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Sample records for affinity tagging method

  1. Comparison of a Label-Free Quantitative Proteomic Method Based on Peptide Ion Current Area to the Isotope Coded Affinity Tag Method

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    Young Ah Goo

    2008-01-01

    Full Text Available Recently, several research groups have published methods for the determination of proteomic expression profiling by mass spectrometry without the use of exogenously added stable isotopes or stable isotope dilution theory. These so-called label-free, methods have the advantage of allowing data on each sample to be acquired independently from all other samples to which they can later be compared in silico for the purpose of measuring changes in protein expression between various biological states. We developed label free software based on direct measurement of peptide ion current area (PICA and compared it to two other methods, a simpler label free method known as spectral counting and the isotope coded affinity tag (ICAT method. Data analysis by these methods of a standard mixture containing proteins of known, but varying, concentrations showed that they performed similarly with a mean squared error of 0.09. Additionally, complex bacterial protein mixtures spiked with known concentrations of standard proteins were analyzed using the PICA label-free method. These results indicated that the PICA method detected all levels of standard spiked proteins at the 90% confidence level in this complex biological sample. This finding confirms that label-free methods, based on direct measurement of the area under a single ion current trace, performed as well as the standard ICAT method. Given the fact that the label-free methods provide ease in experimental design well beyond pair-wise comparison, label-free methods such as our PICA method are well suited for proteomic expression profiling of large numbers of samples as is needed in clinical analysis.

  2. Several Affinity Tags Commonly Used in Chromatographic Purification

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    Xinyu Zhao

    2013-01-01

    Full Text Available Affinity tags have become powerful tools from basic biological research to structural and functional proteomics. They were widely used to facilitate the purification and detection of proteins of interest, as well as the separation of protein complexes. Here, we mainly discuss the benefits and drawbacks of several affinity or epitope tags frequently used, including hexahistidine tag, FLAG tag, Strep II tag, streptavidin-binding peptide (SBP tag, calmodulin-binding peptide (CBP, glutathione S-transferase (GST, maltose-binding protein (MBP, S-tag, HA tag, and c-Myc tag. In some cases, a large-size affinity tag, such as GST or MBP, can significantly impact on the structure and biological activity of the fusion partner protein. So it is usually necessary to excise the tag by protease. The most commonly used endopeptidases are enterokinase, factor Xa, thrombin, tobacco etch virus, and human rhinovirus 3C protease. The proteolysis features of these proteases are described in order to provide a general guidance on the proteolytic removal of the affinity tags.

  3. Differential profiling of breast cancer plasma proteome by isotope-coded affinity tagging method reveals biotinidase as a breast cancer biomarker

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    Yu Myeong-Hee

    2010-03-01

    Full Text Available Abstract Background Breast cancer is one of the leading causes of women's death worldwide. It is important to discover a reliable biomarker for the detection of breast cancer. Plasma is the most ideal source for cancer biomarker discovery since many cells cross-communicate through the secretion of soluble proteins into blood. Methods Plasma proteomes obtained from 6 breast cancer patients and 6 normal healthy women were analyzed by using the isotope-coded affinity tag (ICAT labeling approach and tandem mass spectrometry. All the plasma samples used were depleted of highly abundant 6 plasma proteins by immune-affinity column chromatography before ICAT labeling. Several proteins showing differential abundance level were selected based on literature searches and their specificity to the commercially available antibodies, and then verified by immunoblot assays. Results A total of 155 proteins were identified and quantified by ICAT method. Among them, 33 proteins showed abundance changes by more than 1.5-fold between the plasmas of breast cancer patients and healthy women. We chose 5 proteins for the follow-up confirmation in the individual plasma samples using immunoblot assay. Four proteins, α1-acid glycoprotein 2, monocyte differentiation antigen CD14, biotinidase (BTD, and glutathione peroxidase 3, showed similar abundance ratio to ICAT result. Using a blind set of plasmas obtained from 21 breast cancer patients and 21 normal healthy controls, we confirmed that BTD was significantly down-regulated in breast cancer plasma (Wilcoxon rank-sum test, p = 0.002. BTD levels were lowered in all cancer grades (I-IV except cancer grade zero. The area under the receiver operating characteristic curve of BTD was 0.78. Estrogen receptor status (p = 0.940 and progesterone receptor status (p = 0.440 were not associated with the plasma BTD levels. Conclusions Our study suggests that BTD is a potential serological biomarker for the detection of breast cancer.

  4. Affinity Purification of Protein Complexes Using TAP Tags

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    Gerace, Erica; Moazed, Danesh

    2016-01-01

    This protocol is used for the isolation and analysis of protein complexes using the tandem affinity purification (TAP) tag system. The protocol describes the purification of a protein fused to a TAP tag comprised of two protein A domains and the calmodulin binding peptide separated by a TEV cleavage site. This is a powerful technique for rapid purification of protein complexes and the analysis of their stoichiometric composition, posttranslational modifications, structure, and functional activities. PMID:26096502

  5. Tandem affinity purification of functional TAP-tagged proteins from human cells

    NARCIS (Netherlands)

    Gregan, Juraj; Riedel, Christian G; Petronczki, Mark; Cipak, Lubos; Rumpf, Cornelia; Poser, Ina; Buchholz, Frank; Mechtler, Karl; Nasmyth, Kim

    2007-01-01

    Tandem affinity purification (TAP) is a generic two-step affinity purification protocol for isolation of TAP-tagged proteins together with associated proteins. We used bacterial artificial chromosome to heterologously express TAP-tagged murine Sgo1 protein in human HeLa cells. This allowed us to tes

  6. Identification of Thioredoxin Target Disulfides Using Isotope-Coded Affinity Tags

    DEFF Research Database (Denmark)

    Hägglund, Per; Bunkenborg, Jakob; Maeda, Kenji;

    2014-01-01

    extracts is described. The procedure utilizes the isotope-coded affinity tag (ICAT) reagents containing a thiol reactive iodoacetamide group and a biotin affinity tag to target peptides containing reduced cysteine residues. The identification of substrates for Trx and the extent of target disulfide...

  7. A photocleavable affinity tag for the enrichment of alkyne-modified biomolecules

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    Koopmans, Timo; Dekker, Frank J.; Martin, Nathaniel I.

    2012-01-01

    A new photocleavable affinity tag for use in the enrichment of alkyne-labelled biomolecules is reported. The tag is prepared via a concise synthetic route using readily available materials. The photolytic conditions employed for cleavage of the tag provide for a clean release of enriched biomolecule

  8. Observations on different resin strategies for affinity purification mass spectrometry of a tagged protein.

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    Mali, Sujina; Moree, Wilna J; Mitchell, Morgan; Widger, William; Bark, Steven J

    2016-12-15

    Co-affinity purification mass spectrometry (CoAP-MS) is a highly effective method for identifying protein complexes from a biological sample and inferring important interactions, but the impact of the solid support is usually not considered in design of such experiments. Affinity purification (AP) experiments typically utilize a bait protein expressing a peptide tag such as FLAG, c-Myc, HA or V5 and high affinity antibodies to these peptide sequences to facilitate isolation of a bait protein to co-purify interacting proteins. We observed significant variability for isolation of tagged bait proteins between Protein A/G Agarose, Protein G Dynabeads, and AminoLink resins. While previous research identified the importance of tag sequence and their location, crosslinking procedures, reagents, dilution, and detergent concentrations, the effect of the resin itself has not been considered. Our data suggest the type of solid support is important and, under the conditions of our experiments, AminoLink resin provided a more robust solid-support platform for AP-MS.

  9. The Dac-tag, an affinity tag based on penicillin-binding protein 5.

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    Lee, David Wei; Peggie, Mark; Deak, Maria; Toth, Rachel; Gage, Zoe Olivia; Wood, Nicola; Schilde, Christina; Kurz, Thimo; Knebel, Axel

    2012-09-01

    Penicillin-binding protein 5 (PBP5), a product of the Escherichia coli gene dacA, possesses some β-lactamase activity. On binding to penicillin or related antibiotics via an ester bond, it deacylates and destroys them functionally by opening the β-lactam ring. This process takes several minutes. We exploited this process and showed that a fragment of PBP5 can be used as a reversible and monomeric affinity tag. At ambient temperature (e.g., 22°C), a PBP5 fragment binds rapidly and specifically to ampicillin Sepharose. Release can be facilitated either by eluting with 10mM ampicillin or in a ligand-free manner by incubation in the cold (1-10°C) in the presence of 5% glycerol. The "Dac-tag", named with reference to the gene dacA, allows the isolation of remarkably pure fusion protein from a wide variety of expression systems, including (in particular) eukaryotic expression systems.

  10. A dual-tagging system for the affinity purification of mammalian protein complexes

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    Giannone, Richard J [ORNL; McDonald, W Hayes [ORNL; Hurst, Gregory {Greg} B [ORNL; Huang, Ying [ORNL; Wu, Jun [ORNL; Liu, Yie [National Institute on Aging, Baltimore; Wang, Yisong [ORNL

    2007-01-01

    One popular method to elucidate protein-protein interactions involves the native co-purification of an affinity tagged protein and its interacting partners, which are subsequently identified through mass spectrometry (MS) (1). Although straightforward, reproducible, and broadly employed, this strategy is hampered by the efficacy of protein recoveries both in terms of sensitivity and specificity. This is especially pertinent to methodologies that employ a single-step of purification, where suboptimal enrichment of the bait protein and its partners over background can lead to masking of their signals. Although improvements to MS instrumentation generally increase peptide detection sensitivities, the problem of specificity, i.e. distinguishing specific from non-specific interacting proteins, remains. Thus ultimately, the limiting factor in the identification of specific interacting proteins lies with the purification itself. An effort to resolve this specificity issue has been made with the introduction of the Tandem Affinity Purification (TAP) tag. This construct consists of an IgG-binding domain and calmodulin binding peptide domain separated by a tobacco etch virus (TEV) protease cleavage site (2). The TAP method was originally developed in yeast and has best demonstrated its utility in the systematic identification of numerous multiprotein complexes in the yeast proteome (3). Although modifications to the original TAP have been successful in examining the protein networks of mammalian cells (4-7), the strategy offers a relatively low yield of bait and specific interacting proteins (8), and the success rate are usually on case-by-case basis. In addition, problems inherent to any protein tagging strategy remain, such as variable exposure of the affinity tag, disruption of the bait protein's ability to fold properly, steric exclusion of interacting partners, and/or ectopic overexpression of the fusion protein, which can lead to complications in both the

  11. A Method to Site-Specifically Identify and Quantitate Carbonyl End Products of Protein Oxidation Using Oxidation-Dependent Element Coded Affinity Tags (O-ECAT) and NanoLiquid Chromatography Fourier Transform Mass Spectrometry

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    Lee, S; Young, N L; Whetstone, P A; Cheal, S M; Benner, W H; Lebrilla, C B; Meares, C F

    2005-08-25

    Protein oxidation is linked to cellular stress, aging, and disease. Protein oxidations that result in reactive species are of particular interest, since these reactive oxidation products may react with other proteins or biomolecules in an unmediated and irreversible fashion, providing a potential marker for a variety of disease mechanisms. We have developed a novel system to identify and quantitate, relative to other states, the sites of oxidation on a given protein. A specially designed Oxidation-dependent carbonyl-specific Element-Coded Affinity Mass Tag (O-ECAT), AOD, ((S)-2-(4-(2-aminooxy)-acetamido)-benzyl)-1, 4, 7, 10-tetraazacyclododecane-N, N', N'', N'''-tetraacetic acid, is used to covalently tag the residues of a protein oxidized to aldehyde or keto end products. After proteolysis, the resulting AOD-tagged peptides are affinity purified, and analyzed by nanoLC-FTICR-MS, which provides high specificity in extracting co-eluting AOD mass pairs with a unique mass difference and affords relative quantitation based on isotopic ratios. Using this methodology, we have mapped the surface oxidation sites on a model protein, recombinant human serum albumin (rHSA) in its native form (as purchased) and after FeEDTA oxidation. A variety of modified amino acid residues including lysine, arginine, proline, histidine, threonine, aspartic and glutamic acids, were found to be oxidized to aldehyde and keto end products. The sensitivity of this methodology is shown by the number of peptides identified, twenty peptides on the native protein and twenty-nine after surface oxidation using FeEDTA and ascorbate. All identified peptides map to the surface of the HSA crystal structure validating this method for identifying oxidized amino acids on protein surfaces. In relative quantitation experiments between FeEDTA oxidation and native protein oxidation, identified sites showed different relative propensities towards oxidation independent of amino acid

  12. Application of coupled affinity-sizing chromatography for the detection of proteolyzed HSA-tagged proteins.

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    London, Anne Serdakowski; Patel, Kunal; Quinn, Lisa; Lemmerer, Martin

    2015-04-01

    Coupled affinity liquid chromatography and size exclusion chromatography (ALC-SEC) is a technique that has been shown to successfully report product quality of proteins during cell expression and prior to the commencement of downstream processing chromatography steps. This method was applied to monitoring the degradation and subsequent partial remediation of a HSA-tagged protein which showed proteolysis, allowing for rapid cell line development to address this product quality dilemma. This paper outlines the novel application of this method for measuring and addressing protease-induced proteolysis.

  13. Robotic high-throughput purification of affinity-tagged recombinant proteins.

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    Wiesler, Simone C; Weinzierl, Robert O J

    2015-01-01

    Affinity purification of recombinant proteins has become the method of choice to obtain good quantities and qualities of proteins for a variety of downstream biochemical applications. While manual or FPLC-assisted purification techniques are generally time-consuming and labor-intensive, the advent of high-throughput technologies and liquid handling robotics has simplified and accelerated this process significantly. Additionally, without the human factor as a potential source of error, automated purification protocols allow for the generation of large numbers of proteins simultaneously and under directly comparable conditions. The delivered material is ideal for activity comparisons of different variants of the same protein. Here, we present our strategy for the simultaneous purification of up to 24 affinity-tagged proteins for activity measurements in biochemical assays. The protocol described is suitable for the scale typically required in individual research laboratories.

  14. A tailor-made "tag-receptor" affinity pair for the purification of fusion proteins.

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    Pina, Ana S; Guilherme, Márcia; Pereira, Alice S; Fernandes, Cláudia S F M; Branco, Ricardo J F; El Khoury, Graziella; Lowe, Christopher R; Roque, A Cecília A

    2014-07-07

    A novel affinity "tag-receptor" pair was developed as a generic platform for the purification of fusion proteins. The hexapeptide RKRKRK was selected as the affinity tag and fused to green fluorescent protein (GFP). The DNA fragments were designed, cloned in Pet-21c expression vector and expressed in E. coli host as soluble protein. A solid-phase combinatorial library based on the Ugi reaction was synthesized: 64 affinity ligands displaying complementary functionalities towards the designed tag. The library was screened by affinity chromatography in a 96-well format for binding to the RKRKRK-tagged GFP protein. Lead ligand A7C1 was selected for the purification of RKRKRK fusion proteins. The affinity pair RKRKRK-tagged GFP with A7C1 emerged as a promising solution (Ka of 2.45×10(5)  M(-1) ). The specificity of the ligand towards the tag was observed experimentally and theoretically through automated docking and molecular dynamics simulations.

  15. Dual-tagging system for the affinity purification of mammalian protein complexes

    Energy Technology Data Exchange (ETDEWEB)

    Giannone, Richard J [ORNL; McDonald, W Hayes [ORNL; Hurst, Gregory {Greg} B [ORNL; Huang, Ying [ORNL; Wu, Jun [ORNL; Liu, Yie [National Institute on Aging, Baltimore; Wang, Yisong [ORNL

    2007-01-01

    Although affinity purification coupled with mass spectrometry (MS) provides a powerful tool to study protein-protein interactions, this strategy has encountered numerous difficulties when adapted to mammalian cells. Here we describe a Gateway{reg_sign}-compatible dual-tag affinity purification system that integrates regulatable expression, tetracysteine motifs, and various combinations of affinity tags to facilitate the cloning, detection, and purification of bait proteins and their interacting partners. Utilizing the human telomere binding protein TRF2 as a benchmark, we demonstrate bait protein recoveries upwards of approximately 16% from as little as 1-7 x 10{sup 7} cells and successfully identify known TRF2 interacting proteins, suggesting that our dual-tag affinity purification approach is a capable new tool for expanding the capacity to explore mammalian proteomic networks.

  16. Batch affinity adsorption of His-tagged proteins with EDTA-based chitosan.

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    Hua, Weiwei; Lou, Yimin; Xu, Weiyuan; Cheng, Zhixian; Gong, Xingwen; Huang, Jianying

    2016-01-01

    Affinity adsorption purification of hexahistidine-tagged (His-tagged) proteins using EDTA-chitosan-based adsorption was designed and carried out. Chitosan was elaborated with ethylenediaminetetraacetic acid (EDTA), and the resulting polymer was characterized by FTIR, TGA, and TEM. Different metals including Ni(2+), Cu(2+), and Zn(2+) were immobilized with EDTA-chitosan, and their capability to the specific adsorption of His-tagged proteins were then investigated. The results showed that Ni(2+)-EDTA-chitosan and Zn(2+)-EDTA-chitosan had high affinity toward the His-tagged proteins, thus isolating them from protein mixture. The target fluorescent-labeled hexahistidine protein remained its fluorescent characteristic throughout the purification procedure when Zn(2+)-EDTA-chitosan was used as a sorbent, wherein the real-time monitor was performed to examine the immigration of fluorescent-labeled His-tagged protein. Comparatively, Zn(2+)-EDTA-chitosan showed more specific binding ability for the target protein, but with less binding capacity. It was further proved that this purification system could be recovered and reused at least for 5 times and could run on large scales. The presented M(2+)-EDTA-chitosan system, with the capability to specifically bind His-tagged proteins, make the purification of His-tagged proteins easy to handle, leaving out fussy preliminary treatment, and with the possibility of continuous processing and a reduction in operational cost in relation to the costs of conventional processes.

  17. Quantitative proteome analysis using isotope-coded affinity tags and mass spectrometry.

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    Shiio, Yuzuru; Aebersold, Ruedi

    2006-01-01

    A main objective of proteomics research is to systematically identify and quantify proteins in a given proteome (cells, subcellular fractions, protein complexes, tissues or body fluids). Protein labeling with isotope-coded affinity tags (ICAT) followed by tandem mass spectrometry allows sequence identification and accurate quantification of proteins in complex mixtures, and has been applied to the analysis of global protein expression changes, protein changes in subcellular fractions, components of protein complexes, protein secretion and body fluids. This protocol describes protein-sample labeling with ICAT reagents, chromatographic fractionation of the ICAT-labeled tryptic peptides, and protein identification and quantification using tandem mass spectrometry. The method is suitable for both large-scale analysis of complex samples including whole proteomes and small-scale analysis of subproteomes, and allows quantitative analysis of proteins, including those that are difficult to analyze by gel-based proteomics technology.

  18. MHC class II tetramers made from isolated recombinant α and β chains refolded with affinity-tagged peptides

    DEFF Research Database (Denmark)

    Braendstrup, Peter; Justesen, Sune Frederik Lamdahl; Osterbye, Thomas;

    2013-01-01

    Targeting CD4+ T cells through their unique antigen-specific, MHC class II-restricted T cell receptor makes MHC class II tetramers an attractive strategy to identify, validate and manipulate these cells at the single cell level. Currently, generating class II tetramers is a specialized undertaking...... effectively limiting their use and emphasizing the need for improved methods of production. Using class II chains expressed individually in E. coli as versatile recombinant reagents, we have previously generated peptide-MHC class II monomers, but failed to generate functional class II tetramers. Adding...... a monomer purification principle based upon affinity-tagged peptides, we here provide a robust method to produce class II tetramers and demonstrate staining of antigen-specific CD4+ T cells. We also provide evidence that both MHC class II and T cell receptor molecules largely accept affinity-tagged peptides...

  19. Microbial polyhydroxyalkanote synthesis repression protein PhaR as an affinity tag for recombinant protein purification

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    Chen Guo

    2010-05-01

    Full Text Available Abstract Background PhaR which is a repressor protein for microbial polyhydroxyalkanoates (PHA biosynthesis, is able to attach to bacterial PHA granules in vivo, was developed as an affinity tag for in vitro protein purification. Fusion of PhaR-tagged self-cleavable Ssp DnaB intein to the N-terminus of a target protein allowed protein purification with a pH and temperature shift. During the process, the target protein was released to the supernatant while PhaR-tagged intein was still immobilized on the PHA nanoparticles which were then separated by centrifugation. Results Fusion protein PhaR-intein-target protein was expressed in recombinant Escherichia coli. The cell lysates after sonication and centrifugation were collected and then incubated with PHA nanoparticles to allow sufficient absorption onto the PHA nanoparticles. After several washing processes, self-cleavage of intein was triggered by pH and temperature shift. As a result, the target protein was released from the particles and purified after centrifugation. As target proteins, enhanced green fluorescent protein (EGFP, maltose binding protein (MBP and β-galactosidase (lacZ, were successfully purified using the PhaR based protein purification method. Conclusion The successful purification of EGFP, MBP and LacZ indicated the feasibility of this PhaR based in vitro purification system. Moreover, the elements used in this system can be easily obtained and prepared by users themselves, so they can set up a simple protein purification strategy by themselves according to the PhaR method, which provides another choice instead of expensive commercial protein purification systems.

  20. The Monitoring and Affinity Purification of Proteins Using Dual-Tags with Tetracysteine Motifs

    Energy Technology Data Exchange (ETDEWEB)

    Giannone, Richard J [ORNL; Liu, Yie [ORNL; Wang, Yisong [ORNL

    2009-01-01

    Identification and characterization of protein-protein interaction networks is essential for the elucidation of biochemical mechanisms and cellular function. Affinity purification in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) has emerged as a very powerful tactic for the identification of specific protein-protein interactions. In this chapter we describe a comprehensive methodology that utilizes our recently developed dual-tag affinity purification system for the enrichment and identification of mammalian protein complexes. The protocol covers a series of separate but sequentially related techniques focused on the facile monitoring and purification of a dual-tagged protein of interest and its interacting partners via a system built with tetracysteine motifs and various combinations of affinity tags. Using human telomeric repeat binding factor 2 (TRF2) as an example, we have demonstrated the power of the system in terms of bait protein recovery after dual-tag affinity purification, detection of bait protein subcellular localization and expression, and successful identification of known and potentially novel TRF2 interacting proteins. Although the protocol described here has been optimized for the identification and characterization of TRF2-associated proteins, it is, in principle, applicable to the study of any other mammalian protein complexes that may be of interest to the research community.

  1. The Monitoring and Affinity Purification of Proteins Using Dual Tags with Tetracysteine Motifs

    Science.gov (United States)

    Giannone, Richard J.; Liu, Yie; Wang, Yisong

    Identification and characterization of protein-protein interaction networks is essential for the elucidation of biochemical mechanisms and cellular function. Affinity purification in combination with liquid chromatography-tandem mass spectrometry (LC-MS/MS) has emerged as a very powerful tactic for the identification of specific protein-protein interactions. In this chapter, we describe a comprehensive methodology that uses our recently developed dual-tag affinity purification system for the enrichment and identification of mammalian protein complexes. The protocol covers a series of separate but sequentially related techniques focused on the facile monitoring and purification of a dual-tagged protein of interest and its interacting partners via a system built with tetracysteine motifs and various combinations of affinity tags. Using human telomeric repeat binding factor 2 (TRF2) as an example, we demonstrate the power of the system in terms of bait protein recovery after dual-tag affinity purification, detection of bait protein subcellular localization and expression, and successful identification of known and potentially novel TRF2 interacting proteins. Although the protocol described here has been optimized for the identification and characterization of TRF2-associated proteins, it is, in principle, applicable to the study of any other mammalian protein complexes that may be of interest to the research community.

  2. Crystallographic structure of Ni-Co coating on the affinity adsorption of histidine-tagged protein.

    Science.gov (United States)

    Chang, Yaw-Jen; Chen, Sheng-Zheng; Ho, Ching-Yuan

    2015-04-01

    The principle of immobilized metal affinity chromatography (IMAC) has been recently implemented for protein microarrays for the study of protein abundance and function. Ni-Co film fabricated by electrodeposition is a novel microarray surface in an alloy type for immobilizing histidine-tagged proteins based on IMAC. In this paper, the effects of crystallographic structures and surface properties of Ni-Co coatings, with and without the annealing process, on the immobilization of histidine-tagged proteins were systematically investigated. The experimental results reveal that the stronger hcp texture, due to a higher Co content, results in better affinity adsorption for histidine-tagged biotin. Nevertheless, the allotropic phase transformation from hcp to fcc, due to the annealing process, leads to the decrease of affinity adsorption. The wettability property and the surface roughness of Ni-Co coating are, however, not important factors. Obviously, the crystallographic structure of Ni-Co coating is the dominant factor for the specific affinity adsorption of histidine-tagged protein.

  3. A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana.

    Science.gov (United States)

    Stotz, Henrik U; Findling, Simone; Nukarinen, Ella; Weckwerth, Wolfram; Mueller, Martin J; Berger, Susanne

    2014-01-01

    Tandem affinity purification (TAP) tagging provides a powerful tool for isolating interacting proteins in vivo. TAP-tag purification offers particular advantages for the identification of stimulus-induced protein interactions. Type II bZIP transcription factors (TGA2, TGA5 and TGA6) play key roles in pathways that control salicylic acid, ethylene, xenobiotic and reactive oxylipin signaling. Although proteins interacting with these transcription factors have been identified through genetic and yeast 2-hybrid screening, others are still elusive. We have therefore generated a C-terminal TAP-tag of TGA2 to isolate additional proteins that interact with this transcription factor. Three lines most highly expressing TAP-tagged TGA2 were functional in that they partially complemented reactive oxylipin-responsive gene expression in a tga2 tga5 tga6 triple mutant. TAP-tagged TGA2 in the most strongly overexpressing line was proteolytically less stable than in the other 2 lines. Only this overexpressing line could be used in a 2-step purification process, resulting in isolation of co-purifying bands of larger molecular weight than TGA2. TAP-tagged TGA2 was used to pull down NPR1, a protein known to interact with this transcription factor. Mass spectrometry was used to identify peptides that co-purified with TAP-tagged TGA2. Having generated this TGA2 TAP-tag line will therefore be an asset to researchers interested in stimulus-induced signal transduction processes.

  4. A tandem affinity purification tag of TGA2 for isolation of interacting proteins in Arabidopsis thaliana.

    Science.gov (United States)

    Stotz, Henrik U; Findling, Simone; Nukarinen, Ella; Weckwerth, Wolfram; Mueller, Martin J; Berger, Susanne

    2014-08-19

    Tandem affinity purification (TAP) tagging provides a powerful tool for isolating interacting proteins in vivo. TAP-tag purification offers particular advantages for the identification of stimulus-induced protein interactions. Type II bZIP transcription factors (TGA2, TGA5 and TGA6) play key roles in pathways that control salicylic acid, ethylene, xenobiotic and reactive oxylipin signaling. Although proteins interacting with these transcription factors have been identified through genetic and yeast two-hybrid screening, others are still elusive. We have therefore generated a C-terminal TAP-tag of TGA2 to isolate additional proteins that interact with this transcription factor. Three lines most highly expressing TAP-tagged TGA2 were functional in that they partially complemented reactive oxylipin-responsive gene expression in a tga2 tga5 tga6 triple mutant. TAP-tagged TGA2 in the most strongly overexpressing line was proteolytically less stable than in the other two lines. Only this overexpressing line could be used in a two-step purification process, resulting in isolation of co-purifying bands of larger molecular weight than TGA2. TAP-tagged TGA2 was used to pull down NPR1, a protein known to interact with this transcription factor. Mass spectrometry was used to identify peptides that co-purified with TAP-tagged TGA2. Having generated this TGA2 TAP-tag line will therefore be an asset to researchers interested in stimulus-induced signal transduction processes.

  5. Single-step affinity and cost-effective purification of recombinant proteins using the Sepharose-binding lectin-tag from the mushroom Laetiporus sulphureus as fusion partner.

    Science.gov (United States)

    Li, Xiao-Jing; Liu, Jin-Ling; Gao, Dong-Sheng; Wan, Wen-Yan; Yang, Xia; Li, Yong-Tao; Chang, Hong-Tao; Chen, Lu; Wang, Chuan-Qing; Zhao, Jun

    2016-03-01

    Previous research showed that a lectin from the mushroom Laetiporus sulphureus, designed LSL, bound to Sepharose and could be eluted by lactose. In this study, by taking advantage of the strong affinity of LSL-tag for Sepharose, we developed a single-step purification method for LSL-tagged fusion proteins. We utilized unmodified Sepharose-4B as a specific adsorbent and 0.2 M lactose solution as an elution buffer. Fusion proteins of LSL-tag and porcine circovirus capsid protein, designated LSL-Cap was recovered with purity of 90 ± 4%, and yield of 87 ± 3% from crude extract of recombinant Escherichia coli. To enable the remove of LSL-tag, tobacco etch virus (TEV) protease recognition sequence was placed downstream of LSL-tag in the expression vector, and LSL-tagged TEV protease, designated LSL-TEV, was also expressed in E. coli., and was recovered with purity of 82 ± 5%, and yield of 85 ± 2% from crude extract of recombinant E. coli. After digestion of LSL-tagged recombinant proteins with LSL-TEV, the LSL tag and LSL-TEV can be easily removed by passing the digested products through the Sepharose column. It is of worthy noting that the Sepharose can be reused after washing with PBS. The LSL affinity purification method enables rapid and inexpensive purification of LSL-tagged fusion proteins and scale-up production of native proteins.

  6. Evaluation of Affinity-Tagged Protein Expression Strategies using Local and Global Isotope Ratio Measurements

    Energy Technology Data Exchange (ETDEWEB)

    Hervey, IV, William Judson [ORNL; Khalsa-Moyers, Gurusahai K [ORNL; Lankford, Patricia K [ORNL; Owens, Elizabeth T [ORNL; McKeown, Catherine K [ORNL; Lu, Tse-Yuan S [ORNL; Foote, Linda J [ORNL; Morrell-Falvey, Jennifer L [ORNL; McDonald, W Hayes [ORNL; Pelletier, Dale A [ORNL; Hurst, Gregory {Greg} B [ORNL

    2009-01-01

    Protein enrichments of engineered, affinity-tagged (or bait ) fusion proteins with interaction partners are often laden with background, non-specific proteins, due to interactions that occur in vitro as an artifact of the technique. Furthermore, the in vivo expression of the bait protein may itself affect physiology or metabolism. In this study, intrinsic affinity purification challenges were investigated in a model protein complex, DNA-dependent RNA polymerase (RNAP), encompassing chromosome- and plasmid-encoding strategies for bait proteins in two different microbial species: Escherichia coli and Rhodopseudomonas palustris. Isotope ratio measurements of bait protein expression strains relative to native, wild-type strains were performed by liquid chromatography tandem mass spectrometry (LC-MS-MS) to assess bait protein expression strategies in each species. Authentic interacting proteins of RNAP were successfully discerned from artifactual co-isolating proteins by the isotopic differentiation of interactions as random or targeted (I-DIRT) method (A. J. Tackett et al. J. Proteome Res. 2005, 4 (5), 1752-1756). To investigate broader effects of bait protein production in the bacteria, we compared proteomes from strains harboring a plasmid that encodes an affinity-tagged subunit (RpoA) of the RNAP complex with the corresponding wild-type strains using stable isotope metabolic labeling. The ratio of RpoA abundance in plasmid strains versus wild type was 0.8 for R. palustris and 1.7 for E. coli. While most other proteins showed no appreciable difference, proteins significantly increased in abundance in plasmid-encoded bait-expressing strains of both species included the plasmid encoded antibiotic resistance protein, GenR and proteins involved in amino acid biosynthesis. Together, these local, complex-specific and more global, whole proteome isotopic abundance ratio measurements provided a tool for evaluating both in vivo and in vitro effects of plasmid

  7. A chimeric affinity tag for efficient expression and chromatographic purification of heterologous proteins from plants

    Directory of Open Access Journals (Sweden)

    Frank eSainsbury

    2016-02-01

    Full Text Available The use of plants as expression hosts for recombinant proteins is an increasingly attractive option for the production of complex and challenging biopharmaceuticals. Tools are needed at present to marry recent developments in high-yielding gene vectors for heterologous expression with routine protein purification techniques. In this study we designed the Cysta-tag, a new purification tag for immobilized metal affinity chromatography (IMAC of plant-made proteins based on the protein-stabilizing fusion partner SlCYS8. We show that the Cysta-tag may be used to rapidly purify proteins under native conditions, and then be removed enzymatically to isolate the protein of interest. We also show that commonly used protease recognition sites for linking purification tags are differentially stable in leaves of the commonly used expression host Nicotiana benthamiana, with those linkers susceptible to cysteine proteases being less stable then serine protease-cleavable linkers. As an example we describe a Cysta-tag experimental scheme for the one-step purification of a clinically useful protein, human α1-antitrypsin, transiently expressed in N. benthamiana. With potential applicability to the variety of chromatography formats commercially available for IMAC-based protein purification, the Cysta-tag provides a convenient means for the efficient and cost-effective purification of recombinant proteins from plant tissues.

  8. A novel purification method for histidine-tagged proteins containing a thrombin cleavage site

    NARCIS (Netherlands)

    Hefti, M.H.; Vugt-Toorn, van der C.J.; Dixon, R.; Vervoort, J.J.M.

    2001-01-01

    A general procedure for the purification of histidine-tagged proteins has been developed using immobilized metal-ion affinity chromatography. This two-step purification method can be used for proteins containing a hexahistidine tag and a thrombin cleavage site, yielding high amounts of purified prot

  9. A cleavable silica-binding affinity tag for rapid and inexpensive protein purification.

    Science.gov (United States)

    Coyle, Brandon L; Baneyx, François

    2014-10-01

    We describe a new affinity purification tag called Car9 that confers proteins to which it is fused micromolar affinity for unmodified silica. When appended to the C-terminus of GFPmut2 through a flexible linker, Car9 promotes efficient adsorption to silica gel and the fusion protein can be released from the particles by incubation with L-lysine. Using a silica gel column and the lysine elution approach in fast protein liquid chromatography (FPLC) mode, Car9-tagged versions of GFPmut2, mCherry and maltose binding protein (MBP) can be recovered from clarified lysates with a purity of 80-90%. Capitalizing on silica's ability to handle large pressure drops, we further show that it is possible to go from cell lysates to purified protein in less than 15 min using a fully disposable device. Finally, we demonstrate that the linker-Car9 region is susceptible to proteolysis by E. coli OmpT and take advantage of this observation to excise the C-terminal extension of GFPmut2-Car9 by incubating purified fusion protein with cells that overproduce the outer membrane protease OmpT. The set of strategies described herein, should reduce the cost of affinity purification by at least 10-fold, cut down purification times to minutes, and allow for the production of proteins with native (or nearly native) termini from their C-terminally-tagged versions.

  10. In-house preparation of hydrogels for batch affinity purification of glutathione S-transferase tagged recombinant proteins

    Directory of Open Access Journals (Sweden)

    Buhrman Jason S

    2012-09-01

    Full Text Available Abstract Background Many branches of biomedical research find use for pure recombinant proteins for direct application or to study other molecules and pathways. Glutathione affinity purification is commonly used to isolate and purify glutathione S-transferase (GST-tagged fusion proteins from total cellular proteins in lysates. Although GST affinity materials are commercially available as glutathione immobilized on beaded agarose resins, few simple options for in-house production of those systems exist. Herein, we describe a novel method for the purification of GST-tagged recombinant proteins. Results Glutathione was conjugated to low molecular weight poly(ethylene glycol diacrylate (PEGDA via thiol-ene “click” chemistry. With our in-house prepared PEGDA:glutathione (PEGDA:GSH homogenates, we were able to purify a glutathione S-transferase (GST green fluorescent protein (GFP fusion protein (GST-GFP from the soluble fraction of E. coli lysate. Further, microspheres were formed from the PEGDA:GSH hydrogels and improved protein binding to a level comparable to purchased GSH-agarose beads. Conclusions GSH containing polymers might find use as in-house methods of protein purification. They exhibited similar ability to purify GST tagged proteins as purchased GSH agarose beads.

  11. A strategy to enhance the binding affinity of fluorophore-aptamer pairs for RNA tagging with neomycin conjugation.

    Science.gov (United States)

    Jeon, Jongho; Lee, Kyung Hyun; Rao, Jianghong

    2012-10-14

    Fluorogenic sulforhodamine-neomycin conjugates have been designed and synthesized for RNA tagging. Conjugates were fluorescently activated by binding to RNA aptamers and exhibited greater than 250-400 fold enhancement in binding affinity relative to corresponding unconjugated fluorophores.

  12. A novel gigaporous GSH affinity medium for high-speed affinity chromatography of GST-tagged proteins.

    Science.gov (United States)

    Huang, Yongdong; Zhang, Rongyue; Li, Juan; Li, Qiang; Su, Zhiguo; Ma, Guanghui

    2014-03-01

    Novel GSH-AP (phenoxyl agarose coated gigaporous polystyrene, Agap-co-PSt) microspheres were successfully prepared by introducing GSH ligand into hydrophilic AP microspheres pre-activated with 1,4-butanediol diglycidyl ether. The gigaporous structure and chromatographic properties of GSH-AP medium were evaluated and compared with commercial GSH Sepharose FF (GSH-FF) medium. The macropores (100-500nm) of gigaporous PSt microspheres were well maintained after coating with agarose and functionalized with GSH ligand. Hydrodynamic experiments showed that GSH-AP column had less backpressure and plate height than those of GSH-FF column at high flow velocity, which was beneficial for its use in high-speed chromatography. The presence of flow-through pores in GSH-AP microspheres also accelerated the mass transfer rate of biomolecules induced by convective flow, leading to high protein resolution and high dynamic binding capacity (DBC) of glutathione S-transferase (GST) at high flow velocity. High purity of GST and GST-tagged recombinant human interleukin-1 receptor antagonist (rhIL-1RA) were obtained from crude extract with an acceptable recovery yield within 1.5min at a velocity up to 1400cm/h. GSH-AP medium is promising for high-speed affinity chromatography for the purification of GST and GST-tagged proteins.

  13. SNAP-Tag Technology: A Useful Tool To Determine Affinity Constants and Other Functional Parameters of Novel Antibody Fragments.

    Science.gov (United States)

    Niesen, Judith; Sack, Markus; Seidel, Melanie; Fendel, Rolf; Barth, Stefan; Fischer, Rainer; Stein, Christoph

    2016-08-17

    Antibody derivatives, such as the single chain fragment variable (scFv), can be developed as diagnostic and therapeutic tools in cancer research, especially in the form of fusion proteins. Such derivatives are easier to produce and modify than monoclonal antibodies (mAbs) and achieve better tissue/tumor penetration. The genetic modification of scFvs is also much more straightforward than the challenging chemical modification of mAbs. Therefore, we constructed two scFvs derived from the approved monoclonal antibodies cetuximab (scFv2112) and panitumumab (scFv1711), both of which are specific for the epidermal growth factor receptor (EGFR), a well-characterized solid tumor antigen. Both scFvs were genetically fused to the SNAP-tag, an engineered version of the human DNA repair enzyme O(6)-alkylguanine DNA alkyltransferase that allows the covalent coupling of benzylguanine (BG)-modified substrates such as fluorescent dyes. The SNAP-tag achieves controllable and irreversible protein modification and is an important tool for experimental studies in vitro and in vivo. The affinity constant of a scFv is a key functional parameter, especially in the context of a fusion protein. Therefore, we developed a method to define the affinity constants of scFv-SNAP fusion proteins by surface plasmon resonance (SPR) spectroscopy. We could confirm that both scFvs retained their functionality after fusion to the SNAP-tag in a variety of procedures and assays, including ELISA, flow cytometry, and confocal microscopy. The experimental procedures described herein, and the new protocol for affinity determination by SPR spectroscopy, are suitable for the preclinical evaluation of diverse antibody formats and derivatives.

  14. Proteomic analysis of Trypanosoma cruzi developmental stages using isotope-coded affinity tag reagents.

    Science.gov (United States)

    Paba, Jaime; Ricart, Carlos A O; Fontes, Wagner; Santana, Jaime M; Teixeira, Antonio R L; Marchese, Jason; Williamson, Brian; Hunt, Tony; Karger, Barry L; Sousa, Marcelo V

    2004-01-01

    Comparative proteome analysis of developmental stages of the human pathogen Trypanosoma cruzi was carried out by isotope-coded affinity tag technology (ICAT) associated with liquid cromatography-mass spectrometry peptide sequencing (LC-MS/MS). Protein extracts of the protozoan trypomastigote and amastigote stages were labeled with heavy (D8) and light (D0) ICAT reagents and subjected to cation exchange and avidin affinity chromatographies followed by LC-MS/MS analysis. High confidence sequence information and expression levels for 41 T. cruzi polypeptides, including metabolic enzymes, paraflagellar rod components, tubulins, and heat-shock proteins were reported. Twenty-nine proteins displayed similar levels of expression in both forms of the parasite, nine proteins presented higher levels in trypomastigotes, whereas three were more expressed in amastigotes.

  15. A photo-cleavable biotin affinity tag for the facile release of a photo-crosslinked carbohydrate-binding protein.

    Science.gov (United States)

    Chang, Tsung-Che; Adak, Avijit K; Lin, Ting-Wei; Li, Pei-Jhen; Chen, Yi-Ju; Lai, Chain-Hui; Liang, Chien-Fu; Chen, Yu-Ju; Lin, Chun-Cheng

    2016-03-15

    The use of photo-crosslinking glycoprobes represents a powerful strategy for the covalent capture of labile protein complexes and allows detailed characterization of carbohydrate-mediated interactions. The selective release of target proteins from solid support is a key step in functional proteomics. We envisaged that light activation can be exploited for releasing labeled protein in a dual photo-affinity probe-based strategy. To investigate this possibility, we designed a trifunctional, galactose-based, multivalent glycoprobe for affinity labeling of carbohydrate-binding proteins. The resulting covalent protein-probe adduct is attached to a photo-cleavable biotin affinity tag; the biotin moiety enables specific presentation of the conjugate on streptavidin-coated beads, and the photolabile linker allows the release of the labeled proteins. This dual probe promotes both the labeling and the facile cleavage of the target protein complexes from the solid surfaces and the remainder of the cell lysate in a completely unaltered form, thus eliminating many of the common pitfalls associated with traditional affinity-based purification methods.

  16. Cellulose affinity purification of fusion proteins tagged with fungal family 1 cellulose-binding domain.

    Science.gov (United States)

    Sugimoto, Naohisa; Igarashi, Kiyohiko; Samejima, Masahiro

    2012-04-01

    N- or C-terminal fusions of red-fluorescent protein (RFP) with various fungal cellulose-binding domains (CBDs) belonging to carbohydrate binding module (CBM) family 1 were expressed in a Pichia pastoris expression system, and the resulting fusion proteins were used to examine the feasibility of large-scale affinity purification of CBD-tagged proteins on cellulose columns. We found that RFP fused with CBD from Trichoderma reesei CBHI (CBD(Tr)(CBHI)) was expressed at up to 1.2g/l in the culture filtrate, which could be directly injected into the cellulose column. The fusion protein was tightly adsorbed on the cellulose column in the presence of a sufficient amount of ammonium sulfate and was efficiently eluted with pure water. Bovine serum albumin (BSA) was not captured under these conditions, whereas both BSA and the fusion protein were adsorbed on a phenyl column, indicating that the cellulose column can be used for the purification of not only hydrophilic proteins but also for hydrophobic proteins. Recovery of various fusion proteins exceeded 80%. Our results indicate that protein purification by expression of a target protein as a fusion with a fungal family 1 CBD tag in a yeast expression system, followed by affinity purification on a cellulose column, is simple, effective and easily scalable.

  17. Single-Step Affinity Purification of ERK Signaling Complexes Using the Streptavidin-Binding Peptide (SBP) Tag.

    Science.gov (United States)

    Yang, Liu; Veraksa, Alexey

    2017-01-01

    Elucidation of biological functions of signaling proteins is facilitated by studying their protein-protein interaction networks. Affinity purification combined with mass spectrometry (AP-MS) has become a favorite method to study protein complexes. Here we describe a procedure for single-step purification of ERK (Rolled) and associated proteins from Drosophila cultured cells. The use of the streptavidin-binding peptide (SBP) tag allows for a highly efficient isolation of native ERK signaling complexes, which are suitable for subsequent analysis by mass spectrometry. Our analysis of the ERK interactome has identified both known and novel signaling components. This method can be easily adapted for SBP-based purification of protein complexes in any expression system.

  18. Immobilized palladium(II) ion affinity chromatography for recovery of recombinant proteins with peptide tags containing histidine and cysteine.

    Science.gov (United States)

    Kikot, Pamela; Polat, Aise; Achilli, Estefania; Fernandez Lahore, Marcelo; Grasselli, Mariano

    2014-11-01

    Fusion of peptide-based tags to recombinant proteins is currently one of the most used tools for protein production. Also, immobilized metal ion affinity chromatography (IMAC) has a huge application in protein purification, especially in research labs. The combination of expression systems of recombinant tagged proteins with this robust chromatographic system has become an efficient and rapid tool to produce milligram-range amounts of proteins. IMAC-Ni(II) columns have become the natural partners of 6xHis-tagged proteins. The Ni(II) ion is considered as the best compromise of selectivity and affinity for purification of a recombinant His-tagged protein. The palladium(II) ion is also able to bind to side chains of amino acids and form ternary complexes with iminodiacetic acid and free amino acids and other sulfur-containing molecules. In this work, we evaluated two different cysteine- and histidine-containing six amino acid tags linked to the N-terminal group of green fluorescent protein (GFP) and studied the adsorption and elution conditions using novel eluents. Both cysteine-containing tagged GFPs were able to bind to IMAC-Pd(II) matrices and eluted successfully using a low concentration of thiourea solution. The IMAC-Ni(II) system reaches less than 20% recovery of the cysteine-containing tagged GFP from a crude homogenate of recombinant Escherichia coli, meanwhile the IMAC-Pd(II) yields a recovery of 45% with a purification factor of 13.

  19. Proton Affinity Calculations with High Level Methods.

    Science.gov (United States)

    Kolboe, Stein

    2014-08-12

    Proton affinities, stretching from small reference compounds, up to the methylbenzenes and naphthalene and anthracene, have been calculated with high accuracy computational methods, viz. W1BD, G4, G3B3, CBS-QB3, and M06-2X. Computed and the currently accepted reference proton affinities are generally in excellent accord, but there are deviations. The literature value for propene appears to be 6-7 kJ/mol too high. Reported proton affinities for the methylbenzenes seem 4-5 kJ/mol too high. G4 and G3 computations generally give results in good accord with the high level W1BD. Proton affinity values computed with the CBS-QB3 scheme are too low, and the error increases with increasing molecule size, reaching nearly 10 kJ/mol for the xylenes. The functional M06-2X fails markedly for some of the small reference compounds, in particular, for CO and ketene, but calculates methylbenzene proton affinities with high accuracy.

  20. Evaluation of Intercontinental Transport of Ozone Using Full-tagged, Tagged-N and Sensitivity Methods

    Science.gov (United States)

    Guo, Y.; Liu, J.; Mauzerall, D. L.; Emmons, L. K.; Horowitz, L. W.; Fan, S.; Li, X.; Tao, S.

    2014-12-01

    Long-range transport of ozone is of great concern, yet the source-receptor relationships derived previously depend strongly on the source attribution techniques used. Here we describe a new tagged ozone mechanism (full-tagged), the design of which seeks to take into account the combined effects of emissions of ozone precursors, CO, NOx and VOCs, from a particular source, while keeping the current state of chemical equilibrium unchanged. We label emissions from the target source (A) and background (B). When two species from A and B sources react with each other, half of the resulting products are labeled A, and half B. Thus the impact of a given source on downwind regions is recorded through tagged chemistry. We then incorporate this mechanism into the Model for Ozone and Related chemical Tracers (MOZART-4) to examine the impact of anthropogenic emissions within North America, Europe, East Asia and South Asia on ground-level ozone downwind of source regions during 1999-2000. We compare our results with two previously used methods -- the sensitivity and tagged-N approaches. The ozone attributed to a given source by the full-tagged method is more widely distributed spatially, but has weaker seasonal variability than that estimated by the other methods. On a seasonal basis, for most source/receptor pairs, the full-tagged method estimates the largest amount of tagged ozone, followed by the sensitivity and tagged-N methods. In terms of trans-Pacific influence of ozone pollution, the full-tagged method estimates the strongest impact of East Asian (EA) emissions on the western U.S. (WUS) in MAM and JJA (~3 ppbv), which is substantially different in magnitude and seasonality from tagged-N and sensitivity studies. This difference results from the full-tagged method accounting for the maintenance of peroxy radicals (e.g., CH3O2, CH3CO3, and HO2), in addition to NOy, as effective reservoirs of EA source impact across the Pacific, allowing for a significant contribution to

  1. Evaluation of immobilized metal affinity chromatography kits for the purification of histidine-tagged recombinant CagA protein.

    Science.gov (United States)

    Karakus, Cebrail; Uslu, Merve; Yazici, Duygu; Salih, Barik A

    2016-05-15

    Immobilized metal affinity chromatography (IMAC) technique is used for fast and reliable purification of histidine(His)-tagged recombinant proteins. The technique provides purification under native and denaturing conditions. The aim of this study is to evaluate three commercially available IMAC kits (Thermo Scientific, GE Healthcare and Qiagen) for the purification of a 6xHis-tagged recombinant CagA (cytotoxin-associated gene A) protein from IPTG-induced Escherichia coli BL21(DE3) culture. The kits were tested according to the manufacturer instructions and the protein was purified with only GE Healthcare and Qiagen kits under denaturing conditions. 1% (w/v) SDS was used as denaturing agent in PBS instead of extraction reagent of Thermo Scientific kit to lyse bacterial cells from 100ml culture. The 6xHis-tagged recombinant protein was purified by the three kits equally.

  2. Identification of lncRNA MEG3 Binding Protein Using MS2-Tagged RNA Affinity Purification and Mass Spectrometry.

    Science.gov (United States)

    Liu, Shanshan; Zhu, Juanjuan; Jiang, Taifeng; Zhong, Yiran; Tie, Yi; Wu, Yongge; Zheng, Xiaofei; Jin, Yinghua; Fu, Hanjiang

    2015-08-01

    Long noncoding RNAs (lncRNAs) are nonprotein coding transcripts longer than 200 nucleotides. Recently in mammals, thousands of long noncoding RNAs have been identified and studied as key molecular players in different biological processes with protein complexes. As a long noncoding RNA, maternally expressed gene 3 (MEG3) plays an important role in many cellular processes. However, the mechanism underlying MEG3 regulatory effects remains enigmatic. By using the specific interaction between MS2 coat protein and MS2 RNA hairpin, we developed a method (MS2-tagged RNA affinity purification and mass spectrometry (MTRAP-MS)) to identify proteins that interact with MEG3. Mass spectrometry and gene ontology (GO) analysis showed that MEG3 binding proteins possess nucleotide binding properties and take part in transport, translation, and other biological processes. In addition, interleukin enhancer binding factor 3 (ILF3) and poly(A) binding protein, cytoplasmic 3 (PABPC3) were validated for their interaction with MEG3. These findings indicate that the newly developed method can effectively enrich lncRNA binding proteins and provides a strong basis for studying MEG3 functions.

  3. [Archival tags and geolocation methods for marine animals: A review].

    Science.gov (United States)

    Zhang, Tian-feng; Fan, Wei; Dai, Yang

    2015-11-01

    Archival tags, a group of data storable electronic tags, are widely used as strong tools for obtaining long term and large scale activity information of marine animals, specifically highly migratory oceanic fishes, and corresponding environmental data. Though retrieving tags is an indispensable step for obtaining data, which is a shortage of archival tags, a series of achievements have been made on marine animals by using archival tags since the 1990s. With the appearance of pop-up satellite tag, which solved the problem of data retrieving and was fully independent of the fishing, both breadth and depth of marine animals' studies are extended by the end of the 1990s. Geolocation based on light intensity is the key to estimate marine animals' movement and has achieved some progress in the past 20 years. However, the accuracy of geolocation for latitude is not high enough, and there is still much room for improvement. To date, most geolocation methods that use ambient daylight involve identifying the times when the sun is at a precisely known zenith angle (e.g., sunrise and sunset). The problem of estimating longitude has been proved easy to solve, but accurate latitude estimates remain elusive. This paper mainly introduced two tags, i. e., archival tags and pop-up tags, and three geolocation methods, i.e. , 1) the "fixed reference" method, 2) the "variable reference" method, and 3) the "reflection" method. We also presented a prospect analysis on archival tags and possible research direction of geolocation methods. We believed that miniaturization and multi-sensor integration are the trends for electronic tags while more environmental factors such as depth, SST (sea surface temperature) or magnetic field intensity, instead of single factor, as auxiliary parameters would be used for improving the geolocation accuracy in the future.

  4. Multiple ant colony algorithm method for selecting tag SNPs.

    Science.gov (United States)

    Liao, Bo; Li, Xiong; Zhu, Wen; Li, Renfa; Wang, Shulin

    2012-10-01

    The search for the association between complex disease and single nucleotide polymorphisms (SNPs) or haplotypes has recently received great attention. Finding a set of tag SNPs for haplotyping in a great number of samples is an important step to reduce cost for association study. Therefore, it is essential to select tag SNPs with more efficient algorithms. In this paper, we model problem of selection tag SNPs by MINIMUM TEST SET and use multiple ant colony algorithm (MACA) to search a smaller set of tag SNPs for haplotyping. The various experimental results on various datasets show that the running time of our method is less than GTagger and MLR. And MACA can find the most representative SNPs for haplotyping, so that MACA is more stable and the number of tag SNPs is also smaller than other evolutionary methods (like GTagger and NSGA-II). Our software is available upon request to the corresponding author.

  5. G196 epitope tag system: a novel monoclonal antibody, G196, recognizes the small, soluble peptide DLVPR with high affinity

    Science.gov (United States)

    Tatsumi, Kasumi; Sakashita, Gyosuke; Nariai, Yuko; Okazaki, Kosuke; Kato, Hiroaki; Obayashi, Eiji; Yoshida, Hisashi; Sugiyama, Kanako; Park, Sam-Yong; Sekine, Joji; Urano, Takeshi

    2017-01-01

    The recognition specificity of monoclonal antibodies (mAbs) has made mAbs among the most frequently used tools in both basic science research and in clinical diagnosis and therapies. Precise determination of the epitope allows the development of epitope tag systems to be used with recombinant proteins for various purposes. Here we describe a new family of tag derived from the epitope recognized by a highly specific mAb G196. The minimal epitope was identified as the five amino acid sequence Asp-Leu-Val-Pro-Arg. Permutation analysis was used to characterize the binding requirements of mAb G196, and the variable regions of the mAb G196 were identified and structurally analyzed by X-ray crystallography. Isothermal titration calorimetry revealed the high affinity (Kd = 1.25 nM) of the mAb G196/G196-epitope peptide interaction, and G196-tag was used to detect several recombinant cytosolic and nuclear proteins in human and yeast cells. mAb G196 is valuable for developing a new peptide tagging system for cell biology and biochemistry research. PMID:28266535

  6. Hepatitis C virus expressing flag-tagged envelope protein 2 has unaltered infectivity and density, is specifically neutralized by flag antibodies and can be purified by affinity chromatography

    DEFF Research Database (Denmark)

    Prentø, Jannick Cornelius; Bukh, Jens

    2011-01-01

    Hepatitis C virus (HCV) purification by ultracentrifugation is difficult because of the low and heterogeneous density of native and cultured viruses. It was recently shown that inserting flag tag into envelope protein 2 (E2) of HCV permitted virus purification by affinity chromatography. However...... to the original virus. Flag-tagged virus was susceptible to flag-specific antibody neutralization, and infected cells could be immuno-stained by anti-flag antibodies. Using affinity chromatography with anti-flag resin we repeatedly obtained ~30% recovery of infectious particles. The full viability and unaltered...

  7. Affinity Purification of a Recombinant Protein Expressed as a Fusion with the Maltose-Binding Protein (MBP) Tag

    Science.gov (United States)

    Duong-Ly, Krisna C.; Gabelli, Sandra B.

    2015-01-01

    Expression of fusion proteins such as MBP fusions can be used as a way to improve the solubility of the expressed protein in E. coli (Fox and Waugh, 2003; Nallamsetty et al., 2005; Nallamsetty and Waugh, 2006) and as a way to introduce an affinity purification tag. The protocol that follows was designed by the authors as a first step in the purification of a recombinant protein fused with MBP, using fast protein liquid chromatography (FPLC). Cells should have been thawed, resuspended in binding buffer, and lysed by sonication or microfluidization before mixing with the amylose resin or loading on the column. Slight modifications to this protocol may be made to accommodate both the protein of interest and the availability of equipment. PMID:26096500

  8. Expression of recombinant West Nile virus prM protein fused to an affinity tag for use as a diagnostic antigen.

    Science.gov (United States)

    Setoh, Y X; Hobson-Peters, J; Prow, N A; Young, P R; Hall, R A

    2011-07-01

    Previous studies have concluded that the Flavivirus prM protein is a suitable viral antigen to distinguish serologically between infections with closely related Flaviviruses (Cardosa et al., 2002). To express the recombinant West Nile virus (WNV) prM antigen fused to a suitable affinity tag for purification, a series of prM-His-tag and prM-V5-tag fusion proteins were generated. Analysis of the prM-His-tag fusion proteins revealed that either prM epitopes were disrupted or the His-tag was not presented properly depending on the location of the His tag and the presence of the prM transmembrane domains in these constructs. This identified domains critical for proper folding of prM, and arrangements that allowed the correct presentation of the His-tag. However, the inclusion of the V5 epitope tag fused to the C terminus of prM allowed formation of the authentic antigenic structure of prM and the proper presentation of the V5 epitope. Capture of tagged recombinant WNV(NY99) prM antigen to the solid phase with anti-V5 antibody in ELISA enabled the detection of prM-specific antibodies in WNV(NY99)-immune horse serum, confirming its potential as a useful diagnostic reagent.

  9. A generic protocol for the purification and characterization of water-soluble complexes of affinity-tagged proteins and lipids.

    Science.gov (United States)

    Maeda, Kenji; Poletto, Mattia; Chiapparino, Antonella; Gavin, Anne-Claude

    2014-09-01

    Interactions between lipids and proteins in the aqueous phases of cells contribute to many aspects of cell physiology. Here we describe a detailed protocol to systematically characterize in vivo-assembled complexes of soluble proteins and lipids. Saccharomyces cerevisiae strains expressing physiological amounts of a protein of interest fused to the tandem-affinity purification (TAP) tag are first lysed in the absence of detergent to capture intact protein-lipid complexes. The affinity-purified complexes (typically 30-50 kDa) are subjected to analytical size-exclusion chromatography (SEC) to remove contaminating lipids that elute at the void volume (>600 kDa), in order to achieve sufficient signal-to-background lipid ratios. Proteins in the SEC fractions are then analyzed by denaturing gel electrophoresis. Lipidomics techniques such as high-performance thin-layer chromatography or gas or liquid chromatography-mass spectrometry can then be applied to measure the elution profiles of lipids and to pinpoint the true interactors co-eluting with the TAP fusions. The procedure (starting from cell lysis) requires 2 d, and it can easily be adapted to other organisms.

  10. Immobilised metal-ion affinity chromatography purification of histidine-tagged recombinant proteins : a wash step with a low concentration of EDTA

    NARCIS (Netherlands)

    Westra, DF; Welling, GW; Koedijk, DGAM; Scheffer, AJ; The, TH; Welling-Wester, S

    2001-01-01

    Immobilised metal-ion affinity chromatography (IMAC) is widely used for the purification of recombinant proteins in which a poly-histidine tag is introduced. However, other proteins may also bind to IMAC columns. We describe the use of a washing buffer with a low concentration of EDTA (0.5 mM) for t

  11. Ni(2+)-zeolite/ferrosphere and Ni(2+)-silica/ferrosphere beads for magnetic affinity separation of histidine-tagged proteins.

    Science.gov (United States)

    Vereshchagina, T A; Fedorchak, M A; Sharonova, O M; Fomenko, E V; Shishkina, N N; Zhizhaev, A M; Kudryavtsev, A N; Frank, L A; Anshits, A G

    2016-01-28

    Magnetic Ni(2+)-zeolite/ferrosphere and Ni(2+)-silica/ferrosphere beads (Ni-ferrosphere beads - NFB) of a core-shell structure were synthesized starting from coal fly ash ferrospheres having diameters in the range of 0.063-0.050 mm. The strategy of NFB fabrication is an oriented chemical modification of the outer surface preserving the magnetic core of parent beads with the formation of micro-mesoporous coverings. Two routes of ferrosphere modification were realized, such as (i) hydrothermal treatment in an alkaline medium resulting in a NaP zeolite layer and (ii) synthesis of micro-mesoporous silica on the glass surface using conventional methods. Immobilization of Ni(2+) ions in the siliceous porous shell of the magnetic beads was carried out via (i) the ion exchange of Na(+) for Ni(2+) in the zeolite layer or (ii) deposition of NiO clusters in the zeolite and silica pores. The final NFB were tested for affinity in magnetic separation of the histidine-tagged green fluorescent protein (GFP) directly from a cell lysate. Results pointed to the high affinity of the magnetic beads towards the protein in the presence of 10 mM EDTA. The sorption capacity of the ferrosphere-based Ni-beads with respect to GFP was in the range 1.5-5.7 mg cm(-3).

  12. Method for Flavor Tagging in Neutral B Meson Decays

    OpenAIRE

    1992-01-01

    A method is proposed for tagging the flavor of neutral $B$ mesons in the study of CP-violating decay asymmetries. The method makes use of a possible difference in interactions in $B \\pi$ or $B^* \\pi$ systems with isospins 1/2 and 3/2, and would be particularly clean if the $I = 1/2$ systems can be detected as ``$B^{**}$'' resonances.

  13. Method for flavor tagging in neutral B meson decays

    CERN Document Server

    Gronau, Michael; Rosner, Jonathan L; Gronau, Michael; Nippe, Alex; Rosner, Jonathan L.

    1993-01-01

    A method is proposed for tagging the flavor of neutral $B$ mesons in the study of CP-violating decay asymmetries. The method makes use of a possible difference in interactions in $B \\pi$ or $B^* \\pi$ systems with isospins 1/2 and 3/2, and would be particularly clean if the $I = 1/2$ systems can be detected as ``$B^{**}$'' resonances.

  14. Method for flavor tagging in neutral B meson decays

    Science.gov (United States)

    Gronau, Michael; Nippe, Alex; Rosner, Jonathan L.

    1993-03-01

    A method is proposed for tagging the flavor of neutral B mesons in the study of CP-violating decay asymmetries. The method makes use of a possible difference in interactions in Bπ or B*π systems with isospins 1/2 and 3/2, and would be particularly clean if the I=1/2 systems can be detected as ``B**'' resonances.

  15. Communication methods, systems, apparatus, and devices involving RF tag registration

    Science.gov (United States)

    Burghard, Brion J.; Skorpik, James R.

    2008-04-22

    One technique of the present invention includes a number of Radio Frequency (RF) tags that each have a different identifier. Information is broadcast to the tags from an RF tag interrogator. This information corresponds to a maximum quantity of tag response time slots that are available. This maximum quantity may be less than the total number of tags. The tags each select one of the time slots as a function of the information and a random number provided by each respective tag. The different identifiers are transmitted to the interrogator from at least a subset of the RF tags.

  16. Moment-Based Method to Estimate Image Affine Transform

    Institute of Scientific and Technical Information of China (English)

    FENG Guo-rui; JIANG Ling-ge

    2005-01-01

    The estimation of affine transform is a crucial problem in the image recognition field. This paper resorted to some invariant properties under translation, rotation and scaling, and proposed a simple method to estimate the affine transform kernel of the two-dimensional gray image. Maps, applying to the original, produce some correlative points that can accurately reflect the affine transform feature of the image. Furthermore, unknown variables existing in the kernel of the transform are calculated. The whole scheme only refers to one-order moment,therefore, it has very good stability.

  17. Integrative refolding and purification of histidine-tagged protein by like-charge facilitated refolding and metal-chelate affinity adsorption.

    Science.gov (United States)

    Liu, Hu; Du, Wen-Jie; Dong, Xiao-Yan; Sun, Yan

    2014-05-30

    This work proposed an integrative method of protein refolding and purification by like-charged resin facilitated refolding and metal-chelate affinity adsorption. Hexahistidine-tagged enhanced green fluorescence protein (EGFP) was overexpressed in Escherichia coli as inclusion bodies (IBs), and then the protein was refolded and purified from urea-solubilized IBs by this method. A metal-chelating resin was fabricated by coupling iminodiacetic acid (IDA) to agarose gel (Sepharose FF). The anionic resin was used to facilitate the refolding of like-charged EGFP from IBs. After refolding, nickel ions were introduced for the affinity purification of the target protein by metal-chelating adsorption. It was found that the resin was effective in facilitating EGFP refolding. For 0.1mg/mL EGFP IBs refolding, the fluorescence recovery (FR) by direct dilution was only 64%; addition of only 0.05 g/mL resin increased the FR to over 90%. Moreover, the FR increased with increasing resin concentration. Owning to the shielding effect of the oppositely charged impurities embedded in IBs on the surface charges of the IDA resin, more resin particles were required to exert an aggregation inhibition effect in the IBs protein refolding. Additionally, compared with direct-dilution refolding, inclusion of like-charged resins not only offered an enhanced FR of EGFP, but also bound some opposite-charged contaminant proteins, leading to a preliminary purification effect. Afterwards, the refolded EGFP was recovered by metal-chelating adsorption at an FR of 85% and purity of 93%. This work has thus extended the like-charge facilitated protein refolding strategy to the integrative protein refolding and purification.

  18. Studies with an immobilized metal affinity chromatography cassette system involving binuclear triazacyclononane-derived ligands: automation of batch adsorption measurements with tagged recombinant proteins.

    Science.gov (United States)

    Petzold, Martin; Coghlan, Campbell J; Hearn, Milton T W

    2014-07-18

    This study describes the determination of the adsorption isotherms and binding kinetics of tagged recombinant proteins using a recently developed IMAC cassette system and employing automated robotic liquid handling procedures for IMAC resin screening. These results confirm that these new IMAC resins, generated from a variety of different metal-charged binuclear 1,4,7-triaza-cyclononane (tacn) ligands, interact with recombinant proteins containing a novel N-terminal metal binding tag, NT1A, with static binding capacities similar to those obtained with conventional hexa-His tagged proteins, but with significantly increased association constants. In addition, higher kinetic binding rates were observed with these new IMAC systems, an attribute that can be positively exploited to increase process productivity. The results from this investigation demonstrate that enhancements in binding capacities and affinities were achieved with these new IMAC resins and chosen NT1A tagged protein. Further, differences in the binding performances of the bis(tacn) xylenyl-bridged ligands were consistent with the distance between the metal binding centres of the two tacn moieties, the flexibility of the ligand and the potential contribution from the aromatic ring of the xylenyl group to undergo π/π stacking interactions with the tagged proteins.

  19. Tag, you're it: tagging as an alternative to yes/no recognition in item method directed forgetting.

    Science.gov (United States)

    Thompson, Kate M; Fawcett, Jonathan M; Taylor, Tracy L

    2011-09-01

    The current study contrasted a standard yes/no recognition task with a tagging recognition task in the context of an item-method directed forgetting paradigm. During the study phase, a series of words was presented one at a time, each followed by an instruction to remember (R) or forget (F). The retention of R and F study words was tested using either a typical yes/no recognition task or a tagging recognition task in which participants labeled each word as "R", "F" or "New". The directed forgetting effect observed in each task was equivalent in magnitude. However, the tagging recognition task afforded an additional analysis of the errors of misattribution that was not possible with the more typical yes/no recognition task. Interestingly, when falsely recognizing a Foil word, participants were more likely to assign an "F" tag than an "R" tag. These errors of misattribution are consistent with existing accounts of directed forgetting that suggest R words are better encoded than F words. We argue for the utility of the tagging procedure, given it does not alter the directed forgetting effect normally seen with yes/no recognition but provides additional information about errors of misattribution.

