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Sample records for surface biotinylation exofacial

  1. Metabolic behavior of cell surface biotinylated proteins

    International Nuclear Information System (INIS)

    Hare, J.F.; Lee, E.

    1989-01-01

    The turnover of proteins on the surface of cultured mammalian cells was measured by a new approach. Reactive free amino or sulfhydryl groups on surface-accessible proteins were derivatized with biotinyl reagents and the proteins solubilized from culture dishes with detergent. Solubilized, biotinylated proteins were then adsorbed onto streptavidin-agarose, released with sodium dodecyl sulfate and mercaptoethanol, and separated on polyacrylamide gels. Biotin-epsilon-aminocaproic acid N-hydroxysuccinimide ester (BNHS) or N-biotinoyl-N'-(maleimidohexanoyl)hydrazine (BM) were the derivatizing agents. Only 10-12 bands were adsorbed onto streptavidin-agarose from undervatized cells or from derivatized cells treated with free avidin at 4 degrees C. Two-dimensional isoelectric focusing-sodium dodecyl sulfate gel electrophoresis resolved greater than 100 BNHS-derivatized proteins and greater than 40 BM-derivatized proteins. There appeared to be little overlap between the two groups of derivatized proteins. Short-term pulse-chase studies showed an accumulation of label into both groups of biotinylated proteins up until 1-2 h of chase and a rapid decrease over the next 1-5 h. Delayed appearance of labeled protein at the cell surface was attributed to transit time from site of synthesis. The unexpected and unexplained rapid disappearance of pulse-labeled proteins from the cell surface was invariant for all two-dimensionally resolved proteins and was sensitive to temperature reduction to 18 degrees C. Long-term pulse-chase experiments beginning 4-8 h after the initiation of chase showed the disappearance of derivatized proteins to be a simple first-order process having a half-life of 115 h in the case of BNHS-derivatized proteins and 30 h in the case of BM-derivatized proteins

  2. Binding properties of a streptavidin layer formed on a biotinylated Langmuir–Schaefer film of unfolded protein

    Energy Technology Data Exchange (ETDEWEB)

    Furuno, Taiji, E-mail: t_furuno@a8.keio.jp

    2016-04-01

    A Langmuir monolayer of carbonic anhydrase (CA) unfolded at an air/water interface was transferred onto the hydrophobic surface of a silicon wafer by means of the Langmuir–Schaefer technique. The transferred CA film was biotinylated and was incubated in a streptavidin (SAv) solution to obtain a densely packed SAv layer by biotin–SAv linkage. Biotinylated proteins including ferritin, catalase, alcohol dehydrogenase, and carbonic anhydrase were incubated with the SAv layer and binding of these proteins was examined by atomic force microscopy. High-density binding of the biotinylated proteins was observed, whereas the amount of adsorbed non-biotinylated proteins was low or negligible. The SAv layer on the Langmuir–Schaefer film of unfolded protein could become a basic architecture for protein immobilization studies. - Highlights: • Langmuir–Schaefer film of carbonic anhydrase (LSF-CA) was biotinylated. • A densely packed streptavidin (SAv) layer was formed on the biotinylated LSF-CA. • Biotinylated proteins were bound to the SAv layer at high density. • Nonspecific adsorption of intact proteins to the SAv layer was weak. • Atomic force microscopy showed the binding of proteins at molecular resolution.

  3. Fc-specific biotinylation of antibody using an engineered photoactivatable Z–Biotin and its biosensing application

    International Nuclear Information System (INIS)

    Yang, Hong-Ming; Bao, Ru-Meng; Yu, Chang-Mei; Lv, Yan-Na; Zhang, Wei-Fen; Tang, Jin-Bao

    2017-01-01

    The development of a site-specific and covalent attachment methodology is crucial for antibody–biotin conjugates to preserve the antigen-binding ability of antibodies and yield homogeneous products. In this study, an engineered photoactivatable Z-domain variant [an UV-active amino acid benzoylphenylalanine (Bpa) was genetically incorporated into the Z-domain] carrying one biotin molecule (Z_B_p_a–Biotin) was prepared by employing aminoacyl-tRNA synthetase/suppressor tRNA and Avitag/BirA techniques. The site-specific and covalent attachment of IgG–biotin conjugates, viz. photo-biotinylated IgG, was successfully achieved after UV exposure by combining the inherent Fc-binding capability of the Z-domain with the formation of covalent bond by the photo-crosslinker. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis assay showed that more than 90% of IgGs conjugated with Z_B_p_a–Biotin molecules suffered 3 h UV irradiation. Further pepsin digestion analysis confirmed that the Z_B_p_a–Biotin was conjugated to the Fc fragment of IgG without interference. We took the tumor biomarker carcinoembryoic antigen (CEA) as model to evaluate the detection efficiency of the site-specific photo-biotinylated IgG in biosensing application using surface plasmon resonance (SPR) technology. The photo-biotinylated IgG coated surface gave a limit of detection (LOD) of 2 ng mL"-"1, is 5-fold lower than that of the randomly NHS-biotinylated IgG (10 ng mL"-"1). Given that the (strept)avidin–biotin complex is extensively used in immunoassays, the proposed method for biotinylated IgG provides a powerful approach to further expand related applications. - Highlights: • A photoactivable Z_B_p_a–Biotin was fabricated by aaRS/tRNA and Avitag/BirA techniques. • A approach for Fc-specific photo-biotinylated IgG via Z_B_p_a–Biotin was proposed. • The photo-biotinylated IgG was used to fabricate an immunosensor for detecting CEA. • It gave a LOD of 2 ng mL"-"1 CEA

  4. Fc-specific biotinylation of antibody using an engineered photoactivatable Z–Biotin and its biosensing application

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Hong-Ming; Bao, Ru-Meng; Yu, Chang-Mei; Lv, Yan-Na; Zhang, Wei-Fen; Tang, Jin-Bao, E-mail: tangjb@wfmc.edu.cn

    2017-01-01

    The development of a site-specific and covalent attachment methodology is crucial for antibody–biotin conjugates to preserve the antigen-binding ability of antibodies and yield homogeneous products. In this study, an engineered photoactivatable Z-domain variant [an UV-active amino acid benzoylphenylalanine (Bpa) was genetically incorporated into the Z-domain] carrying one biotin molecule (Z{sub Bpa}–Biotin) was prepared by employing aminoacyl-tRNA synthetase/suppressor tRNA and Avitag/BirA techniques. The site-specific and covalent attachment of IgG–biotin conjugates, viz. photo-biotinylated IgG, was successfully achieved after UV exposure by combining the inherent Fc-binding capability of the Z-domain with the formation of covalent bond by the photo-crosslinker. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis assay showed that more than 90% of IgGs conjugated with Z{sub Bpa}–Biotin molecules suffered 3 h UV irradiation. Further pepsin digestion analysis confirmed that the Z{sub Bpa}–Biotin was conjugated to the Fc fragment of IgG without interference. We took the tumor biomarker carcinoembryoic antigen (CEA) as model to evaluate the detection efficiency of the site-specific photo-biotinylated IgG in biosensing application using surface plasmon resonance (SPR) technology. The photo-biotinylated IgG coated surface gave a limit of detection (LOD) of 2 ng mL{sup -1}, is 5-fold lower than that of the randomly NHS-biotinylated IgG (10 ng mL{sup -1}). Given that the (strept)avidin–biotin complex is extensively used in immunoassays, the proposed method for biotinylated IgG provides a powerful approach to further expand related applications. - Highlights: • A photoactivable Z{sub Bpa}–Biotin was fabricated by aaRS/tRNA and Avitag/BirA techniques. • A approach for Fc-specific photo-biotinylated IgG via Z{sub Bpa}–Biotin was proposed. • The photo-biotinylated IgG was used to fabricate an immunosensor for detecting CEA. • It gave a LOD

  5. Fc-specific biotinylation of antibody using an engineered photoactivatable Z-Biotin and its biosensing application.

    Science.gov (United States)

    Yang, Hong-Ming; Bao, Ru-Meng; Yu, Chang-Mei; Lv, Yan-Na; Zhang, Wei-Fen; Tang, Jin-Bao

    2017-01-01

    The development of a site-specific and covalent attachment methodology is crucial for antibody-biotin conjugates to preserve the antigen-binding ability of antibodies and yield homogeneous products. In this study, an engineered photoactivatable Z-domain variant [an UV-active amino acid benzoylphenylalanine (Bpa) was genetically incorporated into the Z-domain] carrying one biotin molecule (Z Bpa -Biotin) was prepared by employing aminoacyl-tRNA synthetase/suppressor tRNA and Avitag/BirA techniques. The site-specific and covalent attachment of IgG-biotin conjugates, viz. photo-biotinylated IgG, was successfully achieved after UV exposure by combining the inherent Fc-binding capability of the Z-domain with the formation of covalent bond by the photo-crosslinker. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis assay showed that more than 90% of IgGs conjugated with Z Bpa -Biotin molecules suffered 3 h UV irradiation. Further pepsin digestion analysis confirmed that the Z Bpa -Biotin was conjugated to the Fc fragment of IgG without interference. We took the tumor biomarker carcinoembryoic antigen (CEA) as model to evaluate the detection efficiency of the site-specific photo-biotinylated IgG in biosensing application using surface plasmon resonance (SPR) technology. The photo-biotinylated IgG coated surface gave a limit of detection (LOD) of 2 ng mL -1 , is 5-fold lower than that of the randomly NHS-biotinylated IgG (10 ng mL -1 ). Given that the (strept)avidin-biotin complex is extensively used in immunoassays, the proposed method for biotinylated IgG provides a powerful approach to further expand related applications. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Ultrastructural and biochemical detection of biotin and biotinylated polypeptides in Schistosoma mansoni

    Directory of Open Access Journals (Sweden)

    Santos P.R.P.

    1997-01-01

    Full Text Available Biotinylation is proposed for the identification of surface proteins in Schistosoma mansoni using the streptavidin-HRP conjugate for the detection of labeled polypeptides. However, control samples also showed several endogenous biotinylated polypeptides. In an attempt to determine the possibility of nonspecific binding between the streptavidin-HRP conjugate and polypeptides from S. mansoni, the conjugate was blocked with biotinamidecaproate-N-hydroxysuccinimide ester (BcapNHS before biotin-streptavidin blotting. No bands were detected on the nitrocellulose sheet, demonstrating the specific recognition of biotin by the streptavidin present in the conjugate. Whole cercariae and cercarial bodies and tails showed several endogenous biotinylated polypeptides. The biotin concentration was 13 µg/190,000 cercariae. Adult worms presented less endogenous biotinylated polypeptides than cercariae. These results may be due to changes in the environment from aerobic to anaerobic conditions when cercarial bodies (schistosomula are transformed into adult worms and a decrease in CO2 production may occur. Cercariae, cercarial bodies and adult male worms were examined by transmission electron microscopy employing an avidin-colloidal gold conjugate for the detection of endogenous biotin. Gold particles were distributed mainly on the muscle fibers, but dispersed granules were observed in the tegument, mitochondria and cytosol. The discovery of endogenous biotin in S. mansoni should be investigated in order to clarify the function of this vitamin in the parasite

  7. Biotinylated human. beta. -endorphins as probes for the opioid receptor

    Energy Technology Data Exchange (ETDEWEB)

    Hochhaus, G.; Gibson, B.W.; Sadee, W.

    1988-01-05

    The reaction of human ..beta..-endorphin and biotinyl N-hydroxysuccinimide with or without spacer arm, afforded a series of products that were separated by high performance liquid chromatography (HPLC). Liquid secondary ion mass spectrometry of the biotinylated products and their tryptic digests produced abundant protonated molecular ions (MH/sup +/), which specified the number and location of biotinylation. Between 1 and 4 biotinyl residues were incorporated per human ..beta..-endorphin molecule, at Lys-9, -19, -24, -28, and -29, but not at the amino-terminal Try-1. Three HPLC fractions were isolated for receptor binding studies monobiotinylation of Lys-9, Lys-19, and a mixture of Lys-24, Lys-28, and Lys-29 derivatives. IC/sub 50/ values for binding to ..mu.. and delta opioid receptor sites were 3-8 times higher for monobiotinylated derivatives than for the parent human ..beta..-endorphin. Association with avidin decreased opioid receptor affinities for the C/sub 6/ spacer derivative biotinylated at position Lys-9, which is close to the (1-5) enkephalin receptor region. In contrast, avidin did not affect or even increased apparent affinities to ..mu.. and delta sites for derivatives biotinylated at the ..cap alpha..-helical part of the molecule (Lys-19, -24, -28, and -29). Biotinylated human ..beta..-endorphins also bound to low affinity nonopioid binding sites on NG-108-15 cells; however, affinities to these sites were considerably reduced when derivatives were bound to avidin. The ability of biotinylated human ..beta..-endorphin to cross-link the ..mu.. and delta opioid receptors to avidin allows application of the biotin-avidin system as a molecular probe of the opioid receptor.

  8. Simple methods for the 3' biotinylation of RNA.

    Science.gov (United States)

    Moritz, Bodo; Wahle, Elmar

    2014-03-01

    Biotinylation of RNA allows its tight coupling to streptavidin and is thus useful for many types of experiments, e.g., pull-downs. Here we describe three simple techniques for biotinylating the 3' ends of RNA molecules generated by chemical or enzymatic synthesis. First, extension with either the Schizosaccharomyces pombe noncanonical poly(A) polymerase Cid1 or Escherichia coli poly(A) polymerase and N6-biotin-ATP is simple, efficient, and generally applicable independently of the 3'-end sequences of the RNA molecule to be labeled. However, depending on the enzyme and the reaction conditions, several or many biotinylated nucleotides are incorporated. Second, conditions are reported under which splint-dependent ligation by T4 DNA ligase can be used to join biotinylated and, presumably, other chemically modified DNA oligonucleotides to RNA 3' ends even if these are heterogeneous as is typical for products of enzymatic synthesis. Third, we describe the use of 29 DNA polymerase for a template-directed fill-in reaction that uses biotin-dUTP and, thanks to the enzyme's proofreading activity, can cope with more extended 3' heterogeneities.

  9. Biotinylation is a natural, albeit rare, modification of human histones

    Science.gov (United States)

    Kuroishi, Toshinobu; Rios-Avila, Luisa; Pestinger, Valerie; Wijeratne, Subhashinee S. K.; Zempleni, Janos

    2011-01-01

    Previous studies suggest that histones H3 and H4 are posttranslationally modified by binding of the vitamin biotin, catalyzed by holocarboxylase synthetase (HCS). Albeit a rare epigenetic mark, biotinylated histones were repeatedly shown to be enriched in repeat regions and repressed loci, participating in the maintenance of genome stability and gene regulation. Recently, a team of investigators failed to detect biotinylated histones and proposed that biotinylation is not a natural modification of histones, but rather an assay artifact. Here, we describe the results of experiments, including the comparison of various analytical protocols, antibodies, cell lines, classes of histones, and radiotracers. These studies provide unambiguous evidence that biotinylation is a natural, albeit rare, histone modification. Less than 0.001% of human histones H3 and H4 are biotinylated, raising concerns that the abundance might too low to elicit biological effects in vivo. We integrated information from this study, previous studies, and ongoing research efforts to present a new working model in which biological effects are caused by a role of HCS in multiprotein complexes in chromatin. In this model, docking of HCS in chromatin causes the occasional binding of biotin to histones as a tracer for HCS binding sites. PMID:21930408

  10. Synthesis of biotinyl derivatives of peptide hormones and other biological materials

    International Nuclear Information System (INIS)

    Finn, F.M.; Hofmann, K.H.

    1985-01-01

    Methods for the preparation of biotinylated ligands for the avidin-biotin system are described. Also described are procedures for modifying and labelling avidin and for assessing the rate of dissociation of biotin derivatives from avidin. The most widely used procedure for introducing biotin into other molecules involves acylation with N-hydroxysuccinimido-biotinate. Experimental details are given for the synthesis of dethiobiotin, iminobiotin, and biotinylated ligands in which a 6-aminohexanoic acid spacer is interposed between the biotin or biotin derivative and the insulin molecule. The syntheses of biotinylated corticotropins are presented only in principle

  11. Biotinylated liposomes as potential carriers for the oral delivery of insulin.

    Science.gov (United States)

    Zhang, Xingwang; Qi, Jianping; Lu, Yi; He, Wei; Li, Xiaoyang; Wu, Wei

    2014-01-01

    This study aimed to explore biotinylated liposomes (BLPs) as novel carriers to enhance the oral delivery of insulin. Biotinylation was achieved by incorporating biotin-conjugated phospholipids into the liposome membranes. A significant hypoglycemic effect and enhanced absorption were observed after treating diabetic rats with the BLPs with a relative bioavailability of 12.09% and 8.23%, based on the measurement of the pharmacologic effect and the blood insulin level, respectively; this achieved bioavailability was approximately double that of conventional liposomes. The significance of the biotinylation was confirmed by the facilitated absorption of the BLPs through receptor-mediated endocytosis, as well as by the improved physical stability of the liposomes. Increased cellular uptake and quick gastrointestinal transport further verified the ability of the BLPs to enhance absorption. These results provide a proof of concept that BLPs can be used as potential carriers for the oral delivery of insulin. Diabetes remains a major source of mortality in the Western world, and advances in its management are expected to have substantial socioeconomic impact. In this paper, biotinylated liposomes were utilized as carriers of insulin for local delivery, demonstrating the feasibility of this approach in a rat model. © 2014.

  12. Intravenous avidin chase improved localization of radiolabeled streptavidin in intraperitoneal xenograft pretargeted with biotinylated antibody

    International Nuclear Information System (INIS)

    Zhang Meili; Sakahara, Harumi; Yao Zhengsheng; Saga, Tsuneo; Nakamoto, Yuhi; Sato, Noriko; Nakada, Hiroshi; Yamashina, Ikuo; Konishi, Junji

    1997-01-01

    In the present study, we examined the effect of avidin administered intravenously (i.v.) on the biodistribution of radiolabeled streptavidin in mice bearing intraperitoneal (IP) xenografts pretargeted with biotinylated antibody. Tumors were established in nude mice by IP inoculation of LS180 human colon cancer cells. Monoclonal antibody MLS128, which recognizes Tn antigen on mucin, was biotinylated and injected IP into the IP tumor-bearing mice. Radioiodinated streptavidin was administered IP or i.v. 48 h after pretargeting of biotinylated antibody. Avidin was administered i.v. 30 min prior to streptavidin injection. The localization of radioiodinated streptavidin in the tumor pretargeted with biotinylated antibody was significantly higher than that without pretargeting and that of radioiodinated MLS128 by the one-step method. Avidin administration significantly accelerated the clearance of radioiodinated streptavidin in blood and other normal tissues and increased the tumor-to-blood radioactivity ratio regardless of administration route of streptavidin. The i.v. avidin chase improved tumor localization of radiolabeled streptavidin in the IP xenografts pretargeted with biotinylated antibody

  13. Paracellular transport of avidin saturated or not with biotinylated cobalamin through Caco-2 cell epithelium monolayer.

    Science.gov (United States)

    Guy, M; Pons, L; Namour, F; de Nonancourt, M; Michalski, J C; Hatier, R; Guéant, J L

    2001-01-01

    The cationic charge of molecules may promote their uptake across epithelia, which are rich in brush border anionic sites. The transport of unsaturated avidin and avidin saturated with a biotinylated compound was investigated across Caco-2 adenocarcinoma cell with fetal enterocyte phenotype. The unsaturated avidin and avidin saturated with either biotin or a biotinyl-cobalamin conjugate (biotinyl-Cbl) were iodinated to follow their transport through the cell monolayer. Their apparent permeability coefficient (Papp) and transepithelial pathway were determined and compared to those for control radiolabeled markers [3H]-mannitol, [125I]-beta-lactoglobulin and [57Co]-cobalamin/intrinsic factor (Cbl/IF). The Papp of [125I]-avidin estimated at 2.8 x 10(-7) +/- 0.08 cm/s was close to that for mannitol that uses paracellular pathway. The binding of biotin or biotin conjugate to avidin enhanced its tetrameric conformation. The Papp for [125I]-avidin/biotin and [125I]- avidin/biotinyl-Cbl were respectively increased by 2-fold, compared to that for [125I]-avidin and 4-fold, compared to that for [125I]-beta-lactoglobulin and [54Co]-Cbl/IF. The protein was not accumulated in the cell and was found in intact form in the basolateral side, after its transport across the monolayer. Chloroquine (0.66 micromol/ml) did not significantly decrease the Papp for [125I]-avidin/biotinyl-Cbl. Conversely it decreased by 80% the Papp for Cbl/IF, that uses transepithelial pathway. Avidin (either saturated or not with biotin and biotinyl-Cbl) was able to cross the monolayer of Caco-2 cell line through a paracellular pathway. This study pointed out the interest for using this protein as a shuttle for increasing the transport rate of biotinylated compounds through fetal epithelial barriers. Copyright 2001 S. Karger AG, Basel

  14. Magnetic molecularly imprinted polymer for the isolation and detection of biotin and biotinylated biomolecules.

    Science.gov (United States)

    Ben Aissa, A; Herrera-Chacon, A; Pupin, R R; Sotomayor, M D P T; Pividori, M I

    2017-02-15

    Magnetic separation based on biologically-modified magnetic particles is a preconcentration procedure commonly integrated in magneto actuated platforms for the detection of a huge range of targets. However, the main drawback of this material is the low stability and high cost. In this work, a novel hybrid molecularly-imprinted polymer with magnetic properties is presented with affinity towards biotin and biotinylated biomolecules. During the synthesis of the magneto core-shell particles, biotin was used as a template. The characterization of this material by microscopy techniques including SEM, TEM and confocal microscopy is presented. The application of the magnetic-MIPs for the detection of biotin and biotinylated DNA in magneto-actuated platforms is also described for the first time. The magnetic-MIP showed a significant immobilization capacity of biotinylated molecules, giving rise to a cheaper and a robust method (it is not required to be stored at 4°C) with high binding capacity for the separation and purification under magnetic actuation of a wide range of biotinylated molecules, and their downstream application including determination of their specific targets. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Transfer in SDS of biotinylated proteins from acrylamide gels to an avidin-coated membrane filter.

    Science.gov (United States)

    Karlin, Arthur; Wang, Chaojian; Li, Jing; Xu, Qiang

    2004-06-01

    Avidin was covalently linked to aldehyde-derivatized polyethersulfone membrane filters. These filters were used in Western blot analysis of proteins reacted with biotinylation reagents and electrophoresed in sodium dodecyl sulfate (SDS) on polyacrylamide gels. Electrophoretic transfer from the gels to these filters was in 0.1% SDS, in which the covalently bound avidin retained its biotin-binding capacity. We compared Western blots on avidin-coated membrane filters of biotinylated and nonbiotinylated forms of mouse immunoglobulin G (IgG), mouse IgG heavy chain, muscle-type acetylcholine receptor alpha subunit, and fused alpha and beta subunits of receptor. Biotinylated proteins were captured with high specificity compared to their nonbiotinylated counterparts and sensitively detected on the avidin-coated membranes.

  16. In vivo biotinylation of recombinant beta-glucosidase enables simultaneous purification and immobilization on streptavidin coated magnetic particles

    DEFF Research Database (Denmark)

    Alftrén, Johan; Ottow, Kim Ekelund; Hobley, Timothy John

    2013-01-01

    Beta-glucosidase from Bacillus licheniformis was in vivo biotinylated in Escherichia coli and subsequently immobilized directly from cell lysate on streptavidin coated magnetic particles. In vivo biotinylation was mediated by fusing the Biotin Acceptor Peptide to the C-terminal of beta......-glucosidase and co-expressing the BirA biotin ligase. The approach enabled simultaneous purification and immobilization of the enzyme from crude cell lysate on magnetic particles because of the high affinity and strong interaction between biotin and streptavidin. After immobilization of the biotinylated beta...

  17. Enhanced sensitivity in detection of antiviral antibody responses using biotinylation of foot-and-mouth disease virus (FMDV) capsids.

    Science.gov (United States)

    Kenney, Mary; Waters, Ryan A; Rieder, Elizabeth; Pega, Juan; Perez-Filguera, Mariano; Golde, William T

    2017-11-01

    Analysis of the immune response to infection of livestock by foot-and-mouth disease virus (FMDV) is most often reported as the serum antibody response to the virus. While measurement of neutralizing antibody has been sensitive and specific, measurements of the quality of the antibody response are less robust. Determining the immunoglobulin (Ig) isotype of the serum antibody response provides a deeper understanding of the biology of the response and more sensitive methods for these assays will facilitate analyses of B cell mediated immunity. We tested the hypothesis that using the virus as the molecular probe could be achieved by adding tags to the surface of the FMDV capsid, and that would enhance sensitivity in assays for anti-FMDV antibody responses. The use of a FLAG-tagged virus in these assays failed to yield improvement whereas chemically biotinylating the virus capsid resulted in significant enhancement of the signal. Here we describe methods using biotinylated virus for measuring anti-viral antibody in serum and antibody secreting cells (ASCs) in blood that are sensitive and specific. Finally, we describe using the biotinylated virus in flow cytometry where such assays should greatly enhance the analysis of anti-virus antibody producing B cells, allowing the investigator to focus on only the FMDV specific B cells when analyzing the development of the B cell response to either infection or vaccination. Published by Elsevier B.V.

  18. A simple elution strategy for biotinylated proteins bound to streptavidin conjugated beads using excess biotin and heat.

    Science.gov (United States)

    Cheah, Joleen S; Yamada, Soichiro

    2017-12-02

    Protein-protein interactions are the molecular basis of cell signaling. Recently, proximity based biotin identification (BioID) has emerged as an alternative approach to traditional co-immunoprecipitation. In this protocol, a mutant biotin ligase promiscuously labels proximal binding partners with biotin, and resulting biotinylated proteins are purified using streptavidin conjugated beads. This approach does not require preservation of protein complexes in vitro, making it an ideal approach to identify transient or weak protein complexes. However, due to the high affinity bond between streptavidin and biotin, elution of biotinylated proteins from streptavidin conjugated beads requires harsh denaturing conditions, which are often incompatible with downstream processing. To effectively release biotinylated proteins bound to streptavidin conjugated beads, we designed a series of experiments to determine optimal binding and elution conditions. Interestingly, the concentrations of SDS and IGEPAL-CA630 during the incubation with streptavidin conjugated beads were the key to effective elution of biotinylated proteins using excess biotin and heating. This protocol provides an alternative method to isolate biotinylated proteins from streptavidin conjugated beads that is suitable for further downstream analysis. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Development of an efficient signal amplification strategy for label-free enzyme immunoassay using two site-specific biotinylated recombinant proteins

    International Nuclear Information System (INIS)

    Tang, Jin-Bao; Tang, Ying; Yang, Hong-Ming

    2015-01-01

    Highlights: • An efficient signal amplification strategy for label-free EIA is proposed. • Divalent biotinylated AP and monovalent biotinylated ZZ were prepared via Avitag–BirA system. • The above site-specific biotinylated fusion proteins form complex via SA–biotin interaction. • The mechanism relies on the ZZ–Avi-B/SA/AP–(Avi-B) 2 complex. • The analytical signals are enhanced (32-fold) by the proposed strategy. - Abstract: Constructing a recombinant protein between a reporter enzyme and a detector protein to produce a homogeneous immunological reagent is advantageous over random chemical conjugation. However, the approach hardly recombines multiple enzymes in a difunctional fusion protein, which results in insufficient amplification of the enzymatic signal, thereby limiting its application in further enhancement of analytical signal. In this study, two site-specific biotinylated recombinant proteins, namely, divalent biotinylated alkaline phosphatase (AP) and monovalent biotinylated ZZ domain, were produced by employing the Avitag–BirA system. Through the high streptavidin (SA)–biotin interaction, the divalent biotinylated APs were clustered in the SA–biotin complex and then incorporated with the biotinylated ZZ. This incorporation results in the formation of a functional macromolecule that involves numerous APs, thereby enhancing the enzymatic signal, and in the production of several ZZ molecules for the interaction with immunoglobulin G (IgG) antibody. The advantage of this signal amplification strategy is demonstrated through ELISA, in which the analytical signal was substantially enhanced, with a 32-fold increase in the detection sensitivity compared with the ZZ–AP fusion protein approach. The proposed immunoassay without chemical modification can be an alternative strategy to enhance the analytical signals in various applications involving immunosensors and diagnostic chips, given that the label-free IgG antibody is suitable for

  20. Development of an efficient signal amplification strategy for label-free enzyme immunoassay using two site-specific biotinylated recombinant proteins

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Jin-Bao [School of Pharmacy, Weifang Medical University, Weifang 261053 (China); Tang, Ying [Affiliated Hospital of Weifang Medical University, Weifang 261041 (China); Yang, Hong-Ming, E-mail: yanghongming2006@sohu.com [School of Pharmacy, Weifang Medical University, Weifang 261053 (China)

    2015-02-15

    Highlights: • An efficient signal amplification strategy for label-free EIA is proposed. • Divalent biotinylated AP and monovalent biotinylated ZZ were prepared via Avitag–BirA system. • The above site-specific biotinylated fusion proteins form complex via SA–biotin interaction. • The mechanism relies on the ZZ–Avi-B/SA/AP–(Avi-B){sub 2} complex. • The analytical signals are enhanced (32-fold) by the proposed strategy. - Abstract: Constructing a recombinant protein between a reporter enzyme and a detector protein to produce a homogeneous immunological reagent is advantageous over random chemical conjugation. However, the approach hardly recombines multiple enzymes in a difunctional fusion protein, which results in insufficient amplification of the enzymatic signal, thereby limiting its application in further enhancement of analytical signal. In this study, two site-specific biotinylated recombinant proteins, namely, divalent biotinylated alkaline phosphatase (AP) and monovalent biotinylated ZZ domain, were produced by employing the Avitag–BirA system. Through the high streptavidin (SA)–biotin interaction, the divalent biotinylated APs were clustered in the SA–biotin complex and then incorporated with the biotinylated ZZ. This incorporation results in the formation of a functional macromolecule that involves numerous APs, thereby enhancing the enzymatic signal, and in the production of several ZZ molecules for the interaction with immunoglobulin G (IgG) antibody. The advantage of this signal amplification strategy is demonstrated through ELISA, in which the analytical signal was substantially enhanced, with a 32-fold increase in the detection sensitivity compared with the ZZ–AP fusion protein approach. The proposed immunoassay without chemical modification can be an alternative strategy to enhance the analytical signals in various applications involving immunosensors and diagnostic chips, given that the label-free IgG antibody is suitable

  1. Low density lipoprotein sensor based on surface plasmon resonance

    International Nuclear Information System (INIS)

    Matharu, Zimple; Sumana, G.; Pandey, M.K.; Gupta, Vinay; Malhotra, B.D.

    2009-01-01

    Biotinylated heparin has been immobilized onto self-assembled monolayer of 4-aminothiophenol using avidin-biotin specific binding. The modified electrodes have been characterized using surface plasmon resonance technique (SPR), cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM) and contact angle (CA) measurements. The interaction of immobilized biotinylated heparin with low density lipoprotein (LDL) has been studied using surface plasmon resonance technique. The biotinylated heparin modified electrode can be used to detect LDL in the range of 20 to 100 mg/dl with the sensitivity of 513.3 m o /μM.

  2. Low density lipoprotein sensor based on surface plasmon resonance

    Energy Technology Data Exchange (ETDEWEB)

    Matharu, Zimple [Department of Science and Technology Centre on Biomolecular Electronics, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India); Department of Physics and Astrophysics, University of Delhi, New Delhi-110007 (India); Sumana, G.; Pandey, M.K. [Department of Science and Technology Centre on Biomolecular Electronics, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India); Gupta, Vinay [Department of Physics and Astrophysics, University of Delhi, New Delhi-110007 (India); Malhotra, B.D., E-mail: bansi.malhotra@gmail.co [Department of Science and Technology Centre on Biomolecular Electronics, National Physical Laboratory, Dr. K. S. Krishnan Marg, New Delhi-110012 (India)

    2009-11-30

    Biotinylated heparin has been immobilized onto self-assembled monolayer of 4-aminothiophenol using avidin-biotin specific binding. The modified electrodes have been characterized using surface plasmon resonance technique (SPR), cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), atomic force microscopy (AFM) and contact angle (CA) measurements. The interaction of immobilized biotinylated heparin with low density lipoprotein (LDL) has been studied using surface plasmon resonance technique. The biotinylated heparin modified electrode can be used to detect LDL in the range of 20 to 100 mg/dl with the sensitivity of 513.3 m{sup o}/{mu}M.

  3. Biotinylated Photopolymers for 3D-Printed Unibody Lab-on-a-Chip Optical Platforms.

    Science.gov (United States)

    Credi, Caterina; Griffini, Gianmarco; Levi, Marinella; Turri, Stefano

    2018-01-01

    The present work reports the first demonstration of straightforward fabrication of monolithic unibody lab-on-a-chip (ULOCs) integrating bioactive micrometric 3D scaffolds by means of multimaterial stereolithography (SL). To this end, a novel biotin-conjugated photopolymer is successfully synthesized and optimally formulated to achieve high-performance SL-printing resolution, as demonstrated by the SL-fabrication of biotinylated structures smaller than 100 µm. By optimizing a multimaterial single-run SL-based 3D-printing process, such biotinylated microstructures are incorporated within perfusion microchambers whose excellent optical transparency enables real-time optical microscopy analyses. Standard biotin-binding assays confirm the existence of biotin-heads on the surfaces of the embedded 3D microstructures and allow to demonstrate that the biofunctionality of biotin is not altered during the SL-printing, thus making it exploitable for further conjugation with other biomolecules. As a step forward, an in-line optical detection system is designed, prototyped via SL-printing and serially connected to the perfusion microchambers through customized world-to-chip connectors. Such detection system is successfully employed to optically analyze the solution flowing out of the microchambers, thus enabling indirect quantification of the concentration of target interacting biomolecules. The successful application of this novel biofunctional photopolymer as SL-material enables to greatly extend the versatility of SL to directly fabricate ULOCs with intrinsic biofunctionality. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Biotinylation of interleukin-2 (IL-2) for flow cytometric analysis of IL-2 receptor expression. Comparison of different methods

    NARCIS (Netherlands)

    M.O. de Jong (Marg); H. Rozemuller (Henk); J.G.J. Bauman (J. G J); J.W.M. Visser (Jan)

    1995-01-01

    textabstractThe main prerequisites for the use of biotinylated ligands to study the expression of growth factor receptors on heterogeneous cell populations, such as peripheral blood or bone marrow, by flow cytometric methods, are that the biotinylated ligand retains its binding ability and that

  5. Site-specific fab fragment biotinylation at the conserved nucleotide binding site for enhanced Ebola detection.

    Science.gov (United States)

    Mustafaoglu, Nur; Alves, Nathan J; Bilgicer, Basar

    2015-07-01

    The nucleotide binding site (NBS) is a highly conserved region between the variable light and heavy chains at the Fab domains of all antibodies, and a small molecule that we identified, indole-3-butyric acid (IBA), binds specifically to this site. Fab fragment, with its small size and simple production methods compared to intact antibody, is good candidate for use in miniaturized diagnostic devices and targeted therapeutic applications. However, commonly used modification techniques are not well suited for Fab fragments as they are often more delicate than intact antibodies. Fab fragments are of particular interest for sensor surface functionalization but immobilization results in damage to the antigen binding site and greatly reduced activity due to their truncated size that allows only a small area that can bind to surfaces without impeding antigen binding. In this study, we describe an NBS-UV photocrosslinking functionalization method (UV-NBS(Biotin) in which a Fab fragment is site-specifically biotinylated with an IBA-EG11-Biotin linker via UV energy exposure (1 J/cm(2)) without affecting its antigen binding activity. This study demonstrates successful immobilization of biotinylated Ebola detecting Fab fragment (KZ52 Fab fragment) via the UV-NBS(Biotin) method yielding 1031-fold and 2-fold better antigen detection sensitivity compared to commonly used immobilization methods: direct physical adsorption and NHS-Biotin functionalization, respectively. Utilization of the UV-NBS(Biotin) method for site-specific conjugation to Fab fragment represents a proof of concept use of Fab fragment for various diagnostic and therapeutic applications with numerous fluorescent probes, affinity molecules and peptides. © 2015 Wiley Periodicals, Inc.

  6. PET imaging of apoptosis with 64Cu-labeled streptavidin following pretargeting of phosphatidylserine with biotinylated annexin-V

    International Nuclear Information System (INIS)

    Cauchon, Nicole; Langlois, Rejean; Rousseau, Jacques A.; Tessier, Guillaume; Cadorette, Jules; Lecomte, Roger; Hunting, Darel J.; Lier, Johan E. van; Pavan, Roberto A.; Zeisler, Stefan K.

    2007-01-01

    In vivo detection of apoptosis is a diagnostic tool with potential clinical applications in cardiology and oncology. Radiolabeled annexin-V (anxV) is an ideal probe for in vivo apoptosis detection owing to its strong affinity for phosphatidylserine (PS), the molecular flag on the surface of apoptotic cells. Most clinical studies performed to visualize apoptosis have used 99m Tc-anxV; however, its poor distribution profile often compromises image quality. In this study, tumor apoptosis after therapy was visualized by positron emission tomography (PET) using 64 Cu-labeled streptavidin (SAv), following pre-targeting of apoptotic cells with biotinylated anxV. Apoptosis was induced in tumor-bearing mice by photodynamic therapy (PDT) using phthalocyanine dyes as photosensitizers, and red light. After PDT, mice were injected i.v. with biotinylated anxV, followed 2 h later by an avidin chase, and after another 2 h with 64 Cu-DOTA-biotin-SAv. PET images were subsequently recorded up to 13 h after PDT. PET images delineated apoptosis in treated tumors as early as 30 min after 64 Cu-DOTA-biotin-SAv administration, with tumor-to-background ratios reaching a maximum at 3 h post-injection, i.e., 7 h post-PDT. Omitting the administration of biotinylated anxV or the avidin chase failed to provide a clear PET image, confirming that all three steps are essential for adequate visualization of apoptosis. Furthermore, differences in action mechanisms between photosensitizers that target tumor cells directly or via initial vascular stasis were clearly recognized through differences in tracer uptake patterns detecting early or delayed apoptosis. This study demonstrates the efficacy of a three-step 64 Cu pretargeting procedure for PET imaging of apoptosis. Our data also confirm the usefulness of small animal PET to evaluate cancer treatment protocols. (orig.)

  7. Quantitation of biotinylated compounds by a solid-phase assay using a /sup 125/I-labelled biotin derivative

    Energy Technology Data Exchange (ETDEWEB)

    Smith, P J; Warren, R M; Holt, C von

    1987-05-11

    The biotin analogue biotinylglycyltyrosine has been synthesized and labelled to a specific activity of 2000 Ci/mmol with /sup 125/I. This analogue has been used in conjunction with immobilized streptavidin in an assay which detects as little as 1 fmol biotin or biotinylated molecules in solution. The determination of biotinylated insulin in a tissue extract and the quantitation of a transcription assay are given as examples. (Auth.). 12 refs.; 4 figs.; 3 tabs.

  8. Analysis of Biotinylated Generation 4 Poly(amidoamine (PAMAM Dendrimer Distribution in the Rat Brain and Toxicity in a Cellular Model of the Blood-Brain Barrier

    Directory of Open Access Journals (Sweden)

    Heather A. Bullen

    2013-09-01

    Full Text Available Dendrimers are highly customizable nanopolymers with qualities that make them ideal for drug delivery. The high binding affinity of biotin/avidin provides a useful approach to fluorescently label synthesized dendrimer-conjugates in cells and tissues. In addition, biotin may facilitate delivery of dendrimers through the blood-brain barrier (BBB via carrier-mediated endocytosis. The purpose of this research was to: (1 measure toxicity using lactate dehydrogenase (LDH assays of generation (G4 biotinylated and non-biotinylated poly(amidoamine (PAMAM dendrimers in a co-culture model of the BBB, (2 determine distribution of dendrimers in the rat brain, kidney, and liver following systemic administration of dendrimers, and (3 conduct atomic force microscopy (AFM on rat brain sections following systemic administration of dendrimers. LDH measurements showed that biotinylated dendrimers were toxic to cell co-culture after 48 h of treatment. Distribution studies showed evidence of biotinylated and non-biotinylated PAMAM dendrimers in brain. AFM studies showed evidence of dendrimers only in brain tissue of treated rats. These results indicate that biotinylation does not decrease toxicity associated with PAMAM dendrimers and that biotinylated PAMAM dendrimers distribute in the brain. Furthermore, this article provides evidence of nanoparticles in brain tissue following systemic administration of nanoparticles supported by both fluorescence microscopy and AFM.

  9. Local myocardial insulin-like growth factor 1 (IGF-1) delivery with biotinylated peptide nanofibers improves cell therapy for myocardial infarction

    Science.gov (United States)

    Davis, Michael E.; Hsieh, Patrick C. H.; Takahashi, Tomosaburo; Song, Qing; Zhang, Shuguang; Kamm, Roger D.; Grodzinsky, Alan J.; Anversa, Piero; Lee, Richard T.

    2006-05-01

    Strategies for cardiac repair include injection of cells, but these approaches have been hampered by poor cell engraftment, survival, and differentiation. To address these shortcomings for the purpose of improving cardiac function after injury, we designed self-assembling peptide nanofibers for prolonged delivery of insulin-like growth factor 1 (IGF-1), a cardiomyocyte growth and differentiation factor, to the myocardium, using a "biotin sandwich" approach. Biotinylated IGF-1 was complexed with tetravalent streptavidin and then bound to biotinylated self-assembling peptides. This biotin sandwich strategy allowed binding of IGF-1 but did not prevent self-assembly of the peptides into nanofibers within the myocardium. IGF-1 that was bound to peptide nanofibers activated Akt, decreased activation of caspase-3, and increased expression of cardiac troponin I in cardiomyocytes. After injection into rat myocardium, biotinylated nanofibers provided sustained IGF-1 delivery for 28 days, and targeted delivery of IGF-1 in vivo increased activation of Akt in the myocardium. When combined with transplanted cardiomyocytes, IGF-1 delivery by biotinylated nanofibers decreased caspase-3 cleavage by 28% and increased the myocyte cross-sectional area by 25% compared with cells embedded within nanofibers alone or with untethered IGF-1. Finally, cell therapy with IGF-1 delivery by biotinylated nanofibers improved systolic function after experimental myocardial infarction, demonstrating how engineering the local cellular microenvironment can improve cell therapy. engineering | maturation | scaffold

  10. Biotin IgM Antibodies in Human Blood: A Previously Unknown Factor Eliciting False Results in Biotinylation-Based Immunoassays

    Science.gov (United States)

    Chen, Tingting; Hedman, Lea; Mattila, Petri S.; Jartti, Laura; Jartti, Tuomas; Ruuskanen, Olli; Söderlund-Venermo, Maria; Hedman, Klaus

    2012-01-01

    Biotin is an essential vitamin that binds streptavidin or avidin with high affinity and specificity. As biotin is a small molecule that can be linked to proteins without affecting their biological activity, biotinylation is applied widely in biochemical assays. In our laboratory, IgM enzyme immuno assays (EIAs) of µ-capture format have been set up against many viruses, using as antigen biotinylated virus like particles (VLPs) detected by horseradish peroxidase-conjugated streptavidin. We recently encountered one serum sample reacting with the biotinylated VLP but not with the unbiotinylated one, suggesting in human sera the occurrence of biotin-reactive antibodies. In the present study, we search the general population (612 serum samples from adults and 678 from children) for IgM antibodies reactive with biotin and develop an indirect EIA for quantification of their levels and assessment of their seroprevalence. These IgM antibodies were present in 3% adults regardless of age, but were rarely found in children. The adverse effects of the biotin IgM on biotinylation-based immunoassays were assessed, including four inhouse and one commercial virus IgM EIAs, showing that biotin IgM do cause false positivities. The biotin can not bind IgM and streptavidin or avidin simultaneously, suggesting that these biotin-interactive compounds compete for the common binding site. In competitive inhibition assays, the affinities of biotin IgM antibodies ranged from 2.1×10−3 to 1.7×10−4 mol/L. This is the first report on biotin antibodies found in humans, providing new information on biotinylation-based immunoassays as well as new insights into the biomedical effects of vitamins. PMID:22879954

  11. Complexes of Streptavidin-Fused Antigens with Biotinylated Antibodies Targeting Receptors on Dendritic Cell Surface: A Novel Tool for Induction of Specific T-Cell Immune Responses

    Czech Academy of Sciences Publication Activity Database

    Staněk, Ondřej; Linhartová, Irena; Majlessi, L.; Leclerc, C.; Šebo, Peter

    2012-01-01

    Roč. 51, č. 3 (2012), s. 221-232 ISSN 1073-6085 R&D Projects: GA AV ČR KAN200520702; GA ČR GA310/08/0447; GA MŠk 2B06161 Institutional research plan: CEZ:AV0Z50200510 Keywords : Streptavidin * Antigen delivery * Biotinylated antibody Subject RIV: EE - Microbiology, Virology Impact factor: 2.262, year: 2012

  12. Biotinylated dextran amine anterograde tracing of the canine corticospinal tract?

    OpenAIRE

    Han, Xiao; Lv, Guangming; Wu, Huiqun; Ji, Dafeng; Sun, Zhou; Li, Yaofu; Tang, Lemin

    2012-01-01

    In this study, biotinylated dextran amine (BDA) was microinjected into the left cortical motor area of the canine brain. Fluorescence microscopy results showed that a large amount of BDA-labeled pyramidal cells were visible in the left cortical motor area after injection. In the left medulla oblongata, the BDA-labeled corticospinal tract was evenly distributed, with green fluorescence that had a clear boundary with the surrounding tissue. The BDA-positive corticospinal tract entered into the ...

  13. Effects of various spacers between biotin and the phospholipid headgroup on immobilization and sedimentation of biotinylated phospholipid-containing liposomes facilitated by avidin-biotin interactions.

    Science.gov (United States)

    Sakamoto, Yasuhisa; Kikuchi, Koji; Umeda, Kazuaki; Nakanishi, Hiroyuki

    2017-09-01

    Immobilization and sedimentation of liposomes (lipid vesicles) are used in liposome-protein binding assays, facilitated by avidin/streptavidin/NeutrAvidin and biotinylated phospholipid-containing liposomes. Here, we examined the effects of three spacers [six-carbon (X), polyethylene glycol (PEG) 180 (molecular weight 180) and PEG2000 (molecular weight 2,000)] between biotin and the phospholipid headgroup on the immobilization and sedimentation of small unilamellar liposomes/vesicles (SUVs). PEG180 and PEG2000 showed more efficient immobilization of biotinylated SUVs on NeutrAvidin-coated plates than X, but X and PEG180 showed more efficient sedimentation of biotinylated SUVs upon NeutrAvidin addition than PEG2000. Thus, the most appropriate spacers differed between immobilization and sedimentation. A spacer for biotinylated SUVs must be selected according to the particular liposome-protein binding assays examined. © The Authors 2017. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  14. Production of in vivo biotinylated scFv specific to almond (Prunus dulcis) proteins by recombinant Pichia pastoris.

    Science.gov (United States)

    de la Cruz, Silvia; Alcocer, Marcos; Madrid, Raquel; García, Aina; Martín, Rosario; González, Isabel; García, Teresa

    2016-06-10

    The methylotropic yeast Pichia pastoris has demonstrated its suitability for large-scale production of recombinant proteins. As an eukaryotic organism P. pastoris presents a series of advantages at expression and processing of heterologous proteins when compared with Escherichia coli. In this work, P. pastoris has been used to express a scFv from a human synthetic library previously shown to bind almond proteins. In order to facilitate purification and post processing manipulations, the scFv was engineered with a C-terminal tag and biotinylated in vivo. After purification, biotinylated scFv were bound to avidin conjugated with HRP producing a multimeric scFv. The multimeric scFv showed to maintain their ability to recognize almond protein when assayed in ELISA, reaching a LOD of 470mgkg(-1). This study describes an easy method to produce large quantities of in vivo biotinylated scFv in P. pastoris. By substituting the enzyme or fluorochromes linked to avidin, it will be possible to generate a diverse number of multimeric scFv as probes to suit different analytical platforms in the detection of almond in food products. Copyright © 2016 Elsevier B.V. All rights reserved.

  15. Comparison of the performance of the borax buffer-based HRP-enhanced reagent and the 'Lumi-Phos 530' chemiluminescence systems in the detection of biotinylated DNA.

    Science.gov (United States)

    Cercek, B; Roby, K; Siaw, M

    1995-01-01

    A comparison of two chemiluminescence methods, the borax buffer-based HRP-enhanced reagent and Lumi-Phos 530, applied to the detection of a biotinylated 30-mer DNA slot blotted onto a nylon membrane, is presented. A streptavidin-HRP and streptavidin-ALP mediated detection system was used. The HRP-enhanced system is up to 15-fold greater with respect to the signal/background ratios than the Lumi-Phos 530 system at 0.5 microgram biotinylated DNA with at least a two-fold improvement in detection sensitivity for 0.5 ng biotinylated DNA.

  16. Autodisplay of an avidin with biotin-binding activity on the surface of Escherichia coli.

    Science.gov (United States)

    Pardavé-Alejandre, H D; Alvarado-Yaah, J E; Pompa-Mera, E N; Muñoz-Medina, J E; Sárquiz-Martínez, B; Santacruz-Tinoco, C E; Manning-Cela, R G; Ortíz-Navarrete, V; López-Macías, C; González-Bonilla, C R

    2018-03-01

    To display a recombinant avidin fused to the autotransporter ShdA to bind biotinylated molecules on the surface of Escherichia coli. Two chimeric protein constructs containing avidin fused to the autotransporter ShdA were expressed on the surface of Escherichia coli DH5α. One fusion protein contained 476 amino acids of the ShdA α and β domains, whereas the second consisted of a 314 amino acid from α and truncated β domains. Protein production was verified by SDS-PAGE using an antibody to the molecular FLAG-tag. The surface display of the avidin-shdA fusion protein was confirmed by confocal microscopy and flow cytometry analysis, and the biotin-binding activity was evaluated by fluorescence microscopy and flow cytometry using biotin-4-fluorescein and biotinylated-ovalbumin (OVA). Expression of a recombinant avidin with biotin-binding activity on the surface of E. coli was achieved using the autotransporter ShdA. This system is an alternative to bind biotinylated molecules to E. coli.

  17. Multi-step surface functionalization of polyimide based evanescent wave photonic biosensors and application for DNA hybridization by Mach-Zehnder interferometer

    Energy Technology Data Exchange (ETDEWEB)

    Melnik, Eva [Health and Environment Department, Nano Systems, AIT Austrian Institute of Technology GmbH, Donau-City-Strasse 1, 1220 Vienna (Austria); Department of Analytical Chemistry, University of Vienna, Waehringer Strasse 38, 1090 Vienna (Austria); Bruck, Roman [Health and Environment Department, Nano Systems, AIT Austrian Institute of Technology GmbH, Donau-City-Strasse 1, 1220 Vienna (Austria); Hainberger, Rainer, E-mail: rainer.hainberger@ait.ac.at [Health and Environment Department, Nano Systems, AIT Austrian Institute of Technology GmbH, Donau-City-Strasse 1, 1220 Vienna (Austria); Laemmerhofer, Michael, E-mail: michael.laemmerhofer@univie.ac.at [Department of Analytical Chemistry, University of Vienna, Waehringer Strasse 38, 1090 Vienna (Austria)

    2011-08-12

    Highlights: {yields} We realize a biosensing platform for polyimide evanescent photonic wave sensors. {yields} We show that the surface functionalization via silanisation and biotinylation followed by streptavidin immobilization do not destroy or damage the thin polyimide film. {yields} A highly dense streptavidin layer enables the immobilisation of biotinylated ligands such as biotinylated ssDNA for the selective measurement of DNA hybridization. - Abstract: The process of surface functionalization involving silanization, biotinylation and streptavidin bonding as platform for biospecific ligand immobilization was optimized for thin film polyimide spin-coated silicon wafers, of which the polyimide film serves as a wave guiding layer in evanescent wave photonic biosensors. This type of optical sensors make great demands on the materials involved as well as on the layer properties, such as the optical quality, the layer thickness and the surface roughness. In this work we realized the binding of a 3-mercaptopropyl trimethoxysilane on an oxygen plasma activated polyimide surface followed by subsequent derivatization of the reactive thiol groups with maleimide-PEG{sub 2}-biotin and immobilization of streptavidin. The progress of the functionalization was monitored by using different fluorescence labels for optimization of the chemical derivatization steps. Further, X-ray photoelectron spectroscopy and atomic force microscopy were utilized for the characterization of the modified surface. These established analytical methods allowed to derive information like chemical composition of the surface, surface coverage with immobilized streptavidin, as well as parameters of the surface roughness. The proposed functionalization protocol furnished a surface density of 144 fmol mm{sup -2} streptavidin with good reproducibility (13.9% RSD, n = 10) and without inflicted damage to the surface. This surface modification was applied to polyimide based Mach-Zehnder interferometer

  18. A comprehensive platform for the analysis of ubiquitin-like protein modifications using in vivo biotinylation

    DEFF Research Database (Denmark)

    Pirone, Lucia; Xolalpa, Wendy; Sigurdsson, Jón Otti

    2017-01-01

    L conjugates from interactors, and low quantities of modified substrates. Here we describe bioUbLs, a comprehensive set of tools for studying modifications in Drosophila and mammals, based on multicistronic expression and in vivo biotinylation using the E. coli biotin protein ligase BirA. While the bio...

  19. Targeting protein biotinylation enhances tuberculosis chemotherapy.

    Science.gov (United States)

    Tiwari, Divya; Park, Sae Woong; Essawy, Maram M; Dawadi, Surendra; Mason, Alan; Nandakumar, Madhumitha; Zimmerman, Matthew; Mina, Marizel; Ho, Hsin Pin; Engelhart, Curtis A; Ioerger, Thomas; Sacchettini, James C; Rhee, Kyu; Ehrt, Sabine; Aldrich, Courtney C; Dartois, Véronique; Schnappinger, Dirk

    2018-04-25

    Successful drug treatment for tuberculosis (TB) depends on the unique contributions of its component drugs. Drug resistance poses a threat to the efficacy of individual drugs and the regimens to which they contribute. Biologically and chemically validated targets capable of replacing individual components of current TB chemotherapy are a major unmet need in TB drug development. We demonstrate that chemical inhibition of the bacterial biotin protein ligase (BPL) with the inhibitor Bio-AMS (5'-[ N -(d-biotinoyl)sulfamoyl]amino-5'-deoxyadenosine) killed Mycobacterium tuberculosis ( Mtb ), the bacterial pathogen causing TB. We also show that genetic silencing of BPL eliminated the pathogen efficiently from mice during acute and chronic infection with Mtb Partial chemical inactivation of BPL increased the potency of two first-line drugs, rifampicin and ethambutol, and genetic interference with protein biotinylation accelerated clearance of Mtb from mouse lungs and spleens by rifampicin. These studies validate BPL as a potential drug target that could serve as an alternate frontline target in the development of new drugs against Mtb . Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  20. Oriented Immobilization of Fab Fragments by Site-Specific Biotinylation at the Conserved Nucleotide Binding Site for Enhanced Antigen Detection.

    Science.gov (United States)

    Mustafaoglu, Nur; Alves, Nathan J; Bilgicer, Basar

    2015-09-08

    Oriented immobilization of antibodies and antibody fragments has become increasingly important as a result of the efforts to reduce the size of diagnostic and sensor devices to miniaturized dimensions for improved accessibility to the end-user. Reduced dimensions of sensor devices necessitate the immobilized antibodies to conserve their antigen binding activity for proper operation. Fab fragments are becoming more commonly used in small-scaled diagnostic devices due to their small size and ease of manufacture. In this study, we used the previously described UV-NBS(Biotin) method to functionalize Fab fragments with IBA-EG11-Biotin linker utilizing UV energy to initiate a photo-cross-linking reaction between the nucleotide binding site (NBS) on the Fab fragment and IBA-Biotin molecule. Our results demonstrate that immobilization of biotinylated Fab fragments via UV-NBS(Biotin) method generated the highest level of immobilized Fab on surfaces when compared to other typical immobilization methods while preserving antigen binding activity. UV-NBS(Biotin) method provided 432-fold, 114-fold, and 29-fold improved antigen detection sensitivity than physical adsorption, NHS-Biotin, and ε-NH3(+), methods, respectively. Additionally, the limit of detection (LOD) for PSA utilizing Fab fragments immobilized via UV-NBS(Biotin) method was significantly lower than that of the other immobilization methods, with an LOD of 0.4 pM PSA. In summary, site-specific biotinylation of Fab fragments without structural damage or loss in antigen binding activity provides a wide range of application potential for UV-NBS immobilization technique across numerous diagnostic devices and nanotechnologies.

  1. Binding analysis of ferritin with heme using α-casein and biotinylated-hemin: detection of heme-binding capacity of Dpr derived from heme synthesis-deficient Streptococcus mutans.

    Science.gov (United States)

    Mieno, Ayako; Yamamoto, Yuji; Yoshikawa, Yasunaga; Watanabe, Kiyotaka; Mukai, Takao; Orino, Koichi

    2013-01-01

    Bacterial and mammalian ferritins are known to bind heme. The use of α-casein and biotinylated hemin could be applicable to detection of protein-bound heme and of proteins with heme-binding capacity, respectively. Although commercial horse spleen ferritin and purified horse spleen ferritin (L:H subunit ratio=4) bound to an α-casein-coated plate, and this binding could be inhibited by hemin, recombinant iron-binding protein (rDpr), derived from heme-deficient Streptococcus mutans and expressed in Escherichia coli, did not bind to an α-casein-coated plate. Both horse spleen ferritins bound to α-casein-immobilized beads. Commercial horse spleen ferritin and rDpr showed direct binding to hemin-agarose beads. After preincubation of commercial horse spleen ferritin or rDpr with biotinylated hemin, they showed indirect binding to avidin-immobilized beads through biotinylated hemin. These results demonstrate that α-casein is useful for detection of heme-binding ferritin and that both hemin-agarose and the combination of biotinylated hemin and avidin-beads are useful for detection of the heme-binding capacity of ferritin. In addition, this study also revealed that Dpr, a decameric iron-binding protein, from heme-deficient cells binds heme.

  2. Targeting of liposomes to cells bearing nerve growth factor receptors mediated by biotinylated NGF

    International Nuclear Information System (INIS)

    Rosenberg, M.B.

    1986-01-01

    Previous studies of liposome targeting have concentrated on immunological systems, the use of ligand-receptor interactions has received little attention. The protein hormone beta-nerve growth factor (NGF) was modified by biotinylation via carboxyl group substitution (C-bio-NGF) under reaction conditions that yielded an average of 3 biotin additions per NGF subunit. NGF was also biotinylated through amino group substitution to produce derivatives with ratios of 1, 2 and 4 biotin moieties per NGF subunit (N-bio-NGF). These derivatives were compared with native NGF for their ability to compete with 125 I-NGF for binding to NGF receptors on rat pheochromocytoma (PC 12) cells at 4 0 C. C-bio-NGF was as effective as native NGF in binding to NGF receptors, while N-bio-NGF containing 1 biotin per NGF subunit was only 28% as active in binding as native NGF. C-bio-NGF, but not N-bio-NGF, mediated the specific binding of 125 I-streptavidin to PC12 cells. Biocytin-NGF, a derivative of C-bio-NGF with an extended spacer chain, was also synthesized and retained full biological and receptor binding activities. C-bio-NGF and biocytin-NGF were as effective as native NGF in a bioassay involving induction of neurite outgrowth from PC12 cells

  3. Detection of chromogranins A and B in endocrine tissues with radioactive and biotinylated oligonucleotide probes

    International Nuclear Information System (INIS)

    Lloyd, R.V.; Jin, L.; Fields, K.

    1990-01-01

    We analyzed the distribution of chromogranins A and B in normal and neoplastic endocrine tissues with secretory granules using 35 S-labeled and biotin-labeled oligonucleotide probes by in situ hybridization (ISH). Both radioactive and nonradioactive probes detected messenger RNAs (mRNAs) in frozen and paraffin tissue sections. Endocrine tissues with variable immunoreactivities for chromogranin A protein, such as small-cell lung carcinomas, neuroblastomas, insulinomas, and parathyroid adenomas, expressed the mRNA for chromogranins A and B in most cells. Some technical problems with the biotinylated probes included nonspecific nuclear staining and endogenous alkaline phosphatase, which was not completely abolished by levamisole pretreatment. A differential distribution of chromogranins A and B was seen in pituitary prolactinomas, which expressed abundant chromogranin B but not chromogranin A mRNAs, and in parathyroid adenomas, which expressed abundant chromogranin A but only small amounts of chromogranin B mRNAs. These results indicate that ISH can be used to detect chromogranins A and B in endocrine tissues with radioactive and biotinylated oligonucleotide probes and that the mRNAs for chromogranin A and B are demonstrable in some tumors even when the chromogranin proteins cannot be detected by immunohistochemistry

  4. Detection of chromogranins A and B in endocrine tissues with radioactive and biotinylated oligonucleotide probes

    Energy Technology Data Exchange (ETDEWEB)

    Lloyd, R.V.; Jin, L.; Fields, K. (Univ. of Michigan, Ann Arbor (USA))

    1990-01-01

    We analyzed the distribution of chromogranins A and B in normal and neoplastic endocrine tissues with secretory granules using {sup 35}S-labeled and biotin-labeled oligonucleotide probes by in situ hybridization (ISH). Both radioactive and nonradioactive probes detected messenger RNAs (mRNAs) in frozen and paraffin tissue sections. Endocrine tissues with variable immunoreactivities for chromogranin A protein, such as small-cell lung carcinomas, neuroblastomas, insulinomas, and parathyroid adenomas, expressed the mRNA for chromogranins A and B in most cells. Some technical problems with the biotinylated probes included nonspecific nuclear staining and endogenous alkaline phosphatase, which was not completely abolished by levamisole pretreatment. A differential distribution of chromogranins A and B was seen in pituitary prolactinomas, which expressed abundant chromogranin B but not chromogranin A mRNAs, and in parathyroid adenomas, which expressed abundant chromogranin A but only small amounts of chromogranin B mRNAs. These results indicate that ISH can be used to detect chromogranins A and B in endocrine tissues with radioactive and biotinylated oligonucleotide probes and that the mRNAs for chromogranin A and B are demonstrable in some tumors even when the chromogranin proteins cannot be detected by immunohistochemistry.

  5. Biotinylation of lysine method identifies acetylated histone H3 lysine 79 in Saccharomyces cerevisiae as a substrate for Sir2.

    Science.gov (United States)

    Bheda, Poonam; Swatkoski, Stephen; Fiedler, Katherine L; Boeke, Jef D; Cotter, Robert J; Wolberger, Cynthia

    2012-04-17

    Although the biological roles of many members of the sirtuin family of lysine deacetylases have been well characterized, a broader understanding of their role in biology is limited by the challenges in identifying new substrates. We present here an in vitro method that combines biotinylation and mass spectrometry (MS) to identify substrates deacetylated by sirtuins. The method permits labeling of deacetylated residues with amine-reactive biotin on the ε-nitrogen of lysine. The biotin can be utilized to purify the substrate and identify the deacetylated lysine by MS. The biotinyl-lysine method was used to compare deacetylation of chemically acetylated histones by the yeast sirtuins, Sir2 and Hst2. Intriguingly, Sir2 preferentially deacetylates histone H3 lysine 79 as compared to Hst2. Although acetylation of K79 was not previously reported in Saccharomyces cerevisiae, we demonstrate that a minor population of this residue is indeed acetylated in vivo and show that Sir2, and not Hst2, regulates the acetylation state of H3 lysine 79. The in vitro biotinyl-lysine method combined with chemical acetylation made it possible to identify this previously unknown, low-abundance histone acetyl modification in vivo. This method has further potential to identify novel sirtuin deacetylation substrates in whole cell extracts, enabling large-scale screens for new deacetylase substrates.

  6. Buoyancy-activated cell sorting using targeted biotinylated albumin microbubbles.

    Directory of Open Access Journals (Sweden)

    Yu-Ren Liou

    Full Text Available Cell analysis often requires the isolation of certain cell types. Various isolation methods have been applied to cell sorting, including fluorescence-activated cell sorting and magnetic-activated cell sorting. However, these conventional approaches involve exerting mechanical forces on the cells, thus risking cell damage. In this study we applied a novel isolation method called buoyancy-activated cell sorting, which involves using biotinylated albumin microbubbles (biotin-MBs conjugated with antibodies (i.e., targeted biotin-MBs. Albumin MBs are widely used as contrast agents in ultrasound imaging due to their good biocompatibility and stability. For conjugating antibodies, biotin is conjugated onto the albumin MB shell via covalent bonds and the biotinylated antibodies are conjugated using an avidin-biotin system. The albumin microbubbles had a mean diameter of 2 μm with a polydispersity index of 0.16. For cell separation, the MDA-MB-231 cells are incubated with the targeted biotin-MBs conjugated with anti-CD44 for 10 min, centrifuged at 10 g for 1 min, and then allowed 1 hour at 4 °C for separation. The results indicate that targeted biotin-MBs conjugated with anti-CD44 antibodies can be used to separate MDA-MB-231 breast cancer cells; more than 90% of the cells were collected in the MB layer when the ratio of the MBs to cells was higher than 70:1. Furthermore, we found that the separating efficiency was higher for targeted biotin-MBs than for targeted avidin-incorporated albumin MBs (avidin-MBs, which is the most common way to make targeted albumin MBs. We also demonstrated that the recovery rate of targeted biotin-MBs was up to 88% and the sorting purity was higher than 84% for a a heterogenous cell population containing MDA-MB-231 cells (CD44(+ and MDA-MB-453 cells (CD44-, which are classified as basal-like breast cancer cells and luminal breast cancer cells, respectively. Knowing that the CD44(+ is a commonly used cancer

  7. In vitro evaluation of avidin antibody pretargeting using 211At-labeled and biotinylated poly-L-lysine as effector molecule

    DEFF Research Database (Denmark)

    Frost, Sofia H L; Jensen, Holger Lau; Lindegren, Sture

    2010-01-01

    Pretargeting is an approach for enhancing the therapeutic index of radioimmunotherapy by separating the administrations of tumor-targeting substance and radiolabel. In this study, a pretargeting model system of avidin-conjugated monoclonal antibody trastuzumab and biotinylated, (211)At-labeled poly...

  8. Expression of proto-oncogenes in non-Hodgkin's lymphomas by in situ hybridization with biotinylated DNA probes

    International Nuclear Information System (INIS)

    Hamatani, Kiyohiro; Yoshida, Kuniko; Abe, Masumi; Shimaoka, Katsutaro; Shiku, Hiroshi; Akiyama, Mitoshi; Kondo, Hisayoshi.

    1989-11-01

    Expression of six proto-oncogenes (fos, myc, myb, Ki-ras, Ha-ras, and N-ras) in 43 cases of non-Hodgkin's lymphoma was analyzed by means of in situ hybridization. Biotinylated DNA probes of the six oncogenes and those of the immunoglobulin H-chain (IgH) gene and the T cell receptor β-chain (TCRβ) gene were used. The results of in situ hybridization performed under blind conditions by IgH and TCRβ gene probes were compatible with those of typing by cell surface markers. The nuclear protein-related proto-oncogenes, fos myc, and myb, were expressed in about 70 % - 80 % of all cases regardless of phenotypes, histology or histologic grade. On the contrary, genes of the ras family were expressed in fewer cases except for the Ki-ras gene which was more frequently expressed by cases of the T cell immunophenotype with a high malignancy grade. The results of dot hybridization with RNA extracted from some cases were compatible with those of in situ hybridization, further demonstrating the specificity of in situ hybridization. (author)

  9. Binding assays with streptavidin-functionalized superparamagnetic nanoparticles and biotinylated analytes using fluxgate magnetorelaxometry

    International Nuclear Information System (INIS)

    Heim, Erik; Ludwig, Frank; Schilling, Meinhard

    2009-01-01

    Binding assays based on the magnetorelaxation of superparamagnetic nanoparticles as markers are presented utilizing a differential fluxgate system. As ligand and receptor, streptavidin and biotin, respectively, are used. Superparamagnetic nanoparticles are functionalized with streptavidin and bound to two types of biotinylated analytes: agarose beads and bovine serum (BSA) proteins. The size difference of the two analytes causes a different progress of the reaction. As a consequence, the analysis of the relaxation signal is carried out dissimilarly for the two analytes. In addition, we studied the reaction kinetics of the two kinds of analytes with the fluxgate system.

  10. Radiolabeling and biotinylation of internalizing monoclonal antibody chimeric BR96: Potential use of extracorporeal immunoadsorption with enhanced tumor radioactivity retention of Iodine, Indium and Rhenium

    International Nuclear Information System (INIS)

    Chen, JianQing.

    1997-01-01

    In this thesis, methodology of radiolabeling and simultaneous biotinylation for internalizing monoclonal antibody chimeric BR96 have been investigated by using three element groups of potential therapeutic radionuclides iodine, indium and rhenium, and their different labeling methods. The biodistribution and kinetics of biotinylated and radiolabeled chiBR96 have been studied in colon carcinoma isografted rats. The potential use of ECIA, based on the biotin-avidin concept, has been evaluated and compared with the approach of avidin 'chase' in the same animal tumor model with respect to an enhancement of tumor-to-normal tissue (T/N) activity ratio. 131 refs

  11. Radiolabeling and biotinylation of internalizing monoclonal antibody chimeric BR96: Potential use of extracorporeal immunoadsorption with enhanced tumor radioactivity retention of Iodine, Indium and Rhenium

    Energy Technology Data Exchange (ETDEWEB)

    Chen, JianQing

    1997-01-01

    In this thesis, methodology of radiolabeling and simultaneous biotinylation for internalizing monoclonal antibody chimeric BR96 have been investigated by using three element groups of potential therapeutic radionuclides iodine, indium and rhenium, and their different labeling methods. The biodistribution and kinetics of biotinylated and radiolabeled chiBR96 have been studied in colon carcinoma isografted rats. The potential use of ECIA, based on the biotin-avidin concept, has been evaluated and compared with the approach of avidin `chase` in the same animal tumor model with respect to an enhancement of tumor-to-normal tissue (T/N) activity ratio. 131 refs.

  12. Size matters: influence of the size of nanoparticles on their interactions with ligands immobilized on the solid surface.

    Science.gov (United States)

    Piletska, Elena V; Piletsky, Sergey A

    2010-03-16

    The correlation between the size of biotinylated nanoparticles and their affinity in relation to interactions with the solid surface was investigated. The silica particles with a diameter of 50-200 nm containing amino groups on the surface were labeled with different quantities of biotin. The affinity properties of biotinylated nanoparticles were studied using a Biacore 3000 instrument equipped with a streptavidin-coated sensor chip (SA chip). It was shown that the increase in the particle size from 50 to 200 nm reduced the affinity (K(D)) of biotin-streptavidin interactions from 1.2 x 10(-12) to 1.2 x 10(-10) M. It was found that the particles with higher concentrations of immobilized biotin on particle surfaces demonstrated stronger binding with streptavidin.

  13. Effects of Biotin Deficiency on Biotinylated Proteins and Biotin-Related Genes in the Rat Brain.

    Science.gov (United States)

    Yuasa, Masahiro; Aoyama, Yuki; Shimada, Ryoko; Sawamura, Hiromi; Ebara, Shuhei; Negoro, Munetaka; Fukui, Toru; Watanabe, Toshiaki

    2016-01-01

    Biotin is a water-soluble vitamin that functions as a cofactor for biotin-dependent carboxylases. The biochemical and physiological roles of biotin in brain regions have not yet been investigated sufficiently in vivo. Thus, in order to clarify the function of biotin in the brain, we herein examined biotin contents, biotinylated protein expression (e.g. holocarboxylases), and biotin-related gene expression in the brain of biotin-deficient rats. Three-week-old male Wistar rats were divided into a control group, biotin-deficient group, and pair-fed group. Rats were fed experimental diets from 3 wk old for 8 wk, and the cortex, hippocampus, striatum, hypothalamus, and cerebellum were then collected. In the biotin-deficient group, the maintenance of total biotin and holocarboxylases, increases in the bound form of biotin and biotinidase activity, and the expression of an unknown biotinylated protein were observed in the cortex. In other regions, total and free biotin contents decreased, holocarboxylase expression was maintained, and bound biotin and biotinidase activity remained unchanged. Biotin-related gene (pyruvate carboxylase, sodium-dependent multivitamin transporter, holocarboxylase synthetase, and biotinidase) expression in the cortex and hippocampus also remained unchanged among the dietary groups. These results suggest that biotin may be related to cortex functions by binding protein, and the effects of a biotin deficiency and the importance of biotin differ among the different brain regions.

  14. Detection of HCV-RNA by Reverse Transcription Polymerase Chain Reaction Using Biotinylated and Radioiodinated Primers

    International Nuclear Information System (INIS)

    Ryu, Jin Sook; Moon, Dae Hyuk; Cheon, Jun Hong; Chung, Yoon Young; Park, Hung Dong; Chung, Young Hwa; Lee, Young Sang

    1994-01-01

    This study was performed to evaluate the clinical applicability of the reverse transcription polymerase chain reaction (RT-PCR) kit of HCV-RNA using biotinylated and radioiodinated primers. Study subjects were 118 patients with positive anti-HCV. HCV-RNA in patients serum was extracted by guanidium thiocyanate method. After first amplification, the product was reamplified by primers labelled with biotin and I-125. The final amplification product was detected by counting the radioactivity after incubation in avidin coated tubes. In 51 samples, the test was repeated for evaluation of reproducibility. This new method was also compared with conventional RT-PCR methods in 34 samples from patients with chronic liver disease. The results were as follows, 1) HCV-RNA was positive in 85(97%)of 88 patients with chronic liver disease, and in 23 (73%) of 30 patients with normal liver function. 2) In comparison with conventional method, HCV-RNA was detected in 32(94%) of 34 patients with new method, whereas in 27(79% ) of the same group with conventional method 3) Repeated test with new method in 52 samples demonstrated 82% of concordant result. In conclusion, new method with biotinylated and radioiodinated primers was more sensitive than conventional method. However, great care must be taken for quality control because there were considerable interassay variation and possibility of false positivity and false negativity.

  15. Detection of HCV-RNA by Reverse Transcription Polymerase Chain Reaction Using Biotinylated and Radioiodinated Primers

    Energy Technology Data Exchange (ETDEWEB)

    Ryu, Jin Sook; Moon, Dae Hyuk; Cheon, Jun Hong; Chung, Yoon Young; Park, Hung Dong; Chung, Young Hwa; Lee, Young Sang [Asan Medical Center, University of Ulsan, Seoul (Korea, Republic of)

    1994-07-15

    This study was performed to evaluate the clinical applicability of the reverse transcription polymerase chain reaction (RT-PCR) kit of HCV-RNA using biotinylated and radioiodinated primers. Study subjects were 118 patients with positive anti-HCV. HCV-RNA in patients serum was extracted by guanidium thiocyanate method. After first amplification, the product was reamplified by primers labelled with biotin and I-125. The final amplification product was detected by counting the radioactivity after incubation in avidin coated tubes. In 51 samples, the test was repeated for evaluation of reproducibility. This new method was also compared with conventional RT-PCR methods in 34 samples from patients with chronic liver disease. The results were as follows, 1) HCV-RNA was positive in 85(97%)of 88 patients with chronic liver disease, and in 23 (73%) of 30 patients with normal liver function. 2) In comparison with conventional method, HCV-RNA was detected in 32(94%) of 34 patients with new method, whereas in 27(79% ) of the same group with conventional method 3) Repeated test with new method in 52 samples demonstrated 82% of concordant result. In conclusion, new method with biotinylated and radioiodinated primers was more sensitive than conventional method. However, great care must be taken for quality control because there were considerable interassay variation and possibility of false positivity and false negativity.

