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Sample records for single step purification

  1. Separation and purification of nanoparticles in a single step.

    Science.gov (United States)

    Hollamby, Martin J; Eastoe, Julian; Chemelli, Angela; Glatter, Otto; Rogers, Sarah; Heenan, Richard K; Grillo, Isabelle

    2010-05-18

    Reversed-micelle synthesis has been used to generate CTAB-stabilized gold (Au-NPs) and silver nanoparticles (Ag-NPs). By inducing a phase transition and subsequent separation of the background supporting microemulsion, it has been possible to extract and purify the NPs from the reaction medium. After addition of excess water, the NPs concentrate into an upper octane-rich phase, with impurities and reaction debris (in particular CTAB) partitioning into the water-rich lower phase. UV and (1)H NMR showed that 82% of the original mass of Au-NPs can be purified from the excess CTAB and other salt impurities. The concentrated and purified NPs can be dried down, by solvent removal, and then redispersed in octane. Using the complementary techniques small-angle neutron and X-ray scattering (SANS and SAXS), the structures of microemulsions both with and without nanoparticles prior to separation, and in both upper and lower phases after separation, have been elucidated. The approach has also been applied to the synthesis and recovery of silver nanoparticles, but on a larger scale. This new approach compares favorably with existing methods as it uses no additional organic solvents, has a low-energy demand, and requires no specialist surfactants. The new advance here is that by using a colloidal system to prepare and support the nanoparticles as a structured solvent, a simple soft purification method becomes accessible, which is otherwise impossible with a normal molecular solvent.

  2. Single-Step Purification of Ovalbumin from Egg White Using Aqueous Biphasic Systems.

    Science.gov (United States)

    Pereira, Matheus M; Cruz, Rafaela A P; Almeida, Mafalda R; Lima, Álvaro S; Coutinho, João A P; Freire, Mara G

    2016-06-01

    The ability of aqueous biphasic systems (ABS) composed of polyethylene glycols of different molecular weights (PEG 400, 600 and 1000) and buffered aqueous solutions of potassium citrate/citric acid (pH = 5.0 - 8.0) to selectively extract ovalbumin from egg white was here investigated. Phase diagrams, tie-lines and tie-line lengths were determined at 25ºC and the partitioning of ovalbumin in these systems was then evaluated. Aiming at optimizing the selective extraction of ovalbumin in the studied ABS, factors such as pH, PEG molecular weight and amount of the phase-forming components were initially investigated with pure commercial ovalbumin. In almost all ABS, it was observed a preferential partitioning of ovalbumin to the polymer-rich phase, with extraction efficiencies higher than 90%. The best ABS were then applied in the purification of ovalbumin from the real egg white matrix. In order to ascertain on the ovalbumin purity and yield, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and size exclusion high performance liquid chromatography (SE-HPLC) analyses were conducted, confirming that the isolation/purification of ovalbumin from egg white was completely achieved in a single-step with a recovery yield of 65%. The results obtained show that polymer-salt-based ABS allow the selective extraction of ovalbumin from egg white with a simpler approach and better performance than previously reported. Finally, it is shown that ovalbumin can be completely recovered from the PEG-rich phase by an induced precipitation using an inexpensive and sustainable separation platform which can be easily applied on an industrial scale.

  3. 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.

  4. Engineering Escherichia coli for Soluble Expression and Single Step Purification of Active Human Lysozyme

    Science.gov (United States)

    Lamppa, John W.; Tanyos, Sam A.; Griswold, Karl E.

    2012-01-01

    Genetically engineered variants of human lysozyme represent promising leads in the battle against drug-resistant bacterial pathogens, but early stage development and testing of novel lysozyme variants is constrained by the lack of a robust, scalable and facile expression system. While wild type human lysozyme is reportedly produced at 50 – 80 kg per hectare of land in recombinant rice, this plant-based system is not readily scaled down to bench top production, and it is therefore not suitable for development and characterization of novel lysozyme variants. Here, we describe a novel and efficient expression system capable of producing folded, soluble and functional human lysozyme in E. coli cells. To achieve this goal, we simultaneously co-express multiple protein folding chaperones as well as harness the lysozyme inhibitory protein, Ivy. Our strategy exploits E. coli’s ease of culture, short doubling time, and facile genetics to yield upwards of 30 mg/L of soluble lysozyme in a bioreactor system, a 3000-fold improvement over prior efforts in E. coli. Additionally, molecular interactions between lysozyme and a his-tagged Ivy allows for one-step purification by IMAC chromatography, yielding as much as 21 mg/L of purified enzyme. We anticipate that our expression and purification platform will facilitate further development of engineered lysozymes having utility in disease treatment and other practical applications. PMID:23220215

  5. Single-Step Purification and Characterization of A Recombinant Serine Proteinase Inhibitor from Transgenic Plants.

    Science.gov (United States)

    Jha, Shweta; Agarwal, Saurabh; Sanyal, Indraneel; Amla, D V

    2016-05-01

    Expression of recombinant therapeutic proteins in transgenic plants has a tremendous impact on safe and economical production of biomolecules for biopharmaceutical industry. The major limitation in their production is downstream processing of recombinant protein to obtain higher yield and purity of the final product. In this study, a simple and rapid process has been developed for purification of therapeutic recombinant α1-proteinase inhibitor (rα1-PI) from transgenic tomato plants, which is an abundant serine protease inhibitor in human serum and chiefly inhibits the activity of neutrophil elastase in lungs. We have expressed rα1-PI with modified synthetic gene in transgenic tomato plants at a very high level (≃3.2 % of total soluble protein). The heterologous protein was extracted with (NH4)2SO4 precipitation, followed by chromatographic separation on different matrices. However, only immunoaffinity chromatography resulted into homogenous preparation of rα1-PI with 54 % recovery. The plant-purified rα1-PI showed molecular mass and structural conformation comparable to native serum α1-PI, as shown by mass spectrometry and optical spectroscopy. The results of elastase inhibition assay revealed biological activity of the purified rα1-PI protein. This work demonstrates a simple and efficient one-step purification of rα1-PI from transgenic plants, which is an essential prerequisite for further therapeutic development.

  6. Engineering Escherichia coli for soluble expression and single step purification of active human lysozyme.

    Science.gov (United States)

    Lamppa, John W; Tanyos, Sam A; Griswold, Karl E

    2013-03-10

    Genetically engineered variants of human lysozyme represent promising leads in the battle against drug-resistant bacterial pathogens, but early stage development and testing of novel lysozyme variants is constrained by the lack of a robust, scalable and facile expression system. While wild type human lysozyme is reportedly produced at 50–80 kg per hectare of land in recombinant rice, this plant-based system is not readily scaled down to bench top production, and it is therefore not suitable for development and characterization of novel lysozyme variants. Here, we describe a novel and efficient expression system capable of producing folded, soluble and functional human lysozyme in Escherichia coli cells. To achieve this goal, we simultaneously co-express multiple protein folding chaperones as well as harness the lysozyme inhibitory protein, Ivy. Our strategy exploits E. coli's ease of culture, short doubling time, and facile genetics to yield upwards of 30 mg/l of soluble lysozyme in a bioreactor system, a 3000-fold improvement over prior efforts in E. coli. Additionally, molecular interactions between lysozyme and a his-tagged Ivy allows for one-step purification by IMAC, yielding as much as 21 mg/l of purified enzyme. We anticipate that our expression and purification platform will facilitate further development of engineered lysozymes having utility in disease treatment and other practical applications. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. A novel multimodal chromatography based single step purification process for efficient manufacturing of an E. coli based biotherapeutic protein product.

    Science.gov (United States)

    Bhambure, Rahul; Gupta, Darpan; Rathore, Anurag S

    2013-11-01

    Methionine oxidized, reduced and fMet forms of a native recombinant protein product are often the critical product variants which are associated with proteins expressed as bacterial inclusion bodies in E. coli. Such product variants differ from native protein in their structural and functional aspects, and may lead to loss of biological activity and immunogenic response in patients. This investigation focuses on evaluation of multimodal chromatography for selective removal of these product variants using recombinant human granulocyte colony stimulating factor (GCSF) as the model protein. Unique selectivity in separation of closely related product variants was obtained using combined pH and salt based elution gradients in hydrophobic charge induction chromatography. Simultaneous removal of process related impurities was also achieved in flow-through leading to single step purification process for the GCSF. Results indicate that the product recovery of up to 90.0% can be obtained with purity levels of greater than 99.0%. Binding the target protein at pHpurification step. Copyright © 2013 Elsevier B.V. All rights reserved.

  8. Using guanidine-hydrochloride for fast and efficient protein digestion and single-step affinity-purification mass spectrometry

    DEFF Research Database (Denmark)

    Poulsen, Jon Wriedt; Madsen, Christian Toft; Young, Clifford

    2013-01-01

    be optimally completed within 30 min with endoprotease Lys-C. No chemical artifacts were introduced when samples were incubated in Gnd-HCl at 95 °C, making Gnd-HCl an appropriate digestion buffer for shotgun proteomics. Current methodologies for investigating protein-protein interactions (PPIs) often require...... several preparation steps, which prolongs any parallel operation and high-throughput interaction analysis. Gnd-HCl allow the efficient elution and subsequent fast digestion of PPIs to provide a convenient high-throughput methodology for affinity-purification mass spectrometry (AP-MS) experiments...

  9. Single-step affinity purification of recombinant proteins using a self-excising module from Neisseria meningitidis FrpC

    Czech Academy of Sciences Publication Activity Database

    Sadílková, Lenka; Osička, Radim; Šulc, Miroslav; Linhartová, Irena; Novák, Petr; Šebo, Peter

    2008-01-01

    Roč. 17, č. 10 (2008), s. 1834-1843 ISSN 0961-8368 R&D Projects: GA AV ČR KAN200520702; GA ČR GA310/06/0720 Institutional research plan: CEZ:AV0Z50200510 Keywords : asp-pro bond * frpc * purification Subject RIV: EE - Microbiology, Virology Impact factor: 3.115, year: 2008

  10. Fibrous polymer grafted magnetic chitosan beads with strong poly(cation-exchange) groups for single step purification of lysozyme.

    Science.gov (United States)

    Bayramoglu, Gulay; Tekinay, Turgay; Ozalp, V Cengiz; Arica, M Yakup

    2015-05-15

    Lysozyme is an important polypetide used in medical and food applications. We report a novel magnetic strong cation exchange beads for efficient purification of lysozyme from chicken egg white. Magnetic chitosan (MCHT) beads were synthesized via phase inversion method, and then grafted with poly(glycidyl methacrylate) (p(GMA)) via the surface-initiated atom transfer radical polymerization (SI-ATRP). Epoxy groups of the grafted polymer, were modified into strong cation-exchange groups (i.e., sulfonate groups) in the presence of sodium sulfite. The MCTH and MCTH-g-p(GMA)-SO3H beads were characterized by ATR-FTIR, SEM, and VSM. The sulphonate groups content of the modified MCTH-g-p(GMA)-4 beads was found to be 0.53mmolg(-1) of beads by the potentiometric titration method. The MCTH-g-p(GMA)-SO3H beads were first used as an ion-exchange support for adsorption of lysozyme from aqueous solution. The influence of different experimental parameters such as pH, contact time, and temperature on the adsorption process was evaluated. The maximum adsorption capacity was found to be 208.7mgg(-1) beads. Adsorption of lysozyme on the MCTH-g-p(GMA)-SO3H beads fitted to Langmuir isotherm model and followed the pseudo second-order kinetic. More than 93% of the adsorbed lysozyme was desorbed using Na2CO3 solution (pH 11.0). The purity of the lysozyme was checked by HPLC and SDS gel electrophoresis. In addition, the MCTH-g-p(GMA)-SO3H beads prepared in this work showed promising potential for separation of various anionic molecules. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Modulation of protein A binding allows single-step purification of mouse bispecific antibodies that retain FcRn binding

    Science.gov (United States)

    Armstrong, Anthony A.; Pardinas, Jose R.; Zheng, Songmao; Brosnan, Kerry; Emmell, Eva; Luo, Jeffrey; Chiu, Mark L.

    2017-01-01

    ABSTRACT The increased number of bispecific antibodies (BsAb) under therapeutic development has resulted in a need for mouse surrogate BsAbs. Here, we describe a one-step method for generating highly pure mouse BsAbs suitable for in vitro and in vivo studies. We identify two mutations in the mouse IgG2a and IgG2b Fc region: one that eliminates protein A binding and one that enhances protein A binding by 8-fold. We show that BsAbs harboring these mutations can be purified from the residual parental monoclonal antibodies in one step using protein A affinity chromatography. The structural basis for the effects of these mutations was analyzed by X-ray crystallography. While the mutation that disrupted protein A binding also inhibited FcRn interaction, a bispecific mutant in which one subunit retained the ability to bind protein A could still interact with FcRn. Pharmacokinetic analysis of the serum half-lives of the mutants showed that the mutant BsAb had a serum half-life comparable to a wild-type Ab. The results describe a rapid method for generating panels of mouse BsAbs that could be used in mouse studies. PMID:28898162

  12. Rapid one-step purification of single-cells encapsulated in alginate microcapsules from oil to aqueous phase using a hydrophobic filter paper: implications for single-cell experiments.

    Science.gov (United States)

    Lee, Do-Hyun; Jang, Miran; Park, Je-Kyun

    2014-10-01

    By virtue of the biocompatibility and physical properties of hydrogel, picoliter-sized hydrogel microcapsules have been considered to be a biometric signature containing several features similar to that of encapsulated single cells, including phenotype, viability, and intracellular content. To maximize the experimental potential of encapsulating cells in hydrogel microcapsules, a method that enables efficient hydrogel microcapsule purification from oil is necessary. Current methods based on centrifugation for the conventional stepwise rinsing of oil, are slow and laborious and decrease the monodispersity and yield of the recovered hydrogel microcapsules. To remedy these shortcomings we have developed a simple one-step method to purify alginate microcapsules, containing a single live cell, from oil to aqueous phase. This method employs oil impregnation using a commercially available hydrophobic filter paper without multistep centrifugal purification and complicated microchannel networks. The oil-suspended alginate microcapsules encapsulating single cells from mammalian cancer cell lines (MCF-7, HepG2, and U937) and microorganisms (Chlorella vulgaris) were successfully exchanged to cell culture media by quick (~10 min) depletion of the surrounding oil phase without coalescence of neighboring microcapsules. Cell proliferation and high integrity of the microcapsules were also demonstrated by long-term incubation of microcapsules containing a single live cell. We expect that this method for the simple and rapid purification of encapsulated single-cell microcapsules will attain widespread adoption, assisting cell biologists and clinicians in the development of single-cell experiments. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. High yield expression and single-step purification of Toxoplasma gondii SAG1, GRA1, and GRA7 antigens in Escherichia coli

    DEFF Research Database (Denmark)

    Hiszczynska-Sawicka, E.; Brillowska-Dabrowska, A.; Dabrowski, Slawomir

    2003-01-01

    This report describes a simple, highly efficient and reproducible method for obtaining large quantities of highly pure recombinant Toxoplasma gondii antigens, which can be used for diagnostic application. The obtained T gondii SAG1, GRA1, and GRA7 antigens (as fusion proteins), expressed...... in Escherichia coli, contained polyhistidine tags at the N- and C-ends that allowed single-step isolation by metal-affinity chromatography on Ni2+-IDA-Sepharose columns. The immunoreactivity of the recombinant antigens was tested in an enzyme-linked immunosorbent assay (ELISA) format for potential application...

  14. Single step purification and characterization of a thermostable and calcium independent α-amylase from Bacillus amyloliquifaciens TSWK1-1 isolated from Tulsi Shyam hot spring reservoir, Gujarat (India).

    Science.gov (United States)

    Kikani, B A; Singh, S P

    2011-05-01

    A thermophilic bacteria, identified and designated as Bacillus amyloliquifaciens TSWK1-1 (16S rRNA gene sequence, GenBank: GQ121033), was isolated from a hot water reservoir located at Tulsi Shyam, Gujarat, India. The optimum temperature and pH for amylase production were 50 °C and 7.0, respectively. The crude enzyme was partially purified by ammonium sulphate fractionation followed by dialysis. However, single step purification was achieved on Phenyl Sepharose 6FF affinity column with 45.71% yield, 8000 U/mg specific activity and 13.33 fold purification. The molecular weight of the purified α-amylase was 43 kD. The optimal temperature and pH for amylase activity were 70 °C and 7.0, respectively; however, the purified amylase was stable at broad temperature and pH range. The purified amylase did not require Ca(++) and K(+); however, it was moderately affected by Mg(++) and Cu(++) and significantly inhibited by Na(+) and Fe(++). The amylase was highly thermostable and remained active for 24h at 60 °C, for 12h at 70 °C and up to 3h even at 90 °C. Other unique features of the enzyme were calcium independent nature and resistance against chemical denaturation by Urea and Guanidine-HCl. The data on the enzymatic stability at different levels of purity would add significantly to the knowledge of amylases. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. Two-Step Vapor/Liquid/Solid Purification

    Science.gov (United States)

    Holland, L. R.

    1986-01-01

    Vertical distillation system combines in single operation advantages of multiple zone refining with those of distillation. Developed specifically to load Bridgman-Stockbarger (vertical-solidification) growth ampoules with ultrapure tellurium and cadmium, system, with suitable modifications, serves as material refiner. In first phase of purification process, ampoule heated to drive off absorbed volatiles. Second phase, evaporator heated to drive off volatiles in charge. Third phase, slowly descending heater causes distillation from evaporator to growing crystal in ampoule.

  16. Influence of the drying method in chitosans purification step

    International Nuclear Information System (INIS)

    Fonseca, Ana C.M.; Batista, Jorge G.S.; Bettega, Antonio; Lima, Nelson B. de

    2015-01-01

    Currently, the study of extracellular biopolymers properties has received prominence for being easy extraction and purification. Chitosan has been an attractive proposition for applications in various fields such as engineering, biotechnology, medicine and pharmacology. For such applications, it is necessary purification of chitosan to obtain a product more concentrated and free of undesirable impurities. However, at this stage of the process of obtaining the biopolymer may occur morphological and physicochemical changes. This study evaluated the influence of the drying process after purification of a commercial chitosan sample and the importance of this step and its cost/benefit in applications requiring a high degree of purity. The method of drying influenced in the organoleptic properties and in the main characteristics of material. Analysis of the crystal structure by X-ray diffraction showed that the degree of crystallinity, X (%), in the purified chitosan samples was lower when compared with the unpurified sample. The degree of acetylation, DA (%), was analyzed by spectroscopy infrared with no significant changes on the three drying methods assessed, unlike the viscosimetric molecular weight, M v , determined by capillary viscometry. (author)

  17. Enhancing recombinant interleukin-6 production yield by fermentation optimization, two-step denaturing, and one-step purification.

    Science.gov (United States)

    Ahmed, Nadeem; Abbas, Rabbia; Khan, Mohsin Ahmad; Bashir, Hamid; Tahir, Saad; Zafar, Ahmad Usman

    2017-08-22

    Interleukin-6 a pleiotropic cytokine involved in a wide range of biological activities. So the large-scale production of biologically active recombinant human interleukin-6 is important for its structural and functional studies. Here, we report an optimized method for shake flask fermentation and a simplified high-yield purification procedure for the recombinant interleukin-6. This high-yield expression method not only involves the optimization of the fermentation condition but also the single step purification method as well as a two-step denaturing and one-step refolding process. This approach replaces the more conventional procedure of protein solubilization and refolding. Through applying these strategies, the final cell density and overall product yield of the recombinant human interleukin-6 were obtained as 20.4 g as cell biomass and 150 mg as purified active protein from the I-L of the culture. The purified protein was characterized by HPLC and SDS-PAGE. The results of the current work demonstrate that the described method may be used to develop the process for industrial-scale production of the biologically active recombinant interleukin-6 protein. © 2017 International Union of Biochemistry and Molecular Biology, Inc.

  18. One-step purification of E. coli elongation factor Tu

    DEFF Research Database (Denmark)

    Knudsen, Charlotte Rohde; Clark, Brian F. C.; Degn, B

    1993-01-01

    The tuf A gene, encoding the E. coli elongation factor Tu, was cloned in the pGEX gene fusion system. Upon expression EF-Tu is fused to glutathione-S-transferase serving as a purification handle with affinity for glutathione immobilised on agarose. This allows purification of EF-Tu in a one...

  19. Simultaneous purification of DNA and RNA from microbiota in a single colonic mucosal biopsy.

    Science.gov (United States)

    Moen, Aina E F; Tannæs, Tone M; Vatn, Simen; Ricanek, Petr; Vatn, Morten Harald; Jahnsen, Jørgen

    2016-06-28

    Nucleic acid purification methods are of importance when performing microbiota studies and especially when analysing the intestinal microbiota as we here find a wide range of different microbes. Various considerations must be taken to lyse the microbial cell wall of each microbe. In the present article, we compare several tissue lysis steps and commercial purification kits, to achieve a joint RNA and DNA purification protocol for the purpose of investigating the microbiota and the microbiota-host interactions in a single colonic mucosal tissue sample. A further optimised tissue homogenisation and lysis protocol comprising mechanical bead beating, lysis buffer replacement and enzymatic treatment, in combination with the AllPrep DNA/RNA Mini Kit (Qiagen, Hilden, Germany) resulted in efficient and simultaneous purification of microbial and human RNA and DNA from a single mucosal colonic tissue sample. The present work provides a unique possibility to study RNA and DNA from the same mucosal biopsy sample, making a direct comparison between metabolically active microbes and total microbial DNA. The protocol also offers an opportunity to investigate other members of a microbiota such as viruses, fungi and micro-eukaryotes, and moreover the possibility to extract data on microbiota and host interactions from one single mucosal biopsy.

  20. Single-step purification and characterization of an extreme halophilic, ethanol tolerant and acidophilic xylanase from Aureobasidium pullulans NRRL Y-2311-1 with application potential in the food industry.

    Science.gov (United States)

    Yegin, Sirma

    2017-04-15

    An extracellular xylanase from Aureobasidium pullulans NRRL Y-2311-1 produced on wheat bran was purified by a single-step chromatographic procedure. The enzyme had a molecular weight of 21.6kDa. The optimum pH and temperature for xylanase activity were 4.0 and 30-50°C, respectively. The enzyme was stable in the pH range of 3.0-8.0. The inactivation energy of the enzyme was calculated as 218kJmol -1 . The xylanase was ethanol tolerant and kept complete activity in the presence of 10% ethanol. Likewise, it retained almost complete activity at a concentration range of 0-20% NaCl. In general, the enzyme was resistant to several metal ions and reagents. Mg 2+ , Zn 2+ , Cu 2+ , K 1+ , EDTA and β-mercaptoethanol resulted in enhanced xylanase activity. The K m and V max values on beechwood xylan were determined to be 19.43mgml -1 and 848.4Uml -1 , respectively. The enzyme exhibits excellent characteristics and could, therefore, be a promising candidate for application in food and bio-industries. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. TCA precipitation and ethanol/HCl single-step purification evaluation: One-dimensional gel electrophoresis, bradford assays, spectrofluorometry and Raman spectroscopy data on HSA, Rnase, lysozyme - Mascots and Skyline data

    Directory of Open Access Journals (Sweden)

    Balkis Eddhif

    2018-04-01

    Full Text Available The data presented here are related to the research paper entitled “Study of a Novel Agent for TCA Precipitated Proteins Washing - Comprehensive Insights into the Role of Ethanol/HCl on Molten Globule State by Multi-Spectroscopic Analyses” (Eddhif et al., submitted for publication [1]. The suitability of ethanol/HCl for the washing of TCA-precipitated proteins was first investigated on standard solution of HSA, cellulase, ribonuclease and lysozyme. Recoveries were assessed by one-dimensional gel electrophoresis, Bradford assays and UPLC-HRMS. The mechanistic that triggers protein conformational changes at each purification stage was then investigated by Raman spectroscopy and spectrofluorometry. Finally, the efficiency of the method was evaluated on three different complex samples (mouse liver, river biofilm, loamy soil surface. Proteins profiling was assessed by gel electrophoresis and by UPLC-HRMS.

  2. Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

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    Brenton James D

    2004-01-01

    Full Text Available Abstract Background Expression microarrays have evolved into a powerful tool with great potential for clinical application and therefore reliability of data is essential. RNA amplification is used when the amount of starting material is scarce, as is frequently the case with clinical samples. Purification steps are critical in RNA amplification and labelling protocols, and there is a lack of sufficient data to validate and optimise the process. Results Here the purification steps involved in the protocol for indirect labelling of amplified RNA are evaluated and the experimentally determined best method for each step with respect to yield, purity, size distribution of the transcripts, and dye coupling is used to generate targets tested in replicate hybridisations. DNase treatment of diluted total RNA samples followed by phenol extraction is the optimal way to remove genomic DNA contamination. Purification of double-stranded cDNA is best achieved by phenol extraction followed by isopropanol precipitation at room temperature. Extraction with guanidinium-phenol and Lithium Chloride precipitation are the optimal methods for purification of amplified RNA and labelled aRNA respectively. Conclusion This protocol provides targets that generate highly reproducible microarray data with good representation of transcripts across the size spectrum and a coefficient of repeatability significantly better than that reported previously.

  3. Effects of Two Purification Pretreatments on Electroless Copper Coating over Single-Walled Carbon Nanotubes

    Directory of Open Access Journals (Sweden)

    Zhong Zheng

    2014-01-01

    Full Text Available To achieve the reinforcement of copper matrix composite by single-walled carbon nanotubes, a three-step-refluxing purification of carbon nanotubes sample with HNO3-NaOH-HCl was proposed and demonstrated. A previously reported purification process using an electromagnetic stirring with H2O2/HCl mixture was also repeated. Then, the purified carbon nanotubes were coated with copper by the same electroless plating process. At the end, the effects of the method on carbon nanotubes themselves and on copper coating were determined by transmission electron microscope spectroscopy, scanning electron microscope spectroscopy, X-ray diffractometry, thermogravimetric analysis, Fourier transformed infrared spectroscopy, and energy dispersive spectrometry. It was clearly confirmed that both of the two processes could remove most of iron catalyst particles and carbonaceous impurities without significant damage to carbon nanotubes. The thermal stability of the sample purified by H2O2/HCl treatment was slightly higher than that purified by HNO3-NaOH-HCl treatment. Nevertheless, the purification by HNO3-NaOH-HCl treatment was more effective for carboxyl functionalization on nanotubes than that by H2O2/HCl treatment. The Cu-coating on carbon nanotubes purified by both purification processes was complete, homogenous, and continuous. However, the Cu-coating on carbon nanotubes purified by H2O2/HCl was oxidized more seriously than those on carbon nanotubes purified by HNO3-NaOH-HCl treatment.

  4. One-step chromatographic purification of Helicobacter pylori neutrophil-activating protein expressed in Bacillus subtilis.

    Directory of Open Access Journals (Sweden)

    Kuo-Shun Shih

    Full Text Available Helicobacter pylori neutrophil-activating protein (HP-NAP, a major virulence factor of Helicobacter pylori (H. pylori, is capable of activating human neutrophils to produce reactive oxygen species (ROS and secrete inammatory mediators. HP-NAP is a vaccine candidate, a possible drug target, and a potential in vitro diagnostic marker for H. pylori infection. HP-NAP has also been shown to be a novel therapeutic agent for the treatment of allergic asthma and bladder cancer. Hence, an efficient way to obtain pure HP-NAP needs to be developed. In this study, one-step anion-exchange chromatography in negative mode was applied to purify the recombinant HP-NAP expressed in Bacillus subtilis (B. subtilis. This purification technique was based on the binding of host cell proteins and/or impurities other than HP-NAP to DEAE Sephadex resins. At pH 8.0, almost no other proteins except HP-NAP passed through the DEAE Sephadex column. More than 60% of the total HP-NAP with purity higher than 91% was recovered in the flow-through fraction from this single-step DEAE Sephadex chromatography. The purified recombinant HP-NAP was further demonstrated to be a multimeric protein with a secondary structure of α-helix and capable of activating human neutrophils to stimulate ROS production. Thus, this one-step negative chromatography using DEAE Sephadex resin can efficiently yield functional HP-NAP from B. subtilis in its native form with high purity. HP-NAP purified by this method could be further utilized for the development of new drugs, vaccines, and diagnostics for H. pylori infection.

  5. One-Step Chromatographic Purification of Helicobacter pylori Neutrophil-Activating Protein Expressed in Bacillus subtilis

    Science.gov (United States)

    Shih, Kuo-Shun; Lin, Chih-Chang; Hung, Hsiao-Fang; Yang, Yu-Chi; Wang, Chung-An; Jeng, Kee-Ching; Fu, Hua-Wen

    2013-01-01

    Helicobacter pylori neutrophil-activating protein (HP-NAP), a major virulence factor of Helicobacter pylori (H. pylori), is capable of activating human neutrophils to produce reactive oxygen species (ROS) and secrete inammatory mediators. HP-NAP is a vaccine candidate, a possible drug target, and a potential in vitro diagnostic marker for H. pylori infection. HP-NAP has also been shown to be a novel therapeutic agent for the treatment of allergic asthma and bladder cancer. Hence, an efficient way to obtain pure HP-NAP needs to be developed. In this study, one-step anion-exchange chromatography in negative mode was applied to purify the recombinant HP-NAP expressed in Bacillus subtilis (B. subtilis). This purification technique was based on the binding of host cell proteins and/or impurities other than HP-NAP to DEAE Sephadex resins. At pH 8.0, almost no other proteins except HP-NAP passed through the DEAE Sephadex column. More than 60% of the total HP-NAP with purity higher than 91% was recovered in the flow-through fraction from this single-step DEAE Sephadex chromatography. The purified recombinant HP-NAP was further demonstrated to be a multimeric protein with a secondary structure of α-helix and capable of activating human neutrophils to stimulate ROS production. Thus, this one-step negative chromatography using DEAE Sephadex resin can efficiently yield functional HP-NAP from B. subtilis in its native form with high purity. HP-NAP purified by this method could be further utilized for the development of new drugs, vaccines, and diagnostics for H. pylori infection. PMID:23577158

  6. Purification of Single-Wall carbon nanotubes by heat treatment and supercritical extraction

    Directory of Open Access Journals (Sweden)

    Mariana Bertoncini

    2011-09-01

    Full Text Available Arc discharge is the most practical method for the synthesis of single wall carbon nanotubes (SWCNT. However, the production of SWCNT by this technique has low selectivity and yield, requiring further purification steps. This work is a study of purification of SWCNT by heat treatment in an inert atmosphere followed by supercritical fluid extraction. The raw arc discharge material was first heat-treated at 1250 °C under argon. The nanotubes were further submitted to an extraction process using supercritical CO2 as solvent. A surfactant (tributylphosphate, TBP and a chelating agent (hexafluoroacetylacetone, HFA were used together to eliminate metallic impurities from the remaining arc discharge catalysts. Analysis of Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP-AES showed an efficient removal of iron and cobalt (>80%. The purified nanotubes were further analyzed by TGA and Raman spectroscopy.

  7. Development of an affinity cryogel for one step purification of lysozyme from chicken egg white.

    Science.gov (United States)

    Mól, Paula Chequer Gouveia; Veríssimo, Lizzy Ayra Alcântara; Eller, Monique Renon; Minim, Valéria Paula Rodrigues; Minim, Luis Antonio

    2017-02-15

    In this study, a supermacroporous polyacrylamide cryogel was produced by cryo-polymerization and activated with Tris(hydroxymethyl)aminomethane (Tris-cryogel) to be applied as an affinity ligand for a one step purification of lysozyme (LYZ), directly from chicken egg white (EW). The Tris-cryogel presented interconnected pores with size varying in the range of 20-80μm and swelling capacity of 19.6±0.9g/g. The axial dispersion of the Tris-cryogel was analyzed at different flow velocities and mobile phase viscosities. It was verified that higher viscosity resulted in a higher degree of dispersion, causing the HETP values to increase from 0.04cm to 0.8cm. Adsorption isotherms were measured at 15°C and 35°C at pH 7.5. A Langmuir model was fitted to the equilibrium data, with a maximum adsorptive capacity of 285mg/g at 15°C and 363mg/g at 35°C. Thermodynamic analysis based on the Van't Hoff relationship showed that the process was spontaneous and enthalpically driven. Lysozyme was purified directly from egg white in a one step purification process at different pH values (7.5, 8.5 and 9.5). Independent of the pH, the specificity of Tris-cryogel for lysozyme adsorption was confirmed. At pH 7.5, yield and purification fold were higher (30% and 45). In addition, the effect of the dilution rate on egg white and flow velocity were also analyzed and it was shown that flow velocity did not affected purification and column efficiency, and that diluting the egg white increased yield to 70% with a purification fold of 23. Results show Tris-cryogel is a promising matrix for use in high throughput purification of lysozyme from egg white. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Column chromatography as a useful step in purification of diatom pigments.

    Science.gov (United States)

    Tokarek, Wiktor; Listwan, Stanisław; Pagacz, Joanna; Leśniak, Piotr; Latowski, Dariusz

    2016-01-01

    Fucoxanthin, diadinoxanthin and diatoxanthin are carotenoids found in brown algae and most other heterokonts. These pigments are involved in photosynthetic and photoprotective reactions, and they have many potential health benefits. They can be extracted from diatom Phaeodactylum tricornutum by sonication, extraction with chloroform : methanol and preparative thin layer chromatography. We assessed the utility of an additional column chromatography step in purification of these pigments. This novel addition to the isolation protocol increased the purity of fucoxanthin and allowed for concentration of diadinoxanthin and diatoxanthin before HPLC separation. The enhanced protocol is useful for obtaining high purity pigments for biochemical studies.

  9. Nucleotide-mimetic synthetic ligands for DNA-recognizing enzymes One-step purification of Pfu DNA polymerase.

    Science.gov (United States)

    Melissis, S; Labrou, N E; Clonis, Y D

    2006-07-28

    The commercial availability of DNA polymerases has revolutionized molecular biotechnology and certain sectors of the bio-industry. Therefore, the development of affinity adsorbents for purification of DNA polymerases is of academic interest and practical importance. In the present study we describe the design, synthesis and evaluation of a combinatorial library of novel affinity ligands for the purification of DNA polymerases (Pols). Pyrococcus furiosus DNA polymerase (Pfu Pol) was employed as a proof-of-principle example. Affinity ligand design was based on mimicking the natural interactions between deoxynucleoside-triphosphates (dNTPs) and the B-motif, a conserved structural moiety found in Pol-I and Pol-II family of enzymes. Solid-phase 'structure-guided' combinatorial chemistry was used to construct a library of 26 variants of the B-motif-binding 'lead' ligand X-Trz-Y (X is a purine derivative and Y is an aliphatic/aromatic sulphonate or phosphonate derivative) using 1,3,5-triazine (Trz) as the scaffold for assembly. The 'lead' ligand showed complementarity against a Lys and a Tyr residue of the polymerase B-motif. The ligand library was screened for its ability to bind and purify Pfu Pol from Escherichia coli extract. One immobilized ligand (oABSAd), bearing 9-aminoethyladenine (AEAd) and sulfanilic acid (oABS) linked on the triazine scaffold, displayed the highest purifying ability and binding capacity (0,55 mg Pfu Pol/g wet gel). Adsorption equilibrium studies with this affinity ligand and Pfu Pol determined a dissociation constant (K(D)) of 83 nM for the respective complex. The oABSAd affinity adsorbent was exploited in the development of a facile Pfu Pol purification protocol, affording homogeneous enzyme (>99% purity) in a single chromatography step. Quality control tests showed that Pfu Pol purified on the B-motif-complementing ligand is free of nucleic acids and contaminating nuclease activities, therefore, suitable for experimental use.

  10. Process simulation of single-step dimethyl ether production via biomass gasification.

    Science.gov (United States)

    Ju, Fudong; Chen, Hanping; Ding, Xuejun; Yang, Haiping; Wang, Xianhua; Zhang, Shihong; Dai, Zhenghua

    2009-01-01

    In this study, we simulated the single-step process of dimethyl ether (DME) synthesis via biomass gasification using ASPEN Plus. The whole process comprised four parts: gasification, water gas shift reaction, gas purification, and single-step DME synthesis. We analyzed the influence of the oxygen/biomass and steam/biomass ratios on biomass gasification and synthesis performance. The syngas H(2)/CO ratio after water gas shift process was modulated to 1, and the syngas was then purified to remove H(2)S and CO(2), using the Rectisol process. Syngas still contained trace amounts of H(2)S and about 3% CO(2) after purification, which satisfied the synthesis demands. However, the high level of cold energy consumption was a problem during the purification process. The DME yield in this study was 0.37, assuming that the DME selectivity was 0.91 and that CO was totally converted. We performed environmental and economic analyses, and propose the development of a poly-generation process based on economic considerations.

  11. Direct current injection and thermocapillary flow for purification of aligned arrays of single-walled carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Xu; Islam, Ahmad E.; Seabron, Eric; Dunham, Simon N.; Du, Frank; Lin, Jonathan; Wilson, William L.; Rogers, John A., E-mail: jrogers@illinois.edu [Department of Materials Science and Engineering, Frederick Seitz Materials Research Laboratory, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801 (United States); Wahab, Muhammad A.; Alam, Muhammad A. [School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Li, Yuhang [Institute of Solid Mechanics, Beihang University, Beijing 100191 (China); Tomic, Bojan [Department of Electrical Engineering, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801 (United States); Huang, Jiyuan [Department of Physics, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801 (United States); Burns, Branden [Department of Physics, Purdue University, West Lafayette, Indiana 47907 (United States); Song, Jizhou [Department of Engineering Mechanics and Soft Matter Research Center, Zhejiang University, Hangzhou 310027 (China); Huang, Yonggang [Department of Civil and Environmental Engineering, Department of Mechanical Engineering, Center for Engineering and Health, and Skin Disease Research Center, Northwestern University, Evanston, Illinois 60208 (United States)

    2015-04-07

    Aligned arrays of semiconducting single-walled carbon nanotubes (s-SWNTs) represent ideal configurations for use of this class of material in high performance electronics. Development of means for removing the metallic SWNTs (m-SWNTs) in as-grown arrays represents an essential challenge. Here, we introduce a simple scheme that achieves this type of purification using direct, selective current injection through interdigitated electrodes into the m-SWNTs, to allow their complete removal using processes of thermocapillarity and dry etching. Experiments and numerical simulations establish the fundamental aspects that lead to selectivity in this process, thereby setting design rules for optimization. Single-step purification of arrays that include thousands of SWNTs demonstrates the effectiveness and simplicity of the procedures. The result is a practical route to large-area aligned arrays of purely s-SWNTs with low-cost experimental setups.

  12. Direct current injection and thermocapillary flow for purification of aligned arrays of single-walled carbon nanotubes

    International Nuclear Information System (INIS)

    Xie, Xu; Islam, Ahmad E.; Seabron, Eric; Dunham, Simon N.; Du, Frank; Lin, Jonathan; Wilson, William L.; Rogers, John A.; Wahab, Muhammad A.; Alam, Muhammad A.; Li, Yuhang; Tomic, Bojan; Huang, Jiyuan; Burns, Branden; Song, Jizhou; Huang, Yonggang

    2015-01-01

    Aligned arrays of semiconducting single-walled carbon nanotubes (s-SWNTs) represent ideal configurations for use of this class of material in high performance electronics. Development of means for removing the metallic SWNTs (m-SWNTs) in as-grown arrays represents an essential challenge. Here, we introduce a simple scheme that achieves this type of purification using direct, selective current injection through interdigitated electrodes into the m-SWNTs, to allow their complete removal using processes of thermocapillarity and dry etching. Experiments and numerical simulations establish the fundamental aspects that lead to selectivity in this process, thereby setting design rules for optimization. Single-step purification of arrays that include thousands of SWNTs demonstrates the effectiveness and simplicity of the procedures. The result is a practical route to large-area aligned arrays of purely s-SWNTs with low-cost experimental setups

  13. Two-step purification of scutellarin from Erigeron breviscapus (vant.) Hand. Mazz. by high-speed counter-current chromatography.

    Science.gov (United States)

    Gao, Min; Gu, Ming; Liu, Chun-Zhao

    2006-07-11

    Scutellarin, a flavone glycoside, popularly applied for the treatment of cardiopathy, has been purified in two-step purification by high-speed counter-current chromatography (HSCCC) from Erigeron breviscapus (vant.) Hand. Mazz. (Deng-zhan-hua in Chinese), a well-known traditional Chinese medicinal plant for heart disease. Two solvent systems, n-hexane-ethyl acetate-methanol-acetic acid-water (1:6:1.5:1:4, v/v/v/v/v) and ethyl acetate-n-butanol-acetonitrile-0.1% HCl (5:2:5:10, v/v/v/v) were used for the two-step purification. The purity of the collected fraction of scutellarin was 95.6%. This study supplies a new alternative method for purification of scutellarin.

  14. Single-step digital backpropagation for nonlinearity mitigation

    DEFF Research Database (Denmark)

    Secondini, Marco; Rommel, Simon; Meloni, Gianluca

    2015-01-01

    Nonlinearity mitigation based on the enhanced split-step Fourier method (ESSFM) for the implementation of low-complexity digital backpropagation (DBP) is investigated and experimentally demonstrated. After reviewing the main computational aspects of DBP and of the conventional split-step Fourier...... is experimentally demonstrated by using a single-step DBP based on the ESSFM. The proposed DBP implementation requires only a single step of the ESSFM algorithm to achieve a transmission distance of 3200 km over a dispersion-unmanaged link. In comparison, a conventional DBP implementation requires 20 steps...

  15. Semi-automated hydrophobic interaction chromatography column scouting used in the two-step purification of recombinant green fluorescent protein.

    Science.gov (United States)

    Stone, Orrin J; Biette, Kelly M; Murphy, Patrick J M

    2014-01-01

    Hydrophobic interaction chromatography (HIC) most commonly requires experimental determination (i.e., scouting) in order to select an optimal chromatographic medium for purifying a given target protein. Neither a two-step purification of untagged green fluorescent protein (GFP) from crude bacterial lysate using sequential HIC and size exclusion chromatography (SEC), nor HIC column scouting elution profiles of GFP, have been previously reported. Bacterial lysate expressing recombinant GFP was sequentially adsorbed to commercially available HIC columns containing butyl, octyl, and phenyl-based HIC ligands coupled to matrices of varying bead size. The lysate was fractionated using a linear ammonium phosphate salt gradient at constant pH. Collected HIC eluate fractions containing retained GFP were then pooled and further purified using high-resolution preparative SEC. Significant differences in presumptive GFP elution profiles were observed using in-line absorption spectrophotometry (A395) and post-run fluorimetry. SDS-PAGE and western blot demonstrated that fluorometric detection was the more accurate indicator of GFP elution in both HIC and SEC purification steps. Comparison of composite HIC column scouting data indicated that a phenyl ligand coupled to a 34 µm matrix produced the highest degree of target protein capture and separation. Conducting two-step protein purification using the preferred HIC medium followed by SEC resulted in a final, concentrated product with >98% protein purity. In-line absorbance spectrophotometry was not as precise of an indicator of GFP elution as post-run fluorimetry. These findings demonstrate the importance of utilizing a combination of detection methods when evaluating purification strategies. GFP is a well-characterized model protein, used heavily in educational settings and by researchers with limited protein purification experience, and the data and strategies presented here may aid in development other of HIC-compatible protein

  16. Semi-Automated Hydrophobic Interaction Chromatography Column Scouting Used in the Two-Step Purification of Recombinant Green Fluorescent Protein

    Science.gov (United States)

    Murphy, Patrick J. M.

    2014-01-01

    Background Hydrophobic interaction chromatography (HIC) most commonly requires experimental determination (i.e., scouting) in order to select an optimal chromatographic medium for purifying a given target protein. Neither a two-step purification of untagged green fluorescent protein (GFP) from crude bacterial lysate using sequential HIC and size exclusion chromatography (SEC), nor HIC column scouting elution profiles of GFP, have been previously reported. Methods and Results Bacterial lysate expressing recombinant GFP was sequentially adsorbed to commercially available HIC columns containing butyl, octyl, and phenyl-based HIC ligands coupled to matrices of varying bead size. The lysate was fractionated using a linear ammonium phosphate salt gradient at constant pH. Collected HIC eluate fractions containing retained GFP were then pooled and further purified using high-resolution preparative SEC. Significant differences in presumptive GFP elution profiles were observed using in-line absorption spectrophotometry (A395) and post-run fluorimetry. SDS-PAGE and western blot demonstrated that fluorometric detection was the more accurate indicator of GFP elution in both HIC and SEC purification steps. Comparison of composite HIC column scouting data indicated that a phenyl ligand coupled to a 34 µm matrix produced the highest degree of target protein capture and separation. Conclusions Conducting two-step protein purification using the preferred HIC medium followed by SEC resulted in a final, concentrated product with >98% protein purity. In-line absorbance spectrophotometry was not as precise of an indicator of GFP elution as post-run fluorimetry. These findings demonstrate the importance of utilizing a combination of detection methods when evaluating purification strategies. GFP is a well-characterized model protein, used heavily in educational settings and by researchers with limited protein purification experience, and the data and strategies presented here may aid in

  17. Development of adsorptive hybrid filters to enable two-step purification of biologics

    Science.gov (United States)

    Peck, Michael; Voloshin, Alexei M.; Moreno, Angela M.; Tan, Zhijun; Hester, Jonathan; Borys, Michael C.; Li, Zheng Jian

    2017-01-01

    ABSTRACT Recent progress in mammalian cell culture process has resulted in significantly increased product titers, but also a substantial increase in process- and product-related impurities. Due to the diverse physicochemical properties of these impurities, there is constant need for new technologies that offer higher productivity and improved economics without sacrificing the process robustness required to meet final drug substance specifications. Here, we examined the use of new synthetic adsorptive hybrid filters (AHF) modified with the high binding capacity of quaternary amine (Emphaze™ AEX) and salt-tolerant biomimetic (Emphaze™ ST-AEX) ligands for clearance of process-related impurities like host cell protein (HCP), residual DNA, and virus. The potential to remove soluble aggregates was also examined. Our aim was to develop a mechanistic understanding of the interactions governing adsorptive removal of impurities during filtration by evaluating the effect of various filter types, feed streams, and process conditions on impurity removal. The ionic capacity of these filters was measured and correlated with their ability to remove impurities for multiple molecules. The ionic capacity of AHF significantly exceeded that of traditional adsorptive depth filters (ADF) by 40% for the Emphaze™ AEX and by 700% for the Emphaze™ ST-AEX, providing substantially higher reduction of soluble anionic impurities, including DNA, HCPs and model virus. Nevertheless, we determined that ADF with filter aid provided additional hydrophobic functionality that resulted in removal of higher molecular weight species than AHF. Implementing AHF demonstrated improved process-related impurity removal and viral clearance after Protein A chromatography and enabled a two-step purification process. The consequences of enhanced process performance are far reaching because it allows the downstream polishing train to be restructured and simplified, and chromatographic purity standards to be

  18. Accuracy of Single-Step versus 2-Step Double-Mix Impression Technique

    DEFF Research Database (Denmark)

    Franco, Eduardo Batista; da Cunha, Leonardo Fernandes; Herrera, Francyle Simões

    2011-01-01

    Objective. To investigate the accuracy of dies obtained from single-step and 2-step double-mix impressions. Material and Methods. Impressions (n = 10) of a stainless steel die simulating a complete crown preparation were performed using a polyether (Impregum Soft Heavy and Light body) and a vinyl...

  19. One-step fabrication of multifunctional composite polyurethane spider-web-like nanofibrous membrane for water purification

    Energy Technology Data Exchange (ETDEWEB)

    Pant, Hem Raj, E-mail: hempant@jbnu.ac.kr [Department of Bio-nano System Engineering, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Department of Engineering Science and Humanities, Institute of Engineering, Pulchowk Campus, Tribhuvan University, Kathmandu (Nepal); Kim, Han Joo [Division of Mechanical Design Engineering, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Joshi, Mahesh Kumar; Pant, Bishweshwar; Park, Chan Hee; Kim, Jeong In [Department of Bio-nano System Engineering, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Hui, K.S., E-mail: kshui@hanyang.ac.kr [Department of Mechanical Engineering, Hanyang University, 17 Haengdang-dong, Seongdong-gu, Seoul 133-791 (Korea, Republic of); Kim, Cheol Sang, E-mail: chskim@jbnu.ac.kr [Department of Bio-nano System Engineering, Chonbuk National University, Jeonju 561-756 (Korea, Republic of); Division of Mechanical Design Engineering, Chonbuk National University, Jeonju 561-756 (Korea, Republic of)

    2014-01-15

    Highlights: • A single mat having varieties of performance for water treatment is simply introduced. • Cost effective Ag-doped fly ash/PU nanofibers are fabricated in one-step. • Solvent reduction of AgNO{sub 3} could produce Ag-loaded spider-web nets. • Size of Ag NPs on fiber surface can be controlled by controlling stirring time. • Fabrication of nanocomposite using pollutant material to control other pollutents. -- Abstract: A stable silver-doped fly ash/polyurathene (Ag-FA/PU) nanocomposite multifunctional membrane is prepared by a facile one-step electrospinning process using fly ash particles (FAPs). Colloidal solution of PU with FAPs and Ag metal precursor was subjected to fabricate nanocomposite spider-web-like membrane using electrospinning process. Presence of N,N-dimethylformamide (solvent of PU) led to reduce silver nitrate into Ag NPs. Incorporation of Ag NPs and FAPs through electrospun PU fibers is proven through electron microscopy and spectroscopic techniques. Presence of these NPs on PU nanofibers introduces several potential physicochemical properties such as spider-web-like nano-neeting for NPs separation, enhanced absorption capacity to remove carcinogenic arsenic (As) and toxic organic dyes, and antibacterial properties with reduce bio-fouling for membrane filter application. Preliminary observations used for above-mentioned applications for water treatment showed that it will be an economically and environmentally friendly nonwoven matrix for water purification. This simple approach highlights new avenues about the utilization of one pollutant material to control other pollutants in scalable and inexpensive ways.

  20. One-step fabrication of multifunctional composite polyurethane spider-web-like nanofibrous membrane for water purification

    International Nuclear Information System (INIS)

    Pant, Hem Raj; Kim, Han Joo; Joshi, Mahesh Kumar; Pant, Bishweshwar; Park, Chan Hee; Kim, Jeong In; Hui, K.S.; Kim, Cheol Sang

    2014-01-01

    Highlights: • A single mat having varieties of performance for water treatment is simply introduced. • Cost effective Ag-doped fly ash/PU nanofibers are fabricated in one-step. • Solvent reduction of AgNO 3 could produce Ag-loaded spider-web nets. • Size of Ag NPs on fiber surface can be controlled by controlling stirring time. • Fabrication of nanocomposite using pollutant material to control other pollutents. -- Abstract: A stable silver-doped fly ash/polyurathene (Ag-FA/PU) nanocomposite multifunctional membrane is prepared by a facile one-step electrospinning process using fly ash particles (FAPs). Colloidal solution of PU with FAPs and Ag metal precursor was subjected to fabricate nanocomposite spider-web-like membrane using electrospinning process. Presence of N,N-dimethylformamide (solvent of PU) led to reduce silver nitrate into Ag NPs. Incorporation of Ag NPs and FAPs through electrospun PU fibers is proven through electron microscopy and spectroscopic techniques. Presence of these NPs on PU nanofibers introduces several potential physicochemical properties such as spider-web-like nano-neeting for NPs separation, enhanced absorption capacity to remove carcinogenic arsenic (As) and toxic organic dyes, and antibacterial properties with reduce bio-fouling for membrane filter application. Preliminary observations used for above-mentioned applications for water treatment showed that it will be an economically and environmentally friendly nonwoven matrix for water purification. This simple approach highlights new avenues about the utilization of one pollutant material to control other pollutants in scalable and inexpensive ways

  1. Rational design of a host cell protein heat precipitation step simplifies the subsequent purification of recombinant proteins from tobacco

    OpenAIRE

    Buyel, J.F.; Gruchow, H.M.; Boes, A.; Fischer, R.

    2014-01-01

    The benefits of plants for the production of biopharmaceutical proteins include the ease of scale-up and the low pathogen burden, but the purification of target proteins is often affected by low expression levels and the abundance of host cell proteins in leaf extracts. These factors can significantly diminish the competitiveness of plant-based expression systems. We therefore compared three different heat precipitation steps that remove most host cell proteins while leaving a heat-stable can...

  2. Pelvifemoral kinematics while ascending single steps of different heights.

    Science.gov (United States)

    Bohannon, Richard W; Smutnick, Jason

    2010-08-01

    Motion of the femur and pelvis during hip flexion has been examined previously, but principally in the sagittal plane and during nonfunctional activities. In this study we examined femoral elevation in the sagittal plane and pelvic rotation in the sagittal and frontal planes while subjects flexed their hips to ascend single steps. Fourteen subjects ascended single steps of 4 different heights leading with each lower limb. Motion of the lead femur and pelvis during the flexion phase of step ascent was tracked using an infrared motion capture system. Depending on step height and lead limb, step ascent involved elevation of the femur (mean 47.2 degrees to 89.6 degrees) and rotation of the pelvis in both the sagittal plane (tilting: mean 2.6 degrees to 9.7 degrees) and frontal plane (listing: mean 4.2 degrees to 11.9 degrees). Along with maximum femoral elevation, maximum pelvic rotation increased significantly (pelevation and pelvic rotation during the flexion phase of step ascent were synergistic (r=.852-.999). Practitioners should consider pelvic rotation in addition to femoral motion when observing individuals' ascent of steps.

  3. High quality potassium phosphate production through step-by-step glycerol purification: a strategy to economize biodiesel production.

    Science.gov (United States)

    Javani, Azita; Hasheminejad, Meisam; Tahvildari, Kambiz; Tabatabaei, Meisam

    2012-01-01

    The cost of biodiesel production can be reduced by a number of strategies such as utilization of waste cooking oils and non-edible plant oils as well as implementation of improved separation technologies. In addition, processes dealing with the glycerol by-product can have economic benefits. In the present study, acidification of crude glycerol with phosphoric acid to pH 9.67 followed by acidification to 4.67 was implemented to produce high quality potassium phosphate during glycerol purification. KH(2)PO(4), K(2)HPO(4), glycerol and free fatty acids (FFAs) with a purity of 98%, 98.05%, 96.08% and 99.58% were obtained, respectively. Copyright © 2011 Elsevier Ltd. All rights reserved.

  4. Influence of the drying method in chitosans purification step; Influencia do metodo de secagem na etapa de purificacao de quitosanas

    Energy Technology Data Exchange (ETDEWEB)

    Fonseca, Ana C.M.; Batista, Jorge G.S.; Bettega, Antonio; Lima, Nelson B. de, E-mail: acmfonseca.acf@gmail.com [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)

    2015-07-01

    Currently, the study of extracellular biopolymers properties has received prominence for being easy extraction and purification. Chitosan has been an attractive proposition for applications in various fields such as engineering, biotechnology, medicine and pharmacology. For such applications, it is necessary purification of chitosan to obtain a product more concentrated and free of undesirable impurities. However, at this stage of the process of obtaining the biopolymer may occur morphological and physicochemical changes. This study evaluated the influence of the drying process after purification of a commercial chitosan sample and the importance of this step and its cost/benefit in applications requiring a high degree of purity. The method of drying influenced in the organoleptic properties and in the main characteristics of material. Analysis of the crystal structure by X-ray diffraction showed that the degree of crystallinity, X (%), in the purified chitosan samples was lower when compared with the unpurified sample. The degree of acetylation, DA (%), was analyzed by spectroscopy infrared with no significant changes on the three drying methods assessed, unlike the viscosimetric molecular weight, M{sub v}, determined by capillary viscometry. (author)

  5. A rapid and efficient two-step gel electrophoresis method for the purification of major rye grass pollen allergens.

    Science.gov (United States)

    Levy, D; Davies, J; O'Hehir, R; Suphioglu, C

    2001-06-01

    Purified proteins are mandatory for molecular, immunological and cellular studies. However, purification of proteins from complex mixtures requires specialised chromatography methods (i.e., gel filtration, ion exchange, etc.) using fast protein liquid chromatography (FPLC) or high-performance liquid chromatography (HPLC) systems. Such systems are expensive and certain proteins require two or more different steps for sufficient purity and generally result in low recovery. The aim of this study was to develop a rapid, inexpensive and efficient gel-electrophoresis-based protein purification method using basic and readily available laboratory equipment. We have used crude rye grass pollen extract to purify the major allergens Lol p 1 and Lol p 5 as the model protein candidates. Total proteins were resolved on large primary gel and Coomassie Brilliant Blue (CBB)-stained Lol p 1/5 allergens were excised and purified on a secondary "mini"-gel. Purified proteins were extracted from unstained separating gels and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses. Silver-stained SDS-PAGE gels resolved pure proteins (i.e., 875 microg of Lol p 1 recovered from a 8 mg crude starting material) while immunoblot analysis confirmed immunological reactivity of the purified proteins. Such a purification method is rapid, inexpensive, and efficient in generating proteins of sufficient purity for use in monoclonal antibody (mAb) production, protein sequencing and general molecular, immunological, and cellular studies.

  6. Two-step Purification Method for Aging Pigments A2E and Iso-A2E Using Medium Pressure Liquid Chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Park, Sang Il; Park, Sang Cheol [Kyunghee Univ., Seoul (Korea, Republic of); Kim, So Ra; Jang, Young Pyo [Seoul National Univ., Seoul (Korea, Republic of)

    2016-09-15

    A newly modified method for the efficient purification of A2E and iso-A2E using reverse phase silica gel resin with medium pressure liquid chromatography (MPLC) was suggested. MPLC is one of the various preparative column chromatography techniques and separation under pressure renders the use of smaller particle size resins possible and increases the diversity of available stationary phases. A simplified two-step purification method was developed for the purification of aging pigments in eye, A2E and iso-A2E. With simple two-step elution of mobile phase in reverse phase MPLC, A2E and iso-A2E were successfully purified with great efficiency compare with previous column chromatography and HPLC method. This method provides more simple, convenient, cost-effective, and less time-consuming procedure for mass purification of aging pigments A2E and iso-A2E.

  7. Single-centre comparison of a novel single-step balloon inflation ...

    African Journals Online (AJOL)

    Subject. Single-centre comparison of a novel single-step balloon inflation device and Amplatz sheath dilatation during percutaneous nephrolithotomy – a pilot study. Outcome measures. Single procedure success rates, retreatment rates, hospital stay, haemoglobin concentration, calculi volume, calculi configuration, patient ...

  8. One-step affinity purification of the yeast ribosome and its associated proteins and mRNAs.

    Science.gov (United States)

    Inada, Toshifumi; Winstall, Eric; Tarun, Salvador Z; Yates, John R; Schieltz, Dave; Sachs, Alan B

    2002-07-01

    We describe a one-step affinity method for purifying ribosomes from the budding yeast Saccharomyces cerevisiae. Extracts from yeast strains expressing only C-terminally tagged Rpl25 protein or overexpressing this protein in the presence of endogenous Rpl25p were used as the starling materials. The purification was specific for tagged 60S subunits, and resulted in the copurification of 80S subunits and polysomes, as well as ribosome-associated proteins and mRNAs. Two of these associated proteins, Mpt4p and Asc1p, were nearly stoichiometrically bound to the ribosome. In addition, the degree of mRNA association with the purified ribosomes was found to reflect the mRNA's translational status within the cell. The one-step purification of ribosome and its associated components from a crude extract should provide an important tool for future structural and biochemical studies of the ribosome, as well as for expression profiling of translated mRNAs.

  9. Two-step purification of a highly thermostable alkaline protease from salt-tolerant alkaliphilic Streptomyces clavuligerus strain Mit-1.

    Science.gov (United States)

    Thumar, Jignasha; Singh, S P

    2007-07-01

    An alkaline protease from a salt-tolerant alkaliphilic Streptomyces clavuligerus was purified to homogeneity by 141-fold with a yield of 12% using two-step method of salt precipitation and ion exchange chromatography on DEAE cellulose. The apparent molecular mass was 49+/-2 kDa and the enzyme appeared as monomer based on SDS and Native-PAGE. The temperature optimum was 70 degrees C with significant stability at 60-80 degrees C for more than 60 min. The enzyme was active over the pH range of 8.5-11, with an optimum at 10-11. The serine nature of the protease was confirmed by PMSF inhibition. The enzyme was highly resistant against chemical denaturation and displayed varied effects towards metal ions. The results are significant as extremozymes are difficult to purify and therefore, a two-step purification of alkaline protease from relatively less explored group of actinomycetes is quite appealing.

  10. Flavourzyme, an Enzyme Preparation with Industrial Relevance: Automated Nine-Step Purification and Partial Characterization of Eight Enzymes.

    Science.gov (United States)

    Merz, Michael; Eisele, Thomas; Berends, Pieter; Appel, Daniel; Rabe, Swen; Blank, Imre; Stressler, Timo; Fischer, Lutz

    2015-06-17

    Flavourzyme is sold as a peptidase preparation from Aspergillus oryzae. The enzyme preparation is widely and diversely used for protein hydrolysis in industrial and research applications. However, detailed information about the composition of this mixture is still missing due to the complexity. The present study identified eight key enzymes by mass spectrometry and partially by activity staining on native polyacrylamide gels or gel zymography. The eight enzymes identified were two aminopeptidases, two dipeptidyl peptidases, three endopeptidases, and one α-amylase from the A. oryzae strain ATCC 42149/RIB 40 (yellow koji mold). Various specific marker substrates for these Flavourzyme enzymes were ascertained. An automated, time-saving nine-step protocol for the purification of all eight enzymes within 7 h was designed. Finally, the purified Flavourzyme enzymes were biochemically characterized with regard to pH and temperature profiles and molecular sizes.

  11. Purification and alignment of arc-synthesis single-walled carbon nanotube bundles

    Science.gov (United States)

    Huang, Houjin; Kajiura, Hisashi; Yamada, Atsuo; Ata, Masafumi

    2002-04-01

    We report here a scalable method for purification and alignment of single-walled carbon nanotubes (SWNT) in an aqueous solution. Arc-synthesis soot containing SWNTs is first treated with a concentrated nitric acid. After removal of most of the impurities and water, macroscopic and well-aligned SWNT bundles up to several centimeters long are formed in a rotary evaporator. Alignment of the SWNT bundles is ascribed to the liquid flow induced by rotary evaporation and van der Waals interactions among the bundles. The aligned SWNT bundles are further purified by ultrasonic Soxhlet extraction and annealing.

  12. Purification and Characterization of Lipase from Aspergillus flavus ...

    African Journals Online (AJOL)

    USER

    Abstract. Lipase from Aspergillus flavus was purified in a single step purification using MnFeO4 magnetic nano particles to achieve a 20.53- fold purification with specific activity of. 11.29 U/mg and a 59% recovery yield. SDS-PAGE of lipase showed a single pure band with corresponding molecular weight of 35 kDa.

  13. Single-step colony assay for screening antibody libraries.

    Science.gov (United States)

    Kato, Mieko; Hanyu, Yoshiro

    2017-08-10

    We describe a method, single-step colony assay, for simple and rapid screening of single-chain Fv fragment (scFv) libraries. Colonies of Escherichia coli expressing the scFv library are formed on a hydrophilic filter that is positioned in contact with a membrane coated with an antigen. scFv expression is triggered upon treatment of colonies with an induction reagent, following which scFvs are secreted from the cells and diffused to the antigen-coated membrane. scFvs that exhibit binding affinity for the antigen are captured by the membrane-immobilized antigen. Lastly, detection of scFv binding of the antigen on the membrane allows identification of the clones on the filter that express antigen-specific scFvs. We tested this methodology by using an anti-rabbit IgG scFv, scFv(A10B), and a rat immune scFv library. Experiments conducted using scFv(A10B) revealed that this method improves scFv expression during the colony assay. By using our method to screen an immune library of 3×10 3 scFv clones, we established several clones exhibiting affinity for the antigen. Moreover, we tested 7 other antigens, including peptides, and successfully identified positive clones. We believe that this simple procedure and controlled scFv expression of the single-step colony assay could make the antibody screening both rapid and reliable and lead to successful isolation of positive clones from antibody libraries. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Efficient and versatile one-step affinity purification of in vivo biotinylated proteins: Expression, characterization and structure analysis of recombinant human glutamate carboxypeptidase II

    Energy Technology Data Exchange (ETDEWEB)

    Tykvart, J.; Sacha, P.; Barinka, C.; Knedlik, T.; Starkova, J.; Lubkowski, J.; Konvalinka, J. (Gilead); (NCI); (Czech Academy)

    2012-02-07

    Affinity purification is a useful approach for purification of recombinant proteins. Eukaryotic expression systems have become more frequently used at the expense of prokaryotic systems since they afford recombinant eukaryotic proteins with post-translational modifications similar or identical to the native ones. Here, we present a one-step affinity purification set-up suitable for the purification of secreted proteins. The set-up is based on the interaction between biotin and mutated streptavidin. Drosophila Schneider 2 cells are chosen as the expression host, and a biotin acceptor peptide is used as an affinity tag. This tag is biotinylated by Escherichia coli biotin-protein ligase in vivo. We determined that localization of the ligase within the ER led to the most effective in vivo biotinylation of the secreted proteins. We optimized a protocol for large-scale expression and purification of AviTEV-tagged recombinant human glutamate carboxypeptidase II (Avi-GCPII) with milligram yields per liter of culture. We also determined the 3D structure of Avi-GCPII by X-ray crystallography and compared the enzymatic characteristics of the protein to those of its non-tagged variant. These experiments confirmed that AviTEV tag does not affect the biophysical properties of its fused partner. Purification approach, developed here, provides not only a sufficient amount of highly homogenous protein but also specifically and effectively biotinylates a target protein and thus enables its subsequent visualization or immobilization.

  15. Algal-based, single-step treatment of urban wastewaters.

    Science.gov (United States)

    Henkanatte-Gedera, S M; Selvaratnam, T; Caskan, N; Nirmalakhandan, N; Van Voorhies, W; Lammers, Peter J

    2015-08-01

    Currently, urban wastewaters (UWW) laden with organic carbon (BOD) and nutrients (ammoniacal nitrogen, N, and phosphates, P) are treated in multi-stage, energy-intensive process trains to meet the mandated discharge standards. This study presents a single-step process based on mixotrophic metabolism for simultaneous removal of carbon and nutrients from UWWs. The proposed system is designed specifically for hot, arid environments utilizing an acidophilic, thermotolerant algal species, Galdieria sulphuraria, and an enclosed photobioreactor to limit evaporation. Removal rates of BOD, N, and P recorded in this study (14.93, 7.23, and 1.38 mg L(-1) d(-1), respectively) are comparable to literature reports. These results confirm that the mixotrophic system can reduce the energy costs associated with oxygen supply in current UWW treatment systems, and has the potential to generate more energy-rich biomass for net energy extraction from UWW. Copyright © 2015 Elsevier Ltd. All rights reserved.

  16. Simplified method to obtain enhanced expression of tau protein from E. coli and one-step purification by direct boiling.

    Science.gov (United States)

    KrishnaKumar, V Guru; Gupta, Sharad

    2017-05-28

    Tau is an intrinsically disordered protein responsible for maintaining the structure and stability of axonal microtubules. However, in certain disease conditions including Alzheimer's disease, tau protein may undergo biochemical and structural changes to form intracellular aggregates. Since tau is a proline- and arginine-rich eukaryotic protein, heterologous expression in Escherichia coli often results in poor yield and has been a major technical challenge. In the current work, we have improved the expressed yield of tau by overcoming codon bias problem and established a simplified protocol for efficient extraction. The reported method has two distinct features: (i) enhanced tau expression (upto eightfold) by supplementing deficient tRNAs that aid in rapid translation and (ii) direct boiling of expressed E. coli cells to extract tau with no separate cell lysis step. We further demonstrate that tau extracted by the direct boiling method is similar to tau purified by size-exclusion chromatography exhibiting similar structural and biophysical characteristics including aggregation propensity. Since morphologies and in vitro toxicity of fibrillar tau aggregates were also similar, tau extracted by the one-step direct boiling method can be used for tau aggregation assays without any additional purification.

  17. Virtual substitution scan via single-step free energy perturbation.

    Science.gov (United States)

    Chiang, Ying-Chih; Wang, Yi

    2016-02-05

    With the rapid expansion of our computing power, molecular dynamics (MD) simulations ranging from hundreds of nanoseconds to microseconds or even milliseconds have become increasingly common. The majority of these long trajectories are obtained from plain (vanilla) MD simulations, where no enhanced sampling or free energy calculation method is employed. To promote the 'recycling' of these trajectories, we developed the Virtual Substitution Scan (VSS) toolkit as a plugin of the open-source visualization and analysis software VMD. Based on the single-step free energy perturbation (sFEP) method, VSS enables the user to post-process a vanilla MD trajectory for a fast free energy scan of substituting aryl hydrogens by small functional groups. Dihedrals of the functional groups are sampled explicitly in VSS, which improves the performance of the calculation and is found particularly important for certain groups. As a proof-of-concept demonstration, we employ VSS to compute the solvation free energy change upon substituting the hydrogen of a benzene molecule by 12 small functional groups frequently considered in lead optimization. Additionally, VSS is used to compute the relative binding free energy of four selected ligands of the T4 lysozyme. Overall, the computational cost of VSS is only a fraction of the corresponding multi-step FEP (mFEP) calculation, while its results agree reasonably well with those of mFEP, indicating that VSS offers a promising tool for rapid free energy scan of small functional group substitutions. This article is protected by copyright. All rights reserved. © 2016 Wiley Periodicals, Inc.

  18. Two step procedure for purification of enzymatically active prostate-specific antigen from seminal plasma.

    Science.gov (United States)

    Bindukumar, B; Kawinski, Elzbieta; Cherrin, Craig; Gambino, Leah M; Nair, Madhavan P N; Schwartz, Stanley A; Chadha, Kailash C

    2004-12-25

    The role of prostate-specific antigen (PSA) during the onset of prostate cancer and subsequent tumor growth and metastasis is not well understood. We have developed a simple two step procedure, based on principles of hydrophobic charge-induction chromatography and molecular size chromatography to provide pure free-PSA (f-PSA) preparation that is free from all other known PSA complexes as well as human kallikrein 2 (hK2). The overall recovery of f-PSA is 72%. The isolated f-PSA consists of three known isoforms that corresponds to pI of 6.2, 6.4 and 7.2. f-PSA is enzymatically active and its enzymatic activity can be effectively neutralized by a serine protease inhibitor.

  19. A new application of aptamer: One-step purification and immobilization of enzyme from cell lysates for biocatalysis.

    Science.gov (United States)

    Qiao, Lifeng; Lv, Bo; Feng, Xudong; Li, Chun

    2015-06-10

    Aptamers are nucleic acid-based high affinity ligands that are able to capture their corresponding target through molecular recognition. In this study, several DNA aptamers with high affinity and specificity for β-glucuronidases (PGUS-E) were obtained by our modified SELEX method. Among them, Apt5 and Apt9 were selected as representatives and covalently linked to magnetic beads, respectively. The aptamer-modified magnetic beads were characterized and successfully applied to one-step purification and immobilization of PGUS-E from the complex cell lysates. By conveniently adjusting the pH and ion strength, the PGUS-E purities reached 84% for Apt5-modified beads and 88% for Apt9-modified beads. Moreover, the maximum PGUS-E capturing capacity of the Apt5 and Apt9 modified magnetic beads were found to be 31.75μg/mg and 32.95μg/mg, respectively. The immobilized PGUS-E on aptamer-based magnetic beads showed good reusability, and the conversion of glycyrrhizin still remained more than 70% after 7 cycles. In addition, the aptamer-modified beads support can be easily regenerated, and the conversion rate of glycyrrhizin (GL) was still 62% after the 7th cycle of regeneration. This investigation can be easily extended to other enzyme systems and may help open a generic route to develop a novel enzyme immobilization technology for biocatalysis based on aptamer. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. EXPERIMENTAL DATA, THERMODYNAMIC MODELING AND SENSITIVITY ANALYSES FOR THE PURIFICATION STEPS OF ETHYL BIODIESEL FROM FODDER RADISH OIL PRODUCTION

    Directory of Open Access Journals (Sweden)

    R. C. Basso

    Full Text Available Abstract The goals of this work were to present original liquid-liquid equilibrium data of the system containing glycerol + ethanol + ethyl biodiesel from fodder radish oil, including the individual distribution of each ethyl ester; to adjust binary parameters of the NRTL; to compare NRTL and UNIFAC-Dortmund in the LLE representation of the system containing glycerol; to simulate different mixer/settler flowsheets for biodiesel purification, evaluating the ratio water/biodiesel used. In thermodynamic modeling, the deviations between experimental data and calculated values were 0.97% and 3.6%, respectively, using NRTL and UNIFAC-Dortmund. After transesterification, with 3 moles of excess ethanol, removal of this component until a content equal to 0.08 before an ideal settling step allows a glycerol content lower than 0.02% in the ester-rich phase. Removal of ethanol, glycerol and water from biodiesel can be performed with countercurrent mixer/settler, using 0.27% of water in relation to the ester amount in the feed stream.

  1. Preparative isolation and purification of phlorotannins from Ecklonia cava using centrifugal partition chromatography by one-step.

    Science.gov (United States)

    Lee, Ji-Hyeok; Ko, Ju-Young; Oh, Jae-Young; Kim, Chul-Young; Lee, Hee-Ju; Kim, Jaeil; Jeon, You-Jin

    2014-09-01

    Various bioactive phlorotannins of Ecklonia cava (e.g., dieckol, eckol, 6,6-bieckol, phloroglucinol, phloroeckol, and phlorofucofuroeckol-A) are reported. However, their isolation and purification are not easy. Centrifugal partition chromatography (CPC) can be used to efficiently purify the various bioactive-compounds efficiently from E. cava. Phlorotannins are successfully isolated from the ethyl acetate (EtOAc) fraction of E. cava by CPC with a two-phase solvent system comprising n-hexane:EtOAc:methanol:water (2:7:3:7, v/v) solution. The dieckol (fraction I, 40.2mg), phlorofucofuroeckol-A (fraction III, 31.1mg), and fraction II (34.1mg) with 2,7-phloroglucinol-6,6-bieckol and pyrogallol-phloroglucinol-6,6-bieckol are isolated from the crude extract (500 mg) by a one-step CPC system. The purities of the isolated dieckol and phlorofucofuroeckol-A are ⩾90% according to high performance liquid chromatography (HPLC) and electrospray ionization multi stage tandem mass spectrometry analyses. The purified 2,7-phloroglucinol-6,6-bieckol and pyrogallol-phloroglucinol-6,6-bieckol are collected from fraction II by recycle-HPLC. Thus, the CPC system is useful for easy and simple isolation of phlorotannins from E. cava. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Bentonite clay purification for development of polymeric nan composites using a single screw extruder

    International Nuclear Information System (INIS)

    Carvalho, Ana C.M. de; Ito, Edson N.; Costa, Maria C.B.; Barbosa, Maria I.R.