  20. A statistical method for assessing peptide identification confidence in accurate mass and time tag proteomics.

    Science.gov (United States)

    Stanley, Jeffrey R; Adkins, Joshua N; Slysz, Gordon W; Monroe, Matthew E; Purvine, Samuel O; Karpievitch, Yuliya V; Anderson, Gordon A; Smith, Richard D; Dabney, Alan R

    2011-08-15

    Current algorithms for quantifying peptide identification confidence in the accurate mass and time (AMT) tag approach assume that the AMT tags themselves have been correctly identified. However, there is uncertainty in the identification of AMT tags, because this is based on matching LC-MS/MS fragmentation spectra to peptide sequences. In this paper, we incorporate confidence measures for the AMT tag identifications into the calculation of probabilities for correct matches to an AMT tag database, resulting in a more accurate overall measure of identification confidence for the AMT tag approach. The method is referenced as Statistical Tools for AMT Tag Confidence (STAC). STAC additionally provides a uniqueness probability (UP) to help distinguish between multiple matches to an AMT tag and a method to calculate an overall false discovery rate (FDR). STAC is freely available for download, as both a command line and a Windows graphical application.

  1. A Quick and Affine Invariance Matching Method for Oblique Images

    Directory of Open Access Journals (Sweden)

    XIAO Xiongwu

    2015-04-01

    Full Text Available This paper proposed a quick, affine invariance matching method for oblique images. It calculated the initial affine matrix by making full use of the two estimated camera axis orientation parameters of an oblique image, then recovered the oblique image to a rectified image by doing the inverse affine transform, and left over by the SIFT method. We used the nearest neighbor distance ratio(NNDR, normalized cross correlation(NCC measure constraints and consistency check to get the coarse matches, then used RANSAC method to calculate the fundamental matrix and the homography matrix. And we got the matches that they were interior points when calculating the homography matrix, then calculated the average value of the matches' principal direction differences. During the matching process, we got the initial matching features by the nearest neighbor(NN matching strategy, then used the epipolar constrains, homography constrains, NCC measure constrains and consistency check of the initial matches' principal direction differences with the calculated average value of the interior matches' principal direction differences to eliminate false matches. Experiments conducted on three pairs of typical oblique images demonstrate that our method takes about the same time as SIFT to match a pair of oblique images with a plenty of corresponding points distributed evenly and an extremely low mismatching rate.

  2. Parameters of explosives detection through tagged neutron method

    Energy Technology Data Exchange (ETDEWEB)

    Bagdasaryan, Kh.E.; Batyaev, V.F.; Belichenko, S.G., E-mail: consul757@mail.ru; Bestaev, R.R.; Gavryuchenkov, A.V.; Karetnikov, M.D.

    2015-06-01

    The potentialities of tagged neutron method (TNM) for explosives detection are examined on the basis of an idealized geometrical model. The model includes ING-27 14 MeV neutron generator with a built-in α-detector, a LYSO γ-detector and samples of material to be identified of approximately 0.3 kg each: explosives imitators (trinitrotoluene - TNT, tetryl, RDX and ammonium nitrate), legal materials (sugar, water, silk and polyethylene). The samples were unshielded or shielded by a paper layer of various thicknesses. The experimental data were interpreted by numerical simulation using a Poisson distribution of signals with the statistical parameters defined experimentally. The detection parameters were obtained by a pattern classification theory and a Bayes classifier.

  3. Peculiarities of Thermodynamic Simulation with the Method of Bound Affinity

    CERN Document Server

    Zilbergleyt, B

    2004-01-01

    Thermodynamic simulation of chemical and metallurgical systems is the only method to predict their equilibrium composition and is the most important application of chemical thermodynamics. The conventional strategy of simulation is always to find the most probable composition of the system, corresponding to thermodynamic equilibrium. Traditional simulation methods do not account for interactions within the chemical system. The Method of Bound Affinity (MBA) is based on the theory that explicitly takes into account interactions between subsystems of a complex chemical system and leads sometimes to essential differences in simulation results. This article discusses peculiarities of MBA application, exemplified by results for a complex system with a set of subsystems.

  4. A novel affinity purification method to isolate peptide specific antibodies

    DEFF Research Database (Denmark)

    Karlsen, Alan E; Lernmark, A; Kofod, Hans

    1990-01-01

    Site-specific, high affinity polyclonal antisera are effectively and successfully produced by immunizing rabbits with synthetic peptides. The use of these antisera in subsequent immune analysis is often limited because of non-specific binding. We describe a new and simple method to effectively...... affinity-purify anti-peptide antibodies. To test our system, rabbits were immunized with model peptides representing sequences of the putative rabbit growth hormone receptor and several HLA-DQ beta-chain molecules. Polystyrene plastic beads were coated with peptides. Immune serum was incubated...... with the beads and after a wash step the bound antibodies were eluted in 1 M acetic acid. The eluted material was composed predominantly of intact immunoglobulin as evidenced by the presence of heavy and light chain bands in SDS-PAGE. The eluted antibodies were peptide specific in ELISA and bound only to intact...

  5. An angular momentum conserving Affine-Particle-In-Cell method

    CERN Document Server

    Jiang, Chenfanfu; Teran, Joseph

    2016-01-01

    We present a new technique for transferring momentum and velocity between particles and grid with Particle-In-Cell (PIC) calculations which we call Affine-Particle-In-Cell (APIC). APIC represents particle velocities as locally affine, rather than locally constant as in traditional PIC. We show that this representation allows APIC to conserve linear and angular momentum across transfers while also dramatically reducing numerical diffusion usually associated with PIC. Notably, conservation is achieved with lumped mass, as opposed to the more commonly used Fluid Implicit Particle (FLIP) transfers which require a 'full' mass matrix for exact conservation. Furthermore, unlike FLIP, APIC retains a filtering property of the original PIC and thus does not accumulate velocity modes on particles as FLIP does. In particular, we demonstrate that APIC does not experience velocity instabilities that are characteristic of FLIP in a number of Material Point Method (MPM) hyperelasticity calculations. Lastly, we demonstrate th...

  6. Efficient production and purification of recombinant human interleukin-12 (IL-12) overexpressed in mammalian cells without affinity tag

    Science.gov (United States)

    Jayanthi, Srinivas; Koppolu, Bhanu prasanth; Smith, Sean G.; Jalah, Rashmi; Bear, Jenifer; Rosati, Margherita; Pavlakis, George N.; Felber, Barbara K.; Zaharoff, David A.; Kumar, Thallapuranam Krishnaswamy Suresh

    2014-01-01

    Interleukin-12 is a heterodimeric, pro-inflammatory cytokine that is a key driver of cell-mediated immunity. Clinical interest in IL-12 is significant due to its potent anti-tumor activity and efficacy in controlling certain infectious diseases such as Leishmaniasis and Listeria infection. For clinical applications, the ease of production and purification of IL-12 and the associated cost continues to be a consideration. In this context, we report a simple and effective heparin-affinity based purification of recombinant human IL-12 (hIL-12) from the serum-free supernatants of stable IL-12-transduced HEK293 cells. Fractionation of culture supernatants on heparin Sepharose columns revealed that hIL-12 elutes as a single peak in 500 mM NaCl. Coomassie staining and Western blot analysis showed that hIL-12 eluted in 500 mM NaCl is homogeneous.Purity of hIL-12 was ascertained by RP-HPLC and ESI-MS analysis, and found to be ~98%. Western blot analysis, using monoclonal antibodies, demonstrated that the crucial inter-subunit disulfide bond linking the p35 and p40 subunits is intact in the purified hIL-12. Results of far UV circular dichrosim, steady-state tryptophan fluorescence, and differential scanning calorimetry experiments suggest that purified hIL-12 is in its stable native conformation. Enzyme linked immunosorbent assays (ELISAs) and bioactivity studies demonstrate that hIL-12 is obtained in high yields (0.31 ± 0.05 mg/ mL of the culture medium) and is also fully bioactive. Isothermal titration calorimetry data show that IL-12 exhibits a moderate binding affinity (Kd(app) = 69 ± 1 μM) to heparin. The purification method described in this study is expected to provide greater impetus for research on the role of heparin in the regulation of the function of IL-12. In addition, the results of this study provide an avenue to obtain high amounts of IL-12 required for structural studies which are aimed at the development of novel IL-12-based therapeutics. PMID:25123642

  7. System Virtualization and Efficient ID Transmission Method for RFID Tag Infrastructure Network

    Directory of Open Access Journals (Sweden)

    Shin-ichi Kuribayashi

    2010-11-01

    Full Text Available The use of RFID tag which identifies a thing and an object will be expanded with progress of ubiquitoussociety, and it is necessary to study how to construct RFID network system as a social infrastructure likethe Internet. First, this paper proposes the virtualization method of RFID tag network system to enablethe same physical RFID network system to be used by multiple different service systems. The systemvirtualization not only reduces the system cost but also can dramatically reduce the installation space ofphysical readers and the operation cost. It is proposed that all equipments in the RFID network systemexcept RFID tag could be shared with the conventional virtual technologies. Then, this paper proposesthe conditional tag ID processing and the efficient tag ID transmission method which can greatly reducethe processing time and processing load in RFID tag Infrastructure network The conditional tag IDprocessing allows that tag ID is valid only at a certain time zone of day or in a certain area. The efficienttag ID transmission method uses the virtual network address of the service center as a part of the ID of anRF tag, which allows the direct ID forwarding to the service center.

  8. Design, synthesis, and application of a hydrazide-functionalized isotope-coded affinity tag for the quantification of oxylipid-protein conjugates.

    Science.gov (United States)

    Han, Bingnan; Stevens, Jan F; Maier, Claudia S

    2007-05-01

    An isotopically coded affinity probe was developed and evaluated for the characterization and quantification of proteins adducted by 2-alkenals derived from lipid peroxidation (LPO) processes. Lipid-derived 2-alkenals, such as acrolein and 4-hydroxy-2-nonenal (HNE), have the ability to react with cysteine, histidine, and lysine residues in proteins, thus causing protein damage and loss of protein function. Such modifications of proteins are difficult to characterize in biological samples by mass spectrometry due to the complexity of protein extracts and the low abundance of adducted proteins. The novel aldehyde-reactive, hydrazide-functionalized, isotope-coded affinity tag (HICAT) described in this study was found effective for the selective isolation, detection, and quantification of Michael-type adducts of 2-alkenals with proteins using a combination of affinity isolation, nanoLC, and matrix-assisted laser desorption ionization tandem mass spectrometry (MALDI-MS/MS). The chemical and mass spectrometric properties of the new probe are demonstrated on a model protein treated with HNE. The efficacy of HICAT for the analysis of complex samples was tested using preparations of mitochondrial proteins that were modified in vitro with HNE. The potential of the HICAT strategy for the identification, characterization, and quantification of in vivo oxylipid-protein conjugates is demonstrated on cardiac mitochondrial protein preparations, in which, for example, the ADP/ATP translocase 1 was found adducted to the 2-alkenals, acrolein and 4-hydroxy-2-hexenal, at Cys-256.

  9. Detection of protein-protein interactions using tandem affinity purification.

    Science.gov (United States)

    Goodfellow, Ian; Bailey, Dalan

    2014-01-01

    Tandem affinity purification (TAP) is an invaluable technique for identifying interaction partners for an affinity tagged bait protein. The approach relies on the fusion of dual tags to the bait before separate rounds of affinity purification and precipitation. Frequently two specific elution steps are also performed to increase the specificity of the overall technique. In the method detailed here, the two tags used are protein G and a short streptavidin binding peptide; however, many variations can be employed. In our example the tags are separated by a cleavable tobacco etch virus protease target sequence, allowing for specific elution after the first round of affinity purification. Proteins isolated after the final elution step in this process are concentrated before being identified by mass spectrometry. The use of dual affinity tags and specific elution in this technique dramatically increases both the specificity and stringency of the pull-downs, ensuring a low level of background nonspecific interactions.

  10. Immobilized metal affinity chromatography in open-loop simulated moving bed technology: purification of a heat stable histidine tagged beta-glucosidase.

    Science.gov (United States)

    Sahoo, Deepti; Andersson, Jonatan; Mattiasson, Bo

    2009-06-01

    Open-loop simulated moving bed (SMB) has been used for immobilized metal affinity chromatographic (IMAC) purification of his-tagged beta-glucosidase expressed in E. coli. A simplified approach based on an optimized single column protocol is used to design the open-loop SMB. A set of columns in the SMB represent one step in the chromatographic cycle i.e. there will be one set each of columns for load, wash, elution etc within the SMB. Only the wash and elution are operated with columns in sequence. The beta-glucosidase was purified to almost single band purity with a purification factor of 15 and a recovery of 91%. SMB-performance showed reduced buffer consumption, higher purification fold, a better yield and higher productivity.

  11. System Virtualization and Efficient ID Transmission Method for RFID Tag Infrastructure Network

    CERN Document Server

    Kuribayashi, Shin-ichi

    2010-01-01

    The use of RFID tag which identifies a thing and an object will be expanded with progress of ubiquitous society, and it is necessary to study how to construct RFID network system as a social infrastructure like the Internet. First, this paper proposes the virtualization method of RFID tag network system to enable the same physical RFID network system to be used by multiple different service systems. The system virtualization not only reduces the system cost but also can dramatically reduce the installation space of physical readers and the operation cost. It is proposed that all equipments in the RFID network system except RFID tag could be shared with the conventional virtual technologies. Then, this paper proposes the conditional tag ID processing and the efficient tag ID transmission method which can greatly reduce the processing time and processing load in RFID tag Infrastructure network The conditional tag ID processing allows that tag ID is valid only at a certain time zone of day or in a certain area. ...

  12. Methods for determining the genetic affinity of microorganisms and viruses

    Science.gov (United States)

    Fox, George E. (Inventor); Willson, III, Richard C. (Inventor); Zhang, Zhengdong (Inventor)

    2012-01-01

    Selecting which sub-sequences in a database of nucleic acid such as 16S rRNA are highly characteristic of particular groupings of bacteria, microorganisms, fungi, etc. on a substantially phylogenetic tree. Also applicable to viruses comprising viral genomic RNA or DNA. A catalogue of highly characteristic sequences identified by this method is assembled to establish the genetic identity of an unknown organism. The characteristic sequences are used to design nucleic acid hybridization probes that include the characteristic sequence or its complement, or are derived from one or more characteristic sequences. A plurality of these characteristic sequences is used in hybridization to determine the phylogenetic tree position of the organism(s) in a sample. Those target organisms represented in the original sequence database and sufficient characteristic sequences can identify to the species or subspecies level. Oligonucleotide arrays of many probes are especially preferred. A hybridization signal can comprise fluorescence, chemiluminescence, or isotopic labeling, etc.; or sequences in a sample can be detected by direct means, e.g. mass spectrometry. The method's characteristic sequences can also be used to design specific PCR primers. The method uniquely identifies the phylogenetic affinity of an unknown organism without requiring prior knowledge of what is present in the sample. Even if the organism has not been previously encountered, the method still provides useful information about which phylogenetic tree bifurcation nodes encompass the organism.

  13. Identification of protein partners in mycobacteria using a single-step affinity purification method.

    Directory of Open Access Journals (Sweden)

    Przemysław Płociński

    Full Text Available Tuberculosis is a leading cause of death in developing countries. Efforts are being made to both prevent its spread and improve curability rates. Understanding the biology of the bacteria causing the disease, Mycobacterium tuberculosis (M. tuberculosis, is thus vital. We have implemented improved screening methods for protein-protein interactions based on affinity purification followed by high-resolution mass spectrometry. This method can be efficiently applied to both medium- and high-throughput studies aiming to characterize protein-protein interaction networks of tubercle bacilli. Of the 4 tested epitopes FLAG, enhanced green fluorescent protein (eGFP, protein A and haemagglutinin, the eGFP tag was found to be most useful on account of its easily monitored expression and its ability to function as a simultaneous tool for subcellular localization studies. It presents a relatively low background with cost-effective purification. RNA polymerase subunit A (RpoA was used as a model for investigation of a large protein complex. When used as bait, it co-purified with all remaining RNA polymerase core subunits as well as many accessory proteins. The amount of RpoA strongly correlated with the amount of quantification peptide used as part of the tagging system in this study (SH, making it applicable for semi-quantification studies. Interactions between the components of the RpoA-eGFP protein complex were further confirmed using protein cross-linking. Dynamic changes in the composition of protein complexes under induction of UV damage were observed when UvrA-eGFP expressing cells treated with UV light were used to co-purify UvrA interaction partners.

  14. Identification of protein partners in mycobacteria using a single-step affinity purification method.

    Science.gov (United States)

    Płociński, Przemysław; Laubitz, Daniel; Cysewski, Dominik; Stoduś, Krystian; Kowalska, Katarzyna; Dziembowski, Andrzej

    2014-01-01

    Tuberculosis is a leading cause of death in developing countries. Efforts are being made to both prevent its spread and improve curability rates. Understanding the biology of the bacteria causing the disease, Mycobacterium tuberculosis (M. tuberculosis), is thus vital. We have implemented improved screening methods for protein-protein interactions based on affinity purification followed by high-resolution mass spectrometry. This method can be efficiently applied to both medium- and high-throughput studies aiming to characterize protein-protein interaction networks of tubercle bacilli. Of the 4 tested epitopes FLAG, enhanced green fluorescent protein (eGFP), protein A and haemagglutinin, the eGFP tag was found to be most useful on account of its easily monitored expression and its ability to function as a simultaneous tool for subcellular localization studies. It presents a relatively low background with cost-effective purification. RNA polymerase subunit A (RpoA) was used as a model for investigation of a large protein complex. When used as bait, it co-purified with all remaining RNA polymerase core subunits as well as many accessory proteins. The amount of RpoA strongly correlated with the amount of quantification peptide used as part of the tagging system in this study (SH), making it applicable for semi-quantification studies. Interactions between the components of the RpoA-eGFP protein complex were further confirmed using protein cross-linking. Dynamic changes in the composition of protein complexes under induction of UV damage were observed when UvrA-eGFP expressing cells treated with UV light were used to co-purify UvrA interaction partners.

  15. A Practical Method for UHF RFID Interrogation Area Measurement Using Battery Assisted Passive Tag

    Science.gov (United States)

    Mitsugi, Jin; Tokumasu, Osamu

    For the success of a large deployment of UHF RFID, easyto-use and low-cost engineering tools to facilitate the performance evaluation are demanded particularly in installations and for trouble shooting. The measurement of interrogation area is one of the most typical industrial demands to establish the stable readability of UHF RFID. Exhaustive repetition of tag position change with a read operation and a usage of expensive measurement equipment or special interrogators are common practices to measure the interrogation area. In this paper, a practical method to measure the interrogation area of a UHF RFID by using a battery assisted passive tag (BAP) is presented. After introducing the fundamental design and performances of the BAP that we have developed, we introduce the measurement method. In the method, the target tag in the target installation is continuously traversed either manually or automatically while it is subjected to a repetitive read of a commercial interrogator. During the target tag traversal, the interrogator's commands are continuously monitored by a BAP. With an extensive analysis on interrogator commands, the BAP can differentiate between its own read timings and those of the target tag. The read timings of the target tag collected by the BAP are recorded synchronously with the target tag position, yielding a map of the interrogation area. The present method does not entail a measurement burden. It is also independent of the choice of interrogator and tag. The method is demonstrated in a practical UHF RFID installation to show that the method can measure a 40mm resolution interrogation area measurement just by traversing the target tag at a slow walking speed, 300mm/sec.

  16. Development of a Method for Tissue Elasticity Imaging Using Tagged Magnetic Resonance Imaging

    CERN Document Server

    Takeuchi, Tomoki; Murase, Kenya

    2016-01-01

    The purpose of this study was to develop a method for tissue elasticity imaging using tagged magnetic resonance imaging (MRI). First, we developed a cyclic pressure device that used air to remotely transmit the power to generate cyclic deformation in an object. The pressure induced by the cyclic pressure device was measured by MRI-compatible force sensors. Second, we developed a software to calculate Young's modulus from tagged MRI data using the harmonic phase (HARP) method and the finite element method (FEM). We also developed a software to extract tag-cross points from tagged MRI data. Finally, we evaluated the usefulness of our method using three homogeneous silicone gel phantoms with different degrees of stiffness in comparison with Young's moduli measured by a material testing machine. The coefficient of variation of the pressure data measured by MRI-compatible force sensors was within 5 %, indicating that the reproducibility of the pressure generated by our cyclic pressure device was good. The Young's ...

  17. PDZ affinity chromatography: a general method for affinity purification of proteins based on PDZ domains and their ligands.

    Science.gov (United States)

    Walkup, Ward G; Kennedy, Mary B

    2014-06-01

    PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ∼90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized that the specific, strong interactions of PDZ domains with their ligands would make them well suited for use in affinity chromatography. Here we describe a novel affinity chromatography method applicable for the purification of proteins that contain PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We created a series of affinity resins comprised of PDZ domains from the scaffold protein PSD-95, or from neuronal nitric oxide synthase (nNOS), coupled to solid supports. We used them to purify heterologously expressed neuronal proteins or protein domains containing endogenous PDZ domain ligands, eluting the proteins with free PDZ domain peptide ligands. We show that Proteins of Interest (POIs) lacking endogenous PDZ domain ligands can be engineered as fusion products containing C-terminal PDZ domain ligand peptides or internal, N- or C-terminal PDZ domains and then can be purified by the same method. Using this method, we recovered recombinant GFP fused to a PDZ domain ligand in active form as verified by fluorescence yield. Similarly, chloramphenicol acetyltransferase (CAT) and β-Galactosidase (LacZ) fused to a C-terminal PDZ domain ligand or an N-terminal PDZ domain were purified in active form as assessed by enzymatic assay. In general, PDZ domains and ligands derived from PSD-95 were superior to those from nNOS for this method. PDZ Domain Affinity Chromatography promises to be a versatile and effective method for purification of a wide variety of natural and recombinant proteins.

  18. Extended Range Passive Wireless Tag System and Method

    Science.gov (United States)

    Fink, Patrick W. (Inventor); Kennedy, Timothy F. (Inventor); Lin, Gregory Y. (Inventor)

    2013-01-01

    A passive wireless tag assembly comprises a plurality of antennas and transmission lines interconnected with circuitry and constructed and arranged in a Van Atta array or configuration to reflect an interrogator signal in the direction from where it came. The circuitry may comprise at least one surface acoustic wave (SAW)-based circuit that functions as a signal reflector and is operatively connected with an information circuit. In another embodiment, at least one delay circuit and/or at least one passive modulation circuit(s) are utilized. In yet another embodiment, antennas connected to SAW-based devices are mounted to at least one of the orthogonal surfaces of a corner reflector.

  19. Method of remote powering and detecting multiple UWB passive tags in an RFID system

    Science.gov (United States)

    Dowla, Farid U [Castro Valley, CA; Nekoogar, Faranak [San Ramon, CA; Benzel, David M [Livermore, CA; Dallum, Gregory E [Livermore, CA; Spiridon, Alex [Palo Alto, CA

    2012-05-29

    A new Radio Frequency Identification (RFID), tracking, powering apparatus/system and method using coded Ultra-wideband (UWB) signaling is introduced. The proposed hardware and techniques disclosed herein utilize a plurality of passive UWB transponders in a field of an RFID-radar system. The radar system itself enables multiple passive tags to be remotely powered (activated) at about the same time frame via predetermined frequency UWB pulsed formats. Once such tags are in an activated state, an UWB radar transmits specific "interrogating codes" to put predetermined tags in an awakened status. Such predetermined tags can then communicate by a unique "response code" so as to be detected by an UWB system using radar methods.

  20. Modified PCR methods for 3' end amplification from serial analysis of gene expression (SAGE) tags.

    Science.gov (United States)

    Xu, Wang-Jie; Wang, Zhao-Xia; Qiao, Zhong-Dong

    2009-05-01

    Serial analysis of gene expression (SAGE) is a powerful technique to study gene expression at the genome level. However, a disadvantage of the shortness of SAGE tags is that it prevents further study of SAGE library data, thus limiting extensive application of the SAGE method in gene expression studies. However, this problem can be solved by extension of the SAGE tags to 3' cDNAs. Therefore, several methods based on PCR have been developed to generate a 3' longer fragment cDNA corresponding to a SAGE tag. The list of modified methods is extensive, and includes rapid RT-PCR analysis of unknown SAGE tags (RAST-PCR), generation of longer cDNA fragments from SAGE tags for gene identification (GLGI), a high-throughput GLGI procedure, reverse SAGE (rSAGE), two-step analysis of unknown SAGE tags (TSAT-PCR), etc. These procedures are constantly being updated because they have characteristics and advantages that can be shared. Development of these methods has promoted the widespread use of the SAGE technique, and has accelerated the speed of studies of large-scale gene expression.

  1. Prediction of peptide bonding affinity: kernel methods for nonlinear modeling

    CERN Document Server

    Bergeron, Charles; Sundling, C Matthew; Krein, Michael; Katt, Bill; Sukumar, Nagamani; Breneman, Curt M; Bennett, Kristin P

    2011-01-01

    This paper presents regression models obtained from a process of blind prediction of peptide binding affinity from provided descriptors for several distinct datasets as part of the 2006 Comparative Evaluation of Prediction Algorithms (COEPRA) contest. This paper finds that kernel partial least squares, a nonlinear partial least squares (PLS) algorithm, outperforms PLS, and that the incorporation of transferable atom equivalent features improves predictive capability.

  2. On the Finite Convergence of Newton-type Methods for P0 Affine Variational Inequalities

    Institute of Scientific and Technical Information of China (English)

    Li Ping ZHANG; Wen Xun XING

    2007-01-01

    Based on the techniques used in non-smooth Newton methods and regularized smoothing Newton methods, a Newton-type algorithm is proposed for solving the P0 affine variational inequality problem. Under mild conditions, the algorithm can find an exact solution of the P0 affine variational inequality problem in finite steps. Preliminary numerical results indicate that the algorithm is promis-ing.

  3. New Tagging Method of B Flavor of Neutral B Meson in CP Violation Measurement in Asymmetric B-Factory Experiment

    Science.gov (United States)

    Enomoto, Ryoji

    1994-10-01

    In CP violation measurements in asymmetric B-factory experiments, a determination of the B flavor of the neutral B mesons is necessary. A new method to this purpose using only three vectors of charged particles has been developed. This method (weighted charge method) does not require either lepton identification or charged-kaon identification. The tagging efficiency, probability for incorrect tagging, and effective tagging efficiency of this method are 43.1, 18.3, and 17.3%, respectively.

  4. New tagging method of B flavor of neutral B meson in CP violation measurement in asymmetric b-factory experiment

    CERN Document Server

    Enomoto, R

    1994-01-01

    In CP violation measurements in asymmetric B-factory experiments, a determination of the B flavor of the neutral B mesons is necessary. A new method to this purpose using only three vectors of charged particles has been developed. This method (weighted charge method) does not require either lepton identification or charged-kaon identification. The tagging efficiency, probability for incorrect tagging, and effective tagging efficiency of this method are 43.1, 18.3, and 17.3\\%, respectively.

  5. New Tagging Method of B Flavor of Neutral B Meson in CP Violation Measurement in Asymmetric B-Factory Experiment

    OpenAIRE

    1994-01-01

    In CP violation measurements in asymmetric B-factory experiments, a determination of the B flavor of the neutral B mesons is necessary. A new method to this purpose using only three vectors of charged particles has been developed. This method (weighted charge method) does not require either lepton identification or charged-kaon identification. The tagging efficiency, probability for incorrect tagging, and effective tagging efficiency of this method are 43.1, 18.3, and 17.3\\%, respectively.

  6. Cyanine-based probe\\tag-peptide pair fluorescence protein imaging and fluorescence protein imaging methods

    Science.gov (United States)

    Mayer-Cumblidge, M. Uljana; Cao, Haishi

    2013-01-15

    A molecular probe comprises two arsenic atoms and at least one cyanine based moiety. A method of producing a molecular probe includes providing a molecule having a first formula, treating the molecule with HgOAc, and subsequently transmetallizing with AsCl.sub.3. The As is liganded to ethanedithiol to produce a probe having a second formula. A method of labeling a peptide includes providing a peptide comprising a tag sequence and contacting the peptide with a biarsenical molecular probe. A complex is formed comprising the tag sequence and the molecular probe. A method of studying a peptide includes providing a mixture containing a peptide comprising a peptide tag sequence, adding a biarsenical probe to the mixture, and monitoring the fluorescence of the mixture.

  7. SILAC-iPAC: a quantitative method for distinguishing genuine from non-specific components of protein complexes by parallel affinity capture.

    Science.gov (United States)

    Rees, Johanna S; Lilley, Kathryn S; Jackson, Antony P

    2015-02-06

    Pull-down assays can identify members of protein complexes but suffer from co-isolation of contaminants. The problem is particularly acute when the specifically interacting partners are of low-abundance and/or bind transiently with low affinity. To differentiate true interacting partners from contaminants, we have combined SILAC labelling with a proteomic method called "Interactomes by Parallel Affinity Capture" (iPAC). In our method, a cell-line stably expressing a doubly tagged target endogenous protein and its tag-less control cell-line are differentially SILAC labelled. Lysates from the two cell-lines are mixed and the tagged protein is independently purified for MS analysis using multiple affinity resins in parallel. This allows the quantitative identification of tagged proteins and their binding partners. SILAC-iPAC provides a rigorous and sensitive approach that can discriminate between genuine binding partners and contaminants, even when the contaminants in the pull-down are in large excess. We employed our method to examine the interacting partners of phosphatidyl inositol 5-phosphate 4-kinase 2β subunit (PI5P4K2β) and the Fanconi anaemia core complex in the chicken pre-B cell-line DT40. We confirmed known components of these two complexes, and we have identified new potential binding partners. Combining the iPAC approach with SILAC labelling provides a sensitive and fully quantitative method for the discrimination of specific interactions under conditions where low signal to noise ratios are unavoidable. In addition, our work provides the first characterisation of the most abundant proteins within the DT40 proteome and the non-specific DT40 'beadomes' (non-specific proteins binding to beads) for common epitope tags. Given the importance and widespread use of the DT40 cell-line, these will be important resources for the cell biology and immunology communities. Biological significance SILAC-iPAC provides an improved method for the analysis of low-affinity

  8. Measurement of B- -> tau- nu_tau-bar Decay With a Semileptonic Tagging Method

    CERN Document Server

    Adachi, I; Anipko, D; Arinstein, K; Aso, T; Aulchenko, V; Aushev, T; Aziz, T; Bahinipati, S; Bakich, A M; Balagura, V; Ban, Y; Barberio, E; Bay, A; Bedny, I; Belous, K S; Bhardwaj, V; Bitenc, U; Blyth, S; Bondar, A; Bozek, A; Bracko, M; Brodzicka, J; Browder, T E; Chang, M C; Chang, P; Chang, Y W; Chao, Y; Chen, A; Chen, K F; Cheon, B G; Chiang, C C; Chistov, R; Cho, I S; Choi, S K; Choi, Y; Choi, Y K; Cole, S; Dalseno, J; Danilov, M; Das, A; Dash, M; Drutskoy, A; Dungel, W; Eidelman, S; Epifanov, D; Esen, S; Fratina, S; Fujii, H; Fujikawa, M; Gabyshev, N; Garmash, A; Goldenzweig, P; Golob, B; Grosse-Perdekamp, M; Guler, H; Guo, H; Ha, H; Haba, J; Hara, K; Hara, T; Hasegawa, Y; Hastings, N C; Hayasaka, K; Hayashii, H; Hazumi, M; Heffernan, D; Higuchi, T; Hodlmoser, H; Hokuue, T; Horii, Y; Hoshi, Y; Hoshina, K; Hou, W S; Hsiung, Y B; Hyun, H J; Igarashi, Y; Iijima, T; Ikado, K; Inami, K; Ishikawa, A; Ishino, H; Itoh, R; Iwabuchi, M; Iwasaki, M; Iwasaki, Y; Jacoby, C; Joshi, N J; Kaga, M; Kah, D H; Kaji, H; Kakuno, H; Kang, J H; Kapusta, P; Kataoka, S U; Katayama, N; Kawai, H; Kawasaki, T; Kibayashi, A; Kichimi, H; Kim, H J; Kim, H O; Kim, J H; Kim, S K; Kim, Y I; Kim, Y J; Kinoshita, K; Korpar, S; Kozakai, Y; Krizan, P; Krokovny, P; Kumar, R; Kurihara, E; Kuroki, Y; Kuzmin, A; Kwon, Y J; Kyeong, S H; Lange, J S; Leder, G; Lee, J; Lee, J S; Lee, M J; Lee, S E; Lesiak, T; Li, J; Limosani, A; Lin, S W; Liu, C; Liu, Y; Liventsev, D; MacNaughton, J; Mandl, F; Marlow, D; Matsumura, T; Matyja, A; McOnie, S; Medvedeva, T; Mikami, Y; Miyabayashi, K; Miyata, H; Miyazaki, Y; Mizuk, R; Moloney, G R; Mori, T; Nagamine, T; Nagasaka, Y; Nakahama, Y; Nakamura, I; Nakano, E; Nakao, M; Nakayama, H; Nakazawa, H; Natkaniec, Z; Neichi, K; Nishida, S; Nishimura, K; Nishio, Y; Nishizawa, I; Nitoh, O; Noguchi, S; Nozaki, T; Ogawa, A; Ogawa, S; Ohshima, T; Okuno, S; Olsen, S L; Ono, S; Ostrowicz, W; Ozaki, H; Pakhlov, P; Pakhlova, G; Palka, H; Park, C W; Park, H; Park, H K; Park, K S; Parslow, N; Peak, L S; Pernicka, M; Pestotnik, R; Peters, M; Piilonen, L E; Poluektov, A; Rorie, J; Rózanska, M; Sahoo, H; Sakai, Y; Sasao, N; Sayeed, K; Schietinger, T; Schneider, O; Schonmeier, P; Schümann, J; Schwanda, C; Schwartz, A J; Seidl, R; Sekiya, A; Senyo, K; Sevior, M E; Shang, L; Shapkin, M; Shebalin, V; Shen, C P; Shibuya, H; Shinomiya, S; Shiu, J G; Shwartz, B; Sidorov, V; Singh, J B; Sokolov, A; Somov, A; Stanic, S; Staric, M; Stypula, J; Sugiyama, A; Sumisawa, K; Sumiyoshi, T; Suzuki, S; Suzuki, S Y; Tajima, O; Takasaki, F; Tamai, K; Tamura, N; Tanaka, M; Taniguchi, N; Taylor, G N; Teramoto, Y; Tikhomirov, I; Trabelsi, K; Tse, Y F; Tsuboyama, T; Uchida, Y; Uehara, S; Ueki, Y; Ueno, K; Uglov, T; Unno, Y; Uno, S; Urquijo, P; Ushiroda, Y; Usov, Yu; Varner, G; Varvell, K E; Vervink, K; Villa, S; Vinokurova, A; Wang, C C; Wang, C H; Wang, J; Wang, M Z; Wang, P; Wang, X L; Watanabe, M; Watanabe, Y; Wedd, R; Wei, J T; Wicht, J; Widhalm, L; Wiechczynski, J; Won, E; Yabsley, B D; Yamaguchi, A; Yamamoto, H; Yamaoka, M; Yamashita, Y; Yamauchi, M; Yuan, C Z; Yusa, Y; Zhang, C C; Zhang, L M; Zhang, Z P; Zhilich, V; Zhulanov, V; Zivko, T; Zupanc, A; Zwahlen, N; Zyukova, O

    2008-01-01

    We present a new measurement of the decay B- -> tau- nu_tau-bar with a semileptonic B tagging method, using a data sample containing 657*10^6 BB-bar pairs collected at the (4S) resonance with the Belle detector at the KEKB asymmetric e+e- collider. A sample of BB pairs are tagged by reconstructing one B meson decaying semileptonically. We detect the B- -> tau- nu_tau-bar candidate in the recoil. We obtain a signal with a signicance of 3.8 standard deviations including systematics, and measure the branching fraction to be B(B- -> tau- nu_tau-bar)=(1.65+0.38-0.37(stat)+0.35-0.37(syst))*10^4. This result confirms the evidence for B- -> tau- nu_tau-bartained in the previous Belle measurement with a hadronic B tagging method.

  9. A touch probe method of operating an implantable RFID tag for orthopedic implant identification.

    Science.gov (United States)

    Liu, Xiaoyu; Berger, J Lee; Ogirala, Ajay; Mickle, Marlin H

    2013-06-01

    The major problem in operating an implantable radio-frequency identification (RFID) tag embedded on an orthopedic implant is low efficiency because of metallic interference. To improve the efficiency, this paper proposes a method of operating an implantable passive RFID tag using a touch probe at 13.56 MHz. This technology relies on the electric field interaction between two pairs of electrodes, one being a part of the touch probe placed on the surface of tissue and the other being a part of the tag installed under the tissue. Compared with using a conventional RFID antenna such as a loop antenna, this method has a better performance in the near field operation range to reduce interference with the orthopedic implant. Properly matching the touch probe and the tag to the tissue and the implant reduces signal attenuation and increases the overall system efficiency. The experiments have shown that this method has a great performance in the near field transcutaneous operation and can be used for orthopedic implant identification.

  10. Affinity-tagged phosphorylation assay by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (ATPA-MALDI): application to calcium/calmodulin-dependent protein kinase.

    Science.gov (United States)

    Kinumi, Tomoya; Niki, Etsuo; Shigeri, Yasushi; Matsumoto, Hiroyuki

    2005-12-01

    A matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based kinase assay using a peptide substrate tagged with a biotinyl group has been developed. The peptide moiety was designed to serve as an efficient substrate for calcium/calmodulin-dependent protein kinase II, based on the in vivo phosphorylation site of phosrestin I, a Drosophila homolog of arrestin. In the assay, the quantitative relationship was determined from the ratio of the peak areas between the two peaks respectively representing the unphosphorylated and the phosphorylated substrate. Attempts to assay phosphorylated peptides directly from the reaction mixture, gave inaccurate results because of the high noise level caused by the presence of salts and detergents. In contrast, after purifying the substrate peptides with the biotin affinity tag using streptavidin-coated magnetic beads, peak areas accurately represented the ratio between the unphosphorylated and phosphorylated peptide. By changing the substrate peptide to a peptide sequence that serves as a kinase substrate, it is expected that an efficient non-radioactive protein kinase assay using MALDI-TOF MS can be developed for any type of protein kinase. We call this technique "Affinity-Tagged Phosphorylation Assay by MALDI-TOF MS (ATPA-MALDI)." ATPA-MALDI should serve as a quick and efficient non-radioactive protein kinase assay by MALDI-TOF MS.

  11. A comparison of implantation methods for large PIT tags or injectable acoustic transmitters in juvenile Chinook salmon

    Energy Technology Data Exchange (ETDEWEB)

    Cook, Katrina V.; Brown, Richard S.; Deng, Zhiqun; Klett, Ryan S.; Li, Huidong; Seaburg, Adam; Eppard, M. B.

    2014-04-15

    The miniaturization of acoustic transmitters may allow greater flexibility in terms of the size and species of fish available to tag. New downsized injectable acoustic tags similar in shape to passive integrated transponder tags can be rapidly injected rather than surgically implanted through a sutured incision, as is current practice. Before wide-scale field use of these injectable transmitters, standard protocols to ensure the most effective and least damaging methods of implantation must be developed. Three implantation methods were tested in various sizes of juvenile Chinook salmon Oncorhynchus tschawytscha. Methods included a needle bevel-down injection, a needle bevel-up injection with a 90-degree rotation, and tag implantation through an unsutured incision. Tagged fish were compared to untagged control groups. Weight and wound area were measured at tagging and every week for 3 weeks; holding tanks were checked daily for mortalities and tag losses. No differences among treatments were found in growth, tag loss, or survival, but wound area was significantly reduced among incision-treated fish. The bevel-up injection had the worst results in terms of tag loss and wound area and also had high mortality. Implantation through an incision resulted in the lowest tag loss but the highest mortality. Fish from the bevel-down treatment group had the least mortality; wound areas also were smaller than the bevel-up treatment group. Cumulatively, the data suggest that the unsutured incision and bevel-down injection methods were the most effective; the drawbacks of both methods are described in detail. However, we further recommend larger and longer studies to find more robust thresholds for tagging size that include more sensitive measures.

  12. Affinity capillary electrophoresis method for investigation of bile salts complexation with sulfobutyl ether-ß-cyclodextrin

    DEFF Research Database (Denmark)

    Østergaard, Jesper; Jensen, Henrik; Holm, Rene

    2012-01-01

    an influence on the ionic strength of the background electrolyte when the cyclodextrin is used in capillary electrophoresis. Mobility-shift affinity capillary methods for investigation of the complexation of taurocholate and taurochenodeoxycholate with the negatively charged cyclodextrin derivative applying...... constant power and ionic strength conditions as well as constant voltage and varying ionic strength were investigated. A new approach for the correction of background electrolyte ionic strength was developed. Mobility-shift affinity capillary electrophoresis experiments obtained at constant voltage...

  13. Tag Correspondence Model for User Tag Suggestion

    Institute of Scientific and Technical Information of China (English)

    涂存超; 刘知远; 孙茂松

    2015-01-01

    Some microblog services encourage users to annotate themselves with multiple tags, indicating their attributes and interests. User tags play an important role for personalized recommendation and information retrieval. In order to better understand the semantics of user tags, we propose Tag Correspondence Model (TCM) to identify complex correspondences of tags from the rich context of microblog users. The correspondence of a tag is referred to as a unique element in the context which is semantically correlated with this tag. In TCM, we divide the context of a microblog user into various sources (such as short messages, user profile, and neighbors). With a collection of users with annotated tags, TCM can automatically learn the correspondences of user tags from multiple sources. With the learned correspondences, we are able to interpret implicit semantics of tags. Moreover, for the users who have not annotated any tags, TCM can suggest tags according to users’ context information. Extensive experiments on a real-world dataset demonstrate that our method can effciently identify correspondences of tags, which may eventually represent semantic meanings of tags.

  14. Top tagging: a method for identifying boosted hadronically decaying top quarks.

    Science.gov (United States)

    Kaplan, David E; Rehermann, Keith; Schwartz, Matthew D; Tweedie, Brock

    2008-10-01

    A method is introduced for distinguishing top jets (boosted, hadronically decaying top quarks) from light-quark and gluon jets using jet substructure. The procedure involves parsing the jet cluster to resolve its subjets and then imposing kinematic constraints. With this method, light-quark or gluon jets with p{T} approximately 1 TeV can be rejected with an efficiency of around 99% while retaining up to 40% of top jets. This reduces the dijet background to heavy tt[over ] resonances by a factor of approximately 10 000, thereby allowing resonance searches in tt[over ] to be extended into the all-hadronic channel. In addition, top tagging can be used in tt[over ] events when one of the top quarks decays semileptonically, in events with missing energy, and in studies of b-tagging efficiency at high p{T}.

  15. Novel method for digital subtraction of tagged stool in virtual colonoscopy

    Science.gov (United States)

    Guendel, Lutz; Suehling, Michael; Eckert, Helmut

    2008-03-01

    Colon cancer is one of the most frequent causes of death. CT colonography is a novel method for the detection of polyps and early cancer. The general principle of CT colonography includes a cathartic bowel preparation. The resulting discomfort for patients leads to limited patient acceptance and therefore to limited cancer detection rates. Reduced bowel preparation, techniques for stool tagging, and electronic cleansing, however, improve the acceptance rates. Hereby, the high density of oral contrast material highlights residual stool and can be digitally removed. Known subtraction methods cause artifacts: additional 3D objects are introduced and small bowel folds are perforated. We propose a new algorithm that is based on the 2 nd derivative of the image data using the Hessian matrix and the following principal axis transform to detect tiny folds which shall not be subtracted together with tagged stool found by a thresholding method. Since the stool is usually not homogenously tagged with contrast media a detection algorithm for island-like structures is incorporated. The interfaces of air-stool level and colon wall are detected by a 3-dimensional difference of Gaussian module. A 3-dimensional filter smoothes the transitions between removed stool and colon tissue. We evaluated the efficacy of the new algorithm with 10 patient data sets. The results showed no introduced artificial objects and no perforated folds. The artifacts at the air-stool and colon tissue-stool transitions are considerably reduced compared to those known from the literature.

  16. Computational methods for the analysis of tag sequences in metagenomics studies.

    Science.gov (United States)

    Chang, Qin; Luan, Yihui; Chen, Ting; Fuhrman, Jed A; Sun, Fengzhu

    2012-06-01

    Metagenomics commonly refers to the study of genetic materials directly derived from environments without culturing. Several ongoing large-scale metagenomics projects related to human and marine life, as well as pedology studies, have generated enormous amounts of data, posing a key challenge for efficient analysis, as we try to 1) understand microbial organism assemblage under different conditions, 2) compare different communities, and 3) understand how microbial organisms associate with each other and the environment.To address such questions, investigators are using new sequencing technologies, including Sanger, Illumina Solexa, and Roche 454, to sequence either particular genes, called tag sequences, mostly 16S or 18S ribosomal RNA sequences or other conserved genes, or whole metagenome shotgun sequences of all the genetic materials in a given community. In this paper, we review computational methods used for the analysis of tag sequences.

  17. A Statistical Method for Assessing Peptide Identification Confidence in Accurate Mass and Time Tag Proteomics

    Energy Technology Data Exchange (ETDEWEB)

    Stanley, Jeffrey R.; Adkins, Joshua N.; Slysz, Gordon W.; Monroe, Matthew E.; Purvine, Samuel O.; Karpievitch, Yuliya V.; Anderson, Gordon A.; Smith, Richard D.; Dabney, Alan R.

    2011-07-15

    High-throughput proteomics is rapidly evolving to require high mass measurement accuracy for a variety of different applications. Increased mass measurement accuracy in bottom-up proteomics specifically allows for an improved ability to distinguish and characterize detected MS features, which may in turn be identified by, e.g., matching to entries in a database for both precursor and fragmentation mass identification methods. Many tools exist with which to score the identification of peptides from LC-MS/MS measurements or to assess matches to an accurate mass and time (AMT) tag database, but these two calculations remain distinctly unrelated. Here we present a statistical method, Statistical Tools for AMT tag Confidence (STAC), which extends our previous work incorporating prior probabilities of correct sequence identification from LC-MS/MS, as well as the quality with which LC-MS features match AMT tags, to evaluate peptide identification confidence. Compared to existing tools, we are able to obtain significantly more high-confidence peptide identifications at a given false discovery rate and additionally assign confidence estimates to individual peptide identifications. Freely available software implementations of STAC are available in both command line and as a Windows graphical application.

  18. Deformation analysis of 3D tagged cardiac images using an optical flow method

    Directory of Open Access Journals (Sweden)

    Gorman Robert C

    2010-03-01

    Full Text Available Abstract Background This study proposes and validates a method of measuring 3D strain in myocardium using a 3D Cardiovascular Magnetic Resonance (CMR tissue-tagging sequence and a 3D optical flow method (OFM. Methods Initially, a 3D tag MR sequence was developed and the parameters of the sequence and 3D OFM were optimized using phantom images with simulated deformation. This method then was validated in-vivo and utilized to quantify normal sheep left ventricular functions. Results Optimizing imaging and OFM parameters in the phantom study produced sub-pixel root-mean square error (RMS between the estimated and known displacements in the x (RMSx = 0.62 pixels (0.43 mm, y (RMSy = 0.64 pixels (0.45 mm and z (RMSz = 0.68 pixels (1 mm direction, respectively. In-vivo validation demonstrated excellent correlation between the displacement measured by manually tracking tag intersections and that generated by 3D OFM (R ≥ 0.98. Technique performance was maintained even with 20% Gaussian noise added to the phantom images. Furthermore, 3D tracking of 3D cardiac motions resulted in a 51% decrease in in-plane tracking error as compared to 2D tracking. The in-vivo function studies showed that maximum wall thickening was greatest in the lateral wall, and increased from both apex and base towards the mid-ventricular region. Regional deformation patterns are in agreement with previous studies on LV function. Conclusion A novel method was developed to measure 3D LV wall deformation rapidly with high in-plane and through-plane resolution from one 3D cine acquisition.

  19. PINP: a new method of tagging neuronal populations for identification during in vivo electrophysiological recording.

    Directory of Open Access Journals (Sweden)

    Susana Q Lima

    Full Text Available Neural circuits are exquisitely organized, consisting of many different neuronal subpopulations. However, it is difficult to assess the functional roles of these subpopulations using conventional extracellular recording techniques because these techniques do not easily distinguish spikes from different neuronal populations. To overcome this limitation, we have developed PINP (Photostimulation-assisted Identification of Neuronal Populations, a method of tagging neuronal populations for identification during in vivo electrophysiological recording. The method is based on expressing the light-activated channel channelrhodopsin-2 (ChR2 to restricted neuronal subpopulations. ChR2-tagged neurons can be detected electrophysiologically in vivo since illumination of these neurons with a brief flash of blue light triggers a short latency reliable action potential. We demonstrate the feasibility of this technique by expressing ChR2 in distinct populations of cortical neurons using two different strategies. First, we labeled a subpopulation of cortical neurons-mainly fast-spiking interneurons-by using adeno-associated virus (AAV to deliver ChR2 in a transgenic mouse line in which the expression of Cre recombinase was driven by the parvalbumin promoter. Second, we labeled subpopulations of excitatory neurons in the rat auditory cortex with ChR2 based on projection target by using herpes simplex virus 1 (HSV1, which is efficiently taken up by axons and transported retrogradely; we find that this latter population responds to acoustic stimulation differently from unlabeled neurons. Tagging neurons is a novel application of ChR2, used in this case to monitor activity instead of manipulating it. PINP can be readily extended to other populations of genetically identifiable neurons, and will provide a useful method for probing the functional role of different neuronal populations in vivo.

  20. Tagging French comparing a statistical and a constraint-based method

    CERN Document Server

    Chanod, J P; Chanod, Jean-Pierre; Tapanainen, Pasi

    1995-01-01

    In this paper we compare two competing approaches to part-of-speech tagging, statistical and constraint-based disambiguation, using French as our test language. We imposed a time limit on our experiment: the amount of time spent on the design of our constraint system was about the same as the time we used to train and test the easy-to-implement statistical model. We describe the two systems and compare the results. The accuracy of the statistical method is reasonably good, comparable to taggers for English. But the constraint-based tagger seems to be superior even with the limited time we allowed ourselves for rule development.

  1. Predicting the relative binding affinity of mineralocorticoid receptor antagonists by density functional methods

    Science.gov (United States)

    Roos, Katarina; Hogner, Anders; Ogg, Derek; Packer, Martin J.; Hansson, Eva; Granberg, Kenneth L.; Evertsson, Emma; Nordqvist, Anneli

    2015-12-01

    In drug discovery, prediction of binding affinity ahead of synthesis to aid compound prioritization is still hampered by the low throughput of the more accurate methods and the lack of general pertinence of one method that fits all systems. Here we show the applicability of a method based on density functional theory using core fragments and a protein model with only the first shell residues surrounding the core, to predict relative binding affinity of a matched series of mineralocorticoid receptor (MR) antagonists. Antagonists of MR are used for treatment of chronic heart failure and hypertension. Marketed MR antagonists, spironolactone and eplerenone, are also believed to be highly efficacious in treatment of chronic kidney disease in diabetes patients, but is contra-indicated due to the increased risk for hyperkalemia. These findings and a significant unmet medical need among patients with chronic kidney disease continues to stimulate efforts in the discovery of new MR antagonist with maintained efficacy but low or no risk for hyperkalemia. Applied on a matched series of MR antagonists the quantum mechanical based method gave an R2 = 0.76 for the experimental lipophilic ligand efficiency versus relative predicted binding affinity calculated with the M06-2X functional in gas phase and an R2 = 0.64 for experimental binding affinity versus relative predicted binding affinity calculated with the M06-2X functional including an implicit solvation model. The quantum mechanical approach using core fragments was compared to free energy perturbation calculations using the full sized compound structures.

  2. Purification of His-Tagged Proteins.

    Science.gov (United States)

    Spriestersbach, Anne; Kubicek, Jan; Schäfer, Frank; Block, Helena; Maertens, Barbara

    2015-01-01

    Ni-NTA affinity purification of His-tagged proteins is a bind-wash-elute procedure that can be performed under native or denaturing conditions. Here, protocols for purification of His-tagged proteins under native, as well as under denaturing conditions, are given. The choice whether to purify the target protein under native or denaturing conditions depends on protein location and solubility, the accessibility of the His tag, and the desired downstream application. His-tagged proteins can be purified by a single-step affinity chromatography, namely immobilized metal ion affinity chromatography (IMAC), which is commercially available in different kinds of formats, Ni-NTA matrices being the most widely used. The provided protocols describe protein purification in the batch binding mode and apply gravity-assisted flow in disposable columns; this procedure is simple to conduct and extremely robust. IMAC purification can equally be performed in prepacked columns using FPLC or other liquid chromatography instrumentation, or using magnetic bead-based methods (Block et al., 2009).

  3. Protein purification-free method of binding affinity determination by microscale thermophoresis.

    Science.gov (United States)

    Khavrutskii, Lyuba; Yeh, Joanna; Timofeeva, Olga; Tarasov, Sergey G; Pritt, Samuel; Stefanisko, Karen; Tarasova, Nadya

    2013-08-15

    Quantitative characterization of protein interactions is essential in practically any field of life sciences, particularly drug discovery. Most of currently available methods of KD determination require access to purified protein of interest, generation of which can be time-consuming and expensive. We have developed a protocol that allows for determination of binding affinity by microscale thermophoresis (MST) without purification of the target protein from cell lysates. The method involves overexpression of the GFP-fused protein and cell lysis in non-denaturing conditions. Application of the method to STAT3-GFP transiently expressed in HEK293 cells allowed to determine for the first time the affinity of the well-studied transcription factor to oligonucleotides with different sequences. The protocol is straightforward and can have a variety of application for studying interactions of proteins with small molecules, peptides, DNA, RNA, and proteins.

  4. Superlinearly Convergent Affine Scaling Interior Trust-Region Method for Linear Constrained LC1 Minimization

    Institute of Scientific and Technical Information of China (English)

    De Tong ZHU

    2008-01-01

    We extend the classical affine scaling interior trust region algorithm for the linear con-strained smooth minimization problem to the nonsmooth case where the gradient of objective function is only locally Lipschitzian. We propose and analyze a new affine scaling trust-region method in associ-ation with nonmonotonic interior backtracking line search technique for solving the linear constrained LC1 optimization where the second-order derivative of the objective function is explicitly required to be locally Lipschitzian. The general trust region subproblem in the proposed algorithm is defined by minimizing an augmented affine scaling quadratic model which requires both first and second order information of the objective function subject only to an affine scaling ellipsoidal constraint in a null subspace of the augmented equality constraints. The global convergence and fast local convergence rate of the proposed algorithm are established under some reasonable conditions where twice smoothness of the objective function is not required. Applications of the algorithm to some nonsmooth optimization problems are discussed.

  5. Tagged Chromosomal Insertion Site System: A Method to Study Lamina-Associated Chromatin.

    Science.gov (United States)

    Harr, Jennifer C; Reddy, Karen L

    2016-01-01

    The three-dimensional (3D) organization of the genome is important for chromatin regulation. This organization is nonrandom and appears to be tightly correlated with or regulated by chromatin state and scaffolding proteins. To understand how specific DNA and chromatin elements contribute to the functional organization of the genome, we developed a new tool-the tagged chromosomal insertion site (TCIS) system-to identify and study minimal DNA sequences that drive nuclear compartmentalization and applied this system to specifically study the role of cis elements in targeting DNA to the nuclear lamina. The TCIS system allows Cre-recombinase-mediated site-directed integration of any DNA fragment into a locus tagged with lacO arrays, thus enabling both functional molecular studies and positional analysis of the altered locus. This system can be used to study the minimal DNA sequences that target the nuclear periphery (or other nuclear compartments), allowing researchers to understand how genome-wide results obtained, for example, by DNA adenine methyltransferase identification, chromosome conformation capture (HiC), or related methods, connect to the actual organization of DNA and chromosomes at the single-cell level. Finally, TCIS allows one to test roles for specific proteins in chromatin reorganization and to determine how changes in nuclear environment affect chromatin state and gene regulation at a single locus.

  6. Billfish Tagging

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The SWFSC's constituent-based Billfish Tagging Program began in 1963 and since that time has provided conventional spaghetti type tags and tagging supplies to...

  7. The use of external electronic tags on fish: an evaluation of tag retention and tagging effects

    DEFF Research Database (Denmark)

    Jepsen, Niels; Thorstad, Eva B.; Havn, Torgeir

    2015-01-01

    External tagging of fish with electronic tags has been used for decades for a wide range of marine and freshwater species. In the early years of fish telemetry research, it was the most commonly used attachment method, but later internal implants became preferred. Recently, the number of telemetry...... studies using external tagging has increased, especially with the development of archival tags (data storage tags, DSTs), pop-up satellite archival tags (PSATs) and other environment-sensing tags. Scientific evaluations of the tagging method are rather scarce for most species. We identified 89...... growth and survival have also been recorded, but direct mortality caused by external tagging seems rare. Most of the studies reviewed evaluate tag retention, survival, and tissue reactions. There is a general need for more research on the effects of external tagging of fish with electronic tags...

  8. Superlinear Convergence of Affine Scaling Interior Point Newton Method for Linear Inequality Constrained Minimization without Strict Complementarity

    Institute of Scientific and Technical Information of China (English)

    De-tong Zhu

    2009-01-01

    In this paper we extend and improve the classical affine scaling interior-point Newton method for solving nonlinear optimization subject to linear inequality constraints in the absence of the strict complementar-ity assumption. Introducing a computationally efficient technique and employing an identification function for the definition of the new affine scaling matrix, we propose and analyze a new affine scaling interior-point Newton method which improves the Coleman and Li affine scaling matrix in [2] for solving the linear inequality con-strained optimization. Local superlinear and quadratical convergence of the proposed algorithm is established under the strong second order sufficiency condition without assuming strict complementarity of the solution.

  9. A method for rapidly screening functionality of actin mutants and tagged actins

    Directory of Open Access Journals (Sweden)

    Rommelaere Heidi

    2004-01-01

    Full Text Available Recombinant production and biochemical analysis of actin mutants has been hampered by the fact that actin has an absolute requirement for the eukaryotic chaperone CCT to reach its native state. We therefore have developed a method to rapidly screen the folding capacity and functionality of actin variants, by combining in vitro expression of labelled actin with analysis on native gels, band shift assays or copolymerization tests. Additionally, we monitor, using immuno-fluorescence, incorporation of actin variants in cytoskeletal structures in transfected cells. We illustrate the method by two examples. In one we show that tagged versions of actin do not always behave native-like and in the other we study some of the molecular defects of three &bgr;-actin mutants that have been associated with diseases.

  10. Tag questions Tag questions

    Directory of Open Access Journals (Sweden)

    David Brazil

    2008-04-01

    Full Text Available The so-called 'tag' structures of English have received a lot of attention in language teaching programmes, attention that is not hard to justify when one considers the problems and anxiety they can occasion for many foreign learners. Most teachers one speaks to seem fairly willing to agree, however, that traditional treatments of the topic leave much to be desired. It happens, also, that, when considered collectively, the tags and some related phenomena have a special heoretical interest. For they constitute a field in which it seems essential to bring together insights that derive from the study of several aspects of linguistic organisation, aspects which in some recent work have been held to need distinctive kinds of descriptive category to handle. Traditional treatments have found it necessary to recognise different syntactic types (e.g. 'same polarity' and 'reversed polarity' tags and ifferent intonational treatments ("falling'and 'rising' tag; while the way the communicative significance of the various permutations is described normally requires reference to the expectations they signal regarding the immediately following behaviour of the other party (in the common phrase, 'What kind of answer they expect'. This last consideration places the matter squarely in the arena of recent work on the analysis of interactive discourse. The so-called 'tag' structures of English have received a lot of attention in language teaching programmes, attention that is not hard to justify when one considers the problems and anxiety they can occasion for many foreign learners. Most teachers one speaks to seem fairly willing to agree, however, that traditional treatments of the topic leave much to be desired. It happens, also, that, when considered collectively, the tags and some related phenomena have a special heoretical interest. For they constitute a field in which it seems essential to bring together insights that derive from the study of several aspects

  11. A Lyapunov method for stability analysis of piecewise-affine systems over non-invariant domains

    Science.gov (United States)

    Rubagotti, Matteo; Zaccarian, Luca; Bemporad, Alberto

    2016-05-01

    This paper analyses stability of discrete-time piecewise-affine systems, defined on possibly non-invariant domains, taking into account the possible presence of multiple dynamics in each of the polytopic regions of the system. An algorithm based on linear programming is proposed, in order to prove exponential stability of the origin and to find a positively invariant estimate of its region of attraction. The results are based on the definition of a piecewise-affine Lyapunov function, which is in general discontinuous on the boundaries of the regions. The proposed method is proven to lead to feasible solutions in a broader range of cases as compared to a previously proposed approach. Two numerical examples are shown, among which a case where the proposed method is applied to a closed-loop system, to which model predictive control was applied without a-priori guarantee of stability.

  12. Simplified, enhanced protein purification using an inducible, autoprocessing enzyme tag.

    Directory of Open Access Journals (Sweden)

    Aimee Shen

    Full Text Available We introduce a new method for purifying recombinant proteins expressed in bacteria using a highly specific, inducible, self-cleaving protease tag. This tag is comprised of the Vibrio cholerae MARTX toxin cysteine protease domain (CPD, an autoprocessing enzyme that cleaves exclusively after a leucine residue within the target protein-CPD junction. Importantly, V. cholerae CPD is specifically activated by inositol hexakisphosphate (InsP(6, a eukaryotic-specific small molecule that is absent from the bacterial cytosol. As a result, when His(6-tagged CPD is fused to the C-terminus of target proteins and expressed in Escherichia coli, the full-length fusion protein can be purified from bacterial lysates using metal ion affinity chromatography. Subsequent addition of InsP(6 to the immobilized fusion protein induces CPD-mediated cleavage at the target protein-CPD junction, releasing untagged target protein into the supernatant. This method condenses affinity chromatography and fusion tag cleavage into a single step, obviating the need for exogenous protease addition to remove the fusion tag(s and increasing the efficiency of tag separation. Furthermore, in addition to being timesaving, versatile, and inexpensive, our results indicate that the CPD purification system can enhance the expression, integrity, and solubility of intractable proteins from diverse organisms.

  13. Calculating protein-ligand binding affinities with MMPBSA: Method and error analysis.

    Science.gov (United States)

    Wang, Changhao; Nguyen, Peter H; Pham, Kevin; Huynh, Danielle; Le, Thanh-Binh Nancy; Wang, Hongli; Ren, Pengyu; Luo, Ray

    2016-10-15

    Molecular Mechanics Poisson-Boltzmann Surface Area (MMPBSA) methods have become widely adopted in estimating protein-ligand binding affinities due to their efficiency and high correlation with experiment. Here different computational alternatives were investigated to assess their impact to the agreement of MMPBSA calculations with experiment. Seven receptor families with both high-quality crystal structures and binding affinities were selected. First the performance of nonpolar solvation models was studied and it was found that the modern approach that separately models hydrophobic and dispersion interactions dramatically reduces RMSD's of computed relative binding affinities. The numerical setup of the Poisson-Boltzmann methods was analyzed next. The data shows that the impact of grid spacing to the quality of MMPBSA calculations is small: the numerical error at the grid spacing of 0.5 Å is already small enough to be negligible. The impact of different atomic radius sets and different molecular surface definitions was further analyzed and weak influences were found on the agreement with experiment. The influence of solute dielectric constant was also analyzed: a higher dielectric constant generally improves the overall agreement with experiment, especially for highly charged binding pockets. The data also showed that the converged simulations caused slight reduction in the agreement with experiment. Finally the direction of estimating absolute binding free energies was briefly explored. Upon correction of the binding-induced rearrangement free energy and the binding entropy lost, the errors in absolute binding affinities were also reduced dramatically when the modern nonpolar solvent model was used, although further developments were apparently necessary to further improve the MMPBSA methods. © 2016 Wiley Periodicals, Inc.

  14. Cyanine-based probe\\tag-peptide pair for fluorescence protein imaging and fluorescence protein imaging methods

    Science.gov (United States)

    Mayer-Cumblidge, M. Uljana; Cao, Haishi

    2010-08-17

    A molecular probe comprises two arsenic atoms and at least one cyanine based moiety. A method of producing a molecular probe includes providing a molecule having a first formula, treating the molecule with HgOAc, and subsequently transmetallizing with AsCl.sub.3. The As is liganded to ethanedithiol to produce a probe having a second formula. A method of labeling a peptide includes providing a peptide comprising a tag sequence and contacting the peptide with a biarsenical molecular probe. A complex is formed comprising the tag sequence and the molecular probe. A method of studying a peptide includes providing a mixture containing a peptide comprising a peptide tag sequence, adding a biarsenical probe to the mixture, and monitoring the fluorescence of the mixture.

  15. Elliptical modification of trilateration method based on linear model of distance vs power ratio for RFID tags spatial localization

    Directory of Open Access Journals (Sweden)

    Yu. B. Gimpilevich

    2013-12-01

    Full Text Available Introduction. Radio-frequency identification (RFID systems can be applied for a 2D spatial localization of objects in indoor spaces. For implementing the known localization method of trilateration one needs to build a model of ratio of distance between antenna and RFID tag versus power level of tag response signal. To maximize the accuracy of the model, one needs to collect measurement data from RFID tags placed at known positions. Due to the labor intensity of this process we aim to develop an alternative modification of elliptical trilateration method resulting in elimination of the preliminary data collecting stage. Theory part. We propose to use a linear model of distance vs power ratio for localization of passive RFID tags with small read ranges. Additionally, our model takes into account the possible ellipticity of position figures which happens because of antennas directivity. Also we propose some heuristics for solving the estimates ambiguity problem which arises when response signals from RFID tags are received by one or two antennas. Experimental part. We carried out the experimental analysis of the proposed trilateration variant using the previously developed RFID system in the 5 m ´ 5 m localization field. During the experiment, we compared our variant with a classical trilateration which was based on the polynomial model of distance vs power ratio formed by analyzing preliminarily gathered measurement data from RFID tags. The comparison indicated that our variant had a bigger by 1.6 cm mean localization error. Furthermore, taking into account ellipticity of position figures resulted in decrease of localization error for 18 of the 24 analyzed cases. Conclusions. It was determined that the proposed trilateration modification produced a slightly bigger mean localization error compared to the classical variant of trilateration. However, our approach allows one to eliminate the preliminary labor-intensive stage of collecting

  16. Solid support resins and affinity purification mass spectrometry.

    Science.gov (United States)

    Havis, Spencer; Moree, Wilna J; Mali, Sujina; Bark, Steven J

    2017-02-28

    Co-affinity purification-mass spectrometry (CoAP-MS) is a primary technology for elucidating the protein-protein interactions that form the basis of all biological processes. A critical component of CoAP-MS is the affinity purification (AP) of the bait protein, usually by immobilization of an antibody to a solid-phase resin. This Minireview discusses common resins, reagents, tagging methods, and their consideration for successful AP of tagged proteins. We discuss our experiences with different solid supports, their impact in AP experiments, and propose areas where chemistry can advance this important technology.

