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Sample records for copper-tio2-boric acid hybrid

  1. Kit for detecting nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    2001-01-01

    A kit is provided for detecting a target nucleic acid sequence in a sample, the kit comprising: a first hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a first portion of the target sequence, the first hybridization probe including a first complexing agent for forming a binding pair with a second complexing agent; and a second hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a second portion of the target sequence to which the first hybridization probe does not selectively hybridize, the second hybridization probe including a detectable marker; a third hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a first portion of the target sequence, the third hybridization probe including the same detectable marker as the second hybridization probe; and a fourth hybridization probe which includes a nucleic acid sequence that is sufficiently complementary to selectively hybridize to a second portion of the target sequence to which the third hybridization probe does not selectively hybridize, the fourth hybridization probe including the first complexing agent for forming a binding pair with the second complexing agent; wherein the first and second hybridization probes are capable of simultaneously hybridizing to the target sequence and the third and fourth hybridization probes are capable of simultaneously hybridizing to the target sequence, the detectable marker is not present on the first or fourth hybridization probes and the first, second, third, and fourth hybridization probes each include a competitive nucleic acid sequence which is sufficiently complementary to a third portion of the target sequence that the competitive sequences of the first, second, third, and fourth hybridization probes compete with each other to hybridize to the third portion of the

  2. Rapid hybridization of nucleic acids using isotachophoresis

    PubMed Central

    Bercovici, Moran; Han, Crystal M.; Liao, Joseph C.; Santiago, Juan G.

    2012-01-01

    We use isotachophoresis (ITP) to control and increase the rate of nucleic acid hybridization reactions in free solution. We present a new physical model, validation experiments, and demonstrations of this assay. We studied the coupled physicochemical processes of preconcentration, mixing, and chemical reaction kinetics under ITP. Our experimentally validated model enables a closed form solution for ITP-aided reaction kinetics, and reveals a new characteristic time scale which correctly predicts order 10,000-fold speed-up of chemical reaction rate for order 100 pM reactants, and greater enhancement at lower concentrations. At 500 pM concentration, we measured a reaction time which is 14,000-fold lower than that predicted for standard second-order hybridization. The model and method are generally applicable to acceleration of reactions involving nucleic acids, and may be applicable to a wide range of reactions involving ionic reactants. PMID:22733732

  3. Optimizing the specificity of nucleic acid hybridization

    NASA Astrophysics Data System (ADS)

    Zhang, David Yu; Chen, Sherry Xi; Yin, Peng

    2012-03-01

    The specific hybridization of complementary sequences is an essential property of nucleic acids, enabling diverse biological and biotechnological reactions and functions. However, the specificity of nucleic acid hybridization is compromised for long strands, except near the melting temperature. Here, we analytically derived the thermodynamic properties of a hybridization probe that would enable near-optimal single-base discrimination and perform robustly across diverse temperature, salt and concentration conditions. We rationally designed ‘toehold exchange’ probes that approximate these properties, and comprehensively tested them against five different DNA targets and 55 spurious analogues with energetically representative single-base changes (replacements, deletions and insertions). These probes produced discrimination factors between 3 and 100+ (median, 26). Without retuning, our probes function robustly from 10 °C to 37 °C, from 1 mM Mg2+ to 47 mM Mg2+, and with nucleic acid concentrations from 1 nM to 5 µM. Experiments with RNA also showed effective single-base change discrimination.

  4. Optimizing the specificity of nucleic acid hybridization.

    PubMed

    Zhang, David Yu; Chen, Sherry Xi; Yin, Peng

    2012-01-22

    The specific hybridization of complementary sequences is an essential property of nucleic acids, enabling diverse biological and biotechnological reactions and functions. However, the specificity of nucleic acid hybridization is compromised for long strands, except near the melting temperature. Here, we analytically derived the thermodynamic properties of a hybridization probe that would enable near-optimal single-base discrimination and perform robustly across diverse temperature, salt and concentration conditions. We rationally designed 'toehold exchange' probes that approximate these properties, and comprehensively tested them against five different DNA targets and 55 spurious analogues with energetically representative single-base changes (replacements, deletions and insertions). These probes produced discrimination factors between 3 and 100+ (median, 26). Without retuning, our probes function robustly from 10 °C to 37 °C, from 1 mM Mg(2+) to 47 mM Mg(2+), and with nucleic acid concentrations from 1 nM to 5 µM. Experiments with RNA also showed effective single-base change discrimination.

  5. Optimizing the specificity of nucleic acid hybridization

    PubMed Central

    Zhang, David Yu; Chen, Sherry Xi; Yin, Peng

    2014-01-01

    The specific hybridization of complementary sequences is an essential property of nucleic acids, enabling diverse biological and biotechnological reactions and functions. However, the specificity of nucleic acid hybridization is compromised for long strands, except near the melting temperature. Here, we analytically derived the thermodynamic properties of a hybridization probe that would enable near-optimal single-base discrimination and perform robustly across diverse temperature, salt and concentration conditions. We rationally designed ‘toehold exchange’ probes that approximate these properties, and comprehensively tested them against five different DNA targets and 55 spurious analogues with energetically representative single-base changes (replacements, deletions and insertions). These probes produced discrimination factors between 3 and 100+ (median, 26). Without retuning, our probes function robustly from 10 °C to 37 °C, from 1 mM Mg2+ to 47 mM Mg2+, and with nucleic acid concentrations from 1 nM to 5 μM. Experiments with RNA also showed effective single-base change discrimination. PMID:22354435

  6. Fluorescent hybridization probes for nucleic acid detection.

    PubMed

    Guo, Jia; Ju, Jingyue; Turro, Nicholas J

    2012-04-01

    Due to their high sensitivity and selectivity, minimum interference with living biological systems, and ease of design and synthesis, fluorescent hybridization probes have been widely used to detect nucleic acids both in vivo and in vitro. Molecular beacons (MBs) and binary probes (BPs) are two very important hybridization probes that are designed based on well-established photophysical principles. These probes have shown particular applicability in a variety of studies, such as mRNA tracking, single nucleotide polymorphism (SNP) detection, polymerase chain reaction (PCR) monitoring, and microorganism identification. Molecular beacons are hairpin oligonucleotide probes that present distinctive fluorescent signatures in the presence and absence of their target. Binary probes consist of two fluorescently labeled oligonucleotide strands that can hybridize to adjacent regions of their target and generate distinctive fluorescence signals. These probes have been extensively studied and modified for different applications by modulating their structures or using various combinations of fluorophores, excimer-forming molecules, and metal complexes. This review describes the applicability and advantages of various hybridization probes that utilize novel and creative design to enhance their target detection sensitivity and specificity.

  7. Hybridization and sequencing of nucleic acids using base pair mismatches

    DOEpatents

    Fodor, Stephen P. A.; Lipshutz, Robert J.; Huang, Xiaohua

    2001-01-01

    Devices and techniques for hybridization of nucleic acids and for determining the sequence of nucleic acids. Arrays of nucleic acids are formed by techniques, preferably high resolution, light-directed techniques. Positions of hybridization of a target nucleic acid are determined by, e.g., epifluorescence microscopy. Devices and techniques are proposed to determine the sequence of a target nucleic acid more efficiently and more quickly through such synthesis and detection techniques.

  8. Continuously tunable nucleic acid hybridization probes.

    PubMed

    Wu, Lucia R; Wang, Juexiao Sherry; Fang, John Z; Evans, Emily R; Pinto, Alessandro; Pekker, Irena; Boykin, Richard; Ngouenet, Celine; Webster, Philippa J; Beechem, Joseph; Zhang, David Yu

    2015-12-01

    In silico-designed nucleic acid probes and primers often do not achieve favorable specificity and sensitivity tradeoffs on the first try, and iterative empirical sequence-based optimization is needed, particularly in multiplexed assays. We present a novel, on-the-fly method of tuning probe affinity and selectivity by adjusting the stoichiometry of auxiliary species, which allows for independent and decoupled adjustment of the hybridization yield for different probes in multiplexed assays. Using this method, we achieved near-continuous tuning of probe effective free energy. To demonstrate our approach, we enforced uniform capture efficiency of 31 DNA molecules (GC content, 0-100%), maximized the signal difference for 11 pairs of single-nucleotide variants and performed tunable hybrid capture of mRNA from total RNA. Using the Nanostring nCounter platform, we applied stoichiometric tuning to simultaneously adjust yields for a 24-plex assay, and we show multiplexed quantitation of RNA sequences and variants from formalin-fixed, paraffin-embedded samples.

  9. Continuously Tunable Nucleic Acid Hybridization Probes

    PubMed Central

    Wu, Lucia R.; Wang, J. Sherry; Fang, John Z.; Reiser, Emily; Pinto, Alessandro; Pekker, Irena; Boykin, Richard; Ngouenet, Celine; Webster, Philippa J.; Beechem, Joseph; Zhang, David Yu

    2015-01-01

    In silico designed nucleic acid probes and primers often fail to achieve favorable specificity and sensitivity tradeoffs on the first try, and iterative empirical sequence-based optimization is needed, particularly in multiplexed assays. Here, we present a novel, on-the-fly method of tuning probe affinity and selectivity via the stoichiometry of auxiliary species, allowing independent and decoupled adjustment of hybridization yield for different probes in multiplexed assays. Using this method, we achieve near-continuous tuning of probe effective free energy (0.03 kcal·mol−1 granularity). As applications, we enforced uniform capture efficiency of 31 DNA molecules (GC content 0% – 100%), maximized signal difference for 11 pairs of single nucleotide variants, and performed tunable hybrid-capture of mRNA from total RNA. Using the Nanostring nCounter platform, we applied stoichiometric tuning to simultaneously adjust yields for a 24-plex assay, and we show multiplexed quantitation of RNA sequences and variants from formalin-fixed, paraffin-embedded samples (FFPE). PMID:26480474

  10. Nucleic acid in-situ hybridization detection of infectious agents

    NASA Astrophysics Data System (ADS)

    Thompson, Curtis T.

    2000-04-01

    Limitations of traditional culture methods and newer polymerase chain reaction (PCR)-based methods for detection and speciation of infectious agents demonstrate the need for more rapid and better diagnostics. Nucleic acid hybridization is a detection technology that has gained wide acceptance in cancer and prenatal cytogenetics. Using a modification of the nucleic acid hybridization technique known as fluorescence in-situ hybridization, infectious agents can be detected in a variety of specimens with high sensitivity and specificity. The specimens derive from all types of human and animal sources including body fluids, tissue aspirates and biopsy material. Nucleic acid hybridization can be performed in less than one hour. The result can be interpreted either using traditional fluorescence microscopy or automated platforms such as micro arrays. This paper demonstrates proof of concept for nucleic acid hybridization detection of different infectious agents. Interpretation within a cytologic and histologic context is possible with fluorescence microscopic analysis, thereby providing confirmatory evidence of hybridization. With careful probe selection, nucleic acid hybridization promises to be a highly sensitive and specific practical diagnostic alternative to culture, traditional staining methods, immunohistochemistry and complicated nucleic acid amplification tests.

  11. Hybrids of Nucleic Acids and Carbon Nanotubes for Nanobiotechnology

    PubMed Central

    Umemura, Kazuo

    2015-01-01

    Recent progress in the combination of nucleic acids and carbon nanotubes (CNTs) has been briefly reviewed here. Since discovering the hybridization phenomenon of DNA molecules and CNTs in 2003, a large amount of fundamental and applied research has been carried out. Among thousands of papers published since 2003, approximately 240 papers focused on biological applications were selected and categorized based on the types of nucleic acids used, but not the types of CNTs. This survey revealed that the hybridization phenomenon is strongly affected by various factors, such as DNA sequences, and for this reason, fundamental studies on the hybridization phenomenon are important. Additionally, many research groups have proposed numerous practical applications, such as nanobiosensors. The goal of this review is to provide perspective on biological applications using hybrids of nucleic acids and CNTs. PMID:28347014

  12. What controls the hybridization thermodynamics of spherical nucleic acids?

    PubMed

    Randeria, Pratik S; Jones, Matthew R; Kohlstedt, Kevin L; Banga, Resham J; Olvera de la Cruz, Monica; Schatz, George C; Mirkin, Chad A

    2015-03-18

    The hybridization of free oligonucleotides to densely packed, oriented arrays of DNA modifying the surfaces of spherical nucleic acid (SNA)-gold nanoparticle conjugates occurs with negative cooperativity; i.e., each binding event destabilizes subsequent binding events. DNA hybridization is thus an ever-changing function of the number of strands already hybridized to the particle. Thermodynamic quantification of this behavior reveals a 3 orders of magnitude decrease in the binding constant for the capture of a free oligonucleotide by an SNA conjugate as the fraction of pre-hybridized strands increases from 0 to ∼30%. Increasing the number of pre-hybridized strands imparts an increasing enthalpic penalty to hybridization that makes binding more difficult, while simultaneously decreasing the entropic penalty to hybridization, which makes binding more favorable. Hybridization of free DNA to an SNA is thus governed by both an electrostatic barrier as the SNA accumulates charge with additional binding events and an effect consistent with allostery, where hybridization at certain sites on an SNA modify the binding affinity at a distal site through conformational changes to the remaining single strands. Leveraging these insights allows for the design of conjugates that hybridize free strands with significantly higher efficiencies, some of which approach 100%.

  13. Fast hybridization solution for the detection of immobilized nucleic acids.

    PubMed

    Yang, T T; Kain, S R

    1995-03-01

    We have developed a fast hybridization solution, termed ExpressHyb, for the rapid and sensitive detection of nucleic acids immobilized on membrane supports. This solution reduces typical hybridization times of 12-24 h to as little as 1 h while simultaneously increasing the sensitivity of detection in many applications. Using ExpressHyb, human beta-actin mRNA was detected on a human multiple tissue Northern (MTN) blot following a 30-min hybridization, with optimal detection occurring with a 1-h hybridization interval. The moderately abundant human glyceraldehyde-3-phosphate dehydrogenase (G3PDH) mRNA was detected using similar hybridization conditions and yielded improved signal-to-background characteristics relative to overnight hybridizations in conventional solutions. ExpressHyb can be used with either 32P- or digoxigenin-labeled probes and works effectively with both cDNA and oligonucleotide probes. For non-isotopic detection in particular, ExpressHyb reduces the nonspecific background commonly encountered with this technique. In cDNA library screening, ExpressHyb was found to both reduce the time required for effective hybridizations and to increase the number of positive colonies obtained relative to conventional overnight procedures. Taken together, these results illustrate the broad capability of ExpressHyb Hybridization Solution to improve nucleic acid detection in a variety of important techniques.

  14. Hybride magnetic nanostructure based on amino acids functionalized polypyrrole

    SciTech Connect

    Nan, Alexandrina Bunge, Alexander; Turcu, Rodica

    2015-12-23

    Conducting polypyrrole is especially promising for many commercial applications because of its unique optical, electric, thermal and mechanical properties. We report the synthesis and characterization of novel pyrrole functionalized monomers and core-shell hybrid nanostructures, consisting of a conjugated polymer layer (amino acids functionalized pyrrole copolymers) and a magnetic nanoparticle core. For functionalization of the pyrrole monomer we used several amino acids: tryptophan, leucine, phenylalanine, serine and tyrosine. These amino acids were linked via different types of hydrophobic linkers to the nitrogen atom of the pyrrole monomer. The magnetic core-shell hybrid nanostructures are characterized by various methods such as FTIR spectroscopy, transmission electron microscopy (TEM) and magnetic measurements.

  15. Hybride magnetic nanostructure based on amino acids functionalized polypyrrole

    NASA Astrophysics Data System (ADS)

    Nan, Alexandrina; Bunge, Alexander; Turcu, Rodica

    2015-12-01

    Conducting polypyrrole is especially promising for many commercial applications because of its unique optical, electric, thermal and mechanical properties. We report the synthesis and characterization of novel pyrrole functionalized monomers and core-shell hybrid nanostructures, consisting of a conjugated polymer layer (amino acids functionalized pyrrole copolymers) and a magnetic nanoparticle core. For functionalization of the pyrrole monomer we used several amino acids: tryptophan, leucine, phenylalanine, serine and tyrosine. These amino acids were linked via different types of hydrophobic linkers to the nitrogen atom of the pyrrole monomer. The magnetic core-shell hybrid nanostructures are characterized by various methods such as FTIR spectroscopy, transmission electron microscopy (TEM) and magnetic measurements.

  16. Tracing hybrid incompatibilities to single amino acid substitutions.

    PubMed

    Harrison, J Scott; Burton, Ronald S

    2006-03-01

    Deleterious interactions among genes cause reductions in fitness of interpopulation hybrids (hybrid breakdown). Identifying genes involved in hybrid breakdown has proven difficult, and few studies have addressed the molecular basis of this widespread phenomenon. Because proper function of the mitochondrial electron transport system (ETS) requires a coadapted set of nuclear and mitochondrial gene products, ETS genes present an attractive system for studying the evolution of coadapted gene complexes within isolated populations and the loss of fitness in interpopulation hybrids. Here we show the effects of single amino acid substitutions in cytochrome c (CYC) on its functional interaction with another ETS protein, cytochrome c oxidase (COX) in the intertidal copepod Tigriopus californicus. The individual and pairwise consequences of three naturally occurring amino acid substitutions in CYC are examined by site-directed mutagenesis and found to differentially effect the rates of CYC oxidation by COX variants from different source populations. In one case, we show that interpopulation hybrid breakdown in COX activity can be attributed to a single naturally occurring amino acid substitution in CYC.

  17. Molecular beacons: nucleic acid hybridization and emerging applications.

    PubMed

    Antony, T; Subramaniam, V

    2001-12-01

    Molecular beacons (MBs) are a novel class of nucleic acid probes that become fluorescent when bound to a complementary sequence. Because of this characteristic, coupled with the sequence specificity of nucleic acid hybridization and the sensitivity of fluorescence techniques, MBs are very useful probes for a variety of applications requiring the detection of DNA or RNA. We survey various applications of MBs, including the monitoring of DNA triplex formation, and describe recent developments in MB design that enhance their sensitivity.

  18. Caged molecular beacons: controlling nucleic acid hybridization with light.

    PubMed

    Wang, Chunming; Zhu, Zhi; Song, Yanling; Lin, Hui; Yang, Chaoyong James; Tan, Weihong

    2011-05-28

    We have constructed a novel class of light-activatable caged molecular beacons (cMBs) that are caged by locking two stems with a photo-labile biomolecular interaction or covalent bond. With the cMBs, the nucleic acid hybridization process can be easily controlled with light, which offers the possibility for a high spatiotemporal resolution study of intracellular mRNAs.

  19. Synthesis and Characterization of Hybrid Hyaluronic Acid-Gelatin Hydrogels

    PubMed Central

    Camci-Unal, Gulden; Cuttica, Davide; Annabi, Nasim; Demarchi, Danilo; Khademhosseini, Ali

    2013-01-01

    Biomimetic hybrid hydrogels have generated broad interest in tissue engineering and regenerative medicine. Hyaluronic acid (HA) and gelatin (hydrolyzed collagen) are naturally derived polymers and biodegradable under physiological conditions. Moreover, collagen and HA are major components of the extracellular matrix (ECM) in most of the tissues (e.g. cardiovascular, cartilage, neural). When used as a hybrid material, HA-gelatin hydrogels may enable mimicking the ECM of native tissues. Although HA-gelatin hybrid hydrogels are promising biomimetic substrates, their material properties have not been thoroughly characterized in the literature. Herein, we generated hybrid hydrogels with tunable physical and biological properties by using different concentrations of HA and gelatin. The physical properties of the fabricated hydrogels including swelling ratio, degradation, and mechanical properties were investigated. In addition, in vitro cellular responses in both two and three dimensional (2D and 3D) culture conditions were assessed. It was found that the addition of gelatin methacrylate (GelMA) into HA methacrylate (HAMA) promoted cell spreading in the hybrid hydogels. Moreover, the hybrid hydrogels showed significantly improved mechanical properties compared to their single component analogs. The HAMA-GelMA hydrogels exhibited remarkable tunability behavior and may be useful for cardiovascular tissue engineering applications. PMID:23419055

  20. Natural cinnamic acids, synthetic derivatives and hybrids with antimicrobial activity.

    PubMed

    Guzman, Juan David

    2014-11-25

    Antimicrobial natural preparations involving cinnamon, storax and propolis have been long used topically for treating infections. Cinnamic acids and related molecules are partly responsible for the therapeutic effects observed in these preparations. Most of the cinnamic acids, their esters, amides, aldehydes and alcohols, show significant growth inhibition against one or several bacterial and fungal species. Of particular interest is the potent antitubercular activity observed for some of these cinnamic derivatives, which may be amenable as future drugs for treating tuberculosis. This review intends to summarize the literature data on the antimicrobial activity of the natural cinnamic acids and related derivatives. In addition, selected hybrids between cinnamic acids and biologically active scaffolds with antimicrobial activity were also included. A comprehensive literature search was performed collating the minimum inhibitory concentration (MIC) of each cinnamic acid or derivative against the reported microorganisms. The MIC data allows the relative comparison between series of molecules and the derivation of structure-activity relationships.

  1. A homogeneous nucleic acid hybridization assay based on strand displacement.

    PubMed Central

    Vary, C P

    1987-01-01

    A homogeneous nucleic acid hybridization assay which is conducted in solution and requires no separation steps is described. The assay is based on the concept of strand displacement. In the strand displacement assay, an RNA "signal strand" is hybridized within a larger DNA strand termed the "probe strand", which is, in turn, complementary to the target nucleic acid of interest. Hybridization of the target nucleic acid with the probe strand ultimately results in displacement of the RNA signal strand. Strand displacement, therefore, causes conversion of the RNA from double to single-stranded form. The single-strand specificity of polynucleotide phosphorylase (EC 2.7.7.8) allows discrimination between double-helical and single-stranded forms of the RNA signal strand. As displacement proceeds, free RNA signal strands are preferentially phosphorolyzed to component nucleoside diphosphates, including adenosine diphosphate. The latter nucleotide is converted to ATP by pyruvate kinase(EC 2.7.1.40). Luciferase catalyzed bioluminescence is employed to measure the ATP generated as a result of strand displacement. Images PMID:3309890

  2. Nucleic acid hybridization-an alternative tool in diagnostic microbiology.

    PubMed

    Pettersson, U; Hyypiä, T

    1985-09-01

    The use of radioimmunoossays (RIAs) and enzyme-linked immunosorbent assays (ELISAs) has revolutionized diagnostic microbiology. Their high specificity and sensitivity make them versatile, they are simple to carry out either for direct detection of microorganisms in specimens or for serological diagnosis, and they can easily and reliably be standardized. Monoclonal antibodies have further improved these immunoassays. However, the development of simple and highly sensitive detection methods for nucleic acids has nevertheless promoted an interest also in diagnostic methods based on nucleic acid hybridization. Here Ulf Pettersson and Timo Hyypiä discuss methods which are likely to become a useful complement to the immunoassays in the near future.

  3. Bipolar lead-acid battery for hybrid vehicles

    NASA Astrophysics Data System (ADS)

    Saakes, M.; Woortmeijer, R.; Schmal, D.

    Within the framework of the European project bipolar lead-acid power source (BILAPS), a new production route is being developed for the bipolar lead-acid battery. The performance targets are 500 W kg -1, 30 Wh kg -1 and 100 000 power-assist life cycles (PALCs). The operation voltage of the battery can be, according to the requirements, 12, 36 V or any other voltage. Tests with recently developed 4 and 12 V prototypes, each of 30 Ah capacity have demonstrated that the PALC can be operated using 10 C discharge and 9 C charge peaks. The tests show no overvoltage or undervoltage problems during three successive test periods of 16 h with 8 h rest in between. The temperature stabilizes during these tests at 40-45 °C using a thermal-management system. The bipolar lead acid battery is operated at an initial 50% state-of-charge. During the tests, the individual cell voltages display only very small differences. Tests are now in progress to improve further the battery-management system, which has been developed at the cell level, during the period no PALCs are run in order to improve the hybrid behaviour of the battery. The successful tests show the feasibility of operating the bipolar lead-acid battery in a hybrid mode. The costs of the system are estimated to be much lower than those for nickel-metal-hydride or Li-ion based high-power systems. An additional advantage of the lead-acid system is that recycling of lead-acid batteries is well established.

  4. Improved DNA hybridization parameters by Twisted Intercalating Nucleic Acid (TINA).

    PubMed

    Schneider, Uffe Vest

    2012-01-01

    This thesis establishes oligonucleotide design rules and applications of a novel group of DNA stabilizing molecules collectively called Twisted Intercalating Nucleic Acid - TINA. Three peer-reviewed publications form the basis for the thesis. One publication describes an improved and rapid method for determination of DNA melting points and two publications describe the effects of positioning TINA molecules in parallel triplex helix and antiparallel duplex helix forming DNA structures. The third publication establishes that TINA molecules containing oligonucleotides improve an antiparallel duplex hybridization based capture assay's analytical sensitivity compared to conventionel DNA oligonucleotides. Clinical microbiology is traditionally based on pathogenic microorganisms' culture and serological tests. The introduction of DNA target amplification methods like PCR has improved the analytical sensitivity and total turn around time involved in clinical diagnostics of infections. Due to the relatively weak hybridization between the two strands of double stranded DNA, a number of nucleic acid stabilizing molecules have been developed to improve the sensitivity of DNA based diagnostics through superior binding properties. A short introduction is given to Watson-Crick and Hoogsteen based DNA binding and the derived DNA structures. A number of other nucleic acid stabilizing molecules are described. The stabilizing effect of TINA molecules on different DNA structures is discussed and considered in relation to other nucleic acid stabilizing molecules and in relation to future use of TINA containing oligonucleotides in clinical diagnostics and therapy. In conclusion, design of TINA modified oligonucleotides for antiparallel duplex helixes and parallel triplex helixes follows simple purpose dependent rules. TINA molecules are well suited for improving multiplex PCR assays and can be used as part of novel technologies. Future research should test whether combinations of TINA

  5. Nonenzymatic catalytic signal amplification for nucleic acid hybridization assays

    NASA Technical Reports Server (NTRS)

    Fan, Wenhong (Inventor); Cassell, Alan M. (Inventor); Han, Jie (Inventor)

    2006-01-01

    Devices, methods, and kits for amplifying the signal from hybridization reactions between nucleic acid probes and their cognate targets are presented. The devices provide partially-duplexed, immobilized probe complexes, spatially separate from and separately addressable from immobilized docking strands. Cognate target acts catalytically to transfer probe from the site of probe complex immobilization to the site of immobilized docking strand, generating a detectable signal. The methods and kits of the present invention may be used to identify the presence of cognate target in a fluid sample.

  6. A DNA origami nanorobot controlled by nucleic acid hybridization.

    PubMed

    Torelli, Emanuela; Marini, Monica; Palmano, Sabrina; Piantanida, Luca; Polano, Cesare; Scarpellini, Alice; Lazzarino, Marco; Firrao, Giuseppe

    2014-07-23

    A prototype for a DNA origami nanorobot is designed, produced, and tested. The cylindrical nanorobot (diameter of 14 nm and length of 48 nm) with a switchable flap, is able to respond to an external stimulus and reacts by a physical switch from a disarmed to an armed configuration able to deliver a cellular compatible message. In the tested design the robot weapon is a nucleic acid fully contained in the inner of the tube and linked to a single point of the internal face of the flap. Upon actuation the nanorobot moves the flap extracting the nucleic acid that assembles into a hemin/G-quadruplex horseradish peroxidase mimicking DNAzyme catalyzing a colorimetric reaction or chemiluminescence generation. The actuation switch is triggered by an external nucleic acid (target) that interacts with a complementary nucleic acid that is beard externally by the nanorobot (probe). Hybridization of probe and target produces a localized structural change that results in flap opening. The flap movement is studied on a two-dimensional prototype origami using Förster resonance energy transfer and is shown to be triggered by a variety of targets, including natural RNAs. The nanorobot has potential for in vivo biosensing and intelligent delivery of biological activators.

  7. Detection and isolation of nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, J.N.; Straume, T.; Bogen, K.T.

    1997-04-01

    A method for detecting a target nucleic acid sequence in a sample is provided using hybridization probes which competitively hybridize to a target nucleic acid. According to the method, a target nucleic acid sequence is hybridized to first and second hybridization probes which are complementary to overlapping portions of the target nucleic acid sequence, the first hybridization probe including a first complexing agent capable of forming a binding pair with a second complexing agent and the second hybridization probe including a detectable marker. The first complexing agent attached to the first hybridization probe is contacted with a second complexing agent, the second complexing agent being attached to a solid support such that when the first and second complexing agents are attached, target nucleic acid sequences hybridized to the first hybridization probe become immobilized on to the solid support. The immobilized target nucleic acids are then separated and detected by detecting the detectable marker attached to the second hybridization probe. A kit for performing the method is also provided. 7 figs.

  8. Detection and isolation of nucleic acid sequences using competitive hybridization probes

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    1997-01-01

    A method for detecting a target nucleic acid sequence in a sample is provided using hybridization probes which competitively hybridize to a target nucleic acid. According to the method, a target nucleic acid sequence is hybridized to first and second hybridization probes which are complementary to overlapping portions of the target nucleic acid sequence, the first hybridization probe including a first complexing agent capable of forming a binding pair with a second complexing agent and the second hybridization probe including a detectable marker. The first complexing agent attached to the first hybridization probe is contacted with a second complexing agent, the second complexing agent being attached to a solid support such that when the first and second complexing agents are attached, target nucleic acid sequences hybridized to the first hybridization probe become immobilized on to the solid support. The immobilized target nucleic acids are then separated and detected by detecting the detectable marker attached to the second hybridization probe. A kit for performing the method is also provided.

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

    PubMed

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

    2006-01-01

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

  10. Photodegradation of lipopolysaccharides and the inhibition of macrophage activation by anthraquinone-boronic acid hybrids.

    PubMed

    Takahashi, Daisuke; Miura, Takuya; Toshima, Kazunobu

    2012-08-07

    Target-selective photodegradation of 3-deoxy-D-manno-2-octulopyranosonic acid (KDO) was achieved without additives and under neutral conditions using a designed anthraquinone-boronic acid hybrid and long wavelength UV light irradiation. The hybrid can photodegrade lipopolysaccharides (LPS) and inhibit macrophage activation induced by LPS.

