommender, Tour Wizard, and Mobile Tour Guide. Based on the semantically enriched Rijksmuseum Amsterdam2 collection, we show how Semantic Web tech

  7. Nutrient-enrichment effect on plankton composition:

    OpenAIRE

    Malej, Alenka; Mozetič, Patricija; Turk, Valentina

    1998-01-01

    Three enrichment experiments were conduced in order to analyze the development of plankton biomass and species composition as a response to different nutrient regimes. The additions of all nutrients had the most profound effect on phytoplankton biomass increase, whereas picoplankton abundance decreased continuously in all treatments. Initially dominating phytoplanktonic groups and species expanded significantly towards the end of experiments...

  8. The Complexity of Enriched Mu-Calculi

    CERN Document Server

    Bonatti, Piero A; Murano, Aniello; Vardi, Moshe Y

    2008-01-01

    The fully enriched mu-calculus is the extension of the propositional mu-calculus with inverse programs, graded modalities, and nominals. While satisfiability in several expressive fragments of the fully enriched mu-calculus is known to be decidable and ExpTime-complete, it has recently been proved that the full calculus is undecidable. In this paper, we study the fragments of the fully enriched mu-calculus that are obtained by dropping at least one of the additional constructs. We show that, in all fragments obtained in this way, satisfiability is decidable and ExpTime-complete. Thus, we identify a family of decidable logics that are maximal (and incomparable) in expressive power. Our results are obtained by introducing two new automata models, showing that their emptiness problems are ExpTime-complete, and then reducing satisfiability in the relevant logics to these problems. The automata models we introduce are two-way graded alternating parity automata over infinite trees (2GAPTs) and fully enriched automa...

  9. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.

    2016-01-01

    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...

  10. Environmental Enrichment in the 21st Century.

    Science.gov (United States)

    Coleman, Kristine; Novak, Melinda A

    2017-04-21

    More than a quarter of a century has elapsed since the Animal Welfare Act mandated that research facilities develop and follow a plan to promote the psychological well-being of captive primates. Since passage of this law, considerable effort and resources have been directed to designing environmental enrichment strategies in an effort to improve animal welfare. These plans typically consist of environmental enrichment and socialization efforts. While environmental enhancement has undergone a great deal of improvement in the past 25 years, it should be viewed as a continual work in progress, which takes advantage of emergent and future technologies. In this review, we discuss the objectives of the environmental enhancement plan along with relevant outcome measures, as well as ongoing challenges, costs, and benefits. We then review various enrichment strategies and assess their efficacy in meeting goals and objectives. Finally, we look forward to consider what the future might hold for environmental enrichment of nonhuman primates used in research. © The Author 2017. Published by Oxford University Press on behalf of the Institute for Laboratory Animal Research. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  11. Teaching Mathematical Modelling: Demonstrating Enrichment and Elaboration

    Science.gov (United States)

    Warwick, Jon

    2015-01-01

    This paper uses a series of models to illustrate one of the fundamental processes of model building--that of enrichment and elaboration. The paper describes how a problem context is given which allows a series of models to be developed from a simple initial model using a queuing theory framework. The process encourages students to think about the…

  12. Enriching a Descriptive Grammar with Treebank Queries

    NARCIS (Netherlands)

    Bouma, G.; van Koppen, J.M.|info:eu-repo/dai/nl/203188934; Landsbergen, Frank; Odijk, J.E.J.M.|info:eu-repo/dai/nl/082781710; van der Wouden, Ton; van de Camp, Matje

    2015-01-01

    The Syntax of Dutch (SoD) is a descriptive and detailed grammar of Dutch, that provides data for many issues raised in linguistic theory. We present the results of a pilot project that investigated the possibility of enriching the online version of the text with links to queries that provide

  13. Teaching Mathematical Modelling: Demonstrating Enrichment and Elaboration

    Science.gov (United States)

    Warwick, Jon

    2015-01-01

    This paper uses a series of models to illustrate one of the fundamental processes of model building--that of enrichment and elaboration. The paper describes how a problem context is given which allows a series of models to be developed from a simple initial model using a queuing theory framework. The process encourages students to think about the…

  14. Oxygen permeable membrane for oxygen enriched combustion

    Energy Technology Data Exchange (ETDEWEB)

    Asakawa, S. (Matsushita Research Inst., Tokyo, Japan); Saito, Y.; Kawahito, M.; Ito, Y.; Tsuchiya, S.; Sugata, K.

    1983-02-01

    An oxygen enriched air production system using gas separation membranes has been developed to be used for fuel combustion systems. High oxygen permeable scopolymers, including three dimensional structure, have been synthesized through condensation of polyvinylphenol with some, ..cap alpha..,..omega..-bis (diethylamino) polydimethylsiloxanes. The experimental results showed that the oxygen permeability through the copolymer varies as a function of the dimethylsiloxane content of the copolymers. Such composition dependence of the oxygen permeability was explained on the basis of polymer constitution. Typical values of the oxygen permeability, 3.4 x 10/sup -8/ (cc x cm/cm/sup 2/ x sec x cmHg) and ..cap alpha.., 2.1, were obtained at 72% of dimethylsiloxane content. The copolymers are soluble in most common organic solvents and uniform, defect-free membranes as thin as 1000 Angstroms have been formed by spreading solutions of the copolymer on water. Composite membranes fabricated by applying the membranes to porous support materials were used for practical gas separation and 30% oxygen enriched air was produced from ambient air. A new type oxygen enriched combustion system, which is more efficient for energy saving, has been developed by utilizing oxygen enriched air thus produced. 15 references, 15 figures, 1 table.

  15. MICROBIOLOGICAL ASSAY FOR VITAMIN B

    OpenAIRE

    Bishnoi Kapil*, , ,; Kataria Mahesh; Singhal Vipin; Gupta Deepika

    2012-01-01

    Micronutrients added to foods are analyzed using various procedures depending on their nature and properties. The microbiological assays are better than chemical method because any suitable change in vitamin molecule which may not be detected by chemical method will be revealed by change in microbial activity. The microbiological assay of vitamins is based upon the comparison of the stimulation of growth of bacteria by measured concentration of vitamin with that produced by known concentratio...

  16. ChIP-Enrich: gene set enrichment testing for ChIP-seq data.

    Science.gov (United States)

    Welch, Ryan P; Lee, Chee; Imbriano, Paul M; Patil, Snehal; Weymouth, Terry E; Smith, R Alex; Scott, Laura J; Sartor, Maureen A

    2014-07-01

    Gene set enrichment testing can enhance the biological interpretation of ChIP-seq data. Here, we develop a method, ChIP-Enrich, for this analysis which empirically adjusts for gene locus length (the length of the gene body and its surrounding non-coding sequence). Adjustment for gene locus length is necessary because it is often positively associated with the presence of one or more peaks and because many biologically defined gene sets have an excess of genes with longer or shorter gene locus lengths. Unlike alternative methods, ChIP-Enrich can account for the wide range of gene locus length-to-peak presence relationships (observed in ENCODE ChIP-seq data sets). We show that ChIP-Enrich has a well-calibrated type I error rate using permuted ENCODE ChIP-seq data sets; in contrast, two commonly used gene set enrichment methods, Fisher's exact test and the binomial test implemented in Genomic Regions Enrichment of Annotations Tool (GREAT), can have highly inflated type I error rates and biases in ranking. We identify DNA-binding proteins, including CTCF, JunD and glucocorticoid receptor α (GRα), that show different enrichment patterns for peaks closer to versus further from transcription start sites. We also identify known and potential new biological functions of GRα. ChIP-Enrich is available as a web interface (http://chip-enrich.med.umich.edu) and Bioconductor package. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  17. Mechanisms Underlying Mammalian Hybrid Sterility in Two Feline Interspecies Models.

    Science.gov (United States)

    Davis, Brian W; Seabury, Christopher M; Brashear, Wesley A; Li, Gang; Roelke-Parker, Melody; Murphy, William J

    2015-10-01

    The phenomenon of male sterility in interspecies hybrids has been observed for over a century, however, few genes influencing this recurrent phenotype have been identified. Genetic investigations have been primarily limited to a small number of model organisms, thus limiting our understanding of the underlying molecular basis of this well-documented "rule of speciation." We utilized two interspecies hybrid cat breeds in a genome-wide association study employing the Illumina 63 K single-nucleotide polymorphism array. Collectively, we identified eight autosomal genes/gene regions underlying associations with hybrid male sterility (HMS) involved in the function of the blood-testis barrier, gamete structural development, and transcriptional regulation. We also identified several candidate hybrid sterility regions on the X chromosome, with most residing in close proximity to complex duplicated regions. Differential gene expression analyses revealed significant chromosome-wide upregulation of X chromosome transcripts in testes of sterile hybrids, which were enriched for genes involved in chromatin regulation of gene expression. Our expression results parallel those reported in Mus hybrids, supporting the "Large X-Effect" in mammalian HMS and the potential epigenetic basis for this phenomenon. These results support the value of the interspecies feline model as a powerful tool for comparison to rodent models of HMS, demonstrating unique aspects and potential commonalities that underpin mammalian reproductive isolation.

  18. Grafted Cross-Linked Polyolefin Substrates for Peptide Synthesis and Assays

    DEFF Research Database (Denmark)

    1999-01-01

    suited for use in solid-phase biosystems, notably bioassays, such as immunoassays, DNA hybridization assays or PCR amplification. The grafted chains may bear substituents which are such that the polymer-grafted cross-linked polyolefin substrate is swellable by water or aqueous media, in other words...

  19. Electrochemical DNA sandwich assay with a lipase label for attomole detection of DNA

    DEFF Research Database (Denmark)

    Ferapontova, Elena; Hansen, Majken Nørgaard; Saunders, Aaron Marc

    2010-01-01

    A fast and sensitive electrochemical lipase-based sandwich hybridization assay for detection of attomole levels of DNA has been developed. A combination of magnetic beads, used for pre-concentration and bioseparation of the analyte with a lipase catalyst label allowed detection of DNA with a limit...

  20. Stathmin is enriched in the developing corticospinal tract.

    Science.gov (United States)

    Fuller, Heidi R; Slade, Robert; Jovanov-Milošević, Nataša; Babić, Mirjana; Sedmak, Goran; Šimić, Goran; Fuszard, Matthew A; Shirran, Sally L; Botting, Catherine H; Gates, Monte A

    2015-11-01

    Understanding the intra- and extracellular proteins involved in the development of the corticospinal tract (CST) may offer insights into how the pathway could be regenerated following traumatic spinal cord injury. Currently, however, little is known about the proteome of the developing corticospinal system. The present study, therefore, has used quantitative proteomics and bioinformatics to detail the protein profile of the rat CST during its formation in the spinal cord. This analysis identified increased expression of 65 proteins during the early ingrowth of corticospinal axons into the spinal cord, and 36 proteins at the period of heightened CST growth. A majority of these proteins were involved in cellular assembly and organization, with annotations being most highly associated with cytoskeletal organization, microtubule dynamics, neurite outgrowth, and the formation, polymerization and quantity of microtubules. In addition, 22 proteins were more highly expressed within the developing CST in comparison to other developing white matter tracts of the spinal cord of age-matched animals. Of these differentially expressed proteins, only one, stathmin 1 (a protein known to be involved in microtubule dynamics), was both highly enriched in the developing CST and relatively sparse in other developing descending and ascending spinal tracts. Immunohistochemical analyses of the developing rat spinal cord and fetal human brain stem confirmed the enriched pattern of stathmin expression along the developing CST, and in vitro growth assays of rat corticospinal neurons showed a reduced length of neurite processes in response to pharmacological perturbation of stathmin activity. Combined, these findings suggest that stathmin activity may modulate axonal growth during development of the corticospinal projection, and reinforces the notion that microtubule dynamics could play an important role in the generation and regeneration of the CST.

  1. Evaluation and comparison of rapid methods for the detection of Salmonella in naturally contaminated pine nuts using different pre enrichment media.

    Science.gov (United States)

    Wang, Hua; Gill, Vikas S; Cheng, Chorng-Ming; Gonzalez-Escalona, Narjol; Irvin, Kari A; Zheng, Jie; Bell, Rebecca L; Jacobson, Andrew P; Hammack, Thomas S

    2015-04-01

    Foodborne outbreaks, involving pine nuts and peanut butter, illustrate the need to rapidly detect Salmonella in low moisture foods. However, the current Bacteriological Analytical Manual (BAM) culture method for Salmonella, using lactose broth (LB) as a pre enrichment medium, has not reliably supported real-time quantitative PCR (qPCR) assays for certain foods. We evaluated two qPCR assays in LB and four other pre enrichment media: buffered peptone water (BPW), modified BPW (mBPW), Universal Pre enrichment broth (UPB), and BAX(®) MP media to detect Salmonella in naturally-contaminated pine nuts (2011 outbreak). A four-way comparison among culture method, Pathatrix(®) Auto, VIDAS(®) Easy SLM, and qPCR was conducted. Automated DNA extraction techniques were compared with manual extraction methods (boiling or InstaGene™). There were no significant differences (P > 0.05) among the five pre enrichment media for pine nuts using the culture method. While both qPCR assays produced significantly (P ≤ 0.05) higher false negatives in 24 h pre enriched LB than in the other four media, they were as sensitive as the culture method in BPW, mBPW, UPB, and BAX media. The VIDAS Easy and qPCR were equivalent; Pathatrix was the least effective method. The Automatic PrepSEQ™ DNA extraction, using 1000 μL of pre enrichment, was as effective as manual extraction methods.

  2. Rapid hybridization of nucleic acids using isotachophoresis

    Science.gov (United States)

    Bercovici, Moran; Han, Crystal M.; Liao, Joseph C.; Santiago, Juan G.