  17. Research progress of the affinity tag in the field of recombinant protein purification%亲和标签在重组蛋白纯化领域的研究进展

    Institute of Scientific and Technical Information of China (English)

    武文轩; 孙冬琳; 孙海明; 金焰

    2016-01-01

    亲和纯化标签在重组蛋白纯化领域中发挥着重要作用,经过标准化流程即可得到高产量高纯度的靶蛋白。近年来,许多不同种类的纯化标签陆续被设计出来,其中包括短肽标签、表位标签、折叠蛋白结构域标签、非层析法标签,以及最近研究较多的复合标签。除了通过经典的蛋白酶去除靶蛋白所携带标签的方法之外,自我裂解去除标签的方法因其简便廉价等优点近年来逐渐受到重视。本文主要对重组蛋白纯化标签的研究进展进行综述。%Engineered purification tags can very efficiently facilitate the purification of recombi -nant proteins , resulting in high yields and purities in a few standard steps .Over the years , many different purification tags have been developed, including short peptides , epitopes , folded protein domains , non-chromatographic tags and recently developed compound multifunctional tags with optimized capabilities .Al-though classic proteases are still used as a primary method to remove the tags from the target proteins , new self-cleaving methods are gaining more attention as a highly convenient alternative .In this review ,we dis-cuss some of these emerging trends , and examine their potential impacts and novel challenges on recombi -nant protein research .

  18. Development of Decision Making Algorithm for Control of Sea Cargo Containers by ``TAGGED'' Neutron Method

    Science.gov (United States)

    Anan'ev, A. A.; Belichenko, S. G.; Bogolyubov, E. P.; Bochkarev, O. V.; Petrov, E. V.; Polishchuk, A. M.; Udaltsov, A. Yu.

    2009-12-01

    Nowadays in Russia and abroad there are several groups of scientists, engaged in development of systems based on "tagged" neutron method (API method) and intended for detection of dangerous materials, including high explosives (HE). Particular attention is paid to possibility of detection of dangerous objects inside a sea cargo container. Energy gamma-spectrum, registered from object under inspection is used for determination of oxygen/carbon and nitrogen/carbon chemical ratios, according to which dangerous object is distinguished from not dangerous one. Material of filled container, however, gives rise to additional effects of rescattering and moderation of 14 MeV primary neutrons of generator, attenuation of secondary gamma-radiation from reactions of inelastic neutron scattering on objects under inspection. These effects lead to distortion of energy gamma-response from examined object and therefore prevent correct recognition of chemical ratios. These difficulties are taken into account in analytical method, presented in the paper. Method has been validated against experimental data, obtained by the system for HE detection in sea cargo, based on API method and developed in VNIIA. Influence of shielding materials on results of HE detection and identification is considered. Wood and iron were used as shielding materials. Results of method application for analysis of experimental data on HE simulator measurement (tetryl, trotyl, hexogen) are presented.

  19. Agonist high- and low-affinity states of dopamine D-2 receptors : methods of detection and clinical implications

    NARCIS (Netherlands)

    van Wieringen, Jan-Peter; Booij, Jan; Shalgunov, Vladimir; Elsinga, Philip; Michel, Martin C.

    2013-01-01

    Dopamine D-2 receptors, similar to other G-protein-coupled receptors, exist in a high- and low-affinity state for agonists. Based upon a review of the methods for detecting D-2 receptor agonist high-affinity states, we discuss alterations of such states in animal models of disease and the implicatio

  20. Extracting tag hierarchies.

    Directory of Open Access Journals (Sweden)

    Gergely Tibély

    Full Text Available Tagging items with descriptive annotations or keywords is a very natural way to compress and highlight information about the properties of the given entity. Over the years several methods have been proposed for extracting a hierarchy between the tags for systems with a "flat", egalitarian organization of the tags, which is very common when the tags correspond to free words given by numerous independent people. Here we present a complete framework for automated tag hierarchy extraction based on tag occurrence statistics. Along with proposing new algorithms, we are also introducing different quality measures enabling the detailed comparison of competing approaches from different aspects. Furthermore, we set up a synthetic, computer generated benchmark providing a versatile tool for testing, with a couple of tunable parameters capable of generating a wide range of test beds. Beside the computer generated input we also use real data in our studies, including a biological example with a pre-defined hierarchy between the tags. The encouraging similarity between the pre-defined and reconstructed hierarchy, as well as the seemingly meaningful hierarchies obtained for other real systems indicate that tag hierarchy extraction is a very promising direction for further research with a great potential for practical applications. Tags have become very prevalent nowadays in various online platforms ranging from blogs through scientific publications to protein databases. Furthermore, tagging systems dedicated for voluntary tagging of photos, films, books, etc. with free words are also becoming popular. The emerging large collections of tags associated with different objects are often referred to as folksonomies, highlighting their collaborative origin and the "flat" organization of the tags opposed to traditional hierarchical categorization. Adding a tag hierarchy corresponding to a given folksonomy can very effectively help narrowing or broadening the scope of

  1. Extracting tag hierarchies.

    Science.gov (United States)

    Tibély, Gergely; Pollner, Péter; Vicsek, Tamás; Palla, Gergely

    2013-01-01

    Tagging items with descriptive annotations or keywords is a very natural way to compress and highlight information about the properties of the given entity. Over the years several methods have been proposed for extracting a hierarchy between the tags for systems with a "flat", egalitarian organization of the tags, which is very common when the tags correspond to free words given by numerous independent people. Here we present a complete framework for automated tag hierarchy extraction based on tag occurrence statistics. Along with proposing new algorithms, we are also introducing different quality measures enabling the detailed comparison of competing approaches from different aspects. Furthermore, we set up a synthetic, computer generated benchmark providing a versatile tool for testing, with a couple of tunable parameters capable of generating a wide range of test beds. Beside the computer generated input we also use real data in our studies, including a biological example with a pre-defined hierarchy between the tags. The encouraging similarity between the pre-defined and reconstructed hierarchy, as well as the seemingly meaningful hierarchies obtained for other real systems indicate that tag hierarchy extraction is a very promising direction for further research with a great potential for practical applications. Tags have become very prevalent nowadays in various online platforms ranging from blogs through scientific publications to protein databases. Furthermore, tagging systems dedicated for voluntary tagging of photos, films, books, etc. with free words are also becoming popular. The emerging large collections of tags associated with different objects are often referred to as folksonomies, highlighting their collaborative origin and the "flat" organization of the tags opposed to traditional hierarchical categorization. Adding a tag hierarchy corresponding to a given folksonomy can very effectively help narrowing or broadening the scope of search. Moreover

  2. Advances and applications of binding affinity prediction methods in drug discovery.

    Science.gov (United States)

    Parenti, Marco Daniele; Rastelli, Giulio

    2012-01-01

    Nowadays, the improvement of R&D productivity is the primary commitment in pharmaceutical research, both in big pharma and smaller biotech companies. To reduce costs, to speed up the discovery process and to increase the chance of success, advanced methods of rational drug design are very helpful, as demonstrated by several successful applications. Among these, computational methods able to predict the binding affinity of small molecules to specific biological targets are of special interest because they can accelerate the discovery of new hit compounds. Here we provide an overview of the most widely used methods in the field of binding affinity prediction, as well as of our own work in developing BEAR, an innovative methodology specifically devised to overtake some limitations in existing approaches. The BEAR method was successfully validated against different biological targets, and proved its efficacy in retrieving active compounds from virtual screening campaigns. The results obtained so far indicate that BEAR may become a leading tool in the drug discovery pipeline. We primarily discuss advantages and drawbacks of each technique and show relevant examples and applications in drug discovery.

  3. Studies of top tagging identification methods and development of a new heavy object tagger

    Energy Technology Data Exchange (ETDEWEB)

    Lapsien, Tobias

    2016-05-15

    At the Large Hadron Collider (LHC), precision tests of the standard model of particle physics and searches for new phenomena are performed. To make optimal use of the proton-proton collisions delivered by the LHC and its increasing collision rate, both the detectors and the reconstruction algorithms have to be optimized. The identification of heavy quarks is a key component in many measurements. This thesis describes a hardware and a software project which both aim at improving the identification of heavy quarks. In the first part of this thesis, the Phase 1 upgrade of the CMS pixel detector is introduced. One of the main motivations of the replacement of the Pixel detector is the improved b jet identification at large collision rates. The Phase 1 upgrade involves several production and calibration steps. An X-ray calibration procedure has been developed and the corresponding experimental setup is described. Measurements show that the calibration of the pixel modules is temperature independent and can be performed at room temperature. The stability of the setup is tested in order to fulfill the requirements for mass production of the pixel modules. A method to stabilize the calibration is introduced which is shown to reduce the systematic uncertainty. In the second part, algorithms to identify heavily boosted top quarks (''top tagger'') are described and their performance is compared. The OptimalR HEP top tagger and the shower deconstruction tagger show a better performance than existing tagging algorithms. They can be used in Run II with increased centre-of-mass energies of 13 and 14 TeV. It is also shown that existing top tagging algorithms can be improved by the usage of multivariate analysis methods. New algorithms are commissioned using CMS data with a centre-of-mass energy of 8 TeV, corresponding to an integrated luminosity of 19.7 fb{sup -1}. In order to validate these new algorithms in data, two selections are made to measure the

  4. Affinity filtration coupled with capillary-based affinity purification for the isolation of protein complexes.

    Science.gov (United States)

    Qureshi, M S; Sheikh, Q I; Hill, R; Brown, P E; Dickman, M J; Tzokov, S B; Rice, D W; Gjerde, D T; Hornby, D P

    2013-08-01

    The isolation of complex macromolecular assemblies at the concentrations required for structural analysis represents a major experimental challenge. Here we present a method that combines the genetic power of site-specific recombination in order to selectively "tag" one or more components of a protein complex with affinity-based rapid filtration and a final step of capillary-based enrichment. This modified form of tandem affinity purification produces highly purified protein complexes at high concentrations in a highly efficient manner. The application of the method is demonstrated for the yeast Arp2/3 heptameric protein complex involved in mediating reorganization of the actin cytoskeleton.

  5. An Affine Scaling Interior Trust Region Method via Optimal Path for Solving Monotone Variational Inequality Problem with Linear Constraints

    Institute of Scientific and Technical Information of China (English)

    Yunjuan WANG; Detong ZHU

    2008-01-01

    Based on a differentiable merit function proposed by Taji et al.in "Math.Prog. Stud.,58,1993,369-383",the authors propose an affine scaling interior trust region strategy via optimal path to modify Newton method for the strictly monotone variational inequality problem subject to linear equality and inequality constraints.By using the eigensystem decomposition and affine scaling mapping,the authors form an affine scaling optimal curvilinear path very easily in order to approximately solve the trust region subproblem.Theoretical analysis is given which shows that the proposed algorithm is globally convergent and has a local quadratic convergence rate under some reasonable conditions.

  6. Detection of hidden shot balls in a gas-cooled turbine blade with an NRT gadolinium tagging method

    Science.gov (United States)

    Sim, Cheul Muu; Kim, Yi Kyung; Kim, TaeJoo; Lee, Kye Hong; Kim, Jeong Uk

    2009-06-01

    This report provides a preliminary insight into the benefits and effectiveness of neutron radiography in identifying alien materials, namely shot balls hidden in a turbine blade that are otherwise undetected using other methods. The detection of 0.2-mm-diameter shot balls in gas-cooled turbine blades is possible for thermal neutron radiography. A tagging processing is more useful for a distinctive image of newer turbine blades. Areas of concern for the tagging process include the solution concentration and the possibility of a slight washing of the blades. The location of the shot balls within the turbine blades tagged with Gd((2%, 5%)+water) was shown. Shot balls were placed externally on a turbine blade (F100-700, F100-200) surface in order to check for a dead zone from a surface examination. The image is produced from neutron radiography after a 5 min exposure time. When the blade is tagged with 2% and 5% Gd with slight washing, the shot can also be effectively seen on the SR-45 film. Shot balls are more obvious on a neutron image SR-45 film than an image plate or a DR film.

  7. Extracting tag hierarchies

    CERN Document Server

    Tibély, Gergely; Vicsek, Tamás; Palla, Gergely

    2014-01-01

    Tagging items with descriptive annotations or keywords is a very natural way to compress and highlight information about the properties of the given entity. Over the years several methods have been proposed for extracting a hierarchy between the tags for systems with a "flat", egalitarian organization of the tags, which is very common when the tags correspond to free words given by numerous independent people. Here we present a complete framework for automated tag hierarchy extraction based on tag occurrence statistics. Along with proposing new algorithms, we are also introducing different quality measures enabling the detailed comparison of competing approaches from different aspects. Furthermore, we set up a synthetic, computer generated benchmark providing a versatile tool for testing, with a couple of tunable parameters capable of generating a wide range of test beds. Beside the computer generated input we also use real data in our studies, including a biological example with a pre-defined hierarchy betwe...

  8. Methods for Fault Diagnosability Analysis of a Class of Affine Nonlinear Systems

    Directory of Open Access Journals (Sweden)

    Xiafu Peng

    2015-01-01

    Full Text Available The fault diagnosability analysis for a given model, before developing a diagnosis algorithm, can be used to answer questions like “can the fault fi be detected by observed states?” and “can it separate fault fi from fault fj by observed states?” If not, we should redesign the sensor placement. This paper deals with the problem of the evaluation of detectability and separability for the diagnosability analysis of affine nonlinear system. First, we used differential geometry theory to analyze the nonlinear system and proposed new detectability criterion and separability criterion. Second, the related matrix between the faults and outputs of the system and the fault separable matrix are designed for quantitative fault diagnosability calculation and fault separability calculation, respectively. Finally, we illustrate our approach to exemplify how to analyze diagnosability by a certain nonlinear system example, and the experiment results indicate the effectiveness of the fault evaluation methods.

  9. Exploring the interplay between experimental methods and the performance of predictors of binding affinity change upon mutations in protein complexes.

    Science.gov (United States)

    Geng, Cunliang; Vangone, Anna; Bonvin, Alexandre M J J

    2016-08-01

    Reliable prediction of binding affinity changes (ΔΔG) upon mutations in protein complexes relies not only on the performance of computational methods but also on the availability and quality of experimental data. Binding affinity changes can be measured by various experimental methods with different accuracies and limitations. To understand the impact of these on the prediction of binding affinity change, we present the Database of binding Affinity Change Upon Mutation (DACUM), a database of 1872 binding affinity changes upon single-point mutations, a subset of the SKEMPI database (Moal,I.H. and Fernández-Recio,J. Bioinformatics, 2012;28:2600-2607) extended with information on the experimental methods used for ΔΔG measurements. The ΔΔG data were classified into different data sets based on the experimental method used and the position of the mutation (interface and non-interface). We tested the prediction performance of the original HADDOCK score, a newly trained version of it and mutation Cutoff Scanning Matrix (Pires,D.E.V., Ascher,D.B. and Blundell,T.L. Bioinformatics 2014;30:335-342), one of the best reported ΔΔG predictors so far, on these various data sets. Our results demonstrate a strong impact of the experimental methods on the performance of binding affinity change predictors for protein complexes. This underscores the importance of properly considering and carefully choosing experimental methods in the development of novel binding affinity change predictors. The DACUM database is available online at https://github.com/haddocking/DACUM.

  10. 可伸缩TAGS%Scalable TAGS

    Institute of Scientific and Technical Information of China (English)

    闵帆; 张君雁; 杨国纬

    2003-01-01

    In a distributed Web server system where tasks are unpreemptible,the most important issue for improving quality of service (QoS) is how to realize fairness and reduce average slow down. In this paper we present an algorithm named Scalable TAGS by integrating Central Queue algorithm and Task Assignment by Guessing Size (TAGS), together with its performance analysis, system parameter setting algorithm subject to fairness requirement, and optimal grouping method.

  11. Highly specific purification of N-glycans using phosphate-based derivatization as an affinity tag in combination with Ti(4+)-SPE enrichment for mass spectrometric analysis.

    Science.gov (United States)

    Zhang, Ying; Peng, Ye; Bin, Zhichao; Wang, Huijie; Lu, Haojie

    2016-08-31

    N-linked protein glycosylation is involved in regulation of a wide variety of cellular processes and associated with numerous diseases. Highly specific identification of N-glycome remains a challenge while its biological significance is acknowledged. The relatively low abundance of glycan in complex biological mixtures, lack of basic sites for protonation, and suppression by other highly abundant proteins/peptides lead to the particularly poor detection sensitivity of N-glycans in the MS analysis. Therefore, the highly specific purification procedure becomes a crucial step prior to MS analysis of the N-glycome. Herein, a novel N-glycans enrichment approach based on phosphate derivatization combined with Ti(4+)-SPE (solid phase extraction) was developed. Briefly, in this strategy, N-glycans were chemically labeled with a phospho-group at their reducing ends, such that the Ti(4+)-SPE microspheres were able to capture the phospho-containing glycans. The enrichment method was developed and optimized using model oligosaccharides (maltoheptaose DP7 and sialylated glycan A1) and also glycans from a standard glycoprotein (asialofetuin, ASF). This method experimentally showed high derivatization efficiency (almost 100%), excellent selectivity (analyzing DP7 in the digests of bovine serum albumin at a mass ratio of 1:100), high enriching recovery (90%), good reproducibility (CVN-glycome in human serum, in which a total of 31 N-glycan masses were identified.

  12. Electron affinity of p-quinones. Improved method of electrochemical estimation

    Science.gov (United States)

    Jaworski, Jan S.

    1986-06-01

    Electron affinities of four p-quinones are estimated from enthalpy changes obtained on the basis of measured formal potentials and reaction entropies in the electroreduction process. A linear correlation between electron affinities of p-quinones and parent hydrocarbons is found.

  13. Using seed-tagging methods for assessing post-dispersal seed fate in rodent-dispersed trees

    NARCIS (Netherlands)

    Xiao, ZS; Jansen, PA; Zhang, ZB

    2006-01-01

    Seed tagging is widely used for tracking seeds during dispersal by seed-caching animals. No studies, however, have fully examined the effects of seed tagging on post-dispersal seed fate. We studied how two seed tagging techniques - thread-marking and wire tin-tagging - affected seed fate by placing

  14. Using seed-tagging methods for assessing post-dispersal seed fate in rodent-dispersed trees.

    NARCIS (Netherlands)

    Xiao, Z.; Jansen, P.A.; Zhang, Z.

    2006-01-01

    Seed tagging is widely used for tracking seeds during dispersal by seed-caching animals. No studies, however, have fully examined the effects of seed tagging on post-dispersal seed fate. We studied how two seed tagging techniques – thread-marking and wire tin-tagging – affected seed fate by placing

  15. Spectral Tagging

    Energy Technology Data Exchange (ETDEWEB)

    Smartt, Heidi A. [Sandia National Laboratories (United States)

    2003-05-01

    This research examines the feasibility of spectral tagging, which involves modifying the spectral signature of a target, e.g. by mixing an additive with the target's paint. The target is unchanged to the human eye, but the tag is revealed when viewed with a spectrometer. This project investigates a layer of security that is not obvious, and therefore easy to conceal. The result is a tagging mechanism that is difficult to counterfeit. Uniquely tagging an item is an area of need in safeguards and security and non-proliferation. The powdered forms of the minerals lapis lazuli and olivine were selected as the initial test tags due to their availability and uniqueness in the visible to near-infrared spectral region. They were mixed with paints and applied to steel. In order to verify the presence of the tags quantitatively, the data from the spectrometer was input into unmixing models and signal detection algorithms. The mixture with the best results was blue paint mixed with lapis lazuli and olivine. The tag had a 0% probability of false alarm and a 100% probability of detection. The research proved that spectral tagging is feasible, although certain tag/paint mixtures are more detectable than others.

  16. An extended affinity propagation clustering method based on different data density types.

    Science.gov (United States)

    Zhao, XiuLi; Xu, WeiXiang

    2015-01-01

    Affinity propagation (AP) algorithm, as a novel clustering method, does not require the users to specify the initial cluster centers in advance, which regards all data points as potential exemplars (cluster centers) equally and groups the clusters totally by the similar degree among the data points. But in many cases there exist some different intensive areas within the same data set, which means that the data set does not distribute homogeneously. In such situation the AP algorithm cannot group the data points into ideal clusters. In this paper, we proposed an extended AP clustering algorithm to deal with such a problem. There are two steps in our method: firstly the data set is partitioned into several data density types according to the nearest distances of each data point; and then the AP clustering method is, respectively, used to group the data points into clusters in each data density type. Two experiments are carried out to evaluate the performance of our algorithm: one utilizes an artificial data set and the other uses a real seismic data set. The experiment results show that groups are obtained more accurately by our algorithm than OPTICS and AP clustering algorithm itself.

  17. Nonradioactive sequence-tagged microsatellite site analyses: a method transferable to the tropics.

    Science.gov (United States)

    Lagoda, P J; Dambier, D; Grapin, A; Baurens, F C; Lanaud, C; Noyer, J L

    1998-02-01

    Utilization of existing isozyme analysis facilities to detect sequence-tagged microsatellite site (STMS) polymorphism or any simple sequence repeat (SSR) variation is described. Different parameters concerning the difficulties in transferring molecular techniques to less sophisticated laboratory infrastructures (i.e. tropical outstations) are discussed (e.g. reproducibility, efficacy, precision). Nonradioactive STMS analysis is bound to foster collaborative research between "biodiversity" and "biotechnology" centers.

  18. The MM/PBSA and MM/GBSA methods to estimate ligand-binding affinities

    Science.gov (United States)

    Genheden, Samuel; Ryde, Ulf

    2015-01-01

    Introduction: The molecular mechanics energies combined with the Poisson–Boltzmann or generalized Born and surface area continuum solvation (MM/PBSA and MM/GBSA) methods are popular approaches to estimate the free energy of the binding of small ligands to biological macromolecules. They are typically based on molecular dynamics simulations of the receptor–ligand complex and are therefore intermediate in both accuracy and computational effort between empirical scoring and strict alchemical perturbation methods. They have been applied to a large number of systems with varying success. Areas covered: The authors review the use of MM/PBSA and MM/GBSA methods to calculate ligand-binding affinities, with an emphasis on calibration, testing and validation, as well as attempts to improve the methods, rather than on specific applications. Expert opinion: MM/PBSA and MM/GBSA are attractive approaches owing to their modular nature and that they do not require calculations on a training set. They have been used successfully to reproduce and rationalize experimental findings and to improve the results of virtual screening and docking. However, they contain several crude and questionable approximations, for example, the lack of conformational entropy and information about the number and free energy of water molecules in the binding site. Moreover, there are many variants of the method and their performance varies strongly with the tested system. Likewise, most attempts to ameliorate the methods with more accurate approaches, for example, quantum-mechanical calculations, polarizable force fields or improved solvation have deteriorated the results. PMID:25835573

  19. Development of an aptamer-based affinity purification method for vascular endothelial growth factor

    Directory of Open Access Journals (Sweden)

    Maren Lönne

    2015-12-01

    Full Text Available Since aptamers bind their targets with high affinity and specificity, they are promising alternative ligands in protein affinity purification. As aptamers are chemically synthesized oligonucleotides, they can be easily produced in large quantities regarding GMP conditions allowing their application in protein production for therapeutic purposes. Several advantages of aptamers compared to antibodies are described in general within this paper. Here, an aptamer directed against the human Vascular Endothelial Growth Factor (VEGF was used as affinity ligand for establishing a purification platform for VEGF in small scale. The aptamer was covalently immobilized on magnetic beads in a controlled orientation resulting in a functional active affinity matrix. Target binding was optimized by introduction of spacer molecules and variation of aptamer density. Further, salt-induced target elution was demonstrated as well as VEGF purification from a complex protein mixture proving the specificity of protein-aptamer binding.

  20. Pop-up archival transmitting (PAT) tags: A method to investigate the migration and behavior of Pacific halibut (Hippoglossus stenolepis) in the Gulf of Alaska

    Science.gov (United States)

    Seitz, Andrew C; Wilson, Derek; Norcross, Brenda L.; Nielsen, Jennifer L.

    2003-01-01

    Pop-up archival transmitting (PAT) tags provide a fisheries-independent method of collecting environmental preference data (depth and ambient water temperature) and migration distance. In this study, we evaluate the use of pop-up archival transmitting tags as a method to investigate demersal fish. We report the results from eight pop-up archival transmitting tagged Pacific halibut Hippoglossus stenolepis (from 107 to 165 cm FL) that were released in and around Resurrection Bay, Alaska. Commercial fishermen recovered three tags, while five tags transmitted data to Argos satellites. Horizontal migration was not consistent among fish as four Pacific halibut remained in the vicinity of release while the other four traveled up to 358 km from the release site. Vertical movement was not consistent among fish or over time; however, they spent most of their time at depths of 150 to 350 m. The minimum and maximum depths reached by any of the Pacific halibut were 2 m and 502 m, respectively. The fish preferred water temperatures of approximately 6°C, but experienced temperatures between 4.3 and 12.2°C. Light attenuation with depth prevented geolocation software and light sensing hardware from accurately estimating geoposition for the majority of days. The methods, adapted from investigations on large pelagic fish, proved to be effective for studying Pacific halibut in the northern Gulf of Alaska. PAT tags allowed us to obtain high accuracy locations of the fish at the end of the tag deployments as well as preliminary data to identify approximate seasonal locations and to characterize their depth and temperature characteristics. By using PAT tags, we will be able to ensure tag returns during the winter season (which is closed to fishing) and gain valuable biological information even if fish migrate large distances or to unexpected locations.

  1. Methods for optimizing over the efficient and weakly efficient sets of an affine fractional vector optimization program

    DEFF Research Database (Denmark)

    Le, T.H.A.; Pham, D. T.; Canh, Nam Nguyen;

    2010-01-01

    Both the efficient and weakly efficient sets of an affine fractional vector optimization problem, in general, are neither convex nor given explicitly. Optimization problems over one of these sets are thus nonconvex. We propose two methods for optimizing a real-valued function over the efficient a...

  2. Flavour Tagging at LHCb

    CERN Multimedia

    Grabalosa Gandara, M

    2009-01-01

    To do precise CP violation measurements, the most possible accurate knowledge of the flavour at production of the reconstructed B meson is required. This poster summarizes the flavour tagging performances for the LHCb experiment. We use same side an opposite side algorithms to establish wheter the meson contained a b or a b\\bar quark. The final decision is obtained through a combination of several methods. The use of control channels, decays to a flavour specific final state, will allow to determine the wrong tag fraction \\omega (the probability of a tag to be wrong), which can be used as input for the determination of CKM unitary triangle angles.

  3. Proton affinity determinations using the kinetic method in an ion trap mass spectrometer

    Science.gov (United States)

    Nourse, Bobette D.; Graham Cooks, R.

    1991-05-01

    Proton affinities for various compounds have been estimated using a quadrupole ion trap by generating and mass-selecting proton-bound dimers and measuring their dissociation kinetics (A-H+ -B --> AH+ + B and/or BH+ + A). From the relative abundances of the fragment ions ([BH+] and [AH+]), which are related to their relative proton affinities by ln ([AH+]/[BH+]) = [Delta]PA/RT, it is shown that the proton affinities of the alicyclic carboxylic acids decrease in the order: cyclohexane- > cyclopropane- > cyclopentane- > cyclobutanecarboxylic acid. Proton affinity values for these species, measured from their proton-bound dimers with specific ketones, esters and carboxylic acids of known PA, are determined to be 198.3 ± 0.2 kcal mol-1, 198.0 ± 0.2 kcal mol-1, 197.8 ± 0.2 kcal mol-1 and 197.0 ± 0.2 kcal mol-1, respectively. The major contribution to the estimated uncertainties in these values results from the uncertainties in literature proton affinity values for the reference compounds. Proton affinity differences of meta and para deuterated benzoic acid proton-bound to benzoic acid (kH/kD = 1.0 ± 0.1 and 0.9 ± 0.1, respectively), for acetophenone proton-bound to deuterated-acetophenone (C6H5C(O)CD3) (kH/kD = 0.7 ± 0.1) and for 2-pentanone proton-bound to deuterated 2-pentanone (CH3CH2CH2C(O)CH3) (kH/kD = 2.1 ± 0.2). These results, as well as those for the carboxylic acids and benzoic acids given above, are accounted for in terms of stabilizing electronic effects in the protonated molecules.

  4. Method to produce acetyldiacylglycerols (ac-TAGs) by expression of an acetyltransferase gene isolated from Euonymus alatus (burning bush)

    Energy Technology Data Exchange (ETDEWEB)

    Durrett, Timothy; Ohlrogge, John; Pollard, Michael

    2016-05-03

    The present invention relates to novel diacylglycerol acyltransferase genes and proteins, and methods of their use. In particular, the invention describes genes encoding proteins having diacylglycerol acetyltransferase activity, specifically for transferring an acetyl group to a diacylglycerol substrate to form acetyl-Triacylglycerols (ac-TAGS), for example, a 3-acetyl-1,2-diacyl-sn-glycerol. The present invention encompasses both native and recombinant wild-type forms of the transferase, as well as mutants and variant forms. The present invention also relates to methods of using novel diacylglycerol acyltransferase genes and proteins, including their expression in transgenic organisms at commercially viable levels, for increasing production of 3-acetyl-1,2-diacyl-sn-glycerols in plant oils and altering the composition of oils produced by microorganisms, such as yeast, by increasing ac-TAG production. Additionally, oils produced by methods of the present inventions comprising genes and proteins are contemplated for use as biodiesel fuel, in polymer production and as naturally produced food oils with reduced calories.

  5. A new method for improving the cell affinity of poly (D,L-lactide)

    Institute of Scientific and Technical Information of China (English)

    YANG Jian; BEI Jianzhong; WANG Shenguo

    2001-01-01

    @@ Cell affinity is an important factor to be concerned when biodegradable polymeric materials are utilized as cell scaffold in tissue engineering . Many studies haveproved that hydrophilicity , surface energy, charge and roughness of the materialsurface greatly influence the cell attachment and cell growth on the material.

  6. b Tagging in CMS

    CERN Document Server

    Tomalin, Ian R

    2008-01-01

    Many of the exotic particles expected at the LHC, such as SUSY, Higgs bosons and top quarks, will decay to b quarks. This paper presents the methods used to identify b-jets at CMS. The algorithms exploit the long B hadron lifetime, semi-leptonic B decays and jet kinematics. The prospect for measuring the performance of these b-tags directly from CMS data is examined. Finally, the use of b-tagging in the High-Level Trigger is explained.

  7. His6 tag-assisted chemical protein synthesis

    Science.gov (United States)

    Bang, Duhee; Kent, Stephen B. H.

    2005-04-01

    To make more practical the total chemical synthesis of proteins by the ligation of unprotected peptide building blocks, we have developed a method to facilitate the isolation and handling of intermediate products. The synthetic technique makes use of a His6 tag at the C terminus of the target polypeptide chain, introduced during the synthesis of the C-terminal peptide segment building block. The presence of a His6 tag enables the isolation of peptide or protein products directly from ligation reaction mixtures by Ni-NTA affinity column purification. This simple approach enables facile buffer exchange to alternate reaction conditions and is compatible with direct analytical control by protein MS of the multiple ligation steps involved in protein synthesis. We used syntheses of crambin and a modular tetratricopeptide repeat protein of 17 kDa as models to examine the utility of this affinity purification approach. The results show that His6 tag-assisted chemical protein synthesis is a useful method that substantially reduces handling losses and provides for rapid chemical protein syntheses. affinity purification | native chemical ligation

  8. The Method of Selecting tagSNPs Based on Linkage Disequilibrium%一种基于连锁不平衡的tagSNPs选择算法

    Institute of Scientific and Technical Information of China (English)

    王钧峰; 王新赠

    2016-01-01

    进行全基因组关联研究(genome-wide association studies,简记为GWAS)时,我们需要获得一个足够密集的单核苷酸多态性(single-nucleotide polymorphism,简记为SNP)标记集来解释常见疾病遗传风险的一部分.候选基因中SNP的数量是有限的,但是直接分析所有现存的SNPs是无效的,因为在这些位点上的基因型有很强的关联性,会导致大量的冗余信息,并且会造成基因分型成本的增加,消耗大量的时间.所以我们在进行关联检验时,没有必要对所有的SNPs进行基因分型,只需要选择出具有代表性,并且数量很少的SNPs进行分型,并对这些SNPs进行关联检验.这里选择出的SNPs称为标签SNPs(记为tagSNPs),它为SNPs的一个小的子集,在每个单体型区域中足以捕获单体型的信息.选择tagSNPs的方法有很多,本文我们提出了一种新的tagSNPs选择方法,通过使用基于连锁不平衡(linkage disequilibrium,简记为LD)的两两r2准则对单体型分组,分成不相交的组,并在每个组中选择标签SNPs.与基于原始SNPs集的检验方法比较,我们的方法产生了更少的tagSNPs,在最大化所选标记提供信息含量的同时,降低了基因分型成本,提高了效率.

  9. Geometrical and P.D.E. Methods in the Treatment of the Theory of Shells: Comparing Euclidean and Affine Approaches

    Directory of Open Access Journals (Sweden)

    Salvador Gigena

    2014-01-01

    Full Text Available The use of differential equations methods in the approach, treatment, and solution of problems in diverse areas of geometry, particularly in affine differential geometry is well known and prolific, where they have proven to be quite fruitful when it comes to the obtainment of definite results. It is perhaps lesser known that the same kind of those very same methods has been and is currently being used to treat developments in some specific areas of applied sciences, such as the theory of shells where, similarly, they can be proven to be quite effective as well. In this paper we precisely show that such is the case in two particular, related instances: the historic approach of the classical, Euclidean part of the theory pursued by Fritz John, in the past century, and the more recent expositions that we ourselves have dedicated to the affine counterpart of the theory.

  10. EXPRESSION OF SV40 Tag AND FORMATION Tag-p53 AND Tag-Rb COMPLEXES IN CHINESE BRAIN TUMORS

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective: To investigate the expression of SV40 Tag andformation of Tag-p53 and Tag-Rb complexes in Chinese brain tumors. Methods: SV40 large tumor antigen (Tag) were investigated by immunoprecipitation, silver staining and Western blot in 65 cases of Chinese brain tumors and 8 cases of normal brain tissues. Tag-p53 and Tag-Rb complexes were screened by the same way in 20 and 15 Tag positive tumor tissues respectively. Results: Tag was found in all of 8 ependymomas and 2 choroid plexus papillomas, 90% (9/10) of pituitary adenomas, 73% (11/15) of astrocytomas, 70% (7/10) of meningiomas, 50% (4/8) of glioblastoma multiform, 33% (2/6) of medulloblastomas, 5 oligodendrogliomas, 1 pineocytoma and 8 normal brain tissues were negative for Tag. Tag-p53 complex was detected in all of 20 Tag positive tumors as well as Tag-Rb complex in all of 15 Tag positive tumors. Conclusion: SV40 Tag is not only expressed in human brain tumors, but also it can form specific complexes with tumor suppressors p53 and Rb. SV40 is correlated to human brain tumorigenesis. The inactivation of p53 and Rb due to the formation of Tag-p53 and Tag-Rb complexes is possibly an important mechanism in the etiopathogenesis of human brain tumors.

  11. NONMONOTONIC REDUCED PROJECTED HESSIAN METHOD VIA AN AFFINE SCALING INTERIOR MODIFIED GRADIENT PATH FOR BOUNDED-CONSTRAINED OPTIMIZATION

    Institute of Scientific and Technical Information of China (English)

    Peihua GUO; Detong ZHU

    2008-01-01

    The authors propose an affine scaling modified gradient path method in association with reduced projective Hessian and nonmonotonic interior backtracking line search techniques for solving the linear equality constrained optimization subject to bounds on variables. By employing the QR decomposition of the constraint matrix and the eigensystem decomposition of reduced projective Hes-sian matrix in the subproblem, the authors form affine scaling modified gradient curvilinear path very easily. By using interior backtracking line search technique, each iterate switches to trial step of strict interior feasibility. The global convergence and fast local superlinear/quadratical convergence rates of the proposed algorithm are established under some reasonable conditions. A nonmonotonic criterion should bring about speeding up the convergence progress in some ill-conditioned cases. The results of numerical experiments are reported to show the effectiveness of the proposed algorithm.

  12. The expression sequence tag is an effective method for screening DNA segments that predict urinary bladder transitional cell carcinoma prognosis

    Directory of Open Access Journals (Sweden)

    Yang PS

    2014-09-01

    Full Text Available Pei-Shan Yang,1,* Yu-Chao Hsu,1,2,* Yu-Hsiang Lin,1,2 Cheng-Pang Hou,1,2 Chien-Lun Chen,1,2 Phei-Lang Chang,1–3 Horng-Heng Juang,3,4 Ke-Hung Tsui1–3 1Department of Urology, Chang Gung Memorial Hospital at Linkou, Taiwan, Republic of China; 2School of Medicine, Chang Gung University, Taiwan, Republic of China; 3Bioinformation Center, Chang Gung Memorial Hospital, Taiwan, Republic of China; 4Department of Anatomy, School of Medicine, Chang Gung University, Taiwan, Republic of China; Kwei-Shan, Tao-Yuan, Taiwan, Republic of China *These authors contributed equally to this workPurpose: We validated the use of expression sequence tags (ESTs as an effective method of screening for DNA segments that could predict urothelial cell carcinoma and for identifying ESTs with such predictive value.Patients and methods: From 2004 to 2009, eleven patients were enrolled in this study: six with high-grade bladder carcinoma and five with low-grade bladder carcinoma. ESTs were used to screen for differential gene expression in a high-grade cell line (MGH-U1 and in a premalignant cell line (MGH-U4. Immunohistochemistry and real-time reverse transcription-polymerase chain reaction were used to validate the degree of EST expression and the prognostic value of ESTs.Results: Apoferritin H subunit (FTH1 protein exhibited increased expression in high-grade bladder carcinoma compared with that seen in low-grade carcinoma. Immunohistochemistry and real-time reverse transcription-polymerase chain reaction both supported the higher expression of FTH1 in high-grade urothelial carcinoma.Conclusion: ESTs are useful for detecting the FTH1 protein, which predicts the prognosis of patients with bladder carcinoma.Keywords: expression sequence tag, transitional cell carcinoma, FTH1

  13. Optimized E. coli expression strain LOBSTR eliminates common contaminants from His-tag purification.

    Science.gov (United States)

    Andersen, Kasper R; Leksa, Nina C; Schwartz, Thomas U

    2013-11-01

    His-tag affinity purification is one of the most commonly used methods to purify recombinant proteins expressed in E. coli. One drawback of using the His-tag is the co-purification of contaminating histidine-rich E. coli proteins. We engineered a new E. coli expression strain, LOBSTR (low background strain), which eliminates the most abundant contaminants. LOBSTR is derived from the E. coli BL21(DE3) strain and carries genomically modified copies of arnA and slyD, whose protein products exhibit reduced affinities to Ni and Co resins, resulting in a much higher purity of the target protein. The use of LOBSTR enables the pursuit of challenging low-expressing protein targets by reducing background contamination with no additional purification steps, materials, or costs, and thus pushes the limits of standard His-tag purifications.

  14. Inntags: small self-structured epitopes for innocuous protein tagging.

    Science.gov (United States)

    Georgieva, Maya V; Yahya, Galal; Codó, Laia; Ortiz, Raúl; Teixidó, Laura; Claros, José; Jara, Ricardo; Jara, Mònica; Iborra, Antoni; Gelpí, Josep Lluís; Gallego, Carme; Orozco, Modesto; Aldea, Martí

    2015-10-01

    Protein tagging is widely used in approaches ranging from affinity purification to fluorescence-based detection in live cells. However, an intrinsic limitation of tagging is that the native function of the protein may be compromised or even abolished by the presence of the tag. Here we describe and characterize a set of small, innocuous protein tags (inntags) that we anticipate will find application in a variety of biological techniques.

  15. Social Tagging of Mission Data

    Science.gov (United States)

    Norris, Jeffrey S.; Wallick, Michael N.; Joswig, Joseph C.; Powell, Mark W.; Torres, Recaredo J.; Mittman, David S.; Abramyan, Lucy; Crockett, Thomas M.; Shams, Khawaja S.; Fox, Jason M.; Pyrzak, Guy; Vaughn, Michael B.

    2010-01-01

    Mars missions will generate a large amount of data in various forms, such as daily plans, images, and scientific information. Often, there is a semantic linkage between images that cannot be captured automatically. Software is needed that will provide a method for creating arbitrary tags for this mission data so that items with a similar tag can be related to each other. The tags should be visible and searchable for all users. A new routine was written to offer a new and more flexible search option over previous applications. This software allows users of the MSLICE program to apply any number of arbitrary tags to a piece of mission data through a MSLICE search interface. The application of tags creates relationships between data that did not previously exist. These tags can be easily removed and changed, and contain enough flexibility to be specifically configured for any mission. This gives users the ability to quickly recall or draw attention to particular pieces of mission data, for example: Give a semantic and meaningful description to mission data; for example, tag all images with a rock in them with the tag "rock." Rapidly recall specific and useful pieces of data; for example, tag a plan as"driving template." Call specific data to a user s attention; for example, tag a plan as "for:User." This software is part of the MSLICE release, which was written in Java. It will run on any current Windows, Macintosh, or Linux system.

  16. Prediction of MHC class II binding affinity using SMM-align, a novel stabilization matrix alignment method

    DEFF Research Database (Denmark)

    Nielsen, Morten; Lundegaard, Claus; Lund, Ole

    2007-01-01

    the correct alignment of a peptide in the binding groove a crucial part of identifying the core of an MHC class II binding motif. Here, we present a novel stabilization matrix alignment method, SMM-align, that allows for direct prediction of peptide:MHC binding affinities. The predictive performance...... of the method is validated on a large MHC class II benchmark data set covering 14 HLA-DR (human MHC) and three mouse H2-IA alleles. RESULTS: The predictive performance of the SMM-align method was demonstrated to be superior to that of the Gibbs sampler, TEPITOPE, SVRMHC, and MHCpred methods. Cross validation...... by favoring binding registers with a minimum PFR length of two amino acids. Visualizing the binding motif as obtained by the SMM-align and TEPITOPE methods highlights a series of fundamental discrepancies between the two predicted motifs. For the DRB1*1302 allele for instance, the TEPITOPE method favors basic...

  17. A Simple Three-Step Method for Design and Affinity Testing of New Antisense Peptides: An Example of Erythropoietin

    Directory of Open Access Journals (Sweden)

    Nikola Štambuk

    2014-05-01

    Full Text Available Antisense peptide technology is a valuable tool for deriving new biologically active molecules and performing peptide–receptor modulation. It is based on the fact that peptides specified by the complementary (antisense nucleotide sequences often bind to each other with a higher specificity and efficacy. We tested the validity of this concept on the example of human erythropoietin, a well-characterized and pharmacologically relevant hematopoietic growth factor. The purpose of the work was to present and test simple and efficient three-step procedure for the design of an antisense peptide targeting receptor-binding site of human erythropoietin. Firstly, we selected the carboxyl-terminal receptor binding region of the molecule (epitope as a template for the antisense peptide modeling; Secondly, we designed an antisense peptide using mRNA transcription of the epitope sequence in the 3'→5' direction and computational screening of potential paratope structures with BLAST; Thirdly, we evaluated sense–antisense (epitope–paratope peptide binding and affinity by means of fluorescence spectroscopy and microscale thermophoresis. Both methods showed similar Kd values of 850 and 816 µM, respectively. The advantages of the methods were: fast screening with a small quantity of the sample needed, and measurements done within the range of physicochemical parameters resembling physiological conditions. Antisense peptides targeting specific erythropoietin region(s could be used for the development of new immunochemical methods. Selected antisense peptides with optimal affinity are potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines.

  18. Chemical ligation methods for the tagging of DNA-encoded chemical libraries.

    Science.gov (United States)

    Keefe, Anthony D; Clark, Matthew A; Hupp, Christopher D; Litovchick, Alexander; Zhang, Ying

    2015-06-01

    The generation of DNA-encoded chemical libraries requires the unimolecular association of multiple encoding oligonucleotides with encoded chemical entities during combinatorial synthesis processes. This has traditionally been achieved using enzymatic ligation. We discuss a range of chemical ligation methods that provide alternatives to enzymatic ligation. These chemical ligation methods include the generation of modified internucleotide linkages that support polymerase translocation and other modified linkages that while not supporting the translocation of polymerases can also be used to generate individual cDNA molecules containing encoded chemical information specifying individual library members. We also describe which of these approaches have been successfully utilized for the preparation of DNA-encoded chemical libraries and those that were subsequently used for the discovery of inhibitors.

  19. Simple Protein Complex Purification and Identification Method Suitable for High- throughput Mapping of Protein Interaction Networks

    Energy Technology Data Exchange (ETDEWEB)

    Markillie, Lye Meng; Lin, Chiann Tso; Adkins, Joshua N.; Auberry, Deanna L.; Hill, Eric A.; Hooker, Brian S.; Moore, Priscilla A.; Moore, Ronald J.; Shi, Liang; Wiley, H. S.; Kery, Vladimir

    2005-04-11

    Most of the current methods for purification and identification of protein complexes use endogenous expression of affinity tagged bait, tandem affinity tag purification of protein complexes followed by specific elution of complexes from beads, gel separation, in-gel digestion and mass spectrometric analysis of protein interactors. We propose a single affinity tag in vitro pulldown assay with denaturing elution, trypsin digestion in organic solvent and LC ESI MS/MS protein identification using SEQUEST analysis. Our method is simple, easy to scale up and automate thus suitable for high throughput mapping of protein interaction networks and functional proteomics.

  20. Adaptor-tagged competitive PCR: a novel method for measuring relative gene expression.

    OpenAIRE

    Kato, K.

    1997-01-01

    A simple and reliable PCR-based method to quantitate gene expression is described. Following the digestion of double-stranded cDNA by a restriction enzyme, an adaptor is ligated to a cDNA from a first RNA sample, and another adaptor to a second RNA sample. The two adaptors share a common sequence at the outer region, but differ in size. Equal amounts of the ligated samples are mixed, and amplified by an adaptor-primer and a primer specific to the gene of interest. Products derived from the tw...

  1. A Review on Chipless RFID Tag Design

    Directory of Open Access Journals (Sweden)

    Ali Hashemi

    2013-06-01

    Full Text Available The demand for RFID tags has recently increased due to the development of the RFID. Over the past decade, great efforts have been devoted to the design of RFID tags without chip inside. The concept of chipless RFID tags appears to be a promising solution for low cost item tagging because the cost of active RFID tag depends mainly on the chip used in them. This paper presents the progress of the chipless RFID tag and introduces major studies in this filed. Advantages and disadvantages of these methods are pointed out.

  2. Two-dimensional Length Extraction of Ballistic Target from ISAR Images Using a New Scaling Method by Affine Registration

    Directory of Open Access Journals (Sweden)

    Jin Guanghu

    2014-09-01

    Full Text Available The length of ballistic target is one of the most important features for target recognition. It can be extracted from ISAR Images. Unlike from the optical image, the length extraction from ISAR image has two difficulties. The first one is that it is hard to get the actual position of scattering centres by the traditional target extraction method. The second one is that the ISAR image’s cross scale is not known because of the target’s complex rotation. Here we propose two methods to solve these problems. Firstly, we use clustering method to get scattering centers. Secondly we propose to get cross scale of the ISAR images by affine registration. Experiments verified that our approach is realisable and has good performance.Defence Science Journal, Vol. 64, No. 5, September 2014, pp.458-463, DOI:http://dx.doi.org/10.14429/dsj.64.5001

  3. A new real-time method for investigation of affinity properties and binding kinetics of magnetic nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Orlov, Alexey V. [Prokhorov General Physics Institute, Russian Academy of Sciences, 38 Vavilov St, 119991 Moscow (Russian Federation); Nikitin, Maxim P. [Prokhorov General Physics Institute, Russian Academy of Sciences, 38 Vavilov St, 119991 Moscow (Russian Federation); Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 16/10 Miklukho-Maklaya St, 117997 Moscow (Russian Federation); Moscow Institute of Physics and Technology, 9 Institutskii per., Dolgoprudny, Moscow Region 141700 (Russian Federation); Bragina, Vera A.; Znoyko, Sergey L.; Zaikina, Marina N.; Ksenevich, Tatiana I.; Gorshkov, Boris G. [Prokhorov General Physics Institute, Russian Academy of Sciences, 38 Vavilov St, 119991 Moscow (Russian Federation); Nikitin, Petr I., E-mail: nikitin@kapella.gpi.ru [Prokhorov General Physics Institute, Russian Academy of Sciences, 38 Vavilov St, 119991 Moscow (Russian Federation); National Research Nuclear University MEPhI (Moscow Engineering Physics Institute), 31 Kashirskoe shosse, 115409 Moscow (Russian Federation)

    2015-04-15

    A method for quantitative investigation of affinity constants of receptors immobilized on magnetic nanoparticles (MP) is developed based on spectral correlation interferometry (SCI). The SCI records with a picometer resolution the thickness changes of a layer of molecules or nanoparticles due to a biochemical reaction on a cover slip, averaged over the sensing area. The method is compatible with other types of sensing surfaces employed in biosensing. The measured values of kinetic association constants of magnetic nanoparticles are 4 orders of magnitude higher than those of molecular antibody association with antigen. The developed method also suggests highly sensitive detection of antigens in a wide dynamic range. The limit of detection of 92 pg/ml has been demonstrated for prostate-specific antigen (PSA) with 50-nm MP employed as labels, which produce 3-order amplification of the SCI signals. The calibration curve features high sensitivity (slope) of 3-fold signal raise per 10-fold increase of PSA concentration within 4-order dynamic range, which is an attractive compromise for precise quantitative and highly sensitive immunoassay. The proposed biosensing technique offers inexpensive disposable sensor chips of cover slips and represents an economically sound alternative to traditional immunoassays for disease diagnostics, detection of pathogens in food and environmental monitoring. - Highlights: • Method for study of affinity constants of magnetic nanoparticles with receptors is proposed. • Association constants of such particles are 4 orders higher than for biomolecules. • Method is compatible with widely used biosensor surfaces and affordable consumables. • It has high sensitivity: 3-fold signal increasing per 10-fold of PSA concentration. • Limit of detection for PSA is 92 pg/ml, dynamic range – 4 orders of concentration.

  4. Prediction of MHC class II binding affinity using SMM-align, a novel stabilization matrix alignment method

    Directory of Open Access Journals (Sweden)

    Lund Ole

    2007-07-01

    Full Text Available Abstract Background Antigen presenting cells (APCs sample the extra cellular space and present peptides from here to T helper cells, which can be activated if the peptides are of foreign origin. The peptides are presented on the surface of the cells in complex with major histocompatibility class II (MHC II molecules. Identification of peptides that bind MHC II molecules is thus a key step in rational vaccine design and developing methods for accurate prediction of the peptide:MHC interactions play a central role in epitope discovery. The MHC class II binding groove is open at both ends making the correct alignment of a peptide in the binding groove a crucial part of identifying the core of an MHC class II binding motif. Here, we present a novel stabilization matrix alignment method, SMM-align, that allows for direct prediction of peptide:MHC binding affinities. The predictive performance of the method is validated on a large MHC class II benchmark data set covering 14 HLA-DR (human MHC and three mouse H2-IA alleles. Results The predictive performance of the SMM-align method was demonstrated to be superior to that of the Gibbs sampler, TEPITOPE, SVRMHC, and MHCpred methods. Cross validation between peptide data set obtained from different sources demonstrated that direct incorporation of peptide length potentially results in over-fitting of the binding prediction method. Focusing on amino terminal peptide flanking residues (PFR, we demonstrate a consistent gain in predictive performance by favoring binding registers with a minimum PFR length of two amino acids. Visualizing the binding motif as obtained by the SMM-align and TEPITOPE methods highlights a series of fundamental discrepancies between the two predicted motifs. For the DRB1*1302 allele for instance, the TEPITOPE method favors basic amino acids at most anchor positions, whereas the SMM-align method identifies a preference for hydrophobic or neutral amino acids at the anchors. Conclusion

  5. Shark Tagging Activities.

    Science.gov (United States)

    Current: The Journal of Marine Education, 1998

    1998-01-01

    In this group activity, children learn about the purpose of tagging and how scientists tag a shark. Using a cut-out of a shark, students identify, measure, record data, read coordinates, and tag a shark. Includes introductory information about the purpose of tagging and the procedure, a data sheet showing original tagging data from Tampa Bay, and…

  6. Tags, micro-tags and tag editing: improving internet search

    Science.gov (United States)

    Rogowitz, Bernice E.; Topkara, Mercan

    2009-02-01

    Social tagging is an emerging methodology that allows individual users to assign semantic keywords to content on the web. Popular web services allow the community of users to search for content based on these user-defined tags. Tags are typically attached to a whole entity such as a web page (e.g., del.icio.us), a video (e.g., YouTube), a product description (e.g., Amazon) or a photograph (e.g., Flickr). However, finding specific information within a whole entity can be a difficult, time-intensive process. This is especially true for content such as video, where the information sought may be a small segment within a very long presentation. Moreover, the tags provided by a community of users may be incorrect, conflicting, or incomplete when used as search terms. In this paper we introduce a system that allows users to create "micro-tags," that is, semantic markers that are attached to subsets of information. These micro-tags give the user the ability to direct attention to specific subsets within a larger and more complex entity, and the set of micro-tags provides a more nuanced description of the full content. Also, when these micro-tags are used as search terms, there is no need to do a serial search of the content, since micro-tags draw attention to the semantic content of interest. This system also provides a mechanism that allows users in the community to edit and delete each others' tags, using the community to refine and improve tag quality. We will also report on empirical studies that demonstrate the value of micro-tagging and tag editing and explore the role micro-tags and tag editing will play in future applications.

  7. Affinity in electrophoresis.

    Science.gov (United States)

    Heegaard, Niels H H

    2009-06-01

    The journal Electrophoresis has greatly influenced my approaches to biomolecular affinity studies. The methods that I have chosen as my main tools to study interacting biomolecules--native gel and later capillary zone electrophoresis--have been the topic of numerous articles in Electrophoresis. Below, the role of the journal in the development and dissemination of these techniques and applications reviewed. Many exhaustive reviews on affinity electrophoresis and affinity CE have been published in the last few years and are not in any way replaced by the present deliberations that are focused on papers published by the journal.

  8. Yellowtail Tagging Data (MRDBS)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Yellowtail Flounder Tagging Program began in 2003 and works with commercial fishermen to tag and release yellowtaiI flounder with pink and yellow disc tags or...

  9. Restriction site extension PCR: a novel method for high-throughput characterization of tagged DNA fragments and genome walking.

    Directory of Open Access Journals (Sweden)

    Jiabing Ji

    Full Text Available BACKGROUND: Insertion mutant isolation and characterization are extremely valuable for linking genes to physiological function. Once an insertion mutant phenotype is identified, the challenge is to isolate the responsible gene. Multiple strategies have been employed to isolate unknown genomic DNA that flanks mutagenic insertions, however, all these methods suffer from limitations due to inefficient ligation steps, inclusion of restriction sites within the target DNA, and non-specific product generation. These limitations become close to insurmountable when the goal is to identify insertion sites in a high throughput manner. METHODOLOGY/PRINCIPAL FINDINGS: We designed a novel strategy called Restriction Site Extension PCR (RSE-PCR to efficiently conduct large-scale isolation of unknown genomic DNA fragments linked to DNA insertions. The strategy is a modified adaptor-mediated PCR without ligation. An adapter, with complementarity to the 3' overhang of the endonuclease (KpnI, NsiI, PstI, or SacI restricted DNA fragments, extends the 3' end of the DNA fragments in the first cycle of the primary RSE-PCR. During subsequent PCR cycles and a second semi-nested PCR (secondary RSE-PCR, touchdown and two-step PCR are combined to increase the amplification specificity of target fragments. The efficiency and specificity was demonstrated in our characterization of 37 tex mutants of Arabidopsis. All the steps of RSE-PCR can be executed in a 96 well PCR plate. Finally, RSE-PCR serves as a successful alternative to Genome Walker as demonstrated by gene isolation from maize, a plant with a more complex genome than Arabidopsis. CONCLUSIONS/SIGNIFICANCE: RSE-PCR has high potential application in identifying tagged (T-DNA or transposon sequence or walking from known DNA toward unknown regions in large-genome plants, with likely application in other organisms as well.

  10. Improved methodology for the affinity isolation of human protein complexes expressed at near endogenous levels

    DEFF Research Database (Denmark)

    Domanski, Michal; Molloy, Kelly; Jiang, Hua;

    2012-01-01

    An efficient and reliable procedure for the capture of affinity-tagged proteins and associated complexes from human cell lines is reported. Through multiple optimizations, high yield and low background affinity-purifications are achieved from modest quantities of human cells expressing endogenous......-level tagged proteins. Isolations of triple-FLAG and GFP-tagged fusion proteins involved in RNA metabolism are presented.......An efficient and reliable procedure for the capture of affinity-tagged proteins and associated complexes from human cell lines is reported. Through multiple optimizations, high yield and low background affinity-purifications are achieved from modest quantities of human cells expressing endogenous...

  11. High-Throughput Melanin-Binding Affinity and In Silico Methods to Aid in the Prediction of Drug Exposure in Ocular Tissue.

    Science.gov (United States)

    Reilly, John; Williams, Sarah L; Forster, Cornelia J; Kansara, Viral; End, Peter; Serrano-Wu, Michael H

    2015-12-01

    Drugs possessing the ability to bind to melanin-rich tissue, such as the eye, are linked with higher ocular exposure, and therefore have the potential to affect the efficacy and safety profiles of therapeutics. A high-throughput melanin chromatographic affinity assay has been developed and validated, which has allowed the rapid melanin affinity assessment for a large number of compounds. Melanin affinity of compounds can be quickly assigned as low, medium, or high melanin binders. A high-throughput chromatographic method has been developed and fully validated to assess melanin affinity of pharmaceuticals and has been useful in predicting ocular tissue distribution in vivo studies. The high-throughput experimental approach has also allowed for a specific training set of 263 molecules for a quantitative structure-affinity relationships (QSAR) method to be developed, which has also been shown to be a predictor of ocular tissue exposure. Previous studies have reported the development of in silico QSAR models based on training sets of relatively small and mostly similar compounds; this model covers a broader range of melanin-binding affinities than what has been previously published and identified several physiochemical descriptors to be considered in the design of compounds where melanin-binding modulation is desired.

  12. A method for the chemical modification of polychlorinated biphenyls for improved affinity towards noble metal surfaces

    DEFF Research Database (Denmark)

    2015-01-01

    The present application discloses a method for the modification and analysis of a field sample suspected of containing contaminant(s) like polychlorinated biphenyls (PCBs). The invention also relates to a corresponding kit for the modification of samples suspected of containing such contaminant(s)....

  13. Cloning and Expression of Ontak Immunotoxin Using Intein Tag

    Directory of Open Access Journals (Sweden)

    SA Moosavizadeh

    2016-06-01

    Full Text Available Introduction: Inteins (INT are internal parts of a number of proteins in yeast and some other unicellular eukaryotes, which can be separated from the immature protein during protein splicing process. After identifying the mechanism of intein action, applications of these sequences are be considered in the single- step purification of recombinant proteins and different intein tags were developed. The most important advantage of using intein tags in purification of recombinant proteins than other affinity tags is no requirement of expensive protease enzymes and following additional steps to remove protease that make intein tags economically are considered more important. In the present study, denileukin diftitox immunotoxin (brand name Ontak, be fused with an intein tag and it was inserted in pTXB1 plasmid. Methods: In this study, with respect to multiple cloning sites (MCS of pTXB1, specific primers were designed. Polymerase Chain Reaction (PCR was performed and encoding sequence of ONTAK was cloned using restriction sites of NdeI and SapI. Recombinant vector (PTX-IDZ was transformed into E. coli strain ER2566 and expression of gene was studied. Results: The accuracy of recombinant construct was confirmed by PCR and enzymatic digestion. The produced recombinant proteins were confirmed by SDS-PAGE and Western blotting. Conclusion: Restriction site of SapI guarantees no additional residues incorporate in primary protein sequence. Also, the expression of this construct was analyzed in compare with fused protein to poly-His tag. According to the appropriate expression of fused protein in both constructs it was expected that one step- purification of considered drug protein will be success in the following steps.

  14. Generation of an affinity column for antibody purification by intein-mediated protein ligation.

    Science.gov (United States)

    Sun, Luo; Ghosh, Inca; Xu, Ming-Qun

    2003-11-01

    Coupling an antigenic peptide to a solid support is a crucial step in the affinity purification of a peptide-specific antibody. Conventional methods for generating reactive agarose, cellulose or other matrices for peptide conjugation are laborious and can result in a significant amount of chemical waste. In this report, we present a novel method for the facile production of a peptide affinity column by employing intein-mediated protein ligation (IPL) in conjunction with chitin affinity chromatography. A reactive thioester was generated at the C-terminal of the chitin binding domain (CBD) from the chitinase A1 of Bacillus circulans WL-2 by thiol-induced cleavage of the peptide bond between the CBD and a modified intein. Peptide epitopes possessing an N-terminal cysteine were ligated to the chitin bound CBD tag. We demonstrate that the resulting peptide columns permit the highly specific and efficient affinity purification of antibodies from animal sera.

  15. Critical evaluation of sequential extraction and sink-float methods used for the determination of Ga and Ge affinity in lignite

    Energy Technology Data Exchange (ETDEWEB)

    Zdenek Klika; Lenka Ambruzova; Ivana Sykorova; Jana Seidlerova; Ivan Kolomaznik [VSB-Technical University Ostrava, Ostrava (Czech Republic)

    2009-10-15

    The affinities of Ga and Ge in lignite were determined using sequential extraction (SE) and element affinity calculation (EAC) based on sink-float data. For this study a bulk lignite sample was fractioned into two sets. The first set of samples (A) consisted of the different grain sizes fractions; the second one set (B) was prepared by density fractionation. Sequential extractions (1) were performed on both sets of fractions with very good agreement between determined organic elements affinities (OEA of Ga evaluated from A data is 32%, from B data 35%; OEA of Ge evaluated from A data is 31% and from B data 26%). The data of B lignite fractions were evaluated using two element affinity calculations: (a) EAC (I) of Klika and Kolomaznk (2) and (b) newly prepared subroutine EAC (II) based on quantitative contents of lignite macerals and minerals. There was also good agreement between both methods obtained (OEA of Ga calculated by EAC (I) is 83% and by EAC (II) 77%; OEA of Ge calculated by EAC (I) is 89% and by EAC (II) 97%). The significant differences of organic elements affinities of Ga and Ge evaluated by sequential extraction and by element affinity calculation based on sink-float data are discussed. 34 refs., 7 figs., 6 tabs.

  16. Fluorinated carbon tag derivatization combined with fluorous solid-phase extraction: a new method for the highly sensitive and selective mass spectrometric analysis of glycans.

    Science.gov (United States)

    Li, Lulu; Jiao, Jing; Cai, Yan; Zhang, Ying; Lu, Haojie

    2015-01-01

    The sensitive and specific detection of glycans via mass spectrometry (MS) remains a significant challenge due to their low abundance in complex biological mixtures, inherent lack of hydrophobicity, and suppression by other, more abundant biological molecules (proteins/peptides) or contaminants. A new strategy for the sensitive and selective MS analysis of glycans based on fluorous chemistry is reported. Glycan reducing ends were derivatized with a hydrophobic fluorinated carbon tag, increasing glycan ionization efficiency during MS by more than an order of magnitude. More importantly, the fluorinated carbon tag enabled efficient fluorous solid-phase extraction (FSPE) to specifically enrich the glycans from contaminated solutions and protein mixtures. Finally, we successfully analyzed the N-glycome in human serum using this new method.

  17. Purification of GST-Tagged Proteins.

    Science.gov (United States)

    Schäfer, Frank; Seip, Nicole; Maertens, Barbara; Block, Helena; Kubicek, Jan

    2015-01-01

    This protocol describes the purification of recombinant proteins fused to glutathione S-transferase (GST, GST-tagged proteins) by Glutathione Affinity purification. The GST tag frequently increases the solubility of the fused protein of interest and thus enables its purification and subsequent functional characterization. The GST-tagged protein specifically binds to glutathione immobilized to a matrix (e.g., agarose) and can be easily separated from a cell lysate by a bind-wash-elute procedure. GST-tagged proteins are often used to study protein-protein interactions, again making use of glutathione affinity in a procedure called a GST pull-down assay. The protocol is designed to process 200 ml of E. coli culture expressing intermediate to high amounts of a GST-tagged protein (~25 mg l(-1)). Depending on the expression rate or the available culture volume, the scale can be increased or decreased linearly. The protocol can also be used to purify GST-tagged proteins from other expression systems, such as insect or mammalian cells. Tips are provided to aid in modifying certain steps if proteins shall be recovered from alternative expression systems.