    2011-01-01

    In this work, a bentonite clay rich in montmorillonite was purified and chemical treat to be used in the development of poly (methyl methacrylate) /clay nanocomposites via melting processes. After the clay treatment and purification, a masterbatch with 25% clay and 75% PMMA was produced by solution technique, using acetone as solvent. For produce samples with 2.5% clay, the masterbatch along with pure polymer were added and mixed in single screw extruder with a diameter of 16 mm and W/D 26. X-rays diffractometry (XRD) and X-rays fluorescence (XRF). Tests were performed to evaluate and characterizing the bentonite clay used in the development of this work and differential scanning calorimetry (DSC) tests were performed to evaluate changes in the thermal properties of the nanocomposites produced. (author)

  3. A new plastic scintillation resin for single-step separation, concentration and measurement of technetium-99.

    Science.gov (United States)

    Barrera, J; Tarancón, A; Bagán, H; García, J F

    2016-09-14

    Technetium is a synthetic element with no stable isotopes, produced as waste in nuclear power plants and in cyclotrons used for nuclear medicine. The element has high mobility, in the form of TcO4(-); its determination is therefore important for environmental protection. Technetium is found in low concentrations and therefore common methods for its analysis include long treatments in several steps and require large amounts of reagents for its purification and preconcentration. Plastic scintillation resins (PSresin) are novel materials used to separate, preconcentrate and measure radionuclides in a single step. The objective of this study is to prepare and characterise a PSresin for the preconcentration and measurement of (99)Tc. The study first evaluates the reproducibility of the production of PSresins between batches and over time; showing good reproducibility and storage stability. Next, we studied the effect of some common non-radioactive interferences, showing small influences on measurement, and radioactive interferences ((36)Cl and (238)U/(234)U). (36)Cl can be removed by a simple treatment with 0.5 M HCl and (238)U/(234)U can be removed from the column by cleaning with a mixture of 0.1 M HNO3 and 0.1 M HF. In the latter case, a slight change in the morphology of the PSresin caused an increase in detection efficiency. Finally, the PSresin was applied to the measurement of real spiked samples (sea water and urine) with deviations lower than 10% in all cases. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. A new plastic scintillation resin for single-step separation, concentration and measurement of technetium-99

    Energy Technology Data Exchange (ETDEWEB)

    Barrera, J. [Department of Analytical Chemistry, Universitat de Barcelona, Martí i Franqués, 1-11, E-08028, Barcelona (Spain); Tarancón, A., E-mail: alex.tarancon@ub.edu [Department of Analytical Chemistry, Universitat de Barcelona, Martí i Franqués, 1-11, E-08028, Barcelona (Spain); Bagán, H. [Department of Pure and Applied Biochemistry, Lund University, Getingevägen 60, hus II, 22100 SE, Lund (Sweden); García, J.F. [Department of Analytical Chemistry, Universitat de Barcelona, Martí i Franqués, 1-11, E-08028, Barcelona (Spain)

    2016-09-14

    Technetium is a synthetic element with no stable isotopes, produced as waste in nuclear power plants and in cyclotrons used for nuclear medicine. The element has high mobility, in the form of TcO{sub 4}{sup −}; its determination is therefore important for environmental protection. Technetium is found in low concentrations and therefore common methods for its analysis include long treatments in several steps and require large amounts of reagents for its purification and preconcentration. Plastic scintillation resins (PSresin) are novel materials used to separate, preconcentrate and measure radionuclides in a single step. The objective of this study is to prepare and characterise a PSresin for the preconcentration and measurement of {sup 99}Tc. The study first evaluates the reproducibility of the production of PSresins between batches and over time; showing good reproducibility and storage stability. Next, we studied the effect of some common non-radioactive interferences, showing small influences on measurement, and radioactive interferences ({sup 36}Cl and {sup 238}U/{sup 234}U). {sup 36}Cl can be removed by a simple treatment with 0.5 M HCl and {sup 238}U/{sup 234}U can be removed from the column by cleaning with a mixture of 0.1 M HNO{sub 3} and 0.1 M HF. In the latter case, a slight change in the morphology of the PSresin caused an increase in detection efficiency. Finally, the PSresin was applied to the measurement of real spiked samples (sea water and urine) with deviations lower than 10% in all cases. - Highlights: • A plastic scintillation resin for selective analysis of {sup 99}Tc has been developed. • The method is valid for analysis of {sup 99}Tc in seawater and urine samples. • Presence of Cl{sup −}, NO{sub 3}{sup −}, SO{sub 4}{sup 2−}, {sup 36}Cl, U and Th not affect retention of {sup 99}Tc.

  5. One-step purification and characterization of alginate lyase from a clinical Pseudomonas aeruginosa with destructive activity on bacterial biofilm

    Directory of Open Access Journals (Sweden)

    Parinaz Ghadam

    2017-05-01

    Full Text Available Objective(s: Pseudomonas aeruginosais a Gram-negative and aerobic rod bacterium that displays mucoid and non-mucoid phenotype. Mucoid strains secrete alginate, which is the main agent of biofilms in chronic P. aeruginosa infections, show high resistance to antibiotics; consequently, the biological disruption of mucoid P. aeruginosa biofilms is an attractive area of study for researchers. Alginate lyase gene (algl is a member of alginate producing operon which by glycosidase activity produces primer for other enzymes in this cluster. Also this activity can destroy the extracellular alginate; therefore this enzyme participates in alginate production and destruction pathway. Alginate lyase causes detachment of a biofilm by reducing its adhesion to the surfaces, and increases phagocytosis and antibiotic susceptibility. In this study, alginate lyase was purified in just one step and its properties were investigated. Materials and Methods: The purification was done by affinity chromatography, analysed by SDS-PAGE, and its effect on P. aeruginosa biofilms was surveyed by micro titer plate assay and SEM. The substrate specificity of the enzyme was determined by PCR. Results: Alginate lyase from isolate 48 was purified in one step. It is more thermally resistant than alginate lyase from Pseudomonas aeruginosa PAO1 and poly M, poly G and poly MG alginate were the substrate of this enzyme. Moreover, it has an eradication effect on biofilms from P. aeruginosa 48 and PAO1. Conclusion: In this study an alginate lyase with many characteristics suitable in medicine such as thermal stability, effective on poly M alginate, and bacterial biofilm destructive was introduced and purified.

  6. The Effects of Multiple-Step and Single-Step Directions on Fourth and Fifth Grade Students' Grammar Assessment Performance

    Science.gov (United States)

    Mazerik, Matthew B.

    2006-01-01

    The mean scores of English Language Learners (ELL) and English Only (EO) students in 4th and 5th grade (N = 110), across the teacher-administered Grammar Skills Test, were examined for differences in participants' scores on assessments containing single-step directions and assessments containing multiple-step directions. The results indicated no…

  7. Online hyphenation of extraction, Sephadex LH-20 column chromatography, and high-speed countercurrent chromatography: A highly efficient strategy for the preparative separation of andrographolide from Andrographis paniculata in a single step.

    Science.gov (United States)

    Zhang, Ying-Qi; Wang, Shan-Shan; Han, Chao; Xu, Jin-Fang; Luo, Jian-Guang; Kong, Ling-Yi

    2017-12-01

    A novel isolation strategy, online hyphenation of ultrasonic extraction, Sephadex LH-20 column chromatography combined with high-speed countercurrent chromatography, was developed for pure compounds extraction and purification. Andrographolide from Andrographis paniculata was achieved only in a single step purification protocol via the present strategy. The crude powder was ultrasonic extracted and extraction was pumped into Sephadex LH-20 column directly to cut the nontarget fractions followed by the second-dimensional high-speed countercurrent chromatography, hyphenated by a six-port valve equipped at the post-end of Sephadex LH-20 column, for the final purification. The results yielded andrographolide with the amount of 1.02 mg and a purity of 98.5% in a single step, indicating that the present method is effective to harvest target compound from medicinal plant. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Caprylic acid-induced impurity precipitation from protein A capture column elution pool to enable a two-chromatography-step process for monoclonal antibody purification.

    Science.gov (United States)

    Zheng, Ji; Wang, Lu; Twarowska, Barbara; Laino, Sarah; Sparks, Colleen; Smith, Timothy; Russell, Reb; Wang, Michelle

    2015-01-01

    This article presents the use of caprylic acid (CA) to precipitate impurities from the protein A capture column elution pool for the purification of monoclonal antibodies (mAbs) with the objective of developing a two chromatography step antibody purification process. A CA-induced impurity precipitation in the protein A column elution pool was evaluated as an alternative method to polishing chromatography techniques for use in the purification of mAbs. Parameters including pH, CA concentrations, mixing time, mAb concentrations, buffer systems, and incubation temperatures were evaluated on their impacts on the impurity removal, high-molecular weight (HMW) formation and precipitation step yield. Both pH and CA concentration, but not mAb concentrations and buffer systems, are key parameters that can affect host-cell proteins (HCPs) clearance, HMW species, and yield. CA precipitation removes HCPs and some HMW species to the acceptable levels under the optimal conditions. The CA precipitation process is robust at 15-25°C. For all five mAbs tested in this study, the optimal CA concentration range is 0.5-1.0%, while the pH range is from 5.0 to 6.0. A purification process using two chromatography steps (protein A capture column and ion exchange polishing column) in combination with CA-based impurity precipitation step can be used as a robust downstream process for mAb molecules with a broad range of isoelectric points. Residual CA can be effectively removed by the subsequent polishing cation exchange chromatography. © 2015 American Institute of Chemical Engineers.

  9. Comparative analysis of single-step and two-step biodiesel production using supercritical methanol on laboratory-scale

    International Nuclear Information System (INIS)

    Micic, Radoslav D.; Tomić, Milan D.; Kiss, Ferenc E.; Martinovic, Ferenc L.; Simikić, Mirko Ð.; Molnar, Tibor T.

    2016-01-01

    Highlights: • Single-step supercritical transesterification compared to the two-step process. • Two-step process: oil hydrolysis and subsequent supercritical methyl esterification. • Experiments were conducted in a laboratory-scale batch reactor. • Higher biodiesel yields in two-step process at milder reaction conditions. • Two-step process has potential to be cost-competitive with the single-step process. - Abstract: Single-step supercritical transesterification and two-step biodiesel production process consisting of oil hydrolysis and subsequent supercritical methyl esterification were studied and compared. For this purpose, comparative experiments were conducted in a laboratory-scale batch reactor and optimal reaction conditions (temperature, pressure, molar ratio and time) were determined. Results indicate that in comparison to a single-step transesterification, methyl esterification (second step of the two-step process) produces higher biodiesel yields (95 wt% vs. 91 wt%) at lower temperatures (270 °C vs. 350 °C), pressures (8 MPa vs. 12 MPa) and methanol to oil molar ratios (1:20 vs. 1:42). This can be explained by the fact that the reaction system consisting of free fatty acid (FFA) and methanol achieves supercritical condition at milder reaction conditions. Furthermore, the dissolved FFA increases the acidity of supercritical methanol and acts as an acid catalyst that increases the reaction rate. There is a direct correlation between FFA content of the product obtained in hydrolysis and biodiesel yields in methyl esterification. Therefore, the reaction parameters of hydrolysis were optimized to yield the highest FFA content at 12 MPa, 250 °C and 1:20 oil to water molar ratio. Results of direct material and energy costs comparison suggest that the process based on the two-step reaction has the potential to be cost-competitive with the process based on single-step supercritical transesterification. Higher biodiesel yields, similar or lower energy

  10. Method and apparatus for single-stepping coherence events in a multiprocessor system under software control

    Science.gov (United States)

    Blumrich, Matthias A.; Salapura, Valentina

    2010-11-02

    An apparatus and method are disclosed for single-stepping coherence events in a multiprocessor system under software control in order to monitor the behavior of a memory coherence mechanism. Single-stepping coherence events in a multiprocessor system is made possible by adding one or more step registers. By accessing these step registers, one or more coherence requests are processed by the multiprocessor system. The step registers determine if the snoop unit will operate by proceeding in a normal execution mode, or operate in a single-step mode.

  11. Single-Step Ironmaking from Ore to Improve Energy Efficiency

    Energy Technology Data Exchange (ETDEWEB)

    S.K. Kawatra; B. Anamerie; T.C. Eisele

    2005-10-01

    The pig iron nugget process was developed as an alternative to the traditional blast furnace process by Kobe Steel. The process aimed to produce pig iron nuggets, which have similar chemical and physical properties to blast furnace pig iron, in a single step. The pig iron nugget process utilizes coal instead of coke and self reducing and fluxing dried green balls instead of pellets and sinters. In this process the environmental emissions caused by coke and sinter production, and energy lost between pellet induration (heat hardening) and transportation to the blast furnace can be eliminated. The objectives of this research were to (1) produce pig iron nuggets in the laboratory, (2) characterize the pig iron nugget produced and compare them with blast furnace pig iron, (3) investigate the furnace temperature and residence time effects on the pig iron nugget production, and (4) optimize the operational furnace temperatures and residence times. The experiments involved heat treatment of self reducing and fluxing dried green balls at various furnace temperatures and residence times. Three chemically and physically different products were produced after the compete reduction of iron oxides to iron depending on the operational furnace temperatures and/or residence times. These products were direct reduced iron (DRI), transition direct reduced iron (TDRI), and pig iron nuggets. The increase in the carbon content of the system as a function of furnace temperature and/or residence time dictated the formation of these products. The direct reduced iron, transition direct reduced iron, and pig iron nuggets produced were analyzed for their chemical composition, degree of metallization, apparent density, microstructure and microhardness. In addition, the change in the carbon content of the system with the changing furnace temperature and/or residence time was detected by optical microscopy and Microhardness measurements. The sufficient carbon dissolution required for the

  12. A single-ligand ultra-microporous MOF for precombustion CO2 capture and hydrogen purification.

    Science.gov (United States)

    Nandi, Shyamapada; De Luna, Phil; Daff, Thomas D; Rother, Jens; Liu, Ming; Buchanan, William; Hawari, Ayman I; Woo, Tom K; Vaidhyanathan, Ramanathan

    2015-12-01

    Metal organic frameworks (MOFs) built from a single small ligand typically have high stability, are rigid, and have syntheses that are often simple and easily scalable. However, they are normally ultra-microporous and do not have large surface areas amenable to gas separation applications. We report an ultra-microporous (3.5 and 4.8 Å pores) Ni-(4-pyridylcarboxylate)2 with a cubic framework that exhibits exceptionally high CO2/H2 selectivities (285 for 20:80 and 230 for 40:60 mixtures at 10 bar, 40°C) and working capacities (3.95 mmol/g), making it suitable for hydrogen purification under typical precombustion CO2 capture conditions (1- to 10-bar pressure swing). It exhibits facile CO2 adsorption-desorption cycling and has CO2 self-diffusivities of ~3 × 10(-9) m(2)/s, which is two orders higher than that of zeolite 13X and comparable to other top-performing MOFs for this application. Simulations reveal a high density of binding sites that allow for favorable CO2-CO2 interactions and large cooperative binding energies. Ultra-micropores generated by a small ligand ensures hydrolytic, hydrostatic stabilities, shelf life, and stability toward humid gas streams.

  13. A single-ligand ultra-microporous MOF for precombustion CO2 capture and hydrogen purification

    Science.gov (United States)

    Nandi, Shyamapada; De Luna, Phil; Daff, Thomas D.; Rother, Jens; Liu, Ming; Buchanan, William; Hawari, Ayman I.; Woo, Tom K.; Vaidhyanathan, Ramanathan

    2015-01-01

    Metal organic frameworks (MOFs) built from a single small ligand typically have high stability, are rigid, and have syntheses that are often simple and easily scalable. However, they are normally ultra-microporous and do not have large surface areas amenable to gas separation applications. We report an ultra-microporous (3.5 and 4.8 Å pores) Ni-(4-pyridylcarboxylate)2 with a cubic framework that exhibits exceptionally high CO2/H2 selectivities (285 for 20:80 and 230 for 40:60 mixtures at 10 bar, 40°C) and working capacities (3.95 mmol/g), making it suitable for hydrogen purification under typical precombustion CO2 capture conditions (1- to 10-bar pressure swing). It exhibits facile CO2 adsorption-desorption cycling and has CO2 self-diffusivities of ~3 × 10−9 m2/s, which is two orders higher than that of zeolite 13X and comparable to other top-performing MOFs for this application. Simulations reveal a high density of binding sites that allow for favorable CO2-CO2 interactions and large cooperative binding energies. Ultra-micropores generated by a small ligand ensures hydrolytic, hydrostatic stabilities, shelf life, and stability toward humid gas streams. PMID:26824055

  14. The Application of Dextran Sedimentation as an Initial Step in Neutrophil Purification Promotes Their Stimulation, due to the Presence of Monocytes

    Science.gov (United States)

    Ferrante, Antonio

    2017-01-01

    The purification of human neutrophils for in vitro studies is challenging as they are easily activated through ex vivo manipulations. The technique of erythrocyte sedimentation combined with density gradient centrifugation remains widely practiced and was the subject of this study. Since in the sedimentation step the leukocytes are incubated with dextran, we have raised the likelihood that cellular activation would occur with mediator release leading to neutrophil activation. By comparing the activity of neutrophils purified from whole blood by the classical 2-step method of dextran sedimentation followed by low-density Ficoll-Hypaque (1.077 g/mL) medium, and the 1-step high-density Ficoll-Hypaque (1.114 g/mL) gradient centrifugation, we found that neutrophils from the 2-step method had a significant increase in cell surface CD11b expression and CD62L shedding and a marked increase in adhesion. Decreased random migration and chemotaxis and raised baseline oxidative burst activity were also observed. The effect was not specific to dextran, as using Ficoll for erythrocyte sedimentation replicated the elevated neutrophil adherence. Through the depletion of monocytes, lymphocytes, and platelets prior to sedimentation, we deduced that monocytes were responsible for the neutrophil activation. Our findings suggest that care needs to be exercised in choosing the method of neutrophil purification for functional studies. PMID:29164154

  15. Development and scale-up of the recovery and purification of a domain antibody Fc fusion protein-comparison of a two and three-step approach.

    Science.gov (United States)

    Herzer, Sibylle; Bhangale, Atul; Barker, Gregory; Chowdhary, Isha; Conover, Matthew; O'Mara, Brian W; Tsang, Lily; Wang, Shi-Yu; Krystek, Stanley R; Yao, Yan; Rieble, Siegfried

    2015-07-01

    A robust, economical process should leverage proven technology, yet be flexible enough to adopt emerging technologies which show significant benefit. Antibody and Fc-fusion processes may capitalize on the high selectivity of an affinity capture step by reducing the total number of chromatographic steps to 2. Risk associated with this approach stems from the potentially increased time frame needed for process development as well as unforeseen changes in impurity profile during first scale-up of drug substance (DS) for animal toxicology and clinical phase I trials (FIH) production, which could challenge a two-step process to the point of failure. Two different purification strategies were pursued during process development for an FIH process of a dAB-Fc fusion protein. A two-step process was compared to a three-step process. The two-step process leveraged additives to maximize impurity reduction during affinity capture. While wash additives in combination with a mixed mode chromatography met all impurity reduction requirements, HCP levels were still higher as compared to the three-step process. The three-step process was implemented for manufacture of clinical material to mitigate risk. © 2015 Wiley Periodicals, Inc.

  16. The possibility of a fully automated procedure for radiosynthesis of fluorine-18-labeled fluoromisonidazole using a simplified single, neutral alumina column purification procedure

    International Nuclear Information System (INIS)

    Nandy, Saikat; Rajan, M.G.R.; Korde, A.; Krishnamurthy, N.V.

    2010-01-01

    A novel fully automated radiosynthesis procedure for [ 18 F]Fluoromisonidazole using a simple alumina cartridge-column for purification instead of conventionally used semi-preparative HPLC was developed. [ 18 F]FMISO was prepared via a one-pot, two-step synthesis procedure using a modified nuclear interface synthesis module. Nucleophilic fluorination of the precursor molecule 1-(2'-nitro-1'-imidazolyl) -2-O-tetrahydropyranyl-3-O-toluenesulphonylpropanediol (NITTP) with no-carrier added [ 18 F]fluoride followed by hydrolysis of the protecting group with 1 M HCl. Purification was carried out using a single neutral alumina cartridge-column instead of semi-preparative HPLC. The maximum overall radiochemical yield obtained was 37.49±1.68% with 10 mg NITTP (n=3, without any decay correction) and the total synthesis time was 40±1 min. The radiochemical purity was greater than 95% and the product was devoid of other chemical impurities including residual aluminum and acetonitrile. The biodistribution study in fibrosarcoma tumor model showed maximum uptake in tumor, 2 h post injection. Finally, PET/CT imaging studies in normal healthy rabbit, showed clear uptake in the organs involved in the metabolic process of MISO. No bone uptake was observed excluding the presence of free [ 18 F]fluoride. The reported method can be easily adapted in any commercial FDG synthesis module.

  17. The stepping behavior analysis of pedestrians from different age groups via a single-file experiment

    Science.gov (United States)

    Cao, Shuchao; Zhang, Jun; Song, Weiguo; Shi, Chang'an; Zhang, Ruifang

    2018-03-01

    The stepping behavior of pedestrians with different age compositions in single-file experiment is investigated in this paper. The relation between step length, step width and stepping time are analyzed by using the step measurement method based on the calculation of curvature of the trajectory. The relations of velocity-step width, velocity-step length and velocity-stepping time for different age groups are discussed and compared with previous studies. Finally effects of pedestrian gender and height on stepping laws and fundamental diagrams are analyzed. The study is helpful for understanding pedestrian dynamics of movement. Meanwhile, it offers experimental data to develop a microscopic model of pedestrian movement by considering stepping behavior.

  18. Two-step ion-exchange chromatographic purification combined with reversed-phase chromatography to isolate C-peptide for mass spectrometric analysis.

    Science.gov (United States)

    Kabytaev, Kuanysh; Durairaj, Anita; Shin, Dmitriy; Rohlfing, Curt L; Connolly, Shawn; Little, Randie R; Stoyanov, Alexander V

    2016-02-01

    A liquid chromatography with mass spectrometry on-line platform that includes the orthogonal techniques of ion exchange and reversed phase chromatography is applied for C-peptide analysis. Additional improvement is achieved by the subsequent application of cation- and anion-exchange purification steps that allow for isolating components that have their isoelectric points in a narrow pH range before final reversed-phase mass spectrometry analysis. The utility of this approach for isolating fractions in the desired "pI window" for profiling complex mixtures is discussed. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Steps to detect catalytic ethylene oxide formation on single crystals

    Energy Technology Data Exchange (ETDEWEB)

    Boecklein, Sebastian; Guenther, Sebastian; Reichelt, Robert; Seibald, Markus; Preimesser, Andreas; Ehrensberger, Martin; Rozsa, Gergely; Wintterlin, Joost [Ludwig-Maximilians-Universitaet, 81377 Muenchen (Germany)

    2010-07-01

    As part of a project to bridge the ''pressure gap'' for the catalytic synthesis of ethylene oxide (EtO) on Ag surfaces we have undertaken extensive studies in a model reactor. The investigations aimed at finding conditions under which the production of EtO can be unambiguously and quantitatively detected on single crystal Ag surfaces, a challenging task because of the extremely low ethylene-to-EtO reaction probability. The experiments were performed in a specially designed reactor, and they involved the variation of partial pressures, temperature, and type of Ag samples (powders and polycrystalline sheets), and great effort was expended for proper background subtraction. We find that for the sheets an essential ingredient is an activation treatment by annealing in oxygen, which raises the activity by more than one order of magnitude. There are indications that subsurface O atoms are created by this pretreatment. The maximum values obtained for activity, selectivity, yield, and reaction probability allow us to predict that EtO produced on a single Ag crystal can indeed be detected under flow conditions in a UHV chamber. Experiments on the deactivation show that sintering plays an important role for the dispersed samples, but that there is an additional deactivation process for the sheets that is not caused by sintering or poisoning.

  20. Production of alpha-amylase from Aspergillus oryzae for several industrial applications in a single step.

    Science.gov (United States)

    Porfirif, María C; Milatich, Esteban J; Farruggia, Beatriz M; Romanini, Diana

    2016-06-01

    A one-step method as a strategy of alpha-amylase concentration and purification was developed in this work. This methodology requires the use of a very low concentration of biodegradable polyelectrolyte (Eudragit(®) E-PO) and represents a low cost, fast, easy to scale up and non-polluting technology. Besides, this methodology allows recycling the polymer after precipitation. The formation of reversible soluble/insoluble complexes between alpha-amylase and the polymer Eudragit(®) E-PO was studied, and their precipitation in selected conditions was applied with bioseparation purposes. Turbidimetric assays allowed to determine the pH range where the complexes are insoluble (4.50-7.00); pH 5.50 yielded the highest turbidity of the system. The presence of NaCl (0.05M) in the medium totally dissociates the protein-polymer complexes. When the adequate concentration of polymer was added under these conditions to a liquid culture of Aspergillus oryzae, purification factors of alpha-amylase up to 7.43 and recoveries of 88% were obtained in a simple step without previous clarification. These results demonstrate that this methodology is suitable for the concentration and production of alpha-amylase from this source and could be applied at the beginning of downstream processing. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Purification and Characterization of Lipase from Aspergillus flavus ...

    African Journals Online (AJOL)

    Lipase from Aspergillus flavus was purified in a single step purification using MnFeO4 magnetic nano particles to achieve a 20.53- fold purification with specific activity of 11.29 U/mg and a 59% recovery yield. SDS-PAGE of lipase showed a single pure band with corresponding molecular weight of 35 kDa. The optimal ...

  2. One-step purification of R-phycoerythrin from the red macroalga Palmaria palmata using preparative polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Galland-Irmouli, A V; Pons, L; Luçon, M; Villaume, C; Mrabet, N T; Guéant, J L; Fleurence, J

    2000-02-28

    Phycoerythrin is a major light-harvesting pigment of red algae and cyanobacteria widely used as a fluorescent probe. In this study, phycoerythrin of the red macroalga Palmaria palmata was extracted by grinding the algal sample in liquid nitrogen, homogenisation in phosphate buffer and centrifugation. Phycoerythrin was then purified from this crude extract using preparative polyacrylamide gel electrophoresis (PAGE) with a continuous elution system and detected by its pink colour and fluorescence. The pigment presented a typical spectrum of R-phycoerythrin, with three absorbance maxima at 499, 545 and 565 nm, and displayed a fluorescence maximum at 578 nm. The absorbance ratio A565/A280, a criterion for purity, was 3.2. A single protein of relative molecular mass 240,000 was detected on native-PAGE with silver staining. Sodium dodecyl sulphate-PAGE demonstrated the presence of two major subunits with Mr 20,000 and 21,000, respectively, and a very minor subunit of Mr 30,000. These observations are consistent with the (alphabeta)6gamma subunit composition characteristic of R-phycoerythrin. Phycoerythrin of Palmaria palmata was determined to be present in larger amounts in autumn and showed a good stability up to 60 degrees C and between pH 3.5 and 9.5. In conclusion, phycoerythrin of Palmaria palmata was purified in a single-step using preparative PAGE. Obtaining pure R-phycoerythrin of Palmaria palmata will allow one to evaluate its fluorescence properties for future applications in biochemical techniques.

  3. A single step solid-phase extraction method for complete separation of sterol oxidation products in food lipids.

    Science.gov (United States)

    Azadmard-Damirchi, Sodeif; Dutta, Paresh C

    2009-01-02

    One of the crucial steps in determination of sterol oxidation products (SOPs) in foods is their enrichment and purifications by various preparative methods for further analysis by GC and GC-MS. Among the preparative methods, SPE of various adsorbents and solvent systems, are being used most widely. At present, no single step SPE method is suitable to completely separate the SOPs. In this study, a SPE (1g silica) method, suitable for both transesterified and cold saponified oil samples, was developed to separate completely SOPs from other lipid components. This method resulted in high recovery from rapeseed oil of added 5beta,6beta-epoxycholestan-3beta-ol (94-96%), cholest-5-en-3beta-ol-7-one(94%), cholestane-3beta,5alpha,6beta-triol (88-91%), cholest-5-en-3beta,7alpha-diol and 5alpha,6alpha-epoxycholestan-3beta-ol (88-90%). The method has a high sample capacity of up to 1g transesterified or cold-saponified oil sample. The method was tested and applied to different vegetable oils and to monitor the effects of refining processes on POPs in hazelnut oil.

  4. Improving Genetic Evaluation of Litter Size Using a Single-step Model

    DEFF Research Database (Denmark)

    Guo, Xiangyu; Christensen, Ole Fredslund; Ostersen, Tage

    A recently developed single-step method allows genetic evaluation based on information from phenotypes, pedigree and markers simultaneously. This paper compared reliabilities of predicted breeding values obtained from single-step method and the traditional pedigree-based method for two litter size...... traits, total number of piglets born (TNB), and litter size at five days after birth (Ls 5) in Danish Landrace and Yorkshire pigs. The results showed that the single-step method combining phenotypic and genotypic information provided more accurate predictions than the pedigree-based method, not only...

  5. Single-step antibody-based affinity cryo-electron microscopy for imaging and structural analysis of macromolecular assemblies.

    Science.gov (United States)

    Yu, Guimei; Vago, Frank; Zhang, Dongsheng; Snyder, Jonathan E; Yan, Rui; Zhang, Ci; Benjamin, Christopher; Jiang, Xi; Kuhn, Richard J; Serwer, Philip; Thompson, David H; Jiang, Wen

    2014-07-01

    Single particle cryo-electron microscopy (cryo-EM) is an emerging powerful tool for structural studies of macromolecular assemblies (i.e., protein complexes and viruses). Although single particle cryo-EM requires less concentrated and smaller amounts of samples than X-ray crystallography, it remains challenging to study specimens that are low-abundance, low-yield, or short-lived. The recent development of affinity grid techniques can potentially further extend single particle cryo-EM to these challenging samples by combining sample purification and cryo-EM grid preparation into a single step. Here we report a new design of affinity cryo-EM approach, cryo-SPIEM, that applies a traditional pathogen diagnosis tool Solid Phase Immune Electron Microscopy (SPIEM) to the single particle cryo-EM method. This approach provides an alternative, largely simplified and easier to use affinity grid that directly works with most native macromolecular complexes with established antibodies, and enables cryo-EM studies of native samples directly from cell cultures. In the present work, we extensively tested the feasibility of cryo-SPIEM with multiple samples including those of high or low molecular weight, macromolecules with low or high symmetry, His-tagged or native particles, and high- or low-yield macromolecules. Results for all these samples (non-purified His-tagged bacteriophage T7, His-tagged Escherichiacoli ribosomes, native Sindbis virus, and purified but low-concentration native Tulane virus) demonstrated the capability of cryo-SPIEM approach in specifically trapping and concentrating target particles on TEM grids with minimal view constraints for cryo-EM imaging and determination of 3D structures. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Purification and characterization of xylanase from Aspergillus ...

    African Journals Online (AJOL)

    Xylanase was subjected to a three-step purification scheme involving ammonium sulphate precipitation, gel filtration chromatography and anion exchange chromatography. Purity was verified by running the extracted protein on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and a single band was ...

  7. Expression, purification and characterization of the interferon ...

    Indian Academy of Sciences (India)

    2012-01-19

    Jan 19, 2012 ... utilizing a single-step affinity purification with an appreciable yield of the highly purified protein. Recombinant RNase L was characterized by SDS-PAGE, immunoblotting and MALDI-TOF analysis. A semi-quantitative agarose-gel-based ribonuclease assay was developed for measuring its 2-5A-dependent ...

  8. Purification and characterisation of Cyclodextrin glycosyltransferase ...

    African Journals Online (AJOL)

    EXPER

    2012-06-05

    Jun 5, 2012 ... affinity chromatography were 25.8 and 17.8%, respectively. ... industrial applications according to its characteristic found in the current study. ..... and single step purification of cyclodextrin glycosyltransferase from alkalophilic Bacillus firmus by ion extchange chromatography. Biochem. Eng. J., 39: 510-515.

  9. Development of a method to extract and purify target compounds from medicinal plants in a single step: online hyphenation of expanded bed adsorption chromatography and countercurrent chromatography

    Science.gov (United States)

    Li, Yang; Wang, Nan; Zhang, Min; Ito, Yoichiro; Zhang, Hongyang; Wang, Yuerong; Guo, Xin; Hu, Ping

    2014-01-01

    Pure compounds extracted and purified from natural sources are crucial to lead discovery and drug screening. This study presents a novel two-dimensional hyphenation of expanded bed adsorption chromatography (EBAC) and high-speed countercurrent chromatography (HSCCC) for extraction and purification of target compounds from medicinal plants in a single step. The EBAC and HSCCC were hyphenated via a six-port injection valve as an interface. Fractionation of ingredients of Salvia miltiorrhiza and Rhizoma coptidis was performed on the hyphenated system to verify its efficacy. An amount each of 52.9 mg of salvianolic acid B and 2.1 mg of rosmarinic acid was obtained from Salvia miltiorrhiza by the two-dimensional system in a single step. The purities of the targets were over 96% of salvianolic acid B and 74% of rosmarinic acid. An amount each of 4.6 mg of coptisine and 4.1 mg of berberine was obtained from Rhizoma coptidis each with 98% and 82% purity, respectively. The processing time was nearly 50% that of the multi-step method. These results indicate that the present method is a rapid and green way to harvest targets from medicinal plants in a single step. PMID:24588208

  10. Two-step purification and partial characterization of an extra cellular α-amylase from Bacillus licheniformis

    Directory of Open Access Journals (Sweden)

    Zare Mirakabadi, A.

    2012-11-01

    Full Text Available The aim of this study was production and partial purification of α-amylase enzyme by Bacillus licheniformis. B. Licheniformis was allowed to grow in broth culture for purpose of inducing α-amylase enzyme. Optimal conditions for amylase production by B. Licheniformis are incubation period of 120 h, temperature of 37 °C and pH 7.0. The α-amylase enzyme was purified by ion exchange chromatography on DEAE-sepharose CL-6B and sephadex G-100 gel filtration with a 19.1-fold increase in specific activity as compared to the concentrated supernatant and with a specific activity of 926.47 U/mg. The α-amylase had the highest activity at pH 7.0 and 65 °C. According to the data on native polyacrylamide gel electrophoresis, the molecular weight of the purified enzyme was 72 kDa.

  11. Large-scale purification of pharmaceutical-grade plasmid DNA using tangential flow filtration and multi-step chromatography.

    Science.gov (United States)

    Sun, Bo; Yu, XiangHui; Yin, Yuhe; Liu, Xintao; Wu, Yongge; Chen, Yan; Zhang, Xizhen; Jiang, Chunlai; Kong, Wei

    2013-09-01

    The demand for pharmaceutical-grade plasmid DNA in vaccine applications and gene therapy has been increasing in recent years. In the present study, a process consisting of alkaline lysis, tangential flow filtration, purification by anion exchange chromatography, hydrophobic interaction chromatography and size exclusion chromatography was developed. The final product met the requirements for pharmaceutical-grade plasmid DNA. The chromosomal DNA content was RNA was not detectable by agarose gel electrophoresis. Moreover, the protein content was bacterial cell paste. The overall yield of the final plasmid DNA reached 48%. Therefore, we have established a rapid and efficient production process for pharmaceutical-grade plasmid DNA. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  12. Two-step versus Single Application of Mitomycin-C in Photorefractive Keratectomy for High Myopia

    Directory of Open Access Journals (Sweden)

    Farhad Fazel

    2012-01-01

    Full Text Available Purpose: To evaluate the long-term outcomes of two-step versus single application of mitomycin-C (MMC during photorefractive keratectomy (PRK for high myopia. Methods: This randomized clinical trial included consecutive patients with high myopia (exceeding 7 D. Patients underwent PRK and were randomized to two methods of MMC 0.02% application as follows: in the single application group, MMC was applied for 45 seconds followed by irrigation; in the two-step group MMC was used identically followed by repeat application for another 15 seconds and corneal surface irrigation. Visual acuity, refractive error, pachymetry, topography, corneal haze and complications were compared between the two groups 18 months after surgery. Results: One hundred and forty patients (70 subjects in either study arm underwent PRK according to the study protocol. Mean spherical equivalent refractive error was significantly reduced from baseline to -1.16΁0.39 D in the single application group and to -1.07΁0.39 D in the two-step group. Sixteen (11.5% versus 8 (5.7% eyes lost one or more line(s of best corrected visual acuity in the single application group as compared to the two-step group (P=0.05. Corneal haze was observed in 18 (12.9% and 8 (5.7% eyes in the single application versus two-step group, respectively (P=0.04. Grade 3 corneal haze was not observed in the two-step group but occurred in five eyes (3.6% in the single application group (P=0.03. No eyes developed corneal ectasia during the follow-up period. Conclusion: Two-step intraoperative application of MMC 0.02% in highly myopic eyes undergoing PRK can reduce the frequency and severity of haze formation.

  13. Compatibility of pedigree-based and marker-based relationships for single-step genomic prediction

    DEFF Research Database (Denmark)

    Christensen, Ole Fredslund

    2012-01-01

    Single-step methods for genomic prediction have recently become popular because they are conceptually simple and in practice such a method can completely replace a pedigree-based method for routine genetic evaluation. An issue with single-step methods is compatibility between the marker-based rel......Single-step methods for genomic prediction have recently become popular because they are conceptually simple and in practice such a method can completely replace a pedigree-based method for routine genetic evaluation. An issue with single-step methods is compatibility between the marker...... that it may be important that a single-step method is based on a model conditional on the observed markers. When data are from routine evaluation systems, selection affects the allele frequencies, and therefore both observed markers and observed phenotypes contain information about allele frequencies...... alpha/2. The parameter alpha should be determined from the markers, but since there is selection in routine evaluation systems the phenotypes in principle also provide information about this parameter. The likelihood function used for inference contains two terms. The first term is the REML...

  14. Preparative isolation and purification of seven compounds from Hibiscus mutabilis L. leaves by two-step high-speed counter-current chromatography

    Directory of Open Access Journals (Sweden)

    Hou Zhuoni

    2015-01-01

    Full Text Available Seven compounds from Hibiscus mutabilis L. leaves were first successfully achieved by two-step high-speed counter-current chromatography with two-phase solvent system composed of n-butanol-ethyl acetate-water (1:6:9, v/v/v and n-hexane-ethyl acetate-methanol-water (3:5:3:5, v/v/v/v/. The critical experimental parameters of first-step separation were optimized with response surface methodology as follows: flow rate was 1.1 mL/min, revolution speed was 800 rpm and temperature was 30°C. Under the optimal conditions, around 5.0 mg of salicylic acid, 13.6 mg of rutin, 5.5 mg of genistein were obtained in 100 mg crude sample. Then, 9.2 mg of potengriffioside A, 4.7 mg of kaempferol 3-O-rutinoside, 3.0 mg of steppogenin and 2.5 mg of emodin were obtained by second-step separation. The purities of the seven compounds determined by UPLC were 96.2%, 93.8%, 95.4%, 94.3%, 98.0%, 94.1% and 90.8%, respectively. Their chemical structures were identified by electron spray ionization mass spectroscopy (ESI-MS and 1H, 13C nuclear magnetic resonance (NMR. Furthermore, compound steppogenin and genistein were first reported from Hibiscus mutabilis L. The purification method was simple, efficient and evaded tedious separation process.

  15. Single-walled carbon nanotube purification, pelletization, and surfactant-assisted dispersion: a combined TEM and resonant micro-raman spectroscopy study.

    Science.gov (United States)

    Shen, Kai; Curran, Seamus; Xu, Huifang; Rogelj, Snezna; Jiang, Yingbing; Dewald, James; Pietrass, Tanja

    2005-03-17

    Resonant Raman spectroscopy and transmission electron microscopy were used to characterize the structural changes of three single-walled carbon nanotube samples processed with purification, pelletization, and surfactant-assisted dispersion. A two-stage purification process selectively removes metallic tubes as well as small-diameter ones, enriching large-diameter semiconducting tubes. Pelletizing reduces the intertube distance but greatly increases the intensity ratio of the D band to the G band. Single-walled nanotube (SWNT) bundle size decreases during ultrasonication dispersion aided by a surfactant. SWNT bundles composed of large-diameter tubes are prone to debundling.

  16. RNA integrity as a quality indicator during the first steps of RNP purifications : A comparison of yeast lysis methods

    Directory of Open Access Journals (Sweden)

    Jansen Ralf-Peter

    2004-10-01

    Full Text Available Abstract Background The completion of several genome-sequencing projects has increased our need to assign functions to newly identified genes. The presence of a specific protein domain has been used as the determinant for suggesting a function for these new genes. In the case of proteins that are predicted to interact with mRNA, most RNAs bound by these proteins are still unknown. In yeast, several protocols for the identification of protein-protein interactions in high-throughput analyses have been developed during the last years leading to an increased understanding of cellular proteomics. If any of these protocols or similar approaches shall be used for the identification of mRNA-protein complexes, the integrity of mRNA is a critical factor. Results We compared the effect of different lysis protocols on RNA integrity. We report dramatic differences in RNA stability depending on the method used for yeast cell lysis. Glass bead milling and French Press lead to degraded mRNAs even in the presence of RNase inhibitors. Thus, they are not suitable to purify intact mRNP complexes or to identify specific mRNAs bound to proteins. Conclusion We suggest a novel protocol, grinding deep-frozen cells, for the preparation of protein extracts that contain intact RNAs, as lysis method for the purification of mRNA-protein complexes from yeast cells.

  17. An improved single-step lysis protocol to measure luciferase bioluminescence in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Hasenkamp Sandra

    2012-02-01

    Full Text Available Abstract This report describes the optimization and evaluation of a simple single-step lysis protocol to measure luciferase bioluminescence from genetically modified Plasmodium falciparum. This protocol utilizes a modified commercial buffer to improve speed of assay and consistency in the bioluminescence signal measured by reducing the manipulation steps required to release the cytoplasmic fraction. The utility of this improved assay protocol is demonstrated in typical assays that explore absolute and temporal gene expression activity.

  18. Comparison of Model Reliabilities from Single-Step and Bivariate Blending Methods

    DEFF Research Database (Denmark)

    Taskinen, Matti; Mäntysaari, Esa; Lidauer, Martin

    2013-01-01

    the production trait evaluation of Nordic Red dairy cattle. Genotyped bulls with daughters are used as training animals, and genotyped bulls and producing cows as candidate animals. For simplicity, size of the data is chosen so that the full inverses of the mixed model equation coefficient matrices can......Model based reliabilities in genetic evaluation are compared between three methods: animal model BLUP, single-step BLUP, and bivariate blending after genomic BLUP. The original bivariate blending is revised in this work to better account animal models. The study data is extracted from...... be calculated. Model reliabilities by the single-step and the bivariate blending methods were higher than by animal model due to genomic information. Compared to the single-step method, the bivariate blending method reliability estimates were, in general, lower. Computationally bivariate blending method was...

  19. Three step synthesis of single diastereoisomers of the vicinal trifluoro motif

    Directory of Open Access Journals (Sweden)

    Vincent A. Brunet

    2009-11-01

    Full Text Available A three step route to single diastereoisomers of the vicinal trifluoromethyl motif is described. The route starts from either syn- or anti-α,β-epoxy alcohols and takes a direct approach in that each of the three steps introduces a fluorine atom in a regio- and stereo-specific manner. Starting from either the syn- or the anti-α,β-epoxy alcohol, stereospecific reactions generate two separate diastereoisomeric series of this motif. The route is a significant improvement on an earlier six step strategy.

  20. A data comparison between a traditional and the single-step β-galactosidase assay

    Directory of Open Access Journals (Sweden)

    Jorrit Schaefer

    2016-09-01

    Full Text Available This article describes reproducibility of a single-step automated β-galactosidase, and the equivalence of its data to the traditional assay (“Experiments in Molecular Genetics” [1]. This was done via a pairwise comparison of both methods using strains with Miller Unit [MU] values ranging from 0 to over 2000. The data presented in this article is associated with the research article entitled “A single-step method for mid to high throughput β-galactosidase assays in Escherichia coli using a microplate reader” [2].

  1. Expression and purification of single cysteine-containing mutant variants of the mouse prion protein by oxidative refolding.

    Science.gov (United States)

    Sengupta, Ishita; Udgaonkar, Jayant B

    2017-12-01

    The folding and aggregation of proteins has been studied extensively, using multiple probes. To facilitate such experiments, introduction of spectroscopically-active moieties in to the protein of interest is often necessary. This is commonly achieved by specifically labelling cysteine residues in the protein, which are either present naturally or introduced artificially by site-directed mutagenesis. In the case of the recombinant prion protein, which is normally expressed in inclusion bodies, the presence of the native disulfide bond complicates the correct refolding of single cysteine-containing mutant variants of the protein. To overcome this major bottleneck, a simple purification strategy for single tryptophan, single cysteine-containing mutant variants of the mouse prion protein is presented, with yields comparable to that of the wild type protein. The protein(s) obtained by this method are correctly folded, with a single reduced cysteine, and the native disulfide bond between residues C178 and C213 intact. The β-sheet rich oligomers formed from these mutant variant protein(s) are identical to the wild type protein oligomer. The current strategy facilitates sample preparation for a number of high resolution spectroscopic measurements for the prion protein, which specifically require thiol labelling. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Composition of single-step media used for human embryo culture.

    Science.gov (United States)

    Morbeck, Dean E; Baumann, Nikola A; Oglesbee, Devin

    2017-04-01

    To determine compositions of commercial single-step culture media and test with a murine model whether differences in composition are biologically relevant. Experimental laboratory study. University-based laboratory. Inbred female mice were superovulated and mated with outbred male mice. Amino acid, organic acid, and ions content were determined for single-step culture media: CSC, Global, G-TL, and 1-Step. To determine whether differences in composition of these media are biologically relevant, mouse one-cell embryos were cultured for 96 hours in each culture media at 5% and 20% oxygen in a time-lapse incubator. Compositions of four culture media were analyzed for concentrations of 30 amino acids, organic acids, and ions. Blastocysts at 96 hours of culture and cell cycle timings were calculated, and experiments were repeated in triplicate. Of the more than 30 analytes, concentrations of glucose, lactate, pyruvate, amino acids, phosphate, calcium, and magnesium varied in concentrations. Mouse embryos were differentially affected by oxygen in G-TL and 1-Step. Four single-step culture media have compositions that vary notably in pyruvate, lactate, and amino acids. Blastocyst development was affected by culture media and its interaction with oxygen concentration. Copyright © 2017 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  3. Fully automated radiosynthesis of [18F]Fluoromisonidazole with single neutral alumina column purification: optimization of reaction parameters

    International Nuclear Information System (INIS)

    Nandy, S.K.; Rajan, M.G.R.

    2010-01-01

    1-H-1-(3-[ 18 F]fluoro-2-hydroxypropyl)-2-nitroimidazole ([ 18 F]FMISO), is the most used hypoxia-imaging agent in oncology and we have recently reported a fully automated procedure for its synthesis using the Nuclear Interface FDG module and a single neutral alumina column for purification. Using 1-(2'-nitro-1'-imidazolyl)-2-O-tetra-hydropyranyl-3-O- toluenesulfonylpropanediol (NITTP) as the precursor, we have investigated the yield of [ 18 F]FMISO using different reaction times, temperatures, and the amount of precursor. The overall yield was 48.4 ± 1.2% (n = 3), (without decay correction) obtained using 10 mg NITTP with the radio-fluorination carried out at 145 deg C for 3 min followed by acid hydrolysis for 3 min at 125 deg C in a total synthesis time of 32 ± 1 min. Increasing the precursor amount to 25 mg did not improve the overall yield under identical reaction conditions, with the decay uncorrected yield being 46.8 ± 1.6% (n = 3), but rather made the production less economical. It was also observed that the yield increased linearly with the amount of NITTP used, from 2.5 to 10 mg and plateaued from 10 to 25 mg. Radio-fluorination efficiency at four different conditions was also compared. It was also observed by radio thin layer chromatography (radio-TLC) that the duration of radio-fluorination of NITTP, not the radio-fluorination temperature favoured the formation of labeled thermally degraded product, but the single neutral alumina column purification was sufficient enough to obtain [ 18 F]FMISO devoid of any radiochemical as well as cold impurities. (author)

  4. Single-Step Resection of an Intraosseous Meningioma and Cranial Reconstruction : Technical Note

    NARCIS (Netherlands)

    Broeckx, Charlotte-Elise; Maal, Thomas J. J.; Vreeken, Rinaldo D.; Bos, Ruud R. M.; ter Laan, Mark

    2017-01-01

    OBJECTIVE: Simultaneous tumor resection and cranial reconstruction can be a challenging task. Surgical navigation is an indispensable tool in making this single-step procedure possible. In this technical note, we describe a new technique for this procedure to ensure a precise resection and optimal

  5. A three-step vehicle detection framework for range estimation using a single camera

    CSIR Research Space (South Africa)

    Kanjee, R

    2015-12-01

    Full Text Available This paper proposes and validates a real-time onroad vehicle detection system, which uses a single camera for the purpose of intelligent driver assistance. A three-step vehicle detection framework is presented to detect and track the target vehicle...

  6. Evaluation of Single-Step Steam Pyrolysis-Activated Carbons from ...

    African Journals Online (AJOL)

    Activated carbon has been widely used worldwide as an effective filtration or adsorption material for removing biological and chemical contaminants from drinking water. The potential of producing activated carbon (AC) from local agroforestry residues by single-step steam pyrolysis processes was investigated. The research ...

  7. A simple two-step purification procedure for the iC3b binding collectin conglutinin

    DEFF Research Database (Denmark)

    Krogh-Meibom, Thomas; Ingvartsen, Klaus Lønne; Tornoe, Ida

    2010-01-01

    Bovine conglutinin is a serum protein involved in innate immunity. It binds calcium dependently to iC3b, a product of the complement component C3 deposited on cell surfaces, immune complexes or artificial surfaces after complement activation. We here present a simple and efficient two-step proced......M). SDS-PAGE and time-resolved immunofluorometric assays showed that the conglutinin was not contaminated with other serum collectins such as collectin-43 or mannan-binding lectin....

  8. Differences in Lower Extremity and Trunk Kinematics between Single Leg Squat and Step Down Tasks.

    Directory of Open Access Journals (Sweden)

    Cara L Lewis

    Full Text Available The single leg squat and single leg step down are two commonly used functional tasks to assess movement patterns. It is unknown how kinematics compare between these tasks. The purpose of this study was to identify kinematic differences in the lower extremity, pelvis and trunk between the single leg squat and the step down. Fourteen healthy individuals participated in this research and performed the functional tasks while kinematic data were collected for the trunk, pelvis, and lower extremities using a motion capture system. For the single leg squat task, the participant was instructed to squat as low as possible. For the step down task, the participant was instructed to stand on top of a box, slowly lower him/herself until the non-stance heel touched the ground, and return to standing. This was done from two different heights (16 cm and 24 cm. The kinematics were evaluated at peak knee flexion as well as at 60° of knee flexion. Pearson correlation coefficients (r between the angles at those two time points were also calculated to better understand the relationship between each task. The tasks resulted in kinematics differences at the knee, hip, pelvis, and trunk at both time points. The single leg squat was performed with less hip adduction (p ≤ 0.003, but more hip external rotation and knee abduction (p ≤ 0.030, than the step down tasks at 60° of knee flexion. These differences were maintained at peak knee flexion except hip external rotation was only significant in the 24 cm step down task (p ≤ 0.029. While there were multiple differences between the two step heights at peak knee flexion, the only difference at 60° of knee flexion was in trunk flexion (p < 0.001. Angles at the knee and hip had a moderate to excellent correlation (r = 0.51-0.98, but less consistently so at the pelvis and trunk (r = 0.21-0.96. The differences in movement patterns between the single leg squat and the step down should be considered when selecting a

  9. Identification of two-step chemical mechanisms using small temperature oscillations and a single tagged species.

    Science.gov (United States)

    Closa, F; Gosse, C; Jullien, L; Lemarchand, A

    2015-05-07

    In order to identify two-step chemical mechanisms, we propose a method based on a small temperature modulation and on the analysis of the concentration oscillations of a single tagged species involved in the first step. The thermokinetic parameters of the first reaction step are first determined. Then, we build test functions that are constant only if the chemical system actually possesses some assumed two-step mechanism. Next, if the test functions plotted using experimental data are actually even, the mechanism is attributed and the obtained constant values provide the rate constants and enthalpy of reaction of the second step. The advantage of the protocol is to use the first step as a probe reaction to reveal the dynamics of the second step, which can hence be relieved of any tagging. The protocol is anticipated to apply to many mechanisms of biological relevance. As far as ligand binding is considered, our approach can address receptor conformational changes or dimerization as well as competition with or modulation by a second partner. The method can also be used to screen libraries of untagged compounds, relying on a tracer whose concentration can be spectroscopically monitored.

  10. One-Step Purification of Human Skeletal Muscle Myoblasts and Subsequent Expansion Using Laminin-Coated Surface.

    Science.gov (United States)

    Chowdhury, Shiplu Roy; binti Ismail, Annis; Chee, Sia Chye; bin Laupa, Mohd Suffian; binti Jaffri, Fadhlun; Saberi, Salfarina Ezrina Mohmad; Idrus, Ruszymah Bt Hj

    2015-11-01

    Skeletal myoblasts have been extensively used to study muscle growth and differentiation, and were recently tested for their application as cell therapy and as a gene delivery system to treat muscle and nonmuscle diseases. However, contamination of fibroblasts in isolated cells from skeletal muscle is one of the long-standing problems for routine expansion. This study aimed to establish a simple one-step process to purify myoblasts and maintain their purity during expansion. Mixed cells were preplated serially on laminin- and collagen type I-coated surfaces in a different array for 5, 10, and 15 min. Immunocytochemical staining with antibodies specific to myoblasts was performed to evaluate myoblast attachment efficiency, purity, and yield. It was found that laminin-coated surface favors the attachment of myoblasts. Highest myoblast purity of 78.9% ± 6.8% was achieved by 5 min of preplating only on the laminin-coated surface with a yield of 56.9% ± 3.3%. Primary cells, isolated from skeletal muscle (n = 4), confirm the enhancement of purity through preplating on laminin-coated surface for 5 min. Subsequent expansion after preplating enhanced myoblast purity due to an increase in myoblast growth than fibroblasts. Myoblast purity of ∼ 98% was achieved when another preplating was performed during passaging. In conclusion, myoblasts can be purified and efficiently expanded in one step by preplating on laminin-coated surface, which is a simple and robust technique.

  11. Integrated Method for Purification and Single-Particle Characterization of Lentiviral Vector Systems by Size Exclusion Chromatography and Tunable Resistive Pulse Sensing.

    Science.gov (United States)

    Heider, Susanne; Muzard, Julien; Zaruba, Marianne; Metzner, Christoph

    2017-07-01

    Elements derived from lentiviral particles such as viral vectors or virus-like particles are commonly used for biotechnological and biomedical applications, for example in mammalian protein expression, gene delivery or therapy, and vaccine development. Preparations of high purity are necessary in most cases, especially for clinical applications. For purification, a wide range of methods are available, from density gradient centrifugation to affinity chromatography. In this study we have employed size exclusion columns specifically designed for the easy purification of extracellular vesicles including exosomes. In addition to viral marker protein and total protein analysis, a well-established single-particle characterization technology, termed tunable resistive pulse sensing, was employed to analyze fractions of highest particle load and purity and characterize the preparations by size and surface charge/electrophoretic mobility. With this study, we propose an integrated platform combining size exclusion chromatography and tunable resistive pulse sensing for monitoring production and purification of viral particles.

  12. Dispersed single-phase-step Michelson interferometer for Doppler imaging using sunlight.

    Science.gov (United States)

    Wan, Xiaoke; Ge, Jian

    2012-09-15

    A Michelson interferometer is dispersed with a fiber array-fed spectrograph, providing 59 Doppler sensing channels using sunlight in the 510-570 nm wavelength region. The interferometer operates at a single-phase-step mode, which is particularly advantageous in multiplexing and data processing compared to the phase-stepping mode of other interferometer spectrometer instruments. Spectral templates are prepared using a standard solar spectrum and simulated interferometer modulations, such that the correlation function with a measured 1D spectrum determines the Doppler shift. Doppler imaging of a rotating cylinder is demonstrated. The average Doppler sensitivity is ~12 m/s, with some channels reaching ~5 m/s.

  13. Modeling single-file diffusion with step fractional Brownian motion and a generalized fractional Langevin equation

    International Nuclear Information System (INIS)

    Lim, S C; Teo, L P

    2009-01-01

    Single-file diffusion behaves as normal diffusion at small time and as subdiffusion at large time. These properties can be described in terms of fractional Brownian motion with variable Hurst exponent or multifractional Brownian motion. We introduce a new stochastic process called Riemann–Liouville step fractional Brownian motion which can be regarded as a special case of multifractional Brownian motion with a step function type of Hurst exponent tailored for single-file diffusion. Such a step fractional Brownian motion can be obtained as a solution of the fractional Langevin equation with zero damping. Various kinds of fractional Langevin equations and their generalizations are then considered in order to decide whether their solutions provide the correct description of the long and short time behaviors of single-file diffusion. The cases where the dissipative memory kernel is a Dirac delta function, a power-law function and a combination of these functions are studied in detail. In addition to the case where the short time behavior of single-file diffusion behaves as normal diffusion, we also consider the possibility of a process that begins as ballistic motion

  14. Modeling single-file diffusion with step fractional Brownian motion and a generalized fractional Langevin equation

    Science.gov (United States)

    Lim, S. C.; Teo, L. P.

    2009-08-01

    Single-file diffusion behaves as normal diffusion at small time and as subdiffusion at large time. These properties can be described in terms of fractional Brownian motion with variable Hurst exponent or multifractional Brownian motion. We introduce a new stochastic process called Riemann-Liouville step fractional Brownian motion which can be regarded as a special case of multifractional Brownian motion with a step function type of Hurst exponent tailored for single-file diffusion. Such a step fractional Brownian motion can be obtained as a solution of the fractional Langevin equation with zero damping. Various kinds of fractional Langevin equations and their generalizations are then considered in order to decide whether their solutions provide the correct description of the long and short time behaviors of single-file diffusion. The cases where the dissipative memory kernel is a Dirac delta function, a power-law function and a combination of these functions are studied in detail. In addition to the case where the short time behavior of single-file diffusion behaves as normal diffusion, we also consider the possibility of a process that begins as ballistic motion.

  15. Data on enhanced expression and purification of camelid single domain antibodies from Escherichia coli classical inclusion bodies.

    Science.gov (United States)

    Maggi, Maristella; Scotti, Claudia

    2017-06-01

    Heterologous expression of high amounts of recombinant proteins is a milestone for research and industrial purposes. Single domain antibodies (sdAbs) are heavy-chain only antibody fragments with applications in the biotechnological, medical and industrial fields. The simple nature and small size of sdAbs allows for efficient expression of the soluble molecule in different hosts. However, in some cases, it results in low functional protein yield. To overcome this limitation, expression of a 6xHistag sdAb was attempted in different conditions in Escherichia coli BL21(DE3) cells. Data showed that high amount of sdAb can be expressed in E. coli classical inclusion bodies, efficiently extracted by urea in a short-time, and properly purified by metal ion affinity chromatography. These data originate from the research article "Enhanced expression and purification of camelid single domain VHH antibodies from classical inclusion bodies" Maggi and Scotti (2017) [1] (DOI: http://dx.doi.org/10.1016/j.pep.2017.02.007).

  16. Single-step fabrication of quantum funnels via centrifugal colloidal casting of nanoparticle films.

    KAUST Repository

    Kim, Jin Young

    2015-07-13

    Centrifugal casting of composites and ceramics has been widely employed to improve the mechanical and thermal properties of functional materials. This powerful method has yet to be deployed in the context of nanoparticles--yet size-effect tuning of quantum dots is among their most distinctive and application-relevant features. Here we report the first gradient nanoparticle films to be constructed in a single step. By creating a stable colloid of nanoparticles that are capped with electronic-conduction-compatible ligands we were able to leverage centrifugal casting for thin-films devices. This new method, termed centrifugal colloidal casting, is demonstrated to form films in a bandgap-ordered manner with efficient carrier funnelling towards the lowest energy layer. We constructed the first quantum-gradient photodiode to be formed in a single deposition step and, as a result of the gradient-enhanced electric field, experimentally measured the highest normalized detectivity of any colloidal quantum dot photodetector.

  17. Fast Step Transition and State Identification (STaSI) for Discrete Single-Molecule Data Analysis.

    Science.gov (United States)

    Shuang, Bo; Cooper, David; Taylor, J Nick; Kisley, Lydia; Chen, Jixin; Wang, Wenxiao; Li, Chun Biu; Komatsuzaki, Tamiki; Landes, Christy F

    2014-09-18

    We introduce a step transition and state identification (STaSI) method for piecewise constant single-molecule data with a newly derived minimum description length equation as the objective function. We detect the step transitions using the Student's t test and group the segments into states by hierarchical clustering. The optimum number of states is determined based on the minimum description length equation. This method provides comprehensive, objective analysis of multiple traces requiring few user inputs about the underlying physical models and is faster and more precise in determining the number of states than established and cutting-edge methods for single-molecule data analysis. Perhaps most importantly, the method does not require either time-tagged photon counting or photon counting in general and thus can be applied to a broad range of experimental setups and analytes.

  18. Two-Step Single Slope/SAR ADC with Error Correction for CMOS Image Sensor

    Directory of Open Access Journals (Sweden)

    Fang Tang

    2014-01-01

    Full Text Available Conventional two-step ADC for CMOS image sensor requires full resolution noise performance in the first stage single slope ADC, leading to high power consumption and large chip area. This paper presents an 11-bit two-step single slope/successive approximation register (SAR ADC scheme for CMOS image sensor applications. The first stage single slope ADC generates a 3-bit data and 1 redundant bit. The redundant bit is combined with the following 8-bit SAR ADC output code using a proposed error correction algorithm. Instead of requiring full resolution noise performance, the first stage single slope circuit of the proposed ADC can tolerate up to 3.125% quantization noise. With the proposed error correction mechanism, the power consumption and chip area of the single slope ADC are significantly reduced. The prototype ADC is fabricated using 0.18 μm CMOS technology. The chip area of the proposed ADC is 7 μm × 500 μm. The measurement results show that the energy efficiency figure-of-merit (FOM of the proposed ADC core is only 125 pJ/sample under 1.4 V power supply and the chip area efficiency is 84 k μm2·cycles/sample.

  19. Peyton’s four-step approach: differential effects of single instructional steps on procedural and memory performance – a clarification study

    Directory of Open Access Journals (Sweden)

    Krautter M

    2015-05-01

    Full Text Available Markus Krautter,1 Ronja Dittrich,2 Annette Safi,2 Justine Krautter,1 Imad Maatouk,2 Andreas Moeltner,2 Wolfgang Herzog,2 Christoph Nikendei2 1Department of Nephrology, 2Department of General Internal and Psychosomatic Medicine, University of Heidelberg Medical Hospital, Heidelberg, Germany Background: Although Peyton’s four-step approach is a widely used method for skills-lab training in undergraduate medical education and has been shown to be more effective than standard instruction, it is unclear whether its superiority can be attributed to a specific single step. Purpose: We conducted a randomized controlled trial to investigate the differential learning outcomes of the separate steps of Peyton’s four-step approach. Methods: Volunteer medical students were randomly assigned to four different groups. Step-1 group received Peyton’s Step 1, Step-2 group received Peyton’s Steps 1 and 2, Step-3 group received Peyton’s Steps 1, 2, and 3, and Step-3mod group received Peyton’s Steps 1 and 2, followed by a repetition of Step 2. Following the training, the first independent performance of a central venous catheter (CVC insertion using a manikin was video-recorded and scored by independent video assessors using binary checklists. The day after the training, memory performance during delayed recall was assessed with an incidental free recall test. Results: A total of 97 participants agreed to participate in the trial. There were no statistically significant group differences with regard to age, sex, completed education in a medical profession, completed medical clerkships, preliminary memory tests, or self-efficacy ratings. Regarding checklist ratings, Step-2 group showed a superior first independent performance of CVC placement compared to Step-1 group (P<0.001, and Step-3 group showed a superior performance to Step-2 group (P<0.009, while Step-2 group and Step-3mod group did not differ (P=0.055. The findings were similar in the incidental

  20. Single-Step Fabrication of High-Density Microdroplet Arrays of Low-Surface-Tension Liquids.

    Science.gov (United States)

    Feng, Wenqian; Li, Linxian; Du, Xin; Welle, Alexander; Levkin, Pavel A

    2016-04-01

    A facile approach for surface patterning that enables single-step fabrication of high-density arrays of low-surface-tension organic-liquid microdroplets is described. This approach enables miniaturized and parallel high-throughput screenings in organic solvents, formation of homogeneous arrays of hydrophobic nanoparticles, polymer micropads of specific shapes, and polymer microlens arrays. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Single step hydrothermal based synthesis of M(II)Sb2O6 (M = Cd ...

    Indian Academy of Sciences (India)

    Abstract. Experiments involving single step hydrothermal reactions of the divalent metal (Zn2+, Cd2+, Pb2+,. Cu2+, Ni2+ and Mn2+) salts with ilmenite NaSbO3 yielded pure divalent antimonates in the case of CdSb2O6 crys- tallizing in the PbSb2O6 type structure and ZnSb2O6 crystallizing in the trirutile structure type.

  2. A single-step method for rapid extraction of total lipids from green microalgae.

    Directory of Open Access Journals (Sweden)

    Martin Axelsson

    Full Text Available Microalgae produce a wide range of lipid compounds of potential commercial interest. Total lipid extraction performed by conventional extraction methods, relying on the chloroform-methanol solvent system are too laborious and time consuming for screening large numbers of samples. In this study, three previous extraction methods devised by Folch et al. (1957, Bligh and Dyer (1959 and Selstam and Öquist (1985 were compared and a faster single-step procedure was developed for extraction of total lipids from green microalgae. In the single-step procedure, 8 ml of a 2∶1 chloroform-methanol (v/v mixture was added to fresh or frozen microalgal paste or pulverized dry algal biomass contained in a glass centrifuge tube. The biomass was manually suspended by vigorously shaking the tube for a few seconds and 2 ml of a 0.73% NaCl water solution was added. Phase separation was facilitated by 2 min of centrifugation at 350 g and the lower phase was recovered for analysis. An uncharacterized microalgal polyculture and the green microalgae Scenedesmus dimorphus, Selenastrum minutum, and Chlorella protothecoides were subjected to the different extraction methods and various techniques of biomass homogenization. The less labour intensive single-step procedure presented here allowed simultaneous recovery of total lipid extracts from multiple samples of green microalgae with quantitative yields and fatty acid profiles comparable to those of the previous methods. While the single-step procedure is highly correlated in lipid extractability (r² = 0.985 to the previous method of Folch et al. (1957, it allowed at least five times higher sample throughput.

  3. Optimization of single-step tapering amplitude and energy detuning for high-gain FELs

    Science.gov (United States)

    Li, He-Ting; Jia, Qi-Ka

    2015-01-01

    We put forward a method to optimize the single-step tapering amplitude of undulator strength and initial energy tuning of electron beam to maximize the saturation power of high gain free-electron lasers (FELs), based on the physics of longitudinal electron beam phase space. Using the FEL simulation code GENESIS, we numerically demonstrate the accuracy of the estimations for parameters corresponding to the linac coherent light source and the Tesla test facility.

  4. Single-step isolation of extracellular vesicles by size-exclusion chromatography

    OpenAIRE

    Böing, Anita N.; van der Pol, Edwin; Grootemaat, Anita E.; Coumans, Frank A. W.; Sturk, Auguste; Nieuwland, Rienk

    2014-01-01

    Background: Isolation of extracellular vesicles from plasma is a challenge due to the presence of proteins and lipoproteins. Isolation of vesicles using differential centrifugation or density-gradient ultracentrifugation results in co-isolation of contaminants such as protein aggregates and incomplete separation of vesicles from lipoproteins, respectively.Aim: To develop a single-step protocol to isolate vesicles from human body fluids.Methods: Platelet-free supernatant, derived from platelet...

  5. Single step synthesis, characterization and applications of curcumin functionalized iron oxide magnetic nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Bhandari, Rohit; Gupta, Prachi; Dziubla, Thomas; Hilt, J. Zach, E-mail: zach.hilt@uky.edu

    2016-10-01

    Magnetic iron oxide nanoparticles have been well known for their applications in magnetic resonance imaging (MRI), hyperthermia, targeted drug delivery, etc. The surface modification of these magnetic nanoparticles has been explored extensively to achieve functionalized materials with potential application in biomedical, environmental and catalysis field. Herein, we report a novel and versatile single step methodology for developing curcumin functionalized magnetic Fe{sub 3}O{sub 4} nanoparticles without any additional linkers, using a simple coprecipitation technique. The magnetic nanoparticles (MNPs) were characterized using transmission electron microscopy, X-ray diffraction, fourier transform infrared spectroscopy and thermogravimetric analysis. The developed MNPs were employed in a cellular application for protection against an inflammatory agent, a polychlorinated biphenyl (PCB) molecule. - Graphical abstract: Novel single step curcumin coated magnetic Fe{sub 3}O{sub 4} nanoparticles without any additional linkers for medical, environmental, and other applications. Display Omitted - Highlights: • A novel and versatile single step methodology for developing curcumin functionalized magnetic Fe{sub 3}O{sub 4} nanoparticles is reported. • The magnetic nanoparticles (MNPs) were characterized using TEM, XRD, FTIR and TGA. • The developed MNPs were employed in a cellular application for protection against an inflammatory agent, a polychlorinated biphenyl (PCB).

  6. Computing single step operators of logic programming in radial basis function neural networks

    Energy Technology Data Exchange (ETDEWEB)

    Hamadneh, Nawaf; Sathasivam, Saratha; Choon, Ong Hong [School of Mathematical Sciences, Universiti Sains Malaysia, 11800 USM, Penang (Malaysia)

    2014-07-10

    Logic programming is the process that leads from an original formulation of a computing problem to executable programs. A normal logic program consists of a finite set of clauses. A valuation I of logic programming is a mapping from ground atoms to false or true. The single step operator of any logic programming is defined as a function (T{sub p}:I→I). Logic programming is well-suited to building the artificial intelligence systems. In this study, we established a new technique to compute the single step operators of logic programming in the radial basis function neural networks. To do that, we proposed a new technique to generate the training data sets of single step operators. The training data sets are used to build the neural networks. We used the recurrent radial basis function neural networks to get to the steady state (the fixed point of the operators). To improve the performance of the neural networks, we used the particle swarm optimization algorithm to train the networks.

  7. Computing single step operators of logic programming in radial basis function neural networks

    Science.gov (United States)

    Hamadneh, Nawaf; Sathasivam, Saratha; Choon, Ong Hong

    2014-07-01

    Logic programming is the process that leads from an original formulation of a computing problem to executable programs. A normal logic program consists of a finite set of clauses. A valuation I of logic programming is a mapping from ground atoms to false or true. The single step operator of any logic programming is defined as a function (Tp:I→I). Logic programming is well-suited to building the artificial intelligence systems. In this study, we established a new technique to compute the single step operators of logic programming in the radial basis function neural networks. To do that, we proposed a new technique to generate the training data sets of single step operators. The training data sets are used to build the neural networks. We used the recurrent radial basis function neural networks to get to the steady state (the fixed point of the operators). To improve the performance of the neural networks, we used the particle swarm optimization algorithm to train the networks.

  8. Computing single step operators of logic programming in radial basis function neural networks

    International Nuclear Information System (INIS)

    Hamadneh, Nawaf; Sathasivam, Saratha; Choon, Ong Hong

    2014-01-01

    Logic programming is the process that leads from an original formulation of a computing problem to executable programs. A normal logic program consists of a finite set of clauses. A valuation I of logic programming is a mapping from ground atoms to false or true. The single step operator of any logic programming is defined as a function (T p :I→I). Logic programming is well-suited to building the artificial intelligence systems. In this study, we established a new technique to compute the single step operators of logic programming in the radial basis function neural networks. To do that, we proposed a new technique to generate the training data sets of single step operators. The training data sets are used to build the neural networks. We used the recurrent radial basis function neural networks to get to the steady state (the fixed point of the operators). To improve the performance of the neural networks, we used the particle swarm optimization algorithm to train the networks

  9. [Embryo development in two single-step media: Analysis of 2059 sibling oocytes].

    Science.gov (United States)

    Durand, M; Sermondade, N; Herbemont, C; Benard, J; Gronier, H; Boujenah, J; Cédrin-Durnerin, I; Poncelet, C; Grynberg, M; Sifer, C

    2016-03-01

    The aim of this study was to compare embryo development cultured in two single-step media commercially available: Fert/Sage One Step® (Origio) and Continuous Single Culture® (CSC) (Irvine Scientific). A prospective auto-controlled study of sibling oocytes from women undergoing conventional in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) was performed in our center from February to June 2015. After fertilization, for every patient, half of oocytes were cultured in the single-step Fert/Sage One Step® (serie SAGE) and the other half in the single-step CSC®(serie CSC). Fertilization and embryo morphology rates were assessed by day 1 to day 5-6 if needed. Embryo presentingtwo attempts of IVF and 133 of ICSI were analyzed, corresponding to 2059 inseminated or micro-injected oocytes. Fertilization rate were not different between the 2 series, respectively SAGE vs CSC (IVF: 73.4% vs 68.3% [P=0.49]; ICSI: 58.9% vs 63.8% [P=0.12]). No difference was found for embryo morphology, respectively SAGE vs CSC, at day 2 (top quality embryo at day 2 IVF: 34.4% vs 33% [P=0.98]; ICSI: 42.4% vs 44.9% [P=0.37]; and good quality embryo at day 2 IVF: 44% vs 50.2% [P=0.07]; ICSI: 64% vs 71% [P=0.35]); no difference at day 3 (top quality embryo at day 3 IVF: 19.4% vs 21.3% [P=0.61]; ICSI: 28.7% vs 27.4% [P=0.54]; and good quality embryo at day 3 IVF: 40.4% vs 50.2% [P=0.91]; ICSI: 51% vs 47.6% [P=0.47]). Blastocyst development rate were not different, respectively SAGE vs CSC (IVF: 39.9% vs 41.5% [P=0.63] with 42.9% vs 42.2% of good quality blastocyst [P=0.70]; ICSI: 41.1% vs 37.8% [P=0.18] with 32.9% vs 40.8% of good quality blastocyst [P=0.13]). No difference was found in the useful embryo rate in the 2 series SAGE vs CSC (IVF: 52.8% vs 55.2% [P=0.83]; ICSI: 62.4% vs 61.7% [P=0.70]). Embryo development and rate of useful embryos, transferred or frozen, were not different according to the embryo culture in single-step media Fert/Sage One Step® vs single-step

  10. New purification process of fungal immunomodulatory protein, FIP ...

    African Journals Online (AJOL)

    cy l

    parameters can be amplified linearly. Concentrated supernatant from ultrafiltration was further separated with two-step chromatographic purification of strong cation-exchange and size-exclusion, purified product shows as single band at 13 kDa in SDS-PAGE gel, the analysis result of reversed phase HPLC C4 column also ...