    2012-01-01

    We use isotachophoresis (ITP) to control and increase the rate of nucleic acid hybridization reactions in free solution. We present a new physical model, validation experiments, and demonstrations of this assay. We studied the coupled physicochemical processes of preconcentration, mixing, and chemical reaction kinetics under ITP. Our experimentally validated model enables a closed form solution for ITP-aided reaction kinetics, and reveals a new characteristic time scale which correctly predicts order 10,000-fold speed-up of chemical reaction rate for order 100 pM reactants, and greater enhancement at lower concentrations. At 500 pM concentration, we measured a reaction time which is 14,000-fold lower than that predicted for standard second-order hybridization. The model and method are generally applicable to acceleration of reactions involving nucleic acids, and may be applicable to a wide range of reactions involving ionic reactants. PMID:22733732

  3. Correlation between the genotoxicity endpoints measured by two different genotoxicity assays: comet assay and CBMN assay

    Directory of Open Access Journals (Sweden)

    Carina Ladeira

    2015-06-01

    The results concerning of positive findings by micronuclei and non significant ones by comet assay, are corroborated by Deng et al. (2005 study performed in workers occupationally exposed to methotrexate, also a cytostatic drug. According to Cavallo et al. (2009, the comet assay seems to be more suitable for the prompt evaluation of the genotoxic effects, for instance, of polycyclic aromatic hydrocarbons mixtures containing volatile substances, whereas the micronucleus test seems more appropriate to evaluate the effects of exposure to antineoplastic agents. However, there are studies that observed an increase in both the comet assay and the micronucleus test in nurses handling antineoplastic drugs, although statistical significance was only seen in the comet assay, quite the opposite of our results (Maluf & Erdtmann, 2000; Laffon et al. 2005.

  4. "Bringing Taxonomy to the Service of Genetics": Edgar Anderson and Introgressive Hybridization.

    Science.gov (United States)

    Kleinman, Kim

    2016-12-01

    In introgressive hybridization (the repeated backcrossing of hybrids with parental populations), Edgar Anderson found a source for variation upon which natural selection could work. In his 1953 review article "Introgressive Hybridization," he asserted that he was "bringing taxonomy to the service of genetics" whereas distinguished colleagues such as Theodosius Dobzhansky and Ernst Mayr did the precise opposite. His work as a geneticist particularly focused on linkage and recombination and was enriched by collaborations with Missouri Botanical Garden colleagues interested in taxonomy as well as with cytologists C.D. Darlington and Karl Sax. As the culmination of a biosystemtatic research program, Anderson's views challenged the mainstream of the Evolutionary Synthesis.

  5. Evaluation of Enrichment Protocols for Bacterial Endosymbionts of Ciliates by Real-Time PCR.

    Science.gov (United States)

    Castelli, Michele; Lanzoni, Olivia; Rossi, Leonardo; Potekhin, Alexey; Schrallhammer, Martina; Petroni, Giulio

    2016-06-01

    Large-scale studies on obligate bacterial endosymbionts may frequently require preliminary purification and enrichment protocols, which are often elaborate to set up and to evaluate, especially if the host organism is a protist. The purpose of this study was to develop a real-time PCR-based strategy and employ it for assessing two of such enrichment protocols for Holospora caryophila, hosted by the ciliate Paramecium. Four SSU rRNA gene-targeted real-time PCR assays were designed, which allowed to compare the amount of H. caryophila to other organisms, namely the host, its food bacterium (Raoultella planticola), and free-living bacteria present in the culture medium. By the use of the real-time PCR assays in combination, it was possible to conclude that the "cell fractionation" protocol was quite successful in the enrichment of the symbiont, while the "Percoll gradient" protocol will need further refinements to be fully repeatable. The proposed approach has the potential to facilitate and encourage future studies on the yet underexplored field of bacterial endosymbionts of ciliates and other protists. It can also find valuable applications for experimental questions other than those tested, such as fast and precise assessment of symbiont abundance in natural populations and comparison among multiple coexisting symbionts.

  6. Lipase assay in soils by copper soap colorimetry.

    Science.gov (United States)

    Saisuburamaniyan, N; Krithika, L; Dileena, K P; Sivasubramanian, S; Puvanakrishnan, R

    2004-07-01

    A simple and sensitive method for the estimation of lipase activity in soils is reported. In this method, 50mg of soil is incubated with emulsified substrate, the fatty acids liberated are treated with cupric acetate-pyridine reagent, and the color developed is measured at 715 nm. Use of olive oil in this protocol leads to an estimation of true lipase activity in soils. The problem of released fatty acids getting adsorbed onto the soil colloids is obviated by the use of isooctane, and separate standards for different soils need not be developed. Among the various surfactants used for emulsification, polyvinyl alcohol is found to be the most effective. Incubation time of 20 min, soil concentration of 50 mg, pH 6.5, and incubation temperature of 37 degrees C were found to be the most suitable conditions for this assay. During the process of enrichment of the soils with oil, interference by the added oil is avoided by the maintenance of a suitable control, wherein 50 mg of soil is added after stopping the reaction. This assay is sensitive and it could be adopted to screen for lipase producers from enriched soils and oil-contaminated soils before resorting to isolation of the microbes by classical screening methods.

  7. Hybrid electric vehicles TOPTEC

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1994-06-21

    This one-day TOPTEC session began with an overview of hybrid electric vehicle technology. Updates were given on alternative types of energy storage, APU control for low emissions, simulation programs, and industry and government activities. The keynote speech was about battery technology, a key element to the success of hybrids. The TOPEC concluded with a panel discussion on the mission of hybrid electric vehicles, with a perspective from industry and government experts from United States and Canada on their view of the role of this technology.

  8. Hybrid systems with constraints

    CERN Document Server

    Daafouz, Jamal; Sigalotti, Mario

    2013-01-01

    Control theory is the main subject of this title, in particular analysis and control design for hybrid dynamic systems.The notion of hybrid systems offers a strong theoretical and unified framework to cope with the modeling, analysis and control design of systems where both continuous and discrete dynamics interact. The theory of hybrid systems has been the subject of intensive research over the last decade and a large number of diverse and challenging problems have been investigated. Nevertheless, many important mathematical problems remain open.This book is dedicated mainly to

  9. Hybrid Bloch Brane

    CERN Document Server

    Bazeia, D; Losano, L

    2016-01-01

    This work reports on models described by two real scalar fields coupled with gravity in the five-dimensional spacetime, with a warped geometry involving one infinite extra dimension. Through a mechanism that smoothly changes a thick brane into a hybrid brane, one investigates the appearance of hybrid branes hosting internal structure, characterized by the splitting on the energy density and the volcano potential, induced by the parameter which controls interactions between the two scalar fields. In particular, we investigate distinct symmetric and asymmetric hybrid brane scenarios.

  10. Hybrid Bloch brane

    Energy Technology Data Exchange (ETDEWEB)

    Bazeia, D.; Lima, Elisama E.M.; Losano, L. [Universidade Federal da Paraiba, Departamento de Fisica, Joao Pessoa, PB (Brazil)

    2017-02-15

    This work reports on models described by two real scalar fields coupled with gravity in the five-dimensional spacetime, with a warped geometry involving one infinite extra dimension. Through a mechanism that smoothly changes a thick brane into a hybrid brane, one investigates the appearance of hybrid branes hosting internal structure, characterized by the splitting on the energy density and the volcano potential, induced by the parameter which controls interactions between the two scalar fields. In particular, we investigate distinct symmetric and asymmetric hybrid brane scenarios. (orig.)

  11. Hybrid silicon evanescent devices

    Directory of Open Access Journals (Sweden)

    Alexander W. Fang

    2007-07-01

    Full Text Available Si photonics as an integration platform has recently been a focus of optoelectronics research because of the promise of low-cost manufacturing based on the ubiquitous electronics fabrication infrastructure. The key challenge for Si photonic systems is the realization of compact, electrically driven optical gain elements. We review our recent developments in hybrid Si evanescent devices. We have demonstrated electrically pumped lasers, amplifiers, and photodetectors that can provide a low-cost, scalable solution for hybrid integration on a Si platform by using a novel hybrid waveguide architecture, consisting of III-V quantum wells bonded to Si waveguides.

  12. Synthesis and reducing power assay of methyl semicarbazone derivatives

    Directory of Open Access Journals (Sweden)

    Manmohan Singhal

    2014-04-01

    Full Text Available In the present study we have designed a new pharmacophore ‘Chalconesemicarbazone’ by pharmacophore hybridization approach of drug design. A series of novel chalconesemicarbazones was synthesized and evaluated for their antioxidant activity by reducing power assay. Most of the compounds were found to be potent antioxidants. Free radicals play an important role in various pathological and xenotoxic effects so antioxidant may have protective role in these pathological conditions. Based on the results of reducing power assay 1-[1-(2,4-dihydroxyphenyl-3-(2-hydroxyphenylallylidene]-4-(4-methylphenylsemicarbazide (compound 18 and 1-[1-(2,5-dihydroxyphenyl-3-(6-hydroxyphenylallylidene]-4-(4-methylphenylsemicarbazide (compound 21 were the most active lead compounds. It was found that methoxy and hydroxyl substituted chalconesemicarbazones exhibited potent reducing power and unsubstituted compound showed less reducing potential.

  13. Characterization of hybridization between synthetic oligodeoxynucleotides and RNA in living cells.

    Science.gov (United States)

    Politz, J C; Taneja, K L; Singer, R H

    1995-01-01

    Cells internalized synthetic oligonucleotides (oligos) in culture. The hybridization of these molecules to target RNA in the living cell was subsequently detected and characterized after fixation of the cells, with or without previous detergent extraction. Hybridized oligo was distinguished from free oligo in the cell using an in situ reverse transcription technique. This assay exploited the ability of the hybridized oligo to prime synthesis of a specific cDNA strand; unhybridized oligo present in the cell could not act as a primer for reverse transcription. Phosphorothioate and fluorochrome-labeled phosphodiester oligo dT were found to enter cells rapidly and hybridize to poly (A) RNA within 30 min. Hybrids containing phosphorothioate oligo dT were detectable in cells after up to 4 h of efflux time. Phosphodiester bonded oligo dT containing covalently-linked fluorochromes appeared more stable in the cell than unmodified phosphodiester oligo dT; hybrids containing these oligos could be detected in cells as long as 18h after efflux began. The in situ transcription assay was also sensitive enough to detect hybridization of anti-actin oligos to actin mRNA in the cell. It is probable, therefore, that this assay can be used to help assess the efficacy of antisense oligos by their hybridization to a target mRNA in cells or tissues; hybridized oligos are more likely to induce a specific antisense effect. Additionally, this assay will help to identify probes that would be useful as stable hybridization tags to follow RNA movement in living cells. Images PMID:8559650

  14. Quantitative Proteomics of Zea mays Hybrids Exhibiting Different Levels of Heterosis.

    Science.gov (United States)

    Dahal, Diwakar; Newton, Kathleen J; Mooney, Brian P

    2016-08-01

    Maize hybrids exhibiting heterosis (hybrid vigor) were generated from inbred parents with increasing genetic distance. B73 was used as the common female parent in crosses with N192 (low heterosis), MO17 (high-heterosis 1), and NC350 (high-heterosis 2). Total and mitochondria-enriched proteomes were analyzed from ear shoots of field-grown hybrids and their inbred parents. GeLCMS (1D SDS-PAGE fractionation, trypsin digestion, LTQ Orbitrap nano-RP-LC MS/MS) was used to analyze proteins, and spectral counting was used for quantitation. In total, 3,568 proteins were identified and quantified in hybrids including 2,489 in the mitochondria-enriched fraction and 2,162 in the total protein fraction. Sixty-one proteins were differentially abundant (p hybrids compared with the low-heterosis hybrid. For the total proteome, eight of these showed similar trends in abundance in both of the higher-heterosis hybrids. Nine proteins showed this heterosis-correlated pattern in the mitochondrial proteome, including a mitochondria-associated target of rapamycin (TOR) protein. Although differentially abundant proteins belong to various pathways, protein, and RNA metabolism, and stress responsive proteins were the major classes changed in response to increasing heterosis.

  15. Reproductive isolation and the expansion of an invasive hybrid swarm

    Science.gov (United States)

    Blum, Michael J.; Walters, David M.; Burkhead, Noel M.; Freeman, Byron J.; Porter, Brady A.

    2010-01-01

    Biological invasions involving hybridization proceed according to prezygotic and postzygotic reproductive isolating mechanisms. Yet few comparisons of reproductive isolation have been carried out to understand how different mechanisms prevent or promote invasions involving hybridization. Here we present a study of prezygotic and postzygotic isolation between non-native red shiner (Cyprinella lutrensis) and native blacktail shiner (C. venusta stigmatura) from the Coosa River basin (USA) to better understand the formation and expansion of invasive hybrid swarms. We conducted spawning trials to measure mating preferences and raised broods from crosses to assay hybrid viability through early juvenile development. Females of both species were more responsive to conspecific mates, although blacktail shiner females responded more often to heterospecific mates than did red shiner females. Fecundity of red shiner females was also higher than blacktail shiner females. Heterospecific crosses resulted in lower fertilization and egg hatching rates, but we found no other evidence of inviability. Rather, we found comparatively low larval mortality of F1 hybrids, which is suggestive of heterosis. These findings support prior inferences of assortative mating from genetic descriptions of hybridization, and that the invasion in the Coosa River is likely proceeding due to interspecific competition and intrinsic hybrid viability.

  16. Chaotic mixer improves microarray hybridization.

    Science.gov (United States)

    McQuain, Mark K; Seale, Kevin; Peek, Joel; Fisher, Timothy S; Levy, Shawn; Stremler, Mark A; Haselton, Frederick R

    2004-02-15

    Hybridization is an important aspect of microarray experimental design which influences array signal levels and the repeatability of data within an array and across different arrays. Current methods typically require 24h and use target inefficiently. In these studies, we compare hybridization signals obtained in conventional static hybridization, which depends on diffusional target delivery, with signals obtained in a dynamic hybridization chamber, which employs a fluid mixer based on chaotic advection theory to deliver targets across a conventional glass slide array. Microarrays were printed with a pattern of 102 identical probe spots containing a 65-mer oligonucleotide capture probe. Hybridization of a 725-bp fluorescently labeled target was used to measure average target hybridization levels, local signal-to-noise ratios, and array hybridization uniformity. Dynamic hybridization for 1h with 1 or 10ng of target DNA increased hybridization signal intensities approximately threefold over a 24-h static hybridization. Similarly, a 10- or 60-min dynamic hybridization of 10ng of target DNA increased hybridization signal intensities fourfold over a 24h static hybridization. In time course studies, static hybridization reached a maximum within 8 to 12h using either 1 or 10ng of target. In time course studies using the dynamic hybridization chamber, hybridization using 1ng of target increased to a maximum at 4h and that using 10ng of target did not vary over the time points tested. In comparison to static hybridization, dynamic hybridization reduced the signal-to-noise ratios threefold and reduced spot-to-spot variation twofold. Therefore, we conclude that dynamic hybridization based on a chaotic mixer design improves both the speed of hybridization and the maximum level of hybridization while increasing signal-to-noise ratios and reducing spot-to-spot variation.