  11. Detection and isolation of nucleic acid sequences using a bifunctional hybridization probe

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    2000-01-01

    A method for detecting and isolating a target sequence in a sample of nucleic acids is provided using a bifunctional hybridization probe capable of hybridizing to the target sequence that includes a detectable marker and a first complexing agent capable of forming a binding pair with a second complexing agent. A kit is also provided for detecting a target sequence in a sample of nucleic acids using a bifunctional hybridization probe according to this method.

  12. Method for nucleic acid hybridization using single-stranded DNA binding protein

    DOEpatents

    Tabor, Stanley; Richardson, Charles C.

    1996-01-01

    Method of nucleic acid hybridization for detecting the presence of a specific nucleic acid sequence in a population of different nucleic acid sequences using a nucleic acid probe. The nucleic acid probe hybridizes with the specific nucleic acid sequence but not with other nucleic acid sequences in the population. The method includes contacting a sample (potentially including the nucleic acid sequence) with the nucleic acid probe under hybridizing conditions in the presence of a single-stranded DNA binding protein provided in an amount which stimulates renaturation of a dilute solution (i.e., one in which the t.sub.1/2 of renaturation is longer than 3 weeks) of single-stranded DNA greater than 500 fold (i.e., to a t.sub.1/2 less than 60 min, preferably less than 5 min, and most preferably about 1 min.) in the absence of nucleotide triphosphates.

  13. Nucleic Acid-Peptide Complex Phase Controlled by DNA Hybridization

    NASA Astrophysics Data System (ADS)

    Vieregg, Jeffrey; Lueckheide, Michael; Leon, Lorraine; Marciel, Amanda; Tirrell, Matthew

    When polyanions and polycations are mixed, counterion release drives formation of polymer-rich complexes that can either be solid (precipitates) or liquid (coacervates) depending on the properties of the polyelectrolytes. These complexes are important in many fields, from encapsulation of industrial polymers to membrane-free segregation of biomolecules such as nucleic acids and proteins. Condensation of long double-stranded DNA has been studied for several decades, but comparatively little attention has been paid to the polyelectrolyte behavior of oligonucleotides. We report here studies of DNA oligonucleotides (10 - 88 nt) complexed with polylysine (10 - 100 aa). Unexpectedly, we find that the phase of the resulting complexes is controlled by the hybridization state of the nucleic acid, with double-stranded DNA forming precipitates and single-stranded DNA forming coacervates. Stability increases with polyelectrolyte length and decreases with solution salt concentration, with complexes of the longer double-stranded polymers undergoing precipitate/coacervate/soluble transitions as ionic strength is increased. Mixing coacervates formed by complementary single-stranded oligonucleotides results in precipitate formation, raising the possibility of stimulus-responsive material design.

  14. Selection of fluorophore and quencher pairs for fluorescent nucleic acid hybridization probes.

    PubMed

    Marras, Salvatore A E

    2006-01-01

    With the introduction of simple and relatively inexpensive methods for labeling nucleic acids with nonradioactive labels, doors have been opened that enable nucleic acid hybridization probes to be used for research and development, as well as for clinical diagnostic applications. The use of fluorescent hybridization probes that generate a fluorescence signal only when they bind to their target enables real-time monitoring of nucleic acid amplification assays. The use of hybridization probes that bind to the amplification products in real-time markedly improves the ability to obtain quantitative results. Furthermore, real-time nucleic acid amplification assays can be carried out in sealed tubes, eliminating carryover contamination. Because fluorescent hybridization probes are available in a wide range of colors, multiple hybridization probes, each designed for the detection of a different nucleic acid sequence and each labeled with a differently colored fluorophore, can be added to the same nucleic acid amplification reaction, enabling the development of high-throughput multiplex assays. It is therefore important to carefully select the labels of hybridization probes, based on the type of hybridization probe used in the assay, the number of targets to be detected, and the type of apparatus available to perform the assay. This chapter outlines different aspects of choosing appropriate labels for the different types of fluorescent hybridization probes used with different types of spectrofluorometric thermal cyclers.

  15. Hybridization properties of long nucleic acid probes for detection of variable target sequences, and development of a hybridization prediction algorithm.

    PubMed

    Ohrmalm, Christina; Jobs, Magnus; Eriksson, Ronnie; Golbob, Sultan; Elfaitouri, Amal; Benachenhou, Farid; Strømme, Maria; Blomberg, Jonas

    2010-11-01

    One of the main problems in nucleic acid-based techniques for detection of infectious agents, such as influenza viruses, is that of nucleic acid sequence variation. DNA probes, 70-nt long, some including the nucleotide analog deoxyribose-Inosine (dInosine), were analyzed for hybridization tolerance to different amounts and distributions of mismatching bases, e.g. synonymous mutations, in target DNA. Microsphere-linked 70-mer probes were hybridized in 3M TMAC buffer to biotinylated single-stranded (ss) DNA for subsequent analysis in a Luminex® system. When mismatches interrupted contiguous matching stretches of 6 nt or longer, it had a strong impact on hybridization. Contiguous matching stretches are more important than the same number of matching nucleotides separated by mismatches into several regions. dInosine, but not 5-nitroindole, substitutions at mismatching positions stabilized hybridization remarkably well, comparable to N (4-fold) wobbles in the same positions. In contrast to shorter probes, 70-nt probes with judiciously placed dInosine substitutions and/or wobble positions were remarkably mismatch tolerant, with preserved specificity. An algorithm, NucZip, was constructed to model the nucleation and zipping phases of hybridization, integrating both local and distant binding contributions. It predicted hybridization more exactly than previous algorithms, and has the potential to guide the design of variation-tolerant yet specific probes.

  16. Hybridization properties of long nucleic acid probes for detection of variable target sequences, and development of a hybridization prediction algorithm

    PubMed Central

    Öhrmalm, Christina; Jobs, Magnus; Eriksson, Ronnie; Golbob, Sultan; Elfaitouri, Amal; Benachenhou, Farid; Strømme, Maria; Blomberg, Jonas

    2010-01-01

    One of the main problems in nucleic acid-based techniques for detection of infectious agents, such as influenza viruses, is that of nucleic acid sequence variation. DNA probes, 70-nt long, some including the nucleotide analog deoxyribose-Inosine (dInosine), were analyzed for hybridization tolerance to different amounts and distributions of mismatching bases, e.g. synonymous mutations, in target DNA. Microsphere-linked 70-mer probes were hybridized in 3M TMAC buffer to biotinylated single-stranded (ss) DNA for subsequent analysis in a Luminex® system. When mismatches interrupted contiguous matching stretches of 6 nt or longer, it had a strong impact on hybridization. Contiguous matching stretches are more important than the same number of matching nucleotides separated by mismatches into several regions. dInosine, but not 5-nitroindole, substitutions at mismatching positions stabilized hybridization remarkably well, comparable to N (4-fold) wobbles in the same positions. In contrast to shorter probes, 70-nt probes with judiciously placed dInosine substitutions and/or wobble positions were remarkably mismatch tolerant, with preserved specificity. An algorithm, NucZip, was constructed to model the nucleation and zipping phases of hybridization, integrating both local and distant binding contributions. It predicted hybridization more exactly than previous algorithms, and has the potential to guide the design of variation-tolerant yet specific probes. PMID:20864443

  17. Variability in coconut (Cocos nucifera L.) germplasm and hybrids for fatty acid profile of oil.

    PubMed

    Kumar, S Naresh

    2011-12-28

    Coconut oil, the main product of coconut fruit, is the richest source of glycerol and lauric acid and hence is called lauric oil. This paper reports the fatty acid profile of oil from 60 Talls, 14 Dwarfs, and 34 hybrids. These include collections from 13 countries covering a large coconut-growing area of the world, apart from the indigenous ones. Capillary gas chromatography analysis of oil indicated a wider variation for the fatty acid profile than earlier reported. Apart from this, for the first time other fatty acids such as behenic and lignoceric acids were detected. Oil from cultivars and hybrids of coconut has significantly differed, particularly for commercially important fatty acids such as lauric acid and unsaturated fatty acids. However, coconut oil seems to have a conserved fatty acid profile, mainly because of low unsaturated fatty acids, indicating the possibility of grouping cultivars on the basis of their fatty acid profiles. The cluster analysis based on fatty acid profile indicated grouping together of geographically and typically closely related cultivars. Cultivars with high concentrations of specific fatty acids can be of potential use for industrial exploitation, whereas those with high concentrations of short- and medium-chain fatty acids and unsaturated fatty acids are more suitable for human consumption. Cultivars and hybrids with high and low values for each of the fatty acids are also identified.

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

    PubMed

    Li, Xu; Morgenroth, Eberhard; Raskin, Lutgarde

    2008-12-01

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

  19. Design, Synthesis and Microbiological Evaluation of Ampicillin Tetramic acid Hybrid Antibiotics

    PubMed Central

    Cherian, Philip T.; Deshpande, Aditi; Cheramie, Martin N.; Bruhn, David F.; Hurdle, Julian G.; Lee, Richard E.

    2016-01-01

    Exploiting iron-uptake pathways by conjugating β-lactam antibiotics with iron-chelators such as catechol and hydroxamic acid is a proven strategy to overcome permeability-related resistance in Gram-negative bacteria. Since naturally occurring iron chelating tetramic acids have not been previously examined for this purpose, an exploratory series of novel ampicillin-tetramic acid hybrids that structurally resemble ureidopenicillins was designed and synthesized. The new analogs were evaluated for the ability to chelate iron and their MIC activities determined against a representative panel of clinically significant bacterial pathogens. The tetramic acid β-lactam hybrids demonstrated a high affinity to iron in the order of 10−30 M3. The hybrids were less active against Gram-positive bacteria. However, against Gram-negative bacteria, their activity was species dependent with several hybrids displaying improved activity over ampicillin against wild-type Pseudomonas aeruginosa. The anti-Gram-negative activities of the hybrids improved in the presence of clavulanic acid revealing that the tetramic acid moiety did not provide added protection against β-lactamases. Additionally, the hybrids were found to be efflux pump substrates as their activities markedly improved against pump-inactivated strains. Unlike the catechol and hydroxamic acid siderophore β-lactam conjugates, the activities of the hybrids did not improve under iron-deficient conditions. These results suggest that the tetramic acid hybrids gain permeability via different membrane receptors, or they are out competed by native bacterial siderophores with stronger affinities for iron. This study provides a foundation for the further exploitation of the tetramic acid moiety to achieve novel β-lactam anti-Gram-negative agents, providing that efflux and β-lactamase mediated resistance is addressed. PMID:27189120

  20. Design, synthesis and microbiological evaluation of ampicillin-tetramic acid hybrid antibiotics.

    PubMed

    Cherian, Philip T; Deshpande, Aditi; Cheramie, Martin N; Bruhn, David F; Hurdle, Julian G; Lee, Richard E

    2017-01-01

    Exploiting iron-uptake pathways by conjugating β-lactam antibiotics with iron-chelators, such as catechol and hydroxamic acid is a proven strategy to overcome permeability-related resistance in Gram-negative bacteria. As naturally occurring iron-chelating tetramic acids have not been previously examined for this purpose, an exploratory series of novel ampicillin-tetramic acid hybrids that structurally resemble ureidopenicillins was designed and synthesized. The new analogs were evaluated for the ability to chelate iron and their MIC activities determined against a representative panel of clinically significant bacterial pathogens. The tetramic acid β-lactam hybrids demonstrated a high affinity to iron in the order of 10(-30) M(3). The hybrids were less active against Gram-positive bacteria. However, against Gram-negative bacteria, their activity was species dependent with several hybrids displaying improved activity over ampicillin against wild-type Pseudomonas aeruginosa. The anti-Gram-negative activities of the hybrids improved in the presence of clavulanic acid revealing that the tetramic acid moiety did not provide added protection against β-lactamases. In addition, the hybrids were found to be efflux pump substrates as their activities markedly improved against pump-inactivated strains. Unlike the catechol and hydroxamic acid siderophore β-lactam conjugates, the activities of the hybrids did not improve under iron-deficient conditions. These results suggest that the tetramic acid hybrids gain permeability via different membrane receptors, or they are outcompeted by native bacterial siderophores with stronger affinities for iron. This study provides a foundation for the further exploitation of the tetramic acid moiety to achieve novel β-lactam anti-Gram-negative agents, providing that efflux and β-lactamase mediated resistance is addressed.

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

    PubMed

    Marras, Salvatore A E

    2008-03-01

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

  2. Synthesis of organic/inorganic hybrid gel with acid activated clay after γ-ray radiation.

    PubMed

    Kim, Donghyun; Lee, Hoik; Sohn, Daewon

    2014-08-01

    A hybrid gel was prepared from acid activated clay (AA clay) and acrylic acid by gamma ray irradiation. Irradiated inorganic particles which have peroxide groups act as initiator because it generates oxide radicals by increasing temperature. Inorganic nanoparticles which are rigid part in hybrid gel also contribute to increase the mechanical property as a crosslinker. We prepared two hybrid gels to compare the effect of acid activated treatment of clay; one is synthesized with raw clay particles and another is synthesized with AA clay particles. The composition and structure of AA clay particles and raw clay particles were confirmed by X-ray diffraction (XRD), X-ray fluorescence instrument and surface area analyzer. And chemical and physical property of hybrid gel with different ratios of acrylic acid and clay particle was tested by Raman spectroscope and universal testing machine (UTM). The synthesized hydrogel with 76% gel contents can elongated approximately 1000% of its original size.

  3. Catalytic performance of hybrid nanocatalyst for levulinic acid production from glucose

    NASA Astrophysics Data System (ADS)

    Ya'aini, Nazlina; Amin, Nor Aishah Saidina

    2012-11-01

    Levulinic acid is one of the potential and versatile biomass-derived chemicals. Product analysis via HPLC revealed that the heterogeneous dehydration of glucose over hybrid nanocatalyst exhibited better performance compared to single catalyst. Hybrid nanocatalyst containing H-Y zeolite and CrCl3 could substitute homogenous acid catalyst for attaining high levulinic acid yield. Different CrC3 and H-Y zeolite weight ratios of 1:1, 1:2 and 2:1 were prepared according to the wetness impregnation method. The hybrid catalyst with a 1:1 weight ratio performed better compared to others with the highest levulinic acid yield reported (93.5%) at 140 °C, 180 min reaction time, 0.1 g catalyst loading and 0.1 g glucose feed. Characterization results revealed that properties such as surface area, mesoporosity and acidic strength of the catalyst have significant effects on glucose dehydration for levulinic acid production.

  4. Oligonucleotide microarrays with stem-loop probes: enhancing the hybridization of nucleic acids for sensitive analysis.

    PubMed

    Mahajan, Shweta; Swami, Archana; Sethi, Dalip; Kumar, P; Gupta, K C

    2008-06-15

    We have demonstrated that the dynamics of nucleic acid hybridization in microarrays depend on the physical structure of immobilized probes. We have immobilized oligonucleotide-3'-phosphates with and without stem-loop structure on epoxylated glass surface, followed by hybridization under different conditions, viz., hybridization buffer, pH condition, temperature and ionic strength. In a comparative study, we have established that array constructed using probes with stem-loop structure displayed approximately 2.2 times higher hybridization signals than the probes without it. The stem-loop DNA array format is simple and flexible in design and thus potentially useful in various DNA diagnostic tests.

  5. Lead-acid batteries in micro-hybrid vehicles

    NASA Astrophysics Data System (ADS)

    Albers, Joern; Meissner, Eberhard; Shirazi, Sepehr

    More and more vehicles hit the European automotive market, which comprise some type of micro-hybrid functionality to improve fuel efficiency and reduce emissions. Most carmakers already offer at least one of their vehicles with an optional engine start/stop system, while some other models are sold with micro-hybrid functions implemented by default. But these car concepts show a wide variety in detail-the term "micro-hybrid" may mean a completely different functionality in one vehicle model compared to another. Accordingly, also the battery technologies are not the same. There is a wide variety of batteries from standard flooded and enhanced flooded to AGM which all are claimed to be "best choice" for micro-hybrid applications. A technical comparison of micro-hybrid cars available on the European market has been performed. Different classes of cars with different characteristics have been identified. Depending on the scope and characteristics of micro-hybrid functions, as well as on operational strategies implemented by the vehicle makers, the battery operating duties differ significantly between these classes of vehicles. Additional laboratory investigations have been carried out to develop an understanding of effects observed in batteries operated in micro-hybrid vehicles pursuing different strategies, to identify limitations for applications of different battery technologies.

  6. Optimization of levulinic acid from lignocellulosic biomass using a new hybrid catalyst.

    PubMed

    Ya'aini, Nazlina; Amin, Nor Aishah Saidina; Asmadi, Mohd

    2012-07-01

    Conversion of glucose, empty fruit bunch (efb) and kenaf to levulinic acid over a new hybrid catalyst has been investigated in this study. The characterization and catalytic performance results revealed that the physico-chemical properties of the new hybrid catalyst comprised of chromium chloride and HY zeolite increased the levulinic acid production from glucose compared to the parent catalysts. Optimization of the glucose conversion process using two level full factorial designs (2(3)) with two center points reported 55.2% of levulinic acid yield at 145.2 °C, 146.7 min and 12.0% of reaction temperature, reaction time and catalyst loading, respectively. Subsequently, the potential of efb and kenaf for producing levulinic acid at the optimum conditions was established after 53.2% and 66.1% of efficiencies were reported. The observation suggests that the hybrid catalyst has a potential to be used in biomass conversion to levulinic acid.

  7. Cassava interspecific hybrids with increased protein content and improved amino acid profiles.

    PubMed

    Gomes, P T C; Nassar, N M A

    2013-04-12

    Cassava (Manihot esculenta) is a principal food for large populations of poor people in the tropics and subtropics. Its edible roots are poor in protein and lack several essential amino acids. Interspecific hybrids may acquire high protein characteristics from wild species. We analyzed 19 hybrids of M. esculenta with its wild relative, M. oligantha, for crude protein, amino acid profile, and total cyanide. Some hybrids produced roots with high protein content of up to 5.7%, while the common cultivar that we examined had just 2.3% crude protein. The essential amino acids alanine, phenylalanine, and valine were detected in the hybrids. The sulfur-containing amino acids cysteine and methionine were found at relatively high concentrations in the roots of 4 hybrids. The proportion of lysine in one hybrid was 20 times higher than in the common cultivar. The levels of total cyanide ranged from 19.73 to 172.56 mg/kg and most of the roots analyzed were classified as "non-toxic" and "low toxic". Furthermore, 2 progenies showed reasonable levels of cyanide, but higher protein content and amino acid profile more advantageous than the common cassava.

  8. Pyrrolidinyl peptide nucleic acid homologues: effect of ring size on hybridization properties.

    PubMed

    Mansawat, Woraluk; Vilaivan, Chotima; Balázs, Árpád; Aitken, David J; Vilaivan, Tirayut

    2012-03-16

    The effect of ring size of four- to six-membered cyclic β-amino acid on the hybridization properties of pyrrolidinyl peptide nucleic acid with an alternating α/β peptide backbone is reported. The cyclobutane derivatives (acbcPNA) show the highest T(m) and excellent specificity with cDNA and RNA.

  9. Cativic acid-caffeic acid hybrid exerts cytotoxic effects and induces apoptotic death in human neuroblastoma cells.

    PubMed

    Alza, Natalia P; Murray, Ana P; Salvador, Gabriela A

    2017-09-05

    The development of hybrids from natural products is a promising strategy for drug discovery. In cancer therapy, there is a need to discover novel agents that can induce apoptosis in cancer cells. To contribute to this field of interest, we investigated the effect of a synthetic hybrid from cativic acid and caffeic acid (5) on viability, proliferation, and apoptosis in human neuroblastoma cells (IMR-32). Three hybrids were prepared via Mitsunobu esterification from 17-hydroxycativic acid (1) and natural phenols. Cell viability was analyzed by MTT assay. SYTOX green and LDH leakage were used to determine the cytotoxic effect. Caspase-3 activity, cell cycle phases, and proliferation were analyzed in order to characterize the biological effects of hybrid 5. The mitogen-activated protein kinase (MAPK) status was evaluated for elucidating the potential mechanisms involved in hybrid 5 effect. Hybrid 5 reduced the viability of IMR-32 cells in a time- and concentration-dependent manner (IC50 = 18.0 ± 1.3 μM) as a result of its antiproliferative effect through changes in the cell cycle distribution and induction of apoptosis associated with activation of caspase-3. Exposure to 5 triggered ERK1/2 activation and nuclear translocation. Hybrid 5 also promoted an increase in nuclear localization of the transcription factor c-Jun. Inhibition of ERK1/2 and JNK potentiated 5-induced inhibition of IMR-32 viability. Hybrid 5 displays cell growth inhibition by promoting cell cycle arrest and apoptosis, through ERK1/2 and JNK participation.

  10. Producing a trimethylpentanoic acid using hybrid polyketide synthases

    DOEpatents

    Katz, Leonard; Fortman, Jeffrey L; Keasling, Jay D

    2014-10-07

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing trimethylpentanoic acid. The present invention also provides for a host cell comprising the PKS and when cultured produces the trimethylpentanoic acid. The present invention also provides for a method of producing the trimethylpentanoic acid, comprising: providing a host cell of the present invention, and culturing said host cell in a suitable culture medium such that the trimethylpentanoic acid is produced, optionally isolating the trimethylpentanoic acid, and optionally, reducing the isolated trimethylpentanoic acid into a trimethylpentanol or an iso-octane.

  11. Nucleic acid hybridization with RNA immobilized on filter paper.

    NASA Technical Reports Server (NTRS)

    Saxinger, W. C.; Ponnamperuma, C.; Gillespie, D.

    1972-01-01

    RNA has been immobilized in a manner suitable for use in molecular hybridization experiments with dissolved RNA or DNA by a nonaqueous solid-phase reaction with carbonyldiimidazole and RNA 'dry coated' on cellulose or, preferably, on previously activated phosphocellulose filters. Immobilization of RNA does not appear to alter its chemical character or cause it to acquire affinity for unspecific RNA or DNA. The versatility and efficiency of this method make it potentially attractive for use in routine analytical or preparative hybridization experiments, among other applications.

  12. Nucleic acid hybridization with RNA immobilized on filter paper.

    NASA Technical Reports Server (NTRS)

    Saxinger, W. C.; Ponnamperuma, C.; Gillespie, D.

    1972-01-01

    RNA has been immobilized in a manner suitable for use in molecular hybridization experiments with dissolved RNA or DNA by a nonaqueous solid-phase reaction with carbonyldiimidazole and RNA 'dry coated' on cellulose or, preferably, on previously activated phosphocellulose filters. Immobilization of RNA does not appear to alter its chemical character or cause it to acquire affinity for unspecific RNA or DNA. The versatility and efficiency of this method make it potentially attractive for use in routine analytical or preparative hybridization experiments, among other applications.

  13. Design of an innovative lead-acid battery for electric and hybrid vehicles

    NASA Astrophysics Data System (ADS)

    Baxa, M. S.; Weinlein, C. E.

    An innovative lead acid battery was designed specifically for use in a hybrid car. The battery has exceeded all of the minimum performance goals, and in many cases the optimistic target goals. The eV-1300, which features electrolyte circulation, has excellent energy density, power characteristics, efficiency, and cycle life. It is a good candidate battery for other hybrid vehicles and electric vehicles which require a relatively small cell size.

  14. Design and Synthesis of Novel Isoxazole Tethered Quinone-Amino Acid Hybrids

    PubMed Central

    Ravi Kumar, P.; Sambaiah, M.; Kandula, Venu; Payili, Nagaraju; Jaya Shree, A.; Yennam, Satyanarayana

    2014-01-01

    A new series of isoxazole tethered quinone-amino acid hybrids has been designed and synthesized involving 1,3-dipolar cycloaddition reaction followed by an oxidation reaction using cerium ammonium nitrate (CAN). Using this method, for the first time various isoxazole tethered quinone-phenyl alanine and quinone-alanine hybrids were synthesized from simple commercially available 4-bromobenzyl bromide, propargyl bromide, and 2,5-dimethoxybenzaldehyde in good yield. PMID:25709839

  15. Visual, base-specific detection of nucleic acid hybridization using polymerization-based amplification.

    PubMed

    Hansen, Ryan R; Johnson, Leah M; Bowman, Christopher N

    2009-03-15

    Polymerization-based signal amplification offers sensitive visualization of biotinylated biomolecules functionalized to glass microarrays in a manner suitable for point-of-care use. Here we report using this method for visual detection of multiplexed nucleic acid hybridizations from complex media and develop an application toward point mutation detection and single nucleotide polymorphism (SNP) typing. Primer extension reactions were employed to label selectively and universally all complementary surface DNA hybrids with photoinitiators, permitting simultaneous and dynamic photopolymerization from positive sites to 0.5-nM target concentrations. Dramatic improvements in signal ratios between complementary and mismatched hybrids enabled visual discrimination of single base differences in KRAS codon-12 biomarkers.

  16. Synthesis of hybrid hydrazino peptides: protected vs unprotected chiral α-hydrazino acids.

    PubMed

    Suć, Josipa; Jerić, Ivanka

    2015-01-01

    Peptidomimetics based on hydrazino derivatives of α-amino acids represent an important class of peptidic foldamers with promising biological activities, like protease inhibition and antimicrobial activity. However, the lack of straightforward method for the synthesis of optically pure hydrazino acids and efficient incorporation of hydrazino building blocks into peptide sequence hamper wider exploitation of hydrazino peptidomimetics. Here we described the utility of N (α)-benzyl protected and unprotected hydrazino derivatives of natural α-amino acids in synthesis of peptidomimetics. While incorporation of N (α)-benzyl-hydrazino acids into peptide chain and deprotection of benzyl moiety proceeded with difficulties, unprotected hydrazino acids allowed fast and simple construction of hybrid peptidomimetics.

  17. The chromatin remodeler DDM1 promotes hybrid vigor by regulating salicylic acid metabolism.

    PubMed

    Zhang, Qingzhu; Li, Yanqiang; Xu, Tao; Srivastava, Ashish Kumar; Wang, Dong; Zeng, Liang; Yang, Lan; He, Li; Zhang, Heng; Zheng, Zhimin; Yang, Dong-Lei; Zhao, Cheng; Dong, Juan; Gong, Zhizhong; Liu, Renyi; Zhu, Jian-Kang

    2016-01-01

    In plants, hybrid vigor is influenced by genetic and epigenetic mechanisms; however, the molecular pathways are poorly understood. We investigated the potential contributions of epigenetic regulators to heterosis in Arabidposis and found that the chromatin remodeler DECREASED DNA METHYLATION 1 (DDM1) affects early seedling growth heterosis in Col/C24 hybrids. ddm1 mutants showed impaired heterosis and increased expression of non-additively expressed genes related to salicylic acid metabolism. Interestingly, our data suggest that salicylic acid is a hormetic regulator of seedling growth heterosis, and that hybrid vigor arises from crosses that produce optimal salicylic acid levels. Although DNA methylation failed to correlate with differential non-additively expressed gene expression, we uncovered DDM1 as an epigenetic link between salicylic acid metabolism and heterosis, and propose that the endogenous salicylic acid levels of parental plants can be used to predict the heterotic outcome. Salicylic acid protects plants from pathogens and abiotic stress. Thus, our findings suggest that stress-induced hormesis, which has been associated with increased longevity in other organisms, may underlie specific hybrid vigor traits.

  18. The chromatin remodeler DDM1 promotes hybrid vigor by regulating salicylic acid metabolism

    PubMed Central

    Zhang, Qingzhu; Li, Yanqiang; Xu, Tao; Srivastava, Ashish Kumar; Wang, Dong; Zeng, Liang; Yang, Lan; He, Li; Zhang, Heng; Zheng, Zhimin; Yang, Dong-Lei; Zhao, Cheng; Dong, Juan; Gong, Zhizhong; Liu, Renyi; Zhu, Jian-Kang

    2016-01-01

    In plants, hybrid vigor is influenced by genetic and epigenetic mechanisms; however, the molecular pathways are poorly understood. We investigated the potential contributions of epigenetic regulators to heterosis in Arabidposis and found that the chromatin remodeler DECREASED DNA METHYLATION 1 (DDM1) affects early seedling growth heterosis in Col/C24 hybrids. ddm1 mutants showed impaired heterosis and increased expression of non-additively expressed genes related to salicylic acid metabolism. Interestingly, our data suggest that salicylic acid is a hormetic regulator of seedling growth heterosis, and that hybrid vigor arises from crosses that produce optimal salicylic acid levels. Although DNA methylation failed to correlate with differential non-additively expressed gene expression, we uncovered DDM1 as an epigenetic link between salicylic acid metabolism and heterosis, and propose that the endogenous salicylic acid levels of parental plants can be used to predict the heterotic outcome. Salicylic acid protects plants from pathogens and abiotic stress. Thus, our findings suggest that stress-induced hormesis, which has been associated with increased longevity in other organisms, may underlie specific hybrid vigor traits. PMID:27551435

  19. Nanofiltration, bipolar electrodialysis and reactive extraction hybrid system for separation of fumaric acid from fermentation broth.

    PubMed

    Prochaska, Krystyna; Staszak, Katarzyna; Woźniak-Budych, Marta Joanna; Regel-Rosocka, Magdalena; Adamczak, Michalina; Wiśniewski, Maciej; Staniewski, Jacek

    2014-09-01

    A novel approach based on a hybrid system allowing nanofiltration, bipolar electrodialysis and reactive extraction, was proposed to remove fumaric acid from fermentation broth left after bioconversion of glycerol. The fumaric salts can be concentrated in the nanofiltration process to a high yield (80-95% depending on pressure), fumaric acid can be selectively separated from other fermentation components, as well as sodium fumarate can be conversed into the acid form in bipolar electrodialysis process (stack consists of bipolar and anion-exchange membranes). Reactive extraction with quaternary ammonium chloride (Aliquat 336) or alkylphosphine oxides (Cyanex 923) solutions (yield between 60% and 98%) was applied as the final step for fumaric acid recovery from aqueous streams after the membrane techniques. The hybrid system permitting nanofiltration, bipolar electrodialysis and reactive extraction was found effective for recovery of fumaric acid from the fermentation broth.