  11. Novel 1:1 labeling and purification process for C-terminal thioester and single cysteine recombinant proteins using generic peptidic toolbox reagents.

    Science.gov (United States)

    Portal, Christophe F; Seifert, Jan-Marcus; Buehler, Christof; Meisner-Kober, Nicole-Claudia; Auer, Manfred

    2014-07-16

    We developed a versatile set of chemical labeling reagents which allow dye ligation to the C-terminus of a protein or a single internal cysteine and target purification in a simple two-step process. This simple process results in a fully 1:1 labeled conjugate suitable for all quantitative fluorescence spectroscopy and imaging experiments. We refer to a "generic labeling toolbox" because of the flexibility to choose one of many available dyes, spacers of different lengths and compositions which increase the target solubility, a variety of affinity purification tags, and different cleavage chemistries to release the 1:1 labeled proteins. Studying protein function in vitro or in the context of live cells and organisms is of vital importance in biological research. Although label free detection technologies gain increasing interest in molecular recognition science, fluorescence spectroscopy is still the most often used detection technique for assays and screens both in academic as well as in industrial groups. For generations, fluorescence spectroscopists have labeled their proteins of interest with small fluorescent dyes by random chemical linking on the proteins' exposed lysines and cysteines. Chemical reactions with a certain excess of activated esters or maleimides of longer wavelength dyes hardly ever result in quantitative labeling of the target protein. Most of the time, more than one exposed amino acid side chain reacts. This results in a mixture of dye-protein complexes of different labeling stoichiometries and labeling sites. Only mass spectrometry allows resolving the precise chemical composition of the conjugates. In "classical" ensemble averaging fluorescent experiments, these labeled proteins are still useful, and quantification of, e.g., ligand binding experiments, is achieved via knowledge of the overall protein concentration and a fluorescent signal change which is proportional to the amount of complex formed. With the development of fluorescence

  12. One-step affinity tag purification of full-length recombinant human AP-1 complexes from bacterial inclusion bodies using a polycistronic expression system.

    Science.gov (United States)

    Wang, Wei-Ming; Lee, A-Young; Chiang, Cheng-Ming

    2008-05-01

    The AP-1 transcription factor is a dimeric protein complex formed primarily between Jun (c-Jun, JunB, JunD) and Fos (c-Fos, FosB, Fra-1, Fra-2) family members. These distinct AP-1 complexes are expressed in many cell types and modulate target gene expression implicated in cell proliferation, differentiation, and stress responses. Although the importance of AP-1 has long been recognized, the biochemical characterization of AP-1 remains limited in part due to the difficulty in purifying full-length, reconstituted dimers with active DNA-binding and transcriptional activity. Using a combination of bacterial coexpression and epitope-tagging methods, we successfully purified all 12 heterodimers (3 Junx4 Fos) of full-length human AP-1 complexes as well as c-Jun/c-Jun, JunD/JunD, and c-Jun/JunD dimers from bacterial inclusion bodies using one-step nickel-NTA affinity tag purification following denaturation and renaturation of coexpressed AP-1 subunits. Coexpression of two constitutive components in a dimeric AP-1 complex helps stabilize the proteins when compared with individual protein expression in bacteria. Purified dimeric AP-1 complexes are functional in sequence-specific DNA binding, as illustrated by electrophoretic mobility shift assays and DNase I footprinting, and are also active in transcription with in vitro-reconstituted human papillomavirus (HPV) chromatin containing AP-1-binding sites in the native configuration of HPV nucleosomes. The availability of these recombinant full-length human AP-1 complexes has greatly facilitated mechanistic studies of AP-1-regulated gene transcription in many biological systems.

  13. Bridge flap technique as a single-step solution to mucogingival problems: A case series

    Directory of Open Access Journals (Sweden)

    Vivek Gupta

    2011-01-01

    Full Text Available Shallow vestibule, gingival recession, inadequate width of attached gingiva (AG and aberrant frenum pull are an array of mucogingival problems for which several independent and effective surgical solutions are reported in the literature. This case series reports the effectiveness of the bridge flap technique as a single-step surgical entity for increasing the depth of the vestibule, root coverage, increasing the width of the AG and solving the problem of abnormal frenum pull. Eight patients with 18 teeth altogether having Millers class I, II or III recession along with problems of shallow vestibule, inadequate width of AG and with or without frenum pull underwent this surgical procedure and were followed-up till 9 months post-operatively. The mean root coverage obtained was 55% and the mean average gain in width of the AG was 3.5 mm. The mean percentage gain in clinical attachment level was 41%. The bridge flap technique can be an effective single-step solution for the aforementioned mucogingival problems if present simultaneously in any case, and offers considerable advantages over other mucogingival surgical techniques in terms of simplicity, limited chair-time for the patient and the operator, single surgical intervention for manifold mucogingival problems and low morbidity because of the absence of palatal donor tissue.

  14. Fast Synthesis of High Quality Biodiesel from ‘Waste Fish Oil’ by Single Step Transesterification

    Directory of Open Access Journals (Sweden)

    Yogesh C. Sharma

    2014-09-01

    Full Text Available A large volume of fish wastes is produced on a daily basis in the Indian sub-continent. This abundant waste source could serve as an economic feedstock for bioenergy generation. In the present study, oil extracted from discarded fish parts was used for high quality biodiesel production. More specifically, a single step transesterification of ‘waste fishoil’ with methanol using sodium methoxide (CH3ONa as homogeneous catalyst under moderate operational conditions resulted in highly pure biodiesel of > 98% of fatty acid methyl ester (FAME content. Characterization was performed by Fourier Transform-Nuclear Magnetic Resonance (FT-NMR.

  15. Real-time, single-step bioassay using nanoplasmonic resonator with ultra-high sensitivity

    Science.gov (United States)

    Zhang, Xiang; Ellman, Jonathan A; Chen, Fanqing Frank; Su, Kai-Hang; Wei, Qi-Huo; Sun, Cheng

    2014-04-01

    A nanoplasmonic resonator (NPR) comprising a metallic nanodisk with alternating shielding layer(s), having a tagged biomolecule conjugated or tethered to the surface of the nanoplasmonic resonator for highly sensitive measurement of enzymatic activity. NPRs enhance Raman signals in a highly reproducible manner, enabling fast detection of protease and enzyme activity, such as Prostate Specific Antigen (paPSA), in real-time, at picomolar sensitivity levels. Experiments on extracellular fluid (ECF) from paPSA-positive cells demonstrate specific detection in a complex bio-fluid background in real-time single-step detection in very small sample volumes.

  16. Binary Factorization in Hopfield-Like Neural Networks: Single-Step Approximation and Computer Simulations

    Czech Academy of Sciences Publication Activity Database

    Frolov, A. A.; Sirota, A.M.; Húsek, Dušan; Muraviev, I. P.

    2004-01-01

    Roč. 14, č. 2 (2004), s. 139-152 ISSN 1210-0552 R&D Projects: GA ČR GA201/01/1192 Grant - others:BARRANDE(EU) 99010-2/99053; Intellectual computer Systems(EU) Grant 2.45 Institutional research plan: CEZ:AV0Z1030915 Keywords : nonlinear binary factor analysis * feature extraction * recurrent neural network * Single-Step approximation * neurodynamics simulation * attraction basins * Hebbian learning * unsupervised learning * neuroscience * brain function modeling Subject RIV: BA - General Mathematics

  17. Stress induced growth of Sn nanowires in a single step by sputtering method

    Science.gov (United States)

    Yadav, A.; Patel, N.; Miotello, A.; Kothari, D. C.

    2015-06-01

    Sn nanowires in aluminum film have been synthesized in a single step by co-sputtering of Al and Sn targets. Due to immiscibility of Sn and Al, co-sputtering leads to generation of stress in the composite film. In order to attain thermodynamic equilibrium, Sn separates from Al and diffuses towards the grain boundaries. External perturbation due to ambient atmosphere leads to corrosion at the grain boundaries forming pits which provide path for Sn to evolve. Owing to this, extrusion of Sn nanowires from Al film occurs to release the residual stress in the film.

  18. "Silicon millefeuille": From a silicon wafer to multiple thin crystalline films in a single step

    Science.gov (United States)

    Hernández, David; Trifonov, Trifon; Garín, Moisés; Alcubilla, Ramon

    2013-04-01

    During the last years, many techniques have been developed to obtain thin crystalline films from commercial silicon ingots. Large market applications are foreseen in the photovoltaic field, where important cost reductions are predicted, and also in advanced microelectronics technologies as three-dimensional integration, system on foil, or silicon interposers [Dross et al., Prog. Photovoltaics 20, 770-784 (2012); R. Brendel, Thin Film Crystalline Silicon Solar Cells (Wiley-VCH, Weinheim, Germany 2003); J. N. Burghartz, Ultra-Thin Chip Technology and Applications (Springer Science + Business Media, NY, USA, 2010)]. Existing methods produce "one at a time" silicon layers, once one thin film is obtained, the complete process is repeated to obtain the next layer. Here, we describe a technology that, from a single crystalline silicon wafer, produces a large number of crystalline films with controlled thickness in a single technological step.

  19. Stability of cell-free DNA from maternal plasma isolated following a single centrifugation step.

    Science.gov (United States)

    Barrett, Angela N; Thadani, Henna A; Laureano-Asibal, Cecille; Ponnusamy, Sukumar; Choolani, Mahesh

    2014-12-01

    Cell-free fetal DNA can be used for prenatal testing with no procedure-related risk to the fetus. However, yield of fetal DNA is low compared with maternal cell-free DNA fragments, resulting in technical challenges for some downstream applications. To maximize the fetal fraction, careful blood processing procedures are essential. We demonstrate that fetal fraction can be preserved using a single centrifugation step followed by postage of plasma to the laboratory for further processing. Digital PCR was used to quantify copies of total, maternal, and fetal DNA present in single-spun plasma at time points over a two-week period, compared with immediately processed double-spun plasma, with storage at room temperature, 4°C, and -80°C representing different postage scenarios. There was no significant change in total, maternal, or fetal DNA copy numbers when single-spun plasma samples were stored for up to 1 week at room temperature and 2 weeks at -80°C compared with plasma processed within 4 h. Following storage at 4°C no change in composition of cell-free DNA was observed. Single-spun plasma can be transported at room temperature if the journey is expected to take one week or less; shipping on dry ice is preferable for longer journeys. © 2014 John Wiley & Sons, Ltd.

  20. Single-Stage Step up/down Driver for Permanent-Magnet Synchronous Machines

    Science.gov (United States)

    Chen, T. R.; Juan, Y. L.; Huang, C. Y.; Kuo, C. T.

    2017-11-01

    The two-stage circuit composed of a step up/down dc converter and a three-phase voltage source inverter is usually adopted as the electric vehicle’s motor driver. The conventional topology is more complicated. Additional power loss resulted from twice power conversion would also cause lower efficiency. A single-stage step up/down Permanent-Magnet Synchronous Motor driver for Brushless DC (BLDC) Motor is proposed in this study. The number components and circuit complexity are reduced. The low frequency six-step square-wave control is used to reduce the switching losses. In the proposed topology, only one active switch is gated with a high frequency PWM signal for adjusting the rotation speed. The rotor position signals are fed back to calculate the motor speed for digital close-loop control in a MCU. A 600W prototype circuit is constructed to drive a BLDC motor with rated speed 3000 rpm, and can control the speed of six sections.

  1. High step-up isolated efficient single switch DC-DC converter for renewable energy source

    Directory of Open Access Journals (Sweden)

    A. Gopi

    2014-12-01

    Full Text Available In this paper, an isolated high step-up single switch DC-DC converter for renewable energy source is proposed. In the proposed converter high step-up voltage is obtained by single power switching technique that operates low duty cycle with isolated transformer inductors and switched capacitors and power diodes. The disadvantage of conventional converters is that it has high duty ratio and high voltage stress on power devices with less efficiency. The proposed converter eliminates the switching losses and recycles the leakage energy which includes reverse recovery energy of the power diode by using passive clamp circuit. To achieve high output voltage gain, the isolated transformer primary terminal and secondary terminal are connected in series during switching operation. PSIM software has been used for simulation. Simulation circuit is analyzed at 40Vdc/400Vdc, 200 W and this operation is validated by implementing in the hardware model at 12Vdc/120Vdc, 60 W.

  2. Single-step fabrication of electrodes with controlled nanostructured surface roughness using optically-induced electrodeposition

    Science.gov (United States)

    Liu, N.; Li, M.; Liu, L.; Yang, Y.; Mai, J.; Pu, H.; Sun, Y.; Li, W. J.

    2018-02-01

    The customized fabrication of microelectrodes from gold nanoparticles (AuNPs) has attracted much attention due to their numerous applications in chemistry and biomedical engineering, such as for surface-enhanced Raman spectroscopy (SERS) and as catalyst sites for electrochemistry. Herein, we present a novel optically-induced electrodeposition (OED) method for rapidly fabricating gold electrodes which are also surface-modified with nanoparticles in one single step. The electrodeposition mechanism, with respect to the applied AC voltage signal and the elapsed deposition time, on the resulting morphology and particle sizes was investigated. The results from SEM and AFM analysis demonstrated that 80-200 nm gold particles can be formed on the surface of the gold electrodes. Simultaneously, both the size of the nanoparticles and the roughness of the fabricated electrodes can be regulated by the deposition time. Compared to state-of-the-art methods for fabricating microelectrodes with AuNPs, such as nano-seed-mediated growth and conventional electrodeposition, this OED technique has several advantages including: (1) electrode fabrication and surface modification using nanoparticles are completed in a single step, eliminating the need for prefabricating micro electrodes; (2) the patterning of electrodes is defined using a digitally-customized, projected optical image rather than using fixed physical masks; and (3) both the fabrication and surface modification processes are rapid, and the entire fabrication process only requires less than 6 s.

  3. Single-step synthesis of PtRu/N-doped graphene for methanol electrocatalytic oxidation

    International Nuclear Information System (INIS)

    Xu, Xiao; Zhou, Yingke; Lu, Jiming; Tian, Xiaohui; Zhu, Hongxi; Liu, Jiangbo

    2014-01-01

    In this work, a single-step route was applied to achieve the reduction of graphene oxide, functional doping of graphene with nitrogen, and deposition of well-dispersed PtRu alloy nanoparticles on the doped graphene simultaneously in the solvent mixture of ethylene glycol and N-methyl-2-pyrrolidone. Transmission electron microscopy, X-ray powder diffraction, Raman spectroscopy and X-ray photoelectron spectroscopy were used to characterize the morphology and microstructure, while cyclic voltammetry, chronoamperometry and electrochemical impedance techniques were carried out to evaluate the electrocatalytic methanol oxidation activity and durability of the obtained PtRu/nitrogen-doped graphene catalysts. Compared to undoped PtRu/graphene, the nitrogen-doped PtRu/graphene catalyst presented better particle size distribution and improved activity and durability of methanol electrocatalytic oxidation, which could be further improved by optimization of the reaction time, temperature, as well as the composition of the reaction medium. The convenient single-step synthesis process for the PtRu/nitrogen-doped graphene catalysts is promising for the potential application of direct methanol fuel cells

  4. Enhanced expression and purification of camelid single domain VHH antibodies from classical inclusion bodies.

    Science.gov (United States)

    Maggi, Maristella; Scotti, Claudia

    2017-08-01

    Single domain antibodies (sdAbs) are small antigen-binding domains derived from naturally occurring, heavy chain-only immunoglobulins isolated from camelid and sharks. They maintain the same binding capability of full-length IgGs but with improved thermal stability and permeability, which justifies their scientific, medical and industrial interest. Several described recombinant forms of sdAbs have been produced in different hosts and with different strategies. Here we present an optimized method for a time-saving, high yield production and extraction of a poly-histidine-tagged sdAb from Escherichia coli classical inclusion bodies. Protein expression and extraction were attempted using 4 different methods (e.g. autoinducing or IPTG-induced soluble expression, non-classical and classical inclusion bodies). The best method resulted to be expression in classical inclusion bodies and urea-mediated protein extraction which yielded 60-70 mg/l bacterial culture. The method we here describe can be of general interest for an enhanced and efficient heterologous expression of sdAbs for research and industrial purposes. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Preparation and photoluminescence characterization of high-purity CdTe single crystals: purification effect of normal freezing on tellurium and cadmium telluride

    Science.gov (United States)

    Song, S. H.; Wang, J.; Isshiki, M.

    2002-03-01

    Extremely high-purity CdTe single crystals have been obtained by the traditional vertical Bridgman technique, beginning with the refining of tellurium material by the normal freezing method. The purification effect of normal freezing on tellurium has been confirmed to be very effective. This effect was also used to prepare extremely high-purity CdTe single crystals. The crystals were characterized by low temperature high-resolution photoluminescence (PL) spectroscopy. Only a sharp peak at 1.5896 eV was detected in the PL spectrum. The full-width at half-maximum is prepared by the Bridgman method.

  6. Comparison on genomic predictions using three GBLUP methods and two single-step blending methods in the Nordic Holstein population

    Directory of Open Access Journals (Sweden)

    Gao Hongding

    2012-07-01

    Full Text Available Abstract Background A single-step blending approach allows genomic prediction using information of genotyped and non-genotyped animals simultaneously. However, the combined relationship matrix in a single-step method may need to be adjusted because marker-based and pedigree-based relationship matrices may not be on the same scale. The same may apply when a GBLUP model includes both genomic breeding values and residual polygenic effects. The objective of this study was to compare single-step blending methods and GBLUP methods with and without adjustment of the genomic relationship matrix for genomic prediction of 16 traits in the Nordic Holstein population. Methods The data consisted of de-regressed proofs (DRP for 5 214 genotyped and 9 374 non-genotyped bulls. The bulls were divided into a training and a validation population by birth date, October 1, 2001. Five approaches for genomic prediction were used: 1 a simple GBLUP method, 2 a GBLUP method with a polygenic effect, 3 an adjusted GBLUP method with a polygenic effect, 4 a single-step blending method, and 5 an adjusted single-step blending method. In the adjusted GBLUP and single-step methods, the genomic relationship matrix was adjusted for the difference of scale between the genomic and the pedigree relationship matrices. A set of weights on the pedigree relationship matrix (ranging from 0.05 to 0.40 was used to build the combined relationship matrix in the single-step blending method and the GBLUP method with a polygenetic effect. Results Averaged over the 16 traits, reliabilities of genomic breeding values predicted using the GBLUP method with a polygenic effect (relative weight of 0.20 were 0.3% higher than reliabilities from the simple GBLUP method (without a polygenic effect. The adjusted single-step blending and original single-step blending methods (relative weight of 0.20 had average reliabilities that were 2.1% and 1.8% higher than the simple GBLUP method, respectively. In

  7. Rapid single-step upconversion-linked immunosorbent assay for diclofenac

    Czech Academy of Sciences Publication Activity Database

    Hlaváček, Antonín; Peterek, M.; Farka, Z.; Mickert, M. J.; Prechtl, L.; Knopp, D.; Gorris, H H.

    2017-01-01

    Roč. 184, č. 10 (2017), s. 4159-4165 ISSN 0026-3672 R&D Projects: GA ČR(CZ) GBP206/12/G014; GA MŠk(CZ) LM2015043 Institutional support: RVO:68081715 Keywords : bioconjugation * electrophoretic purification * immunoassay * luminescence Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 4.580, year: 2016

  8. Rapid single-step upconversion-linked immunosorbent assay for diclofenac

    Czech Academy of Sciences Publication Activity Database

    Hlaváček, Antonín; Peterek, M.; Farka, Z.; Mickert, M. J.; Prechtl, L.; Knopp, D.; Gorris, H H.

    2017-01-01

    Roč. 184, č. 10 (2017), s. 4159-4165 ISSN 0026-3672 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : bioconjugation * electrophoretic purification * immunoassay * luminescence Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 4.580, year: 2016

  9. Magnetic purification of curcumin from Curcuma longa rhizome by novel naked maghemite nanoparticles.

    Science.gov (United States)

    Magro, Massimiliano; Campos, Rene; Baratella, Davide; Ferreira, Maria Izabela; Bonaiuto, Emanuela; Corraducci, Vittorino; Uliana, Maíra Rodrigues; Lima, Giuseppina Pace Pereira; Santagata, Silvia; Sambo, Paolo; Vianello, Fabio

    2015-01-28

    Naked maghemite nanoparticles, namely, surface active maghemite nanoparticles (SAMNs), characterized by a diameter of about 10 nm, possessing peculiar colloidal stability, surface chemistry, and superparamagnetism, present fundamental requisites for the development of effective magnetic purification processes for biomolecules in complex matrices. Polyphenolic molecules presenting functionalities with different proclivities toward iron chelation were studied as probes for testing SAMN suitability for magnetic purification. Thus, the binding efficiency and reversibility on SAMNs of phenolic compounds of interest in the pharmaceutical and food industries, namely, catechin, tyrosine, hydroxytyrosine, ferulic acid, coumaric acid, rosmarinic acid, naringenin, curcumin, and cyanidin-3-glucoside, were evaluated. Curcumin emerged as an elective compound, suitable for magnetic purification by SAMNs from complex matrices. A combination of curcumin, demethoxycurcumin, and bis-demethoxycurcumin was recovered by a single magnetic purification step from extracts of Curcuma longa rhizomes, with a purity >98% and a purification yield of 45%, curcumin being >80% of the total purified curcuminoids.

  10. Comparison of single-entry and double-entry two-step couple screening for cystic fibrosis carriers

    NARCIS (Netherlands)

    tenKate, LP; Verheij, JBGM; Wildhagen, MF; Hilderink, HBM; Kooij, L; Verzijl, JG; Habbema, JDF

    1996-01-01

    Both single-entry two-step (SETS) couple screening and double-entry two-step (DETS) couple screening have been recommended as methods to screen for cystic fibrosis gene carriers. In this paper we compare the expected results from both types of screening. In general, DETS results in a higher

  11. Analytic observations for the d=1+ 1 bridge site (or single-step) deposition model

    International Nuclear Information System (INIS)

    Evans, J.W.; Kang, H.C.

    1991-01-01

    Some exact results for a reversible version of the d=1+1 bridge site (or single-step) deposition model are presented. Exact steady-state properties are determined directly for finite systems with various mean slopes. These show explicitly how the asymptotic growth velocity and fluctuations are quenched as the slope approaches its maximum allowed value. Next, exact hierarchial equations for the dynamics are presented. For the special case of ''equilibrium growth,'' these are analyzed exactly at the pair-correlation level directly for an infinite system. This provided further insight into asymptotic scaling behavior. Finally, the above hierarchy is compared with one generated from a discrete form of the Kardar--Parisi--Zhang equations. Some differences are described

  12. Preparation of orthorhombic Ba2YCu3O7 powder by single-step calcining

    International Nuclear Information System (INIS)

    Spann, J.R.; Kahn, M.; Lloyd, I.K.

    1990-01-01

    A single calcination step, solid-state process that provides orthohombic Ba 2 YCu 3 O 7 powder is described. BaCO 3 , Y 2 O 3 , and CuO are used as precursor materials. The only phase identifiable by x-ray diffraction is the orthorhombic Ba 2 YCu 3 O 7 . The use of a vacuum during the initial stages of the calcining process promotes complete decomposition of the carbonate, and no residual carbonate is observed. An oxygen atmosphere during the later stages of calcining ensures proper oxidation to Ba 2 YCu 3 O 7 . The use of a similar combination vacuum-oxygen calcining schedule should also be beneficial in the preparation of chemically derived powders

  13. A single-step lithography system based on an enhanced robotic adhesive dispenser.

    Science.gov (United States)

    Xing, Jiyao; Rong, Weibin; Sun, Ding; Wang, Lefeng; Sun, Lining

    2016-09-01

    In the paper, we present a single-step lithography system whereby the robotically controlled micro-extrusion of resist adhesive onto a substrate surface to directly create resist adhesive patterns of interest. This system is modified from a robotic adhesive dispenser by shrinking the aperture of the nozzle to a few micrometers aiming to realize patterns at microscale. From experimental investigation, it is found that working factors including writing speed, working time, and applied pressure can be adopted to conveniently regulate the feature size (the width of the line features and the diameter of the dot features). To test its functionality, the system was used to pattern line features on silicon dioxide (SiO 2 ) and generate an array of square-like silicon microstructure by combining with wet etching. It provides a simple and flexible alternative tool to facilitate the development of microfabrication.

  14. A facile single-step synthesis of polyvinylpyrrolidone-silver nanocomposites using a conventional spray dryer.

    Science.gov (United States)

    Kim, Byung-Ho; Kim, Yoon Hyuck; Lee, Young Jin; Lee, Mi Jai; Kim, Jin-Ho; Hwang, Jonghee; Jeon, Dae-Woo

    2018-01-19

    We have developed a facile single-step synthesis of silver nanocomposite using a conventional spray dryer. We investigated the synthetic conditions by controlling the concentrations of the chemical reactants. Further, we confirmed the effect of the molecular weight of polyvinylpyrrolidones, and revealed that the molecular weight significantly affected the properties of the resultant silver nanocomposites. The long-term stability of the silver nanocomposites was tested, and little change was observed, even after storage for three months. Most of all, the simple commercial implementation, in combination with large-scale synthesis, possesses a variety of advantages, compared to conventional complicated and costly dry-process synthesis methods. Thus, our method presents opportunities for further investigation, for both lab-scale studies and large-scale industrial applications.

  15. A facile single-step synthesis of polyvinylpyrrolidone‑silver nanocomposites using a conventional spray dryer

    Science.gov (United States)

    Kim, Byung-Ho; Hyuck Kim, Yoon; Lee, Young Jin; Lee, Mi Jai; Kim, Jin-Ho; Hwang, Jonghee; Jeon, Dae-Woo

    2018-01-01

    We have developed a facile single-step synthesis of silver nanocomposite using a conventional spray dryer. We investigated the synthetic conditions by controlling the concentrations of the chemical reactants. Further, we confirmed the effect of the molecular weight of polyvinylpyrrolidones, and revealed that the molecular weight significantly affected the properties of the resultant silver nanocomposites. The long-term stability of the silver nanocomposites was tested, and little change was observed, even after storage for three months. Most of all, the simple commercial implementation, in combination with large-scale synthesis, possesses a variety of advantages, compared to conventional complicated and costly dry-process synthesis methods. Thus, our method presents opportunities for further investigation, for both lab-scale studies and large-scale industrial applications.

  16. Shapes of Exotic Nuclei by Single- and Multi-Step Coulomb Excitation

    Science.gov (United States)

    Allmond, J. M.; Clarion-Bareball Collaboration; Gretina-Chico2 Collaboration

    2017-09-01

    Coulomb excitation is a powerful technique for probing the shell structure, collectivity, and shapes of atomic nuclei through the measurement of electromagnetic moments. Single- and multi-step Coulomb excitation results from CLARION-BareBall at HRIBF-ORNL (e.g., 136Te) and GRETINA-CHICO2 at CARIBU-ANL (e.g., 106Mo and 106,110Ru) will be presented. A survey of the equipment, techniques, and results will be given. An emphasis will be placed on unique opportunities with 3-MeV/u beams and future directions at CARIBU and ReA3. This material is based upon work supported by the U.S. Department of Energy, Office of Science, Office of Nuclear Physics.

  17. Single step high-speed printing of continuous silver lines by laser-induced forward transfer

    Energy Technology Data Exchange (ETDEWEB)

    Puerto, D., E-mail: puerto@lp3.univ-mrs.fr [Aix-Marseille University, CNRS, LP3 laboratory Campus de Luminy, C.917, Marseille (France); Biver, E. [Aix-Marseille University, CNRS, LP3 laboratory Campus de Luminy, C.917, Marseille (France); Oxford Lasers Ltd., Unit 8, Moorbrook Park, Didcot, OX11 7HP (United Kingdom); Alloncle, A.-P.; Delaporte, Ph. [Aix-Marseille University, CNRS, LP3 laboratory Campus de Luminy, C.917, Marseille (France)

    2016-06-30

    Highlights: • We have performed an experimental study on laser micro-printing of silver nanoparticle inks. • We have achieved the printing of lines in a single pass at velocities of 17 m/s (1 MHz laser). • The ejection dynamics has been investigated by means of a time-resolved imaging technique. • The control of the donor film properties is of prime importance to print lines at high velocities. • Continuous conductive lines of silver inks are laser-printed on PET flexible substrates. - Abstract: The development of high-speed ink printing process by Laser-Induced Forward Transfer (LIFT) is of great interest for the printing community. To address the problems and the limitations of this process that have been previously identified, we have performed an experimental study on laser micro-printing of silver nanoparticle inks by LIFT and demonstrated for the first time the printing of continuous conductive lines in a single pass at velocities of 17 m/s using a 1 MHz repetition rate laser. We investigated the printing process by means of a time-resolved imaging technique to visualize the ejection dynamics of single and adjacent jets. The control of the donor film properties is of prime importance to achieve single step printing of continuous lines at high velocities. We use a 30 ps pulse duration laser with a wavelength of 343 nm and a repetition rate from 0.2 to 1 MHz. A galvanometric mirror head controls the distance between two consecutives jets by scanning the focused beam along an ink-coated donor substrate at different velocities. Droplets and lines of silver inks are laser-printed on glass and PET flexible substrates and we characterized their morphological quality by atomic force microscope (AFM) and optical microscope.

  18. Single step biotransformation of corn oil phytosterols to boldenone by a newly isolated Pseudomonas aeruginosa

    Directory of Open Access Journals (Sweden)

    Mohamed Eisa

    2016-09-01

    Full Text Available A new potent Pseudomonas aeruginosa isolate capable for biotransformation of corn oil phytosterol (PS to 4-androstene-3, 17-dione (AD, testosterone (T and boldenone (BOL was identified by phenotypic analysis and 16S rRNA gene sequencing. Sequential statistical strategy was used to optimize the biotransformation process mainly concerning BOL using Factorial design and response surface methodology (RSM. The production of BOL in single step microbial biotransformation from corn oil phytosterols by P. aeruginosa was not previously reported. Results showed that the pH concentration of the medium, (NH42SO4 and KH2PO4 were the most significant factors affecting BOL production. By analyzing the statistical model of three-dimensional surface plot, BOL production increased from 36.8% to 42.4% after the first step of optimization, and the overall biotransformation increased to 51.9%. After applying the second step of the sequential statistical strategy BOL production increased to 53.6%, and the overall biotransformation increased to 91.9% using the following optimized medium composition (g/l distilled water (NH42SO4, 2; KH2PO4, 4; Na2HPO4. 1; MgSO4·7H2O, 0.3; NaCl, 0.1; CaCl2·2H2O, 0.1; FeSO4·7H2O, 0.001; ammonium acetate 0.001; Tween 80, 0.05%; corn oil 0.5%; 8-hydroxyquinoline 0.016; pH 8; 200 rpm agitation speed and incubation time 36 h at 30 °C. Validation experiments proved the adequacy and accuracy of model, and the results showed the predicted value agreed well with the experimental values.

  19. A novel single step percutaneous access sheath: the initial human experience.

    Science.gov (United States)

    Baldwin, D Duane; Maynes, Lincoln J; Desai, Premal J; Jellison, Forrest C; Tsai, Christopher K; Barker, Gary R

    2006-01-01

    A novel 1-step percutaneous access sheath NS has been developed that allows the insertion of a dilating balloon and renal access sheath in a single step. We present the initial human experience with this sheath. We performed a retrospective chart and database review of the initial 30 consecutive patients undergoing percutaneous nephrostolithotomy using the NS. Data collected included patient demographics, operative and recovery parameters, and complications. Mean patient age was 50.4 years (range 11 to 81), mean body mass index was 31.63 kg/m(2) (range 17.1 to 65) and mean preoperative stone area was 6.23 cm(2) (range 1 to 14.6). Six and 3 patients had full and partial staghorn calculi, respectively. Access was achieved via the upper pole in 16 patients, middle pole in 7 and lower pole in 7. Mean operative time was 114.8 minutes (range 61 to 237). Mean estimated blood loss was 145.5 cc (range 10 to 500) and mean postoperative hospital stay was 4.89 days (range 2 to 14). A total of 23 patients (76.7%) had no residual calculi on postoperative computerized tomography, 5 (16.7%) had residual fragments 4 mm or less and 2 (6.7%) had residual stone fragments greater than 4 mm. There were no complications related to the NS. The NS is safe, easy to use and has potential advantages compared to currently available renal access sheaths.

  20. Robust nanoplasmonic substrates for aptamer macroarrays with single-step detection of PDGF-BB.

    Science.gov (United States)

    Zhu, Dong; Yang, Rui Xiang; Tang, Yu-Ping; Li, Wei; Miao, Zhao Yi; Hu, Yue; Chen, Jun; Yu, Sheng; Wang, Juan; Xu, Chen Yang

    2016-11-15

    An aptamer macroarray on a robust nanoplasmonic substrate with fluorescence enhancement is developed for a single-step sensitive detection of human platelet-derived growth factor-BB (PDGF-BB), a predominant cancer biomarker in cancer angiogenesis. A hybrid Au-nanoparticles-poly (dimethylsiloxane) (PDMS) as nanoplasmonic substrate is prepared via the in-situ reduction of AuCl4(-) ions in PDMS matrixes onto 96 or 384 well plates. In the absence of target molecules, unfolded PDGF-BB aptamer conjugated with dye TAMRA is electrostatically bound to a positively charged poly-L-lysine (PLL)-coated Au nanocomposites film surface, and the fluorescence enhancement effects can be optimized by varying the distance between TAMRA and the Au nanocomposites film, which is easily adjusted by varying the thickness of the biocompatible poly-(acrylic acid) (PAA/PLL) multilayers, and thus metal-enhanced fluorescence of dye TAMRA conjugated with the aptamer is generated up to 15.2-fold. The interaction of the aptamer to its target induces the reversible conformation change of the aptamer, and consequently, the electrostatic potential is overcome by binding force. As a result, the target-binding interaction of the aptamer causes the irreversible detachment of the aptamer from the nanostructured Au film surface to decrease fluorescence of TAMRA. The aptamer macroarray provides not only the appropriate sensitivity for clinical diagnostics with a wide range of linear detection from 10pg/mL to 10μg/mL, high specificity for PDGF-BB against VEGF-165, VEGF-121, NaCl and IgG, and temporal biological stability, but also a single-step detection. We envision that the efficient and robust aptamer macroarray can be extended to the detection of other biomarkers. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. 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

  2. Purification optimization for a recombinant single-chain variable fragment against type 1 insulin-like growth factor receptor (IGF-1R) by using design of experiment (DoE).

    Science.gov (United States)

    Song, Yong-Hong; Sun, Xue-Wen; Jiang, Bo; Liu, Ji-En; Su, Xian-Hui

    2015-12-01

    Design of experiment (DoE) is a statistics-based technique for experimental design that could overcome the shortcomings of traditional one-factor-at-a-time (OFAT) approach for protein purification optimization. In this study, a DoE approach was applied for optimizing purification of a recombinant single-chain variable fragment (scFv) against type 1 insulin-like growth factor receptor (IGF-1R) expressed in Escherichia coli. In first capture step using Capto L, a 2-level fractional factorial analysis and successively a central composite circumscribed (CCC) design were used to identify the optimal elution conditions. Two main effects, pH and trehalose, were identified, and high recovery (above 95%) and low aggregates ratio (below 10%) were achieved at the pH range from 2.9 to 3.0 with 32-35% (w/v) trehalose added. In the second step using cation exchange chromatography, an initial screening of media and elution pH and a following CCC design were performed, whereby the optimal selectivity of the scFv was obtained on Capto S at pH near 6.0, and the optimal conditions for fulfilling high DBC and purity were identified as pH range of 5.9-6.1 and loading conductivity range of 5-12.5 mS/cm. Upon a further gel filtration, the final purified scFv with a purity of 98% was obtained. Finally, the optimized conditions were verified by a 20-fold scale-up experiment. The purities and yields of intermediate and final products all fell within the regions predicted by DoE approach, suggesting the robustness of the optimized conditions. We proposed that the DoE approach described here is also applicable in production of other recombinant antibody constructs. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. Single-step colloidal quantum dot films for infrared solar harvesting

    KAUST Repository

    Kiani, Amirreza

    2016-11-01

    Semiconductors with bandgaps in the near- to mid-infrared can harvest solar light that is otherwise wasted by conventional single-junction solar cell architectures. In particular, colloidal quantum dots (CQDs) are promising materials since they are cost-effective, processed from solution, and have a bandgap that can be tuned into the infrared (IR) via the quantum size effect. These characteristics enable them to harvest the infrared portion of the solar spectrum to which silicon is transparent. To date, IR CQD solar cells have been made using a wasteful and complex sequential layer-by-layer process. Here, we demonstrate ∼1 eV bandgap solar-harvesting CQD films deposited in a single step. By engineering a fast-drying solvent mixture for metal iodide-capped CQDs, we deposited active layers greater than 200 nm in thickness having a mean roughness less than 1 nm. We integrated these films into infrared solar cells that are stable in air and exhibit power conversion efficiencies of 3.5% under illumination by the full solar spectrum, and 0.4% through a simulated silicon solar cell filter.

  4. First steps towards fuel cells testing harmonisation: Procedures and parameters for single cell performance evaluation

    Energy Technology Data Exchange (ETDEWEB)

    Lunghi, P. [Department of Industrial Engineering, University of Perugia, Via Duranti 93, Perugia (Italy); Ubertini, S. [Department of Mechanical Engineering, University of Rome ' ' Tor Vergata' ' , Via di Torvergata, 110, Rome (Italy)

    2004-01-01

    The great interest in Fuel Cell Systems, combined with the innovation of the device itself, has led to a huge developmental effort to make the steps necessary for future FC plant commissioning. The clearest and most effective way to evaluate the performance of a fuel cell is to measure it directly and, since few fuel cell test rigs are available at the moment, standard experimental procedures have not been realized so far. Our research group is currently performing single cell testing at the University of Perugia fuel cell laboratory and particular emphasis has been put on the definition of procedures and the testing of parameterisation. The work team strongly believes that this is the key to effective system testing and reliable performance evaluation. In this work, the test parameterisation developed by the team, and the resulting advanced control procedure used for a single MCFC experimental characterization are presented. Efforts have been dedicated to obtain some relevant non-dimensional parameters to allow an easy understanding of the results and quick comparisons with other tests under different operating conditions, or with results obtained on different cells eventually tested in different laboratories. The authors strongly emphasise this topic to avoid the data that developers and research institutions collect being of no practical use due to a lack of shared rules. (Abstract Copyright [2003], Wiley Periodicals, Inc.)

  5. Single mode step-index polymer optical fiber for humidity insensitive high temperature fiber Bragg grating sensors

    OpenAIRE

    Woyessa, Getinet; Fasano, Andrea; Stefani, Alessio; Markos, Christos; Nielsen, Kristian; Rasmussen, Henrik K.; Bang, Ole

    2016-01-01

    We have fabricated the first single-mode step-index and humidity insensitive polymer optical fiber operating in the 850 nm wavelength ranges. The step-index preform is fabricated using injection molding, which is an efficient method for cost effective, flexible and fast preparation of the fiber preform. The fabricated single-mode step-index (SI) polymer optical fiber (POF) has a 4.8µm core made from TOPAS grade 5013S-04 with a glass transition temperature of 134°C and a 150 µm cladding made f...

  6. One single catalyst, Pd(OAc)2, for two sequential very different steps: allylic alcohol oxidation-Heck reaction. Access to functionalised alpha,beta-unsaturated ketones.

    Science.gov (United States)

    Batt, Frédéric; Gozzi, Christel; Fache, Fabienne

    2008-11-30

    A single addition of the catalyst, Pd(OAc)(2), was realised to mediate two transformations as different as allylic alcohol oxidation under O(2) and C-C bond formation of the Heck type, to give substituted alpha,beta-unsaturated ketones without intermediate purification.

  7. Direct Uniaxial Alignment of a Donor-Acceptor Semiconducting Polymer Using Single-Step Solution Shearing.

    Science.gov (United States)

    Shaw, Leo; Hayoz, Pascal; Diao, Ying; Reinspach, Julia Antonia; To, John W F; Toney, Michael F; Weitz, R Thomas; Bao, Zhenan

    2016-04-13

    The alignment of organic semiconductors (OSCs) in the active layers of electronic devices can confer desirable properties, such as enhanced charge transport properties due to better ordering, charge transport anisotropy for reduced device cross-talk, and polarized light emission or absorption. The solution-based deposition of highly aligned small molecule OSCs has been widely demonstrated, but the alignment of polymeric OSCs in thin films deposited directly from solution has typically required surface templating or complex pre- or postdeposition processing. Therefore, single-step solution processing and the charge transport enhancement afforded by alignment continue to be attractive. We report here the use of solution shearing to tune the degree of alignment in poly(diketopyrrolopyrrole-terthiophene) thin films by controlling the coating speed. A maximum dichroic ratio of ∼7 was achieved on unpatterned substrates without any additional pre- or postdeposition processing. The degree of polymer alignment was found to be a competition between the shear alignment of polymer chains in solution and the complex thin film drying process. Contrary to previous reports, no charge transport anisotropy was observed because of the small crystallite size relative to the channel length, a meshlike morphology, and the likelihood of increased grain boundaries in the direction transverse to coating. In fact, the lack of aligned morphological structures, coupled with observed anisotropy in X-ray diffraction data, suggests the alignment of polymer molecules in both the crystalline and the amorphous regions of the films. The shear speed at which maximum dichroism is achieved can be controlled by altering deposition parameters such as temperature and substrate treatment. Modest changes in molecular weight showed negligible effects on alignment, while longer polymer alkyl side chains were found to reduce the degree of alignment. This work demonstrates that solution shearing can be used

  8. Efficient and versatile one-step affinity purification of in vivo biotinylated proteins: Expression, characterization and structure analysis of recombinant human glutamate carboxypeptidase II

    Czech Academy of Sciences Publication Activity Database

    Tykvart, Jan; Šácha, Pavel; Bařinka, Cyril; Knedlík, Tomáš; Starková, Jana; Lubkowski, J.; Konvalinka, Jan

    2012-01-01

    Roč. 82, č. 1 (2012), s. 106-115 ISSN 1046-5928 R&D Projects: GA MŠk 1M0508; GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506; CEZ:AV0Z50520701 Keywords : affinity purification * biotin acceptor peptide * recombinant protein expression * biotin-protein ligase (BirA) * co-localization * PSMA Subject RIV: CE - Biochemistry Impact factor: 1.429, year: 2012

  9. Well-Defined Silica Supported Aluminum Hydride: Another Step Towards the Utopian Single Site Dream?

    KAUST Repository

    Werghi, Baraa

    2015-07-17

    Reaction of triisobutylaluminum with SBA15700 at room temperature occurs by two parallel pathways involving either silanol or siloxane bridges. It leads to the formation of a well-defined bipodal [(≡SiO)2Al-CH2CH(CH3)2] 1a, silicon isobutyl [≡Si-CH2CH(CH3)2] 1b and a silicon hydride [≡Si-H] 1c. Their structural identity was characterized by FT-IR and advance solid-state NMR spectroscopies (1H, 13C, 29Si, 27Al and 2D multiple quantum), elemental and gas phase analysis, and DFT calculations. The reaction involves the formation of a highly reactive monopodal intermediate: [≡SiO-Al-[CH2CH(CH3)2]2], with evolution of isobutane. This intermediate undergoes two parallel routes: Transfer of either one isobutyl fragment or of one hydride to an adjacent silicon atom. Both processes occur by opening of a strained siloxane bridge, ≡Si-O-Si≡ but with two different mechanisms, showing that the reality of “single site” catalyst may be an utopia: DFT calculations indicate that isobutyl transfer occurs via a simple metathesis between the Al-isobutyl and O-Si bonds, while hydride transfer occurs via a two steps mechanism, the first one is a ß-H elimination to Al with elimination of isobutene, whereas the second is a metathesis step between the formed Al-H bond and a O-Si bond. Thermal treatment of 1a (at 250 °C) under high vacuum (10-5 mbar) generates Al-H through a ß-H elimination of isobutyl fragment. These supported well-defined Al-H which are highly stable with time, are tetra, penta and octa coordinated as demonstrated by IR and 27Al–1H J-HMQC NMR spectroscopy. All these observations indicate that surfaces atoms around the site of grafting play a considerable role in the reactivity of a single site system.

  10. Well-defined silica supported aluminum hydride: another step towards the utopian single site dream?

    Science.gov (United States)

    Werghi, Baraa; Bendjeriou-Sedjerari, Anissa; Sofack-Kreutzer, Julien; Jedidi, Abdesslem; Abou-Hamad, Edy; Cavallo, Luigi; Basset, Jean-Marie

    2015-10-01

    processes occur by opening of a strained siloxane bridge, Si-O-Si but with two different mechanisms, showing that the reality of "single site" catalyst may be an utopia: DFT calculations indicate that isobutyl transfer occurs via a simple metathesis between the Al-isobutyl and O-Si bonds, while hydride transfer occurs via a two steps mechanism, the first one is a β-H elimination to Al with elimination of isobutene, whereas the second is a metathesis step between the formed Al-H bond and a O-Si bond. Thermal treatment of 1a (at 250 °C) under high vacuum (10 -5 mbar) generates Al-H through a β-H elimination of isobutyl fragment. These supported well-defined Al-H which are highly stable with time, are tetra, penta and octa coordinated as demonstrated by IR and 27 Al- 1 H J-HMQC NMR spectroscopy. All these observations indicate that surfaces atoms around the site of grafting play a considerable role in the reactivity of a single site system.

  11. Evaluating tamsulosin hydrochloride-released microparticles prepared using single-step matrix coating.

    Science.gov (United States)

    Maeda, Atsushi; Shinoda, Tatsuki; Ito, Naoki; Baba, Keizo; Oku, Naoto; Mizumoto, Takao

    2011-04-15

    The objective of the present study was to determine the optimum composition for sustained-release of tamsulosin hydrochloride from microparticles intended for orally disintegrating tablets. Microparticles were prepared from an aqueous ethylcellulose dispersion (Aquacoa®), and an aqueous copolymer based on ethyl acrylate and methyl methacrylate dispersion (Eudragit®) NE30D), with microcrystalline cellulose as core particles with a fluidized bed coating process. Prepared microparticles were about 200 μm diameter and spherical. The microparticles were evaluated for in vitro drug release and in vivo absorption to assess bioequivalence in a commercial product, Harnal® pellets. The optimum ratio of Aquacoat® and Eudragit® NE30D in the matrix was 9:1. We observed similar drug release profiles in microparticles and Harnal® pellets. Higuchi model analysis of the in vitro drug release from microparticles was linear up to 80% release, typical of Fickian diffusion sustained-release profile. The in vivo absorption properties from microparticles were comparable to Harnal® pellets, and there was a linear relationship between in vitro drug release and in vivo drug release. In conclusion, this development produces microparticles in single-step coating, that provided a sustained-release of tamsulosin hydrochloride comparable to Harnal® pellets. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Structural Studies of Silver Nanoparticles Obtained Through Single-Step Green Synthesis

    International Nuclear Information System (INIS)

    Peddi, Siva Prasad; Sadeh, Bilal Abdallah

    2015-01-01

    Green synthesis of silver Nanoparticles (AGNP's) has been the most prominent among the metallic nanoparticles for research for over a decade and half now due to both the simplicity of preparation and the applicability of biological species with extensive applications in medicine and biotechnology to reduce and trap the particles. The current article uses Eclipta Prostrata leaf extract as the biological species to cap the AGNP's through a single step process. The characterization data obtained was used for the analysis of the sample structure. The article emphasizes the disquisition of their shape and size of the lattice parameters and proposes a general scheme and a mathematical model for the analysis of their dependence. The data of the synthesized AGNP's has been used to advantage through the introduction of a structural shape factor for the crystalline nanoparticles. The properties of the structure of the AGNP's proposed and evaluated through a theoretical model was undeviating with the experimental consequences. This modus operandi gives scope for the structural studies of ultrafine particles prepared using biological methods. (paper)

  13. Synthetic lethality between gene defects affecting a single non-essential molecular pathway with reversible steps.

    Directory of Open Access Journals (Sweden)

    Andrei Zinovyev

    2013-04-01

    Full Text Available Systematic analysis of synthetic lethality (SL constitutes a critical tool for systems biology to decipher molecular pathways. The most accepted mechanistic explanation of SL is that the two genes function in parallel, mutually compensatory pathways, known as between-pathway SL. However, recent genome-wide analyses in yeast identified a significant number of within-pathway negative genetic interactions. The molecular mechanisms leading to within-pathway SL are not fully understood. Here, we propose a novel mechanism leading to within-pathway SL involving two genes functioning in a single non-essential pathway. This type of SL termed within-reversible-pathway SL involves reversible pathway steps, catalyzed by different enzymes in the forward and backward directions, and kinetic trapping of a potentially toxic intermediate. Experimental data with recombinational DNA repair genes validate the concept. Mathematical modeling recapitulates the possibility of kinetic trapping and revealed the potential contributions of synthetic, dosage-lethal interactions in such a genetic system as well as the possibility of within-pathway positive masking interactions. Analysis of yeast gene interaction and pathway data suggests broad applicability of this novel concept. These observations extend the canonical interpretation of synthetic-lethal or synthetic-sick interactions with direct implications to reconstruct molecular pathways and improve therapeutic approaches to diseases such as cancer.

  14. Single-step generation of fluorophore-encapsulated gold nanoparticle core-shell materials

    International Nuclear Information System (INIS)

    Sardar, R; Shem, P M; Pecchia-Bekkum, C; Bjorge, N S; Shumaker-Parry, J S

    2010-01-01

    We report a simple route to produce fluorophore-encapsulated gold nanoparticles (AuNPs) in a single step under aqueous conditions using the fluorophore 1-pyrenemethylamine (PMA). Different amounts of PMA were used and the resulting core-shell gold nanoparticles were analyzed using UV-visible absorption spectroscopy, fluorescence spectroscopy, and transmission and scanning electron microscopy. Electron microscopy analysis shows nanoparticles consisting of a gold nanoparticle core which is encapsulated with a lower contrast shell. In the UV-visible spectra, we observed a significant red shift (37 nm) of the localized surface plasmon resonance (LSPR) absorption maximum (λ max ) compared to citrate-stabilized AuNPs of a similar size. We attribute the prominent LSPR wavelength shift for PMA-AuNP conjugates to the increase in the local dielectric environment near the gold nanoparticles due to the shell formation. This simple, aqueous-based synthesis is a new approach to the production of fluorophore-encapsulated AuNPs that could be applicable in biological sensing systems and photonic device fabrication.

  15. Single-step gas phase synthesis of stable iron aluminide nanoparticles with soft magnetic properties

    Energy Technology Data Exchange (ETDEWEB)

    Vernieres, Jerome, E-mail: Jerome.vernieres@oist.jp; Benelmekki, Maria; Kim, Jeong-Hwan; Grammatikopoulos, Panagiotis; Diaz, Rosa E. [Nanoparticles by Design Unit, Okinawa Institute of Science and Technology (OIST) Graduate University, 1919-1 Tancha, Onna Son, Okinawa 904-0495 (Japan); Bobo, Jean-François [Centre d’Elaboration de Materiaux et d’Etudes Structurales (CEMES), 29 rue Jeanne Marvig, 31055 Toulouse Cedex 4 (France); Sowwan, Mukhles, E-mail: Mukhles@oist.jp [Nanoparticles by Design Unit, Okinawa Institute of Science and Technology (OIST) Graduate University, 1919-1 Tancha, Onna Son, Okinawa 904-0495 (Japan); Nanotechnology Research Laboratory, Al-Quds University, P.O. Box 51000, East Jerusalem, Palestine (Country Unknown)

    2014-11-01

    Soft magnetic alloys at the nanoscale level have long generated a vivid interest as candidate materials for technological and biomedical purposes. Consequently, controlling the structure of bimetallic nanoparticles in order to optimize their magnetic properties, such as high magnetization and low coercivity, can significantly boost their potential for related applications. However, traditional synthesis methods stumble upon the long standing challenge of developing true nanoalloys with effective control over morphology and stability against oxidation. Herein, we report on a single-step approach to the gas phase synthesis of soft magnetic bimetallic iron aluminide nanoparticles, using a versatile co-sputter inert gas condensation technique. This method allowed for precise morphological control of the particles; they consisted of an alloy iron aluminide crystalline core (DO{sub 3} phase) and an alumina shell, which reduced inter-particle interactions and also prevented further oxidation and segregation of the bimetallic core. Remarkably, the as-deposited alloy nanoparticles show interesting soft magnetic properties, in that they combine a high saturation magnetization (170 emu/g) and low coercivity (less than 20 Oe) at room temperature. Additional functionality is tenable by modifying the surface of the particles with a polymer, to ensure their good colloidal dispersion in aqueous environments.

  16. Pharmaceutical 3D printing: Design and qualification of a single step print and fill capsule.

    Science.gov (United States)

    Smith, Derrick M; Kapoor, Yash; Klinzing, Gerard R; Procopio, Adam T

    2018-03-29

    Fused deposition modeling (FDM) 3D printing (3DP) has a potential to change how we envision manufacturing in the pharmaceutical industry. A more common utilization for FDM 3DP is to build upon existing hot melt extrusion (HME) technology where the drug is dispersed in the polymer matrix. However, reliable manufacturing of drug-containing filaments remains a challenge along with the limitation of active ingredients which can sustain the processing risks involved in the HME process. To circumvent this obstacle, a single step FDM 3DP process was developed to manufacture thin-walled drug-free capsules which can be filled with dry or liquid drug product formulations. Drug release from these systems is governed by the combined dissolution of the FDM capsule 'shell' and the dosage form encapsulated in these shells. To prepare the shells, the 3D printer files (extension '.gcode') were modified by creating discrete zones, so-called 'zoning process', with individual print parameters. Capsules printed without the zoning process resulted in macroscopic print defects and holes. X-ray computed tomography, finite element analysis and mechanical testing were used to guide the zoning process and printing parameters in order to manufacture consistent and robust capsule shell geometries. Additionally, dose consistencies of drug containing liquid formulations were investigated in this work. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Single-step collision-free trajectory planning of biped climbing robots in spatial trusses.

    Science.gov (United States)

    Zhu, Haifei; Guan, Yisheng; Chen, Shengjun; Su, Manjia; Zhang, Hong

    For a biped climbing robot with dual grippers to climb poles, trusses or trees, feasible collision-free climbing motion is inevitable and essential. In this paper, we utilize the sampling-based algorithm, Bi-RRT, to plan single-step collision-free motion for biped climbing robots in spatial trusses. To deal with the orientation limit of a 5-DoF biped climbing robot, a new state representation along with corresponding operations including sampling, metric calculation and interpolation is presented. A simple but effective model of a biped climbing robot in trusses is proposed, through which the motion planning of one climbing cycle is transformed to that of a manipulator. In addition, the pre- and post-processes are introduced to expedite the convergence of the Bi-RRT algorithm and to ensure the safe motion of the climbing robot near poles as well. The piecewise linear paths are smoothed by utilizing cubic B-spline curve fitting. The effectiveness and efficiency of the presented Bi-RRT algorithm for climbing motion planning are verified by simulations.

  18. One-step separation and purification of three lignans and one flavonol from Sinopodophyllum emodi by medium-pressure liquid chromatography and high-speed counter-current chromatography.

    Science.gov (United States)

    Wang, Ping; Liu, Yongling; Chen, Tao; Xu, Wenhua; You, Jinmao; Liu, Yongjun; Li, Yulin

    2013-01-01

    Lignans and flavonols are the primary constituents of Sinopodophyllum emodi and have been used as cathartic, anthelmintic, chemotherapeutic and anti-hypertensive compounds. Although these compounds have been isolated, there have been no reports on the separation of 4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin and kaempferol in one step by medium-pressure liquid chromatography (MPLC) and high-speed counter-current chromatography (HSCCC). Development of an efficient method for the preparative separation and purification of three lignans and one flavonol from S. emodi. The precipitate of crude extracts was first separated by MPLC into four parts, numbered GJ-1, GJ-2, GJ-3 and GJ-4. GJ-1 was separated and purified by HSCCC using a solvent system composed of n-hexane:ethyl acetate:methanol:water (1.75:1.5:1:0.75, v/v/v/v). The purities of the target compounds were assessed using high-performance liquid chromatography (HPLC) and chemical structures were identified by (1) H-NMR and (13) C-NMR. The HSCCC and MPLC methods were successfully used for the preparative separation and purification of 4'-demethyl podophyllotoxin (8.5 mg, 92.4%), podophyllotoxin (40.1 mg, 92.1%), deoxypodophyllotoxin (4.6 mg, 98.1%), and kaempferol (1.6 mg, 96.7%) from a 100 mg sample. Three lignans (4'-demethyl podophyllotoxin, podophyllotoxin, deoxypodophyllotoxin) and one flavonol (kaempferol) were successfully isolated by HSCCC and MPLC in one step. Copyright © 2013 John Wiley & Sons, Ltd.

  19. Compatibility of pedigree-based and marker-based relationship matrices for single-step genetic evaluation

    DEFF Research Database (Denmark)

    Christensen, Ole Fredslund

    2012-01-01

    Single-step methods for genomic prediction have recently become popular because they are conceptually simple and in practice such a method can completely replace a pedigree-based method for routine genetic evaluation. An issue with single-step methods is compatibility between the marker...... that it may be important that a single-step method is based on a model conditional on the observed markers. When data are from routine evaluation systems, selection affects the allele frequencies, and therefore both observed markers and observed phenotypes contain information about allele frequencies...... the marker-based relationship matrix is constructed assuming all allele frequencies equal to 0.5 and the pedigree-based relationship matrix is constructed using the unusual assumption that animals in the base population are related and inbreed with relationship coefficient alpha and inbreeding coefficient...

  20. Single-step reinitialization and extending algorithms for level-set based multi-phase flow simulations

    Science.gov (United States)

    Fu, Lin; Hu, Xiangyu Y.; Adams, Nikolaus A.

    2017-12-01

    We propose efficient single-step formulations for reinitialization and extending algorithms, which are critical components of level-set based interface-tracking methods. The level-set field is reinitialized with a single-step (non iterative) "forward tracing" algorithm. A minimum set of cells is defined that describes the interface, and reinitialization employs only data from these cells. Fluid states are extrapolated or extended across the interface by a single-step "backward tracing" algorithm. Both algorithms, which are motivated by analogy to ray-tracing, avoid multiple block-boundary data exchanges that are inevitable for iterative reinitialization and extending approaches within a parallel-computing environment. The single-step algorithms are combined with a multi-resolution conservative sharp-interface method and validated by a wide range of benchmark test cases. We demonstrate that the proposed reinitialization method achieves second-order accuracy in conserving the volume of each phase. The interface location is invariant to reapplication of the single-step reinitialization. Generally, we observe smaller absolute errors than for standard iterative reinitialization on the same grid. The computational efficiency is higher than for the standard and typical high-order iterative reinitialization methods. We observe a 2- to 6-times efficiency improvement over the standard method for serial execution. The proposed single-step extending algorithm, which is commonly employed for assigning data to ghost cells with ghost-fluid or conservative interface interaction methods, shows about 10-times efficiency improvement over the standard method while maintaining same accuracy. Despite their simplicity, the proposed algorithms offer an efficient and robust alternative to iterative reinitialization and extending methods for level-set based multi-phase simulations.

  1. Purification and characterization of angiotensin-1 converting enzyme

    African Journals Online (AJOL)

    user

    2013-04-10

    Apr 10, 2013 ... al., 1999), meat (Jang and Lee, 2005) and marine resources (Je et al., ... species that have been blooming on the offshore areas of. Korea, China and ..... Lim et al. 1893. Table 1. Purification yield and ACE-inhibitory activity of each step. Fraction. Purification step. Purification yield (%). IC50 (μg/ml). W/H.

  2. Site-selective substitutional doping with atomic precision on stepped Al (111) surface by single-atom manipulation.

    Science.gov (United States)

    Chen, Chang; Zhang, Jinhu; Dong, Guofeng; Shao, Hezhu; Ning, Bo-Yuan; Zhao, Li; Ning, Xi-Jing; Zhuang, Jun

    2014-01-01

    In fabrication of nano- and quantum devices, it is sometimes critical to position individual dopants at certain sites precisely to obtain the specific or enhanced functionalities. With first-principles simulations, we propose a method for substitutional doping of individual atom at a certain position on a stepped metal surface by single-atom manipulation. A selected atom at the step of Al (111) surface could be extracted vertically with an Al trimer-apex tip, and then the dopant atom will be positioned to this site. The details of the entire process including potential energy curves are given, which suggests the reliability of the proposed single-atom doping method.

  3. Development of single step RT-PCR for detection of Kyasanur forest disease virus from clinical samples

    Directory of Open Access Journals (Sweden)

    Gouri Chaubal

    2018-02-01

    Discussion and conclusion: The previously published sensitive real time RT-PCR assay requires higher cost in terms of reagents and machine setup and technical expertise has been the primary reason for development of this assay. A single step RT-PCR is relatively easy to perform and more cost effective than real time RT-PCR in smaller setups in the absence of Biosafety Level-3 facility. This study reports the development and optimization of single step RT-PCR assay which is more sensitive and less time-consuming than nested RT-PCR and cost effective for rapid diagnosis of KFD viral RNA.

  4. Single-step brazing process for mono-block joints and mechanical testing

    International Nuclear Information System (INIS)

    Casalegno, V.; Ferraris, M.; Salvo, M.; Rizzo, S.; Merola, M.

    2007-01-01

    Full text of publication follows: Plasma facing components act as actively cooled thermal shields to sustain thermal and particle loads during normal and transient operations in ITER (International Thermonuclear Experimental Reactor). The plasma-facing layer is referred to as 'armour', which is made of either carbon fibre reinforced carbon composite (CFC) or tungsten (W). CFC is the reference design solution for the lower part of the vertical target of the ITER divertor. The armour is joined onto an actively cooled substrate, the heat sink, made of precipitation hardened copper alloy CuCrZr through a thin pure copper interlayer to decrease, by plastic deformation, the joint interface stresses; in fact, the CFC to Cu joint is affected by the CTE mismatch between the ceramic and metallic material. A new method of joining CFC to copper and CFC/Cu to CuCrZr alloy was effectively developed for the flat-type configuration; the feasibility of this process also for mono-block geometry and the development of a procedure for testing mono-block-type mock-ups is described in this work. The mono-block configuration consists of copper alloy pipe shielded by CFC blocks. It is worth noting that in mono-block configuration, the large thermal expansion mismatch between CFC and copper alloy is more significant than for flat-tile configuration, due to curved interfaces. The joining technique foresees a single-step brazing process: the brazing of the three materials (CFC-Cu-CuCrZr) can be performed in a single heat treatment using the same Cu/Ge based braze. The composite surface was modified by solid state reaction with chromium with the purpose of increasing the wettability of CFC by the brazing alloy. The CFC substrate reacts with Cr which, forming a carbide layer, allows a large reduction of the contact angle; then, the brazing of CFC to pure copper and pure copper to CuCrZr by the same treatment is feasible. This process allows to obtain good joints using a non-active brazing

  5. Single-Step BLUP with Varying Genotyping Effort in Open-Pollinated Picea glauca

    Directory of Open Access Journals (Sweden)

    Blaise Ratcliffe

    2017-03-01

    Full Text Available Maximization of genetic gain in forest tree breeding programs is contingent on the accuracy of the predicted breeding values and precision of the estimated genetic parameters. We investigated the effect of the combined use of contemporary pedigree information and genomic relatedness estimates on the accuracy of predicted breeding values and precision of estimated genetic parameters, as well as rankings of selection candidates, using single-step genomic evaluation (HBLUP. In this study, two traits with diverse heritabilities [tree height (HT and wood density (WD] were assessed at various levels of family genotyping efforts (0, 25, 50, 75, and 100% from a population of white spruce (Picea glauca consisting of 1694 trees from 214 open-pollinated families, representing 43 provenances in Québec, Canada. The results revealed that HBLUP bivariate analysis is effective in reducing the known bias in heritability estimates of open-pollinated populations, as it exposes hidden relatedness, potential pedigree errors, and inbreeding. The addition of genomic information in the analysis considerably improved the accuracy in breeding value estimates by accounting for both Mendelian sampling and historical coancestry that were not captured by the contemporary pedigree alone. Increasing family genotyping efforts were associated with continuous improvement in model fit, precision of genetic parameters, and breeding value accuracy. Yet, improvements were observed even at minimal genotyping effort, indicating that even modest genotyping effort is effective in improving genetic evaluation. The combined utilization of both pedigree and genomic information may be a cost-effective approach to increase the accuracy of breeding values in forest tree breeding programs where shallow pedigrees and large testing populations are the norm.

  6. Single-step production of a recyclable nanobiocatalyst for organophosphate pesticides biodegradation using functionalized bacterial magnetosomes.

    Directory of Open Access Journals (Sweden)

    Nicolas Ginet

    Full Text Available Enzymes are versatile catalysts in laboratories and on an industrial scale; improving their immobilization would be beneficial to broadening their applicability and ensuring their (reuse. Lipid-coated nano-magnets produced by magnetotactic bacteria are suitable for a universally applicable single-step method of enzyme immobilization. By genetically functionalizing the membrane surrounding these magnetite particles with a phosphohydrolase, we engineered an easy-to-purify, robust and recyclable biocatalyst to degrade ethyl-paraoxon, a commonly used pesticide. For this, we genetically fused the opd gene from Flavobacterium sp. ATCC 27551 encoding a paraoxonase to mamC, an abundant protein of the magnetosome membrane in Magnetospirillum magneticum AMB-1. The MamC protein acts as an anchor for the paraoxonase to the magnetosome surface, thus producing magnetic nanoparticles displaying phosphohydrolase activity. Magnetosomes functionalized with Opd were easily recovered from genetically modified AMB-1 cells: after cellular disruption with a French press, the magnetic nanoparticles are purified using a commercially available magnetic separation system. The catalytic properties of the immobilized Opd were measured on ethyl-paraoxon hydrolysis: they are comparable with the purified enzyme, with K(m (and k(cat values of 58 µM (and 178 s(-1 and 43 µM (and 314 s(-1 for the immobilized and purified enzyme respectively. The Opd, a metalloenzyme requiring a zinc cofactor, is thus properly matured in AMB-1. The recycling of the functionalized magnetosomes was investigated and their catalytic activity proved to be stable over repeated use for pesticide degradation. In this study, we demonstrate the easy production of functionalized magnetic nanoparticles with suitably genetically modified magnetotactic bacteria that are efficient as a reusable nanobiocatalyst for pesticides bioremediation in contaminated effluents.

  7. Single-step isolation of extracellular vesicles by size-exclusion chromatography

    Science.gov (United States)

    Böing, Anita N.; van der Pol, Edwin; Grootemaat, Anita E.; Coumans, Frank A. W.; Sturk, Auguste; Nieuwland, Rienk

    2014-01-01

    Background Isolation of extracellular vesicles from plasma is a challenge due to the presence of proteins and lipoproteins. Isolation of vesicles using differential centrifugation or density-gradient ultracentrifugation results in co-isolation of contaminants such as protein aggregates and incomplete separation of vesicles from lipoproteins, respectively. Aim To develop a single-step protocol to isolate vesicles from human body fluids. Methods Platelet-free supernatant, derived from platelet concentrates, was loaded on a sepharose CL-2B column to perform size-exclusion chromatography (SEC; n=3). Fractions were collected and analysed by nanoparticle tracking analysis, resistive pulse sensing, flow cytometry and transmission electron microscopy. The concentrations of high-density lipoprotein cholesterol (HDL) and protein were measured in each fraction. Results Fractions 9–12 contained the highest concentrations of particles larger than 70 nm and platelet-derived vesicles (46%±6 and 61%±2 of totals present in all collected fractions, respectively), but less than 5% of HDL and less than 1% of protein (4.8%±1 and 0.65%±0.3, respectively). HDL was present mainly in fractions 18–20 (32%±2 of total), and protein in fractions 19–21 (36%±2 of total). Compared to the starting material, recovery of platelet-derived vesicles was 43%±23 in fractions 9–12, with an 8-fold and 70-fold enrichment compared to HDL and protein. Conclusions SEC efficiently isolates extracellular vesicles with a diameter larger than 70 nm from platelet-free supernatant of platelet concentrates. Application SEC will improve studies on the dimensional, structural and functional properties of extracellular vesicles. PMID:25279113

  8. Single-step isolation of extracellular vesicles by size-exclusion chromatography.

    Science.gov (United States)

    Böing, Anita N; van der Pol, Edwin; Grootemaat, Anita E; Coumans, Frank A W; Sturk, Auguste; Nieuwland, Rienk

    2014-01-01

    Isolation of extracellular vesicles from plasma is a challenge due to the presence of proteins and lipoproteins. Isolation of vesicles using differential centrifugation or density-gradient ultracentrifugation results in co-isolation of contaminants such as protein aggregates and incomplete separation of vesicles from lipoproteins, respectively. To develop a single-step protocol to isolate vesicles from human body fluids. Platelet-free supernatant, derived from platelet concentrates, was loaded on a sepharose CL-2B column to perform size-exclusion chromatography (SEC; n=3). Fractions were collected and analysed by nanoparticle tracking analysis, resistive pulse sensing, flow cytometry and transmission electron microscopy. The concentrations of high-density lipoprotein cholesterol (HDL) and protein were measured in each fraction. Fractions 9-12 contained the highest concentrations of particles larger than 70 nm and platelet-derived vesicles (46%±6 and 61%±2 of totals present in all collected fractions, respectively), but less than 5% of HDL and less than 1% of protein (4.8%±1 and 0.65%±0.3, respectively). HDL was present mainly in fractions 18-20 (32%±2 of total), and protein in fractions 19-21 (36%±2 of total). Compared to the starting material, recovery of platelet-derived vesicles was 43%±23 in fractions 9-12, with an 8-fold and 70-fold enrichment compared to HDL and protein. SEC efficiently isolates extracellular vesicles with a diameter larger than 70 nm from platelet-free supernatant of platelet concentrates. Application SEC will improve studies on the dimensional, structural and functional properties of extracellular vesicles.

  9. Single-step blood direct PCR: A robust and rapid method to diagnose triplet repeat disorders.

    Science.gov (United States)

    Singh, Inder; Swarup, Vishnu; Shakya, Sunil; Goyal, Vinay; Faruq, Mohammed; Srivastava, Achal Kumar

    2017-08-15

    DNA extraction prior to polymerase chain reaction (PCR) amplification in genetic diagnoses of triplet repeat disorders (TRDs) is tedious and labour-intensive and has the limitations of sample contamination with foreign DNA, including that from preceding samples. Therefore, we aimed to develop a rapid, robust, and cost-effective method for expeditious genetic investigation of TRDs from whole blood as a DNA template. Peripheral blood samples were collected from 70 clinically suspected patients of progressive ataxia. The conventional method using genomic DNA and single-step Blood-Direct PCR (BD-PCR) method with just 2μl of whole blood sample were tested to amplify triplet repeat expansion in genes related to spinocerebellar ataxia (SCA) types 1, 2, 3, 12 and Friedreich's ataxia (FRDA). Post-PCR, the allele sizes were mapped and repeat numbers were calculated using GeneMapper and macros run in Microsoft Excel programmes. Successful amplification of target regions was achieved in all samples by both methods. The frequency of the normal and mutated allele was concordant between both methods, diagnosing 37% positive for a mutation in either of the candidate genes. The BD-PCR resulted in higher intensities of product peaks of normal and pathogenic alleles. The nearly-accurate sizing of the normal and expanded allele was achieved in a shorter time (4-5h), without DNA extraction and any risk of cross contamination, which suggests the BD-PCR to be a reliable, inexpensive, and rapid method to confirm TRDs. This technique can be introduced in routine diagnostic procedures of other tandem repeat disorders. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. An efficient single-step scheme for manipulating quantum information of two trapped ions beyond the Lamb-Dicke limit

    International Nuclear Information System (INIS)

    Wei, L.F.; Nori, Franco

    2003-01-01

    Based on the exact conditional quantum dynamics for a two-ion system, we propose an efficient single-step scheme for coherently manipulating quantum information of two trapped cold ions by using a pair of synchronous laser pulses. Neither the auxiliary atomic level nor the Lamb-Dicke approximation are needed

  11. Mechanical stability of a locked step-plate versus single compression screw fixation for medial displacement calcaneal osteotomy.

    Science.gov (United States)

    Konan, Sujith; Meswania, Jay; Blunn, Gordon W; Madhav, Rohit T; Oddy, Michael J

    2012-08-01

    Reconstruction of a flatfoot commonly involves a calcaneal Medial Displacement Osteotomy (MDO) to correct hindfoot valgus in combination with soft tissue procedures. We compared fixation of an MDO using either a single, large cannulated screw versus a locking step-plate in load to failure in a cadaveric model. Eight matched pairs of cadaveric limbs were loaded using a mechanical testing rig. Two pairs served as non-operated controls. The remaining paired limbs underwent a 10-mm MDO stabilized either with a single 7-mm screw or a step-plate with four locking screws. One pair was used as a pilot study and the remaining five pairs were loaded up to 4500 N to failure. In the five pairs loaded to failure, the median (with 95% CI) maximum force were 1779 N (1099-2312) and 826 N (288-1607) for the plate and screw, respectively (p = 0.043). With single screw fixation, the tuberosity fragment consistently failed by rotation and angulation into varus. With plate fixation, failure occurred as the screws cut through the internal surfaces of the tuberosity and body with no failure at the screw-plate interface. In this cadaveric model, a locked step-plate supported a significantly higher maximum force than a single large cannulated screw. The magnitude of the load supported by the locking step-plate suggests that allowing early weightbearing post-operation may be safe in clinical practice before union of the osteotomy.

  12. Single mode step-index polymer optical fiber for humidity insensitive high temperature fiber Bragg grating sensors

    DEFF Research Database (Denmark)

    Woyessa, Getinet; Fasano, Andrea; Stefani, Alessio

    2016-01-01

    We have fabricated the first single-mode step-index and humidity insensitive polymer optical fiber operating in the 850 nm wavelength ranges. The step-index preform is fabricated using injection molding, which is an efficient method for cost effective, flexible and fast preparation of the fiber...... preform. The fabricated single-mode step-index (SI) polymer optical fiber (POF) has a 4.8µm core made from TOPAS grade 5013S-04 with a glass transition temperature of 134°C and a 150 µm cladding made from ZEONEX grade 480R with a glass transition temperature of 138°C. The key advantages of the proposed...... SIPOF are low water absorption, high operating temperature and chemical inertness to acids and bases and many polar solvents as compared to the conventional poly-methyl-methacrylate (PMMA) and polystyrene based POFs. In addition, the fiber Bragg grating writing time is short compared to microstructured...