  17. Bio-bar-code functionalized magnetic nanoparticle label for ultrasensitive flow injection chemiluminescence detection of DNA hybridization.

    Science.gov (United States)

    Bi, Sai; Zhou, Hong; Zhang, Shusheng

    2009-10-07

    A signal amplification strategy based on bio-bar-code functionalized magnetic nanoparticles as labels holds promise to improve the sensitivity and detection limit of the detection of DNA hybridization and single-nucleotide polymorphisms by flow injection chemiluminescence assays.

  18. Cage-enrichment: rabbit does prefer straw or a compressed wooden block

    Directory of Open Access Journals (Sweden)

    Jorine M. Rommers

    2014-12-01

    Full Text Available The effect of different food related materials on the behaviour of commercial meat rabbit does was investigated to provide them enrichment. Five different treatments were tested. Control (pens without additional enrichment, C was compared with pens containing a pinewood stick (Pine, straw in a plastic bin (Straw, a compressed wooden block (Ply or a combination of straw and a pinewood stick (Straw+Pine. The experiment was conducted on a commercial rabbit farm using 80 cages with multiparous lactating hybrid (Hycole rabbit does. Behavioural observations were conducted in the first 4 wk of 2 successive lactations of 6 wk each, twice a week from 15:00 to 18:30 h. Once every week the consumption of gnawing materials and soiling of the cages was scored. More does were significantly occupied with Straw and Ply than with Pine (24±20, 11±9 and 4±3% of does, respectively for a longer duration (4±4, 2±2, 0.1±0.2% of observed time, respectively. In does of Straw+Pine group, the pinewood was barely touched and straw was preferred. It can be concluded that straw (loose material and wooden block are used by the animals as enrichment material to gnaw or chew on. The materials remain attractive for the 2 lactations which were measured. The pinewood stick as provided in this study was rarely used and it may be questioned whether it is sufficient as enrichment material or if it should be provided in another way than hanging on the roof of the cage. This study provides a first step towards a positive list of enrichment materials that can be used in commercial rabbit farming in The Netherlands.

  19. The effects of the formula of amino acids enriched BCAA on nutritional support in traumatic patients

    Institute of Scientific and Technical Information of China (English)

    Xin-Ying Wang; Ning Li; Jun Gu; Wei-Qin Li; Jie-Shou Li

    2003-01-01

    AIM: To investigate the formula of amino acid enriched BCAA on nutritional support in traumatic patients after operation.METHODS: 40 adult patients after moderate or large abdominal operations were enrolled in a prospective,randomly and single-blind-controlled study, and total parenteral nutrition (TPN) with either formula of amino acid (AA group, 20 cases) or formula of amino acid enriched BCAA (BCAA group, 20 cases). From the second day after operation, total parenteral nutrition was infused to the patients in both groups with equal calorie and equal nitrogen by central or peripheral vein during more than 12 hours per day for 6 days. Meanwhile, nitrogen balance was assayed by collecting 24 hours urine for 6 days. The markers of protein metabolism were investigated such as amino acid patterns, levels of total protein, albumin, prealbumin,transferrin and fibronectin in serum.RESULTS: The positive nitrogen balance in BCAA group occurred two days earlier than that in AA group. The serum levels oftotal protein and albumin in BCAA group were increased more obviously than that in AA group. The concentration of valine was notably increased and the concentration of arginine was markedly decreased in BCAA group after the formula of amino acids enriched BCAA transfusion.CONCLUSION: The formula of amino acid enriched BCAA may normalize the levels of serum amino acids, reduce the proteolysis, increase the synthesis of protein, improve the nutritional status of traumatic patients after operation.

  20. Oxidative stability and chemical safety of mayonnaise enriched with grape seed extract.

    Science.gov (United States)

    Altunkaya, Arzu; Hedegaard, Rikke V; Harholt, Jesper; Brimer, Leon; Gökmen, Vural; Skibsted, Leif H

    2013-11-01

    Grape seed extract (GSE), a by-product from the wine industry, was explored for its use as enrichment to mayonnaise, due to its potential health effects. Mayonnaises were enriched with 0 mg GSE per mL, 0.5 mg GSE per mL (~0.050%), 0.9 mg GSE per mL (~0.10%) and 1.4 mg GSE per mL (~0.15%) during preparation and stored in the dark at room temperature for 8 weeks. The antioxidative capacity of the mayonnaises was evaluated by their ability to scavenge the stable radical TEMPO by electron spin resonance (ESR) spectroscopy. The oxidative stability of the mayonnaises was determined by the content of lipid hydroperoxides (peroxide value, POV), the content of conjugated diene hydroperoxides and the content of thiobarbituric acid reactive substances (TBARS). The highest antioxidative capacity and the lowest content of lipid hydroperoxides and TBARS were found in the mayonnaise with the highest percentage of GSE (0.15%). Therefore, the oxidative stability of the mayonnaises enriched with GSE was slightly improved through storage. However, mayonnaise without GSE had the highest sensorial acceptability compared to mayonnaise enriched with GSE. In the Artemia salina assay, a fast screening method for overall toxicity, the death rate of brine shrimps larvae was found to increase for increasing percentage of GSE. A level of 0.05% GSE in mayonnaise is concluded not to constitute any toxicological risks, but to provide significant protection against oxidation during storage.

  1. Food enrichment with marine phospholipid emulsions

    DEFF Research Database (Denmark)

    Lu, Henna Fung Sieng; Nielsen, Nina Skall; Baron, Caroline P.

    . The main objective of this study was to explore the possibilities of using marine PL for food enrichment. In order to achieve the objective, the study was divided into 4 stages: i) evaluation of physico-chemical properties of marine PL emulsions, ii) evaluation of hydrolytic and oxidative stability...... of marine PL emulsions, iii) evaluation of non-enzymatic browning reactions in marine PL emulsions, iv) evaluation of sensory properties and oxidative stability of yoghurt enriched with marine PL. The obtained results showed that marine PL have good emulsifying properties and it was feasible to prepare...... marine PL emulsions with and without addition of fish oil. The oxidative stability of marine PL emulsions was significantly influenced by the chemical composition of marine PL used for emulsions preparation. For instance, emulsions with good oxidative stability could be obtained when using raw materials...

  2. Uranium enrichment activities: the SILVA program

    Energy Technology Data Exchange (ETDEWEB)

    Guyot, J.; Cazalet, J.; Camarcat, N.; Figuet, J.

    1994-12-31

    Through its commitment to a nuclear electricity generation policy, France holds today a specific position in the uranium enrichment market thanks to the modern multinational EURODIF gaseous diffusion plant. France has, altogether, a long-term goal in developing SILVA, a laser uranium enrichment process, based on the selective photo-ionization of U-235. After reviewing the fundamentals of SILVA (the laser system with copper vapor lasers and dye lasers and the separator system), a description of the general organization of the R and D program is provided going through basic research, subsystems assessment, production demonstrations and simulations (with the LACAN code), plant design and economics. The general schedule of SILVA is outlined, leading to the possible construction of a commercial plant. 7 figs., 11 refs.

  3. Challenges when developing omega-3 enriched foods

    Directory of Open Access Journals (Sweden)

    JACOBSEN Charlotte

    2010-07-01

    Full Text Available Due to the polyunsaturated nature of omega-3 fatty acids, lipid oxidation is a major challenge when developing omega-3 enriched foods. In multiphase food systems, several factors can affect lipid oxidation and efficacy of antioxidants, added to prevent lipid oxidation. This review discusses the influence of important factors such as oil quality, delivery systems for omega-3 fatty acids, processing conditions, composition of the food matrix on lipid oxidation in different omega-3 enriched foods (milk, yoghurt, mayonnaise and mayonnaise-based salads, dressing, energy bar and fish paté. Moreover, the effect of different antioxidants (tocopherol, EDTA, lactoferrin, caffeic acid, ascorbic acid, ascorbyl palmitate, propyl gallate, gallic acid, as well as lipophilized antioxidants is compared in different food systems.

  4. Development Issues on Linked Data Weblog Enrichment

    Science.gov (United States)

    Ruiz-Rube, Iván; Cornejo, Carlos M.; Dodero, Juan Manuel; García, Vicente M.

    In this paper, we describe the issues found during the development of LinkedBlog, a Linked Data extension for WordPress blogs. This extension enables to enrich text-based and video information contained in blog entries with RDF triples that are suitable to be stored, managed and exploited by other web-based applications. The issues have to do with the generality, usability, tracking, depth, security, trustiness and performance of the linked data enrichment process. The presented annotation approach aims at maintaining web-based contents independent from the underlying ontological model, by providing a loosely coupled RDFa-based approach in the linked data application. Finally, we detail how the performance of annotations can be improved through a semantic reasoner.

  5. Enriched model categories and diagram categories

    CERN Document Server

    Guillou, Bertrand

    2011-01-01

    We collect in one place a variety of known and folklore results in enriched model category theory and add a few new twists. One twist is a new perspective on equivariant model categories. A central theme is a general procedure for constructing a Quillen adjunction, often a Quillen equivalence, between a given V-model category and a category of diagrams in V, where V is any good enriching category. From this perspective, we rederive the result of Schwede and Shipley that reasonable stable model categories are Quillen equivalent to diagram categories of spectra (alias categories of module spectra). The general theory will be applied to G-spectra in a sequel, and for that we need quite a few technical improvements and modifications of general model categorical results. We collect those here. They are bound to have applications in a variety of other contexts.

  6. Challenges when developing omega-3 enriched foods

    DEFF Research Database (Denmark)

    Jacobsen, Charlotte

    2010-01-01

    Due to the polyunsaturated nature of omega-3 fatty acids, lipid oxidation is a major challenge when developing omega-3 enriched foods. In multiphase food systems, several factors can affect lipid oxidation and efficacy of antioxidants, added to prevent lipid oxidation. This review discusses...... the influence of important factors such as oil quality, delivery systems for omega-3 fatty acids, processing conditions, composition of the food matrix on lipid oxidation in different omega-3 enriched foods (milk, yoghurt, mayonnaise and mayonnaise-based salads, dressing, energy bar and fish paté). Moreover......, the effect of different antioxidants (tocopherol, EDTA, lactoferrin, caffeic acid, ascorbic acid, ascorbyl palmitate, propyl gallate, gallic acid, as well as lipophilized antioxidants) is compared in different food systems....

  7. High Purity Germanium Gamma-PHA Assay of Uranium Scrap Cans Used in 321-M Facility

    Science.gov (United States)

    Salaymeh, S. R.; Dewberry, R. A.; Casella, V.

    2001-12-01

    The Analytical Development Section of SRTC was requested by the Facilities Disposition Division (FDD) to determine the holdup of enriched uranium in the 321-M facility as part of an overall deactivation project of the facility. The 321-M facility was used to fabricate enriched uranium fuel assemblies, lithium-aluminum target tubes, neptunium assemblies, and miscellaneous components for the production reactors. The facility also includes the 324-M storage building and the passageway connecting it to 321-M. The results of the holdup assays are essential for determining compliance with the Solid Waste's Waste Acceptance Criteria, Material Control & Accountability, and to meet criticality safety controls. This report describes and documents the use of a portable HPGe detector and EG&G DART system that contains a high voltage power supply, signal processing electronics, a personal computer with Gamma-Vision software, and space to store and manipulate multiple 4096-channel gamma-ray spectra to assay for 235U content. The system was used to assay a large number of scrap cans used to store highly enriched uranium (HEU) chips and filings. This report includes a description of two efficiency calibration configurations and also the results of the assay. A description of the quality control checks is included as well.

  8. Barcoded microchips for biomolecular assays.

    Science.gov (United States)

    Zhang, Yi; Sun, Jiashu; Zou, Yu; Chen, Wenwen; Zhang, Wei; Xi, Jianzhong Jeff; Jiang, Xingyu

    2015-01-20

    Multiplexed assay of analytes is of great importance for clinical diagnostics and other analytical applications. Barcode-based bioassays with the ability to encode and decode may realize this goal in a straightforward and consistent manner. We present here a microfluidic barcoded chip containing several sets of microchannels with different widths, imitating the commonly used barcode. A single barcoded microchip can carry out tens of individual protein/nucleic acid assays (encode) and immediately yield all assay results by a portable barcode reader or a smartphone (decode). The applicability of a barcoded microchip is demonstrated by human immunodeficiency virus (HIV) immunoassays for simultaneous detection of three targets (anti-gp41 antibody, anti-gp120 antibody, and anti-gp36 antibody) from six human serum samples. We can also determine seven pathogen-specific oligonucleotides by a single chip containing both positive and negative controls.

  9. Radioreceptor assay method for insulin

    Energy Technology Data Exchange (ETDEWEB)

    Mori, K.F.; Wood, R.J. (Bureau of Drug Research, Health and Welfare Canada, Ottawa, Ontario. Health Protection Branch)

    1984-01-01

    A sensitive practical radioreceptor assay method for pharmaceutical insulin products has been developed with partially purified rat liver plasma membranes and the optimal conditions under which the best overall assay performance is obtainable have been defined. Intra- and inter-assay variations of the method averaged 7.3 and 12.2% of the man, respectively, when expressed as the coefficient of variation. Potency estimates of an insulin product obtained with the proposed method correlated well with those determined by the mouse convulsion bioassay method. Liver membranes prepared according to the method could be stored for up to ten weeks at 4/sup 0/C and for 6 months or more at -18/sup 0/C without losing insulin-binding ability.