  20. Interactions of hybrid gold-tannic acid nanoparticles with human serum albumin.

    PubMed

    Sekowski, Szymon; Tomaszewska, Emilia; Soliwoda, Katarzyna; Celichowski, Grzegorz; Grobelny, Jaroslaw

    2017-01-01

    Nanoparticles present a wide spectrum of chemical, biological, and physical properties which result in their usage in many branches of science. We present an investigation of the interaction between human serum albumin and hybrid gold-tannic acid nanoparticles synthesized via a chemical reduction method. The results obtained demonstrate that tannic acid can be a very effective reducing and stabilizing agent and allows monodisperse hybrid gold nanomaterial to be obtained. The synthesized hybrid gold-tannic acid nanoparticles strongly interact with human serum albumin by formation of protein-corona complexes. The strength of the interaction with albumin depends on the number of tannic acid molecules on the surface of the nanoparticles and the presence of citric acid. Nanoparticles of large size and rich in tannic acid react more strongly with the protein [K SV = (8.00 ± 0.2) × 10(5) M(-1)] compared with smaller ones [K SV = (6.83 ± 0.5) × 10(4) M(-1)] containing citric acid and low concentration of tannic acid.

  1. Arginine-responsive terbium luminescent hybrid sensors triggered by two crown ether carboxylic acids.

    PubMed

    Jiang, Lasheng; Tang, Ke; Ding, Xiaoping; Wang, Qianming; Zhou, Zhan; Xiao, Rui

    2013-12-01

    Crown ether carboxylic acids constitute main building blocks for the synthesis of terbium containing covalent cross-linked luminescent materials. Both the complexes and the hybrid nanomaterials could exhibit remarkable green emissions in pure water. More importantly, they were found to have a profound effect on the luminescence responses to arginine compared with glutamic acid, histidine, tryptophan, threonine, tyrosine and phenylalanine in aqueous environment. The present study provided the possibility of using a host-guest mechanism as a way of signal transduction based on lanthanide supramolecular hybrid materials. © 2013.

  2. Synthesis of high erucic acid rapeseed (Brassica napus L.) somatic hybrids with improved agronomic characters.

    PubMed

    Heath, D W; Earle, E D

    1995-11-01

    Novel Brassica napus somatic hybrids have been created through protoplast fusion of B. oleracea var. botrytis and B. rapa var. oleifera genotypes selected for high erucic acid (22:1) content in the seed oil. Fifty amphidiploids (aacc) and one putative hexaploid (aacccc) hybrid were recovered in one fusion experiment. Conversely, only one amphidiploid and numerous regenerates with higher DNA contents were produced in a similar fusion using a different B. rapa partner. Hybridity was confirmed by morphology, isozyme expression, flow cytometry, and DNA hybridization. Analysis of organellar DNA revealed a distinct bias toward the inheritance of chloroplasts from the B. rapa (aa) genome. All amphidiploids set self-pollinated seed. A erucic acid content as high as 57.4% was found in the seed oil of one regenerated plant. Fatty acid composition was stable in the R1 generation and was coupled with increased female fertility. Other novel agronomic characters in the hybrids recovered include large seed size, lodging resistance, and non-shattering seed pods.

  3. Biomimetic growth of gallic acid-ZnO hybrid assemblies and their applications

    NASA Astrophysics Data System (ADS)

    Sarker, Nazmul H.; Barnaby, Stacey N.; Fath, Karl R.; Frayne, Stephen H.; Nakatsuka, Nako; Banerjee, Ipsita A.

    2012-03-01

    In this study, we probed the biomimetic formation of gallic acid (GA)-ZnO nanoparticle hybrids. It was found that the morphologies formed were dependent upon pH values, resulting in GA-ZnO hybrids of varying shapes such as micro or nanoplates or fibers. The formed supramolecular GA-ZnO hybrids were found to be luminescent as indicated by confocal microscopy and were utilized for the photocatalytic degradation of the organic dye methylene blue. We also explored the bactericidal effects of the hybrids on Staphylococcus aureus ( S. aureus) as well as Escherichia Coli ( E. Coli). Thus, we have developed a new class of shape-controlled nanohybrid assemblies via mild, green synthetic methods that may be utilized for photocatalytic degradation for environmental remediation as well as for antibacterial applications.

  4. Novel blue-light-emitting hybrid materials based on oligothiophene acids and ZnO

    NASA Astrophysics Data System (ADS)

    Jiu, Tonggang; Liu, Huibiao; Fu, Liming; He, Xiaorong; Wang, Ning; Li, Yuliang; Ai, Xicheng; Zhu, Daoben

    2004-11-01

    Novel blue-light-emitting materials based on ZnO and 2,2'-bithiophene-5,5'-dicarboxylic acid (DTDA), 4',3″-dipentyl-5,2': 5',2″: 5″,2‴-quaterthiophene-2,5‴-dicarboxylic acid (QTDA) have been prepared. The hybrid materials show that the PL λmax are at 450 and 425 nm for DTDA-ZnO and QTDA-ZnO, respectively.

  5. Probing the transition state for nucleic acid hybridization using phi-value analysis.

    PubMed

    Kim, Jandi; Shin, Jong-Shik

    2010-04-27

    Genetic regulation by noncoding RNA elements such as microRNA and small interfering RNA (siRNA) involves hybridization of a short single-stranded RNA with a complementary segment in a target mRNA. The physical basis of the hybridization process between the structured nucleic acids is not well understood primarily because of the lack of information about the transition-state structure. Here we use transition-state theory, inspired by phi-value analysis in protein folding studies, to provide quantitative analysis of the relationship between changes in the secondary structure stability and the activation free energy. Time course monitoring of the hybridization reaction was performed under pseudo-steady-state conditions using a single fluorophore. The phi-value analysis indicates that the native secondary structure remains intact in the transition state. The nativelike transition state was confirmed via examination of the salt dependence of the hybridization kinetics, indicating that the number of sodium ions associated with the transition state was not substantially affected by changes in the native secondary structure. These results propose that hybridization between structured nucleic acids undergoes a transition state leading to formation of a nucleation complex and then is followed by sequential displacement of preexisting base pairings involving successive small energy barriers. The proposed mechanism might provide new insight into physical processes during small RNA-mediated gene silencing, which is essential to selection of a target mRNA segment for siRNA design.

  6. Permeabilization of mycolic-acid-containing actinomycetes for in situ hybridization with fluorescently labelled oligonucleotide probes.

    PubMed

    Macnaughton, S J; O'Donnell, A G; Embley, T M

    1994-10-01

    The application of whole-cell hybridization using labelled oligonucleotide probes in microbial systematics and ecology is limited by difficulties in permeabilizing many Gram-positive organisms. In this investigation paraformaldehyde treatment, acid methanolysis and acid hydrolysis were evaluated as a means of permeabilizing mycolic-acid-containing actinomycetes prior to hybridization with a fluorescently labelled oligonucleotide probe designed to bind to a conserved sequence of bacterial 16S rRNA. Methods were evaluated on stationary-phase cultures of Gordona bronchialis, Mycobacterium fortuitum, Nocardia asteroides, N. brasiliensis, Rhodococcus equi, R. erythropolis, R. fascians, R. rhodochrous and Tsukamurella paurometabola, none of which could be probed following 4% (w/v) paraformaldehyde fixation. For comparison and to test the general applicability of mild acid pretreatments, Bacillus subtilis, Lactobacillus plantarum, Escherichia coli and Pseudomonas putida were also studied. The data showed that most of the mycolic-acid-containing organisms were successfully permeabilized by mild acid hydrolysis in 1 M HCl at 37 degrees C. Cells were treated for different lengths of time. In general, the mycolic-acid-containing organisms required between 30 and 50 min hydrolysis, whereas B. subtilis, E. coli and P. putida were rendered permeable in only 10 min. Interestingly, L. plantarum could not be permeabilized using acid hydrolysis even after 60 min exposure to 1 M HCl.

  7. [Fatty acids profile characterization of white maize hybrids grown in Venezuela].

    PubMed

    Alezones, Jesús; Avila, Manuel; Chassaigne, Alberto; Barrientos, Venancio

    2010-12-01

    In Venezuela, white corn is the most important crop regarding production, harvest area and consumption. One of its main by-products is corn oil, whose positive effect on health caused by the high content of unsaturated fatty acids has been widely recognized. In order to characterize the fatty acids profile of twelve white grained maize hybrids extensively grown in Venezuela, and the effect that divergent localities has on this profile, three semi commercial scale trials where established in Portuguesa, Yaracuy and Guárico states. Proportions of the main fatty acids in the raw oil of the different grain samples were determined using gas chromatography. Significant differences (p < 0,01) between hybrids were found for arachidic, palmitic, stearic, oleic, gadoleic and linoleic acids; non significant differences were found for linolenic acid. Significant differences between localities were found for all the fatty acids evaluated. High and significant correlations between fatty acids content were found; the most important relations were: linoleic-oleic (Rho = -0,98**), arachidic-palmitic (Rho = -0,61**), linoleic-stearic (Rho = -0,61**) and oleic-stearic (Rho = 0,58**). Corn produced in Venezuela presents lower levels of linoleic and higher levels of palmitic, stearic and oleic acids than the levels found in temperate corn. These differences involve significant changes in the nutritional properties of Venezuelan corn oil that should be considered in the development of new cultivars and industrial processes for oil production.

  8. Molecular hybridization between rat liver deoxyribonucleic acid and complementary ribonucleic acid

    PubMed Central

    Melli, Marialuisa; Bishop, J. O.

    1970-01-01

    RNA (cRNA) was synthesized in vitro on a template of rat liver DNA and its hybridization with rat liver DNA was studied by using the nitrocellulose-filter method. Sonication of the DNA diminished its apparent capacity to hybridize with RNA by about 50%. This is not due to cross-linkage of DNA molecules, because it could be shown that less than 2% of the sonicated DNA was cross-linked. The effect is due instead to the small size of the sonicated DNA molecules. Below a single-stranded molecular weight of 5×105 the DNA showed a progressive loss of capacity to hybridize with decrease in molecular weight. Evidence is presented suggesting that the apparently diminished capacity of the DNA to hybridize is due to loss of hybridized DNA from the membrane filters. When cRNA at concentrations of up to 25μg/ml is annealed with sonicated total DNA, an apparent hybridization saturation value is found at which about 2.5% of the DNA is hybridized with RNA. Increasing the cRNA concentration tenfold brought about the hybridization of a second component of the DNA approximately equal in amount to the first. The renaturation of rat liver DNA was studied by measuring the fall in the extinction at 260nm and two different components of renaturation were observed within the reiterated fraction of DNA. By hybridizing cRNA with different fractions of rat DNA the two components of the hybridization curve are shown to correspond to the two components of the renaturation curve. The conclusion is drawn that at a cRNA concentration of 250μg/ml most of the reiterated fraction of rat liver DNA is hybridized after annealing for 16h under standard conditions (0.30m-sodium chloride–30mm-sodium citrate at 65°C). Even with such a high cRNA concentration little or no hybridization of the slowly renaturing DNA fraction occurs. It is suggested that the most highly reiterated DNA component is poorly transcribed in vitro. PMID:5493851

  9. A collagen-poly(lactic acid-co-ɛ-caprolactone) hybrid scaffold for bladder tissue regeneration.

    PubMed

    Engelhardt, Eva-Maria; Micol, Lionel A; Houis, Stephanie; Wurm, Florian M; Hilborn, Jöns; Hubbell, Jeffrey A; Frey, Peter

    2011-06-01

    Scaffold materials should favor cell attachment and proliferation, and provide designable 3D structures with appropriate mechanical strength. Collagen matrices have proven to be beneficial scaffolds for tissue regeneration. However, apart from small intestinal submucosa, they offer a limited mechanical strength even if crosslinking can enhance their mechanical properties. A more cell-friendly way to increase material strength is to combine synthetic polymer meshes with plastic compressed collagen gels. This work describes the potential of plastic compressed collagen-poly(lactic acid-co-ɛ-caprolactone) (PLAC) hybrids as scaffolds for bladder tissue regeneration. Human bladder smooth muscle and urothelial cells were cultured on and inside collagen-PLAC hybrids in vitro. Scaffolds were analyzed by electron microscopy, histology, immunohistochemistry, and AlamarBlue assay. Both cell types proliferated in and on the hybrid, forming dense cell layers on top after two weeks. Furthermore, hybrids were implanted subcutaneously in the backs of nude mice. Host cell infiltration, scaffold degradation, and the presence of the seeded bladder cells were analyzed. Hybrids showed a lower inflammatory reaction in vivo than PLAC meshes alone, and first signs of polymer degradation were visible at six months. Collagen-PLAC hybrids have potential for bladder tissue regeneration, as they show efficient cell seeding, proliferation, and good mechanical properties.

  10. The vector homology problem in diagnostic nucleic acid hybridization of clinical specimens.

    PubMed Central

    Ambinder, R F; Charache, P; Staal, S; Wright, P; Forman, M; Hayward, S D; Hayward, G S

    1986-01-01

    Nucleic acid hybridization techniques using cloned probes are finding application in assays of clinical specimens in research and diagnostic laboratories. The probes that we and others have used are recombinant plasmids composed of viral inserts and bacterial plasmid vectors such as pBR322. We suspected that there was material homologous to pBR322 present in many clinical samples. because hybridization occurred in samples which lacked evidence of virus by other techniques. If the presence of this vector-homologous material was unrecognized, hybridization in the test sample might erroneously be interpreted as indicating the presence of viral sequences. In this paper we demonstrate specific hybridization of labeled pBR322 DNA with DNA from various clinical samples. Evidence is presented that nonspecific probe trapping could not account for this phenomenon. In mixing experiments, it is shown that contamination of clinical samples with bacteria would explain such a result. Approaches tested to circumvent this problem included the use of isolated insert probes, alternate cloning vectors, and cold competitor pBR322 DNA in prehybridization and hybridization mixes. None proved entirely satisfactory. We therefore emphasize that it is essential that all hybridization detection systems use a control probe of the vector alone in order to demonstrate the absence of material with vector homology in the specimen tested. Images PMID:3013928

  11. Novel hybrid DHPM-fatty acids: synthesis and activity against glioma cell growth in vitro.

    PubMed

    Treptow, Tamara G M; Figueiró, Fabrício; Jandrey, Elisa H F; Battastini, Ana M O; Salbego, Christianne G; Hoppe, Juliana B; Taborda, Priscila S; Rosa, Sabrina B; Piovesan, Luciana A; Montes D'Oca, Caroline Da R; Russowsky, Dennis; Montes D'Oca, Marcelo G

    2015-05-05

    We described the first synthesis of fatty acid 3,4-dihydropyrimidinones (DHPM-fatty acids) using the Biginelli multicomponent reaction. Antiproliferative activity on two glioma cell lines (C6 rat and U-138-MG human) was also reported. The novel DHPM-fatty acids reduced glioma cell viability relative to temozolomide. Hybrid oxo-monastrol-palmitic acid was the most potent, reducing U-138-MG human cell viability by ca. 50% at 10 μM. In addition, the DHPM-fatty acids showed a large safety range to neural cells, represented by the organotypic hippocampal culture. These results suggest that the increased lipophilicity of DHPM-fatty acids offer a promising approach to overcoming resistance to chemotherapy and may play an important role in the development of new antitumor drugs. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  12. Hydroxyapatite-phosphonoformic acid hybrid compounds prepared by hydrothermal method

    NASA Astrophysics Data System (ADS)

    Turki, Thouraya; Othmani, Masseoud; Bantignies, Jean-Louis; Bouzouita, Khaled

    2014-01-01

    Hydroxyapatites were prepared in the presence of different amounts of phosphonoformic acid (PFA) via the hydrothermal method. The obtained powders were characterized through chemical analysis, XRD, IR, 31P MAS-NMR, TEM, and TG-TDA. The XRD showed that the PFA did not affect the apatite composition. Indeed, only a reduction of the crystallite size was noted. After grafting of PFA, the IR spectroscopy revealed the appearance of new bands belonging to HPO42- and carboxylate groups of the apatite and organic moiety, respectively. Moreover, the 31P MAS-NMR spectra exhibited a peak with a low intensity assigned to the terminal phosphonate group of the organic moiety in addition to that of the apatite. Based on these results, a reaction mechanism involving the surface hydroxyl groups (tbnd Casbnd OH) of the apatite and the carboxyl group of the acid was proposed.

  13. A novel biocompatible hyaluronic acid-chitosan hybrid hydrogel for osteoarthrosis therapy.

    PubMed

    Kaderli, S; Boulocher, C; Pillet, E; Watrelot-Virieux, D; Rougemont, A L; Roger, T; Viguier, E; Gurny, R; Scapozza, L; Jordan, O

    2015-04-10

    A conventional therapy for the treatment of osteoarthrosis is intra-articular injection of hyaluronic acid, which requires repeated, frequent injections. To extend the viscosupplementation effect of hyaluronic acid, we propose to associate it with another biopolymer in the form of a hybrid hydrogel. Chitosan was chosen because of its structural similarity to synovial glycosaminoglycans, its anti-inflammatory effects and its ability to promote cartilage growth. To avoid polyelectrolyte aggregation and obtain transparent, homogeneous gels, chitosan was reacetylated to a 50% degree, and different salts and formulation buffers were investigated. The biocompatibility of the hybrid gels was tested in vitro on human arthrosic synoviocytes, and in vivo assessments were made 1 week after subcutaneous injection in rats and 1 month after intra-articular injection in rabbits. Hyaluronic acid-chitosan polyelectrolyte complexes were prevented by cationic complexation of the negative charges of hyaluronic acid. The different salts tested were found to alter the viscosity and thermal degradation of the gels. Good biocompatibility was observed in rats, although the calcium-containing formulation induced calcium deposits after 1 week. The sodium chloride formulation was further tested in rabbits and did not show acute clinical signs of pain or inflammation. Hybrid HA-Cs hydrogels may be a valuable alternative viscosupplementation agent. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Heteropoly Acid/Nitrogen Functionalized Onion-like Carbon Hybrid Catalyst for Ester Hydrolysis Reactions.

    PubMed

    Liu, Wei; Qi, Wei; Guo, Xiaoling; Su, Dangsheng

    2016-02-18

    A novel heteropoly acid (HPA)/nitrogen functionalized onion-like carbon (NOLC) hybrid catalyst was synthesized through supramolecular (electrostatic and hydrogen bond) interactions between the two components. The chemical structure and acid strength of the HPA/NOLC hybrid have been fully characterized by thermogravimetric analysis, IR spectroscopy, X-ray photoelectron spectroscopy, NH3 temperature-programmed desorption and acid-base titration measurements. The proposed method for the fabrication of the HPA/NOLC hybrid catalyst is a universal strategy for different types of HPAs to meet various requirements of acidic or redox catalysis. The hydrophobic environment of NOLC effectively prevents the deactivation of HPA in an aqueous system, and the combination of uniformly dispersed HPA clusters and the synergistic effect between NOLC and HPA significantly promotes its activity in ester hydrolysis reactions, which is higher than that of bare PWA as homogeneous catalyst. The kinetics of the hydrolysis reactions indicate that the aggregation status of the catalyst particles has great influence on the apparent activity. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Stereoselectivity of formation of monoterpene - Amino acids hybrid molecules in the reaction of monoterpene nitroso chlorides with α-amino acid derivatives.

    PubMed

    Marenin, K S; Gatilov, Yu V; Agafontsev, A M; Tkachev, A V

    2017-01-01

    Reaction of nitrosochlorides of natural monoterpene hydrocarbons (+)-3-carene and (-)-α-pinene with L-amino acids and their methyl esters results in stereoselective formation of terpene-amino acids hybrids, which belong to the series of α-substituted amino oximes. The reaction with an excess of racemic DL-amino acids and their derivatives induces partial resolution of the amino acid components and formation of the diastereomeric mixtures of the terpene-amino acids hybrids, with diastereomeric excess varying from 0 to 100%. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Hierarchical assembly of viral nanotemplates with encoded microparticles via nucleic acid hybridization.

    PubMed

    Tan, Wui Siew; Lewis, Christina L; Horelik, Nicholas E; Pregibon, Daniel C; Doyle, Patrick S; Yi, Hyunmin

    2008-11-04

    We demonstrate hierarchical assembly of tobacco mosaic virus (TMV)-based nanotemplates with hydrogel-based encoded microparticles via nucleic acid hybridization. TMV nanotemplates possess a highly defined structure and a genetically engineered high density thiol functionality. The encoded microparticles are produced in a high throughput microfluidic device via stop-flow lithography (SFL) and consist of spatially discrete regions containing encoded identity information, an internal control, and capture DNAs. For the hybridization-based assembly, partially disassembled TMVs were programmed with linker DNAs that contain sequences complementary to both the virus 5' end and a selected capture DNA. Fluorescence microscopy, atomic force microscopy (AFM), and confocal microscopy results clearly indicate facile assembly of TMV nanotemplates onto microparticles with high spatial and sequence selectivity. We anticipate that our hybridization-based assembly strategy could be employed to create multifunctional viral-synthetic hybrid materials in a rapid and high-throughput manner. Additionally, we believe that these viral-synthetic hybrid microparticles may find broad applications in high capacity, multiplexed target sensing.

  17. Wollastonite hybridizing stearic acid as thermal energy storage material

    NASA Astrophysics Data System (ADS)

    Xu, Dawei; Yang, Huaming

    2014-11-01

    This paper reported on the preparation of a novel stearic acid (SA)/wollastonite (W) composite as a form-stable phase change material (PCM) for thermal energy-storage (TES) by vacuum impregnation, and especially investigated the effect of the size grade of W on the thermal properties of the SA/W composite. Samples were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), laser particle-size analysis, and differential scanning calorimetry (DSC). Natural W (Wr) was classified into four size grades by wet screening. The results indicate that no chemical reaction took place between SA and W, and the SA load in the SA/W composite increased with an increase in the length/diameter (L/D) ratio of the W. The SA/W composite with a W L/D ratio of 22.5 exhibited latent heats of melting and freezing of 58.64 J/g and 56.95 J/g, respectively, which was higher than those of the composite incorporating natural W. We believe that the as-prepared form-stable PCM composite could provide a potential means of TES for the concentrated solar power.

  18. The Use of Atomic Force Microscopy for 3D Analysis of Nucleic Acid Hybridization on Microarrays.

    PubMed

    Dubrovin, E V; Presnova, G V; Rubtsova, M Yu; Egorov, A M; Grigorenko, V G; Yaminsky, I V

    2015-01-01

    Oligonucleotide microarrays are considered today to be one of the most efficient methods of gene diagnostics. The capability of atomic force microscopy (AFM) to characterize the three-dimensional morphology of single molecules on a surface allows one to use it as an effective tool for the 3D analysis of a microarray for the detection of nucleic acids. The high resolution of AFM offers ways to decrease the detection threshold of target DNA and increase the signal-to-noise ratio. In this work, we suggest an approach to the evaluation of the results of hybridization of gold nanoparticle-labeled nucleic acids on silicon microarrays based on an AFM analysis of the surface both in air and in liquid which takes into account of their three-dimensional structure. We suggest a quantitative measure of the hybridization results which is based on the fraction of the surface area occupied by the nanoparticles.

  19. Synthesis of hollow hybrid hydroxyapatite microspheres based on chitosan-poly(acrylic acid) microparticles.

    PubMed

    Zhang, Haibin; Zhou, Kechao; Li, Zhiyou; Huang, Suping

    2009-06-01

    Core-template-free hybrid hydroxyapatite (HA) hollow microspheres based on a chitosan-poly acrylic acid (CS-PAA) complex were prepared. The amine groups on chitosan can interact with the carboxyl groups of poly(acrylic acid) to form hollow microspheres. The hollow HA microspheres of about 1.0 microm are obtained by heterogeneous nucleation of HA on CS-PAA hollow spheres. Gelatin (Gel), acting as a novel cross-link agent, is introduced to bind the spheres of CS-PAA and HA. The forming mechanism of hollow spheres and the influencing factors on the size of microspheres are investigated. In addition, the role of Gel is elucidated in the forming process of the hollow hybrid sphere.

  20. A new class of homogeneous nucleic acid probes based on specific displacement hybridization

    PubMed Central

    Li, Qingge; Luan, Guoyan; Guo, Qiuping; Liang, Jixuan

    2002-01-01

    We have developed a new class of probes for homogeneous nucleic acid detection based on the proposed displacement hybridization. Our probes consist of two complementary oligodeoxyribonucleotides of different length labeled with a fluorophore and a quencher in close proximity in the duplex. The probes on their own are quenched, but they become fluorescent upon displacement hybridization with the target. These probes display complete discrimination between a perfectly matched target and single nucleotide mismatch targets. A comparison of double-stranded probes with corresponding linear probes confirms that the presence of the complementary strand significantly enhances their specificity. Using four such probes labeled with different color fluorophores, each designed to recognize a different target, we have demonstrated that multiple targets can be distinguished in the same solution, even if they differ from one another by as little as a single nucleotide. Double-stranded probes were used in real-time nucleic acid amplifications as either probes or as primers. In addition to its extreme specificity and flexibility, the new class of probes is simple to design and synthesize, has low cost and high sensitivity and is accessible to a wide range of labels. This class of probes should find applications in a variety of areas wherever high specificity of nucleic acid hybridization is relevant. PMID:11788731

  1. Diagnostics based on nucleic acid sequence variant profiling: PCR, hybridization, and NGS approaches.

    PubMed

    Khodakov, Dmitriy; Wang, Chunyan; Zhang, David Yu

    2016-10-01

    Nucleic acid sequence variations have been implicated in many diseases, and reliable detection and quantitation of DNA/RNA biomarkers can inform effective therapeutic action, enabling precision medicine. Nucleic acid analysis technologies being translated into the clinic can broadly be classified into hybridization, PCR, and sequencing, as well as their combinations. Here we review the molecular mechanisms of popular commercial assays, and their progress in translation into in vitro diagnostics. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  2. Mismatch discrimination in fluorescent in situ hybridization using different types of nucleic acids.

    PubMed

    Fontenete, Silvia; Silvia, Fontenete; Barros, Joana; Joana, Barros; Madureira, Pedro; Pedro, Madureira; Figueiredo, Céu; Céu, Figueiredo; Wengel, Jesper; Jesper, Wengel; Azevedo, Nuno Filipe; Filipe, Azevedo Nuno

    2015-05-01

    In the past few years, several researchers have focused their attention on nucleic acid mimics due to the increasing necessity of developing a more robust recognition of DNA or RNA sequences. Fluorescence in situ hybridization (FISH) is an example of a method where the use of these novel nucleic acid monomers might be crucial to the success of the analysis. To achieve the expected accuracy in detection, FISH probes should have high binding affinity towards their complementary strands and discriminate effectively the noncomplementary strands. In this study, we investigate the effect of different chemical modifications in fluorescent probes on their ability to successfully detect the complementary target and discriminate the mismatched base pairs by FISH. To our knowledge, this paper presents the first study where this analysis is performed with different types of FISH probes directly in biological targets, Helicobacter pylori and Helicobacter acinonychis. This is also the first study where unlocked nucleic acids (UNA) were used as chemistry modification in oligonucleotides for FISH methodologies. The effectiveness in detecting the specific target and in mismatch discrimination appears to be improved using locked nucleic acids (LNA)/2'-O-methyl RNA (2'OMe) or peptide nucleic acid (PNA) in comparison to LNA/DNA, LNA/UNA, or DNA probes. Further, the use of LNA modifications together with 2'OMe monomers allowed the use of shorter fluorescent probes and increased the range of hybridization temperatures at which FISH would work.

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

    SciTech Connect

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

    2002-04-17

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

  4. Naturally occurring hybrids derived from γ-amino acids and sugars with potential tail to tail ether-bonds

    NASA Astrophysics Data System (ADS)

    Feng, Zi-Ming; Zhan, Zhi-Lai; Yang, Ya-Nan; Jiang, Jian-Shuang; Zhang, Pei-Cheng

    2016-05-01

    The basic substances of life include various amino acids and sugars. To search such molecules is the precondition to understand the essential nature. Here we reported four unprecedented hybrids of γ-amino acids and sugars from the roots of Ranunculus ternatus, which possess potential tail to tail ether-connected (6,6-ether-connected) modes in the sugar moiety. The structures of these hybrids were elucidated by extensive analyses of spectra and calculated electronic circular dichroism (ECD) method.