  16. Affinity cytochemistry of vascular endothelia in brain tumors by biotinylated Ulex europaeus type I lectin (UEA I).

    Science.gov (United States)

    Weber, T; Seitz, R J; Liebert, U G; Gallasch, E; Wechsler, W

    1985-01-01

    The vascularization of 50 tumors of the central nervous system (CNS) including 17 meningiomas, 25 neuroectodermal tumors, i.e., astrocytomas, oligodendrogliomas, mixed gliomas, glioblastomas, medulloblastomas, seven metastatic carcinomas, and one malignant hemangioendothelioma were investigated using biotinylated Ulex europaeus type I lectin (UEA I) in an indirect avidinbiotin-peroxidase procedure. The cytochemical staining pattern of UEA I on paraffin sections was compared with that of biotinylated Dolichos biflorus lectin (DBA), and with the immunocytochemical staining of factor VIII related antigen (F VIII/RAG) by polyclonal antisera using the PAP technique. UEA I visualized the endothelia of blood vessels with equal intensity, sensitivity, and reliability in normal brain and in tumor tissue with neovascularization. While large, medium, and small vessels were equally well demonstrated by UEA I and antibodies against FVIII/RAG, capillaries and endothelial sprouts were stained more consistently and intensely by UEA I. No reliable cytochemical staining could be obtained by DBA regardless of tissue or cell type investigated. It is concluded that UEA I is a highly useful cytochemical marker for the identification of vascular endothelia in paraffin sections of human brain tumors.

  17. Selective radiolabeling of cell surface proteins to a high specific activity

    International Nuclear Information System (INIS)

    Thompson, J.A.; Lau, A.L.; Cunningham, D.D.

    1987-01-01

    A procedure was developed for selective radiolabeling of membrane proteins on cells to higher specific activities than possible with available techniques. Cell surface amino groups were derivatized with 125 I-(hydroxyphenyl)propionyl groups via 125 I-sulfosuccinimidyl (hydroxyphenyl)propionate ( 125 II-sulfo-SHPP). This reagent preferentially labeled membrane proteins exposed at the cell surface of erythrocytes as assessed by the degree of radiolabel incorporation into erythrocyte ghost proteins and hemoglobin. Comparison with the lactoperoxidase-[ 125 I]iodide labeling technique revealed that 125 I-sulfo-SHPP labeled cell surface proteins to a much higher specific activity and hemoglobin to a much lower specific activity. Additionally, this reagent was used for selective radiolabeling of membrane proteins on the cytoplasmic face of the plasma membrane by blocking exofacial amino groups with uniodinated sulfo-SHPP, lysing the cells, and then incubating them with 125 I-sulfo-SHPP. Exclusive labeling of either side of the plasma membrane was demonstrated by the labeling of some marker proteins with well-defined spacial orientations on erythroctyes. Transmembrane proteins such as the epidermal growth factor receptor on cultured cells could also be labeled differentially from either side of the plasma membrane

  18. Biotinylated N-Acetyllactosamine- and N,N-Diacetyllactosamine-Based Oligosaccharides as Novel Ligands for Human Galectin-3

    Directory of Open Access Journals (Sweden)

    Sophia Böcker

    2017-04-01

    Full Text Available Galectin inhibitor design is an emerging research field due to the involvement of galectins in cancer. Galectin-3, in particular, plays an important role in tumor progression. To generate inhibitors, modifications of the glycan structure can be introduced. Conjugation of hydrophobic compounds to saccharides has proven to be promising as increased binding of galectin-3 can be observed. In the present study, we report on neo-glycans carrying hydrophobic biotin as novel ligands for human galectin-3. We modified N-acetyllactosamine- and N,N-diacetyllactosamine-based tetrasaccharides at the C6-position of the terminal saccharide unit using selective enzymatic oxidation and subsequent chemical conjugation of biotinamidohexanoic acid hydrazide. These neo-glycans were much better bound by galectin-3 than the unmodified counterparts. High selectivity for galectin-3 over galectin-1 was also proven. We generated multivalent neo-glycoproteins by conjugation of neo-glycans to bovine serum albumin showing high affinity for galectin-3. Compared to non-biotinylated neo-glycoproteins, we achieved high binding levels of galectin-3 with a lesser amount of conjugated neo-glycans. Multivalent ligand presentation of neo-glycoproteins significantly increased the inhibitory potency towards galectin-3 binding to asialofetuin when compared to free monovalent glycans. Our findings show the positive impact of 6-biotinylation of tetrasaccharides on galectin-3 binding, which broadens the recent design approaches for producing high-affinity ligands.

  19. Influence of impaired lipoprotein biogenesis on surface and exoproteome of Streptococcus pneumoniae.

    Science.gov (United States)

    Pribyl, Thomas; Moche, Martin; Dreisbach, Annette; Bijlsma, Jetta J E; Saleh, Malek; Abdullah, Mohammed R; Hecker, Michael; van Dijl, Jan Maarten; Becher, Dörte; Hammerschmidt, Sven

    2014-02-07

    Surface proteins are important for the fitness and virulence of the Gram-positive pathogen Streptococcus pneumoniae. They are crucial for interaction of the pathogen with its human host during infection. Therefore, the analysis of the pneumococcal surface proteome is an important task that requires powerful tools. In this study, two different methods, an optimized biotinylation approach and shaving with trypsin beads, were applied to study the pneumococcal surface proteome and to identify surface-exposed protein domains, respectively. The identification of nearly 95% of the predicted lipoproteins and 75% of the predicted sortase substrates reflects the high coverage of the two classical surface protein classes accomplished in this study. Furthermore, the biotinylation approach was applied to study the impact of an impaired lipoprotein maturation pathway on the cell envelope proteome and exoproteome. Loss of the lipoprotein diacylglyceryl transferase Lgt leads to striking changes in the lipoprotein distribution. Many lipoproteins disappear from the surface proteome and accumulate in the exoproteome. Further insights into lipoprotein processing in pneumococci are provided by immunoblot analyses of bacterial lysates and corresponding supernatant fractions. Taken together, the first comprehensive overview of the pneumococcal surface and exoproteome is presented, and a model for lipoprotein processing in S. pneumoniae is proposed.

  20. Synthesis, kinetic evaluation, and utilization of a biotinylated dipeptide proline diphenyl phosphonate for the disclosure of dipeptidyl peptidase IV-like serine proteases.

    Science.gov (United States)

    Gilmore, Brendan F; Carson, Louise; McShane, Laura L; Quinn, Derek; Coulter, Wilson A; Walker, Brian

    2006-08-18

    In this study, we report on the synthesis, kinetic characterisation, and application of a novel biotinylated and active site-directed inactivator of dipeptidyl peptidase IV (DPP-IV). Thus, the dipeptide-derived proline diphenyl phosphonate NH(2)-Glu(biotinyl-PEG)-Pro(P)(OPh)(2) has been prepared by a combination of classical solution- and solid-phase methodologies and has been shown to be an irreversible inhibitor of porcine DPP-IV, exhibiting an over all second-order rate constant (k(i)/K(i)) for inhibition of 1.57 x 10(3) M(-1) min(-1). This value compares favourably with previously reported rates of inactivation of DPP-IV by dipeptides containing a P(1) proline diphenyl phosphonate grouping [B. Boduszek, J. Oleksyszyn, C.M. Kam, J. Selzler, R.E. Smith, J.C. Powers, Dipeptide phophonates as inhibitors of dipeptidyl peptidase IV, J. Med. Chem. 37 (1994) 3969-3976; B.F. Gilmore, J.F. Lynas, C.J. Scott, C. McGoohan, L. Martin, B. Walker, Dipeptide proline diphenyl phosphonates are potent, irreversible inhibitors of seprase (FAPalpha), Biochem, Biophys. Res. Commun. 346 (2006) 436-446.], thus demonstrating that the incorporation of the side-chain modified (N-biotinyl-3-(2-(2-(3-aminopropyloxy)-ethoxy)-ethoxy)-propyl) glutamic acid residue at the P(2) position is compatible with inhibitor efficacy. The utilisation of this probe for the detection of both purified dipeptidyl peptidase IV and the disclosure of a dipeptidyl peptidase IV-like activity from a clinical isolate of Porphyromonas gingivalis, using established electrophoretic and Western blotting techniques previously developed by our group, is also demonstrated.

  1. Direct Detection of Biotinylated Proteins by Mass Spectrometry

    Science.gov (United States)

    2015-01-01

    Mass spectrometric strategies to identify protein subpopulations involved in specific biological functions rely on covalently tagging biotin to proteins using various chemical modification methods. The biotin tag is primarily used for enrichment of the targeted subpopulation for subsequent mass spectrometry (MS) analysis. A limitation of these strategies is that MS analysis does not easily discriminate unlabeled contaminants from the labeled protein subpopulation under study. To solve this problem, we developed a flexible method that only relies on direct MS detection of biotin-tagged proteins called “Direct Detection of Biotin-containing Tags” (DiDBiT). Compared with conventional targeted proteomic strategies, DiDBiT improves direct detection of biotinylated proteins ∼200 fold. We show that DiDBiT is applicable to several protein labeling protocols in cell culture and in vivo using cell permeable NHS-biotin and incorporation of the noncanonical amino acid, azidohomoalanine (AHA), into newly synthesized proteins, followed by click chemistry tagging with biotin. We demonstrate that DiDBiT improves the direct detection of biotin-tagged newly synthesized peptides more than 20-fold compared to conventional methods. With the increased sensitivity afforded by DiDBiT, we demonstrate the MS detection of newly synthesized proteins labeled in vivo in the rodent nervous system with unprecedented temporal resolution as short as 3 h. PMID:25117199

  2. Cell surface localization of the 78 kD glucose regulated protein (GRP 78) induced by thapsigargin.

    Science.gov (United States)

    Delpino, A; Piselli, P; Vismara, D; Vendetti, S; Colizzi, V

    1998-01-01

    In the present study it was found that the synthesis of the 78 kD glucose-regulated protein (GRP 78 or BIP) is vigorously induced in human rabdomiosarcoma cells (TE 671/RD) following both short-term (1 h) and prolonged (18 h) exposure to 100 nM thapsigargin (Tg). Flow cytometric analysis with a specific anti-GRP 78 polyclonal antibody showed that Tg-treated cells express the GRP 78 on the plasma membrane. Cell surface localization of the Tg-induced GRP 78 was confirmed by biotinylation of membrane-exposed proteins and subsequent isolation of the biotin-labelled proteins by streptavidin/agarose affinity chromatography. It was found that a fraction of the Tg-induced GRP 78 is present among the biotin-labelled, surface-exposed, proteins. Conversely, the GRP 78 immunoprecipitated from unfractionated lysates of Tg-treated and biotin-reacted cells was found to be biotinylated. This is the first report demonstrating surface expression of GRP 78 in cells exposed to a specific GRP 78-inducing stimulus.

  3. A novel biotinylated lipid raft reporter for electron microscopic imaging of plasma membrane microdomains[S

    Science.gov (United States)

    Krager, Kimberly J.; Sarkar, Mitul; Twait, Erik C.; Lill, Nancy L.; Koland, John G.

    2012-01-01

    The submicroscopic spatial organization of cell surface receptors and plasma membrane signaling molecules is readily characterized by electron microscopy (EM) via immunogold labeling of plasma membrane sheets. Although various signaling molecules have been seen to segregate within plasma membrane microdomains, the biochemical identity of these microdomains and the factors affecting their formation are largely unknown. Lipid rafts are envisioned as submicron membrane subdomains of liquid ordered structure with differing lipid and protein constituents that define their specific varieties. To facilitate EM investigation of inner leaflet lipid rafts and the localization of membrane proteins therein, a unique genetically encoded reporter with the dually acylated raft-targeting motif of the Lck kinase was developed. This reporter, designated Lck-BAP-GFP, incorporates green fluorescent protein (GFP) and biotin acceptor peptide (BAP) modules, with the latter allowing its single-step labeling with streptavidin-gold. Lck-BAP-GFP was metabolically biotinylated in mammalian cells, distributed into low-density detergent-resistant membrane fractions, and was readily detected with avidin-based reagents. In EM images of plasma membrane sheets, the streptavidin-gold-labeled reporter was clustered in 20–50 nm microdomains, presumably representative of inner leaflet lipid rafts. The utility of the reporter was demonstrated in an investigation of the potential lipid raft localization of the epidermal growth factor receptor. PMID:22822037

  4. Preparation and biological evaluation of an astatine-211 labeled biotin conjugate: Biotinyl-3-[211At]astatoanilide

    International Nuclear Information System (INIS)

    Foulon, Catherine F.; Schoultz, Bent W.; Zalutsky, Michael R.

    1997-01-01

    Biotinyl-3-[ 211 At]astatoanilide ([ 211 At]AtBA) was prepared in more than 80% yield by destannylation. In vitro, [ 211 At]AtBA exhibited a high affinity for streptavidin, and was stable after incubation in human serum, cerebrospinal fluid and distilled water, whereas it was rapidly degraded in mouse serum. HPLC analysis showed that the main degradation pathway in mouse serum was the cleavage of [ 211 At]astatoaniline. In mice, [ 211 At]AtBA and its 125 I-labeled analogue cleared rapidly from most tissues; however, there was some evidence for dehalogenation of both tracers

  5. Structure-Guided Design of an Engineered Streptavidin with Reusability to Purify Streptavidin-Binding Peptide Tagged Proteins or Biotinylated Proteins

    OpenAIRE

    Wu, Sau-Ching; Wong, Sui-Lam

    2013-01-01

    Development of a high-affinity streptavidin-binding peptide (SBP) tag allows the tagged recombinant proteins to be affinity purified using the streptavidin matrix without the need of biotinylation. The major limitation of this powerful technology is the requirement to use biotin to elute the SBP-tagged proteins from the streptavidin matrix. Tight biotin binding by streptavidin essentially allows the matrix to be used only once. To address this problem, differences in interactions of biotin an...

  6. Quantification of alpha-tubulin isotypes by sandwich ELISA with signal amplification through biotinyl-tyramide or immuno-PCR

    Czech Academy of Sciences Publication Activity Database

    Dráberová, Eduarda; Stegurová, Lucie; Sulimenko, Vadym; Hájková, Zuzana; Dráber, Petr; Dráber, Pavel

    2013-01-01

    Roč. 395, 1-2 (2013), s. 63-70 ISSN 0022-1759 R&D Projects: GA AV ČR KAN200520701; GA ČR GAP302/12/1673; GA ČR GPP302/11/P709; GA ČR GAP302/10/1759; GA ČR GA301/09/1826; GA MŠk(CZ) LD13015; GA MŠk LD12073 Institutional support: RVO:68378050 Keywords : alpha-tubulin isotypes * biotinyl-tyramide * ELISA * immuno-PCR * mast cells Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.005, year: 2013

  7. New aspects of galectin functionality in nuclei of cultured bone marrow stromal and epidermal cells: biotinylated galectins as tool to detect specific bindong sites

    Czech Academy of Sciences Publication Activity Database

    Purkrábková, T.; Smetana Jr., K.; Dvořánková, B.; Holíková, Z.; Böck, C.; Lensch, M.; André, S.; Pytlík, R.; Liu, F.; Klíma, Jiří; Smetana, K.; Motlík, Jan; Gabius, H. J.

    2003-01-01

    Roč. 95, - (2003), s. 535-545 ISSN 0248-4900 R&D Projects: GA MŠk LN00A065 Institutional research plan: CEZ:AV0Z5045916 Keywords : biotinylation * bone marrow stromal cells * galectin Subject RIV: ED - Physiology Impact factor: 2.159, year: 2003

  8. Combining magnetic nanoparticle with biotinylated nanobodies for rapid and sensitive detection of influenza H3N2

    Science.gov (United States)

    Zhu, Min; Hu, Yonghong; Li, Guirong; Ou, Weijun; Mao, Panyong; Xin, Shaojie; Wan, Yakun

    2014-09-01

    Our objective is to develop a rapid and sensitive assay based on magnetic beads to detect the concentration of influenza H3N2. The possibility of using variable domain heavy-chain antibodies (nanobody) as diagnostic tools for influenza H3N2 was investigated. A healthy camel was immunized with inactivated influenza H3N2. A nanobody library of 8 × 108 clones was constructed and phage displayed. After three successive biopanning steps, H3N2-specific nanobodies were successfully isolated, expressed in Escherichia coli, and purified. Sequence analysis of the nanobodies revealed that we possessed four classes of nanobodies against H3N2. Two nanobodies were further used to prepare our rapid diagnostic kit. Biotinylated nanobody was effectively immobilized onto the surface of streptavidin magnetic beads. The modified magnetic beads with nanobody capture specifically influenza H3N2 and can still be recognized by nanobodies conjugated to horseradish peroxidase (HRP) conjugates. Under optimized conditions, the present immunoassay exhibited a relatively high sensitive detection with a limit of 50 ng/mL. In conclusion, by combining magnetic beads with specific nanobodies, this assay provides a promising influenza detection assay to develop a potential rapid, sensitive, and low-cost diagnostic tool to screen for influenza infections.

  9. Synthesis and in Vitro evaluation of ''1''8''8Re-biotinyl-hydrazino-etda

    International Nuclear Information System (INIS)

    Pervez, S; Mushtaq, A.; Jehangir, M.

    2002-01-01

    Pretargeting strategies have overcome many drawbacks associated with the use of directly labelled MoAbs in the diagnosis / treatment of various solid tumors. In particular the avidin-biotin system has received much attention due to extremely high affinity between avidin and biotin. An EDTA derivative of biotin has been synthesized (yield-35%). In order or label biotin derivative (biotinyl-hydrazino-EDTA) , stannous ion was used to reduce ''1''8''8ReO 4 (VII) to lower oxidation state and weak chelating agent glucoheptonate as stabilizer and trans chelating agent. Thin layer chromatography and high performance liquid chromatography techniques were employed to monitor the radiolabeling efficiency. The radiolabeling yield of ''1''8''8Re-EDTA B1 was >95%. The radiolabeled product was found to bind to avidin (70-80%), thereby demonstrating retention of its biological integrity

  10. Reaction of [3H]-taurine maleimide with platelet surface thiols

    International Nuclear Information System (INIS)

    Karl, D.W.; Mills, D.C.B.

    1986-01-01

    Taurine Maleimide (2-maleimidoethanesulfonate, TM) was synthesized from [2- 3 H]-taurine and methoxycarbonylmaleimide (MCM). The yield of a 1 μmol synthesis approached 100% (based on taurine) when MCM was used in 4-fold excess. The product (TM*) was purified by ion exchange chromatography. TM* reacted irreversibly with thiol groups on the surface of washed human platelets, leading to incorporation of radioactivity into platelet pellets. Incorporation was blocked by cysteine, mercuribenzenesulfonate (MBS), dithiobisnitrobenzoate, and N-ethylmaleimide, but not by taurine or by inhibitors of anion transport. Reaction of TM* with platelets showed the dependence on time and concentration characteristics of a bimolecular reaction. The number of reactive sites ranged from 1 to 5 x 10 5 /platelet, and the apparent rate constant from 1 to 3 x 10 3 /(M x min). TM was less effective than MBS as an inhibitor of platelet aggregation induced by several agents. TM had no effect on the uptake of serotonin, taurine, or phosphate by the platelets, processes which are sensitive to MBS. These differences, considered with the similarity in size and charge of TM and MBS, suggest that classes of thiols defined as exofacial by their accessibility to MBS can differ substantially in their reactivity with other impermeant reagents

  11. Nucleolin: acharan sulfate–binding protein on the surface of cancer cells

    Science.gov (United States)

    Joo, Eun Ji; ten Dam, Gerdy B.; van Kuppevelt, Toin H.; Toida, Toshihiko; Linhardt, Robert J.; Kim, Yeong Shik

    2005-01-01

    Glycosaminoglycans (GAGs) are complex polysaccharides that participate in the regulation of physiological processes through the interactions with a wide variety of proteins. Acharan sulfate (AS), isolated from the giant African snail Achatina fulica, primarily consists of the repeating disaccharide structure α-D-N-acetylglucosaminyl (1→4) 2-sulfoiduronic acid. Exogenous AS was injected subcutaneously near the tumor tissue in C57BL/6 mice that had been implanted with Lewis lung carcinoma cells (LLCs). The location of AS in the tumor was assessed by staining of sectioned tissues with alcian blue and periodic acid–Schiff (PAS) reagent. In vitro assays indicated binding of cells to 50 μg/ml AS (or heparin) after a 5-h incubation. Immunofluorescence assays, using anti-AS antibody, detected AS at the cell surface. The outer-surface of LLCs were next biotinylated to identify the AS-binding proteins. Biotinylated cells were lysed, and the lysates were fractionated on the AS affinity column using a stepwise salt gradient (0, 0.1, 0.3, 0.5, 0.7, 1.0, and 2.0 M). The fractions were analyzed by SDS–PAGE with silver staining and western blotting. We focused on the proteins with high affinity for AS (eluting at 1 M NaCl) and detected only two bands by western blotting. ESI Q-TOF MS analysis of one of these bands, molecular weight ~110 kDa, showed it to be nucleolin. A phosphorylated form of nucleolin on the surface of cells acts as a cell surface receptor for a variety of ligands, including growth factors (i.e., basic fibroblast growth factor) and chemokines (i.e., midkine). These results show that nucleolin is one of several AS-binding proteins and suggest that AS might demonstrate its tumor growth inhibitory activity by binding the nucleolin receptor protein on the surface of cancer cells. PMID:15329357

  12. Chemical modifications of Au/SiO2 template substrates for patterned biofunctional surfaces.

    Science.gov (United States)

    Briand, Elisabeth; Humblot, Vincent; Landoulsi, Jessem; Petronis, Sarunas; Pradier, Claire-Marie; Kasemo, Bengt; Svedhem, Sofia

    2011-01-18

    The aim of this work was to create patterned surfaces for localized and specific biochemical recognition. For this purpose, we have developed a protocol for orthogonal and material-selective surface modifications of microfabricated patterned surfaces composed of SiO(2) areas (100 μm diameter) surrounded by Au. The SiO(2) spots were chemically modified by a sequence of reactions (silanization using an amine-terminated silane (APTES), followed by amine coupling of a biotin analogue and biospecific recognition) to achieve efficient immobilization of streptavidin in a functional form. The surrounding Au was rendered inert to protein adsorption by modification by HS(CH(2))(10)CONH(CH(2))(2)(OCH(2)CH(2))(7)OH (thiol-OEG). The surface modification protocol was developed by testing separately homogeneous SiO(2) and Au surfaces, to obtain the two following results: (i) SiO(2) surfaces which allowed the grafting of streptavidin, and subsequent immobilization of biotinylated antibodies, and (ii) Au surfaces showing almost no affinity for the same streptavidin and antibody solutions. The surface interactions were monitored by quartz crystal microbalance with dissipation monitoring (QCM-D), and chemical analyses were performed by polarization modulation-reflexion absorption infrared spectroscopy (PM-RAIRS) and X-ray photoelectron spectroscopy (XPS) to assess the validity of the initial orthogonal assembly of APTES and thiol-OEG. Eventually, microscopy imaging of the modified Au/SiO(2) patterned substrates validated the specific binding of streptavidin on the SiO(2)/APTES areas, as well as the subsequent binding of biotinylated anti-rIgG and further detection of fluorescent rIgG on the functionalized SiO(2) areas. These results demonstrate a successful protocol for the preparation of patterned biofunctional surfaces, based on microfabricated Au/SiO(2) templates and supported by careful surface analysis. The strong immobilization of the biomolecules resulting from the described

  13. Ultrasensitive Detection of Single-Walled Carbon Nanotubes Using Surface Plasmon Resonance.

    Science.gov (United States)

    Jang, Daeho; Na, Wonhwi; Kang, Minwook; Kim, Namjoon; Shin, Sehyun

    2016-01-05

    Because single-walled carbon nanotubes (SWNTs) are known to be a potentially dangerous material, inducing cancers and other diseases, any possible leakage of SWNTs through an aquatic medium such as drinking water will result in a major public threat. To solve this problem, for the present study, a highly sensitive, quantitative detection method of SWNTs in an aqueous solution was developed using surface plasmon resonance (SPR) spectroscopy. For a highly sensitive and specific detection, a strong affinity conjugation with biotin-streptavidin was adopted on an SPR sensing mechanism. During the pretreatment process, the SWNT surface was functionalized and hydrophilized using a thymine-chain based biotinylated single-strand DNA linker (B-ssDNA) and bovine serum albumin (BSA). The pretreated SWNTs were captured on a sensing film, the surface of which was immobilized with streptavidin on biotinylated gold film. The captured SWNTs were measured in real-time using SPR spectroscopy. Specific binding with SWNTs was verified through several validation experiments. The present method using an SPR sensor is capable of detecting SWNTs of as low as 100 fg/mL, which is the lowest level reported thus far for carbon-nanotube detection. In addition, the SPR sensor showed a linear characteristic within the range of 100 pg/mL to 200 ng/mL. These findings imply that the present SPR sensing method can detect an extremely low level of SWNTs in an aquatic environment with high sensitivity and high specificity, and thus any potential leakage of SWNTs into an aquatic environment can be precisely monitored within a couple of hours.

  14. Immobilization of oligonucleotide probes on silicon surfaces using biotin–streptavidin system examined with microscopic and spectroscopic techniques

    Energy Technology Data Exchange (ETDEWEB)

    Awsiuk, K., E-mail: kamil.awsiuk@uj.edu.pl [M. Smoluchowski Institute of Physics, Jagiellonian University, Reymonta 4, Kraków 30-059 (Poland); Rysz, J. [M. Smoluchowski Institute of Physics, Jagiellonian University, Reymonta 4, Kraków 30-059 (Poland); Petrou, P. [Institute of Nuclear and Radiological Sciences and Technology, Energy and Safety, NCSR “Demokritos”, End Patriarchou Gregoriou Str., Aghia Paraskevi 15310 (Greece); Budkowski, A. [M. Smoluchowski Institute of Physics, Jagiellonian University, Reymonta 4, Kraków 30-059 (Poland); Bernasik, A. [Faculty of Physics and Applied Computer Science, AGH-University of Science and Technology, Al. Mickiewicza 30, Kraków 30-059 (Poland); Kakabakos, S. [Institute of Nuclear and Radiological Sciences and Technology, Energy and Safety, NCSR “Demokritos”, End Patriarchou Gregoriou Str., Aghia Paraskevi 15310 (Greece); Marzec, M.M. [Faculty of Physics and Applied Computer Science, AGH-University of Science and Technology, Al. Mickiewicza 30, Kraków 30-059 (Poland); Raptis, I. [Institute for Advanced Materials, Physicochemical Processes, Nanotechnology and Microsystems, NCSR “Demokritos”, End Patriarchou Gregoriou Str., Aghia Paraskevi 15310 (Greece)

    2014-01-30

    To immobilize effectively oligonucleotide probes on SiO{sub 2} modified with (3-aminopropyl)triethoxysilane, four procedures based on streptavidin–biotin system are compared with Atomic Force Microscopy, Angle-Resolved X-ray Photoelectron Spectroscopy and Time-of-Flight Secondary Ion Mass Spectrometry. The first approach involves: adsorption of biotinylated Bovine Serum Albumin, blocking free surface sites with BSA, binding of streptavidin and biotinylated oligonucleotide (b-oligo). Final steps are exchanged in the second procedure with immobilization of preformed streptavidin–b-oligo conjugate. The third approach consists of streptavidin adsorption, blocking with BSA and b-oligo binding. Finally, streptavidin–b-oligo conjugate is immobilized directly within the fourth method. Surface coverage with biomolecules, determined from ARXPS, accords with average AFM height, and is anti-correlated with the intensity of Si+ ions. Higher biomolecular coverage was achieved during the last steps of the first (2.45(±0.38) mg/m{sup 2}) and second (1.31(±0.22) mg/m{sup 2}) approach, as compared to lower surface density resulting from the third (0.58(±0.20) mg/m{sup 2}) and fourth (0.41(±0.11) mg/m{sup 2}) method. Phosphorus atomic concentration indicates effectiveness of oligonucleotide immobilization. Secondary ions intensities, characteristic for oligonucleotides, streptavidin, BSA, and proteins, allow additional insight into overlayer composition. These measurements verify the ARXPS results and show the superiority of the first two immobilization approaches in terms of streptavidin and oligonucleotide density achieved onto the surface.

  15. Immobilization of oligonucleotide probes on silicon surfaces using biotin–streptavidin system examined with microscopic and spectroscopic techniques

    International Nuclear Information System (INIS)

    Awsiuk, K.; Rysz, J.; Petrou, P.; Budkowski, A.; Bernasik, A.; Kakabakos, S.; Marzec, M.M.; Raptis, I.

    2014-01-01

    To immobilize effectively oligonucleotide probes on SiO 2 modified with (3-aminopropyl)triethoxysilane, four procedures based on streptavidin–biotin system are compared with Atomic Force Microscopy, Angle-Resolved X-ray Photoelectron Spectroscopy and Time-of-Flight Secondary Ion Mass Spectrometry. The first approach involves: adsorption of biotinylated Bovine Serum Albumin, blocking free surface sites with BSA, binding of streptavidin and biotinylated oligonucleotide (b-oligo). Final steps are exchanged in the second procedure with immobilization of preformed streptavidin–b-oligo conjugate. The third approach consists of streptavidin adsorption, blocking with BSA and b-oligo binding. Finally, streptavidin–b-oligo conjugate is immobilized directly within the fourth method. Surface coverage with biomolecules, determined from ARXPS, accords with average AFM height, and is anti-correlated with the intensity of Si+ ions. Higher biomolecular coverage was achieved during the last steps of the first (2.45(±0.38) mg/m 2 ) and second (1.31(±0.22) mg/m 2 ) approach, as compared to lower surface density resulting from the third (0.58(±0.20) mg/m 2 ) and fourth (0.41(±0.11) mg/m 2 ) method. Phosphorus atomic concentration indicates effectiveness of oligonucleotide immobilization. Secondary ions intensities, characteristic for oligonucleotides, streptavidin, BSA, and proteins, allow additional insight into overlayer composition. These measurements verify the ARXPS results and show the superiority of the first two immobilization approaches in terms of streptavidin and oligonucleotide density achieved onto the surface.

  16. Targeted DNA delivery to cancer cells using a biotinylated chitosan carrier.

    Science.gov (United States)

    Darvishi, Mohammad H; Nomani, Alireza; Hashemzadeh, Hadi; Amini, Mohsen; Shokrgozar, Mohammad A; Dinarvand, Rassoul

    2017-05-01

    A novel biotinylated chitosan-graft-polyethyleneimine (Bio-Chi-g-PEI) copolymer was synthesized and evaluated as a nonviral gene delivery carrier for improvement of the transfection efficiency, endosomal escape, and targeted gene delivery of a plasmid encoding green fluorescent protein N1 (pEGFP-N1) into two different biotin-overexpressing cell lines including HeLa and OVCAR-3 cells. The structure of the obtained copolymers was confirmed by 1 H nuclear magnetic resonance ( 1 H NMR) and Fourier transform infrared spectroscopy. Physicochemical properties of the Bio-Chi-g-PEI/plasmid DNA (pDNA) complexes such as complex stability, size, zeta potential, and their morphology were investigated at various weight ratios of copolymer to pDNA. Bio-Chi-g-PEI copolymers could effectively condense pDNA into small particles with average diameters less than 164 nm and the zeta potential of +34.8 mV at the N/P ratio of 40/1. As revealed by flow cytometry, Bio-Chi-g-PEI/pDNA complexes had lower cytotoxicity than that of PEI 25 kDa/pDNA complexes in both cell lines. In vitro experiments revealed that the Bio-Chi-gPEI/pDNA complexes not only had much lower cytotoxicity, but also displayed higher transfection efficiency than that of PEI 25kDa/pDNA complexes. High percentage of cancer cells was successfully transfected by Bio-Chi-g-PEI/pDNA and properly expressed GFP protein. This study indicates that this copolymer complex can be a promising gene delivery carrier. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

  17. Country report: Poland. 1. Determination of 90Sr in 90YCL3 solution using DGA and SR-Spec resins (extraction chromatography) and by TLC according to USP. 2. Preliminary investigation on antibody biotinylation. 3. The development of method for preparation of human albumin microspheres as potential radionuclide carriers for diagnostic and therapeutic use. 4. Preparation of radiocolloids for radiosynovectomy using radionuclides of various beta energies

    International Nuclear Information System (INIS)

    Pawlak, D.; Mikolajczak, R.; Korsak, A.; Dziel, T.; Parus, J.L.; Byszewska-Szpocinska, E.; Jakubowska, E.; Mikolajczak, R.; Jaroń, A.; Pijarowska, J.; Iller, E.; Mikolajczak, W.

    2010-01-01

    1. • It seems that strontium can be more efficiently eluted for the DGA resin than form the Sr-spec resin. Paper chromatography (USP) method provides good resolution of Y and Sr, more results are needed to evaluate results statistically. Comparison of paper chromatography (USP) with paper extraction chromatography was not possible due to the problems in purchasing of the KSM-17. Further works in order to determine detection and determination limits of each of proposed methods are planned. 2. • The preliminary investigation on antibody biotinylation has been performed using commercially available polyclonal human immunoglobulin IgG, mw 150 kDa (Biomed Sp.z o.o, Lublin). Sulfo-NHS –Biotin (Pierce Biotechnology) was used as biotinylating agent. Activated biotin as sulfosuccinimidyl-6-[biotin-amido]hexanoate. is one of the commonly used reagent for proteins biotinylation. Sulfo-NHS-biotin effectively reacts at pH 7-9 with primary amino groups (- NH2) of proteins provided by lysine residues and Nterminus of polypeptide, forming stable amide bond. This reagent dissolves well in water. 3. • The average production yield of HAM for receiving the desired size range between 10-32 μm amounts to 84% and the mean size of particles was estimated to about 15 μm. Optical micrographs show microspheres as very regular spherical forms with quite smooth surfaces and rather narrow spread of size in the selected range. The labelling yields determined as relation of measured radioactivity remaining in the membrane filter to the total radioactivity were > 95% for 99mTc-HAM complex and > 98% for 90Y–X-HAM and 177Lu-X-HAM complexes. Labelled modified microspheres showed high in vitro stability in human plasma with only 4-5% loss of radioactivity from the surface after 48 h of incubation. 4. • The aim of the project is synthesis of monodispersive particles of yttrium citrate and characterization of its surface with instrumental techniques (collaboration with Maria Curie

  18. Structure-guided design of an engineered streptavidin with reusability to purify streptavidin-binding peptide tagged proteins or biotinylated proteins.

    Directory of Open Access Journals (Sweden)

    Sau-Ching Wu

    Full Text Available Development of a high-affinity streptavidin-binding peptide (SBP tag allows the tagged recombinant proteins to be affinity purified using the streptavidin matrix without the need of biotinylation. The major limitation of this powerful technology is the requirement to use biotin to elute the SBP-tagged proteins from the streptavidin matrix. Tight biotin binding by streptavidin essentially allows the matrix to be used only once. To address this problem, differences in interactions of biotin and SBP with streptavidin were explored. Loop3-4 which serves as a mobile lid for the biotin binding pocket in streptavidin is in the closed state with biotin binding. In contrast, this loop is in the open state with SBP binding. Replacement of glycine-48 with a bulkier residue (threonine in this loop selectively reduces the biotin binding affinity (Kd from 4 × 10(-14 M to 4.45 × 10(-10 M without affecting the SBP binding affinity. Introduction of a second mutation (S27A to the first mutein (G48T results in the development of a novel engineered streptavidin SAVSBPM18 which could be recombinantly produced in the functional form from Bacillus subtilis via secretion. To form an intact binding pocket for tight binding of SBP, two diagonally oriented subunits in a tetrameric streptavidin are required. It is vital for SAVSBPM18 to be stably in the tetrameric state in solution. This was confirmed using an HPLC/Laser light scattering system. SAVSBPM18 retains high binding affinity to SBP but has reversible biotin binding capability. The SAVSBPM18 matrix can be applied to affinity purify SBP-tagged proteins or biotinylated molecules to homogeneity with high recovery in a reusable manner. A mild washing step is sufficient to regenerate the matrix which can be reused for multiple rounds. Other applications including development of automated protein purification systems, lab-on-a-chip micro-devices, reusable biosensors, bioreactors and microarrays, and strippable

  19. Structure-guided design of an engineered streptavidin with reusability to purify streptavidin-binding peptide tagged proteins or biotinylated proteins.

    Science.gov (United States)

    Wu, Sau-Ching; Wong, Sui-Lam

    2013-01-01

    Development of a high-affinity streptavidin-binding peptide (SBP) tag allows the tagged recombinant proteins to be affinity purified using the streptavidin matrix without the need of biotinylation. The major limitation of this powerful technology is the requirement to use biotin to elute the SBP-tagged proteins from the streptavidin matrix. Tight biotin binding by streptavidin essentially allows the matrix to be used only once. To address this problem, differences in interactions of biotin and SBP with streptavidin were explored. Loop3-4 which serves as a mobile lid for the biotin binding pocket in streptavidin is in the closed state with biotin binding. In contrast, this loop is in the open state with SBP binding. Replacement of glycine-48 with a bulkier residue (threonine) in this loop selectively reduces the biotin binding affinity (Kd) from 4 × 10(-14) M to 4.45 × 10(-10) M without affecting the SBP binding affinity. Introduction of a second mutation (S27A) to the first mutein (G48T) results in the development of a novel engineered streptavidin SAVSBPM18 which could be recombinantly produced in the functional form from Bacillus subtilis via secretion. To form an intact binding pocket for tight binding of SBP, two diagonally oriented subunits in a tetrameric streptavidin are required. It is vital for SAVSBPM18 to be stably in the tetrameric state in solution. This was confirmed using an HPLC/Laser light scattering system. SAVSBPM18 retains high binding affinity to SBP but has reversible biotin binding capability. The SAVSBPM18 matrix can be applied to affinity purify SBP-tagged proteins or biotinylated molecules to homogeneity with high recovery in a reusable manner. A mild washing step is sufficient to regenerate the matrix which can be reused for multiple rounds. Other applications including development of automated protein purification systems, lab-on-a-chip micro-devices, reusable biosensors, bioreactors and microarrays, and strippable detection agents for

  20. Identification of Litopenaeus vannamei BiP as a novel cellular attachment protein for white spot syndrome virus by using a biotinylation based affinity chromatography method.

    Science.gov (United States)

    Yuan, Zengzhi; Chen, Meng; Wang, Jingting; Li, Zhuoyu; Geng, Xuyun; Sun, Jinsheng

    2018-05-05

    White spot syndrome virus (WSSV) is a dangerous threat to shrimp farming that also attacks a wide range of crustaceans. Knowledge of the surface protein-protein interactions between the pathogen and host is very crucial to unraveling the molecular pathogenesis mechanisms of WSSV. In this study, LvBiP (Litopenaeus vannamei immunoglobulin heavy-chain-binding protein) was identified as a novel WSSV binding protein of L. vannamei by a biotinylation based affinity chromatography method. By using pull-down and ELISA assays, the binding of recombinant LvBiP to WSSV was proved to be specific and ATP- dependent. The interaction was also confirmed by the result of co-immunoprecipitation assay. Immunofluorescence studies revealed the co-localization of LvBiP with WSSV on the cell surface of shrimp haemocytes. Additionally, LvBiP is likely to play an important role in WSSV infection. Treatment of gill cellular membrane proteins (CMPs) with purified rLvBiP and antibody that specifically recognizes LvBiP, led to a significant reduction in the binding of WSSV to gill CMPs. In the in vivo neutralization assay, rLvBiP and anti-LvBiP polyclonal antibody partially blocked the infection of WSSV. Taken together, the results indicate that LvBiP, a molecular chaperon of the HSP70 family, is a novel host factor involved at the step of attachment of the WSSV to the host cells and a potential candidate of therapeutic target. Copyright © 2018. Published by Elsevier Ltd.

  1. Country report: Poland. 1. Determination of {sup 90}Sr in {sup 90}YCL{sub 3} solution using DGA and SR-Spec resins (extraction chromatography) and by TLC according to USP. 2. Preliminary investigation on antibody biotinylation. 3. The development of method for preparation of human albumin microspheres as potential radionuclide carriers for diagnostic and therapeutic use. 4. Preparation of radiocolloids for radiosynovectomy using radionuclides of various beta energies

    Energy Technology Data Exchange (ETDEWEB)

    Pawlak, D.; Mikolajczak, R.; Korsak, A.; Dziel, T.; Parus, J. L.; Byszewska-Szpocinska, E.; Jakubowska, E.; Mikolajczak, R.; Jaroń, A.; Pijarowska, J.; Iller, E.; Mikolajczak, W.

    2010-07-01

    1. • It seems that strontium can be more efficiently eluted for the DGA resin than form the Sr-spec resin. Paper chromatography (USP) method provides good resolution of Y and Sr, more results are needed to evaluate results statistically. Comparison of paper chromatography (USP) with paper extraction chromatography was not possible due to the problems in purchasing of the KSM-17. Further works in order to determine detection and determination limits of each of proposed methods are planned. 2. • The preliminary investigation on antibody biotinylation has been performed using commercially available polyclonal human immunoglobulin IgG, mw 150 kDa (Biomed Sp.z o.o, Lublin). Sulfo-NHS –Biotin (Pierce Biotechnology) was used as biotinylating agent. Activated biotin as sulfosuccinimidyl-6-[biotin-amido]hexanoate. is one of the commonly used reagent for proteins biotinylation. Sulfo-NHS-biotin effectively reacts at pH 7-9 with primary amino groups (- NH2) of proteins provided by lysine residues and Nterminus of polypeptide, forming stable amide bond. This reagent dissolves well in water. 3. • The average production yield of HAM for receiving the desired size range between 10-32 μm amounts to 84% and the mean size of particles was estimated to about 15 μm. Optical micrographs show microspheres as very regular spherical forms with quite smooth surfaces and rather narrow spread of size in the selected range. The labelling yields determined as relation of measured radioactivity remaining in the membrane filter to the total radioactivity were > 95% for 99mTc-HAM complex and > 98% for 90Y–X-HAM and 177Lu-X-HAM complexes. Labelled modified microspheres showed high in vitro stability in human plasma with only 4-5% loss of radioactivity from the surface after 48 h of incubation. 4. • The aim of the project is synthesis of monodispersive particles of yttrium citrate and characterization of its surface with instrumental techniques (collaboration with Maria Curie

  2. Modern trends in radioimmunotherapy of cancer. Pre targeting strategies for the treatment of ovarian cancer

    International Nuclear Information System (INIS)

    Mcquarrie, S.A.; Xiao, Z.; Mercer, J. R.; Suresh, M. R.

    2001-01-01

    A review of published data on some of the problems associated in treating cancer using radioimmunotherapy is presented. Potential improvements for this type of therapy using pretargeting strategies are discussed and preliminary results on a novel multistep regimen to treat human ovarian cancer are presented. A pretargeting strategy using ovarian cancer are presented. A pretargeting strategy using a biotinylated, anti-CA 125 monoclonal antibody (MAb) to attract biotinylated long-circulating liposomes to the surface of CA 125-expressing ovarian cancer cells, was employed. Confocal laser scanning microscopy and fluorescent labels were used to establish the biodistribution patterns in NIH:OVCAR-3 (CA-125 positive) and SK-OV-3 (CA-125 negative) human ovarian cancer cells. Shedding kinetics of the pretargeted stage were measured using 125 I labeled MAbs. No significant internalization of the MAb used in the pretargeting step was observed by 4 hrs. The antibody was gradually internalized starting at 6 hrs, and most of the labelled MAb was detected in cytoplasm by 24 hrs. Shedding and exocytosis of the antigen-MAb complex was not significant for up to 6-hours following administration of the iodinated MAb. Biotinylated liposomes were shown to specifically target the biotinylated MAb/streptavidin complex on the cell surface. It has been demonstrated that by a three-step pretargeting approach, biotinylated liposomes can be specifically delivered to cells pretargeted with biotinylated MAb/SAv complex. The slow internalization and shedding properties of the two MAbs are ideal for multistep pretargeting methods. A successful multistep linkage was established with the biotinylated MAb B27.1, streptavidin and biotinylated liposomes to OVCAR-3 cells, but not to SK-OV-3 cells

  3. Utilizing Biotinylated Proteins Expressed in Yeast to Visualize DNA–Protein Interactions at the Single-Molecule Level

    Directory of Open Access Journals (Sweden)

    Huijun Xue

    2017-10-01

    Full Text Available Much of our knowledge in conventional biochemistry has derived from bulk assays. However, many stochastic processes and transient intermediates are hidden when averaged over the ensemble. The powerful technique of single-molecule fluorescence microscopy has made great contributions to the understanding of life processes that are inaccessible when using traditional approaches. In single-molecule studies, quantum dots (Qdots have several unique advantages over other fluorescent probes, such as high brightness, extremely high photostability, and large Stokes shift, thus allowing long-time observation and improved signal-to-noise ratios. So far, however, there is no convenient way to label proteins purified from budding yeast with Qdots. Based on BirA–Avi and biotin–streptavidin systems, we have established a simple method to acquire a Qdot-labeled protein and visualize its interaction with DNA using total internal reflection fluorescence microscopy. For proof-of-concept, we chose replication protein A (RPA and origin recognition complex (ORC as the proteins of interest. Proteins were purified from budding yeast with high biotinylation efficiency and rapidly labeled with streptavidin-coated Qdots. Interactions between proteins and DNA were observed successfully at the single-molecule level.

  4. The cell-based L-glutathione protection assays to study endocytosis and recycling of plasma membrane proteins.

    Science.gov (United States)

    Cihil, Kristine M; Swiatecka-Urban, Agnieszka

    2013-12-13

    Membrane trafficking involves transport of proteins from the plasma membrane to the cell interior (i.e. endocytosis) followed by trafficking to lysosomes for degradation or to the plasma membrane for recycling. The cell based L-glutathione protection assays can be used to study endocytosis and recycling of protein receptors, channels, transporters, and adhesion molecules localized at the cell surface. The endocytic assay requires labeling of cell surface proteins with a cell membrane impermeable biotin containing a disulfide bond and the N-hydroxysuccinimide (NHS) ester at 4 ºC - a temperature at which membrane trafficking does not occur. Endocytosis of biotinylated plasma membrane proteins is induced by incubation at 37 ºC. Next, the temperature is decreased again to 4 ºC to stop endocytic trafficking and the disulfide bond in biotin covalently attached to proteins that have remained at the plasma membrane is reduced with L-glutathione. At this point, only proteins that were endocytosed remain protected from L-glutathione and thus remain biotinylated. After cell lysis, biotinylated proteins are isolated with streptavidin agarose, eluted from agarose, and the biotinylated protein of interest is detected by western blotting. During the recycling assay, after biotinylation cells are incubated at 37 °C to load endocytic vesicles with biotinylated proteins and the disulfide bond in biotin covalently attached to proteins remaining at the plasma membrane is reduced with L-glutathione at 4 ºC as in the endocytic assay. Next, cells are incubated again at 37 °C to allow biotinylated proteins from endocytic vesicles to recycle to the plasma membrane. Cells are then incubated at 4 ºC, and the disulfide bond in biotin attached to proteins that recycled to the plasma membranes is reduced with L-glutathione. The biotinylated proteins protected from L-glutathione are those that did not recycle to the plasma membrane.

  5. Gold island films on indium tin oxide for localized surface plasmon sensing

    International Nuclear Information System (INIS)

    Szunerits, Sabine; Praig, Vera G; Manesse, Mael; Boukherroub, Rabah

    2008-01-01

    Mechanically, chemically and optically stable gold island films were prepared on indium tin oxide (ITO) substrates by direct thermal evaporation of thin gold films (2-6 nm) without the need for pre- or post-coating. The effect of mild thermal annealing (150 deg. C, 12 h) or short high temperature annealing (500 deg. C, 1 min) on the morphology of the gold nanostructures was investigated. ITO covered with 2 nm gold nanoislands and annealed at 500 deg. C for 1 min was investigated for its ability to detect the adsorption of biotinylated bovine serum albumin using local surface plasmon resonance (LSPR), and its subsequent molecular recognition of avidin

  6. Osseoconductivity of a Specific Streptavidin-Biotin-Fibronectin Surface Coating of Biotinylated Titanium Implants - A Rabbit Animal Study.

    Science.gov (United States)

    Kämmerer, Peer W; Lehnert, Michael; Al-Nawas, Bilal; Kumar, Vinay V; Hagmann, Sebastien; Alshihri, Abdulmonem; Frerich, Bernhard; Veith, Michael

    2015-10-01

    Biofunctionalized implant surfaces may accelerate bony integration and increase long-term stability. The aim of the study was to evaluate the osseous reaction toward biomimetic titanium implants surfaces coated with quasicovalent immobilized fibronectin in an in vivo animal model. A total of 84 implants (uncoated [control 1, n = 36], streptavidin-biotin coated [test 1, n = 24], streptavidin-biotin-fibronectin coated [test 2, n = 24]) were inserted 1 mm supracortically in the proximal tibia of 12 rabbits. The samples were examined after 3 and 6 weeks. Total bone-implant contact (tBIC; %), bone-implant contact in the cortical (cBIC; %) and in the spongious bone (sBIC; %) as well as the percentage of linear bone fill (PLF; %) were evaluated. After 3 weeks, streptavidin-biotin-fibronectin implants had a significant higher sBIC (p = .043) and PLF (p = .007) compared with the uncoated samples. After 6 weeks, this difference was significant for tBIC (p = .016) and cBIC (p biotin-coated implants showed less bone growth at both time points of all examined parameters when compared with their counterparts (all p biotin-fibronectin system on smooth surface titanium shows a beneficial faster osseous healing in vivo. Besides, an antifouling effect of the streptavidin-biotin coating was proven. © 2015 Wiley Periodicals, Inc.

  7. Biomolecular strategies for cell surface engineering

    Science.gov (United States)

    Wilson, John Tanner

    backbone molecular weight, PEG chain length, and grafting ratio, PLL-g-PEG copolymers were rendered cytocompatible and used to initiate and propagate the growth of cell surface-supported PEM films. Planar characterization of this novel class of PEM films indicated that film thickness and composition may be tailored through appropriate control of layer number and copolymer properties. Furthermore, these investigations have helped establish a conceptual framework for the rational design of cell surface-supported thin films, with the objective of translating the diverse biomedical and biotechnological applications of PEM films to cellular interfaces. Important to the development of effective conformal islet coatings is an inherent strategy through which to incorporate bioactive molecules for directing desired biochemical or cellular responses. Towards this end, PLL-g-PEG copolymers functionalized with biotin, azide, and hydrazide moieties were synthesized and used, either alone or in combination, to capture streptavidin-, triphenylphosphine-, and aldehyde-labeled probes, respectively, on the islet surface. Additionally, PEM films assembled using alginate chemically modified to contain aldehyde groups could be used to introduce hydrazide-functionalized molecules to the islet surface. Hence, modified film constituents may be used as modular elements for controlling the chemical composition cell and tissue surfaces. Finally, we report a strategy for tethering thrombomodulin (TM) to the islet surface. Through site-specific, C-terminal biotinylation of TM and optimization of cell surface biotinylation, TM could be integrated with the islet surface. Re-engineering of islet surfaces with TM resulted in an increased catalytic capacity of islets to generate the powerful anti-inflammatory agent, activated protein C (APC), thereby providing a facile strategy for increasing the local concentration of APC at the site of transplantation.

  8. A simple and robust approach to immobilization of antibody fragments.

    Science.gov (United States)

    Ikonomova, Svetlana P; He, Ziming; Karlsson, Amy J

    2016-08-01

    Antibody fragments, such as the single-chain variable fragment (scFv), have much potential in research and diagnostics because of their antigen-binding ability similar to a full-sized antibody and their ease of production in microorganisms. Some applications of antibody fragments require immobilization on a surface, and we have established a simple immobilization method that is based on the biotin-streptavidin interaction and does not require a separate purification step. We genetically fused two biotinylation tags-the biotin carboxyl carrier protein (BCCP) or the AviTag minimal sequence-to six different scFvs (scFv13R4, scFvD10, scFv26-10, scFv3, scFv5, and scFv12) for site-specific biotinylation in vivo by endogenous biotin ligases produced by Escherichia coli. The biotinylated scFvs were immobilized onto streptavidin-coated plates directly from cell lysates, and immobilization was detected through enzyme-linked immunosorbent assays. All scFvs fusions were successfully immobilized, and scFvs biotinylated via the BCCP tag tended to immobilize better than those biotinylated via the AviTag, even when biotinylation efficiency was improved with the biotin ligase BirA. The ability of immobilized scFvs to bind antigens was confirmed using scFv13R4 and scFvD10 with their respective targets β-galactosidase and bacteriophage lambda head protein D (gpD). The immobilized scFv13R4 bound to β-galactosidase at the same level for both biotinylation tags when the surface was saturated with the scFv, and immobilized scFvs retained their functionality for at least 100days after immobilization. The simplicity and robustness of our method make it a promising approach for future applications that require antibody fragment immobilization. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Synthesis of Biotin Linkers with the Activated Triple Bond Donor [p-(N-propynoylaminotoluic Acid] (PATA for Efficient Biotinylation of Peptides and Oligonucleotides

    Directory of Open Access Journals (Sweden)

    Martina Jezowska

    2012-11-01

    Full Text Available Biotin is an important molecule for modern biological studies including, e.g., cellular transport. Its exclusive affinity to fluorescent streptavidin/avidin proteins allows ready and specific detection. As a consequence methods for the attachment of biotin to various biological targets are of high importance, especially when they are very selective and can also proceed in water. One useful method is Hüisgen dipolar [3+2]-cycloaddition, commonly referred to as “click chemistry”. As we reported recently, the activated triple bond donor p-(N-propynoylaminotoluic acid (PATA gives excellent results when used for conjugations at submicromolar concentrations. Thus, we have designed and synthesized two biotin linkers, with different lengths equipped with this activated triple bond donor and we proceeded with biotinylation of oligonucleotides and C-myc peptide both in solution and on solid support with excellent yields of conversion.

  10. Improving the Application of High Molecular Weight Biotinylated Dextran Amine for Thalamocortical Projection Tracing in the Rat.

    Science.gov (United States)

    Xu, Dongsheng; Cui, Jingjing; Wang, Jia; Zhang, Zhiyun; She, Chen; Bai, Wanzhu

    2018-04-12

    High molecular weight biotinylated dextran amine (BDA) has been used as a highly sensitive neuroanatomical tracer for many decades. Since the quality of its labeling was affected by various factors, here, we provide a refined protocol for the application of high molecular weight BDA for studying optimal neural labeling in the central nervous system. After stereotactic injection of BDA into the ventral posteromedial nucleus (VPM) of the thalamus in the rat through a delicate glass pipette, BDA was stained with fluorescent streptavidin-Alexa (AF) 594 and counterstained with fluorescent Nissl stain AF500/525. On the background of green Nissl staining, the red BDA labeling, including neuronal cell bodies and axonal terminals, was more distinctly demonstrated in the somatosensory cortex. Furthermore, double fluorescent staining for BDA and the calcium-binding protein parvalbumin (PV) was carried out to observe the correlation of BDA labeling and PV-positive interneurons in the cortical target, providing the opportunity to study the local neural circuits and their chemical characteristics. Thus, this refined method is not only suitable for visualizing high quality neural labeling with the high molecular weight BDA through reciprocal neural pathways between the thalamus and cerebral cortex, but also will permit the simultaneous demonstration of other neural markers with fluorescent histochemistry or immunochemistry.

  11. How nanoparticles encapsulating fluorophores allow a double detection of biomolecules by localized surface plasmon resonance and luminescence

    International Nuclear Information System (INIS)

    Barbillon, G; Faure, A C; Kork, N El; Moretti, P; Roux, S; Tillement, O; Ou, M G; Descamps, A; Perriat, P; Vial, A; Bijeon, J-L; Marquette, C A; Jacquier, B

    2008-01-01

    The paper shows how polysiloxane particles encapsulating fluorophores can be successfully used to detect biotin-streptavidin binding by two types of technique. After functionalization of the particles by streptavidin, the fixation of the biomolecule can indeed be detected by a shift of the localized surface plasmon resonance of the biotinylated gold dots used as substrate and by the luminescence of the fluorophores evidenced by scanning near-field optical microscopy. The development of particles allowing such a double detection opens a route for increasing the reliability of biological detection and for multi-labelling strategies crossing both detection principles

  12. How nanoparticles encapsulating fluorophores allow a double detection of biomolecules by localized surface plasmon resonance and luminescence

    Energy Technology Data Exchange (ETDEWEB)

    Barbillon, G; Faure, A C; Kork, N El; Moretti, P [Universite de Lyon, Universite Lyon 1, CNRS UMR 5620, Laboratoire de Physico-Chimie des Materiaux Luminescents (LPCML), Domaine Scientifique de La Doua, Bat Kastler, 10 rue Andre Marie Ampere 69622 Villeurbanne Cedex (France); Roux, S; Tillement, O [Universite de Lyon, Universite Lyon 1, CNRS UMR 5620, Laboratoire de Physico-Chimie des Materiaux Luminescents (LPCML), Domaine Scientifique de La Doua, Bat Kastler, 10 rue Andre Marie Ampere 69622 Villeurbanne Cedex (France); Ou, M G; Descamps, A; Perriat, P [Materiaux, Ingenierie et Sciences (MATEIS), CNRS UMR 5510, Universite de Lyon, INSA-Lyon, Domaine Scientifique de La Doua, 7 avenue Jean Capelle 69621 Villeurbanne Cedex (France); Vial, A; Bijeon, J-L [Institut Charles Delaunay, CNRS FRE 2848, Laboratoire de Nanotechnologie et d' Instrumentation Optique (LNIO), Universite de Technologie de Troyes, 12 rue Marie Curie BP 2060 10010 Troyes Cedex (France); Marquette, C A [Laboratoire de Genie Enzymatique et Biomoleculaire, UMR 5246 CNRS-ICBMS, Universite de Lyon, Universite Lyon 1, 69622 Villeurbanne Cedex (France); Jacquier, B [Universite de Lyon, Universite Lyon 1, CNRS UMR 5620, Laboratoire de Physico-Chimie des Materiaux Luminescents (LPCML), Domaine Scientifique de La Doua, Bat Kastler, 10 rue Andre Marie Ampere 69622 Villeurbanne Cedex (France)

    2008-01-23

    The paper shows how polysiloxane particles encapsulating fluorophores can be successfully used to detect biotin-streptavidin binding by two types of technique. After functionalization of the particles by streptavidin, the fixation of the biomolecule can indeed be detected by a shift of the localized surface plasmon resonance of the biotinylated gold dots used as substrate and by the luminescence of the fluorophores evidenced by scanning near-field optical microscopy. The development of particles allowing such a double detection opens a route for increasing the reliability of biological detection and for multi-labelling strategies crossing both detection principles.

  13. Cell-based lipid flippase assay employing fluorescent lipid derivatives

    DEFF Research Database (Denmark)

    Jensen, Maria Stumph; Costa, Sara; Günther-Pomorski, Thomas

    2016-01-01

    P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases, s...... flippase activities in the plasma membrane of cells, using yeast as an example.......P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases......, studies of individual P4-ATPase family members from fungi, plants, and animals show that P4-ATPases differ in their substrate specificities and mediate transport of a broader range of lipid substrates. Here, we describe an assay based on fluorescent lipid derivatives to monitor and characterize lipid...

  14. Detection of Biomolecular Binding Through Enhancement of Localized Surface Plasmon Resonance (LSPR by Gold Nanoparticles

    Directory of Open Access Journals (Sweden)

    Min-Gon Kim

    2009-03-01

    Full Text Available To amplify the difference in localized surface plasmon resonance (LSPR spectra of gold nano-islands due to intermolecular binding events, gold nanoparticles were used. LSPR-based optical biosensors consisting of gold nano-islands were readily made on glass substrates using evaporation and heat treatment. Streptavidin (STA and biotinylated bovine serum albumin (Bio-BSA were chosen as the model receptor and the model analyte, respectively, to demonstrate the effectiveness of this detection method. Using this model system, we were able to enhance the sensitivity in monitoring the binding of Bio-BSA to gold nano-island surfaces functionalized with STA through the addition of gold nanoparticle-STA conjugates. In addition, SU-8 well chips with gold nano-island surfaces were fabricated through a conventional UV patterning method and were then utilized for image detection using the attenuated total reflection mode. These results suggest that the gold nano-island well chip may have the potential to be used for multiple and simultaneous detection of various bio-substances.

  15. Immobilization of streptavidin on 4H-SiC for biosensor development

    International Nuclear Information System (INIS)

    Williams, Elissa H.; Davydov, Albert V.; Motayed, Abhishek; Sundaresan, Siddarth G.; Bocchini, Peter; Richter, Lee J.; Stan, Gheorghe; Steffens, Kristen; Zangmeister, Rebecca; Schreifels, John A.; Rao, Mulpuri V.

    2012-01-01

    A sequential layer formation chemistry is demonstrated for the functionalization of silicon carbide (SiC) appropriate to biosensing applications. (0 0 0 1) 4H-SiC was functionalized with 3-aminopropyltriethoxysilane (APTES) and subsequently biotinylated for the selective immobilization of streptavidin. Atomic force microscopy, X-ray photoelectron spectroscopy, ellipsometry, fluorescence microscopy, and contact angle measurements were utilized to determine the structure, thickness, wettability, and reactivity of the resulting surface after each functionalization step. Optimization of the APTES layer was found to be critical to the success of the subsequent steps; multilayer, polymeric films resulted in irreproducible behavior. It was shown that there was significant non-specific (electrostatic) binding of streptavidin to APTES functionalized SiC, thus revealing the importance of a uniform biotinylation step prior to streptavidin attachment. The experimental results demonstrate that the APTES functionalized and biotinylated SiC surface has the potential to be employed as a biosensing platform for the selective detection of streptavidin molecules.

  16. Silica encapsulation of fluorescent nanodiamonds for colloidal stability and facile surface functionalization.

    Science.gov (United States)

    Bumb, Ambika; Sarkar, Susanta K; Billington, Neil; Brechbiel, Martin W; Neuman, Keir C

    2013-05-29

    Fluorescent nanodiamonds (FNDs) emit in the near-IR and do not photobleach or photoblink. These properties make FNDs better suited for numerous imaging applications compared with commonly used fluorescence agents such as organic dyes and quantum dots. However, nanodiamonds do not form stable suspensions in aqueous buffer, are prone to aggregation, and are difficult to functionalize. Here we present a method for encapsulating nanodiamonds with silica using an innovative liposome-based encapsulation process that renders the particle surface biocompatible, stable, and readily functionalized through routine linking chemistries. Furthermore, the method selects for a desired particle size and produces a monodisperse agent. We attached biotin to the silica-coated FNDs and tracked the three-dimensional motion of a biotinylated FND tethered by a single DNA molecule with high spatial and temporal resolution.

  17. Development of Robust and Standardized Cantilever Sensors Based on Biotin/Neutravidin Coupling for Antibody Detection

    Directory of Open Access Journals (Sweden)

    Christoph Gerber

    2013-04-01

    Full Text Available A cantilever-based protein biosensor has been developed providing a customizable multilayer platform for the detection of antibodies. It consists of a biotin-terminated PEG layer pre-functionalized on the gold-coated cantilever surface, onto which NeutrAvidin is adsorbed through biotin/NeutrAvidin specific binding. NeutrAvidin is used as a bridge layer between the biotin-coated surface and the biotinylated biomolecules, such as biotinylated bovine serum albumin (biotinylated BSA, forming a multilayer sensor for direct antibody capture. The cantilever biosensor has been successfully applied to the detection of mouse anti-BSA (m-IgG and sheep anti-BSA(s-IgG antibodies. As expected, the average differential surface stress signals of about 5.7 ± 0.8 ´ 10−3 N/m are very similar for BSA/m-IgG and BSA/s-IgG binding, i.e., they are independent of the origin of the antibody. A statistic evaluation of 112 response curves confirms that the multilayer protein cantilever biosensor shows high reproducibility. As a control test, a biotinylated maltose binding protein was used for detecting specificity of IgG, the result shows a signal of bBSA layer in response to antibody is 5.8 ´ 10−3 N/m compared to bMBP. The pre-functionalized biotin/PEG cantilever surface is found to show a long shelf-life of at least 40 days and retains its responsivity of above 70% of the signal when stored in PBS buffer at 4 °C. The protein cantilever biosensor represents a rapid, label-free, sensitive and reliable detection technique for a real-time protein assay.

  18. Correlative Light-Electron Microscopy of Lipid-Encapsulated Fluorescent Nanodiamonds for Nanometric Localization of Cell Surface Antigens.

    Science.gov (United States)

    Hsieh, Feng-Jen; Chen, Yen-Wei; Huang, Yao-Kuan; Lee, Hsien-Ming; Lin, Chun-Hung; Chang, Huan-Cheng

    2018-02-06

    Containing an ensemble of nitrogen-vacancy centers in crystal matrices, fluorescent nanodiamonds (FNDs) are a new type of photostable markers that have found wide applications in light microscopy. The nanomaterial also has a dense carbon core, making it visible to electron microscopy. Here, we show that FNDs encapsulated in biotinylated lipids (bLs) are useful for subdiffraction imaging of antigens on cell surface with correlative light-electron microscopy (CLEM). The lipid encapsulation enables not only good dispersion of the particles in biological buffers but also high specific labeling of live cells. By employing the bL-encapsulated FNDs to target CD44 on HeLa cell surface through biotin-mediated immunostaining, we obtained the spatial distribution of these antigens by CLEM with a localization accuracy of ∼50 nm in routine operations. A comparative study with dual-color imaging, in which CD44 was labeled with FND and MICA/MICB was labeled with Alexa Fluor 488, demonstrated the superior performance of FNDs as fluorescent fiducial markers for CLEM of cell surface antigens.

  19. Development of Piezoelectric DNA-Based Biosensor for Direct Detection of Mycobacterium Tuberculosis in Clinical Specimens

    Directory of Open Access Journals (Sweden)

    Thongchai KAEWPHINIT

    2010-02-01

    Full Text Available This study was focused on establishment of piezoelectric biosensor for direct detection of Mycobacterium tuberculosis (MTB in clinical specimens. The quartz crystal immobilized via 3-mercaptopropionic acid (MPA/avidin/DNA biotinylated probe on gold surface and hybridization of the DNA target to DNA biotinylated probe. The optimal concentration of MPA, avidin and 5’-biotinylated DNA probe for immobilization of specific DNA probe on gold surface were 15 mM, 0.1 mg/ml and 1.5 μM, respectively. The detection of genomic DNA digestion in the range from 0.5 to 30 μg/ml. The fabricated biosensor was evaluated through an examination of 200 samples. No cross hybridization were observed against M. avium complex (MAC and other microorganism. This target DNA preparation without amplification will reduce time consuming, costs, and the tedious step of amplification. This study can be extended to develop the new method which is high sensitivity, specificity, cheap, easy to use, and rapid for detection of MTB in many fields.

  20. An SH2 domain-based tyrosine kinase assay using biotin ligase modified with a terbium(III) complex.

    Science.gov (United States)

    Sueda, Shinji; Shinboku, Yuki; Kusaba, Takeshi

    2013-01-01

    Src homology 2 (SH2) domains are modules of approximately 100 amino acids and are known to bind phosphotyrosine-containing sequences with high affinity and specificity. In the present work, we developed an SH2 domain-based assay for Src tyrosine kinase using a unique biotinylation reaction from archaeon Sulfolobus tokodaii. S. tokodaii biotinylation has a unique property that biotin protein ligase (BPL) forms a stable complex with its biotinylated substrate protein (BCCP). Here, an SH2 domain from lymphocyte-specific tyrosine kinase was genetically fused to a truncated BCCP, and the resulting fusion protein was labeled through biotinylation with BPL carrying multiple copies of a luminescent Tb(3+) complex. The labeled SH2 fusion proteins were employed to detect a phosphorylated peptide immobilized on the surface of the microtiter plate, where the phosphorylated peptide was produced by phosphorylation to the substrate peptide by Src tyrosine kinase. Our assay allows for a reliable determination of the activity of Src kinase lower than 10 pg/μL by a simple procedure.

  1. Aptasensor development for detection of virus in water

    DEFF Research Database (Denmark)

    Kirkegaard, Julie

    of three types of waterborne viruses: norovirus, rotavirus and hepatitis A virus. The development of the aptasensor involved sample preparation for aptamer selection of rotavirus and hepatitis A virus, an iterative design process of the aptasensor, investigation of the surface immobilisation of aptamers...... and finally an impedimetric electrical characterisation of the sensor. The sample preparation of the rotavirus was based on purification and biotinylation of the virus to meet the requirements of the aptamer selection process. The selection process, performed by an external collaborator, was based...... on streptavidin coated magnetic bead separation, hence the needed biotinylation. It was found that the BPH linker gave the highest yield when the biotinylated rotavirus were immobilised onto the beads. The design of the viral aptasensor was determined by an iterative design process. The final chip design...

  2. Chemical Reactive Anchoring Lipids with Different Performance for Cell Surface Re-engineering Application.

    Science.gov (United States)

    Vabbilisetty, Pratima; Boron, Mallorie; Nie, Huan; Ozhegov, Evgeny; Sun, Xue-Long

    2018-02-28

    Introduction of selectively chemical reactive groups at the cell surface enables site-specific cell surface labeling and modification opportunity, thus facilitating the capability to study the cell surface molecular structure and function and the molecular mechanism it underlies. Further, it offers the opportunity to change or improve a cell's functionality for interest of choice. In this study, two chemical reactive anchor lipids, phosphatidylethanolamine-poly(ethylene glycol)-dibenzocyclooctyne (DSPE-PEG 2000 -DBCO) and cholesterol-PEG-dibenzocyclooctyne (CHOL-PEG 2000 -DBCO) were synthesized and their potential application for cell surface re-engineering via lipid fusion were assessed with RAW 264.7 cells as a model cell. Briefly, RAW 264.7 cells were incubated with anchor lipids under various concentrations and at different incubation times. The successful incorporation of the chemical reactive anchor lipids was confirmed by biotinylation via copper-free click chemistry, followed by streptavidin-fluorescein isothiocyanate binding. In comparison, the cholesterol-based anchor lipid afforded a higher cell membrane incorporation efficiency with less internalization than the phospholipid-based anchor lipid. Low cytotoxicity of both anchor lipids upon incorporation into the RAW 264.7 cells was observed. Further, the cell membrane residence time of the cholesterol-based anchor lipid was evaluated with confocal microscopy. This study suggests the potential cell surface re-engineering applications of the chemical reactive anchor lipids.

  3. Chemical Reactive Anchoring Lipids with Different Performance for Cell Surface Re-engineering Application

    Science.gov (United States)

    2018-01-01

    Introduction of selectively chemical reactive groups at the cell surface enables site-specific cell surface labeling and modification opportunity, thus facilitating the capability to study the cell surface molecular structure and function and the molecular mechanism it underlies. Further, it offers the opportunity to change or improve a cell’s functionality for interest of choice. In this study, two chemical reactive anchor lipids, phosphatidylethanolamine–poly(ethylene glycol)–dibenzocyclooctyne (DSPE–PEG2000–DBCO) and cholesterol–PEG–dibenzocyclooctyne (CHOL–PEG2000–DBCO) were synthesized and their potential application for cell surface re-engineering via lipid fusion were assessed with RAW 264.7 cells as a model cell. Briefly, RAW 264.7 cells were incubated with anchor lipids under various concentrations and at different incubation times. The successful incorporation of the chemical reactive anchor lipids was confirmed by biotinylation via copper-free click chemistry, followed by streptavidin-fluorescein isothiocyanate binding. In comparison, the cholesterol-based anchor lipid afforded a higher cell membrane incorporation efficiency with less internalization than the phospholipid-based anchor lipid. Low cytotoxicity of both anchor lipids upon incorporation into the RAW 264.7 cells was observed. Further, the cell membrane residence time of the cholesterol-based anchor lipid was evaluated with confocal microscopy. This study suggests the potential cell surface re-engineering applications of the chemical reactive anchor lipids. PMID:29503972

  4. Rapid and specific biotin labelling of the erythrocyte surface antigens of both cultured and ex-vivo Plasmodium parasites

    Directory of Open Access Journals (Sweden)

    Thompson Joanne

    2007-05-01

    Full Text Available Abstract Background Sensitive detection of parasite surface antigens expressed on erythrocyte membranes is necessary to further analyse the molecular pathology of malaria. This study describes a modified biotin labelling/osmotic lysis method which rapidly produces membrane extracts enriched for labelled surface antigens and also improves the efficiency of antigen recovery compared with traditional detergent extraction and surface radio-iodination. The method can also be used with ex-vivo parasites. Methods After surface labelling with biotin in the presence of the inhibitor furosemide, detergent extraction and osmotic lysis methods of enriching for the membrane fractions were compared to determine the efficiency of purification and recovery. Biotin-labelled proteins were identified on silver-stained SDS-polyacrylamide gels. Results Detergent extraction and osmotic lysis were compared for their capacity to purify biotin-labelled Plasmodium falciparum and Plasmodium chabaudi erythrocyte surface antigens. The pellet fraction formed after osmotic lysis of P. falciparum-infected erythrocytes is notably enriched in suface antigens, including PfEMP1, when compared to detergent extraction. There is also reduced co-extraction of host proteins such as spectrin and Band 3. Conclusion Biotinylation and osmotic lysis provides an improved method to label and purify parasitised erythrocyte surface antigen extracts from both in vitro and ex vivo Plasmodium parasite preparations.

  5. ASIC subunit ratio and differential surface trafficking in the brain.

    Science.gov (United States)

    Wu, Junjun; Xu, Yuanyuan; Jiang, Yu-Qing; Xu, Jiangping; Hu, Youjia; Zha, Xiang-ming

    2016-01-08

    Acid-sensing ion channels (ASICs) are key mediators of acidosis-induced responses in neurons. However, little is known about the relative abundance of different ASIC subunits in the brain. Such data are fundamental for interpreting the relative contribution of ASIC1a homomers and 1a/2 heteromers to acid signaling, and essential for designing therapeutic interventions to target these channels. We used a simple biochemical approach and semi-quantitatively determined the molar ratio of ASIC1a and 2 subunits in mouse brain. Further, we investigated differential surface trafficking of ASIC1a, ASIC2a, and ASIC2b. ASIC1a subunits outnumber the sum of ASIC2a and ASIC2b. There is a region-specific variation in ASIC2a and 2b expression, with cerebellum and striatum expressing predominantly 2b and 2a, respectively. Further, we performed surface biotinylation and found that surface ASIC1a and ASIC2a ratio correlates with their total expression. In contrast, ASIC2b exhibits little surface presence in the brain. This result is consistent with increased co-localization of ASIC2b with an ER marker in 3T3 cells. Our data are the first semi-quantitative determination of relative subunit ratio of various ASICs in the brain. The differential surface trafficking of ASICs suggests that the main functional ASICs in the brain are ASIC1a homomers and 1a/2a heteromers. This finding provides important insights into the relative contribution of various ASIC complexes to acid signaling in neurons.