  18. b-tagging in DELPHI at LEP

    CERN Document Server

    Abdallah, J; Adam, W; Adye, T; Adzic, P; Albrecht, T; Alderweireld, T; Alemany-Fernandez, R; Allmendinger, T; Allport, P P; Almehed, S; Amaldi, Ugo; Amapane, N; Amato, S; Anashkin, E; Andreazza, A; Andringa, S; Anjos, N; Antilogus, P; Apel, W D; Arnoud, Y; Ask, S; Åsman, B; Augustin, J E; Augustinus, A; Baillon, Paul; Ballestrero, A; Bambade, P; Barbier, R; Bardin, Dimitri Yuri; Barker, G; Baroncelli, A; Bates, M; Battaglia, Marco; Baubillier, M; Becks, K H; Begalli, M; Behrmann, A; Benekos, N C; Benvenuti, Alberto C; Bérat, C; Berggren, M; Berntzon, L; Bertrand, D; Besançon, M; Besson, N; Bibby, J; Biffi, P; Bloch, D; Blom, M; Bonesini, M; Boonekamp, M; Booth, P S L; Borisov, G; Botner, O; Bouquet, B; Bowcock, T J V; Boyko, I; Bracko, M; Branchini, P; Brenner, R; Brodet, E; Brückman, P; Brunet, J M; Bugge, L; Buschmann, P; Caccia, M; Calvi, M; Camporesi, T; Canale, V; Carena, F; Castro, N; Cavallo, F R; Chabaud, V; Chapkin, M M; Charpentier, P; Checchia, P; Chierici, R; Shlyapnikov, P; Chudoba, J; Chung, S U; Cieslik, K; Collins, P; Contri, R; Cosme, G; Cossutti, F; Costa, M J; Couchot, F; Crawley, B; Crennell, D J; Cuevas-Maestro, J; D'Almagne, B; D'Hondt, J; Dalmau, J; Da Silva, T; Da Silva, W; Della Ricca, G; De Angelis, A; de Boer, Wim; De Clercq, C; De Lotto, B; De Maria, N; De Min, A; De Paula, L S; Di Ciaccio, Lucia; Dijkstra, H; Di Simone, A; Doroba, K; Drees, J; Dris, M; Eigen, G; Ekelöf, T J C; Ellert, M; Elsing, M; Espirito-Santo, M C; Fanourakis, G K; Fassouliotis, D; Feindt, M; Fernández, J; Ferrer, A; Ferro, F; Flagmeyer, U; Föth, H; Fokitis, E; Fulda-Quenzer, F; Fuster, J A; Gandelman, M; García, C; Gavillet, P; Gazis, E N; Geralis, T; Gokieli, R; Golob, B; Gómez-Cadenas, J J; Gómez-Ceballos, G; Gonçalves, P; Graziani, E; Grosdidier, G; Grzelak, K; Guy, J; Haag, C; Hallgren, A; Hamacher, K; Hamilton, K; Hansen, J; Haug, S; Hauler, F; Hedberg, V; Hennecke, M; Hernando, J A; Herr, H; Heuser, J M; Holmgren, S O; Holt, P J; Houlden, M A; Hultqvist, K; Jackson, J N; Jalocha, P; Jarlskog, C; Jarlskog, G; Jarry, P; Jeans, D; Johansson, E K; Johansson, P D; Jonsson, P; Joram, C; Jungermann, L; Kapusta, F; Karlsson, M; Katsanevas, S; Katsoufis, E C; Keränen, R; Kernel, G; Kersevan, Borut P; Kiiskinen, A P; King, B T; Kjaer, N J; Kluit, P; Kokkinias, P; Kourkoumelis, C; Kuznetsov, O; Krumshtein, Z; Kucharczyk, M; Kucewicz, W; Kurowska, J; Lamsa, J; Leder, G; Ledroit, F; Leinonen, L; Leitner, R; Lemonne, J; Lepeltier, V; Lesiak, T; Liebig, W; Liko, D; Lipniacka, A; Lopes, J H; López, J M; Loukas, D; Lutz, P; Lyons, L; MacNaughton, J; Malek, A; Maltezos, S; Mandl, F; Marco, J; Marco, R; Maréchal, B; Margoni, M; Marin, J C; Mariotti, C; Markou, A; Martínez-Rivero, C; Martínez-Vidal, F; Masik, J; Mastroyiannopoulos, N; Matorras, F; Matteuzzi, C; Mazzucato, F; Mazzucato, M; McNulty, R; Meroni, C; Meyer, W T; Migliore, E; Mitaroff, W A; Mjörnmark, U; Moa, T; Moch, M; Mönig, K; Monge, R; Montenegro, J; Moraes, D; Moreno, S; Morettini, P; Müller, U; Münich, K; Mulders, M; Mundim, L; Murray, W; Muryn, B; Myatt, Gerald; Myklebust, T; Nassiakou, M; Navarria, Francesco Luigi; Nawrocki, K; Nicolaidou, R; Niezurawski, P; Nikolenko, M; Nomerotski, A; Norman, A; Nygren, A; Oblakowska-Mucha, A; Obraztsov, V F; Olshevskii, A G; Onofre, A; Orava, Risto; Österberg, K; Ouraou, A; Oyanguren, A; Paganoni, M; Paiano, S; Palacios, J P; Palka, H; Papadopoulou, T D; Pape, L; Parkes, C; Parodi, F; Parzefall, U; Passeri, A; Passon, O; Peralta, L; Perepelitsa, V F; Perrotta, A; Petrolini, A; Piedra, J; Pieri, L; Pierre, F; Pimenta, M; Piotto, E; Podobnik, T; Poireau, V; Pol, M E; Polok, G; Poropat, P; Pozdnyakov, V; Pukhaeva, N; Pullia, Antonio; Rames, J; Ramler, L; Read, A; Rebecchi, P; Rehn, J; Reid, D; Reinhardt, R; Renton, P B; Richard, F; Rídky, J; Rivero, M; Rodríguez, D; Romero, A; Ronchese, P; Rosenberg, E I; Roudeau, Patrick; Rovelli, T; Ruhlmann-Kleider, V; Ryabtchikov, D; Sadovskii, A; Salmi, L; Salt, J; Savoy-Navarro, A; Schwickerath, U; Segar, A; Sekulin, R L; Siebel, M; Sissakian, A N; Smadja, G; Smirnova, O G; Sokolov, A; Sopczak, A; Sosnowski, R; Spassoff, Tz; Stanitzki, M; Stavitski, I; Stocchi, A; Strauss, J; Stugu, B; Szczekowski, M; Szeptycka, M; Szumlak, T; Tabarelli de Fatis, T; Taffard, A C; Tegenfeldt, F; Timmermans, J; Tinti, N; Tkatchev, L G; Tobin, M; Todorovova, S; Tomaradze, A G; Tomé, B; Tonazzo, A; Tortosa, P; Travnicek, P; Treille, D; Trischuk, W; Tristram, G; Trochimczuk, M; Troncon, C; Turluer, M L; Tyapkin, I A; Tyapkin, P; Tyndel, M; Tzamarias, S; Uvarov, V; Valenti, G; van Dam, P; Van Eldik, J; Van Lysebetten, A; Van Remortel, N; Van Vulpen, I B; Vegni, G; Veloso, F; Venus, W A; Verbeure, F; Verdier, P; Verzi, V; Vilanova, D; Vitale, L; Vrba, V; Wahlen, H; Washbrook, A J; Weilhammer, Peter; Weiser, C; Wicke, D; Wickens, J H; Wilkinson, G; Winter, M; Witek, M; Yushchenko, O P; Zalewska-Bak, A; Zalewski, Piotr; Zavrtanik, D; Zimin, N I; Zinchenko, A I; Zupan, M

    2004-01-01

    The standard method used for tagging b-hadrons in the DELPHI experiment at the CERN LEP Collider is discussed in detail. The main ingredient of b-tagging is the impact parameters of tracks, which relies mostly on the vertex detector. Additional information, such as the mass of particles associated to a secondary vertex, significantly improves the selection efficiency and the background suppression. The paper describes various discriminating variables used for the tagging and the procedure of their combination. In addition, applications of b-tagging to some physics analyses, which depend crucially on the performance and reliability of b-tagging, are described briefly.

  19. LOMA: A fast method to generate efficient tagged-random primers despite amplification bias of random PCR on pathogens

    Directory of Open Access Journals (Sweden)

    Lee Wah

    2008-09-01

    Full Text Available Abstract Background Pathogen detection using DNA microarrays has the potential to become a fast and comprehensive diagnostics tool. However, since pathogen detection chips currently utilize random primers rather than specific primers for the RT-PCR step, bias inherent in random PCR amplification becomes a serious problem that causes large inaccuracies in hybridization signals. Results In this paper, we study how the efficiency of random PCR amplification affects hybridization signals. We describe a model that predicts the amplification efficiency of a given random primer on a target viral genome. The prediction allows us to filter false-negative probes of the genome that lie in regions of poor random PCR amplification and improves the accuracy of pathogen detection. Subsequently, we propose LOMA, an algorithm to generate random primers that have good amplification efficiency. Wet-lab validation showed that the generated random primers improve the amplification efficiency significantly. Conclusion The blind use of a random primer with attached universal tag (random-tagged primer in a PCR reaction on a pathogen sample may not lead to a successful amplification. Thus, the design of random-tagged primers is an important consideration when performing PCR.

  20. Engineering Escherichia coli BL21(DE3) derivative strains to minimize E. coli protein contamination after purification by immobilized metal affinity chromatography.

    Science.gov (United States)

    Robichon, Carine; Luo, Jianying; Causey, Thomas B; Benner, Jack S; Samuelson, James C

    2011-07-01

    Recombinant His-tagged proteins expressed in Escherichia coli and purified by immobilized metal affinity chromatography (IMAC) are commonly coeluted with native E. coli proteins, especially if the recombinant protein is expressed at a low level. The E. coli contaminants display high affinity to divalent nickel or cobalt ions, mainly due to the presence of clustered histidine residues or biologically relevant metal binding sites. To improve the final purity of expressed His-tagged protein, we engineered E. coli BL21(DE3) expression strains in which the most recurring contaminants are either expressed with an alternative tag or mutated to decrease their affinity to divalent cations. The current study presents the design, engineering, and characterization of two E. coli BL21(DE3) derivatives, NiCo21(DE3) and NiCo22(DE3), which express the endogenous proteins SlyD, Can, ArnA, and (optionally) AceE fused at their C terminus to a chitin binding domain (CBD) and the protein GlmS, with six surface histidines replaced by alanines. We show that each E. coli CBD-tagged protein remains active and can be efficiently eliminated from an IMAC elution fraction using a chitin column flowthrough step, while the modification of GlmS results in loss of affinity for nickel-containing resin. The "NiCo" strains uniquely complement existing methods for improving the purity of recombinant His-tagged protein.

  1. Study of the interactions between fluoroquinolones and human serum albumin by affinity capillary electrophoresis and fluorescence method

    Energy Technology Data Exchange (ETDEWEB)

    Zhang Liwei; Wang Kun [Beijing National Laboratory for Molecular Sciences (BNLMS), College of Chemistry, Peking University, Beijing 100871 (China); Zhang Xinxiang [Beijing National Laboratory for Molecular Sciences (BNLMS), College of Chemistry, Peking University, Beijing 100871 (China)], E-mail: zxx@pku.edu.cn

    2007-11-05

    The interactions between fluoroquinolones and human serum albumin (HSA) were investigated by affinity capillary electrophoresis (ACE) and fluorescence quenching technique. Based on the efficient separation of several fluoroquinolones using a simple phosphate buffer, the binding constants of fluoroquinolones with HSA were determined simultaneously during one set of electrophoresis by ACE method. The thermodynamic parameters were obtained from data at different temperatures, and the negative {delta}H and {delta}S values showed that both hydrogen bonds and van der Waals interaction played major roles in the binding of fluoroquinolones to HSA. The interactions were also studied by fluorescence quenching technique. The results of fluorescence titration revealed that fluoroquinolones had the strong ability to quenching the intrinsic fluorescence of HSA through the static quenching procedure. The binding site number n, apparent binding constant K{sub b} and the Stern-Volmer quenching constant K{sub sv} were determined. The thermodynamic parameters were also studied by fluorescence method, and the results were consonant with that of ACE.

  2. Measurement of the CP-Violation Parameter sin⁡2ϕ1 with a New Tagging Method at the Υ(5S) Resonance

    Science.gov (United States)

    Sato, Y.; Yamamoto, H.; Aihara, H.; Asner, D. M.; Aulchenko, V.; Aushev, T.; Aziz, T.; Bakich, A. M.; Bhardwaj, V.; Bhuyan, B.; Bischofberger, M.; Bondar, A.; Bozek, A.; Bračko, M.; Browder, T. E.; Chang, P.; Chen, P.; Cheon, B. G.; Chilikin, K.; Chistov, R.; Cho, I.-S.; Cho, K.; Choi, S.-K.; Choi, Y.; Dalseno, J.; Doležal, Z.; Drásal, Z.; Eidelman, S.; Epifanov, D.; Fast, J. E.; Gaur, V.; Gabyshev, N.; Goh, Y. M.; Golob, B.; Haba, J.; Hara, T.; Hayasaka, K.; Hayashii, H.; Horii, Y.; Hoshi, Y.; Hou, W.-S.; Hyun, H. J.; Ishikawa, A.; Itoh, R.; Iwabuchi, M.; Iwasaki, Y.; Iwashita, T.; Julius, T.; Kapusta, P.; Kawasaki, T.; Kichimi, H.; Kiesling, C.; Kim, H. J.; Kim, H. O.; Kim, J. B.; Kim, J. H.; Kim, K. T.; Kim, M. J.; Kim, S. K.; Kim, Y. J.; Kinoshita, K.; Ko, B. R.; Kobayashi, N.; Kodyš, P.; Korpar, S.; Križan, P.; Krokovny, P.; Kuhr, T.; Kumar, R.; Kumita, T.; Kuzmin, A.; Kwon, Y.-J.; Lange, J. S.; Lee, S.-H.; Li, J.; Li, Y.; Liu, C.; Liu, Z. Q.; Louvot, R.; McOnie, S.; Miyabayashi, K.; Miyata, H.; Mizuk, R.; Mohanty, G. B.; Moll, A.; Muramatsu, N.; Nakano, E.; Nakao, M.; Nakazawa, H.; Natkaniec, Z.; Nishida, S.; Nishimura, K.; Nitoh, O.; Ogawa, S.; Ohshima, T.; Okuno, S.; Olsen, S. L.; Onuki, Y.; Ostrowicz, W.; Pakhlov, P.; Pakhlova, G.; Park, C. W.; Park, H.; Park, H. K.; Pedlar, T. K.; Petrič, M.; Piilonen, L. E.; Poluektov, A.; Röhrken, M.; Ryu, S.; Sahoo, H.; Sakai, Y.; Sanuki, T.; Schneider, O.; Schwanda, C.; Schwartz, A. J.; Seidl, R.; Senyo, K.; Seon, O.; Sevior, M. E.; Shapkin, M.; Shen, C. P.; Shibata, T.-A.; Shiu, J.-G.; Shwartz, B.; Sibidanov, A.; Simon, F.; Smerkol, P.; Sohn, Y.-S.; Sokolov, A.; Solovieva, E.; Stanič, S.; Starič, M.; Stypula, J.; Sumihama, M.; Sumiyoshi, T.; Tanaka, S.; Tatishvili, G.; Teramoto, Y.; Trabelsi, K.; Uchida, M.; Uglov, T.; Unno, Y.; Uno, S.; Urquijo, P.; Varner, G.; Varvell, K. E.; Wang, C. H.; Wang, M.-Z.; Wang, P.; Wang, X. L.; Watanabe, M.; Watanabe, Y.; Wicht, J.; Won, E.; Yabsley, B. D.; Yamashita, Y.; Yusa, Y.; Zhang, Z. P.; Zhilich, V.; Zhulanov, V.; Zupanc, A.

    2012-04-01

    We report a measurement of the CP-violation parameter sin⁡2ϕ1 at the Υ(5S) resonance using a new tagging method, called “B-π tagging.” In Υ(5S) decays containing a neutral B meson, a charged B, and a charged pion, the neutral B is reconstructed in the J/ψKS0 CP-eigenstate decay channel. The initial flavor of the neutral B meson at the moment of the Υ(5S) decay is opposite to that of the charged B and may thus be inferred from the charge of the pion without reconstructing the charged B. From the asymmetry between B-π+ and B-π- tagged J/ψKS0 yields, we determine sin⁡2ϕ1=0.57±0.58(stat)±0.06(syst). The results are based on 121fb-1 of data recorded by the Belle detector at the KEKB e+e- collider.

  3. Measurement of the CP-violation Parameter sin2$\\phi_1$ with a New Tagging Method at the $\\Upsilon(5S)$ Resonance

    CERN Document Server

    Sato, Y; Aihara, H; Asner, D M; Aulchenko, V; Aushev, T; Aziz, T; Bakich, A M; Bhardwaj, V; Bhuyan, B; Bischofberger, M; Bondar, A; Bozek, A; Bračko, M; Browder, T E; Chang, P; Chen, P; Cheon, B G; Chilikin, K; Chistov, R; Cho, I -S; Cho, K; Choi, S -K; Choi, Y; Dalseno, J; Doležal, Z; Drásal, Z; Eidelman, S; Epifanov, D; Fast, J E; Gaur, V; Gabyshev, N; Goh, Y M; Golob, B; Haba, J; Hara, T; Hayasaka, K; Hayashii, H; Horii, Y; Hoshi, Y; Hou, W -S; Hyun, H J; Ishikawa, A; Itoh, R; Iwabuchi, M; Iwasaki, Y; Iwashita, T; Julius, T; Kapusta, P; Kawasaki, T; Kichimi, H; Kiesling, C; Kim, H J; Kim, H O; Kim, J B; Kim, J H; Kim, K T; Kim, M J; Kim, S K; Kim, Y J; Kinoshita, K; Ko, B R; Kobayashi, N; Kodyš, P; Korpar, S; Križan, P; Krokovny, P; Kuhr, T; Kumar, R; Kumita, T; Kuzmin, A; Kwon, Y -J; Lange, J S; Lee, S -H; Li, J; Li, Y; Liu, C; Liu, Z Q; Louvot, R; McOnie, S; Miyabayashi, K; Miyata, H; Mizuk, R; Mohanty, G B; Moll, A; Muramatsu, N; Nakano, E; Nakao, M; Natkaniec, Z; Nishida, S; Nishimura, K; Nitoh, O; Ogawa, S; Ohshima, T; Okuno, S; Olsen, S L; Onuki, Y; Ostrowicz, W; Pakhlov, P; Pakhlova, G; Park, C W; Park, H; Park, H K; Pedlar, T K; Petrič, M; Piilonen, L E; Poluektov, A; Röhrken, M; Ryu, S; Sahoo, H; Sakai, Y; Sanuki, T; Schneider, O; Schwanda, C; Schwartz, A J; Seidl, R; Senyo, K; Seon, O; Sevior, M E; Shapkin, M; Shen, C P; Shibata, T -A; Shiu, J -G; Shwartz, B; Sibidanov, A; Simon, F; Smerkol, P; Sohn, Y -S; Sokolov, A; Solovieva, E; Stanič, S; Starič, M; Stypula, J; Sumihama, M; Sumiyoshi, T; Tanaka, S; Tatishvili, G; Teramoto, Y; Trabelsi, K; Uchida, M; Uglov, T; Unno, Y; Uno, S; Urquijo, P; Varner, G; Varvell, K E; Wang, C H; Wang, M -Z; Wang, P; Wang, X L; Watanabe, M; Watanabe, Y; Wicht, J; Won, E; Yabsley, B D; Yamashita, Y; Yusa, Y; Zhang, Z P; Zhilich, V; Zhulanov, V; Zupanc, A

    2012-01-01

    We report a measurement of the CP-violation parameter sin2$\\phi_1$ at the $\\Upsilon(5S)$ resonance using a new tagging method, called "$B$-$\\pi$ tagging." In $\\Upsilon(5S)$ decays containing a neutral $B$ meson, a charged $B$, and a charged pion, the neutral $B$ is reconstructed in the $J/\\psi K_S^0$ CP-eigenstate decay channel. The initial flavor of the neutral $B$ meson at the moment of the $\\Upsilon(5S)$ decay is opposite to that of the charged $B$ and may thus be inferred from the charge of the pion without reconstructing the charged $B$. From the asymmetry between $B$-$\\pi^+$ and $B$-$\\pi^-$ tagged $J/\\psi K_S^0$ yields, we determine sin2$\\phi_1$ = 0.57 $\\pm$ 0.58(stat) $\\pm$ 0.06(syst). The results are based on 121 fb$^{-1}$ of data recorded by the Belle detector at the KEKB $e^+ e^-$ collider.

  4. Measurement of the CP-violation parameter sin2φ1 with a new tagging method at the Υ(5S) resonance.

    Science.gov (United States)

    Sato, Y; Yamamoto, H; Aihara, H; Asner, D M; Aulchenko, V; Aushev, T; Aziz, T; Bakich, A M; Bhardwaj, V; Bhuyan, B; Bischofberger, M; Bondar, A; Bozek, A; Bračko, M; Browder, T E; Chang, P; Chen, P; Cheon, B G; Chilikin, K; Chistov, R; Cho, I-S; Cho, K; Choi, S-K; Choi, Y; Dalseno, J; Doležal, Z; Drásal, Z; Eidelman, S; Epifanov, D; Fast, J E; Gaur, V; Gabyshev, N; Goh, Y M; Golob, B; Haba, J; Hara, T; Hayasaka, K; Hayashii, H; Horii, Y; Hoshi, Y; Hou, W-S; Hyun, H J; Ishikawa, A; Itoh, R; Iwabuchi, M; Iwasaki, Y; Iwashita, T; Julius, T; Kapusta, P; Kawasaki, T; Kichimi, H; Kiesling, C; Kim, H J; Kim, H O; Kim, J B; Kim, J H; Kim, K T; Kim, M J; Kim, S K; Kim, Y J; Kinoshita, K; Ko, B R; Kobayashi, N; Kodyš, P; Korpar, S; Križan, P; Krokovny, P; Kuhr, T; Kumar, R; Kumita, T; Kuzmin, A; Kwon, Y-J; Lange, J S; Lee, S-H; Li, J; Li, Y; Liu, C; Liu, Z Q; Louvot, R; McOnie, S; Miyabayashi, K; Miyata, H; Mizuk, R; Mohanty, G B; Moll, A; Muramatsu, N; Nakano, E; Nakao, M; Nakazawa, H; Natkaniec, Z; Nishida, S; Nishimura, K; Nitoh, O; Ogawa, S; Ohshima, T; Okuno, S; Olsen, S L; Onuki, Y; Ostrowicz, W; Pakhlov, P; Pakhlova, G; Park, C W; Park, H; Park, H K; Pedlar, T K; Petrič, M; Piilonen, L E; Poluektov, A; Röhrken, M; Ryu, S; Sahoo, H; Sakai, Y; Sanuki, T; Schneider, O; Schwanda, C; Schwartz, A J; Seidl, R; Senyo, K; Seon, O; Sevior, M E; Shapkin, M; Shen, C P; Shibata, T-A; Shiu, J-G; Shwartz, B; Sibidanov, A; Simon, F; Smerkol, P; Sohn, Y-S; Sokolov, A; Solovieva, E; Stanič, S; Starič, M; Stypula, J; Sumihama, M; Sumiyoshi, T; Tanaka, S; Tatishvili, G; Teramoto, Y; Trabelsi, K; Uchida, M; Uglov, T; Unno, Y; Uno, S; Urquijo, P; Varner, G; Varvell, K E; Wang, C H; Wang, M-Z; Wang, P; Wang, X L; Watanabe, M; Watanabe, Y; Wicht, J; Won, E; Yabsley, B D; Yamashita, Y; Yusa, Y; Zhang, Z P; Zhilich, V; Zhulanov, V; Zupanc, A

    2012-04-27

    We report a measurement of the CP-violation parameter sin2φ1 at the Υ(5S) resonance using a new tagging method, called "B-π tagging." In Υ(5S) decays containing a neutral B meson, a charged B, and a charged pion, the neutral B is reconstructed in the J/ψK(S)(0) CP-eigenstate decay channel. The initial flavor of the neutral B meson at the moment of the Υ(5S) decay is opposite to that of the charged B and may thus be inferred from the charge of the pion without reconstructing the charged B. From the asymmetry between B-π(+) and B-π(-) tagged J/ψK(S)(0) yields, we determine sin2φ1=0.57±0.58(stat)±0.06(syst). The results are based on 121 fb(-1) of data recorded by the Belle detector at the KEKB e(+)e(-) collider.

  5. Microplanktonic community structure in a coastal system relative to a Phaeocystis bloom inferred from morphological and tag pyrosequencing methods.

    Directory of Open Access Journals (Sweden)

    Sébastien Monchy

    Full Text Available BACKGROUND: Massive phytoplankton blooms, like the recurrent Phaeocystis proliferation observed every year in the Eastern English Channel (EEC, have a significant influence on the overall planktonic community structure and their food web dynamics. As well as being an important area for local fisheries, the EEC is an ideal ecosystem for work on microbial diversity. This is because, although its environmental context is relatively complex, it is reasonably well understood due to several years of monitoring and morphological observations of its planktonic organisms. The objective of our study was to better understand the under-explored microbial eukaryotic diversity relative to the Phaeocystis bloom. METHODOLOGY AND PRINCIPAL FINDINGS: The community structure of microplankton (diatoms, haptophytes, ciliates and dinoflagellates was studied through morphological observations and tag pyrosequencing. During the annual Phaeocystis spring bloom, the phytoplankton biomass increased by 34-fold, while the microzooplankton biomass showed a 4-fold increase, representing on average about 4.6% of the biomass of their phytoplankton prey. Tag pyrosequencing unveiled an extensive diversity of Gymnodiniaceae, with G. spirale and G. fusiformis representing the most abundant reads. An extended diversity of Phaeocystales, with partial 18S rDNA genes sequence identity as low as 85% was found, with taxa corresponding to P. globosa, but also to unknown Phaeocystaceae. CONCLUSIONS: Morphological analyses and pyrosequencing were generally in accordance with capturing frequency shifts of abundant taxa. Tag pyrosequencing allowed highlighting the maintenance of microplankton diversity during the Phaeocystis bloom and the increase of the taxa presenting low number of reads (minor taxa along with the dominant ones in response to biotic and/or abiotic changing conditions. Although molecular approaches have enhanced our perception on diversity, it has come to light that the

  6. 利用生物信息学方法挑选MCPH1基因标签单核苷酸多态性(tag-SNPs)位点%Bioinformatics methods for selecting the MCHPH1 gene tag single nucleotide polymorphisms(tag-SNPs)sites

    Institute of Scientific and Technical Information of China (English)

    刘跃亮; 曾照芳

    2011-01-01

    目的:为考察汉族人群MCPH1基因与原发性小头畸形病的关系,利用生物信息学方法挑选汉族人群中的MCPH1基因标签单核苷酸多态性(tag-SNPs)位点。方法:利用NCBI数据库,确定MCPHl基因研究范围:利用Hapmap数据库获取汉族人群的MCPHl基因SNPs数据;应用Haploview4.2对MCPH1基因进行连锁不平衡分析;在D′值95%可信区间内构建单倍域(haplotype block):根据单倍域内SNPs之间的r2值和LOD值,挑选标签SNP。结果:在汉族人群中,MCPH1全基因范围内共构建35个单倍域,挑选出122个标签SNPs,确定了各个单倍域内的代表单倍型。结论:汉族人群MCPH1基因的122个SNPs位点是较具代表性的标志性位点,可被选为标签SNPs,并作为汉族人群MCPHl基因与原发性小头畸形病的关联研究。%Objective:Han population for the study MCPH1 primary microcephaly genes and the relationship between disease,the use of bioinformatics methods for selecting the Han population MCPH1 gene tag single nucleotide polymorphisms(tag-SNPs) sites.Methods:Using NCBI databases,to determine the scope of genetic research MCPH1;use Hapmap database for the Han population MCPH1 gene SNPs data;application Haploview4.2 MCPH1 gene on linkage disequilibrium analysis;in the D'value of the 95% confidence interval constructed single-fold domain(haplotype block);based on the single-fold within the r2 values between SNPs and the LOD value,tag SNP selection.Results:In the Han population,MCPH 1 genome were constructed within the domain of 35 haplotypes,selected 122 tag SNPs,identified representatives of the various haplotypes within haplotype.Conclusion:MCPH1 gene in Han population of 122 SNPs loci are more representative of the landmark site,can be selected as tag SNPs,Han population MCPH1 favor of primary microcephaly genes and disease association studies.

  7. Accurate approximation method for prediction of class I MHC affinities for peptides of length 8, 10 and 11 using prediction tools trained on 9mers

    DEFF Research Database (Denmark)

    Lundegaard, Claus; Lund, Ole; Nielsen, Morten

    2008-01-01

    Several accurate prediction systems have been developed for prediction of class I major histocompatibility complex (MHC):peptide binding. Most of these are trained on binding affinity data of primarily 9mer peptides. Here, we show how prediction methods trained on 9mer data can be used for accurate...

  8. A novel method to isolate protein N-terminal peptides from proteome samples using sulfydryl tagging and gold-nanoparticle-based depletion.

    Science.gov (United States)

    Li, Lanting; Wu, Runqing; Yan, Guoquan; Gao, Mingxia; Deng, Chunhui; Zhang, Xiangmin

    2016-01-01

    A novel method to isolate global N-termini using sulfydryl tagging and gold-nanoparticle-based depletion (STagAu method) is presented. The N-terminal and lysine amino groups were first completely dimethylated at the protein level, after which the proteins were digested. The newly generated internal peptides were tagged with sulfydryl by Traut's reagent through digested N-terminal amines in yields of 96%. The resulting sulfydryl peptides were depleted through binding onto nano gold composite materials. The Au-S bond is stable and widely used in materials science. Nano gold composite materials showed nearly complete depletion of sulfydryl peptides. A set of the acetylated and dimethylated N-terminal peptides were analyzed by liquid chromatography-tandem mass spectrometry. This method was demonstrated to be an efficient N-terminus enrichment method because of the use of an effective derivatization reaction, in combination with robust and relative easy to implement Au-S coupling. We identified 632 N-terminal peptides from 386 proteins in a mouse liver sample. The STagAu approach presented is therefore a facile and efficient method for mass-spectrometry-based analysis of proteome N-termini or protease-generated cleavage products.

  9. TagCombine:Recommending Tags to Contents in Software Information Sites

    Institute of Scientific and Technical Information of China (English)

    王新宇; 夏鑫; David Lo

    2015-01-01

    Nowadays, software engineers use a variety of online media to search and become informed of new and interesting technologies, and to learn from and help one another. We refer to these kinds of online media which help software engineers improve their performance in software development, maintenance, and test processes as software information sites. In this paper, we propose TagCombine, an automatic tag recommendation method which analyzes objects in software information sites. TagCombine has three different components: 1) multi-label ranking component which considers tag recommendation as a multi-label learning problem; 2) similarity-based ranking component which recommends tags from similar objects; 3) tag-term based ranking component which considers the relationship between different terms and tags, and recommends tags after analyzing the terms in the objects. We evaluate TagCombine on four software information sites, Ask Different, Ask Ubuntu, Freecode, and Stack Overflow. On averaging across the four projects, TagCombine achieves recall@5 and recall@10 to 0.619 8 and 0.762 5 respectively, which improves TagRec proposed by Al-Kofahi et al. by 14.56% and 10.55% respectively, and the tag recommendation method proposed by Zangerle et al. by 12.08% and 8.16% respectively.

  10. Objective evaluation of methods to track motion from clinical cardiac-gated tagged MRI without the use of a gold standard

    Science.gov (United States)

    Parages, Felipe M.; Denney, Thomas S.; Brankov, Jovan G.

    2015-03-01

    Cardiac-gated MRI is widely used for the task of measuring parameters related to heart motion. More specifically, gated tagged MRI is the preferred modality to estimate local deformation (strain) and rotational motion (twist) of myocardial tissue. Many methods have been proposed to estimate cardiac motion from gated MRI sequences. However, when dealing with clinical data, evaluation of these methods is problematic due to the absence of gold-standards for cardiac motion. To overcome that, a linear regression scheme known as regression-without-truth (RWT) was proposed in the past. RWT uses priors to model the distribution of true values, thus enabling us to assess image-analysis algorithms without knowledge of the ground-truth. Furthermore, it allows one to rank methods by means of an objective figure-of-merit γ (i.e. precision). In this work we apply RWT to compare the performance of several gated MRI motion-tracking methods (e.g. non-rigid registration, feature based, harmonic phase) at the task of estimating myocardial strain and left-ventricle (LV) twist, from a population of 18 clinical human cardiac-gated tagged MRI studies.

  11. A Phosphorylation Tag for Uranyl Mediated Protein Purification and Photo Assisted Tag Removal

    DEFF Research Database (Denmark)

    Zhang, Qiang; Jørgensen, Thomas. J. D.; Nielsen, Peter E;

    2014-01-01

    enables target protein purification from an E. coli extract by immobilized uranyl affinity chromatography. Subsequently, the tag can be efficiently removed by UV-irradiation assisted uranyl photocleavage. We therefore suggest that the divalent uranyl ion (UO22+) may provide a dual function in protein...

  12. Cooperative Tagging Center (CTC)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Cooperative Tagging Center (CTC) began as the Cooperative Game Fish Tagging Program (GTP) at Woods Hole Oceanographic Institute (WHOI) in 1954. The GTP was...

  13. Satellite Tags- Hawaii EEZ

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Satellite tagging was implemented in 2013. Satellite tagging is conducted using a Dan Inject air rifle and deployment arrows designed by Wildlife Computers. Two...

  14. Donor Tag Game

    Science.gov (United States)

    ... Games > Donor Tag Game Printable Version Donor Tag Game This feature requires version 6 or later of ... LGBTQ+ Donors Blood Donor Community Real Stories SleevesUp Games Facebook Avatars and Badges Banners eCards Enter your ...

  15. North Pacific Albacore Tagging

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Conventional tagging data are available from 1971 to 1996. Electronic tagging data are available from 2000 to present. The data are managed by SWFSC in Access...

  16. Gillnet Tag Program

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Certain fishery management programs require vessels to obtain gillnet tags to be used with their gillnet gear. Gillnet tag data is a collection of requests and...

  17. Using Interference to Block RFID Tags

    DEFF Research Database (Denmark)

    Krigslund, Rasmus; Popovski, Petar; Pedersen, Gert Frølund

    We propose a novel method to block RFID tags from responding, using intentional interference. We focus on the experimental evaluation, where we impose interference on the download and uplink, respectively. The results are positive, where modulated CCI shows most effective to block a tag.......We propose a novel method to block RFID tags from responding, using intentional interference. We focus on the experimental evaluation, where we impose interference on the download and uplink, respectively. The results are positive, where modulated CCI shows most effective to block a tag....

  18. Simple method for Shiga toxin 2e purification by affinity chromatography via binding to the divinyl sulfone group.

    Directory of Open Access Journals (Sweden)

    Hideyuki Arimitsu

    Full Text Available Here we describe a simple affinity purification method for Shiga toxin 2e (Stx2e, a major causative factor of edema disease in swine. Escherichia coli strain MV1184 transformed with the expression plasmid pBSK-Stx2e produced Stx2e when cultivated in CAYE broth containing lincomycin. Stx2e bound to commercial D-galactose gel, containing α-D-galactose immobilized on agarose resin via a divinyl sulfone linker, and was eluted with phosphate-buffered saline containing 4.5 M MgCl2. A small amount of Stx2e bound to another commercial α-galactose-immobilized agarose resin, but not to β-galactose-immobilized resin. In addition, Stx2e bound to thiophilic adsorbent resin containing β-mercaptoethanol immobilized on agarose resin via a divinyl sulfone, and was purified in the same manner as from D-galactose gel, but the Stx2e sample contained some contamination. These results indicate that Stx2e bound to D-galactose gel mainly through the divinyl sulfone group on the resin and to a lesser extent through α-D-galactose. With these methods, the yields of Stx2e and attenuated mutant Stx2e (mStx2e from 1 L of culture were approximately 36 mg and 27.7 mg, respectively, and the binding capacity of the D-galactose gel and thiophilic adsorbent resin for Stx2e was at least 20 mg per 1 ml of resin. In addition, using chimeric toxins with prototype Stx2 which did not bind to thiophilic adsorbent resin and some types of mutant Stx2e and Stx2 which contained inserted mutations in the B subunits, we found that, at the least, asparagine (amino acid 17 of the B subunits was associated with Stx2e binding to the divinyl sulfone group. The mStx2e that was isolated exhibited vaccine effects in ICR mice, indicating that these methods are beneficial for large-scale preparation of Stx2e toxoid, which protects swine from edema disease.

  19. ePAT: a simple method to tag adenylated RNA to measure poly(A)-tail length and other 3' RACE applications.

    Science.gov (United States)

    Jänicke, Amrei; Vancuylenberg, John; Boag, Peter R; Traven, Ana; Beilharz, Traude H

    2012-06-01

    The addition of a poly(A)-tail to the 3' termini of RNA molecules influences stability, nuclear export, and efficiency of translation. In the cytoplasm, dynamic changes in the length of the poly(A)-tail have long been recognized as reflective of the switch between translational silence and activation. Thus, measurement of the poly(A)-tail associated with any given mRNA at steady-state can serve as a surrogate readout of its translation-state. Here, we describe a simple new method to 3'-tag adenylated RNA in total RNA samples using the intrinsic property of Escherichia coli DNA polymerase I to extend an RNA primer using a DNA template. This tag can serve as an anchor for cDNA synthesis and subsequent gene-specific PCR to assess poly(A)-tail length. We call this method extension Poly(A) Test (ePAT). The ePAT approach is as efficient as traditional Ligation-Mediated Poly(A) Test (LM-PAT) assays, avoids problems of internal priming associated with oligo-dT-based methods, and allows for the accurate analysis of both the poly(A)-tail length and alternate 3' UTR usage in 3' RACE applications.

  20. Cutaneous skin tag

    Science.gov (United States)

    Skin tag; Acrochordon; Fibroepithelial polyp ... have diabetes. They are thought to occur from skin rubbing against skin. ... The tag sticks out of the skin and may have a short, narrow stalk connecting it to the surface of the skin. Some skin tags are as long as ...

  1. Mobile Application Partition Method Based on Tag in Cloud Environment%云环境下基于标记的移动应用划分方法

    Institute of Scientific and Technical Information of China (English)

    樊新; 高曙

    2015-01-01

    In order to solve the resource -constrained problem to mobile devices and save the energy consumption , a tag-based mobile application partition method was proposed .It is a way to partition the mobile application according to its functional structure in advance and tag the transferable application module .The abundant resources and strong ability of cloud computing technology were employed to process the information .And combining with transfer energy consumption model , it was determined whether the mobile application tag module was transferred to the cloud to remote execution or not .At last, through wireless net-work, the cloud execution results were return so as to extend equipment resources and save the energy consumption .%针对移动设备资源受限及节省其能耗的问题,提出了一种基于标记的移动应用划分方法。该方法根据移动应用功能结构对其进行划分,将可转移执行的应用模块进行标记,利用云计算丰富的资源和强大的信息处理能力,结合转移能耗模型,决定移动应用标记模块是否转移到云端远程执行,通过无线网络,返回云端执行结果,从而达到扩展移动设备资源,节省其能耗的目的。

  2. Comparative Study of Three Methods for Affinity Measurements: Capillary Electrophoresis Coupled with UV Detection and Mass Spectrometry, and Direct Infusion Mass Spectrometry

    Science.gov (United States)

    Mironov, Gleb G.; Logie, Jennifer; Okhonin, Victor; Renaud, Justin B.; Mayer, Paul M.; Berezovski, Maxim V.

    2012-07-01

    We present affinity capillary electrophoresis and mass spectrometry (ACE-MS) as a comprehensive separation technique for label-free solution-based affinity analysis. The application of ACE-MS for measuring affinity constants between eight small molecule drugs [ibuprofen, s-flurbiprofen, diclofenac, phenylbutazone, naproxen, folic acid, resveratrol, and 4,4'-(propane-1,3-diyl) dibenzoic acid] and β-cyclodextrin is described. We couple on-line ACE with MS to combine the separation and kinetic capability of ACE together with the molecular weight and structural elucidation of MS in one system. To understand the full potential of ACE-MS, we compare it with two other methods: Direct infusion mass spectrometry (DIMS) and ACE with UV detection (ACE-UV). After the evaluation, DIMS provides less reliable equilibrium dissociation constants than separation-based ACE-UV and ACE-MS, and cannot be used solely for the study of noncovalent interactions. ACE-MS determines apparent dissociation constants for all reacting small molecules in a mixture, even in cases when drugs overlap with each other during separation. The ability of ACE-MS to interact, separate, and rapidly scan through m/z can facilitate the simultaneous affinity analysis of multiple interacting pairs, potentially leading to the high-throughput screening of drug candidates.

  3. Semantic Analysis of Tag Similarity Measures in Collaborative Tagging Systems

    CERN Document Server

    Cattuto, Ciro; Hotho, Andreas; Stumme, Gerd

    2008-01-01

    Social bookmarking systems allow users to organise collections of resources on the Web in a collaborative fashion. The increasing popularity of these systems as well as first insights into their emergent semantics have made them relevant to disciplines like knowledge extraction and ontology learning. The problem of devising methods to measure the semantic relatedness between tags and characterizing it semantically is still largely open. Here we analyze three measures of tag relatedness: tag co-occurrence, cosine similarity of co-occurrence distributions, and FolkRank, an adaptation of the PageRank algorithm to folksonomies. Each measure is computed on tags from a large-scale dataset crawled from the social bookmarking system del.icio.us. To provide a semantic grounding of our findings, a connection to WordNet (a semantic lexicon for the English language) is established by mapping tags into synonym sets of WordNet, and applying there well-known metrics of semantic similarity. Our results clearly expose differe...

  4. Hierarchical Affinity Propagation

    CERN Document Server

    Givoni, Inmar; Frey, Brendan J

    2012-01-01

    Affinity propagation is an exemplar-based clustering algorithm that finds a set of data-points that best exemplify the data, and associates each datapoint with one exemplar. We extend affinity propagation in a principled way to solve the hierarchical clustering problem, which arises in a variety of domains including biology, sensor networks and decision making in operational research. We derive an inference algorithm that operates by propagating information up and down the hierarchy, and is efficient despite the high-order potentials required for the graphical model formulation. We demonstrate that our method outperforms greedy techniques that cluster one layer at a time. We show that on an artificial dataset designed to mimic the HIV-strain mutation dynamics, our method outperforms related methods. For real HIV sequences, where the ground truth is not available, we show our method achieves better results, in terms of the underlying objective function, and show the results correspond meaningfully to geographi...

  5. Affitins as robust tailored reagents for affinity chromatography purification of antibodies and non-immunoglobulin proteins

    OpenAIRE

    Béhar, Ghislaine; Renodon-Cornière, Axelle; Mouratou, Barbara; Pecorari, Frédéric

    2016-01-01

    International audience; Affinity chromatography is a convenient way of purifying proteins, as a high degree of purity can be reached in one step. The use of tags has greatly contributed to the popularity of this technique. However, the addition of tags may not be desirable or possible for the production of biopharmaceuticals. There is thus a need for tailored artificial affinity ligands. We have developed the use of archaeal extremophilic proteins as scaffolds to generate affinity proteins (A...

  6. Measurement of tag confidence in user generated contents retrieval

    Science.gov (United States)

    Lee, Sihyoung; Min, Hyun-Seok; Lee, Young Bok; Ro, Yong Man

    2009-01-01

    As online image sharing services are becoming popular, the importance of correctly annotated tags is being emphasized for precise search and retrieval. Tags created by user along with user-generated contents (UGC) are often ambiguous due to the fact that some tags are highly subjective and visually unrelated to the image. They cause unwanted results to users when image search engines rely on tags. In this paper, we propose a method of measuring tag confidence so that one can differentiate confidence tags from noisy tags. The proposed tag confidence is measured from visual semantics of the image. To verify the usefulness of the proposed method, experiments were performed with UGC database from social network sites. Experimental results showed that the image retrieval performance with confidence tags was increased.

  7. Highly specific purification of N-glycans using phosphate-based derivatization as an affinity tag in combination with Ti{sup 4+}-SPE enrichment for mass spectrometric analysis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Ying [Shanghai Cancer Center and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032 (China); Key Laboratory of Glycoconjugates Research Ministry of Public Health, Fudan University, Shanghai 200032 (China); Peng, Ye; Bin, Zhichao [Department of Chemistry, Fudan University, Shanghai 200032 (China); Wang, Huijie [Shanghai Cancer Center and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032 (China); Lu, Haojie, E-mail: luhaojie@fudan.edu.cn [Shanghai Cancer Center and Institutes of Biomedical Sciences, Fudan University, Shanghai 200032 (China); Department of Chemistry, Fudan University, Shanghai 200032 (China); Key Laboratory of Glycoconjugates Research Ministry of Public Health, Fudan University, Shanghai 200032 (China)

    2016-08-31

    N-linked protein glycosylation is involved in regulation of a wide variety of cellular processes and associated with numerous diseases. Highly specific identification of N-glycome remains a challenge while its biological significance is acknowledged. The relatively low abundance of glycan in complex biological mixtures, lack of basic sites for protonation, and suppression by other highly abundant proteins/peptides lead to the particularly poor detection sensitivity of N-glycans in the MS analysis. Therefore, the highly specific purification procedure becomes a crucial step prior to MS analysis of the N-glycome. Herein, a novel N-glycans enrichment approach based on phosphate derivatization combined with Ti{sup 4+}-SPE (solid phase extraction) was developed. Briefly, in this strategy, N-glycans were chemically labeled with a phospho-group at their reducing ends, such that the Ti{sup 4+}-SPE microspheres were able to capture the phospho-containing glycans. The enrichment method was developed and optimized using model oligosaccharides (maltoheptaose DP7 and sialylated glycan A1) and also glycans from a standard glycoprotein (asialofetuin, ASF). This method experimentally showed high derivatization efficiency (almost 100%), excellent selectivity (analyzing DP7 in the digests of bovine serum albumin at a mass ratio of 1:100), high enriching recovery (90%), good reproducibility (CV<15%) as well as high sensitivity (LOD at fmol level). At last, the proposed method was successfully applied in the profiling of N-glycome in human serum, in which a total of 31 N-glycan masses were identified. - Graphical abstract: A selective enrichment method for the N-glycome is reported. N-glycans were chemically labeled with a phosphate derivatization reagent (AMS), then the phospho-containing glycans were enriched using Ti{sup 4+}-microspheres. - Highlights: • A highly specific N-glycans purification method based on phosphate derivatization combined with Ti{sup 4+}-SPE was developed

  8. Tag Based Audio Search Engine

    Directory of Open Access Journals (Sweden)

    Parameswaran Vellachu

    2012-03-01

    Full Text Available The volume of the music database is increasing day by day. Getting the required song as per the choice of the listener is a big challenge. Hence, it is really hard to manage this huge quantity, in terms of searching, filtering, through the music database. It is surprising to see that the audio and music industry still rely on very simplistic metadata to describe music files. However, while searching audio resource, an efficient "Tag Based Audio Search Engine" is necessary. The current research focuses on two aspects of the musical databases 1. Tag Based Semantic Annotation Generation using the tag based approach.2. An audio search engine, using which the user can retrieve the songs based on the users choice. The proposed method can be used to annotation and retrieve songs based on musical instruments used , mood of the song, theme of the song, singer, music director, artist, film director, instrument, genre or style and so on.

  9. Site-specific conjugation of an antibody-binding protein catalyzed by horseradish peroxidase creates a multivalent protein conjugate with high affinity to IgG.

    Science.gov (United States)

    Minamihata, Kosuke; Goto, Masahiro; Kamiya, Noriho

    2015-01-01

    Cross-linking proteins offers an approach to enhance the distinct function of proteins due to the multivalent effect. In this study, we demonstrated the preparation of a multivalent antibody-binding protein possessing high affinity to IgG by conjugating a number of antibody-binding proteins using the horseradish peroxidase (HRP)-mediated protein conjugation method. By introducing a peptide tag containing a tyrosine (Y-tag) to the C-terminus of the model protein, a chimera protein of protein G and protein A (pG2 pA), the Tyr residue in the Y-tag was efficiently recognized by HRP and cross-linked with each other to yield a pG2 pA conjugate, composed of mainly two to three units of pG2 pA. The cross-linking occurred site specifically at the Tyr residue in the Y-tag and introduction of the Y-tag showed no effect on the function of pG2 pA. The affinity of the Y-tagged pG2 pA conjugate against IgG clearly increased because of the multivalent effect, demonstrating the benefit of this protein cross-linking reaction, which yields functional protein oligomers. Such multivalent protein conjugates created by this reaction should have potential to be used in ELISA and Western blotting applications in which highly sensitive detection of target molecules is desired.

  10. An AIL/IL-based liquid/liquid extraction system for the purification of His-tagged proteins.

    Science.gov (United States)

    Xu, Weiyuan; Cao, Huazhen; Ren, Guangwei; Xie, Hujun; Huang, Jianying; Li, Shijun

    2014-06-01

    A sorbent based on affinity ionic liquid (AIL), triazacyclononane-ionic liquid, was synthesized, characterized, and applied to the extraction of histidine (His)-tagged proteins from aqueous buffer to ionic liquid (IL) phase. The adsorbed His-tagged proteins could be back-extracted from the IL phase to the aqueous buffer with an imidazole solution. The specific binding of His-tagged proteins with AIL/IL could be affected by a few factors including the ionic strength and coordinated metal ions. In the case of His-tagged enhanced green fluorescent protein (EGFP), the maximum binding capacity of Cu(2+)-AIL/IL reached 2.58 μg/μmol under the optimized adsorption conditions. The eluted His-tagged EGFP kept fluorescent and remained active through the purification process. Moreover, a tandem extraction process successively using Cu(2+)-AIL/IL and Zn(2+)-AIL/IL systems was developed, which was proven very efficient to obtain the ultimate protein with a purity of about 90 %. An effective reclamation method for the AIL/IL extraction system was further established. The sorbent could be easily regenerated by removing metal ions with EDTA and the followed reimmobilization of metal ions. Easy handling of the presented M(2+)-AIL/IL system and highly specific ability to absorb His-tagged proteins make it attractive and potentially applicable in biomolecular separation.

  11. One-step purification of twin-strep-tagged proteins and their complexes on strep-tactin resin cross-linked with bis(sulfosuccinimidyl) suberate (BS3).

    Science.gov (United States)

    Ivanov, Konstantin I; Bašić, Marta; Varjosalo, Markku; Mäkinen, Kristiina

    2014-04-20

    Affinity purification of Strep-tagged fusion proteins on resins carrying an engineered streptavidin (Strep-Tactin) has become a widely used method for isolation of protein complexes under physiological conditions. Fusion proteins containing two copies of Strep-tag II, designated twin-Strep-tag or SIII-tag, have the advantage of higher affinity for Strep-Tactin compared to those containing only a single Strep-tag, thus allowing more efficient protein purification. However, this advantage is offset by the fact that elution of twin-Strep-tagged proteins with biotin may be incomplete, leading to low protein recovery. The recovery can be dramatically improved by using denaturing elution with sodium dodecyl sulfate (SDS), but this leads to sample contamination with Strep-Tactin released from the resin, making the assay incompatible with downstream proteomic analysis. To overcome this limitation, we have developed a method whereby resin-coupled tetramer of Strep-Tactin is first stabilized by covalent cross-linking with Bis(sulfosuccinimidyl) suberate (BS3) and the resulting cross-linked resin is then used to purify target protein complexes in a single batch purification step. Efficient elution with SDS ensures good protein recovery, while the absence of contaminating Strep-Tactin allows downstream protein analysis by mass spectrometry. As a proof of concept, we describe here a protocol for purification of SIII-tagged viral protein VPg-Pro from nuclei of virus-infected N. benthamiana plants using the Strep-Tactin polymethacrylate resin cross-linked with BS3. The same protocol can be used to purify any twin-Strep-tagged protein of interest and characterize its physiological binding partners.

  12. Evaluation of a method using survey counts and tag data to estimate the number of Pacific walruses (Odobenus rosmarus divergens) using a coastal haulout in northwestern Alaska

    Science.gov (United States)

    Battaile, Brian; Jay, Chadwick V.; Udevitz, Mark S.; Fischbach, Anthony S.

    2017-01-01

    Increased periods of sparse sea ice over the continental shelf of the Chukchi Sea in late summer have reduced offshore haulout habitat for Pacific walruses (Odobenus rosmarus divergens) and increased opportunities for human activities in the region. Knowing how many walruses could be affected by human activities would be useful to conservation decisions. Currently, there are no adequate estimates of walrus abundance in the northeastern Chukchi Sea during summer–early autumn. Estimating abundance in autumn might be possible from coastal surveys of hauled out walruses during periods when offshore sea ice is unavailable to walruses. We evaluated methods to estimate the size of the walrus population that was using a haulout on the coast of northwestern Alaska in autumn by using aerial photography to count the number of hauled out walruses (herd size) and data from 37 tagged walruses to estimate availability (proportion of population hauled out). We used two methods to estimate availability, direct proportions of hauled out tagged walruses and smoothed proportions using local polynomial regression. Point estimates of herd size (4200–38,000 walruses) and total population size (76,000–287,000 walruses) ranged widely among days and between the two methods of estimating availability. Estimates of population size were influenced most by variation in estimates of availability. Coastal surveys might be improved most by counting walruses when the greatest numbers are hauled out, thereby reducing the influence of availability on population size estimates. The chance of collecting data during peak haulout periods would be improved by conducting multiple surveys.

  13. Multilabel Learning for Automatic Web Services Tagging

    Directory of Open Access Journals (Sweden)

    Mustapha AZNAG

    2014-08-01

    Full Text Available Recently, some web services portals and search engines as Biocatalogue and Seekda!, have allowed users to manually annotate Web services using tags. User Tags provide meaningful descriptions of services and allow users to index and organize their contents. Tagging technique is widely used to annotate objects in Web 2.0 applications. In this paper we propose a novel probabilistic topic model (which extends the CorrLDA model - Correspondence Latent Dirichlet Allocation- to automatically tag web services according to existing manual tags. Our probabilistic topic model is a latent variable model that exploits local correlation labels. Indeed, exploiting label correlations is a challenging and crucial problem especially in multi-label learning context. Moreover, several existing systems can recommend tags for web services based on existing manual tags. In most cases, the manual tags have better quality. We also develop three strategies to automatically recommend the best tags for web services. We also propose, in this paper, WS-Portal; An Enriched Web Services Search Engine which contains 7063 providers, 115 sub-classes of category and 22236 web services crawled from the Internet. In WS-Portal, severals technologies are employed to improve the effectiveness of web service discovery (i.e. web services clustering, tags recommendation, services rating and monitoring. Our experiments are performed out based on real-world web services. The comparisons of Precision@n, Normalised Discounted Cumulative Gain (NDCGn values for our approach indicate that the method presented in this paper outperforms the method based on the CorrLDA in terms of ranking and quality of generated tags.

  14. Production of biologically active IgG hinge-tag soluble epidermal growth factor receptors (ErbB).

    Science.gov (United States)

    Otani, Takayuki; Hashizume, Toshihiro; Nagaoka, Tadahiro; Fukuda, Tomoko; Tang, Careen K; Salomon, David S; Seno, Masaharu

    2010-03-01

    The extracellular domains (ECD) of epidermal growth factor receptors, ErbB1, 2, 3 and 4, were designed as soluble dimeric forms. Each ECD was fused to a short hinge region derived from IgG, such that the stable dimer could be formed with disulfide bridges. This hinge-tagged design minimized the molecular weight to approximately 50% of the conventional Fc-fusion design without an Fc domain of IgG. The refolded dimers could be easily analyzed and characterized by SDS-PAGE. Hinge-tagged soluble ErbBs demonstrated significant affinity for betacellulin and heregulin. The IgG hinge-tag should be a simple method to design soluble dimers that would be useful for high throughput screening of ligands, antagonists or derivatives.

  15. Realization of Fractal Affine Transformation

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    This paper gives the definition of fractal affine transformation and presents a specific method for its realization and its cor responding mathematical equations which are essential in fractal image construction.

  16. Study on method of solving ambiguity in Tibetan part of speech tagging%藏文词性自动标注中歧义问题处理方法研究

    Institute of Scientific and Technical Information of China (English)

    羊毛卓玛

    2013-01-01

    Tibetan language Part-Of-Speech(POS)tagging is the subsequent parsing of Tibetan language information processing. POS tagging is an essential foundation work for semantic analysis and text analysis. POS ambiguity problem solving is the key to Tibetan POS tagging, is also one of the difficulties in the Tibetan automatic POS tagging. This paper analyzes and studies POS ambiguity problem in the Tibetan POS tagging, and puts forward a method of solving POS ambiguity problem suitable for Tibetan grammar rules. Experiments prove that this method on speech disambiguation in the Tibetan POS tagging has achieved better results and has definitely increased the accuracy of the Tibetan POS tagging.%藏文词性自动标注是藏文信息处理后续句法分析、语义分析及篇章分析必不可少的基础工作。词性歧义问题的处理是藏文词性自动标注的关键所在,也是藏文信息处理的难点问题。对藏文词性标注中词性歧义问题进行了分析研究,提出了符合藏文语法规则实用于藏文词性标注的解决词性排岐方法。实验证明:该处理方法在藏文词性自动标注中对词性排岐方面有较好的效果,使藏文词性标注正确率有了一定的提高。

  17. Comparing the hierarchy of author given tags and repository given tags in a large document archive

    CERN Document Server

    Tibély, Gergely; Palla, Gergely

    2015-01-01

    Folksonomies - large databases arising from collaborative tagging of items by independent users - are becoming an increasingly important way of categorizing information. In these systems users can tag items with free words, resulting in a tripartite item-tag-user network. Although there are no prescribed relations between tags, the way users think about the different categories presumably has some built in hierarchy, in which more special concepts are descendants of some more general categories. Several applications would benefit from the knowledge of this hierarchy. Here we apply a recent method to check the differences and similarities of hierarchies resulting from tags given by independent individuals and from tags given by a centrally managed repository system. The results from out method showed substantial differences between the lower part of the hierarchies, and in contrast, a relatively high similarity at the top of the hierarchies.

  18. Prediction of binding affinity and efficacy of thyroid hormone receptor ligands using QSAR and structure-based modeling methods

    Energy Technology Data Exchange (ETDEWEB)

    Politi, Regina [Laboratory for Molecular Modeling, Division of Chemical Biology and Medicinal Chemistry, University of North Carolina, Chapel Hill, NC 27599 (United States); Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC 27599 (United States); Rusyn, Ivan, E-mail: iir@unc.edu [Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC 27599 (United States); Tropsha, Alexander, E-mail: alex_tropsha@unc.edu [Laboratory for Molecular Modeling, Division of Chemical Biology and Medicinal Chemistry, University of North Carolina, Chapel Hill, NC 27599 (United States)

    2014-10-01

    The thyroid hormone receptor (THR) is an important member of the nuclear receptor family that can be activated by endocrine disrupting chemicals (EDC). Quantitative Structure–Activity Relationship (QSAR) models have been developed to facilitate the prioritization of THR-mediated EDC for the experimental validation. The largest database of binding affinities available at the time of the study for ligand binding domain (LBD) of THRβ was assembled to generate both continuous and classification QSAR models with an external accuracy of R{sup 2} = 0.55 and CCR = 0.76, respectively. In addition, for the first time a QSAR model was developed to predict binding affinities of antagonists inhibiting the interaction of coactivators with the AF-2 domain of THRβ (R{sup 2} = 0.70). Furthermore, molecular docking studies were performed for a set of THRβ ligands (57 agonists and 15 antagonists of LBD, 210 antagonists of the AF-2 domain, supplemented by putative decoys/non-binders) using several THRβ structures retrieved from the Protein Data Bank. We found that two agonist-bound THRβ conformations could effectively discriminate their corresponding ligands from presumed non-binders. Moreover, one of the agonist conformations could discriminate agonists from antagonists. Finally, we have conducted virtual screening of a chemical library compiled by the EPA as part of the Tox21 program to identify potential THRβ-mediated EDCs using both QSAR models and docking. We concluded that the library is unlikely to have any EDC that would bind to the THRβ. Models developed in this study can be employed either to identify environmental chemicals interacting with the THR or, conversely, to eliminate the THR-mediated mechanism of action for chemicals of concern. - Highlights: • This is the largest curated dataset for ligand binding domain (LBD) of the THRβ. • We report the first QSAR model for antagonists of AF-2 domain of THRβ. • A combination of QSAR and docking enables

  19. Identification of protein interacting partners using tandem affinity purification.

    Science.gov (United States)

    Bailey, Dalan; Urena, Luis; Thorne, Lucy; Goodfellow, Ian

    2012-02-25

    A critical and often limiting step in understanding the function of host and viral proteins is the identification of interacting cellular or viral protein partners. There are many approaches that allow the identification of interacting partners, including the yeast two hybrid system, as well as pull down assays using recombinant proteins and immunoprecipitation of endogenous proteins followed by mass spectrometry identification(1). Recent studies have highlighted the utility of double-affinity tag mediated purification, coupled with two specific elution steps in the identification of interacting proteins. This approach, termed Tandem Affinity Purification (TAP), was initially used in yeast(2,3) but more recently has been adapted to use in mammalian cells(4-8). As proof-of-concept we have established a tandem affinity purification (TAP) method using the well-characterized eukaryotic translation initiation factor eIF4E(9,10).The cellular translation factor eIF4E is a critical component of the cellular eIF4F complex involved in cap-dependent translation initiation(10). The TAP tag used in the current study is composed of two Protein G units and a streptavidin binding peptide separated by a Tobacco Etch Virus (TEV) protease cleavage sequence. The TAP tag used in the current study is composed of two Protein G units and a streptavidin binding peptide separated by a Tobacco Etch Virus (TEV) protease cleavage sequence(8). To forgo the need for the generation of clonal cell lines, we developed a rapid system that relies on the expression of the TAP-tagged bait protein from an episomally maintained plasmid based on pMEP4 (Invitrogen). Expression of tagged murine eIF4E from this plasmid was controlled using the cadmium chloride inducible metallothionein promoter. Lysis of the expressing cells and subsequent affinity purification via binding to rabbit IgG agarose, TEV protease cleavage, binding to streptavidin linked agarose and subsequent biotin elution identified numerous

  20. Study on Method of Purification of GST-tagged Proteins from Inclusion Bodies%GST融合蛋白包涵体纯化方法的探索

    Institute of Scientific and Technical Information of China (English)

    季林; 刘佩娟; 王毅; 赵勇; 毛峰峰; 赵善民; 尚淑琴; 师长宏

    2012-01-01

    The aim was to study the method of purification of GST-tagged proteins from inclusion bodies. The vector PGEX-4T1-Rpfd expressed Rpfd -GST fusion protein was transformed into E. Coli DH5a and protein expression was induced with IPTG. The effect of these factors is observed on the impact of protein purification by comparing lysis with guanidine hydrochloride and pretreatment with Novagen Protein Refolding Kit, and adjusting the different types of protein refolding solution. The active GST fusion protein can be efficiently purified, through removing soluble protein after ultrasonic treatment, and then using the reaction mixture containing 20 mmol Tris-HCl and 0.1 mmol DTT for rehabilitation. To remove soluble protein expression and change the formula of protein refolding solutions can, effectively improve protein purification of GST-tagged proteins from inclusion bodies.%探讨GST融合蛋白包涵体纯化的方法.将表达Rpfd与GST融合蛋白的质粒PGEX-4T1 -Rpfd转入大肠杆菌DH5a,IPTG诱导表达目的蛋白.通过比较盐酸胍裂解和Novagen蛋白折叠试剂盒预处理,及调整不同类型蛋白复性液,观察上述因素对GST融合蛋白包涵体纯化效果的影响.结果:超声后先去除可溶性表达,再使用20 mmolris-HC1和0.1 mmol DTT进行复性,可获得高效纯化,具有活性的GST融合蛋白.对于GST包涵体蛋白进行纯化时,去除可溶性表达蛋白,改变蛋白复性液的成分,可有效提高蛋白纯化效果.

  1. Endotoxin depletion of recombinant protein preparations through their preferential binding to histidine tags.

    Science.gov (United States)

    Mack, Laura; Brill, Boris; Delis, Natalia; Groner, Bernd

    2014-12-01

    The presence of endotoxins in preparations of recombinantly produced therapeutic proteins poses serious problems for patients. Endotoxins can cause fever, respiratory distress syndromes, intravascular coagulation, or endotoxic shock. A number of methods have been devised to remove endotoxins from protein preparations using separation procedures based on molecular mass or charge properties. Most of the methods are limited in their endotoxin removal capacities and lack general applicability. We are describing a biotechnological approach for endotoxin removal. This strategy exploits the observation that endotoxins form micelles that expose negative charges on their surface, leading to preferential binding of endotoxins to cationic surfaces, allowing the separation from their resident protein. Endotoxins exhibit high affinity to stretches of histidines, which are widely used tools to facilitate the purification of recombinant proteins. They bind to nickel ions and are the basis for protein purification from cellular extracts by immobilized metal affinity chromatography. We show that the thrombin-mediated cleavage of two histidine tags from the purified recombinant protein and the adsorption of these histidine tags and their associated endotoxins to a nickel affinity column result in an appreciable depletion of the endotoxins in the purified protein fraction.

  2. Influence of material properties upon immobilization of histidine-tagged protein on Ni-Co coated chip.

    Science.gov (United States)

    Chang, Yaw-Jen; Ho, Ching-Yuan; Chang, Cheng-Hao

    2014-04-01

    In protein research, protein microarray facilitates high-throughput study of protein abundance and function. An appropriate microarray surface that can be used to immobilize protein samples is a prerequisite for the investigation of molecular interactions. Ni-Co alloy coated protein microarray chip has been found to adsorb histidine-tagged proteins effectively based on the method of immobilized metal affinity chromatography. Due to the ingredient of bi-metallic elements, different electroplating conditions resulted in distinct binding affinities. Therefore, the influence of Ni-Co material properties on the immobilization of histidine-tagged protein was systematically investigated in this study. In the experiments, the contact angle measurement suggested that no strong relationship can be established between the wettability of chip surface and its corresponding protein immobilization. ESCA test demonstrated that the major ingredients of the Ni-Co alloy coated protein microarray chip were Ni and Co. In addition, the XRD test concluded that a Ni-Co protein chip that consists mostly of hcp lattice has better binding capability. SEM micrographs provide direct image evidence. These material tests summarize that the Ni-Co alloy coated protein microarray chip adsorbs His-tagged proteins through its surface morphology. Therefore, it can provide specific binding due to the affinity adsorption between the intermediate metals and the protein.

  3. Genetic tagging of humpback whales.

    Science.gov (United States)

    Palsbøll, P J; Allen, J; Bérubé, M; Clapham, P J; Feddersen, T P; Hammond, P S; Hudson, R R; Jørgensen, H; Katona, S; Larsen, A H; Larsen, F; Lien, J; Mattila, D K; Sigurjónsson, J; Sears, R; Smith, T; Sponer, R; Stevick, P; Oien, N

    1997-08-21

    The ability to recognize individual animals has substantially increased our knowledge of the biology and behaviour of many taxa. However, not all species lend themselves to this approach, either because of insufficient phenotypic variation or because tag attachment is not feasible. The use of genetic markers ('tags') represents a viable alternative to traditional methods of individual recognition, as they are permanent and exist in all individuals. We tested the use of genetic markers as the primary means of identifying individuals in a study of humpback whales in the North Atlantic Ocean. Analysis of six microsatellite loci among 3,060 skin samples collected throughout this ocean allowed the unequivocal identification of individuals. Analysis of 692 'recaptures', identified by their genotype, revealed individual local and migratory movements of up to 10,000 km, limited exchange among summer feeding grounds, and mixing in winter breeding areas, and also allowed the first estimates of animal abundance based solely on genotypic data. Our study demonstrates that genetic tagging is not only feasible, but generates data (for example, on sex) that can be valuable when interpreting the results of tagging experiments.

  4. 基于标签和因子分析的协同推荐方法%Collaborative Recommendation Method Based on Tags and Factor Analysis

    Institute of Scientific and Technical Information of China (English)

    蔡国永; 吕瑞; 樊永显

    2015-01-01

    根据在线社区中群体的历史行为进行物品(或信息)推荐是当前研究热点之一,传统推荐算法都面临数据稀疏性问题的挑战。针对传统推荐算法知识表示的局限性进行了研究,提出了一种基于标签系统的用户行为知识表示法,把用户在物品上历史行为的统计,转化为对用户在物品标签上的统计,从而缓解数据稀疏的情况。为了降低标签维度过高导致的计算复杂性问题,提出了采用因子分析法,抽取出潜在重要且稳定的特征因子向量来最终表示用户的历史行为,并据此度量用户行为在特征因子向量上的相似性。最后采用协同过滤的思想给出了一种新的协同推荐方法。通过在真实数据集上的大量对比实验,表明该方法在处理具有稀疏性的数据集时,总是能保持更高且更稳定的推荐准确率。%Item (or information) recommendation is one of hot research topics currently. However the is-sue of sparseness in dataset challenges all traditional recommendation algorithms. Limitations of knowl-edge representation in traditional recommendation algorithms were studied. The tag-system-based knowl-edge to represent information of each user’s behavior was proposed. That it the account on user’s behav-ior on items is transferred to an account on a user’s behavior on tags. To decrease the computation com-plexity on high dimensional tag-based datasets, a factor analysis method was taken to extract those most important latent factors to represent users’ behaviors. Based on each user’s representing vector of latent factors, a new way was given to compute similarities among users. By incorporating this similarity meas-ure, a new collaborative recommendation method with low sensitivity to sparseness was built to meet the need of practical and dynamic datasets. Experiments were carried on real-world datasets to compare the proposed method with other state

  5. A novel thiol-affinity micropipette tip method using zinc(II)-cyclen-attached agarose beads for enrichment of cysteine-containing molecules.

    Science.gov (United States)

    Kusamoto, Hiroshi; Shiba, Akio; Koretake, Norinao; Fujioka, Haruto; Hieda, Yuhzo; Kinoshita-Kikuta, Emiko; Kinoshita, Eiji; Koike, Tohru

    2016-09-15

    Cysteine-containing biomolecules are attractive targets in the study of thiol biology. Here we introduce a novel method for the selective enrichment of thiol-containing molecules using a thiol-capture zinc(II) complex of 1,4,7,10-tetraazacyclododecane (Zn(2+)-cyclen). Recognition of N-acetylcysteine amide by Zn(2+)-cyclen has been studied by potentiometric pH titration, revealing formation of a 1:1 thiolate-bound Zn(2+)-cyclen complex with a large thiolate-affinity constant of 10(6.2)M(-1) at 25°C and I=0.10M (NaCl). The Zn(2+)-bound thiolate anion is unexpectedly stable in aqueous solution at pH 7.8 under atmospheric conditions for a few days. These findings have contributed to the development of a convenient method for separation of thiol compounds by using a micropipette tip. A 200μL micropipette tip containing 10μL of hydrophilic cross-linked agarose beads attached to Zn(2+)-cyclen moieties was prepared. All steps for thiol-affinity separation (binding, washing, and eluting) are conducted using aqueous buffers at room temperature. The entire separation protocol requires less than 15min per sample. We demonstrate practical example separations of cysteine-containing molecules. This micropipette tip method would be used preferentially as an alternative to existing tools for reliable enrichment of thiol-containing molecules.