  5. Organic/inorganic hybrid amine and sulfonic acid tethered silica materials: Synthesis, characterization and application

    NASA Astrophysics Data System (ADS)

    Hicks, Jason Christopher

    The major goals of this thesis were to: (1) create a site-isolated aminosilica material with higher amine loadings than previously reported isolation methods, (2) use spectroscopic, reactivity, and catalytic (olefin polymerization precatalysts) probes to determine isolation of amine groups on these organic/inorganic hybrid materials, (3) synthesize an organic/inorganic hybrid material capable of activating Group 4 olefin polymerization precatalysts, and (4) synthesize a high amine loaded organic/inorganic hybrid material capable of reversibly capturing CO2 in a simulated flue gas stream. The underlying motivation of this research involved the synthesis and design of novel amine and sulfonic acid materials. Traditional routes to synthesize aminosilicas have led to the formation of a high loading of multiple types of amine sites on the silica surface. Part of this research involved the creation of a new aminosilica material via a protection/deprotection method designed to prevent multiple sites, while maintaining a relatively high loading. As a characterization technique, fluorescence spectroscopy of pyrene-based fluorophores loaded on traditional aminosilicas and site-isolated aminosilicas was used to probe the degree of site-isolation obtained with these methods. Also, this protection/deprotection method was compared to other reported isolation techniques with heterogeneous Group 4 constrained-geometry inspired catalysts (CGCs). It was determined that the degree of separation of the amine sites could be controlled with protection/deprotection methods. Furthermore, an increase in the reactivity of the amines and the catalytic activity of CGCs built off of the amines was determined for aminosilicas synthesized by a protection/deprotection method. The second part of this work involved developing organic/inorganic hybrid materials as heterogeneous Bronsted acidic cocatalysts for activation of olefin polymerization precatalysts. This was the first reported organic

  6. Label-Free Potentiometry for Detecting DNA Hybridization Using Peptide Nucleic Acid and DNA Probes

    PubMed Central

    Goda, Tatsuro; Singi, Ankit Balram; Maeda, Yasuhiro; Matsumoto, Akira; Torimura, Masaki; Aoki, Hiroshi; Miyahara, Yuji

    2013-01-01

    Peptide nucleic acid (PNA) has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM)-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry. PMID:23435052

  7. Label-free potentiometry for detecting DNA hybridization using peptide nucleic acid and DNA probes.

    PubMed

    Goda, Tatsuro; Singi, Ankit Balram; Maeda, Yasuhiro; Matsumoto, Akira; Torimura, Masaki; Aoki, Hiroshi; Miyahara, Yuji

    2013-02-07

    Peptide nucleic acid (PNA) has outstanding affinity over DNA for complementary nucleic acid sequences by forming a PNA-DNA heterodimer upon hybridization via Watson-Crick base-pairing. To verify whether PNA probes on an electrode surface enhance sensitivity for potentiometric DNA detection or not, we conducted a comparative study on the hybridization of PNA and DNA probes on the surface of a 10-channel gold electrodes microarray. Changes in the charge density as a result of hybridization at the solution/electrode interface on the self-assembled monolayer (SAM)-formed microelectrodes were directly transformed into potentiometric signals using a high input impedance electrometer. The charge readout allows label-free, reagent-less, and multi-parallel detection of target oligonucleotides without any optical assistance. The differences in the probe lengths between 15- to 22-mer dramatically influenced on the sensitivity of the PNA and DNA sensors. Molecular type of the capturing probe did not affect the degree of potential shift. Theoretical model for charged rod-like duplex using the Gouy-Chapman equation indicates the dominant effect of electrostatic attractive forces between anionic DNA and underlying electrode at the electrolyte/electrode interface in the potentiometry.

  8. Preparation of Phytic Acid/Silane Hybrid Coating on Magnesium Alloy and Its Corrosion Resistance in Simulated Body Fluid

    NASA Astrophysics Data System (ADS)

    Wang, Fengwu; Cai, Shu; Shen, Sibo; Yu, Nian; Zhang, Feiyang; Ling, Rui; Li, Yue; Xu, Guohua

    2017-09-01

    In order to decrease the corrosion rate and improve the bioactivity of magnesium alloy, phytic acid/saline hybrid coatings were synthesized on AZ31 magnesium alloys by sol-gel dip-coating method. It was found that the mole ratio of phytic acid to γ-APS had a great influence on coating morphology and the corresponding corrosion resistance of the coated magnesium alloys. When the mole ratio of phytic acid to γ-APS was 1:1, the obtained hybrid coating was integral and without cracks, which was ascribed to the strong chelate capability of phytic acid and Si-O-Si network derived from silane. Electrochemical test result indicated that the corrosion resistance of the coated magnesium alloy was about 27 times larger than that of the naked counterpart. In parallel, immersion test showed that the phytic acid/silane hybrid coating could induce CaP-mineralized product deposition, which offered another protection for magnesium alloy.

  9. Porous Zirconium-Phytic Acid Hybrid: a Highly Efficient Catalyst for Meerwein-Ponndorf-Verley Reductions.

    PubMed

    Song, Jinliang; Zhou, Baowen; Zhou, Huacong; Wu, Lingqiao; Meng, Qinglei; Liu, Zhimin; Han, Buxing

    2015-08-03

    The utilization of compounds from natural sources to prepare functional materials is of great importance. Herein, we describe for the first time the preparation of organic-inorganic hybrid catalysts by using natural phytic acid as building block. Zirconium phosphonate (Zr-PhyA) was synthesized by reaction of phytic acid and ZrCl4 and was obtained as a mesoporous material with pore sizes centered around 8.5 nm. Zr-PhyA was used to catalyze the mild and selective Meerwein-Ponndorf-Verley (MPV) reduction of various carbonyl compounds, e.g., of levulinic acid and its esters into γ-valerolactone. Further studies indicated that both Zr and phosphate groups contribute significantly to the excellent performance of Zr-PhyA.

  10. Phosphoric acid doped polybenzimidazole/imidazolium-modified silsesquioxane hybrid proton conducting membranes for anhydrous proton exchange membrane application

    NASA Astrophysics Data System (ADS)

    Lin, Bencai; Chu, Fuqiang; Yuan, Ningyi; Shang, Hui; Ren, Yurong; Gu, Zongzong; Ding, Jianning; Wei, Yingqiang; Yu, Xiaomin

    2014-04-01

    Phosphoric acid doped polybenzimidazole (PBI)/imidazolium-modified silsesquioxane (Im-SiO3/2) hybrid membranes with high proton conductivity at high temperature under anhydrous conditions are synthesized and characterized. The presence of Im-SiO3/2 is confirmed by FT-IR and energy-dispersive X-ray spectroscopy (EDS) mapping of silicon element. The phosphoric acid uptake and proton conductivity of the hybrid membranes increase with the Im-SiO3/2 content, and the conductivity of PBI/Im-SiO3/2-20 reaching 6.3 × 10-2 S cm-1 at 180 °C. Compared with pure PBI membranes, the introduction of Im-SiO3/2 is effective in preventing the release of the phosphoric acid component from the hybrid membranes. The properties of the prepared hybrid membranes indicate their promising prospects in anhydrous proton exchange membrane applications.

  11. Efficient Solar-Driven Nitrogen Fixation over Carbon-Tungstic-Acid Hybrids.

    PubMed

    Li, Xiaoman; Wang, Wenzhong; Jiang, Dong; Sun, Songmei; Zhang, Ling; Sun, Xiang

    2016-09-19

    Ammonia synthesis under mild conditions is of supreme interest. Photocatalytic nitrogen fixation with water at room temperature and atmospheric pressure is an intriguing strategy. However, the efficiency of this method has been far from satisfied for industrialization, mainly due to the sluggish cleavage of the N≡N bond. Herein, we report a carbon-tungstic-acid (WO3 ⋅H2 O) hybrid for the co-optimization of N2 activation as well as subsequent photoinduced protonation. Efficient ammonia evolution reached 205 μmol g(-1)  h(-1) over this hybrid under simulated sunlight. Nitrogen temperature-programmed desorption revealed the decisive role of carbon in N2 adsorption. Photoactive WO3 ⋅H2 O guaranteed the supply of electrons and protons for subsequent protonation. The universality of carbon modification for enhancing the N2 reduction was further verified over various photocatalysts, shedding light on future materials design for ideal solar energy utilization.

  12. Evaluation of three-dimensional microchannel glass biochips for multiplexed nucleic acid fluorescence hybridization assays.

    PubMed

    Benoit, V; Steel, A; Torres, M; Yu, Y Y; Yang, H; Cooper, J

    2001-06-01

    Three-dimensional, flow-through microchannel glass substrates have a potential for enhanced performance, including increased sensitivity and dynamic range, over traditional planar substrates used in medium-density microarray platforms. This paper presents a methodology for the implementation of multiplexed nucleic acid hybridization fluorescence assays on microchannel glass substrates. Fluorescence detection was achieved, in a first instance, using conventional low-magnification microscope objective lenses, as imaging optics whose depth-of-field characteristics match the thickness of the microchannel glass chip. The optical properties of microchannel glass were shown, through experimental results and simulations, to be compatible with the quantitative detection of heterogeneous hybridization events taking place along the microchannel sidewalls, with detection limits for oligonucleotide targets in the low-attomole range.

  13. Detection of Huanglongbing (citrus greening) disease by nucleic acid spot hybridization.

    PubMed

    Gopal, Kuraba; Sudarsan, Sundeep; Gopi, Venati; Naidu, Latchireddy Naram; Ramaiah, Maniyaram; Sreenivasulu, Yasodam; Wesley, Edward

    2009-01-01

    Polymerase chain reaction (PCR) amplification with primers specific to the rDNA region successfully amplified the 1160-bp DNA fragment from a Huanglongbing (HLB)-infected sweet orange sample with mottling symptoms leaves, but not from healthy sweet orange plants. The PCR product of 1160-bp was used as probe labeled with biotin for detection of the HLB pathogen in the nucleic acid spot hybridization (NASH) test. It was found that the HLB pathogen could be detected up to 1:100 dilution in HLB-infected tissue. Total DNA extracted from HLB-infected tissue was diluted 2-fold as 900 ng in TE buffer and spotted on a nitrocellulose membrane. Strong signals were observed up to 225 ng of DNA dilution, whereas a moderate signal was recorded at 112 ng. No hybridization signal was observed in the healthy samples, while strong signals were observed in the positive control.

  14. Effect of acidic solutions on the surface degradation of a micro-hybrid composite resin.

    PubMed

    Münchow, Eliseu A; Ferreira, Ana Cláudia A; Machado, Raissa M M; Ramos, Tatiana S; Rodrigues-Junior, Sinval A; Zanchi, Cesar H

    2014-01-01

    Composite resins may undergo wear by the action of chemical substances (e.g., saliva, alcohol, bacterial acids) of the oral environment, which may affect the material's structure and surface properties. This study evaluated the effect of acidic substances on the surface properties of a micro-hybrid composite resin (Filtek Z-250). Eighty specimens were prepared, and baseline hardness and surface roughness (KMN0 and Ra0, respectively) were measured. The specimens were subjected to sorption (SO) and solubility (SL) tests according to ISO 4049:2009, but using different storage solutions: deionized water; 75/25 vol% ethanol/water solution; lactic acid; propionic acid; and acetic acid. The acids were used in two concentrations: PA and 0.02 N. pH was measured for all solutions and final hardness (KMN1) and surface roughness (Ra1) were measured. Data were analyzed with paired t-tests and one-way ANOVA and Tukey's test (a=5%). All solutions decreased hardness and increased the Ra values, except for the specimens stored in water and 0.02 N lactic acid, which maintained the hardness. All solutions produced similar SO and SL phenomena, except for the 0.02 N lactic acid, which caused lower solubility than the other solutions. Ethanol showed the highest pH (6.6) and the 0.02 N lactic acid the lowest one (2.5). The solutions affected negatively the surface properties of the composite resin; in addition, an acidic pH did not seem to be a significant factor that intensifies the surface degradation phenomena.

  15. Time-Resolved Nucleic Acid Hybridization Beacons Utilizing Unimolecular and Toehold-Mediated Strand Displacement Designs.

    PubMed

    Massey, Melissa; Ancona, Mario G; Medintz, Igor L; Algar, W Russ

    2015-12-01

    Nucleic acid hybridization probes are sought after for numerous assay and imaging applications. These probes are often limited by the properties of fluorescent dyes, prompting the development of new probes where dyes are paired with novel or nontraditional luminescent materials. Luminescent terbium complexes are an example of such a material, and these complexes offer several unique spectroscopic advantages. Here, we demonstrate two nonstem-loop designs for light-up nucleic acid hybridization beacons that utilize time-resolved Förster resonance energy transfer (TR-FRET) between a luminescent Lumi4-Tb cryptate (Tb) donor and a fluorescent reporter dye, where time-resolved emission from the dye provides an analytical signal. Both designs are based on probe oligonucleotides that are labeled at their opposite termini with Tb and a fluorescent reporter dye. In one design, a probe is partially blocked with a quencher dye-labeled oligonucleotide, and target hybridization is signaled through toehold-mediated strand displacement and loss of a competitive FRET pathway. In the other design, the intrinsic folding properties of an unblocked probe are utilized in combination with a temporal mechanism for signaling target hybridization. This temporal mechanism is based on a recently elucidated "sweet spot" for TR-FRET measurements and exploits distance control over FRET efficiencies to shift the Tb lifetime within or outside the time-gated detection window for measurements. Both the blocked and unblocked beacons offer nanomolar (femtomole) detection limits, response times on the order of minutes, multiplexing through the use of different reporter dyes, and detection in complex matrices such as serum and blood. The blocked beacons offer better mismatch selectivity, whereas the unblocked beacons are simpler in design. The temporal mechanism of signaling utilized with the unblocked beacons also plays a significant role with the blocked beacons and represents a new and effective

  16. Variability in fatty acid and triacylglycerol composition of the oil of coconut (Cocos nucifera L.) hybrids and their parentals.

    PubMed

    Laureles, Lucita R; Rodriguez, Felicito M; Reaño, Consorcia E; Santos, Gerardo A; Laurena, Antonio C; Mendoza, Evelyn Mae Tecson

    2002-03-13

    The fatty acid profiles and triacylglycerol (TAG) compositions of oils from the solid endosperm of different Philippine coconut hybrids and their parentals were determined by using gas chromatography (GC) and high-performance liquid chromatography (HPLC). In general, varietal differences in fatty acid composition were observed. Lauric acid (C12) content was significantly higher in the hybrids PCA 15-8 (50.45%) and PCA 15-9 (50.26%) by about 3.16% points as compared to other hybrids, and higher in Tacunan Green Dwarf (50.50%) among the parentals. Among the fatty acids, lauric acid exhibited the least variation. In general, none of the hybrids had higher fatty acid content than their parentals. The HPLC chromatogram of triacylglycerols (TAG) showed 8 major peaks which differ in carbon number (CN) by two: identified as TAG CN 30, 32, 34, 36, 38, 40, 42, and 44. TAGs CN 30 (4.08%) and CN 34 (19.20%) were found to be significantly higher in PCA 15-9 than in the other hybrids. CN 36 was highest (21.94-23.66%) in all hybrids and parentals. The TAG CNs varied significantly among hybrids and parents, i.e., in CN 30, 32, and 34, which are high in medium chain triacylglycerols (MCTs), and in CN 30 (for parentals only), 40, 42, and 44 (the latter two for parentals only), and none in CN 36. MCTs calculated for two hybrids and their parents ranged from 13.81% to 20.55%.

  17. Synthesis and anti-tumor activity evaluation of gallic acid-mangiferin hybrid molecule.

    PubMed

    Hu, Xiang-yu; Deng, Jia-gang; Wang, Lin; Yuan, Ye-fei

    2013-12-01

    To improve the anti-tumor effects of gallic acid and mangiferin, a gallic acid-mangiferin hybrid molecule (GAMA) was synthesized from gallic acid with mangiferin in the presence of ionic liquid ChC1(choline chloride)·2SnC12. Chemical and spectroscopic methods, such as (1)H and (13)C NMR spectroscopy, and HR-ESIMS were used for the structure identification of GA-MA. Using the cell counting kit-8 (CCK-8) assay, the in vitro anti-tumor effects were compared between GA-MA, gallic acid and mangiferin on human hepatoma HepG2, human nasopharyngeal carcinoma CNE, human lung cancer NCI-H460, human ovarian cancer SK-OV-3, and human cervical cancer Hela cells. The results showed that the half inhibitory concentration (IC50) of GA-MA on HepG2, CNE, NCI-H460, SK-OV-3, and Hela cells was significantly lower than that of gallic acid or mangiferin. This showed that GA-MA has a better in vitro anti-tumor effect than gallic acid and mangi-ferin.

  18. A high power spiral wound lead-acid battery for hybrid electric vehicles

    SciTech Connect

    Olson, J.B.; Sexton, E.D.

    1997-12-01

    Optima Batteries, Inc. is currently in development of a high power (660 W/kg) spiral wound lead-acid 6V battery with a nominal capacity of 15 Ah. Its exceptional power and excellent thermal characteristics make it a promising choice for hybrid electric vehicle applications. The hybrid electric vehicle presents a new and unique challenge for energy storage systems. The batteries require high power for acceleration and hill climbing and good charge acceptance for regenerative braking and overall energy efficiency. Since the on board auxiliary power unit results in much lower demands for battery energy capacity, lead-acid batteries fit quite well into these performance requirements. Many of the remaining challenges involve the development of battery management systems which must function to maintain the battery pack at peak performance and achieve an economical cycle life. Related to the issue of battery management is information about conditions that may cause damage or unbalance of the pack. Experiments are described investigating the effects of extreme cell reversal on battery capacity and cycle life. The results demonstrate the amazing robustness of the lead-acid battery for tolerating over discharge.

  19. Application of locked nucleic acid-based probes in fluorescence in situ hybridization.

    PubMed

    Fontenete, Sílvia; Carvalho, Daniel; Guimarães, Nuno; Madureira, Pedro; Figueiredo, Céu; Wengel, Jesper; Azevedo, Nuno Filipe

    2016-07-01

    Fluorescence in situ hybridization (FISH) employing nucleic acid mimics as probes is becoming an emerging molecular tool in the microbiology area for the detection and visualization of microorganisms. However, the impact that locked nucleic acid (LNA) and 2'-O-methyl (2'-OMe) RNA modifications have on the probe that is targeting microorganisms is unknown. In this study, the melting and hybridization efficiency properties of 18 different probes in regards to their use in FISH for the detection of the 16S rRNA of Helicobacter pylori were compared. For the same sequence and target, probe length and the type of nucleic acid mimics used as mixmers in LNA-based probes strongly influence the efficiency of detection. LNA probes with 10 to 15 mers showed the highest efficiency. Additionally, the combination of 2'-OMe RNA with LNA allowed an increase on the fluorescence intensities of the probes. Overall, these results have significant implications for the design and applications of LNA probes for the detection of microorganisms.

  20. Identification of random nucleic acid sequence aberrations using dual capture probes which hybridize to different chromosome regions

    DOEpatents

    Lucas, Joe N.; Straume, Tore; Bogen, Kenneth T.

    1998-01-01

    A method is provided for detecting nucleic acid sequence aberrations using two immobilization steps. According to the method, a nucleic acid sequence aberration is detected by detecting nucleic acid sequences having both a first nucleic acid sequence type (e.g., from a first chromosome) and a second nucleic acid sequence type (e.g., from a second chromosome), the presence of the first and the second nucleic acid sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. In the method, immobilization of a first hybridization probe is used to isolate a first set of nucleic acids in the sample which contain the first nucleic acid sequence type. Immobilization of a second hybridization probe is then used to isolate a second set of nucleic acids from within the first set of nucleic acids which contain the second nucleic acid sequence type. The second set of nucleic acids are then detected, their presence indicating the presence of a nucleic acid sequence aberration.

  1. Identification of random nucleic acid sequence aberrations using dual capture probes which hybridize to different chromosome regions

    DOEpatents

    Lucas, J.N.; Straume, T.; Bogen, K.T.

    1998-03-24

    A method is provided for detecting nucleic acid sequence aberrations using two immobilization steps. According to the method, a nucleic acid sequence aberration is detected by detecting nucleic acid sequences having both a first nucleic acid sequence type (e.g., from a first chromosome) and a second nucleic acid sequence type (e.g., from a second chromosome), the presence of the first and the second nucleic acid sequence type on the same nucleic acid sequence indicating the presence of a nucleic acid sequence aberration. In the method, immobilization of a first hybridization probe is used to isolate a first set of nucleic acids in the sample which contain the first nucleic acid sequence type. Immobilization of a second hybridization probe is then used to isolate a second set of nucleic acids from within the first set of nucleic acids which contain the second nucleic acid sequence type. The second set of nucleic acids are then detected, their presence indicating the presence of a nucleic acid sequence aberration. 14 figs.

  2. The dietary branched chain amino acid requirements of hybrid striped bass(Morone chrysops x M. saxatilis)

    USDA-ARS?s Scientific Manuscript database

    The requirements for branched chain amino acids (BCAAs) are unknown in hybrid striped bass and necessary for formulating efficient and nutritious diets. Moreover, the dietary balance among these three amino acids can substantially influence the performance of meat animals fed those diets. The diet...

  3. Principles of nucleic acid hybridization and comparison with monoclonal antibody technology for the diagnosis of infectious diseases.

    PubMed Central

    Edberg, S. C.

    1985-01-01

    Until the 1980s the diagnosis of specific etiologic agents of infectious diseases rested with their isolation in vitro and identification by analysis of their phenotypic characteristics. In the 1970s the concept of a microbial species evolved from phenotypic analysis to nucleic acid homology. Currently, nucleic acid sequences specific for a given species are being isolated and amplified and utilized not only to identify the pathogen after it has been grown in vitro but also elucidate it directly in biological material. The procedures for making nucleic acid hybridization probes are analogous to the generation of monoclonal antibody tests. Currently, research and development are centered in choosing the particular nucleic acid to analyze, establishing the most efficient vector system for amplifying the nucleic acid, generating an efficient means of selecting the particular nucleic acid fragment specific for the microorganism, and in measuring the hybridization reaction. While immunological techniques have been utilized in the clinical laboratory for over thirty years, the means of detecting nucleic acid hybridization reactions are just beginning to be usable in the clinical diagnostic laboratory. Much of nucleic acid hybridization research is proprietary, and a particular challenge is to develop a means whereby information can be used for the progress of science as a whole when generated by private ownership. Images FIG. 4 PMID:3004048

  4. Kinetics for exchange of imino protons in deoxyribonucleic acid, ribonucleic acid, and hybrid oligonucleotide helices

    SciTech Connect

    Pardi, A.; Tinoco, I. Jr.

    1982-09-14

    The lifetimes for opening of individual base pairs in a DNA (dCA/sub 5/G + dCT/sub 5/G), an RNA (rCA/sub 5/G + rCU/sub 5/G), and a hybrid DNA-RNA (rCA/sub 5/G + dCT/sub 5/G) helix have been measured by proton nuclear magnetic resonance. The lifetimes were obtained by saturation recovery experiments performed on the hydrogen-bonding imino protons of the Watson-Crick base pairs. In these oligonucleotide helices the observed relaxation rates were dominated by exchange with water, with the magnetic spin-lattice relaxation time of the imino protons possibly being important only at the lowest temperatures in the DNA helix. It was shown that three interior base pairs in the DNA heptamer dCA/sub 5/G + dCT/sub 5/G were in the open-limited region, which means that these imino protons exchange every time the base pair opens. The lifetimes of the terminal G-C base pairs in the DNA helix are much shorter than the interior A-T base pairs. The pH dependence of the terminal base pairs indicated that the ends of the helix open and close many times before exchange of the imino protons with water takes place. The temperature dependence of the lifetimes of the interior A-T imino protons in the DNA helix showed that these protons exchange only when the double helix was dissociated into single strands. Thus, these lifetimes measure the rate for dissociation of the double helix. The activation energy for this process was found to be 47 kcal/mol. Comparison of the lifetimes of the interior protons in the DNA, RNA, and hybrid helices showed that the rates of dissociation of the RNA and hybrid helices are very similar at 5/sup 0/ C, whereas the rate for the DNA helix was approximately 1 order of magnitude smaller than that for the other two helices.

  5. A novel chaotic based image encryption using a hybrid model of deoxyribonucleic acid and cellular automata

    NASA Astrophysics Data System (ADS)

    Enayatifar, Rasul; Sadaei, Hossein Javedani; Abdullah, Abdul Hanan; Lee, Malrey; Isnin, Ismail Fauzi

    2015-08-01

    Currently, there are many studies have conducted on developing security of the digital image in order to protect such data while they are sending on the internet. This work aims to propose a new approach based on a hybrid model of the Tinkerbell chaotic map, deoxyribonucleic acid (DNA) and cellular automata (CA). DNA rules, DNA sequence XOR operator and CA rules are used simultaneously to encrypt the plain-image pixels. To determine rule number in DNA sequence and also CA, a 2-dimension Tinkerbell chaotic map is employed. Experimental results and computer simulations, both confirm that the proposed scheme not only demonstrates outstanding encryption, but also resists various typical attacks.

  6. Intertribal somatic hybrids between Brassica napus and Thlaspi perfoliatum with high content of the T. perfoliatum-specific nervonic acid.

    PubMed

    Fahleson, J; Eriksson, I; Landgren, M; Stymne, S; Glimelius, K

    1994-02-01

    Protoplast fusions were performed between hypocotyl protoplasts of Brassica napus and mesophyll protoplasts of Thlaspi perfoliatum. The two species are members of the Lepidieae and Brassiceae tribes, respectively, in the family of Brassicaceae. Seeds of T. perfoliatum are rich in the fatty acid C24∶1 (nervonic acid), an oil valuable for technical purposes. In the search for renewable oils to replace the mineral oils, plant breeders have been trying to develop oil crops with a high content of long-chain fatty acids. After fusion of B. napus protoplasts with non-irradiated as well as irradiated protoplasts of T. perfoliatum selection was carried out by flow cytometry and cell sorting. Of the shoots regenerated from different calli 27 were verified as hybrids or partial hybrids using the isoenzyme phosphoglucose isomerase (PGI) as a marker. Another 6 plants were identified as partial hybrids using a T. perfoliatum-specific repetitive DNA sequence. Slot blot experiments were performed to estimate the copy number of the repetitive DNA sequence in the parental species and in the hybrids. In T. perfoliatum there were approximately 10(5) copies per haploid genome, and the range in the hybrids was 1-37% of the value in T. perfoliatum. When the nuclear DNA content of the regenerated shoots was analysed we found partial as well as symmetric hybrids. Even though the rooting and establishment of hybrid shoots in the greenhouse were difficult, resulting in the death of many plants, 19 plants were cultured to full maturity. Seeds obtained from 15 plants were analysed to determine whether they contained nervonic acid, and 5 of the hybrids were found to contain significantly greater amounts of nervonic acid than B. napus.

  7. High affinity γPNA sandwich hybridization assay for rapid detection of short nucleic acid targets with single mismatch discrimination.

    PubMed

    Goldman, Johnathan M; Zhang, Li Ang; Manna, Arunava; Armitage, Bruce A; Ly, Danith H; Schneider, James W

    2013-07-08

    Hybridization analysis of short DNA and RNA targets presents many challenges for detection. The commonly employed sandwich hybridization approach cannot be implemented for these short targets due to insufficient probe-target binding strengths for unmodified DNA probes. Here, we present a method capable of rapid and stable sandwich hybridization detection for 22 nucleotide DNA and RNA targets. Stable hybridization is achieved using an n-alkylated, polyethylene glycol γ-carbon modified peptide nucleic acid (γPNA) amphiphile. The γPNA's exceptionally high affinity enables stable hybridization of a second DNA-based probe to the remaining bases of the short target. Upon hybridization of both probes, an electrophoretic mobility shift is measured via interaction of the n-alkane modification on the γPNA with capillary electrophoresis running buffer containing nonionic surfactant micelles. We find that sandwich hybridization of both probes is stable under multiple binding configurations and demonstrate single base mismatch discrimination. The binding strength of both probes is also stabilized via coaxial stacking on adjacent hybridization to targets. We conclude with a discussion on the implementation of the proposed sandwich hybridization assay as a high-throughput microRNA detection method.

  8. Synthesis and structural characterization of sialic acid-glutamic acid hybrid foldamers as conformational surrogates of alpha-2,8-linked polysialic acid.

    PubMed

    Saludes, Jonel P; Ames, James B; Gervay-Hague, Jacquelyn

    2009-04-22

    Surface expression of alpha-(2,8)-linked polymers of sialic acid in adult tissues has been correlated with metastasis of several human cancers. One approach to chemotherapeutic intervention against the spread of these cancers involves the development of immunogenic molecules that elicit an antibody response against alpha-(2,8)-linked polysialic acids. Naturally occurring polysialic acids are not viable candidates because they are present during embryonic development and are recognized as self by the immune system. These natural polymers also have poor pharmacokinetic properties because they are readily degraded by neuraminidase enzymes. We have been interested in developing structural surrogates of polysialic acids in an effort to overcome these limitations. Reported herein are microwave-assisted solid-phase peptide syntheses and structural characterization studies of a series of alpha/delta hybrid peptides derived from Fmoc-Neu2en and Fmoc-Glu(OtBu)-OH. Conformational experiments including circular dichroism, NH/ND exchange, and ROESY in aqueous solution were performed to study the secondary structures of these hybrid foldamers. ROESY data were analyzed with the assistance of XPLOR-NIH that was modified to include parameter and topology files to accommodate unnatural amino acids and the delta amide linkages. The results indicate that stable secondary structure is dependent upon both the amino acid sequence and the configuration of Glu. The most stable foldamer was composed of a total of 6 residues beginning with L-Glu at the carboxy terminus and alternating Neu2en and L-Glu residues. In water, this foldamer adopts a right-handed helical conformation with 3.7 residues per turn, 7.4 A pitch, 5.8 A diameter, and a length of 18.5 A, which is stabilized by both classical C=O...H-N backbone interactions and by pyranose ring O and L-Glu HN H-bonding. These structural features orient the L-Glu carboxylates along the helical backbone with a periodicity that matches the

  9. Fabrication of gelatin-hyaluronic acid hybrid scaffolds with tunable porous structures for soft tissue engineering.

    PubMed

    Zhang, Fan; He, Chuanglong; Cao, Lijun; Feng, Wei; Wang, Hongsheng; Mo, Xiumei; Wang, Jinwu

    2011-04-01

    The development of three-dimensional (3-D) scaffolds with highly open porous structure is one of the most important issues in tissue engineering. In this study, 3-D macroporous gelatin/hyaluronic acid (GE/HA) hybrid scaffolds with varying porous morphology were prepared by freeze-drying their blending solutions and subsequent chemical crosslinking by using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). The resulting scaffolds were characterized by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). Their swelling, in vitro degradation properties and compressive strength were also investigated. To evaluate in vitro cytocompatibility of scaffolds, mouse L929 fibroblasts were seeded onto the scaffolds for cell morphology and cell viability studies. It was found that the porous structure of scaffolds can be tailored by varying the ratios of gelatin to HA, both the swelling ratios and degradation rate increased with the increase of HA content in hybrid scaffolds, and crosslinking the scaffolds with EDC improved the degradation resistance of the scaffold in culture media and increased the mechanical strength of scaffolds. The in vitro results revealed that the prepared scaffolds do not induce cytotoxic effects and suitable for cell growth, especially in the case of scaffolds with higher gelatin content. The combined results of the physicochemical and biological studies suggested that the developed GE/HA hybrid scaffolds exhibit good potential and biocompatibility for soft tissue engineering applications. Copyright © 2011 Elsevier B.V. All rights reserved.