  6. Uniform Orientation of Biotinylated Nanobody as an Affinity Binder for Detection of Bacillus thuringiensis (Bt) Cry1Ac Toxin

    Science.gov (United States)

    Li, Min; Zhu, Min; Zhang, Cunzheng; Liu, Xianjin; Wan, Yakun

    2014-01-01

    Nanobodies are the smallest natural fragments with useful properties such as high affinity, distinct paratope and high stability, which make them an ideal tool for detecting target antigens. In this study, we generated and characterized nanobodies against the Cry1Ac toxin and applied them in a biotin-streptavidin based double antibodies (nanobodies) sandwich-ELISA (DAS-ELISA) assay. After immunizing a camel with soluble Cry1Ac toxin, a phage displayed library was constructed to generate Nbs against the Cry1Ac toxin. Through successive rounds of affinity bio-panning, four nanobodies with greatest diversity in CDR3 sequences were obtained. After affinity determination and conjugating to HRP, two nanobodies with high affinity which can recognize different epitopes of the same antigen (Cry1Ac) were selected as capture antibody (Nb61) and detection antibody (Nb44). The capture antibody (Nb61) was biotinylated in vivo for directional immobilization on wells coated with streptavidin matrix. Both results of specificity analysis and thermal stability determination add support for reliability of the following DAS-ELISA with a minimum detection limit of 0.005 μg·mL−1 and a working range 0.010–1.0 μg·mL−1. The linear curve displayed an acceptable correlation coefficient of 0.9976. These results indicated promising applications of nanobodies for detection of Cry1Ac toxin with biotin-streptavidin based DAS-ELISA system. PMID:25474492

  7. Uniform Orientation of Biotinylated Nanobody as an Affinity Binder for Detection of Bacillus thuringiensis (Bt Cry1Ac Toxin

    Directory of Open Access Journals (Sweden)

    Min Li

    2014-12-01

    Full Text Available Nanobodies are the smallest natural fragments with useful properties such as high affinity, distinct paratope and high stability, which make them an ideal tool for detecting target antigens. In this study, we generated and characterized nanobodies against the Cry1Ac toxin and applied them in a biotin-streptavidin based double antibodies (nanobodies sandwich-ELISA (DAS-ELISA assay. After immunizing a camel with soluble Cry1Ac toxin, a phage displayed library was constructed to generate Nbs against the Cry1Ac toxin. Through successive rounds of affinity bio-panning, four nanobodies with greatest diversity in CDR3 sequences were obtained. After affinity determination and conjugating to HRP, two nanobodies with high affinity which can recognize different epitopes of the same antigen (Cry1Ac were selected as capture antibody (Nb61 and detection antibody (Nb44. The capture antibody (Nb61 was biotinylated in vivo for directional immobilization on wells coated with streptavidin matrix. Both results of specificity analysis and thermal stability determination add support for reliability of the following DAS-ELISA with a minimum detection limit of 0.005 μg·mL−1 and a working range 0.010–1.0 μg·mL−1. The linear curve displayed an acceptable correlation coefficient of 0.9976. These results indicated promising applications of nanobodies for detection of Cry1Ac toxin with biotin-streptavidin based DAS-ELISA system.

  8. Ultrasensitive detection and quantification of E. coli O157:H7 using a giant magneto impedance sensor in an open-surface micro fluidic cavity covered with an antibody-modified gold surface

    International Nuclear Information System (INIS)

    Yang, Zhen; Liu, Yan; Lei, Chong; Sun, Xue-cheng; Zhou, Yong

    2016-01-01

    We report on a method for ultrasensitive detection and quantification of the pathogen Escherichia coli (E. coli), type O157:H7. It is using a tortuous-shaped giant magneto impedance (GMI) sensor in combination with an open-surface micro fluidic system coated with a gold film for performing the sandwich immuno binding on its surface. Streptavidin-coated super magnetic Dynabeads were loaded with biotinylated polyclonal antibody to capture E. coli O157:H7. The E. coli-loaded Dynabeads are then injected into the microfluidics system where it comes into contact with the surface of gold nanofilm carrying the monoclonal antibody to form the immuno complex. As a result, the GMI ratio is strongly reduced at high frequencies if E. coli O157:H7 is present. The sensor has a linear response in the 50 to 500 cfu·mL"−"1 concentration range, and the detection limit is 50 cfu·mL"−"1 at a working frequency of 2.2 MHz. In our perception, this method provides a valuable tool for developing GMI-based micro fluidic sensors systems for ultrasensitive and quantitative analysis of pathogenic bacteria. The method may also be extended to other sensing applications by employing respective immuno reagents. (author)

  9. Heat shock proteins on the human sperm surface.

    Science.gov (United States)

    Naaby-Hansen, Soren; Herr, John C

    2010-01-01

    The sperm plasma membrane is known to be critical to fertilization and to be highly regionalized into domains of head, mid- and principal pieces. However, the molecular composition of the sperm plasma membrane and its alterations during genital tract passage, capacitation and the acrosome reaction remains to be fully dissected. A two-dimensional gel-based proteomic study previously identified 98 human sperm proteins which were accessible for surface labelling with both biotin and radioiodine. In this report twelve dually labelled protein spots were excised from stained gels or PDVF membranes and analysed by mass spectrometry (MS) and Edman degradation. Seven members from four different heat shock protein (HSP) families were identified including HYOU1 (ORP150), HSPC1 (HSP86), HSPA5 (Bip), HSPD1 (HSP60), and several isoforms of the two testis-specific HSP70 chaperones HSPA2 and HSPA1L. An antiserum raised against the testis-specific HSPA2 chaperone reacted with three 65kDa HSPA2 isoforms and three high molecular weight surface proteins (78-79kDa, 84kDa and 90-93kDa). These proteins, together with seven 65kDa HSP70 forms, reacted with human anti-sperm IgG antibodies that blocked in vitro fertilization in humans. Three of these surface biotinylated human sperm antigens were immunoprecipitated with a rabbit antiserum raised against a linear peptide epitope in Chlamydia trachomatis HSP70. The results indicate diverse HSP chaperones are accessible for surface labelling on human sperm. Some of these share epitopes with C. trachomatis HSP70, suggesting an association between genital tract infection, immunity to HSP70 and reproductive failure. 2009 Elsevier Ireland Ltd. All rights reserved.

  10. Visual detection and microplate assay for Staphylococcus aureus based on aptamer recognition coupled to tyramine signal amplification

    International Nuclear Information System (INIS)

    Yuan, Jinglei; Li, Can; Ma, Xiaoyuan; Xia, Yu; Chen, Jie; Wang, Zhouping; Yu, Ye

    2014-01-01

    We have developed a specific method for the visual detection of Staphylococcus aureus based on aptamer recognition coupled to tyramine signal amplification technology. A biotinylated aptamer specific for S. aureus was immobilized on the surface of the wells of a microplate via biotin-avidin binding. Then, the target bacteria (S. aureus), the biotinylated-aptamer-streptavidin-HRP conjugates, biotinylated tyramine, hydrogen peroxide and streptavidin-HRP were successively placed in the wells of the microplate. After adding TMB reagent and stop solution, the intensity of the yellow reaction product can be visually inspected or measured with a plate reader. Under optimized conditions, there is a linear relationship between absorbance at 450 nm and the concentration of S. aureus in the 10 to 107 cfu mL −1 concentration range (with an R 2 of 0.9976). The limit of detection is 8 cfu mL −1 . (author)

  11. Resolving the detailed structure of cortical and thalamic neurons in the adult rat brain with refined biotinylated dextran amine labeling.

    Science.gov (United States)

    Ling, Changying; Hendrickson, Michael L; Kalil, Ronald E

    2012-01-01

    Biotinylated dextran amine (BDA) has been used frequently for both anterograde and retrograde pathway tracing in the central nervous system. Typically, BDA labels axons and cell somas in sufficient detail to identify their topographical location accurately. However, BDA labeling often has proved to be inadequate to resolve the fine structural details of axon arbors or the dendrites of neurons at a distance from the site of BDA injection. To overcome this limitation, we varied several experimental parameters associated with the BDA labeling of neurons in the adult rat brain in order to improve the sensitivity of the method. Specifically, we compared the effect on labeling sensitivity of: (a) using 3,000 or 10,000 MW BDA; (b) injecting different volumes of BDA; (c) co-injecting BDA with NMDA; and (d) employing various post-injection survival times. Following the extracellular injection of BDA into the visual cortex, labeled cells and axons were observed in both cortical and thalamic areas of all animals studied. However, the detailed morphology of axon arbors and distal dendrites was evident only under optimal conditions for BDA labeling that take into account the: molecular weight of the BDA used, concentration and volume of BDA injected, post-injection survival time, and toning of the resolved BDA with gold and silver. In these instances, anterogradely labeled axons and retrogradely labeled dendrites were resolved in fine detail, approximating that which can be achieved with intracellularly injected compounds such as biocytin or fluorescent dyes.

  12. Fluorescent polymeric nanocomposite films generated by surface-mediated photoinitiation of polymerization

    International Nuclear Information System (INIS)

    Avens, Heather J.; Chang, Erin L.; May, Allison M.; Berron, Brad J.; Seedorf, Gregory J.; Balasubramaniam, Vivek; Bowman, Christopher N.

    2011-01-01

    Incorporation of nanoparticles (NPs) into polymer films represents a valuable strategy for achieving a variety of desirable physical, optical, mechanical, and electrical attributes. Here, we describe and characterize the creation of highly fluorescent polymer films by entrapment of fluorescent NPs into polymer matrices through surface-mediated eosin photoinitiation reactions. Performing surface-mediated polymerizations with NPs combines the benefits of a covalently anchored film with the unique material properties afforded by NPs. The effects of monomer type, crosslinker content, NP size, and NP surface chemistry were investigated to determine their impact on the relative amount of NPs entrapped in the surface-bound films. The density of entrapped NPs was increased up to 6-fold by decreasing the NP diameter. Increasing the crosslinking agent concentration enabled a greater than 2-fold increase in the amount of NPs entrapped. Additionally, the monomer chemistry played a significant role as poly(ethylene glycol) diacrylate (PEGDA)-based monomer formulations entrapped a 10-fold higher density of carboxy-functionalized NPs than did acrylamide/bisacrylamide formulations, though the latter formulations ultimately immobilized more fluorophores by generating thicker films. In the context of a polymerization-based microarray biodetection platform, these findings enabled tailoring of the monomer and NP selection to yield a 200-fold improvement in sensitivity from 31 (±1) to 0.16 (±0.01) biotinylated target molecules per square micron. Similarly, in polymerization-based cell staining applications, appropriate monomer and NP selection enabled facile visualization of microscale, sub-cellular features. Careful consideration of monomer and NP selection is critical to achieve the desired properties in applications that employ surface-mediated polymerization to entrap NPs.

  13. Quantitative proteomic view on secreted, cell surface-associated, and cytoplasmic proteins of the methicillin-resistant human pathogen Staphylococcus aureus under iron-limited conditions.

    Science.gov (United States)

    Hempel, Kristina; Herbst, Florian-Alexander; Moche, Martin; Hecker, Michael; Becher, Dörte

    2011-04-01

    Staphylococcus aureus is capable of colonizing and infecting humans by its arsenal of surface-exposed and secreted proteins. Iron-limited conditions in mammalian body fluids serve as a major environmental signal to bacteria to express virulence determinants. Here we present a comprehensive, gel-free, and GeLC-MS/MS-based quantitative proteome profiling of S. aureus under this infection-relevant situation. (14)N(15)N metabolic labeling and three complementing approaches were combined for relative quantitative analyses of surface-associated proteins. The surface-exposed and secreted proteome profiling approaches comprise trypsin shaving, biotinylation, and precipitation of the supernatant. By analysis of the outer subproteomic and cytoplasmic protein fraction, 1210 proteins could be identified including 221 surface-associated proteins. Thus, access was enabled to 70% of the predicted cell wall-associated proteins, 80% of the predicted sortase substrates, two/thirds of lipoproteins and more than 50% of secreted and cytoplasmic proteins. For iron-deficiency, 158 surface-associated proteins were quantified. Twenty-nine proteins were found in altered amounts showing particularly surface-exposed proteins strongly induced, such as the iron-regulated surface determinant proteins IsdA, IsdB, IsdC and IsdD as well as lipid-anchored iron compound-binding proteins. The work presents a crucial subject for understanding S. aureus pathophysiology by the use of methods that allow quantitative surface proteome profiling.

  14. Profiling the outer membrane proteome during growth and development of the social bacterium Myxococcus xanthus by selective biotinylation and analyses of outer membrane vesicles.

    Science.gov (United States)

    Kahnt, Jörg; Aguiluz, Kryssia; Koch, Jürgen; Treuner-Lange, Anke; Konovalova, Anna; Huntley, Stuart; Hoppert, Michael; Søgaard-Andersen, Lotte; Hedderich, Reiner

    2010-10-01

    Social behavior in the bacterium Myxococcus xanthus relies on contact-dependent activities involving cell-cell and cell-substratum interactions. To identify outer membrane proteins that have a role in these activities, we profiled the outer membrane proteome of growing and starving cells using two strategies. First, outer membrane proteins were enriched by biotinylation of intact cells using the reagent NHS (N-hydroxysuccinimide)-PEO(12) (polyethylene oxide)-biotin with subsequent membrane solubilization and affinity chromatography. Second, the proteome of outer membrane vesicles (OMV) was determined. Comparisons of detected proteins show that these methods have different detection profiles and together provide a comprehensive view of the outer membrane proteome. From 362 proteins identified, 274 (76%) were cell envelope proteins including 64 integral outer membrane proteins and 85 lipoproteins. The majority of these proteins were of unknown function. Among integral outer membrane proteins with homologues of known function, TonB-dependent transporters comprise the largest group. Our data suggest novel functions for these transporters. Among lipoproteins with homologues of known function, proteins with hydrolytic functions comprise the largest group. The luminal load of OMV was enriched for proteins with hydrolytic functions. Our data suggest that OMV have functions in predation and possibly in transfer of intercellular signaling molecules between cells.

  15. Perfringolysin O Theta Toxin as a Tool to Monitor the Distribution and Inhomogeneity of Cholesterol in Cellular Membranes.

    Science.gov (United States)

    Maekawa, Masashi; Yang, Yanbo; Fairn, Gregory D

    2016-03-08

    Cholesterol is an essential structural component of cellular membranes in eukaryotes. Cholesterol in the exofacial leaflet of the plasma membrane is thought to form membrane nanodomains with sphingolipids and specific proteins. Additionally, cholesterol is found in the intracellular membranes of endosomes and has crucial functions in membrane trafficking. Furthermore, cellular cholesterol homeostasis and regulation of de novo synthesis rely on transport via both vesicular and non-vesicular pathways. Thus, the ability to visualize and detect intracellular cholesterol, especially in the plasma membrane, is critical to understanding the complex biology associated with cholesterol and the nanodomains. Perfringolysin O (PFO) theta toxin is one of the toxins secreted by the anaerobic bacteria Clostridium perfringens and this toxin forms pores in the plasma membrane that causes cell lysis. It is well understood that PFO recognizes and binds to cholesterol in the exofacial leaflets of the plasma membrane, and domain 4 of PFO (D4) is sufficient for the binding of cholesterol. Recent studies have taken advantage of this high-affinity cholesterol-binding domain to create a variety of cholesterol biosensors by using a non-toxic PFO or the D4 in isolation. This review highlights the characteristics and usefulness of, and the principal findings related to, these PFO-derived cholesterol biosensors.

  16. Cholesterol asymmetry in synaptic plasma membranes.

    Science.gov (United States)

    Wood, W Gibson; Igbavboa, Urule; Müller, Walter E; Eckert, Gunter P

    2011-03-01

    Lipids are essential for the structural and functional integrity of membranes. Membrane lipids are not randomly distributed but are localized in different domains. A common characteristic of these membrane domains is their association with cholesterol. Lipid rafts and caveolae are examples of cholesterol enriched domains, which have attracted keen interest. However, two other important cholesterol domains are the exofacial and cytofacial leaflets of the plasma membrane. The two leaflets that make up the bilayer differ in their fluidity, electrical charge, lipid distribution, and active sites of certain proteins. The synaptic plasma membrane (SPM) cytofacial leaflet contains over 85% of the total SPM cholesterol as compared with the exofacial leaflet. This asymmetric distribution of cholesterol is not fixed or immobile but can be modified by different conditions in vivo: (i) chronic ethanol consumption; (ii) statins; (iii) aging; and (iv) apoE isoform. Several potential candidates have been proposed as mechanisms involved in regulation of SPM cholesterol asymmetry: apoE, low-density lipoprotein receptor, sterol carrier protein-2, fatty acid binding proteins, polyunsaturated fatty acids, P-glycoprotein and caveolin-1. This review examines cholesterol asymmetry in SPM, potential mechanisms of regulation and impact on membrane structure and function. © 2011 The Authors. Journal of Neurochemistry © 2011 International Society for Neurochemistry.

  17. Immobilization of serum albumin and peptide aptamer for EPC on polydopamine coated titanium surface for enhanced in-situ self-endothelialization

    International Nuclear Information System (INIS)

    Chen, Zhuoyue; Li, Quanli; Chen, Jialong; Luo, Rifang; Maitz, Manfred F.; Huang, Nan

    2016-01-01

    Restenosis and thrombosis are two major complications associated with vascular stents and grafts. The homing of circulating endothelial progenitor cells (EPCs) onto implant surfaces brings a new strategy to solve these problems by accelerating self -endothelialization in situ. Peptide aptamers with high affinity and specific recognition of EPCs can be immobilized to capture EPCs from the circulating blood. In this study, a biotinylated peptide aptamer (TPSLEQRTVYAK-GGGC-K-Biotin) for EPC, and bovine serum albumin (BSA) were co-immobilized onto titanium surface through avidin–biotin recognition to endow the surface with specific affinity for EPC and anti-platelet adhesion properties. Quartz crystal microbalance with dissipation (QCM-D), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and water contact angle measuring were adopted for coating characterization. EPC affinity and hemocompatibility of the coating were also investigated in vitro. The results demonstrated that aptamer and BSA co-immobilized surface significantly reduced platelet adhesion and fibrinogen adsorption/activation. Besides, such functional surface could remarkably enhance EPC adhesion, without affecting the behavior of endothelial cells (ECs) and smooth muscle cells (SMCs) obviously. The result shows the possibility of utilizing such a multifunctional surface in cardiovascular implants. - Highlights: • We construct a multifunctional surface based on immobilization of BSA and aptamer. • It can significantly reduce platelet adhesion and fibrinogen adsorption/activation. • Such functional surface could remarkably enhance EPC adhesion in vitro. • It can induce rapid self-endothelialization of the implant surface in situ in vivo. • It is possible to use such a multifunctional surface in cardiovascular implants.

  18. Immobilization of serum albumin and peptide aptamer for EPC on polydopamine coated titanium surface for enhanced in-situ self-endothelialization

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zhuoyue, E-mail: 362947953@qq.com [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu, 610031 (China); RegeMed Lab of Tissue Engineering, Faculty of Life Science, Northwest University, Xi' an, 710069 (China); Li, Quanli [College of Stomology, Anhui Medical University, Hefei, 230032 (China); Chen, Jialong [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu, 610031 (China); College of Stomology, Anhui Medical University, Hefei, 230032 (China); Luo, Rifang [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu, 610031 (China); Maitz, Manfred F. [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu, 610031 (China); Leibniz Institute of Polymer Research Dresden, Max Bergmann Center of Biomaterials, Dresden (Germany); Huang, Nan, E-mail: huangnan1956@163.com [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu, 610031 (China)

    2016-03-01

    Restenosis and thrombosis are two major complications associated with vascular stents and grafts. The homing of circulating endothelial progenitor cells (EPCs) onto implant surfaces brings a new strategy to solve these problems by accelerating self -endothelialization in situ. Peptide aptamers with high affinity and specific recognition of EPCs can be immobilized to capture EPCs from the circulating blood. In this study, a biotinylated peptide aptamer (TPSLEQRTVYAK-GGGC-K-Biotin) for EPC, and bovine serum albumin (BSA) were co-immobilized onto titanium surface through avidin–biotin recognition to endow the surface with specific affinity for EPC and anti-platelet adhesion properties. Quartz crystal microbalance with dissipation (QCM-D), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and water contact angle measuring were adopted for coating characterization. EPC affinity and hemocompatibility of the coating were also investigated in vitro. The results demonstrated that aptamer and BSA co-immobilized surface significantly reduced platelet adhesion and fibrinogen adsorption/activation. Besides, such functional surface could remarkably enhance EPC adhesion, without affecting the behavior of endothelial cells (ECs) and smooth muscle cells (SMCs) obviously. The result shows the possibility of utilizing such a multifunctional surface in cardiovascular implants. - Highlights: • We construct a multifunctional surface based on immobilization of BSA and aptamer. • It can significantly reduce platelet adhesion and fibrinogen adsorption/activation. • Such functional surface could remarkably enhance EPC adhesion in vitro. • It can induce rapid self-endothelialization of the implant surface in situ in vivo. • It is possible to use such a multifunctional surface in cardiovascular implants.

  19. Streptavidin-coated gold nanoparticles: critical role of oligonucleotides on stability and fractal aggregation

    Directory of Open Access Journals (Sweden)

    Roberta D'Agata

    2017-01-01

    Full Text Available Gold nanoparticles (AuNPs exhibit unique properties that can be modulated through a tailored surface functionalization, enabling their targeted use in biochemical sensing and medical diagnostics. In particular, streptavidin-modified AuNPs are increasingly used for biosensing purposes. We report here a study of AuNPs surface-functionalized with streptavidin-biotinylated oligonucleotide, focussing on the role played by the oligonucleotide probes in the stabilization/destabilization of the functionalized nanoparticle dispersion. The behaviour of the modified AuNP dispersion as a consequence of the competitive displacement of the biotinylated oligonucleotide has been investigated and the critical role of displaced oligonucletides in triggering the quasi one-dimensional aggregation of nanoparticles is demonstrated for the first time. The thorough understanding of the fundamental properties of bioconjugated AuNPs is of great importance for the design of highly sensitive and reliable functionalized AuNP-based assays.

  20. Biotin-tagged proteins: Reagents for efficient ELISA-based serodiagnosis and phage display-based affinity selection.

    Science.gov (United States)

    Verma, Vaishali; Kaur, Charanpreet; Grover, Payal; Gupta, Amita; Chaudhary, Vijay K

    2018-01-01

    The high-affinity interaction between biotin and streptavidin has opened avenues for using recombinant proteins with site-specific biotinylation to achieve efficient and directional immobilization. The site-specific biotinylation of proteins carrying a 15 amino acid long Biotin Acceptor Peptide tag (BAP; also known as AviTag) is effected on a specific lysine either by co-expressing the E. coli BirA enzyme in vivo or by using purified recombinant E. coli BirA enzyme in the presence of ATP and biotin in vitro. In this paper, we have designed a T7 promoter-lac operator-based expression vector for rapid and efficient cloning, and high-level cytosolic expression of proteins carrying a C-terminal BAP tag in E. coli with TEV protease cleavable N-terminal deca-histidine tag, useful for initial purification. Furthermore, a robust three-step purification pipeline integrated with well-optimized protocols for TEV protease-based H10 tag removal, and recombinant BirA enzyme-based site-specific in vitro biotinylation is described to obtain highly pure biotinylated proteins. Most importantly, the paper demonstrates superior sensitivities in indirect ELISA with directional and efficient immobilization of biotin-tagged proteins on streptavidin-coated surfaces in comparison to passive immobilization. The use of biotin-tagged proteins through specific immobilization also allows more efficient selection of binders from a phage-displayed naïve antibody library. In addition, for both these applications, specific immobilization requires much less amount of protein as compared to passive immobilization and can be easily multiplexed. The simplified strategy described here for the production of highly pure biotin-tagged proteins will find use in numerous applications, including those, which may require immobilization of multiple proteins simultaneously on a solid surface.

  1. A sensitive gold nanoparticle-based colorimetric aptasensor for Staphylococcus aureus.

    Science.gov (United States)

    Yuan, Jinglei; Wu, Shijia; Duan, Nuo; Ma, Xiaoyuan; Xia, Yu; Chen, Jie; Ding, Zhansheng; Wang, Zhouping

    2014-09-01

    In this study, a gold nanoparticle-based colorimetric aptasensor for Staphylococcus aureus (S. aureus) using tyramine signal amplification (TSA) technology has been developed. First, the biotinylated aptamer specific for S. aureus was immobilized on the surface of the wells of the microtiter plate via biotin-avidin binding. Then, the target bacteria (S. aureus), biotinylated-aptamer-streptavidin-HRP conjugates, biotinylated tyramine, hydrogen peroxide and avidin-catalase were successively introduced into the wells of the microtiter plate. After that, the existing catalase consumed the hydrogen peroxide. Finally, the freshly prepared gold (III) chloride trihydrate was added, the color of the reaction production would be changed and the absorbance at 550 nm could be measured with a plate reader. Under optimized conditions, there was a linear relationship between the absorbance at 550 nm and the concentration of S. aureus over the range from 10 to 10(6) cfu mL(-1) (with an R² of 0.9947). The limit of the developed method was determined to be 9 cfu mL(-1). Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Identification of outer membrane proteins of Yersinia pestis through biotinylation

    NARCIS (Netherlands)

    Smither, S.J.; Hill, J.; Baar, B.L.M. van; Hulst, A.G.; Jong, A.L. de; Titball, R.W.

    2007-01-01

    The outer membrane of Gram-negative bacteria contains proteins that might be good targets for vaccines, antimicrobials or detection systems. The identification of surface located proteins using traditional methods is often difficult. Yersinia pestis, the causative agent of plague, was labelled with

  3. Time-Resolved Analysis of Cytosolic and Surface-Associated Proteins of Staphylococcus aureus HG001 under Planktonic and Biofilm Conditions.

    Science.gov (United States)

    Moche, Martin; Schlüter, Rabea; Bernhardt, Jörg; Plate, Kristina; Riedel, Katharina; Hecker, Michael; Becher, Dörte

    2015-09-04

    Staphylococcal biofilms are associated with persistent infections due to their capacity to protect bacteria against the host's immune system and antibiotics. Cell-surface-associated proteins are of great importance during biofilm formation. In the present study, an optimized biotinylation approach for quantitative GeLC-MS-based analysis of the staphylococcal cell-surface proteome was applied and the cytoplasmic protein fraction was analyzed to elucidate proteomic differences between colony biofilms and planktonic cells. The experimental setup enabled a time-resolved monitoring of the proteome under both culture conditions and the comparison of biofilm cells to planktonic cells at several time points. This allowed discrimination of differences attributed to delayed growth phases from responses provoked by biofilm conditions. Biofilm cells expressed CcpA-dependent catabolic proteins earlier than planktonic cells and strongly accumulated proteins that belong to the SigB stress regulon. The amount of the cell-surface protein and virulence gene regulator Rot decreased within biofilms and MgrA-dependent regulations appeared more pronounced. Biofilm cells simultaneously up-regulated activators (e.g., SarZ) as well as repressors (e.g., SarX) of RNAIII. A decreased amount of high-affinity iron uptake systems and an increased amount of the iron-storage protein FtnA possibly indicated a lower demand of iron in biofilms.

  4. Incorporating functionalized polyethylene glycol lipids into reprecipitated conjugated polymer nanoparticles for bioconjugation and targeted labeling of cells

    Science.gov (United States)

    Kandel, Prakash K.; Fernando, Lawrence P.; Ackroyd, P. Christine; Christensen, Kenneth A.

    2011-03-01

    We report a simple and rapid method to prepare extremely bright, functionalized, stable, and biocompatible conjugated polymer nanoparticles incorporating functionalized polyethylene glycol (PEG) lipids by reprecipitation. These nanoparticles retain the fundamental spectroscopic properties of conjugated polymer nanoparticles prepared without PEG lipid, but demonstrate greater hydrophilicity and quantum yield compared to unmodified conjugated polymer nanoparticles. The sizes of these nanoparticles, as determined by TEM, were 21-26 nm. Notably, these nanoparticles were prepared with several PEG lipid functional end groups, including biotin and carboxy moieties that can be easily conjugated to biomolecules. We have demonstrated the availability of these end groups for functionalization using the interaction of biotin PEG lipid conjugated polymer nanoparticles with streptavidin. Biotinylated PEG lipid conjugated polymer nanoparticles bound streptavidin-linked magnetic beads, while carboxy and methoxy PEG lipid modified nanoparticles did not. Similarly, biotinylated PEG lipid conjugated polymer nanoparticles bound streptavidin-coated glass slides and could be visualized as diffraction-limited spots, while nanoparticles without PEG lipid or with non-biotin PEG lipid end groups were not bound. To demonstrate that nanoparticle functionalization could be used for targeted labelling of specific cellular proteins, biotinylated PEG lipid conjugated polymer nanoparticles were bound to biotinylated anti-CD16/32 antibodies on J774A.1 cell surface receptors, using streptavidin as a linker. This work represents the first demonstration of targeted delivery of conjugated polymer nanoparticles and demonstrates the utility of these new nanoparticles for fluorescence based imaging and sensing.We report a simple and rapid method to prepare extremely bright, functionalized, stable, and biocompatible conjugated polymer nanoparticles incorporating functionalized polyethylene glycol (PEG

  5. Localization of urokinase-type plasminogen activator receptor on U937 cells

    DEFF Research Database (Denmark)

    Hansen, S H; Behrendt, N; Danø, K

    1990-01-01

    The binding of human urokinase-type plasminogen activator (u-PA) to the surface of the human monocytic cell line U937 was studied by immunological detection of bound u-PA or binding of biotinylated diisopropyl fluorophosphate-inactivated human u-PA visualized by light or electron microscopy...

  6. Mpl traffics to the cell surface through conventional and unconventional routes.

    Science.gov (United States)

    Cleyrat, Cédric; Darehshouri, Anza; Steinkamp, Mara P; Vilaine, Mathias; Boassa, Daniela; Ellisman, Mark H; Hermouet, Sylvie; Wilson, Bridget S

    2014-09-01

    Myeloproliferative neoplasms (MPNs) are often characterized by JAK2 or calreticulin (CALR) mutations, indicating aberrant trafficking in pathogenesis. This study focuses on Mpl trafficking and Jak2 association using two model systems: human erythroleukemia cells (HEL; JAK2V617F) and K562 myeloid leukemia cells (JAK2WT). Consistent with a putative chaperone role for Jak2, Mpl and Jak2 associate on both intracellular and plasma membranes (shown by proximity ligation assay) and siRNA-mediated knockdown of Jak2 led to Mpl trapping in the endoplasmic reticulum (ER). Even in Jak2 sufficient cells, Mpl accumulates in punctate structures that partially colocalize with ER-tracker, the ER exit site marker (ERES) Sec31a, the autophagy marker LC3 and LAMP1. Mpl was fused to miniSOG, a genetically encoded tag for correlated light and electron microscopy. Results suggest that a fraction of Mpl is taken up into autophagic structures from the ER and routed to autolyososomes. Surface biotinylation shows that both immature and mature Mpl reach the cell surface; in K562 cells Mpl is also released in exosomes. Both forms rapidly internalize upon ligand addition, while recovery is primarily attributed to immature Mpl. Mpl appears to reach the plasma membrane via both conventional ER-Golgi and autolysosome secretory pathways, as well as recycling. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. A streptavidin linker layer that functions after drying.

    Science.gov (United States)

    Xia, Nan; Shumaker-Parry, Jennifer S; Zareie, M Hadi; Campbell, Charles T; Castner, David G

    2004-04-27

    The ability of streptavidin (SA) to simultaneously bind four biotins is often used in linker layers, where a biotinylated molecule is linked to a biotin-functionalized surface via SA. For biosensor and array applications, it is desirable that the SA linker layer be stable to drying and rehydration. In this study it was observed that a significant decrease in binding capacity of a SA layer occurred when that layer was dried. For this study a SA linker layer was constructed by binding SA to a biotin-containing alkylthiolate monolayer (BAT/OEG) self-assembled onto gold. Its stability after drying was investigated using surface plasmon resonance (SPR). Approximately a quarter of the SA layer was removed from the BAT/OEG surface upon drying and rehydration, suggesting disruption of SA-biotin binding when dry. This resulted in the dried SA layer losing approximately 40% of its biotinylated ferritin (BF) binding capacity. Coating the layer with trehalose before drying was found to inhibit the loss of SA from the BAT/OEG surface. SPR showed that the trehalose-protected SA linker layer retained approximately 91% of its original BF binding capacity after drying and rehydration. Atomic force microscopy, which was used to image individual surface-bound SA and BF molecules, qualitatively confirmed these observations.

  8. Proteomic plasma membrane profiling reveals an essential role for gp96 in the cell surface expression of LDLR family members, including the LDL receptor and LRP6.

    Science.gov (United States)

    Weekes, Michael P; Antrobus, Robin; Talbot, Suzanne; Hör, Simon; Simecek, Nikol; Smith, Duncan L; Bloor, Stuart; Randow, Felix; Lehner, Paul J

    2012-03-02

    The endoplasmic reticulum chaperone gp96 is required for the cell surface expression of a narrow range of proteins, including toll-like receptors (TLRs) and integrins. To identify a more comprehensive repertoire of proteins whose cell surface expression is dependent on gp96, we developed plasma membrane profiling (PMP), a technique that combines SILAC labeling with selective cell surface aminooxy-biotinylation. This approach allowed us to compare the relative abundance of plasma membrane (PM) proteins on gp96-deficient versus gp96-reconstituted murine pre-B cells. Analysis of unfractionated tryptic peptides initially identified 113 PM proteins, which extended to 706 PM proteins using peptide prefractionation. We confirmed a requirement for gp96 in the cell surface expression of certain TLRs and integrins and found a marked decrease in cell surface expression of four members of the extended LDL receptor family (LDLR, LRP6, Sorl1 and LRP8) in the absence of gp96. Other novel gp96 client proteins included CD180/Ly86, important in the B-cell response to lipopolysaccharide. We highlight common structural motifs in these client proteins that may be recognized by gp96, including the beta-propeller and leucine-rich repeat. This study therefore identifies the extended LDL receptor family as an important new family of proteins whose cell surface expression is regulated by gp96.

  9. Characterization of cleavage events in the multifunctional cilium adhesin Mhp684 (P146) reveals a mechanism by which Mycoplasma hyopneumoniae regulates surface topography.

    Science.gov (United States)

    Bogema, Daniel R; Deutscher, Ania T; Woolley, Lauren K; Seymour, Lisa M; Raymond, Benjamin B A; Tacchi, Jessica L; Padula, Matthew P; Dixon, Nicholas E; Minion, F Chris; Jenkins, Cheryl; Walker, Mark J; Djordjevic, Steven P

    2012-01-01

    Mycoplasma hyopneumoniae causes enormous economic losses to swine production worldwide by colonizing the ciliated epithelium in the porcine respiratory tract, resulting in widespread damage to the mucociliary escalator, prolonged inflammation, reduced weight gain, and secondary infections. Protein Mhp684 (P146) comprises 1,317 amino acids, and while the N-terminal 400 residues display significant sequence identity to the archetype cilium adhesin P97, the remainder of the molecule is novel and displays unusual motifs. Proteome analysis shows that P146 preprotein is endogenously cleaved into three major fragments identified here as P50(P146), P40(P146), and P85(P146) that reside on the cell surface. Liquid chromatography with tandem mass spectrometry (LC-MS/MS) identified a semitryptic peptide that delineated a major cleavage site in Mhp684. Cleavage occurred at the phenylalanine residue within sequence (672)ATEF↓QQ(677), consistent with a cleavage motif resembling S/T-X-F↓X-D/E recently identified in Mhp683 and other P97/P102 family members. Biotinylated surface proteins recovered by avidin chromatography and separated by two-dimensional gel electrophoresis (2-D GE) showed that more-extensive endoproteolytic cleavage of P146 occurs. Recombinant fragments F1(P146)-F3(P146) that mimic P50(P146), P40(P146), and P85(P146) were constructed and shown to bind porcine epithelial cilia and biotinylated heparin with physiologically relevant affinity. Recombinant versions of F3(P146) generated from M. hyopneumoniae strain J and 232 sequences strongly bind porcine plasminogen, and the removal of their respective C-terminal lysine and arginine residues significantly reduces this interaction. These data reveal that P146 is an extensively processed, multifunctional adhesin of M. hyopneumoniae. Extensive cleavage coupled with variable cleavage efficiency provides a mechanism by which M. hyopneumoniae regulates protein topography. Vaccines used to control Mycoplasma

  10. MHJ_0125 is an M42 glutamyl aminopeptidase that moonlights as a multifunctional adhesin on the surface of Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Robinson, Mark W; Buchtmann, Kyle A; Jenkins, Cheryl; Tacchi, Jessica L; Raymond, Benjamin B A; To, Joyce; Roy Chowdhury, Piklu; Woolley, Lauren K; Labbate, Maurizio; Turnbull, Lynne; Whitchurch, Cynthia B; Padula, Matthew P; Djordjevic, Steven P

    2013-04-17

    Bacterial aminopeptidases play important roles in pathogenesis by providing a source of amino acids from exogenous proteins, destroying host immunological effector peptides and executing posttranslational modification of bacterial and host proteins. We show that MHJ_0125 from the swine respiratory pathogen Mycoplasma hyopneumoniae represents a new member of the M42 class of bacterial aminopeptidases. Despite lacking a recognizable signal sequence, MHJ_0125 is detectable on the cell surface by fluorescence microscopy and LC-MS/MS of (i) biotinylated surface proteins captured by avidin chromatography and (ii) peptides released by mild trypsin shaving. Furthermore, surface-associated glutamyl aminopeptidase activity was detected by incubation of live M. hyopneumoniae cells with the diagnostic substrate H-Glu-AMC. MHJ_0125 moonlights as a multifunctional adhesin, binding to both heparin and plasminogen. Native proteomics and comparative modelling studies suggest MHJ_0125 forms a dodecameric, homopolymeric structure and provide insight into the positions of key residues that are predicted to interact with heparin and plasminogen. MHJ_0125 is the first aminopeptidase shown to both bind plasminogen and facilitate its activation by tissue plasminogen activator. Plasmin cleaves host extracellular matrix proteins and activates matrix metalloproteases, generating peptide substrates for MHJ_0125 and a source of amino acids for growth of M. hyopneumoniae. This unique interaction represents a new paradigm in microbial pathogenesis.

  11. Fulltext PDF

    Indian Academy of Sciences (India)

    SEARCHU

    Figure 1. GAF of erythrocytes belonging to young, intermediate and old age groups. Circulating erythrocytes were labelled by DIB tech- nique where the second step of low intensity biotinylation was performed, 25 days after the initial step high intensity biotinylation. After. 15 days of the second biotinylation step, erythrocytes ...

  12. Peptides for Specific Intracellular Delivery and Targeting of Nanoparticles: Implications for Developing Nanoparticle-Mediated Drug Delivery

    Science.gov (United States)

    2010-01-01

    less in size and span an array of compositions including met- als, semiconductor quantum dots (QDs), oxides, polymers , vesicles (e.g., micelles...93,98] Polymers Poly(lactic-co-glycolic acid), poly(amido amine) and dendrimers Biocompatibility, biodegradability and controlled release High drug...Secondary interactions Covalent atachment to surface ligand Thiolated peptide Biotinylated peptide Aminated peptide Sulfo-NHS Tetravalent streptavidin

  13. Ligand binding to G protein-coupled receptors in tethered cell membranes

    DEFF Research Database (Denmark)

    Martinez, Karen L.; Meyer, Bruno H.; Hovius, Ruud

    2003-01-01

    for the surface immobilization of membrane proteins was developed using the prototypic seven transmembrane neurokinin-1 receptor. The receptor was expressed as a biotinylated protein in mammalian cells. Membranes from cell homogenates were selectively immobilized on glass surfaces covered with streptavidin. TIRF...... measurements showed that a fluorescent agonist binds to the receptor on the sensor surface with similar affinity as to the receptor in live cells. This approach offers the possibility to investigate minute amounts of membrane protein in an active form and in its native environment without purification....

  14. Avidin-biotin system: a small library of cysteine biotinylated derivatives designed for the [99mTc(N)(PNP)]2+ metal fragment

    International Nuclear Information System (INIS)

    Bolzati, Cristina; Caporale, Andrea; Agostini, Stefania; Carta, Davide; Cavazza-Ceccato, Mario; Refosco, Fiorenzo; Tisato, Francesco; Schievano, Elisabetta; Bandoli, Giuliano

    2007-01-01

    Using the avidin-biotin system as model, we investigate here the effective application of [Tc(N)L(PNP)] +/0 technology (L=N-functionalized cysteine [O - ,S - ]; PNP=aminodiphosphine) to the preparation of target-specific radiopharmaceuticals. A series of 99m Tc-nitrido complexes containing functionalized biotin ligands was prepared and their biological profile was determined. To minimize the steric and the electronic influences of the Tc-carrying complex on the biotin-avidin receptor interaction, the following N-functionalized cysteine-biotin derivatives were synthesized: (1) Biot-CysOSH; (2) Biot-Abu-CysOSH; (3) Biot-Abz-CysOSH; (4) Biot-L-(Ac)Lys-CysOSH; (5) Biot-D-(Ac)Lys-CysOSH; (6) Biot-Glu-CysOSH. The asymmetrical nitrido-Tc(V) 99g/99m Tc(N)(Biot-X-CysOS)(PNP3) (X=spacer) complexes, where PNP3 was N,N-bis-[(dimethoxypropyl)phosphinoethyl] methoxy-ethylamine, were obtained by simultaneous addition of PNP3 and the relevant biotinylated ligand to a solution containing a 99m Tc-nitrido precursor (yields >95%). In all cases, a mixture of syn- and anti isomers was observed. In vitro challenge experiments with glutathione and cysteine indicated that no transchelation reactions occurred. Assessment of the in vitro binding to avidin of the complexes revealed that only the complexes containing Biot-Abu-CysOS and Biot-Glu-CysOS ligand maintained a good affinity for the concentrator. Stability studies carried out in human and mouse plasma as well as in rat and mouse liver homogenate evidenced a rapid enzymatic degradation for the 99m Tc(N)(Biot-Abu-CysOS)(PNP3) complex, whereas the 99m Tc(N)(Biot-Glu-CysOS)(PNP3) one was stable in all conditions. Tissue biodistribution in normal Balb/C mice of the most stable candidate showed a rapid clearance both from the blood and the other tissues. The activity was eliminated both through the hepatobiliary system and the urinary tract

  15. A role for host cell exocytosis in InlB-mediated internalisation of Listeria monocytogenes.

    Science.gov (United States)

    Van Ngo, Hoan; Bhalla, Manmeet; Chen, Da-Yuan; Ireton, Keith

    2017-11-01

    The bacterial surface protein InlB mediates internalisation of Listeria monocytogenes into human cells through interaction with the host receptor tyrosine kinase, Met. InlB-mediated entry requires localised polymerisation of the host actin cytoskeleton. Apart from actin polymerisation, roles for other host processes in Listeria entry are unknown. Here, we demonstrate that exocytosis in the human cell promotes InlB-dependent internalisation. Using a probe consisting of VAMP3 with an exofacial green fluorescent protein tag, focal exocytosis was detected during InlB-mediated entry. Exocytosis was dependent on Met tyrosine kinase activity and the GTPase RalA. Depletion of SNARE proteins by small interfering RNA demonstrated an important role for exocytosis in Listeria internalisation. Depletion of SNARE proteins failed to affect actin filaments during internalisation, suggesting that actin polymerisation and exocytosis are separable host responses. SNARE proteins were required for delivery of the human GTPase Dynamin 2, which promotes InlB-mediated entry. Our results identify exocytosis as a novel host process exploited by Listeria for infection. © 2017 John Wiley & Sons Ltd.

  16. Characterization of antigen association with accessory cells: specific removal of processed antigens from the cell surface by phospholipases

    International Nuclear Information System (INIS)

    Falo, L.D. Jr.; Haber, S.I.; Herrmann, S.; Benacerraf, B.; Rock, K.L.

    1987-01-01

    To characterize the basis for the cell surface association of processed antigen with the antigen-presenting cell (APC) the authors analyzed its sensitivity to enzymatic digestion. Antigen-exposed APC that are treated with phospholipase and then immediately fixed lose their ability to stimulate antigen-plus-Ia-specific T-T hybridomas. This effect is seen with highly purified phospholipase A 2 and phospholipase C. In addition it is observed with three distinct antigens - ovalbumin, bovine insulin, and poly(LGlu 56 LLys 35 LPhe 9 )[(GluLysPhe)/sub n/]. The effect of phospholipases is highly specific. Identically treated APC are equivalent to control in their ability to stimulate alloreactive hybridomas specific for precisely the same Ia molecule that is corecognized by antigen-plus-Ia-specific hybrids. Furthermore, the antigen-presenting function of enzyme-treated, fixed APC can be reconstituted by the addition of exogenous in vitro processed or processing independent antigens. In parallel studies 125 I-labeled avidin was shown to specifically bind to APC that were previously exposed and allowed to process biotin-insulin. Biotin-insulin-exposed APC that are pretreated with phospholipase bind significantly less 125 I-labeled avidin than do untreated, exposed APC. Identical enzyme treatment does not reduce the binding of avidin to a biotinylated antibody already bound to class II major histocompatibility complex molecules of APC. These studies demonstrate that phospholipase effectively removes processed cell surface antigen

  17. An investigation of interactions between hypocretin/orexin signaling and glutamate receptor surface expression in the rat nucleus accumbens under basal conditions and after cocaine exposure.

    Science.gov (United States)

    Plaza-Zabala, Ainhoa; Li, Xuan; Milovanovic, Mike; Loweth, Jessica A; Maldonado, Rafael; Berrendero, Fernando; Wolf, Marina E

    2013-12-17

    Hypocretin peptides are critical for the effects of cocaine on excitatory synaptic strength in the ventral tegmental area (VTA). However, little is known about their role in cocaine-induced synaptic plasticity in the nucleus accumbens (NAc). First, we tested whether hypocretin-1 by itself could acutely modulate glutamate receptor surface expression in the NAc, given that hypocretin-1 in the VTA reproduces cocaine's effects on glutamate transmission. We found no effect of hypocretin-1 infusion on AMPA or NMDA receptor surface expression in the NAc, measured by biotinylation, either 30 min or 3h after the infusion. Second, we were interested in whether changes in hypocretin receptor-2 (Hcrtr-2) expression contribute to cocaine-induced plasticity in the NAc. As a first step towards addressing this question, Hcrtr-2 surface expression was compared in the NAc after withdrawal from extended-access self-administration of saline (control) versus cocaine. We found that surface Hcrtr-2 levels remain unchanged following 14, 25 or 48 days of withdrawal from cocaine, a time period in which high conductance GluA2-lacking AMPA receptors progressively emerge in the NAc. Overall, our results fail to support a role for hypocretins in acute modulation of glutamate receptor levels in the NAc or a role for altered Hcrtr-2 expression in withdrawal-dependent synaptic adaptations in the NAc following cocaine self-administration. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  18. Differential biotin labelling of the cell envelope proteins in lipopolysaccharidic diderm bacteria: Exploring the proteosurfaceome of Escherichia coli using sulfo-NHS-SS-biotin and sulfo-NHS-PEG4-bismannose-SS-biotin.

    Science.gov (United States)

    Monteiro, Ricardo; Chafsey, Ingrid; Leroy, Sabine; Chambon, Christophe; Hébraud, Michel; Livrelli, Valérie; Pizza, Mariagrazia; Pezzicoli, Alfredo; Desvaux, Mickaël

    2018-06-15

    Surface proteins are the major factor for the interaction between bacteria and its environment, playing an important role in infection, colonisation, virulence and adaptation. However, the study of surface proteins has proven difficult mainly due to their hydrophobicity and/or relatively low abundance compared with cytoplasmic proteins. To overcome these issues new proteomic strategies have been developed, such as cell-surface protein labelling using biotinylation reagents. Sulfo-NHS-SS-biotin is the most commonly used reagent to investigate the proteins expressed at the cell surface of various organisms but its use in lipopolysaccharidic diderm bacteria (archetypical Gram-negative bacteria) remains limited to a handful of species. While generally pass over in silence, some periplasmic proteins, but also some inner membrane lipoproteins, integral membrane proteins and cytoplasmic proteins (cytoproteins) are systematically identified following this approach. To limit cell lysis and diffusion of the sulfo-NHS-SS-biotin through the outer membrane, biotin labelling was tested over short incubation times and proved to be as efficient for 1 min at room temperature. To further limit labelling of protein located below the outer membrane, the use of high-molecular weight sulfo-NHS-PEG4-bismannose-SS-biotin appeared to recover differentially cell-envelope proteins compared to low-molecular weight sulfo-NHS-SS-biotin. Actually, the sulfo-NHS-SS-biotin recovers at a higher extent the proteins completely or partly exposed in the periplasm than sulfo-NHS-PEG4-bismannose-SS-biotin, namely periplasmic and integral membrane proteins as well as inner membrane and outer membrane lipoproteins. These results highlight that protein labelling using biotinylation reagents of different sizes provides a sophisticated and accurate way to differentially explore the cell envelope proteome of lipopolysaccharidic diderm bacteria. While generally pass over in silence, some periplasmic proteins

  19. Targeting histone abnormality in triple negative breast cancer

    Science.gov (United States)

    2017-08-01

    mass spectrometry analysis. Briefly, we used the biotinylated primers to prepare longer double stranded biotinylated HDAC5 promoter probes...streptavidin-agarose bead suspension was added to a mixture of 400 µg of nuclear extract proteins and 4 µg of double- strand biotinylated oligonucleotides in...20 Wu Y, Wang Y, Yang XH, Kang T, Zhao Y, Wang C et al. The deubiquitinase USP28 stabilizes LSD1 and confers stem -cell-like traits to breast cancer

  20. Dendritic cell targeted chitosan nanoparticles for nasal DNA immunization against SARS CoV nucleocapsid protein.

    Science.gov (United States)

    Raghuwanshi, Dharmendra; Mishra, Vivek; Das, Dipankar; Kaur, Kamaljit; Suresh, Mavanur R

    2012-04-02

    This work investigates the formulation and in vivo efficacy of dendritic cell (DC) targeted plasmid DNA loaded biotinylated chitosan nanoparticles for nasal immunization against nucleocapsid (N) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) as antigen. The induction of antigen-specific mucosal and systemic immune response at the site of virus entry is a major challenge for vaccine design. Here, we designed a strategy for noninvasive receptor mediated gene delivery to nasal resident DCs. The pDNA loaded biotinylated chitosan nanoparticles were prepared using a complex coacervation process and characterized for size, shape, surface charge, plasmid DNA loading and protection against nuclease digestion. The pDNA loaded biotinylated chitosan nanoparticles were targeted with bifunctional fusion protein (bfFp) vector for achieving DC selective targeting. The bfFp is a recombinant fusion protein consisting of truncated core-streptavidin fused with anti-DEC-205 single chain antibody (scFv). The core-streptavidin arm of fusion protein binds with biotinylated nanoparticles, while anti-DEC-205 scFv imparts targeting specificity to DC DEC-205 receptor. We demonstrate that intranasal administration of bfFp targeted formulations along with anti-CD40 DC maturation stimuli enhanced magnitude of mucosal IgA as well as systemic IgG against N protein. The strategy led to the detection of augmented levels of N protein specific systemic IgG and nasal IgA antibodies. However, following intranasal delivery of naked pDNA no mucosal and systemic immune responses were detected. A parallel comparison of targeted formulations using intramuscular and intranasal routes showed that the intramuscular route is superior for induction of systemic IgG responses compared with the intranasal route. Our results suggest that targeted pDNA delivery through a noninvasive intranasal route can be a strategy for designing low-dose vaccines.

  1. Improving the Immunogenicity of the Mycobacterium bovis BCG Vaccine by Non-Genetic Bacterial Surface Decoration Using the Avidin-Biotin System.

    Directory of Open Access Journals (Sweden)

    Ting-Yu Angela Liao

    Full Text Available Current strategies to improve the current BCG vaccine attempt to over-express genes encoding specific M. tuberculosis (Mtb antigens and/or regulators of antigen presentation function, which indeed have the potential to reshape BCG in many ways. However, these approaches often face serious difficulties, in particular the efficiency and stability of gene expression via nucleic acid complementation and safety concerns associated with the introduction of exogenous DNA. As an alternative, we developed a novel non-genetic approach for rapid and efficient display of exogenous proteins on bacterial cell surface. The technology involves expression of proteins of interest in fusion with a mutant version of monomeric avidin that has the feature of reversible binding to biotin. Fusion proteins are then used to decorate the surface of biotinylated BCG. Surface coating of BCG with recombinant proteins was highly reproducible and stable. It also resisted to the freeze-drying shock routinely used in manufacturing conventional BCG. Modifications of BCG surface did not affect its growth in culture media neither its survival within the host cell. Macrophages phagocytized coated BCG bacteria, which efficiently delivered their surface cargo of avidin fusion proteins to MHC class I and class II antigen presentation compartments. Thereafter, chimeric proteins corresponding to a surrogate antigen derived from ovalbumin and the Mtb specific ESAT6 antigen were generated and tested for immunogenicity in vaccinated mice. We found that BCG displaying ovalbumin antigen induces an immune response with a magnitude similar to that induced by BCG genetically expressing the same surrogate antigen. We also found that BCG decorated with Mtb specific antigen ESAT6 successfully induces the expansion of specific T cell responses. This novel technology, therefore, represents a practical and effective alternative to DNA-based gene expression for upgrading the current BCG vaccine.

  2. Improving the Immunogenicity of the Mycobacterium bovis BCG Vaccine by Non-Genetic Bacterial Surface Decoration Using the Avidin-Biotin System.

    Science.gov (United States)

    Liao, Ting-Yu Angela; Lau, Alice; Joseph, Sunil; Hytönen, Vesa; Hmama, Zakaria

    2015-01-01

    Current strategies to improve the current BCG vaccine attempt to over-express genes encoding specific M. tuberculosis (Mtb) antigens and/or regulators of antigen presentation function, which indeed have the potential to reshape BCG in many ways. However, these approaches often face serious difficulties, in particular the efficiency and stability of gene expression via nucleic acid complementation and safety concerns associated with the introduction of exogenous DNA. As an alternative, we developed a novel non-genetic approach for rapid and efficient display of exogenous proteins on bacterial cell surface. The technology involves expression of proteins of interest in fusion with a mutant version of monomeric avidin that has the feature of reversible binding to biotin. Fusion proteins are then used to decorate the surface of biotinylated BCG. Surface coating of BCG with recombinant proteins was highly reproducible and stable. It also resisted to the freeze-drying shock routinely used in manufacturing conventional BCG. Modifications of BCG surface did not affect its growth in culture media neither its survival within the host cell. Macrophages phagocytized coated BCG bacteria, which efficiently delivered their surface cargo of avidin fusion proteins to MHC class I and class II antigen presentation compartments. Thereafter, chimeric proteins corresponding to a surrogate antigen derived from ovalbumin and the Mtb specific ESAT6 antigen were generated and tested for immunogenicity in vaccinated mice. We found that BCG displaying ovalbumin antigen induces an immune response with a magnitude similar to that induced by BCG genetically expressing the same surrogate antigen. We also found that BCG decorated with Mtb specific antigen ESAT6 successfully induces the expansion of specific T cell responses. This novel technology, therefore, represents a practical and effective alternative to DNA-based gene expression for upgrading the current BCG vaccine.

  3. Comparison of adenovirus fiber, protein IX, and hexon capsomeres as scaffolds for vector purification and cell targeting

    International Nuclear Information System (INIS)

    Campos, Samuel K.; Barry, Michael A.

    2006-01-01

    The direct genetic modification of adenoviral capsid proteins with new ligands is an attractive means to confer targeted tropism to adenoviral vectors. Although several capsid proteins have been reported to tolerate the genetic fusion of foreign peptides and proteins, direct comparison of cell targeting efficiencies through the different capsomeres has been lacking. Likewise, direct comparison of with one or multiple ligands has not been performed due to a lack of capsid-compatible ligands available for retargeting. Here we utilize a panel of metabolically biotinylated Ad vectors to directly compare targeted transduction through the fiber, protein IX, and hexon capsomeres using a variety of biotinylated ligands including antibodies, transferrin, EGF, and cholera toxin B. These results clearly demonstrate that cell targeting with a variety of high affinity receptor-binding ligands is only effective when transduction is redirected through the fiber protein. In contrast, protein IX and hexon-mediated targeting by the same set of ligands failed to mediate robust vector targeting, perhaps due to aberrant trafficking at the cell surface or inside targeted cells. These data suggest that vector targeting by genetic incorporation of high affinity ligands will likely be most efficient through modification of the adenovirus fiber rather than the protein IX and hexon capsomeres. In contrast, single-step monomeric avidin affinity purification of Ad vectors using the metabolic biotinylation system is most effective through capsomeres like protein IX and hexon

  4. Characterization, cell-surface expression and ligand-binding properties of different truncated N-terminal extracellular domains of the ionotropic glutamate receptor subunit GluR1.

    Science.gov (United States)

    McIlhinney, R A; Molnár, E

    1996-04-01

    To identify the location of the first transmembrane segment of the GluR1 glutamate receptor subunit artificial stop codons have been introduced into the N-terminal domain at amino acid positions 442, 510, and 563, namely just before and spanning the proposed first two transmembrane regions. The resultant truncated N-terminal fragments of GluR1, termed NT1, NT2, and NT3 respectively were expressed in Cos-7 cells and their cellular distribution and cell-surface expression analysed using an N-terminal antibody to GluR1. All of the fragments were fully glycosylated and were found to be associated with cell membranes but none was secreted. Differential extraction of the cell membranes indicated that both NT1 and NT2 behave as peripheral membrane proteins. In contrast NT3, like the full subunit, has integral membrane protein properties. Furthermore only NT3 is expressed at the cell surface as determined by immunofluorescence and cell-surface biotinylation. Protease protection assays indicated that only NT3 had a cytoplasmic tail. Binding studies using the selective ligand [(3)H]alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate ([(3)H]AMPA) demonstrated that NT3 does not bind ligand. Together these results indicate that the first transmembrane domain of the GluR1 subunit lies between residues 509 and 562, that the N-terminal domain alone cannot form a functional ligand-binding site and that this domain can be targeted to the cell surface provided that it has a transmembrane-spanning region.

  5. Hemiuroid trematode sporocysts are undetected by hemocytes of their intermediate host, the ark cockle Anadara trapezia: potential role of surface carbohydrates in successful parasitism.

    Science.gov (United States)

    Kawasaki, Minami; Delamare-Deboutteville, Jerome; Dang, Cecile; Barnes, Andrew C

    2013-12-01

    In order to establish a successful relationship with their hosts, parasites must subvert or evade immune defences. Cockle Anadara trapezia and Sydney Rock oyster (SRO) Saccostrea glomerata live in the same location but only ark cockles are infected by sporocysts of hemiuroid trematode. This provides an opportunity to explore differing interactions between the parasite and the immune system of susceptible and refractive hosts. Rapid migration and encapsulation of sporocysts was observed by SRO hemocytes but not by cockle hemocytes. This migration/encapsulation was inhibited by N-acetylglucosamine or N-acetylgalactosamine but not by the other sugars, implicating specific surface carbohydrates in immune detection. Effector responses of hemocytes were investigated in vitro in terms of production of reactive oxygen production (ROS). Hemocytes of both species strongly reacted to Zymosan, but only SRO hemocytes responded to live sporocysts. Neither species' hemocytes produced ROS in the presence of dead/fixed sporocysts, and there was no suppression of Zymosan-induced respiratory burst by sporocysts. This suggests that immune escape is mediated by avoiding encapsulation, perhaps through molecular mimicry. Membrane-shaving with proteases indicated that sporocyst surface proteins are not a key factors in hemocytic detection. Surface carbohydrates of SRO and cockle hemocytes and of sporocysts were profiled with a panel of biotinylated lectins. This revealed substantial differences between cockle and SRO hemocytes, but greater similarity between cockle hemocytes and sporocysts. Results suggest that surface carbohydrates play an integral role in hemocyte immunorecognition and that surface carbohydrate molecular mimicry is a potential strategy for immune evasion in cockles by hemiuroid trematode sporocysts. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

  6. Enzymatic shaving of the tegument surface of live schistosomes for proteomic analysis: a rational approach to select vaccine candidates.

    Directory of Open Access Journals (Sweden)

    William Castro-Borges

    2011-03-01

    Full Text Available The membrane-associated and membrane-spanning constituents of the Schistosoma mansoni tegument surface, the parasite's principal interface with the host bloodstream, have recently been characterized using proteomic techniques. Biotinylation of live worms using membrane-impermeant probes revealed that only a small subset of the proteins was accessible to the reagents. Their position within the multilayered architecture of the surface has not been ascertained.An enzymatic shaving approach on live worms has now been used to release the most accessible components, for analysis by MS/MS. Treatment with trypsin, or phosphatidylinositol-specific phospholipase C (PiPLC, only minimally impaired membrane integrity. PiPLC-enriched proteins were distinguished from those released in parasite vomitus or by handling damage, using isobaric tagging. Trypsin released five membrane proteins, Sm200, Sm25 and three annexins, plus host CD44 and the complement factors C3 and C4. Nutrient transporters and ion channels were absent from the trypsin fraction, suggesting a deeper location in the surface complex; surprisingly, two BAR-domain containing proteins were released. Seven parasite and two host proteins were enriched by PiPLC treatment, the vaccine candidate Sm29 being the most prominent along with two orthologues of human CD59, potentially inhibitors of complement fixation. The enzymes carbonic anhydrase and APD-ribosyl cyclase were also enriched, plus Sm200 and alkaline phosphatase. Host GPI-anchored proteins CD48 and CD90, suggest 'surface painting' during worm peregrination in the portal system.Our findings suggest that the membranocalyx secreted over the tegument surface is not the inert barrier previously proposed, some tegument proteins being externally accessible to enzymes and thus potentially located within it. Furthermore, the detection of C3 and C4 indicates that the complement cascade is initiated, while two CD59 orthologues suggest a potential

  7. Ligand-receptor assay for evaluation of functional activity of human recombinant VEGF and VEGFR-1 extracellular fragment.

    Science.gov (United States)

    Leopol'd, A V; Baklaushev, V P; Korchagina, A A; Shein, S A; Grinenko, N F; Pavlov, K A; Ryabukhin, I A; Chekhonin, V P

    2012-04-01

    cDNA encoding VEGF and Ig-like extracellular domains 2-4 of VEGFR-1 (sFlt-1(2-4)) were cloned into prokaryotic expression vectors pET32a and pQE60. Recombinant proteins were purified (metal affinity chromatography) and renatured. Chemiluminescent study for the interaction of recombinant VEGF and sFlt-1(2-4) showed that biotinylated VEGF specifically binds to the polystyrene-immobilized receptor extracellular fragment. Biotinylated recombinant sFlt-1 interacts with immobilized VEGF. Analysis of the interaction of immobilized recombinant VEGFR-1 and VEGF with C6 glioma cells labeled with CFDA-SE (vital fluorescent dye) showed that recombinant VEGFR-1 also binds to native membrane-associated VEGF. Recombinant VEGF was shown to bind to specific receptors expressed on the surface of C6 glioma cells. Functional activity of these proteins was confirmed by ligand-receptor assay for VEGF and VEGFR-1 (sFlt-1) and quantitative chemiluminescent detection.

  8. Single domain antibody–quantum dot conjugates for ricin detection by both fluoroimmunoassay and surface plasmon resonance

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, George P. [Center for Bio/Molecular Science and Engineering, Code 6900, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States); Glaven, Richard H. [Nova Research, Inc., 1900 Elkin Street, Suite 230, Alexandria, VA 22308 (United States); Algar, W. Russ [Center for Bio/Molecular Science and Engineering, Code 6900, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States); College of Science, George Mason University, Fairfax, VA 22030 (United States); Susumu, Kimihiro [Optical Sciences Division, Code 5600, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States); Sotera Defense Solutions, Annapolis Junction, MD 20701 (United States); Stewart, Michael H. [Optical Sciences Division, Code 5600, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States); Medintz, Igor L. [Center for Bio/Molecular Science and Engineering, Code 6900, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States); Goldman, Ellen R., E-mail: ellen.goldman@nrl.navy.mil [Center for Bio/Molecular Science and Engineering, Code 6900, U. S. Naval Research Laboratory, 4555 Overlook Ave. S.W., Washington, DC 20375 (United States)

    2013-07-05

    Graphical abstract: -- Highlights: •Anti-ricin single domain antibodies (sdAb) were self-assembled on quantum dots (QDs). •Conjugates were prepared using dihydrolipoic acid-capped CdSe–ZnS core–shell QDs. •The sdAb–QD conjugates functioned in fluoroimmunoassays for ricin detection. •The conjugates provided signal amplification in surface plasmon resonance assays. •Conjugates provided sensitive detection compared to unconjugated sdAb reporters. -- Abstract: The combination of stable biorecognition elements and robust quantum dots (QDs) has the potential to yield highly effective reporters for bioanalyses. Llama-derived single domain antibodies (sdAb) provide small thermostable recognition elements that can be easily manipulated using standard DNA methods. The sdAb was self-assembled on dihydrolipoic acid (DHLA) ligand-capped CdSe–ZnS core–shell QDs made in our laboratory through the polyhistidine tail of the protein, which coordinated to zinc ions on the QD surface. The sdAb–QD bioconjugates were then applied in both fluorometric and surface plasmon resonance (SPR) immunoassays for the detection of ricin, a potential biothreat agent. The sdAb–QD conjugates functioned in fluoroimmunoassays for the detection of ricin, providing equivalent limits of detection when compared to the same anti-ricin sdAb labeled with a conventional fluorophore. In addition, the DHLA-QD–sdAb conjugates were very effective reporter elements in SPR sandwich assays, providing more sensitive detection with a signal enhancement of ∼10-fold over sdAb reporters and 2–4 fold over full sized antibody reporters. Commercially prepared streptavidin-modified polymer-coated QDs also amplified the SPR signal for the detection of ricin when applied to locations where biotinylated anti-ricin sdAb was bound to target; however, we observed a 4-fold greater amplification when using the DHLA-QD–sdAb conjugates in this format.

  9. Single domain antibody–quantum dot conjugates for ricin detection by both fluoroimmunoassay and surface plasmon resonance

    International Nuclear Information System (INIS)

    Anderson, George P.; Glaven, Richard H.; Algar, W. Russ; Susumu, Kimihiro; Stewart, Michael H.; Medintz, Igor L.; Goldman, Ellen R.

    2013-01-01

    Graphical abstract: -- Highlights: •Anti-ricin single domain antibodies (sdAb) were self-assembled on quantum dots (QDs). •Conjugates were prepared using dihydrolipoic acid-capped CdSe–ZnS core–shell QDs. •The sdAb–QD conjugates functioned in fluoroimmunoassays for ricin detection. •The conjugates provided signal amplification in surface plasmon resonance assays. •Conjugates provided sensitive detection compared to unconjugated sdAb reporters. -- Abstract: The combination of stable biorecognition elements and robust quantum dots (QDs) has the potential to yield highly effective reporters for bioanalyses. Llama-derived single domain antibodies (sdAb) provide small thermostable recognition elements that can be easily manipulated using standard DNA methods. The sdAb was self-assembled on dihydrolipoic acid (DHLA) ligand-capped CdSe–ZnS core–shell QDs made in our laboratory through the polyhistidine tail of the protein, which coordinated to zinc ions on the QD surface. The sdAb–QD bioconjugates were then applied in both fluorometric and surface plasmon resonance (SPR) immunoassays for the detection of ricin, a potential biothreat agent. The sdAb–QD conjugates functioned in fluoroimmunoassays for the detection of ricin, providing equivalent limits of detection when compared to the same anti-ricin sdAb labeled with a conventional fluorophore. In addition, the DHLA-QD–sdAb conjugates were very effective reporter elements in SPR sandwich assays, providing more sensitive detection with a signal enhancement of ∼10-fold over sdAb reporters and 2–4 fold over full sized antibody reporters. Commercially prepared streptavidin-modified polymer-coated QDs also amplified the SPR signal for the detection of ricin when applied to locations where biotinylated anti-ricin sdAb was bound to target; however, we observed a 4-fold greater amplification when using the DHLA-QD–sdAb conjugates in this format

  10. International Workshop on Structural and Functional Aspects of the Cholinergic Synapse Held in Jerusalem, Isreal on 30 August-4 September 1987

    Science.gov (United States)

    1987-09-01

    16 vesaaicol analogues and 9 other compounds of widely varying structures. Only for a weakly binding diner of vesamicol, hexsmethonium and chloroquine ...surface of the vesicle menbrane by use cf a Lambrars-impermaant analogue of vesamicol. Rate processes for association and diissociation of vesamicol...vesamicol analogue occupied the receptor and inhibited ( 14 C]ACh transport. Addition of excess avidin "neutralized’ the biotinylated veasai:ol end freed the

  11. Surface force analysis of molecular interfacial interactions of proteins and lipids with polymeric biomaterials

    International Nuclear Information System (INIS)

    Hamilton-Brown, P.; Griesser, H.J.; Meagher, L.

    2001-01-01

    also performed with a biotinylated PEO coating in order to access the biospecific, high-affinity binding capability of biotin. AFM surface force measurements have been shown to enable characterisation of the range and magnitude of interfacial forces on various surfaces, and to establish how, for instance, the density of PEO coatings affects the forces involved and thereby lipid and protein adsorption. The resulting data provide I molecular information towards tailoring PEO and other polymer coatings for rational control of interfacial behaviour of biomedical devices in biological media. Copyright (2001) Australasian College of Physical Scientists and Engineers in Medicine

  12. Bovine Pancreatic Trypsin Inhibitor-Trypsin Complex as a Detection System for Recombinant Proteins

    Science.gov (United States)

    Borjigin, Jimo; Nathans, Jeremy

    1993-01-01

    Bovine pancreatic trypsin inhibitor (BPTI) binds to trypsin and anhydrotrypsin (an enzymatically inactive derivative of trypsin) with affinities of 6 x 10-14 and 1.1 x 10-13 M, respectively. We have taken advantage of the high affinity and specificity of this binding reaction to develop a protein tagging system in which biotinylated trypsin or biotinylated anhydrotrypsin is used as the reagent to detect recombinant fusion proteins into which BPTI has been inserted. Two proteins, opsin and growth hormone, were used as targets for insertional mutagenesis with BPTI. In each case, both domains of the fusion protein appear to be correctly folded. The fusion proteins can be specifically and efficiently detected by biotinylated trypsin or biotinylated anhydrotrypsin, as demonstrated by staining of transfected cells, protein blotting, affinity purification, and a mobility shift assay in SDS/polyacrylamide gels.

  13. Cytoadherence and sequestration in Plasmodium falciparum: defining the ties that bind.

    Science.gov (United States)

    Sherman, Irwin W; Eda, Shigetoshi; Winograd, Enrique

    2003-08-01

    Infected erythrocytes containing the more mature stages of the human malaria Plasmodium falciparum may adhere to endothelial cells and uninfected red cells. These phenomena, called sequestration and rosetting, respectively, are involved in both host pathogenesis and parasite survival. This review provides a critical summary of recent advances in the characterization of the molecules of the infected red blood cell involved in adhesion, i.e. parasite-encoded molecules (PfEMP1, MESA, rifins, stevor, clag 9, histidine-rich protein), a modified host membrane protein (band 3) and exofacial exposure of phosphatidylserine, as well as receptors on the endothelium, i.e. thrombospondin, CD36, ICAM-1 (intercellular adhesion molecule), and chondroitin sulfate.

  14. Interaction of Mycobacterium leprae with human airway epithelial cells: adherence, entry, survival, and identification of potential adhesins by surface proteome analysis.

    Science.gov (United States)

    Silva, Carlos A M; Danelishvili, Lia; McNamara, Michael; Berredo-Pinho, Márcia; Bildfell, Robert; Biet, Franck; Rodrigues, Luciana S; Oliveira, Albanita V; Bermudez, Luiz E; Pessolani, Maria C V

    2013-07-01

    This study examined the in vitro interaction between Mycobacterium leprae, the causative agent of leprosy, and human alveolar and nasal epithelial cells, demonstrating that M. leprae can enter both cell types and that both are capable of sustaining bacterial survival. Moreover, delivery of M. leprae to the nasal septum of mice resulted in macrophage and epithelial cell infection in the lung tissue, sustaining the idea that the airways constitute an important M. leprae entry route into the human body. Since critical aspects in understanding the mechanisms of infection are the identification and characterization of the adhesins involved in pathogen-host cell interaction, the nude mouse-derived M. leprae cell surface-exposed proteome was studied to uncover potentially relevant adhesin candidates. A total of 279 cell surface-exposed proteins were identified based on selective biotinylation, streptavidin-affinity purification, and shotgun mass spectrometry; 11 of those proteins have been previously described as potential adhesins. In vitro assays with the recombinant forms of the histone-like protein (Hlp) and the heparin-binding hemagglutinin (HBHA), considered to be major mycobacterial adhesins, confirmed their capacity to promote bacterial attachment to epithelial cells. Taking our data together, they suggest that the airway epithelium may act as a reservoir and/or portal of entry for M. leprae in humans. Moreover, our report sheds light on the potentially critical adhesins involved in M. leprae-epithelial cell interaction that may be useful in designing more effective tools for leprosy control.

  15. Chronic zinc exposure decreases the surface expression of NR2A-containing NMDA receptors in cultured hippocampal neurons.

    Directory of Open Access Journals (Sweden)

    Jia Zhu

    Full Text Available Zinc distributes widely in the central nervous system, especially in the hippocampus, amygdala and cortex. The dynamic balance of zinc is critical for neuronal functions. Zinc modulates the activity of N-methyl-D-aspartate receptors (NMDARs through the direct inhibition and various intracellular signaling pathways. Abnormal NMDAR activities have been implicated in the aetiology of many brain diseases. Sustained zinc accumulation in the extracellular fluid is known to link to pathological conditions. However, the mechanism linking this chronic zinc exposure and NMDAR dysfunction is poorly understood.We reported that chronic zinc exposure reduced the numbers of NR1 and NR2A clusters in cultured hippocampal pyramidal neurons. Whole-cell and synaptic NR2A-mediated currents also decreased. By contrast, zinc did not affect NR2B, suggesting that chronic zinc exposure specifically influences NR2A-containg NMDARs. Surface biotinylation indicated that zinc exposure attenuated the membrane expression of NR1 and NR2A, which might arise from to the dissociation of the NR2A-PSD-95-Src complex.Chronic zinc exposure perturbs the interaction of NR2A to PSD-95 and causes the disorder of NMDARs in hippocampal neurons, suggesting a novel action of zinc distinct from its acute effects on NMDAR activity.

  16. Biotin-tagged platinum(iv) complexes as targeted cytostatic agents against breast cancer cells.

    Science.gov (United States)

    Muhammad, Nafees; Sadia, Nasreen; Zhu, Chengcheng; Luo, Cheng; Guo, Zijian; Wang, Xiaoyong

    2017-09-05

    A biotin-guided platinum IV complex is highly cytotoxic against breast cancer cells but hypotoxic against mammary epithelial cells. The mono-biotinylated Pt IV complex is superior to the di-biotinylated one and hence a promising drug candidate for the targeted therapy of breast cancer.

  17. Relationship between molecular structure and supramolecular morphology of DODA-EO2-biotin and related lipids

    NARCIS (Netherlands)

    Huetz, P.; van Neuren, S.; Ringler, P.; Kremer, F.; van Breemen, J.F.L.; Wagenaar, A.; Engberts, J.B.F.N.; Fraaije, J.G E M; Brisson, A.

    1997-01-01

    We have recently reported that a biotinylated lipid molecule, called DODA-EO2-biotin, forms tubular lipid structures upon hydration, which act as a matrix for the formation of ordered helical arrays of streptavidin as well as for secondary macromolecular recognition reactions involving biotinylated

  18. Applying biotin-streptavidin binding for iscom (immunostimulating complex) association of recombinant immunogens.

    Science.gov (United States)

    Wikman, Maria; Friedman, Mikaela; Pinitkiatisakul, Sunan; Hemphill, Andrew; Lövgren-Bengtsson, Karin; Lundén, Anna; Ståhl, Stefan

    2005-04-01

    We have previously reported strategies for Escherichia coli production of recombinant immunogens fused to hydrophobic peptide or lipid tags to improve their capacity to be incorporated into an adjuvant formulation. In the present study, we have explored the strong interaction between biotin and SA (streptavidin) (K(D) approximately 10(-15) M) to couple recombinant immunogens to iscoms (immunostimulating complexes). Two different concepts were evaluated. In the first concept, a His(6)-tagged SA fusion protein (His(6)-SA) was bound to Ni(2+)-loaded iscom matrix (iscom without associated protein), and biotinylated immunogens were thereafter associated with the SA-coated iscoms. The immunogens were either biotinylated in vivo on E. coli expression or double biotinylated in vivo and in vitro. In the second concept, the recombinant immunogens were expressed as SA fusion proteins, which were directly bound to a biotinylated iscom matrix. A 53-amino-acid malaria peptide (M5), derived from the central repeat region of the Plasmodium falciparum blood-stage antigen Pf155/RESA, and a 232-amino-acid segment (SRS2') from the central region (from Pro-97 to Lys-328) of the major surface antigen NcSRS2 of the protozoan parasite Neospora caninum, served as model immunogens in the present study. All fusion proteins generated were found to be efficiently expressed and could be recovered to high purity using affinity chromatography. The association between the different immunogen-containing fusion proteins and the corresponding iscom matrix was demonstrated by analytical ultracentrifugation in a sucrose density gradient. However, some fusion proteins were, to a certain extent, also found to associate unspecifically with a regular iscom matrix. Furthermore, selected iscom fractions were demonstrated to induce high-titre antigen-specific antibody responses on immunization of mice. For the particular target immunogen SRS2', the induced antibodies demonstrated reactivity to the native

  19. Lectins binding during alloxan-induced diabetes in rat soleus muscle

    African Journals Online (AJOL)

    Membrane structural changes of soleus muscle of alloxan-diabetic rats were detected with a panel of six biotinylated lectins. Samples of muscles were obtained from normal and diabetic rats. The biotinylated lectins in staining were detected by avidin-peroxidase complex. Lectin stainning of soleus muscle cryostat sections ...