  10. Nutritional Properties of Enriched Local Complementary Flours

    Directory of Open Access Journals (Sweden)

    W.R. Compaoré

    2011-02-01

    Full Text Available This study aimed to identify the nutritional, functional, sensory and microbiological profile of experimental nutritional flours, produced with local products in Burkina Faso. The raw materials included maize (Zea mays, millet (Pennisetum glaucum and rice (Oryza sativa. Local ingredients were pulps of Adansonia digitata and Parkia biglobosa and seeds of Cucurbita maxima and Moringa oleifera. Three formula were developed, the first (F1 with maize, the second (F2 with rice and the last (F3 with millet. Each of these cereals was mixed with predetermined portions of seeds and pulps in order to obtain enriched flour. Nutritional, microbiological and functional analysis and the acceptability criteria of these enriched flours were assessed and compared to Misola (F4, the existing local complementary flour. The fat content of experimental flours were respectively in the first (F1, second (F2 and third formula (F3 15.91±0.01%, 11.82±0.02% and 17.02±0.02%. The carbohydrate range was 65.46±0.06%, 70.81±0.01% and 64.51±0.01% for F1, F2 and F3, while the energetic value is higher than recommended (453.07±0.05, 424.56±0.03 and 458.96±0.05 kcal respectively for F1, F2 and F3. Functional characteristics indicated the good viscosity (117, 119 and 121 mm/30 sec for F1, F2 and F3 least gelation (9, 6 and 7% and water absorption capacity (2, 4 and 1 g/g. Trained sensory evaluation panellists gore the enriched flour porridge a score of acceptable. These enriched flours have great potential as a weaning food in resource-poor and technologically under-developed countries.

  11. Hybrid polymer microspheres

    Science.gov (United States)

    Rembaum, A.

    1980-01-01

    Techniques have been successfully tested for bonding polymeric spheres, typically 0.1 micron in diameter, to spheres with diameter up to 100 microns. Hybrids are being developed as improved packing material for ion-exchange columns, filters, and separators.

  12. Hybrid adsorptive membrane reactor

    Science.gov (United States)

    Tsotsis, Theodore T. (Inventor); Sahimi, Muhammad (Inventor); Fayyaz-Najafi, Babak (Inventor); Harale, Aadesh (Inventor); Park, Byoung-Gi (Inventor); Liu, Paul K. T. (Inventor)

    2011-01-01

    A hybrid adsorbent-membrane reactor in which the chemical reaction, membrane separation, and product adsorption are coupled. Also disclosed are a dual-reactor apparatus and a process using the reactor or the apparatus.

  13. Hybrid photon detectors

    CERN Document Server

    D'Ambrosio, C

    2003-01-01

    Hybrid photon detectors detect light via vacuum photocathodes and accelerate the emitted photoelectrons by an electric field towards inversely polarized silicon anodes, where they are absorbed, thus producing electron-hole pairs. These, in turn, are collected and generate electronic signals on their ohmic contacts. This review first describes the characteristic properties of the main components of hybrid photon detectors: light entrance windows, photocathodes, and silicon anodes. Then, essential relations describing the trajectories of photoelectrons in electric and magnetic fields and their backscattering from the silicon anodes are derived. Depending on their anode configurations, three families of hybrid photon detectors are presented: hybrid photomultiplier tubes with single anodes for photon counting with high sensitivity and for gamma spectroscopy; multi-anode photon detector tubes with anodes subdivided into square or hexagonal pads for position-sensitive photon detection; imaging silicon pixel array t...

  14. Functional hybrid materials

    National Research Council Canada - National Science Library

    Fahmi, Amir; Pietsch, Torsten; Mendoza, Cesar; Cheval, Nicolas

    2009-01-01

    .... This paper describes our group's achievements towards the development of multifunctional nanostructures via self-assembly of hybrid systems based on the block copolymer PS-b-P4VP and inorganic nanoparticles (NPs...

  15. Hybrid Rocket Technology

    Directory of Open Access Journals (Sweden)

    Sankaran Venugopal

    2011-04-01

    Full Text Available With their unique operational characteristics, hybrid rockets can potentially provide safer, lower-cost avenues for spacecraft and missiles than the current solid propellant and liquid propellant systems. Classical hybrids can be throttled for thrust tailoring, perform in-flight motor shutdown and restart. In classical hybrids, the fuel is stored in the form of a solid grain, requiring only half the feed system hardware of liquid bipropellant engines. The commonly used fuels are benign, nontoxic, and not hazardous to store and transport. Solid fuel grains are not highly susceptible to cracks, imperfections, and environmental temperature and are therefore safer to manufacture, store, transport, and use for launch. The status of development based on the experience of the last few decades indicating the maturity of the hybrid rocket technology is given in brief.Defence Science Journal, 2011, 61(3, pp.193-200, DOI:http://dx.doi.org/10.14429/dsj.61.518

  16. Nitrous Paraffin Hybrid Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The Nitrous Oxide Paraffin Hybrid engine (N2OP) is a proposed technology designed to provide small launch vehicles with high specific impulse, indefinitely storable...

  17. Hybrid adsorptive membrane reactor

    Science.gov (United States)

    Tsotsis, Theodore T.; Sahimi, Muhammad; Fayyaz-Najafi, Babak; Harale, Aadesh; Park, Byoung-Gi; Liu, Paul K. T.

    2011-03-01

    A hybrid adsorbent-membrane reactor in which the chemical reaction, membrane separation, and product adsorption are coupled. Also disclosed are a dual-reactor apparatus and a process using the reactor or the apparatus.

  18. Status report on the conceptual design of a commercial tokamak hybrid reactor (CTHR)

    Energy Technology Data Exchange (ETDEWEB)

    None

    1979-09-01

    A preliminary conceptual design is presented for an early twenty-first century fusion hybrid reactor called the Commercial Tokamak Hybrid Reactor (CTHR). This design was developed as a first generation commercial plant producing fissile fuel to support a significant number of client Light Water Reactor (LWR) plants. The study has been made in sufficient depth to indicate no insurmountable technical problems exist and has provided a basis for valid cost estimates of the hybrid plants as well as the hybrid/LWR system busbar electricity costs. This energy system can be optimized to have a net cost of busbar electricity that is equivalent to the conventional LWR plant, yet is not dependent on uranium ore prices or standard enrichment costs, since the fusion hybrid can be fueled by numerous fertile fuel resources.

  19. Supply of enriched uranium for research reactors

    Energy Technology Data Exchange (ETDEWEB)

    Mueller, H. [NUKEM GmbH, Alzenau (Germany)

    1997-08-01

    Since the RERTR-meeting In Newport/USA in 1990 the author delivered a series of papers in connection with the fuel cycle for research reactors dealing with its front-end. In these papers the author underlined the need for unified specifications for enriched uranium metal suitable for the production of fuel elements and made proposals with regard to the re-use of in Europe reprocessed highly enriched uranium. With regard to the fuel cycle of research reactors the research reactor community was since 1989 more concentrating on the problems of its back-end since the USA stopped the acceptance of spent research reactor fuel on December 31, 1988. Now, since it is apparent that these back-end problem have been solved by AEA`s ability to reprocess and the preparedness of the USA to again accept physically spent research reactor fuel the author is focusing with this paper again on the front-end of the fuel cycle on the question whether there is at all a safe supply of low and high enriched uranium for research reactors in the future.

  20. Enriching Earthdata by Improving Content Curation

    Science.gov (United States)

    Bagwell, R.; Wong, M. M.; Murphy, K. J.

    2014-12-01

    Since the launch of Earthdata in the later part of 2011, there has been an emphasis on improving the user experience and providing more enriched content to the user, ultimately with the focus to bring the "pixels to the people" or to ensure that a user clicks the fewest amount of times to get to the data, tools, or information which they seek. Earthdata was founded to be a single source of information for Earth Observing System Data and Information System (EOSDIS) components and services as a conglomeration between over 15 different websites. With an increased focus on access to Earth science data, the recognition is now on transforming Earthdata from a static website to one that is a dynamic, data-driven site full of enriched content.In the near future, Earthdata will have a number of components that will drive the access to the data, such as Earthdata Search, the Common Metadata Repository (CMR), and a redesign of the Earthdata website. The focus on content curation will be to leverage the use of these components to provide an enriched content environment and a better overall user experience, with an emphasis on Earthdata being "powered by EOSDIS" components and services.

  1. Inhomogeneous chemical enrichment in the Galactic Halo

    Science.gov (United States)

    Kobayashi, Chiaki

    2016-08-01

    In a galaxy, chemical enrichment takes place in an inhomogeneous fashion, and the Galactic Halo is one of the places where the inhomogeneous effects are imprinted and can be constrained from observations. I show this using my chemodynamical simulations of Milky Way type galaxies. The scatter in the elemental abundances originate from radial migration, merging/accretion of satellite galaxies, local variation of star formation and chemical enrichment, and intrinsic variation of nucleosynthesis yields. In the simulated galaxies, there is no strong age-metallicity relation. This means that the most metal-poor stars are not always the oldest stars, and can be formed in chemically unevolved clouds at later times. The long-lifetime sources of chemical enrichment such as asymptotic giant branch stars or neutron star mergers can contribute at low metallicities. The intrinsic variation of yields are important in the early Universe or metal-poor systems such as in the Galactic halo. The carbon enhancement of extremely metal-poor (EMP) stars can be best explained by faint supernovae, the low [α/Fe] ratios in some EMP stars naturally arise from low-mass (~ 13 - 15M ⊙) supernovae, and finally, the [α/Fe] knee in dwarf spheroidal galaxies can be produced by subclasses of Type Ia supernovae such as SN 2002cx-like objects and sub-Chandrasekhar mass explosions.

  2. Enriched $^{82}$Se for the LUCIFER experiment

    CERN Document Server

    Beeman, J W; Benetti, P; Cardani, L; Casali, N; Chiesa, D; Clemenza, M; Dafinei, I; Di Domizio, S; Ferroni, F; Gironi, L; Giuliani, A; Gotti, C; Laubenstein, M; Maino, M; Nagorny, S; Nisi, S; Nones, C; Orio, F; Pagnanini, L; Pattavina, L; Pessina, G; Piperno, G; Pirro, S; Previtali, E; Rusconi, C; Schäffner, K; Tomei, C; Vignati, M

    2015-01-01

    The LUCIFER project aims at deploying the first array of enriched scintillating bolometers for the investigation of neutrinoless double-beta decay of $^{82}$Se. The matrix which embeds the source is an array of ZnSe crystals, where enriched $^{82}$Se is used as decay isotope. The radiopurity of the initial components employed for manufacturing crystals, that can be operated as bolometers, is crucial for achieving a null background level in the region of interest for double-beta decay investigations. In this work, we evaluated the radioactive content in 2.5 kg of 96.3% enriched $^{82}$Se metal, measured with a high-purity germanium detector at the Gran Sasso deep underground laboratory. The limits on internal contaminations of primordial decay chain elements of $^{232}$Th, $^{238}$U and $^{235}$U are respectively: $<$61 $\\mu$Bq/kg, $<$110 $\\mu$Bq/kg and $<$74 $\\mu$Bq/kg at 90% C.L.. The extremely low-background conditions in which the measurement was carried out and the high radiopurity of the $^{82}$...

  3. On Heavy Element Enrichment in Classical Novae

    CERN Document Server

    Alexakis, A; Heger, A; Brown, E F; Dursi, L J; Truran, J W; Rosner, R; Lamb, D Q; Timmes, F X; Fryxell, B; Zingale, M A; Ricker, P M; Olson, K

    2004-01-01

    Many classical nova ejecta are enriched in CNO and Ne. Rosner et al. recently suggested that the enrichment might originate in the resonant interaction between large-scale shear flows in the accreted H/He envelope and gravity waves at the interface between the envelope and the underlying C/O white dwarf. The shear flow amplifies the waves, which eventually form cusps and break. This wave breaking injects a spray of C/O into the superincumbent H/He. Using two-dimensional simulations, we formulate a quantitative expression for the amount of C/O that can be entrained into the H/He at saturation. The fraction of the envelope that is enriched depends on the shear velocity and density contrast between the C/O white dwarf and the H/He layer but is roughly independent of the shape of the shear profile. Using this parameterization for the mixed mass, we then perform several one-dimensional Lagrangian calculations of an accreting white dwarf envelope and consider two scenarios: that the wave breaking and mixing is driv...

  4. Evaluation of environmental genotoxicity by comet assay in Columba livia.

    Science.gov (United States)

    González-Acevedo, Anahi; García-Salas, Juan A; Gosálvez, Jaime; Fernández, José Luis; Dávila-Rodríguez, Martha I; Cerda-Flores, Ricardo M; Méndez-López, Luis F; Cortés-Gutiérrez, Elva I

    2016-01-01

    The concentrations of recognized or suspected genotoxic and carcinogenic agents found in the air of large cities and, in particular, developing countries, have raised concerns about the potential for chronic health effects in the populations exposed to them. The biomonitoring of environmental genotoxicity requires the selection of representative organisms as "sentinels," as well as the development of suitable and sensitive assays, such as those aimed at assessing DNA damage. The aim of this study was to evaluate DNA damage levels in erythrocytes from Columba livia living in the metropolitan area of Monterrey, Mexico, compared with control animals via comet assay, and to confirm the results via Micronuclei test (MN) and DNA breakage detection-fluorescence in situ hybridization (DBD-FISH). Our results showed a significant increase in DNA migration in animals from the area assayed compared with that observed in control animals sampled in non-contaminated areas. These results were confirmed by MN test and DBD-FISH. In conclusion, these observations confirm that the examination of erythrocytes from Columba livia via alkaline comet assay provides a sensitive and reliable end point for the detection of environmental genotoxicants.