  10. Fibrillar networks of glycyrrhizic acid for hybrid nanomaterials with catalytic features.

    PubMed

    Saha, Abhijit; Adamcik, Jozef; Bolisetty, Sreenath; Handschin, Stephan; Mezzenga, Raffaele

    2015-04-27

    Self-assembly of the naturally occurring sweetening agent, glycyrrhizic acid (GA) in water is studied by small-angle X-ray scattering and microscopic techniques. Statistical analysis on atomic force microscopy images reveals the formation of ultralong GA fibrils with uniform thickness of 2.5 nm and right-handed twist with a pitch of 9 nm, independently of GA concentration. Transparent nematic GA hydrogels are exploited to create functional hybrid materials. Two-fold and three-fold hybrids are developed by introducing graphene oxide (GO) and in situ-synthesized gold nanoparticles (Au NPs) in the hydrogel matrix for catalysis applications. In the presence of GO, the catalytic efficiency of Au NPs in the reduction of p-nitrophenol to p-aminophenol is enhanced by 2.5 times. Gold microplate single crystals are further synthesized in the GA hydrogel, expanding the scope of these hybrids and demonstrating their versatility in materials design. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Direct RNA detection without nucleic acid purification and PCR: Combining sandwich hybridization with signal amplification based on branched hybridization chain reaction.

    PubMed

    Xu, Yao; Zheng, Zhi

    2016-05-15

    We have developed a convenient, robust and low-cost RNA detection system suitable for high-throughput applications. This system uses a highly specific sandwich hybridization to capture target RNA directly onto solid support, followed by on-site signal amplification via 2-dimensional, branched hybridizing chain polymerization through toehold-mediated strand displacement reaction. The assay uses SYBR Green to detect targets at concentrations as low as 1 pM, without involving nucleic acid purification or any enzymatic reaction, using ordinary oligonucleotides without modification or labeling. The system was demonstrated in the detection of malaria RNA in blood and GAPDH gene expression in cell lysate.

  12. Tartaric Acid-Assisted Self-Assembly of Hybrid Block Copolymer Composites

    NASA Astrophysics Data System (ADS)

    Yao, Li; Lin, Ying; Watkins, James

    2014-03-01

    Enantiopure tartaric acid was used as an additive to increase the segregation strength of poly(ethylene oxide-block-tert-butyl acrylate) (PEO-b-PtBA) copolymers through strong, selective interactions with one of the polymer chain segments. Addition of tartaric acid to PEO-b-PtBA exhibiting cylindrical morphologies resulted in the formation of helical superstructures as observed by transmission electron microscopy. It was also found that this small acid additive can also enable phase-selective ultra-high loading of nanoparticles (NPs) into target domains of the block copolymer composites. The loading of tartaric acid can increase enthalpically favorable interactions between the nanoparticle ligands and the host domain and mitigate entropic penalties associated with NP incorporation into the target domain. A metal content of over 40 weight percent by mass of the resulting well ordered composites was achieved as measured by thermal gravimetric analysis in PEO-b-PtBA/tartaric acid/4-hydroxythiophenol functionalized Au NP hybrid system. Funding from Center for Hierarchical Manufacturing (CHM); Facility support from Materials Research Science and Engineering Center at UMass Amherst.

  13. Chlorogenic acid-arabinose hybrid domains in coffee melanoidins: Evidences from a model system.

    PubMed

    Moreira, Ana S P; Coimbra, Manuel A; Nunes, Fernando M; Passos, Cláudia P; Santos, Sónia A O; Silvestre, Armando J D; Silva, André M N; Rangel, Maria; Domingues, M Rosário M

    2015-10-15

    Arabinose from arabinogalactan side chains was hypothesized as a possible binding site for chlorogenic acids in coffee melanoidins. To investigate this hypothesis, a mixture of 5-O-caffeoylquinic acid (5-CQA), the most abundant chlorogenic acid in green coffee beans, and (α1 → 5)-L-arabinotriose, structurally related to arabinogalactan side chains, was submitted to dry thermal treatments. The compounds formed during thermal processing were identified by electrospray ionization mass spectrometry (ESI-MS) and characterized by tandem MS (ESI-MS(n)). Compounds composed by one or two CQAs covalently linked with pentose (Pent) residues (1-12) were identified, along with compounds bearing a sugar moiety but composed exclusively by the quinic or caffeic acid moiety of CQAs. The presence of isomers was demonstrated by liquid chromatography online coupled to ESI-MS and ESI-MS(n). Pent1-2CQA were identified in coffee samples. These results give evidence for a diversity of chlorogenic acid-arabinose hybrids formed during roasting, opening new perspectives for their identification in melanoidin structures. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Radical generating coordination complexes as tools for rapid and effective fragmentation and fluorescent labeling of nucleic acids for microchip hybridization.

    SciTech Connect

    Kelly, J. J.; Chernov, B. N.; Mirzabekov, A. D.; Bavykin, S. G.; Biochip Technology Center; Northwestern Univ.; Engelhardt Inst. of Molecular Biology

    2002-01-01

    DNA microchip technology is a rapid, high-throughput method for nucleic acid hybridization reactions. This technology requires random fragmentation and fluorescent labeling of target nucleic acids prior to hybridization. Radical-generating coordination complexes, such as 1,10-phenanthroline-Cu(II) (OP-Cu) and Fe(II)-EDTA (Fe-EDTA), have been commonly used as sequence nonspecific 'chemical nucleases' to introduce single-strand breaks in nucleic acids. Here we describe a new method based on these radical-generating complexes for random fragmentation and labeling of both single- and double-stranded forms of RNA and DNA. Nucleic acids labeled with the OP-Cu and the Fe-EDTA protocols revealed high hybridization specificity in hybridization with DNA microchips containing oligonucleotide probes selected for identification of 16S rRNA sequences of the Bacillus group microorganisms.We also demonstrated cDNA- and cRNA-labeling and fragmentation with this method. Both the OP-Cu and Fe-EDTA fragmentation and labeling procedures are quick and inexpensive compared to other commonly used methods. A column-based version of the described method does not require centrifugation and therefore is promising for the automation of sample preparations in DNA microchip technology as well as in other nucleic acid hybridization studies.

  15. Particle-rod hybrids: growth of arachidic acid molecular rods from capped cadmium selenide nanoparticles.

    PubMed

    Chen, Dongzhong; Wang, Ruomiao; Arachchige, Indika; Mao, Guangzhao; Brock, Stephanie L

    2004-12-22

    This communication describes a spin-coating method to nucleate organic molecular rods of uniform size from an inorganic nanoparticle at a solid surface. The particle-rod hybrid structure spontaneously forms when a film is spin coated from a mixed 2-propanol solution of arachidic acid (AA) and nanoparticles of cadmium selenide capped by mercaptoundecanoic acid (MUA-CdSe) on graphite. AFM images show that MUA-CdSe nanoparticles nucleate single crystalline rods of AA with a cross section of a single unit cell of the C-form. The solution-based process potentially allows the precise tuning of the wetting profile of the solution on the surface-attached nanoparticle, which provides the reservoir for the growth of the single crystalline rods. The results suggest that nanoparticles can be regarded as nanoseeds for the nucleation of guest crystals. It should be possible to further functionalize the AA rods by electrostatic complexation with metal or organic ions.

  16. Spiral wound valve-regulated lead-acid batteries for hybrid vehicles

    NASA Astrophysics Data System (ADS)

    Soria, M. L.; Trinidad, F.; Lacadena, J. M.; Valenciano, J.; Arce, G.

    Future vehicle applications require the development of reliable and long life batteries operating under high-rate partial-state-of-charge (HRPSoC) working conditions. This paper updates work carried out to develop spiral wound valve-regulated lead-acid (VRLA) batteries for vehicles with different hybridisation degrees, ranging from stop-start to mild hybrid applications. Former work on design optimisation and active material formulations has been implemented in two spiral wound VRLA batteries, rated 12 V 50 Ah and 6 V 24 Ah, and these two products are currently being tested both in benches and in vehicles with different hybridisation degrees within a demonstration project funded by the Advanced Lead Acid Battery Consortium and in collaboration with several European vehicle and electrical component manufacturers.

  17. Detection of bluetongue virus serotype 17 in Culicoides variipennis by nucleic acid blot and sandwich hybridization techniques.

    PubMed Central

    Schoepp, R J; Bray, J F; Olson, K E; el-Hussein, A; Holbrook, F R; Blair, C D; Roy, P; Beaty, B J

    1990-01-01

    Molecular hybridization techniques were developed for the detection and surveillance of bluetongue virus (BTV) serotype 17 in the insect vector Culicoides variipennis, a biting midge. Radiolabeled RNA and cDNA probes were generated from sequences of the L3 segment of BTV serotype 17. These probes were used to detect BTV RNA in pools of infected C. variipennis by hybridizing the probes directly to analyte immobilized on nylon membranes or by using a nucleic acid sandwich hybridization test. Hybridization procedures were able to detect 1 infected C. variipennis in a pool of 50 and as little as 3.55 log10 50% tissue culture infective doses per ml of virus. These hybridization techniques provide an alternative to virus isolation for the surveillance of BTV in vector populations. Images PMID:2172294

  18. Design of hybrid multimodal poly(lactic-co-glycolic acid) polymer nanoparticles for neutrophil labeling, imaging and tracking

    NASA Astrophysics Data System (ADS)

    Qiu, Yuan; Palankar, Raghavendra; Echeverría, María; Medvedev, Nikolay; Moya, Sergio E.; Delcea, Mihaela

    2013-11-01

    Poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) incorporating quantum dots (QDs), superparamagnetic iron oxide nanoparticles (SPIONs) and gold (Au) NPs for neutrophil labeling were fabricated via the w/o/w double emulsion method. QDs and SPIONs were entrapped in the PLGA core during emulsification while Au NPs were assembled on top of the PLGA NPs via electrostatic interactions. Transmission Electron Microscopy, Scanning Electron Microscopy and Confocal Scanning Laser Microscopy (CLSM) were applied to characterize the hybrid PLGA NPs. The uptake of the hybrid PLGA NPs by human neutrophils was studied by Flow Cytometry and confocal microscopy. In addition, the induction of reactive oxygen species (ROS) in neutrophils after incubation with the hybrid PLGA NPs was assessed. Magnetophoresis experiments showed that neutrophils with internalized hybrid PLGA NPs can be effectively laterally displaced towards the magnetic field. Magnetic Resonance Imaging of the hybrid PLGA NPs resulted in images with a contrast enhancement linearly dependent on the concentration of the hybrid PLGA NPs. Research reported in this work is relevant for imaging, tracking and manipulating neutrophils and has potential for in vivo applications, e.g., tumor visualization and localized photothermal treatment.Poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) incorporating quantum dots (QDs), superparamagnetic iron oxide nanoparticles (SPIONs) and gold (Au) NPs for neutrophil labeling were fabricated via the w/o/w double emulsion method. QDs and SPIONs were entrapped in the PLGA core during emulsification while Au NPs were assembled on top of the PLGA NPs via electrostatic interactions. Transmission Electron Microscopy, Scanning Electron Microscopy and Confocal Scanning Laser Microscopy (CLSM) were applied to characterize the hybrid PLGA NPs. The uptake of the hybrid PLGA NPs by human neutrophils was studied by Flow Cytometry and confocal microscopy. In addition, the induction of

  19. p-Aminophenylacetic acid-mediated synthesis of monodispersed titanium oxide hybrid microspheres in ethanol solution.

    PubMed

    Zhang, Hongye; Xie, Yun; Liu, Zhimin; Tao, Ranting; Sun, Zhenyu; Ding, Kunlun; An, Guimin

    2009-10-15

    Monodispersed TiO2 hybrid microspheres were prepared via the hydrolysis of titanium isopropoxide (TTIP) in ethanol solution containing p-aminophenylacetic acid (APA). The effects of the APA:TTIP molar ratio, water content, reaction time and reaction temperature on the morphology of the resultant spheres were investigated. The products were characterized by scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, thermogravimetric analysis, and X-ray diffraction. It was demonstrated that the diameters of the resultant TiO2 spheres could be tuned in the range of 380-800 nm by changing the APA:TTIP molar ratio (1:3 to 3:1) and water content (1-3 v/v%) in the reaction medium, and that increasing the APA:TTIP molar ratio led to larger TiO2 hybrid spheres while increasing the water content decreased their size. The loading content of APA in the hybrid spheres could reach 20 wt.% as they were prepared with the APA:TTIP ratio of 3:1. The possible formation mechanism of the hybrid spheres was also investigated. It was found that APA slowed down the hydrolysis rate of the titanium precursor so that resulted in the formation of the TiO2 spheres. In addition, the APA present in TiO2 spheres acted as a reducing agent to in situ convert HAuCl4 into metallic Au on the surface of the TiO2 spheres. The catalytic activity of the resultant Au/APA-TiO2 composite was examined using transfer hydrogenation of phenylacetone with 2-propanol, and it was indicated that the catalyst displayed high efficiency for this reaction.

  20. Development of rapeseed with high erucic acid content by asymmetric somatic hybridization between Brassica napus and Crambe abyssinica.

    PubMed

    Wang, Y P; Sonntag, K; Rudloff, E

    2003-05-01

    PEG-induced asymmetric somatic hybridization between Brassica napus and Crambe abyssinica was carried out. C. abyssinica is an annual cruciferous oil crop with a high content of erucic acid in the seed oil valuable for technical purposes. UV-irradiated mesophyll protoplasts of C. abyssinica cv 'Carmen' and cv 'Galactica' were fused with hypocotyl protoplasts of different genotypes of B. napus cv 'Maplus' and breeding line '11502'. Shoot regeneration frequency varied between 6.1% and 20.8% among the different doses of UV-irradiation, ranging from 0.05 J/cm(2) to 0.30 J/cm(2). In total, 124 shoots were regenerated, of which 20 asymmetric somatic hybrids were obtained and verified by nuclear DNA content and AFLP analysis. AFLP data showed that some of the characteristic bands from C. abyssinica were present in the hybrids. Cytological analysis of these hybrids showed that 9 out of 20 asymmetric hybrids had 38 chromosomes, the others contained 40-78 chromosomes, having additional chromosomes between 2 and 40 beyond the 38 expected for B. napus. The investigation into the fertility of asymmetric somatic hybrids indicated that the fertility increased with increasing UV-doses ranging from 0.05 J/cm(2) to 0.15 J/cm(2). All of the hybrids were cultured to full maturity, and could be fertilized and set seeds after self-pollination or backcrosses with B. napus. An analysis of fatty acid composition in the seeds was conducted and found to contain significantly greater amounts of erucic acid than B. napus. This study indicates that UV-irradiation could be used as a tool to produce asymmetric somatic hybrids and to promote the fertility of the hybrids.

  1. Fatty acid membrane assembly on coacervate microdroplets as a step towards a hybrid protocell model

    NASA Astrophysics Data System (ADS)

    Dora Tang, T.-Y.; Rohaida Che Hak, C.; Thompson, Alexander J.; Kuimova, Marina K.; Williams, D. S.; Perriman, Adam W.; Mann, Stephen

    2014-06-01

    Mechanisms of prebiotic compartmentalization are central to providing insights into how protocellular systems emerged on the early Earth. Protocell models are based predominantly on the membrane self-assembly of fatty-acid vesicles, although membrane-free scenarios that involve liquid-liquid microphase separation (coacervation) have also been considered. Here we integrate these alternative models of prebiotic compartmentalization and develop a hybrid protocell model based on the spontaneous self-assembly of a continuous fatty-acid membrane at the surface of preformed coacervate microdroplets prepared from cationic peptides/polyelectrolytes and adenosine triphosphate or oligo/polyribonucleotides. We show that the coacervate-supported membrane is multilamellar, and mediates the selective uptake or exclusion of small and large molecules. The coacervate interior can be disassembled without loss of membrane integrity, and fusion and growth of the hybrid protocells can be induced under conditions of high ionic strength. Our results highlight how notions of membrane-mediated compartmentalization, chemical enrichment and internalized structuration can be integrated in protocell models via simple chemical and physical processes.

  2. Amplified detection of nucleic acid by G-quadruplex based hybridization chain reaction.

    PubMed

    Dong, Juan; Cui, Xin; Deng, Yun; Tang, Zhuo

    2012-01-01

    A protein-free, isothermal, self-amplified nucleic acid sensing system which was a G-quadruplex integrated hybridization chain reaction (GQ-HCR) system was developed. The G-quadruplex was closed two-thirds in the loop and one-third in the stem of one of the GQ-HCR hairpin probes. In the absence of the target molecule, the GQ-HCR probes stayed as inactive meta-stable hairpin structures and the G-quadruplex was inert. Reversely, the GQ-HCR probes could be cross-opened to start a hybridization chain reaction and the closed G-quadruplex could be released to be free when the GQ-HCR probes came across the target molecule. The GQ-HCR nucleic acid sensing system could detect as low as 7.5 nM ssDNA or RNA by the colorimetric method and 4 nM ssDNA by the fluorometric method. Less than 10 copies of dsDNA template could also be detected when PCR was combined with the GQ-HCR system (PCR+GQ-HCR). Because of these advantages, the GQ-HCR system was also studied for application in visual chip detection to obtain a satisfactory repeatable and specific result.

  3. Hybrid Processes Combining Photocatalysis and Ceramic Membrane Filtration for Degradation of Humic Acids in Saline Water

    PubMed Central

    Song, Lili; Zhu, Bo; Gray, Stephen; Duke, Mikel; Muthukumaran, Shobha

    2016-01-01

    This study explored the combined effects of photocatalysis with ceramic membrane filtration for the removal of humic acid in the presence of salt; to simulate saline wastewater conditions. The effects of operating parameters, such as salinity and TiO2 concentration on permeate fluxes, total organic carbon (TOC), and UV absorbance removal, were investigated. The interaction between the humic acids and TiO2 photocatalyst played an important role in the observed flux change during ceramic membrane filtration. The results for this hybrid system showed that the TOC removal was more than 70% for both without NaCl and with the 500 ppm NaCl concentration, and 62% and 66% for 1000 and 2000 ppm NaCl concentrations. The reduction in UV absorbance was more complete in the absence of NaCl compared to the presence of NaCl. The operation of the integrated photoreactor-ceramic membrane filter over five repeat cycles is described. It can be concluded that the overall removal performance of the hybrid system was influenced by the presence of salts, as salt leads to agglomeration of TiO2 particles by suppressing the stabilising effects of electrostatic repulsion and thereby reduces the effective surface contact between the pollutant and the photocatalyst. PMID:26938568

  4. Hybrid Processes Combining Photocatalysis and Ceramic Membrane Filtration for Degradation of Humic Acids in Saline Water.

    PubMed

    Song, Lili; Zhu, Bo; Gray, Stephen; Duke, Mikel; Muthukumaran, Shobha

    2016-03-01

    This study explored the combined effects of photocatalysis with ceramic membrane filtration for the removal of humic acid in the presence of salt; to simulate saline wastewater conditions. The effects of operating parameters, such as salinity and TiO₂ concentration on permeate fluxes, total organic carbon (TOC), and UV absorbance removal, were investigated. The interaction between the humic acids and TiO₂ photocatalyst played an important role in the observed flux change during ceramic membrane filtration. The results for this hybrid system showed that the TOC removal was more than 70% for both without NaCl and with the 500 ppm NaCl concentration, and 62% and 66% for 1000 and 2000 ppm NaCl concentrations. The reduction in UV absorbance was more complete in the absence of NaCl compared to the presence of NaCl. The operation of the integrated photoreactor-ceramic membrane filter over five repeat cycles is described. It can be concluded that the overall removal performance of the hybrid system was influenced by the presence of salts, as salt leads to agglomeration of TiO₂ particles by suppressing the stabilising effects of electrostatic repulsion and thereby reduces the effective surface contact between the pollutant and the photocatalyst.

  5. Characterization of hybrid plasmids carrying individual ribosomal ribonucleic acid transcription units of Escherichia coli.

    PubMed Central

    Kenerley, M E; Morgan, E A; Post, L; Lindahl, L; Nomura, M

    1977-01-01

    We have screened the strains with ColE1 hybrid plasmids constructed by Clarke and Carbon (Cell 9:91-99, 1976) for the presence of ribosomal ribonucleic acid (rRNA) genes on the plasmids and identified 16 strains whose plasmids carry rRNA genes. The structures of these 16 plasmids were compared by heteroduplex analysis, and the plasmids were classified into six groups on the basis of their chromosomal origins. Homology with known transducing-phage deoxyribonucleic acids and genetic mapping have assigned locations on the Escherichia coli chromosome to three of the six groups. These are rrnB near rif at 88 min, rrnC near ilvE at 83 min, and rrnD near aroE at 71 min. A fourth group is probably rrnA at 85 min (T. Ikemura and M. Nomura, Cell, 11:779-793, 1977). We conclude that the minimum number of rRNA transcription units per haploid chromosomes is seven, that is, the six groups identified in this work plus a known operon (rrnE near metA at 89 min) that we failed to find among the hybrid plasmids. This heteroduplex analysis also suggests that there are only two kinds of rRNA operons with respect to their spacer region; three of the six rRNA operon groups studied here have one kind, whereas the remaining three have the other kind. Images PMID:336613

  6. Fatty acid membrane assembly on coacervate microdroplets as a step towards a hybrid protocell model.

    PubMed

    Dora Tang, T-Y; Rohaida Che Hak, C; Thompson, Alexander J; Kuimova, Marina K; Williams, D S; Perriman, Adam W; Mann, Stephen

    2014-06-01

    Mechanisms of prebiotic compartmentalization are central to providing insights into how protocellular systems emerged on the early Earth. Protocell models are based predominantly on the membrane self-assembly of fatty-acid vesicles, although membrane-free scenarios that involve liquid-liquid microphase separation (coacervation) have also been considered. Here we integrate these alternative models of prebiotic compartmentalization and develop a hybrid protocell model based on the spontaneous self-assembly of a continuous fatty-acid membrane at the surface of preformed coacervate microdroplets prepared from cationic peptides/polyelectrolytes and adenosine triphosphate or oligo/polyribonucleotides. We show that the coacervate-supported membrane is multilamellar, and mediates the selective uptake or exclusion of small and large molecules. The coacervate interior can be disassembled without loss of membrane integrity, and fusion and growth of the hybrid protocells can be induced under conditions of high ionic strength. Our results highlight how notions of membrane-mediated compartmentalization, chemical enrichment and internalized structuration can be integrated in protocell models via simple chemical and physical processes.

  7. SNBRFinder: A Sequence-Based Hybrid Algorithm for Enhanced Prediction of Nucleic Acid-Binding Residues

    PubMed Central

    Sun, Jun; Liu, Rong

    2015-01-01

    Protein-nucleic acid interactions are central to various fundamental biological processes. Automated methods capable of reliably identifying DNA- and RNA-binding residues in protein sequence are assuming ever-increasing importance. The majority of current algorithms rely on feature-based prediction, but their accuracy remains to be further improved. Here we propose a sequence-based hybrid algorithm SNBRFinder (Sequence-based Nucleic acid-Binding Residue Finder) by merging a feature predictor SNBRFinderF and a template predictor SNBRFinderT. SNBRFinderF was established using the support vector machine whose inputs include sequence profile and other complementary sequence descriptors, while SNBRFinderT was implemented with the sequence alignment algorithm based on profile hidden Markov models to capture the weakly homologous template of query sequence. Experimental results show that SNBRFinderF was clearly superior to the commonly used sequence profile-based predictor and SNBRFinderT can achieve comparable performance to the structure-based template methods. Leveraging the complementary relationship between these two predictors, SNBRFinder reasonably improved the performance of both DNA- and RNA-binding residue predictions. More importantly, the sequence-based hybrid prediction reached competitive performance relative to our previous structure-based counterpart. Our extensive and stringent comparisons show that SNBRFinder has obvious advantages over the existing sequence-based prediction algorithms. The value of our algorithm is highlighted by establishing an easy-to-use web server that is freely accessible at http://ibi.hzau.edu.cn/SNBRFinder. PMID:26176857

  8. Crude palm oil from interspecific hybrid Elaeis oleifera×Elaeis guineensis: fatty acid regiodistribution and molecular species of glycerides.

    PubMed

    Mozzon, Massimo; Pacetti, Deborah; Lucci, Paolo; Balzano, Michele; Frega, Natale Giuseppe

    2013-11-01

    The composition and structure of triacylglycerols (TAGs) and partial glycerides of crude palm oil obtained from interspecific hybrid Elaeis oleifera×Elaeis guineensis, grown in Colombia, were fully characterised and compared to data obtained by analysing crude African palm oil. Hybridisation appears to substantially modify the biosynthesis of fatty acids (FAs) rather than their assembly in TAGs. In fact, total FAs analysis showed significant differences between these two types of oil, with hybrid palm oil having a higher percentage of oleic acid (54.6 ± 1.0 vs 41.4 ± 0.3), together with a lower saturated fatty acid content (33.5 ± 0.5 vs 47.3 ± 0.1), while the percentage of essential fatty acid, linoleic acid, does not undergo significant changes. Furthermore, 34 TAG types were identified, with no qualitative differences between African and E. guineensis×E. oleifera hybrid palm oil samples. Short and medium chain FAs (8:0, 10:0, 12:0, 14:0) were utilised, together, to build a restricted number of TAG molecular species. Oil samples from the E. guineensis×E. oleifera hybrid showed higher contents of monosaturated TAGs (47.5-51.0% vs 36.7-37.1%) and triunsaturated TAGs (15.5-15.6% vs 5.2-5.4%). The sn-2 position of TAGs in hybrid palm oil was shown to be predominantly esterified with oleic acid (64.7-66.0 mol% vs 55.1-58.2 mol% in African palm oil) with only 10-15% of total palmitic acid and 6-20% of stearic acid acylated in the secondary position. The total amount of diacylglycerols (DAGs) was in agreement with the values of free acidity; DAG types found were in agreement with the representativeness of different TAG species. Copyright © 2013 Elsevier Ltd. All rights reserved.

  9. Effect of Backbone Design on Hybridization Thermodynamics of Oligo-nucleic Acids: A Coarse-Grained Molecular Dynamics Simulation Study

    NASA Astrophysics Data System (ADS)

    Ghobadi, Ahmadreza F.; Jayaraman, Arthi

    DNA hybridization is the basis of various bio-nano technologies, such as DNA origami and assembly of DNA-functionalized nanoparticles. A hybridized double stranded (ds) DNA is formed when complementary nucleobases on hybridizing strands exhibit specific and directional hydrogen bonds through canonical Watson-Crick base-pairing interactions. In recent years, the need for cheaper alternatives and significant synthetic advances have driven design of DNA mimics with new backbone chemistries. However, a fundamental understanding of how these backbone modifications in the oligo-nucleic acids impact the hybridization and melting behavior of the duplex is still lacking. In this talk, we present our recent findings on impact of varying backbone chemistry on hybridization of oligo-nucleic acid duplexes. We use coarse-grained molecular dynamics simulations to isolate the effect of strand flexibility, electrostatic interactions and nucleobase spacing on the melting curves for duplexes with various strand sequences and concentrations. Since conjugation of oligo-nucleic acids with polymers serve as building blocks for thermo-responsive polymer networks and gels, we also present the effect of such conjugation on hybridization thermodynamics and polymer conformation.

  10. Salinomycin Hydroxamic Acids: Synthesis, Structure, and Biological Activity of Polyether Ionophore Hybrids.

    PubMed

    Borgström, Björn; Huang, Xiaoli; Chygorin, Eduard; Oredsson, Stina; Strand, Daniel

    2016-06-09

    The polyether ionophore salinomycin has recently gained attention due to its exceptional ability to selectively reduce the proportion of cancer stem cells within a number of cancer cell lines. Efficient single step strategies for the preparation of hydroxamic acid hybrids of this compound varying in N- and O-alkylation are presented. The parent hydroxamic acid, salinomycin-NHOH, forms both inclusion complexes and well-defined electroneutral complexes with potassium and sodium cations via 1,3-coordination by the hydroxamic acid moiety to the metal ion. A crystal structure of an cationic sodium complex with a noncoordinating anion corroborates this finding and, moreover, reveals a novel type of hydrogen bond network that stabilizes the head-to-tail conformation that encapsulates the cation analogously to the native structure. The hydroxamic acid derivatives display down to single digit micromolar activity against cancer cells but unlike salinomycin selective reduction of ALDH(+) cells, a phenotype associated with cancer stem cells was not observed. Mechanistic implications are discussed.

  11. Development of SSR Markers Linked to Low Hydrocyanic Acid Content in Sorghum-Sudan Grass Hybrid Based on BSA Method.

    PubMed

    Xiao-Xia, Yu; Zhi-Hua, Liu; Zhuo, Yu; Yue, Shi; Xiao-Yu, Li

    2016-01-01

    Sorghum-Sudan grass hybrid containing high hydrocyanic acid content can cause hydrocyanic acid poisoning to the livestock and limit the popularization of this forage crop. Molecular markers associated with low hydrocyanic acid content can speed up the process of identification of genotypes with low hydrocyanic acid content. In the present study, 11 polymorphic SSR primers were screened and used for bulked segregant analysis and single marker analysis. Three SSR markers Xtxp7230, Xtxp7375 and Bnlg667960 associated with low hydrocyanic acid content were rapidly identified by BSA. In single marker analysis, six markers Xtxp7230, Xtxp7375, Bnlg667960, Xtxp67-11, Xtxp295-7 and Xtxp12-9 were linked to low hydrocyanic acid content, which explained the proportion of phenotypic variation from 7.6 % to 41.2 %. The markers identified by BSA were also verified by single marker analysis. The three SSR marker bands were then cloned and sequenced for sequence homology analysis in NCBI. It is the first report on the development of molecular markers associated with low hydrocyanic acid content in sorghum- Sudan grass hybrid. These markers will be useful for genetic improvement of low hydrocyanic acid sorghum-Sudan grass hybrid by marker-assisted breeding.

  12. One-pot transformation of cellobiose to formic acid and levulinic acid over ionic-liquid-based polyoxometalate hybrids.