  20. Function of the SpoVAEa and SpoVAF Proteins of Bacillus subtilis Spores

    Science.gov (United States)

    2014-06-01

    outer surface of the spore’s inner membrane, as SpoVAEa was accessible to an external biotinylation agent in spores and SpoVAEa disappeared in parallel...codon was PCR amplified from PS832 chromosomal DNA with primers that inserted BamHI and PstI restriction sites upstream and downstream, respectively... chromosomal structure, and this strain was termed PS4348 (spoVAEa mutant). A B. subtilis strain with a deletion of the spoVF gene was constructed by a two

  1. Detection of fumonisin b1 and ochratoxin a in grain products using microsphere-based fluid array immunoassays.

    Science.gov (United States)

    Anderson, George P; Kowtha, Vasudha A; Taitt, Chris R

    2010-02-01

    Grain products are a staple of diets worldwide and therefore, the ability to accurately and efficiently detect foodborne contaminants such as mycotoxins is of importance to everyone. Here we describe an indirect competitive fluid array fluoroimmunoassay to quantify the mycotoxins, fumonisin B1 and ochratoxin A. Both toxins were immobilized to the surface of microspheres using a variety of intermediate molecules and binding of biotinylated "tracer" antibody tracers determined through flow cytometry using streptavidin-phycoerythrin conjugates and the Luminex100 flow cytometer. Competitive assays were developed where the binding of biotinylated monoclonal antibodies to fumonisin B and ochratoxin A was competitively inhibited by different concentrations of those toxins in solution. Concentrations of fumonisin giving 50% inhibition were 300 pg/mL in buffer, 100 ng/g in spiked oats, and 1 μg/g in spiked cornmeal; analogous concentrations for ochratoxin A were 30 ng/mL in buffer, 30 ng/g in spiked oats, and 10 ng/g in spiked corn. The future challenge will be to expand the number of mycotoxins tested both individually and in multiplexed format using this platform.

  2. Combinatorial chemoenzymatic synthesis and high-throughput screening of sialosides.

    Science.gov (United States)

    Chokhawala, Harshal A; Huang, Shengshu; Lau, Kam; Yu, Hai; Cheng, Jiansong; Thon, Vireak; Hurtado-Ziola, Nancy; Guerrero, Juan A; Varki, Ajit; Chen, Xi

    2008-09-19

    Although the vital roles of structures containing sialic acid in biomolecular recognition are well documented, limited information is available on how sialic acid structural modifications, sialyl linkages, and the underlying glycan structures affect the binding or the activity of sialic acid-recognizing proteins and related downstream biological processes. A novel combinatorial chemoenzymatic method has been developed for the highly efficient synthesis of biotinylated sialosides containing different sialic acid structures and different underlying glycans in 96-well plates from biotinylated sialyltransferase acceptors and sialic acid precursors. By transferring the reaction mixtures to NeutrAvidin-coated plates and assaying for the yields of enzymatic reactions using lectins recognizing sialyltransferase acceptors but not the sialylated products, the biotinylated sialoside products can be directly used, without purification, for high-throughput screening to quickly identify the ligand specificity of sialic acid-binding proteins. For a proof-of-principle experiment, 72 biotinylated alpha2,6-linked sialosides were synthesized in 96-well plates from 4 biotinylated sialyltransferase acceptors and 18 sialic acid precursors using a one-pot three-enzyme system. High-throughput screening assays performed in NeutrAvidin-coated microtiter plates show that whereas Sambucus nigra Lectin binds to alpha2,6-linked sialosides with high promiscuity, human Siglec-2 (CD22) is highly selective for a number of sialic acid structures and the underlying glycans in its sialoside ligands.

  3. Biomolecular detection using a metal semiconductor field effect transistor

    Science.gov (United States)

    Estephan, Elias; Saab, Marie-Belle; Buzatu, Petre; Aulombard, Roger; Cuisinier, Frédéric J. G.; Gergely, Csilla; Cloitre, Thierry

    2010-04-01

    In this work, our attention was drawn towards developing affinity-based electrical biosensors, using a MESFET (Metal Semiconductor Field Effect Transistor). Semiconductor (SC) surfaces must be prepared before the incubations with biomolecules. The peptides route was adapted to exceed and bypass the limits revealed by other types of surface modification due to the unwanted unspecific interactions. As these peptides reveal specific recognition of materials, then controlled functionalization can be achieved. Peptides were produced by phage display technology using a library of M13 bacteriophage. After several rounds of bio-panning, the phages presenting affinities for GaAs SC were isolated; the DNA of these specific phages were sequenced, and the peptide with the highest affinity was synthesized and biotinylated. To explore the possibility of electrical detection, the MESFET fabricated with the GaAs SC were used to detect the streptavidin via the biotinylated peptide in the presence of the bovine Serum Albumin. After each surface modification step, the IDS (current between the drain and the source) of the transistor was measured and a decrease in the intensity was detected. Furthermore, fluorescent microscopy was used in order to prove the specificity of this peptide and the specific localisation of biomolecules. In conclusion, the feasibility of producing an electrical biosensor using a MESFET has been demonstrated. Controlled placement, specific localization and detection of biomolecules on a MESFET transistor were achieved without covering the drain and the source. This method of functionalization and detection can be of great utility for biosensing application opening a new way for developing bioFETs (Biomolecular Field-Effect Transistor).

  4. Comparative analysis of the surface exposed proteome of two canine osteosarcoma cell lines and normal canine osteoblasts.

    Science.gov (United States)

    Milovancev, Milan; Hilgart-Martiszus, Ian; McNamara, Michael J; Goodall, Cheri P; Seguin, Bernard; Bracha, Shay; Wickramasekara, Samanthi I

    2013-06-13

    Osteosarcoma (OSA) is the most common primary bone tumor of dogs and carries a poor prognosis despite aggressive treatment. An improved understanding of the biology of OSA is critically needed to allow for development of novel diagnostic, prognostic, and therapeutic tools. The surface-exposed proteome (SEP) of a cancerous cell includes a multifarious array of proteins critical to cellular processes such as proliferation, migration, adhesion, and inter-cellular communication. The specific aim of this study was to define a SEP profile of two validated canine OSA cell lines and a normal canine osteoblast cell line utilizing a biotinylation/streptavidin system to selectively label, purify, and identify surface-exposed proteins by mass spectrometry (MS) analysis. Additionally, we sought to validate a subset of our MS-based observations via quantitative real-time PCR, Western blot and semi-quantitative immunocytochemistry. Our hypothesis was that MS would detect differences in the SEP composition between the OSA and the normal osteoblast cells. Shotgun MS identified 133 putative surface proteins when output from all samples were combined, with good consistency between biological replicates. Eleven of the MS-detected proteins underwent analysis of gene expression by PCR, all of which were actively transcribed, but varied in expression level. Western blot of whole cell lysates from all three cell lines was effective for Thrombospondin-1, CYR61 and CD44, and indicated that all three proteins were present in each cell line. Semi-quantitative immunofluorescence indicated that CD44 was expressed at much higher levels on the surface of the OSA than the normal osteoblast cell lines. The results of the present study identified numerous differences, and similarities, in the SEP of canine OSA cell lines and normal canine osteoblasts. The PCR, Western blot, and immunocytochemistry results, for the subset of proteins evaluated, were generally supportive of the mass spectrometry data

  5. Development of an enzyme-linked immunosorbent assay (ELISA)-like fluorescence assay to investigate the interactions of glycosaminoglycans to cells

    Energy Technology Data Exchange (ETDEWEB)

    Boucas, Rodrigo Ippolito [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Trindade, Edvaldo S. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Departamento de Biologia Celular, Universidade Federal do Parana, Curitiba, Parana (Brazil); Tersariol, Ivarne L.S. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Centro Interdisciplinar de Investigacao Bioquimica, Universidade de Mogi das Cruzes, Mogi das Cruzes, SP (Brazil); Dietrich, Carl P. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil); Nader, Helena B. [Disciplina de Biologia Molecular, Departamento de Bioquimica, Universidade Federal de Sao Paulo, SP (Brazil)], E-mail: hbnader.bioq@epm.br

    2008-06-23

    Sulfated glycosaminoglycans were labeled with biotin to study their interaction with cells in culture. Thus, heparin, heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate and dermatan sulfate were labeled using biotin-hydrazide, under different conditions. The structural characteristics of the biotinylated products were determined by chemical (molar ratios of hexosamine, uronic acid, sulfate and biotin) and enzymatic methods (susceptibility to degradation by chondroitinases and heparitinases). The binding of biotinylated glycosaminoglycans was investigated both in endothelial and smooth muscle cells in culture, using a novel time resolved fluorometric method based on interaction of europium-labeled streptavidin with the biotin covalently linked to the compounds. The interactions of glycosaminoglycans were saturable and number of binding sites could be obtained for each individual compound. The apparent dissociation constant varied among the different glycosaminoglycans and between the two cell lines. The interactions of the biotinylated glycosaminoglycans with the cells were also evaluated using confocal microscopy. We propose a convenient and reliable method for the preparation of biotinylated glycosaminoglycans, as well as a sensitive non-competitive fluorescence-based assay for studies of the interactions and binding of these compounds to cells in culture.

  6. Development of an enzyme-linked immunosorbent assay (ELISA)-like fluorescence assay to investigate the interactions of glycosaminoglycans to cells

    International Nuclear Information System (INIS)

    Boucas, Rodrigo Ippolito; Trindade, Edvaldo S.; Tersariol, Ivarne L.S.; Dietrich, Carl P.; Nader, Helena B.

    2008-01-01

    Sulfated glycosaminoglycans were labeled with biotin to study their interaction with cells in culture. Thus, heparin, heparan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate and dermatan sulfate were labeled using biotin-hydrazide, under different conditions. The structural characteristics of the biotinylated products were determined by chemical (molar ratios of hexosamine, uronic acid, sulfate and biotin) and enzymatic methods (susceptibility to degradation by chondroitinases and heparitinases). The binding of biotinylated glycosaminoglycans was investigated both in endothelial and smooth muscle cells in culture, using a novel time resolved fluorometric method based on interaction of europium-labeled streptavidin with the biotin covalently linked to the compounds. The interactions of glycosaminoglycans were saturable and number of binding sites could be obtained for each individual compound. The apparent dissociation constant varied among the different glycosaminoglycans and between the two cell lines. The interactions of the biotinylated glycosaminoglycans with the cells were also evaluated using confocal microscopy. We propose a convenient and reliable method for the preparation of biotinylated glycosaminoglycans, as well as a sensitive non-competitive fluorescence-based assay for studies of the interactions and binding of these compounds to cells in culture

  7. An impedimetric study of DNA hybridization on paper-supported inkjet-printed gold electrodes

    International Nuclear Information System (INIS)

    Ihalainen, Petri; Määttänen, Anni; Peltonen, Jouko; Pettersson, Fredrik; Pesonen, Markus; Österbacka, Ronald; Viitala, Tapani

    2014-01-01

    In this study, two different supramolecular recognition architectures for impedimetric detection of DNA hybridization have been formed on disposable paper-supported inkjet-printed gold electrodes. The gold electrodes were fabricated using a gold nanoparticle based ink. The first recognition architecture consists of subsequent layers of biotinylated self-assembly monolayer (SAM), streptavidin and biotinylated DNA probe. The other recognition architecture is constructed by immobilization of thiol-functionalized DNA probe (HS-DNA) and subsequent backfill with 11-mercapto-1-undecanol (MUOH) SAM. The binding capacity and selectivity of the recognition architectures were examined by surface plasmon resonance (SPR) measurements. SPR results showed that the HS-DNA/MUOH system had a higher binding capacity for the complementary DNA target. Electrochemical impedance spectroscopy (EIS) measurements showed that the hybridization can be detected with impedimetric spectroscopy in picomol range for both systems. EIS signal indicated a good selectivity for both recognition architectures, whereas SPR showed very high unspecific binding for the HS-DNA/MUOH system. The factors affecting the impedance signal were interpreted in terms of the complexity of the supramolecular architecture. The more complex architecture acts as a less ideal capacitive sensor and the impedance signal is dominated by the resistive elements. (paper)

  8. Overexpression of biotin synthase and biotin ligase is required for efficient generation of sulfur-35 labeled biotin in E. coli.

    Science.gov (United States)

    Delli-Bovi, Teegan A; Spalding, Maroya D; Prigge, Sean T

    2010-10-11

    Biotin is an essential enzyme cofactor that acts as a CO2 carrier in carboxylation and decarboxylation reactions. The E. coli genome encodes a biosynthetic pathway that produces biotin from pimeloyl-CoA in four enzymatic steps. The final step, insertion of sulfur into desthiobiotin to form biotin, is catalyzed by the biotin synthase, BioB. A dedicated biotin ligase (BirA) catalyzes the covalent attachment of biotin to biotin-dependent enzymes. Isotopic labeling has been a valuable tool for probing the details of the biosynthetic process and assaying the activity of biotin-dependent enzymes, however there is currently no established method for 35S labeling of biotin. In this study, we produced [35S]-biotin from Na35SO4 and desthiobiotin with a specific activity of 30.7 Ci/mmol, two orders of magnitude higher than previously published methods. The biotinylation domain (PfBCCP-79) from the Plasmodium falciparum acetyl-CoA carboxylase (ACC) was expressed in E. coli as a biotinylation substrate. We found that overexpression of the E. coli biotin synthase, BioB, and biotin ligase, BirA, increased PfBCCP-79 biotinylation 160-fold over basal levels. Biotinylated PfBCCP-79 was purified by affinity chromatography, and free biotin was liberated using acid hydrolysis. We verified that we had produced radiolabeled biologically active [D]-biotin that specifically labels biotinylated proteins through reuptake in E. coli. The strategy described in our report provides a simple and effective method for the production of [35S]-biotin in E. coli based on affinity chromatography.

  9. Three different signal amplification strategies for the impedimetric sandwich detection of thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Ocaña, Cristina; Valle, Manel del, E-mail: manel.delvalle@uab.cat

    2016-03-17

    In this work, we report a comparative study on three highly specific amplification strategies for the ultrasensitive detection of thrombin with the use of aptamer sandwich protocol. The protocol consisted on the use of a first thrombin aptamer immobilized on the electrode surface, the recognition of thrombin protein, and the reaction with a second biotinylated thrombin aptamer forming the sandwich. Through the exposed biotin end, three variants have been tested to amplify the electrochemical impedance signal. The strategies included (a) silver enhancement treatment, (b) gold enhancement treatment and (c) insoluble product produced by the combination of the enzyme horseradish peroxidase (HRP) and 3-amino-9-ethylcarbazole (AEC). The properties of the sensing surface were probed by electrochemical impedance measurements in the presence of the ferrocyanide/ferricyanide redox marker. Insoluble product strategy and silver enhancement treatment resulted in the lowest detection limit (0.3 pM), while gold enhancement method resulted in the highest reproducibility, 8.8% RSD at the pM thrombin concentration levels. Results of silver and gold enhancement treatment also permitted direct inspection by scanning electron microscopy (SEM). - Highlights: • Aptasensor to detect thrombin reaching the femtomolar level. • Biosensing protocol employs two thrombin aptamers in a sandwich capture scheme. • Use of second biotinylated aptamer allows many amplification and detection variants. • Precipitation reaction provides the highest signal amplification of ca. 3 times. • Double recognition event improves remarkably selectivity for thrombin detection.

  10. Catalase coupled gold nanoparticles: Comparison between carbodiimide and biotin-streptavidin methods

    Science.gov (United States)

    Chirra, Hariharasudhan D.; Sexton, Travis; Biswal, Dipti; Hersh, Louis B.; Hilt, J. Zach

    2011-01-01

    The use of proteins for therapeutic applications requires the protein to maintain sufficient activity for the period of in vivo treatment. Many proteins exhibit a short half-life in vivo and, thus, require delivery systems for them to be applied as therapeutics. The relative biocompatibility and the ability to form functionalized bioconjugates via simple chemistry make gold nanoparticles excellent candidates as protein delivery systems. Herein, two protocols for coupling proteins to gold nanoparticles were compared. In the first, the strong biomolecular binding between biotin and streptavidin was used to couple catalase to the surface of gold nanoparticles. In the second protocol, the formation of an amide bond between carboxylic acid coated gold nanoparticles and free surface amines of catalase using carbodiimide chemistry was performed. The stability and kinetics of the different steps involved in these protocols were studied using UV-Visible spectroscopy, dynamic light scattering, and transmission electron microscopy. The addition of mercaptoundecanoic acid in conjugation with (N-(6-(biotinamido)hexyl)-3′-(2′-pyridyldithio)-propionamide increased the stability of biotinylated gold nanoparticles. Although the carbodiimide chemistry based bioconjugation approach exhibited a decrease in catalase activity, the carbodiimide chemistry based bioconjugation approach resulted in more active catalase per gold nanoparticle compared to that of mercaptoundecanoic acid stabilized biotinylated gold nanoparticles. Both coupling protocols resulted in gold nanoparticles loaded with active catalase. Thus, these gold nanoparticle systems and coupling protocols represent promising methods for the application of gold nanoparticles for protein delivery. PMID:21232642

  11. Three different signal amplification strategies for the impedimetric sandwich detection of thrombin

    International Nuclear Information System (INIS)

    Ocaña, Cristina; Valle, Manel del

    2016-01-01

    In this work, we report a comparative study on three highly specific amplification strategies for the ultrasensitive detection of thrombin with the use of aptamer sandwich protocol. The protocol consisted on the use of a first thrombin aptamer immobilized on the electrode surface, the recognition of thrombin protein, and the reaction with a second biotinylated thrombin aptamer forming the sandwich. Through the exposed biotin end, three variants have been tested to amplify the electrochemical impedance signal. The strategies included (a) silver enhancement treatment, (b) gold enhancement treatment and (c) insoluble product produced by the combination of the enzyme horseradish peroxidase (HRP) and 3-amino-9-ethylcarbazole (AEC). The properties of the sensing surface were probed by electrochemical impedance measurements in the presence of the ferrocyanide/ferricyanide redox marker. Insoluble product strategy and silver enhancement treatment resulted in the lowest detection limit (0.3 pM), while gold enhancement method resulted in the highest reproducibility, 8.8% RSD at the pM thrombin concentration levels. Results of silver and gold enhancement treatment also permitted direct inspection by scanning electron microscopy (SEM). - Highlights: • Aptasensor to detect thrombin reaching the femtomolar level. • Biosensing protocol employs two thrombin aptamers in a sandwich capture scheme. • Use of second biotinylated aptamer allows many amplification and detection variants. • Precipitation reaction provides the highest signal amplification of ca. 3 times. • Double recognition event improves remarkably selectivity for thrombin detection.

  12. A multiwell format assay for heparanase.

    Science.gov (United States)

    Behzad, Farhad; Brenchley, Paul E C

    2003-09-15

    This assay employs a biotinylated heparan sulfate glycosaminoglycan (HSGAG) substrate that is covalently linked to the surface of 96-well immunoassay plates. The ratio of biotin:HSGAG and the coating concentration of substrate bound to the wells have been optimized and allow removal of biotin HSGAG within 60 min of incubation at 37 degrees C in assay buffer with a standard dilution of bacterial heparitinase or platelet heparanase. Loss of biotin signal from the well surface is detected on incubation with peroxidase-streptavidin followed by color development using 3,3',5,5'-tetramethylbenzidine as the peroxidase substrate. The new assay allows specific detection of heparanase activity in multiple samples in a total time of 3 h including a 1-h substrate digestion step and is a significant improvement with regard to sensitivity, specificity, and ease of handling of multiple samples compared to other described assays. Heparanase specifically degrades the biotinylated HSGAG substrate, when used with an optimized assay buffer. A range of enzymes including collagenase, trypsin, plasmin, pepsin, chondroitinases, hyaluronidase, and neuraminidase show no effect on the substrate under optimized assay conditions. The covalent linkage of the substrate to the well prevents leaching of substrate and allows preparation and long-term storage of substrate-coated plates. The assay can be used to detect heparanase levels in clinical samples and cell culture supernatants and is ideal as a screening method for antagonists of enzyme activity.

  13. Alternating current impedance spectroscopic analysis of biofunctionalized vertically-aligned silica nanospring surface for biosensor applications

    Science.gov (United States)

    Timalsina, Yukta P.

    In this dissertation, a process of vertically-aligned (silica) nanosprings (VANS) based biosensor development is presented. Alternating current (AC) impedance spectroscopy has been used to analyze sensor response as a function of saline phosphate (SP) buffer and biological solutions. The sensor is a parallel plate capacitor consisting of two glass substrates coated with indium tin oxide (ITO), where the VANS [or randomly-aligned nanosprings (RANS)] grown on one substrate serve as the dielectric spacer layer. The response of a VANS device as a function of ionic concentration in SP buffer was examined and an equivalent circuit model was developed. The results demonstrated that VANS sensors exhibited greater sensitivity to the changes in SP concentration relative to the ITO sensors, which serve as controls. The biofunctionalized VANS surface via physisorption and the cross-linker method demonstrates the repeatability, specificity, and selectivity of the binding. The physisorption of biotinylated immunoglobulin G (B-IgG) onto the VANS surface simplifies the whole sensing procedure for the detection of glucose oxidase, since the avidin-conjugated glucose oxidase (Av-GOx) can directly be immobilized on the B-IgG. The cross linker method involves the covalent attachment of antibodies onto the functionalized VANS surface via imine bond. The experiments revealed that the VANS sensor response is solely the result of the interaction of target molecule i.e. mouse IgG with the probe layer, i.e. goat antimouse IgG (GalphaM IgG). It was determined that VANS-based sensors exhibit a greater magnitude of change between successive bio-layers relative to the controls above 100 Hz, which indicates that the addition of biomolecules inhibits the diffusion of ions and changes the effective dielectric response of the VANS via biomolecular polarization. The study of ionic transport in nanosprings suggested that conductance follows a scaling law. It was demonstrated that a VANS-based device

  14. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen

    DEFF Research Database (Denmark)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other...... preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious...

  15. Regulation of immunological and inflammatory functions by biotin.

    Science.gov (United States)

    Kuroishi, Toshinobu

    2015-12-01

    Biotin is a water-soluble B-complex vitamin and is well-known as a co-factor for 5 indispensable carboxylases. Holocarboxylase synthetase (HLCS) catalyzes the biotinylation of carboxylases and other proteins, whereas biotinidase catalyzes the release of biotin from biotinylated peptides. Previous studies have reported that nutritional biotin deficiency and genetic defects in either HLCS or biotinidase induces cutaneous inflammation and immunological disorders. Since biotin-dependent carboxylases involve various cellular metabolic pathways including gluconeogenesis, fatty acid synthesis, and the metabolism of branched-chain amino acids and odd-chain fatty acids, metabolic abnormalities may play important roles in immunological and inflammatory disorders caused by biotin deficiency. Transcriptional factors, including NF-κB and Sp1/3, are also affected by the status of biotin, indicating that biotin regulates immunological and inflammatory functions independently of biotin-dependent carboxylases. An in-vivo analysis with a murine model revealed the therapeutic effects of biotin supplementation on metal allergies. The novel roles of biotinylated proteins and their related enzymes have recently been reported. Non-carboxylase biotinylated proteins induce chemokine production. HLCS is a nuclear protein involved in epigenetic and chromatin regulation. In this review, comprehensive knowledge on the regulation of immunological and inflammatory functions by biotin and its potential as a therapeutic agent is discussed.

  16. Dendrimer D5 is a vector for peptide transport to brain cells.

    Science.gov (United States)

    Sarantseva, S V; Bolshakova, O I; Timoshenko, S I; Kolobov, A A; Schwarzman, A L

    2011-02-01

    Dendrimers are a new class of nonviral vectors for gene or drug transport. Dendrimer capacity to penetrate through the blood-brain barrier remaines little studied. Biotinylated polylysine dendrimer D5, similarly to human growth hormone biotinylated fragment covalently bound to D5 dendrimer, penetrates through the blood-brain barrier and accumulates in Drosophila brain after injection into the abdomen. Hence, D5 dendrimer can serve as a vector for peptide transport to brain cells.

  17. Association of Biotin Ingestion With Performance of Hormone and Nonhormone Assays in Healthy Adults.

    Science.gov (United States)

    Li, Danni; Radulescu, Angela; Shrestha, Rupendra T; Root, Matthew; Karger, Amy B; Killeen, Anthony A; Hodges, James S; Fan, Shu-Ling; Ferguson, Angela; Garg, Uttam; Sokoll, Lori J; Burmeister, Lynn A

    2017-09-26

    Biotinylated antibodies and analogues, with their strong binding to streptavidin, are used in many clinical laboratory tests. Excess biotin in blood due to supplemental biotin ingestion may affect biotin-streptavidin binding, leading to potential clinical misinterpretation. However, the degree of interference remains undefined in healthy adults. To assess performance of specific biotinylated immunoassays after 7 days of ingesting 10 mg/d of biotin, a dose common in over-the-counter supplements for healthy adults. Nonrandomized crossover trial involving 6 healthy adults who were treated at an academic medical center research laboratory. Administration of 10 mg/d of biotin supplementation for 7 days. Analyte concentrations were compared with baseline (day 0) measures on the seventh day of biotin treatment and 7 days after treatment had stopped (day 14). The 11 analytes included 9 hormones (ie, thyroid-stimulating hormone, total thyroxine, total triiodothyronine, free thyroxine, free triiodothyronine, parathyroid hormone, prolactin, N-terminal pro-brain natriuretic peptide, 25-hydroxyvitamin D) and 2 nonhormones (prostate-specific antigen and ferritin). A total of 37 immunoassays for the 11 analytes were evaluated on 4 diagnostic systems, including 23 assays that incorporated biotin and streptavidin components and 14 assays that did not include biotin and streptavidin components and served as negative controls. Among the 2 women and 4 men (mean age, 38 years [range, 31-45 years]) who took 10 mg/d of biotin for 7 days, biotin ingestion-associated interference was found in 9 of the 23 (39%) biotinylated assays compared with none of the 14 nonbiotinylated assays (P = .007). Results from 5 of 8 biotinylated (63%) competitive immunoassays tested falsely high and results from 4 out of 15 (27%) biotinylated sandwich immunoassays tested falsely low. In this preliminary study of 6 healthy adult participants and 11 hormone and nonhormone analytes measured by 37 immunoassays

  18. Decoding P4-ATPase substrate interactions.

    Science.gov (United States)

    Roland, Bartholomew P; Graham, Todd R

    Cellular membranes display a diversity of functions that are conferred by the unique composition and organization of their proteins and lipids. One important aspect of lipid organization is the asymmetric distribution of phospholipids (PLs) across the plasma membrane. The unequal distribution of key PLs between the cytofacial and exofacial leaflets of the bilayer creates physical surface tension that can be used to bend the membrane; and like Ca 2+ , a chemical gradient that can be used to transduce biochemical signals. PL flippases in the type IV P-type ATPase (P4-ATPase) family are the principle transporters used to set and repair this PL gradient and the asymmetric organization of these membranes are encoded by the substrate specificity of these enzymes. Thus, understanding the mechanisms of P4-ATPase substrate specificity will help reveal their role in membrane organization and cell biology. Further, decoding the structural determinants of substrate specificity provides investigators the opportunity to mutationally tune this specificity to explore the role of particular PL substrates in P4-ATPase cellular functions. This work reviews the role of P4-ATPases in membrane biology, presents our current understanding of P4-ATPase substrate specificity, and discusses how these fundamental aspects of P4-ATPase enzymology may be used to enhance our knowledge of cellular membrane biology.

  19. STANDARDIZATION OF A FLUORESCENT-BASED QUANTITATIVE ADHESION ASSAY TO STUDY ATTACHMENT OF Taenia solium ONCOSPHERE TO EPITHELIAL CELLS In Vitro

    Science.gov (United States)

    Chile, Nancy; Evangelista, Julio; Gilman, Robert H.; Arana, Yanina; Palma, Sandra; Sterling, Charles R; Garcia, Hector H.; Gonzalez, Armando; Verastegui, Manuela

    2012-01-01

    To fully understand the preliminary stages of Taenia solium oncosphere attachment in the gut, adequate tools and assays are necessary to observe and quantify this event that leads to infection. A fluorescent-based quantitative adhesion assay, using biotinylated activated-oncospheres and monolayers of Chinese hamster ovary cells (CHO-K1) or human intestinal monolayer cells (INT-407, HCT-8 or HT-29), was developed to study initial events during the infection of target cells and to rapidly quantify the in vitro adhesion of T. solium oncospheres. Fluorescein streptavidin was used to identify biotinylated activated-oncospheres adhered to cells. This adherence was quantified using an automated fluorescence plate reader, and the results were expressed as fluorescence intensity values. A series of three assays were performed. The first was to identify the optimum number of biotinylated activated-oncospheres to be used in the adhesion assay. The goal of the second assay was to validate this novel method with the established oncosphere-binding system using the immunofluorescent-antibody assay (IFA) method to quantify oncosphere adhesion. A total of 10,000 biotinylated activated-oncospheres were utilized to assess the role of sera and laminin (LM) in oncosphere adherence to a CHO-K1 cell monolayer. The findings that sera and LM increase the adhesion of oncospheres to monolayer cells were similar to results that were previously obtained using the IFA method. The third assay compared the adherence of biotinylated activated-oncospheres to different types of human intestinal monolayer cells. In this case, the fluorescence intensity was greatest when using the INT-407 cell monolayer. We believe this new method of quantification offers the potential for rapid, large-scale screening to study and elucidate specific molecules and mechanisms involved in oncosphere-host cell attachment. PMID:22178422

  20. Design and Proof-of-Concept Use of a Circular PMMA Platform with 16-Well Sample Capacity for Microwave-Accelerated Bioassays.

    Science.gov (United States)

    Mohammed, Muzaffer; Aslan, Kadir

    2013-01-01

    We demonstrate the design and the proof-of-concept use of a new, circular poly(methyl methacrylate)-based bioassay platform (PMMA platform), which affords for the rapid processing of 16 samples at once. The circular PMMA platform (5 cm in diameter) was coated with a silver nanoparticle film to accelerate the bioassay steps by microwave heating. A model colorimetric bioassay for biotinylated albumin (using streptavidin-labeled horse radish peroxidase) was performed on the PMMA platform coated with and without silver nanoparticles (a control experiment), and at room temperature and using microwave heating. It was shown that the simulated temperature profile of the PMMA platform during microwave heating were comparable to the real-time temperature profile during actual microwave heating of the constructed PMMA platform in a commercial microwave oven. The model colorimetric bioassay for biotinylated albumin was successfully completed in ~2 min (total assay time) using microwave heating, as compared to 90 min at room temperature (total assay time), which indicates a ~45-fold decrease in assay time. Our PMMA platform design afforded for significant reduction in non-specific interactions and low background signal as compared to non-silvered PMMA surfaces when employed in a microwave-accelerated bioassay carried out in a conventional microwave cavity.

  1. Aligned deposition and electrical measurements on single DNA molecules

    International Nuclear Information System (INIS)

    Eidelshtein, Gennady; Kotlyar, Alexander; Hashemi, Mohtadin; Gurevich, Leonid

    2015-01-01

    A reliable method of deposition of aligned individual dsDNA molecules on mica, silicon, and micro/nanofabricated circuits is presented. Complexes of biotinylated double stranded poly(dG)–poly(dC) DNA with avidin were prepared and deposited on mica and silicon surfaces in the absence of Mg 2+ ions. Due to its positive charge, the avidin attached to one end of the DNA anchors the complex to negatively charged substrates. Subsequent drying with a directional gas flow yields DNA molecules perfectly aligned on the surface. In the avidin–DNA complex only the avidin moiety is strongly and irreversibly bound to the surface, while the DNA counterpart interacts with the substrates much more weakly and can be lifted from the surface and realigned in any direction. Using this technique, avidin–DNA complexes were deposited across platinum electrodes on a silicon substrate. Electrical measurements on the deposited DNA molecules revealed linear IV-characteristics and exponential dependence on relative humidity. (paper)

  2. Preparation of Conjugates of Cytotoxic Lupane Triterpenes with Biotin.

    Science.gov (United States)

    Soural, Miroslav; Hodon, Jiri; Dickinson, Niall J; Sidova, Veronika; Gurska, Sona; Dzubak, Petr; Hajduch, Marian; Sarek, Jan; Urban, Milan

    2015-12-16

    To better understand the mechanism of action of antitumor triterpenes, we are developing methods to identify their molecular targets. A promising method is based on combination of quantitative proteomics with SILAC and uses active compounds anchored to magnetic beads via biotin-streptavidin interaction. We developed a simple and fast solid-phase synthetic technique to connect terpenes to biotin through a linker. Betulinic acid was biotinylated from three different conjugation sites for use as a standard validation tool since many molecular targets of this triterpene are already known. Then, a set of four other cytotoxic triterpenoids was biotinylated. Biotinylated terpenes were similarly cytotoxic to their nonbiotinylated parents, which suggests that the target identification should not be influenced by linker or biotin. The developed solid-phase synthetic approach is the first attempt to use solid-phase synthesis to connect active triterpenes to biotin and is applicable as a general procedure for routine conjugation of triterpenes with other molecules of choice.

  3. HaloTag protein-mediated specific labeling of living cells with quantum dots

    International Nuclear Information System (INIS)

    So, Min-kyung; Yao Hequan; Rao Jianghong

    2008-01-01

    Quantum dots emerge as an attractive alternative to small molecule fluorophores as fluorescent tags for in vivo cell labeling and imaging. This communication presents a method for specific labeling of live cells using quantum dots. The labeling is mediated by HaloTag protein expressed at the cell surface which forms a stable covalent adduct with its ligand (HaloTag ligand). The labeling can be performed in one single step with quantum dot conjugates that are functionalized with HaloTag ligand, or in two steps with biotinylated HaloTag ligand first and followed by streptavidin coated quantum dots. Live cell fluorescence imaging indicates that the labeling is specific and takes place at the cell surface. This HaloTag protein-mediated cell labeling method should facilitate the application of quantum dots for live cell imaging

  4. Poly(amido)amine (PAMAM) dendrimer-cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    Science.gov (United States)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath

    2013-09-01

    Dendrimer-cisplatin complexes were prepared using PAMAM dendrimers with terminal -NH2 and -COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer-cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was 11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC50 values in ovarian cancer cells when compared with carboxylate surface dendrimers ( p cisplatin complexes resulted in a 7.0-fold increase ( p cisplatin chemotherapy of ovarian cancer.

  5. Synthesis and evaluation of 99mTc/99Tc-MAG3-biotin conjugates for antibody pretargeting strategies

    International Nuclear Information System (INIS)

    Gog, Frank B. van; Visser, Gerard W.M.; Gowrising, Radjish W.A.; Snow, Gordon B.; Dongen, Guus A.M.S. van

    1998-01-01

    Four 99m Tc-MAG3-biotin conjugates were synthesized to determine their potential use in antibody pretargeting strategies for radioimmunoscintigraphy (RIS). To use these 99m Tc-MAG3-biotin conjugates as model compounds for 186 Re-MAG3-biotin conjugates for radioimmunotherapy (RIT), nanomolar amounts of 99 Tc were added as carrier to 99m Tc. The biotin derivatives used for the preparation of the conjugates - biocytin, biotin hydrazide, biotinyl-piperazine, and biotinyl-diaminosuccinic acid - differed at the site that is regarded to be susceptible to hydrolysis by biotinidase present in human plasma. All four conjugates were produced with high radiochemical purity, were stable in PBS, and demonstrated full binding capacity to streptavidin. The 99m Tc/ 99 Tc-MAG3-labeled biotinyl-piperazine and biotinyl-diaminosuccinic acid conjugates were stable in mouse as well as human plasma, whereas the corresponding biocytin and biotin hydrazide conjugates were rapidly degraded. The biodistribution in nude mice at 30 min after injection was similar for all conjugates, and a rapid blood clearance and high intestinal excretion were both observed. It is concluded that the metabolic routing of a conjugate containing biotin and MAG3 is dominated by these two moieties. For this reason, MAG3-biotin conjugates do not seem suited for pretargeted RIT, for which quantitative and fast renal excretion is a prerequisite to minimize radiation toxicity. However, in a pretargeted RIS approach the 99m Tc-MAG3-biotin conjugates might have potential

  6. Effects of Lectins on initial attachment of cariogenic Streptococcus mutans.

    Science.gov (United States)

    Ito, Takashi; Yoshida, Yasuhiro; Shiota, Yasuyoshi; Ito, Yuki; Yamamoto, Tadashi; Takashiba, Shogo

    2018-02-01

    Oral bacteria initiate biofilm formation by attaching to tooth surfaces via an interaction of a lectin-like bacterial protein with carbohydrate chains on the pellicle. This study aimed to find naturally derived lectins that inhibit the initial attachment of a cariogenic bacterial species, Streptococcus mutans (S. mutans), to carbohydrate chains in saliva in vitro. Seventy kinds of lectins were screened for candidate motifs that inhibit the attachment of S. mutans ATCC 25175 to a saliva-coated culture plate. The inhibitory effect of the lectins on attachment of the S. mutans to the plates was quantified by crystal violet staining, and the biofilm was observed under a scanning electron microscope (SEM). Surface plasmon resonance (SPR) analysis was performed to examine the binding of S. mutans to carbohydrate chains and the binding of candidate lectins to carbohydrate chains, respectively. Moreover, binding assay between the biotinylated-lectins and the saliva components was conducted to measure the lectin binding. Lectins recognizing a salivary carbohydrate chain, Galβ1-3GalNAc, inhibited the binding of S. mutans to the plate. In particular, Agaricus bisporus agglutinin (ABA) markedly inhibited the binding. This inhibition was confirmed by SEM observation. SPR analysis indicated that S. mutans strongly binds to Galβ1-3GalNAc, and ABA binds to Galβ1-3GalNAc. Finally, the biotinylated Galβ1-3GalNAc-binding lectins including ABA demonstrated marked binding to the saliva components. These results suggest that ABA lectin inhibited the attachment of S. mutans to Galβ1-3GalNAc in saliva and ABA can be useful as a potent inhibitor for initial attachment of oral bacteria and biofilm formation.

  7. Enhanced neutralization potency of botulinum neurotoxin antibodies using a red blood cell-targeting fusion protein.

    Directory of Open Access Journals (Sweden)

    Sharad P Adekar

    2011-03-01

    Full Text Available Botulinum neurotoxin (BoNT potently inhibits cholinergic signaling at the neuromuscular junction. The ideal countermeasures for BoNT exposure are monoclonal antibodies or BoNT antisera, which form BoNT-containing immune complexes that are rapidly cleared from the general circulation. Clearance of opsonized toxins may involve complement receptor-mediated immunoadherence to red blood cells (RBC in primates or to platelets in rodents. Methods of enhancing immunoadherence of BoNT-specific antibodies may increase their potency in vivo. We designed a novel fusion protein (FP to link biotinylated molecules to glycophorin A (GPA on the RBC surface. The FP consists of an scFv specific for murine GPA fused to streptavidin. FP:mAb:BoNT complexes bound specifically to the RBC surface in vitro. In a mouse model of BoNT neutralization, the FP increased the potency of single and double antibody combinations in BoNT neutralization. A combination of two antibodies with the FP gave complete neutralization of 5,000 LD50 BoNT in mice. Neutralization in vivo was dependent on biotinylation of both antibodies and correlated with a reduction of plasma BoNT levels. In a post-exposure model of intoxication, FP:mAb complexes gave complete protection from a lethal BoNT/A1 dose when administered within 2 hours of toxin exposure. In a pre-exposure prophylaxis model, mice were fully protected for 72 hours following administration of the FP:mAb complex. These results demonstrate that RBC-targeted immunoadherence through the FP is a potent enhancer of BoNT neutralization by antibodies in vivo.

  8. Analysis of membrane proteome and secretome in cells over-expressing ADAM17 using quantitative proteomics

    International Nuclear Information System (INIS)

    Kawahara, R.; Simabuco, F.M.; Yokoo, S.; Paes Leme, A.F.; Sherman, N.

    2012-01-01

    Full text: A disintegrin and metalloproteinase (ADAM) protease is involved in proteolytic ectodomain shedding of several membrane-associated proteins and modulation of key cell signaling pathways in the tumor microenvironment. In this study, we examined the effect of over-expressing the full length human ADAM17 in membrane and secreted proteins. To this end, we constructed a stable Flp-In T-RExHEK293 cells expressing ADAM17 by tetracycline induction. These cells were grown in Dulbeccos modified Eagles medium containing light lysine, arginine or heavy, L-Arg-13C615N4 and L-Lys -13C615N2 (SILAC: stable isotope labeling with amino acid in cell culture) media and they were treated with an ADAM17 activator, phorbolester (PMA). Controls such as Flp-In T-RExHEK293 cell without PMA treatment and without ADAM17 cloned were cultivated in light medium. The ADAM17 overexpression was induced with tetracycline 500 ng/ml for 24 hours. Cells in a heavy condition were treated with PMA 50 ng/ml for 1 hour and vehicle DMSO was used as control in a light cell condition. The extracellular media were collected, concentrated and used to evaluate the secretome and a cell surface biotinylation-based approach was used to capture cell surface-associated proteins. The biotinylated proteins were eluted with dithiothreitol, alkylated with iodoacetamide and then digested with trypsin. The resulting peptides were subjected to LC-MS/MS analysis on an ETD enabled Orbitrap Velos instrument. The results showed different proteins up or down regulated in membrane and secretome analysis which might represent potential molecules involved in signaling or ADAM17 regulation events. (author)

  9. Extracellular Actin Is a Receptor for Mycoplasma hyopneumoniae

    Directory of Open Access Journals (Sweden)

    Benjamin B. A. Raymond

    2018-02-01

    Full Text Available Mycoplasma hyopneumoniae, an agriculturally important porcine pathogen, disrupts the mucociliary escalator causing ciliostasis, loss of cilial function, and epithelial cell death within the porcine lung. Losses to swine production due to growth rate retardation and reduced feed conversion efficiency are severe, and antibiotics are used heavily to control mycoplasmal pneumonia. Notably, little is known about the repertoire of host receptors that M. hyopneumoniae targets to facilitate colonization. Here we show, for the first time, that actin exists extracellularly on porcine epithelial monolayers (PK-15 using surface biotinylation and 3D-Structured Illumination Microscopy (3D-SIM, and that M. hyopneumoniae binds to the extracellular β-actin exposed on the surface of these cells. Consistent with this hypothesis we show: (i monoclonal antibodies that target β-actin significantly block the ability of M. hyopneumoniae to adhere and colonize PK-15 cells; (ii microtiter plate binding assays show that M. hyopneumoniae cells bind to monomeric G-actin in a dose dependent manner; (iii more than 100 M. hyopneumoniae proteins were recovered from affinity-chromatography experiments using immobilized actin as bait; and (iv biotinylated monomeric actin binds directly to M. hyopneumoniae proteins in ligand blotting studies. Specifically, we show that the P97 cilium adhesin possesses at least two distinct actin-binding regions, and binds monomeric actin with nanomolar affinity. Taken together, these observations suggest that actin may be an important receptor for M. hyopneumoniae within the swine lung and will aid in the future development of intervention strategies against this devastating pathogen. Furthermore, our observations have wider implications for extracellular actin as an important bacterial receptor.

  10. Extracellular Actin Is a Receptor for Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Raymond, Benjamin B A; Madhkoor, Ranya; Schleicher, Ina; Uphoff, Cord C; Turnbull, Lynne; Whitchurch, Cynthia B; Rohde, Manfred; Padula, Matthew P; Djordjevic, Steven P

    2018-01-01

    Mycoplasma hyopneumoniae , an agriculturally important porcine pathogen, disrupts the mucociliary escalator causing ciliostasis, loss of cilial function, and epithelial cell death within the porcine lung. Losses to swine production due to growth rate retardation and reduced feed conversion efficiency are severe, and antibiotics are used heavily to control mycoplasmal pneumonia. Notably, little is known about the repertoire of host receptors that M. hyopneumoniae targets to facilitate colonization. Here we show, for the first time, that actin exists extracellularly on porcine epithelial monolayers (PK-15) using surface biotinylation and 3D-Structured Illumination Microscopy (3D-SIM), and that M. hyopneumoniae binds to the extracellular β-actin exposed on the surface of these cells. Consistent with this hypothesis we show: (i) monoclonal antibodies that target β-actin significantly block the ability of M. hyopneumoniae to adhere and colonize PK-15 cells; (ii) microtiter plate binding assays show that M. hyopneumoniae cells bind to monomeric G-actin in a dose dependent manner; (iii) more than 100 M. hyopneumoniae proteins were recovered from affinity-chromatography experiments using immobilized actin as bait; and (iv) biotinylated monomeric actin binds directly to M. hyopneumoniae proteins in ligand blotting studies. Specifically, we show that the P97 cilium adhesin possesses at least two distinct actin-binding regions, and binds monomeric actin with nanomolar affinity. Taken together, these observations suggest that actin may be an important receptor for M. hyopneumoniae within the swine lung and will aid in the future development of intervention strategies against this devastating pathogen. Furthermore, our observations have wider implications for extracellular actin as an important bacterial receptor.

  11. Single Lipid Molecule Dynamics on Supported Lipid Bilayers with Membrane Curvature

    Directory of Open Access Journals (Sweden)

    Philip P. Cheney

    2017-03-01

    Full Text Available The plasma membrane is a highly compartmentalized, dynamic material and this organization is essential for a wide variety of cellular processes. Nanoscale domains allow proteins to organize for cell signaling, endo- and exocytosis, and other essential processes. Even in the absence of proteins, lipids have the ability to organize into domains as a result of a variety of chemical and physical interactions. One feature of membranes that affects lipid domain formation is membrane curvature. To directly test the role of curvature in lipid sorting, we measured the accumulation of two similar lipids, 1,2-Dihexadecanoyl-sn-glycero-3-phosphoethanolamine (DHPE and hexadecanoic acid (HDA, using a supported lipid bilayer that was assembled over a nanopatterned surface to obtain regions of membrane curvature. Both lipids studied contain 16 carbon, saturated tails and a head group tag for fluorescence microscopy measurements. The accumulation of lipids at curvatures ranging from 28 nm to 55 nm radii was measured and fluorescein labeled DHPE accumulated more than fluorescein labeled HDA at regions of membrane curvature. We then tested whether single biotinylated DHPE molecules sense curvature using single particle tracking methods. Similar to groups of fluorescein labeled DHPE accumulating at curvature, the dynamics of single molecules of biotinylated DHPE was also affected by membrane curvature and highly confined motion was observed.

  12. Effect of administration route and dose of streptavidin or biotin on the tumor uptake of radioactivity in intraperitoneal tumor with multistep targeting

    International Nuclear Information System (INIS)

    Zhang Meili; Yao Zhengsheng; Sakahara, Harumi; Saga, Tsuneo; Nakamoto, Yuji; Sato, Noriko; Zhao Songji; Nakada, Hiroshi; Yamashina, Ikuo; Konishi, Junji

    1998-01-01

    The effect of the administration route and dose of streptavidin or biotin on the biodistribution of radioactivity in multistep targeting was studied in nude mice bearing intraperitoneal (IP) colon cancer xenograft. The multistep targeting included a two-step method using biotinylated antibody and radiolabeled streptavidin and a three-step method with radiolabeled biotin based on the two-step method. A monoclonal antibody, MLS128, which recognizes Tn antigen on mucin, was biotinylated and injected intravenously (IV) or IP in nude mice bearing human colon cancer LS180 IP xenografts for pretargeting. In the two-step method, IP-injected streptavidin showed a higher tumor uptake and tumor-to-nontumor ratios than IV-injected streptavidin regardless of administration route of pretargeting. The tumor uptake of radiolabeled streptavidin was increased with a high dose of biotinylated antibody pretargeting, but decreased with an increasing dose of streptavidin. In the three-step targeting, IP injection also gave a higher tumor uptake of radiolabeled biotin than IV injection. In conclusion, IP administration of radiolabeled streptavidin or biotin resulted in more efficient IP tumor targeting with the multistep methods

  13. Radioimmunocytochemistry with [3H]biotin

    International Nuclear Information System (INIS)

    Hunt, S.P.; Mantyh, P.W.

    1984-01-01

    The authors exploit the high affinity of biotin for avidin in the design of radioimmunocytochemical methods using [ 3 H]biotin. [ 3 H]Biotin and avidin D form a radioactive complex which can be linked onto a primary antibody by means of a biotinylated anti-rabbit IgG or biotinylated protein A link. With both approaches it was possible to localize a number of antigens such as somatostatin, substance P, avian pancreatic polypeptide, tyrosine hydroxylase, and enkephalin-like immunoreactivity in various regions of the rat and human brain. By using tritium-sensitive film, large regions of the brain could be studied and analyzed semiquantitatively using computerized microdensitometry. The technique was also taken to the electron microscope level, and in the case of substance P immunoreactivity within the rat substantia nigra silver grains were found to be highly localized over axons and axon terminals. It was also possible to demonstrate co-existence or lack of co-existence of a number of different antigens within neurones. The first primary antibody was localized with biotinylated protein A followed by avidin-peroxidase, while the second primary antibody was linked to the [ 3 H]biotin again with biotinylated protein A. As an example of the potential of these methods for semiquantification, the distribution of substance P within postmortem human spinal cord was examined 24 months after amputation. A 49% loss of peptide was found in the corresponding dorsal horn. In summary these methods using [ 3 H]biotin have proved successful in quantification, electron microscopy and double labelling studies. (Auth.)

  14. The Motion of a Single Molecule, the Lambda-Receptor, in the Bacterial Outer Membrane

    DEFF Research Database (Denmark)

    Oddershede, Lene; Dreyer, Jakob Kisbye; Grego, Sonia

    2002-01-01

    Using optical tweezers and single particle tracking, we have revealed the motion of a single protein, the lambda-receptor, in the outer membrane of living Escherichia coli bacteria. We genetically modified the lambda-receptor placing a biotin on an extracellular site of the receptor in vivo....... The efficiency of this in vivo biotinylation is very low, thus enabling the attachment of a streptavidin-coated bead binding specifically to a single biotinylated lambda-receptor. The bead was used as a handle for the optical tweezers and as a marker for the single particle tracking routine. We propose a model...

  15. Peptides for functionalization of InP semiconductors.

    Science.gov (United States)

    Estephan, Elias; Saab, Marie-belle; Larroque, Christian; Martin, Marta; Olsson, Fredrik; Lourdudoss, Sebastian; Gergely, Csilla

    2009-09-15

    The challenge is to achieve high specificity in molecular sensing by proper functionalization of micro/nano-structured semiconductors by peptides that reveal specific recognition for these structures. Here we report on surface modification of the InP semiconductors by adhesion peptides produced by the phage display technique. An M13 bacteriophage library has been used to screen 10(10) different peptides against the InP(001) and the InP(111) surfaces to finally isolate specific peptides for each orientation of the InP. MALDI-TOF/TOF mass spectrometry has been employed to study real affinity of the peptide towards the InP surfaces. The peptides serve for controlled placement of biotin onto InP to bind then streptavidin. Our Atomic Force Microscopy study revealed a total surface coverage of molecules when the InP surface was functionalized by its specific biotinylated peptide (YAIKGPSHFRPS). Finally, fluorescence microscopy has been employed to demonstrate the preferential attachment of the peptide onto a micro-patterned InP surface. Use of substrate specific peptides could present an alternative solution for the problems encountered in the actually existing sensing methods and molecular self-assembly due to the unwanted unspecific interactions.

  16. Magnetically triggered clustering of biotinylated iron oxide nanoparticles in the presence of streptavidinylated enzymes

    International Nuclear Information System (INIS)

    Hodenius, Michael; De Cuyper, Marcel; Hieronymus, Thomas; Zenke, Martin; Becker, Christiane; Elling, Lothar; Bornemann, Jörg; Wong, John E; Richtering, Walter; Himmelreich, Uwe

    2012-01-01

    This work deals with the production and characterization of water-compatible, iron oxide based nanoparticles covered with functional poly(ethylene glycol) (PEG)–biotin surface groups (SPIO–PEG–biotin). Synthesis of the functionalized colloids occurred by incubating the oleate coated particles used as precursor magnetic fluid with anionic liposomes containing 14 mol% of a phospholipid–PEG–biotin conjugate. The latter was prepared by coupling dimyristoylphosphatidylethanolamine (DC 14:0 PE) to activated α-biotinylamido-ω –N-hydroxy-succinimidcarbonyl–PEG (NHS–PEG–biotin). Physical characterization of the oleate and PEG–biotin iron oxide nanocolloids revealed that they appear as colloidal stable clusters with a hydrodynamic diameter of 160 nm and zeta potentials of − 39 mV (oleate coated particles) and − 14 mV (PEG–biotin covered particles), respectively, as measured by light scattering techniques. Superconducting quantum interference device (SQUID) measurements revealed specific saturation magnetizations of 62–73 emu g −1 Fe 3 O 4 and no hysteresis was observed at 300 K. MR relaxometry at 3 T revealed very high r 2 relaxivities and moderately high r 1 values. Thus, both nanocolloids can be classified as small, superparamagnetic, negative MR contrast agents. The capacity to functionalize the particles was illustrated by binding streptavidin alkaline phosphatase (SAP). It was found, however, that these complexes become highly aggregated after capturing them on the magnetic filter device during high-gradient magnetophoresis, thereby reducing the accessibility of the SAP. (paper)

  17. Estradiol-induced estrogen receptor-alpha trafficking.

    Science.gov (United States)

    Bondar, Galyna; Kuo, John; Hamid, Naheed; Micevych, Paul

    2009-12-02

    Estradiol has rapid actions in the CNS that are mediated by membrane estrogen receptors (ERs) and activate cell signaling pathways through interaction with metabotropic glutamate receptors (mGluRs). Membrane-initiated estradiol signaling increases the free cytoplasmic calcium concentration ([Ca(2+)](i)) that stimulates the synthesis of neuroprogesterone in astrocytes. We used surface biotinylation to demonstrate that ERalpha has an extracellular portion. In addition to the full-length ERalpha [apparent molecular weight (MW), 66 kDa], surface biotinylation labeled an ERalpha-immunoreactive protein (MW, approximately 52 kDa) identified by both COOH- and NH(2)-directed antibodies. Estradiol treatment regulated membrane levels of both proteins in parallel: within 5 min, estradiol significantly increased membrane levels of the 66 and 52 kDa ERalpha. Internalization, a measure of membrane receptor activation, was also increased by estradiol with a similar time course. Continuous treatment with estradiol for 24-48 h reduced ERalpha levels, suggesting receptor downregulation. Estradiol also increased mGluR1a trafficking and internalization, consistent with the proposed ERalpha-mGluR1a interaction. Blocking ER with ICI 182,780 or mGluR1a with LY 367385 prevented ERalpha trafficking to and from the membrane. Estradiol-induced [Ca(2+)](i) flux was also significantly increased at the time of peak ERalpha activation/internalization. These results demonstrate that ERalpha is present in the membrane and has an extracellular portion. Furthermore, membrane levels and internalization of ERalpha are regulated by estradiol and mGluR1a ligands. The pattern of trafficking into and out of the membrane suggests that the changing concentration of estradiol during the estrous cycle regulates ERalpha to augment and then terminate membrane-initiated signaling.

  18. Estradiol-induced estrogen receptor-α trafficking

    Science.gov (United States)

    Bondar, Galyna; Kuo, John; Hamid, Naheed; Micevych, Paul

    2010-01-01

    Estradiol has rapid actions in the central nervous system, which are mediated by membrane estrogen receptors (ERs) and activate cell signaling pathways through interaction with metabotropic glutamate receptors (mGluRs). Membrane-initiated estradiol signaling increases the free cytoplasmic calcium concentration ([Ca2+]i) that stimulates the synthesis of neuroprogesterone in astrocytes. We used surface biotinylation to demonstrate that ERα has an extracellular portion. In addition to the full length ERα (apparent M.W. 66 kDa), surface biotinylation labeled an ERα-immunoreactive protein (M.W. ~ 52 kDa) identified by both COOH- and NH2-directed antibodies. Estradiol treatment regulated membrane levels of both proteins in parallel: within 5 min, estradiol significantly increased membrane levels of the 66 kDa and 52 kDa ERα. Internalization, a measure of membrane receptor activation, was also increased by estradiol with a similar time course. Continuous treatment with estradiol for 24–48 hr reduced ERα levels, suggesting receptor down-regulation. Estradiol also increased mGluR1a trafficking and internalization, consistent with the proposed ERα-mGluR1a interaction. Blocking ER with ICI 182,780 or mGluR1a with LY 367385 prevented ERα trafficking to and from the membrane. Estradiol-induced [Ca2+]i flux was also significantly increased at the time of peak ERα activation/internalization. These results demonstrate that ERα is present in the membrane and has an extracellular portion. Furthermore, membrane levels and internalization of ERα are regulated by estradiol and mGluR1a ligands. The pattern of trafficking into and out of the membrane suggests that the changing concentration of estradiol during the estrous cycle regulates ERα to augment and then terminate membrane-initiated signaling. PMID:19955385

  19. Tailor-making a protein a-derived domain for efficient site-specific photocoupling to Fc of mouse IgG₁.

    Directory of Open Access Journals (Sweden)

    Feifan Yu

    Full Text Available Affinity proteins binding to antibody constant regions have proved to be invaluable tools in biotechnology. Here, protein engineering was used to expand the repertoire of available immunoglobulin binding proteins via improvement of the binding strength between the widely used staphylococcal protein A-derived Z domain and the important immunoglobulin isotype mouse IgG₁ (mIgG₁. Addressing seven positions in the 58-residue three-helix bundle Z domain by single or double amino acid substitutions, a total of 170 variants were individually constructed, produced in E. coli and tested for binding to a set of mouse IgG₁ monoclonal antibodies (mAbs. The best variant, denoted Z(F5I corresponding to a Phe to Ile substitution at position 5, showed a typical ten-fold higher affinity than the wild-type as determined by biosensor technology. Eight amino acid positions in the Z(F5I variant were separately mutated to cysteine for incorporation of a photoactivable maleimide-benzophenone (MBP group as a probe for site-specific photoconjugation to Fc of mIgG₁, The best photocoupling efficiency to mIgG₁ Fc was seen when the MBP group was coupled to Cys at position 32, resulting in adduct formation to more than 60% of all heavy chains, with no observable non-selective conjugation to the light chains. A similar coupling yield was obtained for a panel of 19 different mIgG₁ mAbs, indicating a general characteristic. To exemplify functionalization of a mIgG₁ antibody via site-specific biotinylation, the Z(F5I-Q32C-MBP protein was first biotinylated using an amine reactive reagent and subsequently photoconjugated to an anti-human interferon-gamma mIgG₁ mAb. When comparing the specific antigen binding ability of the probe-biotinylated mAb to that of the directly biotinylated mAb, a significantly higher bioactivity was observed for the sample biotinylated using the Z(F5I-Q32C-MBP probe. This result indicates that the use of a site-specific and affinity probe

  20. Structural and functional studies of the biotin protein ligase from Aquifex aeolicus reveal a critical role for a conserved residue in target specificity.

    Science.gov (United States)

    Tron, Cecile M; McNae, Iain W; Nutley, Margaret; Clarke, David J; Cooper, Alan; Walkinshaw, Malcolm D; Baxter, Robert L; Campopiano, Dominic J

    2009-03-20

    Biotin protein ligase (BPL; EC 6.3.4.15) catalyses the formation of biotinyl-5'-AMP from biotin and ATP, and the succeeding biotinylation of the biotin carboxyl carrier protein. We describe the crystal structures, at 2.4 A resolution, of the class I BPL from the hyperthermophilic bacteria Aquifex aeolicus (AaBPL) in its ligand-free form and in complex with biotin and ATP. The solvent-exposed beta- and gamma-phosphates of ATP are located in the inter-subunit cavity formed by the N- and C-terminal domains. The Arg40 residue from the conserved GXGRXG motif is shown to interact with the carboxyl group of biotin and to stabilise the alpha- and beta-phosphates of the nucleotide. The structure of the mutant AaBPL R40G in both the ligand-free and biotin-bound forms reveals that the mutated loop has collapsed, thus hindering ATP binding. Isothermal titration calorimetry indicated that the presence of biotin is not required for ATP binding to wild-type AaBPL in the absence of Mg(2+), and the binding of biotin and ATP has been determined to occur via a random but cooperative process. The affinity for biotin is relatively unaffected by the R40G mutation. In contrast, the thermodynamic data indicate that binding of ATP to AaBPL R40G is very weak in the absence or in the presence of biotin. The AaBPL R40G mutant remains catalytically active but shows poor substrate specificity; mass spectrometry and Western blot studies revealed that the mutant biotinylates both the target A. aeolicus BCCPDelta67 fragment and BSA, and is subject to self-biotinylation.

  1. pH and Protein Sensing with Functionalized Semiconducting Oxide Nanobelt FETs

    Science.gov (United States)

    Cheng, Yi; Yun, C. S.; Strouse, G. F.; Xiong, P.; Yang, R. S.; Wang, Z. L.

    2008-03-01

    We report solution pH sensing and selective protein detection with high-performance channel-limited field-effect transistors (FETs) based on single semiconducting oxide (ZnO and SnO2) nanobelts^1. The devices were integrated with PDMS microfluidic channels for analyte delivery and the source/drain contacts were passivated for in-solution sensing. pH sensing experiments were performed on FETs with functionalized and unmodified nanobelts. Functionalization of the nanobelts by APTES was found to greatly improve the pH sensitivity. The change in nanobelt conductance as functions of pH values at different gate voltages and ionic strengths showed high sensitivity and consistency. For the protein detection, we achieved highly selective biotinylation of the nanobelt channel with through APTES linkage. The specific binding of fluorescently-tagged streptavidin to the biotinylated nanobelt was verified by fluorescence microscopy; non-specific binding to the substrate was largely eliminated using PEG-silane passivation. The electrical responses of the biotinylated FETs to the streptavidin binding in PBS buffers of different pH values were systematically measured. The results will be presented and discussed. ^1Y. Cheng et al., Appl. Phys. Lett. 89, 093114 (2006). *Supported by NSF NIRT Grant ECS-0210332.

  2. Optimizing Production of Antigens and Fabs in the Context of Generating Recombinant Antibodies to Human Proteins.

    Directory of Open Access Journals (Sweden)

    Nan Zhong

    Full Text Available We developed and optimized a high-throughput project workflow to generate renewable recombinant antibodies to human proteins involved in epigenetic signalling. Three different strategies to produce phage display compatible protein antigens in bacterial systems were compared, and we found that in vivo biotinylation through the use of an Avi tag was the most productive method. Phage display selections were performed on 265 in vivo biotinylated antigen domains. High-affinity Fabs (<20nM were obtained for 196. We constructed and optimized a new expression vector to produce in vivo biotinylated Fabs in E. coli. This increased average yields up to 10-fold, with an average yield of 4 mg/L. For 118 antigens, we identified Fabs that could immunoprecipitate their full-length endogenous targets from mammalian cell lysates. One Fab for each antigen was converted to a recombinant IgG and produced in mammalian cells, with an average yield of 15 mg/L. In summary, we have optimized each step of the pipeline to produce recombinant antibodies, significantly increasing both efficiency and yield, and also showed that these Fabs and IgGs can be generally useful for chromatin immunoprecipitation (ChIP protocols.

  3. Preparation, biodistribution, and dosimetry of 188Re-Labeled MoAb ior cea1 and its f(ab')2 fragments by avidin-biotin strategy

    International Nuclear Information System (INIS)

    Ferro-Flores, Guillermina; Pimentel-Gonzalez, Gilmara; Gonzalez-Zavala, Maria Antonia; Murphy, Consuelo Arteaga de; Melendez-Alafort, Laura; Tendilla, Jose I.; Croft, Barbara Y.

    1999-01-01

    The biotinylated monoclonal antibody (MoAb) ior cea1 and its F(ab') 2 fragments were labeled with Re-188 by combination of avidin-biotin strategy. 188 Re-MoAb, 188 Re-MoAb-biotin, 188 Re-F(ab') 2 , and 188 Re-F(ab') 2 -biotin preparations were produced for these studies with specific activities of 1.30±0.18 GBq/mg and from instant freeze-dried kit formulations using ethane-1-hydroxy-1,1-diphosphonic acid (EHDP) as a weak competing ligand. There were no significant differences (p>0.05) between the biodistribution in mice of biotinylated and unbiotinylated 188 Re-labeled immunoconjugates. When avidin was injected as a chase after injection of 188 Re-MoAb-biotin or 188 Re-F(ab') 2 -biotin, the blood radioactivity level decreased approximately 75% (cumulated activity) and the effective dose decreased almost 25% with respect to that of the radioimmunoconjugates in which the chase effect was not used. Our results suggest that 188 Re-labeled biotinylated MoAb ior cea1 and its F(ab') 2 fragments prepared by this method are stable complexes in vivo

  4. Signal amplification for impedimetric genosensing using gold-streptavidin nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Bonanni, A.; Esplandiu, M.J. [Sensors and Biosensors Group, Department of Chemistry, Universitat Autonoma de Barcelona, Edifici Cn, 08193 Bellaterra, Barcelona (Spain); Valle, M. del [Sensors and Biosensors Group, Department of Chemistry, Universitat Autonoma de Barcelona, Edifici Cn, 08193 Bellaterra, Barcelona (Spain)], E-mail: manel.delvalle@uab.es

    2008-04-20

    Streptavidin-coated gold nanoparticles (strept-AuNPs) were used in this work to amplify the impedimetric signal generated in a biosensor detecting the DNA hybridization event. Probe oligomer was adsorbed onto a graphite epoxy composite (GEC) electrode surface and the impedance measurement was performed in a solution containing the redox marker ferrocyanide/ferricyanide. The biotinylated complementary oligomer was used as target. The change of interfacial charge transfer resistance (R{sub ct}), experimented by the redox marker, was recorded to confirm the hybrid formation. The addition of strept-AuNPs, binding to the target due to the strong streptavidin-biotin interaction, led to a further increment of R{sub ct} thus obtaining significant signal amplification. Strept-AuNPs on the electrode surface were observed by scanning electron microscopy (SEM) after silver enhancement treatment. A competitive binding assay was also performed using unlabelled DNA target to demonstrate its applicability to real sample analysis.

  5. Visible-light induced photoelectrochemical biosensor for the detection of microRNA based on Bi2S3 nanorods and streptavidin on an ITO electrode

    International Nuclear Information System (INIS)

    Wang, Mo; Yang, Zhiqing; Guo, Yunlong; Wang, Xinxu; Yin, Huanshun; Ai, Shiyun

    2015-01-01

    We demonstrate a photo-electrochemical biosensor for the sensitive and specific detection of microRNA using Bi 2 S 3 nanorods as a photoactive material and streptavidin as the unit that inhibits photocurrent. Bi 2 S 3 nanorods were synthesized hydrothermally in organic phase and displayed excellent light-to-current conversion efficiency. The Bi 2 S 3 was deposited on an indium tin oxide (ITO) slice and then modified with gold nanoparticles onto which biotinylated hairpin probe DNA was deposited as a monolayer. Following hybridization between the biotinylated probe DNA and the target microRNA, the stem-loop structure of the probe DNA was unfolded and the biotin directed outwards into the solution. Streptavidin was then added to bind to biotin via the strong streptavidin-biotin interactions. This causes the photocurrent of the modified ITO to decrease due to steric hindrance that blocks the transfer of electrons from added ascorbic acid to the surface of the electrode. The method has a detection limit as low as 3.5 fM of microRNA and can excellently discriminate even singly mismatched microRNA. The method was successfully applied to investigate the effect of abscisic acid on the expression level of microRNA-159a in seeds of Arabidopsis thaliana. We conclude that the assay presented here has a large potential as a method for quantification of microRNA and for studying the epigenetic regulation of flowering plants. (author)

  6. In vivo detection of c-Met expression in a rat C6 glioma model.

    Science.gov (United States)

    Towner, R A; Smith, N; Doblas, S; Tesiram, Y; Garteiser, P; Saunders, D; Cranford, R; Silasi-Mansat, R; Herlea, O; Ivanciu, L; Wu, D; Lupu, F

    2008-01-01

    The tyrosine kinase receptor, c-Met, and its substrate, the hepatocyte growth factor (HGF), are implicated in the malignant progression of glioblastomas. In vivo detection of c-Met expression may be helpful in the diagnosis of malignant tumours. The C6 rat glioma model is a widely used intracranial brain tumour model used to study gliomas experimentally. We used a magnetic resonance imaging (MRI) molecular targeting agent to specifically tag the cell surface receptor, c-Met, with an anti-c-Met antibody (Ab) linked to biotinylated Gd (gadolinium)-DTPA (diethylene triamine penta acetic acid)-albumin in rat gliomas to detect overexpression of this antigen in vivo. The anti-c-Met probe (anti-c-Met-Gd-DTPA-albumin) was administered intravenously, and as determined by an increase in MRI signal intensity and a corresponding decrease in regional T(1) relaxation values, this probe was found to detect increased expression of c-Met protein levels in C6 gliomas. In addition, specificity for the binding of the anti-c-Met contrast agent was determined by using fluorescence microscopic imaging of the biotinylated portion of the targeting agent within neoplastic and 'normal'brain tissues following in vivo administration of the anti-c-Met probe. Controls with no Ab or with a normal rat IgG attached to the contrast agent component indicated no non-specific binding to glioma tissue. This is the first successful visualization of in vivo overexpression of c-Met in gliomas.

  7. A new application of scanning electrochemical microscopy for the label-free interrogation of antibody-antigen interactions

    Energy Technology Data Exchange (ETDEWEB)

    Holmes, Joanne L.; Davis, Frank; Collyer, Stuart D. [Cranfield Health, Cranfield University, Cranfield, MK43 0AL (United Kingdom); Higson, Seamus P.J., E-mail: s.p.j.higson@cranfield.ac.uk [Cranfield Health, Cranfield University, Cranfield, MK43 0AL (United Kingdom)

    2011-03-18

    Within this work we present a 'proof of principle' study for the use of scanning electrochemical microscopy (SECM) to detect and image biomolecular interactions in a label-free assay as a potential alternative to current fluorescence techniques. Screen-printed carbon electrodes were used as the substrate for the deposition of a dotted array, where the dots consist of biotinylated polyethyleneimine. These were then further derivatised, first with neutravidin and then with a biotinylated antibody to the protein neuron specific enolase (NSE). SECM using a ferrocene carboxylic acid mediator showed clear differences between the array and the surrounding unmodified carbon. Imaging of the arrays before and following exposure to various concentrations of the antigen showed clear evidence for specific binding of the NSE antigen to the antibody derivatised dots. Non-specific binding was quantified. Control experiments with other proteins showed only non-specific binding across the whole of the substrate, thereby confirming that specific binding does occur between the antibody and antigen at the surface of the dots. Binding of the antigen was accompanied by a measured increase in current response, which may be explained in terms of protein electrostatic interaction and hydrophobic interactions to the mediator, thereby increasing the localised mediator flux. A calibration curve was obtained between 500 fg mL{sup -1} to 200 pg mL{sup -1} NSE which demonstrated a logarithmic relationship between the current change upon binding and antigen concentration without the need for any labelling of the substrate.

  8. A Quantitative Electrochemiluminescence Assay for Clostridium perfringens alpha toxin

    National Research Council Canada - National Science Library

    Merrill, Gerald A; Rivera, Victor R; Neal, Dwayne D; Young, Charles; Poli, Mark A

    2006-01-01

    .... Biotinylated antibodies to C. perfringens alpha toxin bound to streptavidin paramagnetic beads specifically immunoadsorbed soluble sample alpha toxin which subsequently selectively immunoadsorbed ruthenium (Ru...

  9. Surface excitation parameter for rough surfaces

    International Nuclear Information System (INIS)

    Da, Bo; Salma, Khanam; Ji, Hui; Mao, Shifeng; Zhang, Guanghui; Wang, Xiaoping; Ding, Zejun

    2015-01-01

    Graphical abstract: - Highlights: • Instead of providing a general mathematical model of roughness, we directly use a finite element triangle mesh method to build a fully 3D rough surface from the practical sample. • The surface plasmon excitation can be introduced to the realistic sample surface by dielectric response theory and finite element method. • We found that SEP calculated based on ideal plane surface model are still reliable for real sample surface with common roughness. - Abstract: In order to assess quantitatively the importance of surface excitation effect in surface electron spectroscopy measurement, surface excitation parameter (SEP) has been introduced to describe the surface excitation probability as an average number of surface excitations that electrons can undergo when they move through solid surface either in incoming or outgoing directions. Meanwhile, surface roughness is an inevitable issue in experiments particularly when the sample surface is cleaned with ion beam bombardment. Surface roughness alters not only the electron elastic peak intensity but also the surface excitation intensity. However, almost all of the popular theoretical models for determining SEP are based on ideal plane surface approximation. In order to figure out whether this approximation is efficient or not for SEP calculation and the scope of this assumption, we proposed a new way to determine the SEP for a rough surface by a Monte Carlo simulation of electron scattering process near to a realistic rough surface, which is modeled by a finite element analysis method according to AFM image. The elastic peak intensity is calculated for different electron incident and emission angles. Assuming surface excitations obey the Poisson distribution the SEPs corrected for surface roughness are then obtained by analyzing the elastic peak intensity for several materials and for different incident and emission angles. It is found that the surface roughness only plays an

  10. Improved antifouling properties and selective biofunctionalization of stainless steel by employing heterobifunctional silane-polyethylene glycol overlayers and avidin-biotin technology

    Science.gov (United States)

    Hynninen, Ville; Vuori, Leena; Hannula, Markku; Tapio, Kosti; Lahtonen, Kimmo; Isoniemi, Tommi; Lehtonen, Elina; Hirsimäki, Mika; Toppari, J. Jussi; Valden, Mika; Hytönen, Vesa P.

    2016-01-01

    A straightforward solution-based method to modify the biofunctionality of stainless steel (SS) using heterobifunctional silane-polyethylene glycol (silane-PEG) overlayers is reported. Reduced nonspecific biofouling of both proteins and bacteria onto SS and further selective biofunctionalization of the modified surface were achieved. According to photoelectron spectroscopy analyses, the silane-PEGs formed less than 10 Å thick overlayers with close to 90% surface coverage and reproducible chemical compositions. Consequently, the surfaces also became more hydrophilic, and the observed non-specific biofouling of proteins was reduced by approximately 70%. In addition, the attachment of E. coli was reduced by more than 65%. Moreover, the potential of the overlayer to be further modified was demonstrated by successfully coupling biotinylated alkaline phosphatase (bAP) to a silane-PEG-biotin overlayer via avidin-biotin bridges. The activity of the immobilized enzyme was shown to be well preserved without compromising the achieved antifouling properties. Overall, the simple solution-based approach enables the tailoring of SS to enhance its activity for biomedical and biotechnological applications. PMID:27381834

  11. Improved antifouling properties and selective biofunctionalization of stainless steel by employing heterobifunctional silane-polyethylene glycol overlayers and avidin-biotin technology

    Science.gov (United States)

    Hynninen, Ville; Vuori, Leena; Hannula, Markku; Tapio, Kosti; Lahtonen, Kimmo; Isoniemi, Tommi; Lehtonen, Elina; Hirsimäki, Mika; Toppari, J. Jussi; Valden, Mika; Hytönen, Vesa P.