  6. Electrochemical immobilization of Fluorescent labelled probe molecules on a FTO surface for affinity detection based on photo-excited current

    Energy Technology Data Exchange (ETDEWEB)

    Haruyama, Tetsuya; Wakabayashi, Ryo; Cho, Takeshi; Matsuyama, Sho-taro, E-mail: haruyama@life.kyutech.as.jp [Kyushu Institute of Technology, Department of Biological Functions and Engineering, Kitakyushu Science and Research Park, Hibikino, Kitakyushu, Fukuoka 808-0196 (Japan)

    2011-10-29

    Photo-excited current can be generated at a molecular interface between a photo-excited molecules and a semi-conductive material in appropriate condition. The system has been recognized for promoting photo-energy devices such as an organic dye sensitized solar-cell. The photo-current generated reactions are totally dependent on the interfacial energy reactions, which are in a highly fluctuated interfacial environment. The authors investigated the photo-excited current reaction to develop a smart affinity detection method. However, in order to perform both an affinity reaction and a photo-excited current reaction at a molecular interface, ordered fabrications of the functional (affinity, photo-excitation, etc.) molecules layer on a semi-conductive surface is required. In the present research, we would like to present the fabrication and functional performance of photo-excited current-based affinity assay device and its application for detection of endocrine disrupting chemicals. On the FTO surface, fluorescent pigment labelled affinity peptide was immobilized through the EC tag (electrochemical-tag) method. The modified FTO produced a current when it was irradiated with diode laser light. However, the photo current decreased drastically when estrogen (ES) coexisted in the reaction solution. In this case, immobilized affinity probe molecules formed a complex with ES and estrogen receptor (ER). The result strongly suggests that the photo-excited current transduction between probe molecule-labelled cyanine pigment and the FTO surface was partly inhibited by a complex that formed at the affinity oligo-peptide region in a probe molecule on the FTO electrode. The bound bulky complex may act as an impediment to perform smooth transduction of photo-excited current in the molecular interface. The present system is new type of photo-reaction-based analysis. This system can be used to perform simple high-sensitive homogeneous assays.

  7. Sensitive and label-free biosensing of RNA with predicted secondary structures by a triplex affinity capture method

    Science.gov (United States)

    Carrascosa, Laura G.; Gómez-Montes, S.; Aviñó, A.; Nadal, A.; Pla, M.; Eritja, R.; Lechuga, L. M.

    2012-01-01

    A novel biosensing approach for the label-free detection of nucleic acid sequences of short and large lengths has been implemented, with special emphasis on targeting RNA sequences with secondary structures. The approach is based on selecting 8-aminoadenine-modified parallel-stranded DNA tail-clamps as affinity bioreceptors. These receptors have the ability of creating a stable triplex-stranded helix at neutral pH upon hybridization with the nucleic acid target. A surface plasmon resonance biosensor has been used for the detection. With this strategy, we have detected short DNA sequences (32-mer) and purified RNA (103-mer) at the femtomol level in a few minutes in an easy and level-free way. This approach is particularly suitable for the detection of RNA molecules with predicted secondary structures, reaching a limit of detection of 50 fmol without any label or amplification steps. Our methodology has shown a marked enhancement for the detection (18% for short DNA and 54% for RNA), when compared with the conventional duplex approach, highlighting the large difficulty of the duplex approach to detect nucleic acid sequences, especially those exhibiting stable secondary structures. We believe that our strategy could be of great interest to the RNA field. PMID:22241768

  8. Hydroxyl Tagging Velocimetry for Rocket Plumes Project

    Data.gov (United States)

    National Aeronautics and Space Administration — A non-intrusive method for measuring velocities in a rocket exhaust is proposed in a joint effort by MetroLaser and Vanderbilt University. Hydroxyl Tagging...

  9. Tandem Affinity Purification Approach Coupled to Mass Spectrometry to Identify Post-translational Modifications of Histones Associated with Chromatin-Binding Proteins.

    Science.gov (United States)

    Beyer, Sophie; Robin, Philippe; Ait-Si-Ali, Slimane

    2017-01-01

    Protein purification by tandem affinity purification (TAP)-tag coupled to mass spectrometry analysis is usually used to reveal protein complex composition. Here we describe a TAP-tag purification of chromatin-bound proteins along with associated nucleosomes, which allow exhaustive identification of protein partners. Moreover, this method allows exhaustive identification of the post-translational modifications (PTMs) of the associated histones. Thus, in addition to partner characterization, this approach reveals the associated epigenetic landscape that can shed light on the function and properties of the studied chromatin-bound protein.

  10. Engineering a novel multifunctional green fluorescent protein tag for a wide variety of protein research.

    Directory of Open Access Journals (Sweden)

    Takuya Kobayashi

    Full Text Available BACKGROUND: Genetically encoded tag is a powerful tool for protein research. Various kinds of tags have been developed: fluorescent proteins for live-cell imaging, affinity tags for protein isolation, and epitope tags for immunological detections. One of the major problems concerning the protein tagging is that many constructs with different tags have to be made for different applications, which is time- and resource-consuming. METHODOLOGY/PRINCIPAL FINDINGS: Here we report a novel multifunctional green fluorescent protein (mfGFP tag which was engineered by inserting multiple peptide tags, i.e., octa-histidine (8xHis, streptavidin-binding peptide (SBP, and c-Myc tag, in tandem into a loop of GFP. When fused to various proteins, mfGFP monitored their localization in living cells. Streptavidin agarose column chromatography with the SBP tag successfully isolated the protein complexes in a native form with a high purity. Tandem affinity purification (TAP with 8xHis and SBP tags in mfGFP further purified the protein complexes. mfGFP was clearly detected by c-Myc-specific antibody both in immunofluorescence and immuno-electron microscopy (EM. These findings indicate that mfGFP works well as a multifunctional tag in mammalian cells. The tag insertion was also successful in other fluorescent protein, mCherry. CONCLUSIONS AND SIGNIFICANCE: The multifunctional fluorescent protein tag is a useful tool for a wide variety of protein research, and may have the advantage over other multiple tag systems in its higher expandability and compatibility with existing and future tag technologies.

  11. Ontologies and tag-statistics

    CERN Document Server

    Tibely, Gergely; Vicsek, Tamas; Palla, Gergely

    2012-01-01

    Due to the increasing popularity of collaborative tagging systems, the research on tagged networks, hypergraphs, ontologies, folksonomies and other related concepts is becoming an important interdisciplinary topic with great actuality and relevance for practical applications. In most collaborative tagging systems the tagging by the users is completely "flat", while in some cases they are allowed to define a shallow hierarchy for their own tags. However, usually no overall hierarchical organisation of the tags is given, and one of the interesting challenges of this area is to provide an algorithm generating the ontology of the tags from the available data. In contrast, there are also other type of tagged networks available for research, where the tags are already organised into a directed acyclic graph (DAG), encapsulating the "is a sub-category of" type of hierarchy between each other. In this paper we study how this DAG affects the statistical distribution of tags on the nodes marked by the tags in various r...

  12. Tagged Vector Contour (TVC)

    Data.gov (United States)

    Kansas Data Access and Support Center — The Kansas Tagged Vector Contour (TVC) dataset consists of digitized contours from the 7.5 minute topographic quadrangle maps. Coverage for the state is incomplete....

  13. TAG Advertisement Hardware

    Science.gov (United States)

    1994-01-01

    LaRc SI Material Overall photograph showing the material specimens, the graphite composite, the gold composite and the molded gears on a black background. These photos were used for the TAG CO-OP Public Relations and promotions

  14. Efficient biotinylation and single-step purification of tagged transcription factors in mammalian cells and transgenic mice

    Science.gov (United States)

    de Boer, Ernie; Rodriguez, Patrick; Bonte, Edgar; Krijgsveld, Jeroen; Katsantoni, Eleni; Heck, Albert; Grosveld, Frank; Strouboulis, John

    2003-06-01

    Proteomic approaches require simple and efficient protein purification methodologies that are amenable to high throughput. Biotinylation is an attractive approach for protein complex purification due to the very high affinity of avidin/streptavidin for biotinylated templates. Here, we describe an approach for the single-step purification of transcription factor complex(es) based on specific in vivo biotinylation. We expressed the bacterial BirA biotin ligase in mammalian cells and demonstrated very efficient biotinylation of a hematopoietic transcription factor bearing a small (23-aa) artificial peptide tag. Biotinylation of the tagged transcription factor altered neither the factor's protein interactions or DNA binding properties in vivo nor its subnuclear distribution. Using this approach, we isolated the biotin-tagged transcription factor and at least one other known interacting protein from crude nuclear extracts by direct binding to streptavidin beads. Finally, this method works efficiently in transgenic mice, thus raising the prospect of using biotinylation tagging in protein complex purification directly from animal tissues. Therefore, BirA-mediated biotinylation of tagged proteins provides the basis for the single-step purification of proteins from mammalian cells.

  15. New tags for recombinant protein detection and O-glycosylation reporters.

    Directory of Open Access Journals (Sweden)

    Gianluca Petris

    Full Text Available Monoclonal antibodies (mAbs, because of their unique specificity, are irreplaceable tools for scientific research. Precise mapping of the antigenic determinants allows the development of epitope tagging approaches to be used with recombinant proteins for several purposes. Here we describe a new family of tags derived from the epitope recognized by a single highly specific mAb (anti-roTag mAb, which was obtained from a pool of mAbs reacting with the rotavirus nonstructural protein 5 (NSP5. The variable regions of the anti-roTag mAb were identified and their binding capacity verified upon expression as a single-chain/miniAb. The minimal epitope, termed roTag, was identified as a 10 amino acid sequence (SISSSIFKNE. The affinity of the anti-roTag/roTag interaction was found to be comparable to that of the anti-SV5/SV5 tag interaction. roTag was successfully used for detection of several recombinant cytosolic, secretory and membrane proteins. Two additional variants of roTag of 10 and 13 amino acids containing O-glycosylation susceptible sites (termed OG-tag and roTagO were constructed and characterised. These tags were useful to detect proteins passing through the Golgi apparatus, the site of O-glycosylation.

  16. A Tag Ranking Method Based on HITS and Random Walk%一种基于HITS和随机跳转的网页标签排序方法

    Institute of Scientific and Technical Information of China (English)

    汪兆鹏; 胡侠; 倪宁; 王灿

    2011-01-01

    Web 2.0应用的兴起,推进了情报学科由"文献组织"向"知识组织"演化.网页标签作为重要的Web 2 0应用之一,已经成为大众组织知识的常用途径.然而,现有的标签排序方法难以有效满足知识组织的需求.本文在三核协同标签模型的基础上,充分考虑标签和用户、标签和标签、标签和文档之间的关系,提出了一种结合HITS和随机跳转的标签排序方法.该方法利用高质量标签和高质量用户之间的相互加强关系,根据标签之间的相似性来找出高质量相关标签,有效提高标签排序的质量.在Delicious数据集上的实验结果表明,该方法能较大提高标签排序的准确度.%With the rise of Web 2. 0 applications, a new trend in information science, namely the evolution from "organizing document" to "organizing knowledge" , is looming on the horizon. One of' important Web 2. 0 applications,social tag, is making this trend a reality by adding meaningful annotations to Web pages. However, existing tag ranking methods are not efficient in knowledge organization. To improve tag ranking performance , this paper proposes a new ranking algorithm by utilizing relationships among users, tags and Web documents in a tripartite collaborative tagging model. By combining HITS and random walk, we effectively exploit the mutual reinforcement between quality users and quality tags and retrieve related tags by measuring similarity between tags. Experimental results on Delicious dataset demonstrate the effectiveness of our algorithm.

  17. APPECT: An Approximate Backbone-Based Clustering Algorithm for Tags

    DEFF Research Database (Denmark)

    Zong, Yu; Xu, Guandong; Jin, Pin

    2011-01-01

    algorithm for Tags (APPECT). The main steps of APPECT are: (1) we execute the K-means algorithm on a tag similarity matrix for M times and collect a set of tag clustering results Z={C1,C2,…,Cm}; (2) we form the approximate backbone of Z by executing a greedy search; (3) we fix the approximate backbone...... resulting from the severe difficulty of ambiguity, redundancy and less semantic nature of tags. Clustering method is a useful tool to address the aforementioned difficulties. Most of the researches on tag clustering are directly using traditional clustering algorithms such as K-means or Hierarchical...

  18. Facets: Ersatz, Resource and Tag

    Science.gov (United States)

    Frické, Martin H.

    2013-01-01

    Introduction: Faceted classification appears to be of utmost importance. Ersatz facets, resource faceting and tag faceting: The distinctions are drawn between facets and ersatz facets, and between faceted resources and faceted tags. Single tag resource faceting and multiple tag information object faceting: The basic features are explored of single…

  19. Sensitive and rapid method for amino acid quantitation in malaria biological samples using AccQ.Tag ultra performance liquid chromatography-electrospray ionization-MS/MS with multiple reaction monitoring.

    Science.gov (United States)

    Armenta, Jenny M; Cortes, Diego F; Pisciotta, John M; Shuman, Joel L; Blakeslee, Kenneth; Rasoloson, Dominique; Ogunbiyi, Oluwatosin; Sullivan, David J; Shulaev, Vladimir

    2010-01-15

    An AccQ*Tag ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (AccQ*Tag-UPLC-ESI-MS/MS) method for fast, reproducible, and sensitive amino acid quantitation in biological samples, particularly, the malaria parasite Plasmodium falciparum is presented. The Waters Acquity TQD UPLC/MS system equipped with a photodiode array (PDA) detector was used for amino acid separation and detection. The method was developed and validated using amino acid standard mixtures containing acidic, neutral, and basic amino acids. For MS analysis, the optimum cone voltage implemented, based on direct infusion analysis of a few selected AccQ*Tag amino acids with multiple reaction monitoring, varied from 29 to 39 V, whereas the collision energy varied from 15 to 35 V. Calibration curves were built using both internal and external standardization. Typically, a linear response for all amino acids was observed at concentration ranges of 3 x 10(-3)-25 pmol/muL. For some amino acids, concentration limits of detection were as low as 1.65 fmol. The coefficients of variation for retention times were within the range of 0.08-1.08%. The coefficients of variation for amino acid quantitation, determined from triplicate UPLC-MS/MS runs, were below 8% on the average. The developed AccQ*Tag-UPLC-ESI-MS/MS method revealed good technical and biological reproducibility when applied to P. falciparum and human red blood cells samples. This study should provide a valuable insight into the performance of UPLC-ESI-MS/MS for amino acid quantitation using AccQ*Tag derivatization.

  20. How reliable are gas-phase proton affinity values of small carbanions? A comparison of experimental data with values calculated using Gaussian-3 and CBS compound methods

    Science.gov (United States)

    Danikiewicz, Witold

    2009-08-01

    Gas-phase proton affinities (PA) of a series of 25 small, aliphatic carbanions were computed using different Gaussian-3 methods: G3, G3(B3LYP), G3(MP2) and G3(MP2, B3LYP) and Complete Basis Set Extrapolation methods: CBS-4M, CBS-Q, CBS-QB3, and CBS-APNO. The results were compared with critically selected experimental data. The analysis of the results shows that for the majority of the studied molecules all compound methods (Gaussian-3 and CBS), except for CBS-4M, give comparable results, which differ no more than +/-2 kcal mol-1 from the experimental data. Taking into account the calculation time, G3(MP2) and G3(MP2, B3LYP) methods offer the best compromise between accuracy and computational cost. As an additional proof, the results obtained by these two methods were compared with the values obtained using CCSD(T) ab initio method with large basis set. It was found also that some of the published experimental data are erroneous and should be corrected. The results described in this work show that for the majority of the studied compounds PA values calculated using compound methods can be used with the same or even higher confidence as the experimental ones because even the largest differences between Gaussian-3 and CBS methods listed above are still comparable with the accuracy of the typical PA measurements.

  1. Research on and realization of contactless testing method of UHF RFID tags'resonance characteristics%U HF RFID 标签谐振特性非接触测试方法的研究与实现

    Institute of Scientific and Technical Information of China (English)

    李帅; 张雪凡; 任秀方; 孟春阳

    2015-01-01

    While UHF RFID tags being attached to the surface of various materials ,there will be great errors when we adopt the traditional method of cable connection test to get the tag's resonance characteristics .In this paper ,we formed a model of tag's power‐receiving ,analyzed the relationship between the energy obtained by tag chip and frequency ,and then proposed a contactless method to get the resonance characteristics of UHF RFID tags .This paper detailed the principle of using the relationship between frequency and distance to test the resonance characteristics ,illustrated the design concept of testing software , built and completely implemented the contactless hardware test platform . Measurement results demonstrated that the proposed method and platform can test the tag's resonance characteristics effectively . In practical applications , tags for different materials can be designed according to the resonance characteristics .%针对U H F RFID标签贴附在介质表面时,标签的谐振特性用接触式测量方法存在较大的误差问题,通过建立标签的接收功率模型,分析标签芯片获取到的能量与频率之间的关系,提出非接触式测试 U HF RFID谐振特性的方法。详细描述了利用频率和距离关系测试标签谐振特性的原理,说明了非接触式测试方法硬件测试平台的搭建方法和测试软件的编写思想,并完整地实现了该平台。实测实验证明,该测试方法和平台能有效地测试标签的谐振特性。在实际应用中,可以根据标签的谐振特性设计适合不同物体的标签。

  2. Dark-lumen MR colonography with fecal tagging: a comparison of water enema and air methods of colonic distension for detecting colonic neoplasms

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez Gomez, Sonia; Pages Llinas, Mario; Juan Garcia, Carmen de; Rimola Gibert, Jordi; Ayuso Colella, Juan R.; Ayuso Colella, Carmen [Hospital Clinico of Barcelona, Department of Radiology, Barcelona (Spain); Castells Garangou, Antoni; Bordas Alsina, Josep M. [Hospital Clinico of Barcelona, Department of Gastroenterology, Barcelona (Spain)

    2008-07-15

    The purpose was to evaluate MR colonography (MRC) with barium fecal tagging in detecting colorectal pathology and to determine how air-based and water-based colonic distension influences MRC. We studied 83 patients with high risk of colonic neoplasms. All received oral barium sulfate for colonic preparation before unenhanced and enhanced T1-weighted gradient-echo MRC using either water (n=54) or air (n=29) for colonic distension. Fecal tagging, distension, and artifacts were recorded. All patients underwent conventional colonoscopy within 2 weeks of MRC; the techniques were compared for detection of malignant neoplasms and polyps {>=}1 cm, 6-9 mm, and {<=}5 mm. Fecal tagging was ''good'' in 76% of the colonic segments in water-distended patients and 46% of air-distended patients. The degree of distension was ''good'' in 90.7% of water-distended patients and 44% of air-distended patients. Severe artifacts were present in 15% air-distended patients and 0.3% of water-distended patients. Both water-distended and air-distended MRC detected all malignant neoplasms and polyps {>=}1 cm, but more air-distended MRC were excluded for poor quality. MRC with fecal tagging is useful for detecting lesions {>=}1 cm. Air distension was inferior to water distension in most aspects. Water-based colonic distension should be used for barium-tagging MRC. (orig.)

  3. On affine rigidity

    Directory of Open Access Journals (Sweden)

    Steven J. Gortler

    2013-12-01

    Full Text Available We study the properties of affine rigidity of a hypergraph and prove a variety of fundamental results. First, we show that affine rigidity is a generic property (i.e., depends only on the hypergraph, not the particular embedding. Then we prove that a graph is generically neighborhood affinely rigid in d-dimensional space if it is (d+1-vertex-connected. We also show neighborhood affine rigidity of a graph implies universal rigidity of its squared graph.  Our results, and affine rigidity more generally, have natural applications in point registration and localization, as well as connections to manifold learning.

  4. Comparing the effects of Fe(III) and Cu(II) on the binding affinity of erlotinib to bovine serum albumin using spectroscopic methods

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yan [The State Key Laboratory of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China); Chen, Mingmao [Institute of Biomedical and Pharmaceutical Technology, Fuzhou University, Fuzhou, Fujian 350002 (China); Song, Ling, E-mail: songling@fjirsm.ac.cn [The State Key Laboratory of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou, Fujian 350002 (China)

    2013-02-15

    The interactions between erlotinib (ET) and bovine serum albumin (BSA) in the absence and presence of Cu(II) and Fe(III) in aqueous solution were investigated by using fluorescence, circular dichroism and three-dimensional (3D) fluorescence spectroscopic methods under simulative physiological conditions. Erlotinib effectively quenched the intrinsic fluorescence of BSA with slight redshifts in the absence and presence of Cu(II) and Fe(III). Cu(II) decreased the binding affinity and reduced the binding sites of erlotinib to BSA, while Fe(III) increased the binding affinity and binding sites of erlotinib to BSA. The negative values of {Delta}H and {Delta}S illustrate that the binding is mainly driven by the hydrogen bond and van der Waals force. The conformation of BSA was changed through ET binding in the presence of Cu(II) and Fe(III), which was revealed by circular dichroism, synchronous fluorescence and 3D fluorescence spectroscopic methods. The results indicate that the binding capability of erlotinib to BSA is affected by the types of metal ions. Highlights: Black-Right-Pointing-Pointer Interaction of erlotinib (ET) with BSA in the presence of Cu(II) and Fe(III) was studied. Black-Right-Pointing-Pointer Using various spectroscopic methods such as UV, CD, fluorescence and three-dimensional (3D) fluorescence. Black-Right-Pointing-Pointer Effects of metal ions on the binding activity of ET to BSA have not been reported. Black-Right-Pointing-Pointer Ternary system Cu(II)/Fe(III)-ET-BSA induced the conformational change of BSA.

  5. Personalized Tag Recommendation Using Social Influence

    Institute of Scientific and Technical Information of China (English)

    Jun Hu; Bing Wang; Yu Liu; De-Yi Li

    2012-01-01

    Tag recommendation encourages users to add more tags in bridging the semantic gap between human concept and the features of media object,which provides a feasible solution for content-based multimedia information retrieval.In this paper,we study personalized tag recommendation in a popular online photo sharing site Flickr.Social relationship information of users is collected to generate an online social network. From the perspective of network topology,we propose node topological potential to characterize user's social influence.With this metric,we distinguish different social relations between users and find out those who really have influence on the target users.Tag recommendations are based on tagging history and the latent personalized preference learned from those who have most influence in user's social network.We evaluate our method on large scale real-world data.The experimental results demonstrate that our method can outperform the non-personalized global co-occurrence method and other two state-of-the-art personalized approaches using social networks.We also analyze the further usage of our approach for the cold-start problem of tag recommendation.

  6. Comparison of several tag-marked methods in seed dispersal by rats%鼠类扩散种子的几种标签标记法的比较

    Institute of Scientific and Technical Information of China (English)

    常罡

    2012-01-01

    以四川省都江堰市亚热带常绿阔叶林内的优势鼠种——小泡巨鼠为实验动物,通过半自然状态围栏控制实验,检验了几种种子标签法对鼠类扩散植物种子的影响.结果表明:鱼线和细钢丝两种标记线在跟踪扩散种子命运上都是很有效的,但考虑到鱼线有时会被实验鼠咬断,细钢丝是更值得推广应用的一种标记线.3种标记牌(大塑料牌、小塑料牌和金属牌)在跟踪扩散种子命运上没有显著差异,但由于较大的尺寸和较强的可见度,大塑料牌更适合作为标签应用于野外种子扩散标记.%Using Edward' s long-tailed rat (Leopoldamys edwardsi) , a dominant species in sub-tropical evergreen broadleaf forests in Dujiangyan City of Sichuan Province, as the experimental animal, a field study with semi-natural enclosure was conducted to examine the effects of several tag-marked methods on the seed dispersal by rats. Among the methods examined, both the marked lines fishing thread and thin steel wire were effective on tracking the seed fates dispersed by rats, but, in considering that the fishing thread was sometimes bitted off by experimental rats, the steel wire was more worthy of application as a kind of perfect marked line. Three kinds of marked tags, I. E. , large plastic tag, small plastic tag, and wire tag, did not differ on tracking the seed fates dispersed by rats, but the large plastic tag, due to its large size and strong visibili-ty , was more suitable to be a perfect marked tag for field seed dispersal.

  7. Methodologies for Improved Tag Cloud Generation with Clustering

    DEFF Research Database (Denmark)

    Leginus, Martin; Dolog, Peter; Lage, Ricardo Gomes

    2012-01-01

    coverage and overlap. We study several clustering algorithms to generate tag clouds. We show that by extending cloud generation based on tag popularity with clustering we slightly improve coverage. We also show that if the cloud is generated by clustering independently of the tag popularity baseline we......Tag clouds are useful means for navigation in the social web systems. Usually the systems implement the tag cloud generation based on tag popularity which is not always the best method. In this paper we propose methodologies on how to combine clustering into the tag cloud generation to improve...... better on bibsonomy due to its specific focus. The best performing is the hierarchical clustering....

  8. Synthesis of Sulochrin-125I and Its Binding Affinity as α-Glucosidase Inhibitor using Radioligand Binding Assay (RBA Method

    Directory of Open Access Journals (Sweden)

    W. Lestari

    2014-04-01

    Full Text Available Most of diabetics patients have type 2 diabetes mellitus or non insulin dependent diabetes mellitus. Treatment type 2 diabetes mellitus can be done by inhibiting α-glucosidase enzyme which converts carbohydrates into glucose. Sulochrin is one of the potential compounds which can inhibit the function of α-glucosidase enzyme. This study was carried out to obtain data of sulochrin binding with α-glucosidase enzyme as α-glucosidase inhibitor using Radioligand Binding Assay (RBA method. Primary reagent required in RBA method is labeled radioactive ligand (radioligand. In this study, the radioligand was sulochrin-125I and prior to sulochrin-125I synthesis, the sulochrin-I was synthesized. Sulochrin-I and sulochrin-125I were synthesized and their bindings were studied using Radioligand Binding Assay method. Sulochrin-I was synthesized with molecular formula C17H15O7I and molecular weight 457.9940. Sulochrin-125I was synthesized from sulochrin-I by isotope exchange method. From the RBA method, dissociation constant (Kd and maximum binding (Bmax were obtained 26.316 nM and Bmax 9.302 nM respectively. This low Kd indicated that sulochrin was can bind to α-glucosidase

  9. Inclusive Flavour Tagging Algorithm

    Science.gov (United States)

    Likhomanenko, Tatiana; Derkach, Denis; Rogozhnikov, Alex

    2016-10-01

    Identifying the flavour of neutral B mesons production is one of the most important components needed in the study of time-dependent CP violation. The harsh environment of the Large Hadron Collider makes it particularly hard to succeed in this task. We present an inclusive flavour-tagging algorithm as an upgrade of the algorithms currently used by the LHCb experiment. Specifically, a probabilistic model which efficiently combines information from reconstructed vertices and tracks using machine learning is proposed. The algorithm does not use information about underlying physics process. It reduces the dependence on the performance of lower level identification capacities and thus increases the overall performance. The proposed inclusive flavour-tagging algorithm is applicable to tag the flavour of B mesons in any proton-proton experiment.

  10. Double trouble-Buffer selection and His-tag presence may be responsible for nonreproducibility of biomedical experiments.

    Science.gov (United States)

    Majorek, Karolina A; Kuhn, Misty L; Chruszcz, Maksymilian; Anderson, Wayne F; Minor, Wladek

    2014-10-01

    The availability of purified and active protein is the starting point for the majority of in vitro biomedical, biochemical, and drug discovery experiments. The use of polyhistidine affinity tags has resulted in great increases of the efficiency of the protein purification process, but can negatively affect structure and/or activity measurements. Similarly, buffer molecules may perturb the conformational stability of a protein or its activity. During the determination of the structure of a Gcn5-related N-acetyltransferase (GNAT) from Pseudomonas aeruginosa (PA4794), we found that both HEPES and the polyhistidine affinity tag bind (separately) in the substrate-binding site. In the case of HEPES, the molecule induces conformational changes in the active site, but does not significantly affect enzyme activity. In contrast, the uncleaved His-tag does not induce major conformational changes but acts as a weak competitive inhibitor of peptide substrate. In two other GNAT enzymes, we observed that the presence of the His-tag had a strong influence on the activity of these proteins. The influence of protein preparation on functional studies may affect the reproducibility of experiments in other laboratories, even when changes between protocols seem at first glance to be insignificant. Moreover, the results presented here show how critical it is to adjust the experimental conditions for each protein or family of proteins, and investigate the influence of these factors on protein activity and structure, as they may significantly alter the effectiveness of functional characterization and screening methods. Thus, we show that a polyhistidine tag and the buffer molecule HEPES bind in the substrate-binding site and influence the conformation of the active site and the activity of GNAT acetyltransferases. We believe that such discrepancies can influence the reproducibility of some experiments and therefore could have a significant "ripple effect" on subsequent studies.

  11. Protease substrate profiling using bacterial display of self-blocking affinity proteins and flow-cytometric sorting.

    Science.gov (United States)

    Sandersjöö, Lisa; Jonsson, Andreas; Löfblom, John

    2017-01-01

    Proteases are involved in fundamental biological processes and are important tools in both biotechnological and biomedical research. An important property of proteases is to discriminate among potential substrates. Here, a new method for substrate profiling of proteases is presented. The substrates are displayed between two anti-idiotypic affinity domains on the Gram-positive bacterium Staphylococcus carnosus. The first domain functions as a reporter tag and has affinity for a labeled reporter protein, whereas the second domain blocks the reporter tag from interacting with the reporter protein. Site-specific proteolysis of the substrate results in release of the blocking domain, enabling the reporter tag to bind the labeled reporter protein. Proteolysis is therefore reflected in reporter binding, which is quantified by flow cytometry. First, the method with tobacco etch virus protease (TEVp) is evaluated and then the substrate preference of matrix metalloprotease-1 (MMP-1) is determined using two libraries of around three million substrates each. Identified substrate peptides contained the previously reported motif (PXXXHy ) and on-cell determination of apparent kcat /KM revealed that the enriched substrate peptides are hydrolyzed six to eight-fold more efficiently than a previously reported substrate peptide. The method thus works as intended and the authors believe it has potential as an efficient tool for substrate profiling.

  12. 固定RFID电子标签探测定位方法%A Method about Detecting the Fixed Tag to Realize Localization

    Institute of Scientific and Technical Information of China (English)

    汪勇

    2012-01-01

    研究了一种无线通信的定位方法,介绍了利用移动阅读器探测固定RFID(射频识别技术)电子标签的定位技术,建立了系统基本模型。并介绍了该定位方法在车载系统的导航定位系统及室内三维定位系统中的应用。提出的新定位方法是对传统GPS/北斗卫星定位系统的有益补充。可弥补卫星定位在城市“峡谷”区域工作时由于阴影、多路径等因素所导致的定位误差。%The purpose of this paper is to propose a new kind of positioning method based on wireless communication. In this study, attempts were made to use mobile reader to detect the fixed tag to realize localization and build the basic model of the system. At the same time, tlle applications of navigation system on vehicle and indoor three dimensional positioning are presented. This study can also supplement the traditional GPS/Beidou satellite positioning system beneficially, because it can make up the positioning error which led to by shadow, multipath and other factors in the city positioning.

  13. Purification and characterisation of recombinant His-tagged RgpB gingipain from Porphymonas gingivalis.

    Science.gov (United States)

    Veillard, Florian; Potempa, Barbara; Guo, Yonghua; Ksiazek, Miroslaw; Sztukowska, Maryta N; Houston, John A; Koneru, Lahari; Nguyen, Ky-Anh; Potempa, Jan

    2015-04-01

    Gingipain proteases are important virulence factors from the periodontal pathogen Porphyromonas gingivalis and are the target of many in vitro studies. Due to their close biochemical properties, purification of individual gingipains is difficult and requires multiple chromatographic steps. In this study, we demonstrate that insertion of a hexahistidine affinity tag upstream of a C-terminal outer membrane translocation signal in RgpB gingipain leads to the secretion of a soluble, mature form of RgpB bearing the affinity tag that can easily be purified by nickel-chelating affinity chromatography. The final product obtained high yielding high purity is biochemically indistinguishable from the native RgpB enzyme.

  14. Tandem affinity purification to identify cytosolic and nuclear gβγ-interacting proteins.

    Science.gov (United States)

    Campden, Rhiannon; Pétrin, Darlaine; Robitaille, Mélanie; Audet, Nicolas; Gora, Sarah; Angers, Stéphane; Hébert, Terence E

    2015-01-01

    It has become clear in recent years that the Gβγ subunits of heterotrimeric proteins serve broad roles in the regulation of cellular activity and interact with many proteins in different subcellular locations including the nucleus. Protein affinity purification is a common method to identify and confirm protein interactions. When used in conjugation with mass spectrometry it can be used to identify novel protein interactions with a given bait protein. The tandem affinity purification (TAP) technique identifies partner proteins bound to tagged protein bait. Combined with protocols to enrich the nuclear fraction of whole cell lysate through sucrose cushions, TAP allows for purification of interacting proteins found specifically in the nucleus. Here we describe the use of the TAP technique on cytosolic and nuclear lysates to identify candidate proteins, through mass spectrometry, that bind to Gβ1 subunits.

  15. A new affine transformation parameters estimation method%一种新的仿射变换参数估计方法

    Institute of Scientific and Technical Information of China (English)

    何凯; 远中文; 牟聪翀

    2011-01-01

    仿射变换参数的精确估计是进行图像配准的关键,传统基于超定方程的参数估计方法对误匹配点比较敏感,对匹配准确率提出了很高的要求。针对这种情况,本文利用改进后的牛顿法对仿射变换参数估计算法进行了研究,建立了迭代模型,并推导了相关的数学公式,通过与SIFT特征点提取方法相结合,构建了新的图像配准算法仿真实验结果表明,与传统求解超定方程的方法相比,该方法能够有效减少误匹配点对估计结果的影响,参数估计更加准确,且估计时间缩短了一个数量级。利用新的图像配准方法实现了实际航空影像的精确配准,表明本文方法具有很高的精度,能够满足实际工程的需要。%The precise estimation of affine transformation parameter is most significant in image matching. Since the traditional parameters estimation method based on solving over determined equation is sensitive to mismatch points, high precision of matching is required. In view of this situation, the improved Newton method is studied to estimate the affine transformation parameters the relevant mathematical model is established, and the formulas are deduced. Besides that, by combining the parameter estimation method with SIFT feature extraction algorithm, a new image registration algorithm is proposed. The simulation results show that this estimation algorithm is robust to mismatch points, and the estimated parameters are more accurate than traditional method. Besides that, an order of magnitude improvement is made in time consuming. With the improved registration method, precise registration of actual aviation images are realized, therefore this method is high precise and suitable in practical engineering.

  16. Analysis of tag-position bias in MPSS technology

    Directory of Open Access Journals (Sweden)

    Rattray Magnus

    2006-04-01

    Full Text Available Abstract Background Massively Parallel Signature Sequencing (MPSS technology was recently developed as a high-throughput technology for measuring the concentration of mRNA transcripts in a sample. It has previously been observed that the position of the signature tag in a transcript (distance from 3' end can affect the measurement, but this effect has not been studied in detail. Results We quantify the effect of tag-position bias in Classic and Signature MPSS technology using published data from Arabidopsis, rice and human. We investigate the relationship between measured concentration and tag-position using nonlinear regression methods. The observed relationship is shown to be broadly consistent across different data sets. We find that there exist different and significant biases in both Classic and Signature MPSS data. For Classic MPSS data, genes with tag-position in the middle-range have highest measured abundance on average while genes with tag-position in the high-range, far from the 3' end, show a significant decrease. For Signature MPSS data, high-range tag-position genes tend to have a flatter relationship between tag-position and measured abundance. Thus, our results confirm that the Signature MPSS method fixes a substantial problem with the Classic MPSS method. For both Classic and Signature MPSS data there is a positive correlation between measured abundance and tag-position for low-range tag-position genes. Compared with the effects of mRNA length and number of exons, tag-position bias seems to be more significant in Arabadopsis. The tag-position bias is reflected both in the measured abundance of genes with a significant tag count and in the proportion of unexpressed genes identified. Conclusion Tag-position bias should be taken into consideration when measuring mRNA transcript abundance using MPSS technology, both in Classic and Signature MPSS methods.

  17. Tagging behaviour with support from controlled vocabulary

    DEFF Research Database (Denmark)

    Lykke, Marianne; Høj, Anne Lyhne; Madsen, Line Nørgaard

    2011-01-01

    The paper investigates how knowledge structures from a controlled vocabulary affect tagging. The study is a comparative analysis of tags assigned in two tagging systems, a simple tagging system (control system) that provides suggestions from two tag clouds (all users tags and my tags) and an enha...

  18. In Silico Prediction of Estrogen Receptor Subtype Binding Affinity and Selectivity Using Statistical Methods and Molecular Docking with 2-Arylnaphthalenes and 2-Arylquinolines

    Directory of Open Access Journals (Sweden)

    Yonghua Wang

    2010-09-01

    Full Text Available Over the years development of selective estrogen receptor (ER ligands has been of great concern to researchers involved in the chemistry and pharmacology of anticancer drugs, resulting in numerous synthesized selective ER subtype inhibitors. In this work, a data set of 82 ER ligands with ERα and ERβ inhibitory activities was built, and quantitative structure-activity relationship (QSAR methods based on the two linear (multiple linear regression, MLR, partial least squares regression, PLSR and a nonlinear statistical method (Bayesian regularized neural network, BRNN were applied to investigate the potential relationship of molecular structural features related to the activity and selectivity of these ligands. For ERα and ERβ, the performances of the MLR and PLSR models are superior to the BRNN model, giving more reasonable statistical properties (ERα: for MLR, Rtr2 = 0.72, Qte2 = 0.63; for PLSR, Rtr2 = 0.92, Qte2 = 0.84. ERβ: for MLR, Rtr2 = 0.75, Qte2 = 0.75; for PLSR, Rtr2 = 0.98, Qte2 = 0.80. The MLR method is also more powerful than other two methods for generating the subtype selectivity models, resulting in Rtr2 = 0.74 and Qte2 = 0.80. In addition, the molecular docking method was also used to explore the possible binding modes of the ligands and a relationship between the 3D-binding modes and the 2D-molecular structural features of ligands was further explored. The results show that the binding affinity strength for both ERα and ERβ is more correlated with the atom fragment type, polarity, electronegativites and hydrophobicity. The substitutent in position 8 of the naphthalene or the quinoline plane and the space orientation of these two planes contribute the most to the subtype selectivity on the basis of similar hydrogen bond interactions between binding ligands and both ER subtypes. The QSAR models built together with the docking procedure should be of great advantage for screening and designing ER ligands with improved affinity

  19. Ordinary differential equations in affine geometry

    Directory of Open Access Journals (Sweden)

    Salvador Gigena

    1996-05-01

    Full Text Available The method of qualitative analysis is used, as applied to a class of fourth order, nonlinear ordinary differential equations, in order to classify, both locally and globally, two classes of hypersurfaces of decomposable type in affine geometry: those with constant unimodular affine mean curvature L , and those with constant Riemannian scalar curvature R. This allows to provide a large number of new examples of hypersurfaces in affine geometry.

  20. Ordinary differential equations in affine geometry

    OpenAIRE

    Salvador Gigena

    1996-01-01

    The method of qualitative analysis is used, as applied to a class of fourth order, nonlinear ordinary differential equations, in order to classify, both locally and globally, two classes of hypersurfaces of decomposable type in affine geometry: those with constant unimodular affine mean curvature L , and those with constant Riemannian scalar curvature R. This allows to provide a large number of new examples of hypersurfaces in affine geometry.

  1. Isolation of Human Antibodies Against Hepatitis E From Phage Display Library by Metal Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective To isolate human antibodies against hepatitis E virus from phage display library by a new method of panning phage antibody library based on immobilized metal affinity chromatography (IMAC). Methods Phage antibody library was allowed to mix with hex-His tagged expressed HEV specific antigen, NE2, in solution for adequate binding before affinity resin for hex-His was added. The non-specific phage antibodies were removed by extensive washing and the specific bound phage antibodies could then be eluted to infect TG1 or repeat the binding process for subsequent rounds of purification. The specificity of the selected human antibodies were tested by antigen competitive ELISA, human sera blocking ELISA, scFv expression, and sequence analysis. Results His-NE2 specific recombinant phages were successfully enriched after panning procedure. Two individual phage clones, 126 and 138, showed 50% inhibition in NE2 antigen competition ELISA and obvious blocking effect by HEV positive serum in blocking ELISA. Soluble scFv of 126, 138 bound to NE2 specifically. Conclusion Two specific human phage antibodies against hepatitis E virus (HEV) from phage display library were isolated by immobilized metal affinity chromatography. The immobilized metal affinity chromatography applied to phage antibody selection was a helpful supplement to the selection in solution.

  2. Harvesting Intelligence in Multimedia Social Tagging Systems

    Science.gov (United States)

    Giannakidou, Eirini; Kaklidou, Foteini; Chatzilari, Elisavet; Kompatsiaris, Ioannis; Vakali, Athena

    As more people adopt tagging practices, social tagging systems tend to form rich knowledge repositories that enable the extraction of patterns reflecting the way content semantics is perceived by the web users. This is of particular importance, especially in the case of multimedia content, since the availability of such content in the web is very high and its efficient retrieval using textual annotations or content-based automatically extracted metadata still remains a challenge. It is argued that complementing multimedia analysis techniques with knowledge drawn from web social annotations may facilitate multimedia content management. This chapter focuses on analyzing tagging patterns and combining them with content feature extraction methods, generating, thus, intelligence from multimedia social tagging systems. Emphasis is placed on using all available "tracks" of knowledge, that is tag co-occurrence together with semantic relations among tags and low-level features of the content. Towards this direction, a survey on the theoretical background and the adopted practices for analysis of multimedia social content are presented. A case study from Flickr illustrates the efficiency of the proposed approach.

  3. Affine Dynamics with Torsion

    CERN Document Server

    Gultekin, Kemal

    2015-01-01

    In this study, we give a thorough analysis of a general affine gravity with torsion. After a brief exposition of the affine gravities considered by Eddington and Schroedinger, we construct and analyze different affine gravities based on determinants of the Ricci tensor, torsion tensor, Riemann tensor and their combinations. In each case we reduce equations of motion to their simplest forms and give a detailed analysis of their solutions. Our analyses lead to construction of the affine connection in terms of curvature and torsion tensors. Our solutions of the dynamical equations show that curvature tensors at different points are correlated via non-local, exponential rescaling factors determined by the torsion tensor.

  4. LHCb flavour tagging performance

    CERN Document Server

    Calvi, M; Musy, M

    2003-01-01

    In CP violation measurements, the most accurate determination of the B flavour of neutral and charged B-mesons is necessary. In this note we summarize the tagging performances for the LHCb experiment, using different approaches and studying different decay channels.

  5. Personalization of tagging systems

    NARCIS (Netherlands)

    Wang, J.; Clements, M.; Yang, J.; Vries, A.P. de; Reinders, M.J.T.

    2010-01-01

    Social media systems have encouraged end user participation in the Internet, for the purpose of storing and distributing Internet content, sharing opinions and maintaining relationships. Collaborative tagging allows users to annotate the resulting user-generated content, and enables effective retrie

  6. Tagging the Teleman Corpus

    CERN Document Server

    Brants, T; Brants, Thorsten; Samuelsson, Christer

    1995-01-01

    Experiments were carried out comparing the Swedish Teleman and the English Susanne corpora using an HMM-based and a novel reductionistic statistical part-of-speech tagger. They indicate that tagging the Teleman corpus is the more difficult task, and that the performance of the two different taggers is comparable.

  7. Tagging Grammatical Functions

    CERN Document Server

    Brants, T; Krenn, B; Brants, Thorsten; Skut, Wojciech; Krenn, Brigitte

    1997-01-01

    This paper addresses issues in automated treebank construction. We show how standard part-of-speech tagging techniques extend to the more general problem of structural annotation, especially for determining grammatical functions and syntactic categories. Annotation is viewed as an interactive process where manual and automatic processing alternate. Efficiency and accuracy results are presented. We also discuss further automation steps.

  8. Tagging vs. Controlled Vocabulary

    DEFF Research Database (Denmark)

    Bogers, Toine; Petras, Vivien

    2015-01-01

    elements like core bibliographic data, controlled vocabulary terms, reviews, and tags to the retrieval performance. Our comparison is done using a test collection of over 2 million book records with information elements from Amazon, the British Library, the Library of Congress, and LibraryThing. We find...

  9. Ontologies and tag-statistics

    Science.gov (United States)

    Tibély, Gergely; Pollner, Péter; Vicsek, Tamás; Palla, Gergely

    2012-05-01

    Due to the increasing popularity of collaborative tagging systems, the research on tagged networks, hypergraphs, ontologies, folksonomies and other related concepts is becoming an important interdisciplinary area with great potential and relevance for practical applications. In most collaborative tagging systems the tagging by the users is completely ‘flat’, while in some cases they are allowed to define a shallow hierarchy for their own tags. However, usually no overall hierarchical organization of the tags is given, and one of the interesting challenges of this area is to provide an algorithm generating the ontology of the tags from the available data. In contrast, there are also other types of tagged networks available for research, where the tags are already organized into a directed acyclic graph (DAG), encapsulating the ‘is a sub-category of’ type of hierarchy between each other. In this paper, we study how this DAG affects the statistical distribution of tags on the nodes marked by the tags in various real networks. The motivation for this research was the fact that understanding the tagging based on a known hierarchy can help in revealing the hidden hierarchy of tags in collaborative tagging systems. We analyse the relation between the tag-frequency and the position of the tag in the DAG in two large sub-networks of the English Wikipedia and a protein-protein interaction network. We also study the tag co-occurrence statistics by introducing a two-dimensional (2D) tag-distance distribution preserving both the difference in the levels and the absolute distance in the DAG for the co-occurring pairs of tags. Our most interesting finding is that the local relevance of tags in the DAG (i.e. their rank or significance as characterized by, e.g., the length of the branches starting from them) is much more important than their global distance from the root. Furthermore, we also introduce a simple tagging model based on random walks on the DAG, capable of

  10. A combination of docking, QM/MM methods, and MD simulation for binding affinity estimation of metalloprotein ligands.

    Science.gov (United States)

    Khandelwal, Akash; Lukacova, Viera; Comez, Dogan; Kroll, Daniel M; Raha, Soumyendu; Balaz, Stefan

    2005-08-25

    To alleviate the problems in the receptor-based design of metalloprotein ligands due to inadequacies in the force-field description of coordination bonds, a four-tier approach was devised. Representative ligand-metalloprotein interaction energies are obtained by subsequent application of (1) docking with metal-binding-guided selection of modes, (2) optimization of the ligand-metalloprotein complex geometry by combined quantum mechanics and molecular mechanics (QM/MM) methods, (3) conformational sampling of the complex with constrained metal bonds by force-field-based molecular dynamics (MD), and (4) a single point QM/MM energy calculation for the time-averaged structures. The QM/MM interaction energies are, in a linear combination with the desolvation-characterizing changes in the solvent-accessible surface areas, correlated with experimental data. The approach was applied to structural correlation of published binding free energies of a diverse set of 28 hydroxamate inhibitors to zinc-dependent matrix metalloproteinase 9 (MMP-9). Inclusion of steps 3 and 4 significantly improved both correlation and prediction. The two descriptors explained 90% of variance in inhibition constants of all 28 inhibitors, ranging from 0.08 to 349 nM, with the average unassigned error of 0.318 log units. The structural and energetic information obtained from the time-averaged MD simulation results helped understand the differences in binding modes of related compounds.

  11. Evidence of multi-affinity in the Japanese stock market

    Science.gov (United States)

    Katsuragi, Hiroaki

    2000-04-01

    Fluctuations of the Japanese stock market (Tokyo Stock Price Index: TOPIX) are analyzed using a multi-affine analysis method. In the research to date, only some simulated self-affine models have shown multi-affinity. In most experiments using observations of self-affine fractal profiles, multi-affinity has not been found. However, we find evidence of multi-affinity in fluctuations of the Japanese stock market (TOPIX). The qth-order Hurst exponent Hq varies with changes in q. This multi-affinity indicates that there are plural mechanisms that affect the same time scale as stock market price fluctuation dynamics.

  12. Affine connections, midpoint formation, and point reflection

    DEFF Research Database (Denmark)

    Kock, Anders

    2011-01-01

    We describe some differential-geometric structures in combinatorial terms: namely affine connections and their torsion and curvature, and we show that torsion free affine connections may equivalently be presented in terms of some simpler combinatorial structure: midpoint formation, and point refl...... reflection (geodesic symmetry). The method employed is that of synthetic differential geometry, which is briefly explained.......We describe some differential-geometric structures in combinatorial terms: namely affine connections and their torsion and curvature, and we show that torsion free affine connections may equivalently be presented in terms of some simpler combinatorial structure: midpoint formation, and point...

  13. Expression and affinity purification of recombinant proteins from plants

    Science.gov (United States)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  14. Associated Particle Tagging (APT) in Magnetic Spectrometers

    Energy Technology Data Exchange (ETDEWEB)

    Jordan, David V.; Baciak, James E.; Stave, Sean C.; Chichester, David; Dale, Daniel; Kim, Yujong; Harmon, Frank

    2012-10-16

    alpha-particle spectrometer concept, and outlines challenges involved in the magnetic field design. Tagged photon interrogation: • We investigated a method for discriminating fissile from benign cargo-material response to an energy-tagged photon beam. The method relies upon coincident detection of the tagged photon and a photoneutron or photofission neutron produced in the target material. The method exploits differences in the shape of the neutron production cross section as a function of incident photon energy in order to discriminate photofission yield from photoneutrons emitted by non-fissile materials. Computational tests of the interrogation method as applied to material composition assay of a simple, multi-layer target suggest that the tagged-photon information facilitates precise (order 1% thickness uncertainty) reconstruction of the constituent thicknesses of fissile (uranium) and high-Z (Pb) constituents of the test targets in a few minutes of photon-beam exposure. We assumed an 18-MeV endpoint tagged photon beam for these simulations. • The report addresses several candidate design and data analysis issues for beamline infrastructure required to produce a tagged photon beam in a notional AI-dedicated facility, including the accelerator and tagging spectrometer.

  15. The HaloTag: Improving Soluble Expression and Applications in Protein Functional Analysis.

    Science.gov (United States)

    N Peterson, Scott; Kwon, Keehwan

    2012-01-01

    Technological and methodological advances have been critical for the rapidly evolving field of proteomics. The development of fusion tag systems is essential for purification and analysis of recombinant proteins. The HaloTag is a 34 KDa monomeric protein derived from a bacterial haloalkane dehalogenase. The majority of fusion tags in use today utilize a reversible binding interaction with a specific ligand. The HaloTag system is unique in that it forms a covalent linkage to its chloroalkane ligand. This linkage permits attachment of the HaloTag to a variety of functional reporters, which can be used to label and immobilize recombinant proteins. The success rate for HaloTag expression of soluble proteins is very high and comparable to maltose binding protein (MBP) tag. Furthermore, cleavage of the HaloTag does not result in protein insolubility that often is observed with the MBP tag. In the present report, we describe applications of the HaloTag system in our ongoing investigation of protein-protein interactions of the Y. pestis Type 3 secretion system on a custom protein microarray. We also describe the utilization of affinity purification/mass spectroscopy (AP/MS) to evaluate the utility of the Halo Tag system to characterize DNA binding activity and protein specificity.

  16. Development of techniques for tagging precursor and essential chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Swansiger, W.A.; Shepodd, T.J. [Sandia National Labs., Livermore, CA (United States); Phillips, M.L.F. [Sandia National Labs., Albuquerque, NM (United States)

    1994-01-01

    The ability to identify the manufacturers and distributors of chemicals seized in raids of illicit drug labs would be of great value in controlling the diversion of these chemicals. We developed a tagging scheme based on the addition of sub-ppM concentrations of various combinations of rare-earth elements to the target chemicals and evaluated a number of techniques for detecting the tags. We developed soluble tags for tagging liquids and selected Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) as the preferred detection technique. We developed insoluble tags for tagging solids and developed methods to analyze them and mix them into solid precursors. We have successfully demonstrated the tagging of several solvents and two of the precursor chemicals used in one of the most popular clandestine methamphetamine syntheses (ephedrine reacting with hydriodic acid/red phosphorus). The tagging scheme is capable of yielding tens of thousands of signatures (using holmium as an internal standard and up to 9 rare-earths at up to 3 concentrations yields 3{sup 9} {minus} 1 = 19,682 signatures) and is applicable to most of the chemicals on the precursor and essential chemicals list. In the concentrations employed, the tags are safe enough to be added to pharmaceuticals and cheap enough to tag tanker loads of chemicals.

  17. Flavor Tagging at Tevatron incl. calibration and control

    OpenAIRE

    2007-01-01

    This report summarizes the flavor tagging techniques developed at the CDF and D{\\O}experiments. Flavor tagging involves identification of the B meson flavor atproduction, whether its constituent is a quark or an anti-quark. It is crucial for measuring the oscillation frequency of neutral B mesons, both in the B^0 and B_S system. The two experiments have developed their unique approaches to flavor tagging, using neural networks, and likelihood methods to disentangle tracks from $b$ decays from...

  18. Single step purification of recombinant proteins using the metal ion-inducible autocleavage (MIIA) domain as linker for tag removal.

    Science.gov (United States)

    Ibe, Susan; Schirrmeister, Jana; Zehner, Susanne

    2015-08-20

    For fast and easy purification, proteins are typically fused with an affinity tag, which often needs to be removed after purification. Here, we present a method for the removal of the affinity tag from the target protein in a single step protocol. The protein VIC_001052 of the coral pathogen Vibrio coralliilyticus ATCC BAA-450 contains a metal ion-inducible autocatalytic cleavage (MIIA) domain. Its coding sequence was inserted into an expression vector for the production of recombinant fusion proteins. Following, the target proteins MalE and mCherry were produced as MIIA-Strep fusion proteins in Escherichia coli. The target proteins could be separated from the MIIA-Strep part simply by the addition of calcium or manganese(II) ions within minutes. The cleavage is not affected in the pH range from 5.0 to 9.0 or at low temperatures (6°C). Autocleavage was also observed with immobilized protein on an affinity column. The protein yield was similar to that achieved with a conventional purification protocol.

  19. 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...

  20. Self-Powered Wireless Affinity-Based Biosensor Based on Integration of Paper-Based Microfluidics and Self-Assembled RFID Antennas.

    Science.gov (United States)

    Yuan, Mingquan; Alocilja, Evangelyn C; Chakrabartty, Shantanu

    2016-08-01

    This paper presents a wireless, self-powered, affinity-based biosensor based on the integration of paper-based microfluidics with our previously reported method for self-assembling radio-frequency (RF) antennas. At the core of the proposed approach is a silver-enhancement technique that grows portions of a RF antenna in regions where target antigens hybridize with target specific affinity probes. The hybridization regions are defined by a network of nitrocellulose based microfluidic channels which implement a self-powered approach to sample the reagent and control its flow and mixing. The integration substrate for the biosensor has been constructed using polyethylene and the patterning of the antenna on the substrate has been achieved using a low-cost ink-jet printing technique. The substrate has been integrated with passive radio-frequency identification (RFID) tags to demonstrate that the resulting sensor-tag can be used for continuous monitoring in a food supply-chain where direct measurement of analytes is typically considered to be impractical. We validate the proof-of-concept operation of the proposed sensor-tag using IgG as a model analyte and using a 915 MHz Ultra-high-frequency (UHF) RFID tagging technology.

  1. Tag-Aware Recommender Systems: A State-of-the-art Survey

    CERN Document Server

    Zhang, Zi-Ke; Zhang, Yi-Cheng; 10.1007/s11390-011-0176-1

    2012-01-01

    In the past decade, Social Tagging Systems have attracted increasing attention from both physical and computer science communities. Besides the underlying structure and dynamics of tagging systems, many efforts have been addressed to unify tagging information to reveal user behaviors and preferences, extract the latent semantic relations among items, make recommendations, and so on. Specifically, this article summarizes recent progress about tag-aware recommender systems, emphasizing on the contributions from three mainstream perspectives and approaches: network-based methods, tensor-based methods, and the topic-based methods. Finally, we outline some other tag-related works and future challenges of tag-aware recommendation algorithms.

  2. An improved toolbox to unravel the plant cellular machinery by tandem affinity purification of Arabidopsis protein complexes.

    Science.gov (United States)

    Van Leene, Jelle; Eeckhout, Dominique; Cannoot, Bernard; De Winne, Nancy; Persiau, Geert; Van De Slijke, Eveline; Vercruysse, Leen; Dedecker, Maarten; Verkest, Aurine; Vandepoele, Klaas; Martens, Lennart; Witters, Erwin; Gevaert, Kris; De Jaeger, Geert

    2015-01-01

    Tandem affinity purification coupled to mass spectrometry (TAP-MS) is one of the most advanced methods to characterize protein complexes in plants, giving a comprehensive view on the protein-protein interactions (PPIs) of a certain protein of interest (bait). The bait protein is fused to a double affinity tag, which consists of a protein G tag and a streptavidin-binding peptide separated by a very specific protease cleavage site, allowing highly specific protein complex isolation under near-physiological conditions. Implementation of this optimized TAP tag, combined with ultrasensitive MS, means that these experiments can be performed on small amounts (25 mg of total protein) of protein extracts from Arabidopsis cell suspension cultures. It is also possible to use this approach to isolate low abundant protein complexes from Arabidopsis seedlings, thus opening perspectives for the exploration of protein complexes in a plant developmental context. Next to protocols for efficient biomass generation of seedlings (∼7.5 months), we provide detailed protocols for TAP (1 d), and for sample preparation and liquid chromatography-tandem MS (LC-MS/MS; ∼5 d), either from Arabidopsis seedlings or from cell cultures. For the identification of specific co-purifying proteins, we use an extended protein database and filter against a list of nonspecific proteins on the basis of the occurrence of a co-purified protein among 543 TAP experiments. The value of the provided protocols is illustrated through numerous applications described in recent literature.

  3. 基于词共现的社会化标签研究热点可视化分析%Visualization of Hot Topics in Social Tagging Based on Co-words Analysis Method

    Institute of Scientific and Technical Information of China (English)

    卢小宾; 孟玺; 张进

    2012-01-01

    The paper chose Web of Science database as the data source, used co-word method and distance-basedsimilarity measure to process the original matrix, through multidimensional scaling analysis and clustering analysis to visualize the research subjects and hot spots of social tagging research field. Analysis results showed that the social tagging research focused on the semantic representation of tags、tag conceptualizations ( ontology)、information filtering,recommend system,how to be compatible with traditional subject heading and social networking platform.%以ISI的Web of Science数据库为数据来源,采用词共现方法和基于距离的相似性度量算法对原始矩阵进行处理,通过多维尺度和系统聚类分析对社会化标签研究领域的研究主题和热点进行可视化揭示.分析结果表明,对社会化标签的研究主要集中在大众标签的语义表达、标签概念化(本体)、信息过滤、推荐系统、与传统主题词表的兼容问题、对社交网络平台的研究.

  4. 仿射变换内点Levenberg-Marquardt法解KKT系统%Affine scaling interior Levenberg-Marquardt method for KKT systems

    Institute of Scientific and Technical Information of China (English)

    王云娟; 朱德通

    2013-01-01

    提供了一类新的结合非单调内点回代线搜索技术的仿射变换Levenberg-Marquardt法解Karush-Kuhn-Tucker (KKT)系统.基于由KKT系统转化得到的等价的部分变量具有非负约束的最小化问题,建立了Levenberg-Maxquardt方程.证明了算法不仅具有整体收敛性,而且在合理的假设条件下,算法具有超线性收敛速率.数值结果验证了算法的实际有效性.%We develop and analyze a new affine scaling Levenberg-Marquardt method with nonmonotonic interior backtracking line search technique for solving Karush-KuhnTucker (KKT) system.By transforming the KKT system into an equivalent minimization problem with nonnegativity constraints on some of the variables,we establish the Levenberg-Marquardt equation based on this reformulation.Theoretical analysis are given which prove that the proposed algorithm is globally convergent and has a local superlinear convergent rate under some reasonable conditions.The results of numerical experiments are reported to show the effectiveness of the proposed algorithm.

  5. Interest Modeling Method for Web Users Based on Social Tagging Systems%一种社会化标签系统的用户兴趣建模方法

    Institute of Scientific and Technical Information of China (English)

    赵蒙; 宋俊德; 鄂海红

    2013-01-01

    With the development of Internet technology, vast amounts of information presented simultaneously, making it dififcult for users interested in information effectively discover itself, and a lot of network information dark.It is dififcult to be obtained by ordinary users, in order to deal with information overloading problem, there has been personalized user system to compensate for the social tagging system modeling deifciencies, this paper describes the social tagging tag systems, analysis of user interest model representation of the four discussion based on interest in social tagging system modeling method.%随着互联网技术的发展,海量信息同时呈现,使得用户难以有效发现本身感兴趣信息,并且大量的网络暗信息少人问津,难以被普通用户获取,为了处理信息过载问题,出现了个性化用户系统,以弥补海量信息中用户很难找到有用信息的问题。而只有具备了精准的用户兴趣模型,个性化用户系统才得以真正存在。因此用户兴趣建模的研究与探索具有深远的意义。从而,本文首先介绍了社会化标签Tag系统,其次分析了用户兴趣建模的四种表示方法,最后讨论了一种基于社会化标签系统的兴趣建模方法。

  6. Tag cloud generation for results of multiple keywords queries

    DEFF Research Database (Denmark)

    2013-01-01

    In this paper we study tag cloud generation for retrieved results of multiple keyword queries. It is motivated by many real world scenarios such as personalization tasks, surveillance systems and information retrieval tasks defined with multiple keywords. We adjust the state-of-the-art tag cloud...... generation techniques for multiple keywords query results. Consequently, we conduct the extensive evaluation on top of three distinct collaborative tagging systems. The graph-based methods perform significantly better for the Movielens and Bibsonomy datasets. Tag cloud generation based on maximal coverage...

  7. Versatile protein tagging in cells with split fluorescent protein

    OpenAIRE

    Kamiyama, Daichi; Sekine, Sayaka; Barsi-Rhyne, Benjamin; Hu, Jeffrey; Chen, Baohui; Gilbert, Luke A.; Ishikawa, Hiroaki; Leonetti, Manuel D.; Marshall, Wallace F.; Weissman, Jonathan S.; Huang, Bo

    2016-01-01

    In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respec...

  8. Characterization of receptor proteins using affinity cross-linking with biotinylated ligands.

    Science.gov (United States)

    Shinya, Tomonori; Osada, Tomohiko; Desaki, Yoshitake; Hatamoto, Masahiro; Yamanaka, Yuko; Hirano, Hisashi; Takai, Ryota; Che, Fang-Sik; Kaku, Hanae; Shibuya, Naoto

    2010-02-01

    The plant genome encodes a wide range of receptor-like proteins but the function of most of these proteins is unknown. We propose the use of affinity cross-linking of biotinylated ligands for a ligand-based survey of the corresponding receptor molecules. Biotinylated ligands not only enable the analysis of receptor-ligand interactions without the use of radioactive compounds but also the isolation and identification of receptor molecules by a simple affinity trapping method. We successfully applied this method for the characterization, isolation and identification of the chitin elicitor binding protein (CEBiP). A biocytin hydrazide conjugate of N-acetylchitooctaose (GN8-Bio) was synthesized and used for the detection of CEBiP in the plasma or microsomal membrane preparations from rice and carrot cells. Binding characteristics of CEBiP analyzed by inhibition studies were in good agreement with the previous results obtained with the use of a radiolabeled ligand. The biotin-tagged CEBiP could be purified by avidin affinity chromatography and identified by LC-MALDI-MS/MS after tryptic digestion. We also used this method to detect OsFLS2, a rice receptor-like kinase for the perception of the peptide elicitor flg22, in membrane preparations from rice cells overexpressing OsFLS2. This work demonstrates the applicability of this method to the purification and identification of plant receptor proteins.

  9. Security Techniques for Prevention of Rank Manipulation in Social Tagging Services including Robotic Domains

    Directory of Open Access Journals (Sweden)

    Okkyung Choi

    2014-01-01

    Full Text Available With smartphone distribution becoming common and robotic applications on the rise, social tagging services for various applications including robotic domains have advanced significantly. Though social tagging plays an important role when users are finding the exact information through web search, reliability and semantic relation between web contents and tags are not considered. Spams are making ill use of this aspect and put irrelevant tags deliberately on contents and induce users to advertise contents when they click items of search results. Therefore, this study proposes a detection method for tag-ranking manipulation to solve the problem of the existing methods which cannot guarantee the reliability of tagging. Similarity is measured for ranking the grade of registered tag on the contents, and weighted values of each tag are measured by means of synonym relevance, frequency, and semantic distances between tags. Lastly, experimental evaluation results are provided and its efficiency and accuracy are verified through them.

  10. Security techniques for prevention of rank manipulation in social tagging services including robotic domains.

    Science.gov (United States)

    Choi, Okkyung; Jung, Hanyoung; Moon, Seungbin

    2014-01-01

    With smartphone distribution becoming common and robotic applications on the rise, social tagging services for various applications including robotic domains have advanced significantly. Though social tagging plays an important role when users are finding the exact information through web search, reliability and semantic relation between web contents and tags are not considered. Spams are making ill use of this aspect and put irrelevant tags deliberately on contents and induce users to advertise contents when they click items of search results. Therefore, this study proposes a detection method for tag-ranking manipulation to solve the problem of the existing methods which cannot guarantee the reliability of tagging. Similarity is measured for ranking the grade of registered tag on the contents, and weighted values of each tag are measured by means of synonym relevance, frequency, and semantic distances between tags. Lastly, experimental evaluation results are provided and its efficiency and accuracy are verified through them.

  11. Tag-elese or The Language of Tags

    Directory of Open Access Journals (Sweden)

    Jan Simons

    2008-01-01

    Full Text Available The core "meme" of Web 2.0 from which almost all other memes radiated was: 'You control your own data' (O'Reilly, 2005, 3. Key instruments for this user control are tagging systems that allow users to freely assign keywords of their own choosing to Internet resources of their own making as well as to documents produced by others. Of course, freely chosen keywords tags do not necessarily follow prefixed taxonomies or classification systems. But going by the maxim that interaction creates similarity and similarity creates interaction, the idea - or hope - is, however, that the tagging practices of individual users will eventually converge into an emergent common vocabulary or folksonomy (Merholz, 2004; Shirky, 2005; Vander Wal, 2005b; Mika, 2007. It is far from clear, however, that free tagging systems will eventually yield controlled vocabularies, and there are many incentives for idiosyncratic, ambiguous, and inconsistent uses of tags. Left to themselves, free tagging systems seem to be too wild and too chaotic for any order to emerge. But are these free tagging systems really as "feral" as they seem to be, or do they only look uncontrolled because one has been looking for order in the wrong place? I have done a quick-and-dirty" analysis of Flickr's tag cloud. The concept was: if folksonomies encourage users to tap on their own vernacular, everyday natural language must somehow "guide" the tagging practices of users of tagging systems. Flickr's tag cloud has been choosen because it may teach us something about tagging systems and folksonomies, and not - or not primarily - because of what tags may tell us about pictures.