    PubMed

    Li, Kaixin; Bai, Linlu; Amaniampong, Prince Nana; Jia, Xinli; Lee, Jong-Min; Yang, Yanhui

    2014-09-01

    Currently, levulinic acid (LA) and formic acid (FA) are considered as important carbohydrates for the production of value-added chemicals. Their direct production from biomass will open up a new opportunity for the transformation of biomass resource to valuable chemicals. In this study, one-pot transformation of cellobiose into LA and FA was demonstrated, using a series of multiple-functional ionic liquid-based polyoxometalate (IL-POM) hybrids as catalytic materials. These IL-POMs not only markedly promoted the production of valuable chemicals including LA, FA and monosaccharides with high selectivities, but also provided great convenience of the recovery and the reuse of the catalytic materials in an environmentally friendly manner. Cellobiose conversion of 100%, LA selectivity of 46.3%, and FA selectivity of 26.1% were obtained at 423 K and 3 MPa for 3 h in presence of oxygen. A detailed catalytic mechanism for the one-pot transformation of cellobiose was also presented. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Effect of backbone chemistry on hybridization thermodynamics of oligonucleic acids: a coarse-grained molecular dynamics simulation study.

    PubMed

    Ghobadi, Ahmadreza F; Jayaraman, Arthi

    2016-02-28

    In this paper we study how varying oligonucleic acid backbone chemistry affects the hybridization/melting thermodynamics of oligonucleic acids. We first describe the coarse-grained (CG) model with tunable parameters that we developed to enable the study of both naturally occurring oligonucleic acids, such as DNA, and their chemically-modified analogues, such as peptide nucleic acids (PNAs) and locked nucleic acids (LNAs). The DNA melting curves obtained using such a CG model and molecular dynamics simulations in an implicit solvent and with explicit ions match with the melting curves obtained using the empirical nearest-neighbor models. We use these CG simulations to then elucidate the effect of backbone flexibility, charge, and nucleobase spacing along the backbone on the melting curves, potential energy and conformational entropy change upon hybridization and base-pair hydrogen bond residence time. We find that increasing backbone flexibility decreases duplex thermal stability and melting temperature mainly due to increased conformational entropy loss upon hybridization. Removing charges from the backbone enhances duplex thermal stability due to the elimination of electrostatic repulsion and as a result a larger energetic gain upon hybridization. Lastly, increasing nucleobase spacing decreases duplex thermal stability due to decreasing stacking interactions that are important for duplex stability.

  14. Synthesis and conformational analysis of hybrid α/β-dipeptides incorporating S-glycosyl-β(2,2)-amino acids.

    PubMed

    García-González, Iván; Mata, Lara; Corzana, Francisco; Jiménez-Osés, Gonzalo; Avenoza, Alberto; Busto, Jesús H; Peregrina, Jesús M

    2015-01-12

    We synthesized and carried out the conformational analysis of several hybrid dipeptides consisting of an α-amino acid attached to a quaternary glyco-β-amino acid. In particular, we combined a S-glycosylated β(2,2)-amino acid and two different types of α-amino acid, namely, aliphatic (alanine) and aromatic (phenylalanine and tryptophan) in the sequence of hybrid α/β-dipeptides. The key step in the synthesis involved the ring-opening reaction of a chiral cyclic sulfamidate, inserted in the peptidic sequence, with a sulfur-containing nucleophile by using 1-thio-β-D-glucopyranose derivatives. This reaction of glycosylation occurred with inversion of configuration at the quaternary center. The conformational behavior in aqueous solution of the peptide backbone and the glycosidic linkage for all synthesized hybrid glycopeptides was analyzed by using a protocol that combined NMR experiments and molecular dynamics with time-averaged restraints (MD-tar). Interestingly, the presence of the sulfur heteroatom at the quaternary center of the β-amino acid induced θ torsional angles close to 180° (anti). Notably, this value changed to 60° (gauche) when the peptidic sequence displayed aromatic α-amino acids due to the presence of CH-π interactions between the phenyl or indole ring and the methyl groups of the β-amino acid unit. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. A hybrid Li-air battery with buckypaper air cathode and sulfuric acid electrolyte

    SciTech Connect

    Li, YF; Huang, K; Xing, YC

    2012-10-30

    We demonstrate a type of carbon nanotube based buckypaper cathode in a hybrid electrolyte Li-air battery (HyLAB) that showed outstanding discharging performances. The HyLAB has sulfuric acid as the catholyte and a large active electrode area (10 cm(2)). The active cathode layer was made from a buckypaper with 5 wt.% Pt supported on carbon nanotubes (Pt/CNTs) for oxygen reduction and evolution. A similar cathode was constructed with a catalyst of 5 wt.% Pt supported on carbon black (Pt/CB). It is demonstrated that sulfuric acid can achieve high discharging current densities while maintaining relatively high cell potentials. The cell with Pt/CNTs showed a much better performance than with Pt/CB at high current densities. The HyLAB with Pt/CNTs achieved a discharging capacity of 306 mAh/g and a cell voltage of 3.15 V at 0.2 mA/cm(2). The corresponding specific energy is 1067 Wh/kg based on the total weight of the sulfuric acid. Slow decrease in performance was observed, but it can be recovered by refilling the cell with new electrolyte after continuous discharging of more than 75 h. A charge-discharge experiment at 0.2 mA/cm(2) showed that the cell was rechargeable with a capacity of more than 300 mAh/g. (c) 2012 Elsevier Ltd. All rights reserved.

  16. Development and operation of a hybrid acid-alkaline advanced water electrolysis cell

    NASA Astrophysics Data System (ADS)

    Teschke, O.; Zwanziger, M.

    A hybrid acid-alkaline water electrolysis cell has been developed for hydrogen production. The cell is based on the use of an acidic solution at the cathode and a basic solution at the anode to reduce the minimum theoretical voltage for water decomposition from the thermoneutral potential of 1.47 V to close to 1.4 V at 25 C and 1 atm. The pH differential is maintained by the removal of OH ions from the cathode section and water removal from the anode section, which can be driven by heat energy. A practical cell has been built using a solid polymer electrolyte in which, however, the cathodic compartment is not acidic but neutral. Tests with a platinum black cathode catalyst and a platinum-iridium anode catalyst have resulted in steady-state water hydrolysis at an applied voltage of 0.9 V, and a V-I diagram with a considerably lower slope than that of a conventional cell has been obtained at 90 C.

  17. Nanoleakage in Hybrid Layer and Acid-Base Resistant Zone at the Adhesive/Dentin Interface.

    PubMed

    Nikaido, Toru; Nurrohman, Hamid; Takagaki, Tomohiro; Sadr, Alireza; Ichinose, Shizuko; Tagami, Junji

    2015-10-01

    The aim of interfacial nanoleakage evaluation is to gain a better understanding of degradation of the adhesive-dentin interface. The acid-base resistant zone (ABRZ) is recognized at the bonded interface under the hybrid layer (HL) in self-etch adhesive systems after an acid-base challenge. The purpose of this study was to evaluate nanoleakage in HL and ABRZ using three self-etch adhesives; Clearfil SE Bond (SEB), Clearfil SE One (SEO), and G-Bond Plus (GBP). One of the three adhesives was applied on the ground dentin surface and light cured. The specimens were longitudinally divided into two halves. One half remained as the control group. The others were immersed in ammoniacal silver nitrate solution, followed by photo developing solution under fluorescent light. Following this, the specimens were subjected to acid-base challenges with an artificial demineralization solution (pH4.5) and sodium hypochlorite, and prepared in accordance with common procedures for transmission electron microscopy (TEM) examination. The TEM images revealed silver depositions in HL and ABRZ due to nanoleakage in all the adhesives; however, the extent of nanoleakage was material dependent. Funnel-shaped erosion beneath the ABRZ was observed only in the all-in-one adhesive systems; SEO and GBP, but not in the two-step self-etch adhesive system; SEB.

  18. Rheological, microstructural, and in vitro characterization of hybrid chitosan-polylactic acid/hydroxyapatite composites.

    PubMed

    Araújo, A B A; Lemos, A F; Ferreira, J M F

    2009-03-15

    In this work, hybrid chitosan/hydroxyapatite composites material were developed and characterized. The polymer matrix was first dissolved in polylactic acid, and then hydroxyapatite (HA) was added as filler material. The effects of the added amounts of a crosslinking agent (genipin) and of the concentrations of lactic acid, and of the presence of HA powder on the evolution of rheological properties were evaluated. A significant decrease of gelation time with increasing amounts of crosslinking agent was observed, the effect being even more pronounced in the presence of HA. The chitosan matrix and the composites with a chitosan/HA weight ratio of 2/5 were characterized using microstructural analysis and in vitro tests. The formation of large pore sizes in the chitosan-based scaffolds was favored by low concentrations of lactic acid and genipin. The in vitro tests in synthetic body fluid revealed an extensive formation of an apatitic layer onto the surface of the chitosan/HA composite scaffolds crosslinked with genipin.

  19. Enzymatic hybridization of α-lipoic acid with bioactive compounds in ionic solvents.

    PubMed

    Papadopoulou, Athena A; Katsoura, Maria H; Chatzikonstantinou, Alexandra; Kyriakou, Eleni; Polydera, Angeliki C; Tzakos, Andreas G; Stamatis, Haralambos

    2013-05-01

    The lipase-catalyzed molecular hybridization of α-lipoic acid (LA) with bioactive compounds pyridoxine, tyrosol and tyramine was performed in ionic solvents and deep eutectic solvents. The biocatalytic reactions were catalyzed by Candida antarctica lipase B immobilized onto various functionalized multi-walled carbon nanotubes (f-CNTs-CaLB), as well as by commercial Novozym 435. The use of f-CNTs-CaLB leads, in most cases, to higher conversion yields as compared to Novozym 435. The nature and ion composition of ionic solvents affect the performance of the biocatalytic process. The highest conversion yield was observed in (mtoa)NTf2. The high enzyme stability and the relatively low solubility of substrates in specific media account for the improved biocatalytic synthesis of molecular hybrids of LA. Principal component analysis was used to screen for potential lipoxygenase inhibitors. In vitro studies showed that the synthesized compounds exhibit up to 10-fold increased inhibitory activity on lipoxygenase mediated lipid peroxidation as compared to parent molecules.

  20. Assessment of methods for covalent binding of nucleic acids to magnetic beads, Dynabeads, and the characteristics of the bound nucleic acids in hybridization reactions.

    PubMed Central

    Lund, V; Schmid, R; Rickwood, D; Hornes, E

    1988-01-01

    Dynabeads are magnetic monosized beads with high stability, high uniformity, unique paramagnetic properties, low particle-particle interaction, and high dispersibility. Different reactive groups; hydroxyl, carboxyl and amino groups can be attached to the surface. Several methods for covalent attachment of DNA or oligonucleotides to the beads were investigated. Best coupling yields were obtained by carbodiimide-mediated end-attachment of 5'-phosphate and 5'-NH2 modified nucleic acids to respectively amino and carboxyl beads. The carboxyl beads showed a low degree of non-specific binding, while a better yield of end-attached nucleic acids was obtained using the amino beads. The DNA-beads worked efficiently in hybridization experiments, and the kinetics of hybridization approach those of solution hybridization. PMID:3205723

  1. Nucleic acid hybridization analyses confirm the presence of a hitherto unknown morbillivirus in Mediterranean dolphins.

    PubMed

    Bolt, G; Blixenkrone-Møller, M

    1994-08-15

    In 1990 an epidemic caused by a morbillivirus was noticed among Mediterranean dolphins. RNA was extracted from the tissues of dolphins and from cell cultures infected with a corresponding dolphin morbillivirus isolate. By nucleic acid hybridization this RNA was compared to RNA extracted from animal tissue or cell cultures infected with canine distemper virus (CDV), phocine distemper virus (PDV) or measles virus (MV). The presence of morbillivirus RNA in the dolphin tissue was demonstrated. Morbillivirus N, P, M and F gene mRNAs were detected in the RNA from dolphin morbillivirus infected cells. These mRNA species seemed to be of approximately the same size as the corresponding mRNA species of CDV, PDV and MV. The results of the comparison demonstrated that the dolphin morbillivirus is genetically different from CDV, PDV and MV. No indication of a close relationship between the dolphin isolate and either CDV, PDV or MV was found.

  2. Lead-acid batteries in micro-hybrid applications. Part II. Test proposal

    NASA Astrophysics Data System (ADS)

    Schaeck, S.; Stoermer, A. O.; Albers, J.; Weirather-Koestner, D.; Kabza, H.

    In the first part of this work [1] selected key parameters for applying lead-acid (LA) batteries in micro-hybrid power systems (MHPS) were investigated. Main results are integrated in an accelerated, comprehensive test proposal presented here. The test proposal aims at a realistic representation of the pSoC operation regime, which is described in Refs. [1,6]. The test is designed to be sensitive with respect to dynamic charge acceptance (DCA) at partially discharged state (critical for regenerative braking) and the internal resistance at high-rate discharge (critical for idling stop applications). First results are presented for up-to-date valve-regulated LA batteries with absorbent glass mat (AGM) separators. The batteries are close to the limits of the first proposal of pass/fail-criteria. Also flooded batteries were tested; the first out of ten units failed already.

  3. Fabrication of Uniform DNA-Conjugated Hydrogel Microparticles via Replica Molding for Facile Nucleic Acid Hybridization Assays

    PubMed Central

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

    2010-01-01

    We identify and investigate several critical parameters in the fabrication of single-stranded DNA conjugated poly(ethylene glycol) (PEG) microparticles based on replica molding (RM) for highly uniform and robust nucleic acid hybridization assays. The effects of PEG-diacrylate, probe DNA, and photoinitiator concentrations on the overall fluorescence and target DNA penetration depth upon hybridization are examined. Fluorescence and confocal microscopy results illustrate high conjugation capacity of probe and target DNA, femtomole sensitivity, and sequence specificity. Combined these findings demonstrate a significant step toward simple, robust, and scalable procedures to manufacture highly uniform and high capacity hybridization assay particles in a well-controlled manner by exploiting many advantages that the batch processing-based RM technique offers. We envision that the results presented here may be readily applied to rapid and high throughput hybridization assays for a wide variety of applications in bioprocess monitoring, food safety, and biological threat detection. PMID:20527819

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

    PubMed

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

    2010-07-01

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

  5. Nanoporous gold on three-dimensional nickel foam: An efficient hybrid electrode for hydrogen peroxide electroreduction in acid media

    NASA Astrophysics Data System (ADS)

    Ke, Xi; Xu, Yantong; Yu, Changchun; Zhao, Jie; Cui, Guofeng; Higgins, Drew; Li, Qing; Wu, Gang

    2014-12-01

    A hybrid structure of nanoporous gold (NPG) on three-dimensional (3D) macroporous Ni foam has been synthesized by electrodeposition of Au-Sn alloy film followed by a facile chemical dealloying process under free corrosion conditions. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) are used to characterize the morphology and structure of the NPG/Ni foam hybrids. It is shown that the Ni foam skeletons are uniformly wrapped by the NPG film which is composed of bicontinuous nanostructures consisting of interconnected ligaments and nanopores. Electroreduction of H2O2 on the NPG/Ni foam hybrid electrode in acid media is investigated by linear scan voltammetry, chronoamperometry and electrochemical impedance spectroscopy. It is found that such hierarchical porous electrode displays superior activity, durability and mass transport property for H2O2 electroreduction. These results demonstrate the potential of the NPG/Ni foam hybrid electrodes for the applications in fuel cell technology.

  6. Lead-acid batteries in micro-hybrid applications. Part I. Selected key parameters

    NASA Astrophysics Data System (ADS)

    Schaeck, S.; Stoermer, A. O.; Kaiser, F.; Koehler, L.; Albers, J.; Kabza, H.

    Micro-hybrid electric vehicles were launched by BMW in March 2007. These are equipped with brake energy regeneration (BER) and the automatic start and stop function (ASSF) of the internal combustion engine. These functions are based on common 14 V series components and lead-acid (LA) batteries. The novelty is given by the intelligent onboard energy management, which upgrades the conventional electric system to the micro-hybrid power system (MHPS). In part I of this publication the key factors for the operation of LA batteries in the MHPS are discussed. Especially for BER one is high dynamic charge acceptance (DCA) for effective boost charging. Vehicle rest time is identified as a particular negative parameter for DCA. It can be refreshed by regular fully charging at elevated charge voltage. Thus, the batteries have to be outstandingly robust against overcharge and water loss. This can be accomplished for valve-regulated lead-acid (VRLA) batteries at least if they are mounted in the trunk. ASSF goes along with frequent high-rate loads for warm cranking. The internal resistance determines the drop of the power net voltage during cranking and is preferably low for reasons of power net stability even after years of operation. Investigations have to be done with aged 90 Ah VRLA-absorbent glass mat (AGM) batteries. Battery operation at partial state-of-charge gives a higher risk of deep discharging (overdischarging). Subsequent re-charging then is likely to lead to the formation of micro-short circuits in the absorbent glass mat separator.

  7. Aminopropyl-Silica Hybrid Particles as Supports for Humic Acids Immobilization

    PubMed Central

    Sándor, Mónika; Nistor, Cristina Lavinia; Szalontai, Gábor; Stoica, Rusandica; Nicolae, Cristian Andi; Alexandrescu, Elvira; Fazakas, József; Oancea, Florin; Donescu, Dan

    2016-01-01

    A series of aminopropyl-functionalized silica nanoparticles were prepared through a basic two step sol-gel process in water. Prior to being aminopropyl-functionalized, silica particles with an average diameter of 549 nm were prepared from tetraethyl orthosilicate (TEOS), using a Stöber method. In a second step, aminopropyl-silica particles were prepared by silanization with 3-aminopropyltriethoxysilane (APTES), added drop by drop to the sol-gel mixture. The synthesized amino-functionalized silica particles are intended to be used as supports for immobilization of humic acids (HA), through electrostatic bonds. Furthermore, by inserting beside APTES, unhydrolysable mono-, di- or trifunctional alkylsilanes (methyltriethoxy silane (MeTES), trimethylethoxysilane (Me3ES), diethoxydimethylsilane (Me2DES) and 1,2-bis(triethoxysilyl)ethane (BETES)) onto silica particles surface, the spacing of the free amino groups was intended in order to facilitate their interaction with HA large molecules. Two sorts of HA were used for evaluating the immobilization capacity of the novel aminosilane supports. The results proved the efficient functionalization of silica nanoparticles with amino groups and showed that the immobilization of the two tested types of humic acid substances was well achieved for all the TEOS/APTES = 20/1 (molar ratio) silica hybrids having or not having the amino functions spaced by alkyl groups. It was shown that the density of aminopropyl functions is low enough at this low APTES fraction and do not require a further spacing by alkyl groups. Moreover, all the hybrids having negative zeta potential values exhibited low interaction with HA molecules. PMID:28787834

  8. Visual detection of nucleic acids based on lateral flow biosensor and hybridization chain reaction amplification.

    PubMed

    Ying, Na; Ju, Chuanjing; Li, Zhongyi; Liu, Wensen; Wan, Jiayu

    2017-03-01

    In this study, a new lateral flow nucleic acid biosensor (LFNAB) using hybridization chain reaction (HCR) for signal amplification was developed for visual detection of nucleic acids with high sensitivity and low cost. A "sandwich-type" detection strategy was employed in our design. The sandwich system of capture probe (CP)/target DNA/reporter probe (RP)-HCR complexes was fabricated as the sensing platform. As the initiator strand, reporter probe propagated a chain reaction of hybridization events between the two hairpin probes modified with biotin, and determined whether long nicked DNA polymers were formed. The biotin-labeled double-strand DNA polymers then introduced numerous Streptavidin (SA)-labeled gold nanoparticles (AuNPs) on the lateral flow device. The CP/target DNA/RP-HCR complexes were captured on the test zone by the specific reaction between anti-Fam monoclonal antibody (anti-Fam mAb) on the test zone and Fam of the complexes. The accumulation of AuNPs on the test zone of the biosensor enabled the visual detection of specific sequences. The detection limit of specific DNA was as low as 1.76pM, which was about 2 orders lower than that of the LFNAB without HCR amplification. And the detection limit of Salmonella was 3×10(3)cfumL(-1). In conclusion, this visual detection system, HCR-LFNAB, is suitable for non-specialist personnel and point-of-care (POC) diagnosis in low-resource settings.

  9. A non-radioactive in situ hybridization method based on mercurated nucleic acid probes and sulfhydryl-hapten ligands.

    PubMed Central

    Hopman, A H; Wiegant, J; Tesser, G I; Van Duijn, P

    1986-01-01

    Mercurated nucleic acid probes can be used for non-radioactive in situ hybridization. The principle of the method is based on the reaction of the mercurated pyrimidine residues of the in situ hybridized probe with the sulfhydryl group of a ligand which contains a hapten. Next, the hapten is immunocytochemically detected. Previous experiments showed that stable coupling of the sulfhydryl ligands could only be obtained when positively charged amino groups are present in the ligand. On basis of this finding, ligands were synthesized containing a sulfhydryl group, two lysyl residues and hapten groups such as trinitrophenyl, fluorescyl and biotinyl. The ligands, free or bound to mercurated nucleic acids, were immunochemically characterized in ELISAs. The method was shown to be specific and sensitive in the detection of target DNA in situ on microscopic preparations and in dot-blot hybridization reactions on nitrocellulose. Images PMID:3748817

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

    PubMed

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

    2015-02-01

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

  11. Novel conducting polymer-heteropoly acid hybrid material for artificial photosynthetic membranes.

    PubMed

    McDonald, Michael B; Freund, Michael S

    2011-04-01

    Artificial photosynthetic (AP) approaches to convert and store solar energy will require membranes capable of conducting both ions and electrons while remaining relatively transparent and chemically stable. A new approach is applied herein involving previously described in situ chemical polymerization of electronically conducting poly(3,4-ethylenedioxythiophene) (PEDOT) in the presence of proton conducting heteropoly acid (HPA) phosphomolybdic acid (PMA). The electrochemical behaviour of the PEDOT/PMA hybrid material was investigated and it was found that the conducting polymer (CP) is susceptible to irreversible oxidative processes at potentials where water is oxidized. This will be problematic in AP devices should the process occur in very close proximity to a conducting polymer-based membrane. It was found that PEDOT grants the system good electrical performance in terms of conductivity and stability over a large pH window; however, the presence of PMA was not found to provide sufficient proton conductivity. This was addressed in an additional study by tuning the ionic (and in turn, electronic) conductivity in creating composites with the proton-permselective polymer Nafion. It was found that a material of this nature with near-equal conductivity for optimal chemical conversion efficiency will consist of roughly three parts Nafion and one part PEDOT/PMA.

  12. Synthesis and characterization of hybrid composite aerogels from alginic acid and graphene oxide

    NASA Astrophysics Data System (ADS)

    Co, C. J. U.; Quitain, A. T.; Borja, J. Q.; Dugos, N. P.; Takafuji, M.; Kida, T.

    2017-06-01

    Aerogels are one class of solid adsorbents that are gaining considerable attention because of their very high porosity, high specific surface area, and extremely low density. However, most aerogels being studied and used recently are synthetic in nature, which are usually mesoporous silica and metal-organic frameworks (MOFs). As research focus is geared towards sustainable engineering, it is desired to utilize biomass to synthesize aerogels. This study thus aims to produce alginic acid-graphene oxide hybrid composite aerogels and compare them with its existing synthetic counterparts. Alginic acid (AA) is an abundant marine biopolymer that easily forms gels, while graphene oxide (GO) is a nanomaterial consisting of many functional groups. Aerogels made up of AA and GO were successfully synthesized using a sol-gel method. The hydrogel was converted into an aerogel by drying with supercritical carbon dioxide. The percentage of graphene oxide was varied from 0 to 20%. The aerogels were characterized by scanning electron microscopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA) and nitrogen adsorption-desorption measurements. The addition of GO increased the specific surface area of the aerogel up to a certain point, after which it decreased. The 10% GO-AA aerogel showed the most favourable porosity characteristics with a specific surface area of 177.26 m2/g and average pore diameter of 53.2 nm. There had been no observable difference in the thermal behaviour of the aerogels with a change in the concentration of graphene oxide.

  13. Murine cytomegalovirus: detection of latent infection by nucleic acid hybridization technique.

    PubMed Central

    Cheung, K S; Huang, E S; Lang, D J

    1980-01-01

    The technique of nucleic acid hybridization was used to detect the presence of murine cytomegalovirus (MCMV)-specific deoxyribonucleic acid (DNA) in cell cultures and salivary gland tissues. The presence of approximately 4.5 and 0.2 genome equivalents per cell of MCMV-specific DNA was identified in cultures of salivary (ISG2) and prostate gland (IP) cells, respectively. These cells, derived from animals with experimentally induced latent infections, were negative for virus-specific antigens by immunofluorescence and on electron microscopy revealed no visible evidence of the presence of herpesviruses. A cell line derived from the salivary gland of an uninoculated animal (NSG2) was also found to possess MCMV-specific DNA (0.2 genome equivalents per cell). For this reason, salivary gland tissues from uninoculated animals supplied as "specific pathogen-free" mice by three commercial sources were tested upon arrival for the presence of MCMC-specific DNA. MCMV-specific DNA was detectable in pooled salivary gland extracts from uninoculated animals derived from two commercial sources. All of these animals were seronegative and virus negative by conventional infectivity assays. PMID:6247281

  14. Development of a lead-acid battery for a hybrid electric vehicle

    NASA Astrophysics Data System (ADS)

    Cooper, A.

    In September 2000, a project reliable, highly optimized lead-acid battery (RHOLAB) started under the UK Foresight Vehicle Programme with the objective of developing an optimized lead-acid battery solution for hybrid electric vehicles. The work is based on a novel, individual, spirally-wound valve-regulated lead-acid 2 V cell optimized for HEV use and low variability. This cell is being used as a building block for the development of a complete battery pack that is managed at the cell level. Following bench testing, this battery pack is to be thoroughly evaluated by substituting it for the Ni-MH pack in a Honda Insight. The RHOLAB cell is based on the 8 Ah Hawker Cyclon cell which has been modified to have current take-off at both ends—the dual-tab design. In addition, a variant has been produced with modified cell chemistry to help deal with problems that can occur when these valve-regulated lead-acid battery (VRLA) cells operate in a partial-state-of-charge condition. The cells have been cycled to a specially formulated test cycle based on real vehicle data derived from testing the Honda Insight on the various test tracks at the Millbrook Proving Grounds in the UK. These cycling tests have shown that the lead-acid pack can be successfully cycled when subjected to the high current demands from the vehicle, which have been measured at up to 15 C on discharge and 8 C during regenerative recharging, and cycle life is looking very promising under this arduous test regime. Concurrent with this work, battery development has been taking place. It was decided early on to develop the 144 V battery as four 36 V modules. Data collection and control has been built-in and special steps taken to minimize the problems of interconnect in this complex system. Development of the battery modules is now at an advanced stage. The project plan then allows for extensive testing of the vehicle with its lead-acid battery at Millbrook so it can be compared with the benchmark tests which

  15. Exploring the Hybridization Thermodynamics of Spherical Nucleic Acids to Tailor Probes for Diagnostic and Therapeutic Applications

    NASA Astrophysics Data System (ADS)

    Randeria, Pratik Shailesh

    Spherical nucleic acids (SNAs), three-dimensional nanoparticle conjugates composed of densely packed and highly oriented oligonucleotides around organic or inorganic nanoparticles, are an emergent class of nanostructures that show promise as single-entity agents for intracellular messenger RNA (mRNA) detection and gene regulation. SNAs exhibit superior biocompatibility and biological properties compared to linear oligonucleotides, enabling them to overcome many of the limitations of linear oligonucleotides for use in biomedical applications. However, the origins of these biologically attractive properties are not well understood. In this dissertation, the chemistry underlying one such property is studied in detail, and the findings are applied towards the rational design of more effective SNAs for diagnostic and therapeutic applications. Chapter 1 introduces the synthesis of SNAs, the unique properties that make them superior to linear nucleic acids for biomedicine, and previously studied applications of these structures. Chapter 2 focuses on quantitatively studying the impact of the chemical structure of the SNA on its ability to hybridize multiple complementary nucleic acids. This chapter lays the groundwork for understanding the factors that govern SNA hybridization thermodynamics and how to tailor SNAs to increase their binding affinity to target mRNA strands. Chapters 3 and 4 capitalize on this knowledge to engineer probes for intracellular mRNA detection and gene regulation applications. Chapter 3 reports the development of an SNA-based probe that can simultaneously report the expression level of two different mRNA transcripts in live cells and differentiate diseased cells from non-diseased cells. Chapter 4 investigates the use of topically-applied SNAs to down-regulate a critical mediator of impaired wound healing in diabetic mice to accelerate wound closure. This study represents the first topical therapeutic application of SNA nanotechnology to treat open

  16. THE EFFECT OF ANOLYTE PRODUCT ACID CONCENTRATION ON HYBRID SULFUR CYCLE PERFORMANCE

    SciTech Connect

    Gorensek, M.; Summers, W.