    2016-07-01

    A straightforward solution-based method to modify the biofunctionality of stainless steel (SS) using heterobifunctional silane-polyethylene glycol (silane-PEG) overlayers is reported. Reduced nonspecific biofouling of both proteins and bacteria onto SS and further selective biofunctionalization of the modified surface were achieved. According to photoelectron spectroscopy analyses, the silane-PEGs formed less than 10 Å thick overlayers with close to 90% surface coverage and reproducible chemical compositions. Consequently, the surfaces also became more hydrophilic, and the observed non-specific biofouling of proteins was reduced by approximately 70%. In addition, the attachment of E. coli was reduced by more than 65%. Moreover, the potential of the overlayer to be further modified was demonstrated by successfully coupling biotinylated alkaline phosphatase (bAP) to a silane-PEG-biotin overlayer via avidin-biotin bridges. The activity of the immobilized enzyme was shown to be well preserved without compromising the achieved antifouling properties. Overall, the simple solution-based approach enables the tailoring of SS to enhance its activity for biomedical and biotechnological applications.

  12. Enzyme-labeled Antigen Method: Development and Application of the Novel Approach for Identifying Plasma Cells Locally Producing Disease-specific Antibodies in Inflammatory Lesions

    International Nuclear Information System (INIS)

    Mizutani, Yasuyoshi; Shiogama, Kazuya; Onouchi, Takanori; Sakurai, Kouhei; Inada, Ken-ichi; Tsutsumi, Yutaka

    2016-01-01

    In chronic inflammatory lesions of autoimmune and infectious diseases, plasma cells are frequently observed. Antigens recognized by antibodies produced by the plasma cells mostly remain unclear. A new technique identifying these corresponding antigens may give us a breakthrough for understanding the disease from a pathophysiological viewpoint, simply because the immunocytes are seen within the lesion. We have developed an enzyme-labeled antigen method for microscopic identification of the antigen recognized by specific antibodies locally produced in plasma cells in inflammatory lesions. Firstly, target biotinylated antigens were constructed by the wheat germ cell-free protein synthesis system or through chemical biotinylation. Next, proteins reactive to antibodies in tissue extracts were screened and antibody titers were evaluated by the AlphaScreen method. Finally, with the enzyme-labeled antigen method using the biotinylated antigens as probes, plasma cells producing specific antibodies were microscopically localized in fixed frozen sections. Our novel approach visualized tissue plasma cells that produced 1) autoantibodies in rheumatoid arthritis, 2) antibodies against major antigens of Porphyromonas gingivalis in periodontitis or radicular cyst, and 3) antibodies against a carbohydrate antigen, Strep A, of Streptococcus pyogenes in recurrent tonsillitis. Evaluation of local specific antibody responses expectedly contributes to clarifying previously unknown processes in inflammatory disorders

  13. tRNA-mediated labelling of proteins with biotin. A nonradioactive method for the detection of cell-free translation products.

    Science.gov (United States)

    Kurzchalia, T V; Wiedmann, M; Breter, H; Zimmermann, W; Bauschke, E; Rapoport, T A

    1988-03-15

    We have developed a new method for the rapid and sensitive detection of cell-free translation products. Biotinylated lysine is incorporated into newly synthesized proteins by means of lysyl-tRNA that is modified in the epsilon-position. After electrophoresis in a dodecyl sulfate gel and blotting onto nitrocellulose, the translation products can be identified by probing with streptavidin and biotinylated alkaline phosphatase, followed by incubation with a chromogenic enzyme substrate. The non-radioactive labelling by biotin approaches in its sensitivity that obtained by radioactive amino acids. The products are absolutely stable and can be rapidly identified. The new method has been tested with different mRNAs in the cell-free translation systems of wheat germ and reticulocytes. Neither the interaction of secretory proteins with the signal recognition particle nor the in vitro translocation across the endoplasmic reticulum membrane or core glycosylation of nascent polypeptides are prevented by the incorporation of biotinylated lysine residues. The results indicate that both the ribosome and the endoplasmic reticulum membrane permit the passage of polypeptides carrying bulky groups attached to the amino acids (by atomic models it was estimated that the size of the side chain of lysine changes from approximately equal to 0.8 nm to approximately equal to 2 nm after modification.

  14. Control of biotin biosynthesis in mycobacteria by a pyruvate carboxylase dependent metabolic signal.

    Science.gov (United States)

    Lazar, Nathaniel; Fay, Allison; Nandakumar, Madhumitha; Boyle, Kerry E; Xavier, Joao; Rhee, Kyu; Glickman, Michael S

    2017-12-01

    Biotin is an essential cofactor utilized by all domains of life, but only synthesized by bacteria, fungi and plants, making biotin biosynthesis a target for antimicrobial development. To understand biotin biosynthesis in mycobacteria, we executed a genetic screen in Mycobacterium smegmatis for biotin auxotrophs and identified pyruvate carboxylase (Pyc) as required for biotin biosynthesis. The biotin auxotrophy of the pyc::tn strain is due to failure to transcriptionally induce late stage biotin biosynthetic genes in low biotin conditions. Loss of bioQ, the repressor of biotin biosynthesis, in the pyc::tn strain reverted biotin auxotrophy, as did reconstituting the last step of the pathway through heterologous expression of BioB and provision of its substrate DTB. The role of Pyc in biotin regulation required its catalytic activities and could be supported by M. tuberculosis Pyc. Quantitation of the kinetics of depletion of biotinylated proteins after biotin withdrawal revealed that Pyc is the most rapidly depleted biotinylated protein and metabolomics revealed a broad metabolic shift in wild type cells upon biotin withdrawal which was blunted in cell lacking Pyc. Our data indicate that mycobacterial cells monitor biotin sufficiency through a metabolic signal generated by dysfunction of a biotinylated protein of central metabolism. © 2017 John Wiley & Sons Ltd.

  15. The synthesis and characterization of biotin-silver-dendrimer nanocomposites as novel bioselective labels

    Energy Technology Data Exchange (ETDEWEB)

    Maly, J; Lampova, H; Semeradtova, A; Stofik, M [Faculty of Science, University of J E Purkynje, 40096 Usti nad Labem (Czech Republic); Kovacik, L, E-mail: malyjalga@seznam.c [Institute of Cellular Biology and Pathology, First Faculty of Medicine, Charles University in Prague (Czech Republic)

    2009-09-23

    This paper presents a synthesis of a novel nanoparticle label with selective biorecognition properties based on a biotinylated silver-dendrimer nanocomposite (AgDNC). Two types of labels, a biotin-AgDNC (bio-AgDNC) and a biotinylated AgDNC with a poly(ethylene)glycol spacer (bio-PEG-AgDNC), were synthesized from a generation 7 (G7) hydroxyl-terminated ethylenediamine-core-type (2-carbon core) PAMAM dendrimer (DDM) by an N,N'-dicyclohexylcarbodiimide (DDC) biotin coupling and a NaBH{sub 4} silver reduction method. Synthesized conjugates were characterized by several analytical methods, such as UV-vis, FTIR, AFM, TEM, ELISA, HABA assay and SPR. The results show that stable biotinylated nanocomposites can be formed either with internalized silver nanoparticles (AgNPs) in a DMM polymer backbone ('type I') or as externally protected ('type E'), depending on the molar ratio of the silver/DMM conjugate and type of conjugate. Furthermore, the selective biorecognition function of the biotin is not affected by the AgNPs' synthesis step, which allows a potential application of silver nanocomposite conjugates as biospecific labels in various bioanalytical assays, or potentially as fluorescence cell biomarkers. An exploitation of the presented label in the development of electrochemical immunosensors is anticipated.

  16. Artificial Metalloenzymes Based on the Biotin-Streptavidin Technology: Challenges and Opportunities.

    Science.gov (United States)

    Heinisch, Tillmann; Ward, Thomas R

    2016-09-20

    The biotin-streptavidin technology offers an attractive means to engineer artificial metalloenzymes (ArMs). Initiated over 50 years ago by Bayer and Wilchek, the biotin-(strept)avidin techonology relies on the exquisite supramolecular affinity of either avidin or streptavidin for biotin. This versatile tool, commonly referred to as "molecular velcro", allows nearly irreversible anchoring of biotinylated probes within a (strept)avidin host protein. Building upon a visionary publication by Whitesides from 1978, several groups have been exploiting this technology to create artificial metalloenzymes. For this purpose, a biotinylated organometallic catalyst is introduced within (strept)avidin to afford a hybrid catalyst that combines features reminiscent of both enzymes and organometallic catalysts. Importantly, ArMs can be optimized by chemogenetic means. Combining a small collection of biotinylated organometallic catalysts with streptavidin mutants allows generation of significant diversity, thus allowing optimization of the catalytic performance of ArMs. Pursuing this strategy, the following reactions have been implemented: hydrogenation, alcohol oxidation, sulfoxidation, dihydroxylation, allylic alkylation, transfer hydrogenation, Suzuki cross-coupling, C-H activation, and metathesis. In this Account, we summarize our efforts in the latter four reactions. X-ray analysis of various ArMs based on the biotin-streptavidin technology reveals the versatility and commensurability of the biotin-binding vestibule to accommodate and interact with transition states of the scrutinized organometallic transformations. In particular, streptavidin residues at positions 112 and 121 recurrently lie in close proximity to the biotinylated metal cofactor. This observation led us to develop a streamlined 24-well plate streptavidin production and screening platform to optimize the performance of ArMs. To date, most of the efforts in the field of ArMs have focused on the use of purified

  17. In vivo evaluation of an anti-PSMA antibody conjugated with varying numbers of biotin molecules in a pretargeting protocol

    International Nuclear Information System (INIS)

    Wilbur, D.S.; Hamlin, D.K.; Quinn, J.; Vessella, R.L.

    2003-01-01

    An investigation has been conducted to determine the effect of varying the number of biotin molecules conjugated with an anti-PSMA antibody (mAb) as part of our studies to optimize biotinylated antibodies and radiolabeled streptavidin in pretargeting protocols for Targeted Radionuclide Therapy of prostate cancer. In the investigation, the anti-PSMA antibody 107-1A4 was biotinylated with varying amounts of biotinamidocaproate N-hydroxysuccinimide ester. This procedure resulted in obtaining 107-1A4 with 2.3, 4.5, and 6.8 biotin conjugated as measured by the standard HABA assay. The biotinylated 107-1A4 was radioiodinated and was evaluated in a pretargeting protocol in athymic mice bearing LNCaP human tumor xenografts. In the protocol, 50 μg biotinylated [ 125 I]107-1A4 was injected, followed 48h later by 25 μg of avidin for blood clearance, and 1h after that 20 μg of radiolabeled succinylated recombinant streptavidin ([ 13 1I]sSAv) was administered. The tumor localization and tissue distribution was evaluated at 24, 48, and 72h post [ 131 I]sSAv injection. With 2.3 biotin/mAb, an approximate 1:1 molar ratio (4-5 pmol/g) of sSAv/mAb was obtained at all three time points. With 4.5 biotin/mAb, a 1:1 ratio was observed at 24h, but approx. 2: 1 was observed at 48 and 72h pi. With 6.8 biotin/mAb, sSAv/mAb ratios of approximately 1.5:1; 2:1; and 3:1 were obtained at 24, 48, and 72h pi respectively. The amount of sSAv localized in the tumor was nearly the same (4-5 pmol/g) when 107-1A4 had 2.3 or 4.5 biotin conjugated, but decreased to 3-4.5 pmol/g with 6.8 biotin conjugated. Because the highest levels of co-localized sSAv was found with the lowest number of biotin conjugates, the observed differences in ratios of sSAv/mAb may be best explained as differences in internalization, and degradation of mAb and protease resistant sSAv. In duplicate experiments, similar results were obtained with biotinylated 107-1A4 F(ab') 2 , but not with an mAb to a non-internalizing antigen

  18. Imaging Early Steps of Sindbis Virus Infection by Total Internal Reflection Fluorescence Microscopy

    Directory of Open Access Journals (Sweden)

    Youling Gu

    2011-01-01

    Full Text Available Sindbis virus (SINV is an alphavirus that has a broad host range and has been widely used as a vector for recombinant gene transduction, DNA-based vaccine production, and oncolytic cancer therapy. The mechanism of SINV entry into host cells has yet to be fully understood. In this paper, we used single virus tracking under total internal reflection fluorescence microscopy (TIRFM to investigate SINV attachment to cell surface. Biotinylated viral particles were labeled with quantum dots, which retained viral viability and infectivity. By time-lapse imaging, we showed that the SINV exhibited a heterogeneous dynamics on the surface of the host cells. Analysis of SINV motility demonstrated a two-step attachment reaction. Moreover, dual color TIRFM of GFP-Rab5 and SINV suggested that the virus was targeted to the early endosomes after endocytosis. These findings demonstrate the utility of quantum dot labeling in studying the early steps and behavior of SINV infection.

  19. A Rhizavidin Monomer with Nearly Multimeric Avidin-Like Binding Stability Against Biotin Conjugates.

    Science.gov (United States)

    Lee, Jeong Min; Kim, Jung A; Yen, Tzu-Chi; Lee, In Hwan; Ahn, Byungjun; Lee, Younghoon; Hsieh, Chia-Lung; Kim, Ho Min; Jung, Yongwon

    2016-03-01

    Developing a monomeric form of an avidin-like protein with highly stable biotin binding properties has been a major challenge in biotin-avidin linking technology. Here we report a monomeric avidin-like protein-enhanced monoavidin-with off-rates almost comparable to those of multimeric avidin proteins against various biotin conjugates. Enhanced monoavidin (eMA) was developed from naturally dimeric rhizavidin by optimally maintaining protein rigidity during monomerization and additionally shielding the bound biotin by diverse engineering of the surface residues. eMA allowed the monovalent and nonperturbing labeling of head-group-biotinylated lipids in bilayer membranes. In addition, we fabricated an unprecedented 24-meric avidin probe by fusing eMA to a multimeric cage protein. The 24-meric avidin and eMA were utilized to demonstrate how artificial clustering of cell-surface proteins greatly enhances the internalization rates of assembled proteins on live cells. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Enzyme-enhanced fluorescence detection of DNA on etched optical fibers.

    Science.gov (United States)

    Niu, Shu-yan; Li, Quan-yi; Ren, Rui; Zhang, Shu-sheng

    2009-05-15

    A novel DNA biosensor based on enzyme-enhanced fluorescence detection on etched optical fibers was developed. The hybridization complex of DNA probe and biotinylated target was formed on the etched optical fiber, and was then bound with streptavidin labeled horseradish peroxidase (streptavidin-HRP). The target DNA was quantified through the fluorescent detection of bi-p,p'-4-hydroxyphenylacetic acid (DBDA) generated from the substrate 4-hydroxyphenylacetic acid (p-HPA) under the catalysis of HRP, with a detection limit of 1 pM and a linear range from 1.69 pM to 169 pM. It is facile to regenerate this sensor through surface treatment with concentrated urea solution. It was discovered that the sensor can retain 70% of its original activity after three detection-regeneration cycles.

  1. Application of magnetic poly(styrene-glycidyl methacrylate) microspheres for immunomagnetic separation of bone marrow cells

    Energy Technology Data Exchange (ETDEWEB)

    Chung, T.-H.; Chang, J.-Y. [Department of Chemical Engineering, National Chung Cheng University, Chiayi 621, Taiwan (China); Lee, W.-C. [Department of Chemical Engineering, National Chung Cheng University, Chiayi 621, Taiwan (China)], E-mail: chmwcl@ccu.edu.tw

    2009-05-15

    Surface-functionalized magnetic poly(styrene-glycidyl methacrylate) (PS-GMA) microspheres were prepared and coupled with Sca-1 antibody for cell selection from murine bone marrow mononuclear cells (MNCs). Biotinylated Sca-1 antibody could be directly coupled to avidin-bound magnetic microspheres. Alternatively, oxidized goat anti-mouse antibody was covalently bound onto the amino group-containing magnetic microspheres in a site-directed manner, and the resultant conjugate was coupled with non-modified Sca-1 antibody. Using the indirect antibody-bound magnetic microspheres, the purity of isolated Sca-1{sup +} cells increased with bead-to-cell ratio. Using a bead-to-cell ratio of 10 beads/cell, a purity of 85% Sca-1{sup +} cells corresponding to a 17-fold enrichment was achieved.

  2. Matrix-immobilized BMP-2 on microcontact printed fibronectin as in vitro tool to study BMP-mediated signaling and cell migration

    Directory of Open Access Journals (Sweden)

    Kristin eHauff

    2015-05-01

    Full Text Available During development, bone morphogenetic proteins (BMPs exert important functions in several tissues by regulating signaling for cell differentiation and migration. In vivo the extracellular matrix (ECM not only provides a support for adherent cells, but also presents a reservoir of growth factors (GFs. Several constituents of the ECM provide adhesive cues, which serve as binding sites for cell transmembrane receptors, such as integrins, which convey adhesion-mediated signaling to the intracellular compartment. Integrins do not function alone but rather crosstalk and cooperate with other receptors, such as GF receptors, in regulating cell responses to extracellular signals. To this, we present here the immobilization of BMP-2 onto cellular fibronectin (cFN, a key protein of the ECM, to investigate their impact on GF-mediated signaling and migration.Following biotinylation, BMP-2 was linked to biotinylated cFN using NeutrAvidin (NA as cross-linker. Characterization with QCM-D and ELISA confirmed the efficient immobilization of BMP-2 on cFN over a period of 24 h.To validate the bioactivity of matrix-immobilized BMP-2 (iBMP-2 we investigated short- and long-term responses of C2C12 myoblasts in comparison to soluble BMP-2 (sBMP-2 or in absence of GFs. Similarly to sBMP-2, iBMP-2 triggered Smad 1/5 phosphorylation and translocation into the nucleus corresponding to the activation of BMP-mediated Smad-dependent pathway. Additionally, successful suppression of myotube formation was observed after six days.We next implemented this approach to fabricate cFN micro patterned stripes by soft lithography. These stripes only allowed cell-surface interaction on the pattern due to passivation of the surface in between, thus serving as platform for studies on directed cell migration. During a 10 h-period, cells showed an increased migratory activity upon BMP-2 exposure.Thus, this versatile tool retains the GF's bioactivity and allows the presentation of ECM

  3. Surface Water & Surface Drainage

    Data.gov (United States)

    Earth Data Analysis Center, University of New Mexico — This data set contains boundaries for all surface water and surface drainage for the state of New Mexico. It is in a vector digital data structure digitized from a...

  4. Modification of rubber surface by UV surface grafting

    International Nuclear Information System (INIS)

    Shanmugharaj, A.M.; Kim, Jin Kuk; Ryu, Sung Hun

    2006-01-01

    Rubber surface is subjected to ultraviolet radiation (UV) in the presence of allylamine and radiation sensitizer benzophenone (BP). Fourier transform infrared spectral studies reveal the presence of allylamine on the surface. The presence of irregular needle shapes on the surface as observed in scanning electron micrographs also confirms the polymerized allylamine on the surface. Allylamine coatings have been further confirmed from atomic force microscopy (AFM) analysis. Thermogravimetric analysis (TGA) reveals that allylamine coating on the rubber surface lowers the thermal degradation rate. The contact angle between the water and rubber surface decreases for the modified rubber surface confirming the surface modification due to UV surface grafting

  5. Surface Topography Hinders Bacterial Surface Motility.

    Science.gov (United States)

    Chang, Yow-Ren; Weeks, Eric R; Ducker, William A

    2018-03-21

    We demonstrate that the surface motility of the bacterium, Pseudomonas aeruginosa, is hindered by a crystalline hemispherical topography with wavelength in the range of 2-8 μm. The motility was determined by the analysis of time-lapse microscopy images of cells in a flowing growth medium maintained at 37 °C. The net displacement of bacteria over 5 min is much lower on surfaces containing 2-8 μm hemispheres than on flat topography, but displacement on the 1 μm hemispheres is not lower. That is, there is a threshold between 1 and 2 μm for response to the topography. Cells on the 4 μm hemispheres were more likely to travel parallel to the local crystal axis than in other directions. Cells on the 8 μm topography were less likely to travel across the crowns of the hemispheres and were also more likely to make 30°-50° turns than on flat surfaces. These results show that surface topography can act as a significant barrier to surface motility and may therefore hinder surface exploration by bacteria. Because surface exploration can be a part of the process whereby bacteria form colonies and seek nutrients, these results help to elucidate the mechanism by which surface topography hinders biofilm formation.

  6. Detecting Extracellular Carbonic Anhydrase Activity Using Membrane Inlet Mass Spectrometry

    Science.gov (United States)

    Delacruz, Joannalyn; Mikulski, Rose; Tu, Chingkuang; Li, Ying; Wang, Hai; Shiverick, Kathleen T.; Frost, Susan C.; Horenstein, Nicole A.; Silverman, David N.

    2010-01-01

    Current research into the function of carbonic anhydrases in cell physiology emphasizes the role of membrane-bound carbonic anhydrases, such as carbonic anhydrase IX that has been identified in malignant tumors and is associated with extracellular acidification as a response to hypoxia. We present here a mass spectrometric method to determine the extent to which total carbonic anhydrase activity is due to extracellular carbonic anhydrase in whole cell preparations. The method is based on the biphasic rate of depletion of 18O from CO2 measured by membrane inlet mass spectrometry. The slopes of the biphasic depletion are a sensitive measure of the presence of carbonic anhydrase outside and inside of the cells. This property is demonstrated here using suspensions of human red cells in which external carbonic anhydrase was added to the suspending solution. It is also applied to breast and prostate cancer cells which both express exofacial carbonic anhydrase IX. Inhibition of external carbonic anhydrase is achieved by use of a membrane impermeant inhibitor that was synthesized for this purpose, p-aminomethylbenzenesulfonamide attached to a polyethyleneglycol polymer. PMID:20417171

  7. Theory of quasiparticle surface states in semiconductor surfaces

    International Nuclear Information System (INIS)

    Hybertsen, M.S.; Louie, S.G.

    1988-01-01

    A first-principles theory of the quasiparticle surface-state energies on semiconductor surfaces is developed. The surface properties are calculated using a repeated-slab geometry. Many-body effects due to the electron-electron interaction are represented by the electron self-energy operator including the full surface Green's function and local fields and dynamical screening effects in the Coulomb interaction. Calculated surface-state energies for the prototypical Si(111):As and Ge(111):As surfaces are presented. The calculated energies and dispersions for the occupied surface states (resonances) are in excellent agreement with recent angle-resolved photoemission data. Predictions are made for the position of empty surface states on both surfaces which may be experimentally accessible. The resulting surface state gap at Gamma-bar for Si(111):As agrees with recent scanning-tunneling-spectroscopy measurements. Comparison of the present results to eigenvalues from the local-density-functional calculation reveals substantial corrections for the gaps between empty and occupied surface states. This correction is found to depend on the character of the surface states involved

  8. Effect of microbubble ligation to cells on ultrasound signal enhancement: implications for targeted imaging.

    Science.gov (United States)

    Lankford, Miles; Behm, Carolyn Z; Yeh, James; Klibanov, Alexander L; Robinson, Peter; Lindner, Jonathan R

    2006-10-01

    Molecular imaging with contrast-enhanced ultrasound (CEU) relies on the detection of microbubbles retained in regions of disease. The aim of this study was to determine whether microbubble attachment to cells influences their acoustic signal generation and stability. Biotinylated microbubbles were attached to streptavidin-coated plates to derive density versus intensity relations during low- and high-power imaging. To assess damping from microbubble attachment to solid or cell surfaces, in vitro imaging was performed for microbubbles charge-coupled to methacrylate spheres and for vascular cell adhesion molecule-1-targeted microbubbles attached to endothelial cells. Signal enhancement on plates increased according to acoustic power and microbubble site density up to 300 mm. Microbubble signal was reduced by attachment to solid spheres during high- and low-power imaging but was minimally reduced by attachment to endothelial cells and only at low power. Attachment of targeted microbubbles to rigid surfaces results in damping and a reduction of their acoustic signal, which is not seen when microbubbles are attached to cells. A reliable concentration versus intensity relationship can be expected from microbubble attachment to 2-dimensional surfaces until a very high site density is reached.

  9. Fabrication of protein microarrays for alpha fetoprotein detection by using a rapid photo-immobilization process

    Directory of Open Access Journals (Sweden)

    Sirasa Yodmongkol

    2016-03-01

    Full Text Available In this study, protein microarrays based on sandwich immunoassays are generated to quantify the amount of alpha fetoprotein (AFP in blood serum. For chip generation a mixture of capture antibody and a photoactive copolymer consisting of N,N-dimethylacrylamide (DMAA, methacryloyloxy benzophenone (MaBP, and Na-4-styrenesulfonate (SSNa was spotted onto unmodified polymethyl methacrylate (PMMA substrates. Subsequently to printing of the microarray, the polymer and protein were photochemically cross-linked and the forming, biofunctionalized hydrogels simultaneously bound to the chip surface by short UV- irradiation. The obtained biochip was incubated with AFP antigen, followed by biotinylated AFP antibody and streptavidin-Cy5 and the fluorescence signal read-out. The developed microarray biochip covers the range of AFP in serum samples such as maternal serum in the range of 5 and 100 ng/ml. The chip production process is based on a fast and simple immobilization process, which can be applied to conventional plastic surfaces. Therefore, this protein microarray production process is a promising method to fabricate biochips for AFP screening processes. Keywords: Photo-immobilization, Protein microarray, Alpha fetoprotein, Hydrogel, 3D surface, Down syndrome

  10. Identification of redox-sensitive cysteines in the arabidopsis proteome using OxiTRAQ, a quantitative redox proteomics method

    KAUST Repository

    Liu, Pei; Zhang, Huoming; Wang, Hai; Xia, Yiji

    2014-01-01

    -throughput quantitative proteomic approach termed OxiTRAQ for identifying proteins whose thiols undergo reversible oxidative modifications in Arabidopsis cells subjected to oxidative stress. In this approach, a biotinylated thiol-reactive reagent is used for differential

  11. Characterization of newly established colorectal cancer cell lines ...

    Indian Academy of Sciences (India)

    Unknown

    2000-12-19

    Dec 19, 2000 ... ... Medicine, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, New York, NY 10021, USA .... with biotinylated horse anti-mouse IgG (Vector Labora- ...... development of new therapies or prognosis of the disease,.

  12. PREFACE: Vibrations at surfaces Vibrations at surfaces

    Science.gov (United States)

    Rahman, Talat S.

    2011-12-01

    This special issue is dedicated to the phenomenon of vibrations at surfaces—a topic that was indispensible a couple of decades ago, since it was one of the few phenomena capable of revealing the nature of binding at solid surfaces. For clean surfaces, the frequencies of modes with characteristic displacement patterns revealed how surface geometry, as well as the nature of binding between atoms in the surface layers, could be different from that in the bulk solid. Dispersion of the surface phonons provided further measures of interatomic interactions. For chemisorbed molecules on surfaces, frequencies and dispersion of the vibrational modes were also critical for determining adsorption sites. In other words, vibrations at surfaces served as a reliable means of extracting information about surface structure, chemisorption and overlayer formation. Experimental techniques, such as electron energy loss spectroscopy and helium-atom-surface scattering, coupled with infra-red spectroscopy, were continually refined and their resolutions enhanced to capture subtleties in the dynamics of atoms and molecules at surfaces. Theoretical methods, whether based on empirical and semi-empirical interatomic potential or on ab initio electronic structure calculations, helped decipher experimental observations and provide deeper insights into the nature of the bond between atoms and molecules in regions of reduced symmetry, as encountered on solid surfaces. Vibrations at surfaces were thus an integral part of the set of phenomena that characterized surface science. Dedicated workshops and conferences were held to explore the variety of interesting and puzzling features revealed in experimental and theoretical investigations of surface vibrational modes and their dispersion. One such conference, Vibrations at Surfaces, first organized by Harald Ibach in Juelich in 1980, continues to this day. The 13th International Conference on Vibrations at Surfaces was held at the University of

  13. An Impedance Aptasensor with Microfluidic Chips for Specific Detection of H5N1 Avian Influenza Virus

    Directory of Open Access Journals (Sweden)

    Jacob Lum

    2015-07-01

    Full Text Available In this research a DNA aptamer, which was selected through SELEX (systematic evolution of ligands by exponential enrichment to be specific against the H5N1 subtype of the avian influenza virus (AIV, was used as an alternative reagent to monoclonal antibodies in an impedance biosensor utilizing a microfluidics flow cell and an interdigitated microelectrode for the specific detection of H5N1 AIV. The gold surface of the interdigitated microelectrode embedded in a microfluidics flow cell was modified using streptavidin. The biotinylated aptamer against H5N1 was then immobilized on the electrode surface using biotin–streptavidin binding. The target virus was captured on the microelectrode surface, causing an increase in impedance magnitude. The aptasensor had a detection time of 30 min with a detection limit of 0.0128 hemagglutinin units (HAU. Scanning electron microscopy confirmed the binding of the target virus onto the electrode surface. The DNA aptamer was specific to H5N1 and had no cross-reaction to other subtypes of AIV (e.g., H1N1, H2N2, H7N2. The newly developed aptasensor offers a portable, rapid, low-cost alternative to current methods with the same sensitivity and specificity.

  14. Glycine transporter dimers: evidence for occurrence in the plasma membrane.

    Science.gov (United States)

    Bartholomäus, Ingo; Milan-Lobo, Laura; Nicke, Annette; Dutertre, Sébastien; Hastrup, Hanne; Jha, Alok; Gether, Ulrik; Sitte, Harald H; Betz, Heinrich; Eulenburg, Volker

    2008-04-18

    Different Na(+)/Cl(-)-dependent neurotransmitter transporters of the SLC6a family have been shown to form dimers or oligomers in both intracellular compartments and at the cell surface. In contrast, the glycine transporters (GlyTs) GlyT1 and -2 have been reported to exist as monomers in the plasma membrane based on hydrodynamic and native gel electrophoretic studies. Here, we used cysteine substitution and oxidative cross-linking to show that of GlyT1 and GlyT2 also form dimeric complexes within the plasma membrane. GlyT oligomerization at the cell surface was confirmed for both GlyT1 and GlyT2 by fluorescence resonance energy transfer microscopy. Endoglycosidase treatment and surface biotinylation further revealed that complex-glycosylated GlyTs form dimers located at the cell surface. Furthermore, substitution of tryptophan 469 of GlyT2 by an arginine generated a transporter deficient in dimerization that was retained intracellulary. Based on these results and GlyT structures modeled by using the crystal structure of the bacterial homolog LeuT(Aa), as a template, residues located within the extracellular loop 3 and at the beginning of transmembrane domain 6 are proposed to contribute to the dimerization interface of GlyTs.

  15. Induced surface stress at crystal surfaces

    International Nuclear Information System (INIS)

    Dahmen, K.

    2002-05-01

    Changes of the surfaces stress Δτ (s) can be studied by observing the bending of thin crystalline plates. With this cantilever method one can gain the induced change of surface stress Δτ (s) from the bending of plates with the help of elasticity theory. For elastic isotropic substrates the relevant relations are known. Here the relations are generalized to elastic anisotropic crystals with a C 2v - Symmetry. The equilibrium shapes of crystalline plates oriented along the (100)-, (110)-, or (111)-direction which are clamped along one edge are calculated with a numeric method under the load of a homogeneous but pure isotropic or anisotropic surface stress. The results can be displayed with the dimensionality, so that the effect of clamping can be described in a systematic way. With these tabulated values one can evaluate cantilever experiments exactly. These results are generalized to cantilever methods for determining magnetoelastic constants. It is shown which magnetoelastic constants are measured in domains of thin films with ordered structures. The eigenshape and the eigenfrequency of plates constraint through a clamping at one side are calculated. These results give a deeper understanding of the elastic anisotropy. The induced surface stress of oxygen on the (110)-surface of molybdenum is measured along the principle directions Δτ [001] and Δτ [ anti 110] . The anisotropy of the surface stress is found for the p(2 x 2)-reconstruction. Lithium induces a tensile surface stress on the Molybdenum (110)-surface up to a coverage of Θ = 0, 3 monolayer. For a higher coverage the induced stress drops and reaches a level of less than -1, 2 N/m at one monolayer. It is shown, that cobalt induces a linear increasing stress with respect to the coverage on the (100)-surface of copper with a value of 2, 4GPa. The copper (100)-surface is bombarded with accelerated ions in the range between 800-2200 eV. The resulting induced compressive stress (Δτ (s) < 0) of the order

  16. Surface-Activated Coupling Reactions Confined on a Surface.

    Science.gov (United States)

    Dong, Lei; Liu, Pei Nian; Lin, Nian

    2015-10-20

    Chemical reactions may take place in a pure phase of gas or liquid or at the interface of two phases (gas-solid or liquid-solid). Recently, the emerging field of "surface-confined coupling reactions" has attracted intensive attention. In this process, reactants, intermediates, and products of a coupling reaction are adsorbed on a solid-vacuum or a solid-liquid interface. The solid surface restricts all reaction steps on the interface, in other words, the reaction takes place within a lower-dimensional, for example, two-dimensional, space. Surface atoms that are fixed in the surface and adatoms that move on the surface often activate the surface-confined coupling reactions. The synergy of surface morphology and activity allow some reactions that are inefficient or prohibited in the gas or liquid phase to proceed efficiently when the reactions are confined on a surface. Over the past decade, dozens of well-known "textbook" coupling reactions have been shown to proceed as surface-confined coupling reactions. In most cases, the surface-confined coupling reactions were discovered by trial and error, and the reaction pathways are largely unknown. It is thus highly desirable to unravel the mechanisms, mechanisms of surface activation in particular, of the surface-confined coupling reactions. Because the reactions take place on surfaces, advanced surface science techniques can be applied to study the surface-confined coupling reactions. Among them, scanning tunneling microscopy (STM) and X-ray photoelectron spectroscopy (XPS) are the two most extensively used experimental tools. The former resolves submolecular structures of individual reactants, intermediates, and products in real space, while the latter monitors the chemical states during the reactions in real time. Combination of the two methods provides unprecedented spatial and temporal information on the reaction pathways. The experimental findings are complemented by theoretical modeling. In particular, density

  17. Coupling between bulk ordering and surface segregation: from alloy surfaces to surface alloys

    International Nuclear Information System (INIS)

    Gallis, Coralie

    1997-01-01

    -The knowledge of the alloy surfaces is of prime interest to understand their catalytic properties. On the one hand, the determination of the stability of the surface alloys depends very strongly on the behaviours of the A c B 1-c alloy surfaces. On the other hand, the knowledge of the kinetics of the formation-dissolution of surface alloys can allow to understand the equilibrium segregation isotherm. We have then studied the relation between the equilibrium surface segregation in an alloy A c B 1-c and the kinetics of dissolution of a few metallic layers of A/B and the inverse deposit. We have used an energetic model derived from the electronic structure (T.I.B.M.) allowing us to study the surface segregation both in the disordered state and in the ordered one. The kinetics of dissolution were studied using the kinetic version of this model (K.T.I.B.M.) consistent with the equilibrium model. To illustrate our study, we have chosen the Cu-Pd system, a model for the formation of surface alloys and for which a great number of studies, both experimental and theoretical, are in progress. We then have shown for the (111) surface of this system that the surface alloys obtained during the dissolution are related to the alloy surfaces observed for the equilibrium segregation. The Cu-Pd system is characteristic of systems which have a weak segregation energy. Then, we have performed an equivalent study for a system with a strong segregation energy. Our choice was directly put on the Pt-Sn system. The surface behaviour, both in equilibrium and during the kinetics of dissolution, is very different from the Cu-Pd case. In particular, we have found pure 2-D surface alloys. Finally, a quenched molecular dynamics study has allowed us to determine the relative stability of various possible surface superstructures. (author) [fr

  18. Surface impedance and optimum surface resistance of a superconductor with an imperfect surface

    Science.gov (United States)

    Gurevich, Alex; Kubo, Takayuki

    2017-11-01

    We calculate a low-frequency surface impedance of a dirty, s -wave superconductor with an imperfect surface incorporating either a thin layer with a reduced pairing constant or a thin, proximity-coupled normal layer. Such structures model realistic surfaces of superconducting materials which can contain oxide layers, absorbed impurities, or nonstoichiometric composition. We solved the Usadel equations self-consistently and obtained spatial distributions of the order parameter and the quasiparticle density of states which then were used to calculate a low-frequency surface resistance Rs(T ) and the magnetic penetration depth λ (T ) as functions of temperature in the limit of local London electrodynamics. It is shown that the imperfect surface in a single-band s -wave superconductor results in a nonexponential temperature dependence of Z (T ) at T ≪Tc which can mimic the behavior of multiband or d -wave superconductors. The imperfect surface and the broadening of the gap peaks in the quasiparticle density of states N (ɛ ) in the bulk give rise to a weakly temperature-dependent residual surface resistance. We show that the surface resistance can be optimized and even reduced below its value for an ideal surface by engineering N (ɛ ) at the surface using pair-breaking mechanisms, particularly by incorporating a small density of magnetic impurities or by tuning the thickness and conductivity of the normal layer and its contact resistance. The results of this work address the limit of Rs in superconductors at T ≪Tc , and the ways of engineering the optimal density of states by surface nanostructuring and impurities to reduce losses in superconducting microresonators, thin-film strip lines, and radio-frequency cavities for particle accelerators.

  19. On-surface synthesis on a bulk insulator surface

    Science.gov (United States)

    Richter, Antje; Floris, Andrea; Bechstein, Ralf; Kantorovich, Lev; Kühnle, Angelika

    2018-04-01

    On-surface synthesis has rapidly emerged as a most promising approach to prepare functional molecular structures directly on a support surface. Compared to solution synthesis, performing chemical reactions on a surface offers several exciting new options: due to the absence of a solvent, reactions can be envisioned that are otherwise not feasible due to the insolubility of the reaction product. Perhaps even more important, the confinement to a two-dimensional surface might enable reaction pathways that are not accessible otherwise. Consequently, on-surface synthesis has attracted great attention in the last decade, with an impressive number of classical reactions transferred to a surface as well as new reactions demonstrated that have no classical analogue. So far, the majority of the work has been carried out on conducting surfaces. However, when aiming for electronic decoupling of the resulting structures, e.g. for the use in future molecular electronic devices, non-conducting surfaces are highly desired. Here, we review the current status of on-surface reactions demonstrated on the (10.4) surface of the bulk insulator calcite. Besides thermally induced C-C coupling of halogen-substituted aryls, photochemically induced [2  +  2] cycloaddition has been proven possible on this surface. Moreover, experimental evidence exists for coupling of terminal alkynes as well as diacetylene polymerization. While imaging of the resulting structures with dynamic atomic force microscopy provides a direct means of reaction verification, the detailed reaction pathway often remains unclear. Especially in cases where the presence of metal atoms is known to catalyze the corresponding solution chemistry reaction (e.g. in the case of the Ullmann reaction), disclosing the precise reaction pathway is of importance to understand and generalize on-surface reactivity on a bulk insulator surface. To this end, density-functional theory calculations have proven to provide atomic

  20. Surface Energy and Setting Process of Contacting Surfaces

    Directory of Open Access Journals (Sweden)

    M. V. Musokhranov

    2014-01-01

    Full Text Available The paper deals with a challenge in terms of ensuring an accuracy of the relative position of the conjugated surfaces that is to determine a coefficient of friction. To solve it, there is a proposal to use the surface energy, as a tool that influences the contacting parts nature. Presently, energy of the surface layers at best is only stated, but not used in practice.Analysis of the conditions of interaction between two contacting surfaces, such as seizing and setting cannot be explained only from the position of the roughness parameters. It is found that these phenomena are explained by the appearing gripe (setting bridges, which result from the energy of interaction between two or more adjacent surfaces. The emerging phenomenon such as micro welding, i.e. occurring bonds, is caused by the overflow of energy, according to the theory of physics, from the surface with a high level of energy to the surface with the smaller one to balance the system as a whole.The paper shows that through the use of process, controlling the depth of the surface layer and creating a certain structure, the energy level of the material as a whole can be specified. And this will allow us to provide the necessary performance and mechanical properties. It means to create as many gripe bridges as possible to ensure continuous positioning i.e. a fixed connection of the contacting surfaces.It was determined that to increase a value of the friction coefficient, the physical and mechanical properties of the surface layer of the parts material must be taken into account, namely, in the part body accumulate the energy to be consumed for forming the surface.The paper gives recommendations for including the parts of the surface energy in the qualitative indicators of characteristics. This will make a technologist, when routing a process, to choose such operations and modes to provide the designer-specified parameters not only of the accuracy and surface finish, but also of the

  1. Study on the preparation and stability of 188Re biomolecules via EHDP

    International Nuclear Information System (INIS)

    Ferro-Flores, G.; Garcia-Salinas, L.; Paredes-Gutierrez, L.; Hashimoto, K.; Melendez-Alafort, L.; Murphy, C.A.

    2001-01-01

    A direct labelling technique via ethane-1-hydroxy-1,1-diphosphonic acid (EHDP) as a weak competing ligand was developed for the preparation of several biomolecules: 188 Re-monoclonal antibody ior cea1 against carcinoembryonic antigen ( 188 Re-MoAb), biotinylated 188 Re-MoAb ( 188 Re-MoAb-biotin), 188 Re-polyclonal IgG ( 188 Re-IgG), 188 Re-peptide (somatostatine analogue peptide b-(2-naphtyl)-D-Ala-Cys-Tyr-D-Trp-Lys-Val-Cys-Thr-amide), 188 Re-MoAb fragments ( 188 Re-F(ab') 2 ) and biotinylated 188 Re-F(ab') 2 ( 188 Re-F(ab') 2 -biotin). The reaction conditions such as pH, temperature, weak ligand concentration and stannous chloride concentration were optimized during the radiolabelling of each biomolecule. Before the labelling procedure, disulphide bridge groups of the biomolecules were reduced with 2-mercaptoethanol (2-ME). To obtain 188 Re labelled antibodies and peptides in high radiochemical yields (>90%) via EHDP, it was necessary to use acidic conditions and a high concentration of stannous chloride to allow the redox reaction Re +7 →Re +5 :Re +4 . The labelling of MoAb and F(ab') 2 with 188 Re via EHDP was also evaluated employing a pretargeted technique by avidin-biotin strategy in normal mice, demonstrating that the 188 Re-labelled biotinylated antibodies are stable complexes in vivo. The 188 Re-peptide complex prepared by this method, was stable for 24 h and no radiolytic degradation was observed. (author)

  2. Tumor delivery of antisense oligomer using trastuzumab within a streptavidin nanoparticle

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yi [University of Massachusetts Medical School, Division of Nuclear Medicine, Department of Radiology, Worcester, MA (United States); Yale University, Yale PET Center, Department of Diagnostic Radiology, New Haven, CT (United States); Liu, Xinrong; Chen, Ling; Cheng, Dengfeng; Rusckowski, Mary [University of Massachusetts Medical School, Division of Nuclear Medicine, Department of Radiology, Worcester, MA (United States); Hnatowich, Donald J. [University of Massachusetts Medical School, Division of Nuclear Medicine, Department of Radiology, Worcester, MA (United States); Umass Medical School, Department of Radiology, Worcester, MA (United States)

    2009-12-15

    Trastuzumab (Herceptin trademark) is often internalized following binding to Her2+ tumor cells. The objective of this study was to investigate whether trastuzumab can be used as a specific carrier to deliver antisense oligomers into Her2+ tumor cells both in vitro and in vivo. A biotinylated MORF oligomer antisense to RhoC mRNA and its biotinylated sense control were labeled with either lissamine for fluorescence detection or {sup 99m}Tc for radioactivity detection and were linked to biotinylated trastuzumab via streptavidin. The nanoparticles were studied in SUM190 (RhoC+, Her2+) study and SUM149 (RhoC+, Her2-) control cells in culture and as xenografts in mice. As evidence of unimpaired Her2+ binding of trastuzumab within the nanoparticle, accumulations were clearly higher in SUM190 compared to SUM149 cells and, by whole-body imaging, targeting of SUM190 tumor was similar to that expected for a radiolabeled trastuzumab. As evidence of internalization, fluorescence microscopy images of cells grown in culture and obtained from xenografts showed uniform cytoplasm distribution of the lissamine-MORF. An invasion assay showed decreased RhoC expression in SUM190 cells when incubated with the antisense MORF nanoparticles at only 100 nM. Both in cell culture and in animals, the nanoparticle with trastuzumab as specific carrier greatly improved tumor delivery of the antisense oligomer against RhoC mRNA into tumor cells overexpressing Her2 and may be of general utility. (orig.)

  3. The 78 kDa glucose-regulated protein (GRP78/BIP) is expressed on the cell membrane, is released into cell culture medium and is also present in human peripheral circulation.

    Science.gov (United States)

    Delpino, Andrea; Castelli, Mauro

    2002-01-01

    In human rabdomiosarcoma cells (TE671/RD) chronic exposure to 500 nM thapsigargin (a powerful inhibitor of the endoplasmic reticulum Ca2+-ATPases) resulted in the induction of the stress protein GRP78/BIP. Making use of the surface biotinylation method, followed by the isolation of the GRP78 using ATP-agarose affinity chromatography, it was found that a fraction of the thapsigargin-induced GRP78 is expressed on the cell surface. The presence of GRP78 on the membrane of thapsigargin-treated cells was confirmed by fractionation of cell lysates into a soluble and a membrane fraction, followed by Western blot analysis with an anti-GRP78 antibody. It was also found that conspicuous amounts of GRP78 are present in the culture medium collected from thapsigargin-treated cultures. This extracellular GRP78 originates mostly by an active release from intact cells and does not result solely from the leakage of proteins from dead cells. Moreover, small amounts of circulating, free GRP78 and naturally-occurring anti-GRP78 autoantibodies were detected in the peripheral circulation of healthy human individuals.

  4. Characterisation of surface roughness for ultra-precision freeform surfaces

    International Nuclear Information System (INIS)

    Li Huifen; Cheung, C F; Lee, W B; To, S; Jiang, X Q

    2005-01-01

    Ultra-precision freeform surfaces are widely used in many advanced optics applications which demand for having surface roughness down to nanometer range. Although a lot of research work has been reported on the study of surface generation, reconstruction and surface characterization such as MOTIF and fractal analysis, most of them are focused on axial symmetric surfaces such as aspheric surfaces. Relative little research work has been found in the characterization of surface roughness in ultra-precision freeform surfaces. In this paper, a novel Robust Gaussian Filtering (RGF) method is proposed for the characterisation of surface roughness for ultra-precision freeform surfaces with known mathematic model or a cloud of discrete points. A series of computer simulation and measurement experiments were conducted to verify the capability of the proposed method. The experimental results were found to agree well with the theoretical results

  5. Microbial biotin protein ligases aid in understanding holocarboxylase synthetase deficiency.

    Science.gov (United States)

    Pendini, Nicole R; Bailey, Lisa M; Booker, Grant W; Wilce, Matthew C; Wallace, John C; Polyak, Steven W

    2008-01-01

    The attachment of biotin onto the biotin-dependent enzymes is catalysed by biotin protein ligase (BPL), also known as holocarboxylase synthase HCS in mammals. Mammals contain five biotin-enzymes that participate in a number of important metabolic pathways such as fatty acid biogenesis, gluconeogenesis and amino acid catabolism. All mammalian biotin-enzymes are post-translationally biotinylated, and therefore activated, through the action of a single HCS. Substrate recognition by BPLs occurs through conserved structural cues that govern the specificity of biotinylation. Defects in biotin metabolism, including HCS, give rise to multiple carboxylase deficiency (MCD). Here we review the literature on this important enzyme. In particular, we focus on the new information that has been learned about BPL's from a number of recently published protein structures. Through molecular modelling studies insights into the structural basis of HCS deficiency in MCD are discussed.

  6. A systematic first-principles study of surface energies, surface relaxation and Friedel oscillation of magnesium surfaces

    International Nuclear Information System (INIS)

    Tang, Jia-Jun; Yang, Xiao-Bao; Zhao, Yu-Jun; OuYang, LiuZhang; Zhu, Min

    2014-01-01

    We systematically study the surface energies and surface relaxations of various low-index and high-index Mg surfaces. It is found that low-index surfaces are not necessarily stable as Mg(1 0  1-bar  0) is the most unstable surface in the series of Mg(1 0  1-bar  n) (n = 0–9). A surface-energy predicting model based on the bond cutting is proposed to explain the relative surface stabilities. The local relaxations of the low-index surfaces could be explained by the Friedel oscillation. For the high-index surfaces, the combination of charge smoothing effect and dramatic charge depletion influences the relaxations, which show a big difference from the low-index ones. Our findings provide theoretical data for considerable insights into the surface energies of hexagonal close-packed metals. (paper)

  7. Evaluation of Surface Fatigue Strength Based on Surface Temperature

    Science.gov (United States)

    Deng, Gang; Nakanishi, Tsutomu

    Surface temperature is considered to be an integrated index that is dependent on not only the load and the dimensions at the contact point but also the sliding velocity, rolling velocity, surface roughness, and lubrication conditions. Therefore, the surface durability of rollers and gears can be evaluated more exactly and simply by the use of surface temperature rather than Hertzian stress. In this research, surface temperatures of rollers under different rolling and sliding conditions are measured using a thermocouple. The effects of load P, mean velocity Vm and sliding velocity Vs on surface temperature are clarified. An experimental formula, which expresses the linear relationship between surface temperature and the P0.86Vs1.31Vm-0.83 value, is used to determine surface temperature. By comparing calculated and measured temperature on the tooth surface of a gear, this formula is confirmed to be applicable for gear tooth surface temperature calculation.

  8. Surface phonons and elastic surface waves

    Science.gov (United States)

    Büscher, H.; Klein-Heßling, W.; Ludwig, W.

    Theoretical investigations on the dynamics of the (001), (110) and (111) surfaces of some cubic metals (Ag, Cu, Ni) will be reviewed. Both, lattice dynamical and continuum theoretical results are obtained via a Green's function formalism. The main attitude of this paper is the comparison of our results with experiments and with results obtained via slab-calculations. The calculation of elastic surface waves has been performed using a modified surface-green-function-matching method. We have used two different approaches of calculation the bulk Green's function (a) using the spectral representation and (b) a method, what works on residues. The investigations are carried out using shortrange phenomenological potentials. The atomic force constants in the first surface layers are modified to describe surface phonon anomalies, observed by experiments. In the case of Ag (100) and Ag(110) we conclude that the detection of odd symmetry shear modes by Erskine et al. [1 a, b] was not very accurate.

  9. Surface phonons and elastic surface waves

    International Nuclear Information System (INIS)

    Buescher, H.; Klein-Hessling, W.; Ludwig, W.

    1993-01-01

    Theoretical investigations on the dynamics of the (001), (110) and (111) surfaces of some cubic metals (Ag, Cu, Ni) will be reviewed. Both, lattice dynamical and continuum theoretical results are obtained via a Green's function formalism. The main attitude of this paper is the comparison of our results with experiments and with results obtained via slab-calculations. The calculation of elastic surface waves has been performed using a modified surface-green-function-matching method. We have used two different approaches of calculation the bulk Green's function (a) using the spectral representation and (b) a method, what works on residues. The investigations are carried out using shortrange phenomenological potentials. The atomic force constants in the first surface layers are modified to describe surface phonon anomalies, observed by experiments. In the case of Ag(100) and Ag(110) we conclude that the detection of odd symmetry shear modes by Erskine et al. was not very accurate. (orig.)

  10. Evaporation rates and surface profiles on heterogeneous surfaces with mass transfer and surface reaction

    Energy Technology Data Exchange (ETDEWEB)

    Flytzani-Stephanopoulos, M; Schmidt, L D

    1979-01-01

    Simple models incorporating surface reaction and diffusion of volatile products through a boundary layer are developed to calculate effective rates of evaporation and local surface profiles on surfaces having active and inactive regions. The coupling between surface heterogeneities with respect to a particular reaction and external mass transfer may provide a mechanism for the surface rearrangement and metal loss encountered in several catalytic systems of practical interest. Calculated transport rates for the volatilization of platinum in oxidizing environments and the rearrangement of this metal during the ammonia oxidation reaction agree well with published experimental data.

  11. Femtosecond laser-induced surface wettability modification of polystyrene surface

    Science.gov (United States)

    Wang, Bing; Wang, XinCai; Zheng, HongYu; Lam, YeeCheong

    2016-12-01

    In this paper, we demonstrated a simple method to create either a hydrophilic or hydrophobic surface. With femtosecond laser irradiation at different laser parameters, the water contact angle (WCA) on polystyrene's surface can be modified to either 12.7° or 156.2° from its original WCA of 88.2°. With properly spaced micro-pits created, the surface became hydrophilic probably due to the spread of the water droplets into the micro-pits. While with properly spaced micro-grooves created, the surface became rough and more hydrophobic. We investigated the effect of laser parameters on WCAs and analyzed the laser-treated surface roughness, profiles and chemical bonds by surface profilometer, scanning electron microscope (SEM) and X-ray photoelectron spectroscopy (XPS). For the laser-treated surface with low roughness, the polar (such as C—O, C=O, and O—C=O bonds) and non-polar (such as C—C or C—H bonds) groups were found to be responsible for the wettability changes. While for a rough surface, the surface roughness or the surface topography structure played a more significant role in the changes of the surface WCA. The mechanisms involved in the laser surface wettability modification process were discussed.

  12. In-surface confinement of topological insulator nanowire surface states

    International Nuclear Information System (INIS)

    Chen, Fan W.; Jauregui, Luis A.; Tan, Yaohua; Manfra, Michael; Klimeck, Gerhard; Chen, Yong P.; Kubis, Tillmann

    2015-01-01

    The bandstructures of [110] and [001] Bi 2 Te 3 nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects

  13. In-surface confinement of topological insulator nanowire surface states

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Fan W., E-mail: fanchen@purdue.edu [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); Jauregui, Luis A. [School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Tan, Yaohua [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Manfra, Michael [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); School of Materials Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Klimeck, Gerhard [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Chen, Yong P. [Department of Physics and Astronomy, Purdue, West Lafayette, Indiana 47907 (United States); School of Electrical and Computer Engineering, Purdue University, West Lafayette, Indiana 47907 (United States); Birck Nanotechnology Center, Purdue University, West Lafayette, Indiana 47907 (United States); Kubis, Tillmann [Network for Computational Nanotechnology, Purdue, West Lafayette, Indiana 47907 (United States)

    2015-09-21

    The bandstructures of [110] and [001] Bi{sub 2}Te{sub 3} nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects.

  14. In-surface confinement of topological insulator nanowire surface states

    Science.gov (United States)

    Chen, Fan W.; Jauregui, Luis A.; Tan, Yaohua; Manfra, Michael; Klimeck, Gerhard; Chen, Yong P.; Kubis, Tillmann

    2015-09-01

    The bandstructures of [110] and [001] Bi2Te3 nanowires are solved with the atomistic 20 band tight binding functionality of NEMO5. The theoretical results reveal: The popular assumption that all topological insulator (TI) wire surfaces are equivalent is inappropriate. The Fermi velocity of chemically distinct wire surfaces differs significantly which creates an effective in-surface confinement potential. As a result, topological insulator surface states prefer specific surfaces. Therefore, experiments have to be designed carefully not to probe surfaces unfavorable to the surface states (low density of states) and thereby be insensitive to the TI-effects.

  15. Validation of in vitro probabilistic tractography

    DEFF Research Database (Denmark)

    Dyrby, Tim B.; Sogaard, L.V.; Parker, G.J.

    2007-01-01

    assessed the anatomical validity and reproducibility of in vitro multi-fiber probabilistic tractography against two invasive tracers: the histochemically detectable biotinylated dextran amine and manganese enhanced magnetic resonance imaging. Post mortern DWI was used to ensure that most of the sources...

  16. Superhydrophobic surfaces fabricated by surface modification of alumina particles

    Science.gov (United States)

    Richard, Edna; Aruna, S. T.; Basu, Bharathibai J.

    2012-10-01

    The fabrication of superhydrophobic surfaces has attracted intense interest because of their widespread potential applications in various industrial fields. Recently, some attempts have been carried out to prepare superhydrophobic surfaces using metal oxide nanoparticles. In the present work, superhydrophobic surfaces were fabricated with low surface energy material on alumina particles with different sizes. It was found that particle size of alumina is an important factor in achieving stable superhydrophobic surface. It was possible to obtain alumina surface with water contact angle (WCA) of 156° and a sliding angle of Superhydrophobicity of the modified alumina is attributed to the combined effect of the micro-nanostructure and low surface energy of fatty acid on the surface. The surface morphology of the alumina powder and coatings was determined by FESEM. The stability of the coatings was assessed by conducting water immersion test. Effect of heat treatment on WCA of the coating was also studied. The transition of alumina from hydrophilic to superhydrophobic state was explained using Wenzel and Cassie models. The method is shown to have potential application for creating superhydrophobic surface on cotton fabrics.

  17. Disposable inkjet-printed electrochemical platform for detection of clinically relevant HER-2 breast cancer biomarker.

    Science.gov (United States)

    Carvajal, Susanita; Fera, Samantha N; Jones, Abby L; Baldo, Thaisa A; Mosa, Islam M; Rusling, James F; Krause, Colleen E

    2018-05-01

    Rapidly fabricated, disposable sensor platforms hold tremendous promise for point-of-care detection. Here, we present an inexpensive (Receptor 2 (HER-2). Capture antibodies were bound to a chemically modified surface on the WEA and placed into a microfluidic device. A full sandwich immunoassay was constructed following a simultaneous injection of target protein, biotinylated antibody, and polymerized horseradish peroxide labels into the microfluidic device housing the WEA. With an ultra fast assay time, of only 15mins a clinically relevant limit of detection of 12pgmL -1 was achieved. Excellent reproducibility and sensitivity were observed through recovery assays preformed in human serum with recoveries ranging from 76% to 103%. These easily fabricated and scalable electrochemical sensor platforms can be readily adapted for multiplex detection following this rapid assay protocol for cancer diagnostics. Copyright © 2018 Elsevier B.V. All rights reserved.

  18. Physical manipulation of single-molecule DNA using microbead and its application to analysis of DNA-protein interaction

    International Nuclear Information System (INIS)

    Kurita, Hirofumi; Yasuda, Hachiro; Takashima, Kazunori; Katsura, Shinji; Mizuno, Akira

    2009-01-01

    We carried out an individual DNA manipulation using an optical trapping for a microbead. This manipulation system is based on a fluorescent microscopy equipped with an IR laser. Both ends of linear DNA molecule were labeled with a biotin and a thiol group, respectively. Then the biotinylated end was attached to a microbead, and the other was immobilized on a thiol-linkable glass surface. We controlled the form of an individual DNA molecule by moving the focal point of IR laser, which trapped the microbead. In addition, we applied single-molecule approach to analyze DNA hydrolysis. We also used microchannel for single-molecule observation of DNA hydrolysis. The shortening of DNA in length caused by enzymatic hydrolysis was observed in real-time. The single-molecule DNA manipulation should contribute to elucidate detailed mechanisms of DNA-protein interactions

  19. Minimal surfaces

    CERN Document Server

    Dierkes, Ulrich; Sauvigny, Friedrich; Jakob, Ruben; Kuster, Albrecht

    2010-01-01

    Minimal Surfaces is the first volume of a three volume treatise on minimal surfaces (Grundlehren Nr. 339-341). Each volume can be read and studied independently of the others. The central theme is boundary value problems for minimal surfaces. The treatise is a substantially revised and extended version of the monograph Minimal Surfaces I, II (Grundlehren Nr. 295 & 296). The first volume begins with an exposition of basic ideas of the theory of surfaces in three-dimensional Euclidean space, followed by an introduction of minimal surfaces as stationary points of area, or equivalently

  20. Multilayer Relaxation and Surface Energies of Metallic Surfaces

    Science.gov (United States)

    Bozzolo, Guillermo; Rodriguez, Agustin M.; Ferrante, John

    1994-01-01

    The perpendicular and parallel multilayer relaxations of fcc (210) surfaces are studied using equivalent crystal theory (ECT). A comparison with experimental and theoretical results is made for AI(210). The effect of uncertainties in the input parameters on the magnitudes and ordering of surface relaxations for this semiempirical method is estimated. A new measure of surface roughness is proposed. Predictions for the multilayer relaxations and surface energies of the (210) face of Cu and Ni are also included.

  1. MR imaging of brain surface structures: Surface anatomy scanning

    International Nuclear Information System (INIS)

    Katada, K.; Koga, S.; Asahina, M.; Kanno, T.; Asahina, K.

    1987-01-01

    Preoperative evaluation of brain surface anatomy, including cortical sulci and veins, relative to cerebral and cerebellar lesions is an important subject for surgeons. Until now, no imaging modality existed that allowed direct visualization of brain surface anatomy. A new MR imaging technique (surface anatomy scanning) was developed to visualize brain surface structures. The technique uses a spin-echo pulse sequence with long repetition and echo times, thick sections and a surface coil. Cortical sulci, fissures, veins, and intracranial lesions were clearly identified with this technique. Initial clinical results indicate that surface anatomy scanning is useful for lesion localization and for detailed evaluation of cortical and subcortical lesions

  2. Investigation of surface roughness on etched glass surfaces

    International Nuclear Information System (INIS)

    Papa, Z.; Budai, J.; Farkas, B.; Toth, Z.

    2011-01-01

    Roughening the surface of solar cells is a common practice within the photovoltaic industry as it reduces reflectance, and thus enhances the performance of devices. In this work the relationship between reflectance characterized by the haze parameter, surface roughness and optical properties was investigated. To achieve this goal, model samples were prepared by hydrofluoric acid etching of glass for various times and measured by optical microscopy, spectroscopic ellipsometry, scanning electron microscopy, and atomic force microscopy. Our investigation showed that the surface reflectance was decreased not only by the roughening of the surface but also by the modification of the depth profile and lowering of the refractive index of the surface domain of the samples.

  3. Surface-to-surface registration using level sets

    DEFF Research Database (Denmark)

    Hansen, Mads Fogtmann; Erbou, Søren G.; Vester-Christensen, Martin

    2007-01-01

    This paper presents a general approach for surface-to-surface registration (S2SR) with the Euclidean metric using signed distance maps. In addition, the method is symmetric such that the registration of a shape A to a shape B is identical to the registration of the shape B to the shape A. The S2SR...... problem can be approximated by the image registration (IR) problem of the signed distance maps (SDMs) of the surfaces confined to some narrow band. By shrinking the narrow bands around the zero level sets the solution to the IR problem converges towards the S2SR problem. It is our hypothesis...... that this approach is more robust and less prone to fall into local minima than ordinary surface-to-surface registration. The IR problem is solved using the inverse compositional algorithm. In this paper, a set of 40 pelvic bones of Duroc pigs are registered to each other w.r.t. the Euclidean transformation...

  4. Isolation of microRNA targets using biotinylated synthetic microRNAs

    DEFF Research Database (Denmark)

    Ørom, Ulf Andersson; Lund, Anders H

    2007-01-01

    MicroRNAs are small regulatory RNAs found in multicellular organisms where they post-transcriptionally regulate gene expression. In animals, microRNAs bind mRNAs via incomplete base pairings making the identification of microRNA targets inherently difficult. Here, we present a detailed method...... for experimental identification of microRNA targets based on affinity purification of tagged microRNAs associated with their targets. Udgivelsesdato: 2007-Oct...

  5. Improvement of photodynamic activity of aluminium sulphophthalocyanine due to biotinylation

    Science.gov (United States)

    Meerovich, Irina G.; Jerdeva, Victoria V.; Derkacheva, Valentina M.; Meerovich, Gennadii A.; Lukyanets, Eugeny A.; Kogan, Eugenia A.; Savitsky, Alexander P.

    2003-09-01

    The photodynamic activity of dibiotinylated aluminium sulphophthalocyanine in vitro and in vivo were studied. It was obtained that in vitro dibiotinylated aluminium sulphophthalocyanine provides the effective damage of small cell lung carcinoma OAT-75. In vivo dibiotinylated aluminium sulphophthalocyanine causes destruction of tumor (Erlich carcinoma), results in total necrosis of tumor tissue and expresses vascular damage (trombosis and destruction of vascular walls) even in concentration 0.25 mg/kg of a body weight.

  6. The utilization of SA-gal in pre-targeting RIT of colon carcinoma xenograft models

    International Nuclear Information System (INIS)

    Wu Hubing; Huang Zuhan; Peng Wuhe; Gao Xiao

    2001-01-01

    To investigate the improved clearance effect of avidin (Av) and streptavidin-gal (SA - gal) for blood radiolabeled biotinylated antibody in pre-targeting radio-immuno therapy (RIT) of colon carcinoma xenograft models. Biotinylated antibody radiolabeled with 131 I was injected into the nude mice bearing the colon carcinoma via the tail vein. 24 h later, in 2 test groups, SA-gal or Av at a 10 - fold molar excess of antibody was intraperitoneally injected into the animals whereas no any chase agents were given to the control animals. Animals were killed for biodistribution study at 30h after 131 I-labelled biotinylated antibody administration. Results showed that Av and SA-gal took the effect of chase very fast. At 6h after injection, the blood level of radioactivity decreased very fast. The tumor/blood ratios of control group, Av chase group, SA-gal chase group were 0.42, 2.09, 5.23 respectively, P < 0.05 and P < 0.01 for the latter two groups as compared other control groups. Compared with Av, SA-gal showed better chase effect, its T/B ratios was 5.23, significantly higher than 2.09 of Av (P < 0.01). In the tissue biodistributions, relatively high non-specific radioactivity uptakes in non-liver organs were seen in Av chase group. Utilized in pre-targeting RIT, both Av and SA-gal could make the blood level of radioactivity decrease quickly and considerably improves tumor T/NT ratios. The chase effect of SA-gal was superior to that of avidin

  7. On-chip magnetic bead-based DNA melting curve analysis using a magnetoresistive sensor

    International Nuclear Information System (INIS)

    Rizzi, Giovanni; Østerberg, Frederik W.; Henriksen, Anders D.; Dufva, Martin; Hansen, Mikkel F.

    2015-01-01

    We present real-time measurements of DNA melting curves in a chip-based system that detects the amount of surface-bound magnetic beads using magnetoresistive magnetic field sensors. The sensors detect the difference between the amount of beads bound to the top and bottom sensor branches of the differential sensor geometry. The sensor surfaces are functionalized with wild type (WT) and mutant type (MT) capture probes, differing by a single base insertion (a single nucleotide polymorphism, SNP). Complementary biotinylated targets in suspension couple streptavidin magnetic beads to the sensor surface. The beads are magnetized by the field arising from the bias current passed through the sensors. We demonstrate the first on-chip measurements of the melting of DNA hybrids upon a ramping of the temperature. This overcomes the limitation of using a single washing condition at constant temperature. Moreover, we demonstrate that a single sensor bridge can be used to genotype a SNP. - Highlights: • We apply magnetoresistive sensors to study solid-surface hybridization kinetics of DNA. • We measure DNA melting profiles for perfectly matching DNA duplexes and for a single base mismatch. • We present a procedure to correct for temperature dependencies of the sensor output. • We reliably extract melting temperatures for the DNA hybrids. • We demonstrate direct measurement of differential binding signal for two probes on a single sensor

  8. Insertion of tetracysteine motifs into dopamine transporter extracellular domains.

    Directory of Open Access Journals (Sweden)

    Deanna M Navaroli

    Full Text Available The neuronal dopamine transporter (DAT is a major determinant of extracellular dopamine (DA levels and is the primary target for a variety of addictive and therapeutic psychoactive drugs. DAT is acutely regulated by protein kinase C (PKC activation and amphetamine exposure, both of which modulate DAT surface expression by endocytic trafficking. In order to use live imaging approaches to study DAT endocytosis, methods are needed to exclusively label the DAT surface pool. The use of membrane impermeant, sulfonated biarsenic dyes holds potential as one such approach, and requires introduction of an extracellular tetracysteine motif (tetraCys; CCPGCC to facilitate dye binding. In the current study, we took advantage of intrinsic proline-glycine (Pro-Gly dipeptides encoded in predicted DAT extracellular domains to introduce tetraCys motifs into DAT extracellular loops 2, 3, and 4. [(3H]DA uptake studies, surface biotinylation and fluorescence microscopy in PC12 cells indicate that tetraCys insertion into the DAT second extracellular loop results in a functional transporter that maintains PKC-mediated downregulation. Introduction of tetraCys into extracellular loops 3 and 4 yielded DATs with severely compromised function that failed to mature and traffic to the cell surface. This is the first demonstration of successful introduction of a tetracysteine motif into a DAT extracellular domain, and may hold promise for use of biarsenic dyes in live DAT imaging studies.

  9. Surface thermodynamics

    International Nuclear Information System (INIS)

    Garcia-Moliner, F.

    1975-01-01

    Basic thermodynamics of a system consisting of two bulk phases with an interface. Solid surfaces: general. Discussion of experimental data on surface tension and related concepts. Adsorption thermodynamics in the Gibbsian scheme. Adsorption on inert solid adsorbents. Systems with electrical charges: chemistry and thermodynamics of imperfect crystals. Thermodynamics of charged surfaces. Simple models of charge transfer chemisorption. Adsorption heat and related concepts. Surface phase transitions

  10. The Role of the Co-Chaperone, CHIP, in Androgen Independent Prostate Cancer

    Science.gov (United States)

    2008-02-01

    C4-2 there was no evidence of lysosomal congregration, the hallmark of autophagy, rather vaculolization of cytoplasm and disintergration of cellular...RNA polymerase promoter at the 5′ end. Second-strand synthesis was followed by cRNA production incorporating a biotinylated base. Hybridization to

  11. Morphology of protein polymer hybrid films studied by AFM and scanning confocal fluorescence microscopy

    NARCIS (Netherlands)

    Foubert, P.; Hannink, J.M.; Köhn, F.; Gronheid, R.; Sommerdijk, N.A.J.M.; Nolte, R.J.M.; Feyter, de S.; Schryver, de F.C.

    2003-01-01

    Protein–polymer hybrids can act as giant monolayer-forming amphiphiles at the air–water interface. Using biotinylated polystyrene (PSb) as the hydrophobic part and streptavidin (SAv) as a hydrophilic end of the giant amphiphile, monolayer formation and subsequent deposition leads to a well defined

  12. Open algebraic surfaces

    CERN Document Server

    Miyanishi, Masayoshi

    2000-01-01

    Open algebraic surfaces are a synonym for algebraic surfaces that are not necessarily complete. An open algebraic surface is understood as a Zariski open set of a projective algebraic surface. There is a long history of research on projective algebraic surfaces, and there exists a beautiful Enriques-Kodaira classification of such surfaces. The research accumulated by Ramanujan, Abhyankar, Moh, and Nagata and others has established a classification theory of open algebraic surfaces comparable to the Enriques-Kodaira theory. This research provides powerful methods to study the geometry and topology of open algebraic surfaces. The theory of open algebraic surfaces is applicable not only to algebraic geometry, but also to other fields, such as commutative algebra, invariant theory, and singularities. This book contains a comprehensive account of the theory of open algebraic surfaces, as well as several applications, in particular to the study of affine surfaces. Prerequisite to understanding the text is a basic b...

  13. Characterization of changes in composition and function of erythrocyte membrane proteins in patients with bone marrow form of acute radiation sickness

    International Nuclear Information System (INIS)

    Li Jinying; Wei Shanjian; Hu Xiaojian

    1998-01-01

    Objective: The delayed effect of radiation on erythrocyte membrane protein, the composition and function of the membrane proteins in five patients with bone marrow form of acute radiation sickness (ARS) were follow up at six years after the Shanghai 60 Co irradiation accident. Methods: Percoll centrifugation, SDS-polyacrylamide gel electrophoresis, and analysis of NO 2 - transport rate and DIDS inhibition rate were performed. Results: The injuries of the membrane proteins induced by radiation, characterized by reduced content of band 8 and declined anion transport function of band 3 protein remained the same as initially observed. The further study showed that the inhibition of DIDS on the anion transport of the ARS erythrocytes was decreased and the transport time for NO 2 - by band 3 was significantly prolonged in younger erythrocytes than those in middle-or old-aged cells. Conclusion: It is suggested that the radiation damage to erythrocyte membrane proteins might occur at the stage of erythropoiesis in bone marrow. The exo-facial site in band 3 may be changed after radiation, which could result in the abnormalities in anion transport. It is believed that the aging of erythrocytes might be present in advanced stage of ARS

  14. Smooth surfaces from bilinear patches: Discrete affine minimal surfaces

    KAUST Repository

    Kä ferbö ck, Florian; Pottmann, Helmut

    2013-01-01

    Motivated by applications in freeform architecture, we study surfaces which are composed of smoothly joined bilinear patches. These surfaces turn out to be discrete versions of negatively curved affine minimal surfaces and share many properties

  15. Sulfide Mineral Surfaces

    International Nuclear Information System (INIS)

    Rosso, Kevin M.; Vaughan, David J.

    2006-01-01

    The past twenty years or so have seen dramatic development of the experimental and theoretical tools available to study the surfaces of solids at the molecular (?atomic resolution?) scale. On the experimental side, two areas of development well illustrate these advances. The first concerns the high intensity photon sources associated with synchrotron radiation; these have both greatly improved the surface sensitivity and spatial resolution of already established surface spectroscopic and diffraction methods, and enabled the development of new methods for studying surfaces. The second centers on the scanning probe microscopy (SPM) techniques initially developed in the 1980's with the first scanning tunneling microscope (STM) and atomic force microscope (AFM) experiments. The direct 'observation' of individual atoms at surfaces made possible with these methods has truly revolutionized surface science. On the theoretical side, the availability of high performance computers coupled with advances in computational modeling has provided powerful new tools to complement the advances in experiment. Particularly important have been the quantum mechanics based computational approaches such as density functional theory (DFT), which can now be easily used to calculate the equilibrium crystal structures of solids and surfaces from first principles, and to provide insights into their electronic structure. In this chapter, we review current knowledge of sulfide mineral surfaces, beginning with an overview of the principles relevant to the study of the surfaces of all crystalline solids. This includes the thermodynamics of surfaces, the atomic structure of surfaces (surface crystallography and structural stability, adjustments of atoms at the surface through relaxation or reconstruction, surface defects) and the electronic structure of surfaces. We then discuss examples where specific crystal surfaces have been studied, with the main sulfide minerals organized by structure type

  16. Sulfide Mineral Surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Rosso, Kevin M.; Vaughan, David J.

    2006-08-01

    The past twenty years or so have seen dramatic development of the experimental and theoretical tools available to study the surfaces of solids at the molecular (?atomic resolution?) scale. On the experimental side, two areas of development well illustrate these advances. The first concerns the high intensity photon sources associated with synchrotron radiation; these have both greatly improved the surface sensitivity and spatial resolution of already established surface spectroscopic and diffraction methods, and enabled the development of new methods for studying surfaces. The second centers on the scanning probe microscopy (SPM) techniques initially developed in the 1980's with the first scanning tunneling microscope (STM) and atomic force microscope (AFM) experiments. The direct 'observation' of individual atoms at surfaces made possible with these methods has truly revolutionized surface science. On the theoretical side, the availability of high performance computers coupled with advances in computational modeling has provided powerful new tools to complement the advances in experiment. Particularly important have been the quantum mechanics based computational approaches such as density functional theory (DFT), which can now be easily used to calculate the equilibrium crystal structures of solids and surfaces from first principles, and to provide insights into their electronic structure. In this chapter, we review current knowledge of sulfide mineral surfaces, beginning with an overview of the principles relevant to the study of the surfaces of all crystalline solids. This includes the thermodynamics of surfaces, the atomic structure of surfaces (surface crystallography and structural stability, adjustments of atoms at the surface through relaxation or reconstruction, surface defects) and the electronic structure of surfaces. We then discuss examples where specific crystal surfaces have been studied, with the main sulfide minerals organized by

  17. Removal of arsenate by ferrihydrite via surface complexation and surface precipitation

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Xiuli [Department of Environment Engineering, College of the Environment and Ecology, and The Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystem, Xiamen University, Xiamen (China); Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, and The Key Laboratory for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen (China); Peng, Changjun; Fu, Dun; Chen, Zheng [Department of Environment Engineering, College of the Environment and Ecology, and The Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystem, Xiamen University, Xiamen (China); Shen, Liang [Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, and The Key Laboratory for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen (China); Li, Qingbiao [Department of Environment Engineering, College of the Environment and Ecology, and The Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystem, Xiamen University, Xiamen (China); Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, and The Key Laboratory for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen (China); Ouyang, Tong, E-mail: yz3t@xmu.edu.cn [Department of Environment Engineering, College of the Environment and Ecology, and The Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystem, Xiamen University, Xiamen (China); Wang, Yuanpeng, E-mail: wypp@xmu.edu.cn [Department of Chemical and Biochemical Engineering, College of Chemistry and Chemical Engineering, and The Key Laboratory for Synthetic Biotechnology of Xiamen City, Xiamen University, Xiamen (China)

    2015-10-30

    Graphical abstract: - Highlights: • Surface complexation and surface precipitation of As on ferrihydrite happen at pH 3–6. • The formation of surface precipitation enhanced As(V) adsorption. • The dissolved Fe{sup 3+} had a good linear relationship with the amount of arsenate re-adsorption. - Abstract: In this study, macroscopic and spectroscopic experimental methods accurately modeled the sorption process of arsenate on ferrihydrite. EXAFS, X-ray diffraction and infrared (IR) spectroscopy indicated that the behavior of As(V) adsorption onto ferrihydrite took place mainly via surface complexation and surface precipitation at acidic pH (3.0–6.0), while the surface precipitation was dominated at longer time intervals and higher Fe{sup 3+} concentration. The macroscopic competitive adsorption experiment between arsenate with phosphate indicated two types of adsorption sites existing on the surface of ferrihydrite, i.e., non-exchangeable sites, which are responsible for a rapid surface complex formation; and exchangeable sites for a slow build-up of surface precipitates. In the slow build-up precipitates, the As(V) surface coverage (mmol/g) exhibited a good linear relationship (R{sup 2} = 0.952) with the amount of dissolved Fe{sup 3+}. Three steps are involved during the process of surface precipitation, i.e., (1) an initial uptake of As(V) via surface complexation; (2) re-adsorption of Fe{sup 3+} leaching from ferrihydrite on the surface complex; and (3) As(V) adsorption via surface complexation again and finally forming the surface precipitate.