  5. Allosteric indicator displacement enzyme assay for a cyanogenic glycoside.

    Science.gov (United States)

    Jose, D Amilan; Elstner, Martin; Schiller, Alexander

    2013-10-18

    Indicator displacement assays (IDAs) represent an elegant approach in supramolecular analytical chemistry. Herein, we report a chemical biosensor for the selective detection of the cyanogenic glycoside amygdalin in aqueous solution. The hybrid sensor consists of the enzyme β-glucosidase and a boronic acid appended viologen together with a fluorescent reporter dye. β-Glucosidase degrades the cyanogenic glycoside amygdalin into hydrogen cyanide, glucose, and benzaldehyde. Only the released cyanide binds at the allosteric site of the receptor (boronic acid) thereby inducing changes in the affinity of a formerly bound fluorescent indicator dye at the other side of the receptor. Thus, the sensing probe performs as allosteric indicator displacement assay (AIDA) for cyanide in water. Interference studies with inorganic anions and glucose revealed that cyanide is solely responsible for the change in the fluorescent signal. DFT calculations on a model compound revealed a 1:1 binding ratio of the boronic acid and cyanide ion. The fluorescent enzyme assay for β-glucosidase uses amygdalin as natural substrate and allows measuring Michaelis-Menten kinetics in microtiter plates. The allosteric indicator displacement assay (AIDA) probe can also be used to detect cyanide traces in commercial amygdalin samples. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Hybridity in Disgrace

    Institute of Scientific and Technical Information of China (English)

    刘建平

    2015-01-01

    John Maxwell Coetzee's masterpiece-Disgrace is the representative work about post colonialism.The novel describes a series of disgraceful events happened between the white and the black in the post apartheid South Africa.The famous literature theory-hybridity of Homi K.Bhabha is the very key theory to analyze the work.In post apartheid South Africa,hybridity is the only way for the white and the black to coexist.

  7. Hybrid Baryon Signatures

    CERN Document Server

    Page, P R

    2000-01-01

    We discuss whether a low-lying hybrid baryon should be defined as a three quark - gluon bound state or as three quarks moving on an excited adiabatic potential. We show that the latter definition becomes exact, not only for very heavy quarks, but also for specific dynamics. We review the literature on the signatures of hybrid baryons, with specific reference to strong hadronic decays, electromagnetic couplings, diffractive production and production in psi decay.

  8. Hybrid vertical cavity laser

    DEFF Research Database (Denmark)

    Chung, Il-Sug; Mørk, Jesper

    2010-01-01

    A new hybrid vertical cavity laser structure for silicon photonics is suggested and numerically investigated. It incorporates a silicon subwavelength grating as a mirror and a lateral output coupler to a silicon ridge waveguide.......A new hybrid vertical cavity laser structure for silicon photonics is suggested and numerically investigated. It incorporates a silicon subwavelength grating as a mirror and a lateral output coupler to a silicon ridge waveguide....

  9. Requirements for Hybrid Cosimulation

    Science.gov (United States)

    2014-08-16

    hybrid cosimulation version of the Functional Mockup Interface (FMI) standard. A cosimulation standard de nes interfaces that enable diverse simulation...cosimulation standards, and specifically provides guidance for development of a hybrid cosimulation version of the Functional Mockup Interface (FMI) standard...V. Peetz, and S. Wolf. The functional mockup interface for tool independent exchange of simulation models. In Proc. of the 8-th International

  10. Enrichment, isolation and characterization of fungi tolerant to 1-ethyl-3-methylimidazolium acetate

    Energy Technology Data Exchange (ETDEWEB)

    Singer, S.W.; Reddy, A. P.; Gladden, J. M.; Guo, H.; Hazen, T.C.; Simmons, B. A.; VanderGheynst, J. S.

    2010-12-15

    This work aims to characterize microbial tolerance to 1-ethyl-3-methylimidazolium acetate ([C2mim][OAc]), ionic liquid that has emerged as a novel biomass pretreatment for lignocellulosic biomass. Enrichment experiments performed using inocula treated with [C2mim][OAc] under solid and liquid cultivation yielded fungal populationsdominated by Aspergilli. Ionic liquid-tolerant Aspergillus isolates from these enrichments were capable of growing in a radial plate growth assay in the presence of 10% [C2mim][OAc]. When a [C2mim][OAc]-tolerant Aspergillus fumigatus strain was grown in the presence of switchgrass, endoglucanases and xylanases were secreted that retained residual enzymatic activity in the presence of 20% [C2mim][OAc]. The results of the study suggest tolerance to ionic liquids is a general property of Aspergilli. Tolerance to an industrially important ionic liquid was discovered in a fungal genera that is widely used in biotechnology, including biomass deconstruction.

  11. A cleavable biotin tagging reagent that enables the enrichment and identification of carbonylation sites in proteins.

    Science.gov (United States)

    Coffey, Chelsea M; Gronert, Scott

    2016-01-01

    The utility of a new, cleavable tag for identifying and enriching protein carbonyls is examined. Using a model system, human serum albumin modified with acrolein, the EZ-Link alkoxyamine-PEG4-SS-PEG4-biotin affinity tag, was tested for its ability to label protein carbonyls in proteomic analyses of protein carbonylation. The efficiency of the labeling was assayed and compared to standard biotin hydrazide reagents. The label was also tested in liquid chromatography-tandem mass spectrometry (LC/MS/MS) experiments. The quality of the fragmentation spectra was assessed and the relative detection efficiency of various modification sites was compared to standard biotin hydrazide reagents. Finally, the viability of using the label with streptavidin bead enrichment protocols in a standard proteomics workflow was probed.

  12. Assortative mating and the maintenance of population structure in a natural hybrid zone.

    Science.gov (United States)

    Culumber, Zachary W; Ochoa, Olivia M; Rosenthal, Gil G

    2014-08-01

    Understanding the factors that give rise to natural hybrid zones and govern their dynamics and structure is important to predicting the evolutionary consequences of hybridization. Here we use a combination of multigenerational population genetic data, mating patterns from a natural population, behavioral assays, and mark-recapture data within clinal hybrid zones of the genus Xiphophorus to test the role of assortative mating in maintaining population structure and the potential for ongoing genetic exchange between heterospecifics. Our data demonstrate that population structure is temporally robust and driven largely by assortative mating stemming from precopulatory isolation between pure species. Furthermore, mark-recapture data revealed that rates of migration within the same stream reach are far below the level needed to support population structure. In contrast to many empirical studies of natural hybrid zones, there appeared to be no hybrid male dysfunction or discrimination against hybrid males by pure parental females, and hybrid females mated and associated with pure species and hybrid males at random. Despite strong isolation between pure parentals, hybrids therefore can act as a conduit for genetic exchange between heterospecifics, which has been shown to increase the tempo of evolutionary change. Additionally, our findings highlight the complexity of natural hybrid zone dynamics, demonstrating that sexual and ecological selection together can give rise to patterns that do not fit classical models of hybrid zone evolution.

  13. A Resolution of the Paradox of Enrichment

    Science.gov (United States)

    Feng, Z. C.; Li, Y. Charles

    2015-06-01

    The paradox of enrichment was observed by Rosenzweig [1971] in a class of predator-prey models. Two of the parameters in the models are crucial for the paradox. These two parameters are the prey's carrying capacity and prey's half-saturation for predation. Intuitively, increasing the carrying capacity due to enrichment of the prey's environment should lead to a more stable predator-prey system. Analytically, it turns out that increasing the carrying capacity always leads to an unstable predator-prey system that is susceptible to extinction from environmental random perturbations. This is the so-called paradox of enrichment. Our resolution here rests upon a closer investigation on a dimensionless number H formed from the carrying capacity and the prey's half-saturation. By recasting the models into dimensionless forms, the models are in fact governed by a few dimensionless numbers including H. The effects of the two parameters: carrying capacity and half-saturation are incorporated into the number H. In fact, increasing the carrying capacity is equivalent (i.e. has the same effect on H) to decreasing the half-saturation which implies more aggressive predation. Since there is no paradox between more aggressive predation and instability of the predator-prey system, the paradox of enrichment is resolved. The so-called instability of the predator-prey system is characterized by the existence of a stable limit cycle in the phase plane, which gets closer and closer to the predator axis and prey axis. Due to random environmental perturbations, this can lead to extinction. We also further explore spatially dependent models for which the phase space is infinite-dimensional. The spatially independent limit cycle which is generated by a Hopf bifurcation from an unstable steady state, is linearly stable in the infinite-dimensional phase space. Numerical simulations indicate that the basin of attraction of the limit cycle is riddled. This shows that spatial perturbations can

  14. Proteomic analysis of heterosis in the leaves of sorghum-sudangrass hybrids.

    Science.gov (United States)

    Han, Pingan; Lu, Xiaoping; Mi, Fugui; Dong, Jing; Xue, Chunlei; Li, Jianke; Han, Bin; Zhang, Xiaoyu

    2016-02-01

    Sorghum-sudangrass hybrids are widely used for forage and silage in the animal husbandry industry due to their hardiness. The heterozygous first generation of sorghum-sudangrass hybrids displays performance superior to their homozygous, parental inbred lines. In order to study the molecular details underlying its heterosis, the leaves of sorghum-sudangrass hybrids and their parents were compared using mass spectrometry-based proteomics. Results showed that among the 996 proteins that were identified, 32 proteins showed 'additive accumulation expression patterns', indicating that the protein abundance in sorghum-sudangrass hybrids showed no significant difference from the average of their parents. Additionally, 74 proteins showed 'nonadditive accumulation expression patterns' (the proteins abundance in the hybrids showed significant difference from the average of their parents). Both additive and nonadditive proteins were mainly involved in photosynthesis and carbohydrate metabolism. More upregulated additive and nonadditive proteins were in the hybrids than in their parents, suggesting that additive and nonadditive proteins are essential to the vigor of sorghum-sudangrass hybrids. The nonadditive proteins were enriched in photosynthesis, carbohydrate metabolism, and protein oligomerization, but the additive proteins were not enriched in any pathway, which indicated that the nonadditive proteins could be greater contributors to heterosis than additive proteins. Furthermore, the highly activated photosynthetic pathway in nonadditive proteins implies that photosynthesis in hybrids is heightened to assimilate more organic matter, resulting in an increased yield. Our results provide a proof-of-concept that reveals the molecular components of heterosis in sorghum-sudangrass hybrid leaves and serves as an important step for future genetic manipulation of specific proteins to improve the performance of hybrids.

  15. Filtration parameters influencing circulating tumor cell enrichment from whole blood.

    Directory of Open Access Journals (Sweden)

    Frank A W Coumans

    Full Text Available Filtration can achieve circulating tumor cell (CTC enrichment from blood. Key parameters such as flow-rate, applied pressure, and fixation, vary largely between assays and their influence is not well understood. Here, we used a filtration system, to monitor these parameters and determine their relationships. Whole blood, or its components, with and without spiked tumor cells were filtered through track-etched filters. We characterize cells passing through filter pores by their apparent viscosity; the viscosity of a fluid that would pass with the same flow. We measured a ratio of 5·10(4∶10(2∶1 for the apparent viscosities of 15 µm diameter MDA-231 cells, 10 µm white cells and 90 fl red cells passing through a 5 µm pore. Fixation increases the pressure needed to pass cells through 8 µm pores 25-fold and halves the recovery of spiked tumor cells. Filtration should be performed on unfixed samples at a pressure of ∼10 mbar for a 1 cm(2 track-etched filter with 5 µm pores. At this pressure MDA-231 cells move through the filter in 1 hour. If fixation is needed for sample preservation, a gentle fixative should be selected. The difference in apparent viscosity between CTC and blood cells is key in optimizing recovery of CTC.

  16. Quality assurance in the enriched uranium operations NDA facility

    Energy Technology Data Exchange (ETDEWEB)

    May, P.K.; Ceo, R.N. [Oak Ridge Y-12 Plant, TN (United States)

    1997-11-01

    The Nondestructive Analysis (NDA) Facility at the Oak Ridge Y-12 Plant has characterized process wastes for Enriched Uranium Operations since 1978. Since that time, over 50,000 items have been analyzed. Analysis results are used to determine whether or not recovery of uranium from process wastes is economically feasible. Our instrument complement includes one large segmented gamma scanner (SGS), two smaller SGS, two solution assay systems (SAS), and Active Well Coincidence Counter (AWCC). The large SGS is used for analyzing High Efficiency Particulate Air (HEPA) filters ant 208-L drums filled with combustible contaminated waste. The smaller SGS are used to analyze 4-L containers of ash and leached residues. The SAS are used to analyze 125 ml bottles of aqueous or organic waste solutions that may contain uranium. The gamma-based NDA techniques are used to identify which process wastes can be discarded, and which must be recycled. The AWCC is used to analyze high-density materials which are not amenable to gamma-ray analysis. 1 ref., 4 figs.

  17. Chromosome aberration assays in Allium

    Energy Technology Data Exchange (ETDEWEB)

    Grant, W.F.

    1982-01-01

    The common onion (Allium cepa) is an excellent plant for the assay of chromosome aberrations after chemical treatment. Other species of Allium (A. cepa var. proliferum, A. carinatum, A. fistulosum and A. sativum) have also been used but to a much lesser extent. Protocols have been given for using root tips from either bulbs or seeds of Allium cepa to study the cytological end-points, such as chromosome breaks and exchanges, which follow the testing of chemicals in somatic cells. It is considered that both mitotic and meiotic end-points should be used to a greater extent in assaying the cytogenetic effects of a chemical. From a literature survey, 148 chemicals are tabulated that have been assayed in 164 Allium tests for their clastogenic effect. Of the 164 assays which have been carried out, 75 are reported as giving a positive reaction, 49 positive and with a dose response, 1 positive and temperature-related, 9 borderline positive, and 30 negative; 76% of the chemicals gave a definite positive response. It is proposed that the Allium test be included among those tests routinely used for assessing chromosomal damage induced by chemicals.

  18. 21 CFR 866.2330 - Enriched culture medium.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Enriched culture medium. 866.2330 Section 866.2330... medium. (a) Identification. An enriched culture medium is a device that consists primarily of liquid or... medium enriched by the addition of such nutritional components as blood, blood serum, vitamins, and...

  19. 21 CFR 139.122 - Enriched nonfat milk macaroni products.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 2 2010-04-01 2010-04-01 false Enriched nonfat milk macaroni products. 139.122... Macaroni and Noodle Products § 139.122 Enriched nonfat milk macaroni products. (a) Each of the enriched macaroni products made with nonfat milk for which a definition and standard of identity is prescribed...

  20. 78 FR 75579 - Low Enriched Uranium From France

    Science.gov (United States)

    2013-12-12

    ... COMMISSION Low Enriched Uranium From France Determination On the basis of the record \\1\\ developed in the... antidumping duty order on low enriched uranium from France would be likely to lead to continuation or...), entitled Low Enriched Uranium from France: Investigation No. 731-TA-909 (Second Review). By order of...