    2010-03-24

    The Hybrid Sulfur (HyS) cycle (Fig. 1) is one of the simplest, all-fluids thermochemical cycles that has been devised for splitting water with a high-temperature nuclear or solar heat source. It was originally patented by Brecher and Wu in 1975 and extensively developed by Westinghouse in the late 1970s and early 1980s. As its name suggests, the only element used besides hydrogen and oxygen is sulfur, which is cycled between the +4 and +6 oxidation states. HyS comprises two steps. One is the thermochemical (>800 C) decomposition of sulfuric acid (H{sub 2}SO{sub 4}) to sulfur dioxide (SO{sub 2}), oxygen (O{sub 2}), and water. H{sub 2}SO{sub 4} = SO{sub 2} + 1/2 O{sub 2} + H{sub 2}O. The other is the SO{sub 2}-depolarized electrolysis of water to H{sub 2}SO{sub 4} and hydrogen (H{sub 2}), SO{sub 2} + 2 H{sub 2}O = H{sub 2}SO{sub 4} + H{sub 2}, E{sup o} = -0.156 V, explaining the 'hybrid' designation. These two steps taken together split water into H{sub 2} and O{sub 2} using heat and electricity. Researchers at the Savannah River National Laboratory (SRNL) and at the University of South Carolina (USC) have successfully demonstrated the use of proton exchange membrane (PEM) electrolyzers (Fig. 2) for the SO{sub 2}-depolarized electrolysis (sulfur oxidation) step, while Sandia National Laboratories (SNL) successfully demonstrated the high-temperature sulfuric acid decomposition (sulfur reduction) step using a bayonet-type reactor (Fig. 3). This latter work was performed as part of the Sulfur-Iodine (SI) cycle Integrated Laboratory Scale demonstration at General Atomics (GA). The combination of these two operations results in a simple process that will be more efficient and cost-effective for the massive production of hydrogen than alkaline electrolysis. Recent developments suggest that the use of PEMs other than Nafion will allow sulfuric acid to be produced at higher concentrations (>60 wt%), offering the possibility of net thermal efficiencies around 50% (HHV basis

  17. Synthesis and utilization of chitin humic acid hybrid as sorbent for Cr(III)

    NASA Astrophysics Data System (ADS)

    Santosa, Sri Juari; Siswanta, Dwi; Sudiono, Sri; Sehol, Muhamad

    2007-11-01

    New types of hybrid material have been synthesized by using four different methods of immobilization of humic acid (HA) on chitin. The most stable hybrid material toward the change of medium acidity was then utilized as sorbent for Cr(III). The HA was extracted from peat soil of Gambut District, South Kalimantan, Indonesia, using the recommended procedure of International Humic Substances Society (IHSS), while the chitin was isolated from crab shell waste through deproteination using 3.5% (w/v) NaOH and followed by removal of inorganic impurities using 1 M HCl. The four methods of immobilization of HA on chitin were (i) Method A: chitin powder (4 g) was gently poured into the stirred solution of 0.4 g HA in 40 mL of 0.01 M NaOH. After overnight stirring, the solid was separated, washed with water, and dried in oven at 70 °C. (ii) Method B: gelatinous chitin (40 g) in 250 mL of 0.5 M HCl was reacted with HA (4 g) in 500 mL of 0.5 M NaOH and aged for 24 h. The product was washed with water and dried. (iii) Method C: HA powder (0.5 g) was mixed with the stirred gel of chitin (2.5 g) in 60 mL of CaCl 2 saturated methanol and the mixture was then washed with the mixed solution of 25 mL of 2 M sodium citrate and ethylene glycol 1:1. The solid was separated, washed with water, and dried. (iv) Method D: the solution of HA (0.056 g) in 10 mL of 0.01 M NaOH was reacted with the gel of chitin (0.2 g) in 10 mL of CaCl 2 saturated methanol. After 24 h stirring, the solid was separated from the reaction medium, washed with the mixed solution of 2 M sodium citrate and ethylene glycol 1:1, and followed by washing with water and drying. Parameters investigated in this study consisted of the stability test of the immobilized HA, as well as the rate constant ( k1), capacity ( b), and energy ( E) of sorption as well as the rate constant of desorption ( k-1). The k1 and k-1 were determined according to a kinetic model of first order sorption reaching equilibrium, while the b and E

  18. Universal Dynamic DNA Assembly-Programmed Surface Hybridization Effect for Single-Step, Reusable, and Amplified Electrochemical Nucleic Acid Biosensing.

    PubMed

    Liu, Shufeng; Fang, Li; Wang, Yanqun; Wang, Li

    2017-03-07

    The traditional sensitive electrochemical biosensors are commonly confronted with the cumbersome interface operation and washing procedures and the inclusion of extra exogenous reagents, which impose the challenge on the detection simplicity, reliability, and reusability. Herein, we present the proof-of-principle of a unique biosensor architecture based on dynamic DNA assembly programmed surface hybridization, which confers the single-step, reusable, and enzyme-free amplified electrochemical nucleic acid analysis. To demonstrate the fabrication universality three dynamic DNA assembly strategies including DNA-fueled target recycling, catalytic hairpin DNA assembly, and hybridization chain reaction were flexibly harnessed to convey the homogeneous target recognition and amplification events into various DNA scaffolds for the autonomous proximity-based surface hybridization. The current biosensor architecture features generalizability, simplicity, low cost, high sensitivity, and specificity over the traditional nucleic acid-related amplified biosensors. The lowest detection limit of 50 aM toward target DNA could be achieved by hybridization chain reaction-programmed surface hybridization. The reliable working ability for both homogeneous solution and heterogeneous inteface facilitates the target analysis with a robust reliability and reproducibility, also making it to be readily extended for the integration with the kinds of detecting platforms. Thus, it may hold great potential for the biosensor fabrication served for the point-of-care applications in resource constrained regions.

  19. Design of hybrid multimodal poly(lactic-co-glycolic acid) polymer nanoparticles for neutrophil labeling, imaging and tracking.

    PubMed

    Qiu, Yuan; Palankar, Raghavendra; Echeverría, María; Medvedev, Nikolay; Moya, Sergio E; Delcea, Mihaela

    2013-12-21

    Poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) incorporating quantum dots (QDs), superparamagnetic iron oxide nanoparticles (SPIONs) and gold (Au) NPs for neutrophil labeling were fabricated via the w/o/w double emulsion method. QDs and SPIONs were entrapped in the PLGA core during emulsification while Au NPs were assembled on top of the PLGA NPs via electrostatic interactions. Transmission Electron Microscopy, Scanning Electron Microscopy and Confocal Scanning Laser Microscopy (CLSM) were applied to characterize the hybrid PLGA NPs. The uptake of the hybrid PLGA NPs by human neutrophils was studied by Flow Cytometry and confocal microscopy. In addition, the induction of reactive oxygen species (ROS) in neutrophils after incubation with the hybrid PLGA NPs was assessed. Magnetophoresis experiments showed that neutrophils with internalized hybrid PLGA NPs can be effectively laterally displaced towards the magnetic field. Magnetic Resonance Imaging of the hybrid PLGA NPs resulted in images with a contrast enhancement linearly dependent on the concentration of the hybrid PLGA NPs. Research reported in this work is relevant for imaging, tracking and manipulating neutrophils and has potential for in vivo applications, e.g., tumor visualization and localized photothermal treatment.

  20. Interfacial transduction of nucleic acid hybridization using immobilized quantum dots as donors in fluorescence resonance energy transfer.

    PubMed

    Algar, W Russ; Krull, Ulrich J

    2009-01-06

    Fluorescence resonance energy transfer (FRET) using immobilized quantum dots (QDs) as energy donors was explored as a transduction method for the detection of nucleic acid hybridization at an interface. This research was motivated by the success of the QD-FRET-based transduction of nucleic acid hybridization in solution-phase assays. This new work represents a fundamental step toward the assembly of a biosensor, where immobilization of the selective chemistry on a surface is desired. After immobilizing QD-probe oligonucleotide conjugates on optical fibers, a demonstration of the retention of selectivity was achieved by the introduction of acceptor (Cy3)-labeled single-stranded target oligonucleotides. Hybridization generated the proximity required for FRET, and the resulting fluorescence spectra provided an analytical signal proportional to the amount of target. This research provides an important framework for the future development of nucleic acid biosensors based on QDs and FRET. The most important findings of this work are that (1) a QD-FRET solid-phase hybridization assay is viable and (2) a passivating layer of denatured bovine serum albumin alleviates nonspecific adsorption, ultimately resulting in (3) the potential for a reusable assay format and mismatch discrimination. In this, the first incarnation of a solid-phase QD-FRET hybridization assay, the limit of detection was found to be 5 nM, and the dynamic range was almost 2 orders of magnitude. Selective discrimination of the target was shown using a three-base-pairs mismatch from a fully complementary sequence. Despite a gradual loss of signal, reuse of the optical fibers over multiple cycles of hybridization and dehybridization was possible. Directions for further improvement of the analytical performance by optimizing the design of the QD-probe oligonucleotide interface are identified.

  1. Intrinsically Labeled Fluorescent Oligonucleotide Probes on Quantum Dots for Transduction of Nucleic Acid Hybridization.

    PubMed

    Shahmuradyan, Anna; Krull, Ulrich J

    2016-03-15

    Quantum dots (QDs) have been widely used in chemical and biosensing due to their unique photoelectrical properties and are well suited as donors in fluorescence resonance energy transfer (FRET). Selective hybridization interactions of oligonucleotides on QDs have been determined by FRET. Typically, the QD-FRET constructs have made use of labeled targets or have implemented labeled sandwich format assays to introduce dyes in proximity to the QDs for the FRET process. The intention of this new work is to explore a method to incorporate the acceptor dye into the probe molecule. Thiazole orange (TO) derivatives are fluorescent intercalating dyes that have been used for detection of double-stranded nucleic acids. One such dye system has been reported in which single-stranded oligonucleotide probes were doubly labeled with adjacent thiazole orange derivatives. In the absence of the fully complementary (FC) oligonucleotide target, the dyes form an H-aggregate, which results in quenching of fluorescence emission due to excitonic interactions between the dyes. The hybridization of the FC target to the probe provides for dissociation of the aggregate as the dyes intercalate into the double stranded duplex, resulting in increased fluorescence. This work reports investigation of the dependence of the ratiometric signal on the type of linkage used to conjugate the dyes to the probe, the location of the dye along the length of the probe, and the distance between adjacent dye molecules. The limit of detection for 34mer and 90mer targets was found to be identical and was 10 nM (2 pmol), similar to analogous QD-FRET using labeled oligonucleotide target. The detection system could discriminate a one base pair mismatch (1BPM) target and was functional without substantial compromise of the signal in 75% serum. The 1BPM was found to reduce background signal, indicating that the structure of the mismatch affected the environment of the intercalating dyes.

  2. Cationic liposome-hyaluronic acid hybrid nanoparticles for intranasal vaccination with subunit antigens

    PubMed Central

    Ochyl, Lukasz J.; Akerberg, Jonathan; Moon, James J.

    2015-01-01

    Here we report the development of a new cationic liposome-hyaluronic acid (HA) hybrid nanoparticle (NP) system and present our characterization of these NPs as an intranasal vaccine platform using a model antigen and F1-V, a candidate recombinant antigen for Yersinia pestis, the causative agent of plague. Incubation of cationic liposomes composed of DOTAP and DOPE with anionic HA biopolymer led to efficient ionic complexation and formation of homogenous liposome-polymer hybrid NPs, as evidenced by fluorescence resonance energy transfer, dynamic light scattering, and nanoparticle tracking analyses. Incorporation of cationic liposomes with thiolated HA allowed for facile surface decoration of NPs with thiol-PEG, resulting in the formation of DOTAP/HA core-PEG shell nanostructures. These NPs, termed DOTAP-HA NPs, exhibited improved colloidal stability and prolonged antigen release. In addition, cytotoxicity associated with DOTAP liposomes (LC50 ~0.2 mg/ml) was significantly reduced by at least 20-fold with DOTAP-HA NPs (LC50 > 4 mg/ml), as measured with bone marrow dendritic cells (BMDCs). Furthermore, NPs co-loaded with ovalbumin (OVA) and a molecular adjuvant, monophosphoryl lipid A (MPLA) promoted BMDC maturation and upregulation of co-stimulatory markers, including CD40, CD86, and MHC-II, and C57BL/6 mice vaccinated with NPs via intranasal route generated robust OVA-specific CD8+ T cell and antibody responses. Importantly, intranasal vaccination with NPs co-loaded with F1-V and MPLA induced potent humoral immune responses with 11-, 23-, and 15-fold increases in F1-V-specific total IgG, IgG1, and IgG2c titers in immune sera by day 77, respectively, and induced balanced Th1/Th2 humoral immune responses, compared with the lack of sero-conversion in mice immunized with the equivalent doses of soluble F1-V vaccine. Overall, these results suggest that liposome-polymer hybrid NPs may serve as a promising vaccine delivery platform for intranasal vaccination against Y

  3. Cationic liposome-hyaluronic acid hybrid nanoparticles for intranasal vaccination with subunit antigens.

    PubMed

    Fan, Yuchen; Sahdev, Preety; Ochyl, Lukasz J; J Akerberg, Jonathan; Moon, James J

    2015-06-28

    Here we report the development of a new cationic liposome-hyaluronic acid (HA) hybrid nanoparticle (NP) system and present our characterization of these NPs as an intranasal vaccine platform using a model antigen and F1-V, a candidate recombinant antigen for Yersinia pestis, the causative agent of plague. Incubation of cationic liposomes composed of DOTAP and DOPE with anionic HA biopolymer led to efficient ionic complexation and formation of homogenous liposome-polymer hybrid NPs, as evidenced by fluorescence resonance energy transfer, dynamic light scattering, and nanoparticle tracking analyses. Incorporation of cationic liposomes with thiolated HA allowed for facile surface decoration of NPs with thiol-PEG, resulting in the formation of DOTAP/HA core-PEG shell nanostructures. These NPs, termed DOTAP-HA NPs, exhibited improved colloidal stability and prolonged antigen release. In addition, cytotoxicity associated with DOTAP liposomes (LC50~0.2mg/ml) was significantly reduced by at least 20-fold with DOTAP-HA NPs (LC50>4mg/ml), as measured with bone marrow derived dendritic cells (BMDCs). Furthermore, NPs co-loaded with ovalbumin (OVA) and a molecular adjuvant, monophosphoryl lipid A (MPLA) promoted BMDC maturation and upregulation of co-stimulatory markers, including CD40, CD86, and MHC-II, and C57BL/6 mice vaccinated with NPs via intranasal route generated robust OVA-specific CD8(+) T cell and antibody responses. Importantly, intranasal vaccination with NPs co-loaded with F1-V and MPLA induced potent humoral immune responses with 11-, 23-, and 15-fold increases in F1-V-specific total IgG, IgG1, and IgG2c titers in immune sera by day 77, respectively, and induced balanced Th1/Th2 humoral immune responses, whereas mice immunized with the equivalent doses of soluble F1-V vaccine failed to achieve sero-conversion. Overall, these results suggest that liposome-polymer hybrid NPs may serve as a promising vaccine delivery platform for intranasal vaccination against Y

  4. Characterization of a Chinese hamster-human hybrid cell line with increased system L amino acid transport activity.

    PubMed Central

    Lobaton, C D; Moreno, A; Oxender, D L

    1984-01-01

    We have studied leucine transport in several Chinese hamster-human hybrid cell lines obtained by fusion of a temperature-sensitive line of Chinese hamster ovary cells, ts025C1, and normal human leukocytes. A hybrid cell line exhibiting a twofold increase in L-leucine uptake over that in the parental cell line was found. This hybrid cell line, 158CnpT-1, was temperature resistant, whereas the parental Chinese hamster ovary mutant, ts025C1, contained a temperature-sensitive leucyl-tRNA synthetase mutation. An examination of the different amino acid transport systems in this hybrid cell line revealed a specific increase of system L activity with no significant changes in systems A and ASC. The Vmax for L-leucine uptake exhibited by the hybrid 158CnpT-1 was twice that in the CHO parental mutant, ts025C1. Cytogenetic analysis showed that the hybrid 158CnpT-1 contains four complete human chromosomes (numbers 4, 5, 10, and 21) and three interspecific chromosomal translocations in a total complement of 34 chromosomes. Biochemical and cytogenetic analysis of segregant clones obtained from hybrid 158CnpT-1 showed that the primary temperature resistance and high system L transport phenotypes can be segregated from this hybrid independently. The loss of the primary temperature resistance was associated with the loss of the human chromosome 5, as previously reported by other laboratories, whereas the loss of the high leucine transport phenotype, which is associated with a lesser degree of temperature resistance, was correlated with the loss of human chromosome 20. Images PMID:6717430

  5. Characterization of a Chinese hamster-human hybrid cell line with increased system L amino acid transport activity.

    PubMed

    Lobaton, C D; Moreno, A; Oxender, D L

    1984-03-01

    We have studied leucine transport in several Chinese hamster-human hybrid cell lines obtained by fusion of a temperature-sensitive line of Chinese hamster ovary cells, ts025C1, and normal human leukocytes. A hybrid cell line exhibiting a twofold increase in L-leucine uptake over that in the parental cell line was found. This hybrid cell line, 158CnpT-1, was temperature resistant, whereas the parental Chinese hamster ovary mutant, ts025C1, contained a temperature-sensitive leucyl-tRNA synthetase mutation. An examination of the different amino acid transport systems in this hybrid cell line revealed a specific increase of system L activity with no significant changes in systems A and ASC. The Vmax for L-leucine uptake exhibited by the hybrid 158CnpT-1 was twice that in the CHO parental mutant, ts025C1. Cytogenetic analysis showed that the hybrid 158CnpT-1 contains four complete human chromosomes (numbers 4, 5, 10, and 21) and three interspecific chromosomal translocations in a total complement of 34 chromosomes. Biochemical and cytogenetic analysis of segregant clones obtained from hybrid 158CnpT-1 showed that the primary temperature resistance and high system L transport phenotypes can be segregated from this hybrid independently. The loss of the primary temperature resistance was associated with the loss of the human chromosome 5, as previously reported by other laboratories, whereas the loss of the high leucine transport phenotype, which is associated with a lesser degree of temperature resistance, was correlated with the loss of human chromosome 20.

  6. Silylated melamine and cyanuric acid as precursors for imprinted and hybrid silica materials with molecular recognition properties.

    PubMed

    Arrachart, Guilhem; Carcel, Carole; Trens, Philippe; Moreau, Jöel J E; Wong Chi Man, Michel

    2009-06-15

    Two monotrialkoxysilylated compounds that consist of complementary fragments of melamine (M) and cyanuric acid (CA) have been synthesised. The molecular recognition properties of the M and CA fragments through complementary hydrogen bonds (DAD and ADA; D=donor, A=acceptor) are the key factor used to direct the formation of hybrid silica materials by using a sol-gel process. These materials were synthesised following two methods: First, an organo-bridged silsesquioxane was obtained by the hydrolysis of the two complementary monotrialkoxysilylated melamine and cyanuric acid derivatives, with fluoride ions as a catalyst. The hydrogen-bonding interactions between the two organic fragments are responsible for the formation of the bridging unit. The transcription of the assembly into the hybrid material was characterised and evidenced by solid-state NMR (29Si, 13C) and FTIR spectroscopic experiments. Second, the molecular recognition was exploited to synthesise an imprinted hybrid silica. This material was prepared by co-condensation of tetraethyl orthosilicate (TEOS) with the monosilylated cyanuric acid derivative (CA) templated by nonsilylated melamine. The melamine template was completely removed by treating the solid material with hydrochloric acid. The reintroduction of the template was performed by treating the resulting material with an aqueous suspension of melamine. These steps were monitored and analysed by several techniques, such as solid-state NMR (29Si, 13C) and FTIR spectroscopic analysis and nitrogen adsorption-desorption isotherms.

  7. Hybrid films of chitosan, cellulose nanofibrils and boric acid: Flame retardancy, optical and thermo-mechanical properties.

    PubMed

    Uddin, Khan M A; Ago, Mariko; Rojas, Orlando J

    2017-12-01

    Chitosan (CS), cellulose nanofibrils (CNF) and boric acid, the latter of which was used as flame retardant, were combined in transparent, hybrid films that were produced by solvent casting. The flammability and the thermal stability of the films were studied with respect to the loading of the inorganic component. Chitosan films displayed fire retardancy properties, which were enhanced in the presence of boric acid. CNF films, in contrast to those from chitosan, were readily flammable; however, when combined with boric acid (30w%), they became self-extinguishing. Most remarkably, bicomponent films comprising CNF and chitosan, displayed better fire retardancy than that of neat CS films. Moreover, boric acid improved the thermal stability of the bicomponent films. The tensile strength and Young's modulus of CS, CNF and CS-CNF films improved at intermediate boric acid addition, although a negative effect on elongation was observed. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Increased stability and specificity through combined hybridization of peptide nucleic acid (PNA) and locked nucleic acid (LNA) to supercoiled plasmids for PNA-anchored "Bioplex" formation.

    PubMed

    Lundin, Karin E; Hasan, Maroof; Moreno, Pedro M; Törnquist, Elisabeth; Oprea, Iulian; Svahn, Mathias G; Simonson, E Oscar; Smith, C I Edvard

    2005-12-01

    Low cellular uptake and poor nuclear transfer hamper the use of non-viral vectors in gene therapy. Addition of functional entities to plasmids using the Bioplex technology has the potential to improve the efficiency of transfer considerably. We have investigated the possibility of stabilizing sequence-specific binding of peptide nucleic acid (PNA) anchored functional peptides to plasmid DNA by hybridizing PNA and locked nucleic acid (LNA) oligomers as "openers" to partially overlapping sites on the opposite DNA strand. The PNA "opener" stabilized the binding of "linear" PNA anchors to mixed-base supercoiled DNA in saline. For higher stability under physiological conditions, bisPNA anchors were used. To reduce nonspecific interactions when hybridizing highly cationic constructs and to accommodate the need for increased amounts of bisPNA when the molecules are uncharged, or negatively charged, we used both PNA and LNA oligomers as "openers" to increase binding kinetics. To our knowledge, this is the first time that LNA has been used together with PNA to facilitate strand invasion. This procedure allows hybridization at reduced PNA-to-plasmid ratios, allowing greater than 80% hybridization even at ratios as low as 2:1. Using significantly lower amounts of PNA-peptides combined with shorter incubation times reduces unspecific binding and facilitates purification.

  9. Cloning and characterization of the gene for L-amino acid oxidase in hybrid tilapia.

    PubMed

    Shen, Yubang; Fu, Gui Hong; Liu, Feng; Yue, Gen Hua

    2015-12-01

    Tilapia is the common name for a group of cichlid fishes. Identification of DNA markers significantly associated with important traits in candidate genes may speed up genetic improvement. L-Amino acid oxidase (LAO) plays a crucial role in the innate immune defences of animals. Previously, whether LAO variants were associated with economic traits had not been studied in fish. We characterized the cDNA sequence of the LAO gene of hybrid tilapia (Oreochromis spp.). Its ORF was 1536 bp, encoding a flavoenzyme of 511 amino acids. This gene consisted of seven exons and six introns. Its expression was detected in the intestine, blood, kidney, skin, liver. It was highly expressed in the intestine. After a challenge with a bacterial pathogen, Streptococcus agalactiae, its expression was up-regulated significantly in the liver, intestine and spleen (P < 0.05). We identified one SNP in the genomic sequence of the gene and found that this SNP was associated significantly with body length (P < 0.05), but not with resistance to S. agalactiae. The results of this study suggest that the LAO gene plays an important role in innate immune responses to the bacterial pathogen in tilapia. The investigation of relationship between polymorphism of LAO gene and disease resistance and growth in tilapia showed that one SNP was associated significantly with body length. Further experiments on whether SNPs in the LAO gene are associated with growth in tilapia and other populations could be useful in understanding more functions of the LAO gene.

  10. Mapping the regioisomeric distribution of fatty acids in triacylglycerols by hybrid mass spectrometry.

    PubMed

    Nagy, Kornél; Sandoz, Laurence; Destaillats, Frédéric; Schafer, Olivier

    2013-01-01

    This study describes the use of hybrid mass spectrometry for the mapping, identification, and semi-quantitation of triacylglycerol regioisomers in fats and oils. The identification was performed based on the accurate mass and fragmentation pattern obtained by data-dependent fragmentation. Quantitation was based on the high-resolution ion chromatograms, and relative proportion of sn-1(3)/sn-2 regioisomers was calculated based on generalized fragmentation models and the relative intensities observed in the product ion spectra. The key performance features of the developed method are inter-batch mass accuracy < 1 ppm (n = 10); lower limit of detection (triggering threshold) 0.1 μg/ml (equivalent to 0.2 weight % in oil); lower limit of quantitation 0.2 μg/ml (equivalent to 0.4 weight % in oil); peak area precision 6.5% at 2 μg/ml concentration and 15% at 0.2 μM concentration; inter-batch precision of fragment intensities < 1% (n = 10) independent of the investigated concentration; and averaged accuracy using the generic calibration 3.8% in the 1-10 μg/ml range and varies between 1-23% depending on analytes. Inter-esterified fat, beef tallow, pork lard, and butter fat samples were used to show how well regioisomeric distribution of palmitic acid can be captured by this method.

  11. Ternary surface monolayers for ultrasensitive (zeptomole) amperometric detection of nucleic acid hybridization without signal amplification.

    PubMed

    Wu, Jie; Campuzano, Susana; Halford, Colin; Haake, David A; Wang, Joseph

    2010-11-01

    A ternary surface monolayer, consisting of coassembled thiolated capture probe, mercaptohexanol and dithiothreitol, is shown to offer dramatic improvements in the signal-to-noise characteristics of electrochemical DNA hybridization biosensors based on common self-assembled monolayers. Remarkably low detection limits down to 40 zmol (in 4 μL samples) as well as only 1 CFU Escherichia coli per sensor are thus obtained without any additional amplification step in connection to the commonly used horseradish peroxidase/3,3',5,5'-tetramethylbenzidine system. Such dramatic improvements in the detection limits (compared to those of common binary alkanethiol interfaces and to those of most electrochemical DNA sensing strategies without target or signal amplification) are attributed primarily to the remarkably higher resistance to nonspecific adsorption. This reflects the highly compact layer (with lower pinhole density) produced by the coupling of the cyclic- and linear-configuration "backfillers" that leads to a remarkably low background noise even in the presence of complex sample matrixes. A wide range of surface compositions have been investigated, and the ternary mixed monolayer has been systematically optimized. Detailed impedance spectroscopy and cyclic voltammetric studies shed useful insights into the surface coverage. The impressive sensitivity and high specificity of the simple developed methodology indicate great promise for a wide range of nucleic acid testing, including clinical diagnostics, biothreat detection, food safety, and forensic analysis.

  12. Yeasts identification in microfluidic devices using peptide nucleic acid fluorescence in situ hybridization (PNA-FISH).

    PubMed

    Ferreira, André M; Cruz-Moreira, Daniela; Cerqueira, Laura; Miranda, João M; Azevedo, Nuno F

    2017-03-01

    Peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) is a highly specific molecular method widely used for microbial identification. Nonetheless, and due to the detection limit of this technique, a time-consuming pre-enrichment step is typically required before identification. In here we have developed a lab-on-a-chip device to concentrate cell suspensions and speed up the identification process in yeasts. The PNA-FISH protocol was optimized to target Saccharomyces cerevisiae, a common yeast that is very relevant for several types of food industries. Then, several coin-sized microfluidic devices with different geometries were developed. Using Computational fluid dynamics (CFD), we modeled the hydrodynamics inside the microchannels and selected the most promising options. SU-8 structures were fabricated based on the selected designs and used to produce polydimethylsiloxane-based microchips by soft lithography. As a result, an integrated approach combining microfluidics and PNA-FISH for the rapid identification of S. cerevisiae was achieved. To improve fluid flow inside microchannels and the PNA-FISH labeling, oxygen plasma treatment was applied to the microfluidic devices and a new methodology to introduce the cell suspension and solutions into the microchannels was devised. A strong PNA-FISH signal was observed in cells trapped inside the microchannels, proving that the proposed methodology works as intended. The microfluidic designs and PNA-FISH procedure described in here should be easily adaptable for detection of other microorganisms of similar size.

  13. Synthesis, in vitro and in vivo antitumor activity of scopoletin-cinnamic acid hybrids.

    PubMed

    Li, Linhu; Zhao, Peng; Hu, Jinglin; Liu, Jinhong; Liu, Yan; Wang, Zhiqiang; Xia, Yufeng; Dai, Yue; Chen, Li

    2015-03-26

    A series of hybrids of scopoletin and substituted cinnamic acid were designed, synthesized and evaluated in vitro and in vivo against five human tumor cell lines [MCF-7, MDA-MB-231, A549, HCT-116, and HeLa] with doxorubicin as the positive control. Compounds 17a, 17b, 17c and 17g exhibited potent cytotoxic activity. Especially, compound 17b displayed broad spectrum activity with IC50 values ranging from 0.249 μM to 0.684 μM. Moreover, in a preliminary pharmacological study, 17b not only remarkably induced cellular apoptosis, but also clearly induced A549 cells cycle arrest at S phase. In vivo study showed that 17b significantly suppressed tumor growth in a dose-dependent manner without causing the loss of the mean body weight of mice, which was superior to doxorubicin. These preliminary results indicate that 17b is an optimal anti-cancer leading compound and merit further structural modification. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  14. Diagnosis of bacterial vaginosis by a new multiplex peptide nucleic acid fluorescence in situ hybridization method

    PubMed Central

    Machado, António; Castro, Joana; Cereija, Tatiana; Almeida, Carina

    2015-01-01

    Bacterial vaginosis (BV) is one of most common vaginal infections. However, its diagnosis by classical methods reveals low specificity. Our goal was to evaluate the accuracy diagnosis of 150 vaginal samples with research gold standard methods and our Peptide Nucleic Acid (PNA) probes by Fluorescence in situ Hybridization (FISH) methodology. Also, we described the first PNA-FISH methodology for BV diagnosis, which provides results in approximately 3 h. The results showed a sensitivity of 84.6% (95% confidence interval (CI), from 64.3 to 95.0%) and a specificity of 97.6% (95% CI [92.6–99.4%]), demonstrating the higher specificity of the PNA-FISH method and showing false positive results in BV diagnosis commonly obtained by the classical methods. This methodology combines the specificity of PNA probes for Lactobacillus species and G. vaginalis visualization and the calculation of the microscopic field by Nugent score, allowing a trustful evaluation of the bacteria present in vaginal microflora and avoiding the occurrence of misleading diagnostics. Therefore, the PNA-FISH methodology represents a valuable alternative for BV diagnosis. PMID:25737820

  15. Biomedical nanocomposites of poly(lactic acid) and calcium phosphate hybridized with modified carbon nanotubes for hard tissue implants.