  13. Single mode step-index polymer optical fiber for humidity insensitive high temperature fiber Bragg grating sensors.

    Science.gov (United States)

    Woyessa, Getinet; Fasano, Andrea; Stefani, Alessio; Markos, Christos; Nielsen, Kristian; Rasmussen, Henrik K; Bang, Ole

    2016-01-25

    We have fabricated the first single-mode step-index and humidity insensitive polymer optical fiber operating in the 850 nm wavelength ranges. The step-index preform is fabricated using injection molding, which is an efficient method for cost effective, flexible and fast preparation of the fiber preform. The fabricated single-mode step-index (SI) polymer optical fiber (POF) has a 4.8µm core made from TOPAS grade 5013S-04 with a glass transition temperature of 134°C and a 150 µm cladding made from ZEONEX grade 480R with a glass transition temperature of 138°C. The key advantages of the proposed SIPOF are low water absorption, high operating temperature and chemical inertness to acids and bases and many polar solvents as compared to the conventional poly-methyl-methacrylate (PMMA) and polystyrene based POFs. In addition, the fiber Bragg grating writing time is short compared to microstructured POFs.

  14. Application of single step genomic BLUP under different uncertain paternity scenarios using simulated data.

    Directory of Open Access Journals (Sweden)

    Rafael Lara Tonussi

    Full Text Available The objective of this study was to investigate the application of BLUP and single step genomic BLUP (ssGBLUP models in different scenarios of paternity uncertainty with different strategies of scaling the G matrix to match the A22 matrix, using simulated data for beef cattle. Genotypes, pedigree, and phenotypes for age at first calving (AFC and weight at 550 days (W550 were simulated using heritabilities based on real data (0.12 for AFC and 0.34 for W550. Paternity uncertainty scenarios using 0, 25, 50, 75, and 100% of multiple sires (MS were studied. The simulated genome had a total length of 2,333 cM, containing 735,293 biallelic markers and 7,000 QTLs randomly distributed over the 29 BTA. It was assumed that QTLs explained 100% of the genetic variance. For QTL, the amount of alleles per loci randomly ranged from two to four. The BLUP model that considers phenotypic and pedigree data, and the ssGBLUP model that combines phenotypic, pedigree and genomic information were used for genetic evaluations. Four ways of scaling the mean of the genomic matrix (G to match to the mean of the pedigree relationship matrix among genotyped animals (A22 were tested. Accuracy, bias, and inflation were investigated for five groups of animals: ALL = all animals; BULL = only bulls; GEN = genotyped animals; FEM = females; and YOUNG = young males. With the BLUP model, the accuracies of genetic evaluations decreased for both traits as the proportion of unknown sires in the population increased. The EBV accuracy reduction was higher for GEN and YOUNG groups. By analyzing the scenarios for YOUNG (from 0 to 100% of MS, the decrease was 87.8 and 86% for AFC and W550, respectively. When applying the ssGBLUP model, the accuracies of genetic evaluation also decreased as the MS in the pedigree for both traits increased. However, the accuracy reduction was less than those observed for BLUP model. Using the same comparison (scenario 0 to 100% of MS, the accuracies reductions

  15. One-step formation of a single atomic-layer transistor by the selective fluorination of a graphene film.

    Science.gov (United States)

    Ho, Kuan-I; Liao, Jia-Hong; Huang, Chi-Hsien; Hsu, Chang-Lung; Zhang, Wenjing; Lu, Ang-Yu; Li, Lain-Jong; Lai, Chao-Sung; Su, Ching-Yuan

    2014-03-12

    In this study, the scalable and one-step fabrication of single atomic-layer transistors is demonstrated by the selective fluorination of graphene using a low-damage CF4 plasma treatment, where the generated F-radicals preferentially fluorinated the graphene at low temperature (semiconductor/insulator can be directly formed in a single layer of graphene using a one-step fluorination process by introducing a Au thin-film as a buffer layer. With this heterojunction structure, it would be possible to fabricate transistors in a single graphene film via a one-step fluorination process, in which pristine graphene, partial F-graphene, and highly F-graphene serve as the source/drain contacts, the channel, and the channel isolation in a transistor, respectively. The demonstrated graphene transistor exhibits an on-off ratio above 10, which is 3-fold higher than that of devices made from pristine graphene. This efficient transistor fabrication method produces electrical heterojunctions of graphene over a large area and with selective patterning, providing the potential for the integration of electronics down to the single atomic-layer scale. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. High-Level Soluble Expression and One-step Purification of HTLV-I P19 Protein in Escherichia coli by Fusion Expression.

    Science.gov (United States)

    Mosadeghi, Parvin; Zarnagh, Hafez Heydari; Mohammad-Zadeh, Mohammad; Salehi Moghaddam, Masoud

    2015-12-01

    Expression of HTLV-I p19 protein in an Escherichia coli expression system always leads to the formation of inclusion body. Solubilisation and refolding of the inclusion bodies is complex, time consuming and difficult during large-scale preparation. This study aimed to express and purify a soluble form of recombinant HTLV-I p19 protein in an E. coli expression system. The synthetic DNA encoding the p19 was subcloned into a pGS21a vector along with a His-GST solubility/purification tag. The recombinant pGS21a-p19 vector was then transformed into chemically competent E. coli BL21 (DE3) cells, and expression of the recombinant His-GST-p19 protein was induced by IPTG. Expression and distribution of the His-GST-p19 protein in soluble and insoluble fractions were evaluated using SDS-PAGE. Antigenicity of the His-GST-p19 protein was evaluated using ELISA after purifying the protein using Ni-NTA affinity chromatography, then compared to the results of synthetic immunodominant p19 peptide ELISA. The fusion His-GST-p19 protein accounted for 30% of the total cellular proteins. The SDS-PAGE results indicated that approximately 50% of the expressed His-GST-p19 proteins were soluble and accounted for 50% of the total soluble proteins. ELISA showed that the His-GST tag did not impair the antigenicity of the p19 protein and that the fusion protein reacted with HTLV-I antibodies in a concentration-dependent manner. The results of His-GST-p19 ELISA indicated that specificity of p19 reactivity was compatible to the results of p19 peptide ELISA. Combination of key strategies for the soluble expresion of proteins, like fusion with solubility/purification tags, low IPTG concentration and induction at low temperature, provide an efficient and facile platform for producing soluble  HTLV-I p19 protein.

  17. Isolation, one-step affinity purification, and characterization of a polyextremotolerant laccase from the halophilic bacterium Aquisalibacillus elongatus and its application in the delignification of sugar beet pulp.

    Science.gov (United States)

    Rezaei, Shahla; Shahverdi, Ahmad Reza; Faramarzi, Mohammad Ali

    2017-04-01

    The aim of the present work was to study the ability of a halophilic bacterial laccase to efficient delignification in extreme conditions. Here, a highly stable extracellular laccase showing ligninolytic activity from halophilic Aquisalibacillus elongatus is described. The laccase production was strongly influenced by NaCl and CuSO 4 and under optimal conditions reached 4.8UmL -1 . The monomeric enzyme of 75kDa was purified by a synthetic affinity column with 68.2% yield and 99.8-fold purification. The enzyme showed some valuable features viz. stability against a wide range of organic solvents, salts, metals, inhibitors, and surfactants and specificity to a wide spectrum of substrates diverse in structure and redox potential. It retained more than 50% of the original activity at 25-75°C and pH 5.0-10.0. Furthermore, the enzyme was found to be effective in the delignification of sugar beet pulp in an ionic liquid that makes it useful for industrial applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Colloidal Quantum Dot Inks for Single-Step-Fabricated Field-Effect Transistors: The Importance of Postdeposition Ligand Removal.

    Science.gov (United States)

    Balazs, Daniel M; Rizkia, Nisrina; Fang, Hong-Hua; Dirin, Dmitry N; Momand, Jamo; Kooi, Bart J; Kovalenko, Maksym V; Loi, Maria Antonietta

    2018-02-14

    Colloidal quantum dots are a class of solution-processed semiconductors with good prospects for photovoltaic and optoelectronic applications. Removal of the surfactant, so-called ligand exchange, is a crucial step in making the solid films conductive, but performing it in solid state introduces surface defects and cracks in the films. Hence, the formation of thick, device-grade films have only been possible through layer-by-layer processing, limiting the technological interest for quantum dot solids. Solution-phase ligand exchange before the deposition allows for the direct deposition of thick, homogeneous films suitable for device applications. In this work, fabrication of field-effect transistors in a single step is reported using blade-coating, an upscalable, industrially relevant technique. Most importantly, a postdeposition washing step results in device properties comparable to the best layer-by-layer processed devices, opening the way for large-scale fabrication and further interest from the research community.

  19. A permeation theory for single-file ion channels: one- and two-step models.

    Science.gov (United States)

    Nelson, Peter Hugo

    2011-04-28

    How many steps are required to model permeation through ion channels? This question is investigated by comparing one- and two-step models of permeation with experiment and MD simulation for the first time. In recent MD simulations, the observed permeation mechanism was identified as resembling a Hodgkin and Keynes knock-on mechanism with one voltage-dependent rate-determining step [Jensen et al., PNAS 107, 5833 (2010)]. These previously published simulation data are fitted to a one-step knock-on model that successfully explains the highly non-Ohmic current-voltage curve observed in the simulation. However, these predictions (and the simulations upon which they are based) are not representative of real channel behavior, which is typically Ohmic at low voltages. A two-step association/dissociation (A/D) model is then compared with experiment for the first time. This two-parameter model is shown to be remarkably consistent with previously published permeation experiments through the MaxiK potassium channel over a wide range of concentrations and positive voltages. The A/D model also provides a first-order explanation of permeation through the Shaker potassium channel, but it does not explain the asymmetry observed experimentally. To address this, a new asymmetric variant of the A/D model is developed using the present theoretical framework. It includes a third parameter that represents the value of the "permeation coordinate" (fractional electric potential energy) corresponding to the triply occupied state n of the channel. This asymmetric A/D model is fitted to published permeation data through the Shaker potassium channel at physiological concentrations, and it successfully predicts qualitative changes in the negative current-voltage data (including a transition to super-Ohmic behavior) based solely on a fit to positive-voltage data (that appear linear). The A/D model appears to be qualitatively consistent with a large group of published MD simulations, but no

  20. Balanced Photodetection in One-Step Liquid-Phase-Synthesized CsPbBr3 Micro-/Nanoflake Single Crystals.

    Science.gov (United States)

    Zheng, Wei; Xiong, Xufan; Lin, Richeng; Zhang, Zhaojun; Xu, Cunhua; Huang, Feng

    2018-01-17

    Here, we reported a low-cost and high-compatibility one-step liquid-phase synthesis method for synthesizing high-purity CsPbBr 3 micro-/nanoflake single crystals. On the basis of the high-purity CsPbBr 3 , we further prepared a low-dimensional photodetector capable of balanced photodetection, involving both high external quantum efficiency and rapid temporal response, which is barely realized in previously reported low-dimensional photodetectors.

  1. Development and evaluation of one step single tube multiplex RT-PCR for rapid detection and typing of dengue viruses

    OpenAIRE

    Parida Manmohan; Shrivastava Ambuj; Santhosh SR; Dash Paban; Saxena Parag; Rao PV

    2008-01-01

    Abstract Background Dengue is emerging as a major public health concern in many parts of the world. The development of a one-step, single tube, rapid, and multiplex reverse transcription polymerase chain reaction (M-RT-PCR) for simultaneous detection and typing of dengue virus using serotype specific primers during acute phase of illness is reported. Results An optimal assay condition with zero background was established having no cross-reaction with closely related members of flavivirus (Jap...

  2. A simple two step procedure for purification of the catalytic domain of chicken tryptophan hydroxylase 1 in a form suitable for crystallization

    DEFF Research Database (Denmark)

    Windahl, Michael Skovbo; Petersen, Charlotte R.; Munch, Astrid

    2008-01-01

    Tryptophan hydroxylase (TPH) [EC 1.14.16.4] catalyzes the conversion of tryptophan to 5-hydroxytryptophan, which is the first and rate-determining step in the biosynthesis of the neurotransmitter serotonin. We have expressed the catalytic domain of chicken (Gallus gallus) TPH isoform 1 in Escheri......Tryptophan hydroxylase (TPH) [EC 1.14.16.4] catalyzes the conversion of tryptophan to 5-hydroxytryptophan, which is the first and rate-determining step in the biosynthesis of the neurotransmitter serotonin. We have expressed the catalytic domain of chicken (Gallus gallus) TPH isoform 1...

  3. Single particle analysis of thylakoid proteins from Thermosynechococcus elongatus and Synechocystis 6803 : Localization of the CupA subunit of NDH-1

    NARCIS (Netherlands)

    Folea, I. Mihaela; Zhang, Pengpeng; Nowaczyk, Marc M.; Ogawa, Teruo; Aro, Eva-Marl; Boekema, Egbert J.; Aro, Eva-Mari

    The larger protein complexes of the cyanobacterial photosynthetic membrane of Thermosynechoccus elongatus and Synechocystis 6803 were studied by single particle electron microscopy after detergent solubilization, without any purification steps. Besides the "standard" L-shaped NDH-1L complex, related

  4. Hamiltonian purification

    Energy Technology Data Exchange (ETDEWEB)

    Orsucci, Davide [Scuola Normale Superiore, I-56126 Pisa (Italy); Burgarth, Daniel [Department of Mathematics, Aberystwyth University, Aberystwyth SY23 3BZ (United Kingdom); Facchi, Paolo; Pascazio, Saverio [Dipartimento di Fisica and MECENAS, Università di Bari, I-70126 Bari (Italy); INFN, Sezione di Bari, I-70126 Bari (Italy); Nakazato, Hiromichi; Yuasa, Kazuya [Department of Physics, Waseda University, Tokyo 169-8555 (Japan); Giovannetti, Vittorio [NEST, Scuola Normale Superiore and Istituto Nanoscienze-CNR, I-56126 Pisa (Italy)

    2015-12-15

    The problem of Hamiltonian purification introduced by Burgarth et al. [Nat. Commun. 5, 5173 (2014)] is formalized and discussed. Specifically, given a set of non-commuting Hamiltonians (h{sub 1}, …, h{sub m}) operating on a d-dimensional quantum system ℋ{sub d}, the problem consists in identifying a set of commuting Hamiltonians (H{sub 1}, …, H{sub m}) operating on a larger d{sub E}-dimensional system ℋ{sub d{sub E}} which embeds ℋ{sub d} as a proper subspace, such that h{sub j} = PH{sub j}P with P being the projection which allows one to recover ℋ{sub d} from ℋ{sub d{sub E}}. The notions of spanning-set purification and generator purification of an algebra are also introduced and optimal solutions for u(d) are provided.

  5. Blastocyst utilization rates after continuous culture in two commercial single-step media: a prospective randomized study with sibling oocytes.

    Science.gov (United States)

    Sfontouris, Ioannis A; Kolibianakis, Efstratios M; Lainas, George T; Venetis, Christos A; Petsas, George K; Tarlatzis, Basil C; Lainas, Tryfon G

    2017-10-01

    The aim of this study is to determine whether blastocyst utilization rates are different after continuous culture in two different commercial single-step media. This is a paired randomized controlled trial with sibling oocytes conducted in infertility patients, aged ≤40 years with ≥10 oocytes retrieved assigned to blastocyst culture and transfer. Retrieved oocytes were randomly allocated to continuous culture in either Sage one-step medium (Origio) or Continuous Single Culture (CSC) medium (Irvine Scientific) without medium renewal up to day 5 post oocyte retrieval. Main outcome measure was the proportion of embryos suitable for clinical use (utilization rate). A total of 502 oocytes from 33 women were randomly allocated to continuous culture in either Sage one-step medium (n = 250) or CSC medium (n = 252). Fertilization was performed by either in vitro fertilization or intracytoplasmic sperm injection, and embryo transfers were performed on day 5. Two patients had all blastocysts frozen due to the occurrence of severe ovarian hyperstimulation syndrome. Fertilization and cleavage rates, as well as embryo quality on day 3, were similar in the two media. Blastocyst utilization rates (%, 95% CI) [55.4% (46.4-64.1) vs 54.7% (44.9-64.6), p = 0.717], blastocyst formation rates [53.6% (44.6-62.5) vs 51.9 (42.2-61.6), p = 0.755], and proportion of good quality blastocysts [36.8% (28.1-45.4) vs 36.1% (27.2-45.0), p = 0.850] were similar in Sage one-step and CSC media, respectively. Continuous culture of embryos in Sage one-step and CSC media is associated with similar blastocyst development and utilization rates. Both single-step media appear to provide adequate support during in vitro preimplantation embryo development. Whether these observations are also valid for other continuous single medium protocols remains to be determined. NCT02302638.

  6. Determination of oil reservoir radiotracer (S{sup 14}CN{sup -}) in a single step using a plastic scintillator extractive resin

    Energy Technology Data Exchange (ETDEWEB)

    Bagan, H.; Tarancon, A. [Departament de Quimica Analitica, Universitat de Barcelona, Diagonal 645, E-08028 Barcelona (Spain); Stavsetra, L. [Department for Reservoir and Exploration Technology, Institute for Energy Technology (IFE), Instituttveien 18, N-2027 Kjeller (Norway); Rauret, G. [Departament de Quimica Analitica, Universitat de Barcelona, Diagonal 645, E-08028 Barcelona (Spain); Garcia, J.F., E-mail: jfgarcia@ub.edu [Departament de Quimica Analitica, Universitat de Barcelona, Diagonal 645, E-08028 Barcelona (Spain)

    2012-07-29

    Highlights: Black-Right-Pointing-Pointer A new procedure for S{sup 14}CN{sup -} radiotracer determination using PS resin was established. Black-Right-Pointing-Pointer The minimum detectable activity for a 100 mL sample is 0.08 Bq L{sup -1}. Black-Right-Pointing-Pointer The minimum quantifiable activity for a 100 mL sample is 0.31 Bq L{sup -1}. Black-Right-Pointing-Pointer PS resin is capable to quantify S{sup 14}CN{sup -} radiotracer samples with errors lower than 5%. Black-Right-Pointing-Pointer PS resin is also capable to quantify complex matrices obtained from oil reservoirs. - Abstract: The analysis of radiotracers is important in the study of oil reservoir dynamics. One of the most widely used radiotracer is S{sup 14}CN{sup -}. Prior to activity measurements by Liquid Scintillation (LS), routine determinations require the pretreatment steps of purification and concentration of the samples using anion exchange columns. The final elution media produces samples with high salt concentration that may lead to problems with phase separation during the LS measurement. Plastic Scintillation (PS) is an alternative technique that provides a solid surface that can be used as a platform for the immobilisation of selective extractants to obtain a PS resin. The proposed procedure unifies chemical separation and sample measurement preparation in a single step, serving to reduce the number of reagents needed and manpower required for the analysis while also avoiding mixed waste production by LS. The objective of this study is to develop a PS resin for the determination of {sup 14}C-labelled thiocyanate radiotracer in water samples. For this purpose, the immobilisation procedure was optimised, including optimisation of the proportion of PS microspheres:extractant and the use of a control blank to monitor the PS resin immobilisation process. The breakthrough volume was studied and the detection and quantification limits for 100 mL of sample were determined to be 0.08 Bq L{sup -1

  7. Engineering a recyclable elastin-like polypeptide capturing scaffold for non-chromatographic protein purification.

    Science.gov (United States)

    Liu, Fang; Chen, Wilfred

    2013-01-01

    Previously, we reported a non-chromatographic protein purification method exploiting the highly specific interaction between the dockerin and cohesin domains from Clostridium thermocellum and the reversible aggregation property of elastin-like polypeptide (ELP) to provide fast and cost-effective protein purification. However, the bound dockerin-intein tag cannot be completely dissociated from the ELP-cohesin capturing scaffold due to the high binding affinity, resulting in a single-use approach. In order to further reduce the purification cost by recycling the ELP capturing scaffold, a truncated dockerin domain with the calcium-coordinating function partially impaired was employed. We demonstrated that the truncated dockerin domain was sufficient to function as an effective affinity tag, and the target protein was purified directly from cell extracts in a single binding step followed by intein cleavage. The efficient EDTA-mediated dissociation of the bound dockerin-intein tag from the ELP-cohesin capturing scaffold was realized, and the regenerated ELP capturing scaffold was reused in another purification cycle without any decrease in the purification efficiency. This recyclable non-chromatographic based affinity method provides an attractive approach for efficient and cost-effective protein purification. © 2013 American Institute of Chemical Engineers.

  8. A two-step model for senescence triggered by a single critically short telomere

    DEFF Research Database (Denmark)

    Abdallah, Pauline; Luciano, Pierre; Runge, Kurt W

    2009-01-01

    Telomeres protect chromosome ends from fusion and degradation. In the absence of a specific telomere elongation mechanism, their DNA shortens progressively with every round of replication, leading to replicative senescence. Here, we show that telomerase-deficient cells bearing a single, very shor...

  9. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction.

    Science.gov (United States)

    Chomczynski, P; Sacchi, N

    1987-04-01

    A new method of total RNA isolation by a single extraction with an acid guanidinium thiocyanate-phenol-chloroform mixture is described. The method provides a pure preparation of undegraded RNA in high yield and can be completed within 4 h. It is particularly useful for processing large numbers of samples and for isolation of RNA from minute quantities of cells or tissue samples.

  10. Single crystal Fe elements patterned by one-step selective chemical wet etching

    NARCIS (Netherlands)

    Sun, Li; Wong, P.K.J.; Niu, Daxin; Zou, Xiao; Zhai, Ya; Wu, Jing; Xu, Yongbing; Zhai, Hongru

    2010-01-01

    A technique has been developed to pattern single crystal ultrathin Fe films by selective chemical wet etching of the Au capping layer and then simultaneous oxidization of the ferromagnetic Fe layer underneath. The focused magneto-optical Kerr effect and ferromagnetic resonance measurements

  11. Penerapan Metode Diagnosis Cepat Virus Avian Influenza H5N1 dengan Metode Single Step Multiplex RT-PCR

    Directory of Open Access Journals (Sweden)

    Aris Haryanto

    2010-12-01

    Full Text Available Avian influenza (AI virus is a segmented single stranded (ss RNA virus with negative polarity andbelong to the Orthomyxoviridae family. Diagnose of AI virus can be performed using conventional methodsbut it has low sensitivity and specificity. The objective of the research was to apply rapid, precise, andaccurate diagnostic method for AI virus and also to determine its type and subtype based on the SingleStep Multiplex Reverse Transcriptase-Polymerase Chain Reaction targeting M, H5, and N1 genes. In thismethod M, H5 and NI genes were simultaneously amplified in one PCR tube. The steps of this researchconsist of collecting viral RNAs from 10 different AI samples originated from Maros Disease InvestigationCenter during 2007. DNA Amplification was conducted by Simplex RT-PCR using M primer set. Then, bysingle step multiplex RT-PCR were conducted simultaneously using M, H5 and N1 primers set. The RTPCRproducts were then separated on 1.5% agarose gel, stained by ethidum bromide and visualized underUV transilluminator. Results showed that 8 of 10 RNA virus samples could be amplified by Simplex RTPCRfor M gene which generating a DNA fragment of 276 bp. Amplification using multiplex RT-PCRmethod showed two of 10 samples were AI positive using multiplex RT-PCR, three DNA fragments weregenerated consisting of 276 bp for M gene, 189 bp for H5 gene, and 131 bp for N1. In this study, rapid andeffective diagnosis method for AI virus can be conducted by using simultaneous Single Step Multiplex RTPCR.By this technique type and subtype of AI virus, can also be determined, especially H5N1.

  12. Single-Cell Genomics: A Stepping Stone for Future Immunology Discoveries.

    Science.gov (United States)

    Giladi, Amir; Amit, Ido

    2018-01-11

    The immunology field has invested great efforts and ingenuity to characterize the various immune cell types and elucidate their functions. However, accumulating evidence indicates that current technologies and classification schemes are limited in their ability to account for the functional heterogeneity of immune processes. Single-cell genomics hold the potential to revolutionize the way we characterize complex immune cell assemblies and study their spatial organization, dynamics, clonal distribution, pathways, function, and crosstalks. In this Perspective, we consider recent and forthcoming technological and analytical advances in single-cell genomics and the potential impact of those advances on the future of immunology research and immunotherapy. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. A novel single-step, multipoint calibration method for instrumented Lab-on-Chip systems

    DEFF Research Database (Denmark)

    Pfreundt, Andrea; Patou, François; Zulfiqar, Azeem

    2014-01-01

    Despite recent and substantial advances in biosensing, information and communication, and Lab-on-Chip (LoC) technologies, the success of Point-of-Care (PoC) diagnostics and monitoring systems is still challenged by stringent requirements for robustness, cost-effectiveness, and system integration....... The pitfalls of PoC system adoption can be addressed early in the system design phase. They require a multidisciplinary design approach supported by systems engineering tools and methods. Considering this, we here present both a model and an implementation of a simple and rapid calibration scheme...... for instrument-based PoC blood biomarker analysis systems. Motivated by the complexity of associating high-accuracy biosensing using silicon nanowire field effect transistors with ease of use for the PoC system user, we propose a novel one-step, multipoint calibration method for LoC-based systems. Our approach...

  14. Rapid, single-step most-probable-number method for enumerating fecal coliforms in effluents from sewage treatment plants

    Science.gov (United States)

    Munoz, E. F.; Silverman, M. P.

    1979-01-01

    A single-step most-probable-number method for determining the number of fecal coliform bacteria present in sewage treatment plant effluents is discussed. A single growth medium based on that of Reasoner et al. (1976) and consisting of 5.0 gr. proteose peptone, 3.0 gr. yeast extract, 10.0 gr. lactose, 7.5 gr. NaCl, 0.2 gr. sodium lauryl sulfate, and 0.1 gr. sodium desoxycholate per liter is used. The pH is adjusted to 6.5, and samples are incubated at 44.5 deg C. Bacterial growth is detected either by measuring the increase with time in the electrical impedance ratio between the innoculated sample vial and an uninnoculated reference vial or by visual examination for turbidity. Results obtained by the single-step method for chlorinated and unchlorinated effluent samples are in excellent agreement with those obtained by the standard method. It is suggested that in automated treatment plants impedance ratio data could be automatically matched by computer programs with the appropriate dilution factors and most probable number tables already in the computer memory, with the corresponding result displayed as fecal coliforms per 100 ml of effluent.

  15. A simple single-step approach towards synthesis of nanofluids containing cuboctahedral cuprous oxide particles using glucose reduction

    Science.gov (United States)

    Shenoy, U. Sandhya; Shetty, A. Nityananda

    2018-01-01

    Enhancement of thermal properties of conventional heat transfer fluids has become one of the important technical challenges. Since nanofluids offer a promising help in this regard, development of simpler and hassle free routes for their synthesis is of utmost importance. Synthesis of nanofluids using a hassle free route with greener chemicals has been reported. The single-step chemical approach reported here overcomes the drawbacks of the two-step procedures in the synthesis of nanofluids. The resulting Newtonian nanofluids prepared contained cuboctahedral particles of cuprous oxide and exhibited a thermal conductivity of 2.852 W·m-1·K-1. Polyvinylpyrrolidone (PVP) used during the synthesis acted as a stabilizing agent rendering the nanofluid a stability of 9 weeks.

  16. Step potential in single wire earth return system; Potencial de passo em sistema monofilar com retorno por terra

    Energy Technology Data Exchange (ETDEWEB)

    Silva Junior, Amancio R.; Caminha Junior, Isidoro C.; Watabe, Celso F.; Romero, Andrea G. [Mato Grosso do Sul Univ., Campo Grande, MS (Brazil). Dept. de Engenharia Eletrica

    1997-12-31

    It was built on the dependence of the campus of Mato Grosso do Sul University, an experimental line single wire earth return (SWER). It was realized, weekly, during the years of 1996, measurement of step potential, with a unique landing (primary only) and, primary and secondary connected. It was verified that: the most critical measurement happened when the earth presented superficial resistivity of 455,31 {Omega} x m, the highest step potential for not causing ventricular fibrillation was 249,93 V, needing a short circuit currents of 11,07 kA, the primary and secondary resistance earth presented values 42 {Omega} and 77 {Omega} respectively. There was no significant alteration when was used only the primary landing. (author) 11 refs., 2 figs.

  17. A sequential two-step near-infrared quantum splitting in Ho3+ singly doped NaYF4

    Directory of Open Access Journals (Sweden)

    D. C. Yu

    2011-12-01

    Full Text Available We demonstrated an efficient sequential two-step near-infrared (NIR quantum splitting (QS in a Ho3+ singly doped β-NaYF4. An incident high-energy ultraviolet (UV-to-visible photon in the wavelength range of 300−560 nm, which enables the Ho3+:5F4,5S2 states excited, could be efficiently split into two NIR photons at 1015 and 1180 nm. Underlying mechanisms for the sequential two-step NIR-QS process are analyzed in terms of the diffuse reflection spectrum, static and dynamic photoemission spectra and monitored excitation spectra. Internal quantum yield is obtained up to 110% on the basis of experimental and theoretical calculation results.

  18. A simple single-step approach towards synthesis of nanofluids containing cuboctahedral cuprous oxide particles using glucose reduction

    Science.gov (United States)

    Shenoy, U. Sandhya; Shetty, A. Nityananda

    2018-03-01

    Enhancement of thermal properties of conventional heat transfer fluids has become one of the important technical challenges. Since nanofluids offer a promising help in this regard, development of simpler and hassle free routes for their synthesis is of utmost importance. Synthesis of nanofluids using a hassle free route with greener chemicals has been reported. The single-step chemical approach reported here overcomes the drawbacks of the two-step procedures in the synthesis of nanofluids. The resulting Newtonian nanofluids prepared contained cuboctahedral particles of cuprous oxide and exhibited a thermal conductivity of 2.852 W·m-1·K-1. Polyvinylpyrrolidone (PVP) used during the synthesis acted as a stabilizing agent rendering the nanofluid a stability of 9 weeks.

  19. Three-dimensional photonic crystals created by single-step multi-directional plasma etching.

    Science.gov (United States)

    Suzuki, Katsuyoshi; Kitano, Keisuke; Ishizaki, Kenji; Noda, Susumu

    2014-07-14

    We fabricate 3D photonic nanostructures by simultaneous multi-directional plasma etching. This simple and flexible method is enabled by controlling the ion-sheath in reactive-ion-etching equipment. We realize 3D photonic crystals on single-crystalline silicon wafers and show high reflectance (>95%) and low transmittance (photonic bandgap. Moreover, our method simply demonstrates Si-based 3D photonic crystals that show the photonic bandgap effect in a shorter wavelength range around 0.6 μm, where further fine structures are required.

  20. From single-species advice to mixed-species management: taking the next step

    DEFF Research Database (Denmark)

    Vinther, Morten; Reeves, S.A.; Patterson, K.R.

    2004-01-01

    that accounts for mixed-fishery effects, but in the short term there is a need for approaches to resolve the conflicting management advice for different species within the same fishery, and to generate catch or effort advice that accounts for the mixed-species nature of the fishery. This paper documents...... a recent approach used to address these problems. The approach takes the single-species advice for each species in the fishery as a starting point, then attempts to resolve it into consistent catch or effort advice using fleet-disaggregated catch forecasts in combination with explicitly stated management...

  1. Bio-Docklets: virtualization containers for single-step execution of NGS pipelines.

    Science.gov (United States)

    Kim, Baekdoo; Ali, Thahmina; Lijeron, Carlos; Afgan, Enis; Krampis, Konstantinos

    2017-08-01

    Processing of next-generation sequencing (NGS) data requires significant technical skills, involving installation, configuration, and execution of bioinformatics data pipelines, in addition to specialized postanalysis visualization and data mining software. In order to address some of these challenges, developers have leveraged virtualization containers toward seamless deployment of preconfigured bioinformatics software and pipelines on any computational platform. We present an approach for abstracting the complex data operations of multistep, bioinformatics pipelines for NGS data analysis. As examples, we have deployed 2 pipelines for RNA sequencing and chromatin immunoprecipitation sequencing, preconfigured within Docker virtualization containers we call Bio-Docklets. Each Bio-Docklet exposes a single data input and output endpoint and from a user perspective, running the pipelines as simply as running a single bioinformatics tool. This is achieved using a "meta-script" that automatically starts the Bio-Docklets and controls the pipeline execution through the BioBlend software library and the Galaxy Application Programming Interface. The pipeline output is postprocessed by integration with the Visual Omics Explorer framework, providing interactive data visualizations that users can access through a web browser. Our goal is to enable easy access to NGS data analysis pipelines for nonbioinformatics experts on any computing environment, whether a laboratory workstation, university computer cluster, or a cloud service provider. Beyond end users, the Bio-Docklets also enables developers to programmatically deploy and run a large number of pipeline instances for concurrent analysis of multiple datasets. © The Authors 2017. Published by Oxford University Press.

  2. Inorganic metallodielectric materials fabricated using two single-step methods based on the Tollen's process.

    Science.gov (United States)

    Peterson, Molly S M; Bouwman, Jason; Chen, Aiqing; Deutsch, Miriam

    2007-02-01

    Two methods for preparing polycrystalline silver shells on colloidal silica spheres are reported. These do not include the use of organic ligands or metal seeding steps and are based on the Tollen's process for silvering glass. Reaction parameters such as temperature and reactant concentrations are adjusted to slow the reaction kinetics, which we find leads to preferential silver growth on the spheres. The resulting shells are polycrystalline and granular, showing highly uniform sphere coverage. Surface morphologies range from sparsely interconnected grains for shells approximately 20 nm thick, to complete (yet porous) shells of interconnected silver clusters which are up to approximately 140 nm in thickness. The extinction spectra of the core-shell materials are markedly different from those of smooth continuous shells, showing clear evidence that the granular shell geometry influences the plasmon resonance of the composite system. Spheres coated with shells 20-40 nm thick are also suitable for colloidal crystallization. Monolayers of self-assembled spheres with long-range ordering are demonstrated.

  3. Single-Step Fabrication of Computationally Designed Microneedles by Continuous Liquid Interface Production.

    Directory of Open Access Journals (Sweden)

    Ashley R Johnson

    Full Text Available Microneedles, arrays of micron-sized needles that painlessly puncture the skin, enable transdermal delivery of medications that are difficult to deliver using more traditional routes. Many important design parameters, such as microneedle size, shape, spacing, and composition, are known to influence efficacy, but are notoriously difficult to alter due to the complex nature of microfabrication techniques. Herein, we utilize a novel additive manufacturing ("3D printing" technique called Continuous Liquid Interface Production (CLIP to rapidly prototype sharp microneedles with tuneable geometries (size, shape, aspect ratio, spacing. This technology allows for mold-independent, one-step manufacturing of microneedle arrays of virtually any design in less than 10 minutes per patch. Square pyramidal CLIP microneedles composed of trimethylolpropane triacrylate, polyacrylic acid and photopolymerizable derivatives of polyethylene glycol and polycaprolactone were fabricated to demonstrate the range of materials that can be utilized within this platform for encapsulating and controlling the release of therapeutics. These CLIP microneedles effectively pierced murine skin ex vivo and released the fluorescent drug surrogate rhodamine.

  4. Single-Step Multiplex PCR Assay for Characterization of New World Leishmania Complexes

    Science.gov (United States)

    Harris, Eva; Kropp, Gerald; Belli, Alejandro; Rodriguez, Betzabé; Agabian, Nina

    1998-01-01

    We have developed a PCR assay for one-step differentiation of the three complexes of New World Leishmania (Leishmania braziliensis, Leishmania mexicana, and Leishmania donovani). This multiplex assay is targeted to the spliced leader RNA (mini-exon) gene repeats of these organisms and can detect all three complexes simultaneously, generating differently sized products for each complex. The assay is specific to the Leishmania genus and does not recognize related kinetoplastid protozoa, such as Trypanosoma cruzi, Trypanosoma brucei, and Crithidia fasciculata. It correctly identified Leishmania species with a broad geographic distribution in Central and South America. The sensitivity of the PCR amplification ranged from 1 fg to 10 pg of DNA (0.01 to 100 parasites), depending on the complex detected. Crude extracts of cultured parasites, prepared simply by boiling diluted cultures, served as excellent templates for amplification. Crude preparations of clinical material were also tested. The assay detected L. braziliensis in dermal scrapings from cutaneous leishmanial lesions, Leishmania chagasi in dermal scrapings of atypical cutaneous leishmaniasis, and L. mexicana from lesion aspirates from infected hamsters. We have minimized the material requirements and maximized the simplicity, rapidity, and informative content of this assay to render it suitable for use in laboratories in countries where leishmaniasis is endemic. This assay should be useful for rapid in-country identification of Leishmania parasites, particularly where different Leishmania complexes are found in the same geographical area. PMID:9650950

  5. Single Step In Situ Synthesis and Optical Properties of Polyaniline/ZnO Nanocomposites

    Science.gov (United States)

    Kaith, B. S.; Rajput, Jaspreet

    2014-01-01

    Polyaniline/ZnO nanocomposites were prepared by in situ oxidative polymerization of aniline monomer in the presence of different weight percentages of ZnO nanostructures. The steric stabilizer added to prevent the agglomeration of nanostructures in the polymer matrix was found to affect the final properties of the nanocomposite. ZnO nanostructures of various morphologies and sizes were prepared in the absence and presence of sodium lauryl sulphate (SLS) surfactant under different reaction conditions like in the presence of microwave radiation (microwave oven), under pressure (autoclave), under vacuum (vacuum oven), and at room temperature (ambient condition). The conductivity of these synthesized nanocomposites was evaluated using two-probe method and the effect of concentration of ZnO nanostructures on conductivity was observed. X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR), and UV-visible (UV-VIS) spectroscopy techniques were used to characterize nanocomposites. The optical energy band gap of the nanocomposites was calculated from absorption spectra and ranged between 1.5 and 3.21 eV. The reported values depicted the blue shift in nanocomposites as compared to the band gap energies of synthesized ZnO nanostructures. The present work focuses on the one-step synthesis and potential use of PANI/ZnO nanocomposite in molecular electronics as well as in optical devices. PMID:24523653

  6. Thin film complementary metal oxide semiconductor (CMOS) device using a single-step deposition of the channel layer

    KAUST Repository

    Nayak, Pradipta K.

    2014-04-14

    We report, for the first time, the use of a single step deposition of semiconductor channel layer to simultaneously achieve both n-and p-type transport in transparent oxide thin film transistors (TFTs). This effect is achieved by controlling the concentration of hydroxyl groups (OH-groups) in the underlying gate dielectrics. The semiconducting tin oxide layer was deposited at room temperature, and the maximum device fabrication temperature was 350C. Both n and p-type TFTs showed fairly comparable performance. A functional CMOS inverter was fabricated using this novel scheme, indicating the potential use of our approach for various practical applications.

  7. Single Molecule FRET Analysis of the 11 Discrete Steps of a DNA Actuator

    DEFF Research Database (Denmark)

    Hildebrandt, Lasse; Preus, Søren; Zhang, Zhao

    2014-01-01

    DNA hybridization allows the design and assembly of dynamic DNA-based molecular devices. Such structures usually accomplish their function by the addition of fuel strands that drive the structure from one conformation to a new one or by internal changes in DNA hybridization. We report here...... and nonautonomously. The 11 states of the actuator were investigated by single molecule Forster Resonance Energy Transfer (smFRET) microscopy to obtain information on the static and dynamic heterogeneities of the device. Our results show that the DNA actuator can be effectively locked in several conformations...... with the help of well-designed DNA lock strands. However, the device also shows pronounced static and dynamic heterogeneities both in the unlocked and locked modes, and we suggest possible structural models. Our study allows for the direct visualization of the conformational diversity and movement...

  8. Steps towards single source--collecting data about quality of life within clinical information systems.

    Science.gov (United States)

    Fritz, Fleur; Ständer, Sonja; Breil, Bernhard; Dugas, Martin

    2010-01-01

    Information about the quality of life from patients being treated in routine medical care is important for the attending physician. This data is also needed in research for example to evaluate the therapy and the course of the disease respectively. Especially skin diseases often negatively affect the quality of life. Therefore we aimed to design a concept to collect such data during treatment and use it for both medical care and research in the setting of dermatology. We performed a workflow analysis and implemented a designated form using the tools of the local clinical information system. Quality of life data is now collected within the clinical information system during treatment and is used for discharge letters, progress overviews as well as research about the treatment and course of disease. This concept which contributes to the single source approach was feasible within dermatology and is ready to be expanded into other domains.

  9. Single Anastomosis Sleeve Ileal Bypass: New Step in the Evolution of Bariatric Surgeries.

    Science.gov (United States)

    Salama, Tamer M S; Sabry, Karim; Ghamrini, Yaser El

    2017-10-01

    Single anastomosis sleeve ileal bypass (SASI) procedure appears as a new metabolic and bariatric surgery based on Santoro's operation, in which a sleeve gastrectomy is followed by a side-to-side gastroileal anastomosis. The study is aimed to evaluate the short-term outcomes of laparoscopic single anastomosis sleeve ileal bypass with respect to postoperative weight loss and metabolic and nutritional effects on patients. Forty-five morbidly obese patients with a mean age of 31.2 years and a mean body mass index (BMI) of 43.2 kg/m 2 underwent SASI bypass. Postoperatively, all patients were followed up at 1, 3, 6, and 12 months, and during each visit data about their BMI, obesity-related comorbidities, and nutritional status were collected. One year after the surgery, mean BMI decreased to 29.1 kg/m 2 and there was significant decrease in plasma level of fasting blood glucose, insulin, and low-density lipoprotein. On the other hand, there was significant increase in high-density lipoprotein plasma level, whereas hemoglobin and albumin plasma level remained normal. Six months after the surgery, calcium plasma level showed mild decrease in two cases, which was improved with multivitamin tablets. In 93% of our patients, multivitamin supplements were stopped after six months of surgery. Laparoscopic SASI bypass has been shown to be an effective, safe, and simple procedure for the treatment of morbid obesity and its associated metabolic consequences. Moreover, it results in minimal postoperative nutritional complications in comparison to other bariatric procedures.

  10. Pyrolysis kinetic evaluation by single-step for waste wood from reforestation.

    Science.gov (United States)

    da Silva, Jean Constantino Gomes; Alves, José Luiz Francisco; Galdino, Wendell Venicio de Araujo; Andersen, Silvia Layara Floriani; de Sena, Rennio Felix

    2018-02-01

    The objective of this study was to evaluate the kinetic parameters of pyrolysis of waste wood from reforestation: Eucalyptus benthamii (EB), Eucalyptus dunnii (ED) and Pinus elliottii (PN). The kinetic study was performed using the Friedman, Flynn-Wall-Ozawa (FWO), Kissinger-Akahira-Sunose (KAS), Starink, and Vyazovkin methods from the experimental data at four heating rates (5, 10, 20 and 30 °C min -1 ). The Friedman method presented higher activation energy values (E a ) when compared to the other methods (E a EB  = 142.98 kJ mol -1 , E a ED  = 147.71 kJ mol -1 , E a PN  = 155.46 kJ mol -1 ). The KAS, Starink and Vyazovkin methods resulted in approximate values of activation energy (E a EB  = 132.83-133.31 kJ mol -1 , E a ED  = 137.51-137.98 kJ mol -1 , E a PN  = 145.24-145.70 kJ mol -1 ) due to the approximation equations with lowest relative errors. The simulation of curves using the kinetic parameters obtained with the Vyazovkin method showed that the decomposition process of EB and ED occurs as a multi-step process resulting in an unsatisfactory result for the simulation. On the other hand, for PN a satisfactory fit to the experimental data was obtained, which demonstrates its suitability for application to the modeling of thermochemical systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Rapid single step subcloning procedure by combined action of type II and type IIs endonucleases with ligase

    Directory of Open Access Journals (Sweden)

    Klingenspor Martin

    2007-11-01

    Full Text Available Abstract Background The subcloning of a DNA fragment from an entry vector into a destination vector is a routinely performed task in molecular biology labs. Results We here present a novel benchtop procedure to achieve rapid recombination into any destination vector of choice with the sole requirement of an endonuclease recognition site. The method relies on a specifically designed entry vector and the combined action of type II and type IIs endonucleases with ligase. The formulation leads to accumulation of a single stable cloning product representing the desired insert carrying destination vector. Conclusion The described method provides a fast single step procedure for routine subcloning from an entry vector into a series of destination vectors with the same restriction enzyme recognition site.

  12. Rapid single step subcloning procedure by combined action of type II and type IIs endonucleases with ligase.

    Science.gov (United States)

    Fromme, Tobias; Klingenspor, Martin

    2007-11-26

    The subcloning of a DNA fragment from an entry vector into a destination vector is a routinely performed task in molecular biology labs. We here present a novel benchtop procedure to achieve rapid recombination into any destination vector of choice with the sole requirement of an endonuclease recognition site. The method relies on a specifically designed entry vector and the combined action of type II and type IIs endonucleases with ligase. The formulation leads to accumulation of a single stable cloning product representing the desired insert carrying destination vector. The described method provides a fast single step procedure for routine subcloning from an entry vector into a series of destination vectors with the same restriction enzyme recognition site.

  13. Direct-immersion solid-phase microextraction coupled to fast gas chromatography mass spectrometry as a purification step for polycyclic aromatic hydrocarbons determination in olive oil.

    Science.gov (United States)

    Purcaro, Giorgia; Picardo, Massimo; Barp, Laura; Moret, Sabrina; Conte, Lanfranco S

    2013-09-13

    The aim of the present work was to optimize a preparation step for polycyclic aromatic hydrocarbons in a fatty extract. Solid-phase microextraction is an easy preparation technique, which allows to minimize solvent consumption and reduce sample manipulation. A Carbopack Z/polydimethylsiloxane fiber, particularly suitable for extraction of planar compounds, was employed to extract polycyclic aromatic hydrocarbons from a hexane solution obtained after a previous extraction with acetonitrile from oil, followed by a liquid-liquid partition between acetonitrile and hexane. The proposed method was a rapid and sensitive solution to reduce the interference of triglycerides saving the column life and avoiding frequent cleaning of the mass spectrometer ion source. Despite the non-quantitative extraction of polycyclic aromatic hydrocarbons from oil using acetonitrile, the signal-to-noise ratio was significantly improved obtaining a limit of detection largely below the performance criteria required by the European Union legislation. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Purification, crystallization, X-ray diffraction analysis and phasing of an engineered single-chain PvuII restriction endonuclease

    Energy Technology Data Exchange (ETDEWEB)

    Meramveliotaki, Chrysi [Department of Science, Agricultural University of Athens, Athens (Greece); Department of Biology, University of Crete, PO Box 2208, GR-71110 Heraklion, Crete (Greece); Institute of Molecular Biology and Biotechnology (IMBB), PO Box 1527, GR-71110 Heraklion, Crete (Greece); Department of Agricultural Biotechnology, Agricultural University of Athens, Athens (Greece); Kotsifaki, Dina [Institute of Molecular Biology and Biotechnology (IMBB), PO Box 1527, GR-71110 Heraklion, Crete (Greece); Androulaki, Maria [Department of Science, Agricultural University of Athens, Athens (Greece); Department of Biology, University of Crete, PO Box 2208, GR-71110 Heraklion, Crete (Greece); Institute of Molecular Biology and Biotechnology (IMBB), PO Box 1527, GR-71110 Heraklion, Crete (Greece); Department of Agricultural Biotechnology, Agricultural University of Athens, Athens (Greece); Hountas, Athanasios [Department of Science, Agricultural University of Athens, Athens (Greece); Eliopoulos, Elias [Department of Agricultural Biotechnology, Agricultural University of Athens, Athens (Greece); Kokkinidis, Michael, E-mail: kokkinid@imbb.forth.gr [Department of Biology, University of Crete, PO Box 2208, GR-71110 Heraklion, Crete (Greece); Institute of Molecular Biology and Biotechnology (IMBB), PO Box 1527, GR-71110 Heraklion, Crete (Greece); Department of Science, Agricultural University of Athens, Athens (Greece)

    2007-10-01

    PvuII is the first type II restriction endonuclease to be converted from its wild-type homodimeric form into an enzymatically active single-chain variant. The enzyme was crystallized and phasing was successfully performed by molecular replacement. The restriction endonuclease PvuII from Proteus vulgaris has been converted from its wild-type homodimeric form into the enzymatically active single-chain variant scPvuII by tandemly joining the two subunits through the peptide linker Gly-Ser-Gly-Gly. scPvuII, which is suitable for the development of programmed restriction endonucleases for highly specific DNA cleavage, was purified and crystallized. The crystals diffract to a resolution of 2.35 Å and belong to space group P4{sub 2}, with unit-cell parameters a = b = 101.92, c = 100.28 Å and two molecules per asymmetric unit. Phasing was successfully performed by molecular replacement.

  15. Single-step multiplex RT-PCR for simultaneous and colourimetric detection of six RNA viruses in olive trees.

    Science.gov (United States)

    Bertolini, E; Olmos, A; Martínez, M C; Gorris, M T; Cambra, M

    2001-07-01

    A single-step multiplex RT-PCR was developed for the simultaneous and colourimetric detection of six RNA viruses (Cucumber mosaic virus, Cherry leaf roll virus, strawberry latent ringspot virus, Arabis mosaic virus, Olive latent-1 virus and Olive latent-2 virus) which infect olive trees. Six compatible primer set for one-step RT-PCR amplification in a single closed-tube and 3' digoxigenin labelled probes were designed in optimal, specific and conserved regions. The method has been assessed with 195 Spanish field olive trees, suggesting that approximately 1.5% of the tested material was infected by Cucumber mosaic virus and 0.5% by Cherry leaf roll virus. This method saves time and reagent costs compared with monospecific RT-PCR which needs several reactions for the same number of tests. Using colourimetric detection, it is possible to analyse many samples, it increases sensitivity 10-fold, and whilst facilitating the interpretation of results, it avoids the use of gels and the toxic ethidium bromide. The method could be used routinely for sanitary and certification programmes.

  16. Effect of increased exposure times on amount of residual monomer released from single-step self-etch adhesives.

    Science.gov (United States)

    Altunsoy, Mustafa; Botsali, Murat Selim; Tosun, Gonca; Yasar, Ahmet

    2015-10-16

    The aim of this study was to evaluate the effect of increased exposure times on the amount of residual Bis-GMA, TEGDMA, HEMA and UDMA released from single-step self-etch adhesive systems. Two adhesive systems were used. The adhesives were applied to bovine dentin surface according to the manufacturer's instructions and were polymerized using an LED curing unit for 10, 20 and 40 seconds (n = 5). After polymerization, the specimens were stored in 75% ethanol-water solution (6 mL). Residual monomers (Bis-GMA, TEGDMA, UDMA and HEMA) that were eluted from the adhesives (after 10 minutes, 1 hour, 1 day, 7 days and 30 days) were analyzed by high-performance liquid chromatography (HPLC). The data were analyzed using 1-way analysis of variance and Tukey HSD tests. Among the time periods, the highest amount of released residual monomers from adhesives was observed in the 10th minute. There were statistically significant differences regarding released Bis-GMA, UDMA, HEMA and TEGDMA between the adhesive systems (p<0.05). There were no significant differences among the 10, 20 and 40 second polymerization times according to their effect on residual monomer release from adhesives (p>0.05). Increasing the polymerization time did not have an effect on residual monomer release from single-step self-etch adhesives.

  17. A facile one-step method for synthesising a parallelogram-shaped single-crystalline ZnO nanosheet

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Renyun, E-mail: renyun.zhang@miun.se; Hummelgård, Magnus; Olin, Håkan

    2014-05-01

    Graphical abstract: - Highlights: • A simple method to synthesise ZnO nanosheets is described. • Parallelogram-shaped ZnO nanosheets were obtained with single-crystalline structure. • A specific mechanism of the growth was suggested. - Abstract: ZnO nanosheets are found to be useful in many fields such as sensors and electronics. Non-uniform-shaped ZnO nanosheets are synthesised using several methods; moreover, uniformly shaped ones are less studied. Here, we report on a simple one-step method to synthesise parallelogram-shaped single-crystalline ZnO nanosheets. By controlling the reaction of Zn(NO{sub 3}){sub 2} and hexamethylenetetramine (HMT) in ethanol, average 30 nm-thick nanosheets with a high aspect ratio of 1:100 were obtained. The parallelogram angles were between 97° and 99°. Transmission electron microscopy (TEM) diffraction and X-ray diffraction (XRD) showed that the nanosheets were wurtzite-structured single-crystalline ZnO. Moreover, a growth mechanism of these parallelogram nanosheets is suggested based on the experimental results. These results suggest a new simple solution process to synthesise uniformly shaped ZnO nanosheets allowing large-scale production to be employed.

  18. Single step sequential polydimethylsiloxane wet etching to fabricate a microfluidic channel with various cross-sectional geometries

    Science.gov (United States)

    Wang, C.-K.; Liao, W.-H.; Wu, H.-M.; Lo, Y.-H.; Lin, T.-R.; Tung, Y.-C.

    2017-11-01

    Polydimethylsiloxane (PDMS) has become a widely used material to construct microfluidic devices for various biomedical and chemical applications due to its desirable material properties and manufacturability. PDMS microfluidic devices are usually fabricated using soft lithography replica molding methods with master molds made of photolithogrpahy patterned photoresist layers on silicon wafers. The fabricated microfluidic channels often have rectangular cross-sectional geometries with single or multiple heights. In this paper, we develop a single step sequential PDMS wet etching process that can be used to fabricate microfluidic channels with various cross-sectional geometries from single-layer PDMS microfluidic channels. The cross-sections of the fabricated channel can be non-rectangular, and varied along the flow direction. Furthermore, the fabricated cross-sectional geometries can be numerically simulated beforehand. In the experiments, we fabricate microfluidic channels with various cross-sectional geometries using the developed technique. In addition, we fabricate a microfluidic mixer with alternative mirrored cross-sectional geometries along the flow direction to demonstrate the practical usage of the developed technique.

  19. Cloning, expression, purification and characterization of a single chain variable fragment specific to tumor necrosis factor alpha in Escherichia coli.

    Science.gov (United States)

    Sushma, Krishnan; Vijayalakshmi, Mookambeswaran A; Krishnan, Venkataraman; Satheeshkumar, Padikara Kutty

    2011-12-20

    Anti TNF-α molecules have been used as therapeutic agents in a variety of human diseases such as Rheumatoid arthritis, Ankylosing spondylitis, Chron's diseases, Psoriasis, etc., where high levels of TNF-α plays a destructive role. The limitations of the present TNF-α inhibitors in terms of size, tissue penetration and immunogenicity, etc., provoked the search for small anti TNF-α molecules. In the present study, a single chain variable fragment (ScFv) construct was made from a monoclonal antibody of the class IgG raised against TNF-α was used. The anti TNF-α ScFv was well expressed as soluble form in Escherichia coli BL21 (DE3), which was purified to homogeneity by commercial methacrylate monolith-convective interaction media (CIM) supports using two different chemistries, immobilized metal affinity chromatography (IMAC) with copper ions followed by anion exchange chromatography. The anti TNF-α ScFv found to be inhibiting the TNF-α mediated cytotoxicity in MCF-7 cells with an IC(50) of 8μg. Data presented here are promising and encouraging to further optimize anti TNF-α ScFv production in larger scale with higher recovery at a cheaper price for therapeutic purposes. Copyright © 2011 Elsevier B.V. All rights reserved.

  20. Compact Ag@Fe3O4 Core-shell Nanoparticles by Means of Single-step Thermal Decomposition Reaction

    Science.gov (United States)

    Brollo, Maria Eugênia F.; López-Ruiz, Román; Muraca, Diego; Figueroa, Santiago J. A.; Pirota, Kleber R.; Knobel, Marcelo

    2014-10-01

    A temperature pause introduced in a simple single-step thermal decomposition of iron, with the presence of silver seeds formed in the same reaction mixture, gives rise to novel compact heterostructures: brick-like Ag@Fe3O4 core-shell nanoparticles. This novel method is relatively easy to implement, and could contribute to overcome the challenge of obtaining a multifunctional heteroparticle in which a noble metal is surrounded by magnetite. Structural analyses of the samples show 4 nm silver nanoparticles wrapped within compact cubic external structures of Fe oxide, with curious rectangular shape. The magnetic properties indicate a near superparamagnetic like behavior with a weak hysteresis at room temperature. The value of the anisotropy involved makes these particles candidates to potential applications in nanomedicine.

  1. One-step synthesis of a singly bridged biscalix[6]arene and evaluation of its alkali metal recognition properties

    Directory of Open Access Journals (Sweden)

    Abha Naveen Kumar

    2017-10-01

    Full Text Available This article describes one-step synthesis of a singly bridged biscalix[6]arene ( 2 and evaluation of its recognition properties towards alkali metal ions for the first time. Synthesis of 2 was accomplished through coupling of two 4-tert-butylcalix[6]arene units with one molecule of diethyleneglycol ditosylate. Both base and linker were found to play a role in the outcome of coupling reaction for the simple and efficient synthesis of 2 . Complexation ability of 2 was studied by liquid-liquid extraction using alkali metal (Na +, K +, Cs + picrates. Carrier activity of 2 towards the facilitated transport of alkali metal ions across a bulk liquid membrane was also studied and the transport behaviour was mechanistically explained. The predominant role of an ion exchange action mechanism in the transport process was observed.

  2. Single-step synthesis of monolithic comb-like CdS nanostructures with tunable waveguide properties.

    Science.gov (United States)

    Liu, Ruibin; Li, Zi-An; Zhang, Chunhua; Wang, Xiaoxu; Kamran, Muhammad A; Farle, Michael; Zou, Bingsuo

    2013-06-12

    Using a simple in situ seeding chemical vapor deposition (CVD) process, comb-like (branched) monolithic CdS micro/nanostructures were grown. Efficient optical coupling between the backbone and the teeth of the branched architecture is demonstrated by distributing light from an UV-laser-excited spot at one end of the backbone to all branch tips. By varying the deposition conditions, the orientation of the branches with respect to the backbone, their size and density can be tuned as well as the size of the backbone. This in situ seeding CVD method has the potential for a low-cost single-step fabrication of high-quality, micro/nanointegrated photonic devices, with tunable complex waveguiding possibilities.

  3. Examining the rudimentary steps of the oxygen reduction reaction on single-atomic Pt using Ti-based non-oxide supports

    DEFF Research Database (Denmark)

    Tak, Young Joo; Yang, Sungeun; Lee, Hyunjoo

    2018-01-01

    In the attempt to reduce the high-cost and improve the overall durability of Pt-based electrocatalysts for the oxygen reduction reaction (ORR), density-functional theory (DFT) calculations have been performed to study the energetics of the elementary steps that occur during ORR on TiN(100)- and T...... of the single-atom Pt catalyst, and directly influences the rudimentary ORR steps on these single-atom platinized supports....

  4. Single step modified ink staining for Tzanck test: quick detection of herpetic giant cells in Tzanck smear.

    Science.gov (United States)

    Mizutani, Hitoshi; Akeda, Tomoko; Yamanaka, Kei-Ichi; Isoda, Kenichi; Gabazza, Esteban C

    2012-02-01

    Tzanck test has been recently re-evaluated as a method for the diagnosis of herpes virus infection. Giemsa staining for the Tzanck test is time-consuming and laborious. There is a need to develop simple and quick staining methods for bedside diagnosis of this disease. We report a single step and quick method for staining herpes giant cells in Tzanck smears using routinely available inks and physiological saline. A keratinocyte cell line (HaCaT) was cultured on a slide glass and stained with various commercially available blue, blue-black and black inks serially diluted with physiological saline. Clinical smear samples from herpes lesions were also stained with these solutions without specific pretreatment. The nuclei of HaCaT were clearly stained showing high contrast with the cytoplasm using 5% Parker-Quink blue-black ink saline solution. Concentration of ink solution higher or lower than 5% resulted in less contrast. Blue or black inks or other manufacturers' inks can also be used, but staining of the cultured keratinocytes was less clear. Smear of clinical samples from herpes lesions were also stained with 5% ink solution. The nuclei of the multinucleated giant cells were clearly stained, and the sample could be immediately used for microscopic examination. One step staining of Tzanck smear using this diluted ink solution is an inexpensive and a convenient bedside diagnostic tool for the dermatologist. © 2011 Japanese Dermatological Association.

  5. Improving Early Palliative Care with a Scalable, Stepped Peer Navigator and Social Work Intervention: A Single-Arm Clinical Trial.

    Science.gov (United States)

    Bekelman, David B; Johnson-Koenke, Rachel; Bowles, Daniel W; Fischer, Stacy M

    2018-02-20

    Patients with cancer could benefit from early primary (i.e., basic) palliative care. Scalable models of care delivery are needed. Examine the feasibility of a stepped peer navigator and social work intervention developed to improve palliative care outcomes. Single-arm prospective clinical trial. The peer navigator educated patients to advocate for pain and symptom management with their healthcare providers, motivated patients to pursue advance care planning, and discussed the role of hospice. The social worker saw patients with persistent psychosocial distress. Patients with advanced cancer at a VA Medical Center not currently in palliative care or hospice whose oncologist would not be surprised if the patient died in the subsequent year. Participation and retention rates, patient-reported symptoms and quality of life, advance directive documentation, patient satisfaction survey, and semistructured interviews. The participation rate was 38% (17/45), and 35% (7/17) completed final survey measures. Patients had stage IV (81%) and primarily genitourinary (47%) and lung (24%) malignancies. Median Eastern Cooperative Oncology Group performance status was 0. Patient-reported surveys indicated low distress (mean scores: Functional Assessment of Cancer Therapy-General, 75.3 [standard deviation {SD} 17.6]; Edmonton Symptom Assessment Scale symptom scores ranged from 1.6 to 3.8; Patient Health Questionnaire-9, 5.7 [SD 5.2]; and Generalized Anxiety Disorder-7, 2.8 [SD 4.1]). Of those who had not completed advance directives at baseline (n = 11, 65%), five completed them by the end of study (5/11, 45%). Patients who completed satisfaction surveys (n = 7) and interviews (n = 4) provided mixed reviews of the intervention. At a single site, a stepped peer navigator and social work palliative care study had several challenges to feasibility, including low patient-reported distress and loss to follow-up.

  6. Emulsion Liquid Membrane Technology in Organic Acid Purification

    International Nuclear Information System (INIS)

    Norela Jusoh; Norasikin Othman; Nur Alina Nasruddin

    2016-01-01

    Emulsion Liquid Membrane (ELM) process have shown a great potential in wide application of industrial separations such as in removal of many chemicals, organic compounds, metal ions, pollutants and biomolecules. This system promote many advantages including simple operation, high selectivity, low energy requirement, and single stage extraction and stripping process. One potential application of ELM is in the purification of succinic acid from fermentation broth. This study outline steps for developing emulsion liquid membrane process in purification of succinic acid. The steps include liquid membrane formulation, ELM stability and ELM extraction of succinic acid. Several carrier, diluent and stripping agent was screened to find appropriate membrane formulation. After that, ELM stability was investigated to enhance the recovery of succinic acid. Finally, the performance of ELM was evaluated in the extraction process. Results show that formulated liquid membrane using Amberlite LA2 as carrier, palm oil as diluent and sodium carbonate, Na 2 CO 3 as stripping agent provide good performance in purification. On the other hand, the prepared emulsion was observed to be stable up to 1 hour and sufficient for extraction process. In conclusion, ELM has high potential to purify succinic acid from fermentation broth. (author)

  7. Rapid purification of recombinant histones.

    Science.gov (United States)

    Klinker, Henrike; Haas, Caroline; Harrer, Nadine; Becker, Peter B; Mueller-Planitz, Felix

    2014-01-01

    The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP) as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants.

  8. Fully-Polymeric pH Sensor Realized by Means of a Single-Step Soft Embossing Technique

    Directory of Open Access Journals (Sweden)

    Paola Fanzio

    2017-05-01

    Full Text Available We present here an electrochemical sensor microsystem for the monitoring of pH. The all-polymeric device is comprised of a cyclic olefin copolymer substrate, a 200 nm-thin patterned layer of conductive polymer (PEDOT, and a 70 nm electropolymerized layer of a pH sensitive conductive polymer (polyaniline. The patterning of the fluidic (microfluidic channels and conductive (wiring and electrodes functional elements was achieved with a single soft PDMS mold via a single embossing step process. A post-processing treatment with ethylene glycol assured the functional enhancement of the electrodes, as demonstrated via an electrical and electrochemical characterization. A surface modification of the electrodes was carried out, based on voltammetric electropolymerization, to obtain a thin layer of polyaniline. The mechanism for pH sensing is based on the redox reactions of the polyaniline layer caused by protonation. The sensing performance of the microsystem was finally validated by monitoring its potentiometric response upon exposure to a relevant range of pH.

  9. High-throughput purification of recombinant proteins using self-cleaving intein tags.

    Science.gov (United States)

    Coolbaugh, M J; Shakalli Tang, M J; Wood, D W

    2017-01-01

    High throughput methods for recombinant protein production using E. coli typically involve the use of affinity tags for simple purification of the protein of interest. One drawback of these techniques is the occasional need for tag removal before study, which can be hard to predict. In this work, we demonstrate two high throughput purification methods for untagged protein targets based on simple and cost-effective self-cleaving intein tags. Two model proteins, E. coli beta-galactosidase (βGal) and superfolder green fluorescent protein (sfGFP), were purified using self-cleaving versions of the conventional chitin-binding domain (CBD) affinity tag and the nonchromatographic elastin-like-polypeptide (ELP) precipitation tag in a 96-well filter plate format. Initial tests with shake flask cultures confirmed that the intein purification scheme could be scaled down, with >90% pure product generated in a single step using both methods. The scheme was then validated in a high throughput expression platform using 24-well plate cultures followed by purification in 96-well plates. For both tags and with both target proteins, the purified product was consistently obtained in a single-step, with low well-to-well and plate-to-plate variability. This simple method thus allows the reproducible production of highly pure untagged recombinant proteins in a convenient microtiter plate format. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. Molecular Techniques for Dicistrovirus Detection without RNA Extraction or Purification

    Directory of Open Access Journals (Sweden)

    Jailson F. B. Querido

    2013-01-01

    Full Text Available Dicistroviridae is a new family of small, nonenveloped, and +ssRNA viruses pathogenic to both beneficial arthropods and insect pests as well. Triatoma virus (TrV, a dicistrovirus, is a pathogen of Triatoma infestans (Hemiptera: Reduviidae, one of the main vectors of Chagas disease. In this work, we report a single-step method to identify TrV, a dicistrovirus, isolated from fecal samples of triatomines. The identification method proved to be quite sensitive, even without the extraction and purification of RNA virus.

  11. Mechanisms of adaptation from a multiple to a single step recovery strategy following repeated exposure to forward loss of balance in older adults.

    Directory of Open Access Journals (Sweden)

    Christopher P Carty

    Full Text Available When released from an initial, static, forward lean angle and instructed to recover with a single step, some older adults are able to meet the task requirements, whereas others either stumble or fall. The purpose of the present study was to use the concept of margin of stability (MoS to investigate balance recovery responses in the anterior-posterior direction exhibited by older single steppers, multiple steppers and those that are able to adapt from multiple to single steps following exposure to repeated forward loss of balance. One hundred and fifty-one healthy, community dwelling, older adults, aged 65-80 years, participated in the study. Participants performed four trials of the balance recovery task from each of three initial lean angles. Balance recovery responses in the anterior-posterior direction were quantified at three events; cable release (CR, toe-off (TO and foot contact (FC, for trials performed at the intermediate lean angle. MoS was computed as the anterior-posterior distance between the forward boundary of the Base of Support (BoS and the vertical projection of the velocity adjusted centre of mass position (XCoM. Approximately one-third of participants adapted from a multiple to a single step recovery strategy following repeated exposure to the task. MoS at FC for the single and multiple step trials in the adaptation group were intermediate between the exclusively single step group and the exclusively multiple step group, with the single step trials having a significant, 3.7 times higher MoS at FC than the multiple step trials. Consistent with differences between single and multiple steppers, adaptation from multiple to single steps was attributed to an increased BoS at FC, a reduced XCoM at FC and an increased rate of BoS displacement from TO to FC. Adaptations occurred within a single test session and suggest older adults that are close to the threshold of successful recovery can rapidly improve dynamic stability following

  12. Magnetic Beads-Based Sensor with Tailored Sensitivity for Rapid and Single-Step Amperometric Determination of miRNAs

    Directory of Open Access Journals (Sweden)

    Eva Vargas

    2017-11-01

    Full Text Available This work describes a sensitive amperometric magneto-biosensor for single-step and rapid determination of microRNAs (miRNAs. The developed strategy involves the use of direct hybridization of the target miRNA (miRNA-21 with a specific biotinylated DNA probe immobilized on streptavidin-modified magnetic beads (MBs, and labeling of the resulting heteroduplexes with a specific DNA–RNA antibody and the bacterial protein A (ProtA conjugated with an horseradish peroxidase (HRP homopolymer (Poly-HRP40 as an enzymatic label for signal amplification. Amperometric detection is performed upon magnetic capture of the modified MBs onto the working electrode surface of disposable screen-printed carbon electrodes (SPCEs using the H2O2/hydroquinone (HQ system. The magnitude of the cathodic signal obtained at −0.20 V (vs. the Ag pseudo-reference electrode demonstrated linear dependence with the concentration of the synthetic target miRNA over the 1.0 to 100 pM range. The method provided a detection limit (LOD of 10 attomoles (in a 25 μL sample without any target miRNA amplification in just 30 min (once the DNA capture probe-MBs were prepared. This approach shows improved sensitivity compared with that of biosensors constructed with the same anti-DNA–RNA Ab as capture instead of a detector antibody and further labeling with a Strep-HRP conjugate instead of the Poly-HRP40 homopolymer. The developed strategy involves a single step working protocol, as well as the possibility to tailor the sensitivity by enlarging the length of the DNA/miRNA heteroduplexes using additional probes and/or performing the labelling with ProtA conjugated with homopolymers prepared with different numbers of HRP molecules. The practical usefulness was demonstrated by determination of the endogenous levels of the mature target miRNA in 250 ng raw total RNA (RNAt extracted from human mammary epithelial normal (MCF-10A and cancer (MCF-7 cells and tumor tissues.

  13. Purification and characterization of thermostable glucoamylase from ...

    African Journals Online (AJOL)

    Thermostable glucoamylase from Rhizopus oligosporus SK5 mutant was purified in a 3-step purification using Imarsil, activated charcoal and Sephadex-G-100 to achieve a 40-fold purification. The enzyme was optimally active at pH 5.0 and temperature of 80 °C. It exhibited a half-life of 60 minutes at 70 °C. Its stability was ...

  14. A Single-step Process to Convert Karanja Oil to Fatty Acid Methyl Esters Using Amberlyst15 as a Catalyst

    Directory of Open Access Journals (Sweden)

    Arun K. Gupta

    2018-03-01

    Full Text Available Karanja oil was successfully converted to fatty acid methyl esters (FAME in a single- step process using Amberlyst15 as a catalyst. A methanol to oil ratio of 6 was required to retain the physical structure of the Amberlyst15 catalyst. At higher methanol to oil ratios, the Amberlyst15 catalyst disintegrated. Disintegration of Amberlyst15 caused an irreversible loss in catalytic activity. This loss in activity was due to a decrease in surface area of Amberlyst15, which was caused by a decrease in its mesoporous volume. It appeared that the chemical nature of Amberlyst15 was unaffected. Reuse of Amberlyst15 with a methanol to oil ratio of 6:1 also revealed a loss in FAME yield. However, this loss in activity was recovered by heating the used Amberlyst15 catalyst to 393 K. The kinetic parameters of a power law model were successfully determined for a methanol to oil ratio of 6:1. An activation energy of 54.9 kJ mol–1 was obtained.

  15. Single-step production of the simvastatin precursor monacolin J by engineering of an industrial strain of Aspergillus terreus.

    Science.gov (United States)

    Huang, Xuenian; Liang, Yajing; Yang, Yong; Lu, Xuefeng

    2017-07-01

    Monacolin J is a key precursor for the synthesis of simvastatin (Zocor), an important drug for treating hypercholesterolemia. Industrially, monacolin J is manufactured through alkaline hydrolysis of lovastatin, a fungal polyketide produced by Aspergillus terreus. Multistep chemical processes for the conversion of lovastatin to simvastatin are laborious, cost expensive and environmentally unfriendly. A biocatalysis process for monacolin J conversion to simvastatin has been developed. However, direct bioproduction of monacolin J has not yet been achieved. Here, we identified a lovastatin hydrolase from Penicillium chrysogenum, which displays a 232-fold higher catalytic efficiency for the in vitro hydrolysis of lovastatin compared to a previously patented hydrolase, but no activity for simvastatin. Furthermore, we showed that an industrial A. terreus strain heterologously expressing this lovastatin hydrolase can produce monacolin J through single-step fermentation with high efficiency, approximately 95% of the biosynthesized lovastatin was hydrolyzed to monacolin J. Our results demonstrate a simple and green technical route for the production of monacolin J, which makes complete bioproduction of the cholesterol-lowering drug simvastatin feasible and promising. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  16. Molecular Doping of the Hole-Transporting Layer for Efficient, Single-Step Deposited Colloidal Quantum Dot Photovoltaics

    KAUST Repository

    Kirmani, Ahmad R.

    2017-07-31

    Employment of thin perovskite shells and metal halides as surface-passivants for colloidal quantum dots (CQDs) have been important, recent developments in CQD optoelectronics. These have opened the route to single-step deposited high-performing CQD solar cells. These promising architectures employ a QD hole-transporting layer (HTL) whose intrinsically shallow Fermi level (EF) restricts band-bending at maximum power-point during solar cell operation limiting charge collection. Here, we demonstrate a generalized approach to effectively balance band-edge energy levels of the main CQD absorber and charge-transport layer for these high-performance solar cells. Briefly soaking the QD HTL in a solution of the metal-organic p-dopant, molybdenum tris(1-(trifluoroacetyl)-2-(trifluoromethyl)ethane-1,2-dithiolene), effectively deepens its Fermi level, resulting in enhanced band bending at the HTL:absorber junction. This blocks the back-flow of photo-generated electrons, leading to enhanced photocurrent and fill factor compared to undoped devices. We demonstrate 9.0% perovskite-shelled and 9.5% metal-halide-passivated CQD solar cells, both achieving ca. 10% relative enhancements over undoped baselines.

  17. Single step synthesis of rutile TiO2 nanoflower array film by chemical bath deposition method

    Science.gov (United States)

    Dhandayuthapani, T.; Sivakumar, R.; Ilangovan, R.

    2016-05-01

    Titanium oxide (TiO2) nanostructures such as nanorod arrays, nanotube arrays and nanoflower arrays have been extensively investigated by the researchers. Among them nanoflower arrays has shown superior performance than other nanostructures in Dye sensitized solar cell, photocatalysis and energy storage applications. Herein, a single step synthesis for rutile TiO2 nanoflower array films suitable for device applications has been reported. Rutile TiO2 nanoflower thin film was synthesized by chemical bath deposition method using NaCl as an additive. Bath temperature induced evolution of nanoflower thin film arrays was observed from the morphological study. X-ray diffraction study confirmed the presence of rutile phase polycrystalline TiO2. Micro-Raman study revealed the presence of surface phonon mode at 105 cm-1 due to the phonon confinement effect (finite size effect), in addition with the rutile Raman active modes of B1g (143 cm-1), Eg (442 cm-1) and A1g (607 cm-1). Further, the FTIR spectrum confirmed the presence of Ti-O-Ti bonding vibration. The Tauc plot showed the direct energy band gap nature of the film with the value of 2.9 eV.