  12. A model-based approach to selection of tag SNPs

    Directory of Open Access Journals (Sweden)

    Sun Fengzhu

    2006-06-01

    Full Text Available Abstract Background Single Nucleotide Polymorphisms (SNPs are the most common type of polymorphisms found in the human genome. Effective genetic association studies require the identification of sets of tag SNPs that capture as much haplotype information as possible. Tag SNP selection is analogous to the problem of data compression in information theory. According to Shannon's framework, the optimal tag set maximizes the entropy of the tag SNPs subject to constraints on the number of SNPs. This approach requires an appropriate probabilistic model. Compared to simple measures of Linkage Disequilibrium (LD, a good model of haplotype sequences can more accurately account for LD structure. It also provides a machinery for the prediction of tagged SNPs and thereby to assess the performances of tag sets through their ability to predict larger SNP sets. Results Here, we compute the description code-lengths of SNP data for an array of models and we develop tag SNP selection methods based on these models and the strategy of entropy maximization. Using data sets from the HapMap and ENCODE projects, we show that the hidden Markov model introduced by Li and Stephens outperforms the other models in several aspects: description code-length of SNP data, information content of tag sets, and prediction of tagged SNPs. This is the first use of this model in the context of tag SNP selection. Conclusion Our study provides strong evidence that the tag sets selected by our best method, based on Li and Stephens model, outperform those chosen by several existing methods. The results also suggest that information content evaluated with a good model is more sensitive for assessing the quality of a tagging set than the correct prediction rate of tagged SNPs. Besides, we show that haplotype phase uncertainty has an almost negligible impact on the ability of good tag sets to predict tagged SNPs. This justifies the selection of tag SNPs on the basis of haplotype

  13. Fast and scalable image auto-tagging

    CERN Document Server

    Frejaville, Camille; Lepetit, Vincent

    Inside Invenio, the web-based integrated system for handling digital libraries developed at CERN, there is a media module, enabling users to upload photos and videos. Especially in CDS, the Invenio instance used at CERN, people use this digital library to upload pictures of official events that took place at CERN. However, so far, there was no way of tagging what’s inside these photos. This project is meant to solve the problem of tagging persons in a photo in an easy and fast way. First, by implementing a complete tagging interface that allows the user to square parts of the photo, resize them, move them and give them a name. Second, by running face detection so that squares already appear on faces and the user just has to fill the title field. Finally, by running a face recognition system that learned from previous tags created by users. In this report, we will show how we implemented the tagging interface, how we improved the existing face detector to make it more efficient, which face detection methods ...

  14. Affine dynamics with torsion

    Energy Technology Data Exchange (ETDEWEB)

    Gueltekin, Kemal [Izmir Institute of Technology, Department of Physics, Izmir (Turkey)

    2016-03-15

    In this study, we give a thorough analysis of a general affine gravity with torsion. After a brief exposition of the affine gravities considered by Eddington and Schroedinger, we construct and analyze different affine gravities based on the determinants of the Ricci tensor, the torsion tensor, the Riemann tensor, and their combinations. In each case we reduce equations of motion to their simplest forms and give a detailed analysis of their solutions. Our analyses lead to the construction of the affine connection in terms of the curvature and torsion tensors. Our solutions of the dynamical equations show that the curvature tensors at different points are correlated via non-local, exponential rescaling factors determined by the torsion tensor. (orig.)

  15. Upper Tag Ontology (UTO) For Integrating Social Tagging Data

    CERN Document Server

    Ding, Ying; Fried, Michael; Toma, Ioan; Yan, Erjia; Foo, Schubert

    2010-01-01

    Data integration and mediation have become central concerns of information technology over the past few decades. With the advent of the Web and the rapid increases in the amount of data and the number of Web documents and users, researchers have focused on enhancing the interoperability of data through the development of metadata schemes. Other researchers have looked to the wealth of metadata generated by bookmarking sites on the Social Web. While several existing ontologies capitalize on the semantics of metadata created by tagging activities, the Upper Tag Ontology (UTO) emphasizes the structure of tagging activities to facilitate modeling of tagging data and the integration of data from different bookmarking sites as well as the alignment of tagging ontologies. UTO is described and its utility in harvesting, modeling, integrating, searching and analyzing data is demonstrated with metadata harvested from three major social tagging systems (Delicious, Flickr and YouTube).

  16. Top tagging with deep neural networks [Vidyo

    CERN Document Server

    CERN. Geneva

    2017-01-01

    Recent literature on deep neural networks for top tagging has focussed on image based techniques or multivariate approaches using high level jet substructure variables. Here, we take a sequential approach to this task by using anordered sequence of energy deposits as training inputs. Unlike previous approaches, this strategy does not result in a loss of information during pixelization or the calculation of high level features. We also propose new preprocessing methods that do not alter key physical quantities such as jet mass. We compare the performance of this approach to standard tagging techniques and present results evaluating the robustness of the neural network to pileup.

  17. Towards an "Intelligent" Tagging Tool for Blogs

    Science.gov (United States)

    Frank, Juraj; Motschnig, Renate; Homola, Martin

    Tagging allows people to effectively organize web resources such as images, bookmarks or blog articles. Things are found easier by browsing tag clouds relying on the tags that have been assigned before. The success is by large determined by the quality and relevance of tags assigned to content - and so it is dependent on people who do the tagging. We investigate mental processes that underlie tagging. In order to improve quality of tagging, we provide guidelines for users of tagging systems and in addition we suggest features that an "intelligent" tagging tool should bear in order to facilitate the tagging process.

  18. Affine and degenerate affine BMW algebras: Actions on tensor space

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2012-01-01

    The affine and degenerate affine Birman-Murakami-Wenzl (BMW) algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic quantum groups and Lie algebras, respectively. Cyclotomic BMW algebras, affine and cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper we explain how the affine and degenerate affine BMW algebras are tantalizers (tensor power centralizer algebras) by defining actions of the affine braid group and the degenerate affine braid algebra on tensor space and showing that, in important cases, these actions induce actions of the affine and degenerate affine BMW algebras. We then exploit the connection to quantum groups and Lie algebras to determine universal parameters for the affine and degenerate affine BMW algebras. Finally, we show that the universal parameters are central elements--the higher Casimir elements for orthogonal and symplectic enveloping algebras and quantum groups.

  19. Social Tagging in a Scholarly Digital Library Environment: Users' Perspectives

    Science.gov (United States)

    Noorhidawati, A.; Hanum, N. Fariza; Zohoorian-Fooladi, N.

    2013-01-01

    Introduction: This paper reports an exploratory study examining how users participate in social tagging activities in a scholarly digital library environment to learn about their motivations, behaviour, and practices. Method: This study was conducted in two phases: a survey to investigate usage and attitudes of social tagging tool, and a…

  20. Improving image segmentation by learning region affinities

    Energy Technology Data Exchange (ETDEWEB)

    Prasad, Lakshman [Los Alamos National Laboratory; Yang, Xingwei [TEMPLE UNIV.; Latecki, Longin J [TEMPLE UNIV.

    2010-11-03

    We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

  1. Imaging mass spectrometer with mass tags

    Science.gov (United States)

    Felton, James S.; Wu, Kuang Jen J.; Knize, Mark G.; Kulp, Kristen S.; Gray, Joe W.

    2013-01-29

    A method of analyzing biological material by exposing the biological material to a recognition element, that is coupled to a mass tag element, directing an ion beam of a mass spectrometer to the biological material, interrogating at least one region of interest area from the biological material and producing data, and distributing the data in plots.

  2. Improving Web Document Clustering through Employing User-Related Tag Expansion Techniques

    Institute of Scientific and Technical Information of China (English)

    Peng Li; Bin Wang; Wei Jin

    2012-01-01

    As high quality descriptors of web page semantics,social annotations or tags have been used for web document clustering and achieved promising results.However,most web pages have few tags (less than 10).This sparsity seriously limits the usage of tags for clustering.In this work,we propose a user-related tag expansion method to overcome this problem,which incorporates additional useful tags into the original tag document by utilizing user tagging data as background knowledge.Unfortunately,simply adding tags may cause topic drift,i.e.,the dominant topic(s) of the original document may be changed.To tackle this problem,we have designed a novel generative model called Folk-LDA,which jointly models original and expanded tags as independent observations.ExPerimental results show that 1) our user-related tag expansion method can be effectively applied to over 90% tagged web documents; 2) Folk-LDA can alleviate topic drift in expansion,especially for those topic-specific documents; 3) the proposed tag-based clustering methods significantly outperform the word-based methods,which indicates that tags could be a better resource for the clustering task.

  3. An updated measurement of sin 2beta with multiple flavor tags using a tag-combining algorithm

    Science.gov (United States)

    Gao, Tianjie

    This thesis reports an updated measurement of Standard Model CP violation parameter sin 2beta using the CDF detector at Fermilab. The signal samples of B0/B¯ 0 → J/psiKS and B0/B¯0 → psi(2 S)KS are extracted from the 110 pb -1 of p-p¯ collisions at s = 1.8 TeV. The flavor of the neutral B mesons is identified at the time of production by three tagging algorithms: a same-side tag, a soft-lepton tag, and an opposite-side-jet tag. The opposite-side-jet tag is a combination of the previously used jet-charge tag and a few new tags. A generic algorithm is developed to combine the tags to improve the tagging performance. A maximum likelihood fitting method is used to determine sin 2beta = 0.91+0.37-0.36 . The uncertainties are improved over the previous CDF measurement. The value is consistent with the standard model prediction of a large positive CP asymmetry in this decay mode. It is also consistent with the world average value of sin 2beta.

  4. An Updated Measurement Of Sin 2beta With Multiple Flavor Tags Using A Tag-combining Algorithm

    CERN Document Server

    Gao, T

    2002-01-01

    This thesis reports an updated measurement of Standard Model CP violation parameter sin 2β using the CDF detector at Fermilab. The signal samples of B0/B¯ 0 → J/ψKS and B0/B¯0 → ψ(2 S)KS are extracted from the 110 pb −1 of p- p¯ collisions at s = 1.8 TeV. The flavor of the neutral B mesons is identified at the time of production by three tagging algorithms: a same-side tag, a soft-lepton tag, and an opposite-side- jet tag. The opposite-side-jet tag is a combination of the previously used jet-charge tag and a few new tags. A generic algorithm is developed to combine the tags to improve the tagging performance. A maximum likelihood fitting method is used to determine sin 2β = 0.91+0.37-0.36 . The uncertainties are improved over the previous CDF measurement. The value is consistent with the standard model prediction of a large positive CP asymmetry in this decay mode. It is also consistent with the world average valu...

  5. Buddy Tag CONOPS and Requirements.

    Energy Technology Data Exchange (ETDEWEB)

    Brotz, Jay Kristoffer [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Deland, Sharon M. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2015-12-01

    This document defines the concept of operations (CONOPS) and the requirements for the Buddy Tag, which is conceived and designed in collaboration between Sandia National Laboratories and Princeton University under the Department of State Key VerificationAssets Fund. The CONOPS describe how the tags are used to support verification of treaty limitations and is only defined to the extent necessary to support a tag design. The requirements define the necessary functions and desired non-functional features of the Buddy Tag at a high level

  6. Preparation of recombinant firefly luciferase by a simple and rapid expression and purification method and its application in bacterial detection

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    A simple and rapid expression and purification method of recombinant firefly luciferase was developed for bacteria detection. A modified luciferase gene from North American firefly Photinus pyralis was cloned into pET28a expression vector and the recombinant protein was produced in Escherichia coli BL21. The recombinant luciferase,equipped with a polyhistidine affinity tag,was purified by immobilized metal ion affinity chromatography (IMAC). The approach generated an abundant expression and an efficient pur...

  7. Online encyclopedia entities tagging method based on page struc-ture and content%融合页面结构与内容的在线百科实体标注方法

    Institute of Scientific and Technical Information of China (English)

    李晓静; 林海伦; 贾岩涛; 王元卓; 程学旗

    2015-01-01

    Online encyclopedia entity tagging aims to label online encyclopedia pages with standard named entity tags such as person, location and organization. It is crucial for a wide range of applications such as entity disambigu-ation, entity relation inference and knowledge base construction and so on. Features of encyclopedia pages can be divided as structure features (e.g., Infobox, title, and category) and content features (i.e., page content). Existing methods that only take one feature or simply combine both features in one classification cause low F1 value. These methods don’t make full use of the difference of these features. This paper presents an online encyclopedia entities tagging method based on page structure and content. This method firstly builds two classifiers with the two kinds of features respectively, and then builds a new classifier by linear combination of these two classifiers, so this method can accurately realize entities tagging. The experimental results show that this method can achieve F1 value improve-ment over the baseline methods on the task of encyclopedia entity tagging.%在线百科实体标注目的是标注出属于特定类别(如人名、地名、机构名等)的实体。百科实体标注对大量的应用,诸如实体消歧、实体关系挖掘、知识库构建都很重要。百科实体特征可以分为结构特征(属性框、标题、类别等)和内容特征(页面正文)。现有的标注方法大多只考虑一种特征或者一种分类器,导致F1值较低,无法充分发挥两种特征的优势。因此,提出了融合页面结构特征和内容特征的在线百科实体标注方法。该方法考虑了两种特征对标注结果的影响,分别构造分类器,并且对结果进行线性组合,能够更准确地实现百科实体的实体标注。实验表明,该方法在实体标注中F1值较其他对比实验方法均有所提高。

  8. Tag Gardening for Folksonomy Enrichment and Maintenance

    Directory of Open Access Journals (Sweden)

    Katrin Weller

    2008-09-01

    Full Text Available As social tagging applications continuously gain in popularity, it becomes more and more accepted that models and tools for (re-organizing tags are needed. Some first approaches are already practically implemented. Recently, activities to edit and organize tags have been described as "tag gardening". We discuss different ways to subsequently revise and reedit tags and thus introduce different "gardening activities"; among them models that allow gradually adding semantic structures to folksonomies and/or that combine them with more complex forms of knowledge organization systems. Moreover, power tags are introduced as tag gardening candidates and the personal tag repository TagCare is presented.

  9. Study of mast cell count in skin tags

    Directory of Open Access Journals (Sweden)

    Zaher Hesham

    2007-01-01

    Full Text Available Background: Skin tags or acrochordons are common tumors of middle-aged and elderly subjects. They consist of loose fibrous tissue and occur mainly on the neck and major flexures as small, soft, pedunculated protrusions. Objectives: The aim was to compare the mast cells count in skin tags to adjacent normal skin in diabetic and nondiabetic participants in an attempt to elucidate the possible role of mast cells in the pathogenesis of skin tags. Participants and Methods: Thirty participants with skin tags were divided into group I (15 nondiabetic participants and group II (15 diabetic participants. Three biopsies were obtained from each participant: a large skin tag, a small skin tag and adjacent normal skin. Mast cell count from all the obtained sections was carried out, and the mast cell density was expressed as the average mast cell count/high power field (HPF. Results: A statistically significant increase in mast cells count in skin tags in comparison to normal skin was detected in group I and group II. There was no statistically significant difference between mast cell counts in skin tags of both the groups. Conclusion: Both the mast cell mediators and hyperinsulinemia are capable of inducing fibroblast proliferation and epidermal hyperplasia that are the main pathologic abnormalities seen in all types of skin tags. However, the presence of mast cells in all examined skin tags regardless of diabetes and obesity may point to the possible crucial role of mast cells in the etiogenesis of skin tags through its interaction with fibroblasts and keratinocytes.

  10. Novel and efficient tag SNPs selection algorithms.

    Science.gov (United States)

    Chen, Wen-Pei; Hung, Che-Lun; Tsai, Suh-Jen Jane; Lin, Yaw-Ling

    2014-01-01

    SNPs are the most abundant forms of genetic variations amongst species; the association studies between complex diseases and SNPs or haplotypes have received great attention. However, these studies are restricted by the cost of genotyping all SNPs; thus, it is necessary to find smaller subsets, or tag SNPs, representing the rest of the SNPs. In fact, the existing tag SNP selection algorithms are notoriously time-consuming. An efficient algorithm for tag SNP selection was presented, which was applied to analyze the HapMap YRI data. The experimental results show that the proposed algorithm can achieve better performance than the existing tag SNP selection algorithms; in most cases, this proposed algorithm is at least ten times faster than the existing methods. In many cases, when the redundant ratio of the block is high, the proposed algorithm can even be thousands times faster than the previously known methods. Tools and web services for haplotype block analysis integrated by hadoop MapReduce framework are also developed using the proposed algorithm as computation kernels.

  11. Affinity Purification and Characterization of a G-Protein Coupled Receptor, Saccharomyces cerevisiae Ste2p

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Byung-Kwon [University of Tennessee, Knoxville (UTK); Jung, Kyung-Sik [University of Tennessee, Knoxville (UTK); Son, Cagdas D [ORNL; Kim, Heejung [University of Tennessee, Knoxville (UTK); Verberkmoes, Nathan C [ORNL; Arshava, Boris [College of Staten Island; Naider, Fred [College of Staten Island; Becker, Jeffrey Marvin [ORNL

    2007-01-01

    We present a rare example of a biologically active G protein coupled receptor (GPCR) whose purity and identity were verified by mass spectrometry after being purified to near homogeneity from its native system. An overexpression vector was constructed to encode the Saccharomyces cerevisiae GPCR -factor receptor (Ste2p, the STE2 gene product) containing a 9-amino acid sequence of rhodopsin that served as an epitope/affinity tag. In the construct, two glycosylation sites and two cysteine residues were removed to aid future structural and functional studies. The receptor was expressed in yeast cells and was detected as a single band in a western blot indicating the absence of glycosylation. Tests of the epitope-tagged, mutated receptor showed it maintained its full biological activity. For extraction of Ste2p, yeast membranes were solubilized with 0.5 % n-dodecyl maltoside (DM). Approximately 120 g of purified -factor receptor was obtained per liter of culture by single-step affinity chromatography using a monoclonal antibody to the rhodopsin epitope. The binding affinity (Kd) of the purified -factor receptor in DM micelles was 28 nM as compared to Kd = 12.7 nM for Ste2p in cell membranes, and approximately 40 % of the purified receptor was correctly folded as judged by ligand saturation binding. About 50 % of the receptor sequence was retrieved from MALDITOF and nanospray mass spectrometry after CNBr digestion of the purified receptor. The methods described will enable structural studies of the -factor receptor and may provide an efficient technique to purify other GPCRs that have been functionally expressed in yeast.

  12. LHCb Tag Collector

    CERN Document Server

    Fuente Fernàndez, P; Cousin, N

    2011-01-01

    The LHCb physics software consists of hundreds of packages, each of which is developed by one or more physicists. When the developers have some code changes that they would like released, they commit them to the version control system, and enter the revision number into a database. These changes have to be integrated into a new release of each of the physics analysis applications. Tests are then performed by a nightly build system, which rebuilds various configurations of the whole software stack and executes a suite of run-time functionality tests. A Tag Collector system has been developed using solid standard technologies to cover both the use cases of developers and integration managers. A simple Web interface, based on an AJAX-like technology, is available. Integration with software management and Nightly Build programs is possible via a Python API. Data are stored in a relational database with the help of an ORM (Object-Relational Mapping) library.

  13. An Overview of Social Tagging and Applications

    Science.gov (United States)

    Gupta, Manish; Li, Rui; Yin, Zhijun; Han, Jiawei

    Social tagging on online portals has become a trend now. It has emerged as one of the best ways of associating metadata with web objects. With the increase in the kinds of web objects becoming available, collaborative tagging of such objects is also developing along new dimensions. This popularity has led to a vast literature on social tagging. In this survey paper, we would like to summarize different techniques employed to study various aspects of tagging. Broadly, we would discuss about properties of tag streams, tagging models, tag semantics, generating recommendations using tags, visualizations of tags, applications of tags, integration of different tagging systems and problems associated with tagging usage. We would discuss topics like why people tag, what influences the choice of tags, how to model the tagging process, kinds of tags, different power laws observed in tagging domain, how tags are created and how to choose the right tags for recommendation. Metadata generated in the form of tags can be efficiently used to improve web search, for web object classification, for generating ontologies, for enhanced browsing etc. We would discuss these applications and conclude with thoughts on future work in the area.

  14. Methyl-CpG island-associated genome signature tags

    Energy Technology Data Exchange (ETDEWEB)

    Dunn, John J

    2014-05-20

    Disclosed is a method for analyzing the organismic complexity of a sample through analysis of the nucleic acid in the sample. In the disclosed method, through a series of steps, including digestion with a type II restriction enzyme, ligation of capture adapters and linkers and digestion with a type IIS restriction enzyme, genome signature tags are produced. The sequences of a statistically significant number of the signature tags are determined and the sequences are used to identify and quantify the organisms in the sample. Various embodiments of the invention described herein include methods for using single point genome signature tags to analyze the related families present in a sample, methods for analyzing sequences associated with hyper- and hypo-methylated CpG islands, methods for visualizing organismic complexity change in a sampling location over time and methods for generating the genome signature tag profile of a sample of fragmented DNA.

  15. HTML Tags as Extraction Cues for Web Page Description Construction

    Directory of Open Access Journals (Sweden)

    Timothy C. Craven

    2003-01-01

    Full Text Available Using four previously identified samples of Web pages containing meta-tagged descriptions, the value of meta-tagged keywords, the first 200 characters of the body, and text marked with common HTML tags as extracts helpful for writing summaries was estimated by applying two measures: density of description words and density of two-word description phrases. Generally, titles and keywords showed the highest densities. Parts of the body showed densities not much different from the body as a whole: somewhat higher for the first 200 characters and for text tagged with "center" and "font"; somewhat lower for text tagged with "a"; not significantly different for "table" and "div". Evidence of non-random clumping of description words in the body of some pages nevertheless suggests that further pursuit of automatic passage extraction methods from the body may be worthwhile. Implications of the findings for aids to summarization, and specifically the TexNet32 package, are discussed.

  16. Semiotic dynamics and collaborative tagging.

    Science.gov (United States)

    Cattuto, Ciro; Loreto, Vittorio; Pietronero, Luciano

    2007-01-30

    Collaborative tagging has been quickly gaining ground because of its ability to recruit the activity of web users into effectively organizing and sharing vast amounts of information. Here we collect data from a popular system and investigate the statistical properties of tag cooccurrence. We introduce a stochastic model of user behavior embodying two main aspects of collaborative tagging: (i) a frequency-bias mechanism related to the idea that users are exposed to each other's tagging activity; (ii) a notion of memory, or aging of resources, in the form of a heavy-tailed access to the past state of the system. Remarkably, our simple modeling is able to account quantitatively for the observed experimental features with a surprisingly high accuracy. This points in the direction of a universal behavior of users who, despite the complexity of their own cognitive processes and the uncoordinated and selfish nature of their tagging activity, appear to follow simple activity patterns.

  17. Engineering the ATLAS TAG Browser

    CERN Document Server

    Zhang, Q; The ATLAS collaboration

    2011-01-01

    ELSSI is a web-based event metadata (TAG) browser and event-level selection service for ATLAS. TAGs from all ATLAS physics and Monte Carlo data sets are routinely loaded into Oracle databases as an integral part of event processing. As data volumes increase, more and more sites are joining the distributed TAG data hosting topology. Meanwhile, TAG content and database schemata continue to evolve as new user requirements and additional sources of metadata emerge. All of this has posed many challenges to the development of ELSSI, which must support vast amounts of TAG data while source, content, geographic locations, and user query patterns may change over time. In this paper, we describe some of the challenges encountered in the process of developing ELSSI, and the software engineering strategies adopted to address those challenges. Approaches to management of access to data, browsing, data rendering, query building, query validation, execution, connection management, and communication with auxiliary services a...

  18. Affine Sphere Relativity

    Science.gov (United States)

    Minguzzi, E.

    2016-11-01

    We investigate spacetimes whose light cones could be anisotropic. We prove the equivalence of the structures: (a) Lorentz-Finsler manifold for which the mean Cartan torsion vanishes, (b) Lorentz-Finsler manifold for which the indicatrix (observer space) at each point is a convex hyperbolic affine sphere centered on the zero section, and (c) pair given by a spacetime volume and a sharp convex cone distribution. The equivalence suggests to describe (affine sphere) spacetimes with this structure, so that no algebraic-metrical concept enters the definition. As a result, this work shows how the metric features of spacetime emerge from elementary concepts such as measure and order. Non-relativistic spacetimes are obtained replacing proper spheres with improper spheres, so the distinction does not call for group theoretical elements. In physical terms, in affine sphere spacetimes the light cone distribution and the spacetime measure determine the motion of massive and massless particles (hence the dispersion relation). Furthermore, it is shown that, more generally, for Lorentz-Finsler theories non-differentiable at the cone, the lightlike geodesics and the transport of the particle momentum over them are well defined, though the curve parametrization could be undefined. Causality theory is also well behaved. Several results for affine sphere spacetimes are presented. Some results in Finsler geometry, for instance in the characterization of Randers spaces, are also included.

  19. Affine stochastic mortality

    NARCIS (Netherlands)

    D.F. Schrager

    2006-01-01

    We propose a new model for stochastic mortality. The model is based on the literature on affine term structure models. It satisfies three important requirements for application in practice: analytical tractibility, clear interpretation of the factors and compatibility with financial option pricing m

  20. Design and Analysis of Salmonid Tagging Studies in the Columbia Basin, Volume XXI; A Summary of Methods for Conducting Salmonid Fry Mark-Recapture Studies for Estimating Survival in Tributaries, Technical Report 2005-2006.

    Energy Technology Data Exchange (ETDEWEB)

    Skalski, John

    2007-02-01

    Productivity and early fry survival can have a major influence on the dynamics of fish stocks. To investigate the early life history of fish, numerous methods have been developed or adapted to these much smaller fish. Some of the marking techniques provide individual identification; many others, only class identification. Some of the tagging techniques require destructive sampling to identify a mark; other methods permit benign examination and rerelease of captured fish. Sixteen alternative release-recapture designs for conducting fry survival investigations were examined. Eleven approaches were found capable of estimating survival parameters; five were not. Of those methods capable of estimating fry survival, five required unique marks, four required batch-specific marks, and two approaches required remarking and rereleasing captured fry. No approach based on a simple batch mark was capable of statistically estimating survival.

  1. Versatile protein tagging in cells with split fluorescent protein

    Science.gov (United States)

    Kamiyama, Daichi; Sekine, Sayaka; Barsi-Rhyne, Benjamin; Hu, Jeffrey; Chen, Baohui; Gilbert, Luke A.; Ishikawa, Hiroaki; Leonetti, Manuel D.; Marshall, Wallace F.; Weissman, Jonathan S.; Huang, Bo

    2016-01-01

    In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respectively. The small size of FP11-tags enables a cost-effective and scalable way to insert them into endogenous genomic loci via CRISPR-mediated homology-directed repair. Tandem arrangement FP11-tags allows proportional enhancement of fluorescence signal in tracking intraflagellar transport particles, or reduction of photobleaching for live microtubule imaging. Finally, we show the utility of tandem GFP11-tag in scaffolding protein oligomerization. These experiments illustrate the versatility of FP11-tag as a labelling tool as well as a multimerization-control tool for both imaging and non-imaging applications. PMID:26988139

  2. Versatile protein tagging in cells with split fluorescent protein.

    Science.gov (United States)

    Kamiyama, Daichi; Sekine, Sayaka; Barsi-Rhyne, Benjamin; Hu, Jeffrey; Chen, Baohui; Gilbert, Luke A; Ishikawa, Hiroaki; Leonetti, Manuel D; Marshall, Wallace F; Weissman, Jonathan S; Huang, Bo

    2016-03-18

    In addition to the popular method of fluorescent protein fusion, live cell protein imaging has now seen more and more application of epitope tags. The small size of these tags may reduce functional perturbation and enable signal amplification. To address their background issue, we adapt self-complementing split fluorescent proteins as epitope tags for live cell protein labelling. The two tags, GFP11 and sfCherry11 are derived from the eleventh β-strand of super-folder GFP and sfCherry, respectively. The small size of FP11-tags enables a cost-effective and scalable way to insert them into endogenous genomic loci via CRISPR-mediated homology-directed repair. Tandem arrangement FP11-tags allows proportional enhancement of fluorescence signal in tracking intraflagellar transport particles, or reduction of photobleaching for live microtubule imaging. Finally, we show the utility of tandem GFP11-tag in scaffolding protein oligomerization. These experiments illustrate the versatility of FP11-tag as a labelling tool as well as a multimerization-control tool for both imaging and non-imaging applications.

  3. Folksonomic Tag Clouds as an Aid to Content Indexing

    CERN Document Server

    Harvey, Morgan; Ruthven, Ian; Elsweiler, David

    2009-01-01

    Social tagging systems have recently developed as a popular method of data organisation on the Internet. These systems allow users to organise their content in a way that makes sense to them, rather than forcing them to use a pre-determined and rigid set of categorisations. These folksonomies provide well populated sources of unstructured tags describing web resources which could potentially be used as semantic index terms for these resources. However getting people to agree on what tags best describe a resource is a difficult problem, therefore any feature which increases the consistency and stability of terms chosen would be extremely beneficial. We investigate how the provision of a tag cloud, a weighted list of terms commonly used to assist in browsing a folksonomy, during the tagging process itself influences the tags produced and how difficult the user perceived the task to be. We show that illustrating the most popular tags to users assists in the tagging process and encourages a stable and consistent ...

  4. Fully printed flexible and disposable wireless cyclic voltammetry tag.

    Science.gov (United States)

    Jung, Younsu; Park, Hyejin; Park, Jin-Ah; Noh, Jinsoo; Choi, Yunchang; Jung, Minhoon; Jung, Kyunghwan; Pyo, Myungho; Chen, Kevin; Javey, Ali; Cho, Gyoujin

    2015-01-29

    A disposable cyclic voltammetry (CV) tag is printed on a plastic film by integrating wireless power transmitter, polarized triangle wave generator, electrochemical cell and signage through a scalable gravure printing method. By proximity of 13.56 MHz RF reader, the printed CV tag generates 320 mHz of triangular sweep wave from +500 mV to -500 mV which enable to scan a printed electrochemical cell in the CV tag. By simply dropping any specimen solution on the electrochemical cell in the CV tag, the presence of solutes in the solution can be detected and shown on the signage of the CV tag in five sec. 10 mM of N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD) was used as a standard solute to prove the working concept of fully printed disposable wireless CV tag. Within five seconds, we can wirelessly diagnose the presence of TMPD in the solution using the CV tag in the proximity of the 13.56 MHz RF reader. This fully printed and wirelessly operated flexible CV tag is the first of its kind and marks the path for the utilization of inexpensive and disposable wireless electrochemical sensor systems for initial diagnose hazardous chemicals and biological molecules to improve public hygiene and health.

  5. The Cutting Edge of Affinity Electrophoresis Technology

    Directory of Open Access Journals (Sweden)

    Eiji Kinoshita

    2015-03-01

    Full Text Available Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years.

  6. The Cutting Edge of Affinity Electrophoresis Technology

    Science.gov (United States)

    Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

    2015-01-01

    Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years.

  7. A new affinity method for purification of bovine testicular hyaluronidase enzyme and an investigation of the effects of some compounds on this enzyme.

    Science.gov (United States)

    Kaya, Mustafa Oguzhan; Arslan, Oktay; Guler, Ozen Ozensoy

    2015-01-01

    In this study, a new affinity gel for the purification of bovine testicular hyaluronidase (BTH) was synthesized. L-Tyrosine was added as the extension arm to the Sepharose-4B activated with cyanogen bromide. m-Anisidine is a specific inhibitor of BTH enzyme. m-Anisidine was clamped to the newly formed Sepharose-4B-L-tyrosine as a ligand. As a result, an affinity gel having the chemical structure of Sepharose-4B-L-tyrosine-m-anisidine was obtained. BTH purified by ammonium sulfate precipitation and affinity chromatography was obtained with a 16.95% yield and 881.78 degree of purity. The kinetic constants K(M) and V(Max) for BTH were determined by using hyaluronic acid as a substrate. K(M) and V(Max) values obtained from the Lineweaver-Burk graph were found to be 2.23 mM and 19.85 U/mL, respectively. In vitro effects of some chemicals were determined on purified BTH enzyme. Some chemically active ingredients were 1,1-dimethyl piperidinium chloride, β-naphthoxyacetic acid and gibberellic acid. Gibberellic acid showed the best inhibition effect on BTH.

  8. Using an Explicit Emission Tagging Method in Global Modeling of Source-Receptor Relationships for Black Carbon in the Arctic: Variations, Sources and Transport Pathways

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Hailong; Rasch, Philip J.; Easter, Richard C.; Singh, Balwinder; Zhang, Rudong; Ma, Po-Lun; Qian, Yun; Ghan, Steven J.; Beagley, Nathaniel

    2014-11-27

    We introduce an explicit emission tagging technique in the Community Atmosphere Model to quantify source-region-resolved characteristics of black carbon (BC), focusing on the Arctic. Explicit tagging of BC source regions without perturbing the emissions makes it straightforward to establish source-receptor relationships and transport pathways, providing a physically consistent and computationally efficient approach to produce a detailed characterization of the destiny of regional BC emissions and the potential for mitigation actions. Our analysis shows that the contributions of major source regions to the global BC burden are not proportional to the respective emissions due to strong region-dependent removal rates and lifetimes, while the contributions to BC direct radiative forcing show a near-linear dependence on their respective contributions to the burden. Distant sources contribute to BC in remote regions mostly in the mid- and upper troposphere, having much less impact on lower-level concentrations (and deposition) than on burden. Arctic BC concentrations, deposition and source contributions all have strong seasonal variations. Eastern Asia contributes the most to the wintertime Arctic burden. Northern Europe emissions are more important to both surface concentration and deposition in winter than in summer. The largest contribution to Arctic BC in the summer is from Northern Asia. Although local emissions contribute less than 10% to the annual mean BC burden and deposition within the Arctic, the per-emission efficiency is much higher than for major non-Arctic sources. The interannual variability (1996-2005) due to meteorology is small in annual mean BC burden and radiative forcing but is significant in yearly seasonal means over the Arctic. When a slow aging treatment of BC is introduced, the increase of BC lifetime and burden is source-dependent. Global BC forcing-per-burden efficiency also increases primarily due to changes in BC vertical distributions. The

  9. Sediment tracing from small torrential channels to gravel-bed rivers using pit tags method. A case study from the upper Guil catchment.

    Science.gov (United States)

    Graff, Kévin; Viel, Vincent; Carlier, Benoit; Lissak, Candide; Arnaud-Fassetta, Gilles; Fort, Monique; Madelin, Malika

    2016-04-01

    In mountainous areas, especially in large catchments with torrential tributaries, the production and sediment transport significantly increase flood impacts in the valley bottoms. The quantification and characterisation of sedimentary transfers are therefore major challenges to provide better flood risk management. As a part of SAMCO (ANR 12 SENV-0004 SAMCO) project, for mountain hazard assessment in a context of global changes, we tried to improve the knowledge of these hydromorphological systems at both spatial and temporal scales, by identifying sediment supply and sediment dynamics from torrential tributaries to the main channel. A sediment budget was used as a tool for quantifying erosion, transport and deposition processes. This research is focused on the upper Guil catchment (Queyras, Southern French Alps - 317 km2) entrenched in "schistes lustrés" and ophiolitic bedrock. This catchment is prone to catastrophic summer floods [June 1957 (>R.I. 100 yr), June 2000 (R.I. 30 yr)] characterised by huge sediment transport from tributaries to downvalley, very much facilitated by strong hillslope-channel connectivity (about 12,000 m3 volume of sediment aggraded in the Peyronnelle fan during the June 2000 RI-30 year flood event). We intend to highlight sediment dynamics on small torrential channels and its connection with gravel-bed streams. Four study sites characterised by avalanche and debris flow-dominated channels located in the upper Guil catchment were investigated. In order to better assess sediment movement, we used the pit-tags technique. In total, 560 pit-tags (pt) have been implemented in four catchments: Peyronnelle (320pt), Combe Morel (40pt), Bouchouse (120pt), and Maloqueste (80pt). Distances and trajectories of gravels sediments have been monitored since two years during summer periods. We specifically describe results obtained along the Peyronnelle channel affected by a large debris-flow during august 2015. Data are used to discuss lag time

  10. RAC-tagging: Recombineering And Cas9-assisted targeting for protein tagging and conditional analyses.

    Science.gov (United States)

    Baker, Oliver; Gupta, Ashish; Obst, Mandy; Zhang, Youming; Anastassiadis, Konstantinos; Fu, Jun; Stewart, A Francis

    2016-05-24

    A fluent method for gene targeting to establish protein tagged and ligand inducible conditional loss-of-function alleles is described. We couple new recombineering applications for one-step cloning of gRNA oligonucleotides and rapid generation of short-arm (~1 kb) targeting constructs with the power of Cas9-assisted targeting to establish protein tagged alleles in embryonic stem cells at high efficiency. RAC (Recombineering And Cas9)-tagging with Venus, BirM, APEX2 and the auxin degron is facilitated by a recombineering-ready plasmid series that permits the reuse of gene-specific reagents to insert different tags. Here we focus on protein tagging with the auxin degron because it is a ligand-regulated loss-of-function strategy that is rapid and reversible. Furthermore it includes the additional challenge of biallelic targeting. Despite high frequencies of monoallelic RAC-targeting, we found that simultaneous biallelic targeting benefits from long-arm (>4 kb) targeting constructs. Consequently an updated recombineering pipeline for fluent generation of long arm targeting constructs is also presented.

  11. Recombinant Staphylococcal protein A with cysteine residue for preparation of affinity chromatography stationary phase and immunosensor applications

    Directory of Open Access Journals (Sweden)

    Gorbatiuk O. B.

    2015-04-01

    Full Text Available Aim. Engineering of recombinant Staphylococcal protein A with cysteine residue (SPA-Cys for preparation of affinity chromatography stationary phase and formation of bioselective element of immunosensor. Methods. DNA sequences encoding IgG-binding region of SPA, His-tag and cysteine were genetically fused and expressed in E. coli. SPA-Cys was immobilized on maleimide-functionalized silica beads for affinity chromatography stationary phase preparation and on a gold sensor surface as a bioselective element of immunosensor. Results. SPA-Cys was expressed at a high-level in a soluble form. The target protein was purified and showed a high IgG-binding activity. The capacity of the obtained SPA-Cys-based affinity chromatography stationary phase was 10–12 mg of IgG /ml. The purity of eluted IgG was more than 95 % in one-step purification procedure. The developed SPA-Cys-based bioselective element of immunosensor selectively interacted with human IgG and did not interact with the control proteins. Conclusions. The recombinant Staphylococcal protein A with cysteine residue was successfully used for the preparation of affinity chromatography stationary phase and formation of the bioselective element of immunosensor.

  12. Web image annotation using two-step filtering on social tags

    Science.gov (United States)

    Cho, Sunyoung; Cha, Jaeseong; Byun, Hyeran

    2011-03-01

    Web image annotation has become an important issue with exploding web images and the necessity of effective image search. The social tags have recently utilized at image annotation because they can reflect the user's tagging tendency, and reduce the semantic gap. However, an effective filtering procedure is required to extract the relevant tags since the user's subjectivity and noisy tags. In this paper, we propose a two-step filtering on social tags for image annotation. This method conducts the filtering and verification tasks by analyzing the tags of visual neighbor images using voting method and co-occurrence analysis. Our method consists of the following three steps: 1) the tag candidate set is founded by searching the visual neighbor images, 2) from a given tag candidate set, coarse filtering is conducted by tag grouping and voting technique, 3) the dense filtering is conducted by using similarity verification for coarse filtered candidate tag set. To evaluate the performance of our approach, we conduct the experiments on a social-tagged image dataset obtained from Flickr. We compare the annotation accuracy between the voting method and our proposed method. Our experimental results show that our method has an improvement in image annotation.

  13. Affine and degenerate affine BMW algebras: The center

    CERN Document Server

    Daugherty, Zajj; Virk, Rahbar

    2011-01-01

    The degenerate affine and affine BMW algebras arise naturally in the context of Schur-Weyl duality for orthogonal and symplectic Lie algebras and quantum groups, respectively. Cyclotomic BMW algebras, affine Hecke algebras, cyclotomic Hecke algebras, and their degenerate versions are quotients. In this paper the theory is unified by treating the orthogonal and symplectic cases simultaneously; we make an exact parallel between the degenerate affine and affine cases via a new algebra which takes the role of the affine braid group for the degenerate setting. A main result of this paper is an identification of the centers of the affine and degenerate affine BMW algebras in terms of rings of symmetric functions which satisfy a "cancellation property" or "wheel condition" (in the degenerate case, a reformulation of a result of Nazarov). Miraculously, these same rings also arise in Schubert calculus, as the cohomology and K-theory of isotropic Grassmanians and symplectic loop Grassmanians. We also establish new inte...

  14. Affinity driven social networks

    Science.gov (United States)

    Ruyú, B.; Kuperman, M. N.

    2007-04-01

    In this work we present a model for evolving networks, where the driven force is related to the social affinity between individuals of a population. In the model, a set of individuals initially arranged on a regular ordered network and thus linked with their closest neighbors are allowed to rearrange their connections according to a dynamics closely related to that of the stable marriage problem. We show that the behavior of some topological properties of the resulting networks follows a non trivial pattern.

  15. Purely affine Gravity

    CERN Document Server

    Skirzewski, Aureliano

    2014-01-01

    We develop a topological theory of gravity with torsion where metric has a dynamical rather than a kinematical origin. This approach towards gravity resembles pre-geometrical approaches in which a fundamental metric does not exist, but the affine connection gives place to a local inertial structure. Such feature reminds us of Mach's principle, that assumes the inertial forces should have dynamical origin. Additionally, a Newtonian gravitational force is obtained in the non-relativistic limit of the theory.

  16. Search for the Higgs boson in the ZH→v$\\bar{v}$b$\\bar{b}$ channel: Development of a b-tagging method based on soft muons

    Energy Technology Data Exchange (ETDEWEB)

    Jamin, David [Univ. of the Mediterranean, Marseille (France)

    2010-09-30

    In the Standard Model of particle physics, the Higgs boson generates elementary particle masses. Current theoretical and experimental constraints lead to a Higgs boson mass between 114.4 and 158 GeV with 95% confidence level. Moreover, Tevatron has recently excluded the mass ranges between 100 and 109 GeV, 158 and 175 GeV with 95% confidence level. These results gives a clear indication to search for a Higgs boson at low mass. The D0 detector is located near Chicago, at the Tevatron, a proton-antiproton collider with an energy in the center of mass of 1.96 TeV. The topic of this thesis is the search for a Higgs boson in association with a Z boson. This channel is sensitive to low mass Higgs boson (<135 GeV) which has a branching ratio H → bb varies between 50% and 90% in this mass range. The decay channel ZH → v$\\bar{v}$b$\\bar{b}$ studied has in the final state 2 heavy-flavor jets and some missing transverse energy due to escaping neutrinos. The heavy-flavor jets identification ('b-tagging') is done with a new algorithm (SLTNN) developed specifically for semi-leptonic decay of b quarks. The Higgs boson search analysis was performed with 3 fb-1 of data. The use of SLTNN increases by 10% the Higgs boson signal efficiency. The global analysis sensitivity improvement, however, is rather low (<1%) after taking into account the backgrounds and systematic uncertainties.

  17. Extraction of haemoglobin from human blood by affinity precipitation using a haptoglobin-based stimuli-responsive affinity macroligand.

    Science.gov (United States)

    Stocker-Majd, Gisela; Hilbrig, Frank; Freitag, Ruth

    2008-06-13

    Affinity precipitation was compared to affinity chromatography and batch adsorption as the final purification step in a protocol for the isolation of haemoglobin from human blood. Haptoglobin was the affinity ligand. The first steps on the process were realized by traditional methods (lyses of red blood cells followed by ammonium sulphate precipitation). For affinity chromatography (and batch adsorption) the ligand was linked to Sepharose, for affinity precipitation to a thermoresponsive polymer, namely poly(N-isopropylacrylamide). Five haptoglobin-poly(N-isopropylacrylamide) bioconjugates (affinity macroligands) were constructed with different polymer: haptoglobin-coupling ratios. Conjugation of haptoglobin to the soluble poly(N-isopropylacrylamide) apparently does not change the interaction thermodynamics with haemoglobin, as the haemoglobin binding constants calculated by a Scatchard analysis for the affinity macroligand were of the same order of magnitude as those described in the literature for the haemoglobin-haptoglobin complex in solution. Two elution protocols were used for haemoglobin release from the various affinity materials, one at pH 2, the other with 5 M urea at pH 11. Both affinity chromatography and affinity precipitation yielded a pure haemoglobin of high quality. Compared to the affinity chromatography, affinity precipitation showed a significantly higher ligand efficiency (ratio of the experimental capacity to the theoretical one). The method thus makes better use of the expensive affinity ligands. As affinity precipitation only requires small temperature changes to bring about precipitation/redissolution of the affinity complexes and a centrifugation step for recovery of the precipitate, the method in addition has advantages in term of scalability and simplicity.

  18. An approach to sequence DNA without tagging

    Science.gov (United States)

    Niu, Sanjun; Saraf, Ravi F.

    2002-10-01

    Microarray technology is playing an increasingly important role in biology and medicine and its application to genomics for gene expression analysis has already reached the market with a variety of commercially available instruments. In these combinatorial analysis methods, known probe single-strand DNA (ssDNA) 'primers' are attached in clusters of typically 100 µm × 100 µm pixels. Each pixel of the array has a slightly different sequence. On exposure to 'unknown' target ssDNA, the pixels with the right complementary probe ssDNA sequence convert to double-stranded DNA (dsDNA) by a hybridization reaction. To transduct the conversion of the pixel to dsDNA, the target ssDNA is labelled with a photoluminescent tag during the polymerase chain reaction (PCR) amplification process. Due to the statistical distribution of the tags in the target ssDNA, it becomes significantly difficult to implement these methods as a diagnostic tool in a pathology laboratory. A method to sequence DNA without tagging the molecule is developed. The fabrication process is compatible with current microelectronics and (emerging) soft-material fabrication technologies, allowing the method to be integrable with micro-electromechanical systems (MEMS) and lab-on-a-chip devices. An estimated sensitivity of 10-12 g on a 1 cm2 device area is obtained.

  19. Affine morphisms at zero level

    CERN Document Server

    Das, Paramita; Gupta, Ved Prakash

    2010-01-01

    Given a finite index subfactor, we show that the {\\em affine morphisms at zero level} in the affine category over the planar algebra associated to the subfactor is isomorphic to the fusion algebra of the subfactor as a *-algebra.

  20. On the Affine Isoperimetric Inequalities

    Indian Academy of Sciences (India)

    Wuyang Yu; Gangsong Leng

    2011-11-01

    We obtain an isoperimetric inequality which estimate the affine invariant -surface area measure on convex bodies. We also establish the reverse version of -Petty projection inequality and an affine isoperimetric inequality of $_{-p}K$.

  1. Device-free object tracking using passive tags

    CERN Document Server

    Han, Jinsong; Zhao, Kun; Jiang, Zhiping

    2014-01-01

    This SpringerBrief examines the use of cheap commercial passive RFID tags to achieve accurate device-free object-tracking. It presents a sensitive detector, named Twins, which uses a pair of adjacent passive tags to detect uncooperative targets (such as intruders). Twins leverages a newly observed phenomenon called critical state that is caused by interference among passive tags.The author expands on the previous object tracking methods, which are mostly device-based, and reveals a new interference model and their extensive experiments for validation. A prototype implementation of the Twins-ba

  2. Flavor Tagging at the Tevatron, including calibration and control

    CERN Document Server

    Moulik, T

    2007-01-01

    This report summarizes the flavor tagging techniques developed at the CDF and D{\\O}experiments. Flavor tagging involves identification of the B meson flavor atproduction, whether its constituent is a quark or an anti-quark. It is crucial for measuring the oscillation frequency of neutral B mesons, both in the B^0 and B_S system. The two experiments have developed their unique approaches to flavor tagging, using neural networks, and likelihood methods to disentangle tracks from $b$ decays from other tracks. This report discusses these techniques and the measurement of B^0 mixing, as a means to calibrate the taggers.

  3. Influence of Histidine-Containing Tags on the Biodistribution of ADAPT Scaffold Proteins.

    Science.gov (United States)

    Lindbo, Sarah; Garousi, Javad; Åstrand, Mikael; Honarvar, Hadis; Orlova, Anna; Hober, Sophia; Tolmachev, Vladimir

    2016-03-16

    Engineered scaffold proteins (ESP) are high-affinity binders that can be used as probes for radionuclide imaging. Histidine-containing tags enable both efficient purification of ESP and radiolabeling with (99m)Tc(CO)3. Earlier studies demonstrated that the use of a histidine-glutamate-histidine-glutamate-histidine-glutamate (HE)3-tag instead of the commonly used hexahistidine (H6)-tag reduces hepatic uptake of radiolabeled ESP and short peptides. Here, we investigated the influence of histidine-containing tags on the biodistribution of a novel type of ESP, ADAPTs. A series of anti-HER2 ADAPT probes having H6- or (HE)3-tags in the N-termini were prepared. The constructs, (HE)3-ADAPT6 and H6-ADAPT6, were labeled with two different nuclides, (99m)Tc or (111)In. The labeling with (99m)Tc(CO)3 utilized the histidine-containing tags, while (111)In was attached through a maleimido derivative of DOTA conjugated to the N-terminus. For (111)In-labeled ADAPTs, the use of (HE)3 provided a significantly (p histidine-containing tags on the biodistribution of the novel ADAPT scaffold proteins was different compared to its influence on other ESPs studied so far. Apparently, the effect of a histidine-containing tag on the biodistribution is highly dependent on the scaffold composition of the ESP.

  4. Compilation of HPSG to TAG

    CERN Document Server

    Kasper, R; Netter, K; Vijay-Shanker, K; Kasper, Robert; Kiefer, Bernd; Netter, Klaus

    1995-01-01

    We present an implemented compilation algorithm that translates HPSG into lexicalized feature-based TAG, relating concepts of the two theories. While HPSG has a more elaborated principle-based theory of possible phrase structures, TAG provides the means to represent lexicalized structures more explicitly. Our objectives are met by giving clear definitions that determine the projection of structures from the lexicon, and identify maximal projections, auxiliary trees and foot nodes.

  5. Affine Patches on Positroid Varieties and Affine Pipe Dreams (Thesis)

    CERN Document Server

    Snider, Michelle

    2010-01-01

    The objects of interest in this thesis are positroid varieties in the Grassmannian, which are indexed by juggling patterns. In particular, we study affine patches on these positroid varieties. Our main result corresponds these affine patches to Kazhdan-Lusztig varieties in the affine Grassmannian. We develop a new term order and study how these spaces are related to subword complexes and Stanley-Reisner ideals. We define an extension of pipe dreams to the affine case and conclude by showing how our affine pipe dreams are generalizations of Cauchon and Le diagrams.

  6. Affine and quasi-affine frames for rational dilations

    DEFF Research Database (Denmark)

    Bownik, Marcin; Lemvig, Jakob

    2011-01-01

    , the corresponding family of quasi-affine systems are frames with uniform frame bounds. We also prove a similar equivalence result between pairs of dual affine frames and dual quasi-affine frames. Finally, we uncover some fundamental differences between the integer and rational settings by exhibiting an example......In this paper we extend the investigation of quasi-affine systems, which were originally introduced by Ron and Shen [J. Funct. Anal. 148 (1997), 408-447] for integer, expansive dilations, to the class of rational, expansive dilations. We show that an affine system is a frame if, and only if...

  7. Assessment of electron-vibrational interaction (EVI) parameters of YAG:Ce3 +, TAG:Ce3 + and LuAG:Ce3 + garnet phosphors by spectrum fitting method

    Science.gov (United States)

    Nair, Govind B.; Dhoble, S. J.

    2017-02-01

    The electron-vibrational interaction (EVI) in 4f ↔ 5d optical transitions of Ce3 + ions in YAG, TAG and LuAG garnet phosphors have been analysed in this work. The main EVI parameters that have been estimated and reported here are Huang-Rhys factor, effective phonon energy, Stokes shift, red shift and Zero-phonon line position. The EVI parameters were estimated from the room temperature photoluminescence results that were recently reported. The spectrum fitting method was employed to determine the EVI parameters. An emission band was modelled with the aid of the calculated EVI parameters. The agreement between the modelled emission bands with the experimentally obtained ones validated the estimated values of EVI parameters.

  8. A simplified method for purification of recombinant soluble DnaA proteins.

    Science.gov (United States)

    Zawilak-Pawlik, Anna M; Kois, Agnieszka; Zakrzewska-Czerwinska, Jolanta

    2006-07-01

    An improved, simplified method for the purification of recombinant, tagged DnaA proteins is described. The presented protocol allowed us to purify soluble DnaA proteins from two different bacterial species: Helicobacter pylori and Streptomyces coelicolor, but it can most likely also be used for the isolation of DnaA proteins from other bacteria, as it was adapted for Mycobacterium tuberculosis DnaA. The isolation procedure consists of protein precipitation with ammonium sulphate followed by affinity chromatography. The composition of the buffers used at each purification step is crucial for the successful isolation of the recombinant DnaA proteins. The universality of the method in terms of its application to differently tagged proteins (His-tagged or GST-tagged) as well as different properties of purified proteins (e.g., highly aggregating truncated forms) makes the protocol highly useful for all studies requiring purified and active DnaA proteins.

  9. Sex identification and PIT-tagging: tools and prospects for studying intersexual differences in freshwater fishes.

    Science.gov (United States)

    Hulthén, K; Chapman, B B; Nilsson, P A; Hansson, L-A; Skov, C; Baktoft, H; Brodersen, J; Brönmark, C

    2014-02-01

    This study evaluated a technique to allow the long-term monitoring of individual fishes of known sex in the wild using sex confirmation in close proximity to the reproductive period combined with individual tagging. Hundreds of partially migratory roach Rutilus rutilus were tagged with passive integrated transponders (PIT) following sex determination in spring and various performance measures were compared with fish tagged outside the reproductive period in autumn. Short-term survival was >95% for R. rutilus sexed and tagged under natural field conditions. Total length (LT ) did not affect the probability of survival within the size range tagged (119-280 mm), nor were there differences in timing of migration the following season between individuals sexed and tagged in spring and individuals tagged in autumn (i.e. outside the reproductive period). Also, a similar per cent of R. rutilus sexed and tagged in spring and tagged in autumn migrated the following season (34·5 and 34·7%). Moreover, long-term recapture data revealed no significant differences in body condition between R. rutilus individuals sexed and tagged in spring, individuals tagged in autumn and unmanipulated individuals. The observed sex ratio of recaptured fish did not differ from the expected values of equal recapture rates between males and females. Hence, there is no observable evidence for an adverse effect of tagging close to the reproductive period and therefore this method is suitable for studying intersexual differences and other phenotypic traits temporarily expressed during reproduction at the individual level in fishes.

  10. The Purification of Natural and Recombinant Peptide Antibodies by Affinity Chromatographic Strategies.

    Science.gov (United States)

    Ma, Hui; O'Kennedy, Richard

    2015-01-01

    The purification of peptide antibodies (e.g., IgG, IgY, scFv, and Fab) are described in this chapter. Affinity chromatographic purification, a very convenient and effective antibody purification strategy, is used to isolate peptide antibodies based on specific binding, i.e., binding of the antibody to a column on which its specific ligand is immobilized with subsequent elution of the purified antibody. In addition, the application of purification methods based on the use of proteins A, G, and L, each of which bind to specific domains on an antibody/fragment, or the use of specific tags (e.g., histidine and biotin) attached to antibodies or antigens are also described.

  11. Peptide nucleic acid probe for protein affinity purification based on biotin-streptavidin interaction and peptide nucleic acid strand hybridization.

    Science.gov (United States)

    Tse, Jenny; Wang, Yuanyuan; Zengeya, Thomas; Rozners, Eriks; Tan-Wilson, Anna

    2015-02-01

    We describe a new method for protein affinity purification that capitalizes on the high affinity of streptavidin for biotin but does not require dissociation of the biotin-streptavidin complex for protein retrieval. Conventional reagents place both the selectively reacting group (the "warhead") and the biotin on the same molecule. We place the warhead and the biotin on separate molecules, each linked to a short strand of peptide nucleic acid (PNA), synthetic polymers that use the same bases as DNA but attached to a backbone that is resistant to attack by proteases and nucleases. As in DNA, PNA strands with complementary base sequences hybridize. In conditions that favor PNA duplex formation, the warhead strand (carrying the tagged protein) and the biotin strand form a complex that is held onto immobilized streptavidin. As in DNA, the PNA duplex dissociates at moderately elevated temperature; therefore, retrieval of the tagged protein is accomplished by a brief exposure to heat. Using iodoacetate as the warhead, 8-base PNA strands, biotin, and streptavidin-coated magnetic beads, we demonstrate retrieval of the cysteine protease papain. We were also able to use our iodoacetyl-PNA:PNA-biotin probe for retrieval and identification of a thiol reductase and a glutathione transferase from soybean seedling cotyledons.

  12. Preparation of silica coated cobalt ferrite magnetic nanoparticles for the purification of histidine-tagged proteins

    Science.gov (United States)

    Aygar, Gülfem; Kaya, Murat; Özkan, Necati; Kocabıyık, Semra; Volkan, Mürvet

    2015-12-01

    Surface modified cobalt ferrite (CoFe2O4) nanoparticles containing Ni-NTA affinity group were synthesized and used for the separation of histidine tag proteins from the complex matrices through the use of imidazole side chains of histidine molecules. Firstly, CoFe2O4 nanoparticles with a narrow size distribution were prepared in an aqueous solution using the controlled co-precipitation method. In order to obtain small CoFe2O4 agglomerates, oleic acid and sodium chloride were used as dispersants. The CoFe2O4 particles were coated with silica and subsequently the surface of these silica coated particles (SiO2-CoFe2O4) was modified by amine (NH2) groups in order to add further functional groups on the silica shell. Then, carboxyl (-COOH) functional groups were added to the SiO2-CoFe2O4 magnetic nanoparticles through the NH2 groups. After that Nα,Nα-Bis(carboxymethyl)-L-lysine hydrate (NTA) was attached to carboxyl ends of the structure. Finally, the surface modified nanoparticles were labeled with nickel (Ni) (II) ions. Furthermore, the modified SiO2-CoFe2O4 magnetic nanoparticles were utilized as a new system that allows purification of the N-terminal His-tagged recombinant small heat shock protein, Tpv-sHSP 14.3.

  13. Evolving effective behaviours to interact with tag-based populations

    Science.gov (United States)

    Yucel, Osman; Crawford, Chad; Sen, Sandip

    2015-07-01

    Tags and other characteristics, externally perceptible features that are consistent among groups of animals or humans, can be used by others to determine appropriate response strategies in societies. This usage of tags can be extended to artificial environments, where agents can significantly reduce cognitive effort spent on appropriate strategy choice and behaviour selection by reusing strategies for interacting with new partners based on their tags. Strategy selection mechanisms developed based on this idea have successfully evolved stable cooperation in games such as the Prisoner's Dilemma game but relies upon payoff sharing and matching methods that limit the applicability of the tag framework. Our goal is to develop a general classification and behaviour selection approach based on the tag framework. We propose and evaluate alternative tag matching and adaptation schemes for a new, incoming individual to select appropriate behaviour against any population member of an existing, stable society. Our proposed approach allows agents to evolve both the optimal tag for the environment as well as appropriate strategies for existing agent groups. We show that these mechanisms will allow for robust selection of optimal strategies by agents entering a stable society and analyse the various environments where this approach is effective.

  14. Posthandling survival and PIT tag retention by alewives—a comparison of gastric and surgical implants

    Science.gov (United States)

    Castro-Santos, Theodore; Voni, Volney

    2013-01-01

    We compared survival and tag retention of Alewives Alosa pseudoharengus tagged with PIT tags, using intraperitoneal (IP) surgical implants, gastric implants (GI), and untagged controls held for 38 d. Retention was 100% for IP-tagged Alewives and 98% for GI-tagged implants. No significant difference in survival was observed among any of these groups. These results lend support to the use of PIT telemetry for studying fish passage and migration of anadromous herring. Both methods hold promise for improving estimates of freshwater survival of adult anadromous clupeids; further research should make it also possible to refine estimates of adult marine survival.

  15. A Novel Recombinant DNA System for High Efficiency Affinity Purification of Proteins in Saccharomyces cerevisiae.

    Science.gov (United States)

    Carrick, Brian H; Hao, Linxuan; Smaldino, Philip J; Engelke, David R

    2015-12-29

    Isolation of endogenous proteins from Saccharomyces cerevisiae has been facilitated by inserting encoding polypeptide affinity tags at the C-termini of chromosomal open reading frames (ORFs) using homologous recombination of DNA fragments. Tagged protein isolation is limited by a number of factors, including high cost of affinity resins for bulk isolation and low concentration of ligands on the resin surface, leading to low isolation efficiencies and trapping of contaminants. To address this, we have created a recombinant "CelTag" DNA construct from which PCR fragments can be created to easily tag C-termini of S. cerevisiae ORFs using selection for a nat1 marker. The tag has a C-terminal cellulose binding module to be used in the first affinity step. Microgranular cellulose is very inexpensive and has an effectively continuous ligand on its surface, allowing rapid, highly efficient purification with minimal background. Cellulose-bound proteins are released by specific cleavage of an included site for TEV protease, giving nearly pure product. The tag can be lifted from the recombinant DNA construct either with or without a 13x myc epitope tag between the target ORF and the TEV protease site. Binding of CelTag protein fusions to cellulose is stable to high salt, nonionic detergents, and 1 M urea, allowing stringent washing conditions to remove loosely associated components, as needed, before specific elution. It is anticipated that this reagent could allow isolation of protein complexes from large quantities of yeast extract, including soluble, membrane-bound, or nucleic acid-associated assemblies.

  16. Social Image Tag Ranking by Two-View Learning

    Science.gov (United States)

    Zhuang, Jinfeng; Hoi, Steven C. H.

    Tags play a central role in text-based social image retrieval and browsing. However, the tags annotated by web users could be noisy, irrelevant, and often incomplete for describing the image contents, which may severely deteriorate the performance of text-based image retrieval models. In order to solve this problem, researchers have proposed techniques to rank the annotated tags of a social image according to their relevance to the visual content of the image. In this paper, we aim to overcome the challenge of social image tag ranking for a corpus of social images with rich user-generated tags by proposing a novel two-view learning approach. It can effectively exploit both textual and visual contents of social images to discover the complicated relationship between tags and images. Unlike the conventional learning approaches that usually assumes some parametric models, our method is completely data-driven and makes no assumption about the underlying models, making the proposed solution practically more effective. We formulate our method as an optimization task and present an efficient algorithm to solve it. To evaluate the efficacy of our method, we conducted an extensive set of experiments by applying our technique to both text-based social image retrieval and automatic image annotation tasks. Our empirical results showed that the proposed method can be more effective than the conventional approaches.

  17. Human P-glycoprotein exhibits reduced affinity for substrates during a catalytic transition state.

    Science.gov (United States)

    Ramachandra, M; Ambudkar, S V; Chen, D; Hrycyna, C A; Dey, S; Gottesman, M M; Pastan, I

    1998-04-01

    Human P-glycoprotein (Pgp), a plasma membrane protein that confers multidrug resistance, functions as an ATP-dependent drug efflux pump. Pgp contains two ATP binding/utilization sites and exhibits ATPase activity that is stimulated in the presence of substrates and modulating agents. The mechanism of coupling of ATP hydrolysis to drug transport is not known. To understand the role of ATP hydrolysis in drug binding, it is necessary to develop methods for purifying and reconstituting Pgp that retains properties including stimulation of ATPase activity by known substrates to an extent similar to that in the native membrane. In this study, (His)6-tagged Pgp was expressed in Trichoplusia ni (High Five) cells using the recombinant baculovirus system and purified by metal affinity chromatography. Upon reconstitution into phospholipid vesicles, purified Pgp exhibited specific binding to analogues of substrates and ATP in affinity labeling experiments and displayed a high level of drug-stimulated ATPase activity (specific activity ranging from 4.5 to 6.5 micromol min-1 mg-1). The ATPase activity was inhibited by ADP in a competitive manner, and by vanadate and N-ethylmaleimide at low concentrations. Vanadate which is known to inhibit ATPase activity by trapping MgADP at the catalytic site inhibited photoaffinity labeling of Pgp with substrate analogues, [125I]iodoarylazidoprazosin and [3H]azidopine, only under ATP hydrolysis conditions. Because vanadate-trapped Pgp is known to resemble the ADP and phosphate-bound catalytic transition state, our findings indicate that ATP hydrolysis results in a conformation with reduced affinity for substrates. A catalytic transition conformation with reduced affinity would essentially result in substrate dissociation and supports a model for drug transport in which an ATP hydrolysis-induced conformational change leads to drug release toward the extracellular medium.

  18. Myocardial MR tagging. Analysis of regional and global myocardial function; Kardiales MR-Tagging. Analyse regionaler und globaler Myokardfunktion

    Energy Technology Data Exchange (ETDEWEB)

    Kramer, U.; Fenchel, M. [Universitaet Tuebingen, Abt. fuer Diagnostische und Interventionelle Radiologie, Radiologische Klinik, Tuebingen (Germany); Hennemuth, A. [Fraunhofer MEVIS, Bremen (Germany)

    2010-06-15

    Myocardial MR tagging is a powerful method which allows for assessment of myocardial function and may become an important tool for clinical evaluation of cardiac dysfunction, particularly in ischemic heart disease. In addition to visual assessment it allows direct quantification of myocardial deformation and strain to measure contractility. The use of myocardial tagging has provided new insights into the (patho)physiology of regional wall motion, and several parameters have been described as being useful to identify an ischemic response of the myocardium. One challenge encountered with tagging at 1.5 T is the fading of tags at end-diastole, greatly limiting the evaluation of myocardial function during diastole. Due to longer T{sub 1} relaxation times of the myocardium, tagging at 3 T has shown to have a higher CNR{sub Tag} and better tag persistence when compared to current clinical gradient-echo tagging protocols at 1.5 T. As a consequence, tagging at higher field strengths may be well suited for the characterization of the diastolic portion of the cardiac cycle in future applications. (orig.) [German] Das myokardiale Tagging mittels der kardialen Magnetresonanztomographie (MRT) stellt ein spezielles Verfahren dar, das eine quantitative Analyse der regionalen Myokardfunktion erlaubt. Mit der Analyse der regionalen Wandbewegung koennen pathologische Bewegungsablaeufe fruehzeitig erkannt und kardiale Dysfunktionen differenziert werden. Neben der visuellen Analyse ist es in erster Linie die quantitative Bestimmung der aus der Echokardiographie bekannten Funktionsparameter, die den Vorteil des Taggings bei der Charakterisierung der myokardialen Funktion ausmachen. Die quantitative Erfassung des Rotations- und Kontraktionsverhaltens mit dem myokardialen Tagging eroeffnet bei verschiedenen Erkrankungen des Herzens neue Einblicke in die Pathophysiologie. Eine intrinsische Limitation dieses Verfahrens besteht in dem insbesondere in der diastolischen Phase des Herzzyklus

  19. HAMS: High-Affinity Mass Spectrometry Screening. A High-Throughput Screening Method for Identifying the Tightest-Binding Lead Compounds for Target Proteins with No False Positive Identifications

    Science.gov (United States)

    Imaduwage, Kasun P.; Go, Eden P.; Zhu, Zhikai; Desaire, Heather

    2016-09-01

    A major challenge in drug discovery is the identification of high affinity lead compounds that bind a particular target protein; these leads are typically identified by high throughput screens. Mass spectrometry has become a detection method of choice in drug screening assays because the target and the ligand need not be modified. Label-free assays are advantageous because they can be developed more rapidly than assays requiring labels, and they eliminate the risk of the label interfering with the binding event. However, in commonly used MS-based screening methods, detection of false positives is a major challenge. Here, we describe a detection strategy designed to eliminate false positives. In this approach, the protein and the ligands are incubated together, and the non-binders are separated for detection. Hits (protein binders) are not detectable by MS after incubation with the protein, but readily identifiable by MS when the target protein is not present in the incubation media. The assay was demonstrated using three different proteins and hundreds of non-inhibitors; no false positive hits were identified in any experiment. The assay can be tuned to select for ligands of a particular binding affinity by varying the quantity of protein used and the immobilization method. As examples, the method selectively detected inhibitors that have Ki values of 0.2 μM, 50 pM, and 700 pM. These findings demonstrate that the approach described here compares favorably with traditional MS-based screening methods.

  20. HAMS: High-Affinity Mass Spectrometry Screening. A High-Throughput Screening Method for Identifying the Tightest-Binding Lead Compounds for Target Proteins with No False Positive Identifications

    Science.gov (United States)

    Imaduwage, Kasun P.; Go, Eden P.; Zhu, Zhikai; Desaire, Heather

    2016-11-01

    A major challenge in drug discovery is the identification of high affinity lead compounds that bind a particular target protein; these leads are typically identified by high throughput screens. Mass spectrometry has become a detection method of choice in drug screening assays because the target and the ligand need not be modified. Label-free assays are advantageous because they can be developed more rapidly than assays requiring labels, and they eliminate the risk of the label interfering with the binding event. However, in commonly used MS-based screening methods, detection of false positives is a major challenge. Here, we describe a detection strategy designed to eliminate false positives. In this approach, the protein and the ligands are incubated together, and the non-binders are separated for detection. Hits (protein binders) are not detectable by MS after incubation with the protein, but readily identifiable by MS when the target protein is not present in the incubation media. The assay was demonstrated using three different proteins and hundreds of non-inhibitors; no false positive hits were identified in any experiment. The assay can be tuned to select for ligands of a particular binding affinity by varying the quantity of protein used and the immobilization method. As examples, the method selectively detected inhibitors that have Ki values of 0.2 μM, 50 pM, and 700 pM. These findings demonstrate that the approach described here compares favorably with traditional MS-based screening methods.