  1. 31 CFR 540.308 - Low Enriched Uranium (LEU).

    Science.gov (United States)

    2010-07-01

    ... 31 Money and Finance: Treasury 3 2010-07-01 2010-07-01 false Low Enriched Uranium (LEU). 540.308... OF FOREIGN ASSETS CONTROL, DEPARTMENT OF THE TREASURY HIGHLY ENRICHED URANIUM (HEU) AGREEMENT ASSETS CONTROL REGULATIONS General Definitions § 540.308 Low Enriched Uranium (LEU). The term low...

  2. 31 CFR 540.306 - Highly Enriched Uranium (HEU).

    Science.gov (United States)

    2010-07-01

    ... 31 Money and Finance: Treasury 3 2010-07-01 2010-07-01 false Highly Enriched Uranium (HEU). 540...) OFFICE OF FOREIGN ASSETS CONTROL, DEPARTMENT OF THE TREASURY HIGHLY ENRICHED URANIUM (HEU) AGREEMENT ASSETS CONTROL REGULATIONS General Definitions § 540.306 Highly Enriched Uranium (HEU). The term...

  3. A Hybrid Approach for Co-Channel Speech Segregation based on CASA, HMM Multipitch Tracking, and Medium Frame Harmonic Model

    Directory of Open Access Journals (Sweden)

    Ashraf M. Mohy Eldin

    2013-08-01

    Full Text Available This paper proposes a hybrid approach for co-channel speech segregation. HMM (hidden Markov model is used to track the pitches of 2 talkers. The resulting pitch tracks are then enriched with the prominent pitch. The enriched tracks are correctly grouped using pitch continuity. Medium frame harmonics are used to extract the second pitch for frames with only one pitch deduced using the previous steps. Finally, the pitch tracks are input to CASA (computational auditory scene analysis to segregate the mixed speech. The center frequency range of the gamma tone filter banks is maximized to reduce the overlap between the channels filtered for better segregation. Experiments were conducted using this hybrid approach on the speech separation challenge database and compared to the single (non-hybrid approaches, i.e. signal processing and CASA. Results show that using the hybrid approach outperforms the single approaches.

  4. Synthesis and osteo-compatibility of novel reduced graphene oxide-aminosilica hybrid nanosheets.

    Science.gov (United States)

    Chen, Song; Du, Xinxin; Jia, Lan; Chang, Haixin; Ikoma, Toshiyuki; Hanagata, Nobutaka

    2016-04-01

    Combination of silica component with other materials is one of the current strategies to design bone regenerative materials. In this study, novel reduced graphene oxide (RGO)-aminosilica hybrid nanosheets with enhanced osteo-compatibility were synthesized from a mixture of 3-aminopropyltriethoxysilane (APTES), graphene oxides (GO) and water. The presence of APTES in the mixture not only caused the conversion of GO to RGO, but also led to the hydrolysis and condensation of itself. It was for the first time reported the reducing role of APTES in the conversion of GO to RGO. It was found that the silicon (IV) ions were released from the hybrid nanosheets in a sustained way. The in vitro osteo-compatibility was evaluated by incubating the hybrid nanosheets with osteoblast MC3T3-E1 cells. A water soluble tetrazolium salt assay quantitatively indicated that the hybrid nanosheets had no significant toxicity and exhibited good biocompatibility. An alkaline phosphatase assay quantitatively indicated that the hybrid nanosheets enhanced the osteoblast differentiation compared to the GO nanosheets. An immunochemical assay further qualitatively indicated that the hybrid nanosheets stimulated the production of osteopontin as typical marker for osteoblast differentiation. Thus, the resultant hybrids nanosheets had a potential application in the bone regeneration.

  5. Effect of environmental enrichment and group size on behaviour and live weight in growing rabbits

    Directory of Open Access Journals (Sweden)

    Daniela Zucca

    2012-07-01

    Full Text Available The aim of this research was to study the effects of group size and environmental enrichment on behaviour and growth of 108 hybrid growing rabbits (Oryctolagus cuniculus. We compared the behaviour (time budget and reactions to specific behavioural tests: “tonic immobility” and “emergence test” and live weight of growing rabbits housed in cages with a different number of rabbits per cage (2, 3 and 4; same density:14 rabbits/m2. Half of the cages were enriched with a wooden stick (Robinia Pseudoacacia, length: 20 cm – diameter: 6 cm, cylindrical hanging from the ceiling of the cage. The stick and number of animals per cage had no effect on weight gain or on behavioural tests responses. Interaction with the stick was significantly higher at the beginning of the growing period. Principal component analysis performed on the data for the whole period showed significant differences according to the treatments: increasing the number of rabbits per cage and introducing a wooden stick seemed to affect locomotor activity frequency and social interactions. Rabbits housed 3 and 4 per cage showed less lying behaviour and higher locomotor activity and sitting. The larger functional space allowance enabled rabbits to perform more natural behaviours compared to smaller cages (2 rabbits/cage. Environmental enrichment seems to be related to higher allogrooming behaviour frequency, which could indicate a social behaviour related to pheromonal olfactory stimulation and mutual recognition.

  6. Sequence analysis of trinucleotide repeat microsatellites from an enrichment library of the equine genome.

    Science.gov (United States)

    Tozaki, T; Inoue, S; Mashima, S; Ohta, M; Miura, N; Tomita, M

    2000-04-01

    Microsatellites are useful tools for the construction of a linkage map and parentage testing of equines, but only a limited number of equine microsatellites have been elucidated. Thus, we constructed the equine genomic library enriched for DNA fragments containing (CAG)n repeats. The enriched method includes hybridization-capture of repeat regions using biotin-conjugated oligonucleotides, nucleotide substrate-biased polymerase reaction with the oligonucleotides and subsequent PCR amplification, because these procedures are useful for the cloning of less abundant trinucleotide microsatellites. Microsatellites containing (CAG)n repeats were obtained at the ratio of one per 3-4 clones, indicating an enrichment value about 10(4)-fold, resulting in less time consumption and less cost for cloning. In this study, 66 different microsatellites, (CAG)n repeats, were identified. The number of complete simple CAG repeats in our clones ranged 4-33, with an average repeat length of 8.8 units. The microsatellites were useful as sequence-tagged site (STS) markers. In addition, some clones containing (CAG)n repeats showed homology to human (CAG)n-containing genes, which have been previously mapped. These results indicate that the clones might be a useful tool for chromosome comparison between equines and humans.

  7. Impact of responder definition on the enriched enrollment randomized withdrawal trial design for establishing proof of concept in neuropathic pain.

    Science.gov (United States)

    Hewitt, David J; Ho, Tony W; Galer, Bradley; Backonja, Miroslav; Markovitz, Paul; Gammaitoni, Arnold; Michelson, David; Bolognese, James; Alon, Achilles; Rosenberg, Elizabeth; Herman, Gary; Wang, Hao

    2011-03-01

    The objective of this study was to evaluate how enrichment for responders increases assay sensitivity in an enriched enrollment randomized withdrawal (EERW) proof-of-concept (POC) study in neuropathic pain. Adults with moderate to severe peripheral neuropathic pain entered a 3- to 4-day screening period, followed by a 12-day titration to the highest tolerated dose that provided pain control (pregabalin 50-200mg t.i.d.), and then a 9-day maintenance period. Subjects were stratified as primary responders (⩾30%), secondary responders (⩾10% to proof-of-concept study to optimize assay sensitivity and efficiently determine the analgesic potential of a new treatment for neuropathic pain. Copyright © 2010 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  8. Isolation of circulating tumor cells by immunomagnetic enrichment and fluorescence-activated cell sorting (IE/FACS) for molecular profiling.

    Science.gov (United States)

    Magbanua, Mark Jesus M; Park, John W

    2013-12-01

    Circulating tumor cells (CTCs) are cells shed by the primary tumor into the blood stream capable of initiating distant metastasis. In the past decade, numerous assays have been developed to reliably detect these extremely rare cells. However, methods for purification of CTCs with little or no contamination of normal blood cells for molecular profiling are limited. We have developed a novel protocol to isolate CTCs by combining immunomagnetic enrichment and fluorescence-activated cell sorting (IE/FACS). The two-part assay includes (1) immunomagnetic capture using magnetic beads conjugated to monoclonal antibody against an epithelial cell adhesion marker (EpCAM) to enrich for tumor cells; and (2) FACS analysis using EpCAM to purify tumor cells away from mononuclear cells of hematopoietic lineage. Downstream molecular analyses of single and pooled cells confirmed the isolation of highly pure CTCs with characteristics typical that of malignant cells.

  9. Expanded genetic codes in next generation sequencing enable decontamination and mitochondrial enrichment.

    Directory of Open Access Journals (Sweden)

    Kevin J McKernan

    Full Text Available We have developed a PCR method, coined Déjà vu PCR, that utilizes six nucleotides in PCR with two methyl specific restriction enzymes that respectively digest these additional nucleotides. Use of this enzyme-and-nucleotide combination enables what we term a "DNA diode", where DNA can advance in a laboratory in only one direction and cannot feedback into upstream assays. Here we describe aspects of this method that enable consecutive amplification with the introduction of a 5th and 6th base while simultaneously providing methylation dependent mitochondrial DNA enrichment. These additional nucleotides enable a novel DNA decontamination technique that generates ephemeral and easy to decontaminate DNA.

  10. 21 CFR 225.158 - Laboratory assays.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State...

  11. A metaphor-enriched social cognition.

    Science.gov (United States)

    Landau, Mark J; Meier, Brian P; Keefer, Lucas A

    2010-11-01

    Social cognition is the scientific study of the cognitive events underlying social thought and attitudes. Currently, the field's prevailing theoretical perspectives are the traditional schema view and embodied cognition theories. Despite important differences, these perspectives share the seemingly uncontroversial notion that people interpret and evaluate a given social stimulus using knowledge about similar stimuli. However, research in cognitive linguistics (e.g., Lakoff & Johnson, 1980) suggests that people construe the world in large part through conceptual metaphors, which enable them to understand abstract concepts using knowledge of superficially dissimilar, typically more concrete concepts. Drawing on these perspectives, we propose that social cognition can and should be enriched by an explicit recognition that conceptual metaphor is a unique cognitive mechanism that shapes social thought and attitudes. To advance this metaphor-enriched perspective, we introduce the metaphoric transfer strategy as a means of empirically assessing whether metaphors influence social information processing in ways that are distinct from the operation of schemas alone. We then distinguish conceptual metaphor from embodied simulation--the mechanism posited by embodied cognition theories--and introduce the alternate source strategy as a means of empirically teasing apart these mechanisms. Throughout, we buttress our claims with empirical evidence of the influence of metaphors on a wide range of social psychological phenomena. We outline directions for future research on the strength and direction of metaphor use in social information processing. Finally, we mention specific benefits of a metaphor-enriched perspective for integrating and generating social cognitive research and for bridging social cognition with neighboring fields.

  12. Nitrogen enrichment regulates calcium sources in forests.

    Science.gov (United States)

    Hynicka, Justin D; Pett-Ridge, Julie C; Perakis, Steven S

    2016-12-01

    Nitrogen (N) is a key nutrient that shapes cycles of other essential elements in forests, including calcium (Ca). When N availability exceeds ecosystem demands, excess N can stimulate Ca leaching and deplete Ca from soils. Over the long term, these processes may alter the proportion of available Ca that is derived from atmospheric deposition vs. bedrock weathering, which has fundamental consequences for ecosystem properties and nutrient supply. We evaluated how landscape variation in soil N, reflecting long-term legacies of biological N fixation, influenced plant and soil Ca availability and ecosystem Ca sources across 22 temperate forests in Oregon. We also examined interactions between soil N and bedrock Ca using soil N gradients on contrasting basaltic vs. sedimentary bedrock that differed 17-fold in underlying Ca content. We found that low-N forests on Ca-rich basaltic bedrock relied strongly on Ca from weathering, but that soil N enrichment depleted readily weatherable mineral Ca and shifted forest reliance toward atmospheric Ca. Forests on Ca-poor sedimentary bedrock relied more consistently on atmospheric Ca across all levels of soil N enrichment. The broad importance of atmospheric Ca was unexpected given active regional uplift and erosion that are thought to rejuvenate weathering supply of soil minerals. Despite different Ca sources to forests on basaltic vs. sedimentary bedrock, we observed consistent declines in plant and soil Ca availability with increasing N, regardless of the Ca content of underlying bedrock. Thus, traditional measures of Ca availability in foliage and soil exchangeable pools may poorly reflect long-term Ca sources that sustain soil fertility. We conclude that long-term soil N enrichment can deplete available Ca and cause forests to rely increasingly on Ca from atmospheric deposition, which may limit ecosystem Ca supply in an increasingly N-rich world.

  13. Work-family enrichment and psychological health

    Directory of Open Access Journals (Sweden)

    Ameeta Jaga

    2013-03-01

    Full Text Available Orientation: This study examines the beneficial aspects of the interface between work and family and its relationships with psychological health from a positive psychology perspective.Research purpose: The objective of this study was to investigate whether work-family enrichment helps to predict psychological health, specifically increased subjective well-being and decreased feelings of emotional exhaustion and depression.Motivation for the study: The burgeoning literature on the work-family interface contains little on the potentially positive benefits of maintaining work and family roles.Research approach, design and method: The authors used a descriptive research design. Employees in two national organisations in the financial retail and logistics industries completed a self-administered survey questionnaire. The authors analysed responses from those who reported both family and work responsibilities (N = 160.Main findings: Consistent with previous research, factor analysis revealed two distinct directions of work-family enrichment: from work to family (W2FE and from family to work (F2WE. Multiple regression analysis showed that F2WE explained a significant proportion of the variance in subjective wellbeing, whilst W2FE explained a significant proportion of the variance in depression and emotional exhaustion.Practical/managerial implications: The findings of this study revealed the individual and organisational benefits of fostering work-family enrichment. Contributions/value add: This study presents empirical evidence for the need to focus on the positive aspects of the work-family interface, provides further support for a positive organisational psychology perspective in organisations and hopefully will encourage further research on interventions in organisations and families.