  18. Effect of a functional monomer (MDP) on the enamel bond durability of single-step self-etch adhesives.

    Science.gov (United States)

    Tsuchiya, Kenji; Takamizawa, Toshiki; Barkmeier, Wayne W; Tsubota, Keishi; Tsujimoto, Akimasa; Berry, Thomas P; Erickson, Robert L; Latta, Mark A; Miyazaki, Masashi

    2016-02-01

    The present study aimed to determine the effect of the functional monomer, 10-methacryloxydecyl dihydrogen phosphate (MDP), on the enamel bond durability of single-step self-etch adhesives through integrating fatigue testing and long-term water storage. An MDP-containing self-etch adhesive, Clearfil Bond SE ONE (SE), and an experimental adhesive, MDP-free (MF), which comprised the same ingredients as SE apart from MDP, were used. Shear bond strength (SBS) and shear fatigue strength (SFS) were measured with or without phosphoric acid pre-etching. The specimens were stored in distilled water for 24 h, 6 months, or 1 yr. Although similar SBS and SFS values were obtained for SE with pre-etching and for MF after 24 h of storage in distilled water, SE with pre-etching showed higher SBS and SFS values than MF after storage in water for 6 months or 1 yr. Regardless of the pre-etching procedure, SE showed higher SBS and SFS values after 6 months of storage in distilled water than after 24 h or 1 yr. To conclude, MDP might play an important role in enhancing not only bond strength but also bond durability with respect to repeated subcritical loading after long-term water storage. © 2015 Eur J Oral Sci.

  19. One-Step Synthesis of Single-Wall Carbon Nanotube-ZnS Core-Shell Nanocables

    Directory of Open Access Journals (Sweden)

    Yanli Zhang

    2016-08-01

    Full Text Available Nanocables with a single-wall carbon nanotube (SWCNT core and a ZnS shell were directly synthesized in one step through a thermal reaction method by using carbon, Zn, and FeS powder as starting materials. The as-fabricated nanocables were studied using scanning electron microscopy, transmission electron microscopy, and Raman spectroscopy. The nanocables have diameters of ~50 nm, lengths of several micrometers, and shell thickness of ~20 nm. TEM analysis revealed that the shell is polycrystalline wurtzite-type ZnS with good crystallinity, and the core of the nanocables is one to several SWCNTs. Raman results showed that the diameters of SWCNTs core are mainly distributed at 1.28 and 1.16 nm, with high quality and metallic character. A growth mechanism is proposed to explain the formation of the nanocables. This simple method may be applied to other SWCNTs-metal sulfide nanocables, which may have potential applications in photocatalysts, photocurrent, and other optical-electrical devices.

  20. One-Step Synthesis of Single-Wall Carbon Nanotube-ZnS Core-Shell Nanocables.

    Science.gov (United States)

    Zhang, Yanli; He, Xiangming; Wang, Li; Gao, Jian; Li, Jianjun

    2016-08-24

    Nanocables with a single-wall carbon nanotube (SWCNT) core and a ZnS shell were directly synthesized in one step through a thermal reaction method by using carbon, Zn, and FeS powder as starting materials. The as-fabricated nanocables were studied using scanning electron microscopy, transmission electron microscopy, and Raman spectroscopy. The nanocables have diameters of ~50 nm, lengths of several micrometers, and shell thickness of ~20 nm. TEM analysis revealed that the shell is polycrystalline wurtzite-type ZnS with good crystallinity, and the core of the nanocables is one to several SWCNTs. Raman results showed that the diameters of SWCNTs core are mainly distributed at 1.28 and 1.16 nm, with high quality and metallic character. A growth mechanism is proposed to explain the formation of the nanocables. This simple method may be applied to other SWCNTs-metal sulfide nanocables, which may have potential applications in photocatalysts, photocurrent, and other optical-electrical devices.

  1. From soil to leaves--aluminum fractionation by single step extraction procedures in polluted and protected areas.

    Science.gov (United States)

    Frankowski, Marcin; Zioła-Frankowska, Anetta; Siepak, Jerzy

    2013-09-30

    The paper presents the fractionation of aluminum in the samples of soil and plants of different species using a selective single-step extraction method. The study was conducted in the area located near a chemical plant, which for many years served as a post-crystallization leachate disposal site storing chemical waste (sector I), and in the area around the site: in Wielkopolski National Park, Rogalin Landscape Park and toward the infiltration ponds at the "Dębina" groundwater well-field for the city of Poznań (Poland) (sector II). The results of aluminum fractionation in samples of soil, leaves and plants showed heavy pollution with aluminum, especially in the water soluble aluminum fraction - Alsw (maximum concentration of aluminum in soil extract was 234.8 ± 4.8 mg kg(-1), in the leaves of Betula pendula it was 107.4 ± 1.8 mg kg(-1) and in the plants of Artemisia vulgaris (root) and Medicago sativa (leaves) it amounted to 464.7 ± 10.7 mg kg(-1)and 146.8 ± 1.2 mg kg(-1) respectively). In addition, the paper presents the problem of organic aluminum fractionation in biological samples and it shows the relationship between aluminum concentration in soil and the analysed woody and herbaceous species. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Three-year clinical evaluation of posterior composite restorations placed with a single-step self-etch adhesive.

    Science.gov (United States)

    Kurokawa, Hiroyasu; Takamizawa, Toshiki; Rikuta, Akitomo; Tsubota, Keishi; Miyazaki, Masashi

    2015-06-01

    In this clinical study, we evaluated the 3-year clinical performance of a resin composite containing a surface-prereacted glass ionomer (S-PRG) filler (Beautifil II; Shofu Inc., Kyoto, Japan) placed with a single-step self-etch adhesive (BeautiBond; Shofu Inc.) in posterior restorations. Using modified US Public Health Service criteria, two experienced investigators performed clinical evaluations at the baseline, 6 months, 18 months, and 3 years. Color match, marginal adaptation, anatomical form, surface roughness, marginal discoloration, postoperative sensitivity, and secondary caries were evaluated. After 3 years, 26 patients attended the recall and 31 restorations were evaluated. No postoperative sensitivity or secondary caries was observed at any time point, and no restorations failed during the follow-up period. However, surface roughness, marginal adaptation, and marginal discoloration showed deterioration after 3 years. In conclusion, although some clinical changes were observed, resin composite containing S-PRG filler placed with self-etch adhesive exhibited acceptable clinical behavior in posterior restorations.

  3. Single-Step Seeded-Growth of Graphene Nanoribbons (GNRs) via Plasma-Enhanced Chemical Vapor Deposition (PECVD)

    Science.gov (United States)

    Hsu, C.-C.; Yang, K.; Tseng, W.-S.; Li, Yiliang; Li, Yilun; Tour, J. M.; Yeh, N.-C.

    One of the main challenges in the fabrication of GNRs is achieving large-scale low-cost production with high quality. Current techniques, including lithography and unzipped carbon nanotubes, are not suitable for mass production. We have recently developed a single-step PECVD growth process of high-quality graphene sheets without any active heating. By adding some substituted aromatic as seeding molecules, we are able to rapidly grow GNRs vertically on various transition-metal substrates. The morphology and electrical properties of the GNRs are dependent on the growth parameters such as the growth time, gas flow and species of the seeding molecules. On the other hand, all GNRs exhibit strong infrared and optical absorption. From studies of the Raman spectra, scanning electron microscopic images, and x-ray/ultraviolet photoelectron spectra of these GNRs as functions of the growth parameters, we propose a model for the growth mechanism. Our findings suggest that our approach opens up a pathway to large-scale, inexpensive production of GNRs for applications to supercapacitors and solar cells. This work was supported by the Grubstake Award and NSF through IQIM at Caltech.

  4. Fire-through Ag contact formation for crystalline Si solar cells using single-step inkjet printing.

    Science.gov (United States)

    Kim, Hyun-Gang; Cho, Sung-Bin; Chung, Bo-Mook; Huh, Joo-Youl; Yoon, Sam S

    2012-04-01

    Inkjet-printed Ag metallization is a promising method of forming front-side contacts on Si solar cells due to its non-contact printing nature and fine grid resolution. However, conventional Ag inks are unable to punch through the SiN(x) anti-reflection coating (ARC) layer on emitter Si surfaces. In this study, a novel formulation of Ag ink is examined for the formation of fire-through contacts on a SiN(x)-coated Si substrate using the single-step printing of Ag ink, followed by rapid thermal annealing at 800 degrees C. In order to formulate Ag inks with fire-through contact formation capabilities, a liquid etching agent was first formulated by dissolving metal nitrates in an organic solvent and then mixing the resulting solution with a commercial Ag nanoparticle ink at various volume ratios. During the firing process, the dissolved metal nitrates decomposed into metal oxides and acted in a similar manner to the glass frit contained in Ag pastes for screen-printed Ag metallization. The newly formulated ink with a 1 wt% loading ratio of metal oxides to Ag formed finely distributed Ag crystallites on the Si substrate after firing at 800 degrees C for 1 min.

  5. Single step method to fabricate durable superliquiphobic coating on aluminum surface with self-cleaning and anti-fogging properties.

    Science.gov (United States)

    Nanda, D; Varshney, P; Satapathy, M; Mohapatra, S S; Bhushan, B; Kumar, A

    2017-12-01

    The development of self-cleaning and anti-fogging durable superliquiphobic coatings for aluminum surfaces has raised tremendous interest in materials science. In this study, a superliquiphobic coating is fabricated on an aluminum surface by a single-step dip-coating method using 1H,1H,2H,2H-Perfluorooctyltrichlorosilane-modified SiO 2 nanoparticles. The successful implementation of the aforesaid coating in different applications requires extensive investigations of its characteristics and stability. To understand the properties of the coating, surface morphology, contact angle, self-cleaning, anti-fogging, and water repellency were investigated under perturbation conditions. Additionally, the dynamics of water and oil on the coated sample also were studied. Furthermore, the durability of the coating also was examined by performing thermal, chemical, and mechanical stability tests. It was found that the coating is superliquiphobic for water, ethylene glycol, glycerol and hexadecane, and shows thermal, chemical, and mechanical stability. Further, it exhibits self-cleaning and anti-fogging properties. This approach can be applied to any size and shape aluminum surface; thus, it has great industrial applications. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Single-step One-pot Synthesis of Graphene Foam/TiO2 Nanosheet Hybrids for Effective Water Treatment

    Science.gov (United States)

    Wang, Weilin; Wang, Zhaofeng; Liu, Jingjing; Zhang, Zhengguo; Sun, Luyi

    2017-03-01

    Millions of tons of wastewater containing both inorganic and organic pollutants are generated every day, leading to significant social, environmental, and economic issues. Herein, we designed a graphene foam/TiO2 nanosheet hybrid, which is able to effectively remove both chromium (VI) cations and organic pollutants simultaneously. This graphene foam/TiO2 nanosheet hybrid was synthesized via a facile single-step one-pot hydrothermal method. The structure of the hybrid was characterized by scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The hybrid was evaluated for both chromium (VI) and organic pollutants (using methyl blue (MB) as an example) removal, and the removal mechanism was also investigated. During water treatment, graphene and TiO2 nanosheets function complimentarily, leading to a significant synergy. The hybrid exhibited outstanding chromium (VI) and MB removal capacity, much superior to the performance of the individual pure TiO2 sheets or pure graphene foam. The hybrid could also be easily separated after water treatment, and exhibited excellent recycle stability. Considering the very facile synthesis of this graphene foam/TiO2 nanosheet hybrid, and its excellent water treatment performance and recycle stability, such a hybrid is promising for large scale production for practical applications where both chromium (VI) cations and organic dyes are the main pollutants.

  7. Single-step in-situ synthesis and optical properties of ZnSe nanostructured dielectric nanocomposites

    Energy Technology Data Exchange (ETDEWEB)

    Dey, Chirantan; Rahaman Molla, Atiar; Tarafder, Anal; Karmakar, Basudeb, E-mail: basudebk@cgcri.res.in [CSIR-Central Glass and Ceramic Research Institute, Glass Science and Technology Section, Glass Division, 196, Raja S. C. Mullick Road, 700032 Kolkata (India); Kr Mishra, Manish; De, Goutam [CSIR-Central Glass and Ceramic Research Institute, Nano-Structured Materials Division, 196, Raja S. C. Mullick Road, 700032 Kolkata (India); Goswami, Madhumita; Kothiyal, G. P. [Glass and Advanced Ceramics Division, Bhaba Atomic Research Centre, Trombay, 400085 Mumbai (India)

    2014-04-07

    This work provides the evidence of visible red photoluminescent light emission from ZnSe nanocrystals (NCs) grown within a dielectric (borosilicate glass) matrix synthesized by a single step in-situ technique for the first time and the NC sizes were controlled by varying only the concentration of ZnSe in glass matrix. The ZnSe NCs were investigated by UV-Vis optical absorption spectroscopy, Raman spectroscopy, and transmission electron microscopy (TEM). The sizes of the ZnSe NCs estimated from the TEM images are found to alter in the range of 2–53 nm. Their smaller sizes of the NCs were also calculated by using the optical absorption spectra and the effective mass approximation model. The band gap enlargements both for carrier and exciton confinements were evaluated and found to be changed in the range of 0–1.0 eV. The Raman spectroscopic studies showed blue shifted Raman peaks of ZnSe at 295 and 315 cm{sup −1} indicating phonon confinement effect as well as compressive stress effect on the surface atoms of the NCs. Red photoluminescence in ZnSe-glass nanocomposite reveals a broad multiple-peak structure due to overlapping of emission from NC size related electron-hole recombination (∼707 nm) and emissions from defects to traps, which were formed due to Se and Zn vacancies signifying potential application in photonics.

  8. Development and evaluation of one step single tube multiplex RT-PCR for rapid detection and typing of dengue viruses

    Directory of Open Access Journals (Sweden)

    Parida Manmohan

    2008-01-01

    Full Text Available Abstract Background Dengue is emerging as a major public health concern in many parts of the world. The development of a one-step, single tube, rapid, and multiplex reverse transcription polymerase chain reaction (M-RT-PCR for simultaneous detection and typing of dengue virus using serotype specific primers during acute phase of illness is reported. Results An optimal assay condition with zero background was established having no cross-reaction with closely related members of flavivirus (Japanese encephalitis, West Nile, Yellow fever and alphavirus (Chikungunya. The feasibility of M-RT-PCR assay for clinical diagnosis was validated with 620 acute phase dengue patient sera samples of recent epidemics in India. The comparative evaluation vis a vis conventional virus isolation revealed higher sensitivity. None of the forty healthy serum samples screened in the present study revealed any amplification, thereby establishing specificity of the reported assay for dengue virus only. Conclusion These findings clearly suggested that M-RT-PCR assay reported in the present study is the rapid and cost-effective method for simultaneous detection as well as typing of the dengue virus in acute phase patient serum samples. Thus, the M-RT-PCR assay developed in this study will serve as a very useful tool for rapid diagnosis and typing of dengue infections in endemic areas.

  9. Do voluntary step reactions in dual task conditions have an added value over single task for fall prediction? A prospective study.

    Science.gov (United States)

    Melzer, Itshak; Kurz, Ilan; Shahar, Danit; Oddsson, Lars I E

    2010-01-01

    Stepping reactions play a critical role in responding to balance perturbations, whether they are a consequence of external perturbation or self-induced in nature. The aim of the present study was to determine prospectively the capacity of voluntary stepping performance in singleand dual-task conditions, to predict future falls among older community-dwelling persons. We also aimed to assess whether dual task conditions have an added value over single tasks for fall prediction. A total of 100 healthy old volunteers (mean age 78.4±5.7 yrs), from two self-care protected retirement homes for older adults, performed the Voluntary Step Execution Test in single- and dual-task conditions as a reaction time task while standing on a single force platform. Step initiation, preparatory and swing phases, and foot-contact time were extracted from data on center of pressure and ground reaction force. One-year fall incidences were monitored. Ninety-eight subjects completed the one-year follow-up, 49 non-fallers, 32 one-time fallers, and 17 recurrent fallers (two or more falls). Recurrent fallers had significantly slower voluntary step execution times in both single- and dual-task conditions, especially due to a slower preparation phase. Two stepwise (backward) logistic regression models showed that longer step execution times have strong predictive value for falls in both single- and dual-task conditions (odds ratio (OR) 8.7 and 5.4, respectively, ppredict future falls, with no added value to dual- over single-task condition.

  10. Single-step growth of InP/InGaAsP buried stripe MQW lasers on structured InP substrate

    Energy Technology Data Exchange (ETDEWEB)

    Rakovics, V.; Nagy, G.; Koltai, F.; Puespoeki, S.; Serenyi, M. [Hungarian Academy of Sciences, Budapest (Hungary). Research Inst. for Technical Physics; Frigeri, C.; Longo, F. [CNR MASPEC, Parma (Italy)

    1996-12-31

    Single-step LPE growth of DH lasers for 1.1--1.6 {micro}m wavelength range, and MQW lasers for 1.5--1.55 {micro}m have been demonstrated. The separate confinement bulk lasers have similar characteristics to 3QW with similar active layer volume, and both type lasers are better than the DH lasers for the same wavelength. These results indicate, that computer controlled low temperature, single-step LPE growth can be used for preparing low cost MQW devices.

  11. Single step hydrothermal synthesis of carbon nanodot decorated V2O5 nanobelts as hybrid conducting material for supercapacitor application

    Science.gov (United States)

    Narayanan, Remya

    2017-09-01

    Carbon nanodot (C-dot) decorated V2O5 (C-dot@V2O5) nanobelts are synthesized by single step, low cost hydrothermal route at low temperature by using V2O5 and glucose as precursors. We have not added any extra organic solvents or surfactants which are commonly used for the preparation of different nanostructures of V2O5. Electron microscopy analyses demonstrate that C-dot is entrapped inside V2O5 nanobelts which in turn enhance the conductivity and ion propagation property of this composite material. The C-dot@V2O5 nanobelts exhibit an excellent three electrode electrochemical performance in 1 M Na2SO4 and which showed a specific capacitance of 270 F g-1 at 1 A g-1, which is 4.5 times higher than the pristine V2O5 electrode. The electrochemical energy storage capacity of this hybrid is investigated towards solid state supercapacitor application also for the first time by employing electrophoretically deposited C-dot as the counter electrode and Li based gel as the electrolyte. The hybrid material delivers an energy density of 60 W h kg-1 and a reasonably high power density of 4.1 kW kg-1 at 5 A g-1 and good cycling stability and capacitance retention of about 87% was observed even after 5000 cycles. Above mentioned results clearly show that C-dot embedded hybrid, nanostructured transition metal oxides has great potential towards fabrication of electrodes for energy storage devices.

  12. Purification and characterization of a protease from Thermophilic ...

    African Journals Online (AJOL)

    The purification and characterization of a thermophilic neutral protease from Thermophilic bacillus strain HS08, originally isolated from a soil sample collected from the Tulufan Crater of China, is presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose anion ...

  13. Purification and characterization of a novel extracellular chitinase ...

    African Journals Online (AJOL)

    The purification and characterization of a new thermophilic chitinase from thermophilic Bacillus sp. HU1, originally isolated from a soil sample collected from hot spring of XinJiang Province, China, is presented in this paper. The purification steps included ammonium sulfate precipitation, with columns of DEAE-Sepharose ...

  14. The effect of a cognitive-motor intervention on voluntary step execution under single and dual task conditions in older adults: a randomized controlled pilot study

    Directory of Open Access Journals (Sweden)

    Pichierri G

    2012-07-01

    Full Text Available Giuseppe Pichierri,1 Amos Coppe,1 Silvio Lorenzetti,2 Kurt Murer,1 Eling D de Bruin11Institute of Human Movement Sciences and Sport, Department of Health Sciences and Technology, ETH Zurich, Switzerland; 2Institute for Biomechanics, Department of Health Sciences and Technology, ETH Zurich, SwitzerlandBackground: This randomized controlled pilot study aimed to explore whether a cognitive-motor exercise program that combines traditional physical exercise with dance video gaming can improve the voluntary stepping responses of older adults under attention demanding dual task conditions.Methods: Elderly subjects received twice weekly cognitive-motor exercise that included progressive strength and balance training supplemented by dance video gaming for 12 weeks (intervention group. The control group received no specific intervention. Voluntary step execution under single and dual task conditions was recorded at baseline and post intervention (Week 12.Results: After intervention between-group comparison revealed significant differences for initiation time of forward steps under dual task conditions (U = 9, P = 0.034, r = 0.55 and backward steps under dual task conditions (U = 10, P = 0.045, r = 0.52 in favor of the intervention group, showing altered stepping levels in the intervention group compared to the control group.Conclusion: A cognitive-motor intervention based on strength and balance exercises with additional dance video gaming is able to improve voluntary step execution under both single and dual task conditions in older adults.Keywords: fall prevention, exercise, dance, video game

  15. The effect of a cognitive-motor intervention on voluntary step execution under single and dual task conditions in older adults: a randomized controlled pilot study.

    Science.gov (United States)

    Pichierri, Giuseppe; Coppe, Amos; Lorenzetti, Silvio; Murer, Kurt; de Bruin, Eling D

    2012-01-01

    This randomized controlled pilot study aimed to explore whether a cognitive-motor exercise program that combines traditional physical exercise with dance video gaming can improve the voluntary stepping responses of older adults under attention demanding dual task conditions. Elderly subjects received twice weekly cognitive-motor exercise that included progressive strength and balance training supplemented by dance video gaming for 12 weeks (intervention group). The control group received no specific intervention. Voluntary step execution under single and dual task conditions was recorded at baseline and post intervention (Week 12). After intervention between-group comparison revealed significant differences for initiation time of forward steps under dual task conditions (U = 9, P = 0.034, r = 0.55) and backward steps under dual task conditions (U = 10, P = 0.045, r = 0.52) in favor of the intervention group, showing altered stepping levels in the intervention group compared to the control group. A cognitive-motor intervention based on strength and balance exercises with additional dance video gaming is able to improve voluntary step execution under both single and dual task conditions in older adults.

  16. Modelling Plasticity of Ni3Al-Based L12 Intermetallic Single Crystals. II. Two-Step (T1) and T2) Deformation Behaviour (Postprint

    National Research Council Canada - National Science Library

    Choi, Y. S; Dimiduk, D. M; Uchic, M. D; Parthasarathy, T. A

    2007-01-01

    The two-step (T1 and T2) deformation behaviour of Ni3Al-based single crystals was modelled under the framework of a new constitutive model proposed by Y.S. Choi, D.M. Dimiduk, M.D. Uchic, et al. [Phil. Mag. 87 1939 (2007...

  17. Soft-Bake Purification of SWCNTs Produced by Pulsed Laser Vaporization

    Science.gov (United States)

    Yowell, Leonard; Nikolaev, Pavel; Gorelik, Olga; Allada, Rama Kumar; Sosa, Edward; Arepalli, Sivaram

    2013-01-01

    The "soft-bake" method is a simple and reliable initial purification step first proposed by researchers at Rice University for single-walled carbon nanotubes (SWCNT) produced by high-pressure carbon mon oxide disproportionation (HiPco). Soft-baking consists of annealing as-produced (raw) SWCNT, at low temperatures in humid air, in order to degrade the heavy graphitic shells that surround metal particle impurities. Once these shells are cracked open by the expansion and slow oxidation of the metal particles, the metal impurities can be digested through treatment with hydrochloric acid. The soft-baking of SWCNT produced by pulsed-laser vaporization (PLV) is not straightforward, because the larger average SWCNT diameters (.1.4 nm) and heavier graphitic shells surrounding metal particles call for increased temperatures during soft-bake. A part of the technology development focused on optimizing the temperature so that effective cracking of the graphitic shells is balanced with maintaining a reasonable yield, which was a critical aspect of this study. Once the ideal temperature was determined, a number of samples of raw SWCNT were purified using the soft-bake method. An important benefit to this process is the reduced time and effort required for soft-bake versus the standard purification route for SWCNT. The total time spent purifying samples by soft-bake is one week per batch, which equates to a factor of three reduction in the time required for purification as compared to the standard acid purification method. Reduction of the number of steps also appears to be an important factor in improving reproducibility of yield and purity of SWCNT, as small deviations are likely to get amplified over the course of a complicated multi-step purification process.

  18. The development and evaluation of single cell suspension from wheat and barley as a model system; a first step towards functional genomics application

    DEFF Research Database (Denmark)

    Dong, Jing; Bowra, Steve; Vincze, Éva

    2010-01-01

    Background The overall research objective was to develop single cell plant cultures as a model system to facilitate functional genomics of monocots, in particular wheat and barley. The essential first step towards achieving the stated objective was the development of a robust, viable single cell...... suspension culture from both species. Results We established growth conditions to allow routine culturing of somatic cells in 24 well microtiter plate format. Evaluation of the wheat and barley cell suspension as model cell system is a multi step process. As an initial step in the evaluation procedure we...... level of genes (P5CS, P5CR) under various treatments and we suggest that the cells can be used as a model host system to study gene expression and regulation in monocots....

  19. Single-Step Syngas-to-Distillates (S2D) Synthesis via Methanol and Dimethyl Ether Intermediates: Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Dagle, Robert A.; Lebarbier, Vanessa MC; Lizarazo Adarme, Jair A.; King, David L.; Zhu, Yunhua; Gray, Michel J.; Jones, Susanne B.; Biddy, Mary J.; Hallen, Richard T.; Wang, Yong; White, James F.; Holladay, Johnathan E.; Palo, Daniel R.

    2013-11-26

    would be allowed for methanol synthesis alone. Aromatic-rich hydrocarbon liquid (C5+), containing a significant amount of methylated benzenes, was produced under these conditions. However, selectivity control to liquid hydrocarbons was difficult to achieve. Carbon dioxide and methane formation was problematic. Furthermore, saturation of the olefinic intermediates formed in the zeolite, and necessary for gasoline production, occurred over PdZnAl. Thus, yield to desirable hydrocarbon liquid product was limited. Evaluation of other oxygenate-producing catalysts could possibly lead to future advances. Potential exists with discovery of other types of catalysts that suppress carbon dioxide and light hydrocarbon formation. Comparative techno-economics for a single-step syngas-to-distillates process and a more conventional MTG-type process were investigated. Results suggest operating and capital cost savings could only modestly be achieved, given future improvements to catalyst performance. Sensitivity analysis indicated that increased single-pass yield to hydrocarbon liquid is a primary need for this process to achieve cost competiveness.

  20. Improving the extraction and purification of immunoglobulin G by the use of ionic liquids as adjuvants in aqueous biphasic systems

    Science.gov (United States)

    Mondal, Dibyendu; Coutinho, João A. P.; Freire, Mara G.

    2017-01-01

    Immunoglobulins G (IgG) could become widespread biopharmaceuticals if cost-efficient processes for their extraction and purification are available. In this work, aqueous biphasic systems (ABS) composed of polyethylene glycols and a buffered salt, and with ionic liquids (ILs) as adjuvants, have been studied as alternative extraction and purification platforms of IgG from a rabbit serum source. Eleven ILs were investigated to provide insights on the chemical features which maximize the IgG partitioning. It is shown that in polymer-salt systems pure IgG preferentially partitions to the polymer-rich phase; yet, the complete extraction was never attained. Remarkably, after the addition of 5 wt% of adequate ILs to polymer-salt ABS, the complete extraction of pure IgG in a single-step was accomplished. The best systems and conditions were then applied to the extraction and purification of IgG directly from rabbit serum samples. The complete extraction of IgG in a single-step was maintained while its purity in the polymer-rich phase was enhanced by ca. 37% as compared to the IL-free ABS. The antibody stability was also evaluated revealing that appropriate ILs are able to maintain the IgG stability and can be used as phase-forming components of ABS when envisaging the purification of high-cost biopharmaceuticals. PMID:27568168

  1. Dynamics of Nonlinear Excitation of the High-Order Mode in a Single-Mode Step-Index Optical Fiber

    Science.gov (United States)

    Burdin, V.; Bourdine, A.

    2018-04-01

    This work is concerned with approximate model of higher-order mode nonlinear excitation in a singlemode silica optical fiber. We present some results of simulation for step-index optical fiber under femtosecond optical pulse launching, which confirm ability of relatively stable higher-order mode excitation in such singlemode optical fiber over sufficiently narrow range of launched optical power variation.

  2. Purification And Characterization Of Marine Bacillus Thuringiensis ...

    African Journals Online (AJOL)

    Urease was purified to homogeneity from Bacillus thuringiensis N2 using different purification steps namely, 55% acetone precipitation, DEAE-Sephadex A50 anion exchange column and Sephadex G120-200 gel filtration chromatography. The enzyme was purified 95.27 fold and showed a final specific activity of 10.48 ...

  3. Purification and refolding of anti-T-antigen single chain antibodies (scFvs) expressed in Escherichia coli as inclusion bodies.

    Science.gov (United States)

    Yuasa, Noriyuki; Koyama, Tsubasa; Fujita-Yamaguchi, Yoko

    2014-02-01

    T-antigen (Galβ1-3GalNAcα-1-Ser/Thr) is an oncofetal antigen that is commonly expressed as a carbohydrate determinant in many adenocarcinomas. Since it is associated with tumor progression and metastasis, production of recombinant antibodies specific for T-antigen could lead to the development of cancer diagnostics and therapeutics. Previously, we isolated and characterized 11 anti-T-antigen phage clones from a phage library displaying human single-chain antibodies (scFvs) and purified one scFv protein, 1G11. More recently, we purified and characterized 1E8 scFv protein using a Drosophila S2 expression system. In the current study, four anti-T-antigen scFv genes belonging to Groups 1-4 were purified from inclusion bodies expressed in Escherichia coli cells. Inclusion bodies isolated from E. coli cells were denatured in 3.5 M Gdn-HCl. Solubilized His-tagged scFv proteins were purified using Ni(2+)-Sepharose column chromatography in the presence of 3.5 M Gdn-HCl. Purified scFv proteins were refolded according to a previously published method of step-wise dialysis. Two anti-T-antigen scFv proteins, 1E6 and 1E8 that belong to Groups 1 and 2, respectively, were produced in sufficient amounts, thus allowing further characterization of their binding activity with T-antigen. Specificity and affinity constants determined using enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR), respectively, provided evidence that both 1E8 and 1E6 scFv proteins are T-antigen specific and suggested that 1E8 scFv protein has a higher affinity for T-antigen than 1E6 scFv protein.

  4. Influence of light intensity on surface-free energy and dentin bond strength of single-step self-etch adhesives.

    Science.gov (United States)

    Nojiri, Kie; Tsujimoto, Akimasa; Suzuki, Takayuki; Shibasaki, Syo; Matsuyoshi, Saki; Takamizawa, Toshiki; Miyazaki, Masashi

    2015-01-01

    In this study, we investigated the influence of light intensity on the surface-free energy and dentin bond strength of single-step selfetch adhesives. The adhesives were applied to the dentin surfaces of bovine mandibular incisors and cured with light intensities of 0 (no irradiation), 200, 400, and 600 mW/cm(2). Surface-free energies were determined by measuring the contact angles of three test liquids placed on the cured adhesives. Dentin bond strengths of the specimens were also measured. Polymerization with a higher light intensity resulted in a lower surface-free energy of the cured adhesives. The greatest bond strength was achieved when a light intensity of 400 mW/cm(2) or greater was used. Our data suggest that the surface-free energy and dentin bond strength of single-step self-etch adhesives are affected by light intensity of the curing unit.

  5. Single-step immobilization of cell adhesive peptides on a variety of biomaterial substrates via tyrosine oxidation with copper catalyst and hydrogen peroxide.

    Science.gov (United States)

    Kakinoki, Sachiro; Yamaoka, Tetsuji

    2015-04-15

    Immobilization of biologically active peptides which were isolated from extracellular matrix proteins is a powerful strategy for the design and functionalization of biomaterial substrates. However, the method of peptide immobilization was restricted, that is, peptide is often immobilized through the reactive groups inherent in substrates with multistep reactions. Here, we report a single-step immobilization of fibronectin-derived cell adhesive peptide (Arg-Glu-Asp-Val; REDV) onto polymer materials by use of tyrosine oxidation with copper catalyst and hydrogen peroxide. REDV peptide was successfully immobilized on tissue culture polystyrene, poly(ethylene terephthalate), poly(vinyl chloride), expanded-poly(tetrafluoroethylene), and poly(l-lactic acid), resulting in enhanced adhesion of human umbilical vein endothelial cells. This method is a single-step reaction under very mild conditions and is available for the biological functionalization of various medical devices.

  6. Single-step preparation of TiO2/MWCNT Nanohybrid materials by laser pyrolysis and application to efficient photovoltaic energy conversion.

    Science.gov (United States)

    Wang, Jin; Lin, Yaochen; Pinault, Mathieu; Filoramo, Arianna; Fabert, Marc; Ratier, Bernard; Bouclé, Johann; Herlin-Boime, Nathalie

    2015-01-14

    This paper presents the continuous-flowand single-step synthesis of a TiO2/MWCNT (multiwall carbon nanotubes) nanohybrid material. The synthesis method allows achieving high coverage and intimate interface between the TiO2particles and MWCNTs, together with a highly homogeneous distribution of nanotubes within the oxide. Such materials used as active layer in theporous photoelectrode of solid-state dye-sensitized solar cells leads to a substantial performance improvement (20%) as compared to reference devices.

  7. Synthesis of [11C]palmitic acid for PET imaging using a single molecular sieve 13X cartridge for reagent trapping, radiolabeling and selective purification.

    Science.gov (United States)

    Amor-Coarasa, Alejandro; Kelly, James M; Babich, John W

    2015-08-01

    Radiolabeled fatty acids are valuable metabolic tracers for PET imaging. Carbon-11 is widely used in clinical PET studies due to the prevalence of facile techniques enabling the incorporation of [(11)C]CO2 and [(11)C]CH3 into molecules and a short half-life (20.4 min) that translates into low patient dose. However, the short half-life considerably limits the time for radiosynthesis. Furthermore, the majority of the syntheses of [(11)C]palmitic acid in common use employ high starting [(11)C]CO2 activities and/or expensive equipment. [(11)C]CO2 was trapped with greater than 99.99% efficiency by a three stage cartridge packed with molecular sieve 13X, 100-120 mesh. The labeling of n-pentadecylmagnesium bromide took place in 5 min in the cartridge, and the [(11)C]palmitic acid product was selectively eluted in ethanol following alkaline and acidic washes of the column. The system reliably produced more than 925 MBq (25 mCi) of [(11)C]palmitic acid suitable for human use from 7.4 GBq (200 mCi) of [(11)C]CO2 in 8 min from end-of-bombardment. We have exploited the properties of the inexpensive molecular sieve 13X to develop a miniature, disposable and leak tight "gas capture" system for the rapid labeling and purification of [(11)C]fatty acids in good yield and >99% radiochemical purity. The rapidity of the synthesis and purification allows small [(11)C]CO2 starting activities to be used, and with no requirement for expensive synthesis equipment or facilities, the system can be implemented in any radiopharmaceutical center. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. Single-step syngas-to-distillates (S2D) process based on biomass-derived syngas--a techno-economic analysis.

    Science.gov (United States)

    Zhu, Yunhua; Jones, Susanne B; Biddy, Mary J; Dagle, Robert A; Palo, Daniel R

    2012-08-01

    This study compared biomass gasification based syngas-to-distillate (S2D) systems using techno-economic analysis (TEA). Three cases, state of technology (SOT), goal, and conventional, were compared in terms of performance and cost. The SOT case represented the best available experimental results for a process starting with syngas using a single-step dual-catalyst reactor for distillate generation. The conventional case mirrored a conventional two-step S2D process consisting of separate syngas-to-methanol and methanol-to-gasoline (MTG) processes. The goal case assumed the same performance as the conventional, but with a single-step S2D technology. TEA results revealed that the SOT was more expensive than the conventional and goal cases. The SOT case suffers from low one-pass yield and high selectivity to light hydrocarbons, both of which drive up production cost. Sensitivity analysis indicated that light hydrocarbon yield and single pass conversion efficiency were the key factors driving the high cost for the SOT case. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. A simple method for large-scale purification of plasma-derived apo-transferrin.

    Science.gov (United States)

    Ascione, Ester; Muscariello, Livio; Maiello, Valentina; Romano, Vincenza; Giovacchini, Pierangelo; Nardini, Ilaria; Farina, Claudio

    2010-12-01

    We investigated and optimized a purification process, suitable for industrial scale, to obtain pharmaceutical grade apo-Tf (apo-transferrin), preserving its physiological properties and functions. Apo-Tf was obtained from fraction IV subfraction 1 and IV subfraction 4 (fraction IV-1,4), a waste product of the Cohn fractionation process, performing a single chromatographic run and two viral inactivation/removal steps. The structural integrity and the biological activity of the final product were extensively tested. The yield of apo-Tf produced was 80% on laboratory scale and 90% in scale-up lots, and the purity was higher than 95%. The purified protein preserves iron- and receptor-binding activities and shows a normal glycosylation pattern. The single chromatographic step process presented here provides an efficient means to prepare commercial quantities of the protein. The final product is sterile and two viral inactivation/removal steps were introduced into the process.

  10. Necessity of purification during bacterial DNA extraction with environmental soils

    Directory of Open Access Journals (Sweden)

    Hyun Jeong Lim

    2017-08-01

    Full Text Available Complexity and heterogeneity of soil samples have often implied the inclusion of purification steps in conventional DNA extraction for polymerase chain reaction (PCR assays. Unfortunately the purification steps are also time and labor intensive. Therefore the necessity of DNA purification was re-visited and investigated for a variety of environmental soil samples that contained various amounts of PCR inhibitors. Bead beating and centrifugation was used as the baseline (without purification method for DNA extraction. Its performance was compared with that of conventional DNA extraction kit (with purification. The necessity criteria for DNA purification were established with environmental soil samples. Using lysis conditions at 3000 rpm for 3 minutes with 0.1 mm glass beads, centrifugation time of 10 minutes and 1:10 dilution ratio, the baseline method outperformed conventional DNA extraction on cell seeded sand samples. Further investigation with PCR inhibitors (i.e., humic acids, clay, and magnesium [Mg] showed that sand samples containing less than 10 μg/g humic acids and 70% clay may not require purifications. Interestingly, the inhibition pattern of Mg ion was different from other inhibitors due to the complexation interaction of Mg ion with DNA fragments. It was concluded that DNA extraction method without purification is suitable for soil samples that have less than 10 μg/g of humic acids, less than 70% clay content and less than 0.01% Mg ion content.

  11. The treatment of cheese whey wastewater by sequential anaerobic and aerobic steps in a single digester at pilot scale.

    Science.gov (United States)

    Frigon, J-C; Breton, J; Bruneau, T; Moletta, R; Guiot, S R

    2009-09-01

    The treatment of reconstituted whey wastewater was performed in a 400 L digester at 20 degrees C, with an anaerobic digestion step, followed by a step of aerobic treatment at low oxygen concentration in the same digester. In a first set of 48 cycles, total cycle time (T(C)) of 2, 3 and 4 days were tested at varying organic loading rates (OLR). The COD removal reached 89+/-4, 97+/-3 and 98+/-2% at T(C) of 2, 3 and 4 days and OLR of 0.56, 1.04 and 0.78 g COD L(-1) d(-1), respectively. The activity of the biomass decreased for the methanogenic population, while increasing by 400% for the acidogens, demonstrating a displacement in the predominant trophic group in the biomass bed. A second set of 16 cycles was performed with higher soluble oxygen concentration in the bulk liquid (0.5 mg L(-1)) during the aerobic treatment at a T(C) of 2 days and an OLR of 1.55 g COD L(-1) d(-1), with a soluble COD removal of 88+/-3%. The biomass specific activities showed a compartmentalization of the trophic group with methanogenic activity maintained in the biomass bed and a high acidogenic activity in the suspended flocs.

  12. Single-step direct fabrication of pillar-on-pore hybrid nanostructures in anodizing aluminum for superior superhydrophobic efficiency.

    Science.gov (United States)

    Jeong, Chanyoung; Choi, Chang-Hwan

    2012-02-01

    Conventional electrochemical anodizing processes of metals such as aluminum typically produce planar and homogeneous nanopore structures. If hydrophobically treated, such 2D planar and interconnected pore structures typically result in lower contact angle and larger contact angle hysteresis than 3D disconnected pillar structures and, hence, exhibit inferior superhydrophobic efficiency. In this study, we demonstrate for the first time that the anodizing parameters can be engineered to design novel pillar-on-pore (POP) hybrid nanostructures directly in a simple one-step fabrication process so that superior surface superhydrophobicity can also be realized effectively from the electrochemical anodization process. On the basis of the characteristic of forming a self-ordered porous morphology in a hexagonal array, the modulation of anodizing voltage and duration enabled the formulation of the hybrid-type nanostructures having controlled pillar morphology on top of a porous layer in both mild and hard anodization modes. The hybrid nanostructures of the anodized metal oxide layer initially enhanced the surface hydrophilicity significantly (i.e., superhydrophilic). However, after a hydrophobic monolayer coating, such hybrid nanostructures then showed superior superhydrophobic nonwetting properties not attainable by the plain nanoporous surfaces produced by conventional anodization conditions. The well-regulated anodization process suggests that electrochemical anodizing can expand its usefulness and efficacy to render various metallic substrates with great superhydrophilicity or -hydrophobicity by directly realizing pillar-like structures on top of a self-ordered nanoporous array through a simple one-step fabrication procedure.

  13. Purification of Escherichia coli L-asparaginase mutants by a native polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Wei, Yujun; Chen, Jianhua; Jia, Ruibo; Wang, Min; Wu, Wutong

    2008-07-01

    The antigenicity of L-asaparaginase (L-ASP) has been problematic for the treatment of leukemia for many years. In order to establish a relationship between the antigenic epitope of L-asparaginase and its antigenicity, several L-asparaginase mutants (mL-ASPs) are constructed and expressed. To effectively purify these enzyme mutants for further investigation, a native preparative polyacrylamide gel electrophoresis is developed. The simplicity and reproducibility of this approach permits the purification of different mutants from the crude enzyme extracts, with a sufficient activity to perform immunological and biological studies. Furthermore, the newly developed method is efficient and cost-effective compared with other methods, such as column chromatography and affinity chromatography. As a result, the enzyme mutants with specific activity of 300 approximately 400 U/mg are obtained by the single-step purification with a high degree of purity.

  14. Total chemical synthesis of proteins without HPLC purification.

    Science.gov (United States)

    Loibl, S F; Harpaz, Z; Zitterbart, R; Seitz, O

    2016-11-01

    The total chemical synthesis of proteins is a tedious and time-consuming endeavour. The typical steps involve solid phase synthesis of peptide thioesters and cysteinyl peptides, native chemical ligation (NCL) in solution, desulfurization or removal of ligation auxiliaries in the case of extended NCL as well as many intermediary and final HPLC purification steps. With an aim to facilitate and improve the throughput of protein synthesis we developed the first method for the rapid chemical total on-resin synthesis of proteins that proceeds without a single HPLC-purification step. The method relies on the combination of three orthogonal protein tags that allow sequential immobilization ( via the N-terminal and C-terminal ends), extended native chemical ligation and release reactions. The peptide fragments to be ligated are prepared by conventional solid phase synthesis and used as crude materials in the subsequent steps. An N-terminal His 6 unit permits selective immobilization of the full length peptide thioester onto Ni-NTA agarose beads. The C-terminal peptide fragment carries a C-terminal peptide hydrazide and an N-terminal 2-mercapto-2-phenyl-ethyl ligation auxiliary, which serves as a reactivity tag for the full length peptide. As a result, only full length peptides, not truncation products, react in the subsequent on-bead extended NCL. After auxiliary removal the ligation product is liberated into solution upon treatment with mild acid, and is concomitantly captured by an aldehyde-modified resin. This step allows the removal of the most frequently observed by-product in NCL chemistry, i.e. the hydrolysed peptide thioester (which does not contain a C-terminal peptide hydrazide). Finally, the target protein is released with diluted hydrazine or acid. We applied the method in the synthesis of 46 to 126 amino acid long MUC1 proteins comprising 2-6 copies of a 20mer tandem repeat sequence. Only three days were required for the parallel synthesis of 9 MUC1 proteins

  15. Single-fluorophore monitoring of DNA hybridization for investigating the effect of secondary structure on the nucleation step.

    Science.gov (United States)

    Jo, Joon-Jung; Kim, Min-Ji; Son, Jung-Tae; Kim, Jandi; Shin, Jong-Shik

    2009-07-17

    Nucleic acid hybridization is one of the essential biological processes involved in storage and transmission of genetic information. Here we quantitatively determined the effect of secondary structure on the hybridization activation energy using structurally defined oligonucleotides. It turned out that activation energy is linearly proportional to the length of a single-stranded region flanking a nucleation site, generating a 0.18 kcal/mol energy barrier per nucleotide. Based on this result, we propose that the presence of single-stranded segments available for non-productive base pairing with a nucleation counterpart extends the searching process for nucleation sites to find a perfect match. This result may provide insights into rational selection of a target mRNA site for siRNA and antisense gene silencing.

  16. Highly efficient one-step conversion of cyclohexane to adipic acid using single-site heterogeneous catalysts.

    Science.gov (United States)

    Raja, Robert; Thomas, John Meurig; Xu, Mingcan; Harris, Kenneth D M; Greenhill-Hooper, Michael; Quill, Kieran

    2006-01-28

    A solid source of 'active' oxygen (acetylperoxyborate, APB), when dissolved in aqueous solution in the presence of a single-site microporous catalyst containing redox centres (Fe(III)AlPO-31, Mn(III)AlPO-5, Fe(III)AlPO-5), converts cyclohexane with high efficiency (ca. 88%) and exceptionally high selectivity (ca. 81%) to adipic acid at 383 K; this procedure is also effective in converting styrene to styrene oxide and -pinene and (+)-limonene to their corresponding epoxides.

  17. A novel single-step procedure for the calibration of the mounting parameters of a multi-camera terrestrial mobile mapping system

    Science.gov (United States)

    Habib, A.; Kersting, P.; Bang, K.; Rau, J.

    2011-12-01

    Mobile Mapping Systems (MMS) can be defined as moving platforms which integrates a set of imaging sensors and a position and orientation system (POS) for the collection of geo-spatial information. In order to fully explore the potential accuracy of such systems and guarantee accurate multi-sensor integration, a careful system calibration must be carried out. System calibration involves individual sensor calibration as well as the estimation of the inter-sensor geometric relationship. This paper tackles a specific component of the system calibration process of a multi-camera MMS - the estimation of the relative orientation parameters among the cameras, i.e., the inter-camera geometric relationship (lever-arm offsets and boresight angles among the cameras). For that purpose, a novel single step procedure, which is easy to implement and not computationally intensive, will be introduced. The proposed method is implemented in such a way that it can also be used for the estimation of the mounting parameters among the cameras and the IMU body frame, in case of directly georeferenced systems. The performance of the proposed method is evaluated through experimental results using simulated data. A comparative analysis between the proposed single-step and the two-step, which makes use of the traditional bundle adjustment procedure, is demonstrated.

  18. Single molecule measurements of DNA helicase activity with magnetic tweezers and t-test based step-finding analysis

    Science.gov (United States)

    Seol, Yeonee; Strub, Marie-Paule; Neuman, Keir C.

    2016-01-01

    Magnetic tweezers is a versatile and easy to implement single-molecule technique that has become increasingly prevalent in the study of nucleic acid based molecular motors. Here, we provide a description of the magnetic tweezers instrument and guidelines for measuring and analyzing DNA helicase activity. Along with experimental methods, we describe a robust method of single-molecule trajectory analysis based on the Student’s t-test that accommodates continuous transitions in addition to the discrete transitions assumed in most widely employed analysis routines. To illustrate the single-molecule unwinding assay and the analysis routine, we provide DNA unwinding measurements of Escherichia coli RecQ helicase under a variety of conditions (Na+, ATP, temperature, and DNA substrate geometry). These examples reveal that DNA unwinding measurements under various conditions can aid in elucidating the unwinding mechanism of DNA helicase but also emphasize that environmental effects on DNA helicase activity must be considered in relation to in vivo activity and mechanism. PMID:27131595

  19. A novel single-step surgical technique for vestibular deepening using laser in conjunction with periodontal flap surgery

    Directory of Open Access Journals (Sweden)

    Ashu Bhardwaj

    2016-01-01

    Full Text Available Moderate-to-severe chronic periodontitis results in clinical loss of attachment, reduced width of attached gingiva (AG, periodontal pockets beyond mucogingival junction (MGJ, gingival recession, loss of alveolar bone, and decreased vestibular depth (VD. The encroachment of frenal and muscle attachments on marginal gingiva increases the rate of progression of periodontal pockets, prevents healing, and causes their recurrence after therapy. Loss of VD and AG associated with continuous progression of pocket formation and bone loss requires two-stage surgical procedures. In this article, one-stage surgical procedure is being described for the first time, to treat the periodontal pockets extending beyond the MGJ by periodontal flap surgery along with vestibular deepening with diode laser to increase the AG. One-step surgical technique is illustrated whereby pocket therapy with reconstruction of lost periodontal tissues can be done along with gingival augmentation by vestibular deepening.

  20. Synthesis and integration of Fe-soc-MOF cubes into colloidosomes via a single-step emulsion-based approach

    KAUST Repository

    Pang, Maolin

    2013-07-17

    Bottom-up fabrication of complex 3D hollow superstructures from nonspherical building blocks (BBs) poses a significant challenge for scientists in materials chemistry and physics. Spherical colloidal silica or polystyrene particles are therefore often integrated as BBs for the preparation of an emerging class of materials, namely colloidosomes (using colloidal particles for Pickering stabilization and fusing them to form a permeable shell). Herein, we describe for the first time a one-step emulsion-based technique that permits the assembly of metal-organic framework (MOF) faceted polyhedral BBs (i.e., cubes instead of spheres) into 3D hollow superstructures (or "colloidosomes" ). The shell of each resultant hollow MOF colloidosome is constructed from a monolayer of cubic BBs, whose dimensions can be precisely controlled by varying the amount of emulsifier used in the synthesis. © 2013 American Chemical Society.

  1. Square-stepping exercise and fall risk factors in older adults: a single-blind, randomized controlled trial.

    Science.gov (United States)

    Shigematsu, Ryosuke; Okura, Tomohiro; Nakagaichi, Masaki; Tanaka, Kiyoji; Sakai, Tomoaki; Kitazumi, Suguru; Rantanen, Taina

    2008-01-01

    Decreased fitness of the lower extremities is a potentially modifiable fall risk factor. This study aimed to compare two exercise programs--square-stepping exercise (SSE), which is a low-cost indoor program, and walking--for improving the fitness of the lower extremities. We randomly allocated 68 community-dwelling older adults (age 65-74 years) to either the SSE or walking group (W group). During the 12-week regimen, the SSE group participated in 70-minute exercise sessions conducted twice a week at a local health center, and the W group participated in outdoor supervised walking sessions conducted weekly. The W group was instructed to increase the number of daily steps. Prior to and after the program, we obtained information on 11 physical performance tests for known fall risk factors and 3 self-reported scales. The fall incidence was followed-up for 8 months. At 12 weeks postregimen, significant differences were observed between the two exercise groups with respect to leg power (1 item), balance (2 items), agility (2 items), reaction time (2 items), and a self-reported scale (1 item); the SSE group demonstrated a marked improvement in the above-mentioned items with Group x Time interactions. Significant time effects were observed in the tests involving chair stands, functional reach, and standing up from a lying-down position without Group x Time interactions. During the follow-up period, the fall rates per person-year in the SSE and W groups were 23.4% and 33.3%, respectively (p =.31). Although further studies are required, SSE is apparently more effective than walking in reducing fall risk factors, and it appears that it may be recommended as a health promotion exercise in older adults.

  2. Single-Step Access to Long-Chain α,ω-Dicarboxylic Acids by Isomerizing Hydroxycarbonylation of Unsaturated Fatty Acids

    KAUST Repository

    Goldbach, Verena

    2016-11-09

    Dicarboxylic acids are compounds of high value, but to date long-chain alpha,omega-dicarboxylic acids have been difficult to access in a direct way. Unsaturated fatty acids are ideal starting materials with their molecular structure of long methylene sequences and a carboxylate functionality, in addition to a double bond that offers itself for functionalization. Within this paper, we established a direct access to alpha,omega-dicarboxylic acids by combining isomerization and selective terminal carbonylation of the internal double bond with water as a nucleophile on unsaturated fatty acids. We identified the key elements of this reaction: a homogeneous reaction mixture ensuring sufficient contact between all reactants and a catalyst system allowing for activation of the Pd precursor under aqueous conditions. Experiments under pressure reactor conditions with [(dtbpx)Pd(OTf)(2)] as catalyst precursor revealed the importance of nucleophile and reactant concentrations and the addition of the diprotonated diphosphine ligand (dtbpxH(2))(OTf)(2) to achieve turnover numbers >120. A variety of unsaturated fatty acids, including a triglyceride, were converted to valuable long-chain dicarboxylic acids with high turnover numbers and selectivities for the linear product of >90%. We unraveled the activation pathway of the Pd-II precursor, which proceeds via a reductive elimination step forming a Pd species and oxidative addition of the diprotonated diphosphine ligand, resulting in the formation of the catalytically active Pd hydride species. Theoretical calculations identified the hydrolysis as the rate-determining step. A low nucleophile concentration in the reaction mixture in combination with this high energetic barrier limits the potential of this reaction. In conclusion, water can be utilized as a nucleophile in isomerizing functionalization reactions and gives access to long-chain dicarboxylic acids from a variety of unsaturated substrates. The activity of the catalytic

  3. Three-phase partitioning as a rapid and easy method for the purification and recovery of catalase from sweet potato tubers (Solanum tuberosum).

    Science.gov (United States)

    Duman, Yonca Avcı; Kaya, Erdem

    2013-07-01

    Three-phase partitioning (TPP) was used to purify and recover catalase from potato crude extract. The method consists of ammonium sulfate saturation, t-butanol addition, and adjustment of pH, respectively. The best catalase recovery (262 %) and 14.1-fold purification were seen in the interfacial phase in the presence of 40 % (w/v) ammonium sulfate saturation with 1.0:1.0 crude extract/t-butanol ratio (v/v) at pH 7 in a single step. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the enzyme showed comparatively purification and protein molecular weight was nearly found to be 56 kDa. This study shows that TPP is a simple, economical, and quick method for the recovering of catalase and can be used for the purification process.

  4. A rapid automated procedure for laboratory and shipboard spectrophotometric measurements of seawater alkalinity: continuously monitored single-step acid additions

    Science.gov (United States)

    Liu, X.; Byrne, R. H.; Lindemuth, M.; Easley, R. A.; Patsavas, M. C.

    2012-12-01

    An automated system for shipboard and laboratory alkalinity measurements is presented. The simple system, which consists of a Dosimat titrator to deliver acid volumetrically and a USB 4000 spectrophotometer to monitor the titration progress, provides fast, precise and accurate measurements of total alkalinity for oceanographic research. The analytical method is based on single-point HCl titrations of seawater samples of a known volume; bromol cresol purple is used as an indicator to determine the final pH. Field data from an Arctic cruise demonstrates accuracy and precision around 1 micro mol/kg and a sample processing rate of 6 min per sample.

  5. Microchannel-connected SU-8 honeycombs by single-step projection photolithography for positioning cells on silicon oxide nanopillar arrays

    International Nuclear Information System (INIS)

    Larramendy, Florian; Paul, Oliver; Blatche, Marie Charline; Mazenq, Laurent; Laborde, Adrian; Temple-Boyer, Pierre

    2015-01-01

    We report on the fabrication, functionalization and testing of SU-8 microstructures for cell culture and positioning over large areas. The microstructure consists of a honeycomb arrangement of cell containers interconnected by microchannels and centered on nanopillar arrays designed for promoting cell positioning. The containers have been dimensioned to trap single cells and, with a height of 50 µm, prevent cells from escaping. The structures are fabricated using a single ultraviolet photolithography exposure with focus depth in the lower part of the SU-8 resist. With optimized process parameters, microchannels of various aspect ratios are thus produced. The cell containers and microchannels serve for the organization of axonal growth between neurons. The roughly 2 µm-high and 500 nm-wide nanopillars are made of silicon oxide structured by deep reactive ion etching. In future work, beyond their cell positioning purpose, the nanopillars could be functionalized as sensors. The proof of concept of the novel microstructure for organized cell culture is given by the successful growth of interconnected PC12 cells. Promoted by the honeycomb geometry, a dense network of interconnections between the cells has formed and the intended intimate contact of cells with the nanopillar arrays was observed by scanning electron microscopy. This proves the potential of these new devices as tools for the controlled cell growth in an interconnected container system with well-defined 3D geometry. (paper)

  6. The Effect of Phosphoric Acid Pre-etching Times on Bonding Performance and Surface Free Energy with Single-step Self-etch Adhesives.

    Science.gov (United States)

    Tsujimoto, A; Barkmeier, W W; Takamizawa, T; Latta, M A; Miyazaki, M

    2016-01-01

    The purpose of this study was to evaluate the effect of phosphoric acid pre-etching times on shear bond strength (SBS) and surface free energy (SFE) with single-step self-etch adhesives. The three single-step self-etch adhesives used were: 1) Scotchbond Universal Adhesive (3M ESPE), 2) Clearfil tri-S Bond (Kuraray Noritake Dental), and 3) G-Bond Plus (GC). Two no pre-etching groups, 1) untreated enamel and 2) enamel surfaces after ultrasonic cleaning with distilled water for 30 seconds to remove the smear layer, were prepared. There were four pre-etching groups: 1) enamel surfaces were pre-etched with phosphoric acid (Etchant, 3M ESPE) for 3 seconds, 2) enamel surfaces were pre-etched for 5 seconds, 3) enamel surfaces were pre-etched for 10 seconds, and 4) enamel surfaces were pre-etched for 15 seconds. Resin composite was bonded to the treated enamel surface to determine SBS. The SFEs of treated enamel surfaces were determined by measuring the contact angles of three test liquids. Scanning electron microscopy was used to examine the enamel surfaces and enamel-adhesive interface. The specimens with phosphoric acid pre-etching showed significantly higher SBS and SFEs than the specimens without phosphoric acid pre-etching regardless of the adhesive system used. SBS and SFEs did not increase for phosphoric acid pre-etching times over 3 seconds. There were no significant differences in SBS and SFEs between the specimens with and without a smear layer. The data suggest that phosphoric acid pre-etching of ground enamel improves the bonding performance of single-step self-etch adhesives, but these bonding properties do not increase for phosphoric acid pre-etching times over 3 seconds.

  7. Single-Step Assembly of Multi-Modal Imaging Nanocarriers: MRI and Long-Wavelength Fluorescence Imaging

    Science.gov (United States)

    Pinkerton, Nathalie M.; Gindy, Marian E.; Calero-DdelC, Victoria L.; Wolfson, Theodore; Pagels, Robert F.; Adler, Derek; Gao, Dayuan; Li, Shike; Wang, Ruobing; Zevon, Margot; Yao, Nan; Pacheco, Carlos; Therien, Michael J.; Rinaldi, Carlos; Sinko, Patrick J.

    2015-01-01

    MRI and NIR-active, multi-modal Composite NanoCarriers (CNCs) are prepared using a simple, one-step process, Flash NanoPrecipitation (FNP). The FNP process allows for the independent control of the hydrodynamic diameter, co-core excipient and NIR dye loading, and iron oxide-based nanocrystal (IONC) content of the CNCs. In the controlled precipitation process, 10 nm IONCs are encapsulated into poly(ethylene glycol) stabilized CNCs to make biocompatible T2 contrast agents. By adjusting the formulation, CNC size is tuned between 80 and 360 nm. Holding the CNC size constant at an intensity weighted average diameter of 99 ± 3 nm (PDI width 28 nm), the particle relaxivity varies linearly with encapsulated IONC content ranging from 66 to 533 mM-1s-1 for CNCs formulated with 4 to 16 wt% IONC. To demonstrate the use of CNCs as in vivo MRI contrast agents, CNCs are surface functionalized with liver targeting hydroxyl groups. The CNCs enable the detection of 0.8 mm3 non-small cell lung cancer metastases in mice livers via MRI. Incorporating the hydrophobic, NIR dye PZn3 into CNCs enables complementary visualization with long-wavelength fluorescence at 800 nm. In vivo imaging demonstrates the ability of CNCs to act both as MRI and fluorescent imaging agents. PMID:25925128

  8. Functionalized TiO2nanoparticles by single-step hydrothermal synthesis: the role of the silane coupling agents.

    Science.gov (United States)

    Dalod, Antoine R M; Henriksen, Lars; Grande, Tor; Einarsrud, Mari-Ann

    2017-01-01

    A simple, robust and versatile hydrothermal synthesis route to in situ functionalized TiO 2 nanoparticles was developed using titanium(IV) isopropoxide as Ti-precursor and selected silane coupling agents (3-aminopropyltriethoxysilane (APTES), 3-(2-aminoethylamino)propyldimethoxymethylsilane (AEAPS), and n -decyltriethoxysilane (DTES)). Spherical nanoparticles (ca. 9 nm) with narrow size distribution were obtained by using DTES or by synthesis performed without silane coupling agents. Rod-like nanoparticles along with 9 nm spherical nanoparticles were formed using aminosilane coupling agents because of a combination of oriented attachment of nanoparticles and specific adsorption of the aminosilane on crystallographic faces of anatase nanoparticles. The nanoparticles were functionalized in situ and became hydrophobic as silanes reacted to form covalent bonds on the surface of TiO 2 . The versatility of the aqueous synthesis route was demonstrated, and by selecting the type of silane coupling agent the surface properties of the TiO 2 nanoparticles could be tailored. This synthesis route has been further developed into a two-step synthesis to TiO 2 -SiO 2 core-shell nanoparticles. Combustion of the silane coupling agents up to 700 °C leads to the formation of a nanometric amorphous SiO 2 layer, preventing growth and phase transition of the in situ functionalized nanoparticles.

  9. Effect of different air-drying time on the microleakage of single-step self-etch adhesives

    Directory of Open Access Journals (Sweden)

    Horieh Moosavi

    2013-05-01

    Full Text Available Objectives This study evaluated the effect of three different air-drying times on microleakage of three self-etch adhesive systems. Materials and Methods Class I cavities were prepared for 108 extracted sound human premolars. The teeth were divided into three main groups based on three different adhesives: Opti Bond All in One (OBAO, Clearfil S3 Bond (CSB, Bond Force (BF. Each main group divided into three subgroups regarding the air-drying time: without application of air stream, following the manufacturer's instruction, for 10 sec more than manufacturer's instruction. After completion of restorations, specimens were thermocycled and then connected to a fluid filtration system to evaluate microleakage. The data were statistically analyzed using two-way ANOVA and Tukey-test (α = 0.05. Results The microleakage of all adhesives decreased when the air-drying time increased from 0 sec to manufacturer's instruction (p < 0.001. The microleakage of BF reached its lowest values after increasing the drying time to 10 sec more than the manufacturer's instruction (p < 0.001. Microleakage of OBAO and CSB was significantly lower compared to BF in all three drying time (p < 0.001. Conclusions Increasing in air-drying time of adhesive layer in one-step self-etch adhesives caused reduction of microleakage, but the amount of this reduction may be dependent on the adhesive components of self-etch adhesives.

  10. THE PURIFICATION OF HYPERTENSIN I

    Science.gov (United States)

    Skeggs, Leonard T.; Marsh, Walton H.; Kahn, Joseph R.; Shumway, Norman P.

    1954-01-01

    The purification of hypertensin I has been described. The final product which is four times as powerful a pressor agent as l-arterenol, is obtained with an over-all recovery of 40 per cent. The product consists of a single component in countercurrent distribution, having a nitrogen content of 15.97 per cent and a specific activity of 7050 Goldblatt units per mg. of N or 1125 units per mg. of solid. Acid hydrolysis and paper chromatography indicate in a preliminary fashion that there are about nine amino acids present in the intact polypeptide. PMID:13201713

  11. Rapid purification of circular DNA by triplex-mediated affinity capture

    Science.gov (United States)

    Ji, H.; Smith, L.M.

    1997-01-07

    A single-step capture of a target supercoiled double-stranded DNA molecule is accomplished by forming a local triple-helix among two strands of the supercoiled circular DNA and an oligonucleotide probe. The oligonucleotide is bound to an immobilizing support which facilitates the immobilization and purification of target DNA molecules. Non-target DNA molecules and other contaminating cellular material are easily removed by washing. The triple-helical structure is destabilized by raising the pH, leaving purified target DNA in the supernatant and reusable affinity capture oligonucleotide secured to the immobilizing support. 3 figs.

  12. Purification and some properties of pectinesterase from potato (Solanum tuberosum L.) alpha cultivar

    OpenAIRE

    Montañez Sáenz, Julio; Téllez, Alejandro; Garza, Heliodoro de la; Reyes, María de la Luz; Contreras-Esquivel, Juan Carlos; Aguilar, Cristóbal Noé

    2000-01-01

    Pectinesterase was extracted from potato alpha cultivar, purified and partially characterized The used protocol resulted in a 58.8-fold purification (51 850.2 units/mg protein) with 15.5% recovery of pectinesterase activity. The purified enzyme had a molecular weight of 27 kDa and its isoelectric point was around 4.5 with pH and temperature optima of 8.0 and 60°C, respectively. The purified enzyme had a single symmetric peak of specific activity after chromatographic steps. The homogeneity of...

  13. The Borexino purification system

    Science.gov (United States)

    Benziger, Jay

    2014-05-01

    Purification of 278 tons of liquid scintillator and 889 tons of buffer shielding for the Borexino solar neutrino detector is performed with a system of combined distillation, water extraction, gas stripping and filtration. The purification system removed K, U and Th by distillation of the pseudocumene solvent and the PPO fluor. Noble gases, Rn, Kr and Ar were removed by gas stripping. Distillation was also employed to remove optical impurities and reduce the attenuation of scintillation light. The success of the purification system has facilitated the first time real time detection of low energy solar neutrinos.

  14. Detection of 22 common leukemic fusion genes using a single-step multiplex qRT-PCR-based assay.

    Science.gov (United States)

    Lyu, Xiaodong; Wang, Xianwei; Zhang, Lina; Chen, Zhenzhu; Zhao, Yu; Hu, Jieying; Fan, Ruihua; Song, Yongping

    2017-07-25

    Fusion genes generated from chromosomal translocation play an important role in hematological malignancies. Detection of fusion genes currently employ use of either conventional RT-PCR methods or fluorescent in situ hybridization (FISH), where both methods involve tedious methodologies and require prior characterization of chromosomal translocation events as determined by cytogenetic analysis. In this study, we describe a real-time quantitative reverse transcription PCR (qRT-PCR)-based multi-fusion gene screening method with the capacity to detect 22 fusion genes commonly found in leukemia. This method does not require pre-characterization of gene translocation events, thereby facilitating immediate diagnosis and therapeutic management. We performed fluorescent qRT-PCR (F-qRT-PCR) using a commercially-available multi-fusion gene detection kit on a patient cohort of 345 individuals comprising 108 cases diagnosed with acute myeloid leukemia (AML) for initial evaluation; remaining patients within the cohort were assayed for confirmatory diagnosis. Results obtained by F-qRT-PCR were compared alongside patient analysis by cytogenetic characterization. Gene translocations detected by F-qRT-PCR in AML cases were diagnosed in 69.4% of the patient cohort, which was comparatively similar to 68.5% as diagnosed by cytogenetic analysis, thereby demonstrating 99.1% concordance. Overall gene fusion was detected in 53.7% of the overall patient population by F-qRT-PCR, 52.9% by cytogenetic prediction in leukemia, and 9.1% in non-leukemia patients by both methods. The overall concordance rate was calculated to be 99.0%. Fusion genes were detected by F-qRT-PCR in 97.3% of patients with CML, followed by 69.4% with AML, 33.3% with acute lymphoblastic leukemia (ALL), 9.1% with myelodysplastic syndromes (MDS), and 0% with chronic lymphocytic leukemia (CLL). We describe the use of a F-qRT-PCR-based multi-fusion gene screening method as an efficient one-step diagnostic procedure as an

  15. Comparison of single-step reverse transepithelial all-surface laser ablation (ASLA to alcohol-assisted photorefractive keratectomy

    Directory of Open Access Journals (Sweden)

    Aslanides IM

    2012-06-01

    Full Text Available Ioannis M Aslanides,1 Sara Padroni,1 Samuel Arba Mosquera,2 Antonis Ioannides,1 Achyut Mukherjee11Emmetropia Mediterranean Eye Institute, Heraklion, Crete, Greece; 2Schwind eye-tech-solutions GmbH, Kleinostheim, GermanyPurpose: To evaluate postoperative pain, corneal epithelial healing, development of corneal haze, refractive outcomes, and corneal aberrations in a novel one-step, modified transepithelial photorefractive keratectomy (PRK, termed All-surface laser ablation (ASLA, compared to conventional, alcohol-assisted PRK.Materials and methods: Sixty eyes of 30 myopic patients were prospectively recruited to a randomized fellow eye study. Patients underwent conventional alcohol-assisted PRK in one eye (control group and ASLA-modified transepithelial PRK in the other (30 eyes in each treatment arm. Primary endpoints were postoperative pain and haze scores at 1 day, 3 days, 1 week, and 1, 3, 6, and 12 months. Secondary endpoints included visual acuity at 1, 3, 6, and 12 months, corneal aberrations at 3, 6, and 12 months, and early and late onset haze. Refractive predictability, safety, and efficacy of the two methods were considered.Results: The average age of the cohort was 29 years (standard deviation [SD]: 9; range: 18–46, and the average spherical equivalent refractive error was -4.18 diopters (SD: 1.9. At 3 days after surgery, the average pain score was 64% lower in the ASLA group (P < 0.0005. At this point, 96% of ASLA eyes had no epithelial defect, whereas 43% in the alcohol-assisted group did not achieve complete epithelial healing, and required replacement of bandage contact lens. The haze level was consistently lower in the ASLA group at all time points from 1 to 6 months.Conclusion: This study shows that the ASLA technique may have a future role in refractive surgery, due to the fact that it offers faster epithelial healing, lower pain scores, and significantly less haze formation.Keywords: cornea, ASLA, PRK, alcohol

  16. Single step synthesis of ZnS quantum dots and their microstructure characterization and electrical transport below room temperature

    Science.gov (United States)

    Mukherjee, P. S.; Patra, S.; Chakraborty, G.; Pradhan, S. K.; Meikap, A. K.

    2016-09-01

    Low dimensional cubic phase ZnS quantum dots (QDs) are formed by mechanical alloying the stoichiometric mixture of Zn and S powders at room temperature. During milling process the primary mixed phase ZnS is formed at about 3.5 h of milling and strain less single phase (cubic) ZnS QDs are formed with ∼4.5 nm in size after 20 h of milling. Detailed microstructure study has been done by both Rietveld analysis of x-ray diffraction pattern and high resolution transmission electron microscope images. Dc resistivity decreases with increasing temperature which can be explained by three-dimensional hopping conduction mechanisms. Observed negative magnetoconductivity has been analyzed by wave function shrinkage model. Alternating current conductivity can be described by the correlated barrier hopping conduction mechanism. Analysis of complex impedance indicates that the grain boundary resistance is found to be dominating over the grain resistance. Relaxation behavior has been explained by the analysis of the electric modulus.

  17. Exposure of Salmonella enterica serovar Typhimurium to high level biocide challenge can select multidrug resistant mutants in a single step.

    Directory of Open Access Journals (Sweden)

    Rebekah N Whitehead

    Full Text Available Biocides are crucial to the prevention of infection by bacteria, particularly with the global emergence of multiply antibiotic resistant strains of many species. Concern has been raised regarding the potential for biocide exposure to select for antibiotic resistance due to common mechanisms of resistance, notably efflux.Salmonella enterica serovar Typhimurium was challenged with 4 biocides of differing modes of action at both low and recommended-use concentration. Flow cytometry was used to investigate the physiological state of the cells after biocide challenge. After 5 hours exposure to biocide, live cells were sorted by FACS and recovered. Cells recovered after an exposure to low concentrations of biocide had antibiotic resistance profiles similar to wild-type cells. Live cells were recovered after exposure to two of the biocides at in-use concentration for 5 hours. These cells were multi-drug resistant and accumulation assays demonstrated an efflux phenotype of these mutants. Gene expression analysis showed that the AcrEF multidrug efflux pump was de-repressed in mutants isolated from high-levels of biocide.These data show that a single exposure to the working concentration of certain biocides can select for mutant Salmonella with efflux mediated multidrug resistance and that flow cytometry is a sensitive tool for identifying biocide tolerant mutants. The propensity for biocides to select for MDR mutants varies and this should be a consideration when designing new biocidal formulations.

  18. Initial steps toward the realization of large area arrays of single photon counting pixels based on polycrystalline silicon TFTs

    Science.gov (United States)

    Liang, Albert K.; Koniczek, Martin; Antonuk, Larry E.; El-Mohri, Youcef; Zhao, Qihua; Jiang, Hao; Street, Robert A.; Lu, Jeng Ping

    2014-03-01

    The thin-film semiconductor processing methods that enabled creation of inexpensive liquid crystal displays based on amorphous silicon transistors for cell phones and televisions, as well as desktop, laptop and mobile computers, also facilitated the development of devices that have become ubiquitous in medical x-ray imaging environments. These devices, called active matrix flat-panel imagers (AMFPIs), measure the integrated signal generated by incident X rays and offer detection areas as large as ~43×43 cm2. In recent years, there has been growing interest in medical x-ray imagers that record information from X ray photons on an individual basis. However, such photon counting devices have generally been based on crystalline silicon, a material not inherently suited to the cost-effective manufacture of monolithic devices of a size comparable to that of AMFPIs. Motivated by these considerations, we have developed an initial set of small area prototype arrays using thin-film processing methods and polycrystalline silicon transistors. These prototypes were developed in the spirit of exploring the possibility of creating large area arrays offering single photon counting capabilities and, to our knowledge, are the first photon counting arrays fabricated using thin film techniques. In this paper, the architecture of the prototype pixels is presented and considerations that influenced the design of the pixel circuits, including amplifier noise, TFT performance variations, and minimum feature size, are discussed.

  19. Purification and characterization of alkaline proteases from aspergillus terreus

    International Nuclear Information System (INIS)

    Hussain, A.; Mannan, A.; Zubair, H.; Mirza, B.