  18. [Influence of different surface treatments on porcelain surface topography].

    Science.gov (United States)

    Tai, Yinxia; Zhu, Xianchun; Sen, Yan; Liu, Chang; Zhang, Xian; Shi, Xueming

    2013-02-01

    To evaluate the influence of different surface treatments on porcelain surface topography. Metal ceramic prostheses in 6 groups were treated according to the different surface treatment methods, and the surface topography was observed through scanning electron microscope (SEM). Group A was the control one (untreated), group B was etched by 9.6% hydrofluoric acid(HF), group C was deglazed by grinding and then etched by 9.6% HF, group D was treated with Nd: YAG laser irradiation(0.75 W) and HF etching, group E was treated with Nd: YAG laser irradiation (1.05 W) and HF etching, and group F was treated with laser irradiation (1.45 W) and HF etching. Surface topography was different in different groups. A lot of inerratic cracks with the shapes of rhombuses and grid, and crater with a shape of circle were observed on the ceramic surface after treatment with energy parameters of 1.05 W Nd: YAG laser irradiation and 9.6% HF etching (group E). Surface topography showed a lot of concaves on the inner wall of the cracks, and the concaves with diameter of 1-5 microm could be observed on the inner wall of the holes, which had a diameter of 20 microm under SEM. The use of Nd: YAG laser irradiation with the energy parameters of 1.05 W and the HF with a concentration of 9.6% can evenly coarsen the porcelain surface, that is an effective surface treatment method.

  19. Laser surface texturing of tool steel: textured surfaces quality evaluation

    Science.gov (United States)

    Šugár, Peter; Šugárová, Jana; Frnčík, Martin

    2016-05-01

    In this experimental investigation the laser surface texturing of tool steel of type 90MnCrV8 has been conducted. The 5-axis highly dynamic laser precision machining centre Lasertec 80 Shape equipped with the nano-second pulsed ytterbium fibre laser and CNC system Siemens 840 D was used. The planar and spherical surfaces first prepared by turning have been textured. The regular array of spherical and ellipsoidal dimples with a different dimensions and different surface density has been created. Laser surface texturing has been realized under different combinations of process parameters: pulse frequency, pulse energy and laser beam scanning speed. The morphological characterization of ablated surfaces has been performed using scanning electron microscopy (SEM) technique. The results show limited possibility of ns pulse fibre laser application to generate different surface structures for tribological modification of metallic materials. These structures were obtained by varying the processing conditions between surface ablation, to surface remelting. In all cases the areas of molten material and re-cast layers were observed on the bottom and walls of the dimples. Beside the influence of laser beam parameters on the machined surface quality during laser machining of regular hemispherical and elipsoidal dimple texture on parabolic and hemispherical surfaces has been studied.

  20. Test surfaces useful for calibration of surface profilometers

    Science.gov (United States)

    Yashchuk, Valeriy V; McKinney, Wayne R; Takacs, Peter Z

    2013-12-31

    The present invention provides for test surfaces and methods for calibration of surface profilometers, including interferometric and atomic force microscopes. Calibration is performed using a specially designed test surface, or the Binary Pseudo-random (BPR) grating (array). Utilizing the BPR grating (array) to measure the power spectral density (PSD) spectrum, the profilometer is calibrated by determining the instrumental modulation transfer.

  1. Transversal Surfaces of Timelike Ruled Surfaces in Minkowski 3-Space

    OpenAIRE

    Önder, Mehmet

    2012-01-01

    In this study we give definitions and characterizations of transversal surfaces of timelike ruled surfaces. We study some special cases such as the striction curve is a geodesic, an asymptotic line or a line of curvature. Moreover, we obtain developable conditions for transversal surfaces of a timelike ruled surface.

  2. Surface phonons

    CERN Document Server

    Wette, Frederik

    1991-01-01

    In recent years substantial progress has been made in the detection of surface phonons owing to considerable improvements in inelastic rare gas scattering tech­ niques and electron energy loss spectroscopy. With these methods it has become possible to measure surface vibrations in a wide energy range for all wave vectors in the two-dimensional Brillouin zone and thus to deduce the complete surface phonon dispersion curves. Inelastic atomic beam scattering and electron energy loss spectroscopy have started to play a role in the study of surface phonons similar to the one played by inelastic neutron scattering in the investigation of bulk phonons in the last thirty years. Detailed comparison between experimen­ tal results and theoretical studies of inelastic surface scattering and of surface phonons has now become feasible. It is therefore possible to test and to improve the details of interaction models which have been worked out theoretically in the last few decades. At this point we felt that a concise, co...

  3. Analytical purification of a 60-kDa target protein of artemisinin detected in Trypanosoma brucei brucei

    Directory of Open Access Journals (Sweden)

    Benetode Konziase

    2015-12-01

    Full Text Available Here we describe the isolation and purity determination of Trypanosoma brucei (T. b. brucei candidate target proteins of artemisinin. The candidate target proteins were detected and purified from their biological source (T. b. brucei lysate using the diazirine-free biotinylated probe 5 for an affinity binding to a streptavidin-tagged resin and, subsequently, the labeled target proteins were purified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE. We herein showed the electrophoresis gel and the immunoblotting film containing the 60-kDa trypanosomal candidate target protein of artemisinin as a single band, which was visualized on-gel by the reverse-staining method and on a Western blotting film by enhanced chemiluminescence. The data provided in this article are related to the original research article “Biotinylated probes of artemisinin with labeling affinity toward Trypanosoma brucei brucei target proteins”, by Konziase (Anal. Biochem., vol. 482, 2015, pp. 25–31. http://dx.doi.org/10.1016/j.ab.2015.04.020.

  4. Expression and purification of functional JNK2beta2: perspectives on high-level production of recombinant MAP kinases.

    Science.gov (United States)

    Savopoulos, John W; Dowd, Stephen; Armour, Carolyn; Carter, Paul S; Greenwood, Catherine J; Mills, David; Powell, David; Pettman, Gary R; Jenkins, Owen; Walsh, Frank S; Philpott, Karen L

    2002-02-01

    The mitogen-activated protein (MAP) kinases are a group of serine/threonine kinases that mediate intracellular signal transduction in response to environmental stimuli including stress, growth factors, and various cytokines. Of this family, the c-Jun N-terminal kinases (JNKs) are members which, depending on cell type, have been shown to activate the transcription of genes involved in the inflammatory response, apoptosis, and hypertrophy. Here we report the use Baculovirus/Sf9 cells to produce milligram quantities of recombinant JNK2beta2 substrate which could be purified to >90% as judged by SDS-PAGE. In addition, we report a novel method for the site-specific biotinylation for this enzyme and demonstrate that the biotinylated product is an authentic substrate of the upstream kinases MKK4 and 7 and can phosphorylate a downstream target, ATF-2. We also show that the phosphorylated product can be captured efficiently on streptavidin-coated beads for use in scintillation proximity assays. Copyright 2002 Elsevier Science (USA).

  5. EnVision+, a new dextran polymer-based signal enhancement technique for in situ hybridization (ISH).

    Science.gov (United States)

    Wiedorn, K H; Goldmann, T; Henne, C; Kühl, H; Vollmer, E

    2001-09-01

    Seventy paraffin-embedded cervical biopsy specimens and condylomata were tested for the presence of human papillomavirus (HPV) by conventional in situ hybridization (ISH) and ISH with subsequent signal amplification. Signal amplification was performed either by a commercial biotinyl-tyramide-based detection system [GenPoint (GP)] or by the novel two-layer dextran polymer visualization system EnVision+ (EV), in which both EV-horseradish peroxidase (EV-HRP) and EV-alkaline phosphatase (EV-AP) were applied. We could demonstrate for the first time, that EV in combination with preceding ISH results in a considerable increase in signal intensity and sensitivity without loss of specificity compared to conventional ISH. Compared to GP, EV revealed a somewhat lower sensitivity, as measured by determination of the integrated optical density (IOD) of the positively stained cells. However, EV is easier to perform, requires a shorter assay time, and does not raise the background problems that may be encountered with biotinyl-tyramide-based amplification systems. (J Histochem Cytochem 49:1067-1071, 2001)

  6. An innovative pre-targeting strategy for tumor cell specific imaging and therapy.

    Science.gov (United States)

    Qin, Si-Yong; Peng, Meng-Yun; Rong, Lei; Jia, Hui-Zhen; Chen, Si; Cheng, Si-Xue; Feng, Jun; Zhang, Xian-Zheng

    2015-09-21

    A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the "biotin-avidin" interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging and therapeutic agents for tumor treatments.

  7. 3D-Printable Bioactivated Nanocellulose-Alginate Hydrogels.

    Science.gov (United States)

    Leppiniemi, Jenni; Lahtinen, Panu; Paajanen, Antti; Mahlberg, Riitta; Metsä-Kortelainen, Sini; Pinomaa, Tatu; Pajari, Heikki; Vikholm-Lundin, Inger; Pursula, Pekka; Hytönen, Vesa P

    2017-07-05

    We describe herein a nanocellulose-alginate hydrogel suitable for 3D printing. The composition of the hydrogel was optimized based on material characterization methods and 3D printing experiments, and its behavior during the printing process was studied using computational fluid dynamics simulations. The hydrogel was biofunctionalized by the covalent coupling of an enhanced avidin protein to the cellulose nanofibrils. Ionic cross-linking of the hydrogel using calcium ions improved the performance of the material. The resulting hydrogel is suitable for 3D printing, its mechanical properties indicate good tissue compatibility, and the hydrogel absorbs water in moist conditions, suggesting potential in applications such as wound dressings. The biofunctionalization potential was shown by attaching a biotinylated fluorescent protein and a biotinylated fluorescent small molecule via avidin and monitoring the material using confocal microscopy. The 3D-printable bioactivated nanocellulose-alginate hydrogel offers a platform for the development of biomedical devices, wearable sensors, and drug-releasing materials.

  8. Changes in biotin levels during production of natto, Japanese fermented soybean

    Directory of Open Access Journals (Sweden)

    Makoto Muratsugu

    2017-09-01

    Full Text Available The change of biotin level during production of natto (Japanese fermented soybean was investigated in this study.  The total biotin level was measured by an agar plate bioassay using Lactobacillus plantarum ATCC 8014.  The total biotin level decreased during water soaking, but increased after the fermentation of soybeans using Bacillus subtilis var. natto (B. natto and reached a maximum level.  The increase of total biotin was not affected by Asp, Arg, and Ile which promoted the growth of L. plantarum in high concentrations.  The peak level of biotin in the fermented soybeans was significantly higher than that of dry soybeans.  The fermented soybeans at the biotin peak level were adequate for food.  In addition, we detected 9 and 4 biotinylated polypeptides in the soybeans and B. natto used in this study, respectively.  We speculated that the increase of biotin level may depend on the increase of the 4 biotinylated polypeptides and free biotin in B. natto.

  9. Atmosphere-surface interactions over polar oceans and heterogeneous surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Vihma, T.

    1995-12-31

    Processes of interaction between the atmospheric boundary layer and the planetary surface have been studied with special emphasis on polar ocean surfaces: the open ocean, leads, polynyas and sea ice. The local exchange of momentum, heat and moisture has been studied experimentally both in the Weddell Sea and in the Greenland Sea. Exchange processes over heterogeneous surfaces are addressed by modelling studies. Over a homogeneous surface, the local turbulent fluxes can be reasonably well estimated using an iterative flux-profile scheme based on the Monin-Obukhov similarity theory. In the Greenland Sea, the near-surface air temperature and the generally small turbulent fluxes over the open ocean were affected by the sea surface temperature fronts. Over the sea ice cover in the Weddell Sea, the turbulent sensible heat flux was generally downwards, and together with an upward oceanic heat flux through the ice it compensated the heat loss from the surface via long-wave radiation. The wind dominated on time scales of days, while the current became important on longer time scales. The drift dynamics showed apparent spatial differences between the eastern and western regions, as well as between the Antarctic Circumpolar Current and the rest of the Weddell Sea. Inertial motion was present in regions of low ice concentration. The surface heterogeneity, arising e.g. from roughness or temperature distribution, poses a problem for the parameterization of surface exchange processes in large-scale models. In the case of neutral flow over a heterogeneous terrain, an effective roughness length can be used to parameterize the roughness effects

  10. Atmosphere-surface interactions over polar oceans and heterogeneous surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Vihma, T

    1996-12-31

    Processes of interaction between the atmospheric boundary layer and the planetary surface have been studied with special emphasis on polar ocean surfaces: the open ocean, leads, polynyas and sea ice. The local exchange of momentum, heat and moisture has been studied experimentally both in the Weddell Sea and in the Greenland Sea. Exchange processes over heterogeneous surfaces are addressed by modelling studies. Over a homogeneous surface, the local turbulent fluxes can be reasonably well estimated using an iterative flux-profile scheme based on the Monin-Obukhov similarity theory. In the Greenland Sea, the near-surface air temperature and the generally small turbulent fluxes over the open ocean were affected by the sea surface temperature fronts. Over the sea ice cover in the Weddell Sea, the turbulent sensible heat flux was generally downwards, and together with an upward oceanic heat flux through the ice it compensated the heat loss from the surface via long-wave radiation. The wind dominated on time scales of days, while the current became important on longer time scales. The drift dynamics showed apparent spatial differences between the eastern and western regions, as well as between the Antarctic Circumpolar Current and the rest of the Weddell Sea. Inertial motion was present in regions of low ice concentration. The surface heterogeneity, arising e.g. from roughness or temperature distribution, poses a problem for the parameterization of surface exchange processes in large-scale models. In the case of neutral flow over a heterogeneous terrain, an effective roughness length can be used to parameterize the roughness effects

  11. Surface properties of Ti-6Al-4V alloy part I: Surface roughness and apparent surface free energy

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Yingdi; Chibowski, Emil; Szcześ, Aleksandra, E-mail: aszczes@poczta.umcs.lublin.pl

    2017-01-01

    Titanium (Ti) and its alloys are the most often used implants material in dental treatment and orthopedics. Topography and wettability of its surface play important role in film formation, protein adhesion, following osseointegration and even duration of inserted implant. In this paper, we prepared Ti-6Al-4V alloy samples using different smoothing and polishing materials as well the air plasma treatment, on which contact angles of water, formamide and diiodomethane were measured. Then the apparent surface free energy was calculated using four different approaches (CAH, LWAB, O-W and Neumann's Equation of State). From LWAB approach the components of surface free energy were obtained, which shed more light on the wetting properties of samples surface. The surface roughness of the prepared samples was investigated with the help of optical profilometer and AFM. It was interesting whether the surface roughness affects the apparent surface free energy. It was found that both polar interactions the electron donor parameter of the energy and the work of water adhesion increased with decreasing roughness of the surfaces. Moreover, short time plasma treatment (1 min) caused decrease in the surface hydrophilic character, while longer time (10 min) treatment caused significant increase in the polar interactions and the work of water adhesion. Although Ti-6Al-4V alloy has been investigated many times, to our knowledge, so far no paper has been published in which surface roughness and changes in the surface free energy of the alloy were compared in the quantitative way in such large extent. This novel approach deliver better knowledge about the surface properties of differently smoothed and polished samples which may be helpful to facilitate cell adhesion, proliferation and mineralization. Therefore the results obtained present also potentially practical meaning. - Highlights: • Surface of five Ti-6Al-4V alloy samples were smoothed and polished successively. • The

  12. Surfaces of Building Practice

    OpenAIRE

    Surynková, Petra

    2009-01-01

    My diploma thesis Surfaces of Building Practice deals with the basic properties of surfaces, their mathematical description, categorization, and application in technical practice. Each studied surface is defined and its process of construction and parametrical description is listed. The thesis studies selected types of surfaces in details - these surfaces include surfaces of revolution, ruled surfaces, screw surfaces, and translational surfaces. An application of each studied surfaces is show...

  13. Scanning Surface Potential Microscopy of Spore Adhesion on Surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Ida [University of Tennessee, Knoxville (UTK); Chung, Eunhyea [Georgia Institute of Technology; Kweon, Hyojin [Georgia Institute of Technology; Yiacoumi, Sotira [Georgia Institute of Technology; Tsouris, Costas [ORNL

    2012-01-01

    The adhesion of spores of Bacillus anthracis - the cause of anthrax and a likely biological threat - to solid surfaces is an important consideration in cleanup after an accidental or deliberate release. However, because of safety concerns, directly studying B. anthracis spores with advanced instrumentation is problematic. As a first step, we are examining the electrostatic potential of Bacillus thuringiensis (Bt), which is a closely related species that is often used as a simulant to study B. anthracis. Scanning surface potential microscopy (SSPM), also known as Kelvin probe force microscopy (KPFM), was used to investigate the influence of relative humidity (RH) on the surface electrostatic potential of Bt that had adhered to silica, mica, or gold substrates. AFM/SSPM side-by-side images were obtained separately in air, at various values of RH, after an aqueous droplet with spores was applied on each surface and allowed to dry before measurements. In the SSPM images, a negative potential on the surface of the spores was observed compared with that of the substrates. The surface potential decreased as the humidity increased. Spores were unable to adhere to a surface with an extremely negative potential, such as mica.

  14. ASSESSMENT OF SURFACE QUALITY FOR CHOSEN MILLING STRATEGIES WHEN PRODUCING RELIEF SURFACES

    OpenAIRE

    Jan Varga; Jozef Stahovec; Jozef Beno; Marek Vrabeľ

    2014-01-01

    The paper deals with design and modeling of the relief surfaces that are produced in milling. Modeled and real surface quality is presented for the chosen fragments of the relief surfaces. Fragmentation of the relief surfaces has been made by the surface sampling. Milling strategies are compared with regard to surface formation. Surface quality was checked with regard to applied cutting conditions.

  15. Advanced Surface Technology

    DEFF Research Database (Denmark)

    Møller, Per; Nielsen, Lars Pleht

    of the components. It covers everything from biocompatible surfaces of IR absorbent or reflective surfaces to surfaces with specific properties within low friction, hardness, corrosion, colors, etc. The book includes more than 400 pages detailing virtually all analysis methods for examining at surfaces.......This new significant book on advanced modern surface technology in all its variations, is aimed at both teaching at engineering schools and practical application in industry. The work covers all the significant aspects of modern surface technology and also describes how new advanced techniques make...

  16. Surface parameter characterization of surface vibrations in linear chains

    International Nuclear Information System (INIS)

    Majlis, N.; Selzer, S.; Puszkarski, H.; Diep-The-Hung

    1982-12-01

    We consider the vibrations of a linear monatomic chain with a complex surface potential defined by the surface pinning parameter a=Aesup(-i psi). It is found that in the case of a semi-infinite chain a is connected with the surface vibration wave number k=s+it by the exact relations: s=psi, t=lnA. We also show that the solutions found can be regarded as approximate ones (in the limit L>>1) for surface vibrations of a finite chain consisting of L atoms. (author)

  17. The Use of Biotin to Demonstrate Immunohistochemistry, Western Blotting, and Dot Blots in University Practical Classes

    Science.gov (United States)

    Millar, Thomas James; Knighton, Ronald; Chuck, Jo-Anne

    2012-01-01

    Immunological detection of proteins is an essential method to demonstrate to undergraduate biology students, however, is often difficult in resource and time poor student laboratory sessions. This method describes a failsafe method to rapidly and economically demonstrate this technique using biotinylated proteins or biotin itself as targets for…

  18. Two lectin-like receptors for alpha 1-acid glycoprotein in mouse testis

    DEFF Research Database (Denmark)

    Andersen, U O; Kirkeby, S; Bøg-Hansen, T C

    1997-01-01

    Three glycoforms of alpha 1-acid glycoprotein (AGP) were biotinylated to examine their binding in mouse testis by light microscopy. The transition from one stage to another in the spermatogenic cycle is marked with an appearance of a receptor for the Concanavalin A (Con A) non-reactive glycoform...

  19. A Miniaturized Impedimetric Immunosensor for the Competitive Detection of Adrenocorticotropic Hormone

    Directory of Open Access Journals (Sweden)

    Nan Li

    2017-12-01

    Full Text Available Adrenocorticotropic hormone (ACTH plays an essential role in regulating corticosteroid hormone production, which has important functions in a myriad of critical physiological functions. In this proof-of-concept study, a miniaturized immunosensor was developed for the highly sensitive detection of ACTH using electrochemical impedance spectroscopy (EIS in connection with disposable screen-printed gold electrodes (SPGEs. A film of 3,3′-dithiobis[sulfosuccinimidylpropionate] (DTSSP was prepared to immobilize anti-ACTH antibodies covalently on the nanostructured SPGE surface. The surface-immobilized anti-ACTH antibodies captured the biotinylated ACTH (biotin-ACTH and non-labelled ACTH for the competitive immunoassay. After coupling of a streptavidin-alkaline phosphatase conjugate (Streptavidin-ALP, the bio-catalysed precipitation of an insoluble and insulating product onto the sensing interface changed the charge transfer resistance (Rct characteristics significantly. The detection limit of 100 fg/mL was determined for ACTH in a 5 μL sample volume, which indicated that this versatile platform can be easily adapted for miniaturized electrochemical immunosensing of cancer marker biomolecules. High selectivity and sensitivity of our immunoassay to detect ACTH in real samples demonstrated its promising potential for future development and applications using clinical samples.

  20. Poly(amido)amine (PAMAM) dendrimer–cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    International Nuclear Information System (INIS)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath

    2013-01-01

    Dendrimer–cisplatin complexes were prepared using PAMAM dendrimers with terminal –NH 2 and –COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer–cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was ∼11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC 50 values in ovarian cancer cells when compared with carboxylate surface dendrimers (p < 0.05). A correlation was observed among cytotoxicity of the complexes, cellular uptake, and platinum–DNA adduct formation. Treatment with dendrimer–cisplatin complexes resulted in a 7.0-fold increase (p < 0.05) in expression of apoptotic genes (Bcl2, Bax, p53) and 13.2- to 27.1-fold increase (p < 0.05) in the activity of caspases 3, 8, and 9 in vitro. Results suggest that PAMAM dendrimers can be used as potential carrier for cisplatin chemotherapy of ovarian cancer

  1. Poly(amido)amine (PAMAM) dendrimer-cisplatin complexes for chemotherapy of cisplatin-resistant ovarian cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yellepeddi, Venkata Kashyap; Vangara, Kiran Kumar; Palakurthi, Srinath, E-mail: palakurthi@tamhsc.edu [Texas A and M Health Science Center, Irma Lerma Rangel College of Pharmacy (United States)

    2013-09-15

    Dendrimer-cisplatin complexes were prepared using PAMAM dendrimers with terminal -NH{sub 2} and -COOH groups as well as biotin-conjugated dendrimers. Preformulation parameters of dendrimer-cisplatin complexes were studied using differential scanning calorimetry (DSC) and inductively coupled plasma-mass spectrometry (ICP-MS). Cytotoxicity and mechanism of cytotoxicity of dendrimer-cisplatin complexes was investigated in OVCAR-3, SKOV, A2780 and cisplatin-resistant CP70 human ovarian cancer cell lines. The loading of cisplatin in dendrimers was {approx}11 % (w/w). PAMAM G4 dendrimers with amine surface groups (biotinylated and native) have shown 2.5- to 3.0-fold reduction in IC{sub 50} values in ovarian cancer cells when compared with carboxylate surface dendrimers (p < 0.05). A correlation was observed among cytotoxicity of the complexes, cellular uptake, and platinum-DNA adduct formation. Treatment with dendrimer-cisplatin complexes resulted in a 7.0-fold increase (p < 0.05) in expression of apoptotic genes (Bcl2, Bax, p53) and 13.2- to 27.1-fold increase (p < 0.05) in the activity of caspases 3, 8, and 9 in vitro. Results suggest that PAMAM dendrimers can be used as potential carrier for cisplatin chemotherapy of ovarian cancer.

  2. Smooth polyhedral surfaces

    KAUST Repository

    Gü nther, Felix; Jiang, Caigui; Pottmann, Helmut

    2017-01-01

    Polyhedral surfaces are fundamental objects in architectural geometry and industrial design. Whereas closeness of a given mesh to a smooth reference surface and its suitability for numerical simulations were already studied extensively, the aim of our work is to find and to discuss suitable assessments of smoothness of polyhedral surfaces that only take the geometry of the polyhedral surface itself into account. Motivated by analogies to classical differential geometry, we propose a theory of smoothness of polyhedral surfaces including suitable notions of normal vectors, tangent planes, asymptotic directions, and parabolic curves that are invariant under projective transformations. It is remarkable that seemingly mild conditions significantly limit the shapes of faces of a smooth polyhedral surface. Besides being of theoretical interest, we believe that smoothness of polyhedral surfaces is of interest in the architectural context, where vertices and edges of polyhedral surfaces are highly visible.

  3. Smooth polyhedral surfaces

    KAUST Repository

    Günther, Felix

    2017-03-15

    Polyhedral surfaces are fundamental objects in architectural geometry and industrial design. Whereas closeness of a given mesh to a smooth reference surface and its suitability for numerical simulations were already studied extensively, the aim of our work is to find and to discuss suitable assessments of smoothness of polyhedral surfaces that only take the geometry of the polyhedral surface itself into account. Motivated by analogies to classical differential geometry, we propose a theory of smoothness of polyhedral surfaces including suitable notions of normal vectors, tangent planes, asymptotic directions, and parabolic curves that are invariant under projective transformations. It is remarkable that seemingly mild conditions significantly limit the shapes of faces of a smooth polyhedral surface. Besides being of theoretical interest, we believe that smoothness of polyhedral surfaces is of interest in the architectural context, where vertices and edges of polyhedral surfaces are highly visible.

  4. Ion surface collisions on surfaces relevant for fusion devices

    International Nuclear Information System (INIS)

    Rasul, B.; Endstrasser, N.; Zappa, F.; Grill, V.; Scheier, P.; Mark, T.

    2006-01-01

    Full text: One of the great challenges of fusion research is the compatibility of reactor grade plasmas with plasma facing materials coating the inner walls of a fusion reactor. The question of which surface coating should be used is of particular interest for the design of ITER. The impact of energetic plasma particles leads to sputtering of wall material into the plasma. A possible solution for the coating of plasma facing walls would be the use of special carbon surfaces. Investigations of these various surfaces have been started at BESTOF ion-surface collision apparatus. Experiment beam of singly charged molecular ions of hydrocarbon molecules, i.e. C 2 H + 4 , is generated in a Nier-type electron impact ionization source at an electron energy of about 70 eV. In the first double focusing mass spectrometer the ions are mass and energy analyzed and afterwards refocused onto a surface. The secondary reaction products are monitored using a Time Of Flight mass spectrometer. The secondary ion mass spectra are recorded as a function of the collision energy for different projectile ions and different surfaces. A comparison of these spectra show for example distinct changes in the survival probability of the same projectile ion C 2 H + 4 for different surfaces. (author)

  5. Surface Modification and Surface - Subsurface Exchange Processes on Europa

    Science.gov (United States)

    Phillips, C. B.; Molaro, J.; Hand, K. P.

    2017-12-01

    The surface of Jupiter's moon Europa is modified by exogenic processes such as sputtering, gardening, radiolysis, sulfur ion implantation, and thermal processing, as well as endogenic processes including tidal shaking, mass wasting, and the effects of subsurface tectonic and perhaps cryovolcanic activity. New materials are created or deposited on the surface (radiolysis, micrometeorite impacts, sulfur ion implantation, cryovolcanic plume deposits), modified in place (thermal segregation, sintering), transported either vertically or horizontally (sputtering, gardening, mass wasting, tectonic and cryovolcanic activity), or lost from Europa completely (sputtering, plumes, larger impacts). Some of these processes vary spatially, as visible in Europa's leading-trailing hemisphere brightness asymmetry. Endogenic geologic processes also vary spatially, depending on terrain type. The surface can be classified into general landform categories that include tectonic features (ridges, bands, cracks); disrupted "chaos-type" terrain (chaos blocks, matrix, domes, pits, spots); and impact craters (simple, complex, multi-ring). The spatial distribution of these terrain types is relatively random, with some differences in apex-antiapex cratering rates and latitudinal variation in chaos vs. tectonic features. In this work, we extrapolate surface processes and rates from the top meter of the surface in conjunction with global estimates of transport and resurfacing rates. We combine near-surface modification with an estimate of surface-subsurface (and vice versa) transport rates for various geologic terrains based on an average of proposed formation mechanisms, and a spatial distribution of each landform type over Europa's surface area. Understanding the rates and mass balance for each of these processes, as well as their spatial and temporal variability, allows us to estimate surface - subsurface exchange rates over the average surface age ( 50myr) of Europa. Quantifying the timescale

  6. Evaluation of methods to reduce background using the Python-based ELISA_QC program.

    Science.gov (United States)

    Webster, Rose P; Cohen, Cinder F; Saeed, Fatima O; Wetzel, Hanna N; Ball, William J; Kirley, Terence L; Norman, Andrew B

    2018-05-01

    Almost all immunological approaches [immunohistochemistry, enzyme-linked immunosorbent assay (ELISA), Western blot], that are used to quantitate specific proteins have had to address high backgrounds due to non-specific reactivity. We report here for the first time a quantitative comparison of methods for reduction of the background of commercial biotinylated antibodies using the Python-based ELISA_QC program. This is demonstrated using a recombinant humanized anti-cocaine monoclonal antibody. Several approaches, such as adjustment of the incubation time and the concentration of blocking agent, as well as the dilution of secondary antibodies, have been explored to address this issue. In this report, systematic comparisons of two different methods, contrasted with other more traditional methods to address this problem are provided. Addition of heparin (HP) at 1 μg/ml to the wash buffer prior to addition of the secondary biotinylated antibody reduced the elevated background absorbance values (from a mean of 0.313 ± 0.015 to 0.137 ± 0.002). A novel immunodepletion (ID) method also reduced the background (from a mean of 0.331 ± 0.010 to 0.146 ± 0.013). Overall, the ID method generated more similar results at each concentration of the ELISA standard curve to that using the standard lot 1 than the HP method, as analyzed by the Python-based ELISA_QC program. We conclude that the ID method, while more laborious, provides the best solution to resolve the high background seen with specific lots of biotinylated secondary antibody. Copyright © 2018. Published by Elsevier B.V.

  7. Novel multi-biotin grafted poly(lactic acid) and its self-assembling nanoparticles capable of binding to streptavidin

    Science.gov (United States)

    Yan, Hao; Jiang, Weimin; Zhang, Yinxing; Liu, Ying; Wang, Bin; Yang, Li; Deng, Lihong; Singh, Gurinder K; Pan, Jun

    2012-01-01

    Targeted drug delivery requires novel biodegradable, specific binding systems with longer circulation time. The aim of this study was to prepare biotinylated poly(lactic acid) (PLA) nanoparticles (NPs) which can meet regular requirements as well conjugate more biotins in the polymer to provide better binding with streptavidin. A biotin-graft-PLA was synthesized based on previously published biodegradable poly(ethylene glycol) (PEG)-graft-PLA, with one polymer molecule containing three PEG molecules. Newly synthesized biotin-graft-PLA had three biotins per polymer molecule, higher than the previous biotinylated PLA (≤1 biotin per polymer molecule). A PEG with a much lower molecular weight (MW ~1900) than the previous biotinylated PLA (PEG MW ≥ 3800), and thus more biocompatible, was used which supplied good nonspecific protein-resistant property compatible to PEG-graft-PLA, suggesting its possible longer stay in the bloodstream. Biotin-graft-PLA specifically bound to streptavidin and self-assembled into NPs, during which naproxen, a model small molecule (MW 230 Da) and hydrophobic drug, was encapsulated (encapsulation efficiency 51.88%). The naproxen-loaded NPs with particle size and zeta potential of 175 nm and −27.35 mV realized controlled release within 170 hours, comparable to previous studies. The biotin-graft-PLA NPs adhered approximately two-fold more on streptavidin film and on biotin film via a streptavidin arm both in static and dynamic conditions compared with PEG-graft-PLA NPs, the proven nonspecific protein-resistant NPs. The specific binding of biotin-graft-PLA NPs with streptavidin and with biotin using streptavidin arm, as well as its entrapment and controlled release for naproxen, suggest potential applications in targeted drug delivery. PMID:22334778

  8. Surface properties of Ti-6Al-4V alloy part I: Surface roughness and apparent surface free energy.

    Science.gov (United States)

    Yan, Yingdi; Chibowski, Emil; Szcześ, Aleksandra

    2017-01-01

    Titanium (Ti) and its alloys are the most often used implants material in dental treatment and orthopedics. Topography and wettability of its surface play important role in film formation, protein adhesion, following osseointegration and even duration of inserted implant. In this paper, we prepared Ti-6Al-4V alloy samples using different smoothing and polishing materials as well the air plasma treatment, on which contact angles of water, formamide and diiodomethane were measured. Then the apparent surface free energy was calculated using four different approaches (CAH, LWAB, O-W and Neumann's Equation of State). From LWAB approach the components of surface free energy were obtained, which shed more light on the wetting properties of samples surface. The surface roughness of the prepared samples was investigated with the help of optical profilometer and AFM. It was interesting whether the surface roughness affects the apparent surface free energy. It was found that both polar interactions the electron donor parameter of the energy and the work of water adhesion increased with decreasing roughness of the surfaces. Moreover, short time plasma treatment (1min) caused decrease in the surface hydrophilic character, while longer time (10min) treatment caused significant increase in the polar interactions and the work of water adhesion. Although Ti-6Al-4V alloy has been investigated many times, to our knowledge, so far no paper has been published in which surface roughness and changes in the surface free energy of the alloy were compared in the quantitative way in such large extent. This novel approach deliver better knowledge about the surface properties of differently smoothed and polished samples which may be helpful to facilitate cell adhesion, proliferation and mineralization. Therefore the results obtained present also potentially practical meaning. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Characterization of polymer surface structure and surface mechanical behaviour by sum frequency generation surface vibrational spectroscopy and atomic force microscopy

    International Nuclear Information System (INIS)

    Opdahl, Aric; Koffas, Telly S; Amitay-Sadovsky, Ella; Kim, Joonyeong; Somorjai, Gabor A

    2004-01-01

    Sum frequency generation (SFG) vibrational spectroscopy and atomic force microscopy (AFM) have been used to study polymer surface structure and surface mechanical behaviour, specifically to study the relationships between the surface properties of polymers and their bulk compositions and the environment to which the polymer is exposed. The combination of SFG surface vibrational spectroscopy and AFM has been used to study surface segregation behaviour of polyolefin blends at the polymer/air and polymer/solid interfaces. SFG surface vibrational spectroscopy and AFM experiments have also been performed to characterize the properties of polymer/liquid and polymer/polymer interfaces, focusing on hydrogel materials. A method was developed to study the surface properties of hydrogel contact lens materials at various hydration conditions. Finally, the effect of mechanical stretching on the surface composition and surface mechanical behaviour of phase-separated polyurethanes, used in biomedical implant devices, has been studied by both SFG surface vibrational spectroscopy and AFM. (topical review)

  10. Extremal surface barriers

    International Nuclear Information System (INIS)

    Engelhardt, Netta; Wall, Aron C.

    2014-01-01

    We present a generic condition for Lorentzian manifolds to have a barrier that limits the reach of boundary-anchored extremal surfaces of arbitrary dimension. We show that any surface with nonpositive extrinsic curvature is a barrier, in the sense that extremal surfaces cannot be continuously deformed past it. Furthermore, the outermost barrier surface has nonnegative extrinsic curvature. Under certain conditions, we show that the existence of trapped surfaces implies a barrier, and conversely. In the context of AdS/CFT, these barriers imply that it is impossible to reconstruct the entire bulk using extremal surfaces. We comment on the implications for the firewall controversy

  11. Imaging near-surface heterogeneities by natural migration of surface waves

    KAUST Repository

    Liu, Zhaolun; AlTheyab, Abdullah; Hanafy, Sherif M.; Schuster, Gerard T.

    2016-01-01

    We demonstrate that near-surface heterogeneities can be imaged by natural migration of backscattered surface waves in common shot gathers. No velocity model is required because the data are migrated onto surface points with the virtual Green

  12. Inspection of freeform surfaces considering uncertainties in measurement, localization and surface reconstruction

    International Nuclear Information System (INIS)

    Mehrad, Vahid; Xue, Deyi; Gu, Peihua

    2013-01-01

    Inspection of a manufactured freeform surface can be conducted by building its surface model and comparing this manufactured surface model with the ideal design surface model and its tolerance requirement. The manufactured freeform surface model is usually achieved by obtaining measurement points on the manufactured surface, transforming these measurement points from the measurement coordinate system to the design coordinate system through localization, and reconstructing the surface model using the localized measurement points. In this research, a method was developed to estimate the locations and their variances of any selected points on the reconstructed freeform surface considering different sources of uncertainties in measurement, localization and surface reconstruction processes. In this method, first locations and variances of the localized measurement points are calculated considering uncertainties of the measurement points and uncertainties introduced in the localization processes. Then locations and variances of points on the reconstructed freeform surface are obtained considering uncertainties of the localized measurement points and uncertainties introduced in the freeform surface reconstruction process. Two case studies were developed to demonstrate how these three different uncertainty sources influence the quality of the reconstructed freeform curve and freeform surface in inspection. (paper)

  13. Variability of emissivity and surface temperature over a sparsely vegetated surface

    International Nuclear Information System (INIS)

    Humes, K.S.; Kustas, W.P.; Moran, M.S.; Nichols, W.D.; Weltz, M.A.

    1994-01-01

    Radiometric surface temperatures obtained from remote sensing measurements are a function of both the physical surface temperature and the effective emissivity of the surface within the band pass of the radiometric measurement. For sparsely vegetated areas, however, a sensor views significant fractions of both bare soil and various vegetation types. In this case the radiometric response of a sensor is a function of the emissivities and kinetic temperatures of various surface elements, the proportion of those surface elements within the field of view of the sensor, and the interaction of radiation emitted from the various surface components. In order to effectively utilize thermal remote sensing data to quantify energy balance components for a sparsely vegetated area, it is important to examine the typical magnitude and degree of variability of emissivity and surface temperature for such surfaces. Surface emissivity measurements and ground and low-altitude-aircraft-based surface temperature measurements (8-13 micrometer band pass) made in conjunction with the Monsoon '90 field experiment were used to evaluate the typical variability of those quantities during the summer rainy season in a semiarid watershed. The average value for thermal band emissivity of the exposed bare soil portions of the surface was found to be approximately 0.96; the average value measured for most of the varieties of desert shrubs present was approximately 0.99. Surface composite emissivity was estimated to be approximately 0.98 for both the grass-dominated and shrub-dominated portions of the watershed. The spatial variability of surface temperature was found to be highly dependent on the spatial scale of integration for the instantaneous field of view (IFOV) of the instrument, the spatial scale of the total area under evaluation, and the time of day

  14. SURFACE PHOTOMETRY OF LOW SURFACE BRIGHTNESS GALAXIES

    NARCIS (Netherlands)

    DEBLOK, WJG; VANDERHULST, JM; BOTHUN, GD

    1995-01-01

    Low surface brightness (LSB) galaxies are galaxies dominated by an exponential disc whose central surface brightness is much fainter than the value of mu(B)(0) = 21.65 +/- 0.30 mag arcsec(-2) found by Freeman. In this paper we present broadband photometry of a sample of 21 late-type LSB galaxies.

  15. Surface composition and surface properties of water hyacinth ...

    African Journals Online (AJOL)

    Surface composition and surface properties of water hyacinth ( Eichhornia ... (2/1, v/v) followed by ethanol, using Fourier Transform Infra-red (FT-IR) spectroscopy, ... polar organic solvents and non-polar n-alkane hydrocarbons is discussed.

  16. Sliding surface searching method for slopes containing a potential weak structural surface

    Directory of Open Access Journals (Sweden)

    Aijun Yao

    2014-06-01

    Full Text Available Weak structural surface is one of the key factors controlling the stability of slopes. The stability of rock slopes is in general concerned with set of discontinuities. However, in soft rocks, failure can occur along surfaces approaching to a circular failure surface. To better understand the position of potential sliding surface, a new method called simplex-finite stochastic tracking method is proposed. This method basically divides sliding surface into two parts: one is described by smooth curve obtained by random searching, the other one is polyline formed by the weak structural surface. Single or multiple sliding surfaces can be considered, and consequently several types of combined sliding surfaces can be simulated. The paper will adopt the arc-polyline to simulate potential sliding surface and analyze the searching process of sliding surface. Accordingly, software for slope stability analysis using this method was developed and applied in real cases. The results show that, using simplex-finite stochastic tracking method, it is possible to locate the position of a potential sliding surface in the slope.

  17. Smooth surfaces from bilinear patches: Discrete affine minimal surfaces

    KAUST Repository

    Käferböck, Florian

    2013-06-01

    Motivated by applications in freeform architecture, we study surfaces which are composed of smoothly joined bilinear patches. These surfaces turn out to be discrete versions of negatively curved affine minimal surfaces and share many properties with their classical smooth counterparts. We present computational design approaches and study special cases which should be interesting for the architectural application. 2013 Elsevier B.V.

  18. Imaging near-surface heterogeneities by natural migration of backscattered surface waves

    KAUST Repository

    AlTheyab, Abdullah

    2016-02-01

    We present a migration method that does not require a velocity model to migrate backscattered surface waves to their projected locations on the surface. This migration method, denoted as natural migration, uses recorded Green\\'s functions along the surface instead of simulated Green\\'s functions. The key assumptions are that the scattering bodies are within the depth interrogated by the surface waves, and the Green\\'s functions are recorded with dense receiver sampling along the free surface. This natural migration takes into account all orders of multiples, mode conversions and non-linear effects of surface waves in the data. The natural imaging formulae are derived for both active source and ambient-noise data, and computer simulations show that natural migration can effectively image near-surface heterogeneities with typical ambient-noise sources and geophone distributions.

  19. Surface adhesion and confinement variation of Staphylococcus aurius on SAM surfaces

    Science.gov (United States)

    Amroski, Alicia; Olsen, Morgan; Calabrese, Joseph; Senevirathne, Reshani; Senevirathne, Indrajith

    2012-02-01

    Controlled surface adhesion of non - pathogenic gram positive strain, Staphylococcus aureus is interesting as a model system due to possible development of respective biosensors for prevention and detection of the pathogenic strain methicillin resistant Staphylococcus aureus (MRSA) and further as a study for bio-machine interfacing. Self Assembled Monolayers (SAM) with engineered surfaces of linear thiols on Au(111) were used as the substrate. Sub cultured S. aureus were used for the analysis. The SAM layered surfaces were dipped in 2 -- 4 Log/ml S. aureus solution. Subsequent surface adhesion at different bacterial dilutions on surfaces will be discussed, and correlated with quantitative and qualitative adhesion properties of bacteria on the engineered SAM surfaces. The bacteria adhered SAM surfaces were investigated using intermittent contact, noncontact, lateral force and contact modes of Atomic Force Microscopy (AFM).

  20. Surface-enhanced chiroptical spectroscopy with superchiral surface waves.

    Science.gov (United States)

    Pellegrini, Giovanni; Finazzi, Marco; Celebrano, Michele; Duò, Lamberto; Biagioni, Paolo

    2018-07-01

    We study the chiroptical properties of one-dimensional photonic crystals supporting superchiral surface waves by introducing a simple formalism based on the Fresnel reflection matrix. We show that the proposed framework provides useful insights on the behavior of all the relevant chiroptical quantities, allowing for a deeper understanding of surface-enhanced chiral sensing platforms based on one-dimensional photonic crystals. Finally, we analyze and discuss the limitations of such platforms as the surface concentration of the target chiral analytes is gradually increased. © 2018 Wiley Periodicals, Inc.

  1. Bacteria-surface interactions.

    Science.gov (United States)

    Tuson, Hannah H; Weibel, Douglas B

    2013-05-14

    The interaction of bacteria with surfaces has important implications in a range of areas, including bioenergy, biofouling, biofilm formation, and the infection of plants and animals. Many of the interactions of bacteria with surfaces produce changes in the expression of genes that influence cell morphology and behavior, including genes essential for motility and surface attachment. Despite the attention that these phenotypes have garnered, the bacterial systems used for sensing and responding to surfaces are still not well understood. An understanding of these mechanisms will guide the development of new classes of materials that inhibit and promote cell growth, and complement studies of the physiology of bacteria in contact with surfaces. Recent studies from a range of fields in science and engineering are poised to guide future investigations in this area. This review summarizes recent studies on bacteria-surface interactions, discusses mechanisms of surface sensing and consequences of cell attachment, provides an overview of surfaces that have been used in bacterial studies, and highlights unanswered questions in this field.

  2. Connecting DNA Origami Structures Using the Biotin-Streptavidin ...

    African Journals Online (AJOL)

    Abstract. This work made use of the strong interaction between biotin and streptavidin to connect designed DNA origami structures. The caDNAno software was used to design a 6 layer 3D origami cross-like structure. Selected DNA strands at the engineered attachment sites on the DNA origami structure were biotinylated.

  3. Isolation and screening of microsatellite markers from the pearl ...

    Indian Academy of Sciences (India)

    DNA was simultaneously digested with. MseI and ligated to MseI-adaptor. The digestion ligation mixture was diluted (1:10) and directly amplified with. ∗For correspondence. E-mail: pearlydh@163.com. AFLP adaptor-specific primers. The PCR product was then hybridized with a biotinylated (CA)15 probe. DNA molecules.

  4. A corrugated perfect magnetic conductor surface supporting spoof surface magnon polaritons.

    Science.gov (United States)

    Liu, Liang-liang; Li, Zhuo; Gu, Chang-qing; Ning, Ping-ping; Xu, Bing-zheng; Niu, Zhen-yi; Zhao, Yong-jiu

    2014-05-05

    In this paper, we demonstrate that spoof surface magnon polaritons (SSMPs) can propagate along a corrugated perfect magnetic conductor (PMC) surface. From duality theorem, the existence of surface electromagnetic modes on corrugated PMC surfaces are manifest to be transverse electric (TE) mode compared with the transverse magnetic (TM) mode of spoof surface plasmon plaritons (SSPPs) excited on corrugated perfect electric conductor surfaces. Theoretical deduction through modal expansion method and simulation results clearly verify that SSMPs share the same dispersion relationship with the SSPPs. It is worth noting that this metamaterial will have more similar properties and potential applications as the SSPPs in large number of areas.

  5. Surface effect theory in binary alloys: surfaces with cut-off

    International Nuclear Information System (INIS)

    Kumar, V.; Silva, C.E.T.G. da; Moran-Lopez, J.L.

    1981-01-01

    A surface effect theory in binary alloys which ore ordered with surfaces with cut-off is presented. This theory is based in a model of pair interaction between first neighbours and includes long and short range effects. The (120) surface with sup(-) (110) monoatomic cut-off and terrace in the (110) planes of an alloy with body centered cubic structure is presented as example. Results for the concentrations in all the different surface sites are given. (L.C.) [pt

  6. Surface correlations of hydrodynamic drag for transitionally rough engineering surfaces

    Science.gov (United States)

    Thakkar, Manan; Busse, Angela; Sandham, Neil

    2017-02-01

    Rough surfaces are usually characterised by a single equivalent sand-grain roughness height scale that typically needs to be determined from laboratory experiments. Recently, this method has been complemented by a direct numerical simulation approach, whereby representative surfaces can be scanned and the roughness effects computed over a range of Reynolds number. This development raises the prospect over the coming years of having enough data for different types of rough surfaces to be able to relate surface characteristics to roughness effects, such as the roughness function that quantifies the downward displacement of the logarithmic law of the wall. In the present contribution, we use simulation data for 17 irregular surfaces at the same friction Reynolds number, for which they are in the transitionally rough regime. All surfaces are scaled to the same physical roughness height. Mean streamwise velocity profiles show a wide range of roughness function values, while the velocity defect profiles show a good collapse. Profile peaks of the turbulent kinetic energy also vary depending on the surface. We then consider which surface properties are important and how new properties can be incorporated into an empirical model, the accuracy of which can then be tested. Optimised models with several roughness parameters are systematically developed for the roughness function and profile peak turbulent kinetic energy. In determining the roughness function, besides the known parameters of solidity (or frontal area ratio) and skewness, it is shown that the streamwise correlation length and the root-mean-square roughness height are also significant. The peak turbulent kinetic energy is determined by the skewness and root-mean-square roughness height, along with the mean forward-facing surface angle and spanwise effective slope. The results suggest feasibility of relating rough-wall flow properties (throughout the range from hydrodynamically smooth to fully rough) to surface

  7. Surface Habitat Systems

    Science.gov (United States)

    Kennedy, Kriss J.

    2009-01-01

    The Surface Habitat Systems (SHS) Focused Investment Group (FIG) is part of the National Aeronautics and Space Administration (NASA) Johnson Space Center (JSC) effort to provide a focused direction and funding to the various projects that are working on human surface habitat designs and technologies for the planetary exploration missions. The overall SHS-FIG effort focuses on directing and guiding those projects that: 1) develop and demonstrate new surface habitat system concepts, innovations, and technologies to support human exploration missions, 2) improve environmental systems that interact with human habitats, 3) handle and emplace human surface habitats, and 4) focus on supporting humans living and working in habitats on planetary surfaces. The activity areas of the SHS FIG described herein are focused on the surface habitat project near-term objectives as described in this document. The SHS-FIG effort focuses on mitigating surface habitat risks (as identified by the Lunar Surface Systems Project Office (LSSPO) Surface Habitat Element Team; and concentrates on developing surface habitat technologies as identified in the FY08 gap analysis. The surface habitat gap assessment will be updated annually as the surface architecture and surface habitat definition continues to mature. These technologies are mapped to the SHS-FIG Strategic Development Roadmap. The Roadmap will bring to light the areas where additional innovative efforts are needed to support the development of habitat concepts and designs and the development of new technologies to support of the LSSPO Habitation Element development plan. Three specific areas of development that address Lunar Architecture Team (LAT)-2 and Constellation Architecture Team (CxAT) Lunar habitat design issues or risks will be focused on by the SHS-FIG. The SHS-FIG will establish four areas of development that will help the projects prepare in their planning for surface habitat systems development. Those development areas are

  8. Mechanics of curved surfaces, with application to surface-parallel cracks

    Science.gov (United States)

    Martel, Stephen J.

    2011-10-01

    The surfaces of many bodies are weakened by shallow enigmatic cracks that parallel the surface. A re-formulation of the static equilibrium equations in a curvilinear reference frame shows that a tension perpendicular to a traction-free surface can arise at shallow depths even under the influence of gravity. This condition occurs if σ11k1 + σ22k2 > ρg cosβ, where k1 and k2 are the principal curvatures (negative if convex) at the surface, σ11 and σ22 are tensile (positive) or compressive (negative) stresses parallel to the respective principal curvature arcs, ρ is material density, g is gravitational acceleration, and β is the surface slope. The curvature terms do not appear in equilibrium equations in a Cartesian reference frame. Compression parallel to a convex surface thus can cause subsurface cracks to open. A quantitative test of the relationship above accounts for where sheeting joints (prominent shallow surface-parallel fractures in rock) are abundant and for where they are scarce or absent in the varied topography of Yosemite National Park, resolving key aspects of a classic problem in geology: the formation of sheeting joints. Moreover, since the equilibrium equations are independent of rheology, the relationship above can be applied to delamination or spalling caused by surface-parallel cracks in many materials.

  9. A Smart Superwetting Surface with Responsivity in Both Surface Chemistry and Microstructure.

    Science.gov (United States)

    Zhang, Dongjie; Cheng, Zhongjun; Kang, Hongjun; Yu, Jianxin; Liu, Yuyan; Jiang, Lei

    2018-03-26

    Recently, smart surfaces with switchable wettability have aroused much attention. However, only single surface chemistry or the microstructure can be changed on these surfaces, which significantly limits their wetting performances, controllability, and applications. A new surface with both tunable surface microstructure and chemistry was prepared by grafting poly(N-isopropylacrylamide) onto the pillar-structured shape memory polymer on which multiple wetting states from superhydrophilicity to superhydrophobicity can be reversibly and precisely controlled by synergistically regulating the surface microstructure and chemistry. Meanwhile, based on the excellent controllability, we also showed the application of the surface as a rewritable platform, and various gradient wettings can be obtained. This work presents for the first time a surface with controllability in both surface chemistry and microstructure, which starts some new ideas for the design of novel superwetting materials. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. A Surface Temperature Initiated Closure (STIC) for surface energy balance fluxes

    DEFF Research Database (Denmark)

    Mallick, Kaniska; Jarvis, Andrew J.; Boegh, Eva

    2014-01-01

    The use of Penman–Monteith (PM) equation in thermal remote sensing based surface energy balance modeling is not prevalent due to the unavailability of any direct method to integrate thermal data into the PM equation and due to the lack of physical models expressing the surface (or stomatal......) and boundary layer conductances (gS and gB) as a function of surface temperature. Here we demonstrate a new method that physically integrates the radiometric surface temperature (TS) into the PM equation for estimating the terrestrial surface energy balance fluxes (sensible heat, H and latent heat, λ......E). The method combines satellite TS data with standard energy balance closure models in order to derive a hybrid closure that does not require the specification of surface to atmosphere conductance terms. We call this the Surface Temperature Initiated Closure (STIC), which is formed by the simultaneous solution...

  11. 'The surface management system' (SuMS) database: a surface-based database to aid cortical surface reconstruction, visualization and analysis

    Science.gov (United States)

    Dickson, J.; Drury, H.; Van Essen, D. C.

    2001-01-01

    Surface reconstructions of the cerebral cortex are increasingly widely used in the analysis and visualization of cortical structure, function and connectivity. From a neuroinformatics perspective, dealing with surface-related data poses a number of challenges. These include the multiplicity of configurations in which surfaces are routinely viewed (e.g. inflated maps, spheres and flat maps), plus the diversity of experimental data that can be represented on any given surface. To address these challenges, we have developed a surface management system (SuMS) that allows automated storage and retrieval of complex surface-related datasets. SuMS provides a systematic framework for the classification, storage and retrieval of many types of surface-related data and associated volume data. Within this classification framework, it serves as a version-control system capable of handling large numbers of surface and volume datasets. With built-in database management system support, SuMS provides rapid search and retrieval capabilities across all the datasets, while also incorporating multiple security levels to regulate access. SuMS is implemented in Java and can be accessed via a Web interface (WebSuMS) or using downloaded client software. Thus, SuMS is well positioned to act as a multiplatform, multi-user 'surface request broker' for the neuroscience community.

  12. Imaging near-surface heterogeneities by natural migration of surface waves

    KAUST Repository

    Liu, Zhaolun

    2016-09-06

    We demonstrate that near-surface heterogeneities can be imaged by natural migration of backscattered surface waves in common shot gathers. No velocity model is required because the data are migrated onto surface points with the virtual Green\\'s functions computed from the shot gathers. Migrating shot gathers recorded by 2D and 3D land surveys validates the effectiveness of detecting nearsurface heterogeneities by natural migration. The implication is that more accurate hazard maps can be created by migrating surface waves in land surveys.

  13. Surface free energy of polypropylene and polycarbonate solidifying at different solid surfaces

    International Nuclear Information System (INIS)

    Chibowski, Emil; Terpilowski, Konrad

    2009-01-01

    Advancing and receding contact angles of water, formamide, glycerol and diiodomethane were measured on polypropylene (PP) and polycarbonate (PC) sample surfaces which solidified at Teflon, glass or stainless steel as matrix surfaces. Then from the contact angle hystereses (CAH) the apparent free energies γ s tot of the surfaces were evaluated. The original PP surface is practically nonpolar, possessing small electron donor interaction (γ s - =1.91mJ/m 2 ), as determined from the advancing contact angles of these liquids. It may result from impurities of the polymerization process. However, it increases up to 8-10 mJ/m 2 for PP surfaces contacted with the solids. The PC surfaces both original and modified show practically the same γ s - =6.56.7mJ/m 2 . No electron acceptor interaction is found on the surfaces. The γ s tot of modified PP and PC surfaces depend on the kind of probe liquid and contacted solid surface. The modified PP γ s tot values determined from CAH of polar liquids are greater than that of original surface and they increase in the sequence: Teflon, glass, stainless steel surface, at which they solidified. No clear dependence is observed between γ s tot and dielectric constant or dipole moment of the polar probe liquids. The changes in γ s tot of the polymer surfaces are due to the polymer nature and changes in its surface structure caused by the structure and force field of the contacting solid. It has been confirmed by AFM images.

  14. Atomic force microscopy analysis of different surface treatments of Ti dental implant surfaces

    International Nuclear Information System (INIS)

    Bathomarco, R.V.; Solorzano, G.; Elias, C.N.; Prioli, R.

    2004-01-01

    The surface of commercial unalloyed titanium, used in dental implants, was analyzed by atomic force microscopy. The morphology, roughness, and surface area of the samples, submitted to mechanically-induced erosion, chemical etching and a combination of both, were compared. The results show that surface treatments strongly influence the dental implant physical and chemical properties. An analysis of the length dependence of the implant surface roughness shows that, for scan sizes larger than 50 μm, the average surface roughness is independent of the scanning length and that the surface treatments lead to average surface roughness in the range of 0.37 up to 0.48 μm. It is shown that the implant surface energy is sensitive to the titanium surface area. As the area increases there is a decrease in the surface contact angle

  15. Atomic force microscopy analysis of different surface treatments of Ti dental implant surfaces

    Science.gov (United States)

    Bathomarco, Ti R. V.; Solorzano, G.; Elias, C. N.; Prioli, R.

    2004-06-01

    The surface of commercial unalloyed titanium, used in dental implants, was analyzed by atomic force microscopy. The morphology, roughness, and surface area of the samples, submitted to mechanically-induced erosion, chemical etching and a combination of both, were compared. The results show that surface treatments strongly influence the dental implant physical and chemical properties. An analysis of the length dependence of the implant surface roughness shows that, for scan sizes larger than 50 μm, the average surface roughness is independent of the scanning length and that the surface treatments lead to average surface roughness in the range of 0.37 up to 0.48 μm. It is shown that the implant surface energy is sensitive to the titanium surface area. As the area increases there is a decrease in the surface contact angle.

  16. Evaluation of different strategies for magnetic particle functionalization with DNA aptamers.

    Science.gov (United States)

    Pérez-Ruiz, Elena; Lammertyn, Jeroen; Spasic, Dragana

    2016-12-25

    The optimal bio-functionalization of magnetic particles is essential for developing magnetic particle-based bioassays. Whereas functionalization with antibodies is generally well established, immobilization of DNA probes, such as aptamers, is not yet fully explored. In this work, four different types of commercially available magnetic particles, coated with streptavidin, maleimide or carboxyl groups, were evaluated for their surface coverage with aptamer bioreceptors, efficiency in capturing target protein and non-specific protein adsorption on their surface. A recently developed aptamer against the peanut allergen, Ara h 1 protein, was used as a model system. Conjugation of biotinylated Ara h 1 aptamer to the streptavidin particles led to the highest surface coverage, whereas the coverage of maleimide particles was 25% lower. Carboxylated particles appeared to be inadequate for DNA functionalization. Streptavidin particles also showed the greatest target capturing efficiency, comparable to the one of particles functionalized with anti-Ara h 1 antibody. The performance of streptavidin particles was additionally tested in a sandwich assay with the aptamer as a capture receptor on the particle surface. While the limit of detection obtained was comparable to the same assay system with antibody as capture receptor, it was superior to previously reported values using the same aptamer in similar assay schemes with different detection platforms. These results point to the promising application of the Ara h 1 aptamer-functionalized particles in bioassay development. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Solid surface vs. liquid surface: nanoarchitectonics, molecular machines, and DNA origami.

    Science.gov (United States)

    Ariga, Katsuhiko; Mori, Taizo; Nakanishi, Waka; Hill, Jonathan P

    2017-09-13

    The investigation of molecules and materials at interfaces is critical for the accumulation of new scientific insights and technological advances in the chemical and physical sciences. Immobilization on solid surfaces permits the investigation of different properties of functional molecules or materials with high sensitivity and high spatial resolution. Liquid surfaces also present important media for physicochemical innovation and insight based on their great flexibility and dynamicity, rapid diffusion of molecular components for mixing and rearrangements, as well as drastic spatial variation in the prevailing dielectric environment. Therefore, a comparative discussion of the relative merits of the properties of materials when positioned at solid or liquid surfaces would be informative regarding present-to-future developments of surface-based technologies. In this perspective article, recent research examples of nanoarchitectonics, molecular machines, DNA nanotechnology, and DNA origami are compared with respect to the type of surface used, i.e. solid surfaces vs. liquid surfaces, for future perspectives of interfacial physics and chemistry.

  18. The influence of the surface atomic structure on surface diffusion

    International Nuclear Information System (INIS)

    Ghaleb, Dominique

    1984-03-01

    This work represents the first quantitative study of the influence of the surface atomic structure on surface diffusion (in the range: 0.2 Tf up 0.5 Tf; Tf: melting temperature of the substrate). The analysis of our results on a microscopic scale shows low formation and migration energies for adatoms; we can describe the diffusion on surfaces with a very simple model. On (110) surfaces at low temperature the diffusion is controlled by the exchange mechanism; at higher temperature direct jumps of adatoms along the channels contribute also to the diffusion process. (author) [fr

  19. Functionalized polymer film surfaces via surface-initiated atom transfer radical polymerization

    International Nuclear Information System (INIS)

    Hu, Y.; Li, J.S.; Yang, W.T.; Xu, F.J.

    2013-01-01

    The ability to manipulate and control the surface properties of polymer films, without altering the substrate properties, is crucial to their wide-spread applications. In this work, a simple one-step method for the direct immobilization of benzyl chloride groups (as the effective atom transfer radical polymerization (ATRP) initiators) on the polymer films was developed via benzophenone-induced coupling of 4-vinylbenzyl chloride (VBC). Polyethylene (PE) and nylon films were selected as examples of polymer films to illustrate the functionalization of film surfaces via surface-initiated ATRP. Functional polymer brushes of (2-dimethylamino)ethyl methacrylate, sodium 4-styrenesulfonate, 2-hydroxyethyl methacrylate and glycidyl methacrylate, as well as their block copolymer brushes, have been prepared via surface-initiated ATRP from the VBC-coupled PE or nylon film surfaces. With the development of a simple approach to the covalent immobilization of ATRP initiators on polymer film surfaces and the inherent versatility of surface-initiated ATRP, the surface functionality of polymer films can be precisely tailored. - Highlights: ► Atom transfer radical polymerization initiators were simply immobilized. ► Different functional polymer brushes were readily prepared. ► Their block copolymer brushes were also readily prepared

  20. Shape anisotropy enhanced optomagnetic measurement for prostate-specific antigen detection via magnetic chain formation

    DEFF Research Database (Denmark)

    Tian, Bo; Wetterskog, Erik; Qiu, Zhen

    2017-01-01

    anisotropy), and directly increasing the optomagnetic signal (via optical shape anisotropy). We achieve a limit of detection (LOD) of 5.5 pM (0.82 ng/mL) for the detection of a model multivalent molecule, biotinylated anti-streptavidin, in PBS. For the measurements of prostate-specific antigen (PSA) in 50...

  1. Syntaxin 1A interaction with the dopamine transporter promotes amphetamine-induced dopamine efflux

    DEFF Research Database (Denmark)

    Binda, Francesca; Dipace, Concetta; Bowton, Erica

    2008-01-01

    of the dopamine (DA) transporter (DAT) as the site of direct interaction with SYN1A. Amphetamine (AMPH) increases the association of SYN1A with human DAT (hDAT) in a heterologous expression system (hDAT cells) and with native DAT in murine striatal synaptosomes. Immunoprecipitation of DAT from the biotinylated...

  2. Cysteine Protease Inhibitors as Chemotherapy: Lessons from a Parasite Target

    Science.gov (United States)

    Selzer, Paul M.; Pingel, Sabine; Hsieh, Ivy; Ugele, Bernhard; Chan, Victor J.; Engel, Juan C.; Bogyo, Matthew; Russell, David G.; Sakanari, Judy A.; McKerrow, James H.

    1999-09-01

    Papain family cysteine proteases are key factors in the pathogenesis of cancer invasion, arthritis, osteoporosis, and microbial infections. Targeting this enzyme family is therefore one strategy in the development of new chemotherapy for a number of diseases. Little is known, however, about the efficacy, selectivity, and safety of cysteine protease inhibitors in cell culture or in vivo. We now report that specific cysteine protease inhibitors kill Leishmania parasites in vitro, at concentrations that do not overtly affect mammalian host cells. Inhibition of Leishmania cysteine protease activity was accompanied by defects in the parasite's lysosome/endosome compartment resembling those seen in lysosomal storage diseases. Colocalization of anti-protease antibodies with biotinylated surface proteins and accumulation of undigested debris and protease in the flagellar pocket of treated parasites were consistent with a pathway of protease trafficking from flagellar pocket to the lysosome/endosome compartment. The inhibitors were sufficiently absorbed and stable in vivo to ameliorate the pathology associated with a mouse model of Leishmania infection.

  3. Fluorescent Quantification of DNA Based on Core-Shell Fe3O4@SiO2@Au Nanocomposites and Multiplex Ligation-Dependent Probe Amplification.

    Science.gov (United States)

    Fan, Jing; Yang, Haowen; Liu, Ming; Wu, Dan; Jiang, Hongrong; Zeng, Xin; Elingarami, Sauli; Ll, Zhiyang; Li, Song; Liu, Hongna; He, Nongyue

    2015-02-01

    In this research, a novel method for relative fluorescent quantification of DNA based on Fe3O4@SiO2@Au gold-coated magnetic nanocomposites (GMNPs) and multiplex ligation- dependent probe amplification (MLPA) has been developed. With the help of self-assembly, seed-mediated growth and chemical reduction method, core-shell Fe3O4@SiO2@Au GMNPs were synthesized. Through modified streptavidin on the GMNPs surface, we obtained a bead chip which can capture the biotinylated probes. Then we designed MLPA probes which were tagged with biotin or Cy3 and target DNA on the basis of human APP gene sequence. The products from the thermostable DNA ligase induced ligation reactions and PCR amplifications were incubated with SA-GMNPs. After washing, magnetic separation, spotting, the fluorescent scanning results showed our method can be used for the relative quantitative analysis of the target DNA in the concentration range of 03004~0.5 µM.

  4. The hypertonic environment differentially regulates wild-type CFTR and TNR-CFTR chloride channels.

    Science.gov (United States)

    Lassance-Soares, Roberta M; Cheng, Jie; Krasnov, Kristina; Cebotaru, Liudmila; Cutting, Garry R; Souza-Menezes, Jackson; Morales, Marcelo M; Guggino, William B

    2010-01-01

    This study tested the hypotheses that the hypertonic environment of the renal medulla regulates the expression of cystic fibrosis transmembrane conductance regulator protein (CFTR) and its natural splice variant, TNR-CFTR. To accomplish this, Madin-Darby canine kidney (MDCK) stable cell lines expressing TNR-CFTR or CFTR were used. The cells were treated with hypertonic medium made with either NaCl or urea or sucrose (480 mOsm/kg or 560 mOsm/kg) to mimic the tonicity of the renal medulla environment. Western blot data showed that CFTR and TNR-CFTR total cell protein is increased by hypertonic medium, but using the surface biotinylation technique, only CFTR was found to be increased in cell plasma membrane. Confocal microscopy showed TNR-CFTR localization primarily at the endoplasmic reticulum and plasma membrane. In conclusion, CFTR and TNR-CFTR have different patterns of distribution in MDCK cells and they are modulated by a hypertonic environment, suggesting their physiological importance in renal medulla. Copyright © 2010 S. Karger AG, Basel.

  5. Screening for epitope specificity directly on culture supernatants in the early phase of monoclonal antibody production by an ELISA with biotin-labeled antigen.

    Science.gov (United States)

    Andersen, Ditte C; Jensen, Charlotte H; Gregersen, Annemette; Brandt, Jette; Kliem, Anette; Skjødt, Karsten; Koch, Claus; Teisner, Børge

    2004-01-01

    This report describes an assay for comparison of epitope specificity in groups of monoclonal antibodies against a given antigen. The only prerequisite is the biotin-labeled antigen. One of the monoclonal antibodies is captured onto a plastic surface via a rabbit anti-mouse Ig, and the other preincubated with biotinylated antigen. When the two antibodies react with the same epitope subsequent binding of the biotin-labeled antigen is abolished (inhibition). In the cases where no inhibition was observed, the two antibodies were considered to react with distinct, independent epitopes. The obvious advantages using this assay, are that it can be performed directly on culture supernatants in the early phase of monoclonal antibody production, and also works for antigens with repetitive epitopes. Moreover, the bonus effect, i.e., a signal in excess of the reference signal when sets of monoclonal antibodies with different epitope specificity are compared, gives a relative measure of affinity.

  6. Surface for dummies

    CERN Document Server

    Rathbone, Andy

    2014-01-01

    Make Microsoft's Surface work-and play-just the way you want it to Microsoft's Surface tablet has the features and personality you're looking for, with a robust environment for business computing that doesn't skimp on fun. Surface for Dummies, 2nd Edition explains how Windows 8.1 Pro and Windows RT differ, and helps you decide which Surface model is best for you. Step by step, this book walks you through both the hardware and software features of the Surface, including the touch cover and type cover, Windows RT and Windows 8.1 Pro operating systems, and the coveted Office Home & Student 2013 s

  7. Surface channeling

    International Nuclear Information System (INIS)

    Sizmann, R.; Varelas, C.

    1976-01-01

    There is experimental evidence that swift light ions incident at small angles towards single crystalline surfaces can lose an appreciable fraction of their kinetic energy during reflection. It is shown that these projectiles penetrate into the bulk surface region of the crystal. They can travel as channeled particles along long paths through the solid (surface channeling). The angular distribution and the depth history of the re-emerged projectiles are investigated by computer simulations. A considerable fraction of the penetrating projectiles re-emerges from the crystal with constant transverse energy if the angle of incidence is smaller than the critical angle for axial channeling. Analytical formulae are derived based on a diffusion model for surface channeling. A comparison with experimental data exhibits the relevance of the analytical solutions. (Auth.)

  8. Characterizing the effects of regolith surface roughness on photoemission from surfaces in space

    Science.gov (United States)

    Dove, A.; Horanyi, M.; Wang, X.

    2017-12-01

    Surfaces of airless bodies and spacecraft in space are exposed to a variety of charging environments. A balance of currents due to plasma bombardment, photoemission, electron and ion emission and collection, and secondary electron emission determines the surface's charge. Photoelectron emission is the dominant charging process on sunlit surfaces in the inner solar system due to the intense solar UV radiation. This can result in a net positive surface potential, with a cloud of photoelectrons immediately above the surface, called the photoelectron sheath. Conversely, the unlit side of the body will charge negatively due the collection of the fast-moving solar wind electrons. The interaction of charged dust grains with these positively and negatively charged surfaces, and within the photoelectron and plasma sheaths may explain the occurrence of dust lofting, levitation and transport above the lunar surface. The surface potential of exposed objects is also dependent on the material properties of their surfaces. Composition and particle size primarily affect the quantum efficiency of photoelectron generation; however, surface roughness can also control the charging process. In order to characterize these effects, we have conducted laboratory experiments to examine the role of surface roughness in generating photoelectrons in dedicated laboratory experiments using solid and dusty surfaces of the same composition (CeO2), and initial comparisons with JSC-1 lunar simulant. Using Langmuir probe measurements, we explore the measured potentials above insulating surfaces exposed to UV and an electric field, and we show that the photoemission current from a dusty surface is largely reduced due to its higher surface roughness, which causes a significant fraction of the emitted photoelectrons to be re-absorbed within the surface. We will discuss these results in context of similar situations on planetary surfaces.

  9. End-Point Immobilization of Recombinant Thrombomodulin via Sortase-Mediated Ligation

    Science.gov (United States)

    Jiang, Rui; Weingart, Jacob; Zhang, Hailong; Ma, Yong; Sun, Xue-Long

    2012-01-01

    We report an enzymatic end-point modification and immobilization of recombinant human thrombomodulin (TM), a cofactor for activation of anticoagulant protein C pathway via thrombin. First, a truncated TM mutant consisting of epidermal growth factor-like domains 4–6 (TM456) with a conserved pentapeptide LPETG motif at its C-terminal was expressed and purified in E. coli. Next, the truncated TM456 derivative was site-specifically modified with N-terminal diglycine containing molecules such as biotin and the fluorescent probe dansyl via sortase A (SrtA) mediated ligation (SML). The successful ligations were confirmed by SDS-PAGE and fluorescence imaging. Finally, the truncated TM456 was immobilized onto N-terminal diglycine-functionalized glass slide surface via SML directly. Alternatively, the truncated TM456 was biotinylated via SML and then immobilized onto streptavidin-functionalized glass slide surface indirectly. The successful immobilizations were confirmed by fluorescence imaging. The bioactivity of the immobilized truncated TM456 was further confirmed by protein C activation assay, in which enhanced activation of protein C by immobilized recombinant TM was observed. The sortase A-catalyzed surface ligation took place under mild conditions and is rapid occurring in a single step without prior chemical modification of the target protein. This site-specific covalent modification leads to molecules being arranged in a definitively ordered fashion and facilitating the preservation of the protein’s biological activity. PMID:22372933

  10. Comparison of different aminofunctionalization strategies for attachment of single antibodies to AFM cantilevers

    Energy Technology Data Exchange (ETDEWEB)

    Ebner, Andreas [Institute of Biophysics, University of Linz, 4040 Linz (Austria)], E-mail: andreas.ebner@jku.at; Hinterdorfer, Peter; Gruber, Hermann J. [Institute of Biophysics, University of Linz, 4040 Linz (Austria)

    2007-10-15

    Atomic force microscopy (AFM) has developed into a key technique for elucidation of biological systems on the single molecular level. In particular, molecular recognition force microscopy has proven to be a powerful tool for the investigation of biological interactions under near physiological conditions. For this purpose, ligands are tethered to AFM tips and the interaction forces with cognate receptors on the sample surface are measured with pico-Newton accuracy. In the first step of tip functionalization, amino groups are typically introduced on the initially inert AFM tip. Several methods have been developed to reproducibly adjust the desired low density of amino groups on the tip surface, i.e. esterification with ethanolamine, gas-phase silanization with aminopropyl-triethoxysilane (APTES), or treatment with aminophenyl-trimethoxysilane (APhS) in toluene solution. In the present study, the usefulness of these methods for attachments of antibodies to AFM tips was characterized by a standardized test system, in which biotinylated IgG was bound to the tip and a dense monolayer of avidin on mica served as test sample. All three methods of aminofunctionalization were found fully satisfactory for attachment of single antibodies to AFM tips, only in a parallel macroscopic assay on silicon nitride chips a minor difference was found in that APTES appeared to yield a slightly lower surface density of amino groups.