  14. Nitrogen enrichment regulates calcium sources in forests

    Science.gov (United States)

    Hynicka, Justin D.; Pett-Ridge, Julie C; Perakis, Steven

    2016-01-01

    Nitrogen (N) is a key nutrient that shapes cycles of other essential elements in forests, including calcium (Ca). When N availability exceeds ecosystem demands, excess N can stimulate Ca leaching and deplete Ca from soils. Over the long term, these processes may alter the proportion of available Ca that is derived from atmospheric deposition vs. bedrock weathering, which has fundamental consequences for ecosystem properties and nutrient supply. We evaluated how landscape variation in soil N, reflecting long-term legacies of biological N fixation, influenced plant and soil Ca availability and ecosystem Ca sources across 22 temperate forests in Oregon. We also examined interactions between soil N and bedrock Ca using soil N gradients on contrasting basaltic vs. sedimentary bedrock that differed 17-fold in underlying Ca content. We found that low-N forests on Ca-rich basaltic bedrock relied strongly on Ca from weathering, but that soil N enrichment depleted readily weatherable mineral Ca and shifted forest reliance toward atmospheric Ca. Forests on Ca-poor sedimentary bedrock relied more consistently on atmospheric Ca across all levels of soil N enrichment. The broad importance of atmospheric Ca was unexpected given active regional uplift and erosion that are thought to rejuvenate weathering supply of soil minerals. Despite different Ca sources to forests on basaltic vs. sedimentary bedrock, we observed consistent declines in plant and soil Ca availability with increasing N, regardless of the Ca content of underlying bedrock. Thus, traditional measures of Ca availability in foliage and soil exchangeable pools may poorly reflect long-term Ca sources that sustain soil fertility. We conclude that long-term soil N enrichment can deplete available Ca and cause forests to rely increasingly on Ca from atmospheric deposition, which may limit ecosystem Ca supply in an increasingly N-rich world.

  15. Modeling Profiles and Signatures of Enrichments

    Science.gov (United States)

    Ali, A.; Qualls, C.; Lucas, S. G.; Lombari, G.; Appenzeller, O.

    2014-12-01

    Anthropogenic and geochemical enrichment of soils and living matter have been well documented 1, 2, 3.Here we report on geochemical, anthropogenic and biological enrichments with heavy metals in Modern Peru and compared this to Modern and ancient data from New Mexico, USA. We established a signature derived from the quantities of 25 metals in various biological, fossil and soil materials. We also speculate that human adaptation to mercury toxicity may occur in remarkably short time spans during the Holocene. We found mercury concentrations in Modern pigeon feathers and llama wool from free foraging birds and animals in Albuquerque, NM, ranging from 0.006 to 0.019 mg/Kg of tissue. The values for Modern Peru ranged from 22.0 to 556 mg/Kg for the same tissues. We discovered, in 64 million-year-old fossilized plants from New Mexico (Paleocene Nacimiento Formation, San Juan Basin), a mercury concentration of 1.11 mg/Kg of fossil, whereas Modern plant material from the Rio Grande Basin in New Mexico contained no mercury. Profiling of metal content of these samples suggests that mercury is a proxy for anthropogenic rather than geochemical enrichment in the localities we examined. We found no overt signs of mercury toxicity in contemporaneous inhabitants of Huancavelica4, Peru; one of the ten most mercury-polluted places in the world and the mercury concentration in their hair is well below modern admissible levels. However, assessment of their annual scalp hair growth-rate showed marked reduction in growth (~ 5cm/yr) versus ~ 16cm/year for normal scalp hair from other continents4. This is consistent with a toxic effect of heavy metals on human metabolism and especially autonomic nervous system function in Huancavelica, Peru. Contemporaneous anthropogenic activities are known to increase heavy metal content in the biosphere with potentially toxic effects on humans. However, signs of human evolutionary adaptation to such toxins might already be evident in Peru4.

  16. Cheese bread enriched with biofortified cowpea flour

    Directory of Open Access Journals (Sweden)

    Rodrigo Barbosa Monteiro Cavalcante

    2016-02-01

    Full Text Available ABSTRACT The development and enrichment of food are of great importance not only for the industry but also to improve the population's nutrition, where you can create new products or optimize existing ones. The consumption of fortified products in the diet is an option for the control of deficiency diseases. This study aimed to develop enriched cheese bread with whole biofortified cowpea flour and evaluate their acceptance and chemical composition. Two formulations, F1 and F2, were prepared containing 5.6 and 8% of cowpea flour as a substitute for starch, respectively. To check acceptance, three sensory tests were used (Hedonic Scale, Purchase Intent, and Paired Comparison, F1 being sensory viable according to assessors, being chemically analyzed. Minerals were determined by atomic emission spectrometry with inductively coupled plasma source. The moisture was determined by drying at 105 °C, ash by calcination in muffle at 550 °C, proteins by the macro-Kjeldahl method, and lipids by hot extraction in a Soxhlet extractor. Carbohydrates were obtained by difference and the calories were calculated. The addition of cowpea increased the amounts of copper, iron, phosphorus, magnesium, manganese, and zinc, as well as protein and carbohydrate values. On the other hand, there was a reduction of the moisture concerning lipids and the total caloric value compared to the standard formulation. It was concluded , therefore, that the cowpea, a regional raw material in market expansion is presented as an option for the enrichment of baked foods that do not contain gluten, such as cheese bread.

  17. Inference of subgenomic origin of BACs in an interspecific hybrid sugarcane cultivar by overlapping oligonucleotide hybridizations.

    Science.gov (United States)

    Kim, Changsoo; Robertson, Jon S; Paterson, Andrew H

    2011-09-01

    Sugarcane (Saccharum spp.) breeders in the early 20th century made remarkable progress in increasing yield and disease resistance by crossing Saccharum spontaneum L., a wild relative, to Saccharum officinarum L., a traditional cultivar. Modern sugarcane cultivars have approximately 71%-83% of their chromosomes originating from S. officinarum, approximately 10%-21% from S. spontaneum, and approximately 2%-13% recombinant or translocated chromosomes. In the present work, C(0)t-based cloning and sequencing (CBCS) was implemented to further explore highly repetitive DNA and to seek species-specific repeated DNA in both S. officinarum and S. spontaneum. For putatively species-specific sequences, overlappping oligonucleotide probes (overgos) were designed and hybridized to BAC filters from the interspecific hybrid sugarcane cultivar 'R570' to try to deduce parental origins of BAC clones. We inferred that 12 967 BACs putatively originated from S. officinarum and 5117 BACs from S. spontaneum. Another 1103 BACs were hybridized by both species-specific overgos, too many to account for by conventional recombination, thus suggesting ectopic recombination and (or) translocation of DNA elements. Constructing a low C(0)t library is useful to collect highly repeated DNA sequences and to search for potentially species-specific molecular markers, especially among recently diverged species. Even in the absence of repeat families that are species-specific in their entirety, the identification of localized variations within consensus sequences, coupled with the site specificity of short synthetic overgos, permits researchers to monitor species-specific or species-enriched variants.

  18. Development of in vitro assay method with radioisotope

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Chang Woon; Lim, S. M.; An, S. H.; Woo, K. S.; Chung, W. S.; Lim, S. J.; Hong, S. W. [Korea Atomic Energy Research Institute. Korea Cancer Center Hospital, Seoul (Korea, Republic of); Oh, O. D. [Yonsei University, Seoul (Korea, Republic of)

    1999-04-01

    Radioimmunoassay (RIA) and related competitive protein-binding methods began a little over 20 years ago as a cumbersome research methodology in a few specialized laboratories. Endocrinology has been greatly enriched by the new knowledge that has come as a direct result of RIA methods. Establishment of the taxol RIA system will be expected to develop RIA for drug monitoring. Scintillation proximity assay was useful since any separation step is not required, it has the advantage of dealing with multiple samples. The increased sensitivity of the new assay in determining HCV RT([{sup 125}I]dUTP) suggests that it would be worth investigating whether the system can be applied to analysis. [{sup 125}I] lodotyramine with 98.5% radiochemical purity. Optimal background counts was certificated using varied radioactivity of radionuclides. Appropriate standard curve was obtained from SPA method successively, and the concentration of hCG from unknown serum was determined by standard curve. The result concentration of hCG from unknown serum was determined by synthesized successively and purified by HPLC system. Hybridoma reducing monoclonal anti thyroglobulin antibodies titer is measured by ELISA. These studies play an important role in development of in vitro assay with radionuclides.

  19. Sample preparation and assay refinements for pathogen detection platforms

    Science.gov (United States)

    Lim, Daniel V.; Kearns, Elizabeth A.; Leskinen, Stephaney D.; Magaña, Sonia; Stroot, Joyce M.; Hunter, Dawn M.; Schlemmer, Sarah M.

    2009-02-01

    Food-borne and waterborne microbial pathogens are a potential problem in biowarfare and public health. Such pathogens can affect the health, combat readiness, and effectiveness of the warfighter in a battlefield environment and present potential threats to the civilian population through intentional or natural contamination of food and water. Conventional procedures to detect and identify microbial pathogens in food, water, and other materials can take days to perform and may provide inconclusive information. Research at the University of South Florida's Advanced Biosensors Laboratory (ABL) focuses on development of sample processing procedures and biosensor-based assays for rapid detection of biothreat agents. Rapid processing methods, including use of an automated concentrator of microorganisms in water, have been developed for complex matrix samples including ground beef, apple juice, produce, potable water and recreational water, enabling such samples to be directly tested by biosensor assays for target analytes. Bacillus atrophaeus spores and other bacteria can be concentrated from potable and recreational water at low levels with a dead-end hollow-fiber ultrafiltration concentration system. Target bacteria recovered by these processing procedures can be identified by evanescent wave, fiber optic biosensors or other detection platforms. Fiber optic biosensor assays have been improved to include subsequent PCR analysis and viability determination of captured target bacteria using broth enrichment and/or ATP luminescence.

  20. The Hybrids of Postmodernism

    Directory of Open Access Journals (Sweden)

    Dana BĂDULESCU

    2014-09-01

    Full Text Available Hybridization is a fundamental characteristic of postmodernism, included by Ihab Hassan in his “catena” of features. This paper looks into the hybrids of postmodernism, which are the result of migration, displacement and uprooting, the re-visitation of myths, folklore and legends, or projections of their author’s imagination. The hybrids used as examples here are drawn from several novels written by Salman Rushdie, especially The Satanic Verses, two short stories, one by Márquez and the other by Donald Barthelme, Borges’s Book of Imaginary Beings, Cărtărescu’s Encyclopaedia of Dragons and Michelle Cliff’s No Telephone to Heaven. Diverse as they may be, these hybrids emphasize a defining characteristic of postmodernism, which is its pluralism. I conclude that the hybrids of postmodernism are aesthetically or politically subversive. Besides, what makes them difficult to grasp is their unfixed and protean nature. They ask for high leaps of the imagination, a total suspension of disbelief and a complete surrender to the powerful seduction of imagination on the reader’s part.

  1. Enriched vibrational resonance in certain discrete systems

    Indian Academy of Sciences (India)

    A Jeevarekha; M Santhiah; P Philominathan

    2014-10-01

    We wish to report the occurrence of vibrational resonance in certain discrete systems like sine square map and sine circle map, in a unique fashion, comprising of multiple resonant peaks which pave the way for enrichment. As the systems of our choice are capable of exhibiting vibrational resonance behaviour unlike the earlier reports, they are taken for investigation and the necessary numerical and analytical results are presented. Further, we study the effect of external forcing on various attractors of these systems with appropriate bifurcation and Lyapunov exponent diagrams.

  2. Enrich the environment to empower the brain.

    Science.gov (United States)

    Sale, Alessandro; Berardi, Nicoletta; Maffei, Lamberto

    2009-04-01

    Environmental enrichment (EE) has long been exploited to investigate the influence of the environment on brain structure and function. Robust morphological and functional effects elicited by EE at the neuronal level have been reported to be accompanied by improvements in cognitive performance. Recently, EE has been shown to accelerate the development of the visual system and to enhance visual-cortex plasticity in adulthood. These new findings highlight the potential of EE as a promising non-invasive strategy to ameliorate deficits in the maturation of the nervous system and to promote recovery of normal sensory functions in pathological conditions affecting the adult brain.

  3. Small volume hypermethylated DNA enrichment for epigenetics

    NARCIS (Netherlands)

    De, Arpita; Sparreboom, Wouter; de Vreede, Lennart; de Vreede, L.; Carlen, Edwin; van den Berg, Albert; Fujii, T.; Hibara, A.; Takeuchi, S.; Fukuba, T.

    2012-01-01

    We present a small volume microfluidic chip for low concentration capture and elution of hypermethylated DNA (hmDNA) using solid-phase extraction. All steps of the assay have been qualitatively characterized using a real-time surface plasmon resoncance bionsensor. A caputre/elution effiency of 84%

  4. A loop-mediated isothermal amplification assay and sample preparation procedure for sensitive detection of Xanthomonas fragariae in strawberry

    Science.gov (United States)

    Xanthomonas fragariae is a bacterium that causes angular leaf spot of strawberry. Asymptomatic infections are common and contribute to the difficulties in disease management. The aim of this study was to develop a loop-mediated isothermal amplification (LAMP) assay with a bacterial enrichment proced...