    2010-01-01

    Proteases belong to an important class of enzymes known as hydrolases and catalyze hydrolysis of proteins. They act primarily to degrade proteins that are used for energy production and as biosynthetic precursors. In the following study, protease produced from Aspergillus terreus was found to be thermo stable and included in the category of alkaline serine and metallo protease. During partial purification, presence of enzyme in 60% (NH/sub 4/)/sub 2/SO/sub 4/ indicated small molecular weight polypeptide; later purification with Sephadex G-75 fractionation yielded a single proteolytic active molecule. At final purification step, the increase in specific activity of the enzyme was 7.5 fold with 23% yield. SDS-PAGE analysis revealed that alkaline protease of Aspergillus terreus is a monomer with approximate molecular weight of 35 kDa. Optimum pH for protease activity was found in the range of 7.5-11.0 (maximum at pH 8.5), thus apparently classified as an alkaline protease. The enzyme was thermo stable towards high temperature (60 deg. C), however it denatured irreversibly at 70 deg. C showing 80% loss of activity. The maximum proteolytic activity was found at 40 deg. C. The enzyme was effectively inhibited by PMSF, EDTA and urea whereas iodoacetamide and thiourea did not result in any loss in activity while cysteine was found to be activator molecule. The study with metal ions Mg/sup +2/, Mn/sup +2/ and Fe/sup +3/ (1 mM each) showed minute stimulatory effects on enzyme activity. Co/sup +2/ and Ca/sup +2/ (1 mM) had neither excitatory nor inhibitory effect while Hg/sup +2/ and Cu/sup +2/ (1 mM) slightly reduced the enzyme activity. (author)

  20. An Improved Single-Step Cloning Strategy Simplifies the Agrobacterium tumefaciens-Mediated Transformation (ATMT)-Based Gene-Disruption Method for Verticillium dahliae.

    Science.gov (United States)

    Wang, Sheng; Xing, Haiying; Hua, Chenlei; Guo, Hui-Shan; Zhang, Jie

    2016-06-01

    The soilborne fungal pathogen Verticillium dahliae infects a broad range of plant species to cause severe diseases. The availability of Verticillium genome sequences has provided opportunities for large-scale investigations of individual gene function in Verticillium strains using Agrobacterium tumefaciens-mediated transformation (ATMT)-based gene-disruption strategies. Traditional ATMT vectors require multiple cloning steps and elaborate characterization procedures to achieve successful gene replacement; thus, these vectors are not suitable for high-throughput ATMT-based gene deletion. Several advancements have been made that either involve simplification of the steps required for gene-deletion vector construction or increase the efficiency of the technique for rapid recombinant characterization. However, an ATMT binary vector that is both simple and efficient is still lacking. Here, we generated a USER-ATMT dual-selection (DS) binary vector, which combines both the advantages of the USER single-step cloning technique and the efficiency of the herpes simplex virus thymidine kinase negative-selection marker. Highly efficient deletion of three different genes in V. dahliae using the USER-ATMT-DS vector enabled verification that this newly-generated vector not only facilitates the cloning process but also simplifies the subsequent identification of fungal homologous recombinants. The results suggest that the USER-ATMT-DS vector is applicable for efficient gene deletion and suitable for large-scale gene deletion in V. dahliae.

  1. Two-step irradiance schedule versus single-dose cold compress for pain control during 5-aminolevulinic acid-based photodynamic therapy of condyloma acuminatum.

    Science.gov (United States)

    Shao, Xiaonan; Wang, Fang; Xu, Bin

    2017-12-01

    The purpose of this study was to investigate the effect of two-step irradiance schedule in relieving pain during PDT of CA. The study was a prospective, controlled trial of 141 CA patients who were randomly divided into Group A (two-step irradiance schedule) and Group B (single-dose cold compress). The numeric rating scales (NRS) of patients' pain were recorded at 5, 10, 15, and 20 minutes during each PDT. The efficacy of PDT and NRS scores of patients in the two groups were compared. There was no significant difference in gender, age, lesion site, and disease course between the two groups (P > 0.05). In addition, the cure rate of patients in the two groups was not significantly different (97.1% vs. 95.8%, χ 2  = 0.000, P = 1.000). However, the NRS scores at different time points and number of PDT sessions were significantly different (F = 198.233 and 165.224, respectively, P two-step irradiance schedule could more significantly reduce the patients' pain degree during treatment, especially for vulva and crissum. Lasers Surg. Med. 49:908-912, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  2. Surface chemical approach to single-step measurement of antibody in human serum using localized surface plasmon resonance biosensor on microtiter plate system.

    Science.gov (United States)

    Yamamichi, Junta; Ojima, Tetsunori; Iida, Mie; Yurugi, Kimiko; Imamura, Takeshi; Ashihara, Eishi; Kimura, Shinya; Maekawa, Taira

    2014-07-01

    In clinical settings, serum antibody levels serve as markers of pathology. For example, antibodies related to autoimmune diseases are among the conventional targets in laboratory tests. Simple clinical tests can improve the efficacy of laboratory practice. This study describes a single-step, wash-free technique for optically detecting antibodies in human serum through the localized surface plasmon resonance (LSPR) of gold nanoparticles. As a proof-of-concept experiment, the amount of antibiotin dissolved in human serum was measured with a LSPR-based biosensor in a wash-free manner using a conventional 96-well microtiter plate and a plate reader. For an efficient surface modification of biosensors, zwitterionic copolymer was used as a scaffold on the gold nanoparticle surface to immobilize antigen and blocking reagent. Single-step, wash-free measurement of antibiotin in human serum was successfully achieved. In addition, nonspecific responses from serum contents were significantly reduced because both the copolymer and hydrophilic antigen reagent that we employed were composed of poly(ethylene oxide) spacer. Comparative experiments of the antigen-antibody reaction in serum to that in buffered solution revealed that serum is a favorable environment for the biological reaction. In conclusion, our gold-nanoparticle-based LSPR method may provide a rapid and simple way to measure the amount of antibody in serum quantitatively in clinical practice.

  3. Microwave pyrolysis using self-generated pyrolysis gas as activating agent: An innovative single-step approach to convert waste palm shell into activated carbon

    Science.gov (United States)

    Yek, Peter Nai Yuh; Keey Liew, Rock; Shahril Osman, Mohammad; Chung Wong, Chee; Lam, Su Shiung

    2017-11-01

    Waste palm shell (WPS) is a biomass residue largely available from palm oil industries. An innovative microwave pyrolysis method was developed to produce biochar from WPS while the pyrolysis gas generated as another product is simultaneously used as activating agent to transform the biochar into waste palm shell activated carbon (WPSAC), thus allowing carbonization and activation to be performed simultaneously in a single-step approach. The pyrolysis method was investigated over a range of process temperature and feedstock amount with emphasis on the yield and composition of the WPSAC obtained. The WPSAC was tested as dye adsorbent in removing methylene blue. This pyrolysis approach provided a fast heating rate (37.5°/min) and short process time (20 min) in transforming WPS into WPSAC, recording a product yield of 40 wt%. The WPSAC was detected with high BET surface area (≥ 1200 m2/g), low ash content (recording high adsorption efficiency of 440 mg of dye/g. The desirable process features (fast heating rate, short process time) and the recovery of WPSAC suggest the exceptional promise of the single-step microwave pyrolysis approach to produce high-grade WPSAC from WPS.

  4. Development of a scientific torsional system experiment containing controlled single or dual-clearance non-linearities: Examination of step-responses

    Science.gov (United States)

    Krak, Michael D.; Singh, Rajendra

    2017-02-01

    The chief goal of this paper is to propose a new laboratory experiment that exhibits the step-response of a torsional system containing one or two controlled clearances. This work is motivated by the disadvantages of prior large-scale experiments which utilize production vehicle drivelines and their components with significant real-life complexities. The conceptual and physical design features, which include sizing, modal properties, excitation, and instrumentation, are discussed with the goal of creating a controlled experiment. Like prior literature, a step-down torque excitation is selected and all analyses are performed on the acceleration signals to observe vibro-impact in the time domain. Typical measurements (for both the single and dual-clearance configurations) exhibit rich non-linear behavior, including the double-sided impact regime and a time-varying oscillatory period. Additionally, new measurements are compared to predictions from simple reduced order non-linear models to verify the feasibility of the proposed experiment. Finally, the utility of this experiment is demonstrated by comparing its measurements to a prior large-scale experiment that accommodates a production vehicle clutch damper with multiple stages. The hardening and softening effects in both experiments are discussed in the context of double and single-sided impacts as well as the oscillatory periods that vary with time.

  5. Screening Test Procedure For Long Life Single Mode Step Index Separate Confinement Heterostructure Single Quantum Well (Sinsch-Sqw) Laser Diodes

    Science.gov (United States)

    Fritz, William J.

    1989-06-01

    An experiment was conducted to establish an effective burn-in and screening procedure for long life SINSCH-SQW laser diodes. The laser diodes were grown by MOCVD and processed with 20μ wide oxide defined stripes. The devices had a high reflective back facet coating with a small etalon bonded to a passivated front facet to ensure single mode operation. The laser diodes were bonded p-side up to copper heat sinks using indium solder. A total of 48 devices were selected prior to burn-in and were operated at 200mA constant current for 2000 hours at an average heat sink temperature of 55°C. The average initial output power was 45mW per device. At the end of the test, the output powers ranged from lmW to 85mW. Most of the devices with the low final power failed catastrophically within 24 hours from the start of the test. Many laser diodes showed very little change in output power while others degraded gradually by varying amounts. Failure analysis showed that failures were facet, bulk, or heat sink related. The temporal output power degradations (i.e., gradual degradation, etc.) can be explained by the identified failure mechanisms. From the test results and failure analysis, a screening strategy based on inspection and burn-in can be devised to reject devices that may fail early. Also, improvements in processing can provide potential yield improvements. After accounting for degradation related to processing, a very long material lifetime is predicted for these SINSCH-SQW laser diodes.

  6. Can a Single Session of a Community-Based Group Exercise Program Combining Step Aerobics and Bodyweight Resistance Exercise Acutely Reduce Blood Pressure?

    Directory of Open Access Journals (Sweden)

    Mendes Romeu

    2014-12-01

    Full Text Available This study aimed to analyze the acute effects of a single session of a community-based group exercise program combining step aerobics and bodyweight resistance exercise on blood pressure in healthy young adult women. Twentythree healthy young adult women (aged 31.57 ± 7.87 years participated in two experimental sessions (exercise and control in a crossover study design. Blood pressure was monitored before, immediately after and at 10, 20 and 30 min of recovery. The exercise session consisted of four phases: 1 a warm-up (5 min of dance aerobics; 2 aerobic exercise training (30 min of step aerobics; 3 resistance exercise training (six sets of 12 repetitions of three bodyweight exercises in a circuit mode, 10 min; and 4 a cool-down (5 min of breathing and flexibility exercises; totaling 50 min of duration. Systolic blood pressure after exercise was significantly lower compared to control at the 10th min (-10.83 ± 2.13 vs. -2.6 ± 2.13 mmHg; p = 0.009, 20th min (-11.26 ± 2.13 vs. -3.04 ± 2.13 mmHg; p = 0.009 and 30th min of recovery (-10.87 ± 2.39 vs. -0.48 ± 2.39 mmHg; p = 0.004. A single session of a community-based group exercise program combining step aerobics and bodyweight resistance exercise was effective in inducing significant post-exercise hypotension in healthy young adult women. This type of low-cost exercise interventions may have an important role in the prevention of cardiovascular diseases and in community health promotion.

  7. The development and evaluation of single cell suspension from wheat and barley as a model system; a first step towards functional genomics application.

    Science.gov (United States)

    Dong, Jing; Bowra, Steve; Vincze, Eva

    2010-11-05

    The overall research objective was to develop single cell plant cultures as a model system to facilitate functional genomics of monocots, in particular wheat and barley. The essential first step towards achieving the stated objective was the development of a robust, viable single cell suspension culture from both species. We established growth conditions to allow routine culturing of somatic cells in 24 well microtiter plate format. Evaluation of the wheat and barley cell suspension as model cell system is a multi step process. As an initial step in the evaluation procedure we chose to study the impact of selected abiotic stress elicitors at the physiological, biochemical and molecular level. We report the results of osmotic stress imposed by NaCl and PEG. As proline is an important osmoprotectant of the cereal cells, colorimetric assay for proline detection was developed for small volumes (200 μl). We performed RT-PCR experiments to study the change in the expression of the genes encoding Δ1-pyrroline-5-carboxylate synthetase (P5CS) and Δ1-pyrroline-5-carboxylate reductase (PC5R) in response to abiotic stress. We found differences between the wheat and barley suspension cultures, barley being more tolerant to the applied osmotic stresses. We suggested a model to explain the obtained differences in stress tolerance between the two species. The suspension cell cultures have proven useful for determining changes in proline concentration and expression level of genes (P5CS, P5CR) under various treatments and we suggest that the cells can be used as a model host system to study gene expression and regulation in monocots.

  8. Characterizing the Final Steps of Chromosomal Replication at the Single-molecule Level in the Model System Escherichia coli

    KAUST Repository

    Elshenawy, Mohamed M.

    2015-12-01

    In the circular Escherichia coli chromosome, two replisomes are assembled at the unique origin of replication and drive DNA synthesis in opposite directions until they meet in the terminus region across from the origin. Despite the difference in rates of the two replisomes, their arrival at the terminus is synchronized through a highly specialized system consisting of the terminator protein (Tus) bound to the termination sites (Ter). This synchronicity is mediated by the polarity of the Tus−Ter complex that stops replisomes from one direction (non-permissive face) but not the other (permissive face). Two oppositely oriented clusters of five Tus–Ters that each block one of the two replisomes create a “replication fork trap” for the first arriving replisome while waiting for the late arriving one. Despite extensive biochemical and structural studies, the molecular mechanism behind Tus−Ter polar arrest activity remained controversial. Moreover, none of the previous work provided answers for the long-standing discrepancy between the ability of Tus−Ter to permanently stop replisomes in vitro and its low efficiency in vivo. Here, I spearheaded a collaborative project that combined single-molecule DNA replication assays, X-ray crystallography and binding studies to provide a true molecular-level understanding of the underlying mechanism of Tus−Ter polar arrest activity. We showed that efficiency of Tus−Ter is determined by a head-to-head kinetic competition between rate of strand separation by the replisome and rate of rearrangement of Tus−Ter interactions during the melting of the first 6 base pairs of Ter. This rearrangement maintains Tus’s strong grip on the DNA and stops the advancing replisome from breaking into Tus−Ter central interactions, but only transiently. We further showed how this kinetic competition functions within the context of two mechanisms to impose permanent fork stoppage. The rate-dependent fork arrest activity of Tus

  9. Single-Step Arthroscopic Repair With Cell-Free Polymer-Based Scaffold in Osteochondral Lesions of the Talus: Clinical and Radiological Results.

    Science.gov (United States)

    Kanatlı, Ulunay; Eren, Ali; Eren, Toygun Kağan; Vural, Abdurrahman; Geylan, Dilan Ece; Öner, Ali Yusuf

    2017-09-01

    To report the clinical and radiological results of patients with talar osteochondral lesions who were treated by microfracture and cell-free scaffold implantation in a single-step arthroscopic surgery. Forty patients, treated with a single-step arthroscopic surgery, were evaluated in this single-center-based retrospective study. Patients with degenerative arthritis (n = 1), history of ankle fracture (n = 1), kissing lesions (n = 1), lower extremity deformity (n = 1), and lesions 10 mm depth) bone cysts were additionally treated with bone graft. Patients were evaluated clinically, using the American Orthopedic Foot and Ankle Society (AOFAS) hindfoot score. Radiological assessment was performed with magnetic resonance imaging, using the magnetic resonance observation of cartilage repair tissue (MOCART) score. Thirty-two patients with a mean age of 38 ± 12 years were evaluated. The mean defect size was 2.5 ± 0.8 cm 2 and the mean defect volume was 2.4 ± 1.9 cm 3 . The mean preoperative AOFAS score was 52.8 ± 13.9 and increased to 87.1 ± 11.1 postoperatively at the mean follow-up of 33.8 ± 14.0 months (P = .0001). A total of 84.4% of patients had good to excellent clinical scores. Clinical scores had no significant relation with age, lesion size, depth, or body mass index. The mean MOCART score was 64.2 ± 12.0. There was no significant correlation between the total MOCART and AOFAS scores (P = .123). A significant relation was found between the defect filling (the subgroup of the MOCART score) and the clinical outcomes (P = .0001, rho = 0.731). The arthroscopic scaffold implantation technique is a single-step, safe, and effective method for the treatment of talar osteochondral lesions with satisfactory clinical and radiological outcomes. Level IV, therapeutic case series. Copyright © 2017 Arthroscopy Association of North America. Published by Elsevier Inc. All rights reserved.

  10. Gel purification of RNA.

    Science.gov (United States)

    Nilsen, Timothy W

    2013-02-01

    For many applications, including size selection of RNAs and purification of in vitro transcription products, it is necessary to purify RNAs on a denaturing gel. This procedure describes how to purify transcripts that have been synthesized in vitro. It is useful for labeled or unlabeled RNAs when sufficient mass is present. It can also be used to isolate small RNAs. In general, RNA purification by denaturing gel electrophoresis is practical only when the size of the desired RNA is 600 nucleotides or less.

  11. Simultaneous determination of PPCPs, EDCs, and artificial sweeteners in environmental water samples using a single-step SPE coupled with HPLC-MS/MS and isotope dilution.

    Science.gov (United States)

    Tran, Ngoc Han; Hu, Jiangyong; Ong, Say Leong

    2013-09-15

    A high-throughput method for the simultaneous determination of 24 pharmaceuticals and personal care products (PPCPs), endocrine disrupting chemicals (EDCs) and artificial sweeteners (ASs) was developed. The method was based on a single-step solid phase extraction (SPE) coupled with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and isotope dilution. In this study, a single-step SPE procedure was optimized for simultaneous extraction of all target analytes. Good recoveries (≥ 70%) were observed for all target analytes when extraction was performed using Chromabond(®) HR-X (500 mg, 6 mL) cartridges under acidic condition (pH 2). HPLC-MS/MS parameters were optimized for the simultaneous analysis of 24 PPCPs, EDCs and ASs in a single injection. Quantification was performed by using 13 isotopically labeled internal standards (ILIS), which allows correcting efficiently the loss of the analytes during SPE procedure, matrix effects during HPLC-MS/MS and fluctuation in MS/MS signal intensity due to instrument. Method quantification limit (MQL) for most of the target analytes was below 10 ng/L in all water samples. The method was successfully applied for the simultaneous determination of PPCPs, EDCs and ASs in raw wastewater, surface water and groundwater samples collected in a local catchment area in Singapore. In conclusion, the developed method provided a valuable tool for investigating the occurrence, behavior, transport, and the fate of PPCPs, EDCs and ASs in the aquatic environment. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. A rapid, ratiometric, enzyme-free, and sensitive single-step miRNA detection using three-way junction based FRET probes

    Science.gov (United States)

    Luo, Qingying; Liu, Lin; Yang, Cai; Yuan, Jing; Feng, Hongtao; Chen, Yan; Zhao, Peng; Yu, Zhiqiang; Jin, Zongwen

    2018-03-01

    MicroRNAs (miRNAs) are single stranded endogenous molecules composed of only 18-24 nucleotides which are critical for gene expression regulating the translation of messenger RNAs. Conventional methods based on enzyme-assisted nucleic acid amplification techniques have many problems, such as easy contamination, high cost, susceptibility to false amplification, and tendency to have sequence mismatches. Here we report a rapid, ratiometric, enzyme-free, sensitive, and highly selective single-step miRNA detection using three-way junction assembled (or self-assembled) FRET probes. The developed strategy can be operated within the linear range from subnanomolar to hundred nanomolar concentrations of miRNAs. In comparison with the traditional approaches, our method showed high sensitivity for the miRNA detection and extreme selectivity for the efficient discrimination of single-base mismatches. The results reveal that the strategy paved a new avenue for the design of novel highly specific probes applicable in diagnostics and potentially in microscopic imaging of miRNAs in real biological environments.

  13. A high-yield, one-step synthesis of surfactant-free gold nanostars and numerical study for single-molecule SERS application

    Energy Technology Data Exchange (ETDEWEB)

    Chatterjee, S.; Ringane, A. B.; Arya, A.; Das, G. M.; Dantham, V. R., E-mail: dantham@iitp.ac.in; Laha, R. [Indian Institute of Technology Patna, Department of Physics (India); Hussian, S. [Indian Institute of Technology Patna, Department of Chemistry (India)

    2016-08-15

    We report a high-yield synthesis of star-shaped gold nanostructures in one step, using a new surfactant-free wet chemistry method. Compared to the existing reports, these nanostars were found to have longer and sharper spikes anchored uniformly on the surface of the spherical core, allowing at least a few hot spots irrespective of the incident light polarization. The average experimental values of core radius and spike length were found to be 88.5 and 72 nm, respectively. Using these values in numerical simulations, the local electric field enhancement (η) and localized surface plasmon resonance (LSPR) spectrum were obtained. Moreover, the single-molecule surface-enhanced Raman scattering (SERS) enhancement factor was found to vary from 10{sup 10} to 10{sup 13} depending on the excitation wavelengths. Our theoretical calculations suggest that these nanostructures can be used to fabricate efficient SERS-based biosensors for the detection of single molecules in real time and for predicting structural information of single molecules.

  14. Implementation of genomic recursions in single-step genomic best linear unbiased predictor for US Holsteins with a large number of genotyped animals.

    Science.gov (United States)

    Masuda, Y; Misztal, I; Tsuruta, S; Legarra, A; Aguilar, I; Lourenco, D A L; Fragomeni, B O; Lawlor, T J

    2016-03-01

    The objectives of this study were to develop and evaluate an efficient implementation in the computation of the inverse of genomic relationship matrix with the recursion algorithm, called the algorithm for proven and young (APY), in single-step genomic BLUP. We validated genomic predictions for young bulls with more than 500,000 genotyped animals in final score for US Holsteins. Phenotypic data included 11,626,576 final scores on 7,093,380 US Holstein cows, and genotypes were available for 569,404 animals. Daughter deviations for young bulls with no classified daughters in 2009, but at least 30 classified daughters in 2014 were computed using all the phenotypic data. Genomic predictions for the same bulls were calculated with single-step genomic BLUP using phenotypes up to 2009. We calculated the inverse of the genomic relationship matrix GAPY(-1) based on a direct inversion of genomic relationship matrix on a small subset of genotyped animals (core animals) and extended that information to noncore animals by recursion. We tested several sets of core animals including 9,406 bulls with at least 1 classified daughter, 9,406 bulls and 1,052 classified dams of bulls, 9,406 bulls and 7,422 classified cows, and random samples of 5,000 to 30,000 animals. Validation reliability was assessed by the coefficient of determination from regression of daughter deviation on genomic predictions for the predicted young bulls. The reliabilities were 0.39 with 5,000 randomly chosen core animals, 0.45 with the 9,406 bulls, and 7,422 cows as core animals, and 0.44 with the remaining sets. With phenotypes truncated in 2009 and the preconditioned conjugate gradient to solve mixed model equations, the number of rounds to convergence for core animals defined by bulls was 1,343; defined by bulls and cows, 2,066; and defined by 10,000 random animals, at most 1,629. With complete phenotype data, the number of rounds decreased to 858, 1,299, and at most 1,092, respectively. Setting up GAPY(-1

  15. Single-step synthesis process of Ti3SiC2 ohmic contacts on 4H-SiC by sputter-deposition of Ti

    International Nuclear Information System (INIS)

    Fashandi, H.; Andersson, M.; Eriksson, J.; Lu, J.; Smedfors, K.; Zetterling, C.-M.; Lloyd Spetz, A.; Eklund, P.

    2015-01-01

    We report a single-step procedure for growth of ohmic Ti 3 SiC 2 on 4H-SiC by sputter-deposition of Ti at 960 °C, based on the Ti–SiC solid-state reaction during deposition. X-ray diffraction and electron microscopy show the growth of interfacial Ti 3 SiC 2 . The as-deposited contacts are ohmic, in contrast to multistep processes with deposition followed by rapid thermal annealing. This procedure also offers the possibility of direct synthesis of oxygen-barrier capping layers before exposure to air, potentially improving contact stability in high-temperature and high-power devices

  16. One-step fabrication of single-crystalline ZnS nanotubes with a novel hollow structure and large surface area for photodetector devices

    Science.gov (United States)

    An, Qinwei; Meng, Xianquan; Xiong, Ke; Qiu, Yunlei; Lin, Weihua

    2017-03-01

    ZnS nanotubes (NTs) were successfully prepared via a one-step thermal evaporation process without using any templates. The resulting NTs were single crystalline and structurally uniform. Based on experimental analysis, a tube-growth vapor-liquid-solid process was proposed as the growth mechanism of ZnS NTs. A metal-semiconductor-metal full-nanostructured ultraviolet (UV) photodetector with ZnS NTs as the active layer, and Ag nanowires of low resistivity and high transmissivity as electrodes, was fabricated and characterized. The ZnS NT-based device displayed a high I on/I off ratio of up to ˜1.56 × 105 with a high response to UV incident light at low operation voltage. This work is a meaningful exploration for preparing other one-dimensional semiconductor NTs, and developing a high-performance and power-saving UV sensor.

  17. Multicolored Cd1-xZnxSe quantum dots with type-I core/shell structure: single-step synthesis and their use as light emitting diodes

    Science.gov (United States)

    Pu, Ying-Chih; Hsu, Yung-Jung

    2014-03-01

    We developed a single-step hot-injection process to synthesize Cd1-xZnxSe quantum dots (QDs) with tunable emission wavelengths. The multiple emission colors of the Cd1-xZnxSe QDs resulted from the variation in their compositions (x value) with the reaction time. Because of the higher reactivity of the Cd precursor, QDs whose composition was rich in CdSe were generated at the beginning of the reaction. As the reaction proceeded, the later-formed ZnSe shell was simultaneously alloyed with the core, giving rise to a progressive alloying treatment for the grown QDs. During the reaction period, the emission color of the Cd1-xZnxSe QDs shifted from red to orange, to yellow, to green and finally to blue. A light emitting diode (LED) composed of multilayers of ITO/poly(3,4-ethylenedioxythiophene):poly(4-styrenesulfonate)/poly(3-hexylthiophene) blended with Cd1-xZnxSe QDs/Al was fabricated to test the electroluminescence (EL) properties of the QDs. The EL results show high color purity for the emission from LED devices containing Cd1-xZnxSe QDs, revealing that the as-synthesized QDs can be easily processed and integrated into a light-emitting device without using a complicated procedure. The findings from the present work also demonstrate the advantage of using the current single-step synthetic approach to obtain a batch of Cd1-xZnxSe QDs that may emit different colors in prototype LEDs.We developed a single-step hot-injection process to synthesize Cd1-xZnxSe quantum dots (QDs) with tunable emission wavelengths. The multiple emission colors of the Cd1-xZnxSe QDs resulted from the variation in their compositions (x value) with the reaction time. Because of the higher reactivity of the Cd precursor, QDs whose composition was rich in CdSe were generated at the beginning of the reaction. As the reaction proceeded, the later-formed ZnSe shell was simultaneously alloyed with the core, giving rise to a progressive alloying treatment for the grown QDs. During the reaction period

  18. The single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction: twenty-something years on.

    Science.gov (United States)

    Chomczynski, Piotr; Sacchi, Nicoletta

    2006-01-01

    Since its introduction, the 'single-step' method has become widely used for isolating total RNA from biological samples of different sources. The principle at the basis of the method is that RNA is separated from DNA after extraction with an acidic solution containing guanidinium thiocyanate, sodium acetate, phenol and chloroform, followed by centrifugation. Under acidic conditions, total RNA remains in the upper aqueous phase, while most of DNA and proteins remain either in the interphase or in the lower organic phase. Total RNA is then recovered by precipitation with isopropanol and can be used for several applications. The original protocol, enabling the isolation of RNA from cells and tissues in less than 4 hours, greatly advanced the analysis of gene expression in plant and animal models as well as in pathological samples, as demonstrated by the overwhelming number of citations the paper gained over 20 years.

  19. Combination of electromembrane extraction and liquid-phase microextraction in a single step: Simultaneous group separation of acidic and basic drugs

    DEFF Research Database (Denmark)

    Huang, Chuixiu; Seip, Knut Fredrik; Gjelstad, Astrid

    2015-01-01

    Electromembrane extraction (EME) and liquid-phase microextraction (LPME) were combined in a single step for the first time to realize simultaneous and clear group separation of basic and acidic drugs. Using 2-nitrophenyl octyl ether as the supported liquid membrane (SLM) for EME and dihexyl ether...... as the SLM for LPME, basic and acidic drugs were extracted and separated simultaneously from a low pH sample by EME and LPME, respectively. After 15 min of extraction, basic drugs (citalopram and sertraline) were exhaustively extracted, whereas the recoveries for acidic drugs (ketoprofen and ibuprofen) were...... in the range of 76%-86%. Longer extraction time provided higher recoveries for the acidic drugs, but this somewhat deteriorated the group separation. Matrices effects from the coexisting acidic drugs/basic drugs were tested, and we observed that simultaneous EME/LPME was not affected by coexisting drugs...

  20. Single-Step Fabrication of Gd2O3@SiO2 Nanoparticles for use as MRI Contrast Agents by Pulsed Laser Ablation in Liquid

    Science.gov (United States)

    Luo, Ning-Qi; Huang, Zhan-Yun; Li, Li; Shao, Yuan-Zhi; Chen, Di-Hu

    2013-03-01

    Gd2O3@SiO2 nanoparticles with a core-shell structure are synthesized by pulsed laser ablation in liquid (PLAL) in single steps. A Gd2O3 target immersed in tetraethyl orthosilicate (TEOS) is ablated by a microsecond Nd:YAG laser, which induces the generation of a Gd2O3 plasma plume and pyrolysis of the TEOS. We propose that the moment Gd2O3 nanoparticles are formed they will be coated immediately by SiO2 and directly synthesized Gd2O3@SiO2 core-shell nanoparticles. These particles obtain high r1 relaxivity of 5.26s-1mM-1 and are used as T1-weighted magnetic resonance imaging contrast agents. It is shown that the PLAL technique is promising for fabricating core-shell structure nanomaterial with potential medical applications.

  1. Magnetic Material Assessment of a Novel Ultra-High Step-Up Converter with Single Semiconductor Switch and Galvanic Isolation for Fuel-Cell Power System

    Directory of Open Access Journals (Sweden)

    Chih-Lung Shen

    2017-11-01

    Full Text Available In this paper, a novel step-up converter is proposed, which has the particular features of single semiconductor switch, ultra-high conversion ratio, galvanic isolation, and easy control. Therefore, the proposed converter is suitable for the applications of fuel-cell power system. Coupled inductors and switched capacitors are incorporated in the converter to obtain an ultra-high voltage ratio that is much higher than that of a conventional high step-up converter. Even if the turns ratio of coupled inductor and duty ratio are only to be 1 and 0.5, respectively, the converter can readily achieve a voltage gain of up to 18. Owing to this outstanding performance, it can also be applied to any other low voltage source for voltage boosting. In the power stage, only one active switch is used to handle the converter operation. In addition, the leakage energy of the two couple inductors can be totally recycled without any snubber, which simplifies the control mechanism and improves the conversion efficiency. Magnetic material dominates the conversion performance of the converter. Different types of iron cores are discussed for the possibility to serve as a coupled inductor. A 200 W prototype with 400 V output voltage is built to validate the proposed converter. In measurement, it indicates that the highest efficiency can be up to 94%.

  2. Selective Single-Step Separation of a Mixture of Three Metal Ions by a Triphasic Ionic-Liquid-Water-Ionic-Liquid Solvent Extraction System.

    Science.gov (United States)

    Vander Hoogerstraete, Tom; Blockx, Jonas; De Coster, Hendrik; Binnemans, Koen

    2015-08-10

    In a conventional solvent extraction system, metal ions are distributed between two immiscible phases, typically an aqueous and an organic phase. In this paper, the proof-of-principle is given for the distribution of metal ions between three immiscible phases, two ionic liquid phases with an aqueous phase in between them. Three-liquid-phase solvent extraction allows separation of a mixture of three metal ions in a single step, whereas at least two steps are required to separate three metals in the case of two-liquid-phase solvent extraction. In the triphasic system, the lower organic phase is comprised of the ionic liquid betainium- or choline bis(trifluoromethylsulfonyl)imide, whereas the upper organic phase is comprised of the ionic liquid trihexyl(tetradecyl)phosphonium bis(trifluoromethylsulfonyl)imide. The triphasic system was used for the separation of a mixture of tin(II), yttrium(III), and scandium(III) ions. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Next Step for STEP

    Energy Technology Data Exchange (ETDEWEB)

    Wood, Claire [CTSI; Bremner, Brenda [CTSI

    2013-08-09

    The Siletz Tribal Energy Program (STEP), housed in the Tribe’s Planning Department, will hire a data entry coordinator to collect, enter, analyze and store all the current and future energy efficiency and renewable energy data pertaining to administrative structures the tribe owns and operates and for homes in which tribal members live. The proposed data entry coordinator will conduct an energy options analysis in collaboration with the rest of the Siletz Tribal Energy Program and Planning Department staff. An energy options analysis will result in a thorough understanding of tribal energy resources and consumption, if energy efficiency and conservation measures being implemented are having the desired effect, analysis of tribal energy loads (current and future energy consumption), and evaluation of local and commercial energy supply options. A literature search will also be conducted. In order to educate additional tribal members about renewable energy, we will send four tribal members to be trained to install and maintain solar panels, solar hot water heaters, wind turbines and/or micro-hydro.

  4. Purification of functionalized DNA origami nanostructures.

    Science.gov (United States)

    Shaw, Alan; Benson, Erik; Högberg, Björn

    2015-05-26

    The high programmability of DNA origami has provided tools for precise manipulation of matter at the nanoscale. This manipulation of matter opens up the possibility to arrange functional elements for a diverse range of applications that utilize the nanometer precision provided by these structures. However, the realization of functionalized DNA origami still suffers from imperfect production methods, in particular in the purification step, where excess material is separated from the desired functionalized DNA origami. In this article we demonstrate and optimize two purification methods that have not previously been applied to DNA origami. In addition, we provide a systematic study comparing the purification efficacy of these and five other commonly used purification methods. Three types of functionalized DNA origami were used as model systems in this study. DNA origami was patterned with either small molecules, antibodies, or larger proteins. With the results of our work we aim to provide a guideline in quality fabrication of various types of functionalized DNA origami and to provide a route for scalable production of these promising tools.

  5. Solid State Air Purification System

    Data.gov (United States)

    National Aeronautics and Space Administration — The solid state air purification project will explore feasibility of a new air purification system based on a liquid membrane, capable of purifying carbon dioxide...

  6. [Handling G-protein-coupled receptors: expression, purification and in vitro stabilization].

    Science.gov (United States)

    Banères, Jean-Louis; Mouillac, Bernard

    2012-10-01

    Among the different classes of integral membrane proteins, G protein-coupled receptors (GPCR) constitute the largest family. They are involved in most essential physiological functions and particularly play a key role in cell-to-cell communication and sensory signal transduction. They represent targets for approximately 30% of currently marketed drugs. In order to better understand their functioning, define their tridimensional structure and develop novel selective and efficient therapeutic compounds, it is crucial to purify these proteins for a full characterization. However, this biochemical step is not trivial since GPCR are present in membranes at very low levels and they require detergents to be extracted from their natural lipid environment and be handled as functional proteins. No universal strategy for GPCR production, purification and stabilization is currently available; each single GPCR possesses a unique set of physicochemical characteristics, preference for some detergents upon solubilization and specific conditions for purification. During the last decade, major breakthroughs regarding overexpression, purification and above all GPCR stabilization, thanks to amphipols and nanodiscs, opened very exciting perspectives for structural and dynamic investigations of these membrane proteins. The aim of this chapter is to provide an overview of the different aspects of GPCR handling. © 2012 médecine/sciences – Inserm / SRMS.

  7. Characterization and purification of proteins suitable for the production of antibodies against 'Ca. Liberibacter asiaticus'.

    Science.gov (United States)

    Liu, Huawei; Atta, Sagheer; Hartung, John S

    2017-11-01

    The citrus disease huanglongbing (HLB), which is caused by 'Candidatus Liberibacter asiaticus' (CaLas), is one of the most devastating pathogens of citrus, and with no effective method of control, poses a serious threat to citrus production throughout the world. In a previous study we described the production of single chain antibodies against several CaLas proteins that provide the basis for efficient and accurate detection of CaLas in citrus tissues. The isolation of a sufficient amount of purified antigen is a key step in the production of functional antibodies. The current report details purification procedures for six protein antigens used to select recombinant and produce polyclonal antibodies. These proteins include a flagellar biosynthesis protein (FlhA), a dinucleoside polyphosphate hydrolase (InvA), a portion of the major outer membrane protein (OmpA), a component of type IV pilus (CapB), the polysialic acid capsule expression protein (KpsA) and the outer membrane efflux protein (TolC). Results of purification under completely native or denatured conditions were not satisfactory. Therefore different hybrid purification conditions were optimized for each of the different proteins. The results of bioinformatic analysis also indicated that the six proteins contained a great diversity of potential antigenic epitopes, which varied in number, and that the antigenic clusters were not uniformly distributed throughout the proteins. The purified proteins are useful for the development of highly specific antibodies capable of differentiating specific strains of Liberibacter. Published by Elsevier Inc.

  8. Purification of methanol dehydrogenase from mouth methylotrophic bacteria of tropical region

    Directory of Open Access Journals (Sweden)

    Waturangi, D.

    2011-01-01

    Full Text Available Aims: Purification of methanol dehydrogenase (MDH from methylotrophic bacteria was conducted to obtain pure enzyme for further research and industrial applications due to the enzyme’s unique activity that catalyzes oxidation of methanol as an important carbon source in methylotrophic bacteria.Methodology and Results: The enzyme was screened from methylotrophic bacteria isolated from human mouth. Purification of this enzyme was conducted using ammonium sulphate precipitation followed by cation exchange chromatography. Two types of media were used to produce the enzymes: luria broth and standard mineral salts media (MSM. MSM produced MDH with higher specific activity than LB. Specific activity was also increased along with the purification steps. Application of ammonium sulphate increased the purity of enzyme and was more effective for the enzyme produced in LB. Using sepharose increased the enzyme activity 10 -57 folds.Conclusion, significant and impact of this study: With this, ammonium sulphate precipitation coupled with single cation exchange chromatographic system has been proved to provide sufficient purified of methanol dehydrogenase from methylotrophic bacteria origin of human mouth with high specific activity for further application.

  9. Single-Tube Reaction Using Perfluorocarbons: A Prerequisite Step Leading to the Whole-Slide In Situ Technique on Histopathological Slides.

    Directory of Open Access Journals (Sweden)

    Yi-Chang Chen

    Full Text Available Developing a robust, novel method for performing multiple reactions in a single tube is not only time- and cost-saving but also critical for future high-throughput whole-slide in situ techniques on diseased tissues. In this study, we introduce the use of perfluorocarbons and compound-coated magnetic particles to create pseudochambers in a single tube, allowing different reactions to be performed in different phases. Perfluorocarbons also serve as cell lysis buffer and polymerase chain reaction (PCR buffer owing to their highly penetrating, repellent and emulsifiable properties. Using this method, nucleic acids can be isolated and purified from various sample types and sizes, followed by PCR, real-time PCR, or multiplex PCR in the same tube. No incubation or enzyme digesting time is needed and the risk of cross-contamination is reduced. Tests can be performed in microemulsions (water-in-oil droplets containing sequence-specific captures and probes for further high-throughput detection. We present a simple, quick, and robust procedure as a prerequisite step to future high-throughput in situ techniques.

  10. Design and Implementation of a High Efficiency, Low Component Voltage Stress, Single-Switch High Step-Up Voltage Converter for Vehicular Green Energy Systems

    Directory of Open Access Journals (Sweden)

    Yu-En Wu

    2016-09-01

    Full Text Available In this study, a novel, non-isolated, cascade-type, single-switch, high step-up DC/DC converter was developed for green energy systems. An integrated coupled inductor and voltage lift circuit were applied to simplify the converter structure and satisfy the requirements of high efficiency and high voltage gain ratios. In addition, the proposed structure is controllable with a single switch, which effectively reduces the circuit cost and simplifies the control circuit. With the leakage inductor energy recovery function and active voltage clamp characteristics being present, the circuit yields optimizable conversion efficiency and low component voltage stress. After the operating principles of the proposed structure and characteristics of a steady-state circuit were analyzed, a converter prototype with 450 W, 40 V of input voltage, 400 V of output voltage, and 95% operating efficiency was fabricated. The Renesas MCU RX62T was employed to control the circuits. Experimental results were analyzed to validate the feasibility and effectiveness of the proposed system.

  11. Ultra-high-density 3D DNA arrays within nanoporous biocompatible membranes for single-molecule-level detection and purification of circulating nucleic acids

    Science.gov (United States)

    Aramesh, M.; Shimoni, O.; Fox, K.; Karle, T. J.; Lohrmann, A.; Ostrikov, K.; Prawer, S.; Cervenka, J.

    2015-03-01

    Extracellular nucleic acids freely circulating in blood and other physiologic fluids are important biomarkers for non-invasive diagnostics and early detection of cancer and other diseases, yet difficult to detect because they exist in very low concentrations and large volumes. Here we demonstrate a new broad-range sensor platform for ultrasensitive and selective detection of circulating DNA down to the single-molecule level. The biosensor is based on a chemically functionalized nanoporous diamond-like carbon (DLC) coated alumina membrane. The few nanometer-thick, yet perfect and continuous DLC-coating confers the chemical stability and biocompatibility of the sensor, allowing its direct application in biological conditions. The selective detection is based on complementary hybridization of a fluorescently-tagged circulating cancer oncomarker (a 21-mer nucleic acid) with covalently immobilized DNA on the surface of the membrane. The captured DNAs are detected in the nanoporous structure of the sensor using confocal scanning laser microscopy. The flow-through membrane sensor demonstrates broad-range sensitivity, spanning from 1015 molecules per cm2 down to single molecules, which is several orders of magnitude improvement compared to the flat DNA microarrays. Our study suggests that these flow-through type nanoporous sensors represent a new powerful platform for large volume sampling and ultrasensitive detection of different chemical biomarkers.Extracellular nucleic acids freely circulating in blood and other physiologic fluids are important biomarkers for non-invasive diagnostics and early detection of cancer and other diseases, yet difficult to detect because they exist in very low concentrations and large volumes. Here we demonstrate a new broad-range sensor platform for ultrasensitive and selective detection of circulating DNA down to the single-molecule level. The biosensor is based on a chemically functionalized nanoporous diamond-like carbon (DLC) coated

  12. Purification of lipases.

    Science.gov (United States)

    Taipa, M A; Aires-Barros, M R; Cabral, J M

    1992-11-01

    Interest on lipases from different sources (microorganisms, animals and plants) has markedly increased in the last decade due to the potential applications of lipases in industry and in medicine. Microbial and mammalian lipases have been purified to homogeneity, allowing the successful determination of their primary aminoacid sequence and, more recently, of the three-dimensional structure. The X-ray studies of pure lipases will enable the establishment of the structure-function relationships and contribute for a better understanding of the kinetic mechanisms of lipase action on hydrolysis, synthesis and group exchange of esters. This article reviews the separation and purification techniques that were used in the recovery of microbial, mammalian and plant lipases. Several purification procedures are analysed taking into account the sequence of the methods and the number of times each method is used. Novel purification methods based on liquid-liquid extraction, membrane processes and immunopurification are also reviewed.

  13. Purification of yeast alcohol dehydrogenase by using immobilized metal affinity cryogels

    Energy Technology Data Exchange (ETDEWEB)

    Akduman, Begüm [Chemistry Department, Adnan Menderes University, Aydın (Turkey); Uygun, Murat [Koçarlı Vocational and Training School, Adnan Menderes University, Aydın (Turkey); Uygun, Deniz Aktaş, E-mail: daktas@adu.edu.tr [Chemistry Department, Adnan Menderes University, Aydın (Turkey); Akgöl, Sinan [Biochemistry Department, Ege University, İzmir (Turkey); Denizli, Adil [Chemistry Department, Hacettepe University, Ankara (Turkey)

    2013-12-01

    In this study, poly(2-hydroxyethyl methacrylate–glycidylmethacrylate) [poly(HEMA–GMA)] cryogels were prepared by radical cryocopolymerization of HEMA with GMA as a functional comonomer and N,N′-methylene-bisacrylamide (MBAAm) as a crosslinker. Iminodiacetic acid (IDA) functional groups were attached via ring opening of the epoxy group on the poly(HEMA–GMA) cryogels and then Zn(II) ions were chelated with these structures. Characterization of cryogels was performed by FTIR, SEM, EDX and swelling studies. These cryogels have interconnected pores of 30–50 μm size. The equilibrium swelling degree of Zn(II) chelated poly(HEMA–GMA)-IDA cryogels was approximately 600%. Zn(II) chelated poly(HEMA–GMA)-IDA cryogels were used in the adsorption of alcohol dehydrogenase from aqueous solutions and adsorption was performed in continuous system. The effects of pH, alcohol dehydrogenase concentration, temperature, and flow rate on adsorption were investigated. The maximum amount of alcohol dehydrogenase adsorption was determined to be 9.94 mg/g cryogel at 1.0 mg/mL alcohol dehydrogenase concentration and in acetate buffer at pH 5.0 with a flow rate of 0.5 mL/min. Desorption of adsorbed alcohol dehydrogenase was carried out by using 1.0 M NaCI at pH 8.0 phosphate buffer and desorption yield was found to be 93.5%. Additionally, these cryogels were used for purification of alcohol dehydrogenase from yeast with a single-step. The purity of desorbed alcohol dehydrogenase was shown by silver-stained SDS–PAGE. This purification process can successfully be used for the purification of alcohol dehydrogenase from unclarified yeast homogenates and this work is the first report about the usage of the cryogels for purification of alcohol dehydrogenase. - Highlights: • Poly(HEMA–GMA) cryogels were synthesized by radical cryocopolymerization technique. • Prepared cryogels were functionalized with IDA, then Zn(II) ions were chelated to the cryogel. • Zn(II) chelated poly

  14. Purification of yeast alcohol dehydrogenase by using immobilized metal affinity cryogels

    International Nuclear Information System (INIS)

    Akduman, Begüm; Uygun, Murat; Uygun, Deniz Aktaş; Akgöl, Sinan; Denizli, Adil

    2013-01-01

    In this study, poly(2-hydroxyethyl methacrylate–glycidylmethacrylate) [poly(HEMA–GMA)] cryogels were prepared by radical cryocopolymerization of HEMA with GMA as a functional comonomer and N,N′-methylene-bisacrylamide (MBAAm) as a crosslinker. Iminodiacetic acid (IDA) functional groups were attached via ring opening of the epoxy group on the poly(HEMA–GMA) cryogels and then Zn(II) ions were chelated with these structures. Characterization of cryogels was performed by FTIR, SEM, EDX and swelling studies. These cryogels have interconnected pores of 30–50 μm size. The equilibrium swelling degree of Zn(II) chelated poly(HEMA–GMA)-IDA cryogels was approximately 600%. Zn(II) chelated poly(HEMA–GMA)-IDA cryogels were used in the adsorption of alcohol dehydrogenase from aqueous solutions and adsorption was performed in continuous system. The effects of pH, alcohol dehydrogenase concentration, temperature, and flow rate on adsorption were investigated. The maximum amount of alcohol dehydrogenase adsorption was determined to be 9.94 mg/g cryogel at 1.0 mg/mL alcohol dehydrogenase concentration and in acetate buffer at pH 5.0 with a flow rate of 0.5 mL/min. Desorption of adsorbed alcohol dehydrogenase was carried out by using 1.0 M NaCI at pH 8.0 phosphate buffer and desorption yield was found to be 93.5%. Additionally, these cryogels were used for purification of alcohol dehydrogenase from yeast with a single-step. The purity of desorbed alcohol dehydrogenase was shown by silver-stained SDS–PAGE. This purification process can successfully be used for the purification of alcohol dehydrogenase from unclarified yeast homogenates and this work is the first report about the usage of the cryogels for purification of alcohol dehydrogenase. - Highlights: • Poly(HEMA–GMA) cryogels were synthesized by radical cryocopolymerization technique. • Prepared cryogels were functionalized with IDA, then Zn(II) ions were chelated to the cryogel. • Zn(II) chelated poly

  15. Purification of a synthetic pterocarpanquinone by countercurrent chromatography

    International Nuclear Information System (INIS)

    Costa, Fernanda das Neves; Silva, Alcides Jose M. da; Domingos, Jorge L. de Oliveira; Costa, Paulo Roberto R.; Leitao, Gilda G.; Daher Netto, Chaquip

    2012-01-01

    Countercurrent chromatography (CCC) was employed as a useful, fast and economic alternative to conventional chromatography techniques for the purification of a synthetic pterocarpanquinone, LQB-118. The separation was performed in a two-step CCC with the solvent system hexanechloroform- methanol-water 2:1.5:5:2 in both steps. Traditional purification of these reaction products by silica gel column chromatography demanded a large amount of solvent and time, besides allowing the irreversible adsorption of the compound in the column. The use of 1 H NMR for the calculation of KD of target compound is proposed as an alternative for HPLC measurements. (author)

  16. Water purification in Borexino

    Energy Technology Data Exchange (ETDEWEB)

    Giammarchi, M. [Infn Milano (Italy); Balata, M.; Ioannucci, L.; Nisi, S. [Laboratori Nazionali del Gran Sasso (Italy); Goretti, A.; Ianni, A. [Princeton University (United States); Miramonti, L. [Dip. di Fisica dell' Università di Milano e Infn (Italy)

    2013-08-08

    Astroparticle Physics and Underground experiments searching for rare nuclear events, need high purity materials to act as detectors or detector shielding. Water has the advantage of being cheap, dense and easily available. Most of all, water can be purified to the goal of obatining a high level of radiopurity. Water Purification can be achieved by means of a combination of processes, including filtration, reverse osmosis, deionization and gas stripping. The Water Purification System for the Borexino experiment, will be described together with its main performances.

  17. Expression, purification and characterization of a novel double-sites mutant of the single-chain sweet-tasting protein monellin (MNEI) with both improved sweetness and stability.

    Science.gov (United States)

    Zheng, Weiwei; Yang, Liu; Cai, Chenggu; Ni, Jinfeng; Liu, Bo

    2018-03-01

    The sweet protein monellin has high sweet potency with limited stability. In this study, 3 double-sites mutants (E2N/E23A, E2N/Y65R and E23A/Y65R) of the single-chain monellin (MNEI) were constructed. The proteins were expressed in E. coli BL21 and purified to homogeneity by nickel affinity chromatography with yields above 10 mg/L cell culture. Introduction of a sweeter mutant E2N into E23A or Y65R (E2N/E23A and E2N/Y65R) led to about 3-fold increase of sweetness, while addition of a more stable mutant E23A into E2N or Y65R (E2N/E23A and E23A/Y65R) resulted in improved thermal stability (about 10 °C). The results indicate that residues E2 and E23 mediate the sweetness and thermal stability of the protein, respectively. Multiple mutations of different residues (E2N/E23A) led to an additive performance with both improved sweetness and stability, suggesting that the sweetness and stability could be modulated by the independent molecular mechanism. The sweeter and thermal stable variant has a potential in further industrial applications. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Expression and purification of a novel therapeutic single-chain variable fragment antibody against BNP from inclusion bodies of Escherichia coli.

    Science.gov (United States)

    Bu, Dawei; Zhou, Yuwei; Tang, Jian; Jing, Fang; Zhang, Wei

    2013-12-01

    Abnormal brain natriuretic peptide (BNP) secretion is regarded as the dominating mechanism of cerebral salt wasting syndrome (CSW), which results from a renal loss of sodium and water during intracranial disease leading to hyponatremia. Scale preparation of therapeutic single-chain variable fragment (scFv) that can neutralize elevated circulating BNP may have potential value for clinical use. In this report, we used a recently isolated humanized anti-BNP scFv fragment (3C1) as model antibody (Ab) to evaluate the potential of scale production of this therapeutic protein. The truncated gene encoding for scFv fragment cloned in pET22b (+) was mainly overexpressed as inclusion bodies in Escherichia coli (E. coli) Rosetta (DE3) pLysS cells. The insoluble fragment was solubilized and purified by Ni-NTA agarose resin under denaturation conditions, and recovered via an effective refolding buffer containing 50 mM Tris-HCl, pH 8.0, 0.15 M NaCl, 1 mM EDTA, 0.5 M arginine, 2 mM GSH, 1 mM GSSG, and 5% glycerol. The refolded scFv fragment was concentrated by PEG20000, and dialyzed in PBS (containing 5% glycerol, pH 7.4). The final yield was approximately 10.2 mg active scFv fragment per liter of culture (3.4 g wet weight cells). The scFv fragment was more than 95% pure assessed by SDS-PAGE assay. Recombinant scFv fragment with His tag displayed its immunoreactivity with anti-His tag Ab by western blotting. ELISA showed the scFv fragment specifically bound to BNP, and it displayed similar activity as the traditional anti-BNP monoclonal Ab (mAb). Thus, the current strategy allows convenient small-scale production of this therapeutic protein. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. Secretory immunoglobulin purification from whey by chromatographic techniques.

    Science.gov (United States)

    Matlschweiger, Alexander; Engelmaier, Hannah; Himmler, Gottfried; Hahn, Rainer

    2017-08-15

    Secretory immunoglobulins (SIg) are a major fraction of the mucosal immune system and represent potential drug candidates. So far, platform technologies for their purification do not exist. SIg from animal whey was used as a model to develop a simple, efficient and potentially generic chromatographic purification process. Several chromatographic stationary phases were tested. A combination of two anion-exchange steps resulted in the highest purity. The key step was the use of a small-porous anion exchanger operated in flow-through mode. Diffusion of SIg into the resin particles was significantly hindered, while the main impurities, IgG and serum albumin, were bound. In this step, initial purity was increased from 66% to 89% with a step yield of 88%. In a second anion-exchange step using giga-porous material, SIg was captured and purified by step or linear gradient elution to obtain fractions with purities >95%. For the step gradient elution step yield of highly pure SIg was 54%. Elution of SIgA and SIgM with a linear gradient resulted in a step yield of 56% and 35%, respectively. Overall yields for both anion exchange steps were 43% for the combination of flow-through and step elution mode. Combination of flow-through and linear gradient elution mode resulted in a yield of 44% for SIgA and 39% for SIgM. The proposed process allows the purification of biologically active SIg from animal whey in preparative scale. For future applications, the process can easily be adopted for purification of recombinant secretory immunoglobulin species. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Single-step preparation of selected biological fluids for the high performance liquid chromatographic analysis of fat-soluble vitamins and antioxidants.

    Science.gov (United States)

    Lazzarino, Giacomo; Longo, Salvatore; Amorini, Angela Maria; Di Pietro, Valentina; D'Urso, Serafina; Lazzarino, Giuseppe; Belli, Antonio; Tavazzi, Barbara

    2017-12-08

    Fat-soluble vitamins and antioxidants are of relevance in health and disease. Current methods to extract these compounds from biological fluids mainly need use of multi-steps and multi organic solvents. They are time-consuming and difficult to apply to treat simultaneously large sample number. We here describe a single-step, one solvent extraction of fat-soluble vitamins and antioxidants from biological fluids, and the chromatographic separation of all-trans-retinoic acid, 25-hydroxycholecalciferol, all-trans-retinol, astaxanthin, lutein, zeaxanthin, trans-β-apo-8'-carotenal, γ-tocopherol, β-cryptoxanthin, α-tocopherol, phylloquinone, lycopene, α-carotene, β-carotene and coenzyme Q 10 . Extraction is obtained by adding one volume of biological fluid to two acetonitrile volumes, vortexing for 60s and incubating for 60min at 37°C under agitation. HPLC separation occurs in 30min using Hypersil C18, 100×4.6mm, 5μm particle size column, gradient from 70% methanol+30% H 2 O to 100% acetonitrile, flow rate of 1.0ml/min and 37°C column temperature. Compounds are revealed using highly sensitive UV-VIS diode array detector. The HPLC method suitability was assessed in terms of sensitivity, reproducibility and recovery. Using the present extraction and chromatographic conditions we obtained values of the fat-soluble vitamins and antioxidants in serum from 50 healthy controls similar to those found in literature. Additionally, the profile of these compounds was also measured in seminal plasma from 20 healthy fertile donors. Results indicate that this simple, rapid and low cost sample processing is suitable to extract fat-soluble vitamins and antioxidants from biological fluids and can be applied in clinical and nutritional studies. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. A single-step synthesis and the kinetic mechanism for monodisperse and hexagonal-phase NaYF4:Yb, Er upconversion nanophosphors.

    Science.gov (United States)

    Shan, Jingning; Ju, Yiguang

    2009-07-08

    A single-step synthesis for monodisperse and hexagonal-phase (beta) NaYF(4):Yb, Er upconversion nanophosphors (UCNPs) with a consistent hexagonal prism shape in the size range from 18 to 200 nm was achieved. The kinetic mechanisms for the particle phase transition and growth were examined. The beta-UCNPs were obtained via co-thermolysis of trifluoroacetate precursors in octadecene (ODE) with combined ligands of oleic acid (OA) and trioctylphosphine (TOP). The experimental results showed that the combined OA-TOP ligand was crucial for changing the surface energy and controlling the particle shape over a broad size range. It was found that the particle sizes could be controlled by varying the molar ratios of Na(CF(3)COO)/Re(CF(3)COO)(3) (Re = Y, Yb, and Er). A high Na/Re ratio accelerated the cubic-phase (alpha)-->beta transition and promoted the growth of smaller beta-UCNPs. The formation of beta-UCNPs was classified into kinetic and diffusion controlled stages, depending on the reaction temperature and the dominant crystalline phases formed in each stage. In stage I, 250-310 degrees C, NaF generation was the limiting step and alpha-UCNPs were formed via a 'burst of nucleation'. In stage II, above 310 degrees C, the alpha-UCNPs formed were re-dissolved and the growth of beta-UCNPs was a diffusion controlled process governed by the Gibbs-Thompson effect. A quasi-steady-state species assumption for NaF and a chemical potential equilibrium in the solution were introduced to explain the particle size dependence on Na/Re ratios. The study of UC luminescence showed that the UC intensity was proportional to the sizes of the beta-UCNPs.

  2. Single-molecule Imaging Analysis of Elementary Reaction Steps of Trichoderma reesei Cellobiohydrolase I (Cel7A) Hydrolyzing Crystalline Cellulose Iα and IIII*

    Science.gov (United States)

    Shibafuji, Yusuke; Nakamura, Akihiko; Uchihashi, Takayuki; Sugimoto, Naohisa; Fukuda, Shingo; Watanabe, Hiroki; Samejima, Masahiro; Ando, Toshio; Noji, Hiroyuki; Koivula, Anu; Igarashi, Kiyohiko; Iino, Ryota

    2014-01-01

    Trichoderma reesei cellobiohydrolase I (TrCel7A) is a molecular motor that directly hydrolyzes crystalline celluloses into water-soluble cellobioses. It has recently drawn attention as a tool that could be used to convert cellulosic materials into biofuel. However, detailed mechanisms of action, including elementary reaction steps such as binding, processive hydrolysis, and dissociation, have not been thoroughly explored because of the inherent challenges associated with monitoring reactions occurring at the solid/liquid interface. The crystalline cellulose Iα and IIII were previously reported as substrates with different crystalline forms and different susceptibilities to hydrolysis by TrCel7A. In this study, we observed that different susceptibilities of cellulose Iα and IIII are highly dependent on enzyme concentration, and at nanomolar enzyme concentration, TrCel7A shows similar rates of hydrolysis against cellulose Iα and IIII. Using single-molecule fluorescence microscopy and high speed atomic force microscopy, we also determined kinetic constants of the elementary reaction steps for TrCel7A against cellulose Iα and IIII. These measurements were performed at picomolar enzyme concentration in which density of TrCel7A on crystalline cellulose was very low. Under this condition, TrCel7A displayed similar binding and dissociation rate constants for cellulose Iα and IIII and similar fractions of productive binding on cellulose Iα and IIII. Furthermore, once productively bound, TrCel7A processively hydrolyzes and moves along cellulose Iα and IIII with similar translational rates. With structural models of cellulose Iα and IIII, we propose that different susceptibilities at high TrCel7A concentration arise from surface properties of substrate, including ratio of hydrophobic surface and number of available lanes. PMID:24692563

  3. Sodium purification in Rapsodie

    International Nuclear Information System (INIS)

    Giraud, B.

    1968-01-01

    This report is one of a series of publications presenting the main results of tests carried out during the start-up of the first french fast neutron reactor: Rapsodie. The article presents the sodium purification techniques used in the reactor cooling circuits both from the constructional point of view and with respect to results obtained during the first years working. (author) [fr

  4. Development and Evaluation of a Single-Step Duplex PCR for Simultaneous Detection of Fasciola hepatica and Fasciola gigantica (Family Fasciolidae, Class Trematoda, Phylum Platyhelminthes)

    Science.gov (United States)

    Nguyen, Khue Thi; Nguyen, Nga Thi Bich; Doan, Huong Thi Thanh; Le, Xuyen Thi Kim; Hoang, Chau Thi Minh; De, Nguyen Van

    2012-01-01

    A single-step multiplex PCR (here referred to as a duplex PCR) has been developed for simultaneous detection and diagnosis of Fasciola hepatica and F. gigantica. These species overlap in distribution in many countries of North and East Africa and Central and Southeast Asia and are similar in egg morphology, making identification from fecal samples difficult. Based on a comparative alignment of mitochondrial DNA (mtDNA) spanning the region of cox1-trnT-rrnL, two species-specific forward primers were designed, FHF (for F. hepatica) and FGF (for F. gigantica), and a single reverse primer, FHGR (common for both species). Conventional PCR followed by sequencing was applied using species-specific primer pairs to verify the specificity of primers and the identity of Fasciola DNA templates. Duplex PCR (using three primers) was used for testing with the DNA extracted from adult worms, miracidia, and eggs, producing amplicons of 1,031 bp for F. hepatica and 615 bp for F. gigantica. The duplex PCR failed to amplify from DNA of other common liver and intestinal trematodes, including two opisthorchiids, three heterophyids, an echinostomid, another fasciolid, and a taeniid cestode. The sensitivity assay showed that the duplex PCR limit of detection for each Fasciola species was between 0.012 ng and 0.006 ng DNA. Evaluation using DNA templates from 32 Fasciola samples (28 adults and 4 eggs) and from 25 field-collected stools of ruminants and humans revealed specific bands of the correct size and the presence of Fasciola species. This novel mtDNA duplex PCR is a sensitive and fast tool for accurate identification of Fasciola species in areas of distributional and zonal overlap. PMID:22692744

  5. Development and evaluation of a single-step duplex PCR for simultaneous detection of Fasciola hepatica and Fasciola gigantica (family Fasciolidae, class Trematoda, phylum Platyhelminthes).

    Science.gov (United States)

    Le, Thanh Hoa; Nguyen, Khue Thi; Nguyen, Nga Thi Bich; Doan, Huong Thi Thanh; Le, Xuyen Thi Kim; Hoang, Chau Thi Minh; De, Nguyen Van

    2012-08-01

    A single-step multiplex PCR (here referred to as a duplex PCR) has been developed for simultaneous detection and diagnosis of Fasciola hepatica and F. gigantica. These species overlap in distribution in many countries of North and East Africa and Central and Southeast Asia and are similar in egg morphology, making identification from fecal samples difficult. Based on a comparative alignment of mitochondrial DNA (mtDNA) spanning the region of cox1-trnT-rrnL, two species-specific forward primers were designed, FHF (for F. hepatica) and FGF (for F. gigantica), and a single reverse primer, FHGR (common for both species). Conventional PCR followed by sequencing was applied using species-specific primer pairs to verify the specificity of primers and the identity of Fasciola DNA templates. Duplex PCR (using three primers) was used for testing with the DNA extracted from adult worms, miracidia, and eggs, producing amplicons of 1,031 bp for F. hepatica and 615 bp for F. gigantica. The duplex PCR failed to amplify from DNA of other common liver and intestinal trematodes, including two opisthorchiids, three heterophyids, an echinostomid, another fasciolid, and a taeniid cestode. The sensitivity assay showed that the duplex PCR limit of detection for each Fasciola species was between 0.012 ng and 0.006 ng DNA. Evaluation using DNA templates from 32 Fasciola samples (28 adults and 4 eggs) and from 25 field-collected stools of ruminants and humans revealed specific bands of the correct size and the presence of Fasciola species. This novel mtDNA duplex PCR is a sensitive and fast tool for accurate identification of Fasciola species in areas of distributional and zonal overlap.

  6. Purification and characterization of bacteriocin like substance produced from bacillus lentus with perspective of a new biopreservative for food preservation

    International Nuclear Information System (INIS)

    Sharma, N.; Gautam, N.

    2009-01-01

    Molecular weight of bacteriocin like substance (BLIS) of a new strain of Bacillus lentus 121 was found to be approximately 11 kDa. Purification of BLIS was attained by single step gel exclusion chromatography. BLIS was characterized by studying the inhibitory spectrum. It was active at broad pH range, high temperature and high NaCl concentration and showed sensitivity to proteolytic enzymes like trypsin, alpha-chymotrypsin and papain, the characters desirable for food preservation. BLIS extended the shelf stability of milk upto 21 days as a biopreservative. (author)

  7. Application of single-step genomic best linear unbiased prediction with a multiple-lactation random regression test-day model for Japanese Holsteins.

    Science.gov (United States)

    Baba, Toshimi; Gotoh, Yusaku; Yamaguchi, Satoshi; Nakagawa, Satoshi; Abe, Hayato; Masuda, Yutaka; Kawahara, Takayoshi

    2017-08-01

    This study aimed to evaluate a validation reliability of single-step genomic best linear unbiased prediction (ssGBLUP) with a multiple-lactation random regression test-day model and investigate an effect of adding genotyped cows on the reliability. Two data sets for test-day records from the first three lactations were used: full data from February 1975 to December 2015 (60 850 534 records from 2 853 810 cows) and reduced data cut off in 2011 (53 091 066 records from 2 502 307 cows). We used marker genotypes of 4480 bulls and 608 cows. Genomic enhanced breeding values (GEBV) of 305-day milk yield in all the lactations were estimated for at least 535 young bulls using two marker data sets: bull genotypes only and both bulls and cows genotypes. The realized reliability (R 2 ) from linear regression analysis was used as an indicator of validation reliability. Using only genotyped bulls, R 2 was ranged from 0.41 to 0.46 and it was always higher than parent averages. The very similar R 2 were observed when genotyped cows were added. An application of ssGBLUP to a multiple-lactation random regression model is feasible and adding a limited number of genotyped cows has no significant effect on reliability of GEBV for genotyped bulls. © 2016 Japanese Society of Animal Science.

  8. Polymer Nanocomposite Film with Metal Rich Surface Prepared by In Situ Single-Step Formation of Palladium Nanoparticles: An Interesting Way to Combine Specific Functional Properties

    Directory of Open Access Journals (Sweden)

    David Thompson

    2016-10-01

    Full Text Available This paper presents a continuous single-step route that permits preparation of a thermostable polymer/metal nanocomposite film and to combine different functional properties in a unique material. More precisely, palladium nanoparticles are in situ generated in a polyimide matrix thanks to a designed curing cycle which is applied to a polyamic acid/metal precursor solution cast on a glass plate. A metal-rich surface layer which is strongly bonded to the bulk film is formed in addition to homogeneously dispersed metal nanoparticles. This specific morphology leads to obtaining an optically reflective film. The metal nanoparticles act as gas diffusion barriers for helium, oxygen, and carbon dioxide; they induce a tortuosity effect which allows dividing the gas permeation coefficients by a factor near to 2 with respect to the neat polyimide matrix. Moreover, the ability of the in situ synthesized palladium nanoparticles to entrap hydrogen is evidenced. The nanocomposite film properties can be modulated as a function of the location of the film metal-rich surface with respect to the hydrogen feed. The synthesized nanocomposite could represent a major interest for a wide variety of applications, from specific coatings for aerospace or automotive industry, to catalysis applications or sensors.

  9. A single-step synthesis of nitrogen-doped graphene sheets decorated with cobalt hydroxide nanoflakes for the determination of dopamine

    Directory of Open Access Journals (Sweden)

    Muhammad Mehmood Shahid

    2017-10-01

    Full Text Available Nitrogen-doped reduced graphene oxide (NrGO sheets decorated with Co(OH2 nanoflakes were prepared by a single-step hydrothermal process. The morphological and structural characterizations of as synthesized NrGO@Co(OH2 nanoflakes were performed by field emission scanning electron microscopy (FESEM, EDX-mapping and X-ray diffraction (XRD. NrGO@Co(OH2 nanoflakes modified glassy carbon electrode (GCE was used for electrochemical sensing of dopamine in neutral medium. The nanocomposite modified electrode showed enhanced electrochemical sensing ability for the detection of dopamine and the limit of detection (LoD was found to be 0.201 μM with a sensitivity value of 0.0286 ± 0.002 mA mM−1. Interference studies revealed that NrGO@Co(OH2─GCE endow excellent selectivity for DA detection even in the presence of higher concentration of common co-existing physiological interfering analytes. Additionally, proposed sensor demonstrated excellent performance in urine samples with promising reproducibility and stability. Keywords: Nitrogen doped graphene, Dopamine, Electrochemical sensor, Amperometric detection

  10. A Multi-Resolution Mode CMOS Image Sensor with a Novel Two-Step Single-Slope ADC for Intelligent Surveillance Systems

    Directory of Open Access Journals (Sweden)

    Daehyeok Kim

    2017-06-01

    Full Text Available In this paper, we present a multi-resolution mode CMOS image sensor (CIS for intelligent surveillance system (ISS applications. A low column fixed-pattern noise (CFPN comparator is proposed in 8-bit two-step single-slope analog-to-digital converter (TSSS ADC for the CIS that supports normal, 1/2, 1/4, 1/8, 1/16, 1/32, and 1/64 mode of pixel resolution. We show that the scaled-resolution images enable CIS to reduce total power consumption while images hold steady without events. A prototype sensor of 176 × 144 pixels has been fabricated with a 0.18 μm 1-poly 4-metal CMOS process. The area of 4-shared 4T-active pixel sensor (APS is 4.4 μm × 4.4 μm and the total chip size is 2.35 mm × 2.35 mm. The maximum power consumption is 10 mW (with full resolution with supply voltages of 3.3 V (analog and 1.8 V (digital and 14 frame/s of frame rates.

  11. Genetic analysis of allelic variants, single-step mutations, three allelic variants of the 15 STR loci in the population of Northeast Bosnia

    Directory of Open Access Journals (Sweden)

    Hadžiavdić Vesna

    2013-01-01

    Full Text Available Diversity of nuclear DNA microsatellite markers were analyzed in a reference sample of the population of northeast Bosnia. 437 samples taken from unrelated individuals were processed and three samples of paternity proof were shown. Detection effectiveness profile of the research, points to a valid choice of method of extraction, amplification and genotyping STR loci with PowerPlextm16. Genetic analysis of allelic variants of the 15 STR loci detected 17 samples determined as microvariants. Samples were divided into 15 different allelic variants at 7 different loci, and are: in locus D7S820, D16S539, D3S1358, D18S51, PENTA D, PENTA E and in locus vWA. Genetic analysis of mutations in cases of paternity determined three examples of single-step mutations in the loci FGA, Penta D and D3S1358. Genetic analysis of observed STR loci detected three allelic variant of genotype combination 7/10/11.3 in locus D7S820 Type II.

  12. One-step implementation of a hybrid Fredkin gate with quantum memories and single superconducting qubit in circuit QED and its applications

    Science.gov (United States)

    Liu, Tong; Guo, Bao-Qing; Yu, Chang-Shui; Zhang, Wei-Ning

    2018-02-01

    In a recent remarkable experiment [R. B. Patel et al., Science advances 2, e1501531 (2016)], a 3-qubit quantum Fredkin (i.e., controlled-SWAP) gate was demonstrated by using linear optics. Here we propose a simple experimental scheme by utilizing the dispersive interaction in superconducting quantum circuit to implement a hybrid Fredkin gate with a superconducting flux qubit as the control qubit and two separated quantum memories as the target qudits. The quantum memories considered here are prepared by the superconducting coplanar waveguide resonators or nitrogen-vacancy center ensembles. In particular, it is shown that this Fredkin gate can be realized using a single-step operation and more importantly, each target qudit can be in an arbitrary state with arbitrary degrees of freedom. Furthermore, we show that this experimental scheme has many potential applications in quantum computation and quantum information processing such as generating arbitrary entangled states (discrete-variable states or continuous-variable states) of the two memories, measuring the fidelity and the entanglement between the two memories. With state-of-the-art circuit QED technology, the numerical simulation is performed to demonstrate that two-memory NOON states, entangled coherent states, and entangled cat states can be efficiently synthesized.

  13. Biorecovered precious metals from industrial wastes: single-step conversion of a mixed metal liquid waste to a bioinorganic catalyst with environmental application.

    Science.gov (United States)

    Mabbett, Amanda N; Sanyahumbi, Douglas; Yong, Ping; Macaskie, Lynne E

    2006-02-01

    The complete and continuous reduction of 1 mM Cr(VI) to Cr(III) was achieved in a flow-through reactor using a novel bioinorganic catalyst ("MM-bio-Pd(0)"), which was produced by single-step reduction of platinum group metals (PGM) from industrial waste solution onto biomass of Desulfovibrio desulfuricans ATCC 29577. Two flow-through reactor systems were compared using both "MM-bioPd(0)" and chemically reduced Pd(0). Reactors containing the latter removed Cr(VI) for 1 week only at the expense of formate as the electron donor, whereas the former gave complete Cr(VI) removal for 3 months of continuous operation. Mass balance analysis showed 100% reduction of Cr(VI) to soluble Cr(III) in the bioreactor exit solution. With the use of electron paramagnetic resonance (EPR) no intermediate Cr(V) species could be detected. Pd(0) was biodeposited similarly using Escherichia coliMC4100 and "bio-Pd(0)". The latter was used to recover Pd(II) from two acidic industrial waste leachates to generate two types of "MM-bio-Pd(0)": "SI-bio-Pd(0)" and "SII-bio-Pd(0)", respectively. The biomaterial composition was comparable in both cases, and the catalytic activity was related inversely to the amount of chloride in the waste leachate from which it was derived.

  14. Quality CuInSe2 and Cu(In,Ga)Se2 thin films processed by single-step electrochemical deposition techniques

    Science.gov (United States)

    Papadimitriou, D.; Roupakas, G.; Sáez-Araoz, R.; Lux-Steiner, M.-Ch; Nickel, N. H.; Alamé, S.; Vogt, P.; Kneissl, M.