  11. Surface and body waves from surface and underground explosions

    International Nuclear Information System (INIS)

    Kusubov, A.S.

    1976-06-01

    The characteristics of surface and ground waves were recorded for surface and underground explosions up to 100 tons and 40 kt in magnitude, respectively, and a preliminary analysis of these results is presented. The experiments were conducted at NTS in the Yucca Flats, Nevada. Ground motions were detected with triaxial geophones along seismic lines extending up to 16 miles from the point of explosions. A comparison of Rayleigh waves generated by surface and underground explosions in the same lake bed is presented indicating a very different behavior of surface and ground waves from the two types of explosions. The magnitude of the transverse wave for surface shots was smaller by a factor of two than its longitudinal counterpart. The dependence of apparent periods on the blast energy was not apparent at a fixed distance from the explosions. Changes in the apparent period with distance for both types of explosion are compared indicating a strong layering effect of the lake bed. The ground motion study was complimented by excavation of cavities generated by the explosions

  12. Surface engineering with ion beams: from self-organized nanostructures to ultra-smooth surfaces

    International Nuclear Information System (INIS)

    Frost, F.; Ziberi, B.; Schindler, A.; Rauschenbach, B.

    2008-01-01

    Low-energy ion-beam sputtering, i.e. the removal of atoms from a surface due to the impact of energetic ions or atoms, is an inherent part of numerous surface processing techniques. Besides the actual removal of material, this surface erosion process often results in a pronounced alteration of the surface topography. Under certain conditions, sputtering results in the formation of well-ordered patterns. This self-organized pattern formation is related to a surface instability between curvature-dependent sputtering that roughens the surface and smoothing by different surface relaxation mechanisms. If the evolution of surface topography is dominated by relaxation mechanisms, surface smoothing can occur. In this presentation the current status of self-organized pattern formation and surface smoothing by low-energy ion-beam erosion of Si and Ge is summarized. In detail it will be shown that a multitude of patterns as well as ultra-smooth surfaces can develop, particularly on Si surfaces. Additionally, the most important experimental parameters that control these processes are discussed. Finally, examples are given for the application of low-energy ion beams as a novel approach for passive optical device engineering for many advanced optical applications. (orig.)

  13. A method for red blood cell biotinylation in a closed system

    NARCIS (Netherlands)

    de Back, Djuna Z.; Vlaar, Richard; Beuger, Boukje; Daal, Brunette; Lagerberg, Johan; Vlaar, Alexander P. J.; de Korte, Dirk; van Kraaij, Marian; van Bruggen, Robin

    2018-01-01

    Several circumstances require the accurate measurement of red blood cell (RBC) survival and clearance, such as determination of posttransfusion recovery of stored RBCs to investigate the effect of new additive solutions. To this end, biotin as a marker of RBCs to track donor RBCs in the blood of the

  14. Biomaterials surface science

    CERN Document Server

    Taubert, Andreas; Rodriguez-Cabello, José Carlos

    2013-01-01

    The book provides an overview of the highly interdisciplinary field of surface science in the context of biological and biomedical applications. The covered topics range from micro- and nanostructuring for imparting functionality in a top-down manner to the bottom-up fabrication of gradient surfaces by self-assembly, from interfaces between biomaterials and living matter to smart, stimuli-responsive surfaces, and from cell and surface mechanics to the elucidation of cell-chip interactions in biomedical devices.

  15. Influence of surface roughness on the friction property of textured surface

    Directory of Open Access Journals (Sweden)

    Yuankai Zhou

    2015-02-01

    Full Text Available In contrast with dimple textures, surface roughness is a texture at the micro-scale, essentially which will influence the load-bearing capacity of lubricant film. The numerical simulation was carried out to investigate the influence of surface roughness on friction property of textured surface. The lubricant film pressure was obtained using the method of computational fluid dynamics according to geometric model of round dimple, and the renormalization-group k–ε turbulent model was adopted in the computation. The numerical simulation results suggest that there is an optimum dimensionless surface roughness, and near this value, the maximum load-bearing capacity can be achieved. The load-bearing capacity is determined by the surface texture, the surface roughness, and the interaction between them. To get information of friction coefficient, the experiments were conducted. This experiment was used to evaluate the simulation. The experimental results show that for the frequency of 4 and 6 Hz, friction coefficient decreases at first and then increases with decreasing surface roughness, which indicates that there exists the optimum region of surface roughness leading to the best friction reduction effect, and it becomes larger when area fractions increase from 2% to 10%. The experimental results agree well with the simulation results.

  16. Analysis of surface bond lengths reported for chemisorption on metal surfaces

    Science.gov (United States)

    Mitchell, K. A. R.

    1985-01-01

    A review is given of bond length information available from the techniques of surface crystallography (particularly with LEED, SEXAFS and photoelectron diffraction) for chemisorption on well-defined surfaces of metals (M). For adsorbed main-group atoms (X), measured X-M interatomic distances for 38 combinations of X and M have been assessed with a bond order-bond length relation in combination with the Schomaker-Stevenson approach for determining single-bond lengths. When the surface bond orders are fixed primarily by the valency of X, this approach appears to provide a simple framework for predicing X-M surface bond lengths. Further, in cases where agreement has been reached from different surface crystallographic techniques, this framework has the potential for assessing refinements to the surface bonding model (e.g. in determining the roles of the effective surface valency of M, and of coordinate bonding and supplementary π bonding between X and M). Preliminary comparisons of structural data are also given for molecular adsorption (CO and ethylidyne) and for the chemisorption of other metal atoms.

  17. Mechanics of active surfaces

    Science.gov (United States)

    Salbreux, Guillaume; Jülicher, Frank

    2017-09-01

    We derive a fully covariant theory of the mechanics of active surfaces. This theory provides a framework for the study of active biological or chemical processes at surfaces, such as the cell cortex, the mechanics of epithelial tissues, or reconstituted active systems on surfaces. We introduce forces and torques acting on a surface, and derive the associated force balance conditions. We show that surfaces with in-plane rotational symmetry can have broken up-down, chiral, or planar-chiral symmetry. We discuss the rate of entropy production in the surface and write linear constitutive relations that satisfy the Onsager relations. We show that the bending modulus, the spontaneous curvature, and the surface tension of a passive surface are renormalized by active terms. Finally, we identify active terms which are not found in a passive theory and discuss examples of shape instabilities that are related to active processes in the surface.

  18. Radionuclide transfer onto ground surface in surface water flow, 1

    International Nuclear Information System (INIS)

    Mukai, Masayuki; Takebe, Shinichi; Komiya, Tomokazu; Kamiyama, Hideo

    1991-07-01

    Radionuclides migration in ground surface water flow is considered to be one of the important path way in the scenario for environmental migration of radionuclides leaked from low level radioactive waste repository. Simulating the slightly sloped surface on which contaminated solution is flowing downward, testing for radionuclide migration on ground surface had been started. As it's first step, an experiment was carried out under the condition of restricted infiltration in order to elucidate the adsorption behavior of radionuclides onto the loamy soil surface in related with hydraulic conditions. Radionuclides concentration change in effluent solution with time and a concentration distribution of radionuclides adsorbed on the ground surface were obtained from several experimental conditions combining the rate and the duration time of the water flow. The radionuclides concentration in the effluent solution was nearly constant during each experimental period, and was reduced under the condition of lower flow rate. The surface distribution of radionuclides concentration showed two distinctive regions. The one was near the inlet vessel where the concentration was promptly reducing, and the other was following the former where the concentration was nearly constant. The characteristic surface distribution of radionuclides concentration can be explained by a two dimensional diffusion model with a first order adsorption reaction, based on the advection of flow rate distribution in perpendicular direction. (author)

  19. Surface vibrational spectroscopy

    International Nuclear Information System (INIS)

    Erskine, J.L.

    1984-01-01

    A brief review of recent studies which combine measurements of surface vibrational energies with lattice dynamical calculations is presented. These results suggest that surface vibrational spectroscopy offers interesting prospects for use as a molecular-level probe of surface geometry, adsorbate bond distances and molecular orientations

  20. Surface functionalization of dopamine coated iron oxide nanoparticles for various surface functionalities

    Energy Technology Data Exchange (ETDEWEB)

    Sherwood, Jennifer; Xu, Yaolin; Lovas, Kira [Chemical and Biological Engineering, The University of Alabama, Tuscaloosa , AL 35487 (United States); Qin, Ying [Alabama Innovation and Mentoring of Entrepreneurs, The University of Alabama, Tuscaloosa, AL 35487 (United States); Bao, Yuping, E-mail: ybao@eng.ua.edu [Chemical and Biological Engineering, The University of Alabama, Tuscaloosa , AL 35487 (United States)

    2017-04-01

    We present effective conjugation of four small molecules (glutathione, cysteine, lysine, and Tris(hydroxymethyl)aminomethane) onto dopamine-coated iron oxide nanoparticles. Conjugation of these molecules could improve the surface functionality of nanoparticles for more neutral surface charge at physiological pH and potentially reduce non-specific adsorption of proteins to nanoparticles surfaces. The success of conjugation was evaluated with dynamic light scattering by measuring the surface charge changes and Fourier transform infrared spectroscopy for surface chemistry analysis. The stability of dopamine-coated nanoparticles and the ability of conjugated nanoparticles to reduce the formation of protein corona were evaluated by measuring the size and charge of the nanoparticles in biological medium. This facile conjugation method opens up possibilities for attaching various surface functionalities onto iron oxide nanoparticle surfaces for biomedical applications.

  1. Surface functionalization of dopamine coated iron oxide nanoparticles for various surface functionalities

    International Nuclear Information System (INIS)

    Sherwood, Jennifer; Xu, Yaolin; Lovas, Kira; Qin, Ying; Bao, Yuping

    2017-01-01

    We present effective conjugation of four small molecules (glutathione, cysteine, lysine, and Tris(hydroxymethyl)aminomethane) onto dopamine-coated iron oxide nanoparticles. Conjugation of these molecules could improve the surface functionality of nanoparticles for more neutral surface charge at physiological pH and potentially reduce non-specific adsorption of proteins to nanoparticles surfaces. The success of conjugation was evaluated with dynamic light scattering by measuring the surface charge changes and Fourier transform infrared spectroscopy for surface chemistry analysis. The stability of dopamine-coated nanoparticles and the ability of conjugated nanoparticles to reduce the formation of protein corona were evaluated by measuring the size and charge of the nanoparticles in biological medium. This facile conjugation method opens up possibilities for attaching various surface functionalities onto iron oxide nanoparticle surfaces for biomedical applications.

  2. Surface boxplots

    KAUST Repository

    Genton, Marc G.; Johnson, Christopher; Potter, Kristin; Stenchikov, Georgiy L.; Sun, Ying

    2014-01-01

    In this paper, we introduce a surface boxplot as a tool for visualization and exploratory analysis of samples of images. First, we use the notion of volume depth to order the images viewed as surfaces. In particular, we define the median image. We use an exact and fast algorithm for the ranking of the images. This allows us to detect potential outlying images that often contain interesting features not present in most of the images. Second, we build a graphical tool to visualize the surface boxplot and its various characteristics. A graph and histogram of the volume depth values allow us to identify images of interest. The code is available in the supporting information of this paper. We apply our surface boxplot to a sample of brain images and to a sample of climate model outputs.

  3. Surface boxplots

    KAUST Repository

    Genton, Marc G.

    2014-01-22

    In this paper, we introduce a surface boxplot as a tool for visualization and exploratory analysis of samples of images. First, we use the notion of volume depth to order the images viewed as surfaces. In particular, we define the median image. We use an exact and fast algorithm for the ranking of the images. This allows us to detect potential outlying images that often contain interesting features not present in most of the images. Second, we build a graphical tool to visualize the surface boxplot and its various characteristics. A graph and histogram of the volume depth values allow us to identify images of interest. The code is available in the supporting information of this paper. We apply our surface boxplot to a sample of brain images and to a sample of climate model outputs.

  4. Rumble surfaces

    CSIR Research Space (South Africa)

    National Institute for Transport and Road

    1977-01-01

    Full Text Available Rumble surfaces are intermittent short lengths of coarse-textured road surfacings on which vehicle tyres produce a rumbling sound. used in conjunction with appropriate roadsigns and markings, they can reduce accidents on rural roads by alerting...

  5. Comment on 'Surface thermodynamics, surface stress, equations at surfaces and triple lines for deformable bodies'

    International Nuclear Information System (INIS)

    Gutman, E M

    2010-01-01

    In a recent publication by Olives (2010 J. Phys.: Condens. Matter 22 085005) he studied 'the thermodynamics and mechanics of the surface of a deformable body, following and refining the general approach of Gibbs' and believed that 'a new definition of the surface stress is given'. However, using the usual way of deriving the equations of Gibbs-Duhem type the author, nevertheless, has fallen into a mathematical discrepancy because he has tried to unite in one equation different thermodynamic systems and 'a new definition of the surface stress' has appeared known in the usual theory of elasticity. (comment)

  6. Water at surfaces with tunable surface chemistries

    Science.gov (United States)

    Sanders, Stephanie E.; Vanselous, Heather; Petersen, Poul B.

    2018-03-01

    Aqueous interfaces are ubiquitous in natural environments, spanning atmospheric, geological, oceanographic, and biological systems, as well as in technical applications, such as fuel cells and membrane filtration. Where liquid water terminates at a surface, an interfacial region is formed, which exhibits distinct properties from the bulk aqueous phase. The unique properties of water are governed by the hydrogen-bonded network. The chemical and physical properties of the surface dictate the boundary conditions of the bulk hydrogen-bonded network and thus the interfacial properties of the water and any molecules in that region. Understanding the properties of interfacial water requires systematically characterizing the structure and dynamics of interfacial water as a function of the surface chemistry. In this review, we focus on the use of experimental surface-specific spectroscopic methods to understand the properties of interfacial water as a function of surface chemistry. Investigations of the air-water interface, as well as efforts in tuning the properties of the air-water interface by adding solutes or surfactants, are briefly discussed. Buried aqueous interfaces can be accessed with careful selection of spectroscopic technique and sample configuration, further expanding the range of chemical environments that can be probed, including solid inorganic materials, polymers, and water immiscible liquids. Solid substrates can be finely tuned by functionalization with self-assembled monolayers, polymers, or biomolecules. These variables provide a platform for systematically tuning the chemical nature of the interface and examining the resulting water structure. Finally, time-resolved methods to probe the dynamics of interfacial water are briefly summarized before discussing the current status and future directions in studying the structure and dynamics of interfacial water.

  7. Surface-electronic-state effects in electron emission from the Be(0001) surface

    International Nuclear Information System (INIS)

    Archubi, C. D.; Gravielle, M. S.; Silkin, V. M.

    2011-01-01

    We study the electron emission produced by swift protons impinging grazingly on a Be(0001) surface. The process is described within a collisional formalism using the band-structure-based (BSB) approximation to represent the electron-surface interaction. The BSB model provides an accurate description of the electronic band structure of the solid and the surface-induced potential. Within this approach we derive both bulk and surface electronic states, with these latter characterized by a strong localization at the crystal surface. We found that such surface electronic states play an important role in double-differential energy- and angle-resolved electron emission probabilities, producing noticeable structures in the electron emission spectra.

  8. Surface-electronic-state effects in electron emission from the Be(0001) surface

    Energy Technology Data Exchange (ETDEWEB)

    Archubi, C. D. [Instituto de Astronomia y Fisica del Espacio, casilla de correo 67, sucursal 28, C1428EGA, Buenos Aires (Argentina); Gravielle, M. S. [Instituto de Astronomia y Fisica del Espacio, casilla de correo 67, sucursal 28, C1428EGA, Buenos Aires (Argentina); Departamento de Fisica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires (Argentina); Silkin, V. M. [Donostia International Physics Center, E-20018 San Sebastian (Spain); Departamento de Fisica de Materiales, Facultad de Ciencias Quimicas, Universidad del Pais Vasco, Apartado 1072, E-20080 San Sebastian (Spain); IKERBASQUE, Basque Foundation for Science, E-48011 Bilbao (Spain)

    2011-07-15

    We study the electron emission produced by swift protons impinging grazingly on a Be(0001) surface. The process is described within a collisional formalism using the band-structure-based (BSB) approximation to represent the electron-surface interaction. The BSB model provides an accurate description of the electronic band structure of the solid and the surface-induced potential. Within this approach we derive both bulk and surface electronic states, with these latter characterized by a strong localization at the crystal surface. We found that such surface electronic states play an important role in double-differential energy- and angle-resolved electron emission probabilities, producing noticeable structures in the electron emission spectra.

  9. Thermal and UV Hydrosilylation of Alcohol-Based Bifunctional Alkynes on Si (111) surfaces: How surface radicals influence surface bond formation.

    Science.gov (United States)

    Khung, Y L; Ngalim, S H; Scaccabarozi, A; Narducci, D

    2015-06-12

    Using two different hydrosilylation methods, low temperature thermal and UV initiation, silicon (111) hydrogenated surfaces were functionalized in presence of an OH-terminated alkyne, a CF3-terminated alkyne and a mixed equimolar ratio of the two alkynes. XPS studies revealed that in the absence of premeditated surface radical through low temperature hydrosilylation, the surface grafting proceeded to form a Si-O-C linkage via nucleophilic reaction through the OH group of the alkyne. This led to a small increase in surface roughness as well as an increase in hydrophobicity and this effect was attributed to the surficial etching of silicon to form nanosize pores (~1-3 nm) by residual water/oxygen as a result of changes to surface polarity from the grafting. Furthermore in the radical-free thermal environment, a mix in equimolar of these two short alkynes can achieve a high contact angle of ~102°, comparable to long alkyl chains grafting reported in literature although surface roughness was relatively mild (rms = ~1 nm). On the other hand, UV initiation on silicon totally reversed the chemical linkages to predominantly Si-C without further compromising the surface roughness, highlighting the importance of surface radicals determining the reactivity of the silicon surface to the selected alkynes.

  10. Anticancer effects of monocarbonyl analogs of curcumin: oxidative stress, nuclear translocation and modulation of AP-1 and NF-κB

    Directory of Open Access Journals (Sweden)

    Brian Adams

    2015-01-01

    Full Text Available Purpose: In order to elucidate anticancer effects of monocarbonyl analogs of curcumin (MACs, we have undertaken the present study to obtain information regarding drug targets by using a microarray approach, and to study the cellular localization of EF24 and the activity of two key transcription factors, AP-1 and NF-κB, involved in complex cellular responses of cell survival and death. Methods: Cytotoxic activity of various drugs was evaluated using a Neutral Red Dye assay. Cellular localization of biotinylated EF24 (active and reduced EF24 (inactive was determined using light and confocal microscopy. Measurement of transcription factor binding was carried out using Transfactor ELISA kits (BD Clontech, Palo Alto, CA. Gene microarray processing was performed at Expression Analysis, Inc (Durham, NC using Affymetrix Human U133A Gene Chips.Results: In this study, we demonstrated that EF24 and UBS109 exhibit much more potent cytotoxic activity against pancreatic cancer than the current standard chemotherapeutic agent gemcitabine. EF24, rapidly localizes to the cell nucleus. The compound modulates the DNA binding activity of NF-κB and AP-1 in MDA-MB-231 human breast cancer cells and DU-145 human prostate cancer cells. Immunohistochemical studies utilizing biotinylated-EF24 and chemically-reduced EF24 show that the unsaturated compound and biotinylated EF24, but not reduced EF24, translocates to the nucleus within 30 minutes after the addition of drug. Through a gene microarray study, EF24 is shown to affect genes directly involved in cytoprotection, tumor growth, angiogenesis, metastasis and apoptosis. Conclusion: EF24 and UBS109 warrant further investigation for development of pancreatic cancer therapy. The dualistic modulations of gene expression may be a manifestation of the cell responses for survival against oxidative stress by EF24. However, the cytotoxic action of EF24 ultimately prevails to kill the cells.

  11. Computer aided surface representation

    Energy Technology Data Exchange (ETDEWEB)

    Barnhill, R E

    1987-11-01

    The aims of this research are the creation of new surface forms and the determination of geometric and physical properties of surfaces. The full sweep from constructive mathematics through the implementation of algorithms and the interactive computer graphics display of surfaces is utilized. Both three-dimensional and multi- dimensional surfaces are considered. Particular emphasis is given to the scientific computing solution of Department of Energy problems. The methods that we have developed and that we are proposing to develop allow applications such as: Producing smooth contour maps from measured data, such as weather maps. Modeling the heat distribution inside a furnace from sample measurements. Terrain modeling based on satellite pictures. The investigation of new surface forms includes the topics of triangular interpolants, multivariate interpolation, surfaces defined on surfaces and monotone and/or convex surfaces. The geometric and physical properties considered include contours, the intersection of surfaces, curvatures as a interrogation tool, and numerical integration.

  12. Modelling of surface evolution of rough surface on divertor target in fusion devices

    International Nuclear Information System (INIS)

    Dai, Shuyu; Liu, Shengguang; Sun, Jizhong; Kirschner, A.; Kawamura, G.; Tskhakaya, D.; Ding, Rui; Luo, Guangnan; Wang, Dezhen

    2015-01-01

    Highlights: • We study the surface evolution of rough surface on divertor target in fusion devices. • The effects of gyration motion and E × B drift affect 3D angular distribution. • A larger magnetic field angle leads to a reduced net eroded areal density. • The rough surface evolution affects the physical sputtering yield. - Abstract: The 3D Monte-Carlo code SURO has been used to study the surface evolution of rough surface on the divertor target in fusion devices. The edge plasma at divertor region is modelled by the SDPIC code and used as input data for SURO. Coupled with SDPIC, SURO can perform more sophisticated simulations to calculate the local angle and surface evolution of rough surface. The simulation results show that the incident direction of magnetic field, gyration and E × B force has a significant impact on 3D angular distribution of background plasma and accordingly on the erosion of rough surface. The net eroded areal density of rough surface is studied by varying the magnetic field angle with surface normal. The evolution of the microscopic morphology of rough surface can lead to a significant change in the physical sputtering yield

  13. Isolation and characterization of microsatellite DNA loci from Sillago ...

    Indian Academy of Sciences (India)

    A diluted digestion–ligation mixture (1:10) was amplified with adaptor-specific primers (MSEP: 5 -GAT GAG TCC. TGA GTA A-3 ). Amplified DNA fragments, with a size range of 200–1000 bp, were enriched for repeats by mag- netic bead selection with a 5 -biotinylated (AC)15 probes, respectively. Enriched fragments were ...

  14. Regularity of Minimal Surfaces

    CERN Document Server

    Dierkes, Ulrich; Tromba, Anthony J; Kuster, Albrecht

    2010-01-01

    "Regularity of Minimal Surfaces" begins with a survey of minimal surfaces with free boundaries. Following this, the basic results concerning the boundary behaviour of minimal surfaces and H-surfaces with fixed or free boundaries are studied. In particular, the asymptotic expansions at interior and boundary branch points are derived, leading to general Gauss-Bonnet formulas. Furthermore, gradient estimates and asymptotic expansions for minimal surfaces with only piecewise smooth boundaries are obtained. One of the main features of free boundary value problems for minimal surfaces is t

  15. Super Riemann surfaces

    International Nuclear Information System (INIS)

    Rogers, Alice

    1990-01-01

    A super Riemann surface is a particular kind of (1,1)-dimensional complex analytic supermanifold. From the point of view of super-manifold theory, super Riemann surfaces are interesting because they furnish the simplest examples of what have become known as non-split supermanifolds, that is, supermanifolds where the odd and even parts are genuinely intertwined, as opposed to split supermanifolds which are essentially the exterior bundles of a vector bundle over a conventional manifold. However undoubtedly the main motivation for the study of super Riemann surfaces has been their relevance to the Polyakov quantisation of the spinning string. Some of the papers on super Riemann surfaces are reviewed. Although recent work has shown all super Riemann surfaces are algebraic, some areas of difficulty remain. (author)

  16. Rough Surface Contact

    Directory of Open Access Journals (Sweden)

    T Nguyen

    2017-06-01

    Full Text Available This paper studies the contact of general rough curved surfaces having nearly identical geometries, assuming the contact at each differential area obeys the model proposed by Greenwood and Williamson. In order to account for the most general gross geometry, principles of differential geometry of surface are applied. This method while requires more rigorous mathematical manipulations, the fact that it preserves the original surface geometries thus makes the modeling procedure much more intuitive. For subsequent use, differential geometry of axis-symmetric surface is considered instead of general surface (although this “general case” can be done as well in Chapter 3.1. The final formulas for contact area, load, and frictional torque are derived in Chapter 3.2.

  17. PREFACE: Nanostructured surfaces

    Science.gov (United States)

    Palmer, Richard E.

    2003-10-01

    We can define nanostructured surfaces as well-defined surfaces which contain lateral features of size 1-100 nm. This length range lies well below the micron regime but equally above the Ångstrom regime, which corresponds to the interatomic distances on single-crystal surfaces. This special issue of Journal of Physics: Condensed Matter presents a collection of twelve papers which together address the fabrication, characterization, properties and applications of such nanostructured surfaces. Taken together they represent, in effect, a status report on the rapid progress taking place in this burgeoning area. The first four papers in this special issue have been contributed by members of the European Research Training Network ‘NanoCluster’, which is concerned with the deposition, growth and characterization of nanometre-scale clusters on solid surfaces—prototypical examples of nanoscale surface features. The paper by Vandamme is concerned with the fundamentals of the cluster-surface interaction; the papers by Gonzalo and Moisala address, respectively, the optical and catalytic properties of deposited clusters; and the paper by van Tendeloo reports the application of transmission electron microscopy (TEM) to elucidate the surface structure of spherical particles in a catalyst support. The fifth paper, by Mendes, is also the fruit of a European Research Training Network (‘Micro-Nano’) and is jointly contributed by three research groups; it reviews the creation of nanostructured surface architectures from chemically-synthesized nanoparticles. The next five papers in this special issue are all concerned with the characterization of nanostructured surfaces with scanning tunnelling microscopy (STM) and atomic force microscopy (AFM). The papers by Bolotov, Hamilton and Dunstan demonstrate that the STM can be employed for local electrical measurements as well as imaging, as illustrated by the examples of deposited clusters, model semiconductor structures and real

  18. Superhydrophobic surfaces

    Science.gov (United States)

    Wang, Evelyn N; McCarthy, Matthew; Enright, Ryan; Culver, James N; Gerasopoulos, Konstantinos; Ghodssi, Reza

    2015-03-24

    Surfaces having a hierarchical structure--having features of both microscale and nanoscale dimensions--can exhibit superhydrophobic properties and advantageous condensation and heat transfer properties. The hierarchical surfaces can be fabricated using biological nanostructures, such as viruses as a self-assembled nanoscale template.

  19. Ion neutralization at metal surfaces by surface-plasmon excitation

    International Nuclear Information System (INIS)

    Almulhem, A.A.

    1988-01-01

    Electron capture by ions scattered from metal surfaces is usually assumed to occur via resonance tunneling or Auger neutralization. A new mechanism is proposed, wherein a surface plasmon is excited during the electron capture. The Fock-Tani transformation is used to transform the Hamiltonian into a form which explicitly contains a term that corresponds to this process. Using this term, the matrix elements are calculated analytically and used to evaluate the transition rate as a function of distance from the surface. Since this is a rearrangement process, the matrix element contains an orthogonalization term. The theory is applied to the scattering of protons from an aluminum surface in which the proton captures an electron into the 1s state. From the results obtained for the transition rate and neutral fractions, it is concluded that this process is important, at least in the low energy region. When the calculations are done with the orthogonalization term in the matrix element neglected, the transition rate and neutral fraction increased appreciably. This shows the importance of this term, and implies that it cannot be neglected as was done in other theories of neutralization at metal surfaces

  20. Amperometric biosensor based on a single antibody of dual function for rapid detection of Streptococcus agalactiae.

    Science.gov (United States)

    Vásquez, Gersson; Rey, Alba; Rivera, Camilo; Iregui, Carlos; Orozco, Jahir

    2017-01-15

    Pathogenic bacteria are responsible for several diseases in humans and in a variety of hosts. Detection of pathogenic bacteria is imperative to avoid and/or fight their potential harmful effects. This work reports on the first amperometric biosensor for the rapid detection of Streptococcus agalactiae (S. agalactiae). The biosensor relies on a single biotinylated antibody that immobilizes the bacteria on a screen-printed carbon electrode while is further linked to a streptavidin-conjugated HRP reporter. The biotinylated antibody provides selectivity to the biosensor whereas serves as an anchoring point to the reporter for further amplification of the electrochemical signal. The resultant immunosensor is simple, responds rapidly, and allows for the selective and highly sensitive quantification of S. agalactiae cells in a concentration range of 10 1 -10 7 CFUml -1 , with a detection limit of 10CFUml -1 . The approach not only enables a rapid detection and quantification of S. agalactiae in environmental samples but also opens up new opportunities for the simple fabrication of electrochemical immunosensors for different target pathogens. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Sensitive detection of enteropathogenic E. coli using a bfpA gene-based electrochemical sensor

    International Nuclear Information System (INIS)

    Zhang, Wei; Luo, Caihui; Zhong, Liang; Zhao, Dan; Ding, Shijia; Nie, Shichang; Cheng, Wei

    2013-01-01

    We have developed a sensitive assay for enteropathogenic E. coli (EPEC) by integrating DNA extraction, specific polymerase chain reaction (PCR) and DNA detection using an electrode modified with the bundle-forming pilus (bfpA) structural gene. The PCR amplified products are captured on the electrode and hybridized with biotinylated detection probes to form a sandwich hybrid containing two biotinylated detection probes. The sandwich hybridization structure significantly combined the numerous streptavidin alkaline phosphatase on the electrode by biotin-streptavidin connectors. Electrochemical readout is based on dual signal amplification by both the sandwich hybridization structure and the enzyme. The electrode can satisfactorily discriminate complementary and mismatched oligonucleotides. Under optimal conditions, synthetic target DNA can be detected in the 1 pM to 10 nM concentration range, with a detection limit of 0.3 pM. EPEC can be quantified in the 10 to 10 7 CFU mL −1 levels within 3.5 h. The method also is believed to present a powerful platform for the screening of pathogenic microorganisms in clinical diagnostics, food safety and environmental monitoring. (author)

  2. Addition of electrophilic lipids to actin alters filament structure

    International Nuclear Information System (INIS)

    Gayarre, Javier; Sanchez, David; Sanchez-Gomez, Francisco J.; Terron, Maria C.; Llorca, Oscar; Perez-Sala, Dolores

    2006-01-01

    Pathophysiological processes associated with oxidative stress lead to the generation of reactive lipid species. Among them, lipids bearing unsaturated aldehyde or ketone moieties can form covalent adducts with cysteine residues and modulate protein function. Through proteomic techniques we have identified actin as a target for the addition of biotinylated analogs of the cyclopentenone prostaglandins 15-deoxy-Δ 12,14 -PGJ 2 (15d-PGJ 2 ) and PGA 1 in NIH-3T3 fibroblasts. This modification could take place in vitro and mapped to the protein C-terminal end. Other electrophilic lipids, like the isoprostane 8-iso-PGA 1 and 4-hydroxy-2-nonenal, also bound to actin. The C-terminal region of actin is important for monomer-monomer interactions and polymerization. Electron microscopy showed that actin treated with 15d-PGJ 2 or 4-hydroxy-2-nonenal formed filaments which were less abundant and displayed shorter length and altered structure. Streptavidin-gold staining allowed mapping of biotinylated 15d-PGJ 2 at sites of filament disruption. These results shed light on the structural implications of actin modification by lipid electrophiles

  3. siRNA associated with immunonanoparticles directed against cd99 antigen improves gene expression inhibition in vivo in Ewing's sarcoma.

    Science.gov (United States)

    Ramon, A L; Bertrand, J R; de Martimprey, H; Bernard, G; Ponchel, G; Malvy, C; Vauthier, C

    2013-07-01

    Ewing's sarcoma is a rare, mostly pediatric bone cancer that presents a chromosome abnormality called EWS/Fli-1, responsible for the development of the tumor. In vivo, tumor growth can be inhibited specifically by delivering small interfering RNA (siRNA) associated with nanoparticles. The aim of the work was to design targeted nanoparticles against the cell membrane glycoprotein cd99, which is overexpressed in Ewing's sarcoma cells to improve siRNA delivery to tumor cells. Biotinylated poly(isobutylcyanoacrylate) nanoparticles were conceived as a platform to design targeted nanoparticles with biotinylated ligands and using the biotin-streptavidin coupling method. The targeted nanoparticles were validated in vivo for the targeted delivery of siRNA after systemic administration to mice bearing a tumor model of the Ewing's sarcoma. The expression of the gene responsible of Ewing's sarcoma was inhibited at 78% ± 6% by associating the siRNA with the cd99-targeted nanoparticles compared with an inhibition of only 41% ± 9% achieved with the nontargeted nanoparticles. Copyright © 2013 John Wiley & Sons, Ltd.

  4. Computer aided surface representation

    Energy Technology Data Exchange (ETDEWEB)

    Barnhill, R.E.

    1990-02-19

    The central research problem of this project is the effective representation, computation, and display of surfaces interpolating to information in three or more dimensions. If the given information is located on another surface, then the problem is to construct a surface defined on a surface''. Sometimes properties of an already defined surface are desired, which is geometry processing''. Visualization of multivariate surfaces is possible by means of contouring higher dimensional surfaces. These problems and more are discussed below. The broad sweep from constructive mathematics through computational algorithms to computer graphics illustrations is utilized in this research. The breadth and depth of this research activity makes this research project unique.

  5. Surface states in crystals with low-index surfaces

    International Nuclear Information System (INIS)

    Wang Hui-Ping; Tao Rui-Bao

    2015-01-01

    For most of the conventional crystals with low-index surfaces, the hopping between the nearest neighbor (1NN) crystal planes (CPs) is dominant and the ones from the nNN (2 ≤ n < ∞) CPs are relatively weak, considered as small perturbations. The recent theoretical analysis [1] has demonstrated the absence of surface states at the level of the hopping approximation between the 1NN CPs when the original infinite crystal has the geometric reflection symmetry (GRS) for each CP. Meanwhile, based on the perturbation theory, it has also been shown that small perturbations from the hopping between the nNN (2 ≤ n < ∞) CPs and surface relaxation have no impact on the above conclusion. However, for the crystals with strong intrinsic spin-orbit coupling (SOC), the dominant terms of intrinsic SOC associate with two 1NN bond hoppings. Thus SOC will significantly contribute the hoppings from the 1NN and/or 2NN CPs except the ones within each CP. Here, we will study the effect of the hopping between the 2NN CPs on the surface states in model crystals with three different type structures (Type I: “···–P–P–P–P–···”, Type II: “···–P–Q–P–Q–···” and Type III: “···–P=Q–P=Q–···” where P and Q indicate CPs and the signs “−” and “=” mark the distance between the 1NN CPs). In terms of analytical and numerical calculations, we study the behavior of surface states in three types after the symmetric/asymmetric hopping from the 2NN CPs is added. We analytically prove that the symmetric hopping from the 2NN CPs cannot induce surface states in Type I when each CP has only one electron mode. The numerical calculations also provide strong support for the conclusion, even up to 5NN. However, in general, the coupling from the 2NN CPs (symmetric and asymmetric) is favorable to generate surface states except Type I with single electron mode only. (paper)

  6. Surface chemistry essentials

    CERN Document Server

    Birdi, K S

    2013-01-01

    Surface chemistry plays an important role in everyday life, as the basis for many phenomena as well as technological applications. Common examples range from soap bubbles, foam, and raindrops to cosmetics, paint, adhesives, and pharmaceuticals. Additional areas that rely on surface chemistry include modern nanotechnology, medical diagnostics, and drug delivery. There is extensive literature on this subject, but most chemistry books only devote one or two chapters to it. Surface Chemistry Essentials fills a need for a reference that brings together the fundamental aspects of surface chemistry w

  7. Symmetric and asymmetric capillary bridges between a rough surface and a parallel surface.

    Science.gov (United States)

    Wang, Yongxin; Michielsen, Stephen; Lee, Hoon Joo

    2013-09-03

    Although the formation of a capillary bridge between two parallel surfaces has been extensively studied, the majority of research has described only symmetric capillary bridges between two smooth surfaces. In this work, an instrument was built to form a capillary bridge by squeezing a liquid drop on one surface with another surface. An analytical solution that describes the shape of symmetric capillary bridges joining two smooth surfaces has been extended to bridges that are asymmetric about the midplane and to rough surfaces. The solution, given by elliptical integrals of the first and second kind, is consistent with a constant Laplace pressure over the entire surface and has been verified for water, Kaydol, and dodecane drops forming symmetric and asymmetric bridges between parallel smooth surfaces. This solution has been applied to asymmetric capillary bridges between a smooth surface and a rough fabric surface as well as symmetric bridges between two rough surfaces. These solutions have been experimentally verified, and good agreement has been found between predicted and experimental profiles for small drops where the effect of gravity is negligible. Finally, a protocol for determining the profile from the volume and height of the capillary bridge has been developed and experimentally verified.

  8. Plasma-surface interactions

    International Nuclear Information System (INIS)

    Goeckner, M J; Nelson, C T; Sant, S P; Jindal, A K; Joseph, E A; Zhou, B S; Padron-Wells, G; Jarvis, B; Pierce, R; Overzet, L J

    2008-01-01

    Materials processing is at a crossroads. Currently a large fraction of industrially viable materials processing is via plasmas. Until recently it has been economical to just examine the influence the plasma properties on the desired surface processes and through this ultimately optimize manufacturing. For example, it is well known that the surface processes (etch or deposition), occur in the top few mono-layers of the surface. Thus, in film growth one requires that molecules from the gas-phase land and bond on the surface. However as processing has reached the nano-scale, development of viable processes has become more and more difficult. In part, this is because of all of the free parameters that exist in plasmas. To overcome this economic issue, tool vendors and semiconductor companies have turned to complex computational models of processing plasmas. For those models to work, one requires a through understanding of all of the gas-phase and surface-phase processes that are exhibited in plasmas. Unfortunately, these processes, particularly those at the surface, are not well understood. In this article we describe a viable model of the surface-phase based on cross sections for processes that occur. While originally developed of fluorocarbon systems, the model also appears to be applicable to hydrocarbon systems.

  9. Plasma-surface interactions

    Energy Technology Data Exchange (ETDEWEB)

    Goeckner, M J; Nelson, C T; Sant, S P; Jindal, A K; Joseph, E A; Zhou, B S; Padron-Wells, G; Jarvis, B; Pierce, R; Overzet, L J [Department of Electrical Engineering, University of Texas at Dallas (United States)], E-mail: goeckner@utdallas.edu

    2008-10-01

    Materials processing is at a crossroads. Currently a large fraction of industrially viable materials processing is via plasmas. Until recently it has been economical to just examine the influence the plasma properties on the desired surface processes and through this ultimately optimize manufacturing. For example, it is well known that the surface processes (etch or deposition), occur in the top few mono-layers of the surface. Thus, in film growth one requires that molecules from the gas-phase land and bond on the surface. However as processing has reached the nano-scale, development of viable processes has become more and more difficult. In part, this is because of all of the free parameters that exist in plasmas. To overcome this economic issue, tool vendors and semiconductor companies have turned to complex computational models of processing plasmas. For those models to work, one requires a through understanding of all of the gas-phase and surface-phase processes that are exhibited in plasmas. Unfortunately, these processes, particularly those at the surface, are not well understood. In this article we describe a viable model of the surface-phase based on cross sections for processes that occur. While originally developed of fluorocarbon systems, the model also appears to be applicable to hydrocarbon systems.

  10. Surface-ionization field mass-spectrometry studies of nonequilibrium surface ionization

    International Nuclear Information System (INIS)

    Blashenkov, Nikolai M; Lavrent'ev, Gennadii Ya

    2007-01-01

    The ionization of polyatomic molecules on tungsten and tungsten oxide surfaces is considered for quasiequilibrium or essentially nonequilibrium conditions (in the latter case, the term nonequilibrium surface ionization is used for adsorbate ionization). Heterogeneous reactions are supposed to proceed through monomolecular decay of polyatomic molecules or fragments of multimolecular complexes. The nonequilibrium nature of these reactions is established. The dependences of the current density of disordered ions on the surface temperature, electric field strength, and ionized particle energy distribution are obtained in analytical form. Heterogeneous dissociation energies, the ionization potentials of radicals, and the magnitude of reaction departure from equilibrium are determined from experimental data, as are energy exchange times between reaction products and surfaces, the number of molecules in molecular complexes, and the number of effective degrees of freedom in molecules and complexes. In collecting the data a new technique relying on surface-ionization field mass-spectrometry was applied. (instruments and methods of investigation)

  11. Direct detection of near-surface faults by migration of back-scattered surface waves

    KAUST Repository

    Yu, Han; Guo, Bowen; Hanafy, Sherif; Lin, Fan-Chi; Schuster, Gerard T.

    2014-01-01

    We show that diffraction stack migration can be used to estimate the distribution of near-surface faults. The assumption is that near-surface faults generate detectable back-scattered surface waves from impinging surface waves. The processing steps

  12. Surface freezing of water.

    Science.gov (United States)

    Pérez-Díaz, J L; Álvarez-Valenzuela, M A; Rodríguez-Celis, F

    2016-01-01

    Freezing, melting, evaporation and condensation of water are essential ingredients for climate and eventually life on Earth. In the present work, we show how surface freezing of supercooled water in an open container is conditioned and triggered-exclusively-by humidity in air. Additionally, a change of phase is demonstrated to be triggered on the water surface forming surface ice crystals prior to freezing of bulk. The symmetry of the surface crystal, as well as the freezing point, depend on humidity, presenting at least three different types of surface crystals. Humidity triggers surface freezing as soon as it overpasses a defined value for a given temperature, generating a plurality of nucleation nodes. An evidence of simultaneous nucleation of surface ice crystals is also provided.

  13. Surface chemistry theory and applications

    CERN Document Server

    Bikerman, J J

    2013-01-01

    Surface Chemistry Theory and Applications focuses on liquid-gas, liquid-liquid, solid-gas, solid-liquid, and solid-solid surfaces. The book first offers information on liquid-gas surfaces, including surface tension, measurement of surface tension, rate of capillarity rise, capillary attraction, bubble pressure and pore size, and surface tension and temperature. The text then ponders on liquid-liquid and solid-gas surfaces. Discussions focus on surface energy of solids, surface roughness and cleanness, adsorption of gases and vapors, adsorption hysteresis, interfacial tension, and interfacial t

  14. PREFACE: Atom-surface scattering Atom-surface scattering

    Science.gov (United States)

    Miret-Artés, Salvador

    2010-08-01

    It has been a privilege and a real pleasure to organize this special issue or festschrift in the general field of atom-surface scattering (and its interaction) in honor of J R Manson. This is a good opportunity and an ideal place to express our deep gratitude to one of the leaders in this field for his fundamental and outstanding scientific contributions. J R Manson, or Dick to his friends and colleagues, is one of the founding fathers, together with N Cabrera and V Celli, of the 'Theory of surface scattering and detection of surface phonons'. This is the title of the very well-known first theoretical paper by Dick published in Physical Review Letters in 1969. My first meeting with Dick was around twenty years ago in Saclay. J Lapujoulade organized a small group seminar about selective adsorption resonances in metal vicinal surfaces. We discussed this important issue in surface physics and many other things as if we had always known each other. This familiarity and warm welcome struck me from the very beginning. During the coming years, I found this to be a very attractive aspect of his personality. During my stays in Göttingen, we had the opportunity to talk widely about science and life at lunch or dinner time, walking or cycling. During these nice meetings, he showed, with humility, an impressive cultural background. It is quite clear that his personal opinions about history, religion, politics, music, etc, come from considering and analyzing them as 'open dynamical systems'. In particular, with good food and better wine in a restaurant or at home, a happy cheerful soirée is guaranteed with him, or even with only a good beer or espresso, and an interesting conversation arises naturally. He likes to listen before speaking. Probably not many people know his interest in tractors. He has an incredible collection of very old tractors at home. In one of my visits to Clemson, he showed me the collection, explaining to me in great detail, their technical properties

  15. Surfaces with Natural Ridges

    DEFF Research Database (Denmark)

    Brander, David; Markvorsen, Steen

    2015-01-01

    We discuss surfaces with singularities, both in mathematics and in the real world. For many types of mathematical surface, singularities are natural and can be regarded as part of the surface. The most emblematic example is that of surfaces of constant negative Gauss curvature, all of which...

  16. Surface Forces Apparatus measurements of interactions between rough and reactive calcite surfaces.

    Science.gov (United States)

    Dziadkowiec, Joanna; Javadi, Shaghayegh; Bratvold, Jon Einar; Nilsen, Ola; Røyne, Anja

    2018-05-28

    Nm-range forces acting between calcite surfaces in water affect macroscopic properties of carbonate rocks and calcite-based granular materials, and are significantly influenced by calcite surface recrystallization. We suggest that the repulsive mechanical effects related to nm-scale surface recrystallization of calcite in water could be partially responsible for the observed decrease of cohesion in calcitic rocks saturated with water. Using the Surface Forces Apparatus (SFA), we simultaneously followed the calcite reactivity and measured the forces in water in two surface configurations: between two rough calcite surfaces (CC), or between rough calcite and a smooth mica surface (CM). We used nm-scale rough, polycrystalline calcite films prepared by Atomic Layer Deposition (ALD). We measured only repulsive forces in CC in CaCO 3 -saturated water, which was related to roughness and possibly to repulsive hydration effects. Adhesive or repulsive forces were measured in CM in CaCO 3 -saturated water depending on calcite roughness, and the adhesion was likely enhanced by electrostatic effects. The pull-off adhesive force in CM became stronger with time and this increase was correlated with a decrease of roughness at contacts, which parameter could be estimated from the measured force-distance curves. That suggested a progressive increase of real contact areas between the surfaces, caused by gradual pressure-driven deformation of calcite surface asperities during repeated loading-unloading cycles. Reactivity of calcite was affected by mass transport across nm to µm-thick gaps between the surfaces. Major roughening was observed only for the smoothest calcite films, where gaps between two opposing surfaces were nm-thick over µm-sized areas, and led to force of crystallization that could overcome confining pressures of the order of MPa. Any substantial roughening of calcite caused a significant increase of the repulsive mechanical force contribution.

  17. On rationally supported surfaces

    DEFF Research Database (Denmark)

    Gravesen, Jens; Juttler, B.; Sir, Z.

    2008-01-01

    We analyze the class of surfaces which are equipped with rational support functions. Any rational support function can be decomposed into a symmetric (even) and an antisymmetric (odd) part. We analyze certain geometric properties of surfaces with odd and even rational support functions....... In particular it is shown that odd rational support functions correspond to those rational surfaces which can be equipped with a linear field of normal vectors, which were discussed by Sampoli et al. (Sampoli, M.L., Peternell, M., Juttler, B., 2006. Rational surfaces with linear normals and their convolutions...... with rational surfaces. Comput. Aided Geom. Design 23, 179-192). As shown recently, this class of surfaces includes non-developable quadratic triangular Bezier surface patches (Lavicka, M., Bastl, B., 2007. Rational hypersurfaces with rational convolutions. Comput. Aided Geom. Design 24, 410426; Peternell, M...

  18. Polaron-Driven Surface Reconstructions

    Directory of Open Access Journals (Sweden)

    Michele Reticcioli

    2017-09-01

    Full Text Available Geometric and electronic surface reconstructions determine the physical and chemical properties of surfaces and, consequently, their functionality in applications. The reconstruction of a surface minimizes its surface free energy in otherwise thermodynamically unstable situations, typically caused by dangling bonds, lattice stress, or a divergent surface potential, and it is achieved by a cooperative modification of the atomic and electronic structure. Here, we combined first-principles calculations and surface techniques (scanning tunneling microscopy, non-contact atomic force microscopy, scanning tunneling spectroscopy to report that the repulsion between negatively charged polaronic quasiparticles, formed by the interaction between excess electrons and the lattice phonon field, plays a key role in surface reconstructions. As a paradigmatic example, we explain the (1×1 to (1×2 transition in rutile TiO_{2}(110.

  19. Approximation by Cylinder Surfaces

    DEFF Research Database (Denmark)

    Randrup, Thomas

    1997-01-01

    We present a new method for approximation of a given surface by a cylinder surface. It is a constructive geometric method, leading to a monorail representation of the cylinder surface. By use of a weighted Gaussian image of the given surface, we determine a projection plane. In the orthogonal...

  20. Direct detection of near-surface faults by migration of back-scattered surface waves

    KAUST Repository

    Yu, Han

    2014-08-05

    We show that diffraction stack migration can be used to estimate the distribution of near-surface faults. The assumption is that near-surface faults generate detectable back-scattered surface waves from impinging surface waves. The processing steps are to isolate the back-scattered surface waves, and then migrate them by diffraction migration using the surface wave velocity as the migration velocity. Instead of summing events along trial quasi-hyperbolas, surface wave migration sums events along trial quasi-linear trajectories that correspond to the moveout of back-scattered surface waves. A deconvolution filter derived from the data can be used to collapse a dispersive arrival into a non-dispersive event. Results with synthetic data and field records validate the feasibility of this method. Applying this method to USArray data or passively recorded exploration data might open new opportunities in mapping tectonic features over the extent of the array.

  1. Effect of Surface Treatment on the Surface Characteristics of AISI 316L Stainless Steel

    Science.gov (United States)

    Trigwell, Steve; Selvaduray, Guna

    2005-01-01

    The ability of 316L stainless steel to maintain biocompatibility, which is dependent upon the surface characteristics, is critical to its effectiveness as an implant material. The surfaces of mechanically polished (MP), electropolished (EP) and plasma treated 316L stainless steel coupons were characterized by X-ray Photoelectron Spectroscopy (XPS) and Auger Electron Spectroscopy (AES) for chemical composition, Atomic Force Microscopy for surface roughness, and contact angle measurements for critical surface tension. All surfaces had a Ni concentration that was significantly lower than the bulk concentration of -43%. The Cr content of the surface was increased significantly by electropolishing. The surface roughness was also improved significantly by electropolishing. Plasma treatment had the reverse effect - the surface Cr content was decreased. It was also found that the Cr and Fe in the surface exist in both the oxide and hydroxide states, with the ratios varying according to surface treatment.

  2. Workbench surface editor of brain cortical surface

    Science.gov (United States)

    Dow, Douglas E.; Nowinski, Wieslaw L.; Serra, Luis

    1996-04-01

    We have developed a 3D reach-in tool to manually reconstruct 3D cortical surface patches from 2D brain atlas images. The first application of our cortex editor is building 3D functional maps, specifically Brodmann's areas. This tool may also be useful in clinical practice to adjust incorrectly mapped atlas regions due to the deforming effect of lesions. The cortex editor allows a domain expert to control the correlation of control points across slices. Correct correlation has been difficult for 3D reconstruction algorithms because the atlas slices are far apart and because of the complex topology of the cortex which differs so much from slice to slice. Also, higher precision of the resulting surfaces is demanded since these define 3D brain atlas features upon which future stereotactic surgery may be based. The cortex editor described in this paper provides a tool suitable for a domain expert to use in defining the 3D surface of a Brodmann's area.

  3. Surface topography and morphology characterization of PIII irradiated silicon surface

    International Nuclear Information System (INIS)

    Sharma, Satinder K.; Barthwal, Sumit

    2008-01-01

    The effect of plasma immersion ion implantation (PIII) treatment on silicon surfaces was investigated by micro-Raman and atomic force microscopy (AFM) technique. The surface damage was given by the implantation of carbon, nitrogen, oxygen and argon ions using an inductively coupled plasma (ICP) source at low pressure. AFM studies show that surface topography of the PIII treated silicon wafers depend on the physical and chemical nature of the implanted species. Micro-Raman spectra indicate that the significant reduction of intensity of Raman peak after PIII treatment. Plasma immersion ion implantation is a non-line-of-sight ion implantation method, which allows 3D treatment of materials. Therefore, PIII based surface modification and plasma immersion ion deposition (PIID) coatings are applied in a wide range of situations.

  4. Adsorption of surface functionalized silica nanoparticles onto mineral surfaces and decane/water interface

    International Nuclear Information System (INIS)

    Metin, Cigdem O.; Baran, Jimmie R.; Nguyen, Quoc P.

    2012-01-01

    The adsorption of silica nanoparticles onto representative mineral surfaces and at the decane/water interface was studied. The effects of particle size (the mean diameters from 5 to 75 nm), concentration and surface type on the adsorption were studied in detail. Silica nanoparticles with four different surfaces [unmodified, surface modified with anionic (sulfonate), cationic (quaternary ammonium (quat)) or nonionic (polyethylene glycol (PEG)) surfactant] were used. The zeta potential of these silica nanoparticles ranges from −79.8 to 15.3 mV. The shape of silica particles examined by a Hitachi-S5500 scanning transmission electron microscope (STEM) is quite spherical. The adsorption of all the nanoparticles (unmodified or surface modified) on quartz and calcite surfaces was found to be insignificant. We used interfacial tension (IFT) measurements to investigate the adsorption of silica nanoparticles at the decane/water interface. Unmodified nanoparticles or surface modified ones with sulfonate or quat do not significantly affect the IFT of the decane/water interface. It also does not appear that the particle size or concentration influences the IFT. However, the presence of PEG as a surface modifying material significantly reduces the IFT. The PEG surface modifier alone in an aqueous solution, without the nanoparticles, yields the same IFT reduction for an equivalent PEG concentration as that used for modifying the surface of nanoparticles. Contact angle measurements of a decane droplet on quartz or calcite plate immersed in water (or aqueous nanoparticle dispersion) showed a slight change in the contact angle in the presence of the studied nanoparticles. The results of contact angle measurements are in good agreement with experiments of adsorption of nanoparticles on mineral surfaces or decane/water interface. This study brings new insights into the understanding and modeling of the adsorption of surface-modified silica nanoparticles onto mineral surfaces and

  5. Ruled Laguerre minimal surfaces

    KAUST Repository

    Skopenkov, Mikhail; Pottmann, Helmut; Grohs, Philipp

    2011-01-01

    A Laguerre minimal surface is an immersed surface in ℝ 3 being an extremal of the functional ∫ (H 2/K-1)dA. In the present paper, we prove that the only ruled Laguerre minimal surfaces are up to isometry the surfaces ℝ (φλ) = (Aφ, Bφ, Cφ + D cos 2φ

  6. Fabrication of a bionic microstructure on a C/SiC brake lining surface: Positive applications of surface defects for surface wetting control

    Science.gov (United States)

    Wu, M. L.; Ren, C. Z.; Xu, H. Z.; Zhou, C. L.

    2018-05-01

    The material removal processes generate interesting surface topographies, unfortunately, that was usually considered to be surface defects. To date, little attention has been devoted to the positive applications of these interesting surface defects resulted from laser ablation to improve C/SiC surface wettability. In this study, the formation mechanism behind surface defects (residual particles) is discussed first. The results showed that the residual particles with various diameters experienced regeneration and migration, causing them to accumulate repeatedly. The effective accumulation of these residual particles with various diameters provides a new method about fabricating bionic microstructures for surface wetting control. The negligible influence of ablation processes on the chemical component of the subsurface was studied by comparing the C-O-Si weight percentage at the C/SiC subsurface. A group of microstructures were fabricated under different laser trace and different laser parameters. Surface wettability experimental results for different types of microstructures were compared. The results showed that the surface wettability increased as the laser scanning speed decreased. The surface wettability increased with the density of the laser scanning trace. We also demonstrated the application of optimized combination of laser parameters and laser trace to simulate a lotus leaf's microstructure on C/SiC surfaces. The parameter selection depends on the specific material properties.

  7. Surface mining

    Science.gov (United States)

    Robert Leopold; Bruce Rowland; Reed Stalder

    1979-01-01

    The surface mining process consists of four phases: (1) exploration; (2) development; (3) production; and (4) reclamation. A variety of surface mining methods has been developed, including strip mining, auger, area strip, open pit, dredging, and hydraulic. Sound planning and design techniques are essential to implement alternatives to meet the myriad of laws,...

  8. Estimation of surface absorptivity in laser surface heating process with experimental data

    International Nuclear Information System (INIS)

    Chen, H-T; Wu, X-Y

    2006-01-01

    This study applies a hybrid technique of the Laplace transform and finite-difference methods in conjunction with the least-squares method and experimental temperature data inside the test material to predict the unknown surface temperature, heat flux and absorptivity for various surface conditions in the laser surface heating process. In this study, the functional form of the surface temperature is unknown a priori and is assumed to be a function of time before performing the inverse calculation. In addition, the whole time domain is divided into several analysis sub-time intervals and then these unknown estimates on each analysis interval can be predicted. In order to show the accuracy of the present inverse method, comparisons are made among the present estimates, direct results and previous results, showing that the present estimates agree with the direct results for the simulated problem. However, the present estimates of the surface absorptivity deviate slightly from previous estimated results under the assumption of constant thermal properties. The effect of the surface conditions on the surface absorptivity and temperature is not negligible

  9. Surface Modification of α-Fe Metal Particles by Chemical Surface Coating

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The structure of α-Fe metal magnetic recording particles coated with silane coupling agents have been studied by TEM, FT-IR, EXAFS, Mossbauer. The results show that a close, uniform, firm and ultra thin layer, which is beneficial to the magnetic and chemical stability, has been formed by the cross-linked chemical bond Si-O-Si. And the organic molecule has chemically bonded to the particle surface, which has greatly affected the surface Fe atom electronic structure. Furthermore, the covalent bond between metal particle surface and organic molecule has obvious effect on the near edge structure of the surface Fe atoms.

  10. Couple of biomimetic surfaces with different morphologies for remanufacturing nonuniform wear rail surface

    Science.gov (United States)

    Sui, Qi; Zhou, Hong; Yang, Lin; Zhang, Haifeng; Feng, Li; Zhang, Peng

    2018-02-01

    In this work, biomimetic laser treatment was performed on repairing and remanufacturing the nonuniform worn rail surface. The wearing depth distribution of three work regions of a failure rail surface was discussed, and different thickness hardening layers with different microstructure, microhardness and wear resistances were detected from the worm surfaces. Varying wear resistances of the surfaces with different biomimetic morphologies were obtained by biomimetic laser treatments, and the corresponding effect on the lubrication sliding wear of treated and untreated surfaces were studied for comparative study. In addition, the relationship between wear resistance and the spacing of units was also provided, which can lay the important theoretical foundation for avoiding the wear resistance of the serious worn surface is less than that of the slight worn surface in the future practical applications.

  11. Thermodynamics and structure of liquid surfaces investigated directly with surface analytical tools

    Energy Technology Data Exchange (ETDEWEB)

    Andersson, Gunther [Flinders Univ., Adelaide, SA (Australia). Centre for NanoScale Science and Technology; Morgner, Harald [Leipzig Univ. (Germany). Wilhelm Ostwald Inst. for Physical and Theoretical Chemistry

    2017-06-15

    Measuring directly the composition, the distribution of constituents as function of the depth and the orientation of molecules at liquid surfaces is essential for determining physicochemical properties of liquid surfaces. While the experimental tools that have been developed for analyzing solid surfaces can in principal be applied to liquid surfaces, it turned out that they had to be adjusted to the particular challenges imposed by liquid samples, e.g. by the unavoidable vapor pressure and by the mobility of the constituting atoms/molecules. In the present work it is shown, how electron spectroscopy and ion scattering spectroscopy have been used for analyzing liquid surfaces. The emphasis of this review is on using the structural information gained for determining the physicochemical properties of liquid surfaces. (copyright 2017 by WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  12. Low surface brightness galaxies in the Fornax Cluster: automated galaxy surface photometry

    International Nuclear Information System (INIS)

    Davies, J.I.; Phillipps, S.; Disney, M.J.

    1988-01-01

    A sample is presented of low surface brightness galaxies (with extrapolated central surface brightness fainter than 22.0 Bμ) in the Fornax Cluster region which has been measured by the APM machine. Photometric parameters, namely profile shape, scale length, central brightness and total magnitude, are derived for the sample galaxies and correlations between the parameters of low surface brightness dwarf galaxies are discussed, with particular reference to the selection limits. Contrary to previous authors we find no evidence for a luminosity-surface brightness correlation in the sense of lower surface brightness galaxies having lower luminosities and scale sizes. In fact, the present data suggest that it is the galaxies with the largest scale lengths which are more likely to be of very low surface brightness. In addition, the larger scale length galaxies occur preferentially towards the centre of the Cluster. (author)

  13. Structure of stable degeneration of K3 surfaces into pairs of rational elliptic surfaces

    Science.gov (United States)

    Kimura, Yusuke

    2018-03-01

    F-theory/heterotic duality is formulated in the stable degeneration limit of a K3 fibration on the F-theory side. In this note, we analyze the structure of the stable degeneration limit. We discuss whether stable degeneration exists for pairs of rational elliptic surfaces. We demonstrate that, when two rational elliptic surfaces have an identical complex structure, stable degeneration always exists. We provide an equation that systematically describes the stable degeneration of a K3 surface into a pair of isomorphic rational elliptic surfaces. When two rational elliptic surfaces have different complex structures, whether their sum glued along a smooth fiber admits deformation to a K3 surface can be determined by studying the structure of the K3 lattice. We investigate the lattice theoretic condition to determine whether a deformation to a K3 surface exists for pairs of extremal rational elliptic surfaces. In addition, we discuss the configurations of singular fibers under stable degeneration. The sum of two isomorphic rational elliptic surfaces glued together admits a deformation to a K3 surface, the singular fibers of which are twice that of the rational elliptic surface. For special situations, singular fibers of the resulting K3 surface collide and they are enhanced to a fiber of another type. Some K3 surfaces become attractive in these situations. We determine the complex structures and the Weierstrass forms of these attractive K3 surfaces. We also deduce the gauge groups in F-theory compactifications on these attractive K3 surfaces times a K3. E 6, E 7, E 8, SU(5), and SO(10) gauge groups arise in these compactifications.

  14. In-situ surface hardening of cast iron by surface layer metallurgy

    International Nuclear Information System (INIS)

    Fischer, Sebastian F.; Muschna, Stefan; Bührig-Polaczek, Andreas; Bünck, Matthias

    2014-01-01

    Abrasive wear is a serious problem in many cast iron castings used in industry. To minimize failure and repair of these components, different strategies exist to improve their surface microhardness thus enhancing their wear resistance. However, most of these methods lead to very brittle and/or expensive castings. In the current work a new method for surface hardening is presented which utilizes surface layer metallurgy to generate in-situ a boron-enriched white cast iron surface layer with a high microhardness on a gray cast iron casting. To do this, sand molds are coated with a ferroboron suspension and cast with a cast iron melt. After solidification, a 100–900 µm thick layer of boron-enriched ledeburite is formed on the surface of the casting which produces an increase in the average microhardness from 284 HV 0.1 ±52 HV 0.1 to 505 HV 0.1 ±87 HV 0.1 . Analyses of the samples' core reveal a typical cast iron microstructure which leads to the conclusion that the coating mainly affects the castings' surface. By varying the grain size of the ferroboron powder in the coatings, it is shown that a powder size ≤100 µm is most suitable to create a boron-enriched ledeburite surface layer possessing high hardness values

  15. Radioactive surface contamination monitors

    International Nuclear Information System (INIS)

    Aoyama, Kei; Minagoshi, Atsushi; Hasegawa, Toru

    1994-01-01

    To reduce radiation exposure and prevent contamination from spreading, each nuclear power plant has established a radiation controlled area. People and articles out of the controlled area are checked for the surface contamination of radioactive materials with surface contamination monitors. Fuji Electric has repeatedly improved these monitors on the basis of user's needs. This paper outlines typical of a surface contamination monitor, a personal surface contamination monitor, an article surface contamination monitor and a laundry monitor, and the whole-body counter of an internal contamination monitor. (author)

  16. Anodized dental implant surface

    Directory of Open Access Journals (Sweden)

    Sunil Kumar Mishra

    2017-01-01

    Full Text Available Purpose: Anodized implants with moderately rough surface were introduced around 2000. Whether these implants enhanced biologic effect to improve the environment for better osseointegration was unclear. The purpose of this article was to review the literature available on anodized surface in terms of their clinical success rate and bone response in patients till now. Materials and Methods: A broad electronic search of MEDLINE and PubMed databases was performed. A focus was made on peer-reviewed dental journals. Only articles related to anodized implants were included. Both animal and human studies were included. Results: The initial search of articles resulted in 581 articles on anodized implants. The initial screening of titles and abstracts resulted in 112 full-text papers; 40 animal studies, 16 studies on cell adhesion and bacterial adhesion onto anodized surfaced implants, and 47 human studies were included. Nine studies, which do not fulfill the inclusion criteria, were excluded. Conclusions: The long-term studies on anodized surface implants do favor the surface, but in most of the studies, anodized surface is compared with that of machined surface, but not with other surfaces commercially available. Anodized surface in terms of clinical success rate in cases of compromised bone and immediately extracted sockets has shown favorable success.

  17. Antibacterial Au nanostructured surfaces

    Science.gov (United States)

    Wu, Songmei; Zuber, Flavia; Brugger, Juergen; Maniura-Weber, Katharina; Ren, Qun

    2016-01-01

    We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all the Au nanostructures, regardless their shapes, exhibited similar excellent antibacterial properties. A comparison of live cells attached to nanotopographic surfaces showed that the number of live S. aureus cells was flat and rough reference surfaces. Our micro/nanofabrication process is a scalable approach based on cost-efficient self-organization and provides potential for further developing functional surfaces to study the behavior of microbes on nanoscale topographies.We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all

  18. Bacterial resistance of self-assembled surfaces using PPOm-b-PSBMAn zwitterionic copolymer - concomitant effects of surface topography and surface chemistry on attachment of live bacteria.

    Science.gov (United States)

    Hsiao, Sheng-Wen; Venault, Antoine; Yang, Hui-Shan; Chang, Yung

    2014-06-01

    Three well-defined diblock copolymers made of poly(sulfobetaine methacrylate) (poly(SBMA)) and poly(propylene oxide) (PPO) groups were synthesized by atom transfer radical polymerization (ATRP) method. They were physically adsorbed onto three types of surfaces having different topography, including smooth flat surface, convex surface, and indented surface. Chemical state of surfaces was characterized by XPS while the various topographies were examined by SEM and AFM. Hydrophilicity of surfaces was dependent on both the surface chemistry and the surface topography, suggesting that orientation of copolymer brushes can be tuned in the design of surfaces aimed at resisting bacterial attachment. Escherichia coli, Staphylococcus epidermidis, Streptococcus mutans and Escherichia coli with green fluorescent protein (E. coli GFP) were used in bacterial tests to assess the resistance to bacterial attachment of poly(SBMA)-covered surfaces. Results highlighted a drastic improvement of resistance to bacterial adhesion with the increasing of poly(SBMA) to PPO ratio, as well as an important effect of surface topography. The chemical effect was directly related to the length of the hydrophilic moieties. When longer, more water could be entrapped, leading to improved anti-bacterial properties. The physical effect impacted on the orientation of the copolymer brushes, as well as on the surface contact area available. Convex surfaces as well as indented surfaces wafer presented the best resistance to bacterial adhesion. Indeed, bacterial attachment was more importantly reduced on these surfaces compared with smooth surfaces. It was explained by the non-orthogonal orientation of copolymer brushes, resulting in a more efficient surface coverage of zwitterionic molecules. This work suggests that not only the control of surface chemistry is essential in the preparation of surfaces resisting bacterial attachment, but also the control of surface topography and orientation of antifouling

  19. Inclusion of Solar Elevation Angle in Land Surface Albedo Parameterization Over Bare Soil Surface.

    Science.gov (United States)

    Zheng, Zhiyuan; Wei, Zhigang; Wen, Zhiping; Dong, Wenjie; Li, Zhenchao; Wen, Xiaohang; Zhu, Xian; Ji, Dong; Chen, Chen; Yan, Dongdong

    2017-12-01

    Land surface albedo is a significant parameter for maintaining a balance in surface energy. It is also an important parameter of bare soil surface albedo for developing land surface process models that accurately reflect diurnal variation characteristics and the mechanism behind the solar spectral radiation albedo on bare soil surfaces and for understanding the relationships between climate factors and spectral radiation albedo. Using a data set of field observations, we conducted experiments to analyze the variation characteristics of land surface solar spectral radiation and the corresponding albedo over a typical Gobi bare soil underlying surface and to investigate the relationships between the land surface solar spectral radiation albedo, solar elevation angle, and soil moisture. Based on both solar elevation angle and soil moisture measurements simultaneously, we propose a new two-factor parameterization scheme for spectral radiation albedo over bare soil underlying surfaces. The results of numerical simulation experiments show that the new parameterization scheme can more accurately depict the diurnal variation characteristics of bare soil surface albedo than the previous schemes. Solar elevation angle is one of the most important factors for parameterizing bare soil surface albedo and must be considered in the parameterization scheme, especially in arid and semiarid areas with low soil moisture content. This study reveals the characteristics and mechanism of the diurnal variation of bare soil surface solar spectral radiation albedo and is helpful in developing land surface process models, weather models, and climate models.

  20. Iron oxide surfaces

    Science.gov (United States)

    Parkinson, Gareth S.

    2016-03-01

    The current status of knowledge regarding the surfaces of the iron oxides, magnetite (Fe3O4), maghemite (γ-Fe2O3), haematite (α-Fe2O3), and wüstite (Fe1-xO) is reviewed. The paper starts with a summary of applications where iron oxide surfaces play a major role, including corrosion, catalysis, spintronics, magnetic nanoparticles (MNPs), biomedicine, photoelectrochemical water splitting and groundwater remediation. The bulk structure and properties are then briefly presented; each compound is based on a close-packed anion lattice, with a different distribution and oxidation state of the Fe cations in interstitial sites. The bulk defect chemistry is dominated by cation vacancies and interstitials (not oxygen vacancies) and this provides the context to understand iron oxide surfaces, which represent the front line in reduction and oxidation processes. Fe diffuses in and out from the bulk in response to the O2 chemical potential, forming sometimes complex intermediate phases at the surface. For example, α-Fe2O3 adopts Fe3O4-like surfaces in reducing conditions, and Fe3O4 adopts Fe1-xO-like structures in further reducing conditions still. It is argued that known bulk defect structures are an excellent starting point in building models for iron oxide surfaces. The atomic-scale structure of the low-index surfaces of iron oxides is the major focus of this review. Fe3O4 is the most studied iron oxide in surface science, primarily because its stability range corresponds nicely to the ultra-high vacuum environment. It is also an electrical conductor, which makes it straightforward to study with the most commonly used surface science methods such as photoemission spectroscopies (XPS, UPS) and scanning tunneling microscopy (STM). The impact of the surfaces on the measurement of bulk properties such as magnetism, the Verwey transition and the (predicted) half-metallicity is discussed. The best understood iron oxide surface at present is probably Fe3O4(100); the structure is

  1. Mostly surfaces

    CERN Document Server

    Schwartz, Richard Evan

    2011-01-01

    This book presents a number of topics related to surfaces, such as Euclidean, spherical and hyperbolic geometry, the fundamental group, universal covering surfaces, Riemannian manifolds, the Gauss-Bonnet Theorem, and the Riemann mapping theorem. The main idea is to get to some interesting mathematics without too much formality. The book also includes some material only tangentially related to surfaces, such as the Cauchy Rigidity Theorem, the Dehn Dissection Theorem, and the Banach-Tarski Theorem. The goal of the book is to present a tapestry of ideas from various areas of mathematics in a clear and rigorous yet informal and friendly way. Prerequisites include undergraduate courses in real analysis and in linear algebra, and some knowledge of complex analysis.

  2. Surface mobilities on solid materials

    International Nuclear Information System (INIS)

    Binh, V.T.

    1983-01-01

    This book constitutes the proceedings of the NATO Advanced Study Institute on Surface Mobilities on Solid Materials held in France in 1981. The goal of the two-week meeting was to review up-to-date knowledge on surface diffusion, both theoretical and experimental, and to highlight those areas in which much more knowledge needs to be accumulated. Topics include theoretical aspects of surface diffusion (e.g., microscopic theories of D at zero coverage; statistical mechanical models and surface diffusion); surface diffusion at the atomic level (e.g., FIM studies of surface migration of single adatoms and diatomic clusters; field emission studies of surface diffusion of adsorbates); foreign adsorbate mass transport; self-diffusion mass transport (e.g., different driving forces for the matter transport along surfaces; measurements of the morphological evolution of tips); the role of surface diffusion in some fundamental and applied sciences (e.g. adatomadatom pair interactions and adlayer superstructure formation; surface mobility in chemical reactions and catalysis); and recent works on surface diffusion (e.g., preliminary results on surface self-diffusion measurements on nickel and chromium tips)

  3. Effect of surface parameter of interband surface mode frequencies of finite diatomic chain

    International Nuclear Information System (INIS)

    Puszkarski, H.

    1982-07-01

    The surface modes of a finite diatomic chain of alternating atoms (M 1 not= M 2 ) are investigated. The surface force constants are assumed to differ from the bulk ones, with the resulting surface parameter a-tilde identical on both ends of the chain. Criteria, governing the existence of interband surface (IBS) modes with frequencies lying in the forbidden gap between acoustical and optical bulk bands for natural (a = 1) as well as non-natural (a not= 1) surface defect, are analysed by the difference equation method. It is found that the IBS modes localize, depending on the value of the surface parameter a, either at the surface of lighter atoms (if a-tilde is positive), or at that of heavier atoms (if a-tilde is negative). Two, one of no IBS modes are found to exist in the chain depending on the relation between the mass ratio and surface parameter - quantities on which the surface localization increment t-tilde depends. If two modes are present (one acoustical and the other optical), their frequencies are disposed symmetrically with respect to the middle of the forbidden gap, provided the surface defect is natural, or asymmetrically - if it is other than natural. If the localization of the IBS mode exceeds a well defined critical value tsub(c), the mode frequency becomes complex, indicating that the mode undergoes a damping. A comparison of the present results and those obtained by Wallis for the diatomic chain with natural surface defect is also given. (author)

  4. Surface-stabilized gold nanocatalysts

    Science.gov (United States)

    Dai, Sheng [Knoxville, TN; Yan, Wenfu [Oak Ridge, TN

    2009-12-08

    A surface-stabilized gold nanocatalyst includes a solid support having stabilizing surfaces for supporting gold nanoparticles, and a plurality of gold nanoparticles having an average particle size of less than 8 nm disposed on the stabilizing surfaces. The surface-stabilized gold nanocatalyst provides enhanced stability, such as at high temperature under oxygen containing environments. In one embodiment, the solid support is a multi-layer support comprising at least a first layer having a second layer providing the stabilizing surfaces disposed thereon, the first and second layer being chemically distinct.

  5. Nano- and Micro-Scale Oxidative Patterning of Titanium Implant Surfaces for Improved Surface Wettability.

    Science.gov (United States)

    Kim, In-hye; Son, Jun Sik; Choi, Seok Hwa; Kim, Kyo-han; Kwon, Tae-yub

    2016-02-01

    A simple and scalable surface modification treatment is demonstrated, in which nano- and microscale features are introduced into the surface of titanium (Ti) substrates by means of a novel and eco-friendly oxidative aqueous solution composed of hydrogen peroxide (H202) and sodium bicarbonate (NaHCO3). By immersing mirror-polished Ti discs in an aqueous mixture of 30 wt% H2O2/5 wt% NaHCO3 at 23 +/- 3 degrees C for 4 h, it was confirmed that this mixture is capable of generating microscale topographies on Ti surfaces. It also simultaneously formed nanochannels that were regularly arranged in a comb-like pattern on the Ti surface, thus forming a hierarchical surface structure. Further, these nano/micro-textured Ti surfaces showed great surface roughness and excellent wettability when compared with control Ti surfaces. This study demonstrates that a H2O2/NaHCO3 mixture can be effectively utilized to create reproducible nano/microscale topographies on Ti implant surfaces, thus providing an economical new oxidative solution that may be used effectively and safely as a Ti surface modification treatment.