  5. PRELIMINARY EVALUATION OF THE ASTED-XL DIALYSIS SYSTEM TOWARDS ITS APPLICABILITY IN LIGAND-BINDING ASSAYS

    NARCIS (Netherlands)

    HUANG, Z; JANSSEN, MJ; PAULUSSEN, RJA; DEZEEUW, RA; FRANKE, JP; ENSING, FK

    1995-01-01

    In ligand binding assays, the separation of bound and free fraction of the labeled ligand is very important. Dialysis is generally overlooked as separation technique since it requires large volumes and long analysis times. The availability of the ASTED-system (Automated Sequential Trace Enrichment o

  6. Assessment of Listeria sp. Interference Using a Molecular Assay To Detect Listeria monocytogenes in Food.

    Science.gov (United States)

    Zittermann, Sandra I; Stanghini, Brenda; See, Ryan Soo; Melano, Roberto G; Boleszczuk, Peter; Murphy, Allana; Maki, Anne; Mallo, Gustavo V

    2016-01-01

    Detection of Listeria monocytogenes in food is currently based on enrichment methods. When L. monocytogenes is present with other Listeria species in food, the species compete during the enrichment process. Overgrowth competition of the nonpathogenic Listeria species might result in false-negative results obtained with the current reference methods. This potential issue was noted when 50 food samples artificially spiked with L. monocytogenes were tested with a real-time PCR assay and Canada's current reference method, MFHPB-30. Eleven of the samples studied were from foods naturally contaminated with Listeria species other than those used for spiking. The real-time PCR assay detected L. monocytogenes in all 11 of these samples; however, only 6 of these samples were positive by the MFHPB-30 method. To determine whether L. monocytogenes detection can be affected by other species of the same genus due to competition, an L. monocytogenes strain and a Listeria innocua strain with a faster rate of growth in the enrichment broth were artificially coinoculated at different ratios into ground pork meat samples and cultured according to the MFHPB-30 method. L. monocytogenes was detected only by the MFHPB-30 method when L. monocytogenes/L. innocua ratios were 6.0 or higher. In contrast, using the same enrichments, the real-time PCR assay detected L. monocytogenes at ratios as low as 0.6. Taken together, these findings support the hypothesis that L. monocytogenes can be outcompeted by L. innocua during the MFHPB-30 enrichment phase. However, more reliable detection of L. monocytogenes in this situation can be achieved by a PCR-based method mainly because of its sensitivity.

  7. Research on Hybrid Vehicle Drivetrain

    Science.gov (United States)

    Xie, Zhongzhi

    Hybrid cars as a solution to energy saving, emission reduction measures, have received widespread attention. Motor drive system as an important part of the hybrid vehicles as an important object of study. Based on the hybrid electric vehicle powertrain control system for permanent magnet synchronous motor as the object of study. Can be applied to hybrid car compares the characteristics of traction motors, chose permanent magnet synchronous Motors as drive motors for hybrid vehicles. Building applications in hybrid cars in MATLAB/Simulink simulation model of permanent-magnet synchronous motor speed control system and analysis of simulation results.

  8. Weighted Feature Significance: A Simple, Interpretable Model of Compound Toxicity Based on the Statistical Enrichment of Structural Features

    OpenAIRE

    Huang, Ruili; Southall, Noel; Xia, Menghang; Cho, Ming-Hsuang; Jadhav, Ajit; Nguyen, Dac-Trung; Inglese, James; Tice, Raymond R.; Austin, Christopher P.

    2009-01-01

    In support of the U.S. Tox21 program, we have developed a simple and chemically intuitive model we call weighted feature significance (WFS) to predict the toxicological activity of compounds, based on the statistical enrichment of structural features in toxic compounds. We trained and tested the model on the following: (1) data from quantitative high–throughput screening cytotoxicity and caspase activation assays conducted at the National Institutes of Health Chemical Genomics Center, (2) dat...

  9. Comet Assay in Cancer Chemoprevention.

    Science.gov (United States)

    Santoro, Raffaela; Ferraiuolo, Maria; Morgano, Gian Paolo; Muti, Paola; Strano, Sabrina

    2016-01-01

    The comet assay can be useful in monitoring DNA damage in single cells caused by exposure to genotoxic agents, such as those causing air, water, and soil pollution (e.g., pesticides, dioxins, electromagnetic fields) and chemo- and radiotherapy in cancer patients, or in the assessment of genoprotective effects of chemopreventive molecules. Therefore, it has particular importance in the fields of pharmacology and toxicology, and in both environmental and human biomonitoring. It allows the detection of single strand breaks as well as double-strand breaks and can be used in both normal and cancer cells. Here we describe the alkali method for comet assay, which allows to detect both single- and double-strand DNA breaks.

  10. The skin-blanching assay.

    Science.gov (United States)

    Smit, P; Neumann, H A M; Thio, H B

    2012-10-01

    The skin-blanching assay is used for the determination and bioequivalence of dermatologic glucocorticoids (GCs). The exact mechanism of the production of blanching is not fully understood, but it is considered that local vasoconstriction of the skin microvasculature and the consequent blood-flow reduction cause this phenomenon. Several factors influence skin blanching, including drug concentration, duration of application, nature of vehicle, occlusion, posture and location. The intensity of vasoconstriction can be measured in several ways: visual or quantitative methods, such as reflectance spectroscopy, thermography, laser Doppler velocimetry and chromametry. In literature, contradicting results in the correlation of the skin-blanching assay with different tests to determine GC sensitivity have been reported, limiting its clinical usefulness.

  11. Engineering Hybrid Chemotaxis Receptors in Bacteria.

    Science.gov (United States)

    Bi, Shuangyu; Pollard, Abiola M; Yang, Yiling; Jin, Fan; Sourjik, Victor

    2016-09-16

    Most bacteria use transmembrane sensors to detect a wide range of environmental stimuli. A large class of such sensors are the chemotaxis receptors used by motile bacteria to follow environmental chemical gradients. In Escherichia coli, chemotaxis receptors are known to mediate highly sensitive responses to ligands, making them potentially useful for biosensory applications. However, with only four ligand-binding chemotaxis receptors, the natural ligand spectrum of E. coli is limited. The design of novel chemoreceptors to extend the sensing capabilities of E. coli is therefore a critical aspect of chemotaxis-based biosensor development. One path for novel sensor design is to harvest the large natural diversity of chemosensory functions found in bacteria by creating hybrids that have the signaling domain from E. coli chemotaxis receptors and sensory domains from other species. In this work, we demonstrate that the E. coli receptor Tar can be successfully combined with most typical sensory domains found in chemotaxis receptors and in evolutionary-related two-component histidine kinases. We show that such functional hybrids can be generated using several different fusion points. Our work further illustrates how hybrid receptors could be used to quantitatively characterize ligand specificity of chemotaxis receptors and histidine kinases using standardized assays in E. coli.

  12. Bioluminescence assay for cell viability.

    Science.gov (United States)

    Lomakina, G Yu; Modestova, Yu A; Ugarova, N N

    2015-06-01

    Theoretical aspects of the adenosine triphosphate bioluminescence assay based on the use of the firefly luciferin-luciferase system are considered, as well as its application for assessing cell viability in microbiology, sanitation, medicine, and ecology. Various approaches for the analysis of individual or mixed cultures of microorganisms are presented, and capabilities of the method for investigation of biological processes in live cells including necrosis, apoptosis, as well as for investigation of the dynamics of metabolism are described.

  13. Protein binding assay for hyaluronate

    Energy Technology Data Exchange (ETDEWEB)

    Lacy, B.E.; Underhill, C.B.

    1986-11-01

    A relatively quick and simple assay for hyaluronate was developed using the specific binding protein, hyaluronectin. The hyaluronectin was obtained by homogenizing the brains of Sprague-Dawley rats, and then centrifuging the homogenate. The resulting supernatant was used as a source of crude hyaluronectin. In the binding assay, the hyaluronectin was mixed with (/sup 3/H)hyaluronate, followed by an equal volume of saturated (NH/sub 4/)/sub 2/SO/sub 4/, which precipitated the hyaluronectin and any (/sup 3/H)hyaluronate associated with it, but left free (/sup 3/H)hyaluronate in solution. The mixture was then centrifuged, and the amount of bound (/sup 3/H)hyaluronate in the precipitate was determined. Using this assay, the authors found that hyaluronectin specifically bound hyaluronate, since other glycosaminoglycans failed to compete for the binding protein. In addition, the interaction between hyaluronectin and hyaluronate was of relatively high affinity, and the size of the hyaluronate did not appear to substantially alter the amount of binding. To determine the amount of hyaluronate in an unknown sample, they used a competition assay in which the binding of a set amount of (/sup 3/H)hyaluronate was blocked by the addition of unlabeled hyaluronate. By comparing the degree of competition of the unknown samples with that of known amounts of hyaluronate, it was possible to determine the amount of hyaluronate in the unknowns. They have found that this method is sensitive to 1 ..mu..g or less of hyaluronate, and is unaffected by the presence of proteins.

  14. GOMA: functional enrichment analysis tool based on GO modules

    Institute of Scientific and Technical Information of China (English)

    Qiang Huang; Ling-Yun Wu; Yong Wang; Xiang-Sun Zhang

    2013-01-01

    Analyzing the function of gene sets is a critical step in interpreting the results of high-throughput experiments in systems biology.A variety of enrichment analysis tools have been developed in recent years,but most output a long list of significantly enriched terms that are often redundant,making it difficult to extract the most meaningful functions.In this paper,we present GOMA,a novel enrichment analysis method based on the new concept of enriched functional Gene Ontology (GO) modules.With this method,we systematically revealed functional GO modules,i.e.,groups of functionally similar GO terms,via an optimization model and then ranked them by enrichment scores.Our new method simplifies enrichment analysis results by reducing redundancy,thereby preventing inconsistent enrichment results among functionally similar terms and providing more biologically meaningful results.

  15. GOMA: functional enrichment analysis tool based on GO modules

    Science.gov (United States)

    Huang, Qiang; Wu, Ling-Yun; Wang, Yong; Zhang, Xiang-Sun

    2013-01-01

    Analyzing the function of gene sets is a critical step in interpreting the results of high-throughput experiments in systems biology. A variety of enrichment analysis tools have been developed in recent years, but most output a long list of significantly enriched terms that are often redundant, making it difficult to extract the most meaningful functions. In this paper, we present GOMA, a novel enrichment analysis method based on the new concept of enriched functional Gene Ontology (GO) modules. With this method, we systematically revealed functional GO modules, i.e., groups of functionally similar GO terms, via an optimization model and then ranked them by enrichment scores. Our new method simplifies enrichment analysis results by reducing redundancy, thereby preventing inconsistent enrichment results among functionally similar terms and providing more biologically meaningful results. PMID:23237213

  16. for hybrid dynamical systems

    Directory of Open Access Journals (Sweden)

    Wassim M. Haddad

    2001-01-01

    Full Text Available In this paper we develop a unified dynamical systems framework for a general class of systems possessing left-continuous flows; that is, left-continuous dynamical systems. These systems are shown to generalize virtually all existing notions of dynamical systems and include hybrid, impulsive, and switching dynamical systems as special cases. Furthermore, we generalize dissipativity, passivity, and nonexpansivity theory to left-continuous dynamical systems. Specifically, the classical concepts of system storage functions and supply rates are extended to left-continuous dynamical systems providing a generalized hybrid system energy interpretation in terms of stored energy, dissipated energy over the continuous-time dynamics, and dissipated energy over the resetting events. Finally, the generalized dissipativity notions are used to develop general stability criteria for feedback interconnections of left-continuous dynamical systems. These results generalize the positivity and small gain theorems to the case of left-continuous, hybrid, and impulsive dynamical systems.

  17. Hybrid Action Systems

    DEFF Research Database (Denmark)

    Rönnkö, M.; Ravn, Anders Peter; Sere, K.

    2003-01-01

    In this paper we investigate the use of action systems with differential actions in the specifcation of hybrid systems. As the main contribution we generalize the definition of a differential action, allowing the use of arbitrary relations over model variables and their time-derivatives in modell......In this paper we investigate the use of action systems with differential actions in the specifcation of hybrid systems. As the main contribution we generalize the definition of a differential action, allowing the use of arbitrary relations over model variables and their time...... parallel composition. Moreover, as the strength of the action system formalism is the support for stepwise development by refinement, we investigate refinement involving a differential action. We show that, due to the predicate transformer semantics, standard action refinement techniques apply also...... to the differential action, thus, allowing stepwise development of hybrid systems Udgivelsesdato: JAN 1...

  18. Conditional Hybrid Nonclassicality

    Science.gov (United States)

    Agudelo, E.; Sperling, J.; Costanzo, L. S.; Bellini, M.; Zavatta, A.; Vogel, W.

    2017-09-01

    We derive and implement a general method to characterize the nonclassicality in compound discrete- and continuous-variable systems. For this purpose, we introduce the operational notion of conditional hybrid nonclassicality which relates to the ability to produce a nonclassical continuous-variable state by projecting onto a general superposition of discrete-variable subsystem. We discuss the importance of this form of quantumness in connection with interfaces for quantum communication. To verify the conditional hybrid nonclassicality, a matrix version of a nonclassicality quasiprobability is derived and its sampling approach is formulated. We experimentally generate an entangled, hybrid Schrödinger cat state, using a coherent photon-addition process acting on two temporal modes, and we directly sample its nonclassicality quasiprobability matrix. The introduced conditional quantum effects are certified with high statistical significance.

  19. Porosity in hybrid materials

    Energy Technology Data Exchange (ETDEWEB)

    Schaefer, D.W.; Beaucage, G.; Loy, D. [Sandia National Labs., Albuquerque, NM (United States)

    1995-12-31

    Multicomponent, or hybrid composites are emerging as precursors to porous materials. Sacrifice of an ephemeral phase can be used to generate porosity, the nature of which depends on precursor structure. Retention of an organic constituent, on the other hand, can add desirable toughness to an otherwise brittle ceramic. We use small-angle x-ray and neutron scattering to examine porosity in both simple and hybrid materials. We find that microphase separation controls porosity in almost all systems studied. Pore distributions are controlled by the detailed bonding within and between phases as well as the flexibility of polymeric constituents. Thus hybridization opens new regions of pore distributions not available in simple systems. We look at several sacrificial concepts and show that it is possible to generate multimodal pore size distributions due to the complicated phase structure in the precursor.

  20. Photoproduction of Hybrid Mesons

    CERN Document Server

    Barnes, T

    1998-01-01

    In this contribution I discuss prospects for photoproducing hybrid mesons at CEBAF, based on recent model results and experimental indications of possible hybrids. One excellent opportunity appears to be a search for the I=1, JPC=2+-, neutral "(b2)o" hybrid in (a2 pi)o through diffractive photoproduction. Other notable possibilities accessible through pi+ or pio exchange photoproduction are I=1, JPC=1-+, charged "pi1+" in f1 pi+, (b1 pi)+ and (rho pi)+; piJ(1770)+ in f2 pi+ and (b1 pi)+; pi(1800)+ in f0 pi+, f2 pi+, omega rho+ and (rho pi)+; a1 in f1 pi+ and f2 pi+; and omega in (rho pi)o, omega eta and (K1 K)o.