    2015-05-01

    Ternary CuInSe2 and quaternary Cu(In,Ga)Se2 chalcopyrite semiconductor films with potential applications as solar absorbers were deposited by single-step electrochemical deposition (ECD) on molybdenum coated glass substrates. The films have been structurally characterized by x-ray diffraction (XRD), scanning electron microscopy (SEM) combined with energy dispersive x-ray analysis (EDAX), x-ray photoelectron spectroscopy (XPS), and Raman spectroscopy. Chalcopyrite phase formation was confirmed already in as-deposited films. The crystal structure of the films was further improved by thermal treatment. Element interdiffusion at the chalcopyrite/Mo/glass interface has been prevented by retaining moderate temperatures of deposition (70 °C) and subsequent annealing (300 °C). The SEM/EDAX analysis revealed the presence of CuxSe secondary phases on the surface of ternary films and almost stoichiometric growth of quaternary deposited on top of ternary films. The XRD and Raman analysis confirmed the high quality assessment of the films being almost equal to that of chalcopyrite selenide layers grown by physical vapor deposition at high temperatures (550-750 °C). The surface sensitive XPS analysis confirmed the absence of other impurities in the ECD processed films except from oxygen and carbon adsorbents by sample exposure to atmospheric air.

  15. Development of a new Xe-133 single dose multi-step method (SDMM) for muscle blood flow measurement using gamma camera

    International Nuclear Information System (INIS)

    Bunko, Hisashi; Seto, Mikito; Taki, Junichi

    1985-01-01

    In order to measure the muscle blood flow (MBF) during exercise (Ex), a new Xe-133 single dose multi-step method (SDMM) for leg MBF measurement before, during and after Ex using gamma camera was developped. Theoretically, if the activity of Xe-133 in the muscle immediately before and after Ex are known, then the mean MBF during Ex can be calculated. In SDMM, these activities are corrected through correction formula using time delays between end of data aquisition (DA) at rest (R1) and begining of the Ex (TAB), and between end of Ex and begining of the DA after Ex (R2) (TDA). Validity of the SDMM and MBF response on mild and heavy Ex were evaluated in 11 normal volunteers. Ex MBF calculated from 5 and 2.5 min DA (5 sec/frame) both at R1 and R2 were highly correlated (r=.996). Ex MBF by SDMM and direct(measurement by fixed leg exercise were also highly correlated (r=.999). Reproducibility of the R1 and Ex MBF were excellent (r=.999). The highest MBF was seen in GCM on miled walking Ex and in VLM on heavy squatting Ex. After miled Ex, MBF rapidly returned to normal. After heavy Ex, MBF remaind high in VLM In conclusion, SDMM is simple and accurate method for evaluation of dynamic MBF response according to exercise. SDMM is also applicable to the field of sports medicine. (author)

  16. Mesoporous-activated carbon prepared from chitosan flakes via single-step sodium hydroxide activation for the adsorption of methylene blue.

    Science.gov (United States)

    Marrakchi, F; Ahmed, M J; Khanday, W A; Asif, M; Hameed, B H

    2017-05-01

    In this work, mesoporous-activated carbon (CSAC) was prepared from chitosan flakes (CS) via single-step sodium hydroxide activation for the adsorption of methylene blue (MB). CSAC was prepared using different impregnation ratios of NaOH:CS (1:1, 2:1, 3:1, and 4:1) at 800°C for 90min. The adsorption performance of CSAC was evaluated for MB at different adsorption variables, such MB initial concentrations (25-400mg/L), solution pH (3-11), and temperature (30-50°C). The adsorption isotherm data of CSAC-MB were well fitted to Langmuir model with a maximum adsorption capacity 143.53mg/g at 50°C. Best representation of kinetic data was obtained by the pseudo-second order model. CSAC exhibited excellent adsorption uptake for MB and can potentially be used for other cationic dyes. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Multicomponent Aqueous Synthesis of Iodo-1,2,3-triazoles: Single-Step Models for Dual Modification of Free Peptide and Radioactive Iodo Labeling.

    Science.gov (United States)

    Li, Lingjun; Ding, Shengqiang; Yang, Yanping; Zhu, Anlian; Fan, Xincui; Cui, Mengchao; Chen, Changpo; Zhang, Guisheng

    2017-01-23

    Iodo-1,2,3-triazoles are of considerable interest for chemical and biomedical applications. However, current synthetic methods for preparing iodo-1,2,3-triazoles cannot easily be applied to the direct modification of bioactive molecules in water. Through the combination of water-compatible oxidative iodination and the copper-catalyzed alkyne-azide cycloaddition reaction, a novel copper-catalyzed aqueous multicomponent synthetic method for the preparation of 5-iodo-1,2,3-triazoles has been developed. The method is highly effective and selective for substrates including biologically relevant compounds with nucleoside, sugar, and amino acid moieties. Based on this aqueous tandem reaction, a direct single-step multicomponent dual modification of peptide is developed from readily available starting materials. Furthermore, the method could also be applied to concise and fast multicomponent radioactive 125 I labeling from an aqueous solution of commercially available sodium 125 iodide as a starting material. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Genome-wide association mapping including phenotypes from relatives without genotypes in a single-step (ssGWAS for 6-week body weight in broiler chickens

    Directory of Open Access Journals (Sweden)

    Huiyu eWang

    2014-05-01

    Full Text Available The purpose of this study was to compare results obtained from various methodologies for genome-wide association studies, when applied to real data, in terms of number and commonality of regions identified and their genetic variance explained, computational speed, and possible pitfalls in interpretations of results. Methodologies include: two iteratively reweighted single-step genomic BLUP procedures (ssGWAS1 and ssGWAS2, a single-marker model (CGWAS, and BayesB. The ssGWAS methods utilize genomic breeding values (GEBVs based on combined pedigree, genomic and phenotypic information, while CGWAS and BayesB only utilize phenotypes from genotyped animals or pseudo-phenotypes. In this study, ssGWAS was performed by converting GEBVs to SNP marker effects. Unequal variances for markers were incorporated for calculating weights into a new genomic relationship matrix. SNP weights were refined iteratively. The data was body weight at 6 weeks on 274,776 broiler chickens, of which 4553 were genotyped using a 60k SNP chip. Comparison of genomic regions was based on genetic variances explained by local SNP regions (20 SNPs. After 3 iterations, the noise was greatly reduced of ssGWAS1 and results are similar to that of CGWAS, with 4 out of the top 10 regions in common. In contrast, for BayesB, the plot was dominated by a single region explaining 23.1% of the genetic variance. This same region was found by ssGWAS1 with the same rank, but the amount of genetic variation attributed to the region was only 3%. These finding emphasize the need for caution when comparing and interpreting results from various methods, and highlight that detected associations, and strength of association, strongly depends on methodologies and details of implementations. BayesB appears to overly shrink regions to zero, while overestimating the amount of genetic variation attributed to the remaining SNP effects. The real world is most likely a compromise between methods and remains to

  19. Two-Step Single Particle Mass Spectrometry for On-Line Monitoring of Polycyclic Aromatic Hydrocarbons Bound to Ambient Fine Particulate Matter

    Science.gov (United States)

    Zimmermann, R.; Bente, M.; Sklorz, M.

    2007-12-01

    Polycyclic aromatic hydrocarbons (PAH) are formed as trace products in combustion processes and are emitted to the atmosphere. Larger PAH have low vapour pressure and are predominantly bound to the ambient fine particulate matter (PM). Upon inhalation, PAH show both, chronic human toxicity (i.e. many PAH are potent carcinogens) as well as acute human toxicity (i.e. inflammatory effects due to oxi-dative stress) and are discussed to be relevant for the observed health effect of ambient PM. Therefore a better understanding of the occurrence, dynamics and particle size dependence of particle bound-PAH is of great interest. On-line aerosol mass spectrometry in principle is the method of choice to investigate the size resolved changes in the chemical speciation of particles as well the status of internal vs. external mixing of chemical constituents. However the present available aerosol mass spectrometers (ATOFMS and AMS) do not allow detection of PAH from ambient air PM. In order to allow a single particle based monitoring of PAH from ambient PM a new single particle laser ionisation mass spectrometer was built and applied. The system is based on ATOFMS principle but uses a two- step photo-ionization. A tracked and sized particle firstly is laser desorbed (LD) by a IR-laser pulse (CO2-laser, λ=10.2 μm) and subsequently the released PAH are selectively ionized by an intense UV-laser pulse (ArF excimer, λ=248 nm) in a resonance enhanced multiphoton ionisation process (REMPI). The PAH-ions are detected in a time of flight mass spectrometer (TOFMS). A virtual impactor enrichment unit is used to increase the detection frequency of the ambient particles. With the current inlet system particles from about 400 nm to 10 μm are accessible. Single particle based temporal profiles of PAH containing particles ion (size distribution and PAH speciation) have been recorded in Oberschleissheim, Germany from ambient air. Furthermore profiles of relevant emission sources (e

  20. Combination of granular activated carbon adsorption and deep-bed filtration as a single advanced wastewater treatment step for organic micropollutant and phosphorus removal.

    Science.gov (United States)

    Altmann, Johannes; Rehfeld, Daniel; Träder, Kai; Sperlich, Alexander; Jekel, Martin

    2016-04-01

    Adsorption onto granular activated carbon (GAC) is an established technology in water and advanced wastewater treatment for the removal of organic substances from the liquid phase. Besides adsorption, the removal of particulate matter by filtration and biodegradation of organic substances in GAC contactors has frequently been reported. The application of GAC as both adsorbent for organic micropollutant (OMP) removal and filter medium for solids retention in tertiary wastewater filtration represents an energy- and space saving option, but has rarely been considered because high dissolved organic carbon (DOC) and suspended solids concentrations in the influent of the GAC adsorber put a significant burden on this integrated treatment step and might result in frequent backwashing and unsatisfactory filtration efficiency. This pilot-scale study investigates the combination of GAC adsorption and deep-bed filtration with coagulation as a single advanced treatment step for simultaneous removal of OMPs and phosphorus from secondary effluent. GAC was assessed as upper filter layer in dual-media downflow filtration and as mono-media upflow filter with regard to filtration performance and OMP removal. Both filtration concepts effectively removed suspended solids and phosphorus, achieving effluent concentrations of 0.1 mg/L TP and 1 mg/L TSS, respectively. Analysis of grain size distribution and head loss within the filter bed showed that considerable head loss occurred in the topmost filter layer in downflow filtration, indicating that most particles do not penetrate deeply into the filter bed. Upflow filtration exhibited substantially lower head loss and effective utilization of the whole filter bed. Well-adsorbing OMPs (e.g. benzotriazole, carbamazepine) were removed by >80% up to throughputs of 8000-10,000 bed volumes (BV), whereas weakly to medium adsorbing OMPs (e.g. primidone, sulfamethoxazole) showed removals technologies are still largely unknown. Gabapentin showed

  1. Membrane concepts for blood purification : towards improved artificial kidney devices

    NARCIS (Netherlands)

    Tijink, M.S.L.

    2013-01-01

    The research presented in this thesis is about the fabrication and characterization of new membranes for blood purification. A novel membrane concept is proposed to combine diffusion and adsorption in one step to remove uremic retention solutes. A membrane with embedded functionalized particles, a

  2. Purification of bacteriocins using size-exclusion chromatography

    Directory of Open Access Journals (Sweden)

    Vivek K. Bajpai

    2016-06-01

    Full Text Available The bacteriocin purification involves following main steps. a. Extraction of cell-free-supernatant of bacteria. b. Ammonium sulfate precipitation. c. Dialysis. d. Diafiltration using PVP and e. Size-exclusion chromatography. However, depending on the nature of work, the compound could be further analyzed by reverse-phase HPLC, NMR, mass spectrometry and sequencing.

  3. Uranium hexafluoride purification

    International Nuclear Information System (INIS)

    Araujo, Eneas F. de

    1986-01-01

    Uranium hexafluoride might contain a large amount of impurities after manufacturing or handling. Three usual methods of purification of uranium hexafluoride were presented: selective sorption, sublimation, and distillation. Since uranium hexafluoride usually is contaminated with hydrogen fluoride, a theoretical study of the phase equilibrium properties was performed for the binary system UF 6 -HF. A large deviation from the ideal solution behaviour was observed. A purification unity based on a constant reflux batch distillation process was developed. A procedure was established in order to design the re boiler, condenser and packed columns for the UF 6 -HF mixture separation. A bench scale facility for fractional distillation of uranium hexafluoride was described. Basic operations for that facility and results extracted from several batches were discussed. (author)

  4. M13 bacteriophage purification using poly(ionic liquids) as alternative separation matrices.

    Science.gov (United States)

    Jacinto, Maria João; Patinha, David J S; Marrucho, Isabel M; Gonçalves, João; Willson, Richard C; Azevedo, Ana M; Aires-Barros, M Raquel

    2018-01-12

    M13 is a filamentous, non-lytic bacteriophage that infects Escherichia coli via the F pilus. Currently, phage M13 is widely used in phage display technology and bio-nanotechnology, and is considered a possible antibacterial therapeutic agent, among other applications. Conventional phage purification involves 5-7 operational steps, with high operational costs and significant product loss (approximately 60%). In this work, we propose a scalable purification process for M13 bacteriophage using a novel stationary phase based on a polymeric ionic liquid (PIL) with a positively charged backbone structure. Poly (1-vinyl-3-ethyl imidazolium bis(trifluoromethylsulfonyl) imide) - poly(VEIM-TFSI) predominantly acted as an anion exchanger under binding-elution mode. This revealed to be a rapid and simple method for the recovery of phage M13 with an overall separation yield of over 70% after a single downstream step. To the best of our knowledge, PILs have never been used as separation matrices for biological products and the results obtained, together with the large number of cations and anions available to prepare PILs, illustrate well the large potential of the proposed methodology. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Purification of crude glycerol from transesterification reaction of palm oil using direct method and multistep method

    Science.gov (United States)

    Nasir, N. F.; Mirus, M. F.; Ismail, M.

    2017-09-01

    Crude glycerol which produced from transesterification reaction has limited usage if it does not undergo purification process. It also contains excess methanol, catalyst and soap. Conventionally, purification method of the crude glycerol involves high cost and complex processes. This study aimed to determine the effects of using different purification methods which are direct method (comprises of ion exchange and methanol removal steps) and multistep method (comprises of neutralization, filtration, ion exchange and methanol removal steps). Two crude glycerol samples were investigated; the self-produced sample through the transesterification process of palm oil and the sample obtained from biodiesel plant. Samples were analysed using Fourier Transform Infrared Spectroscopy, Gas Chromatography and High Performance Liquid Chromatography. The results of this study for both samples after purification have showed that the pure glycerol was successfully produced and fatty acid salts were eliminated. Also, the results indicated the absence of methanol in both samples after purification process. In short, the combination of 4 purification steps has contributed to a higher quality of glycerol. Multistep purification method gave a better result compared to the direct method as neutralization and filtration steps helped in removing most excess salt, fatty acid and catalyst.

  6. One-step synthesis of single phase micro-sized BaFe12O19 hexaplates via a modified hydrothermal approach

    International Nuclear Information System (INIS)

    Cao, Liangliang; Zeng, Yanwei; Ding, Chuan; Li, Rongjie; Li, Chuanming; Zhang, Chengzhe

    2016-01-01

    Single phase BaFe 12 O 19 ferrite identified by X-ray diffraction and Raman spectroscopy has been successfully synthesized using Fe(NO 3 ) 3 ·9H 2 O and Ba(NO 3 ) 2 as starting materials and NaOH as a precipitant via a modified one-step hydrothermal approach which involves the elimination of carbonate radicals from reaction system based on the stoichiometric ratio of [Ba 2+ ]/[Fe 3+ ]. Hydrothermal products under various synthetic conditions were studied, including different addition amounts of Ba(NO 3 ) 2 in the modified operation, reaction temperatures and times, and hydroxyl concentrations. The BaFe 12 O 19 particles featuring an excellent hexagonal plates shape can be hydrothermally synthesized with the aid of polyethylene glycol. It has been found that the presence of α-Fe 2 O 3 in a traditional hydrothermal process is motivated by the deviation from the desired [Ba 2+ ]/[Fe 3+ ] ratio caused by the negligent precipitation of Ba 2+ ions to BaCO 3 . An investigation on the preferred hydrothermal product through thermodynamic calculation shows that the reduction in Gibbs free energy for the exclusive formation of BaFe 12 O 19 with 1 mol of Fe 3+ ions at 220 °C is approximately 32 kJ higher than that for the complete transformation to α-Fe 2 O 3 with an equal consumption quantity of Fe 3+ ions. - Highlights: • Pure BaFe 12 O 19 was hydrothermally synthesized based on the stoichiometric ratio. • A modified operation was employed to eliminate self-invited carbonate ions. • BaFe 12 O 19 particles feature an excellent micro-sized hexaplates shape. • BaFe 12 O 19 was thermodynamically confirmed to be preferred result instead of α-Fe 2 O 3 .

  7. Single-Step Incubation Determination of miRNAs in Cancer Cells Using an Amperometric Biosensor Based on Competitive Hybridization onto Magnetic Beads

    Directory of Open Access Journals (Sweden)

    Eva Vargas

    2018-03-01

    Full Text Available This work reports an amperometric biosensor for the determination of miRNA-21, a relevant oncogene. The methodology involves a competitive DNA-target miRNA hybridization assay performed on the surface of magnetic microbeads (MBs and amperometric transduction at screen-printed carbon electrodes (SPCEs. The target miRNA competes with a synthetic fluorescein isothiocyanate (FITC-modified miRNA with an identical sequence for hybridization with a biotinylated and complementary DNA probe (b-Cp immobilized on the surface of streptavidin-modified MBs (b-Cp-MBs. Upon labeling, the FITC-modified miRNA attached to the MBs with horseradish peroxidase (HRP-conjugated anti-FITC Fab fragments and magnetic capturing of the MBs onto the working electrode surface of SPCEs. The cathodic current measured at −0.20 V (versus the Ag pseudo-reference electrode was demonstrated to be inversely proportional to the concentration of the target miRNA. This convenient biosensing method provided a linear range between 0.7 and 10.0 nM and a limit of detection (LOD of 0.2 nM (5 fmol in 25 μL of sample for the synthetic target miRNA without any amplification step. An acceptable selectivity towards single-base mismatched oligonucleotides, a high storage stability of the b-Cp-MBs, and usefulness for the accurate determination of miRNA-21 in raw total RNA (RNAt extracted from breast cancer cells (MCF-7 were demonstrated.

  8. Clinical utility of simultaneous whole-body 18F-FDG PET/MRI as a single-step imaging modality in the staging of primary nasopharyngeal carcinoma.

    Science.gov (United States)

    Chan, Sheng-Chieh; Yeh, Chih-Hua; Yen, Tzu-Chen; Ng, Shu-Hang; Chang, Joseph Tung-Chieh; Lin, Chien-Yu; Yen-Ming, Tsang; Fan, Kang-Hsing; Huang, Bing-Shen; Hsu, Cheng-Lung; Chang, Kai-Ping; Wang, Hung-Ming; Liao, Chun-Ta

    2018-03-03

    Both head and neck magnetic resonance imaging (MRI) and 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET)/computed tomography (CT) play a crucial role in the staging of primary nasopharyngeal carcinoma (NPC). In this study, we sought to prospectively investigate the clinical utility of simultaneous whole-body 18F-FDG PET/MRI for primary staging of NPC patients. We examined 113 patients with histologically confirmed NPC who underwent pretreatment, simultaneous whole-body PET/MRI and PET/CT for primary tumor staging. The images obtained with the different imaging modalities were interpreted independently and compared with each other. PET/MRI increased the accuracy of head and neck MRI for assessment of primary tumor extent in four patients via addition of FDG uptake information to increase the conspicuity of morphologically subtle lesions. PET/MR images were more discernible than PET/CT images for mapping tumor extension, especially intracranial invasion. Regarding the N staging assessment, the sensitivity of PET/MRI (99.5%) was higher than that of head and neck MRI (94.2%) and PET/CT (90.9%). PET/MRI was particularly useful for distinguishing retropharyngeal nodal metastasis from adjacent nasopharyngeal tumors. For distant metastasis evaluation, PET/MRI exhibited a similar sensitivity (90% vs. 86.7% vs. 83.3%), but higher positive predictive value (93.1% vs. 78.8% vs. 83.3%) than whole-body MRI and PET/CT, respectively. For tumor staging of NPC, simultaneous whole-body PET/MRI was more accurate than head and neck MRI and PET/CT, and may serve as a single-step staging modality.

  9. Enhanced infrared transmittance properties in ultrafine MgAl2O4 nanoparticles synthesised by a single step combustion method, followed by hybrid microwave sintering

    Science.gov (United States)

    Mathew, C. T.; Vidya, S.; Koshy, Jacob; Solomon, Sam; Thomas, Jijimon K.

    2015-09-01

    Infrared transparent ceramics found to have potential applications as infrared windows and domes in strategic defence and space missions. Synthesis of ultrafine nanostructured MgAl2O4 ceramics by a modified single step auto-igniting combustion technique, followed by sintering of the sample by resistive and resistive-microwave hybrid heating to high density and their excellent infrared transmission characteristics are presented in this paper. Structural characterisations of MgAl2O4 nanoparticles reveal that the as prepared powder is phase pure, with average crystallite size ∼15 nm and possess a cubic structure. Optical band gap calculated using the Kubelka-Munk method is 5.75 eV. The thermal stability of the nanopowder at elevated temperatures has been studied using thermo gravimetric analysis (TGA) and differential thermal analysis (DTA). Hybrid heating yield a substantial reduction in sintering temperature and soaking time relative to the conventional resistive heating, and the samples achieved >99% density by microwave-resistive hybrid heating. Scanning electron micrograph (SEM) showed that the pellets are well sintered. The pellet sintered by hybrid heating showed a better transmittance of ∼79% in the UV-Visible region and ∼82% in the mid IR region compared to pellet sintered by resistive heating which has ∼68% in the UV-Visible region and ∼66% in the mid IR region. The results confirm the effective use of nanocrystalline powders from modified combustion synthesis as starting material for the development of high quality IR transparent windows and domes. In addition the microwave hybrid sintering technique employed in the present study also contributes to the results of better transmittance characteristics in highly densified MgAl2O4 ceramic pellets.

  10. Use of genomic recursions and algorithm for proven and young animals for single-step genomic BLUP analyses--a simulation study.

    Science.gov (United States)

    Fragomeni, B O; Lourenco, D A L; Tsuruta, S; Masuda, Y; Aguilar, I; Misztal, I

    2015-10-01

    The purpose of this study was to examine accuracy of genomic selection via single-step genomic BLUP (ssGBLUP) when the direct inverse of the genomic relationship matrix (G) is replaced by an approximation of G(-1) based on recursions for young genotyped animals conditioned on a subset of proven animals, termed algorithm for proven and young animals (APY). With the efficient implementation, this algorithm has a cubic cost with proven animals and linear with young animals. Ten duplicate data sets mimicking a dairy cattle population were simulated. In a first scenario, genomic information for 20k genotyped bulls, divided in 7k proven and 13k young bulls, was generated for each replicate. In a second scenario, 5k genotyped cows with phenotypes were included in the analysis as young animals. Accuracies (average for the 10 replicates) in regular EBV were 0.72 and 0.34 for proven and young animals, respectively. When genomic information was included, they increased to 0.75 and 0.50. No differences between genomic EBV (GEBV) obtained with the regular G(-1) and the approximated G(-1) via the recursive method were observed. In the second scenario, accuracies in GEBV (0.76, 0.51 and 0.59 for proven bulls, young males and young females, respectively) were also higher than those in EBV (0.72, 0.35 and 0.49). Again, no differences between GEBV with regular G(-1) and with recursions were observed. With the recursive algorithm, the number of iterations to achieve convergence was reduced from 227 to 206 in the first scenario and from 232 to 209 in the second scenario. Cows can be treated as young animals in APY without reducing the accuracy. The proposed algorithm can be implemented to reduce computing costs and to overcome current limitations on the number of genotyped animals in the ssGBLUP method. © 2015 Blackwell Verlag GmbH.

  11. Single-Step Fabrication Using a Phase Inversion Method of Poly(vinylidene fluoride) (PVDF) Activated Carbon Air Cathodes for Microbial Fuel Cells

    KAUST Repository

    Yang, Wulin

    2014-10-14

    Air cathodes used in microbial fuel cells (MFCs) need to have high catalytic activity for oxygen reduction, but they must also be easy to manufacture, inexpensive, and watertight. A simple one-step, phase inversion process was used here to construct an inexpensive MFC cathode using a poly(vinylidene fluoride) (PVDF) binder and an activated carbon catalyst. The phase inversion process enabled cathode preparation at room temperatures, without the need for additional heat treatment, and it produced for the first time a cathode that did not require a separate diffusion layer to prevent water leakage. MFCs using this new type of cathode produced a maximum power density of 1470 ± 50 mW m–2 with acetate as a substrate, and 230 ± 10 mW m–2 with domestic wastewater. These power densities were similar to those obtained using cathodes made using more expensive materials or more complex procedures, such as cathodes with a polytetrafluoroethylene (PTFE) binder and a poly(dimethylsiloxane) (PDMS) diffusion layer, or a Pt catalyst. Even though the PVDF cathodes did not have a diffusion layer, they withstood up to 1.22 ± 0.04 m of water head (∼12 kPa) without leakage, compared to 0.18 ± 0.02 m for cathodes made using PTFE binder and PDMS diffusion layer. The cost of PVDF and activated carbon ($3 m–2) was less than that of the stainless steel mesh current collector ($12 m–2). PVDF-based AC cathodes therefore are inexpensive, have excellent performance in terms of power and water leakage, and they can be easily manufactured using a single phase inversion process at room temperature.

  12. Single-step biosynthesis and characterization of silver nanoparticles using Zornia diphylla leaves: A potent eco-friendly tool against malaria and arbovirus vectors.

    Science.gov (United States)

    Govindarajan, Marimuthu; Rajeswary, Mohan; Muthukumaran, Udaiyan; Hoti, S L; Khater, Hanem F; Benelli, Giovanni

    2016-08-01

    Mosquitoes (Diptera: Culicidae) are vectors of important pathogens and parasites, including malaria, dengue, chikungunya, Japanese encephalitis, lymphatic filariasis and Zika virus. The application of synthetic insecticides causes development of resistance, biological magnification of toxic substances through the food chain, and adverse effects on the environment and human health. In this scenario, eco-friendly control tools of mosquito vectors are a priority. Here single-step fabrication of silver nanoparticles (AgNP) using a cheap aqueous leaf extract of Zornia diphylla as reducing and capping agent pf Ag(+) ions has been carried out. Biosynthesized AgNP were characterized by UV-visible spectrophotometry, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy-dispersive spectroscopy (EDX) and X-ray diffraction analysis (XRD). The acute toxicity of Z. diphylla leaf extract and biosynthesized AgNP was evaluated against larvae of the malaria vector Anopheles subpictus, the dengue vector Aedes albopictus and the Japanese encephalitis vector Culex tritaeniorhynchus. Both the Z. diphylla leaf extract and Ag NP showed dose dependent larvicidal effect against all tested mosquito species. Compared to the leaf aqueous extract, biosynthesized Ag NP showed higher toxicity against An. subpictus, Ae. albopictus, and Cx. tritaeniorhynchus with LC50 values of 12.53, 13.42 and 14.61μg/ml, respectively. Biosynthesized Ag NP were found safer to non-target organisms Chironomus circumdatus, Anisops bouvieri and Gambusia affinis, with the respective LC50 values ranging from 613.11 to 6903.93μg/ml, if compared to target mosquitoes. Overall, our results highlight that Z. diphylla-fabricated Ag NP are a promising and eco-friendly tool against larval populations of mosquito vectors of medical and veterinary importance, with negligible toxicity against other non-target organisms. Copyright © 2016 Elsevier B

  13. Process for recovery and purification of lithium from dilute sources

    International Nuclear Information System (INIS)

    Kunugita, Eiichi; Kim, Jong-Hwa; Komasawa, Isao

    1990-01-01

    A process was studied for recovery and purification of lithium from hot spring water and leachate of coal fly ash in which lithium is present at very low concentration in a mixture of high concentration of less desirable species such as alkali metals, alkaline earth metals and others. Lithium is, as a first step, adsorbed selectively by an ion-sieve type adsorbent, λ-MnO 2 , which can effect separation by adsorbing lithium alone in its intraparticle spaces while adsorbing most metals on its surface. Stagewise elutions are performed, employing two elutrients: NH 4 NO 3 and then HNO 3 solution. The concentration ratio of lithium and sodium is 1/70∼1/120 in the original feed stream, and this is upgraded to 300/1 in the pregnant elutrient. Some impurities in the pregnant elutrient is removed by a solvent extraction technique, employing a mixed extractant of a β-diketone (LIX51) and tri-n-octyl-phosphine oxide in kerosene. A lithium solution with an undetectable amount of impurities is obtained by single extraction followed by single scrubbing with dilute hydrochloric acid of carefully controlled concentration. (author)

  14. Development, modelling, optimisation and scale-up of chromatographic purification of a therapeutic protein

    DEFF Research Database (Denmark)

    Mollerup, Jørgen; Hansen, Thomas Budde; Kidal, Steffen

    2007-01-01

    optimisation of a purification step necessitate simulations and thus models of the adsorption isotherm. A model for ion-exchange is reviewed and the strategy for estimation of model parameters is reported. Examples are shown where computer simulations are used for development and optimisation......Development of a chromatographic purification step proceeds through a number of stages. High-throughput screening techniques are used to identify suitable resins. This technique is also suitable for the design of a capture step and some intermediate chromatographic steps, but development and true...

  15. Estimation of breeding values for uniformity of growth in Atlantic salmon (Salmo salar) using pedigree relationships or single-step genomic evaluation.

    Science.gov (United States)

    Sae-Lim, Panya; Kause, Antti; Lillehammer, Marie; Mulder, Han A

    2017-03-07

    In farmed Atlantic salmon, heritability for uniformity of body weight is low, indicating that the accuracy of estimated breeding values (EBV) may be low. The use of genomic information could be one way to increase accuracy and, hence, obtain greater response to selection. Genomic information can be merged with pedigree information to construct a combined relationship matrix ([Formula: see text] matrix) for a single-step genomic evaluation (ssGBLUP), allowing realized relationships of the genotyped animals to be exploited, in addition to numerator pedigree relationships ([Formula: see text] matrix). We compared the predictive ability of EBV for uniformity of body weight in Atlantic salmon, when implementing either the [Formula: see text] or [Formula: see text] matrix in the genetic evaluation. We used double hierarchical generalized linear models (DHGLM) based either on a sire-dam (sire-dam DHGLM) or an animal model (animal DHGLM) for both body weight and its uniformity. With the animal DHGLM, the use of [Formula: see text] instead of [Formula: see text] significantly increased the correlation between the predicted EBV and adjusted phenotypes, which is a measure of predictive ability, for both body weight and its uniformity (41.1 to 78.1%). When log-transformed body weights were used to account for a scale effect, the use of [Formula: see text] instead of [Formula: see text] produced a small and non-significant increase (1.3 to 13.9%) in predictive ability. The sire-dam DHGLM had lower predictive ability for uniformity compared to the animal DHGLM. Use of the combined numerator and genomic relationship matrix ([Formula: see text]) significantly increased the predictive ability of EBV for uniformity when using the animal DHGLM for untransformed body weight. The increase was only minor when using log-transformed body weights, which may be due to the lower heritability of scaled uniformity, the lower genetic correlation of transformed body weight with its

  16. Single-step transepithelial ASLA (SCHWIND with mitomycin-C for the correction of high myopia: long term follow-up

    Directory of Open Access Journals (Sweden)

    Aslanides IM

    2014-12-01

    Full Text Available Ioannis M Aslanides, Panagiotis N Georgoudis, Vasilis D Selimis, Achyut N Mukherjee Emmetropia Mediterranean Eye Institute, Heraklion, Crete, Greece Purpose: We wanted to compare the outcomes of single-step modified transepithelial photorefractive keratectomy (tPRK termed a SCHWIND all surface laser ablation (ASLA versus conventional alcohol-assisted photorefractive keratectomy (PRK and laser-assisted in situ keratomileusis (LASIK for the correction of higher myopia of 6.00 diopters (D or more, in an area with high risk of haze due to high intensity of sunlight.Methods: We used a prospective interventional cohort with matched retrospective control groups. Patients with >6 D myopia and <3.5 D of astigmatism were included. All treatments were performed with the SCHWIND Amaris system using aspheric ablation profiles. Mitomycin C was used in all PRK and ASLA cases. Outcomes were postoperative refraction, visual acuity, stability, and complications. The follow-up period was up to 12 months.Results: In total, 101 eyes were included after exclusions. Mean preoperative spherical equivalent refraction was −7.9 D, −8.2 D, and −7.4 D in the ASLA (n=41, PRK (n=29, and LASIK (n=31 groups. Mean postoperative spherical equivalent at 12 months postoperatively was −0.1 (standard deviation [SD]: 0.34, −0.2 (SD: 0.59, and −0.08 (SD: 0.36 in the ASLA, PRK, and LASIK groups, with 91.4%, 85.7%, and 83.9% within 0.5 D of target, respectively. Refractive outcomes and regression at 12 months did not vary among groups (P>0.05. Mean logMAR (logarithm of the minimum angle of resolution uncorrected distance visual acuity at 12 months was 0.00 (SD: 0.05, 0.06 (SD: 0.1, and 0.05 (SD: 0.09 in the ASLA, PRK, and LASIK groups, with significantly better vision in the tPRK group versus LASIK (P=0.01 and PRK (P=0.01 groups.Conclusion: ASLA (SCHWIND tPRK with mitomycin C for high myopia demonstrates comparable refractive outcomes to LASIK and PRK, with relatively

  17. A novel single-step GC-MS/MS method for cannabinoids and 11-OH-THC metabolite analysis in hair.

    Science.gov (United States)

    Angeli, Ilaria; Casati, Sara; Ravelli, Alessandro; Minoli, Mauro; Orioli, Marica

    2018-03-16

    THC, CBD, CBN, THC-COOH and 11-OH-THC are the most popular markers of cannabis consumption and abuse. The use of this drug is a serious social problem worldwide. In this study, a method based on gas chromatography-tandem mass spectrometry (GC-MS/MS) operated in electron ionization (EI) with simple and rapid liquid-liquid extraction (LLE) and derivatization was developed and validated for the simultaneous determination of THC, CBD, CBN and 11-OH-THC in hair samples. The detection of all compounds was based on multiple reaction monitoring (MRM) transitions. The most important advantage of this method is the single-step, quick, easy and effective sample extraction procedure for THC, CBD, CBN and 11-OH-THC. The method showed a good linearity with a correlation coefficient (r 2 ) between 0.997 and 0.999 for all substances. The variation coefficient (%CV) was <5% for THC, 11-OH-THC and CBD and <13% for CBN. The limit of detection (LOD) was 0.03 pg/mg for 11-OH-THC and it ranged from 0.3 to 1.4 pg/mg for THC, CBD and CBN. The limit of quantification was 0.1 pg/mg for 11-OH-THC and it ranged from 0.9 to 4.7 pg/mg for THC, CBD and CBN. Analytical recovery was higher than 88% for 11-OH-THC and it ranged between 68 and 97% for THC, CBD and CBN. Intra- and inter-assay precision and accuracy were always lower than 9-14% and 5-9%, respectively. In parallel, we have quantified the THC-COOH level, following the methods previously set-up by us. The whole procedure was successfully applied to more than 200 different hair samples from cannabis consumers, disclosing the presence of 11-OH-THC in a range between 0.2 pg/mg and 27 pg/mg, and the presence of THC-COOH in a range between 0.05 pg/mg and 42.05 pg/mg. These data provided a good start towards the use of 11-THC-OH as alternative hair biomarker of cannabis consumption. Copyright © 2018 Elsevier B.V. All rights reserved.

  18. Fast and efficient protein purification using membrane adsorber systems.

    Science.gov (United States)

    Suck, Kirstin; Walter, Johanna; Menzel, Frauke; Tappe, Alexander; Kasper, Cornelia; Naumann, Claudia; Zeidler, Robert; Scheper, Thomas

    2006-02-10

    The purification of proteins from complex cell culture samples is an essential step in proteomic research. Traditional chromatographic methods often require several steps resulting in time consuming and costly procedures. In contrast, protein purification via membrane adsorbers offers the advantage of fast and gentle but still effective isolation. In this work, we present a new method for purification of proteins from crude cell extracts via membrane adsorber based devices. This isolation procedure utilises the membranes favourable pore structure allowing high flow rates without causing high back pressure. Therefore, shear stress to fragile structures is avoided. In addition, mass transfer takes place through convection rather than diffusion, thus allowing very rapid separation processes. Based on this membrane adsorber technology the separation of two model proteins, human serum albumin (HSA) and immungluboline G (IgG) is shown. The isolation of human growth hormone (hGH) from chinese hamster ovary (CHO) cell culture supernatant was performed using a cation exchange membrane. The isolation of the enzyme penicillin acylase from the crude Escherichia coli supernatant was achieved using an anion exchange spin column within one step at a considerable purity. In summary, the membrane adsorber devices have proven to be suitable tools for the purification of proteins from different complex cell culture samples.

  19. The purification of hypertensin II.

    Science.gov (United States)

    SKEGGS, L T; KAHN, J R; SHUMWAY, N P

    1956-03-01

    The enzymatic conversion of hypertensin I to hypertensin II is described together with the subsequent purification of the product by means of counter-current distribution. Improved methods are also presented for the preparation of renin and its substrate, as well as in methods for the reaction of these materials and the purification of the resulting hypertensin I.

  20. Development of a Chemically Defined Medium for Better Yield and Purification of Enterocin Y31 from Enterococcus faecium Y31

    Directory of Open Access Journals (Sweden)

    Wenli Liu

    2017-01-01

    Full Text Available The macro- and micronutrients in traditional medium, such as MRS, used for cultivating lactic acid bacteria, especially for bacteriocin production, have not been defined, preventing the quantitative monitoring of metabolic flux during bacteriocin biosynthesis. To enhance Enterocin Y31 production and simplify steps of separation and purification, we developed a simplified chemically defined medium (SDM for the growth of Enterococcus faecium Y31 and production of its bacteriocin, Enterocin Y31. We found that the bacterial growth was unrelated to Enterocin Y31 production in MRS; therefore, both the growth rate and the Enterocin Y31 production were set as the index for investigation. Single omission experiments revealed that 5 g/L NaCl, five vitamins, two nucleic acid bases, MgSO4·7H2O, MnSO4·4H2O, KH2PO4, K2HPO4, CH3COONa, fourteen amino acids, and glucose were essential for the strain’s growth and Enterocin Y31 production. Thus, a novel simplified and defined medium (SDM was formulated with 30 components in total. Consequently, Enterocin Y31 production yield was higher in SDM as compared to either MRS or CDM. SDM improved the Enterocin Y31 production and simplified the steps of purification (only two steps, which has broad potential applications.

  1. One step purification of biological active human interleukin-2 protein ...

    African Journals Online (AJOL)

    Administrator

    2011-10-31

    Oct 31, 2011 ... Pharmacological importance of recombinant human interleukin-2 protein has increased the demand to establish effective ... Extracellular expression of mrhIL-2 in the culture supernatant was ~210 mg/L. Cell free culture .... bottomed micro plate (tissue culture grade) starting with 10 ng/ml in a volume of 100 ...

  2. One step purification of biological active human interleukin-2 protein ...

    African Journals Online (AJOL)

    The mrhIL-2 was expressed extracellularly under methanol inducible AOX1 promoter of P. pastoris. Extracellular expression of mrhIL-2 in the culture supernatant was ~210 mg/L. Cell free culture supernatant containing mrhIL-2 protein was concentrated and buffer exchanged by diafiltration by tangential flow filtration system.

  3. Protein production and purification.

    Science.gov (United States)

    Gräslund, Susanne; Nordlund, Pär; Weigelt, Johan; Hallberg, B Martin; Bray, James; Gileadi, Opher; Knapp, Stefan; Oppermann, Udo; Arrowsmith, Cheryl; Hui, Raymond; Ming, Jinrong; dhe-Paganon, Sirano; Park, Hee-won; Savchenko, Alexei; Yee, Adelinda; Edwards, Aled; Vincentelli, Renaud; Cambillau, Christian; Kim, Rosalind; Kim, Sung-Hou; Rao, Zihe; Shi, Yunyu; Terwilliger, Thomas C; Kim, Chang-Yub; Hung, Li-Wei; Waldo, Geoffrey S; Peleg, Yoav; Albeck, Shira; Unger, Tamar; Dym, Orly; Prilusky, Jaime; Sussman, Joel L; Stevens, Ray C; Lesley, Scott A; Wilson, Ian A; Joachimiak, Andrzej; Collart, Frank; Dementieva, Irina; Donnelly, Mark I; Eschenfeldt, William H; Kim, Youngchang; Stols, Lucy; Wu, Ruying; Zhou, Min; Burley, Stephen K; Emtage, J Spencer; Sauder, J Michael; Thompson, Devon; Bain, Kevin; Luz, John; Gheyi, Tarun; Zhang, Fred; Atwell, Shane; Almo, Steven C; Bonanno, Jeffrey B; Fiser, Andras; Swaminathan, Sivasubramanian; Studier, F William; Chance, Mark R; Sali, Andrej; Acton, Thomas B; Xiao, Rong; Zhao, Li; Ma, Li Chung; Hunt, John F; Tong, Liang; Cunningham, Kellie; Inouye, Masayori; Anderson, Stephen; Janjua, Heleema; Shastry, Ritu; Ho, Chi Kent; Wang, Dongyan; Wang, Huang; Jiang, Mei; Montelione, Gaetano T; Stuart, David I; Owens, Raymond J; Daenke, Susan; Schütz, Anja; Heinemann, Udo; Yokoyama, Shigeyuki; Büssow, Konrad; Gunsalus, Kristin C

    2008-02-01

    In selecting a method to produce a recombinant protein, a researcher is faced with a bewildering array of choices as to where to start. To facilitate decision-making, we describe a consensus 'what to try first' strategy based on our collective analysis of the expression and purification of over 10,000 different proteins. This review presents methods that could be applied at the outset of any project, a prioritized list of alternate strategies and a list of pitfalls that trip many new investigators.

  4. Gas purification project

    International Nuclear Information System (INIS)

    Broothaerts, J.; Claes, J.; Collard, G.; Goossens, W.; Harnie, R.; Heylen, P.; Vaesen, J.; Beukelaer, R. de; Dubois, G.; Glibert, R.; Mestrez, J.; Zahlen, A.

    1975-06-01

    Conceptual and experimental studies on LMFBR reprocessing and reactor off-gas purification systems are summarized. Iodine sorption on zeolites, low-temperature adsorption of noble gases on charcoal and catalytic oxidation of hydrogen, simulating tritium, are being studied in laboratory set-ups. A pilot loop with 25 m 3 h -1 throughput has been constructed. Results are quoted from the first phase of the iodine removal programme by scrubbing systems. Further extension of the test loop, comprising off-gases conditioning to removal of krypton in a cryodistillation unit, has been prepared. Delay-bed studies on 133 Xe extraction from LWR off-gases are reported. (author)

  5. Air/Water Purification

    Science.gov (United States)

    1992-01-01

    After 18 years of research into air/water pollution at Stennis Space Center, Dr. B. C. Wolverton formed his own company, Wolverton Environmental Services, Inc., to provide technology and consultation in air and water treatment. Common houseplants are used to absorb potentially harmful materials from bathrooms and kitchens. The plants are fertilized, air is purified, and wastewater is converted to clean water. More than 100 U.S. communities have adopted Wolverton's earlier water hyacinth and artificial marsh applications. Catfish farmers are currently evaluating the artificial marsh technology as a purification system.

  6. Water Purification Product

    Science.gov (United States)

    2004-01-01

    Ecomaster, an affiliate of BioServe Space Technologies, this PentaPure technology has been used to purify water for our nation's Space Shuttle missions since 1981. WTC-Ecomaster of Mirneapolis, Minnesota manufactures water purification systems under the brand name PentaPure (TM). BioServe researcher Dr. George Marchin, of Kansas State University, first demonstrated the superiority of this technology and licensed it to WTC. Marchin continues to perform microgravity research in the development of new technologies for the benefit of life on Earth.

  7. Purification and characterization of riproximin from Ximenia americana fruit kernels.

    Science.gov (United States)

    Bayer, Helene; Ey, Noreen; Wattenberg, Andreas; Voss, Cristina; Berger, Martin R

    2012-03-01

    Highly pure riproximin was isolated from the fruit kernels of Ximenia americana, a defined, seasonally available and potentially unlimited herbal source. The newly established purification procedure included an initial aqueous extraction, removal of lipids with chloroform and subsequent chromatographic purification steps on a strong anion exchange resin and lactosyl-Sepharose. Consistent purity and stable biological properties were shown over several purification batches. The purified, kernel-derived riproximin was characterized in comparison to the African plant material riproximin and revealed highly similar biochemical and biological properties but differences in the electrophoresis pattern and mass spectrometry peptide profile. Our results suggest that although the purified fruit kernel riproximin consists of a mixture of closely related isoforms, it provides a reliable basis for further research and development of this type II ribosome inactivating protein (RIP). Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Purification process of recombinant monoclonal antibodies with mixed mode chromatography.

    Science.gov (United States)

    Maria, Sophie; Joucla, Gilles; Garbay, Bertrand; Dieryck, Wilfrid; Lomenech, Anne-Marie; Santarelli, Xavier; Cabanne, Charlotte

    2015-05-08

    An innovative process to purify mAb from CHO cell culture supernatant was developed. This three-step process involved two mixed mode resins and an anion exchange membrane. We used a human IgG mixture to determine the optimal conditions for each purification step. Thereafter, the whole process was evaluated and improved for the purification of a recombinant mAb produced in the supernatant of CHO cells. Once optimized, yield and purity of 88% and 99.9%, respectively were comparable to those obtained in a conventional process based on a capture step using protein A. In addition, aggregates, HCPs and DNA levels in the purified fraction were below regulatory specifications. Then we used mass spectrometry to identify contaminating proteins in the antibody fraction in order to highlight the behavior of HCPs. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Small scale extraction and purification of human prolactin for the preparation of radioimmunoassay reagents

    International Nuclear Information System (INIS)

    Dias, L.E.M.F.

    1989-01-01

    Purification of human prolactin from pituitaries was carried out in our laboratory to obtain a pure reagent for use in RIA. The extraction and purification procedure was adapted from the method of Mc. Lean et al., and it involves the following steps: 1. Extraction of frozen pituitaries in buffers 0.14M phosphate/citrate pH 4.0 and 0.05M ammonium acetate pH 10.0. 2. Purification by hydrophobic interaction chromatography on Phenyl-Sepharose CL-4B in the presence of acetonitrile. 3. Purification by anion exchange chromatography on DEAE-Sepharose Cl-68. The purification method is considered effective for obtaining a hPrl of the purity needed for radioassay purposes, having the advantage of rapidity and relative simplicity. (author) [pt

  10. Co-adsorption of Cations as the Cause of the Apparent pH Dependence of Hydrogen Adsorption on a Stepped Platinum Single-Crystal Electrode.

    Science.gov (United States)

    Chen, Xiaoting; McCrum, Ian T; Schwarz, Kathleen A; Janik, Michael J; Koper, Marc T M

    2017-11-20

    The successful deployment of advanced energy-conversion systems depends critically on our understanding of the fundamental interactions of the key adsorbed intermediates (hydrogen *H and hydroxyl *OH) at electrified metal-aqueous electrolyte interfaces. The effect of alkali metal cations (Li + , Na + , K + , Cs + ) on the non-Nernstian pH shift of the step-related voltammetric peak of the Pt(553) electrode is investigated over a wide pH window (1 to 13) by means of experimental and computational methods. The co-adsorbed alkali cations along the step weaken the OH adsorption at the step sites, causing a positive shift of the potential of the step-related peak on Pt(553). Density functional calculations explain the observations on the identity and concentration of alkali cations on the non-Nernstian pH shift, and demonstrate that cation-hydroxyl co-adsorption causes the apparent pH dependence of "hydrogen" adsorption in the step sites of platinum electrodes. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Effect of charcoal on water purification

    OpenAIRE

    Suzuki, Hirotaka; Kawahigashi, Tatsuo

    2014-01-01

    [Abstract] A natural basin system purifies water through self-purification, but the water pollution load of a river might exceed its self-purification capacity. Charcoal, which is used for other uses aside from heating, such as air purification, was evaluated experimentally for water quality purification. The experiment described herein is based on simple water quality measurements. Some experimentally obtained results are discussed.

  12. Single-step green synthesis and characterization of gold-conjugated polyphenol nanoparticles with antioxidant and biological activities

    Directory of Open Access Journals (Sweden)

    Sanna V

    2014-10-01

    Full Text Available Vanna Sanna,1,2 Nicolino Pala,1 Giuseppina Dessì,1 Paola Manconi,1 Alberto Mariani,1 Sonia Dedola,3 Mauro Rassu,3 Claudia Crosio,3 Ciro Iaccarino,3 Mario Sechi1,2 1Department of Chemistry and Pharmacy, University of Sassari, Sassari, Italy; 2Laboratory of Nanomedicine, Department of Chemistry and Pharmacy, University of Sassari, c/o Porto Conte Ricerche, Tramariglio, Alghero, Italy; 3Department of Biomedical Sciences, University of Sassari, Sassari, Italy Background: Gold nanoparticles (GNPs are likely to provide an attractive platform for combining a variety of biophysicochemical properties into a unified nanodevice with great therapeutic potential. In this study we investigated the capabilities of three different natural polyphenols, epigallocatechin-3-gallate (EGCG, resveratrol (RSV, and fisetin (FS, to allow synergistic chemical reduction of gold salts to GNPs and stabilization in a single-step green process. Moreover, antioxidant properties of the nanosystems, as well as preliminary antiproliferative activity and apoptotic process investigation of model EGCG-GNPs on stable clones of neuroblastoma SH-SY5Y cells expressing CFP-DEVD-YFP reporter, were examined. Methods: The GNPs were characterized by physicochemical techniques, polyphenol content, and in vitro stability. The antioxidant activity of the GNPs was also determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH and 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid cation (ABTS radical-scavenging assays. Stable clones of neuronal SH-SY5Y-CFP-DEVD-YFP were generated and characterized, and cell viability after treatment with EGCG-GNPs was assessed after 72 hours through a 3(4,5-dimethylthiazol-2yl-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl-2H-tetrazolium assay. Activation of the apoptotic pathways was also investigated by Western blot analysis. Results: With a diameter in the size range of 10–25 nm, the obtained nanoparticles (NPs were found to contain 2.71%, 3.23%, and 5.47% of EGCG

  13. RNA purification by preparative polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Petrov, Alexey; Wu, Tinghe; Puglisi, Elisabetta Viani; Puglisi, Joseph D

    2013-01-01

    Preparative polyacrylamide gel electrophoresis (PAGE) is a powerful tool for purifying RNA samples. Denaturing PAGE allows separation of nucleic acids that differ by a single nucleotide in length. It is commonly used to separate and purify RNA species after in vitro transcription, to purify naturally occurring RNA variants such as tRNAs, to remove degradation products, and to purify labeled RNA species. To preserve RNA integrity following purification, RNA is usually visualized by UV shadowing or stained with ethidium bromide or SYBR green dyes. © 2013 Elsevier Inc. All rights reserved.

  14. Purification technologies for colloidal nanocrystals.

    Science.gov (United States)

    Shen, Yi; Gee, Megan Y; Greytak, A B

    2017-01-10

    Almost all applications of colloidal nanocrystals require some type of purification or surface modification process following nanocrystal growth. Nanocrystal purification - the separation of nanocrystals from undesired solution components - can perturb the surface chemistry and thereby the physical properties of colloidal nanocrystals due to changes in solvent, solute concentrations, and exposure of the nanocrystal surface to oxidation or hydrolysis. For example, nanocrystal quantum dots frequently exhibit decreased photoluminescence brightness after precipitation from the growth solvent and subsequent redissolution. Consequently, purification is an integral part of the synthetic chemistry of colloidal nanocrystals, and the effect of purification methods must be considered in order to accurately compare and predict the behavior of otherwise similar nanocrystal samples. In this Feature Article we examine established and emerging approaches to the purification of colloidal nanoparticles from a nanocrystal surface chemistry viewpoint. Purification is generally achieved by exploiting differences in properties between the impurities and the nanoparticles. Three distinct properties are typically manipulated: polarity (relative solubility), electrophoretic mobility, and size. We discuss precipitation, extraction, electrophoretic methods, and size-based methods including ultracentrifugation, ultrafiltration, diafiltration, and size-exclusion chromatography. The susceptibility of quantum dots to changes in surface chemistry, with changes in photoluminescence decay associated with surface chemical changes, extends even into the case of core/shell structures. Accordingly, the goal of a more complete description of quantum dot surface chemistry has been a driver of innovation in colloidal nanocrystal purification methods. We specifically examine the effect of purification on surface chemistry and photoluminescence in quantum dots as an example of the challenges associated with

  15. Magnetic Purification of Antibodies

    Science.gov (United States)

    Dhadge, Vijaykumar Laxman

    This work aimed at the development of magnetic nanoparticles for antibody purification and at the evaluation of their performance in Magnetic fishing and in a newly developed hybrid technology Magnetic Aqueous Two Phase Systems. Magnetic materials were produced by coprecipitation and solvothermal approaches. Natural polymers such as dextran, extracellular polysaccharide and gum Arabic were employed for coating of iron oxide magnetic supports. Polymer coated magnetic supports were then modified with synthetic antibody specific ligands,namely boronic acid, a triazine ligand (named 22/8) and an Ugi ligand (named A2C7I1). To optimize the efficacy of magnetic nanoparticles for antibody magnetic fishing, various solutions of pure and crude antibody solutions along with BSA as a non-specific binding protein were tested. The selectivity of magnetic nanoparticle for antibody, IgG, was found effective with boronic acid and ligand 22/8. Magnetic supports were then studied for their performance in high gradient magnetic separator for effective separation capability as well as higher volume handling capability. The magnetic materials were also supplemented to aqueous two phase systems, devising a new purification technology. For this purpose, magnetic particles modified with boronic acid were more effective. This alternative strategy reduced the time of operation,maximized separation capability (yield and purity), while reducing the amount of salt required. Boronic acid coated magnetic particles bound 170 +/- 10 mg hIgG/g MP and eluted 160 +/- 5 mg hIgG/g MP, while binding only 15 +/- 5 mg BSA/g MP. The affinity constant for the interaction between hIgG and APBA_MP was estimated as 4.9 x 105 M-1 (Ka) with a theoretical maximum capacity of 492 mg hIgG adsorbed/g MP (Qmax). APBA_MPs were also tested for antibody purification directly from CHO cell supernatants. The particles were able to bind 98% of IgG loaded and to recover 95% of pure IgG (purity greater than 98%) at extremely

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

    Directory of Open Access Journals (Sweden)

    Aimee Shen

    2009-12-01

    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.

  17. Purification and Partial Characterization of Trypanosoma cruzi Triosephosphate Isomerase

    Directory of Open Access Journals (Sweden)

    Bourguignon SC

    1998-01-01

    Full Text Available The enzyme triosephosphate isomerase (TPI, EC 5.3.1.1 was purified from extracts of epimastigote forms of Trypanosoma cruzi. The purification steps included: hydrophobic interaction chromatography on phenyl-Sepharose, CM-Sepharose, and high performance liquid gel filtration chromatography. The CM-Sepharose material contained two bands (27 and 25 kDa with similar isoelectric points (pI 9.3-9.5 which could be separated by gel filtration in high performance liquid chromatography. Polyclonal antibodies raised against the porcine TPI detected one single polypeptide on western blot with a molecular weight (27 kDa identical to that purified from T. cruzi. These antibodies also recognized only one band of identical molecular weight in western blots of several other trypanosomatids (Blastocrithidia culicis, Crithidia desouzai, Phytomonas serpens, Herpertomonas samuelpessoai. The presence of only one enzymatic form of TPI in T. cruzi epimastigotes was confirmed by agarose gel activity assay and its localization was established by immunocytochemical analysis. The T. cruzi purified TPI (as well as other trypanosomatid' TPIs is a dimeric protein, composed of two identical subunits with an approximate mw of 27,000 and it is resolved on two dimensional gel electrophoresis with a pI of 9.3. Sequence analysis of the N-terminal portion of the 27 kDa protein revealed a high homology to Leishmania mexicana and T. brucei proteins

  18. An alternative purification method for human serum paraoxonase 1 and its interaction with methidathion.

    Science.gov (United States)

    Gençer, Nahit; Yavuz, Emre

    2017-07-01

    In this study, an alternative purification method for human Paraoxonase 1 (hPON1) enzyme was developed using two-step procedures, namely ammonium sulphate precipitation and Sepharose-4B-L-tyrosine-1-aminoanthracene hydrophobic interaction chromatography. SDS-polyacrylamide gel electrophoresis of the enzyme indicates a single band with an apparent MW of 43 kDa. The enzyme was purified 674-fold with a yield of 16%. Furthermore, we examined the in vitro effect of methidathion on the enzyme activity to understand the better inhibitory properties of the compound. Methidathion is a highly toxic insecticide used to control a broad spectrum of agricultural insect and mite pests. IC 50 value was found to be 0.130 mM for the pesticide. Methidathion showed a competitive inhibition with Ki of 0.119 mM for paraoxon.

  19. Purification of papain using reactive green 5 attached supermacroporous monolithic cryogel.

    Science.gov (United States)

    Uygun, Deniz Aktaş; Akduman, Begüm; Uygun, Murat; Akgöl, Sinan; Denizli, Adil

    2012-06-01

    Supermacroporous poly(2-hydroxyethyl methacrylate) [poly(HEMA)] monolithic cryogel was prepared by radical cryocopolymerization of HEMA with N,N'-methylene bisacrylamide as crosslinker. Reactive Green 5 dye was immobilized to the cryogel with nucleophilic substitution reaction, and this dye attached cryogel column was used for affinity purification of papain from Carica papaya latex. Reactive Green 5-immobilized poly(HEMA) cryogel was characterized by swelling studies, Fourier transform infrared spectroscopy, scanning electron microscopy, and energy dispersive X-ray analysis. Maximum papain adsorption capacity was found to be 68.5 mg/g polymer while nonspecific papain adsorption onto plain cryogel was negligible (3.07 mg/g polymer). Papain from C. papaya was purified 42-fold in single step with dye attached cryogel, and purity of papain was shown by silver-stained sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

  20. Efficient expression and purification of a protease from the common cold virus, human rhinovirus type 14

    Science.gov (United States)

    Leong, L. E.-C.; Walker, P. A.; Porter, A. G.

    1992-08-01

    The protease (3C pro) from human rhinovirus serotype-14 (HRV-14) has been cloned and efficiently expressed in E. coli. A straightforward single-step purification of the recombinant 3C pro has been achieved by fusing the protein to the car☐y-terminus of the glutathione-S-transferase from Schistosoma japonicum. Modifications made to the 5' end of the PCR fragment coding for the 3C pro have allowed the specific cleavage of the fusion protein using thrombin to yield mature 3C pro with the correct amino-terminal amino acid. This protease has been shown to be active when assayed using synthetic peptides corresponding to the natural cleavage recognition sequences within the polyprotein. Other substrates are being developed for this protease for possible use in the screening of inhibitors of 3C pro. Sufficient protease 3C pro has been purified for initial attempts at crystallization.

  1. Purification of adenoviral vectors using expanded bed chromatography.

    Science.gov (United States)

    Peixoto, C; Ferreira, T B; Carrondo, M J T; Cruz, P E; Alves, P M

    2006-03-01

    The increasing numbers of pre-clinical and clinical trials where recombinant adenoviral vectors are used for gene therapy and vaccination require the development of cost-effective and reproducible large scale purification strategies of the biologically active particles. Alternatives to the traditional laboratory scale CsCl density gradient ultracentrifugation method, such as fixed bed chromatography strategies, have been developed, but the yields of final recovery remain too low due mainly to the capture and concentration steps taking place before and between the chromatographic stages. In this study, a rapid and efficient scale-able purification protocol allowing to obtain concentrated, pure and bioactive adenoviral vectors was developed. This allows efficient levels of binding to the column media and vector purification without centrifugation or filtration steps. Expanded bed chromatography followed by hollow fiber concentration allows the capture of viral particles directly from cellular extracts with high efficiency and vector purification is achieved in less than one working day with a minimal amount of sample handling, thus presenting an improvement over existing processes. The overall process yield reached 32%, representing an eight-fold improvement over results reported previously, while the purity is comparable to that obtained with the CsCl method.

  2. Acrylic purification and coatings

    International Nuclear Information System (INIS)

    Kuzniak, Marcin

    2011-01-01

    Radon (Rn) and its decay daughters are a well-known source of background in direct WIMP detection experiments, as either a Rn decay daughter or an alpha particle emitted from a thin inner surface layer of a detector could produce a WIMP-like signal. Different surface treatment and cleaning techniques have been employed in the past to remove this type of contamination. A new method of dealing with the problem has been proposed and used for a prototype acrylic DEAP-1 detector. Inner surfaces of the detector were coated with a layer of ultra pure acrylic, meant to shield the active volume from alphas and recoiling nuclei. An acrylic purification technique and two coating techniques are described: a solvent-borne (tested on DEAP-1) and solvent-less (being developed for the full scale DEAP-3600 detector).

  3. A green recyclable SO(3)H-carbon catalyst derived from glycerol for the production of biodiesel from FFA-containing karanja (Pongamia glabra) oil in a single step.

    Science.gov (United States)

    Prabhavathi Devi, B L A; Vijai Kumar Reddy, T; Vijaya Lakshmi, K; Prasad, R B N

    2014-02-01

    Simultaneous esterification and transesterification method is employed for the preparation of biodiesel from 7.5% free fatty acid (FFA) containing karanja (Pongamia glabra) oil using water resistant and reusable carbon-based solid acid catalyst derived from glycerol in a single step. The optimum reaction parameters for obtaining biodiesel in >99% yield by simultaneous esterification and transesterification are: methanol (1:45 mole ratio of oil), catalyst 20wt.% of oil, temperature 160°C and reaction time of 4h. After the reaction, the catalyst was easily recovered by filtration and reused for five times with out any deactivation under optimized conditions. This single-step process could be a potential route for biodiesel production from high FFA containing oils by simplifying the procedure and reducing costs and effluent generation. Copyright © 2013 Elsevier Ltd. All rights reserved.

  4. Trace element characterisation and purification of quartz powder of Indian origin

    International Nuclear Information System (INIS)

    Dash, K.; Thangavel, S.; Dhavile, S.M.; Sahayam, A.C.; Chaurasia, S.C.

    2002-11-01

    Analytical methodologies for the trace element characterisation and purification of quartz powder of Indian origin have been described. Metallic impurities (11 elements) in ∼700 quartz samples have been analysed using different instrumental techniques. The values are cross-validated by American and British analytical laboratories. A special multi channel vapour phase digestion (MCVPD) chamber has been designed to reduce the process blank levels for the determination of ultra trace impurities in high purity silicon matrix. In this vessel 21 samples can be digested at a time in a period of 8 hrs. at normal pressure and low temperature (∼ 80 degC). Analytical methodologies for the determination of non-metals (phosphorus, boron and chloride) at very low levels <1 ppm) have also been described. A highly cost effective, single step, room temperature purification procedure is developed based on chemical leaching. Around 300 raw quartz powders (3-4N) from various mines have been purified to 5N pure ( optical grade ). (author)

  5. Purification and characterization of a fibrinolytic enzyme from tempeh bongkrek as an alternative of thrombolytic agents

    Science.gov (United States)

    Sasmita, I. R. A.; Sutrisno, A.; Zubaidah, E.; Wardani, A. K.

    2018-03-01

    Tempeh is one of Indonesia’s traditional foods that contain fibrinolytic enzymes. Tempeh bongkrek shows very strong activity among various tempeh. The fibrinolytic enzymes of bongkrek tempeh are obtained by steps of purification i.e, ammonium sulphate precipitation, ion exchange chromatography and gel filtration chromatography. The fibrinolytic enzymes has been successfully purified with a yield of 4.37%, specific activity of 3,361 U / mg and purification fold of 44.02. SDS PAGE analysis showed that the enzyme was purified in to single band with estimated molecular mass of 75.82 kDa. The purified enzyme has optimum pH of 7 and optimum temperature of 50°C and pH stability between pH 4 - 7 with temperature stability from 30°-50°C. The fibrinolytic activity is increased with addition of CaCl2 but inhibited with CuSO4, phenylmethylsulfonyl fluoride (PMSF), sodium dodecyl sulfate (SDS), and ethylenediaminetetraacetic acid (EDTA).

  6. Step-wise integration of single-port laparoscopic surgery into routine colorectal surgical practice by use of a surgical glove port.

    Science.gov (United States)

    Hompes, R; Lindsey, I; Jones, O M; Guy, R; Cunningham, C; Mortensen, N J; Cahill, R A

    2011-06-01

    The cost associated with single-port laparoscopic access devices may limit utilisation of single-port laparoscopic surgery by colorectal surgeons. This paper describes a simple and cheap access modality that has facilitated the widespread adoption of single-port technology in our practice both as a stand-alone procedure and as a useful adjunct to traditional multiport techniques. A surgical glove port is constructed by applying a standard glove onto the rim of the wound protector/retractor used during laparoscopic resectional colorectal surgery. To illustrate its usefulness, we present our total experience to date and highlight a selection of patients presenting for a range of elective colorectal surgery procedures. The surgical glove port allowed successful completion of 25 single-port laparoscopic procedures (including laparoscopic adhesiolysis, ileo-rectal anastomosis, right hemicolectomy, total colectomy and low anterior resection) and has been used as an adjunct in over 80 additional multiport procedures (including refashioning of a colorectal anastomosis made after specimen extraction during a standard multiport laparoscopic anterior resection). This simple, efficient device can allow use of single-port laparoscopy in a broader spectrum of patients either in isolation or in combination with multiport surgery than may be otherwise possible for economic reasons. By separating issues of cost from utility, the usefulness of the technical advance inherent within single-port laparoscopy for colorectal surgery can be better appreciated. We endorse the creative innovation inherent in this approach as surgical practice continues to evolve for ever greater patient benefit.

  7. Single-step functionalization of vertically aligned MWCNTs with Cu and Ni by chemical reduction of copper and nickel acetyl acetonate in benzyl alcohol

    OpenAIRE

    Ilari Gabriele et al.

    2014-01-01

    We report a one-step functionalization reaction of forests of vertically aligned multi-walled carbon nanotubes (MWCNTs) with Cu or Ni nanoparticles. The benzyl alcohol assisted deposition reaction reduces copper or nickel acetyl acetonate to metallic homogeneously distributed nanoparticles and a 1 mu m thick metallic capping layer normal to the MWCNT forests. Scanning electron microscopy and transmission electron microscopy images reveal a trimodal diameter regime for the Cu nanoparticles and...

  8. A new method for high yield purification of type beta transforming growth factor from human platelets

    NARCIS (Netherlands)

    Eijnden-van Raaij, A.J.M. van den; Koornneef, I.; Zoelen, E.J.J. van

    1988-01-01

    A new method was developed for the purification of type beta transforming growth factor from human platelets. This method is a three-step procedure including gel filtration, weak cation exchange HPLC and reverse phase HPLC. All steps are carried out at low pH using exclusively volatile acidic buffer

  9. Sequential purification and crystal growth for the production of low cost silicon substrates. Quarterly technical progress report No. 1, 15 September 1979-31 December 1979

    Energy Technology Data Exchange (ETDEWEB)

    Liaw, M.; Secco, F.; Ingle, B.; Down, D.

    1980-02-01

    Over the past several years, Motorola's Materials Technology Laboratory (MTL), has been conducting several projects with goals directed at the production of high quality low cost silicon crystals. One of the projects which is being investigated is the direct purification of MG-Si. A unique characteristic of the approach used by this project is the use of a crystal puller to perform both purification and crystal growth. Sequential steps of purification were taken. By the completion of this series of purification, the purified MG-Si melt will be further purified by impurity redistribution using ingot pulling. The final purified silicon will be in an ingot form of desired dimensions for slicing into silicon sheets. The sequential steps of purification include: (1) leaching of MG-Si charge, (2) phase separation, (3) reactive gas treatment, (4) liquid-liquid extraction (called Slagging), and (5) purification by redistribution of impurities using ingot pulling. Progress on items (1) and (2) is reported. (WHK)

  10. One Pot Single Step Synthesis and Biological Evaluation of Some Novel Bis(1,3,4-thiadiazole Derivatives as Potential Cytotoxic Agents

    Directory of Open Access Journals (Sweden)

    Sobhi M. Gomha

    2016-11-01

    Full Text Available A novel series of bis(1,3,4-thiadiazole derivatives were synthesized in one step methodology with good yields by condensation reaction between bis-hydrazonoyl chloride 1 and various reagents. The structures of the prepared compounds were confirmed by spectral data (IR, NMR, and MS, and elemental analysis. The anticancer activity against human breast carcinoma (MCF-7 cancer cell lines was evaluated in MTT assay. The results revealed that the bis-thiadiazole derivatives 5c,d, 7b,c and 9c had higher antitumor activity than the standard drug Imatinib.

  11. Conversion of failed laparoscopic adjustable gastric banding to Roux-en-Y gastric bypass is safe as a single-step procedure

    NARCIS (Netherlands)

    Emous, M.; Apers, J.; Hoff, C.; van Beek, A. P.; Totte, E.

    Several different procedures have been proposed as a revisional procedure for treatment of failed laparoscopic adjustable gastric banding (LAGB). Laparoscopic Roux-en-Y gastric bypass (LRYGB) has been advocated as the procedure of choice for revision. In this study, we compare the single- and

  12. Estimating trematode prevalence in snail hosts using a single-step duplex PCR: how badly does cercarial shedding underestimate infection rates?

    Czech Academy of Sciences Publication Activity Database

    Born-Torrijos, A.; Poulin, R.; Raga, J. A.; Holzer, Astrid S.

    2014-01-01

    Roč. 7, MAY 2014 (2014), s. 243 ISSN 1756-3305 R&D Projects: GA ČR GBP505/12/G112 Institutional support: RVO:60077344 Keywords : prevalence * detection * snail host * double infection * single-steo duplex PCR Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.430, year: 2014

  13. Aqueous Chloride Operations Overview: Plutonium and Americium Purification/Recovery

    Energy Technology Data Exchange (ETDEWEB)

    Gardner, Kyle Shelton [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Kimball, David Bryan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Skidmore, Bradley Evan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2016-09-28

    These are a set of slides intended for an information session as part of recruiting activities at Brigham Young University. It gives an overview of aqueous chloride operations, specifically on plutonium and americium purification/recovery. This presentation details the steps taken perform these processes, from plutonium size reduction, dissolution, solvent extraction, oxalate precipitation, to calcination. For americium recovery, it details the CLEAR (chloride extraction and actinide recovery) Line, oxalate precipitation and calcination.

  14. Over-Expression, Purification and Crystallization of Human Dihydrolipoamide Dehydrogenase

    Science.gov (United States)

    Hong, Y. S.; Ciszak, Ewa; Patel, Mulchand

    2000-01-01

    Dehydrolipoamide dehydrogenase (E3; dihydrolipoan-tide:NAD+ oxidoreductase, EC 1.8.1.4) is a common catalytic component found in pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex, and branched-chain cc-keto acid dehydrogenase complex. E3 is also a component (referred to as L protein) of the glycine cleavage system in bacterial metabolism (2). Active E3 forms a homodimer with four distinctive subdomain structures (FAD binding, NAD+ binding, central and interface domains) with non-covalently but tightly bound FAD in the holoenzyme. Deduced amino acids from cloned full-length human E3 gene showed a total of 509 amino acids with a leader sequence (N-terminal 35 amino acids) that is excised (mature form) during transportation of expressed E3 into mitochondria membrane. So far, three-dimensional structure of human E3 has not been reported. Our effort to achieve the elucidation of the X-ray crystal structure of human E3 will be presented. Recombinant pPROEX-1 expression vector (from GIBCO BRL Life Technologies) having the human E3 gene without leader sequence was constructed by Polymerase Chain Reaction (PCR) and subsequent ligation, and cloned in E.coli XL1-Blue by transformation. Since pPROEX-1 vector has an internal His-tag (six histidine peptide) located at the upstream region of a multicloning site, one-step affinity purification of E3 using nickelnitriloacetic acid (Ni-NTA) agarose resin, which has a strong affinity to His-tag, was feasible. Also a seven-amino-acid spacer peptide and a recombinant tobacco etch virus protease recognition site (seven amino acids peptide) found between His-tag and first amino acid of expressed E3 facilitated the cleavage of His-tag from E3 after the affinity purification. By IPTG induction, ca. 15 mg of human E3 (mature form) was obtained from 1L LB culture with overnight incubation at 25C. Over 98% of purity of E3 from one-step Ni-NTA agarose affinity purification was confirmed by SDS-PAGE analysis. For

  15. Relationships between knee valgus, hip-muscle strength, and hip-muscle recruitment during a single-limb step-down.

    Science.gov (United States)

    Hollman, John H; Ginos, Barbara E; Kozuchowski, Jakub; Vaughn, Amanda S; Krause, David A; Youdas, James W

    2009-02-01

    Reduced strength and activation of hip muscles might correlate with increased weight-bearing knee valgus. To describe relationships among frontal-plane hip and knee angles, hip-muscle strength, and electromyographic (EMG) recruitment in women during a step-down. Exploratory study. Laboratory. 20 healthy women 20 to 30 years of age. Frontal-plane hip and knee angles were measured. Gluteus maximus and medius recruitment were examined with surface EMG. Hip-abduction and -external-rotation strength were quantified with handheld dynamometry. The authors analyzed correlation coefficients between knee and hip angles, gluteus maximus and medius EMG, and hip-abduction and -external-rotation strength. Hip-adduction angles (r = .755, P = .001), gluteus maximus EMG (r = -.451, P = .026), and hip-abduction strength (r = .455, P = .022) correlated with frontal-plane projections of knee valgus. Gluteus maximus recruitment might have greater association with reduced knee valgus in women than does external-rotation strength during step-down tasks. Gluteus medius strength might be associated with increased knee valgus.