  6. Modification of Surface Energy via Direct Laser Ablative Surface Patterning

    Science.gov (United States)

    Wohl, Christopher J., Jr. (Inventor); Belcher, Marcus A. (Inventor); Connell, John W. (Inventor); Hopkins, John W. (Inventor)

    2015-01-01

    Surface energy of a substrate is changed without the need for any template, mask, or additional coating medium applied to the substrate. At least one beam of energy directly ablates a substrate surface to form a predefined topographical pattern at the surface. Each beam of energy has a width of approximately 25 micrometers and an energy of approximately 1-500 microJoules. Features in the topographical pattern have a width of approximately 1-500 micrometers and a height of approximately 1.4-100 micrometers.

  7. Bio-Inspired Functional Surfaces Based on Laser-Induced Periodic Surface Structures.

    Science.gov (United States)

    Müller, Frank A; Kunz, Clemens; Gräf, Stephan

    2016-06-15

    Nature developed numerous solutions to solve various technical problems related to material surfaces by combining the physico-chemical properties of a material with periodically aligned micro/nanostructures in a sophisticated manner. The utilization of ultra-short pulsed lasers allows mimicking numerous of these features by generating laser-induced periodic surface structures (LIPSS). In this review paper, we describe the physical background of LIPSS generation as well as the physical principles of surface related phenomena like wettability, reflectivity, and friction. Then we introduce several biological examples including e.g., lotus leafs, springtails, dessert beetles, moth eyes, butterfly wings, weevils, sharks, pangolins, and snakes to illustrate how nature solves technical problems, and we give a comprehensive overview of recent achievements related to the utilization of LIPSS to generate superhydrophobic, anti-reflective, colored, and drag resistant surfaces. Finally, we conclude with some future developments and perspectives related to forthcoming applications of LIPSS-based surfaces.

  8. Site-directed immobilization of a genetically engineered anti-methotrexate antibody via an enzymatically introduced biotin label significantly increases the binding capacity of immunoaffinity columns.

    Science.gov (United States)

    Davenport, Kaitlynn R; Smith, Christopher A; Hofstetter, Heike; Horn, James R; Hofstetter, Oliver

    2016-05-15

    In this study, the effect of random vs. site-directed immobilization techniques on the performance of antibody-based HPLC columns was investigated using a single-domain camelid antibody (VHH) directed against methotrexate (MTX) as a model system. First, the high flow-through support material POROS-OH was activated with disuccinimidyl carbonate (DSC), and the VHH was bound in a random manner via amines located on the protein's surface. The resulting column was characterized by Frontal Affinity Chromatography (FAC). Then, two site-directed techniques were explored to increase column efficiency by immobilizing the antibody via its C-terminus, i.e., away from the antigen-binding site. In one approach, a tetra-lysine tail was added, and the antibody was immobilized onto DSC-activated POROS. In the second site-directed approach, the VHH was modified with the AviTag peptide, and a biotin-residue was enzymatically incorporated at the C-terminus using the biotin ligase BirA. The biotinylated antibody was subsequently immobilized onto NeutrAvidin-derivatized POROS. A comparison of the FAC analyses, which for all three columns showed excellent linearity (R(2)>0.999), revealed that both site-directed approaches yield better results than the random immobilization; the by far highest efficiency, however, was determined for the immunoaffinity column based on AviTag-biotinylated antibody. As proof of concept, all three columns were evaluated for quantification of MTX dissolved in phosphate buffered saline (PBS). Validation using UV-detection showed excellent linearity in the range of 0.04-12μM (R(2)>0.993). The lower limit of detection (LOD) and lower limit of quantification (LLOQ) were found to be independent of the immobilization strategy and were 40nM and 132nM, respectively. The intra- and inter-day precision was below 11.6%, and accuracy was between 90.7% and 112%. To the best of our knowledge, this is the first report of the AviTag-system in chromatography, and the first

  9. Cryogenic Selective Surfaces

    Data.gov (United States)

    National Aeronautics and Space Administration — Selective surfaces have wavelength dependent emissivity/absorption. These surfaces can be designed to reflect solar radiation, while maximizing infrared emittance,...

  10. Lubrication of oral surfaces by food emulsions: the importance of surface characteristics

    NARCIS (Netherlands)

    Hoog, de E.H.A.; Prinz, J.F.; Huntjens, L.; Dresselhuis, D.M.; Aken, van G.A.

    2006-01-01

    The friction between surfaces in relative motion lubricated by food emulsions has been measured. Different types of surfaces were tested, including metal, glass, rubber, and mucosal surfaces (pig tongue and pig esophagus). We demonstrate that the load-dependent behavior of the coefficient of kinetic

  11. Lubrication of oral surfaces by food emulsions: The importance of surface characteristics

    NARCIS (Netherlands)

    Hoog, E.H.A. de; Prinz, J.F.; Huntjens, L.; Dresselhuis, D.M.; Aken, G.A. van

    2006-01-01

    The friction between surfaces in relative motion lubricated by food emulsions has been measured. Different types of surfaces were tested, including metal, glass, rubber, and mucosal surfaces (pig tongue and pig esophagus). We demonstrate that the load-dependent behavior of the coefficient of kinetic

  12. Ursodeoxycholic acid attenuates colonic epithelial secretory function

    Science.gov (United States)

    Kelly, Orlaith B; Mroz, Magdalena S; Ward, Joseph B J; Colliva, Carolina; Scharl, Michael; Pellicciari, Roberto; Gilmer, John F; Fallon, Padraic G; Hofmann, Alan F; Roda, Aldo; Murray, Frank E; Keely, Stephen J

    2013-01-01

    Dihydroxy bile acids, such as chenodeoxycholic acid (CDCA), are well known to promote colonic fluid and electrolyte secretion, thereby causing diarrhoea associated with bile acid malabsorption. However, CDCA is rapidly metabolised by colonic bacteria to ursodeoxycholic acid (UDCA), the effects of which on epithelial transport are poorly characterised. Here, we investigated the role of UDCA in the regulation of colonic epithelial secretion. Cl− secretion was measured across voltage-clamped monolayers of T84 cells and muscle-stripped sections of mouse or human colon. Cell surface biotinylation was used to assess abundance/surface expression of transport proteins. Acute (15 min) treatment of T84 cells with bilateral UDCA attenuated Cl− secretory responses to the Ca2+ and cAMP-dependent secretagogues carbachol (CCh) and forskolin (FSK) to 14.0 ± 3.8 and 40.2 ± 7.4% of controls, respectively (n= 18, P acid (LCA). Accordingly, LCA (50–200 μm) enhanced agonist-induced secretory responses in vitro and a metabolically stable UDCA analogue, 6α-methyl-UDCA, exerted anti-secretory actions in vitro and in vivo. In conclusion, UDCA exerts direct anti-secretory actions on colonic epithelial cells and metabolically stable derivatives of the bile acid may offer a new approach for treating intestinal diseases associated with diarrhoea. PMID:23507881

  13. Erosion of surface and near surface disposal facilities

    International Nuclear Information System (INIS)

    1988-06-01

    A literature search was undertaken to identify existing data and analytical procedures regarding the processes of gully erosion. The applicability of the available information to the problems of gully erosion potential at surface and near surface disposal sites is evaluated. It is concluded that the existing knowledge regarding gully erosion is insufficient to develop procedures to ensure the long-term stability of disposal sites. Recommendations for further research are presented. 46 refs

  14. Surface-enhanced Raman Spectroscopy of Ethephone Adsorbed on Silver Surface

    International Nuclear Information System (INIS)

    Lee, Chul Jae; Kim, Hee Jin; Karim, Mohammad Rezaul; Lee, Mu Sang

    2006-01-01

    We investigated the Surface-enhanced Raman Spectroscopy (SERS) spectrum of ethephone (2- chloroethylphosphonic acid). We observed significant signals in the ordinary Raman spectrum for solid-state ethephone as well as when it was adsorbed on a colloidal silver surface, strong vibrational signals were obtained at a very low concentration. The SERS spectra were obtained by silver colloids that were prepared by the γ - irradiation method. The influence of pH and the influence of anion (Cl - , Br - , I - ) on the adsorption orientation were investigated. Two different adsorption mechanisms were deduced, depending on the experimental conditions. The chlorine atom or the chlorine and two oxygen atoms were adsorbed on the colloidal silver surface. Among halide ions, Br - and I - were more strongly adsorbed on the colloidal silver surfaces. As a result, the adsorption of ethephone was less effective due to their steric hinderance

  15. Hot Surface Ignition

    OpenAIRE

    Tursyn, Yerbatyr; Goyal, Vikrant; Benhidjeb-Carayon, Alicia; Simmons, Richard; Meyer, Scott; Gore, Jay P.

    2015-01-01

    Undesirable hot surface ignition of flammable liquids is one of the hazards in ground and air transportation vehicles, which primarily occurs in the engine compartment. In order to evaluate the safety and sustainability of candidate replacement fuels with respect to hot surface ignition, a baseline low lead fuel (Avgas 100 LL) and four experimental unleaded aviation fuels recommended for reciprocating aviation engines were considered. In addition, hot surface ignition properties of the gas tu...

  16. Characteristics of Turbulent Airflow Deduced from Rapid Surface Thermal Fluctuations: An Infrared Surface Anemometer

    Science.gov (United States)

    Aminzadeh, Milad; Breitenstein, Daniel; Or, Dani

    2017-12-01

    The intermittent nature of turbulent airflow interacting with the surface is readily observable in fluctuations of the surface temperature resulting from the thermal imprints of eddies sweeping the surface. Rapid infrared thermography has recently been used to quantify characteristics of the near-surface turbulent airflow interacting with the evaporating surfaces. We aim to extend this technique by using single-point rapid infrared measurements to quantify properties of a turbulent flow, including surface exchange processes, with a view towards the development of an infrared surface anemometer. The parameters for the surface-eddy renewal (α and β ) are inferred from infrared measurements of a single-point on the surface of a heat plate placed in a wind tunnel with prescribed wind speeds and constant mean temperatures of the surface. Thermally-deduced parameters are in agreement with values obtained from standard three-dimensional ultrasonic anemometer measurements close to the plate surface (e.g., α = 3 and β = 1/26 (ms)^{-1} for the infrared, and α = 3 and β = 1/19 (ms)^{-1} for the sonic-anemometer measurements). The infrared-based turbulence parameters provide new insights into the role of surface temperature and buoyancy on the inherent characteristics of interacting eddies. The link between the eddy-spectrum shape parameter α and the infrared window size representing the infrared field of view is investigated. The results resemble the effect of the sampling height above the ground in sonic anemometer measurements, which enables the detection of larger eddies with higher values of α . The physical basis and tests of the proposed method support the potential for remote quantification of the near-surface momentum field, as well as scalar-flux measurements in the immediate vicinity of the surface.

  17. Impact of initial surface parameters on the final quality of laser micro-polished surfaces

    Science.gov (United States)

    Chow, Michael; Bordatchev, Evgueni V.; Knopf, George K.

    2012-03-01

    Laser micro-polishing (LμP) is a new laser-based microfabrication technology for improving surface quality during a finishing operation and for producing parts and surfaces with near-optical surface quality. The LμP process uses low power laser energy to melt a thin layer of material on the previously machined surface. The polishing effect is achieved as the molten material in the laser-material interaction zone flows from the elevated regions to the local minimum due to surface tension. This flow of molten material then forms a thin ultra-smooth layer on the top surface. The LμP is a complex thermo-dynamic process where the melting, flow and redistribution of molten material is significantly influenced by a variety of process parameters related to the laser, the travel motions and the material. The goal of this study is to analyze the impact of initial surface parameters on the final surface quality. Ball-end micromilling was used for preparing initial surface of samples from H13 tool steel that were polished using a Q-switched Nd:YAG laser. The height and width of micromilled scallops (waviness) were identified as dominant parameter affecting the quality of the LμPed surface. By adjusting process parameters, the Ra value of a surface, having a waviness period of 33 μm and a peak-to-valley value of 5.9 μm, was reduced from 499 nm to 301 nm, improving the final surface quality by 39.7%.

  18. Mirror reactor surface study

    Energy Technology Data Exchange (ETDEWEB)

    Hunt, A. L.; Damm, C. C.; Futch, A. H.; Hiskes, J. R.; Meisenheimer, R. G.; Moir, R. W.; Simonen, T. C.; Stallard, B. W.; Taylor, C. E.

    1976-09-01

    A general survey is presented of surface-related phenomena associated with the following mirror reactor elements: plasma first wall, ion sources, neutral beams, director converters, vacuum systems, and plasma diagnostics. A discussion of surface phenomena in possible abnormal reactor operation is included. Several studies which appear to merit immediate attention and which are essential to the development of mirror reactors are abstracted from the list of recommended areas for surface work. The appendix contains a discussion of the fundamentals of particle/surface interactions. The interactions surveyed are backscattering, thermal desorption, sputtering, diffusion, particle ranges in solids, and surface spectroscopic methods. A bibliography lists references in a number of categories pertinent to mirror reactors. Several complete published and unpublished reports on surface aspects of current mirror plasma experiments and reactor developments are also included.

  19. Mirror reactor surface study

    International Nuclear Information System (INIS)

    Hunt, A.L.; Damm, C.C.; Futch, A.H.; Hiskes, J.R.; Meisenheimer, R.G.; Moir, R.W.; Simonen, T.C.; Stallard, B.W.; Taylor, C.E.

    1976-01-01

    A general survey is presented of surface-related phenomena associated with the following mirror reactor elements: plasma first wall, ion sources, neutral beams, director converters, vacuum systems, and plasma diagnostics. A discussion of surface phenomena in possible abnormal reactor operation is included. Several studies which appear to merit immediate attention and which are essential to the development of mirror reactors are abstracted from the list of recommended areas for surface work. The appendix contains a discussion of the fundamentals of particle/surface interactions. The interactions surveyed are backscattering, thermal desorption, sputtering, diffusion, particle ranges in solids, and surface spectroscopic methods. A bibliography lists references in a number of categories pertinent to mirror reactors. Several complete published and unpublished reports on surface aspects of current mirror plasma experiments and reactor developments are also included

  20. Inkjet printing for direct micropatterning of a superhydrophobic surface: Toward biomimetic fog harvesting surfaces

    KAUST Repository

    Zhang, Lianbin

    2015-01-01

    The preparation of biomimetic superhydrophobic surfaces with hydrophilic micro-sized patterns is highly desirable, but a one-step, mask-free method to produce such surfaces has not previously been reported. We have developed a direct method to produce superhydrophilic micropatterns on superhydrophobic surfaces based on inkjet printing technology. This work was inspired by the efficient fog-harvesting behavior of Stenocara beetles in the Namib Desert. A mussel-inspired ink consisting of an optimized solution of dopamine was applied directly by inkjet printing to superhydrophobic surfaces. Stable Wenzel\\'s microdroplets of the dopamine solution with well-defined micropatterns were obtained on these surfaces. Superhydrophilic micropatterns with well-controlled dimensions were then readily achieved on the superhydrophobic surfaces by the formation of polydopamine via in situ polymerization. The micropatterned superhydrophobic surfaces prepared by this inkjet printing method showed enhanced water collection efficiency compared with uniform superhydrophilic and superhydrophobic surfaces. This method can be used for the facile large-scale patterning of superhydrophobic surfaces with high precision and superior pattern stability and is therefore a key step toward patterning superhydrophobic surfaces for practical applications. This journal is

  1. Surface characterization

    Science.gov (United States)

    Mandla A. Tshabalala

    2005-01-01

    Surface properties of wood play an important role when wood is used or processed into different commodities such as siding, joinery, textiles, paper, sorption media or wood composites. Thus, for example, the quality and durability of a wood coating are determined by the surface properties of the wood and the coating. The same is true for wood composites, as the...

  2. Bio-Inspired Functional Surfaces Based on Laser-Induced Periodic Surface Structures

    Directory of Open Access Journals (Sweden)

    Frank A. Müller

    2016-06-01

    Full Text Available Nature developed numerous solutions to solve various technical problems related to material surfaces by combining the physico-chemical properties of a material with periodically aligned micro/nanostructures in a sophisticated manner. The utilization of ultra-short pulsed lasers allows mimicking numerous of these features by generating laser-induced periodic surface structures (LIPSS. In this review paper, we describe the physical background of LIPSS generation as well as the physical principles of surface related phenomena like wettability, reflectivity, and friction. Then we introduce several biological examples including e.g., lotus leafs, springtails, dessert beetles, moth eyes, butterfly wings, weevils, sharks, pangolins, and snakes to illustrate how nature solves technical problems, and we give a comprehensive overview of recent achievements related to the utilization of LIPSS to generate superhydrophobic, anti-reflective, colored, and drag resistant surfaces. Finally, we conclude with some future developments and perspectives related to forthcoming applications of LIPSS-based surfaces.

  3. EDITORIAL: Molecular switches at surfaces Molecular switches at surfaces

    Science.gov (United States)

    Weinelt, Martin; von Oppen, Felix

    2012-10-01

    In nature, molecules exploit interaction with their environment to realize complex functionalities on the nanometer length scale. Physical, chemical and/or biological specificity is frequently achieved by the switching of molecules between microscopically different states. Paradigmatic examples are the energy production in proton pumps of bacteria or the signal conversion in human vision, which rely on switching molecules between different configurations or conformations by external stimuli. The remarkable reproducibility and unparalleled fatigue resistance of these natural processes makes it highly desirable to emulate nature and develop artificial systems with molecular functionalities. A promising avenue towards this goal is to anchor the molecular switches at surfaces, offering new pathways to control their functional properties, to apply electrical contacts, or to integrate switches into larger systems. Anchoring at surfaces allows one to access the full range from individual molecular switches to self-assembled monolayers of well-defined geometry and to customize the coupling between molecules and substrate or between adsorbed molecules. Progress in this field requires both synthesis and preparation of appropriate molecular systems and control over suitable external stimuli, such as light, heat, or electrical currents. To optimize switching and generate function, it is essential to unravel the geometric structure, the electronic properties and the dynamic interactions of the molecular switches on surfaces. This special section, Molecular Switches at Surfaces, collects 17 contributions describing different aspects of this research field. They analyze elementary processes, both in single molecules and in ensembles of molecules, which involve molecular switching and concomitant changes of optical, electronic, or magnetic properties. Two topical reviews summarize the current status, including both challenges and achievements in the field of molecular switches on

  4. Topographic characterization of glazed surfaces

    International Nuclear Information System (INIS)

    Froeberg, Linda; Hupa, Leena

    2008-01-01

    Detailed characterization of surface microstructure, i.e. phase composition and surface geometry, has become an important criterion of glazed ceramics. Topographic characterization is an important parameter in, e.g. estimating the influence of additional films on the average roughness of a surface. Also, the microscaled and nanoscaled roughnesses correlate with the cleanability and the self-cleaning properties of the surfaces. In this work the surface geometry of several matte glazes were described by topography and roughness as given by whitelight confocal microscopy and atomic force microscopy. Different measuring parameters were compared to justify the usefulness of the techniques in giving a comprehensive description of the surface microstructure. The results suggest that confocal microscopy is well suited for giving reliable topographical parameters for matte surfaces with microscaled crystals in the surfaces. Atomic force microscopy was better suited for smooth surfaces or for describing the local topographic parameters of closely limited areas, e.g. the surroundings of separate crystals in the surface

  5. Topographic characterization of glazed surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Froeberg, Linda [Process Chemistry Centre, Abo Akademi University, FI-20500 Turku (Finland)], E-mail: lfroberg@abo.fi; Hupa, Leena [Process Chemistry Centre, Abo Akademi University, FI-20500 Turku (Finland)

    2008-01-15

    Detailed characterization of surface microstructure, i.e. phase composition and surface geometry, has become an important criterion of glazed ceramics. Topographic characterization is an important parameter in, e.g. estimating the influence of additional films on the average roughness of a surface. Also, the microscaled and nanoscaled roughnesses correlate with the cleanability and the self-cleaning properties of the surfaces. In this work the surface geometry of several matte glazes were described by topography and roughness as given by whitelight confocal microscopy and atomic force microscopy. Different measuring parameters were compared to justify the usefulness of the techniques in giving a comprehensive description of the surface microstructure. The results suggest that confocal microscopy is well suited for giving reliable topographical parameters for matte surfaces with microscaled crystals in the surfaces. Atomic force microscopy was better suited for smooth surfaces or for describing the local topographic parameters of closely limited areas, e.g. the surroundings of separate crystals in the surface.

  6. Controlling the surface photovoltage on WSe2 by surface chemical modification

    Science.gov (United States)

    Liu, Ro-Ya; Ozawa, Kenichi; Terashima, Naoya; Natsui, Yuto; Feng, Baojie; Ito, Suguru; Chen, Wei-Chuan; Cheng, Cheng-Maw; Yamamoto, Susumu; Kato, Hiroo; Chiang, Tai-Chang; Matsuda, Iwao

    2018-05-01

    The surface photovoltage (SPV) effect is key to the development of opto-electronic devices such as solar-cells and photo-detectors. For the prototypical transition metal dichalcogenide WSe2, core level and valence band photoemission measurements show that the surface band bending of pristine cleaved surfaces can be readily modified by adsorption with K (an electron donor) or C60 (an electron acceptor). Time-resolved pump-probe photoemission measurements reveal that the SPV for pristine cleaved surfaces is enhanced by K adsorption, but suppressed by C60 adsorption, and yet the SPV relaxation time is substantially shortened in both cases. Evidently, adsorbate-induced electronic states act as electron-hole recombination centers that shorten the carrier lifetime.

  7. Effect of surface wettability caused by radiation induced surface activation on leidenfrost condition

    International Nuclear Information System (INIS)

    Takamasa, T.; Hazuku, T.; Tamura, N.; Okamoto, K.; Mishima, K.; Furuya, M.

    2003-01-01

    Improving the limit of boiling heat transfer or critical heat flux requires that the cooling liquid can contact the heating surface, or a high-wettability, highly hydrophilic heating surface, even if a vapor bubble layer is generated on the surface. From this basis, we investigated surface wettability and Leidenfrost condition using metal oxides irradiated by γ-rays. In our previous study, contact angle, an indicator of macroscopic wettability, of a water droplet on metal oxide at room temperature was measured by image processing of the images obtained by a CCD video camera. The results showed that the surface wettability on metal oxide pieces of titanium, Zircaloy No. 4, SUS-304, and copper was improved significantly by the Radiation Induced Surface Activation (RISA) phenomenon. To delineate the effect of Radiation Induced Surface Activation (RISA) on heat transferring phenomena, the Leidenfrost condition and quenching of metal oxides irradiated by γ-rays were investigated. In the Leidenfrost experiment, when the temperature of the heating surface reached the wetting limit temperature, water-solid contact vanished because a stable vapor film existed between the droplet and the metal surface; i.e., a Leidenfrost condition obtained. The wetting limit temperature increased with integrated irradiation dose. After irradiation, the wet length and the duration of contact increased, and the contact angle decreased. In the quenching test, high surface wettability, or a highly hydrophilic condition, of a simulated fuel rod made of SUS was achieved, and the quenching velocities were increased up to 20-30% after 300 kGy 60Co γ-ray irradiation

  8. Effect of surface wettability caused by radiation induced surface activation on leidenfrost condition

    Energy Technology Data Exchange (ETDEWEB)

    Takamasa, T.; Hazuku, T.; Tamura, N.; Okamoto, K. [Tokyo Univ., Tokyo (Japan); Mishima, K. [Kyoto Univ., Kyoto (Japan); Furuya, M. [Central Research Institute of Electric Power Industry, Tokyo (Japan)

    2003-07-01

    Improving the limit of boiling heat transfer or critical heat flux requires that the cooling liquid can contact the heating surface, or a high-wettability, highly hydrophilic heating surface, even if a vapor bubble layer is generated on the surface. From this basis, we investigated surface wettability and Leidenfrost condition using metal oxides irradiated by {gamma}-rays. In our previous study, contact angle, an indicator of macroscopic wettability, of a water droplet on metal oxide at room temperature was measured by image processing of the images obtained by a CCD video camera. The results showed that the surface wettability on metal oxide pieces of titanium, Zircaloy No. 4, SUS-304, and copper was improved significantly by the Radiation Induced Surface Activation (RISA) phenomenon. To delineate the effect of Radiation Induced Surface Activation (RISA) on heat transferring phenomena, the Leidenfrost condition and quenching of metal oxides irradiated by {gamma}-rays were investigated. In the Leidenfrost experiment, when the temperature of the heating surface reached the wetting limit temperature, water-solid contact vanished because a stable vapor film existed between the droplet and the metal surface; i.e., a Leidenfrost condition obtained. The wetting limit temperature increased with integrated irradiation dose. After irradiation, the wet length and the duration of contact increased, and the contact angle decreased. In the quenching test, high surface wettability, or a highly hydrophilic condition, of a simulated fuel rod made of SUS was achieved, and the quenching velocities were increased up to 20-30% after 300 kGy 60Co {gamma}-ray irradiation.

  9. Enhanced photochemistry on metal surfaces

    International Nuclear Information System (INIS)

    Goncher, G.M.; Parsons, C.A.; Harris, C.B.

    1984-01-01

    Due to the fast relaxation of molecular excited states in the vicinity of a metal or semiconductor surface, few observations of surface photochemistry have been reported. The following work concerns the surface-enhanced photo-reactions of a variety of physisorbed molecules on roughened Ag surfaces. In summary, photodecomposition leads to a graphitic surface carbon product which is monitored via surface-enhanced Raman scattering. In most cases an initial two-photon molecular absorption step followed by further absorption and fragmentation is thought to occur. Enhancement of the incident fields occurs through roughness-mediated surface plasmon resonances. This mechanism provides the amplified electromagnetic surface fields responsible for the observed photodecomposition. The photodecomposition experiments are performed under ultra-high vacuum. Surface characterization of the roughened surfaces was done by Scanning Electron Microscopy (SEM), and electron-stimulated emission. The SEM revealed morphology on the order of 300-400 A. This size of roughness feature, when modelled as isolated spheres should exhibit the well-known Mie resonances for light of the correct wavelengths. For protrusions existing on a surface these Mie resonances can be thought of as a coupling of the light with the surface plasmon. Experimental verification of these resonances was provided by the electron-stimulated light emission results. These showed that a polished Ag surface emitted only the expected transition radiation at the frequency of the Ag bulk plasmon. Upon roughening, however, a broad range of lower frequencies extending well into the visible are seen from electron irradiation of the surface. Large enhancements are expected for those frequencies which are able to couple into the surface modes

  10. Surface Prognostic Charts

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Surface Prognostic Charts are historical surface prognostic (forecast) charts created by the United States Weather Bureau. They include fronts, isobars, cloud, and...

  11. Surface structure analysis by means of Rutherford scattering: methods to study surface relaxation

    International Nuclear Information System (INIS)

    Turkenburg, W.C.; Soszka, W.; Saris, F.W.; Kersten, H.H.; Colenbrander, B.G.

    1976-01-01

    The use of Rutherford backscattering for structural analysis of single crystal surfaces is reviewed, and a new method is introduced. With this method, which makes use of the channeling and blocking phenomenon of light ions of medium energy, surface atoms can be located with a precision of 0.02 A. This is demonstrated in a measurement of surface relaxation for the Cu(110) surface. (Auth.)

  12. CYGNSS Surface Wind Observations and Surface Flux Estimates within Low-Latitude Extratropical Cyclones

    Science.gov (United States)

    Crespo, J.; Posselt, D. J.

    2017-12-01

    The Cyclone Global Navigation Satellite System (CYGNSS), launched in December 2016, aims to improve estimates of surface wind speeds over the tropical oceans. While CYGNSS's core mission is to provide better estimates of surface winds within the core of tropical cyclones, previous research has shown that the constellation, with its orbital inclination of 35°, also has the ability to observe numerous extratropical cyclones that form in the lower latitudes. Along with its high spatial and temporal resolution, CYGNSS can provide new insights into how extratropical cyclones develop and evolve, especially in the presence of thick clouds and precipitation. We will demonstrate this by presenting case studies of multiple extratropical cyclones observed by CYGNSS early on in its mission in both Northern and Southern Hemispheres. By using the improved estimates of surface wind speeds from CYGNSS, we can obtain better estimates of surface latent and sensible heat fluxes within and around extratropical cyclones. Surface heat fluxes, driven by surface winds and strong vertical gradients of water vapor and temperature, play a key role in marine cyclogenesis as they increase instability within the boundary layer and may contribute to extreme marine cyclogenesis. In the past, it has been difficult to estimate surface heat fluxes from space borne instruments, as these fluxes cannot be observed directly from space, and deficiencies in spatial coverage and attenuation from clouds and precipitation lead to inaccurate estimates of surface flux components, such as surface wind speeds. While CYGNSS only contributes estimates of surface wind speeds, we can combine this data with other reanalysis and satellite data to provide improved estimates of surface sensible and latent heat fluxes within and around extratropical cyclones and throughout the entire CYGNSS mission.

  13. The effects of sea surface temperature gradients on surface turbulent fluxes

    Science.gov (United States)

    Steffen, John

    A positive correlation between sea surface temperature (SST) and wind stress perturbation near strong SST gradients (DeltaSST) has been observed in different parts of the world ocean, such as the Gulf Stream in the North Atlantic and the Kuroshio Extension east of Japan. These changes in winds and SSTs can modify near-surface stability, surface stress, and latent and sensible heat fluxes. In general, these small scale processes are poorly modeled in Numerical Weather Prediction (NWP) and climate models. Failure to account for these air--sea interactions produces inaccurate values of turbulent fluxes, and therefore a misrepresentation of the energy, moisture, and momentum budgets. Our goal is to determine the change in these surface turbulent fluxes due to overlooking the correlated variability in winds, SSTs, and related variables. To model these air--sea interactions, a flux model was forced with and without SST--induced changes to the surface wind fields. The SST modification to the wind fields is based on a baroclinic argument as implemented by the University of Washington Planetary Boundary-Layer (UWPBL) model. Other input parameters include 2-m air temperature, 2-m dew point temperature, surface pressure (all from ERA--interim), and Reynolds Daily Optimum Interpolation Sea Surface Temperature (OISST). Flux model runs are performed every 6 hours starting in December 2002 and ending in November 2003. From these model outputs, seasonal, monthly, and daily means of the difference between DeltaSST and no DeltaSST effects on sensible heat flux (SHF), latent heat flux (LHF), and surface stress are calculated. Since the greatest impacts occur during the winter season, six additional December-January-February (DJF) seasons were analyzed for 1987--1990 and 1999--2002. The greatest differences in surface turbulent fluxes are concentrated near strong SST fronts associated with the Gulf Stream and Kuroshio Extension. On average, 2002---2003 DJF seasonal differences in SHF

  14. Antibacterial Au nanostructured surfaces.

    Science.gov (United States)

    Wu, Songmei; Zuber, Flavia; Brugger, Juergen; Maniura-Weber, Katharina; Ren, Qun

    2016-02-07

    We present here a technological platform for engineering Au nanotopographies by templated electrodeposition on antibacterial surfaces. Three different types of nanostructures were fabricated: nanopillars, nanorings and nanonuggets. The nanopillars are the basic structures and are 50 nm in diameter and 100 nm in height. Particular arrangement of the nanopillars in various geometries formed nanorings and nanonuggets. Flat surfaces, rough substrate surfaces, and various nanostructured surfaces were compared for their abilities to attach and kill bacterial cells. Methicillin-resistant Staphylococcus aureus, a Gram-positive bacterial strain responsible for many infections in health care system, was used as the model bacterial strain. It was found that all the Au nanostructures, regardless their shapes, exhibited similar excellent antibacterial properties. A comparison of live cells attached to nanotopographic surfaces showed that the number of live S. aureus cells was flat and rough reference surfaces. Our micro/nanofabrication process is a scalable approach based on cost-efficient self-organization and provides potential for further developing functional surfaces to study the behavior of microbes on nanoscale topographies.

  15. Fermi surface contours obtained from scanning tunneling microscope images around surface point defects

    International Nuclear Information System (INIS)

    Khotkevych-Sanina, N V; Kolesnichenko, Yu A; Van Ruitenbeek, J M

    2013-01-01

    We present a theoretical analysis of the standing wave patterns in scanning tunneling microscope (STM) images, which occur around surface point defects. We consider arbitrary dispersion relations for the surface states and calculate the conductance for a system containing a small-size tunnel contact and a surface impurity. We find rigorous theoretical relations between the interference patterns in the real-space STM images, their Fourier transforms and the Fermi contours of two-dimensional electrons. We propose a new method for reconstructing Fermi contours of surface electron states, directly from the real-space STM images around isolated surface defects. (paper)

  16. Wireless Metal Detection and Surface Coverage Sensing for All-Surface Induction Heating

    Directory of Open Access Journals (Sweden)

    Veli Tayfun Kilic

    2016-03-01

    Full Text Available All-surface induction heating systems, typically comprising small-area coils, face a major challenge in detecting the presence of a metallic vessel and identifying its partial surface coverage over the coils to determine which of the coils to power up. The difficulty arises due to the fact that the user can heat vessels made of a wide variety of metals (and their alloys. To address this problem, we propose and demonstrate a new wireless detection methodology that allows for detecting the presence of metallic vessels together with uniquely sensing their surface coverages while also identifying their effective material type in all-surface induction heating systems. The proposed method is based on telemetrically measuring simultaneously inductance and resistance of the induction coil coupled with the vessel in the heating system. Here, variations in the inductance and resistance values for an all-surface heating coil loaded by vessels (made of stainless steel and aluminum at different positions were systematically investigated at different frequencies. Results show that, independent of the metal material type, unique identification of the surface coverage is possible at all freqeuncies. Additionally, using the magnitude and phase information extracted from the coupled coil impedance, unique identification of the vessel effective material is also achievable, this time independent of its surface coverage.

  17. Influence of surface roughness on the friction property of textured surface

    OpenAIRE

    Yuankai Zhou; Hua Zhu; Wenqian Zhang; Xue Zuo; Yan Li; Jianhua Yang

    2015-01-01

    In contrast with dimple textures, surface roughness is a texture at the micro-scale, essentially which will influence the load-bearing capacity of lubricant film. The numerical simulation was carried out to investigate the influence of surface roughness on friction property of textured surface. The lubricant film pressure was obtained using the method of computational fluid dynamics according to geometric model of round dimple, and the renormalization-group k–ε turbulent model was adopted in ...

  18. Friction Surface Treatment Selection: Aggregate Properties, Surface Characteristics, Alternative Treatments, and Safety Effects

    Science.gov (United States)

    2017-07-01

    This study aimed to evaluate the long term performance of the selected surface friction treatments, including high friction surface treatment (HFST) using calcined bauxite and steel slag, and conventional friction surfacing, in particular pavement pr...

  19. Use of structured surfaces for friction and wear control on bearing surfaces

    International Nuclear Information System (INIS)

    Wang, Ling

    2014-01-01

    Surface texturing with purposely made regular micropatterns on flat or curved surfaces, as opposed to random roughness inherited from machining processes, has attracted significant attention in recent years. At the 2013 World Tribology Congress in Turin alone there were over 40 presentations related to surface texturing for tribological applications, from magnetic hard discs and hydrodynamic bearings to artificial joints. Although surface texturing has been reported being successfully applied in industrial applications such as seals, pistons, and thrust pad bearings, the demand for robust design is still high. Etsion has recently reviewed the modeling research mainly conducted by his group Etsion I (2013 Friction 1 195–209). This paper aims to review the state-of-the-art development of surface texturing made by a wider range of researchers. (topical review)

  20. Simultaneous determination of CRP and D-dimer in human blood plasma samples with White Light Reflectance Spectroscopy.

    Science.gov (United States)

    Koukouvinos, Georgios; Petrou, Panagiota; Misiakos, Konstantinos; Drygiannakis, Dimitris; Raptis, Ioannis; Stefanitsis, Gerasimos; Martini, Spyridoula; Nikita, Dimitra; Goustouridis, Dimitrios; Moser, Isabella; Jobst, Gerhard; Kakabakos, Sotirios

    2016-10-15

    A dual-analyte assay for the simultaneous determination of C-reactive protein (CRP) and D-dimer in human blood plasma based on a white light interference spectroscopy sensing platform is presented. Measurement is accomplished in real-time by scanning the sensing surface, on which distinct antibody areas have been created, with a reflection probe used both for illumination of the surface and collection of the reflected interference spectrum. The composition of the transducer, the sensing surface chemical activation and biofunctionalization procedures were optimized with respect to signal magnitude and repeatability. The assay format involved direct detection of CRP whereas for D-dimer a two-site immunoassay employing a biotinylated reporter antibody and reaction with streptavidin was selected. The assays were sensitive with detection limits of 25ng/mL for both analytes, precise with intra- and inter-assay CV values ranging from 3.6% to 7.7%, and from 4.8% to 9.5%, respectively, for both assays, and accurate with recovery values ranging from 88.5% to 108% for both analytes. Moreover, the values determined for the two analytes in 35 human plasma samples were in excellent agreement with those received for the same samples by standard diagnostic laboratory instrumentation employing commercial kits. The excellent agreement of the results supported the validity of the proposed system for clinical application for the detection of multiple analytes since it was demonstrated that up to seven antibody areas can be created on the sensing surface and successfully interrogated with the developed optical set-up. Copyright © 2015. Published by Elsevier B.V.

  1. Surface States and Effective Surface Area on Photoluminescent P-Type Porous Silicon

    Science.gov (United States)

    Weisz, S. Z.; Porras, A. Ramirez; Resto, O.; Goldstein, Y.; Many, A.; Savir, E.

    1997-01-01

    The present study is motivated by the possibility of utilizing porous silicon for spectral sensors. Pulse measurements on the porous-Si/electrolyte system are employed to determine the surface effective area and the surface-state density at various stages of the anodization process used to produce the porous material. Such measurements were combined with studies of the photoluminescence spectra. These spectra were found to shift progressively to the blue as a function of anodization time. The luminescence intensity increases initially with anodization time, reaches a maximum and then decreases with further anodization. The surface state density, on the other hand, increases with anodization time from an initial value of about 2 x 10(exp 12)/sq cm surface to about 1013 sq cm for the anodized surface. This value is attained already after -2 min anodization and upon further anodization remains fairly constant. In parallel, the effective surface area increases by a factor of 10-30. This behavior is markedly different from the one observed previously for n-type porous Si.

  2. Surface treatment of zirconia ceramics

    International Nuclear Information System (INIS)

    1980-01-01

    A method of chemically micropitting and/or microcratering at least a portion of a smooth surface of an impervious zirconia-base ceramic is described, comprising (a) contacting the smooth surface with a liquid leachant selected from concentrated sulphuric acid, ammonium bisulphate, alkali metal bisulphates and mixtures thereof at a temperature of at least 250 0 C for a period of time sufficient to effect micropitting and/or microcratering generally uniformly distributed throughout the microstructure of the resultant leached surface; (b) removing the leached surface from contact with the leachant; (c) contacting the leached surface with hydrochloric acid to effect removal from the leached surface of a residue thereon comprising sulphate of metal elements including zirconium in the ceramic; (d) removing the leached surface from contact with the hydrochloric acid; and (e) rinsing the leached surface with water to effect removal of acid residue from that surface. (author)

  3. Fabrication of surface micro- and nanostructures for superhydrophobic surfaces in electric and electronic applications

    Science.gov (United States)

    Xiu, Yonghao

    In our study, the superhydrophobic surface based on biomimetic lotus leave is explored to maintain the desired properties for self-cleaning. Parameters in controlling bead-up and roll-off characteristics of water droplets were investigated on different model surfaces. The governing equations were proposed. Heuristic study is performed. First, the fundamental understanding of the effect of roughness on superhydrophobicity is performed. The effect of hierarchical roughness, i.e., two scale roughness effect on roughness is investigated using systems of (1) monodisperse colloidal silica sphere (submicron) arrays and Au nanoparticle on top and (2) Si micrometer pyramids and Si nanostructures on top from KOH etching and metal assisted etching of Si. The relation between the contact area fraction and water droplet contact angles are derived based on Wenzel and Cassie-Baxter equation for the systems and the two scale effect is explained regarding the synergistic combination of two scales. Previously the microscopic three-phase-contact line is thought to be the key factor in determining contact angles and hystereses. In our study, Laplace pressure was brought up and related to the three-phase-contact line and taken as a key figure of merit in determining superhydrophobicity. In addition, we are one of the first to study the effect of tapered structures (wall inclination). Combining with a second scale roughness on the tapered structures, stable Cassie state for both water and low surface energy oil may be achieved. This is of great significance for designing both superhydrophobicity and superoleophobicity. Regarding the origin of contact angle hysteresis, study of superhydrophobicity on micrometer Si pillars was performed. The relation between the interface work of function and contact angle hysteresis was proposed and derived mathematically based on the Young-Dupre equation. The three-phase-contact line was further related to a secondary scale roughness induced. Based on

  4. Direct measurement of Cu surface self-diffusion on a checked surface

    International Nuclear Information System (INIS)

    Cousty, Jacques; Peix, Roger; Perraillon, Bernard.

    1976-01-01

    A radiotracer technique ( 64 Cu) was developed to measure surface diffusion on copper surfaces of total impurity concentration not exceeding some 10 -3 monolayers. The apparatus used consists of a slow electron diffraction device, an Auger analysis spectrometer (CMA), an ion gun and an evaporation device assembled in an ultra-vacuum chamber holding a residual pressure below 10 -10 Torr. A sample handler enables the surface studied to be positioned in front of each of these instruments. During the diffusion treatment the chemical composition of the surface is checked intermittently, and afterwards the spread of the deposit is measured outside the ultravacuum chamber. Slices several microns thick are removed and dissolved separately in dishes containing HNO 3 . The activity is then measured with a flow counter [fr

  5. Decontamination of floor surfaces

    International Nuclear Information System (INIS)

    Smirous, F.

    1983-01-01

    Requirements are presented put on the surfaces of floors of radiochemical workplaces. The mechanism is described of retaining the contaminant in the surface of the flooring, ways of reducing the hazards of floor surface contamination, decontamination techniques and used decontamination agents. (J.P.)

  6. Surface freezing of water

    OpenAIRE

    P?rez-D?az, J. L.; ?lvarez-Valenzuela, M. A.; Rodr?guez-Celis, F.

    2016-01-01

    Freezing, melting, evaporation and condensation of water are essential ingredients for climate and eventually life on Earth. In the present work, we show how surface freezing of supercooled water in an open container is conditioned and triggered?exclusively?by humidity in air. Additionally, a change of phase is demonstrated to be triggered on the water surface forming surface ice crystals prior to freezing of bulk. The symmetry of the surface crystal, as well as the freezing point, depend on ...

  7. Surfaces: processing, coating, decontamination, pollution, etc. Surface mastering to prevent component corrosion; Surfaces: traitement, revetements, decontamination, pollution, etc. Maitrise de la surface pour prevenir la corrosion des composants

    Energy Technology Data Exchange (ETDEWEB)

    Foucault, M. [Departement Corrosion Chimie, AREVA Centre Technique, BP 181, 71205 Le Creusot (France)

    2012-07-01

    In the primary and secondary circuits of nuclear Pressurized Water Reactors, AREVA uses several nickel-based alloys or austenitic stainless steels for the manufacture of safety components. The experience feedback of the last twenty years allows us to point out the major role hold by the component surface state in their life duration. In this paper, we present four examples of problem encountered and solved by a surface study and the definition and implementation of processes for the surface control of the repaired components. Then, we propose some ideas about the present needs in term of analysis means to improve the surface knowledge and control of the manufactured components. (author)

  8. Ruled Laguerre minimal surfaces

    KAUST Repository

    Skopenkov, Mikhail

    2011-10-30

    A Laguerre minimal surface is an immersed surface in ℝ 3 being an extremal of the functional ∫ (H 2/K-1)dA. In the present paper, we prove that the only ruled Laguerre minimal surfaces are up to isometry the surfaces ℝ (φλ) = (Aφ, Bφ, Cφ + D cos 2φ) + λ(sin φ, cos φ, 0), where A,B,C,D ε ℝ are fixed. To achieve invariance under Laguerre transformations, we also derive all Laguerre minimal surfaces that are enveloped by a family of cones. The methodology is based on the isotropic model of Laguerre geometry. In this model a Laguerre minimal surface enveloped by a family of cones corresponds to a graph of a biharmonic function carrying a family of isotropic circles. We classify such functions by showing that the top view of the family of circles is a pencil. © 2011 Springer-Verlag.

  9. Designing Pulse Laser Surface Modification of H13 Steel Using Response Surface Method

    Science.gov (United States)

    Aqida, S. N.; Brabazon, D.; Naher, S.

    2011-01-01

    This paper presents a design of experiment (DOE) for laser surface modification process of AISI H13 tool steel in achieving the maximum hardness and minimum surface roughness at a range of modified layer depth. A Rofin DC-015 diffusion-cooled CO2 slab laser was used to process AISI H13 tool steel samples. Samples of 10 mm diameter were sectioned to 100 mm length in order to process a predefined circumferential area. The parameters selected for examination were laser peak power, overlap percentage and pulse repetition frequency (PRF). The response surface method with Box-Behnken design approach in Design Expert 7 software was used to design the H13 laser surface modification process. Metallographic study and image analysis were done to measure the modified layer depth. The modified surface roughness was measured using two-dimensional surface profilometer. The correlation of the three laser processing parameters and the modified surface properties was specified by plotting three-dimensional graph. The hardness properties were tested at 981 mN force. From metallographic study, the laser modified surface depth was between 37 μm and 150 μm. The average surface roughness recorded from the 2D profilometry was at a minimum value of 1.8 μm. The maximum hardness achieved was between 728 and 905 HV0.1. These findings are significant to modern development of hard coatings for wear resistant applications.

  10. Comment on 'Surface thermodynamics, surface stress, equations at surfaces and triple lines for deformable bodies'

    Energy Technology Data Exchange (ETDEWEB)

    Gutman, E M, E-mail: gutman@bgu.ac.i [Department of Materials Engineering, Ben-Gurion University of the Negev, PO Box 653, Beer-Sheva 84105 (Israel)

    2010-10-27

    In a recent publication by Olives (2010 J. Phys.: Condens. Matter 22 085005) he studied 'the thermodynamics and mechanics of the surface of a deformable body, following and refining the general approach of Gibbs' and believed that 'a new definition of the surface stress is given'. However, using the usual way of deriving the equations of Gibbs-Duhem type the author, nevertheless, has fallen into a mathematical discrepancy because he has tried to unite in one equation different thermodynamic systems and 'a new definition of the surface stress' has appeared known in the usual theory of elasticity. (comment)

  11. Influence of surface topography on the surface durability of steam oxidised sintered iron

    Directory of Open Access Journals (Sweden)

    José Daniel Biasoli de Mello

    2005-06-01

    Full Text Available Durability of surfaces has been reported as the main factor affecting tribological behavior of steam oxidised sintered iron. The presence of surface pores and their negative influence on load bearing capacity, suggest that surface topography might play an important role on the durability of the oxide layer. In this paper, the influence of compaction pressure and powder grade on surface topography, and as a consequence, its effect on the tribological behavior of steam oxidised sintered iron has been analysed. Specimens prepared from atomised iron powders with different sizes were compacted using 4 different pressures, sintered, and then subjected to steam treatment. Tribological characterisation was carried out in a reciprocating sliding wear test. Although the processing parameters affected the surface topography to a considerable extent, the main influence may be attributed to powder grade. A strong influence of surface topography on the durability distance, evaluated in terms of the evolution of contact resistance with total sliding distance, has been highlighted. Surfaces which were smoother and had high load-carrying capacity were always associated with a higher durability distance.

  12. Inverse analysis of inner surface temperature history from outer surface temperature measurement of a pipe

    International Nuclear Information System (INIS)

    Kubo, S; Ioka, S; Onchi, S; Matsumoto, Y

    2010-01-01

    When slug flow runs through a pipe, nonuniform and time-varying thermal stresses develop and there is a possibility that thermal fatigue occurs. Therefore it is necessary to know the temperature distributions and the stress distributions in the pipe for the integrity assessment of the pipe. It is, however, difficult to measure the inner surface temperature directly. Therefore establishment of the estimation method of the temperature history on inner surface of pipe is needed. As a basic study on the estimation method of the temperature history on the inner surface of a pipe with slug flow, this paper presents an estimation method of the temperature on the inner surface of a plate from the temperature on the outer surface. The relationship between the temperature history on the outer surface and the inner surface is obtained analytically. Using the results of the mathematical analysis, the inverse analysis method of the inner surface temperature history estimation from the outer surface temperature history is proposed. It is found that the inner surface temperature history can be estimated from the outer surface temperature history by applying the inverse analysis method, even when it is expressed by the multiple frequency components.

  13. Evaluation of surface sampling method performance for Bacillus Spores on clean and dirty outdoor surfaces.

    Energy Technology Data Exchange (ETDEWEB)

    Wilson, Mollye C.; Einfeld, Wayne; Boucher, Raymond M.; Brown, Gary Stephen; Tezak, Matthew Stephen

    2011-06-01

    Recovery of Bacillus atrophaeous spores from grime-treated and clean surfaces was measured in a controlled chamber study to assess sampling method performance. Outdoor surfaces investigated by wipe and vacuum sampling methods included stainless steel, glass, marble and concrete. Bacillus atrophaeous spores were used as a surrogate for Bacillus anthracis spores in this study designed to assess whether grime-coated surfaces significantly affected surface sampling method performance when compared to clean surfaces. A series of chamber tests were carried out in which known amounts of spores were allowed to gravitationally settle onto both clean and dirty surfaces. Reference coupons were co-located with test coupons in all chamber experiments to provide a quantitative measure of initial surface concentrations of spores on all surfaces, thereby allowing sampling recovery calculations. Results from these tests, carried out under both low and high humidity conditions, show that spore recovery from grime-coated surfaces is the same as or better than spore recovery from clean surfaces. Statistically significant differences between method performance for grime-coated and clean surfaces were observed in only about half of the chamber tests conducted.

  14. Solvay Conference on Surface Science

    CERN Document Server

    1988-01-01

    The articles collected in this volume give a broad overview of the current state of surface science. Pioneers in the field and researchers met together at this Solvay Conference to discuss important new developments in surface science, with an emphasis on the common area between solid state physics and physical chemistry. The contributions deal with the following subjects: structure of surfaces, surface science and catalysis, two-dimensional physics and phase transitions, scanning tunneling microscopy, surface scattering and surface dynamics, chemical reactions at surfaces, solid-solid interfaces and superlattices, and surface studies with synchrotron radiation. On each of these subjects an introductory review talk and a number of short research contributions are followed by extensive discussions, which appear in full in the text. This nineteenth Solvay Conference commemorates the 75th anniversary of the Solvay Institutes.

  15. Biomechanical aspects of playing surfaces.

    Science.gov (United States)

    Nigg, B M; Yeadon, M R

    1987-01-01

    The purpose of this paper is to discuss some biomechanical aspects of playing surfaces with special focus on (a) surface induced injuries, (b) methodologies used to assess surfaces and (c) findings from various sports. The paper concentrates primarily on questions related to load on the athlete's body. Data from epidemiological studies suggest strongly that the surface is an important factor in the aetiology of injuries. Injury frequencies are reported to be significantly different for different surfaces in several sports. The methodologies used to assess surfaces with respect to load or performance include material tests and tests using experimental subjects. There is only little correlation between the results of these two approaches. Material tests used in many standardized test procedures are not validated which suggests that one should exercise restraint in the interpretation of these results. Point elastic surfaces are widely studied while area elastic surfaces have received little attention to date. Questions of energy losses on sport surfaces have rarely been studied scientifically.

  16. Surface decontamination

    International Nuclear Information System (INIS)

    Silva, S. da; Teixeira, M.V.

    1986-06-01

    The general methods of surface decontamination used in laboratory and others nuclear installations areas, as well as the procedures for handling radioactive materials and surfaces of work are presented. Some methods for decontamination of body external parts are mentioned. The medical supervision and assistance are required for internal or external contamination involving or not lesion in persons. From this medical radiation protection decontamination procedures are determined. (M.C.K.) [pt

  17. Effects of surface modification on the critical behaviour in multiple-surface-layer ferroelectric thin films

    International Nuclear Information System (INIS)

    Lu, Z X

    2013-01-01

    Using the usual mean-field theory approximation, the critical behaviour (i.e. the Curie temperature T c and the critical surface transverse field Ω sc ) in a multiple-surface-layer ferroelectric thin film is studied on the basis of the spin- 1/2 transverse Ising model. The dependence of the Curie temperature T c on the surface transverse field Ω s and the surface layer number N s are discussed in detail. Meanwhile the dependence of the critical surface transverse field Ω sc on the surface layer number N s is also examined. The numerical results indicate that the critical behaviour of ferroelectric thin films is obviously affected by modifications of the surface transverse field Ω s and surface layer number N s .

  18. Architectural Knitted Surfaces

    DEFF Research Database (Denmark)

    Mossé, Aurélie

    2010-01-01

    WGSN reports from the Architectural Knitted Surfaces workshop recently held at ShenkarCollege of Engineering and Design, Tel Aviv, which offered a cutting-edge insight into interactive knitted surfaces. With the increasing role of smart textiles in architecture, the Architectural Knitted Surfaces...... workshop brought together architects and interior and textile designers to highlight recent developments in intelligent knitting. The five-day workshop was led by architects Ayelet Karmon and Mette Ramsgaard Thomsen, together with Amir Cang and Eyal Sheffer from the Knitting Laboratory, in collaboration...

  19. Cooperativity in Surface Bonding and Hydrogen Bonding of Water and Hydroxyl at Metal Surfaces

    DEFF Research Database (Denmark)

    Schiros, T.; Ogasawara, H.; Naslund, L. A.

    2010-01-01

    of the mixed phase at metal surfaces. The surface bonding can be considered to be similar to accepting a hydrogen bond, and we can thereby apply general cooperativity rules developed for hydrogen-bonded systems. This provides a simple understanding of why water molecules become more strongly bonded...... to the surface upon hydrogen bonding to OH and why the OH surface bonding is instead weakened through hydrogen bonding to water. We extend the application of this simple model to other observed cooperativity effects for pure water adsorption systems and H3O+ on metal surfaces.......We examine the balance of surface bonding and hydrogen bonding in the mixed OH + H2O overlayer on Pt(111), Cu(111), and Cu(110) via density functional theory calculations. We find that there is a cooperativity effect between surface bonding and hydrogen bonding that underlies the stability...

  20. Stretch-minimising stream surfaces

    KAUST Repository

    Barton, Michael; Kosinka, Jin; Calo, Victor M.

    2015-01-01

    We study the problem of finding stretch-minimising stream surfaces in a divergence-free vector field. These surfaces are generated by motions of seed curves that propagate through the field in a stretch minimising manner, i.e., they move without stretching or shrinking, preserving the length of their arbitrary arc. In general fields, such curves may not exist. How-ever, the divergence-free constraint gives rise to these 'stretch-free' curves that are locally arc-length preserving when infinitesimally propagated. Several families of stretch-free curves are identified and used as initial guesses for stream surface generation. These surfaces are subsequently globally optimised to obtain the best stretch-minimising stream surfaces in a given divergence-free vector field. Our algorithm was tested on benchmark datasets, proving its applicability to incompressible fluid flow simulations, where our stretch-minimising stream surfaces realistically reflect the flow of a flexible univariate object. © 2015 Elsevier Inc. All rights reserved.

  1. Stretch-minimising stream surfaces

    KAUST Repository

    Barton, Michael

    2015-05-01

    We study the problem of finding stretch-minimising stream surfaces in a divergence-free vector field. These surfaces are generated by motions of seed curves that propagate through the field in a stretch minimising manner, i.e., they move without stretching or shrinking, preserving the length of their arbitrary arc. In general fields, such curves may not exist. How-ever, the divergence-free constraint gives rise to these \\'stretch-free\\' curves that are locally arc-length preserving when infinitesimally propagated. Several families of stretch-free curves are identified and used as initial guesses for stream surface generation. These surfaces are subsequently globally optimised to obtain the best stretch-minimising stream surfaces in a given divergence-free vector field. Our algorithm was tested on benchmark datasets, proving its applicability to incompressible fluid flow simulations, where our stretch-minimising stream surfaces realistically reflect the flow of a flexible univariate object. © 2015 Elsevier Inc. All rights reserved.

  2. The loaded surface profile: a new technique for the investigation of contact surfaces

    OpenAIRE

    McBride, J.W.

    2006-01-01

    Contact between rough surfaces produces a complex contact profile. The contact area is usually estimated according to roughness statistics in conjunction withsurface models or by examining the surfaces before and after contact. Most of the existing literature on loaded surface profiles is theoretical or numerical in nature. This paper presents a methodology for a new system to measure the loaded surface profile, based on a non-contact 3D laser profiler. The system allows the measurement of...

  3. Viscoelastic Surface Waves

    Science.gov (United States)

    Borcherdt, R. D.

    2007-12-01

    General theoretical solutions for Rayleigh- and Love-Type surface waves in viscoelastic media describe physical characteristics of the surface waves in elastic as well as anelastic media with arbitrary amounts of intrinsic absorption. In contrast to corresponding physical characteristics for Rayleigh waves in elastic media, Rayleigh- Type surface waves in anelastic media demonstrate; 1) tilt of the particle motion orbit that varies with depth, and 2) amplitude and volumetric strain distributions with superimposed sinusoidal variations that decay exponentially with depth. Each characteristic is dependent on the amount of intrinsic absorption and the chosen model of viscoelasticity. Distinguishing characteristics of anelastic Love-Type surface waves include: 1) dependencies of the wave speed and absorption coefficient on the chosen model and amount of intrinsic absorption and frequency, and 2) superimposed sinusoidal amplitude variations with an exponential decay with depth. Numerical results valid for a variety of viscoelastic models provide quantitative estimates of the physical characteristics of both types of viscoelastic surface waves appropriate for interpretations pertinent to models of earth materials ranging from low-loss in the crust to moderate- and high-loss in water-saturated soils.

  4. Electron acceleration by surface plasma waves in double metal surface structure

    Science.gov (United States)

    Liu, C. S.; Kumar, Gagan; Singh, D. B.; Tripathi, V. K.

    2007-12-01

    Two parallel metal sheets, separated by a vacuum region, support a surface plasma wave whose amplitude is maximum on the two parallel interfaces and minimum in the middle. This mode can be excited by a laser using a glass prism. An electron beam launched into the middle region experiences a longitudinal ponderomotive force due to the surface plasma wave and gets accelerated to velocities of the order of phase velocity of the surface wave. The scheme is viable to achieve beams of tens of keV energy. In the case of a surface plasma wave excited on a single metal-vacuum interface, the field gradient normal to the interface pushes the electrons away from the high field region, limiting the acceleration process. The acceleration energy thus achieved is in agreement with the experimental observations.

  5. Ozone-surface reactions in five homes: surface reaction probabilities, aldehyde yields, and trends.

    Science.gov (United States)

    Wang, H; Morrison, G

    2010-06-01

    Field experiments were conducted in five homes during three seasons (summer 2005, summer 2006 and winter 2007) to quantify ozone-initiated secondary aldehyde yields, surface reaction probabilities, and trends any temporal over a 1.5-year interval. Surfaces examined include living room carpets, bedroom carpets, kitchen floors, kitchen counters, and living room walls. Reaction probabilities for all surfaces for all seasons ranged from 9.4 x 10(-8) to 1.0 x 10(-4). There were no significant temporal trends in reaction probabilities for any surfaces from summer 2005 to summer 2006, nor over the entire 1.5-year period, indicating that it may take significantly longer than this period for surfaces to exhibit any 'ozone aging' or lowering of ozone-surface reactivity. However, all surfaces in three houses exhibited a significant decrease in reaction probabilities from summer 2006 to winter 2007. The total yield of aldehydes for the summer of 2005 were nearly identical to that for summer of 2006, but were significantly higher than for winter 2007. We also observed that older carpets were consistently less reactive than in newer carpets, but that countertops remained consistently reactive, probably because of occupant activities such as cooking and cleaning. Ozone reactions taking place at indoor surfaces significantly influence personal exposure to ozone and volatile reaction products. These field studies show that indoor surfaces only slowly lose their ability to react with ozone over several year time frames, and that this is probably because of a combination of large reservoirs of reactive coatings and periodic additions of reactive coatings in the form of cooking, cleaning, and skin-oil residues. When considering exposure to ozone and its reaction products and in the absence of dramatic changes in occupancy, activities or furnishings, indoor surface reactivity is expected to change very slowly.

  6. Ion beam surface treatment: A new capability for rapid melt and resolidification of surfaces

    International Nuclear Information System (INIS)

    Stinnett, R.W.; McIntyre, D.C.; Buchheit, R.G.; Greenly, J.B.; Thompson, M.O.

    1994-01-01

    The emerging capability to produce high average power (5--250 kW) pulsed ion beams at 0.2--2 MeV energies is enabling us to develop a new, commercial-scale thermal surface treatment technology called Ion Beam Surface Treatment (IBEST). This technique uses high energy, pulsed (≤100 ns) ion beams to directly deposit energy in the top 2--20 micrometers of the surface of any material. Depth of treatment is controllable by varying the ion energy and species. Deposition of the energy with short pulses in a thin surface layer allows melting of the layer with relatively small energies and allows rapid cooling of the melted layer by thermal diffusion into the underlying substrate. Typical cooling rates of this process (10 9 10 10 K/sec) cause rapid resolidification, resulting in production of non-equilibrium microstructures (nano-crystalline and metastable phases) that have significantly improved corrosion, wear, and hardness properties. We have conducted IBEST feasibility experiments with results confirming surface hardening, nanocrystaline grain formation, metal surface polishing, controlled melt of ceramic surfaces, and surface cleaning

  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. The surface learned from nature

    Science.gov (United States)

    Lim, H.; Kim, W. D.

    2010-07-01

    In this work, I would like to introduce the emerging surface of nature. The surface in nature, has the multi and optimized function with well organized structure. There are so many examples that we learn and apply to technology. First example is self-cleaning surface. Some plants (such as lotus leaf, taro leaf) and the wings of many large-winged insects (such as moth, butterfly, dragonfly) remain their surface clean in the very dirty environment. This self cleaning effect is accomplished by the superhydrophobic surfaces which exhibit the water contact angle of more than 150° with low sliding angle. Generally, the superhydrophobic surface is made up the two factors. One is the surface composition having the low surface tension energy. The other is the surface morphology of hierarchical structure of micro and nano size. Because almost nature surface have the hierarchical structures range from macro to nano size, their topography strength their function to adjust the life in nature environment. The other example is the surface to use for drag reduction. The skin friction drag causes eruptions of air or water resulting in greater drag as the speed is increased. This drag requires more energy to overcome. The shark skin having the fine sharp-edged grooves about 0.1 mm wide known riblet reduces in skin friction drag by being far away the vortex. Among a lot of fuctional surface, the most exciting surface the back of stenocara a kind of desert beetles. Stenocara use the micrometre-sized patterns of hydrophobic, wax-coated and hydrophilic, non-waxy regions on their backs to capture water from fog. This fog-collecting structure improves the water collection of fog-capture film, condenser, engine, and future building. Here, the efforts to realize these emerging functional surfaces in nature on technology are reported with the fabrication method and their properties, especially for the control of surface wettability.

  9. Surface science an introduction

    CERN Document Server

    Hudson, John

    1991-01-01

    The whole field of surface science is covered in this work. Starting with a description of the structure and thermodynamics of clean surfaces, the book goes on to discuss kinetic theory of gases and molecular beam formation. This is followed by a largesection on gas-surface interactions, and another major section on energetic particle-surface interactions. The final chapter provides the background to crystal nucleation and growth. The approach adopted is interdisciplinary and slanted towards theexperimental side, with practical analytical techniques being used to illustrate general princi

  10. Surface preparation of niobium

    International Nuclear Information System (INIS)

    Kneisel, P.

    1980-01-01

    Any discussion of surface preparation for superconducting rf-surfaces is certainly connected with the question what is the best recipe for achieving high Q-values and high break-down fields. Since the break-down in a cavity is not understood so far and because several mechanisms play a role, it also is not possible to give one recipe which always works. Nevertheless in the past certain preparation techniques for niobium surfaces have been developed and certain rules for preparation can be applied. In the following the to-days state of the art will be described and it is attempted to give a short description of the surface in conjunction with the methods of surface treatments, which generally can be applied to niobium cavities. (orig./WTR)

  11. A novel test method for quantifying surface tack of polypropylene compound surfaces

    Directory of Open Access Journals (Sweden)

    2011-11-01

    Full Text Available While adhesiveness is required for polymer surfaces in special applications, tacky surfaces are generally undesirable in many applications like automotive interior parts. The tackiness of polymer surface results from a combination of composition and additivation, and it can change significantly in natural or accelerated ageing. Since there is no established, uniform method to characterize surface tack, the major focus of the present work was on the development of an objective quantification method. A setup having a soft die tip attached to a standard tensile tester was developed aiming for correlation to the human sense of touch. Three different model thermoplastic polyolefin (TPO compound formulations based on a high-impact isotactic polypropylene (iPP composition with varying amounts and types of anti-scratch additives were used for these investigations. As the surface tack phenomenon is related to ageing and weathering, the material’s examination was also performed after various intervals of weathering. The developed method allows a fast assessment of the effect of polymer composition variations and different additive formulations on surface tack and gives identical rankings as the standardized haptic panel.

  12. Imaging near-surface heterogeneities by natural migration of backscattered surface waves

    KAUST Repository

    AlTheyab, Abdullah; Lin, Fan-Chi; Schuster, Gerard T.

    2016-01-01

    We present a migration method that does not require a velocity model to migrate backscattered surface waves to their projected locations on the surface. This migration method, denoted as natural migration, uses recorded Green's functions along

  13. Study on tribological properties of multi-layer surface texture on Babbitt alloys surface

    Science.gov (United States)

    Zhang, Dongya; Zhao, Feifei; Li, Yan; Li, Pengyang; Zeng, Qunfeng; Dong, Guangneng

    2016-12-01

    To improve tribological properties of Babbitt alloys, multi-layer surface texture consisted of the main grooves and secondary micro-dimples are fabricated on the Babbitt substrate through laser pulse ablation. The tribological behaviors of multi-layer surface texture are investigated using a rotating type pin-on-disc tribo-meter under variation sliding speeds, and the film pressure distributions on the textured surfaces are simulated using computational fluid dynamics (CFD) method for elucidating the possible mechanisms. The results suggest that: (i) the multi-layer surface texture can reduce friction coefficient of Babbitt alloy, which has lowest friction coefficient of 0.03, in case of the groove parameter of 300 μm width and 15% of area density; (ii) the improvement effect may be more sensitive to the groove area density and the siding speed, and the textured surface with lower area density has lower friction coefficient under high sliding speed. Based on the reasons of (i) the secondary micro-dimples on Babbitt alloy possesses a hydrophobicity surface and (ii) the CFD analysis indicates that main grooves enhancing hydrodynamic effect, thus the multi-layer surface texture is regarded as dramatically improve the lubricating properties of the Babbitt alloy.

  14. The metal-ion-dependent adhesion site in the Von Willebrand factor-A domain of α2δ subunits is key to trafficking voltage-gated Ca2+ channels

    Science.gov (United States)

    Cantí, C.; Nieto-Rostro, M.; Foucault, I.; Heblich, F.; Wratten, J.; Richards, M. W.; Hendrich, J.; Douglas, L.; Page, K. M.; Davies, A.; Dolphin, A. C.

    2005-01-01

    All auxiliary α2δ subunits of voltage-gated Ca2+ (CaV) channels contain an extracellular Von Willebrand factor-A (VWA) domain that, in α2δ-1 and -2, has a perfect metal-ion-dependent adhesion site (MIDAS). Modeling of the α2δ-2 VWA domain shows it to be highly likely to bind a divalent cation. Mutating the three key MIDAS residues responsible for divalent cation binding resulted in a MIDAS mutant α2δ-2 subunit that was still processed and trafficked normally when it was expressed alone. However, unlike WT α2δ-2, the MIDAS mutant α2δ-2 subunit did not enhance and, in some cases, further diminished CaV1.2, -2.1, and -2.2 currents coexpressed with β1b by using either Ba2+ or Na+ as a permeant ion. Furthermore, expression of the MIDAS mutant α2δ-2 reduced surface expression and strongly increased the perinuclear retention of CaVα1 subunits at the earliest time at which expression was observed in both Cos-7 and NG108–15 cells. Despite the presence of endogenous α2δ subunits, heterologous expression of α2δ-2 in differentiated NG108–15 cells further enhanced the endogenous high-threshold Ca2+ currents, whereas this enhancement was prevented by the MIDAS mutations. Our results indicate that α2δ subunits normally interact with the CaVα1 subunit early in their maturation, before the appearance of functional plasma membrane channels, and an intact MIDAS motif in the α2δ subunit is required to promote trafficking of the α1 subunit to the plasma membrane by an integrin-like switch. This finding provides evidence for a primary role of a VWA domain in intracellular trafficking of a multimeric complex, in contrast to the more usual roles in binding extracellular ligands in other exofacial VWA domains. PMID:16061813

  15. Surface properties of beached plastics.

    Science.gov (United States)

    Fotopoulou, Kalliopi N; Karapanagioti, Hrissi K

    2015-07-01

    Studying plastic characteristics in the marine environment is important to better understand interaction between plastics and the environment. In the present study, high-density polyethylene (HDPE), polyethylene terephalate (PET), and polyvinyl chloride (PVC) samples were collected from the coastal environment in order to study their surface properties. Surface properties such as surface functional groups, surface topography, point of zero charge, and color change are important factors that change during degradation. Eroded HDPE demonstrated an altered surface topography and color and new functional groups. Eroded PET surface was uneven, yellow, and occasionally, colonized by microbes. A decrease in Fourier transform infrared (FTIR) peaks was observed for eroded PET suggesting that degradation had occurred. For eroded PVC, its surface became more lamellar and a new FTIR peak was observed. These surface properties were obtained due to degradation and could be used to explain the interaction between plastics, microbes, and pollutants.

  16. Surface roughening under ion bombardment

    International Nuclear Information System (INIS)

    Bhatia, C.S.

    1982-01-01

    Ion bombardment can cause roughening of a surface. Inadequate step coverage and poor adhesion of films on such surfaces are of concern. An extreme case of surface roughening results in cone formation under ion bombardment. The results of the investigation, using scanning electron microscopy, is discussed in terms of the role of (a) embedded particles, (b) impurities and (c) surface migration in cone formation on the target surface. (Auth.)

  17. Surface relaxation and surface energy of face –centered Cubic ...

    African Journals Online (AJOL)

    DR. MIKE HORSFALL

    Surface relaxation and surface energy of face –centered Cubic metals. 1AGHEMENLO H E; *2IYAYI, S E; 3AVWIRI ,G O. 1, 3 Department of Physics, Ambrose Alli University, Ekpoma, Nigeria. 2 Department of Physics, University of Benin, Benin City, Nigeria. 3 Department of Physics, University of Port Harcourt, PH, Nigeria.

  18. Surface and subsurface cracks characteristics of single crystal SiC wafer in surface machining

    Energy Technology Data Exchange (ETDEWEB)

    Qiusheng, Y., E-mail: qsyan@gdut.edu.cn; Senkai, C., E-mail: senkite@sina.com; Jisheng, P., E-mail: panjisheng@gdut.edu.cn [School of Electromechanical Engineering, Guangdong University of Technology, Guangzhou, 510006 (China)

    2015-03-30

    Different machining processes were used in the single crystal SiC wafer machining. SEM was used to observe the surface morphology and a cross-sectional cleavages microscopy method was used for subsurface cracks detection. Surface and subsurface cracks characteristics of single crystal SiC wafer in abrasive machining were analysed. The results show that the surface and subsurface cracks system of single crystal SiC wafer in abrasive machining including radial crack, lateral crack and the median crack. In lapping process, material removal is dominated by brittle removal. Lots of chipping pits were found on the lapping surface. With the particle size becomes smaller, the surface roughness and subsurface crack depth decreases. When the particle size was changed to 1.5µm, the surface roughness Ra was reduced to 24.0nm and the maximum subsurface crack was 1.2µm. The efficiency of grinding is higher than lapping. Plastic removal can be achieved by changing the process parameters. Material removal was mostly in brittle fracture when grinding with 325# diamond wheel. Plow scratches and chipping pits were found on the ground surface. The surface roughness Ra was 17.7nm and maximum subsurface crack depth was 5.8 µm. When grinding with 8000# diamond wheel, the material removal was in plastic flow. Plastic scratches were found on the surface. A smooth surface of roughness Ra 2.5nm without any subsurface cracks was obtained. Atomic scale removal was possible in cluster magnetorheological finishing with diamond abrasive size of 0.5 µm. A super smooth surface eventually obtained with a roughness of Ra 0.4nm without any subsurface crack.

  19. How old is surface science?

    International Nuclear Information System (INIS)

    Paparazzo, E.

    2004-01-01

    Philosophical and literary testimonies from the Classical World (5th century B.C. to 3rd century A.D.) involving solid surfaces are reviewed. Plato thought the surface to be a real entity, whereas Aristotle considered it to possess an unqualified existence, i.e. not to be a substance, but just an accidental entity. The Old Stoics asserted that surfaces do not possess any physical existence, although the Stoic philosopher Posidonius--apparently the only exception in his school--held them to exist both in thought and reality. While both the Atomists and the Epicureans were very little interested in them, the Sceptic philosopher Sextus Empiricus considered surfaces to be the limits of a body, although he maintained that both the view that they are corporeal or the view that they are incorporeal present unsurmountable difficulties. Among Roman authors, the testimony from Pliny the Elder is mostly concerned with metallic surfaces, chemical change occurring there, and surface treatments used in antiquity. Besides the philosophical motivations, the implications of the testimonies are discussed in the light of surface science. The purely geometrical surface of Plato is found to compare favorably to single-crystal surface, Posidonius' 'corporeal' surface is best likened to an air-oxidized, or otherwise ambient-modified surface, and ancient accounts on mixture are compared to XPS results obtained in adhesion studies of enameled steels. I argue that the long-standing dominance of Aristotle's view from antiquity onwards may have had a part in delaying theoretical speculation into solid surfaces

  20. Nonlinear optical studies of surfaces

    International Nuclear Information System (INIS)

    Shen, Y.R.

    1994-07-01

    The possibly of using nonlinear optical processes for surface studies has attracted increasing attention in recent years. Optical second harmonic generation (SHG) and sum frequency generation (SFG), in particular, have been well accepted as viable surface probes. They have many advantages over the conventional techniques. By nature, they are highly surface-specific and has a submonolayer sensitivity. As coherent optical processes, they are capable of in-situ probing of surfaces in hostile environment as well as applicable to all interfaces accessible by light. With ultrafast pump laser pulses, they can be employed to study surface dynamic processes with a subpicosecond time resolution. These advantages have opened the door to many exciting research opportunities in surface science and technology. This paper gives a brief overview of this fast-growing new area of research. Optical SHG from a surface was first studied theoretically and experimentally in the sixties. Even the submonolayer surface sensitivity of the process was noticed fairly early. The success was, however, limited because of difficulties in controlling the experimental conditions. It was not until the early 1980's that the potential of the process for surface analysis was duly recognized. The first surface study by SHG was actually motivated by the then active search for an understanding of the intriguing surface enhanced Raman scattering (SERS). It had been suspected that the enhancement in SERS mainly came from the local-field enhancement due to local plasmon resonances and pointing rod effect on rough metal surfaces. In our view, Raman scattering is a two-photon process and is therefore a nonlinear optical effect