  1. Tempting To Tag: An Experimental Comparison Of Four Tagging Input Mechanisms

    Directory of Open Access Journals (Sweden)

    Mark Melenhorst

    2010-01-01

    Full Text Available Tagging helps achieve improved indexing and recommendation of resources (e.g., videos or pictures in large data collections. In order to reap the benefits of tagging, people must be persuaded to label the resources they consume. This paper reports on a study in which four different tagging input mechanisms and their effect on users' motivation to tag were compared. The mechanisms consisted of a standard tag input box, a chatbot-like environment, a bookmarking mechanism, and a "tag and vote" game. The results of our experiment show that the use of the nonstandard tagging input mechanisms does not affect users' motivation to tag. In some instances tagging mechanisms were found to distract users from their primary task: consuming resources. Persuading people to tag might be accomplished more effectively by using other motivating tagging mechanisms (e.g., tagging games, or motivation could be created by explaining the usefulness of tagging.

  2. 大交角近景影像的仿射不变特征匹配方法研究%Method of affine invariant feature matching for large-angle and dose-range images

    Institute of Scientific and Technical Information of China (English)

    梁艳; 盛业华; 杨林

    2013-01-01

    针对近景数字摄影测量中大交角影像的匹配问题,本文提出了一种仿射不变特征提取与匹配方法.该方法集成Hessain-Affine和MSER特征检测算法提取一定数量的仿射不变特征区域,并将提取的椭圆形特征区域归一化处理为圆形区域,再用SIFT特征描述算子对特征区域进行描述,然后进行基于距离的粗匹配,最后在核线约束下进行精匹配.实验表明,本文的方法在对大交角近景图像进行匹配时,可以得到相对多数量的匹配对和较高的正确匹配率,具有很好的稳定性和鲁棒性.%To solve such problem for matching the large-angle and close-range images of digital photogrammetry, the affine invariant feature extraction and matching method was proposed in the paper. The method integrated Hessain-Affine and MSER algorithms to extract certain number of affine invariant regions, normalized irregular feature areas into circle areas, and described them with SIFT descriptor, then roughly matched based on the distance, finally, in order to improve the accuracy of matching, fine matching on the epi-polar constraint was carried out. The experiments showed that this method could get the relatively large amount of matched point pairs and higher matching auuracy rate with good stability and robustness in matching the large-angle and close-range images.

  3. Affinity Pull-Down of Proteins Using Anti-FLAG M2 Agarose Beads

    Science.gov (United States)

    Gerace, Erica; Moazed, Danesh

    2016-01-01

    FLAG is an affinity tag widely used for rapid and highly specific one-step protein purification. Native elution of protein from anti-FLAG antibody resins allows the identification of protein and nucleic acid binding partners and functional analysis using biochemical activity assays. PMID:26096505

  4. Classification of neocortical interneurons using affinity propagation

    Science.gov (United States)

    Santana, Roberto; McGarry, Laura M.; Bielza, Concha; Larrañaga, Pedro; Yuste, Rafael

    2013-01-01

    In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. In fact, neuronal classification is a difficult problem because it is unclear how to designate a neuronal cell class and what are the best characteristics to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological, or molecular characteristics, have provided quantitative and unbiased identification of distinct neuronal subtypes, when applied to selected datasets. However, better and more robust classification methods are needed for increasingly complex and larger datasets. Here, we explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. Affinity propagation outperformed Ward's method, a current standard clustering approach, in classifying the neurons into 4 subtypes. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits. PMID:24348339

  5. Classification of neocortical interneurons using affinity propagation.

    Science.gov (United States)

    Santana, Roberto; McGarry, Laura M; Bielza, Concha; Larrañaga, Pedro; Yuste, Rafael

    2013-01-01

    In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. In fact, neuronal classification is a difficult problem because it is unclear how to designate a neuronal cell class and what are the best characteristics to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological, or molecular characteristics, have provided quantitative and unbiased identification of distinct neuronal subtypes, when applied to selected datasets. However, better and more robust classification methods are needed for increasingly complex and larger datasets. Here, we explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. Affinity propagation outperformed Ward's method, a current standard clustering approach, in classifying the neurons into 4 subtypes. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits.

  6. Affinity Purification of Insulin by Peptide-Ligand Affinity Chromatography

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The affinity heptapeptide (HWWWPAS) for insulin, selected from phage display library,was coupled to EAH Sepharose 4B gel and packed to a 1-mL column. The column was used for the affinity purification of insulin from protein mixture and commercial insulin preparation. It was observed that the minor impurity in the commercial insulin was removed by the affinity chromatography. Nearly 40 mg of insulin could be purified with the 1-mL affinity column. The results revealed the high specificity and capacity of the affinity column for insulin purification. Moreover, based on the analysis of the amino acids in the peptide sequence, shorter peptides were designed and synthesized for insulin chromatography. As a result, HWWPS was found to be a good alternative to HWWWPAS, while the other two peptides with three or four amino acids showed weak affinity for insulin. The results indicated that the peptide sequence of HWWWPAS was quite conservative for specific binding of insulin.

  7. Improving Attachments of Remotely-Deployed Dorsal Fin-Mounted Tags: Tissue Structure, Hydrodynamics, in situ Performance, and Tagged-Animal Follow-up

    Science.gov (United States)

    2014-09-30

    alternative method for satellite tagging beluga whales . In: Abstracts of the Cook Inlet Beluga Whale Conference, April 5, 2014. Anchorage, AK. [oral...G.M. Towers, J.R., Pilkington, J.F., Barrett-Lennard, L.G., and Andrews, R.D. (2013). New insights into the northward migration route of gray whales ...our project includes follow-up studies of whales that have been tagged with a remotely-deployed dorsal fin-mounted tag to accurately quantify wound

  8. Unsupervised mining of frequent tags for clinical eligibility text indexing.

    Science.gov (United States)

    Miotto, Riccardo; Weng, Chunhua

    2013-12-01

    Clinical text, such as clinical trial eligibility criteria, is largely underused in state-of-the-art medical search engines due to difficulties of accurate parsing. This paper proposes a novel methodology to derive a semantic index for clinical eligibility documents based on a controlled vocabulary of frequent tags, which are automatically mined from the text. We applied this method to eligibility criteria on ClinicalTrials.gov and report that frequent tags (1) define an effective and efficient index of clinical trials and (2) are unlikely to grow radically when the repository increases. We proposed to apply the semantic index to filter clinical trial search results and we concluded that frequent tags reduce the result space more efficiently than an uncontrolled set of UMLS concepts. Overall, unsupervised mining of frequent tags from clinical text leads to an effective semantic index for the clinical eligibility documents and promotes their computational reuse.

  9. Jacobi Structures on Affine Bundles

    Institute of Scientific and Technical Information of China (English)

    J. GRABOWSKI; D. IGLESIAS; J. C. MARRERO; E. PADR(O)N; P. URBA(N)SKI

    2007-01-01

    We study affine Jacobi structures (brackets) on an affine bundle π: A→M, i.e. Jacobi brackets that close on affine functions. We prove that if the rank of A is non-zero, there is a one-to- one correspondence between affine Jacobi structures on A and Lie algebroid structures on the vector bundle A+=∪p∈M Aff(Ap, R) of affine functionals. In the case rank A = 0, it is shown that there is a one-to-one correspondence between affins Jacobi structures on A and local Lie algebras on A+. Some examples and applications, also for the linear case, are discussed. For a special type of affine Jacobi structures which are canonically exhibited (strongly-affine or affine-homogeneous Jacobi structures) over a real vector space of finite dimension, we describe the leaves of its characteristic foliation as the orbits of an affine representation. These afline Jacobi structures can be viewed as an analog of the Kostant-Arnold-LiouviUe linear Poisson structure on the dual space of a real finite-dimensional Lie algebra.

  10. Global navigation system with RFID tags

    Science.gov (United States)

    Tsukiyama, Toshifumi

    2002-02-01

    A new navigation system is described for a mobile robot moving around in man-made environments such as hallways in a building. The system is based on a commercial three-wheel mobile platform with the addition of a Linux-based laptop computer, a Radio Frequency Identification (RDID) tag sensor and a vision system. At critical junctions such as the intersection of two passages the navigation system must identify the robot's location on a given map. We propose a method using RFID tags as landmarks. Each RFID tag has a unique ID number corresponding to its location on the map. The navigation system can decide the next movement (left-turn, right-turn and so on) toward a given goal based on this number. The navigation system also can automatically follow walls using the vision system. Since the equipment setup is very simple and the navigation system is easily combined with general mobile robot systems, our proposed technique would be useful for real-world robotic applications such as intelligent navigation for motorized wheelchairs.

  11. Removal of PCR error products and unincorporated primers by metal-chelate affinity chromatography.

    Directory of Open Access Journals (Sweden)

    Indhu Kanakaraj

    Full Text Available Immobilized Metal Affinity Chromatography (IMAC has been used for decades to purify proteins on the basis of amino acid content, especially surface-exposed histidines and "histidine tags" genetically added to recombinant proteins. We and others have extended the use of IMAC to purification of nucleic acids via interactions with the nucleotide bases, especially purines, of single-stranded RNA and DNA. We also have demonstrated the purification of plasmid DNA from contaminating genomic DNA by IMAC capture of selectively-denatured genomic DNA. Here we describe an efficient method of purifying PCR products by specifically removing error products, excess primers, and unincorporated dNTPs from PCR product mixtures using flow-through metal-chelate affinity adsorption. By flowing a PCR product mixture through a Cu(2+-iminodiacetic acid (IDA agarose spin column, 94-99% of the dNTPs and nearly all the primers can be removed. Many of the error products commonly formed by Taq polymerase also are removed. Sequencing of the IMAC-processed PCR product gave base-calling accuracy comparable to that obtained with a commercial PCR product purification method. The results show that IMAC matrices (specifically Cu(2+-IDA agarose can be used for the purification of PCR products. Due to the generality of the base-specific mechanism of adsorption, IMAC matrices may also be used in the purification of oligonucleotides, cDNA, mRNA and micro RNAs.

  12. Determining the ice-binding planes of antifreeze proteins by fluorescence-based ice plane affinity.

    Science.gov (United States)

    Basu, Koli; Garnham, Christopher P; Nishimiya, Yoshiyuki; Tsuda, Sakae; Braslavsky, Ido; Davies, Peter

    2014-01-15

    Antifreeze proteins (AFPs) are expressed in a variety of cold-hardy organisms to prevent or slow internal ice growth. AFPs bind to specific planes of ice through their ice-binding surfaces. Fluorescence-based ice plane affinity (FIPA) analysis is a modified technique used to determine the ice planes to which the AFPs bind. FIPA is based on the original ice-etching method for determining AFP-bound ice-planes. It produces clearer images in a shortened experimental time. In FIPA analysis, AFPs are fluorescently labeled with a chimeric tag or a covalent dye then slowly incorporated into a macroscopic single ice crystal, which has been preformed into a hemisphere and oriented to determine the a- and c-axes. The AFP-bound ice hemisphere is imaged under UV light to visualize AFP-bound planes using filters to block out nonspecific light. Fluorescent labeling of the AFPs allows real-time monitoring of AFP adsorption into ice. The labels have been found not to influence the planes to which AFPs bind. FIPA analysis also introduces the option to bind more than one differently tagged AFP on the same single ice crystal to help differentiate their binding planes. These applications of FIPA are helping to advance our understanding of how AFPs bind to ice to halt its growth and why many AFP-producing organisms express multiple AFP isoforms.

  13. Neutral B meson flavor tagging

    Science.gov (United States)

    Wilson, Robert J.

    2001-07-01

    We present an investigation of the use of net charge and kaon identification to tag the flavor of neutral B mesons. The net charge of the neutral B meson decay products is zero if all charged particles are used and slightly non-zero if only undiscriminated hadronic final states are used. The net charge of the kaons alone correctly tags the identity of the neutral meson in at least a third of all decays. We have parametrized the particle identification capability of several techniques, such as dE/dx in time projection chambers, LEP/SLC ring-imaging chambers and an enhanced BaBar DIRC. Using these parametrisations we compare the relative tagging power of each technique to that of an ideal detector.

  14. Neutral B meson flavor tagging

    CERN Document Server

    Wilson, R J

    2001-01-01

    We present an investigation of the use of net charge and kaon identification to tag the flavor of neutral B mesons. The net charge of the neutral B meson decay products is zero if all charged particles are used and slightly non-zero if only undiscriminated hadronic final states are used. The net charge of the kaons alone correctly tags the identity of the neutral meson in at least a third of all decays. We have parametrized the particle identification capability of several techniques, such as dE/dx in time projection chambers, LEP/SLC ring-imaging chambers and an enhanced BaBar DIRC. Using these parametrisations we compare the relative tagging power of each technique to that of an ideal detector. (8 refs).

  15. 一种新的多尺度仿射几何不变量提取方法%New method for multi-scale affine geometric invariant extraction

    Institute of Scientific and Technical Information of China (English)

    黄波; 赵晓晖; 赵继印; 时公涛; 陈涛

    2012-01-01

    A new method for multi-scale affine geometric invariant extraction is proposed. The method begins with the self-defined multi-scale convolution transformation, combines with gray-scale normalization, and builds a series of affine covariant forms of the object image. After that, a series of extended centroids of each co-variant form are calculated through a set of designed nonlinear functions, and the new multi-scale affine geometric invariants are obtained. Compared with the classical extended centroid features and multi-scale auto-convolution, the introduced invariant only needs cutting once and can construct any number area ratio invariant features. More invariant features can be extracted from a single affine covariant form. All of these can reduce feature errors effectively and improve the efficiency of the feature attainment. A typical "fish" test database is adopted to validate the efficiency of the proposed method from the perspective of computational complexity, noise immunity, anti-blocking and image expansion.%提出了一种新的多尺度仿射几何不变量提取方法.该方法以自定义的多尺度自卷积变换为起点,结合灰度归一化处理,构建出目标图像的一系列仿射协变形式,进而通过设计一组非线性函数计算每个协变形式的一组扩展质 心,由此得到新的多尺度仿射几何不变量.将所得不变量与经典的扩展质心特征、多尺度自卷积相比,由于其仅需一次分割便可构造出任意数量的区域面积比仿射不变特征,且从单个仿射协变形式中即可提取多个不变特征,从而有效减小了特征误差,提高了特征的获取效率.利用典型的“Fish”测试数据库,从计算复杂度、抗噪性、抗遮挡性和图像扩展性等方面验证了所提方法的有效性.

  16. Tissue-specific tagging of endogenous loci in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Kate Koles

    2016-01-01

    Full Text Available Fluorescent protein tags have revolutionized cell and developmental biology, and in combination with binary expression systems they enable diverse tissue-specific studies of protein function. However these binary expression systems often do not recapitulate endogenous protein expression levels, localization, binding partners and/or developmental windows of gene expression. To address these limitations, we have developed a method called T-STEP (tissue-specific tagging of endogenous proteins that allows endogenous loci to be tagged in a tissue specific manner. T-STEP uses a combination of efficient CRISPR/Cas9-enhanced gene targeting and tissue-specific recombinase-mediated tag swapping to temporally and spatially label endogenous proteins. We have employed this method to GFP tag OCRL (a phosphoinositide-5-phosphatase in the endocytic pathway and Vps35 (a Parkinson's disease-implicated component of the endosomal retromer complex in diverse Drosophila tissues including neurons, glia, muscles and hemocytes. Selective tagging of endogenous proteins allows, for the first time, cell type-specific live imaging and proteomics in complex tissues.

  17. Kernel Affine Projection Algorithms

    Directory of Open Access Journals (Sweden)

    José C. Príncipe

    2008-05-01

    Full Text Available The combination of the famed kernel trick and affine projection algorithms (APAs yields powerful nonlinear extensions, named collectively here, KAPA. This paper is a follow-up study of the recently introduced kernel least-mean-square algorithm (KLMS. KAPA inherits the simplicity and online nature of KLMS while reducing its gradient noise, boosting performance. More interestingly, it provides a unifying model for several neural network techniques, including kernel least-mean-square algorithms, kernel adaline, sliding-window kernel recursive-least squares (KRLS, and regularization networks. Therefore, many insights can be gained into the basic relations among them and the tradeoff between computation complexity and performance. Several simulations illustrate its wide applicability.

  18. Kernel Affine Projection Algorithms

    Science.gov (United States)

    Liu, Weifeng; Príncipe, José C.

    2008-12-01

    The combination of the famed kernel trick and affine projection algorithms (APAs) yields powerful nonlinear extensions, named collectively here, KAPA. This paper is a follow-up study of the recently introduced kernel least-mean-square algorithm (KLMS). KAPA inherits the simplicity and online nature of KLMS while reducing its gradient noise, boosting performance. More interestingly, it provides a unifying model for several neural network techniques, including kernel least-mean-square algorithms, kernel adaline, sliding-window kernel recursive-least squares (KRLS), and regularization networks. Therefore, many insights can be gained into the basic relations among them and the tradeoff between computation complexity and performance. Several simulations illustrate its wide applicability.

  19. AUTOMATIC TAGGING OF PERSIAN WEB PAGES BASED ON N-GRAM LANGUAGE MODELS USING MAPREDUCE

    Directory of Open Access Journals (Sweden)

    Saeed Shahrivari

    2015-07-01

    Full Text Available Page tagging is one of the most important facilities for increasing the accuracy of information retrieval in the web. Tags are simple pieces of data that usually consist of one or several words, and briefly describe a page. Tags provide useful information about a page and can be used for boosting the accuracy of searching, document clustering, and result grouping. The most accurate solution to page tagging is using human experts. However, when the number of pages is large, humans cannot be used, and some automatic solutions should be used instead. We propose a solution called PerTag which can automatically tag a set of Persian web pages. PerTag is based on n-gram models and uses the tf-idf method plus some effective Persian language rules to select proper tags for each web page. Since our target is huge sets of web pages, PerTag is built on top of the MapReduce distributed computing framework. We used a set of more than 500 million Persian web pages during our experiments, and extracted tags for each page using a cluster of 40 machines. The experimental results show that PerTag is both fast and accurate

  20. Traceless affinity labeling of endogenous proteins for functional analysis in living cells.

    Science.gov (United States)

    Hayashi, Takahiro; Hamachi, Itaru

    2012-09-18

    Protein labeling and imaging techniques have provided tremendous opportunities to study the structure, function, dynamics, and localization of individual proteins in the complex environment of living cells. Molecular biology-based approaches, such as GFP-fusion tags and monoclonal antibodies, have served as important tools for the visualization of individual proteins in cells. Although these techniques continue to be valuable for live cell imaging, they have a number of limitations that have only been addressed by recent progress in chemistry-based approaches. These chemical approaches benefit greatly from the smaller probe sizes that should result in fewer perturbations to proteins and to biological systems as a whole. Despite the research in this area, so far none of these labeling techniques permit labeling and imaging of selected endogenous proteins in living cells. Researchers have widely used affinity labeling, in which the protein of interest is labeled by a reactive group attached to a ligand, to identify and characterize proteins. Since the first report of affinity labeling in the early 1960s, efforts to fine-tune the chemical structures of both the reactive group and ligand have led to protein labeling with excellent target selectivity in the whole proteome of living cells. Although the chemical probes used for affinity labeling generally inactivate target proteins, this strategy holds promise as a valuable tool for the labeling and imaging of endogenous proteins in living cells and by extension in living animals. In this Account, we summarize traceless affinity labeling, a technique explored mainly in our laboratory. In our overview of the different labeling techniques, we emphasize the challenge of designing chemical probes that allow for dissociation of the affinity module (often a ligand) after the labeling reaction so that the labeled protein retains its native function. This feature distinguishes the traceless labeling approach from the traditional

  1. Social image tagging with diverse semantics.

    Science.gov (United States)

    Qian, Xueming; Hua, Xian-Sheng; Tang, Yuan Yan; Mei, Tao

    2014-12-01

    We have witnessed the popularity of image-sharing websites for sharing personal experiences through photos on the Web. These websites allow users describing the content of their uploaded images with a set of tags. Those user-annotated tags are often noisy and biased. Social image tagging aims at removing noisy tags and suggests new relevant tags. However, most existing tag enrichment approaches predominantly focus on tag relevance and overlook tag diversity problem. How to make the top-ranked tags covering a wide range of semantic is still an opening, yet challenging, issue. In this paper, we propose an approach to retag social images with diverse semantics. Both the relevance of a tag to image as well as its semantic compensations to the already determined tags are fused to determine the final tag list for a given image. Different from existing image tagging approaches, the top-ranked tags are not only highly relevant to the image but also have significant semantic compensations with each other. Experiments show the effectiveness of the proposed approach.

  2. A PERSONALISED RECOMMENDATION METHOD INTEGRATING LBS AND SOCIAL NETWORK TAG%一种综合 LBS 和社会网络标签的个性化推荐方法

    Institute of Scientific and Technical Information of China (English)

    陈平华; 何婕; 梁琼

    2015-01-01

    针对现有的基于 LBS(Location Based Service)个性化推荐系统在构建用户兴趣模型时存在的缺陷,提出一种综合 LBS 和社会网络标签的个性化推荐(LTCF)方法。通过引入网络标签和用户社会关系,从用户标注的标签资源中找到拥有共同兴趣爱好的用户关系以及从社会网络中找到与目标用户关系紧密的用户,同时结合考虑用户兴趣爱好随空间不断变化的特点,依据协同过滤算法,计算用户社会关系度和用户空间相似性,依此得到目标用户的最近邻集合,在最近邻集基础上给出推荐结果。实验结果表明,相比于传统的基于 LBS 推荐方法,LTCF 模型在查全率和产准率有了显著的提升,能更好地反映出用户偏好,显著提高了推荐准确度。%In light of the shortcomings of existing LBS-based personalised recommendation system in building user interests model,this pa-per proposes a personalised recommendation method which integrates the LBS and social network tag (LTCF).By introducing network tag and users social relations,the method finds from the tag resources marked by users the relationship between users having common interests and hobbies as well as finds from social networks the users having intimate relationship with targeted users.Meanwhile,combining the considera-tion of the feature that users'interests constantly change along with the space and in accordance with collaborative filtering algorithm,the method calculates the social relation degree of users and the similarity of user spaces,then according to these it obtains the nearest neighbour sets of target users,and provides the recommendation results based on the nearest neighbour sets.Experimental results show that compared with traditional LBS-based recommendation method,the LTCF model has noticeable improvement in recall rate and precision product rate and can better reflect users'preferences as well as significantly

  3. Tagging, Encoding, and Jones Optimality

    DEFF Research Database (Denmark)

    Danvy, Olivier; Lopez, Pablo E. Martinez

    2003-01-01

    A partial evaluator is said to be Jones-optimal if the result of specializing a self-interpreter with respect to a source program is textually identical to the source program, modulo renaming. Jones optimality has already been obtained if the self-interpreter is untyped. If the selfinterpreter...... is typed, however, residual programs are cluttered with type tags. To obtain the original source program, these tags must be removed. A number of sophisticated solutions have already been proposed. We observe, however, that with a simple representation shift, ordinary partial evaluation is already Jones-optimal...

  4. Adjoint affine fusion and tadpoles

    Science.gov (United States)

    Urichuk, Andrew; Walton, Mark A.

    2016-06-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  5. Adjoint affine fusion and tadpoles

    CERN Document Server

    Urichuk, Andrew

    2016-01-01

    We study affine fusion with the adjoint representation. For simple Lie algebras, elementary and universal formulas determine the decomposition of a tensor product of an integrable highest-weight representation with the adjoint representation. Using the (refined) affine depth rule, we prove that equally striking results apply to adjoint affine fusion. For diagonal fusion, a coefficient equals the number of nonzero Dynkin labels of the relevant affine highest weight, minus 1. A nice lattice-polytope interpretation follows, and allows the straightforward calculation of the genus-1 1-point adjoint Verlinde dimension, the adjoint affine fusion tadpole. Explicit formulas, (piecewise) polynomial in the level, are written for the adjoint tadpoles of all classical Lie algebras. We show that off-diagonal adjoint affine fusion is obtained from the corresponding tensor product by simply dropping non-dominant representations.

  6. WebTag: Web browsing into sensor tags over NFC.

    Science.gov (United States)

    Echevarria, Juan Jose; Ruiz-de-Garibay, Jonathan; Legarda, Jon; Alvarez, Maite; Ayerbe, Ana; Vazquez, Juan Ignacio

    2012-01-01

    Information and Communication Technologies (ICTs) continue to overcome many of the challenges related to wireless sensor monitoring, such as for example the design of smarter embedded processors, the improvement of the network architectures, the development of efficient communication protocols or the maximization of the life cycle autonomy. This work tries to improve the communication link of the data transmission in wireless sensor monitoring. The upstream communication link is usually based on standard IP technologies, but the downstream side is always masked with the proprietary protocols used for the wireless link (like ZigBee, Bluetooth, RFID, etc.). This work presents a novel solution (WebTag) for a direct IP based access to a sensor tag over the Near Field Communication (NFC) technology for secure applications. WebTag allows a direct web access to the sensor tag by means of a standard web browser, it reads the sensor data, configures the sampling rate and implements IP based security policies. It is, definitely, a new step towards the evolution of the Internet of Things paradigm.

  7. WebTag: Web Browsing into Sensor Tags over NFC

    Directory of Open Access Journals (Sweden)

    Juan Jose Echevarria

    2012-06-01

    Full Text Available Information and Communication Technologies (ICTs continue to overcome many of the challenges related to wireless sensor monitoring, such as for example the design of smarter embedded processors, the improvement of the network architectures, the development of efficient communication protocols or the maximization of the life cycle autonomy. This work tries to improve the communication link of the data transmission in wireless sensor monitoring. The upstream communication link is usually based on standard IP technologies, but the downstream side is always masked with the proprietary protocols used for the wireless link (like ZigBee, Bluetooth, RFID, etc.. This work presents a novel solution (WebTag for a direct IP based access to a sensor tag over the Near Field Communication (NFC technology for secure applications. WebTag allows a direct web access to the sensor tag by means of a standard web browser, it reads the sensor data, configures the sampling rate and implements IP based security policies. It is, definitely, a new step towards the evolution of the Internet of Things paradigm.

  8. Dimerization capacities of FGF2 purified with or without heparin-affinity chromatography.

    Directory of Open Access Journals (Sweden)

    Natalia Platonova

    Full Text Available Fibroblast growth factor-2 (FGF2 is a pleiotropic growth factor exhibiting a variety of biological activities. In this article, we studied the capacity of FGF2 purified with or without heparin affinity chromatography to self-associate. Analyzing the NMR HSQC spectra for different FGF2 concentrations, heparin-affinity purified FGF2 showed perturbations that indicate dimerization and are a higher-order oligomerization state. HSQC perturbation observed with different FGF2 concentrations revealed a heparin-binding site and two dimer interfaces. Thus, with increasing protein concentrations, FGF2 monomers make contacts with each other and form dimers or higher order oligomers. On the contrary, FGF2 purified with ion-exchange chromatography did not show similar perturbation indicating that self-association of FGF2 is eliminated if purification is done without heparin-affinity chromatography. The HSQC spectra of heparin-affinity purified FGF2 can be reproduced to some extent by adding heparin tetra-saccharide to ion exchange chromatography purified FGF2. Heparin-affinity purified FGF2 bound to acceptor and donor beads in a tagged form using His-tagged or GST-tagged proteins, also dimerized in the AlphaScreen™ assay. This assay was further validated using different experimental conditions and competitors. The assay constitutes an interesting tool to study dimerization of other FGF forms as well.

  9. Robust segmentation of 4D cardiac MRI-tagged images via spatio-temporal propagation

    Science.gov (United States)

    Qian, Zhen; Huang, Xiaolei; Metaxas, Dimitris N.; Axel, Leon

    2005-04-01

    In this paper we present a robust method for segmenting and tracking cardiac contours and tags in 4D cardiac MRI tagged images via spatio-temporal propagation. Our method is based on two main techniques: the Metamorphs Segmentation for robust boundary estimation, and the tunable Gabor filter bank for tagging lines enhancement, removal and myocardium tracking. We have developed a prototype system based on the integration of these two techniques, and achieved efficient, robust segmentation and tracking with minimal human interaction.

  10. Tagging the European eel Anguilla anguilla (L.) with coded wire tags

    DEFF Research Database (Denmark)

    Thomassen, S.; Pedersen, Michael Ingemann; Holdensgaard, G.

    2000-01-01

    The coded wire tag (CWT) system was examined as a possible tool for tagging European eels (Anguilla anguilla). Two size groups of eels (3.8 and 10.2 g) were tagged with CWTs in the dorsal musculature, Tag loss 28 days after tagging was 3.1% for the small and 0.7% for the large groups of eels....... Of the tag loss in the small eels, 61% were lost during the first 2 h after tagging. In the small group, eels that lost tags were significantly (P...

  11. A Novel Recombinant DNA System for High Efficiency Affinity Purification of Proteins in Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Brian H. Carrick

    2016-03-01

    Full Text Available Isolation of endogenous proteins from Saccharomyces cerevisiae has been facilitated by inserting encoding polypeptide affinity tags at the C-termini of chromosomal open reading frames (ORFs using homologous recombination of DNA fragments. Tagged protein isolation is limited by a number of factors, including high cost of affinity resins for bulk isolation and low concentration of ligands on the resin surface, leading to low isolation efficiencies and trapping of contaminants. To address this, we have created a recombinant “CelTag” DNA construct from which PCR fragments can be created to easily tag C-termini of S. cerevisiae ORFs using selection for a nat1 marker. The tag has a C-terminal cellulose binding module to be used in the first affinity step. Microgranular cellulose is very inexpensive and has an effectively continuous ligand on its surface, allowing rapid, highly efficient purification with minimal background. Cellulose-bound proteins are released by specific cleavage of an included site for TEV protease, giving nearly pure product. The tag can be lifted from the recombinant DNA construct either with or without a 13x myc epitope tag between the target ORF and the TEV protease site. Binding of CelTag protein fusions to cellulose is stable to high salt, nonionic detergents, and 1 M urea, allowing stringent washing conditions to remove loosely associated components, as needed, before specific elution. It is anticipated that this reagent could allow isolation of protein complexes from large quantities of yeast extract, including soluble, membrane-bound, or nucleic acid-associated assemblies.

  12. Affine Invariant Character Recognition by Progressive Removing

    Science.gov (United States)

    Iwamura, Masakazu; Horimatsu, Akira; Niwa, Ryo; Kise, Koichi; Uchida, Seiichi; Omachi, Shinichiro

    Recognizing characters in scene images suffering from perspective distortion is a challenge. Although there are some methods to overcome this difficulty, they are time-consuming. In this paper, we propose a set of affine invariant features and a new recognition scheme called “progressive removing” that can help reduce the processing time. Progressive removing gradually removes less feasible categories and skew angles by using multiple classifiers. We observed that progressive removing and the use of the affine invariant features reduced the processing time by about 60% in comparison to a trivial one without decreasing the recognition rate.

  13. 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...

  14. Graph based techniques for tag cloud generation

    DEFF Research Database (Denmark)

    Leginus, Martin; Dolog, Peter; Lage, Ricardo Gomes

    2013-01-01

    Tag cloud is one of the navigation aids for exploring documents. Tag cloud also link documents through the user defined terms. We explore various graph based techniques to improve the tag cloud generation. Moreover, we introduce relevance measures based on underlying data such as ratings or citat......Tag cloud is one of the navigation aids for exploring documents. Tag cloud also link documents through the user defined terms. We explore various graph based techniques to improve the tag cloud generation. Moreover, we introduce relevance measures based on underlying data such as ratings...

  15. DNA tagged microparticles

    Energy Technology Data Exchange (ETDEWEB)

    Farquar, George Roy; Leif, Roald N; Wheeler, Elizabeth

    2015-05-05

    A simulant that includes a carrier and DNA encapsulated in the carrier. Also a method of making a simulant including the steps of providing a carrier and encapsulating DNA in the carrier to produce the simulant.

  16. Tag-Aware Recommender Systems: A State-of-the-Art Survey

    Institute of Scientific and Technical Information of China (English)

    Zi-Ke Zhang; Tao Zhou; Yi-Cheng Zhang

    2011-01-01

    In the past decade,Social Tagging Systems have attracted increasing attention from both physical and computer science communities.Besides the underlying structure and dynamics of tagging systems,many efforts have been addressed to unify tagging information to reveal user behaviors and preferences,extract the latent semantic relations among items,make recommendations,and so on.Specifically,this article summarizes recent progress about tag-aware recommender systems,emphasizing on the contributions from three mainstream perspectives and approaches:network-based methods,tensor-based methods,and the topic-based methods.Finally,we outline some other tag-related studies and future challenges of tag-aware recommendation algorithms.

  17. SKIN TAGS AND ITS ASSOCIATION WITH SYSTEMIC ILLNESSES

    Directory of Open Access Journals (Sweden)

    Ramya

    2015-05-01

    Full Text Available BACKGROUND AND OBJECTIVES: Skin tags (Acrochordons are very common benign tumor of the skin. It is seen in 25% of population, particularly in women in advancing age and in pregnancy. Skin tags are small, soft, pedunculated and papillomatous lesions usually seen on eyelids and major flexures of the body. Skin tags are enti rely asymptomatic and diagnosis is unmistakable. Many cutaneous lesions have been well established to be associated with some systemic diseases and this has been proven beneficial as it helps in early diagnosis. METHODS: A total no of 50 patients with skin tags were included in the study. Detailed history, thorough physical examination and relevant investigations were done to confirm the systemic manifestations when present. RESULTS: In the study the patient’s age ranged from 18 – 65 yrs with female - male ratio 1:1.17. The number of skin tags among the 50 patients ranged from 1 – 74. In our study we found that 74% of our patients with skin tags were either obese or overweight, 24% of our patients fulfilled the criteria of metabolic syndrome, 36% of the patients were diabetics & 4 of our patients had evidence of thyroid disorder. INTERPRETATION & CONCLUSION: Presence of multiple, hyper pigmented skin tags at more than one site can be a marker for diabetes, obesity and metabolic syndrome. All such patients should be evaluated thoroughly for detecting such illnesses. This study thus emphasizes the need for systemic evaluation of all patients with skin tags for early management and improve d quality of life by life style modification.

  18. EXPLOITING SOCIAL TAGS TO OVERCOME COLD START RECOMMENDATION PROBLEM

    Directory of Open Access Journals (Sweden)

    A. S. Ghabayen

    2014-01-01

    Full Text Available The practice and method of collaboratively creating and managing tags to annotate and categorize content has resulted in the creation of folksonomy. Folksonomies provide new opportunities and challenges in the field of recommender systems. Despite the considerable amount of researches done in the context of recommender systems, the specific problem of integrating tags into standard recommender system algorithms is less explored than the problem of recommending tags. Collaborative filtering is one of the popular approaches for providing recommendation. However, despite the popularity of collaborative filtering, to some extent, it could not recognize the preferences of users in cold-start scenarios, where insufficient preferences are associated to certain users or items, which leads to degraded recommendation quality. This study presents a collaborative filtering approach based on the expansion of users’ tags. In this case, semantics between tags can be unveiled which subsequently resulted in the identification of semantically similar users. Experiment on real-life dataset shows that our approach outperforms the state-of-the-art tag-aware collaborative filtering approaches in terms of recommendation quality particularly in the cold-start situation.

  19. SAGE2Splice: unmapped SAGE tags reveal novel splice junctions.

    Directory of Open Access Journals (Sweden)

    Byron Yu-Lin Kuo

    2006-04-01

    Full Text Available Serial analysis of gene expression (SAGE not only is a method for profiling the global expression of genes, but also offers the opportunity for the discovery of novel transcripts. SAGE tags are mapped to known transcripts to determine the gene of origin. Tags that map neither to a known transcript nor to the genome were hypothesized to span a splice junction, for which the exon combination or exon(s are unknown. To test this hypothesis, we have developed an algorithm, SAGE2Splice, to efficiently map SAGE tags to potential splice junctions in a genome. The algorithm consists of three search levels. A scoring scheme was designed based on position weight matrices to assess the quality of candidates. Using optimized parameters for SAGE2Splice analysis and two sets of SAGE data, candidate junctions were discovered for 5%-6% of unmapped tags. Candidates were classified into three categories, reflecting the previous annotations of the putative splice junctions. Analysis of predicted tags extracted from EST sequences demonstrated that candidate junctions having the splice junction located closer to the center of the tags are more reliable. Nine of these 12 candidates were validated by RT-PCR and sequencing, and among these, four revealed previously uncharacterized exons. Thus, SAGE2Splice provides a new functionality for the identification of novel transcripts and exons. SAGE2Splice is available online at http://www.cisreg.ca.

  20. Genetic tagging of humpback whales

    NARCIS (Netherlands)

    Palsboll, PJ; Allen, J; Berube, M; Clapham, PJ; Feddersen, TP; Hammond, PS; Hudson, RR; Jorgensen, H; Katona, S; Larsen, AH; Larsen, F; Lien, J; Mattila, DK; Sigurjonsson, J; Sears, R; Smith, T; Sponer, R; Stevick, P; Oien, N

    1997-01-01

    The ability to recognize individual animals has substantially increased our knowledge of the biology and behaviour of many taxa(1). However, not all species lend themselves to this approach, either because of insufficient phenotypic variation or because tag attachment is not feasible. The use of gen

  1. MR Colonography with fecal tagging: Barium vs. barium ferumoxsil

    DEFF Research Database (Denmark)

    Achiam, M.P.; Chabanova, E.; Logager, V.B.;

    2008-01-01

    and Methods. Twenty patients referred to CC underwent dark lumen MRC prior to the colonoscopy. Two groups of patients received two different oral contrast agents (barium sulfate and barium sulfate/ferumoxsil) as a laxative-free fecal tagging prior to the MRC. After MRC, the contrast agent was rated...... qualitatively (with the standard method using contrast-to-wall ratio) and subjectively (using a visual analog scale [VAS]) by three different blinded observers. Results. Evaluated both qualitatively and subjectively, the tagging efficiency of barium sulfate/ferumoxsil was significantly better (P ... barium sulfate alone. The VAS method for evaluating the tagging efficiency of contrast agents showed a high correlation (observer 11, r = 0.91) to the standard method using contrast-to-wall ratio and also a high interclass correlation (observer 11 and III = 0.89/0.85). MRC found I of 22 (5%) polyps

  2. Quantifying Visual-Representativeness of Social Image Tags Using Image Tag Clarity

    Science.gov (United States)

    Sun, Aixin; Bhowmick, Sourav S.

    Tags associated with images in various social media sharing web sites are valuable information source for superior image retrieval experiences. Due to the nature of tagging, many tags associated with images are not visually descriptive. In this chapter, we propose Image Tag Clarity to evaluate the effectiveness of a tag in describing the visual content of its annotated images, which is also known as the image tag visual-representativeness. It is measured by computing the zero-mean normalized distance between the tag language model estimated from the images annotated by the tag and the collection language model. The tag/collection language models are derived from the bag of visual-word local content features of the images. The visual-representative tags that are commonly used to annotate visually similar images are given high tag clarity scores. Evaluated on a large real-world dataset containing more than 269K images and their associated tags, we show that the image tag clarity score can effectively identify the visual-representative tags from all tags contributed by users. Based on the tag clarity scores, we have made a few interesting observations that could be used to support many tag-based applications.

  3. HMSRP Hawaiian Monk Seal Tag Data

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — This data set contains records for all tags applied to Hawaiian monk seals since 1981. These tags were applied by PSD personnel and cooperating scientists as part of...

  4. 50 CFR 635.33 - Archival tags.

    Science.gov (United States)

    2010-10-01

    ...) Implantation report. Any person affixing or implanting an archival tag into a regulated species must obtain... catch, possess, retain, and land an Atlantic HMS in which an archival tag has been implanted or...

  5. The Complex Dynamics of Collaborative Tagging

    NARCIS (Netherlands)

    Halpin, H.; Robu, V.; Shepherd, H.

    2007-01-01

    The debate within the Web community over the optimal means by which to organize information often pits formalized classifications against distributed collaborative tagging systems. A number of questions remain unanswered, however, regarding the nature of collaborative tagging systems including wheth

  6. Satellite Tags- Guam/CNMI EEZ

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Satellite tagging was implemented in 2013. Satellite tagging is conducted using a Dan Inject air rifle and deployment arrows designed by Wildlife Computers. Two...

  7. Design and Analysis of Salmonid Tagging Studies in the Columbia Basin, Volume IX; Comparison of Statistical Methods of Estimating Treatment-Control Ratios Using Coded-Wire Tags, Based on Spring Chinook Salmon on the Columbia River, 1986-1988 Technical Report.

    Energy Technology Data Exchange (ETDEWEB)

    Townsend, Richard L.; Skalski, John R. (University of Washington, School of Fisheries, Seattle, WA)

    2000-06-01

    The strength of a salmon run is often measured as the adult return rate from some previous brood year (i.e. the percent of a smolt population returning to spawn or captured in fisheries). The U.S. Army Corp of Engineers (ACOE) program of barge transportation of smolts from collector dams is one mitigation measure used to improve smolt survival. Using Coded Wire-tags, the adult return rates of transported and untransported smolt have been tracked. A ratio of the recovered percentages of adult salmon, those transported in the smolt stage over the salmon not transported (controls), is often used to summarize the program effectiveness. There are a number of ways to estimate this transportation/control (T/C) ratio, and this paper explores six alternative statistical models to improve accuracy and precision of the estimate. Assuming the proportion of adult recoveries are binomially distributed, the data were analyzed using linear regression of arc-sine square-root and logit transformations; general linear model regression (GLM) with logit- and log-links; and a maximum-likelihood estimation (MLE) of the T/C ratio. Profile likelihood intervals were calculated to generate 95% confidence interval estimates of the T/C ratio. Depending on the analytical method, T/C ratios varied greatly. Arc-sine square-root and logit transformations gave individual release T/C ratios which ranged from 1.0934 to 4.0076 and {minus}1.2193 to 1.9057, respectively. The negative T/C ratio is due to the back-transformation properties of the logit transformation. The GLM and MLE approaches produced mean T/C ratios (after adjusting for the individual release batch effects) ranging from 1.4964 to 1.4974. The recommended method from this analysis, a binomial maximum likelihood estimate adjusted for over-dispersion, produced a T/C ratio of 1.4965 with a 95% confidence interval of (1.0618, 1.9312).

  8. Categorical and Specificity Differences between User-Supplied Tags and Search Query Terms for Images. An Analysis of "Flickr" Tags and Web Image Search Queries

    Science.gov (United States)

    Chung, EunKyung; Yoon, JungWon

    2009-01-01

    Introduction: The purpose of this study is to compare characteristics and features of user supplied tags and search query terms for images on the "Flickr" Website in terms of categories of pictorial meanings and level of term specificity. Method: This study focuses on comparisons between tags and search queries using Shatford's categorization…

  9. Using sutures to attach miniature tracking tags to small bats for multimonth movement and behavioral studies

    Science.gov (United States)

    Castle, Kevin T.; Weller, Theodore J.; Cryan, Paul M.; Hein, Cris D.; Schirmacher, Michael R.

    2015-01-01

    1. Determining the detailed movements of individual animals often requires them to carry tracking devices, but tracking broad-scale movement of small bats (brown bats (Eptesicus fuscus) in Colorado and hoary bats (Lasiurus cinereus) in California. 3. GPS tags and data loggers were sutured to 17 bats in this study. Three tagged bats were recaptured seven months after initial deployment, with tags still attached; none of these bats showed ill effects from the tag. No severe injuries were apparent upon recapture of 6 additional bats that carried tags up to 26 days after attachment, however one of the bats exhibited skin chafing. 4. Use of absorbable sutures to affix small tracking devices seems to be a safe, effective method for studying movements of bats over multiple months, although additional testing is warranted. This new attachment method has the potential to quickly advance our understanding of small bats, particularly as more-sophisticated miniature tracking devices (e.g., satellite tags) become available.

  10. ON THE INTERACTIVE FUNCTIONS OF TAG QUESTIONS

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Tag questions are a distinctive feature in spokenEnglish,but seldom used by the non-native speakers.Studies show that tags are employed by native speakersmainly to express their interactive and interpersonalmeaning.In this paper,the writer explores the natureof tag questions,mainly their interactive and inter-personal functions in daily coversation.Suggestionsare made with regard to the teaching of tags in theclassroom.

  11. Multilabel Learning for Automatic Web Services Tagging

    OpenAIRE

    Mustapha AZNAG; Mohamed QUAFAFOU; Jarir, Zahi

    2014-01-01

    Recently, some web services portals and search engines as Biocatalogue and Seekda!, have allowed users to manually annotate Web services using tags. User Tags provide meaningful descriptions of services and allow users to index and organize their contents. Tagging technique is widely used to annotate objects in Web 2.0 applications. In this paper we propose a novel probabilistic topic model (which extends the CorrLDA model - Correspondence Latent Dirichlet Allocation-) to automatically tag we...

  12. Peptide Nucleic Acids Having Enhanced Binding Affinity and Sequence Specificity

    DEFF Research Database (Denmark)

    1998-01-01

    A novel class of compounds, known as peptide nucleic acids, bind complementary DNA and RNA strands more strongly than a corresponding DNA strand, and exhibit increased sequence specificity and binding affinity. Methods of increasing binding affinity and sequence specificity of peptide nucleic aci...

  13. Towards EPC compatible plastic RFID tags

    NARCIS (Netherlands)

    Myny, K.; Steudel, S.; Vicca, P.; Smout, S.; Beenhakkers, M.J.; Aerle, N.A.J.M. van; Furthner, F.; Putten, B. van der; Tripathi, A.K.; Gelinck, G.H.; Genoe, J.; Dehaene, W.; Heremans, P.

    2010-01-01

    A target application for plastic RFID tags is Electronic Product Coding (EPC). The EPC-specifications set some demanding requirements for RFID tags. In this work, we review the work that has been done to fulfill some of these specifications. We describe a complete 64-bit RFID tag that is inductively

  14. Motion analysis of both ventricles using tagged MRI

    Science.gov (United States)

    Ozturk, Cengizhan; McVeigh, Elliot R.

    2000-04-01

    Although several methods exist for the analysis of tagged MRI images of the left ventricle (LV), analysis of the right ventricle (RV) remains challenging due to its complex anatomy and significant through plane motion. We present here preliminary results of our new motion analysis method, both for RV and LV, in healthy human volunteers. In this method, following standard myocardial and tag segmentation of cardiac gated cine tagged MR images; a 4D B-spline based parametric motion field was computed for a volume of interest encompassing both ventricles. Using this motion field, 3D displacements and strains were calculated on the RV and LV. We observed that for both chambers the circumferential strain (Ecc) decreased with a constant rate throughout systole. The systolic strain rate displayed spatial similarity not only for the LV but also for the RV. For RV free wall, mean systolic Ecc was -0.19 +/- 0.05 with an average coefficient of variability of 20%. The 4D B-spline based motion analysis technique for tagged MRI yields compatible results for the LV and gives consistent circumferential strain measures for the RV free wall. Tagged MRI based RV mechanical analysis can be used along with LV results for a more complete cardiac evaluation.

  15. In Vivo Tagging of Lung Epithelial Cells to Define the Early Steps of Tumor Cell Dissemination

    Science.gov (United States)

    2014-10-01

    derived lung cells, only the epithelium will be tagged by this method. Prior to using these animals for our studies, we will ensure that lineage labeled...1 AWARD NUMBER: W81XWH-13-1-0184 TITLE: In Vivo Tagging of Lung Epithelial Cells To Define...REPORT TYPE Annual 3. DATES COVERED 15Sep2013-14Sep2014 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER In Vivo Tagging of Lung

  16. Affitins as robust tailored reagents for affinity chromatography purification of antibodies and non-immunoglobulin proteins.

    Science.gov (United States)

    Béhar, Ghislaine; Renodon-Cornière, Axelle; Mouratou, Barbara; Pecorari, Frédéric

    2016-04-08

    Affinity chromatography is a convenient way of purifying proteins, as a high degree of purity can be reached in one step. The use of tags has greatly contributed to the popularity of this technique. However, the addition of tags may not be desirable or possible for the production of biopharmaceuticals. There is thus a need for tailored artificial affinity ligands. We have developed the use of archaeal extremophilic proteins as scaffolds to generate affinity proteins (Affitins). Here, we explored the potential of Affitins as ligand to design affinity columns. Affitins specific for human immunoglobulin G (hIgG), bacterial PulD protein, and chicken egg lysozyme were immobilized on an agarose matrix. The columns obtained were functional and highly selective for their cognate target, even in the presence of exogenous proteins as found in cell culture media, ascites and bacterial lysates, which result in a high degree of purity (∼95%) and recovery (∼100%) in a single step. Anti-hIgG Affitin columns withstand repetitive cycles of purification and cleaning-in-place treatments with 0.25 M NaOH as well as Protein A does. High levels of Affitin productions in Escherichia coli makes it possible to produce these affinity columns at low cost. Our results validate Affitins as a new class of tailored ligands for the affinity chromatography purification of potentially any proteins of interest including biopharmaceuticals.

  17. Inhibition of Xanthine Oxidase Activity by Gnaphalium Affine Extract

    Institute of Scientific and Technical Information of China (English)

    Wei-qing Lin; Jian-xiang Xie; Xiao-mu Wu; Lin Yang; Hai-dong Wang

    2014-01-01

    Objective To evaluate the inhibitory effect of Gnaphalium affine extracts on xanthine oxidase (XO) activity in vitro and to analyze the mechanism of this effect. Methods In this in vitro study, Kinetic measurements were performed in 4 different inhibitor concentrations and 5 different xanthine concentrations (60, 100, 200, 300, 400 μmol/L). Dixon and Lineweaver-Burk plot analysis were used to determine Ki values and the inhibition mode for the compounds isolated from Gnaphalium affine extract. Results Four potent xanthine oxidase inhibitors were found in 95% ethanolic (v/v) Gnaphalium affine extract. Among them, the flavone Eupatilin exhibited the strongest inhibitory effect on XO with a inhibition constant (Ki) of 0.37μmol/L, lower than the Ki of allopurinol (4.56 mol/L), a known synthetic XO inhibitor. Apigenin (Ki of 0.56μmol/L, a proportion of 0.0053‰in Gnaphalium affine), luteolin (Ki of 2.63 μmol/L, 0.0032‰ in Gnaphalium affine) and 5-hydroxy-6,7,3’,4’-tetramethoxyflavone (Ki of 3.15μmol/L, 0.0043‰ in Gnaphalium affine) also contributed to the inhibitory effect of Gnaphalium affine extract on XO activity. Conclusions These results suggest that the use of Gnaphalium affine in the treatment of gout could be attributed to its inhibitory effect on XO. This study provides a rational basis for the traditional use of Gnaphalium affine against gout.

  18. ATLAS boosted object tagging 2

    CERN Document Server

    Caudron, Julien; The ATLAS collaboration

    2015-01-01

    A detailed study into the optimal techniques for identifying boosted hadronically decaying W or Z bosons is presented. Various algorithms for reconstructing, grooming and tagging bosonic jets are compared for W bosons with a wide range of transverse momenta using 8 TeV data and 8 TeV and 13 TeV MC simulations. In addition, given that a hadronic jet has been identified as resulting from the hadronic decay of a W or Z, a technique is developed to discriminate between W and Z bosons. The modeling of the tagging variables used in this technique is studied using 8 TeV pp collision data and systematic uncertainties for the tagger efficiency and fake rates are evaluated.

  19. Production of tag-free recombinant fusion protein encompassing promiscuous T cell epitope of tetanus toxoid and dog zona pellucida glycoprotein-3 for contraceptive vaccine development.

    Science.gov (United States)

    Gupta, Neha; Shrestha, Abhinav; Panda, Amulya Kumar; Gupta, Satish Kumar

    2013-07-01

    Affinity tags can interfere in various physicochemical properties and immunogenicity of the recombinant proteins. In the present study, tag-free recombinant fusion protein encompassing promiscuous T cell epitope of tetanus toxoid [TT; amino acid (aa) residues 830-844] followed by dilysine linker and dog zona pellucida glycoprotein-3 (ZP3; aa residues 23-348) (TT-KK-ZP3) was expressed in Escherichia coli. The recombinant protein, expressed as inclusion bodies (IBs), was purified by isolation of IBs, processed to remove host cell proteins, followed by solubilization and refolding. A specific 39 kDa protein including ZP3 was identified by SDS-PAGE. CD spectra showed the presence of α-helices and β-sheets, and fluorescent spectroscopy revealed emission maxima of 265 A.U. at 339 nm for refolded protein and showed red shift in the presence of 6 M guanidine hydrochloride. Immunization of inbred FvB/J female mice with purified recombinant TT-KK-ZP3 (25 μg/animal) led to generation of high antibody titers against the recombinant protein. The antibodies reacted specifically with ZP matrix surrounding mouse oocytes. Immunized mice showed significant reduction in fertility as compared to the control group. The studies described herein provide a simple method to produce and purify tag-free recombinant protein for the development of a contraceptive vaccine.

  20. Tag Clusters as Information Retrieval Interfaces

    CERN Document Server

    Knautz, Kathrin; Stock, Wolfgang G

    2010-01-01

    The paper presents our design of a next generation information retrieval system based on tag co-occurrences and subsequent clustering. We help users getting access to digital data through information visualization in the form of tag clusters. Current problems like the absence of interactivity and semantics between tags or the difficulty of adding additional search arguments are solved. In the evaluation, based upon SERVQUAL and IT systems quality indicators, we found out that tag clusters are perceived as more useful than tag clouds, are much more trustworthy, and are more enjoyable to use.

  1. Plastic-casting intrinsic-surface unique identifier (tag)

    Energy Technology Data Exchange (ETDEWEB)

    Palm, R.G.; De Volpi, A.

    1995-04-01

    This report describes the development of an authenticated intrinsic-surf ace tagging method for unique- identification of controlled items. Although developed for control of items limited by an arms control treaty, this method has other potential applications to keep track of critical or high-value items. Each tag (unique-identifier) consists of the intrinsic, microscopic surface topography of a small designated area on a controlled item. It is implemented by making a baseline plastic casting of the designated tag area and usually placing a cover (for example, a bar-code label) over this area to protect the surface from environmental alteration. The plastic casting is returned to a laboratory and prepared for high-resolution scanning electron microscope imaging. Several images are digitized and stored for use as a standard for authentication of castings taken during future inspections. Authentication is determined by numerically comparing digital images. Commercially available hardware and software are used for this tag. Tag parameters are optimized, so unique casting images are obtained from original surfaces, and images obtained from attempted duplicate surfaces are detected. This optimization uses the modulation transfer function, a first principle of image analysis, to determine the parameters. Surface duplication experiments confirmed the optimization.

  2. Approximation properties of haplotype tagging

    Directory of Open Access Journals (Sweden)

    Dreiseitl Stephan

    2006-01-01

    Full Text Available Abstract Background Single nucleotide polymorphisms (SNPs are locations at which the genomic sequences of population members differ. Since these differences are known to follow patterns, disease association studies are facilitated by identifying SNPs that allow the unique identification of such patterns. This process, known as haplotype tagging, is formulated as a combinatorial optimization problem and analyzed in terms of complexity and approximation properties. Results It is shown that the tagging problem is NP-hard but approximable within 1 + ln((n2 - n/2 for n haplotypes but not approximable within (1 - ε ln(n/2 for any ε > 0 unless NP ⊂ DTIME(nlog log n. A simple, very easily implementable algorithm that exhibits the above upper bound on solution quality is presented. This algorithm has running time O((2m - p + 1 ≤ O(m(n2 - n/2 where p ≤ min(n, m for n haplotypes of size m. As we show that the approximation bound is asymptotically tight, the algorithm presented is optimal with respect to this asymptotic bound. Conclusion The haplotype tagging problem is hard, but approachable with a fast, practical, and surprisingly simple algorithm that cannot be significantly improved upon on a single processor machine. Hence, significant improvement in computatational efforts expended can only be expected if the computational effort is distributed and done in parallel.

  3. Enhancing Navigation on Wikipedia with Social Tags

    CERN Document Server

    Zubiaga, Arkaitz

    2012-01-01

    Social tagging has become an interesting approach to improve search and navigation over the actual Web, since it aggregates the tags added by different users to the same resource in a collaborative way. This way, it results in a list of weighted tags describing its resource. Combined to a classical taxonomic classification system such as that by Wikipedia, social tags can enhance document navigation and search. On the one hand, social tags suggest alternative navigation ways, including pivot-browsing, popularity-driven navigation, and filtering. On the other hand, it provides new metadata, sometimes uncovered by documents' content, that can substantially improve document search. In this work, the inclusion of an interface to add user-defined tags describing Wikipedia articles is proposed, as a way to improve article navigation and retrieval. As a result, a prototype on applying tags over Wikipedia is proposed in order to evaluate its effectiveness.

  4. Classification of neocortical interneurons using affinity propagation

    Directory of Open Access Journals (Sweden)

    Roberto eSantana

    2013-12-01

    Full Text Available In spite of over a century of research on cortical circuits, it is still unknown how many classes of cortical neurons exist. Neuronal classification has been a difficult problem because it is unclear what a neuronal cell class actually is and what are the best characteristics are to define them. Recently, unsupervised classifications using cluster analysis based on morphological, physiological or molecular characteristics, when applied to selected datasets, have provided quantitative and unbiased identification of distinct neuronal subtypes. However, better and more robust classification methods are needed for increasingly complex and larger datasets. We explored the use of affinity propagation, a recently developed unsupervised classification algorithm imported from machine learning, which gives a representative example or exemplar for each cluster. As a case study, we applied affinity propagation to a test dataset of 337 interneurons belonging to four subtypes, previously identified based on morphological and physiological characteristics. We found that affinity propagation correctly classified most of the neurons in a blind, non-supervised manner. In fact, using a combined anatomical/physiological dataset, our algorithm differentiated parvalbumin from somatostatin interneurons in 49 out of 50 cases. Affinity propagation could therefore be used in future studies to validly classify neurons, as a first step to help reverse engineer neural circuits.

  5. The use of ammonium sulphate precipitation method for the determination of antigen-binding capacity and affinity of anti-tetanus antibodies in human serum.

    Science.gov (United States)

    Alausa, O K

    1975-01-01

    Radioimmunoassay of anti-tetanus antibodies produced in human serum in response to tetanus toxoid immunization was carried out by employing the non-specific precipitation effect of ammonium sulphate described by Farr (1958). The results showed that the method was sensitive and was able to differentiate between immune and non-immune persons. The effects of booster dose injection on the quantity and quality of antibodies produced during immunization were discussed, and the possible use of the method to predict the amount of immunogen, the timing and number of injections required for optimal host response in immunization schedules was suggested.

  6. Glutamate dehydrogenase isoforms with N-terminal (His)6- or FLAG-tag retain their kinetic properties and cellular localization

    DEFF Research Database (Denmark)

    Pajęcka, Kamilla; Nielsen, Camilla Wendel; Hauge, Anne

    2014-01-01

    containing N-terminal (His)6 tags were successfully expressed in Sf9 cells and the recombinant proteins were isolated to ≥95 % purity in a two-step procedure involving ammonium sulfate precipitation and Ni(2+)-based immobilized metal ion affinity chromatography. To explore whether the presence of the FLAG...

  7. Affinity chromatography of phosphorylated proteins.

    Science.gov (United States)

    Tchaga, Grigoriy S

    2008-01-01

    This chapter covers the use of immobilized metal ion affinity chromatography (IMAC) for enrichment of phosphorylated proteins. Some requirements for successful enrichment of these types of proteins are discussed. An experimental protocol and a set of application data are included to enable the scientist to obtain high-yield results in a very short time with pre-packed phospho-specific metal ion affinity resin (PMAC).

  8. Oriented angles in affine space

    Directory of Open Access Journals (Sweden)

    Włodzimierz Waliszewski

    2004-05-01

    Full Text Available The concept of a smooth oriented angle in an arbitrary affine space is introduced. This concept is based on a kinematics concept of a run. Also, a concept of an oriented angle in such a space is considered. Next, it is shown that the adequacy of these concepts holds if and only if the affine space, in question, is of dimension 2 or 1.

  9. Profile-QSAR: a novel meta-QSAR method that combines activities across the kinase family to accurately predict affinity, selectivity, and cellular activity.

    Science.gov (United States)

    Martin, Eric; Mukherjee, Prasenjit; Sullivan, David; Jansen, Johanna

    2011-08-22

    Profile-QSAR is a novel 2D predictive model building method for kinases. This "meta-QSAR" method models the activity of each compound against a new kinase target as a linear combination of its predicted activities against a large panel of 92 previously studied kinases comprised from 115 assays. Profile-QSAR starts with a sparse incomplete kinase by compound (KxC) activity matrix, used to generate Bayesian QSAR models for the 92 "basis-set" kinases. These Bayesian QSARs generate a complete "synthetic" KxC activity matrix of predictions. These synthetic activities are used as "chemical descriptors" to train partial-least squares (PLS) models, from modest amounts of medium-throughput screening data, for predicting activity against new kinases. The Profile-QSAR predictions for the 92 kinases (115 assays) gave a median external R²(ext) = 0.59 on 25% held-out test sets. The method has proven accurate enough to predict pairwise kinase selectivities with a median correlation of R²(ext) = 0.61 for 958 kinase pairs with at least 600 common compounds. It has been further expanded by adding a "C(k)XC" cellular activity matrix to the KxC matrix to predict cellular activity for 42 kinase driven cellular assays with median R²(ext) = 0.58 for 24 target modulation assays and R²(ext) = 0.41 for 18 cell proliferation assays. The 2D Profile-QSAR, along with the 3D Surrogate AutoShim, are the foundations of an internally developed iterative medium-throughput screening (IMTS) methodology for virtual screening (VS) of compound archives as an alternative to experimental high-throughput screening (HTS). The method has been applied to 20 actual prospective kinase projects. Biological results have so far been obtained in eight of them. Q² values ranged from 0.3 to 0.7. Hit-rates at 10 uM for experimentally tested compounds varied from 25% to 80%, except in K5, which was a special case aimed specifically at finding "type II" binders, where none of the compounds were predicted to be

  10. Affinity purification of influenza virus ribonucleoprotein complexes from the chromatin of infected cells.

    Science.gov (United States)

    Chase, Geoffrey P; Schwemmle, Martin

    2012-06-03

    two salt extraction steps: first using 150 mM NaCl (termed "ch150"), then 500 mM NaCl ("ch500") (Fig. 1). These salt extraction steps were chosen based on our observation that 500 mM NaCl was sufficient to solubilize over 85% of nuclear vRNPs yet still allow binding of tagged vRNPs to the affinity matrix. After subcellular fractionation of infected cells, it is possible to affinity purify PB2-tagged vRNPs from each individual fraction and analyze their protein and RNA components using Western Blot and primer extension, respectively. Recently, we utilized this method to discover that vRNP export complexes form during late points after infection on the chromatin fraction extracted with 500 mM NaCl (ch500).

  11. Identifying novel genes in C. elegans using SAGE tags

    Directory of Open Access Journals (Sweden)

    Chen Nansheng

    2010-12-01

    Full Text Available Abstract Background Despite extensive efforts devoted to predicting protein-coding genes in genome sequences, many bona fide genes have not been found and many existing gene models are not accurate in all sequenced eukaryote genomes. This situation is partly explained by the fact that gene prediction programs have been developed based on our incomplete understanding of gene feature information such as splicing and promoter characteristics. Additionally, full-length cDNAs of many genes and their isoforms are hard to obtain due to their low level or rare expression. In order to obtain full-length sequences of all protein-coding genes, alternative approaches are required. Results In this project, we have developed a method of reconstructing full-length cDNA sequences based on short expressed sequence tags which is called sequence tag-based amplification of cDNA ends (STACE. Expressed tags are used as anchors for retrieving full-length transcripts in two rounds of PCR amplification. We have demonstrated the application of STACE in reconstructing full-length cDNA sequences using expressed tags mined in an array of serial analysis of gene expression (SAGE of C. elegans cDNA libraries. We have successfully applied STACE to recover sequence information for 12 genes, for two of which we found isoforms. STACE was used to successfully recover full-length cDNA sequences for seven of these genes. Conclusions The STACE method can be used to effectively reconstruct full-length cDNA sequences of genes that are under-represented in cDNA sequencing projects and have been missed by existing gene prediction methods, but their existence has been suggested by short sequence tags such as SAGE tags.

  12. Multifunctional g3p-peptide tag for current phage display systems.

    Science.gov (United States)

    Beckmann, C; Haase, B; Timmis, K N; Tesar, M

    1998-03-15

    . The high sensitivity of the antibody for the peptide tag was reflected in the antibody affinity constant K(D) of 6.80 x 10(-10) M, which was determined by real time biomolecular interaction analysis (BIA) based on surface plasmon resonance (SPR) [Karlsson, R., Fält, A., 1997. Experimental design for kinetic analysis of protein-protein interactions with surface plasmon resonance biosensors. J. Immunol. Methods 200, 121-133]. Finally, recombinant proteins in E. coli periplasmic extracts could be purified in a single step by affinity purification using immobilized mAb 10C3. These studies demonstrated that the new peptide-tag and its corresponding mAb represents a versatile tool for the detection of recombinant proteins selected by phage display technology.

  13. Promoting Tag Removal of a MBP-Fused Integral Membrane Protein by TEV Protease.

    Science.gov (United States)

    Chen, Yanke; Li, Qichang; Yang, Jun; Xie, Hao

    2017-03-01

    Tag removal is a prerequisite issue for structural and functional analysis of affinity-purified membrane proteins. The present study took a MBP-fused membrane protein, MrpF, as a model to investigate the tag removal by TEV protease. Influences of the linking sequence between TEV cleavage site and MrpF on protein expression and predicted secondary structure were investigated. The steric accessibility of TEV protease to cleavage site of MBP-fused MrpF was explored. It was found that reducing the size of hydrophilic group of detergents and/or extending the linking sequence between cleavage site and target protein can significantly improve the accessibility of the cleavage site and promote tag removal by TEV protease.

  14. Affinity Monolith-Integrated Microchips for Protein Purification and Concentration.

    Science.gov (United States)

    Gao, Changlu; Sun, Xiuhua; Wang, Huaixin; Qiao, Wei; Hu, Bo

    2016-01-01

    Affinity chromatography is a valuable method to purify and concentrate minute amount of proteins. Monoliths with epoxy groups for affinity immobilization were prepared by direct in-situ photopolymerization of glycidyl methacrylate and ethylene glycol dimethacrylate in porogenic solvents consisting of 1-dodecanol and cyclohexanol. By integrating affinity monoliths onto a microfluidic system, targeted biomolecules can be captured and retained on affinity column, while other biomolecules having no specific interactions toward the immobilized ligands flow through the microchannel. Therefore, proteins which remain on the affinity column are purified and concentrated, and then eluted by appropriate solutions and finally, separated by microchip capillary electrophoresis. This integrated microfluidic device has been applied to the purification and separation of specific proteins (FITC-labeled human serum albumin and IgG) in a mixture.

  15. Decoration of silicon nanostructures with copper particles for simultaneous selective capture and mass spectrometry detection of His-tagged model peptide.