  16. Human granulocyte colony stimulating factor (hG-CSF: cloning, overexpression, purification and characterization

    Directory of Open Access Journals (Sweden)

    Vanz Ana LS

    2008-04-01

    Full Text Available Abstract Background Biopharmaceutical drugs are mainly recombinant proteins produced by biotechnological tools. The patents of many biopharmaceuticals have expired, and biosimilars are thus currently being developed. Human granulocyte colony stimulating factor (hG-CSF is a hematopoietic cytokine that acts on cells of the neutrophil lineage causing proliferation and differentiation of committed precursor cells and activation of mature neutrophils. Recombinant hG-CSF has been produced in genetically engineered Escherichia coli (Filgrastim and successfully used to treat cancer patients suffering from chemotherapy-induced neutropenia. Filgrastim is a 175 amino acid protein, containing an extra N-terminal methionine, which is needed for expression in E. coli. Here we describe a simple and low-cost process that is amenable to scaling-up for the production and purification of homogeneous and active recombinant hG-CSF expressed in E. coli cells. Results Here we describe cloning of the human granulocyte colony-stimulating factor coding DNA sequence, protein expression in E. coli BL21(DE3 host cells in the absence of isopropyl-β-D-thiogalactopyranoside (IPTG induction, efficient isolation and solubilization of inclusion bodies by a multi-step washing procedure, and a purification protocol using a single cationic exchange column. Characterization of homogeneous rhG-CSF by size exclusion and reverse phase chromatography showed similar yields to the standard. The immunoassay and N-terminal sequencing confirmed the identity of rhG-CSF. The biological activity assay, in vivo, showed an equivalent biological effect (109.4% to the standard reference rhG-CSF. The homogeneous rhG-CSF protein yield was 3.2 mg of bioactive protein per liter of cell culture. Conclusion The recombinant protein expression in the absence of IPTG induction is advantageous since cost is reduced, and the protein purification protocol using a single chromatographic step should reduce cost

  17. Water Purification Systems

    Science.gov (United States)

    1994-01-01

    Clearwater Pool Technologies employs NASA-developed silver/copper ionization to purify turtle and dolphin tanks, cooling towers, spas, water recycling systems, etc. The pool purifier consists of a microcomputer to monitor water conditions, a pair of metallic electrodes, and a rheostat controller. Ions are generated by passing a low voltage current through the electrodes; the silver ions kill the bacteria, and the copper ions kill algae. This technology has found broad application because it offers an alternative to chemical disinfectants. It was originally developed to purify water on Apollo spacecraft. Caribbean Clear has been using NASA's silver ionization technology for water purification for more than a decade. Two new products incorporate advancements of the basic technology. One is the AquaKing, a system designed for areas with no source of acceptable drinking water. Another is the Caribbean Clear Controller, designed for commercial pool and water park applications where sanitizing is combined with feedback control of pH and an oxidizer, chlorine or bromine. The technology was originally developed to purify water on Apollo spacecraft.

  18. Reduction of a single granule drying model: An essential step in preparation of a population balance model with a continuous growth term

    DEFF Research Database (Denmark)

    Mortier, Séverine Thérèse F.C.; Daele, Timothy, Van; Gernaey, Krist V.

    2013-01-01

    for a single granule needs reduction in complexity. The starting point is a detailed model that describes the drying behavior of single pharmaceutical granules. A Global Sensitivity Analysis (GSA) was performed to detect the most sensitive degrees of freedom in the model as these need to be retained...... in the reduced model. Simulations of the complex drying model were, in a next phase, used to develop the reduced model, which describes the decrease of the moisture content in function of the gas temperature. The developed reduced model was then included in a Population Balance Equation (PBE) to describe......The development of a Population Balance Model (PBM) for a pharmaceutical granule drying process requires a continuous growth term; the latter actually represents the drying process as the moisture content is the internal coordinate of the PBM. To establish such a PBM, a complex drying model...

  19. A single and two step isomerization process for d-tagatose and l-ribose bioproduction using l-arabinose isomerase and d-lyxose isomerase.

    Science.gov (United States)

    Patel, Manisha J; Akhani, Rekha C; Patel, Arti T; Dedania, Samir R; Patel, Darshan H

    2017-02-01

    l-ribose and d-tagatose are biochemically synthesized using sugar isomerases. The l-arabinose isomerase gene from Shigella flexneri (Sf-AI) was cloned and expressed in Escherichia coli BL-21. Sf-AI was applied for the bioproduction of d-tagatose from d-galactose. l-ribose synthesis was performed by two step isomerization using Sf-AI and d-lyxose/ribose isomerase from Cohnella laevoribosii. The overall 22.3% and 25% conversion rate were observed for d-tagatose and l-ribose production from d-galactose and l-arabinose respectively. In the present manuscript, synthesis of rare sugars from naturally available sugars is discussed along with the biochemical characterization of Sf-AI and its efficiency. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. Affinity purification of the voltage-sensitive sodium channel from electroplax with resins selective for sialic acid

    Energy Technology Data Exchange (ETDEWEB)

    James, W.M.; Emerick, M.C.; Agnew, W.S. (Yale Univ. School of medicine, New Haven, CT (USA))

    1989-07-11

    The voltage-sensitive sodium channel present in the eel (Electrophorus electricus) has an unusually high content of sialic acid, including {alpha}-(2{yields}8)-linked polysialic acid, not found in other electroplax membrane glycopeptides. Lectins from Limax flavus (LFA) and wheat germ (WGA) proved the most effective of 11 lectin resins tried. The most selective resin was prepared from IgM antibodies against Neisseria meningitidis {alpha}-(2{yields}8)-polysialic acid which were affinity purified and coupled to Sepharose 4B. The sodium channel was found to bind to WGA, LFA, and IgM resins and was readily eluted with the appropriate soluble carbohydrates. Experiments with LFA and IgM resins demonstrated binding and unbinding rates and displacement kinetics, which suggest highly specific binding at multiple sites on the sodium channel protein. In preparative-scale purification of protein previously fractionated by anion-exchange chromatography, without stabilizing TTX, high yields were reproducibly obtained. Further, when detergent extracts were prepared from electroplax membranes fractionated by low-speed sedimentation, a single step over the IgM resin provided a 70-fold purification, yielding specific activities of 3,200 pmol of ({sup 3}H)TTX-binding sites/mg of protein and a single polypeptide of {approximately}285,000 Da on SDS-acrylamide gels. No small peptides were observed after this 5-h isolation. The authors describe a cation-dependent stabilization with millimolar levels of monovalent and micromolar levels of divalent species.

  1. Ribonucleoprotein purification and characterization using RNA Mango.

    Science.gov (United States)

    Panchapakesan, Shanker Shyam S; Ferguson, Matthew L; Hayden, Eric J; Chen, Xin; Hoskins, Aaron A; Unrau, Peter J

    2017-10-01

    The characterization of RNA-protein complexes (RNPs) is a difficult but increasingly important problem in modern biology. By combining the compact RNA Mango aptamer with a fluorogenic thiazole orange desthiobiotin (TO1-Dtb or TO3-Dtb) ligand, we have created an RNA tagging system that simplifies the purification and subsequent characterization of endogenous RNPs. Mango-tagged RNP complexes can be immobilized on a streptavidin solid support and recovered in their native state by the addition of free biotin. Furthermore, Mango-based RNP purification can be adapted to different scales of RNP isolation ranging from pull-down assays to the isolation of large amounts of biochemically defined cellular RNPs. We have incorporated the Mango aptamer into the S. cerevisiae U1 small nuclear RNA (snRNA), shown that the Mango-snRNA is functional in cells, and used the aptamer to pull down a U1 snRNA-associated protein. To demonstrate large-scale isolation of RNPs, we purified and characterized bacterial RNA polymerase holoenzyme (HE) in complex with a Mango-containing 6S RNA. We were able to use the combination of a red-shifted TO3-Dtb ligand and eGFP-tagged HE to follow the binding and release of the 6S RNA by two-color native gel analysis as well as by single-molecule fluorescence cross-correlation spectroscopy. Together these experiments demonstrate how the Mango aptamer in conjunction with simple derivatives of its flurophore ligands enables the purification and characterization of endogenous cellular RNPs in vitro. © 2017 Panchapakesan et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  2. High quality protein microarray using in situ protein purification

    Directory of Open Access Journals (Sweden)

    Fleischmann Robert D

    2009-08-01

    protein solubility and denaturation problems caused by buffer exchange steps and freeze-thaw cycles, which are associated with resin-based purification, intermittent protein storage and deposition on microarrays. Conclusion An optimized platform for in situ protein purification on microarray slides using His-tagged recombinant proteins is a desirable tool for the screening of novel protein functions and protein-protein interactions. In the context of immunoproteomics, such protein microarrays are complimentary to approaches using non-recombinant methods to discover and characterize bacterial antigens.

  3. PRELIMINARY MODELING OF AN INDUSTRIAL RECOMBINANT HUMAN ERYTHROPOIETIN PURIFICATION PROCESS BY ARTIFICIAL NEURAL NETWORKS

    Directory of Open Access Journals (Sweden)

    R. H. R. Garcel1

    2015-09-01

    Full Text Available AbstractIn the present study a preliminary neural network modelling to improve our understanding of Recombinant Human Erythropoietin purification process in a plant was explored. A three layer feed-forward back propagation neural network was constructed for predicting the efficiency of the purification section comprising four chromatographic steps as a function of eleven operational variables. The neural network model performed very well in the training and validation phases. Using the connection weight method the predictor variables were ranked based on their estimated explanatory importance in the neural network and five input variables were found to be predominant over the others. These results provided useful information showing that the first chromatographic step and the third chromatographic step are decisive to achieve high efficiencies in the purification section, thus enriching the control strategy of the plant.

  4. Laser material purification of neodymium. Hikari reiki seiseiho ni yoru neodymium no kojundoka

    Energy Technology Data Exchange (ETDEWEB)

    Ogawa, Y.; Ozaki, T.; Yoshimatsu, S. (National Research Institute for Metals, Tokyo (Japan)); Chiba, K.; Umeda, H.; Saeki, M. (Nippon Steel Corp., Tokyo (Japan))

    1991-05-20

    Selective photoexcitation and photoionization of neodymium atoms were studied basically by using laser. Also, using their properties, feasibility of laser material purification (LPM) technology in gaseous phase was presented. In the selective 3-step ionization, laser beam with two kinds of proper wave length causes resonance exitation of the target element, which goes up to the excitation level of the first step, and the element is ionized by the second laser beam and the third laser beam. The 3-step ionization scheme by a single wave length laser beam of 577.612nm was used for the ionization of Nd. Nd ionized selectively by laser beam was recovered as thin layer at the negative potential side of plane electrodes placed at both sides of the laser irradiation area. In the layer formed by the TPD technology with this scheme, it is possible to decrease impuritis such as Pr by 1/16 and to form highly purified thin layer. 13 refs., 12 figs., 1 tab.

  5. The oxidative fermentation of ethanol in Gluconacetobacter diazotrophicus is a two-step pathway catalyzed by a single enzyme: alcohol-aldehyde Dehydrogenase (ADHa).

    Science.gov (United States)

    Gómez-Manzo, Saúl; Escamilla, José E; González-Valdez, Abigail; López-Velázquez, Gabriel; Vanoye-Carlo, América; Marcial-Quino, Jaime; de la Mora-de la Mora, Ignacio; Garcia-Torres, Itzhel; Enríquez-Flores, Sergio; Contreras-Zentella, Martha Lucinda; Arreguín-Espinosa, Roberto; Kroneck, Peter M H; Sosa-Torres, Martha Elena

    2015-01-07

    Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2-C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde.

  6. The Oxidative Fermentation of Ethanol in Gluconacetobacter diazotrophicus Is a Two-Step Pathway Catalyzed by a Single Enzyme: Alcohol-Aldehyde Dehydrogenase (ADHa

    Directory of Open Access Journals (Sweden)

    Saúl Gómez-Manzo

    2015-01-01

    Full Text Available Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH and the aldehyde dehydrogenase (ALDH. We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2–C6 and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde.

  7. Enhanced Water Oxidation Photoactivity of Nano-Architectured α-Fe2O3-WO3 Composite Synthesized by Single-Step Hydrothermal Method

    Science.gov (United States)

    Rahman, Gul; Joo, Oh-Shim; Chae, Sang Youn; Shah, Anwar-ul-Haq Ali; Mian, Shabeer Ahmad

    2018-04-01

    This study reports the one-step in situ synthesis of a hematite-tungsten oxide (α-Fe2O3-WO3) composite on fluorine-doped tin oxide substrate via a simple hydrothermal method. Scanning electron microscopy images indicated that the addition of tungsten (W) precursor into the reaction mixture altered the surface morphology from nanorods to nanospindles. Energy-dispersive x-ray spectroscopy analysis confirmed the presence of W content in the composite. From the ultraviolet-visible spectrum of α-Fe2O3-WO3, it was observed that absorption began at ˜ 600 nm which corresponded to the bandgap energy of ˜ 2.01 eV. The α-Fe2O3-WO3 electrode demonstrated superior performance, with water oxidation photocurrent density of 0.80 mA/cm2 (at 1.6 V vs. reversible hydrogen electrode under standard illumination conditions; AM 1.5G, 100 mW/cm2) which is 2.4 times higher than α-Fe2O3 (0.34 mA/cm2). This enhanced water oxidation performance can be attributed to the better charge separation properties in addition to the large interfacial area of small-sized particles present in the α-Fe2O3-WO3 nanocomposite film.

  8. Development and characterization of a novel, antimicrobial, sterile hydrogel dressing for burn wounds: single-step production with gamma irradiation creates silver nanoparticles and radical polymerization.

    Science.gov (United States)

    Boonkaew, Benjawan; Barber, Philip M; Rengpipat, Sirirat; Supaphol, Pitt; Kempf, Margit; He, Jibao; John, Vijay T; Cuttle, Leila

    2014-10-01

    Patients with burn wounds are susceptible to wound infection and sepsis. This research introduces a novel burn wound dressing that contains silver nanoparticles (SNPs) to treat infection in a 2-acrylamido-2-methylpropane sulfonic acid sodium salt (AMPS-Na(+) ) hydrogel. Silver nitrate was dissolved in AMPS-Na(+) solution and then exposed to gamma irradiation to form SNP-infused hydrogels. The gamma irradiation results in a cross-linked polymeric network of sterile hydrogel dressing and a reduction of silver ions to form SNPs infused in the hydrogel in a one-step process. About 80% of the total silver was released from the hydrogels after 72 h immersion in simulated body fluid solution; therefore, they could be used on wounds for up to 3 days. All the hydrogels were found to be nontoxic to normal human dermal fibroblast cells. The silver-loaded hydrogels had good inhibitory action against Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus. Results from a pilot study on a porcine burn model showed that the 5-mM silver hydrogel was efficient at preventing bacterial colonization of wounds, and the results were comparable to the commercially available silver dressings (Acticoat(TM) , PolyMem Silver(®) ). These results support its use as a potential burn wound dressing. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

  9. The Oxidative Fermentation of Ethanol in Gluconacetobacter diazotrophicus Is a Two-Step Pathway Catalyzed by a Single Enzyme: Alcohol-Aldehyde Dehydrogenase (ADHa)

    Science.gov (United States)

    Gómez-Manzo, Saúl; Escamilla, José E.; González-Valdez, Abigail; López-Velázquez, Gabriel; Vanoye-Carlo, América; Marcial-Quino, Jaime; de la Mora-de la Mora, Ignacio; Garcia-Torres, Itzhel; Enríquez-Flores, Sergio; Contreras-Zentella, Martha Lucinda; Arreguín-Espinosa, Roberto; Kroneck, Peter M. H.; Sosa-Torres, Martha Elena

    2015-01-01

    Gluconacetobacter diazotrophicus is a N2-fixing bacterium endophyte from sugar cane. The oxidation of ethanol to acetic acid of this organism takes place in the periplasmic space, and this reaction is catalyzed by two membrane-bound enzymes complexes: the alcohol dehydrogenase (ADH) and the aldehyde dehydrogenase (ALDH). We present strong evidence showing that the well-known membrane-bound Alcohol dehydrogenase (ADHa) of Ga. diazotrophicus is indeed a double function enzyme, which is able to use primary alcohols (C2–C6) and its respective aldehydes as alternate substrates. Moreover, the enzyme utilizes ethanol as a substrate in a reaction mechanism where this is subjected to a two-step oxidation process to produce acetic acid without releasing the acetaldehyde intermediary to the media. Moreover, we propose a mechanism that, under physiological conditions, might permit a massive conversion of ethanol to acetic acid, as usually occurs in the acetic acid bacteria, but without the transient accumulation of the highly toxic acetaldehyde. PMID:25574602

  10. RELIGION AND PURIFICATION OF SOUL

    Directory of Open Access Journals (Sweden)

    Azam Khodashenas Pelko

    2010-11-01

    Full Text Available The Jainism emphasizes three major teachings about the purification of the soul (jiva, Ahimsa, Aparigrapha and anekantwad. Jainism, The focus of this religion has been purification of the soul by means of right conduct, right faith and right knowledge. The ultimate goal of Hinduism is Moksha or liberation (total freedom. In Hinduism, purification of the soul is a goal that one must work to attain. The Buddhism is the science of pursuing the aim of making the human mind perfect, and of purifying the human soul. The knowledge of purifying of the soul and softening of the hearts is as essential for human. They having the correct motivations means purifying our souls from hypocrisy, caprice, and heedlessness. The primary goal of Taoism may be described as the mystical intuition of the Tao, which is the way, the undivided unity, and the ultimate Reality. According to the Christianity access to truth cannot be conceived without purity of the soul

  11. Purification of Water by Aquatic Plants

    OpenAIRE

    Morimitsu, Katsuhito; Kawahigashi, Tatsuo

    2013-01-01

    [Abstract] Water quality purification of many water systems including those occurring in rivers depends to a great degree on water quality purification activities of aquatic plants and microbes. This paper presents a discussion of results, based on laboratory experiments, of purification by aquatic plants.

  12. Partial Purification and Characterization of Extracellular Protease ...

    African Journals Online (AJOL)

    Nigerian Journal of Basic and Applied Sciences ... Purification of the enzyme by gel filtration chromatography on Sephadex G75 following ammonium sulphate precipitation gave 2.26 fold increase in purification with specific activity of 46.13 units/mg protein while purification on Sephadex CM50 resulted in reduced ...

  13. Preliminary studies on the purification of a monoclonal antibody by affinity precipitation with Eudragit S-100.

    Science.gov (United States)

    Taipa, M A; Kaul, R; Mattiasson, B; Cabral, J M

    1998-01-01

    A simple procedure for the purification of an IgG-type monoclonal antibody by affinity precipitation using Eudragit S-100 is presented. The ligand, a microbial lipase previously used as antigen, was coupled to the polymer at a concentration of 40 mg lipase/g Eudragit. This macroligand was reversibly precipitated by manipulating the pH at values higher and lower than 4.8. The effects of polymer concentration and dilution of hybridoma culture supernatant on the overall precipitation process were evaluated. The best purification factor was achieved with a polymer concentration of 0.1% (w/v) and a supernatant dilution of 1:3. The preliminary studies reported here enabled the purification of a monoclonal antibody in one step with an activity yield (by ELISA) of 50%-55% and a purification factor of ca 6.

  14. Single-step synthesis of Er3+ and Yb3+ ions doped molybdate/Gd2O3 core–shell nanoparticles for biomedical imaging

    Science.gov (United States)

    Kamińska, Izabela; Elbaum, Danek; Sikora, Bożena; Kowalik, Przemysław; Mikulski, Jakub; Felcyn, Zofia; Samol, Piotr; Wojciechowski, Tomasz; Minikayev, Roman; Paszkowicz, Wojciech; Zaleszczyk, Wojciech; Szewczyk, Maciej; Konopka, Anna; Gruzeł, Grzegorz; Pawlyta, Mirosława; Donten, Mikołaj; Ciszak, Kamil; Zajdel, Karolina; Frontczak-Baniewicz, Małgorzata; Stępień, Piotr; Łapiński, Mariusz; Wilczyński, Grzegorz; Fronc, Krzysztof

    2018-01-01

    Nanostructures as color-tunable luminescent markers have become major, promising tools for bioimaging and biosensing. In this paper separated molybdate/Gd2O3 doped rare earth ions (erbium, Er3+ and ytterbium, Yb3+) core–shell nanoparticles (NPs), were fabricated by a one-step homogeneous precipitation process. Emission properties were studied by cathodo- and photoluminescence. Scanning electron and transmission electron microscopes were used to visualize and determine the size and shape of the NPs. Spherical NPs were obtained. Their core–shell structures were confirmed by x-ray diffraction and energy-dispersive x-ray spectroscopy measurements. We postulated that the molybdate rich core is formed due to high segregation coefficient of the Mo ion during the precipitation. The calcination process resulted in crystallization of δ/ξ (core/shell) NP doped Er and Yb ions, where δ—gadolinium molybdates and ξ—molybdates or gadolinium oxide. We confirmed two different upconversion mechanisms. In the presence of molybdenum ions, in the core of the NPs, Yb3+–{{{{MoO}}}4}2- (∣2F7/2, 3T2〉) dimers were formed. As a result of a two 980 nm photon absorption by the dimer, we observed enhanced green luminescence in the upconversion process. However, for the shell formed by the Gd2O3:Er, Yb NPs (without the Mo ions), the typical energy transfer upconversion takes place, which results in red luminescence. We demonstrated that the NPs were transported into cytosol of the HeLa and astrocytes cells by endocytosis. The core–shell NPs are sensitive sensors for the environment prevailing inside (shorter luminescence decay) and outside (longer luminescence decay) of the tested cells. The toxicity of the NPs was examined using MTT assay.

  15. Single-step synthesis of Er3+and Yb3+ions doped molybdate/Gd2O3core-shell nanoparticles for biomedical imaging.

    Science.gov (United States)

    Kamińska, Izabela; Elbaum, Danek; Sikora, Bożena; Kowalik, Przemysław; Mikulski, Jakub; Felcyn, Zofia; Samol, Piotr; Wojciechowski, Tomasz; Minikayev, Roman; Paszkowicz, Wojciech; Zaleszczyk, Wojciech; Szewczyk, Maciej; Konopka, Anna; Gruzeł, Grzegorz; Pawlyta, Mirosława; Donten, Mikołaj; Ciszak, Kamil; Zajdel, Karolina; Frontczak-Baniewicz, Małgorzata; Stępień, Piotr; Łapiński, Mariusz; Wilczyński, Grzegorz; Fronc, Krzysztof

    2018-01-12

    Nanostructures as color-tunable luminescent markers have become major, promising tools for bioimaging and biosensing. In this paper separated molybdate/Gd 2 O 3 doped rare earth ions (erbium, Er 3+ and ytterbium, Yb 3+ ) core-shell nanoparticles (NPs), were fabricated by a one-step homogeneous precipitation process. Emission properties were studied by cathodo- and photoluminescence. Scanning electron and transmission electron microscopes were used to visualize and determine the size and shape of the NPs. Spherical NPs were obtained. Their core-shell structures were confirmed by x-ray diffraction and energy-dispersive x-ray spectroscopy measurements. We postulated that the molybdate rich core is formed due to high segregation coefficient of the Mo ion during the precipitation. The calcination process resulted in crystallization of δ/ξ (core/shell) NP doped Er and Yb ions, where δ-gadolinium molybdates and ξ-molybdates or gadolinium oxide. We confirmed two different upconversion mechanisms. In the presence of molybdenum ions, in the core of the NPs, Yb 3+ -[Formula: see text] (∣ 2 F 7/2 , 3 T 2 〉) dimers were formed. As a result of a two 980 nm photon absorption by the dimer, we observed enhanced green luminescence in the upconversion process. However, for the shell formed by the Gd 2 O 3 :Er, Yb NPs (without the Mo ions), the typical energy transfer upconversion takes place, which results in red luminescence. We demonstrated that the NPs were transported into cytosol of the HeLa and astrocytes cells by endocytosis. The core-shell NPs are sensitive sensors for the environment prevailing inside (shorter luminescence decay) and outside (longer luminescence decay) of the tested cells. The toxicity of the NPs was examined using MTT assay.

  16. Optimization of genetic analysis for single cell

    Directory of Open Access Journals (Sweden)

    hussein mouawia

    2012-03-01

    Full Text Available The molecular genetic analysis of microdissected cells by laser, a method for selecting a starting material of pure DNA or RNA uncontaminated. Our study focuses on technical pre-PCR (polymerase chain reaction for the amplification of DNA from a single cell (leukocyte isolated from human blood after laser microdissection and aims to optimize the yield of DNA extracted of this cell to be amplified without errors and provide reliable genetic analyzes. This study has allowed us to reduce the duration of cell lysis in order to perform the step of expanding genomic PEP (primer extension preamplification directly after lysis the same day and the quality of genomic amplification and eliminate purification step of the product PEP, step with a risk of contamination and risk of loss of genetic material related to manipulation. This approach has shown that the combination of at least 3 STR (short tandem repeat markers for genetic analysis of single cell improves the efficiency and accuracy of PCR and minimizes the loss of allele (allele drop out; ADO. This protocol can be applied to large scale and an effective means suitable for genetic testing for molecular diagnostic from isolated single cell (cancerous - fetal.

  17. Evaluation of effectiveness of intraoral delivery tip on single step impressions using monophase and two-phase impression materials: An in vivo study

    Directory of Open Access Journals (Sweden)

    S Gowri

    2015-01-01

    Full Text Available Background and Rationale: The precise fit of prosthesis depends on the impression materials and the technique used. Addition silicone impression materials being the user-friendly material are used in monophase and two-phase techniques. Considering the paucity of data on the effectiveness of intraoral delivery tip (IODT on void, tear and bubble formation using monophase impression technique in stock tray and two-phase impression technique in custom tray the present study was initiated. Materials and Methods: A total of 50 patients requiring single complete veneer crowns were enrolled. Monophase and two-phase impressions with and without IODT were made. Evaluation of the impression were made for tears, voids, and bubbles and alpha, beta, delta scores were given. Results: A significant reduction in the total number of voids was observed with two-phase technique either with or without IODT. No significant differences were observed in the proportion of overall evaluation into alpha, beta and delta between monophase and two-phase with and without IODT. No significant association was seen in the overall evaluation between the two groups (monophase and two-phase as compared using Chi-square test for independency and a trend in association between overall evaluation as tested using Chi-square for linear trend analysis. Conclusion: Two-phase impression technique with the use of IODT produced decreased number of voids.

  18. Combined Protein A and size exclusion high performance liquid chromatography for the single-step measurement of mAb, aggregates and host cell proteins.

    Science.gov (United States)

    Gjoka, Xhorxhi; Schofield, Mark; Cvetkovic, Aleksandar; Gantier, Rene

    2014-12-01

    Quantification of monoclonal antibody (mAb) monomer, mAb aggregates, and host cell proteins (HCPs) is critical for the optimization of the mAb production process. The present work describes a single high throughput analytical tool capable of tracking the concentration of mAb, mAb aggregate and HCPs in a growing cell culture batch. By combining two analytical HPLC methods, Protein A affinity and size-exclusion chromatography (SEC), it is possible to detect a relative increase or decrease in the concentration of all three entities simultaneously. A comparison of the combined Protein A-SEC assay to SEC alone was performed, demonstrating that it can be useful tool for the quantification of mAb monomer along with trending data for mAb aggregate and HCP. Furthermore, the study shows that the Protein A-SEC method is at least as accurate as other commonly used analytical methods such as ELISA and Bradford. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Propensity approach to nonequilibrium thermodynamics of a chemical reaction network: controlling single E-coli β-galactosidase enzyme catalysis through the elementary reaction steps.

    Science.gov (United States)

    Das, Biswajit; Banerjee, Kinshuk; Gangopadhyay, Gautam

    2013-12-28

    In this work, we develop an approach to nonequilibrium thermodynamics of an open chemical reaction network in terms of the elementary reaction propensities. The method is akin to the microscopic formulation of the dissipation function in terms of the Kullback-Leibler distance of phase space trajectories in Hamiltonian system. The formalism is applied to a single oligomeric enzyme kinetics at chemiostatic condition that leads the reaction system to a nonequilibrium steady state, characterized by a positive total entropy production rate. Analytical expressions are derived, relating the individual reaction contributions towards the total entropy production rate with experimentally measurable reaction velocity. Taking a real case of Escherichia coli β-galactosidase enzyme obeying Michaelis-Menten kinetics, we thoroughly analyze the temporal as well as the steady state behavior of various thermodynamic quantities for each elementary reaction. This gives a useful insight in the relative magnitudes of various energy terms and the dissipated heat to sustain a steady state of the reaction system operating far-from-equilibrium. It is also observed that, the reaction is entropy-driven at low substrate concentration and becomes energy-driven as the substrate concentration rises.

  20. Purification of Sardine Fish Oil Through Degumming and Neutralization

    Directory of Open Access Journals (Sweden)

    Stephanie Bija

    2017-06-01

    Full Text Available The quality of sardine fish oil can be improved by purification method through the step of degumming and neutralization. The aimed of this this study was analysis characteristic of crude sardin fish oil and determined the best method of purification. Degumming was carried out using 30% water and salt at concentration 5%, 8%, 10% b/v. Neutralization process using  NaOH with 16°Be and bleaching using 5% Magnesol XL. All step of refining was done at 50°C, 60°C, 70°C, and 80°C. The result of analysis showed that sardine crude fish oil had 24.86% of palmitic acid as the highest fatty acid, heavy metal was not detected,dencity was 0.92 g/cm3 and viscocity was 51 cPs. The best treatment of purification method was degumming using 5% NaCl at 50°C with rendement 65.37±0.72%; free fatty acid (FFA 0.38±0.03%; peroxide (PV 1.07±0.12 mEq/kg; anisidine (p-AnV 15.18±0.16 mEq/kg; total oxidation value (TOTOX 17.31±0.39 mEq/kg; and clarity was 75.09± 1.20%.

  1. Single molecule TPM analysis of the catalytic pentad mutants of Cre and Flp site-specific recombinases: contributions of the pentad residues to the pre-chemical steps of recombination.

    Science.gov (United States)

    Fan, Hsiu-Fang; Cheng, Yong-Song; Ma, Chien-Hui; Jayaram, Makkuni

    2015-03-31

    Cre and Flp site-specific recombinase variants harboring point mutations at their conserved catalytic pentad positions were characterized using single molecule tethered particle motion (TPM) analysis. The findings reveal contributions of these amino acids to the pre-chemical steps of recombination. They suggest functional differences between positionally conserved residues in how they influence recombinase-target site association and formation of 'non-productive', 'pre-synaptic' and 'synaptic' complexes. The most striking difference between the two systems is noted for the single conserved lysine. The pentad residues in Cre enhance commitment to recombination by kinetically favoring the formation of pre-synaptic complexes. These residues in Flp serve a similar function by promoting Flp binding to target sites, reducing non-productive binding and/or enhancing the rate of assembly of synaptic complexes. Kinetic comparisons between Cre and Flp, and between their derivatives lacking the tyrosine nucleophile, are consistent with a stronger commitment to recombination in the Flp system. The effect of target site orientation (head-to-head or head-to-tail) on the TPM behavior of synapsed DNA molecules supports the selection of anti-parallel target site alignment prior to the chemical steps. The integrity of the synapse, whose establishment/stability is fostered by strand cleavage in the case of Flp but not Cre, appears to be compromised by the pentad mutations. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  2. Single molecule TPM analysis of the catalytic pentad mutants of Cre and Flp site-specific recombinases: contributions of the pentad residues to the pre-chemical steps of recombination

    Science.gov (United States)

    Fan, Hsiu-Fang; Cheng, Yong-Song; Ma, Chien-Hui; Jayaram, Makkuni

    2015-01-01

    Cre and Flp site-specific recombinase variants harboring point mutations at their conserved catalytic pentad positions were characterized using single molecule tethered particle motion (TPM) analysis. The findings reveal contributions of these amino acids to the pre-chemical steps of recombination. They suggest functional differences between positionally conserved residues in how they influence recombinase-target site association and formation of ‘non-productive’, ‘pre-synaptic’ and ‘synaptic’ complexes. The most striking difference between the two systems is noted for the single conserved lysine. The pentad residues in Cre enhance commitment to recombination by kinetically favoring the formation of pre-synaptic complexes. These residues in Flp serve a similar function by promoting Flp binding to target sites, reducing non-productive binding and/or enhancing the rate of assembly of synaptic complexes. Kinetic comparisons between Cre and Flp, and between their derivatives lacking the tyrosine nucleophile, are consistent with a stronger commitment to recombination in the Flp system. The effect of target site orientation (head-to-head or head-to-tail) on the TPM behavior of synapsed DNA molecules supports the selection of anti-parallel target site alignment prior to the chemical steps. The integrity of the synapse, whose establishment/stability is fostered by strand cleavage in the case of Flp but not Cre, appears to be compromised by the pentad mutations. PMID:25765648

  3. Methods in elastic tissue biology: elastin isolation and purification.

    Science.gov (United States)

    Mecham, Robert P

    2008-05-01

    Elastin provides recoil to tissues subjected to repeated stretch, such as blood vessels and the lung. It is encoded by a single gene in mammals and is secreted as a 60-70 kDa monomer called tropoelastin. The functional form of the protein is that of a large, highly crosslinked polymer that organizes as sheets or fibers in the extracellular matrix. Purification of mature, crosslinked elastin is problematic because its insolubility precludes its isolation using standard wet-chemistry techniques. Instead, relatively harsh experimental approaches designed to remove non-elastin 'contaminates' are employed to generate an insoluble product that has the amino acid composition expected of elastin. Although soluble, tropoelastin also presents problems for isolation and purification. The protein's extreme stickiness and susceptibility to proteolysis requires careful attention during purification and in tropoelastin-based assays. This article describes the most common approaches for purification of insoluble elastin and tropoelastin. It also addresses key aspects of studying tropoelastin production in cultured cells, where elastin expression is highly dependent upon cell type, culture conditions, and passage number.

  4. PURIFICATION AND CHARACTERISATION OF ALKALINE ...

    African Journals Online (AJOL)

    Prof. Adipala Ekwamu

    phosphatase activity upon phosphate deprivation with much of the enzyme released into the medium during osmotic shock, indicating .... shaking. Protein purification. Three millilitres of overnight culture (400 ml) was harvested by centrifugation at 6000x g for 1 min. The supernatant was decanted and pellet washed twice in ...

  5. Bioinspired Materials for Water Purification

    Directory of Open Access Journals (Sweden)

    Alfredo Gonzalez-Perez

    2016-06-01

    Full Text Available Water scarcity issues associated with inadequate access to clean water and sanitation is a ubiquitous problem occurring globally. Addressing future challenges will require a combination of new technological development in water purification and environmental remediation technology with suitable conservation policies. In this scenario, new bioinspired materials will play a pivotal role in the development of more efficient and environmentally friendly solutions. The role of amphiphilic self-assembly on the fabrication of new biomimetic membranes for membrane separation like reverse osmosis is emphasized. Mesoporous support materials for semiconductor growth in the photocatalytic degradation of pollutants and new carriers for immobilization of bacteria in bioreactors are used in the removal and processing of different kind of water pollutants like heavy metals. Obstacles to improve and optimize the fabrication as well as a better understanding of their performance in small-scale and pilot purification systems need to be addressed. However, it is expected that these new biomimetic materials will find their way into the current water purification technologies to improve their purification/removal performance in a cost-effective and environmentally friendly way.

  6. The Protein Maker: an automated system for high-throughput parallel purification

    International Nuclear Information System (INIS)

    Smith, Eric R.; Begley, Darren W.; Anderson, Vanessa; Raymond, Amy C.; Haffner, Taryn E.; Robinson, John I.; Edwards, Thomas E.; Duncan, Natalie; Gerdts, Cory J.; Mixon, Mark B.; Nollert, Peter; Staker, Bart L.; Stewart, Lance J.

    2011-01-01

    The Protein Maker instrument addresses a critical bottleneck in structural genomics by allowing automated purification and buffer testing of multiple protein targets in parallel with a single instrument. Here, the use of this instrument to (i) purify multiple influenza-virus proteins in parallel for crystallization trials and (ii) identify optimal lysis-buffer conditions prior to large-scale protein purification is described. The Protein Maker is an automated purification system developed by Emerald BioSystems for high-throughput parallel purification of proteins and antibodies. This instrument allows multiple load, wash and elution buffers to be used in parallel along independent lines for up to 24 individual samples. To demonstrate its utility, its use in the purification of five recombinant PB2 C-terminal domains from various subtypes of the influenza A virus is described. Three of these constructs crystallized and one diffracted X-rays to sufficient resolution for structure determination and deposition in the Protein Data Bank. Methods for screening lysis buffers for a cytochrome P450 from a pathogenic fungus prior to upscaling expression and purification are also described. The Protein Maker has become a valuable asset within the Seattle Structural Genomics Center for Infectious Disease (SSGCID) and hence is a potentially valuable tool for a variety of high-throughput protein-purification applications

  7. Two-step irradiance schedule versus single-dose tramadol sustained-release tablets for pain control during topical 5-aminolevulinic acid-photodynamic therapy of condyloma acuminatum in Chinese patients: a randomized comparative study.

    Science.gov (United States)

    Mchepange, Uwesu O; Huang, Chun-Yan; Sun, Yi; Tu, Ya-Ting; Tao, Juan

    2014-07-01

    Photodynamic therapy with 5-aminolevulinic acid (ALA-PDT) offers promising results for the treatment of condyloma acuminatum. However, patients have to dwell with pain to benefit from this otherwise effective and safe "off-label" treatment modality. Several techniques have been explored to control ALA-PDT-induced pain, but the desperate search for a universally accepted method is still ongoing. This study compares the two-step irradiance approach with single-dose administration of 100 mg tramadol sustained-release tablets for pain induced by ALA-PDT of condyloma acuminatum in Chinese patients. Adult Chinese patients with condyloma acuminatum were enrolled in a randomized comparative study. Pain levels were compared using the Numeric Rating Scale (NRS) at pre-defined assessment points during and after irradiation. The pain was dominated by characteristics such as burning and pricking and was almost always local and superficial. The median pain scores were lower in the two-step irradiance group at 1 minute (U = 621.5, P = 0.002) but higher at 20 minutes (U = 585.5, P = 0.002). The median pain scores between the two groups did not differ significantly at other assessment points. The pain was moderate in both groups and peaked earlier in the analgesics group (median: 5 minutes) but later in the two-step irradiance group (median: 15 minutes). The pain was generally mild. The median pain scores were equal at each assessment point, except at 3 hours where the median was lower in the analgesics group (1.0) as compared with the two-step irradiance group (2.0) (U = 725.0, P = 0.056). Pain in the two-step irradiance protocol is irradiance-dependent. The two-step irradiance approach produces significant benefits over analgesics during the initial stages of therapy but analgesics offer significant benefits thereafter. There are potential benefits of combining the two approaches in minimizing ALA-PDT-induced pain. © 2014 Wiley Periodicals

  8. Application of stepping motor

    International Nuclear Information System (INIS)

    1980-10-01

    This book is divided into three parts, which is about practical using of stepping motor. The first part has six chapters. The contents of the first part are about stepping motor, classification of stepping motor, basic theory og stepping motor, characteristic and basic words, types and characteristic of stepping motor in hybrid type and basic control of stepping motor. The second part deals with application of stepping motor with hardware of stepping motor control, stepping motor control by microcomputer and software of stepping motor control. The last part mentions choice of stepping motor system, examples of stepping motor, measurement of stepping motor and practical cases of application of stepping motor.

  9. Isolation and Purification of Biotechnological Products

    Science.gov (United States)

    Hubbuch, Jürgen; Kula, Maria-Regina

    2007-05-01

    The production of modern pharma proteins is one of the most rapid growing fields in biotechnology. The overall development and production is a complex task ranging from strain development and cultivation to the purification and formulation of the drug. Downstream processing, however, still accounts for the major part of production costs. This is mainly due to the high demands on purity and thus safety of the final product and results in processes with a sequence of typically more than 10 unit operations. Consequently, even if each process step would operate at near optimal yield, a very significant amount of product would be lost. The majority of unit operations applied in downstream processing have a long history in the field of chemical and process engineering; nevertheless, mathematical descriptions of the respective processes and the economical large-scale production of modern pharmaceutical products are hampered by the complexity of the biological feedstock, especially the high molecular weight and limited stability of proteins. In order to develop new operational steps as well as a successful overall process, it is thus a necessary prerequisite to develop a deeper understanding of the thermodynamics and physics behind the applied processes as well as the implications for the product.

  10. Large-Scale Low-Cost NGS Library Preparation Using a Robust Tn5 Purification and Tagmentation Protocol

    Directory of Open Access Journals (Sweden)

    Bianca P. Hennig

    2018-01-01

    Full Text Available Efficient preparation of high-quality sequencing libraries that well represent the biological sample is a key step for using next-generation sequencing in research. Tn5 enables fast, robust, and highly efficient processing of limited input material while scaling to the parallel processing of hundreds of samples. Here, we present a robust Tn5 transposase purification strategy based on an N-terminal His6-Sumo3 tag. We demonstrate that libraries prepared with our in-house Tn5 are of the same quality as those processed with a commercially available kit (Nextera XT, while they dramatically reduce the cost of large-scale experiments. We introduce improved purification strategies for two versions of the Tn5 enzyme. The first version carries the previously reported point mutations E54K and L372P, and stably produces libraries of constant fragment size distribution, even if the Tn5-to-input molecule ratio varies. The second Tn5 construct carries an additional point mutation (R27S in the DNA-binding domain. This construct allows for adjustment of the fragment size distribution based on enzyme concentration during tagmentation, a feature that opens new opportunities for use of Tn5 in customized experimental designs. We demonstrate the versatility of our Tn5 enzymes in different experimental settings, including a novel single-cell polyadenylation site mapping protocol as well as ultralow input DNA sequencing.

  11. Purification and Characterization of BmooAi: A New Toxin from Bothrops moojeni Snake Venom That Inhibits Platelet Aggregation

    Directory of Open Access Journals (Sweden)

    Mayara Ribeiro de Queiroz

    2014-01-01

    Full Text Available In this paper, we describe the purification/characterization of BmooAi, a new toxin from Bothrops moojeni that inhibits platelet aggregation. The purification of BmooAi was carried out through three chromatographic steps (ion-exchange on a DEAE-Sephacel column, molecular exclusion on a Sephadex G-75 column, and reverse-phase HPLC chromatography on a C2/C18 column. BmooAi was homogeneous by SDS-PAGE and shown to be a single-chain protein of 15,000 Da. BmooAi was analysed by MALDI-TOF Spectrometry and revealed two major components with molecular masses 7824.4 and 7409.2 as well as a trace of protein with a molecular mass of 15,237.4 Da. Sequencing of BmooAi by Edman degradation showed two amino acid sequences: IRDFDPLTNAPENTA and ETEEGAEEGTQ, which revealed no homology to any known toxin from snake venom. BmooAi showed a rather specific inhibitory effect on platelet aggregation induced by collagen, adenosine diphosphate, or epinephrine in human platelet-rich plasma in a dose-dependent manner, whereas it had little or no effect on platelet aggregation induced by ristocetin. The effect on platelet aggregation induced by BmooAi remained active even when heated to 100°C. BmooAi could be of medical interest as a new tool for the development of novel therapeutic agents for the prevention and treatment of thrombotic disorders.

  12. Purification and Biochemical Characterization of Three Myotoxins from Bothrops mattogrossensis Snake Venom with Toxicity against Leishmania and Tumor Cells

    Directory of Open Access Journals (Sweden)

    Andréa A. de Moura

    2014-01-01

    Full Text Available Bothrops mattogrossensis snake is widely distributed throughout eastern South America and is responsible for snakebites in this region. This paper reports the purification and biochemical characterization of three new phospholipases A2 (PLA2s, one of which is presumably an enzymatically active Asp49 and two are very likely enzymatically inactive Lys49 PLA2 homologues. The purification was obtained after two chromatographic steps on ion exchange and reverse phase column. The 2D SDS-PAGE analysis revealed that the proteins have pI values around 10, are each made of a single chain, and have molecular masses near 13 kDa, which was confirmed by MALDI-TOF mass spectrometry. The N-terminal similarity analysis of the sequences showed that the proteins are highly homologous with other Lys49 and Asp49 PLA2s from Bothrops species. The PLA2s isolated were named BmatTX-I (Lys49 PLA2-like, BmatTX-II (Lys49 PLA2-like, and BmatTX-III (Asp49 PLA2. The PLA2s induced cytokine release from mouse neutrophils and showed cytotoxicity towards JURKAT (leukemia T and SK-BR-3 (breast adenocarcinoma cell lines and promastigote forms of Leishmania amazonensis. The structural and functional elucidation of snake venoms components may contribute to a better understanding of the mechanism of action of these proteins during envenomation and their potential pharmacological and therapeutic applications.

  13. Purification and Characterization of an Extracellular Dextransucrase from Pediococcus pentosaceus Isolated from the Soil of North East India

    Directory of Open Access Journals (Sweden)

    Damini Kothari

    2011-01-01

    Full Text Available The extracellular dextransucrase produced from Pediococcus pentosaceus, a new isolate from the soil in Assam, India, was purified and characterized. The enzyme activity of cell-free supernatant was 3.4 U/mL and specific activity was 0.6 U/mg. The crude enzyme was purified by a single-step fractionation using polyethylene glycols of different molecular mass. The specific activity achieved was 18 U/mg with 31-fold purification by PEG 400 and 26 U/mg with 45-fold purification by PEG 1500. The molecular mass of dextransucrase determined by non-denaturing SDS-PAGE was approx. 180 kDa. The dextran formation activity of the enzyme was confirmed by activity staining. Optimum conditions for dextransucrase activity were: pH=5.4, reaction temperature 30 °C, 5 % sucrose and 20 mM sodium acetate buffer. A concentration of 1 mM MgCl2 and 6 mM CaCl2 enhanced dextransucrase activity by 5 and 150 %, respectively. The chaotropic agent urea (7 M and chelating agent EDTA (1 mM resulted in the residual enzyme activity of 98 and 80 %, respectively. The organic solvents such as ethanol (50 %, DMSO (90 %, acetone (50 % and acetonitrile (20 % decreased the dextransucrase activity by 80, 91, 94 and 80 %, respectively.

  14. High yield purification of nanobodies from the periplasm of E. coli as fusions with the maltose binding protein.

    Science.gov (United States)

    Salema, Valencio; Fernández, Luis Ángel

    2013-09-01

    Nanobodies (Nbs) are single domain antibodies based on the variable domains of heavy chain only antibodies (HCAbs) found in camelids, also referred to as VHHs. Their small size (ca. 12-15kDa), superior biophysical and antigen binding properties have made Nbs very attractive molecules for multiple biotechnological applications, including human therapy. The most widely used system for the purification of Nbs is their expression in the periplasm of Escherichia coli with a C-terminal hexa-histidine (His6) tag followed by immobilized metal affinity chromatography (IMAC). However, significant variability in the expression levels of different Nbs are routinely observed and a single affinity chromatography step is often not sufficient to obtain Nbs of high purity. Here, we report an alternative method for expression and purification of Nbs from the periplasm of E. coli based on their fusion to maltose binding protein (MBP) in the N-terminus and His6 tag in the C-terminus (MBP-NbHis6). Soluble MBP-NbHis6 fusions were consistently expressed at high levels (⩾12mg/L of induced culture in shake flasks) in the periplasm of E. coli HM140, a strain deficient in several periplasmic proteases. Highly pure MBP-NbHis6 fusions and free NbHis6 (after site specific proteolysis of the fusions), were recovered by amylose and metal affinity chromatography steps. The monomeric nature of the purified NbHis6 was determined by gel filtration chromatography. Lastly, we demonstrated by ELISA that both monomeric NbHis6 and MBP-NbHis6 fusions retained antigen binding activity and specificity, thus facilitating their direct use in antigen recognition assays. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. 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.

  16. Rapid single-step methods for detection of two immune defence gene polymorphisms: the myeloperoxidase (MPO) G-129A and the Fc gamma receptor 2A (FCGR2A) H/R131

    DEFF Research Database (Denmark)

    Mølle, Ingolf; Melsvik, Dorte; Østergaard, Mette

    2007-01-01

    . formalin-fixed paraffin-embedded tissue specimens. Here we describe two new single-step methods for rapid and sensitive analysis of: 1. The G-129A myeloperoxidase (MPO) promoter polymorphism, which affects the amount of myeloperoxidase in neutrophils. 2. The Fc gamma receptor 2A (FCGR2A)-H/R131......Polymorphisms of immune defence genes may act as disease modifiers and are studied by many researchers. A conclusive analysis of the impact of genetic variations typically requires a large number of sample specimens, and in retrospective studies this may include samples of reduced quality, e.g...... polymorphism, which is critical to the binding of IgG2 immune complexes to phagocytes. Udgivelsesdato: 2007-Jul-31...

  17. Isolation and purification of alkaline keratinase from Bacillus sp. 50-3

    African Journals Online (AJOL)

    PAGE, and the molecular weight of the enzyme was found at 27 423 Da by the MALDI-TOF-MS. Its simple purification step and high yield using a cheap medium attest to the great biotechnological potential of keratinase, especially in environment

  18. Methods for Isolation, Purification, and Propagation of Bacteriophages of Campylobacter jejuni

    DEFF Research Database (Denmark)

    Gencay, Yilmaz Emre; Birk, Tina; Sørensen, Martine Camilla Holst

    2017-01-01

    Here, we describe the methods for isolation, purification, and propagation of Campylobacter jejuni bacteriophages from samples expected to contain high number of phages such as chicken feces. The overall steps are (1) liberation of phages from the sample material; (2) observation of plaque-formin...

  19. Some Calculated (p,α Cross-Sections Using the Alpha Particle Knock-On and Triton Pick-Up Reaction Mechanisms: An Optimisation of the Single-Step Feshbach–Kerman–Koonin (FKK Theory

    Directory of Open Access Journals (Sweden)

    Felix S. Olise

    2016-04-01

    Full Text Available The Feshbach–Kerman–Koonin (FKK multi-step direct (MSD theory of pre-equilibrium reactions has been used to compute the single-step cross-sections for some (p,α reactions using the knock-on and pick-up reaction mechanisms at two incident proton energies. For the knock-on mechanism, the reaction was assumed to have taken place by the direct ejection of a preformed alpha cluster in a shell-model state of the target. But the reaction was assumed to have taken place by the pick-up of a preformed triton cluster (also bound in a shell-model state of the target core by the incident proton for the pick-up mechanism. The Yukawa forms of potential were used for the proton-alpha (for the knock-on process and proton-triton (for the pick-up process interaction and several parameter sets for the proton and alpha-particle optical potentials. The calculated cross-sections for both mechanisms gave satisfactory fits to the experimental data. Furthermore, it has been shown that some combinations of the calculated distorted wave Born approximation cross-sections for the two reaction mechanisms in the FKK MSD theory are able to give better fits to the experimental data, especially in terms of range of agreement. In addition, the theory has been observed to be valid over a wider range of energy.

  20. Multicolored Cd(1-x)Zn(x)Se quantum dots with type-I core/shell structure: single-step synthesis and their use as light emitting diodes.

    Science.gov (United States)

    Pu, Ying-Chih; Hsu, Yung-Jung

    2014-04-07

    We developed a single-step hot-injection process to synthesize Cd1-xZnxSe quantum dots (QDs) with tunable emission wavelengths. The multiple emission colors of the Cd1-xZnxSe QDs resulted from the variation in their compositions (x value) with the reaction time. Because of the higher reactivity of the Cd precursor, QDs whose composition was rich in CdSe were generated at the beginning of the reaction. As the reaction proceeded, the later-formed ZnSe shell was simultaneously alloyed with the core, giving rise to a progressive alloying treatment for the grown QDs. During the reaction period, the emission color of the Cd1-xZnxSe QDs shifted from red to orange, to yellow, to green and finally to blue. A light emitting diode (LED) composed of multilayers of ITO/poly(3,4-ethylenedioxythiophene):poly(4-styrenesulfonate)/poly(3-hexylthiophene) blended with Cd1-xZnxSe QDs/Al was fabricated to test the electroluminescence (EL) properties of the QDs. The EL results show high color purity for the emission from LED devices containing Cd1-xZnxSe QDs, revealing that the as-synthesized QDs can be easily processed and integrated into a light-emitting device without using a complicated procedure. The findings from the present work also demonstrate the advantage of using the current single-step synthetic approach to obtain a batch of Cd1-xZnxSe QDs that may emit different colors in prototype LEDs.

  1. A multicenter study for a single, three-step laser treatment for cellulite using a 1440-nm Nd:YAG laser, a novel side-firing fiber, and a temperature-sensing cannula.

    Science.gov (United States)

    DiBernardo, Barry; Sasaki, Gordon; Katz, Bruce E; Hunstad, Joseph P; Petti, Christine; Burns, A Jay

    2013-05-01

    Historically, treatments for cellulite have not been able to address all of its physiological components and require multiple sessions. The authors evaluate the safety and efficacy of a single, subdermal procedure to treat the underlying structure of cellulite. Fifty-seven patients underwent a 3-step cellulite treatment with a 1440-nm Nd:YAG laser with a side-firing fiber and temperature-sensing cannula. Efficacy was measured by the ability of blinded evaluators to distinguish baseline photos from those taken at 3 and 6 months posttreatment, as well as their rating of the results on a 5-point, 2-category ordinal photonumeric scale when comparing baseline photos to those taken at 2, 3, and 6 months posttreatment. Patient and physician satisfaction was assessed based on completion of a satisfaction survey at 2, 3, and 6 months posttreatment. Adverse events (AE) were recorded throughout the study. At 6 months posttreatment, blinded evaluators rated at least a 1-point improvement in the appearance of cellulite in 96% of treated sites. Blinded evaluators were also able to correctly identify baseline versus posttreatment photos in 95% of cases. At least 90% of patients and physicians reported satisfaction with the results of treatment throughout 6 months. AE were mild in intensity and transient to treatment. A single, 3-step, minimally invasive laser treatment using a 1440-nm Nd:YAG laser, side-firing fiber, and temperature-sensing cannula to treat the underlying structure of cellulite proved to be safe and maintained effectiveness at least 6 months posttreatment. 2.

  2. Optimized Expression and Purification for High-Activity Preparations of Algal [FeFe]-Hydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Yacoby, I.; Tegler, L. T.; Pochekailov, S.; Zhang, S.; King, P. W.

    2012-04-01

    Recombinant expression and purification of metallo-enzymes, including hydrogenases, at high-yields is challenging due to complex, and enzyme specific, post-translational maturation processes. Low fidelities of maturation result in preparations containing a significant fraction of inactive, apo-protein that are not suitable for biophysical or crystallographic studies. We describe the construction, overexpression and high-yield purification of a fusion protein consisting of the algal [2Fe2S]-ferredoxin PetF (Fd) and [FeFe]-hydrogenase HydA1. The maturation of Fd-HydA1 was optimized through improvements in culture conditions and media components used for expression. We also demonstrated that fusion of Fd to the N-terminus of HydA1, in comparison to the C-terminus, led to increased expression levels that were 4-fold higher. Together, these improvements led to enhanced HydA1 activity and improved yield after purification. The strong binding-affinity of Fd for DEAE allowed for two-step purification by ion exchange and StrepTactin affinity chromatography. In addition, the incorporation of a TEV protease site in the Fd-HydA1 linker allowed for the proteolytic removal of Fd after DEAE step, and purification of HydA1 alone by StrepTactin. In combination, this process resulted in HydA1 purification yields of 5 mg L{sup -1} of culture from E. coli with specific activities of 1000 U (U = 1 {micro}mol hydrogen evolved mg{sup -1} min{sup -1}). The [FeFe]-hydrogenases are highly efficient enzymes and their catalytic sites provide model structures for synthetic efforts to develop robust hydrogen activation catalysts. In order to characterize their structure-function properties in greater detail, and to use hydrogenases for biotechnological applications, reliable methods for rapid, high-yield expression and purification are required.

  3. Droplet Microfluidics for Compartmentalized Cell Lysis and Extension of DNA from Single-Cells

    Science.gov (United States)

    Zimny, Philip; Juncker, David; Reisner, Walter

    Current single cell DNA analysis methods suffer from (i) bias introduced by the need for molecular amplification and (ii) limited ability to sequence repetitive elements, resulting in (iii) an inability to obtain information regarding long range genomic features. Recent efforts to circumvent these limitations rely on techniques for sensing single molecules of DNA extracted from single-cells. Here we demonstrate a droplet microfluidic approach for encapsulation and biochemical processing of single-cells inside alginate microparticles. In our approach, single-cells are first packaged inside the alginate microparticles followed by cell lysis, DNA purification, and labeling steps performed off-chip inside this microparticle system. The alginate microparticles are then introduced inside a micro/nanofluidic system where the alginate is broken down via a chelating buffer, releasing long DNA molecules which are then extended inside nanofluidic channels for analysis via standard mapping protocols.

  4. Statistical and Judgmental Criteria for Scale Purification

    DEFF Research Database (Denmark)

    Wieland, Andreas; Durach, Christian F.; Kembro, Joakim

    2017-01-01

    of scale purification, to critically analyze the current state of scale purification in supply chain management (SCM) research and to provide suggestions for advancing the scale-purification process. Design/methodology/approach A framework for making scale-purification decisions is developed and used......Purpose “Scale purification” – the process of eliminating items from multi-item scales – is widespread in empirical research, but studies that critically examine the implications of this process are scarce. The goals of this research are threefold: to discuss the methodological underpinning...... to analyze and critically reflect on the application of scale purification in leading SCM journals. Findings This research highlights the need for rigorous scale-purification decisions based on both statistical and judgmental criteria. By applying the proposed framework to the SCM discipline, a lack...

  5. Water purification using organic salts

    Science.gov (United States)

    Currier, Robert P.

    2004-11-23

    Water purification using organic salts. Feed water is mixed with at least one organic salt at a temperature sufficiently low to form organic salt hydrate crystals and brine. The crystals are separated from the brine, rinsed, and melted to form an aqueous solution of organic salt. Some of the water is removed from the aqueous organic salt solution. The purified water is collected, and the remaining more concentrated aqueous organic salt solution is reused.

  6. Purification and characterisation of dsRNA using ion pair reverse phase chromatography and mass spectrometry.

    Science.gov (United States)

    Nwokeoji, Alison O; Kung, An-Wen; Kilby, Peter M; Portwood, David E; Dickman, Mark J

    2017-02-10

    RNA interference has provided valuable insight into a wide range of biological systems and is a powerful tool for the analysis of gene function. The exploitation of this pathway to block the expression of specific gene targets holds considerable promise for the development of novel RNAi-based insect management strategies. In addition, there are a wide number of future potential applications of RNAi to control agricultural insect pests as well as its use for prevention of diseases in beneficial insects. The potential to synthesise large quantities of dsRNA by in-vitro transcription or in bacterial systems for RNA interference applications has generated significant demand for the development and application of high throughput analytical tools for the rapid extraction, purification and analysis of dsRNA. Here we have developed analytical methods that enable the rapid purification of dsRNA from associated impurities from bacterial cells in conjunction with downstream analyses. We have optimised TRIzol extractions in conjunction with a single step protocol to remove contaminating DNA and ssRNA, using RNase T1/DNase I digestion under high-salt conditions in combination with solid phase extraction to purify the dsRNA. In addition, we have utilised and developed IP RP HPLC for the rapid, high resolution analysis of the dsRNA. Furthermore, we have optimised base-specific cleavage of dsRNA by RNase A and developed a novel method utilising RNase T1 for RNase mass mapping approaches to further characterise the dsRNA using liquid chromatography interfaced with mass spectrometry. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  7. Technological assumptions for biogas purification.

    Science.gov (United States)

    Makareviciene, Violeta; Sendzikiene, Egle

    2015-01-01

    Biogas can be used in the engines of transport vehicles and blended into natural gas networks, but it also requires the removal of carbon dioxide, hydrogen sulphide, and moisture. Biogas purification process flow diagrams have been developed for a process enabling the use of a dolomite suspension, as well as for solutions obtained by the filtration of the suspension, to obtain biogas free of hydrogen sulphide and with a carbon dioxide content that does not exceed 2%. The cost of biogas purification was evaluated on the basis of data on biogas production capacity and biogas production cost obtained from local water treatment facilities. It has been found that, with the use of dolomite suspension, the cost of biogas purification is approximately six times lower than that in the case of using a chemical sorbent such as monoethanolamine. The results showed travelling costs using biogas purified by dolomite suspension are nearly 1.5 time lower than travelling costs using gasoline and slightly lower than travelling costs using mineral diesel fuel.

  8. Countercurrent tangential chromatography for large-scale protein purification.

    Science.gov (United States)

    Shinkazh, Oleg; Kanani, Dharmesh; Barth, Morgan; Long, Matthew; Hussain, Daniar; Zydney, Andrew L

    2011-03-01

    Recent advances in cell culture technology have created significant pressure on the downstream purification process, leading to a "downstream bottleneck" in the production of recombinant therapeutic proteins for the treatment of cancer, genetic disorders, and cardiovascular disease. Countercurrent tangential chromatography overcomes many of the limitations of conventional column chromatography by having the resin (in the form of a slurry) flow through a series of static mixers and hollow fiber membrane modules. The buffers used in the binding, washing, and elution steps flow countercurrent to the resin, enabling high-resolution separations while reducing the amount of buffer needed for protein purification. The results obtained in this study provide the first experimental demonstration of the feasibility of using countercurrent tangential chromatography for the separation of a model protein mixture containing bovine serum albumin and myoglobin using a commercially available anion exchange resin. Batch uptake/desorption experiments were used in combination with critical flux data for the hollow fiber filters to design the countercurrent tangential chromatography system. A two-stage batch separation yielded the purified target protein at >99% purity with 94% recovery. The results clearly demonstrate the potential of using countercurrent tangential chromatography for the large-scale purification of therapeutic proteins. Copyright © 2010 Wiley Periodicals, Inc.

  9. Purification of a Multidrug Resistance Transporter for Crystallization Studies

    Directory of Open Access Journals (Sweden)

    Kamela O. Alegre

    2015-03-01

    Full Text Available Crystallization of integral membrane proteins is a challenging field and much effort has been invested in optimizing the overexpression and purification steps needed to obtain milligram amounts of pure, stable, monodisperse protein sample for crystallography studies. Our current work involves the structural and functional characterization of the Escherichia coli multidrug resistance transporter MdtM, a member of the major facilitator superfamily (MFS. Here we present a protocol for isolation of MdtM to increase yields of recombinant protein to the milligram quantities necessary for pursuit of structural studies using X-ray crystallography. Purification of MdtM was enhanced by introduction of an elongated His-tag, followed by identification and subsequent removal of chaperonin contamination. For crystallization trials of MdtM, detergent screening using size exclusion chromatography determined that decylmaltoside (DM was the shortest-chain detergent that maintained the protein in a stable, monodispersed state. Crystallization trials of MdtM performed using the hanging-drop diffusion method with commercially available crystallization screens yielded 3D protein crystals under several different conditions. We contend that the purification protocol described here may be employed for production of high-quality protein of other multidrug efflux members of the MFS, a ubiquitous, physiologically and clinically important class of membrane transporters.

  10. Large-scale functional purification of recombinant HIV-1 capsid.

    Directory of Open Access Journals (Sweden)

    Magdeleine Hung

    Full Text Available During human immunodeficiency virus type-1 (HIV-1 virion maturation, capsid proteins undergo a major rearrangement to form a conical core that protects the viral nucleoprotein complexes. Mutations in the capsid sequence that alter the stability of the capsid core are deleterious to viral infectivity and replication. Recently, capsid assembly has become an attractive target for the development of a new generation of anti-retroviral agents. Drug screening efforts and subsequent structural and mechanistic studies require gram quantities of active, homogeneous and pure protein. Conventional means of laboratory purification of Escherichia coli expressed recombinant capsid protein rely on column chromatography steps that are not amenable to large-scale production. Here we present a function-based purification of wild-type and quadruple mutant capsid proteins, which relies on the inherent propensity of capsid protein to polymerize and depolymerize. This method does not require the packing of sizable chromatography columns and can generate double-digit gram quantities of functionally and biochemically well-behaved proteins with greater than 98% purity. We have used the purified capsid protein to characterize two known assembly inhibitors in our in-house developed polymerization assay and to measure their binding affinities. Our capsid purification procedure provides a robust method for purifying large quantities of a key protein in the HIV-1 life cycle, facilitating identification of the next generation anti-HIV agents.

  11. Biomimetic small peptide functionalized affinity monoliths for monoclonal antibody purification.

    Science.gov (United States)

    Wang, Xiangyu; Xia, Donghai; Han, Hai; Peng, Kun; Zhu, Peijie; Crommen, Jacques; Wang, Qiqin; Jiang, Zhengjin

    2018-08-09

    The rapid development of monoclonal antibodies (mAbs) in therapeutic and diagnostic applications has necessitated the advancement of mAbs purification technologies. In this study, a biomimetic small peptide ligand 3,5-di-tert-butyl-4-hydroxybenzoic acid-Arg-Arg-Gly (DAAG) functionalized monolith was fabricated through a metal ion chelation-based multi-step approach. The resulting monolith showed good chromatographic performance. Compared with the Ni 2+ based IMAC monolith, the DAAG functionalized monolith exhibited not only excellent specificity but also higher dynamic binding capacity (DBC). The 10% DBC and 50% DBC for hIgG reached as high values as 26.0 and 34.6 mg/mL, respectively, at a ligand density of 8.8 μmol/mL, due to the high porosity and accessibility of the monolithic matrix. Moreover, the stability of the DAAG functionalized monolith in successive breakthrough experiments indicates that it has a promising potential for long-term use in mAbs purification. Finally, the DAAG functionalized monolith was successfully applied to the purification of trastuzumab or human immunoglobulin G (hIgG) from biological samples. Copyright © 2018 Elsevier B.V. All rights reserved.

  12. A comprehensive review on biodiesel purification and upgrading

    Directory of Open Access Journals (Sweden)

    Hamed Bateni

    2017-09-01

    Full Text Available Serious environmental concerns regarding the use of fossil-based fuels have raised awareness regarding the necessity of alternative clean fuels and energy carriers. Biodiesel is considered a clean, biodegradable, and non-toxic diesel substitute produced via the transesterification of triglycerides with an alcohol in the presence of a proper catalyst. After initial separation of the by-product (glycerol, the crude biodiesel needs to be purified to meet the standard specifications prior to marketing. The presence of impurities in the biodiesel not only significantly affects its engine performance but also complicates its handling and storage. Therefore, biodiesel purification is an essential step prior to marketing. Biodiesel purification methods can be classified based on the nature of the process into equilibrium-based, affinity-based, membrane-based, reaction-based, and solid-liquid separation processes. The main adverse properties of biodiesel – namely moisture absorption, corrosiveness, and high viscosity – primarily arise from the presence of oxygen. To address these issues, several upgrading techniques have been proposed, among which catalytic (hydrodeoxygenation using conventional hydrotreating catalysts, supported metallic materials, and most recently transition metals in various forms appear promising. Nevertheless, catalyst deactivation (via coking and/or inadequacy of product yields necessitate further research. This paper provides a comprehensive overview on the techniques and methods used for biodiesel purification and upgrading.

  13. [Improved protein-A chromatography for monoclonal antibody purification].

    Science.gov (United States)

    Chen, Quan; Toh, Phyllicia; Hoi, Aina; Xian, Mo; Peng, Xinying; Yang, Yuansheng; Zhang, Haibo; Nian, Rui; Zhang, Wei

    2016-06-25

    Therapeutic monoclonal antibodies become the major product class within the biopharmaceutical market. Protein A as the first capture step is still dominant in current platforms for purification of monoclonal antibodies. In this study, we developed a new antibody harvest process that incorporates acidic treatment of cell harvest, demonstrating high process yield, improved clearance of host cell associated contaminants, like non-histone host cell protein, histone, DNA and heteroaggregates. Host protein contamination was reduced about 10-fold compared to protein A loaded with harvest clarified by centrifugation and microfiltration. Turbidity increase of eluted IgG upon pH neutralization was nearly eliminated. Residual levels of impurities in the protein A eluate were achieved that potentially meet requirements of drug substance and thus alleviate the burden for further impurities removal in subsequent chromatography steps. The mechanism of host cell associated contaminants removal during acidic treatment was also explored. After a polishing step by Capto adhere, host cell protein was reduced to less than 5 ppm, DNA less than 1 ppb, histone to undetectable level, heteroaggregates less than 0.01% with total IgG recovery around 87%. This efficient process can be easily integrated into current IgG purification platforms, and may overcome downstream processing challenges.

  14. High Throughput Identification, Purification and Structural Characterization of Water Soluble Protein Complexes in Desulfovibrio vulgaris

    Energy Technology Data Exchange (ETDEWEB)

    Dong,, Ming; Han, Bong-Gyoon; Liu, Hui-Hai; Malik, J.; Geller, Jil; Yang, Li; Choi, M.; Chandonia, John-Marc; Arbelaez, Pablo; Sterling, H. J.; Typke, Dieter; Shatsky, Max; Brenner, Steve; Fisher, Susan; Williams, Evan; Szakal, Evelin; Allen, S.; Hall, S. C.; Hazen, Terry; Witkowska, H. E.; Jin, Jiming; Glaeser, Robert; Biggin, Mark

    2010-05-17

    Our scheme for the tagless purification of water soluble complexes. 10 g of protein from a crude bacterial extract is first fractionated by ammonium sulfate precipitation and then by a series of chromatographic steps: anion exchange (IEX), hydrophobic interaction (HIC), and finally size exclusion (Gel Filtration). Fractions from the last chromatography step are trypsin digested and peptides labeled with iTRAQ reagents to allow multiplexing and quantitation during mass spectrometric analysis. Elution profiles of identified proteins are then subjected to clustering analysis.

  15. Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

    Science.gov (United States)

    Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

    2016-10-01

    Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a

  16. Fast-Rate Capable Electrode Material with Higher Energy Density than LiFePO4: 4.2V LiVPO4F Synthesized by Scalable Single-Step Solid-State Reaction.

    Science.gov (United States)

    Kim, Minkyung; Lee, Seongsu; Kang, Byoungwoo

    2016-03-01

    Use of compounds that contain fluorine (F) as electrode materials in lithium ion batteries has been considered, but synthesizing single-phase samples of these compounds is a difficult task. Here, it is demonstrated that a simple scalable single-step solid-state process with additional fluorine source can obtain highly pure LiVPO 4 F. The resulting material with submicron particles achieves very high rate capability ≈100 mAh g -1 at 60 C-rate (1-min discharge) and even at 200 C-rate (18 s discharge). It retains superior capacity, ≈120 mAh g -1 at 10 C charge/10 C discharge rate (6-min) for 500 cycles with >95% retention efficiency. Furthermore, LiVPO 4 F shows low polarization even at high rates leading to higher operating potential >3.45 V (≈3.6 V at 60 C-rate), so it achieves high energy density. It is demonstrated for the first time that highly pure LiVPO 4 F can achieve high power capability comparable to LiFePO 4 and much higher energy density (≈521 Wh g -1 at 20 C-rate) than LiFePO 4 even without nanostructured particles. LiVPO 4 F can be a real substitute of LiFePO 4.

  17. Comparing Russian and Finnish standards of water purification

    OpenAIRE

    Maria, Pupkova

    2012-01-01

    The subject of this thesis is water purification. The first aim of this thesis is to consider different ways of water purification. The second aim is to compare Finnish and Russian standards of water purification. The third one is to show water purification methods on the pattern of Mikkeli water purification plan. Water purification methods of water intended for human consumption will be described.Combined tables will be done according to the quality requirement of drinking water of both,...

  18. Purification of highly active alphavirus replication complexes demonstrates altered fractionation of multiple cellular membranes.

    Science.gov (United States)

    Pietilä, Maija K; van Hemert, Martijn J; Ahola, Tero

    2018-01-24

    Positive-strand RNA viruses replicate their genomes in membrane-associated structures; alphaviruses and many other groups induce membrane invaginations called spherules. Here, we established a protocol to purify these membranous replication complexes (RCs) from cells infected with Semliki Forest virus (SFV). We isolated SFV spherules located on the plasma membrane and further purified them using two consecutive density gradients. This revealed that SFV infection strongly modifies cellular membranes. We removed soluble proteins, the Golgi and most of the mitochondria, but plasma membrane, endoplasmic reticulum (ER) and late endosome markers enriched in the membrane fraction that contained viral RNA synthesizing activity, replicase proteins and minus- and plus-strand RNA. Electron microscopy revealed that the purified membranes displayed spherule-like structures with a narrow neck. This membrane enrichment was specific to viral replication as such a distribution of membrane markers was only observed after infection. Besides the plasma membrane, SFV infection remodeled the ER, and the co-fractionation of the RC-carrying plasma membrane and ER suggests that SFV may recruit ER proteins or membrane to the site of replication. The purified RCs were highly active in synthesizing both genomic and subgenomic RNA. Detergent solubilization destroyed the replication activity demonstrating that the membrane association of the complex is essential. Most of the newly made RNA was in double-stranded replicative molecules but the purified complexes also produced single-stranded RNA as well as released newly made RNA. This indicates that the purification established here maintained the functionality of RCs and thus enables further structural and functional studies of active RCs. IMPORTANCE Similar to all positive-strand RNA viruses, the arthropod-borne alphaviruses induce membranous genome factories but little is known about the arrangement of viral replicase proteins and the

  19. Effect of matrices on affinity purification of protein C.

    Science.gov (United States)

    Kang, K; Ryu, D; Drohan, W N; Orthner, C L

    1992-05-01

    One of the critical problems in scale-up of affinity chromatography is the mechanical strength of the support matrix against pressure. Because the costs of both the gel matrix and the ligand for the affinity chromatography are very high, the reusability of gel matrices is directly related to the total production cost. In certain cases, where the source material is viscous (e.g., blood plasma), irreversible deformation of gel matrices can readily occur, necessitating severe constraints in the flow rate. Consequently, productivity is low.We have characterized the system parameters and investigated the performance of various matrices that are commercially available. The experimental system used for this study was the immunoaffinity purification of protein C (an anticoagulant protein) from human blood plasma. The support matrices studied were cross-linked agarose, polymethyl acrylic, cellulose, and polyvinyl alcohol polymers. The major system parameters studied were pressure tolerance, coupling efficiency, adsorption efficiency, and batch adsorption/desorption kinetics of protein C to/from the monoclonal antibody (MAb)-Matrix complex. In addition, the apparent equilibrium constant and bandwidth of the product concentration profile in the eluate were characterized by performing pulse tests.A methodology was developed for evaluating the immunoaffinity column performance for the separation of protein C. By utilizing the experimentally measured parameters, the flow rate limitation for each purification step was computed. Then, the purification performance of the matrices were evaluated in terms of productivity per unit time. Among the matrices tested, cellulose was superior in overall performance for the immunoaffinity purification of protein C using a 10 cm x 10 cm column.

  20. Purification of uranium metal

    International Nuclear Information System (INIS)

    Suzuki, Kenji; Shikama, Tatsuo; Ochiai, Akira.

    1993-01-01

    We developed the system for purifying uranium metal and its metallic compounds and for growing highly pure uranium compounds to study their intrinsic physical properties. Uranium metal was zone refined under low contamination conditions as far as possible. The degree of the purity of uranium metal was examined by the conventional electrical resistivity measurement and by the chemical analysis using the inductive coupled plasma emission spectrometry (ICP). The results show that some metallic impurities evaporated by the r.f. heating and other usual metallic impurities moved to the end of a rod with a molten zone. Therefore, we conclude that the zone refining technique is much effective to the removal of metallic impurities and we obtained high purified uranium metal of 99.99% up with regarding to metallic impurities. The maximum residual resistivity ratio, the r.r.r., so far obtained was about 17-20. Using the purified uranium, we are attempting to grow a highly pure uranium-titanium single crystals. (author)