    Science.gov (United States)

    Coffinier, Yannick; Kurylo, Ievgen; Drobecq, Hervé; Szunerits, Sabine; Melnyk, Oleg; Zaitsev, Vladimir N; Boukherroub, Rabah

    2014-10-21

    We present in this work a simple and fast preparation method of a new affinity surface-assisted laser/desorption ionization mass spectrometry (SALDI-MS) substrate based on silicon nanostructures decorated with copper particles. The silicon nanostructures were fabricated by the metal-assisted chemical etching (MACE) method. Then, superhydrophilic areas surrounded by superhydrophobic regions were formed through hydrosilylation reaction of 1-octadecene, followed by local degradation of the octadecyl layer. After that, copper particles were deposited in the hydrophilic areas by using the electroless method. We have demonstrated that these surfaces were able to perform high selective capture of model His-tag peptide even in a complex mixture such as serum solution. Then, the captured peptide was detected by mass spectrometry at a femtomolar level without the need of organic matrix.

  16. Treelicious: a System for Semantically Navigating Tagged Web Pages

    CERN Document Server

    Mullins, Matt; 10.1109/WI-IAT.2010.289

    2011-01-01

    Collaborative tagging has emerged as a popular and effective method for organizing and describing pages on the Web. We present Treelicious, a system that allows hierarchical navigation of tagged web pages. Our system enriches the navigational capabilities of standard tagging systems, which typically exploit only popularity and co-occurrence data. We describe a prototype that leverages the Wikipedia category structure to allow a user to semantically navigate pages from the Delicious social bookmarking service. In our system a user can perform an ordinary keyword search and browse relevant pages but is also given the ability to broaden the search to more general topics and narrow it to more specific topics. We show that Treelicious indeed provides an intuitive framework that allows for improved and effective discovery of knowledge.

  17. Flavor Tagging at the Tevatron, Including Calibration and Control

    Science.gov (United States)

    Moulik, T.; DØ Collaboration; CDF Collaboration

    2007-08-01

    This report summarizes the flavor tagging techniques developed at the CDF and DØ experiments. Flavor tagging involves identification of the B meson flavor at production, whether its constituent is a b quark or an anti-quark. quark flavor content and hence the decay products do not identify the B flavor content at production. It is crucial for measuring the oscillation frequency of neutral B mesons, both in the B0 and Bs0 system. The two experiments have developed their unique approaches to flavor tagging, using neural networks and likelihood methods to disentangle tracks from b decays from other tracks. This report discusses these techniques and the measurement of B0 mixing as a means to calibrate the taggers.

  18. Inclusive tagging of B-flavour at LHCb [Vidyo

    CERN Document Server

    CERN. Geneva

    2017-01-01

    One of the most important procedure needed for the study of CP violation in Beauty sector is the tagging of the flavour of neutral B-mesons at production. The harsh environment of the Large Hadron Collider makes it particularly hard to succeed in this task. We present a proposal to upgrade current flavour tagging strategy in LHCb experiment. This strategy consists of inclusive tagging ensemble methods (i.e: the use inclusive information about the event without a firm selection rule), which are combined using a probabilistic model for each event. The probabilistic model uses all reconstructed tracks and secondary vertices to obtain well-determined probability of B flavour at production. Such approach reduces the dependence on the performance of lower level identification capacities and thus has the potential to increase the overall performance.

  19. A simplified method to attach antibodies on liposomes by biotin-streptavidin affinity for rapid and economical screening of targeted liposomes.

    Science.gov (United States)

    Papadia, Konstantina; Markoutsa, Eleni; Antimisiaris, Sophia G

    2014-05-01

    The biotin-Streptavidin (STREP) technique for attachment of monoclonal antibodies (mAbs) (or other ligand types) on liposome surface offers high attachment yield, however it is time consuming and expensive due to the number of steps used and the consumption of large quantities of STREP. Herein, a simplified, fast and economic technique, by incubating pre-mixed biotin-mAb/STREP with biotin-liposomes, at a 3:1:1 biotin-mAb/STREP/biotin-LIP ratio (mol/mol/mol) was evaluated. The physichochemical properties, final mAb attachment yield and targeting potential of liposomes decorated with an anti-transferrin receptor mAb (TfR-mAb), prepared by the simple method (SM) and the conventional method (CM), were compared. The vesicle uptake by hCMEC/D3 cells (known to overexpress TfR) were considered as a measure of liposome targeting capability. Results show that both targeted liposome types (SM and CM) have small size (mean diameters around 150 nm), low poly-dispersity (approx. 0.20) and similar mAb attachment yield (between 64-88%). However, the uptake of the SM-liposomes is slightly lower compared to CM-LIP (24-30% decrease), suggesting that the modulated conformation of mAbs on the liposome surface (triplets attached to one single STREP molecule) results in decreased targeting capability. Nevertheless, the simpler and faster one-step preparation procedure which has very high lipid recovery (> 95%) compared to the CM (50-60%) and 15-30 times lower consumption of STREP, may be a good alternative for initial screening of various mAbs as ligands for targeted liposomal or other nanotechnologies, during pre-clinical development.

  20. AFFINE TRANSFORMATION IN RANDOM ITERATED FUNCTION SYSTEMS

    Institute of Scientific and Technical Information of China (English)

    熊勇; 史定华

    2001-01-01

    Random iterated function systems (IFSs) is discussed, which is one of the methods for fractal drawing. A certain figure can be reconstructed by a random IFS. One approach is presented to determine a new random IFS, that the figure reconstructed by the new random IFS is the image of the origin figure reconstructed by old IFS under a given affine transformation. Two particular examples are used to show this approach.

  1. An inexpensive and simple method for thermally stable immobilization of DNA on an unmodified glass surface: UV linking of poly(T)10-poly(C)10-tagged DNA probes

    DEFF Research Database (Denmark)

    Guðnason, Haukur; Dufva, Hans Martin; Bang, Dang Duong;

    2008-01-01

    be linked by UV light irradiation onto a plain, unmodified glass surface. Probes immobilized onto unmodified glass microscope slides performed similarly to probes bound to commercial amino-silane-coated slides and had comparable detection limits. The TC-tagged probes linked to unmodified glass did not show...

  2. Measurement of the tau tag efficiency using the Z ->tau tau -> mu + hadrons +X events

    CERN Document Server

    Kalinowski, Artur

    2006-01-01

    A method for the measurement of the tau tag efficiency for the taus coming from the Z boson decay is presented. Full simulation studies show that the tau tag efficiency can be measured with the precision of 9% including the systematic uncertainties due to the calorimetry scale uncertainty.

  3. ASIFT: An Algorithm for Fully Affine Invariant Comparison

    Directory of Open Access Journals (Sweden)

    Guoshen Yu

    2011-02-01

    Full Text Available If a physical object has a smooth or piecewise smooth boundary, its images obtained by cameras in varying positions undergo smooth apparent deformations. These deformations are locally well approximated by affine transforms of the image plane. In consequence the solid object recognition problem has often been led back to the computation of affine invariant image local features. The similarity invariance (invariance to translation, rotation, and zoom is dealt with rigorously by the SIFT method The method illustrated and demonstrated in this work, Affine-SIFT (ASIFT, simulates a set of sample views of the initial images, obtainable by varying the two camera axis orientation parameters, namely the latitude and the longitude angles, which are not treated by the SIFT method. Then it applies the SIFT method itself to all images thus generated. Thus, ASIFT covers effectively all six parameters of the affine transform.

  4. HAHAHA, HEHEHE, HIHIHI, or HKHKHK: influence of position and composition of histidine containing tags on biodistribution of [(99m)Tc(CO)3](+)-labeled affibody molecules.

    Science.gov (United States)

    Hofström, Camilla; Altai, Mohamed; Honarvar, Hadis; Strand, Joanna; Malmberg, Jennie; Hosseinimehr, Seyed Jalal; Orlova, Anna; Gräslund, Torbjörn; Tolmachev, Vladimir

    2013-06-27

    Engineered affibody molecules can be used for high contrast in vivo molecular imaging. Extending a recombinantly produced HER2 binding affibody molecule with a hexa-histidine tag allows for convenient purification by immobilized metal-ion affinity chromatography and labeling with [(99m)Tc(CO)3](+) but increases radioactivity uptake in the liver. To investigate the impact of charge, lipophilicity, and position on biodistribution, 10 variants of a histidine-based tag was attached to a HER2 binding affibody molecule. The biochemical properties and the HER2 binding affinity appeared to be similar for all variants. In vivo, positive charge promoted liver uptake. For N-terminally placed tags, lipophilicity promoted liver uptake and decreased kidney uptake. Kidney uptake was higher for C-terminally placed tags compared to their N-terminal counterparts. The variant with the amino acid composition HEHEHE placed in the N-terminus gave the lowest nonspecific uptake.

  5. Digital magnetic tagging for multiplexed suspension-based biochemical assays

    Science.gov (United States)

    Mitrelias, T.; Trypiniotis, T.; Palfreyman, J. J.; Hong, B.; Vyas, K.; Hayward, T. J.; Llandro, J.; Kopper, K. P.; Bland, J. A. C.; Robertson, P. A.; Barnes, C. H. W.

    2009-04-01

    Microarrays and suspension (or bead)-based technologies have attracted significant interest for their broad applications in high throughput molecular biology. However, the throughput of microarrays will always be limited by the array density and the slow diffusion of molecules to their binding sites. Suspension-based technologies, in which all the reactions take place directly on the surface of microcarriers functionalized with molecular probes, could offer true multiplexing due to the possibility of extending their detection capability by a straightforward expansion of the size of the chemical library of probes. To fully exploit their potential, the microcarriers must be tagged, but the number of distinct codes available from spectrometric/graphical/physical encoding methods is currently fairly limited. A digital magnetic tagging method based on magnetic microtags, which have been anisotropy engineered to provide stable magnetization directions which correspond to digital codes, is reported. The tags can be suspended in solution and functionalized with a variety of biological molecular probes. Magnetic tagging offers several benefits compared to the traditional optical encoding techniques currently employed. It offers minimal background signals, potential for a large number of distinct codes, miniaturization of devices, and the ability to write a code in situ. Experimental data showing the reading of individual magnetic microbars from samples comprising 50×20 μm2 Ni elements, as well as micromagnetic simulations that show the feasibility of stray field detection, are presented. The stray fields of the magnetic microbars spanning a range of 60 mOe were detected by a microfabricated fluxgate sensor scanned in a raster fashion over the sample that was placed about 70 μm away. Free floating tags have also been fabricated for use in microfluidic systems. A magnetic lab-on-a-chip device could be used for tagging biomolecular probes for applications in genome

  6. In vitro affinity screening of protein and peptide binders by megavalent bead surface display.

    Science.gov (United States)

    Diamante, Letizia; Gatti-Lafranconi, Pietro; Schaerli, Yolanda; Hollfelder, Florian

    2013-10-01

    The advent of protein display systems has provided access to tailor-made protein binders by directed evolution. We introduce a new in vitro display system, bead surface display (BeSD), in which a gene is mounted on a bead via strong non-covalent (streptavidin/biotin) interactions and the corresponding protein is displayed via a covalent thioether bond on the DNA. In contrast to previous monovalent or low-copy bead display systems, multiple copies of the DNA and the protein or peptide of interest are displayed in defined quantities (up to 10(6) of each), so that flow cytometry can be used to obtain a measure of binding affinity. The utility of the BeSD in directed evolution is validated by library selections of randomized peptide sequences for binding to the anti-hemagglutinin (HA) antibody that proceed with enrichments in excess of 10(3) and lead to the isolation of high-affinity HA-tags within one round of flow cytometric screening. On-bead K(d) measurements suggest that the selected tags have affinities in the low nanomolar range. In contrast to other display systems (such as ribosome, mRNA and phage display) that are limited to affinity panning selections, BeSD possesses the ability to screen and rank binders by their affinity in vitro, a feature that hitherto has been exclusive to in vivo multivalent cell display systems (such as yeast display).

  7. Hypergraph model of social tagging networks

    CERN Document Server

    Zhang, Zi-Ke

    2010-01-01

    The past few years have witnessed the great success of a new family of paradigms, so-called folksonomy, which allows users to freely associate tags to resources and efficiently manage them. In order to uncover the underlying structures and user behaviors in folksonomy, in this paper, we propose an evolutionary hypergrah model to explain the emerging statistical properties. The present model introduces a novel mechanism that one can not only assign tags to resources, but also retrieve resources via collaborative tags. We then compare the model with a real-world dataset: \\emph{Del.icio.us}. Indeed, the present model shows considerable agreement with the empirical data in following aspects: power-law hyperdegree distributions, negtive correlation between clustering coefficients and hyperdegrees, and small average distances. Furthermore, the model indicates that most tagging behaviors are motivated by labeling tags to resources, and tags play a significant role in effectively retrieving interesting resources and ...

  8. Onboard tagging for smart medical devices.

    Science.gov (United States)

    Li, Kejia; Warren, Steve

    2011-01-01

    Most medical devices are 'dumb:' their role is to acquire, display, and forward data. They make few if any operational decisions based on those data. Onboard tagging is a means whereby a device can embed information about itself, its data, and the sensibility of those data into its data stream. This diagnostic add-on offers a move toward 'smart' devices that will have the ability to affect changes in operational modes based on onboard contextual decision making, such as decisions to avoid needless wireless transmission of corrupt data. This paper presents a description of three types of onboard tags that relate to device hardware (type I tag), signal statistics (type II tag), and signal viability for the intended application (type III tag). A custom wireless pulse oximeter is presented as a use case to show how type II and III tags that convey photoplethysmogram (PPG) statistics and usability specifiers can be calculated and embedded into the data stream without degrading performance.

  9. Building Tag Clouds in Perl and PHP

    CERN Document Server

    Bumgardner, Jim

    2006-01-01

    Tag clouds are everywhere on the web these days. First popularized by the web sites Flickr, Technorati, and del.icio.us, these amorphous clumps of words now appear on a slew of web sites as visual evidence of their membership in the elite corps of "Web 2.0." This PDF analyzes what is and isn't a tag cloud, offers design tips for using them effectively, and then goes on to show how to collect tags and display them in the tag cloud format. Scripts are provided in Perl and PHP. Yes, some have said tag clouds are a fad. But as you will see, tag clouds, when used properly, have real merits. More

  10. Mass spectrometry-based methods for detection and differentiation of botulinum neurotoxins

    Science.gov (United States)

    Schmidt, Jurgen G.; Boyer, Anne E.; Kalb, Suzanne R.; Moura, Hercules; Barr, John R.; Woolfitt, Adrian R.

    2009-11-03

    The present invention is directed to a method for detecting the presence of clostridial neurotoxins in a sample by mixing a sample with a peptide that can serve as a substrate for proteolytic activity of a clostridial neurotoxin; and measuring for proteolytic activity of a clostridial neurotoxin by a mass spectroscopy technique. In one embodiment, the peptide can have an affinity tag attached at two or more sites.

  11. Understanding why users tag: A survey of tagging motivation literature and results from an empirical study

    Science.gov (United States)

    Strohmaier, Markus; Körner, Christian; Kern, Roman

    2012-01-01

    While recent progress has been achieved in understanding the structure and dynamics of social tagging systems, we know little about the underlying user motivations for tagging, and how they influence resulting folksonomies and tags. This paper addresses three issues related to this question. (1) What distinctions of user motivations are identified by previous research, and in what ways are the motivations of users amenable to quantitative analysis? (2) To what extent does tagging motivation vary across different social tagging systems? (3) How does variability in user motivation influence resulting tags and folksonomies? In this paper, we present measures to detect whether a tagger is primarily motivated by categorizing or describing resources, and apply these measures to datasets from seven different tagging systems. Our results show that (a) users’ motivation for tagging varies not only across, but also within tagging systems, and that (b) tag agreement among users who are motivated by categorizing resources is significantly lower than among users who are motivated by describing resources. Our findings are relevant for (1) the development of tag-based user interfaces, (2) the analysis of tag semantics and (3) the design of search algorithms for social tagging systems. PMID:23471473

  12. Trinity:Walking on a User-Ob ject-Tag Heterogeneous Network for Personalised Recommendations

    Institute of Scientific and Technical Information of China (English)

    Ming-Xin Gan; Lily Sun; Rui Jiang

    2016-01-01

    The rapid evolution of the Internet has been appealing for effective recommender systems to pinpoint useful information from online resources. Although historical rating data has been widely used as the most important information in recommendation methods, recent advancements have been demonstrating the improvement in recommendation performance with the incorporation of tag information. Furthermore, the availability of tag annotations has been well addressed by such fruitful online social tagging applications as CiteULike, MovieLens and BibSonomy, which allow users to express their preferences, upload resources and assign their own tags. Nevertheless, most existing tag-aware recommendation approaches model relationships among users, objects and tags using a tripartite graph, and hence overlook relationships within the same types of nodes. To overcome this limitation, we propose a novel approach, Trinity, to integrate historical data and tag information towards personalised recommendation. Trinity constructs a three-layered object-user-tag network that considers not only interconnections between different types of nodes but also relationships within the same types of nodes. Based on this heterogeneous network, Trinity adopts a random walk with restart model to assign the strength of associations to candidate objects, thereby providing a means of prioritizing the objects for a query user. We validate our approach via a series of large-scale 10-fold cross-validation experiments and evaluate its performance using three comprehensive criteria. Results show that our method outperforms several existing methods, including supervised random walk with restart, simulation of resource allocating processes, and traditional collaborative filtering.

  13. Learner Corpora without Error Tagging

    Directory of Open Access Journals (Sweden)

    Rastelli, Stefano

    2009-01-01

    Full Text Available The article explores the possibility of adopting a form-to-function perspective when annotating learner corpora in order to get deeper insights about systematic features of interlanguage. A split between forms and functions (or categories is desirable in order to avoid the "comparative fallacy" and because – especially in basic varieties – forms may precede functions (e.g., what resembles to a "noun" might have a different function or a function may show up in unexpected forms. In the computer-aided error analysis tradition, all items produced by learners are traced to a grid of error tags which is based on the categories of the target language. Differently, we believe it is possible to record and make retrievable both words and sequence of characters independently from their functional-grammatical label in the target language. For this purpose at the University of Pavia we adapted a probabilistic POS tagger designed for L1 on L2 data. Despite the criticism that this operation can raise, we found that it is better to work with "virtual categories" rather than with errors. The article outlines the theoretical background of the project and shows some examples in which some potential of SLA-oriented (non error-based tagging will be possibly made clearer.

  14. HAHAHA, HEHEHE, HIHIHI, or HKHKHK : Influence of Position and Composition of Histidine Containing Tags on Biodistribution of [99mTc(CO)3]+-Labeled Affibody Molecules

    OpenAIRE

    2013-01-01

    Engineered affibody molecules can be used for high contrast in vivo molecular imaging. Extending a recombinantly produced HER2 binding affibody molecule with a hexa-histidine tag allows for convenient purification by immobilized metal-ion affinity chromatography and labeling with [99mTc(CO)3]+ but increases radioactivity uptake in the liver. To investigate the impact of charge, lipophilicity, and position on biodistribution, 10 variants of a histidine-based tag was attached to a HER2 binding ...

  15. Optimization of tagged MRI for quantification of liver stiffness using computer simulated data.

    Directory of Open Access Journals (Sweden)

    Serena Monti

    Full Text Available The heartbeat has been proposed as an intrinsic source of motion that can be used in combination with tagged Magnetic Resonance Imaging (MRI to measure displacements induced in the liver as an index of liver stiffness. Optimizing a tagged MRI acquisition protocol in terms of sensitivity to these displacements, which are in the order of pixel size, is necessary to develop the method as a quantification tool for staging fibrosis. We reproduced a study of cardiac-induced strain in the liver at 3T and simulated tagged MR images with different grid tag patterns to evaluate the performance of the Harmonic Phase (HARP image analysis method and its dependence on the parameters of tag spacing and grid angle. The Partial Volume Effect (PVE, T1 relaxation, and different levels of noise were taken into account. Four displacement fields of increasing intensity were created and applied to the tagged MR images of the liver. These fields simulated the deformation at different liver stiffnesses. An Error Index (EI was calculated to evaluate the estimation accuracy for various parameter values. In the absence of noise, the estimation accuracy of the displacement fields increased as tag spacings decreased. EIs for each of the four displacement fields were lower at 0° and the local minima of the EI were found to correspond to multiples of pixel size. The accuracy of the estimation decreased for increasing levels of added noise; as the level increased, the improved estimation caused by decreasing the tag spacing tended to zero. The optimal tag spacing turned out to be a compromise between the smallest tag period that is a multiple of the pixel size and is achievable in a real acquisition and the tag spacing that guarantees an accurate liver displacement measure in the presence of realistic levels of noise.

  16. False positive RNA binding activities after Ni-affinity purification from Escherichia coli.

    Science.gov (United States)

    Milojevic, Tetyana; Sonnleitner, Elisabeth; Romeo, Alessandra; Djinović-Carugo, Kristina; Bläsi, Udo

    2013-06-01

    A His-tag is often added by means of recombinant DNA technology to a heterologous protein of interest, which is then over-produced in Escherchia coli and purified by one-step immobilized metal-affinity chromatography (IMAC). Owing to the presence of 24 histidines at the C-termini of the hexameric E. coli RNA chaperone Hfq, the protein co-purifies with His-tagged proteins of interest. As Hfq can bind to distinct RNA substrates with high affinity, its presence can obscure studies performed with (putative) RNA binding activities purified by IMAC. Here, we present results for a seemingly positive RNA-binding activity, exemplifying that false-positive results can be avoided if the protein of interest is either subjected to further purification step(s) or produced in an E. coli hfq- strain.

  17. Effective Personalized Recommendation in Collaborative Tagging Systems

    CERN Document Server

    Zhang, Zi-Ke

    2009-01-01

    Recently, collaborative tagging systems have attracted more and more attention and have been widely applied in web systems. Tags provide highly abstracted information about personal preferences and item content, and are therefore potential to help in improving better personalized recommendations. In this paper, we propose a tag-based recommendation algorithm considering the personal vocabulary and evaluate it in a real-world dataset: Del.icio.us. Experimental results demonstrate that the usage of tag information can significantly improve the accuracy of personalized recommendations.

  18. Chasing polys: Interdisciplinary affinity and its connection to physics identity

    Science.gov (United States)

    Scott, Tyler D.

    This research is based on two motivations that merge by means of the frameworks of interdisciplinary affinity and physics identity. First, a goal of education is to develop interdisciplinary abilities in students' thinking and work. But an often ignored factor is students interests and beliefs about being interdisciplinary. Thus, this work develops and uses a framework called interdisciplinary affinity. It encompasses students interests in making connections across disciplines and their beliefs about their abilities to make those connections. The second motivation of this research is to better understand how to engage more students with physics. Physics identity describes how a student sees themselves in relation to physics. By understanding how physics identity is developed, researchers and educators can identify factors that increase interest and engagement in physics classrooms. Therefore, physics identity was used in conjunction with interdisciplinary affinity. Using a mixed methods approach, this research used quantitative data to identify the relationships interdisciplinary affinity has with physics identity and the physics classroom. These connections were explored in more detail using a case study of three students in a high school physics class. Results showed significant and positive relationships between interdisciplinary affinity and physics identity, including the individual interest and recognition components of identity. It also identified characteristics of physics classrooms that had a significant, positive relationship with interdisciplinary affinity. The qualitative case study highlighted the importance of student interest to the relationship between interdisciplinary affinity and physics identity. It also identified interest and mastery orientation as key to understanding the link between interdisciplinary affinity and the physics classroom. These results are a positive sign that by understanding interdisciplinary affinity and physics identity

  19. Sensitive carbohydrate detection using surface enhanced Raman tagging.

    Science.gov (United States)

    Vangala, Karthikeshwar; Yanney, Michael; Hsiao, Cheng-Te; Wu, Wells W; Shen, Rong-Fong; Zou, Sige; Sygula, Andrzej; Zhang, Dongmao

    2010-12-15

    Glycomic analysis is an increasingly important field in biological and biomedical research as glycosylation is one of the most important protein post-translational modifications. We have developed a new technique to detect carbohydrates using surface enhanced Raman spectroscopy (SERS) by designing and applying a Rhodamine B derivative as the SERS tag. Using a reductive amination reaction, the Rhodamine-based tag (RT) was successfully conjugated to three model carbohydrates (glucose, lactose, and glucuronic acid). SERS detection limits obtained with a 633 nm HeNe laser were ∼1 nM in concentration for all the RT-carbohydrate conjugates and ∼10 fmol in total sample consumption. The dynamic range of the SERS method is about 4 orders of magnitude, spanning from 1 nM to 5 μM. Ratiometric SERS quantification using isotope-substituted SERS internal references allows comparative quantifications of carbohydrates labeled with RT and deuterium/hydrogen substituted RT tags, respectively. In addition to enhancing the SERS detection of the tagged carbohydrates, the Rhodamine tagging facilitates fluorescence and mass spectrometric detection of carbohydrates. Current fluorescence sensitivity of RT-carbohydrates is ∼3 nM in concentration while the mass spectrometry (MS) sensitivity is about 1 fmol, achieved with a linear ion trap electrospray ionization (ESI)-MS instrument. Potential applications that take advantage of the high SERS, fluorescence, and MS sensitivity of this SERS tagging strategy are discussed for practical glycomic analysis where carbohydrates may be quantified with a fluorescence and SERS technique and then identified with ESI-MS techniques.

  20. Artifacts reduction in strain maps of tagged magnetic resonance imaging using harmonic phase

    Directory of Open Access Journals (Sweden)

    Wang Daolei

    2015-01-01

    Full Text Available Tagged Magnetic Resonance Imaging (MRI is a noninvasive technique for examining myocardial function and deformation. Tagged MRI can also be used in quasistatic MR elastography to acquire strain maps of other biological soft tissues. Harmonic phase (HARP provides automatic and rapid analysis of tagged MR images for the quantification and visualization of myocardial strain. We propose a new artifact reduction method in strain maps. Image intensity of the DC component is estimated and subtracted from spatial modulation of magnetization (SPAMM tagged MR images. DC peak interference in harmonic phase extraction is greatly reduced after DC component subtraction. The proposed method is validated using both simulated and MR acquired tagged images. Strain maps are obtained with better accuracy and smoothness after DC component subtraction.

  1. Improved statistical analysis of budding yeast TAG microarrays revealed by defined spike-in pools.

    Science.gov (United States)

    Peyser, Brian D; Irizarry, Rafael A; Tiffany, Carol W; Chen, Ou; Yuan, Daniel S; Boeke, Jef D; Spencer, Forrest A

    2005-09-15

    Saccharomyces cerevisiae knockout collection TAG microarrays are an emergent platform for rapid, genome-wide functional characterization of yeast genes. TAG arrays report abundance of unique oligonucleotide 'TAG' sequences incorporated into each deletion mutation of the yeast knockout collection, allowing measurement of relative strain representation across experimental conditions for all knockout mutants simultaneously. One application of TAG arrays is to perform genome-wide synthetic lethality screens, known as synthetic lethality analyzed by microarray (SLAM). We designed a fully defined spike-in pool to resemble typical SLAM experiments and performed TAG microarray hybridizations. We describe a method for analyzing two-color array data to efficiently measure the differential knockout strain representation across two experimental conditions, and use the spike-in pool to show that the sensitivity and specificity of this method exceed typical current approaches.

  2. External tagging does not affect the feeding behavior of a coral reef fish, Chaetodon vagabundus (Pisces: Chaetodontidae)

    KAUST Repository

    Berumen, Michael L.

    2009-11-10

    Increasingly, the ability to recognize individual fishes is important for studies of population dynamics, ecology, and behavior. Although a variety of methods exist, external tags remain one of the most widely applied because they are both effective and cost efficient. However, a key assumption is that neither the tagging procedure nor the presence of a tag negatively affects the individual. While this has been demonstrated for relatively coarse metrics such as growth and survival, few studies have examined the impact of tags and tagging on more subtle aspects of behavior. We tagged adult vagabond butterflyfish (Chaetodon vagabundus) occupying a 30-ha insular reef in Kimbe Bay, Papua New Guinea, using a commonly-utilized t-bar anchor tag. We quantified and compared feeding behavior (bite rate), which is sensitive to stress, of tagged and untagged individuals over four separate sampling periods spanning 4 months post-tagging. Bite rates did not differ between tagged and untagged individuals at each sampling period and, combined with additional anecdotal observations of normal pairing behavior and successful reproduction, suggest that tagging did not adversely affect individuals. © Springer Science+Business Media B.V. 2009.

  3. Reconciling Knowledge in Social Tagging Web Services

    Science.gov (United States)

    Aranda-Corral, Gonzalo A.; Borrego-Díaz, Joaquín

    Sometimes we want to search for new information about topics but we can not find relevant results using our own knowledge (for example, our personal bookmarks). A potential solution could be the use of knowledge from other users to find what we are searching for. This solution implies that we can achieve some agreement on implicit semantics used by the other users. We call it Reconciliation of Knowledge. The aim of this paper is to show an agent-based method which lets us reconcile two different knowledge basis (associated with tagging systems) into a common language, obtaining a new one that allows the reconcilitiation of (part of) this knowledge. The agents use Formal Concept Analysis concepts and tools and it has been implemented on the JADE multiagent platform.

  4. Affine structures and a tableau model for E_6 crystals

    CERN Document Server

    Jones, Brant

    2009-01-01

    We provide the unique affine crystal structure for type E_6^{(1)} Kirillov-Reshetikhin crystals corresponding to the multiples of fundamental weights s Lambda_1, s Lambda_2, and s Lambda_6 for all s \\geq 1 (in Bourbaki's labeling of the Dynkin nodes, where 2 is the adjoint node). Our methods introduce a generalized tableaux model for classical highest weight crystals of type E and use the order three automorphism of the affine E_6^{(1)} Dynkin diagram. In addition, we provide a conjecture for the affine crystal structure of type E_7^{(1)} Kirillov-Reshetikhin crystals corresponding to the adjoint node.

  5. Extending birthday paradox theory to estimate the number of tags in RFID systems.

    Science.gov (United States)

    Shakiba, Masoud; Singh, Mandeep Jit; Sundararajan, Elankovan; Zavvari, Azam; Islam, Mohammad Tariqul

    2014-01-01

    The main objective of Radio Frequency Identification systems is to provide fast identification for tagged objects. However, there is always a chance of collision, when tags transmit their data to the reader simultaneously. Collision is a time-consuming event that reduces the performance of RFID systems. Consequently, several anti-collision algorithms have been proposed in the literature. Dynamic Framed Slotted ALOHA (DFSA) is one of the most popular of these algorithms. DFSA dynamically modifies the frame size based on the number of tags. Since the real number of tags is unknown, it needs to be estimated. Therefore, an accurate tag estimation method has an important role in increasing the efficiency and overall performance of the tag identification process. In this paper, we propose a novel estimation technique for DFSA anti-collision algorithms that applies birthday paradox theory to estimate the number of tags accurately. The analytical discussion and simulation results prove that the proposed method increases the accuracy of tag estimation and, consequently, outperforms previous schemes.

  6. Improving Recommendations in Tag-based Systems with Spectral Clustering of Tag Neighbors

    DEFF Research Database (Denmark)

    Pan, Rong; Xu, Guandong; Dolog, Peter

    2012-01-01

    Tag as a useful metadata reflects the collaborative and conceptual features of documents in social collaborative annotation systems. In this paper, we propose a collaborative approach for expanding tag neighbors and investigate the spectral clustering algorithm to filter out noisy tag neighbors...

  7. SERS-active nanoparticle aggregate technology for tags and seals

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Leif O [Los Alamos National Laboratory; Montoya, Velma M [Los Alamos National Laboratory; Havrilla, George J [Los Alamos National Laboratory; Doorn, Stephen K [Los Alamos National Laboratory

    2010-06-03

    In this paper, we describe our efforts to create a modern tagging and sealing technology for international safeguards application. Our passive tagging methods are based on SANAs (SERS-Active Nanoparticle Aggregates; SERS: Surface Enhanced Raman Scattering). These SANAs offer robust spectral barcoding capability in an inexpensive tag/seal, with the possibility of rapid in-field verification that requires no human input. At INMM 2009, we introduced SANAs, and showed approaches to integrating our technology with tags under development at Sandia National Laboratories (SNL). Here, we will focus on recent LANL development work, as well as adding additional dimensionality to the barcoding technique. The field of international safeguards employs a broad array of tags, seals, and tamper-indicating devices to assist with identification, tracking, and verification of components and materials. These devices each have unique strengths suited to specific applications, and span a range of technologies from passive metal cup seals and adhesive seals to active, remotely monitored fiber optic seals. Regardless of the technology employed, essential characteristics center around security, environmental and temporal stability, ease of use, and the ability to provide confidence to all parties. Here, we present a new inexpensive tagging technology that will deliver these attributes, while forming the basis of either a new seal, or as a secure layer added to many existing devices. Our approach uses the Surface Enhanced Raman Scattering (SERS) response from SANAs (SERS-Active Nanoparticle Aggregates, Figure 1) to provide a unique identifier or signature for tagging applications. SANAs are formed from gold or silver nanoparticles in the 40-80 nm size range. A chemical dye is installed on the nanoparticle surface, and the nanoparticles are then aggregated into ensembles of {approx}100 to 500 nm diameter, prior to being coated with silica. The silica shell protects the finished SANA from

  8. Strategy for improvement of enteropeptidase efficiency in tag removal processes.

    Science.gov (United States)

    Gasparian, Marine E; Bychkov, Maxim L; Dolgikh, Dmitry A; Kirpichnikov, Mikhail P

    2011-10-01

    Enteropeptidase (synonym: enterokinase, EC 3.4.21.9) is a heterodimeric serine protease of the intestinal brush border that activates trypsinogen by highly specific cleavage of the trypsinogen activation peptide following the sequence (Asp)(4)-Lys. It has also great biotechnological interest because of the unique substrate specificity of the serine protease domain. The high degree of specificity exhibited by enteropeptidase makes it a suitable reagent for cleaving recombinant proteins to remove affinity or other tags. However often unwanted cleavages elsewhere in the protein occurred during cleavage of fusions when high amount of enzyme is required. In this study we have improved the efficiency of fusion proteins cleavage by enteropeptidase by substitution of the Lys residue by Arg in specific cleavage sequence (Asp)(4)-Lys. We have demonstrated that 3-6-fold lower amounts of the catalytic subunit of human and bovine enteropeptidase is required for 95% cleavage of Trx/TRAIL and Trx/FGF-2 fusions with (Asp)(4)-Arg cleavage sequence in comparison to native sequence (Asp)(4)-Lys. As a result, reduced amount of non-specifically cleaved peptide fragments were observed during cleavage of (Asp)(4)-Lys/Arg mutated fusions. These findings overcome limitations of enteropeptidase in tag removal processes during recombinant proteins purification and extend its commercial benefit in the biopharmaceutical industry.

  9. To tag or not to tag: animal welfare, conservation and stakeholder considerations in fish tracking studies that use electronic tags

    Energy Technology Data Exchange (ETDEWEB)

    Cooke, Steven J.; Nguyen, Vivian M.; Murchie, Karen J.; Thiem, Jason D.; Donaldson, Michael R.; Hinch, Scott G.; Brown, Richard S.; Fisk, Aaron

    2013-11-01

    The advent and widespread adoption of electronic tags (including biotelemetry and biologging devices) for tracking animals has provided unprecedented information on the biology, management, and conservation of fish in the world’s oceans and inland waters. However, use of these tools is not without controversy. Even when scientific and management objectives may best be achieved using electronic tags, it is increasingly important to further consider other factors such as the welfare of tagged animals (i.e., the role of training and science-based surgical guidelines, anesthetic use, inability to maintain sterile conditions in field environments), the ethics of tagging threatened species vs. using surrogates, stakeholder perspectives on tagging (including aboriginals), as well as use of data emanating from such studies (e.g., by fishers to facilitate exploitation). Failure to do so will have the potential to create conflict and undermine scientific, management and public confidence in the use of this powerful tool. Indeed, there are already a number of examples of where tracking studies using electronic tags have been halted based on concerns raised by researchers, authorities, or stakeholders. Here we present a candid evaluation of several factors that should be considered when determining when to tag or not to tag fish with electronic devices. It is not our objective to judge the merit of previous studies. Rather, we hope to stimulate debate and discussion regarding the use of electronic tags to study fish. Relatedly, there is a need for more research to address these questions (e.g., what level of cleanliness is needed when conducting surgeries, what type of training should be required for fish surgery) including human dimensions studies to understand perspectives of different actors including society as a whole with respect to tagging and tracking studies.

  10. CT colonography with rectal iodine tagging: Feasibility and comparison with oral tagging in a colorectal cancer screening population

    Energy Technology Data Exchange (ETDEWEB)

    Neri, Emanuele, E-mail: emanuele.neri@med.unipi.it [Diagnostic and Interventional Radiology – Department of Translational Research and New Technologies in Medicine and Surgery, University of Pisa (Italy); Mantarro, Annalisa; Faggioni, Lorenzo; Scalise, Paola; Bemi, Pietro; Pancrazi, Francesca [Diagnostic and Interventional Radiology – Department of Translational Research and New Technologies in Medicine and Surgery, University of Pisa (Italy); D’Ippolito, Giuseppe [Federal University of São Paulo – Sena Madureira 1500 – Vila Mariana, UNIFESP, São Paulo, SP (Brazil); Bartolozzi, Carlo [Diagnostic and Interventional Radiology – Department of Translational Research and New Technologies in Medicine and Surgery, University of Pisa (Italy)

    2015-09-15

    Highlights: • In the group receiving rectal tagging, mean per-polyp sensitivity, specificity were 96.1% and 95.3%; while in the group receiving oral tagging, mean per-polyp sensitivity, specificity were 89.4% and 95.8%. The difference between the two groups was not statistically significant (p = 0.549). • Rectal tagging can be an effective alternative to oral tagging. • Rectal tagging allowed greater patient acceptance and lower overall examination time. - Abstract: Purpose: To evaluate feasibility, diagnostic performance, patient acceptance, and overall examination time of CT colonography (CTC) performed through rectal administration of iodinated contrast material. Materials and methods: Six-hundred asymptomatic subjects (male:female = 270:330; mean 63 years) undergoing CTC for colorectal cancer screening on an individual basis were consecutively enrolled in the study. Out of them, 503 patients (group 1) underwent CTC with rectal tagging, of which 55 had a total of 77 colonic lesions. The remaining 97 patients (group 2) were randomly selected to receive CTC with oral tagging of which 15 had a total of 20 colonic lesions. CTC findings were compared with optical colonoscopy, and per-segment image quality was visually assessed using a semi-quantitative score (1 = poor, 2 = adequate, 3 = excellent). In 70/600 patients (11.7%), CTC was performed twice with both types of tagging over a 5-year follow-up cancer screening program. In this subgroup, patient acceptance was rated via phone interview two weeks after CTC using a semi-quantitative scale (1 = poor, 2 = fair, 3 = average, 4 = good, 5 = excellent). Results: Mean per-polyp sensitivity, specificity, positive and negative predictive values of CTC with rectal vs oral tagging were 96.1% (CI{sub 95%} 85.4 ÷ 99.3%) vs 89.4% (CI{sub 95%} 65.4 ÷ 98.1%), 95.3% (CI{sub 95%} 90.7 ÷ 97.8%) vs 95.8% (CI{sub 95%} 87.6 ÷ 98.9%), 86.0% (CI{sub 95%} 73.6 ÷ 93.3) vs 85.0% (CI{sub 95%} 61.1 ÷ 96.0%), and 98.8% (CI{sub 95

  11. Affine Contractions on the Plane

    Science.gov (United States)

    Celik, D.; Ozdemir, Y.; Ureyen, M.

    2007-01-01

    Contractions play a considerable role in the theory of fractals. However, it is not easy to find contractions which are not similitudes. In this study, it is shown by counter examples that an affine transformation of the plane carrying a given triangle onto another triangle may not be a contraction even if it contracts edges, heights or medians.…

  12. A Radio Tag for Big Whales

    Science.gov (United States)

    Watkins, William A.

    1978-01-01

    Radio tags to track wildlife have been used for years. However, such tagging of whales has been more complicated and less successful. This article explores the latest technology that is designed to give information over a long period of time. (MA)

  13. Baggage Tags for Learning Out of Doors.

    Science.gov (United States)

    Roller, Lib

    The manual provides teachers with not only educational outdoor activities, but also with activities that can be provided on an individual level. The only equipment needed for most of these activities is a bought or homemade "baggage tag". These tags are used for a variety of purposes such as plant and animal identification, nature quiz games, and…

  14. Towards EPC-compatible organic RFID tags

    NARCIS (Netherlands)

    Myny, K.; Steudel, S.; Vicca, P.; Smout, S.; Beenhakkers, M.J.; Aerle, N.A.J.M. van; Furthner, F.; Putten, B. van der; Tripathi, A.K.; Gelinck, G.H.; Genoe, J.; Dehaene, W.; Heremans, P.L.

    2011-01-01

    In this chapter, fully integrated organic RFID tags are demonstrated. These tags are inductively-coupled at a base frequency of 13.56 MHz and can be read out at distances up to 10 cm, which is the expected readout distance for proximity readers. We also demonstrate next generation transponder chips,

  15. Harnessing Collective Knowledge Inherent in Tag Clouds

    Science.gov (United States)

    Cress, U.; Held, C.

    2013-01-01

    Tagging systems represent the conceptual knowledge of a community. We experimentally tested whether people harness this collective knowledge when navigating through the Web. As a within-factor we manipulated people's prior knowledge (no knowledge vs. prior knowledge that was congruent/incongruent to the collective knowledge inherent in the tags).…

  16. Monodisperse REPO4 (RE = Yb, Gd, Y) hollow microspheres covered with nanothorns as affinity probes for selectively capturing and labeling phosphopeptides.

    Science.gov (United States)

    Cheng, Gong; Zhang, Ji-Lin; Liu, Yan-Lin; Sun, De-Hui; Ni, Jia-Zuan

    2012-02-13

    Rare-earth phosphate microspheres with unique structures were developed as affinity probes for the selective capture and tagging of phosphopeptides. Prickly REPO(4) (RE = Yb, Gd, Y) monodisperse microspheres, that have hollow structures, low densities, high specific surface areas, and large adsorptive capacities were prepared by an ion-exchange method. The elemental compositions and crystal structures of these affinity probes were confirmed by energy-dispersive spectroscopy (EDS), powder X-ray diffraction (XRD), and Fourier-transform infrared (FTIR) spectroscopy. The morphologies of these compounds were investigated using scanning electron microscopy (SEM), transmission electron microscopy (TEM), and nitrogen-adsorption isotherms. The potential ability of these microspheres for selectively capturing and labeling target biological molecules was evaluated by using protein-digestion analysis and a real sample as well as by comparison with the widely used TiO(2) affinity microspheres. These results show that these porous rare-earth phosphate microspheres are highly promising probes for the rapid purification and recognition of phosphopeptides.

  17. Affinity purification of sequence-specific DNA binding proteins.

    OpenAIRE

    1986-01-01

    We describe a method for affinity purification of sequence-specific DNA binding proteins that is fast and effective. Complementary chemically synthesized oligodeoxynucleotides that contain a recognition site for a sequence-specific DNA binding protein are annealed and ligated to give oligomers. This DNA is then covalently coupled to Sepharose CL-2B with cyanogen bromide to yield the affinity resin. A partially purified protein fraction is combined with competitor DNA and subsequently passed t...

  18. Sex identification and PIT-tagging: tools and prospects for studying intersexual differences in freshwater fishes

    DEFF Research Database (Denmark)

    Hulthén, K.; Chapman, B.B.; Nilsson, P.A.

    2014-01-01

    . The observed sex ratio of recaptured fish did not differ from the expected values of equal recapture rates between males and females. Hence, there is no observable evidence for an adverse effect of tagging close to the reproductive period and therefore this method is suitable for studying intersexual......This study evaluated a technique to allow the long-term monitoring of individual fishes of known sex in the wild using sex confirmation in close proximity to the reproductive period combined with individual tagging. Hundreds of partially migratory roach Rutilus rutilus were tagged with passive...... integrated transponders (PIT) following sex determination in spring and various performance measures were compared with fish tagged outside the reproductive period in autumn. Short-term survival was >95% for R. rutilus sexed and tagged under natural field conditions. Total length (LT) did not affect...

  19. Flavor tagging TeV jets for physics beyond the Standard Model

    Directory of Open Access Journals (Sweden)

    Sullivan Zack

    2016-01-01

    Full Text Available We present a new scheme for tagging boosted heavy flavor jets called “µx tagging.” At the LHC, the primary method to tag b-jets relies on tracking their charged constituents. However, when highly boosted, track-based b-tags lose efficiency, and the probability to mistag light jets rises dramatically. Using muons from B hadron decay and defining a particular combination “x” of angular information and boost estimation, we find fairly flat efficiencies to tag b-jets, c-jets, light-quark jets, and light-heavy jets (containing B hadrons from gluon splitting of ϵb = 14%, ϵc = 6:5%, ϵlight–light = 0:1%, and ϵlight-heavy = 0:5%, respectively. We demonstrate the usefulness of this new scheme by showing the reach for discovery of a leptophobic Z′ → bb̄ in the dijet channel.

  20. Self-organization in social tagging systems

    CERN Document Server

    Liu, Chuang; Zhang, Zi-Ke

    2011-01-01

    Individuals often imitate each other to fall into the typical group, leading to a self-organized state of typical behaviors in a community. In this paper, we model self-organization in social tagging systems and illustrate the underlying interaction and dynamics. Specifically, we introduce a model in which individuals adjust their own tagging tendency to imitate the average tagging tendency. We found that when users are of low confidence, they tend to imitate others and lead to a self-organized state with active tagging. On the other hand, when users are of high confidence and are stubborn for changes, tagging becomes inactive. We observe a phase transition at a critical level of user confidence when the system changes from one regime to the other. The distributions of post length obtained from the model are compared to real data which show good agreements.

  1. Emergent Community Structure in Social Tagging Systems

    CERN Document Server

    Cattuto, Ciro; Servedio, Vito D P; Loreto, Vittorio

    2008-01-01

    A distributed classification paradigm known as collaborative tagging has been widely adopted in new Web applications designed to manage and share online resources. Users of these applications organize resources (Web pages, digital photographs, academic papers) by associating with them freely chosen text labels, or tags. Here we leverage the social aspects of collaborative tagging and introduce a notion of resource distance based on the collective tagging activity of users. We collect data from a popular system and perform experiments showing that our definition of distance can be used to build a weighted network of resources with a detectable community structure. We show that this community structure clearly exposes the semantic relations among resources. The communities of resources that we observe are a genuinely emergent feature, resulting from the uncoordinated activity of a large number of users, and their detection paves the way for mapping emergent semantics in social tagging systems.

  2. Intrinsic-surface-tag image authentication

    Energy Technology Data Exchange (ETDEWEB)

    Palm, R.G.; DeVolpi, A.

    1991-12-01

    The objective of this work is to further the development of a unique treaty limited item (TLI) intrinsic surface tag for arms control applications. This tag`s unique feature is the ability to capture the sub-micron scale topography of the TLI surface. The surface topography is captured by plastic castings of the surface as digitally imaged by an electron microscope. Tag authentication is accomplished by comparing digital castings images obtained in two different inspections. Surface replication experiments are described, as these experiments from the basis for the authentication algorithm. Both the experiments and the authentication algorithm are analyzed using the modulation transfer function. Recommendations for future improvements in tag authentication are also suggested by the modulation transfer function analysis. 4 refs.

  3. Smart-tag Based Data Dissemination

    DEFF Research Database (Denmark)

    Bonnet, Philippe; Beaufour, Allan; Leopold, Martin

    2002-01-01

    Monitoring wide, hostile areas requires disseminating data between fixed, disconnected clusters of sensor nodes. It is not always possible to install long-range radios in order to cover the whole area. We propose to leverage the movement of mobile individuals, equipped with smart......-tags, to disseminate data across disconnected static nodes spread across a wide area. Static nodes and mobile smart-tags exchange data when they are in the vicinity of each other; smart-tags disseminate data as they move around. In this paper, we propose an algorithm for update propagation and a model for smart......-tag based data dissemination. We use simulation to study the characteristics of the model we propose. Finally, we present an implementation based on Bluetooth smart-tags....

  4. Expression and purification of recombinant proteins in Escherichia coli tagged with the metal-binding protein CusF.

    Science.gov (United States)

    Cantu-Bustos, J Enrique; Vargas-Cortez, Teresa; Morones-Ramirez, Jose Ruben; Balderas-Renteria, Isaias; Galbraith, David W; McEvoy, Megan M; Zarate, Xristo

    2016-05-01

    Production of recombinant proteins in Escherichia coli has been improved considerably through the use of fusion proteins, because they increase protein solubility and facilitate purification via affinity chromatography. In this article, we propose the use of CusF as a new fusion partner for expression and purification of recombinant proteins in E. coli. Using a cell-free protein expression system, based on the E. coli S30 extract, Green Fluorescent Protein (GFP) was expressed with a series of different N-terminal tags, immobilized on self-assembled protein microarrays, and its fluorescence quantified. GFP tagged with CusF showed the highest fluorescence intensity, and this was greater than the intensities from corresponding GFP constructs that contained MBP or GST tags. Analysis of protein production in vivo showed that CusF produces large amounts of soluble protein with low levels of inclusion bodies. Furthermore, fusion proteins can be exported to the cellular periplasm, if CusF contains the signal sequence. Taking advantage of its ability to bind copper ions, recombinant proteins can be purified with readily available IMAC resins charged with this metal ion, producing pure proteins after purification and tag removal. We therefore recommend the use of CusF as a viable alternative to MBP or GST as a fusion protein/affinity tag for the production of soluble recombinant proteins in E. coli.

  5. Design of a covert RFID tag network for target discovery and target information routing.

    Science.gov (United States)

    Pan, Qihe; Narayanan, Ram M

    2011-01-01

    Radio frequency identification (RFID) tags are small electronic devices working in the radio frequency range. They use wireless radio communications to automatically identify objects or people without the need for line-of-sight or contact, and are widely used in inventory tracking, object location, environmental monitoring. This paper presents a design of a covert RFID tag network for target discovery and target information routing. In the design, a static or very slowly moving target in the field of RFID tags transmits a distinct pseudo-noise signal, and the RFID tags in the network collect the target information and route it to the command center. A map of each RFID tag's location is saved at command center, which can determine where a RFID tag is located based on each RFID tag's ID. We propose the target information collection method with target association and clustering, and we also propose the information routing algorithm within the RFID tag network. The design and operation of the proposed algorithms are illustrated through examples. Simulation results demonstrate the effectiveness of the design.

  6. Use of the Nanofitin Alternative Scaffold as a GFP-Ready Fusion Tag.

    Directory of Open Access Journals (Sweden)

    Simon Huet

    Full Text Available With the continuous diversification of recombinant DNA technologies, the possibilities for new tailor-made protein engineering have extended on an on-going basis. Among these strategies, the use of the green fluorescent protein (GFP as a fusion domain has been widely adopted for cellular imaging and protein localization. Following the lead of the direct head-to-tail fusion of GFP, we proposed to provide additional features to recombinant proteins by genetic fusion of artificially derived binders. Thus, we reported a GFP-ready fusion tag consisting of a small and robust fusion-friendly anti-GFP Nanofitin binding domain as a proof-of-concept. While limiting steric effects on the carrier, the GFP-ready tag allows the capture of GFP or its blue (BFP, cyan (CFP and yellow (YFP alternatives. Here, we described the generation of the GFP-ready tag from the selection of a Nanofitin variant binding to the GFP and its spectral variants with a nanomolar affinity, while displaying a remarkable folding stability, as demonstrated by its full resistance upon thermal sterilization process or the full chemical synthesis of Nanofitins. To illustrate the potential of the Nanofitin-based tag as a fusion partner, we compared the expression level in Escherichia coli and activity profile of recombinant human tumor necrosis factor alpha (TNFα constructs, fused to a SUMO or GFP-ready tag. Very similar expression levels were found with the two fusion technologies. Both domains of the GFP-ready tagged TNFα were proved fully active in ELISA and interferometry binding assays, allowing the simultaneous capture by an anti-TNFα antibody and binding to the GFP, and its spectral mutants. The GFP-ready tag was also shown inert in a L929 cell based assay, demonstrating the potent TNFα mediated apoptosis induction by the GFP-ready tagged TNFα. Eventually, we proposed the GFP-ready tag as a versatile capture and labeling system in addition to expected applications of anti

  7. Theoretical proton affinity and fluoride affinity of nerve agent VX.

    Science.gov (United States)

    Bera, Narayan C; Maeda, Satoshi; Morokuma, Keiji; Viggiano, Al A

    2010-12-23

    Proton affinity and fluoride affinity of nerve agent VX at all of its possible sites were calculated at the RI-MP2/cc-pVTZ//B3LYP/6-31G* and RI-MP2/aug-cc-pVTZ//B3LYP/6-31+G* levels, respectively. The protonation leads to various unique structures, with H(+) attached to oxygen, nitrogen, and sulfur atoms; among which the nitrogen site possesses the highest proton affinity of -ΔE ∼ 251 kcal/mol, suggesting that this is likely to be the major product. In addition some H(2), CH(4) dissociation as well as destruction channels have been found, among which the CH(4) + [Et-O-P(═O)(Me)-S-(CH(2))(2)-N(+)(iPr)═CHMe] product and the destruction product forming Et-O-P(═O)(Me)-SMe + CH(2)═N(+)(iPr)(2) are only 9 kcal/mol less stable than the most stable N-protonated product. For fluoridization, the S-P destruction channel to give Et-O-P(═O)(Me)(F) + [S-(CH(2))(2)-N-(iPr)(2)](-) is energetically the most favorable, with a fluoride affinity of -ΔE ∼ 44 kcal. Various F(-) ion-molecule complexes are also found, with the one having F(-) interacting with two hydrogen atoms in different alkyl groups to be only 9 kcal/mol higher than the above destruction product. These results suggest VX behaves quite differently from surrogate systems.

  8. Engineering Protein Hydrogels Using SpyCatcher-SpyTag Chemistry.

    Science.gov (United States)

    Gao, Xiaoye; Fang, Jie; Xue, Bin; Fu, Linglan; Li, Hongbin

    2016-09-12

    Constructing hydrogels from engineered proteins has attracted significant attention within the material sciences, owing to their myriad potential applications in biomedical engineering. Developing efficient methods to cross-link tailored protein building blocks into hydrogels with desirable mechanical, physical, and functional properties is of paramount importance. By making use of the recently developed SpyCatcher-SpyTag chemistry, we successfully engineered protein hydrogels on the basis of engineered tandem modular elastomeric proteins. Our resultant protein hydrogels are soft but stable, and show excellent biocompatibility. As the first step, we tested the use of these hydrogels as a drug carrier, as well as in encapsulating human lung fibroblast cells. Our results demonstrate the robustness of the SpyCatcher-SpyTag chemistry, even when the SpyTag (or SpyCatcher) is flanked by folded globular domains. These results demonstrate that SpyCatcher-SpyTag chemistry can be used to engineer protein hydrogels from tandem modular elastomeric proteins that can find applications in tissue engineering, in fundamental mechano-biological studies, and as a controlled drug release vehicle.

  9. Using DEDICOM for completely unsupervised part-of-speech tagging.

    Energy Technology Data Exchange (ETDEWEB)

    Chew, Peter A.; Bader, Brett William; Rozovskaya, Alla (University of Illinois, Urbana, IL)

    2009-02-01

    A standard and widespread approach to part-of-speech tagging is based on Hidden Markov Models (HMMs). An alternative approach, pioneered by Schuetze (1993), induces parts of speech from scratch using singular value decomposition (SVD). We introduce DEDICOM as an alternative to SVD for part-of-speech induction. DEDICOM retains the advantages of SVD in that it is completely unsupervised: no prior knowledge is required to induce either the tagset or the associations of terms with tags. However, unlike SVD, it is also fully compatible with the HMM framework, in that it can be used to estimate emission- and transition-probability matrices which can then be used as the input for an HMM. We apply the DEDICOM method to the CONLL corpus (CONLL 2000) and compare the output of DEDICOM to the part-of-speech tags given in the corpus, and find that the correlation (almost 0.5) is quite high. Using DEDICOM, we also estimate part-of-speech ambiguity for each term, and find that these estimates correlate highly with part-of-speech ambiguity as measured in the original corpus (around 0.88). Finally, we show how the output of DEDICOM can be evaluated and compared against the more familiar output of supervised HMM-based tagging.

  10. Pop-up Archival Transmitting (PAT) fish tag data

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The cooperative tagging center (CTC) began deploying electronic tags in 2002. To date over 300 tags have been deployed. The following species have been monitored:...

  11. Production of N-acetyl-D-neuraminic acid using two sequential enzymes overexpressed as double-tagged fusion proteins

    Directory of Open Access Journals (Sweden)

    Cheng Chung-Hsien

    2009-07-01

    Full Text Available Abstract Background Two sequential enzymes in the production of sialic acids, N-acetyl-D-glucosamine 2-epimerase (GlcNAc 2-epimerase and N-acetyl-D-neuraminic acid aldolase (Neu5Ac aldolase, were overexpressed as double-tagged gene fusions. Both were tagged with glutathione S-transferase (GST at the N-terminus, but at the C-terminus, one was tagged with five contiguous aspartate residues (5D, and the other with five contiguous arginine residues (5R. Results Both fusion proteins were overexpressed in Escherichia coli and retained enzymatic activity. The fusions were designed so their surfaces were charged under enzyme reaction conditions, which allowed isolation and immobilization in a single step, through a simple capture with either an anionic or a cationic exchanger (Sepharose Q or Sepharose SP that electrostatically bound the 5D or 5R tag. The introduction of double tags only marginally altered the affinity of the enzymes for their substrates, and the double-tagged proteins were enzymatically active in both soluble and immobilized forms. Combined use of the fusion proteins led to the production of N-acetyl-D-neuraminic acid (Neu5Ac from N-acetyl-D-glucosamine (GlcNAc. Conclusion Double-tagged gene fusions were overexpressed to yield two enzymes that perform sequential steps in sialic acid synthesis. The proteins were easily immobilized via ionic tags onto ionic exchange resins and could thus be purified by direct capture from crude protein extracts. The immobilized, double-tagged proteins were effective for one-pot enzymatic production of sialic acid.

  12. Electron affinity of chlorine dioxide

    Energy Technology Data Exchange (ETDEWEB)

    Babcock, L.M.; Pentecost, T.; Koppenol, W.H. (Louisiana State Univ., Baton Rouge (USA))

    1989-12-14

    The flowing afterglow technique was used to determine the electron affinity of chlorine dioxide. A value of 2.37 {plus minus} 0.10 eV was found by bracketing between the electron affinities of HS* and SF{sub 4} as a lower limit and that of NO{sub 2} as an upper limit. This value is in excellent agreement with 2.32 eV predicted from a simple thermodynamic cycle involving the reduction potential of the ClO{sub 2}/ClO{sub 2}{sup {minus}} couple and a Gibbs hydration energy identical with that of SO{sub 2}{sup {sm bullet}{minus}}.

  13. Affine density in wavelet analysis

    CERN Document Server

    Kutyniok, Gitta

    2007-01-01

    In wavelet analysis, irregular wavelet frames have recently come to the forefront of current research due to questions concerning the robustness and stability of wavelet algorithms. A major difficulty in the study of these systems is the highly sensitive interplay between geometric properties of a sequence of time-scale indices and frame properties of the associated wavelet systems. This volume provides the first thorough and comprehensive treatment of irregular wavelet frames by introducing and employing a new notion of affine density as a highly effective tool for examining the geometry of sequences of time-scale indices. Many of the results are new and published for the first time. Topics include: qualitative and quantitative density conditions for existence of irregular wavelet frames, non-existence of irregular co-affine frames, the Nyquist phenomenon for wavelet systems, and approximation properties of irregular wavelet frames.

  14. Enhanced UHF RFID tags for drug tracing.

    Science.gov (United States)

    Catarinucci, Luca; Colella, Riccardo; De Blasi, Mario; Patrono, Luigi; Tarricone, Luciano

    2012-12-01

    Radio Frequency Identification (RFID) technology is playing a crucial role for item-level tracing systems in healthcare scenarios. The pharmaceutical supply chain is a fascinating application context, where RFID can guarantee transparency in the drug flow, supporting both suppliers and consumers against the growing counterfeiting problem. In such a context, the choice of the most adequate RFID tag, in terms of shape, frequency, size and reading range, is crucial. The potential presence of items containing materials hostile to the electromagnetic propagation exasperates the problem. In addition, the peculiarities of the different RFID-based checkpoints make even more stringent the requirements for the tag. In this work, the performance of several commercial UHF RFID tags in each step of the pharmaceutical supply chain has been evaluated, confirming the expected criticality. On such basis, a guideline for the electromagnetic design of new high-performance tags capable to overcome such criticalities has been defined. Finally, driven by such guidelines, a new enhanced tag has been designed, realized and tested. Due to patent pending issues, the antenna shape is not shown. Nevertheless, the optimal obtained results do not lose their validity. Indeed, on the one hand they demonstrate that high performance item level tracing systems can actually be implemented also in critical operating conditions. On the other hand, they encourage the tag designer to follow the identified guidelines so to realize enhanced UHF tags.

  15. Background estimation methods and exclusion limits for scalar top-quark production with top quark tagging in the all-hadronic channel at 13 TeV with the CMS detector

    Science.gov (United States)

    Wei, Hua; CMS Collaboration

    2017-01-01

    Discussed will be estimations of the Z to NuNu and QCD backgrounds with in the search for supersymmetry in all-hadronic events with missing transverse momentum using top quark tagging. This will be done for the 2016 proton-proton running period at a center-of-mass energy of 13 TeV with the CMS detector at the LHC. The exclusion limits are set on the masses of potential new particles in the context of three simplified models, T1tttt, T2tt and T5ttcc.

  16. Satellite tagging and biopsy sampling of killer whales at subantarctic Marion Island: effectiveness, immediate reactions and long-term responses.

    Directory of Open Access Journals (Sweden)

    Ryan R Reisinger

    Full Text Available Remote tissue biopsy sampling and satellite tagging are becoming widely used in large marine vertebrate studies because they allow the collection of a diverse suite of otherwise difficult-to-obtain data which are critical in understanding the ecology of these species and to their conservation and management. Researchers must carefully consider their methods not only from an animal welfare perspective, but also to ensure the scientific rigour and validity of their results. We report methods for shore-based, remote biopsy sampling and satellite tagging of killer whales Orcinus orca at Subantarctic Marion Island. The performance of these methods is critically assessed using 1 the attachment duration of low-impact minimally percutaneous satellite tags; 2 the immediate behavioural reactions of animals to biopsy sampling and satellite tagging; 3 the effect of researcher experience on biopsy sampling and satellite tagging; and 4 the mid- (1 month and long- (24 month term behavioural consequences. To study mid- and long-term behavioural changes we used multievent capture-recapture models that accommodate imperfect detection and individual heterogeneity. We made 72 biopsy sampling attempts (resulting in 32 tissue samples and 37 satellite tagging attempts (deploying 19 tags. Biopsy sampling success rates were low (43%, but tagging rates were high with improved tag designs (86%. The improved tags remained attached for 26±14 days (mean ± SD. Individuals most often showed no reaction when attempts missed (66% and a slight reaction-defined as a slight flinch, slight shake, short acceleration, or immediate dive-when hit (54%. Severe immediate reactions were never observed. Hit or miss and age-sex class were important predictors of the reaction, but the method (tag or biopsy was unimportant. Multievent trap-dependence modelling revealed considerable variation in individual sighting patterns; however, there were no significant mid- or long-term changes

  17. High affinity receptor labeling based on basic leucine zipper domain peptides conjugated with pH-sensitive fluorescent dye: Visualization of AMPA-type glutamate receptor endocytosis in living neurons.

    Science.gov (United States)

    Hayashi, Ayako; Asanuma, Daisuke; Kamiya, Mako; Urano, Yasuteru; Okabe, Shigeo

    2016-01-01

    Techniques to visualize receptor trafficking in living neurons are important, but currently available methods are limited in their labeling efficiency, specificity and reliability. Here we report a method for receptor labeling with a basic leucine zipper domain peptide (ZIP) and a binding cassette specific to ZIP. Receptors are tagged with a ZIP-binding cassette at their extracellular domain. Tagged receptors expressed in cultured cells were labeled with exogenously applied fluorescently labeled ZIP with low background and high affinity. To test if ZIP labeling is useful in monitoring endocytosis and intracellular trafficking, we next conjugated ZIP with a pH-sensitive dye RhP-M (ZIP-RhP-M). ZIP binding to its binding cassette was pH-resistant and RhP-M fluorescence dramatically increased in acidic environment. Thus AMPA-type glutamate receptors (AMPARs) labeled by ZIP-RhP-M can report receptor endocytosis and subsequent intracellular trafficking. Application of ZIP-RhP-M to cultured hippocampal neurons expressing AMPARs tagged with a ZIP-binding cassette resulted in appearance of fluorescent puncta in PSD-95-positive large spines, suggesting local endocytosis and acidification of AMPARs in individual mature spines. This spine pool of AMPARs in acidic environment was distinct from the early endosomes labeled by transferrin uptake. These results suggest that receptor labeling by ZIP-RhP-M is a useful technique for monitoring endocytosis and intracellular trafficking. This article is part of the Special Issue entitled 'Synaptopathy--from Biology to Therapy'.

  18. PATL: A RFID Tag Localization based on Phased Array Antenna

    Science.gov (United States)

    Qiu, Lanxin; Liang, Xiaoxuan; Huang, Zhangqin

    2017-01-01

    In RFID systems, how to detect the position precisely is an important and challenging research topic. In this paper, we propose a range-free 2D tag localization method based on phased array antenna, called PATL. This method takes advantage of the adjustable radiation angle of the phased array antenna to scan the surveillance region in turns. By using the statistics of the tags’ number in different antenna beam directions, a weighting algorithm is used to calculate the position of the tag. This method can be applied to real-time location of multiple targets without usage of any reference tags or additional readers. Additionally, we present an optimized weighting method based on RSSI to increase the locating accuracy. We use a Commercial Off-the-Shelf (COTS) UHF RFID reader which is integrated with a phased array antenna to evaluate our method. The experiment results from an indoor office environment demonstrate the average distance error of PATL is about 21 cm and the optimized approach achieves an accuracy of 13 cm. This novel 2D localization scheme is a simple, yet promising, solution that is especially applicable to the smart shelf visualized management in storage or retail area. PMID:28295014

  19. Inteins and affinity resin substitutes for protein purification and scale up

    Directory of Open Access Journals (Sweden)

    Wood David W

    2005-11-01

    Full Text Available Abstract The development of self-cleaving fusion-tag technology has greatly simplified the purification of recombinant proteins at laboratory scale. The self-cleaving capability of these tags has recently been combined with additional purification tags to generate novel and convenient protein purification methods at a variety of scales. In this review, we describe some of these methods, and provide a rudimentary economic analysis of hypothetical large-scale applications. This work is expected to provide a rough outline for the evaluation of these methods for large-scale bioprocessing of a variety of products.

  20. Lectin affinity chromatography of glycolipids

    Energy Technology Data Exchange (ETDEWEB)

    Torres, B.V.; Smith, D.F.

    1987-05-01

    Since glycolipids (GLs) are either insoluble or form mixed micelles in water, lectin affinity chromatography in aqueous systems has not been applied to their separation. They have overcome this problem by using tetrahydrofuran (THF) in the mobile phase during chromatography. Affinity columns prepared with the GalNAc-specific Helix pomatia agglutinin (HPA) and equilibrated in THF specifically bind the (/sup 3/H)oligosaccharide derived from Forssman GL indicating that the immobilized HPA retained its carbohydrate-binding specificity in this solvent. Intact Forssman GL was bound by the HPA-column equilibrated in THF and was specifically eluted with 0.1 mg/ml GalNAc in THF. Purification of the Forssman GL was achieved when a crude lipid extract of sheep erythrocyte membranes was applied to the HPA-column in THF. Non-specifically bound GLs were eluted from the column using a step gradient of aqueous buffer in THF, while the addition of GalNAc was required to elute the specifically bound GLs. Using this procedure the A-active GLs were purified from a crude lipid extract of type A human erythrocytes in a single chromatographic step. The use of solvents that maintain carbohydrate-binding specificity and lipid solubility will permit the application of affinity chromatography on immobilized carbohydrate-binding proteins to intact GLs.