    PubMed

    Lee, Hae-Hyoung; Sang Shin, Ueon; Lee, Jae-Ho; Kim, Hae-Won

    2011-08-01

    Degradable polymer-based materials are attractive in orthopedics and dentistry as an alternative to metallic implants for use as bone fixatives. Herein, a degradable polymer poly(lactic acid) (PLA) was combined with novel hybrid nanopowder of carbon nanotubes (CNTs)-calcium phosphate (CP) for this application. In particular, CNTs-CP hybrid nanopowders (0.1 and 0.25% CNTs) were prepared from the solution of ionically modified CNTs (mCNTs), which was specifically synthesized to be well-dispersed and thus to effectively adsorb onto the CP nanoparticles. The mCNTs-CP hybrid nanopowders were then mixed with PLA (up to 50%) to produce mCNTs-CP-PLA nanocomposites. The mechanical tensile strength of the nanocomposites was significantly improved by the addition of mCNTs-CP hybrid nanopowders. Moreover, nanocomposites containing low concentration of mCNTs (0.1%) showed significantly stimulated biological responses including cell proliferation and osteoblastic differentiation in terms of gene and protein expressions. Based on this study, the addition of novel mCNT-CP hybrid nanopowders to PLA biopolymer may be considered a new material choice for developing hard tissue implants.

  16. Visualization of the mycelia of wood-rotting fungi by fluorescence in situ hybridization using a peptide nucleic acid probe.

    PubMed

    Nakada, Yuji; Nakaba, Satoshi; Matsunaga, Hiroshi; Funada, Ryo; Yoshida, Makoto

    2013-01-01

    White rot fungus, Phanerochaete chrysosporium, and brown rot fungus, Postia placenta, grown on agar plates, were visualized by fluorescence in situ hybridization (FISH) using a peptide nucleic acid (PNA) probe. Mycelia grown on wood chips were also clearly detected by PNA-FISH following blocking treatment. To the best of our knowledge, this is the first report on the visualization of fungi in wood by FISH.

  17. Degradation of Acid Orange 7 Dye in Two Hybrid Plasma Discharge Reactors

    NASA Astrophysics Data System (ADS)

    Shen, Yongjun; Lei, Lecheng; Zhang, Xingwang; Ding, Jiandong

    2014-11-01

    To get an optimized pulsed electrical plasma discharge reactor and to increase the energy utilization efficiency in the removal of pollutants, two hybrid plasma discharge reactors were designed and optimized. The reactors were compared via the discharge characteristics, energy transfer efficiency, the yields of the active species and the energy utilization in dye wastewater degradation. The results showed that under the same AC input power, the characteristics of the discharge waveform of the point-to-plate reactor were better. Under the same AC input power, the two reactors both had almost the same peak voltage of 22 kV. The peak current of the point-to-plate reactor was 146 A, while that of the wire-to-cylinder reactor was only 48.8 A. The peak powers of the point-to-plate reactor and the wire-to-cylinder reactor were 1.38 MW and 1.01 MW, respectively. The energy per pulse of the point-to-plate reactor was 0.2221 J, which was about 29.4% higher than that of the wire-to-cylinder reactor (0.1716 J). To remove 50% Acid Orange 7 (AO7), the energy utilizations of the point-to-plate reactor and the wire-to-cylinder reactor were 1.02 × 10-9 mol/L and 0.61 × 10-9 mol/L, respectively. In the point-to-plate reactor, the concentration of hydrogen peroxide in pure water was 3.6 mmol/L after 40 min of discharge, which was higher than that of the wire-to-cylinder reactor (2.5 mmol/L). The concentration of liquid phase ozone in the point-to-plate reactor (5.7 × 10-2 mmol/L) was about 26.7% higher than that in the wire-to-cylinder reactor (4.5 × 10-2 mmol/L). The analysis results of the variance showed that the type of reactor and reaction time had significant impacts on the yields of the hydrogen peroxide and ozone. The main degradation intermediates of AO7 identified by gas chromatography and mass spectrometry (GCMS) were acetic acid, maleic anhydride, p-benzoquinone, phenol, benzoic acid, phthalic anhydride, coumarin and 2-naphthol. Proposed degradation pathways were

  18. Fabrication of a palladium nanoparticle/graphene nanosheet hybrid via sacrifice of a copper template and its application in catalytic oxidation of formic acid.

    PubMed

    Zhao, Hong; Yang, Jun; Wang, Lei; Tian, Chungui; Jiang, Baojiang; Fu, Honggang

    2011-02-21

    Small and highly dispersed palladium nanoparticles supported on graphene nanosheets were fabricated via a strategy of etching a copper template with Pd(2+). The obtained hybrid exhibited significant catalytic activity for formic acid oxidation.

  19. Development of a lauric acid/albumin hybrid iron oxide nanoparticle system with improved biocompatibility.

    PubMed

    Zaloga, Jan; Janko, Christina; Nowak, Johannes; Matuszak, Jasmin; Knaup, Sabine; Eberbeck, Dietmar; Tietze, Rainer; Unterweger, Harald; Friedrich, Ralf P; Duerr, Stephan; Heimke-Brinck, Ralph; Baum, Eva; Cicha, Iwona; Dörje, Frank; Odenbach, Stefan; Lyer, Stefan; Lee, Geoffrey; Alexiou, Christoph

    2014-01-01

    The promising potential of superparamagnetic iron oxide nanoparticles (SPIONs) in various nanomedical applications has been frequently reported. However, although many different synthesis methods, coatings, and functionalization techniques have been described, not many core-shell SPION drug delivery systems are available for clinicians at the moment. Here, bovine serum albumin was adsorbed onto lauric acid-stabilized SPIONs. The agglomeration behavior, zeta potential, and their dependence on the synthesis conditions were characterized with dynamic light scattering. The existence and composition of the core-shell-matrix structure was investigated by transmission electron microscopy, Fourier transform infrared spectroscopy, and zeta potential measurements. We showed that the iron oxide cores form agglomerates in the range of 80 nm. Moreover, despite their remarkably low tendency to aggregate even in a complex media like whole blood, the SPIONs still maintained their magnetic properties and were well attractable with a magnet. The magnetic properties were quantified by vibrating sample magnetometry and a superconducting quantum interference device. Using flow cytometry, we further investigated the effects of the different types of nanoparticle coating on morphology, viability, and DNA integrity of Jurkat cells. We showed that by addition of bovine serum albumin, the toxicity of nanoparticles is greatly reduced. We also investigated the effect of the particles on the growth of primary human endothelial cells to further demonstrate the biocompatibility of the particles. As proof of principle, we showed that the hybrid-coated particles are able to carry payloads of up to 800 μg/mL of the cytostatic drug mitoxantrone while still staying colloidally stable. The drug-loaded system exhibited excellent therapeutic potential in vitro, exceeding that of free mitoxantrone. In conclusion, we have synthesized a biocompatible ferrofluid that shows great potential for clinical

  20. Development of a lauric acid/albumin hybrid iron oxide nanoparticle system with improved biocompatibility

    PubMed Central

    Zaloga, Jan; Janko, Christina; Nowak, Johannes; Matuszak, Jasmin; Knaup, Sabine; Eberbeck, Dietmar; Tietze, Rainer; Unterweger, Harald; Friedrich, Ralf P; Duerr, Stephan; Heimke-Brinck, Ralph; Baum, Eva; Cicha, Iwona; Dörje, Frank; Odenbach, Stefan; Lyer, Stefan; Lee, Geoffrey; Alexiou, Christoph

    2014-01-01

    The promising potential of superparamagnetic iron oxide nanoparticles (SPIONs) in various nanomedical applications has been frequently reported. However, although many different synthesis methods, coatings, and functionalization techniques have been described, not many core-shell SPION drug delivery systems are available for clinicians at the moment. Here, bovine serum albumin was adsorbed onto lauric acid-stabilized SPIONs. The agglomeration behavior, zeta potential, and their dependence on the synthesis conditions were characterized with dynamic light scattering. The existence and composition of the core-shell-matrix structure was investigated by transmission electron microscopy, Fourier transform infrared spectroscopy, and zeta potential measurements. We showed that the iron oxide cores form agglomerates in the range of 80 nm. Moreover, despite their remarkably low tendency to aggregate even in a complex media like whole blood, the SPIONs still maintained their magnetic properties and were well attractable with a magnet. The magnetic properties were quantified by vibrating sample magnetometry and a superconducting quantum interference device. Using flow cytometry, we further investigated the effects of the different types of nanoparticle coating on morphology, viability, and DNA integrity of Jurkat cells. We showed that by addition of bovine serum albumin, the toxicity of nanoparticles is greatly reduced. We also investigated the effect of the particles on the growth of primary human endothelial cells to further demonstrate the biocompatibility of the particles. As proof of principle, we showed that the hybrid-coated particles are able to carry payloads of up to 800 μg/mL of the cytostatic drug mitoxantrone while still staying colloidally stable. The drug-loaded system exhibited excellent therapeutic potential in vitro, exceeding that of free mitoxantrone. In conclusion, we have synthesized a biocompatible ferrofluid that shows great potential for clinical

  1. Hybrid molecularly imprinted poly(methacrylic acid-TRIM)-silica chemically modified with (3-glycidyloxypropyl)trimethoxysilane for the extraction of folic acid in aqueous medium.

    PubMed

    de Oliveira, Fernanda Midori; Segatelli, Mariana Gava; Tarley, César Ricardo Teixeira

    2016-02-01

    In the present study a hybrid molecularly imprinted poly(methacrylic acid-trimethylolpropane trimethacrylate)-silica (MIP) was synthesized and modified with (3-glycidyloxypropyl)trimethoxysilane (GPTMS) with posterior opening of epoxy ring to provide hydrophilic properties of material in the extraction of folic acid from aqueous medium. The chemical and structural aggregates of hybrid material were characterized by means of Fourier Transform Infrared (FT-IR), Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Thermogravimetric analysis (TGA) and textural data. Selectivity data of MIP were compared to non-imprinted polymer (NIP) through competitive sorption studies in the presence of caffeine, paracetamol or 4-aminobenzamide yielding relative selectivity coefficients (k′) higher than one unit, thus confirming the selective character of MIP even in the presence of structurally smaller compounds than the folic acid. The lower hydrophobic sorption by bovine serum albumin (BSA) in the MIP as compared to unmodified MIP proves the hydrophilicity of polymer surface by using GPTMS with opening ring. Under acid medium(pH 1.5) the sorption of folic acid onto MIP from batch experiments was higher than the one achieved for NIP. Equilibrium sorption of folic acid was reached at 120 min for MIP, NIP and MIP without GPTMS and kinetic sorption data were well described by pseudo-second-order, Elovich and intraparticle diffusion models. Thus, these results indicate the existence of different binding energy sites in the polymers and a complex mechanism consisting of both surface sorption and intraparticle transport of folic acid within the pores of polymers.

  2. Differentiation of Candida albicans and Candida dubliniensis by Fluorescent In Situ Hybridization with Peptide Nucleic Acid Probes

    PubMed Central

    Oliveira, Kenneth; Haase, Gerhard; Kurtzman, Cletus; Hyldig-Nielsen, Jens Jo/rgen; Stender, Henrik

    2001-01-01

    The recent discovery of Candida dubliniensis as a separate species that traditionally has been identified as Candida albicans has led to the development of a variety of biochemical and molecular methods for the differentiation of these two pathogenic yeasts. rRNA sequences are well-established phylogenetic markers, and probes targeting species-specific rRNA sequences have been used in diagnostic assays for the detection and identification of microorganisms. Peptide nucleic acid (PNA) is a DNA mimic with improved hybridization characteristics, and the neutral backbone of PNA probes offers significant advantages in whole-cell in situ hybridization assays. In this study, we developed PNA probes targeting the rRNAs of C. albicans and C. dubliniensis and applied them to a fluorescence in situ hybridization method (PNA FISH) for differentiation between C. albicans and C. dubliniensis. Liquid cultures were smeared onto microscope slides, heat fixed, and then hybridized for 30 min. Unhybridized PNA probe was removed by washing, and smears were examined by fluorescence microscopy. Evaluation of the PNA FISH method using smears of 79 C. dubliniensis and 70 C. albicans strains showed 100% sensitivity and 100% specificity for both PNA probes. We concluded that PNA FISH is a powerful tool for the differentiation of C. albicans and C. dubliniensis. PMID:11682542

  3. Synthesis and antiproliferative activity of two diastereomeric lignan amides serving as dimeric caffeic acid-l-DOPA hybrids.

    PubMed

    Magoulas, George E; Rigopoulos, Andreas; Piperigkou, Zoi; Gialeli, Chrysostomi; Karamanos, Nikos K; Takis, Panteleimon G; Troganis, Anastassios N; Chrissanthopoulos, Athanassios; Maroulis, George; Papaioannou, Dionissios

    2016-06-01

    Two new diastereomeric lignan amides (4 and 5) serving as dimeric caffeic acid-l-DOPA hybrids were synthesized. The synthesis involved the FeCl3-mediated phenol oxidative coupling of methyl caffeate to afford trans-diester 1a as a mixture of enantiomers, protection of the catechol units, regioselective saponification, coupling with a suitably protected l-DOPA derivative, separation of the two diastereomers thus obtained by flash column chromatography and finally global chemoselective deprotection of the catechol units. The effect of hybrids 4 and 5 and related compounds on the proliferation of two breast cancer cell lines with different metastatic potential and estrogen receptor status (MDA-MB-231 and MCF-7) and of one epithelial lung cancer cell line, namely A-549, was evaluated for concentrations ranging from 1 to 256μM and periods of treatment of 24, 48 and 72h. Both hybrids showed interesting and almost equipotent antiproliferative activities (IC50 64-70μM) for the MDA-MB-231 cell line after 24-48h of treatment, but they were more selective and much more potent (IC50 4-16μM) for the MCF-7 cells after 48h of treatment. The highest activity for both hybrids and both breast cancer lines was observed after 72h of treatment (IC50 1-2μM), probably as the result of slow hydrolysis of their methyl ester functions.

  4. Fluorescence resonance energy transfer from pyrene to perylene labels for nucleic acid hybridization assays under homogeneous solution conditions

    PubMed Central

    Masuko, Masayuki; Ohuchi, Shohkichi; Sode, Koji; Ohtani, Hiroyuki; Shimadzu, Akira

    2000-01-01

    We characterized the fluorescence resonance energy transfer (FRET) from pyrene (donor) to perylene (acceptor) for nucleic acid assays under homogeneous solution conditions. We used the hybridization between a target 32mer and its complementary two sequential 16mer deoxyribonucleotides whose neighboring terminals were each respectively labeled with a pyrene and a perylene residue. A transfer efficiency of ~100% was attained upon the hybridization when observing perylene fluorescence at 459 nm with 347-nm excitation of a pyrene absorption peak. The Förster distance between two dye residues was 22.3 Å (the orientation factor of 2/3). We could change the distance between the residues by inserting various numbers of nucleotides into the center of the target, thus creating a gap between the dye residues on a hybrid. Assuming that the number of inserted nucleotides is proportional to the distance between the dye residues, the energy transfer efficiency versus number of inserted nucleotides strictly obeyed the Förster theory. The mean inter-nucleotide distance of the single-stranded portion was estimated to be 2.1 Å. Comparison between the fluorescent properties of a pyrene–perylene pair with those of a widely used fluorescein–rhodamine pair showed that the pyrene–perylene FRET is suitable for hybridization assays. PMID:10734211

  5. Study of nucleic acid-gold nanorod interactions and detecting nucleic acid hybridization using gold nanorod solutions in the presence of sodium citrate.

    PubMed

    Kanjanawarut, Roejarek; Su, Xiaodi

    2010-09-01

    In this study, the authors report that sodium citrate can aggregate hexadecyl-trimethyl-ammonium ion(+)-coated gold nanorods (AuNRs), and nucleic acids of different charge and structure properties, i.e., single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), single-stranded peptide nucleic acid (PNA), and PNA-DNA complex, can bind to the AuNRs and therefore retard the sodium citrate-induced aggregation to different extents. The discovery that hybridized dsDNA (and the PNA-DNA complex) has a more pronounced protection effect than ssDNA (and PNA) allows the authors to develop a homogeneous phase AuNRs-based UV-visible (UV-vis) spectral assay for detecting specific sequences of oligonucleotides (20 mer) with a single-base-mismatch selectivity and a limit of detection of 5 nM. This assay involves no tedious bioconjugation and on-particle hybridization. The simple "set and test" format allows for a highly efficient hybridization in a homogeneous phase and a rapid display of the results in less than a minute. By measuring the degree of reduction in AuNR aggregation in the presence of different nucleic acid samples, one can assess how different nucleic acids interact with the AuNRs to complement the knowledge of spherical gold nanoparticles. Besides UV-vis characterization, transmission electron microscopy and zeta potential measurements were conduced to provide visual evidence of the particle aggregation and to support the discussion of the assay principle.

  6. Synthesis of Bioactive Chlorogenic Acid-Silica Hybrid Materials via the Sol–Gel Route and Evaluation of Their Biocompatibility

    PubMed Central

    Pacifico, Severina

    2017-01-01

    Natural phenol compounds are gaining a great deal of attention because of their potential use as prophylactic and therapeutic agents in many diseases, as well as in applied science for their preventing role in oxidation deterioration. With the aim to synthetize new phenol-based materials, the sol–gel method was used to embed different content of the phenolic antioxidant chlorogenic acid (CGA) within silica matrices to obtain organic-inorganic hybrid materials. Fourier transform infrared (FTIR) measurements were used to characterize the prepared materials. The new materials were screened for their bioactivity and antioxidant potential. To this latter purpose, direct DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2′-azinobis-(3-ethylbenzothiazolin-6-sulfonic acid) methods were applied: radical scavenging capability appeared strongly dependent on the phenol amount in investigated hybrids, and became pronounced, mainly toward the ABTS radical cation, when materials with CGA content equal to 15 wt% and 20 wt% were analyzed. The in vitro biocompatibility of the synthetized materials was estimated by using the MTT assay towards fibroblast NIH 3T3 cells, human keratinocyte HaCaT cells, and the neuroblastoma SH-SY5Y cell line. As cell viability and morphology of tested cell lines seemed to be unaffected by new materials, the attenuated total reflectance (ATR)-FTIR method was applied to deeply measure the effects of the hybrids in the three different cell lines. PMID:28773198

  7. Nucleic acid sandwich hybridization assay with quantum dot-induced fluorescence resonance energy transfer for pathogen detection.

    PubMed

    Chou, Cheng-Chung; Huang, Yi-Han

    2012-12-04

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

  8. Preparation and characterization of zinc oxide nanoparticles and their sensor applications for electrochemical monitoring of nucleic acid hybridization.

    PubMed

    Yumak, Tugrul; Kuralay, Filiz; Muti, Mihrican; Sinag, Ali; Erdem, Arzum; Abaci, Serdar

    2011-09-01

    In this study, ZnO nanoparticles (ZNP) of approximately 30 nm in size were synthesized by the hydrothermal method and characterized by X-ray diffraction (XRD), Braun-Emmet-Teller (BET) N2 adsorption analysis and transmission electron microscopy (TEM). ZnO nanoparticles enriched with poly(vinylferrocenium) (PVF+) modified single-use graphite electrodes were then developed for the electrochemical monitoring of nucleic acid hybridization related to the Hepatitis B Virus (HBV). Firstly, the surfaces of polymer modified and polymer-ZnO nanoparticle modified single-use pencil graphite electrodes (PGEs) were characterized using scanning electron microscopy (SEM). The electrochemical behavior of these electrodes was also investigated using differential pulse voltammetry (DPV) and electrochemical impedance spectroscopy (EIS). Subsequently, the polymer-ZnO nanoparticle modified PGEs were evaluated for the electrochemical detection of DNA based on the changes at the guanine oxidation signals. Various modifications in DNA oligonucleotides and probe concentrations were examined in order to optimize the electrochemical signals that were generated by means of nucleic acid hybridization. After the optimization studies, the sequence-selective DNA hybridization was investigated in the case of a complementary amino linked probe (target), or noncomplementary (NC) sequences, or target and mismatch (MM) mixture in the ratio of (1:1).

  9. Synthesis of Bioactive Chlorogenic Acid-Silica Hybrid Materials via the Sol-Gel Route and Evaluation of Their Biocompatibility.

    PubMed

    Catauro, Michelina; Pacifico, Severina

    2017-07-21

    Natural phenol compounds are gaining a great deal of attention because of their potential use as prophylactic and therapeutic agents in many diseases, as well as in applied science for their preventing role in oxidation deterioration. With the aim to synthetize new phenol-based materials, the sol-gel method was used to embed different content of the phenolic antioxidant chlorogenic acid (CGA) within silica matrices to obtain organic-inorganic hybrid materials. Fourier transform infrared (FTIR) measurements were used to characterize the prepared materials. The new materials were screened for their bioactivity and antioxidant potential. To this latter purpose, direct DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2'-azinobis-(3-ethylbenzothiazolin-6-sulfonic acid) methods were applied: radical scavenging capability appeared strongly dependent on the phenol amount in investigated hybrids, and became pronounced, mainly toward the ABTS radical cation, when materials with CGA content equal to 15 wt% and 20 wt% were analyzed. The in vitro biocompatibility of the synthetized materials was estimated by using the MTT assay towards fibroblast NIH 3T3 cells, human keratinocyte HaCaT cells, and the neuroblastoma SH-SY5Y cell line. As cell viability and morphology of tested cell lines seemed to be unaffected by new materials, the attenuated total reflectance (ATR)-FTIR method was applied to deeply measure the effects of the hybrids in the three different cell lines.

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

    PubMed Central

    Chou, Cheng-Chung; Huang, Yi-Han

    2012-01-01

    This paper reports a nucleic acid sandwich hybridization assay with a quantum dot (QD)-induced fluorescence resonance energy transfer (FRET) reporter system. Two label-free hemagglutinin H5 sequences (60-mer DNA and 630-nt cDNA fragment) of avian influenza viruses were used as the targets in this work. Two oligonucleotides (16 mers and 18 mers) that specifically recognize two separate but neighboring regions of the H5 sequences were served as the capturing and reporter probes, respectively. The capturing probe was conjugated to QD655 (donor) in a molar ratio of 10:1 (probe-to-QD), and the reporter probe was labeled with Alexa Fluor 660 dye (acceptor) during synthesis. The sandwich hybridization assay was done in a 20 μL transparent, adhesive frame-confined microchamber on a disposable, temperature-adjustable indium tin oxide (ITO) glass slide. The FRET signal in response to the sandwich hybridization was monitored by a homemade optical sensor comprising a single 400 nm UV light-emitting diode (LED), optical fibers, and a miniature 16-bit spectrophotometer. The target with a concentration ranging from 0.5 nM to 1 μM was successfully correlated with both QD emission decrease at 653 nm and dye emission increase at 690 nm. To sum up, this work is beneficial for developing a portable QD-based nucleic acid sensor for on-site pathogen detection. PMID:23211753

  11. A formalin-free method for stabilizing cells for nucleic acid amplification, hybridization and next-generation sequencing.

    PubMed

    Qin, Jianbing; Sanmann, Jennifer N; Kittrell, Jeff S; Althof, Pamela A; Kaspar, Erin E; Hunsley, Bradford A

    2015-12-09

    Formalin has been widely used by pathology laboratories. Its carcinogenicity has led researchers to explore formalin substitutes. Streck Cell Preservative (SCP) is a formalin-free preservative that can preserve cellular antigens. This study was undertaken to investigate the effects of cell preservation using SCP on nucleic acid amplification, hybridization, and next-generation sequencing (NGS) as compared to control frozen cells and cells fixed in the traditional cell and tissue fixative, 10 % neutral buffered formalin (NBF). The breast cancer cell line, SKBR-3, was used as a model system. Prior to nucleic acid extraction and fluorescence in situ hybridization (FISH), cells were fixed in SCP or NBF overnight at room temperature with frozen cells in parallel. Analysis showed that similar DNA extraction yields and amplification profiles determined by PCR in SCP preserved cells and control frozen cells, whereas NBF preserved cells had decreased DNA yield and impaired PCR amplification. Molecular cytogenetic studies by FISH technique indicated that the ratios of ERBB2 (HER-2/neu) signals to the chromosome 17 centromere (CEP17) were comparable for frozen cells and SCP preserved cells. The fluorescence images of both SCP fixed and control frozen cells were also clear and comparable. On the contrary, the same analysis was unsuccessful with NBF preserved cells due to poor hybridization quality. Our data also demonstrated that SCP had negligible effect on NGS testing. We conclude that SCP can be used as an alternative to NBF as a preservative for maintaining the integrity of nucleic acids for nucleic acid amplification, sequencing and FISH analysis.

  12. Human papillomavirus 35 nucleic acid hybridization probes and methods for employing the same

    SciTech Connect

    Lorincz, A.T.

    1989-07-18

    This patent describes an HPV 35 hybridization probe comprising a member selected from the group consisting of (i) HPV 35 DNA or fragments thereof labelled with a marker and (ii) HPV 35 RNA or fragments thereof labelled with a marker.

  13. Human papillomavirus 43 nucleic acid hybridization probes and methods for employing the same

    SciTech Connect

    Lorincz, A.T.

    1989-07-18

    This patent describes an HPV 43 hybridization probe comprising a member selected from the group consisting of (i) HPV 43 DNA or fragments thereof labelled with a marker and (ii) HPV 43 RNA or fragments thereof labelled with a marker.

  14. Human papillomavirus 56 nucleic acid hybridization probes and methods for employing the same

    SciTech Connect

    Lorinez, A.T.

    1990-03-13

    This patent describes an HPV 56 hybridization probe. It comprises: a member selected from the group consisting of HPV 56 DNA or fragments thereof labelled with a marker and HPV 56 RNA or fragments thereof labelled with a marker.

  15. Human papillomavirus 44 nucleic acid hybridization probes and methods for employing the same

    SciTech Connect

    Lorincz, A.T.

    1989-07-18

    This patent describes an HPV 44 hybridization probe comprising a member selected from the group consisting of (1) HPV 44 DNA or fragments thereof labelled with a marker and (ii) HPV 44 RNA or fragments thereof labelled with a marker.

  16. Search for genes responsible for the remarkably high acetic acid tolerance of a Zygosaccharomyces bailii-derived interspecies hybrid strain.

    PubMed

    Palma, Margarida; Roque, Filipa de Canaveira; Guerreiro, Joana Fernandes; Mira, Nuno Pereira; Queiroz, Lise; Sá-Correia, Isabel

    2015-12-16

    Zygosaccharomyces bailii is considered the most problematic acidic food spoilage yeast species due to its exceptional capacity to tolerate high concentrations of weak acids used as fungistatic preservatives at low pH. However, the mechanisms underlying its intrinsic remarkable tolerance to weak acids remain poorly understood. The identification of genes and mechanisms involved in Z. bailii acetic acid tolerance was on the focus of this study. For this, a genomic library from the highly acetic acid tolerant hybrid strain ISA1307, derived from Z. bailii and a closely related species and isolated from a sparkling wine production plant, was screened for acetic acid tolerance genes. This screen was based on the transformation of an acetic acid susceptible Saccharomyces cerevisiae mutant deleted for the gene encoding the acetic acid resistance determinant transcription factor Haa1. The expression of 31 different DNA inserts from ISA1307 strain genome was found to significantly increase the host cell tolerance to acetic acid. The in silico analysis of these inserts was facilitated by the recently available genome sequence of this strain. In total, 65 complete or truncated ORFs were identified as putative determinants of acetic acid tolerance and an S. cerevisiae gene homologous to most of them was found. These include genes involved in cellular transport and transport routes, protein fate, protein synthesis, amino acid metabolism and transcription. The role of strong candidates in Z. bailii and S. cerevisiae acetic acid tolerance was confirmed based on homologous and heterologous expression analyses. ISA1307 genes homologous to S. cerevisiae genes GYP8, WSC4, PMT1, KTR7, RKR1, TIF3, ILV3 and MSN4 are proposed as strong candidate determinants of acetic acid tolerance. The ORF ZBAI_02295 that contains a functional domain associated to the uncharacterised integral membrane proteins of unknown function of the DUP family is also suggested as a relevant tolerance determinant

  17. Amino acids are major energy substrates for tissues of hybrid striped bass and zebrafish.

    PubMed

    Jia, Sichao; Li, Xinyu; Zheng, Shixuan; Wu, Guoyao

    2017-08-29

    Fish generally have much higher requirements for dietary protein than mammals, and this long-standing puzzle remains unsolved. The present study was conducted with zebrafish (omnivores) and hybrid striped bass (HSB, carnivores) to test the hypothesis that AAs are oxidized at a higher rate than carbohydrates (e.g., glucose) and fatty acids (e.g., palmitate) to provide ATP for their tissues. Liver, proximal intestine, kidney, and skeletal muscle isolated from zebrafish and HSB were incubated at 28.5 °C (zebrafish) or 26 °C (HSB) for 2 h in oxygenated Krebs-Henseleit bicarbonate buffer (pH 7.4, with 5 mM D-glucose) containing 2 mM L-[U-(14)C]glutamine, L-[U-(14)C]glutamate, L-[U-(14)C]leucine, or L-[U-(14)C]palmitate, or a trace amount of D-[U-(14)C]glucose. In parallel experiments, tissues were incubated with a tracer and  a mixture of unlabeled substrates [glutamine, glutamate, leucine, and palmitate (2 mM each) plus 5 mM D-glucose]. (14)CO2 was collected to calculate the rates of substrate oxidation. In the presence of glucose or a mixture of substrates, the rates of oxidation of glutamate and ATP production from this AA by the proximal intestine, liver, and kidney of HSB   were much higher than those for glucose and palmitate. This was also true for glutamate in the skeletal muscle and glutamine in the liver of both species, glutamine in the HSB kidney, and leucine in the zebrafish muscle, in the presence of a mixture of substrates. We conclude that glutamate plus glutamine plus leucine contribute to ~80% of ATP production in the liver, proximal intestine, kidney, and skeletal muscle of zebrafish and HSB. Our findings provide the first direct evidence that the major tissues of fish use AAs (mainly glutamate and glutamine) as primary energy sources instead of carbohydrates or lipids.

  18. A hybrid of ant colony optimization and minimization of metabolic adjustment to improve the production of succinic acid in Escherichia coli.

    PubMed

    Chong, Shiue Kee; Mohamad, Mohd Saberi; Mohamed Salleh, Abdul Hakim; Choon, Yee Wen; Chong, Chuii Khim; Deris, Safaai

    2014-06-01

    This paper presents a study on gene knockout strategies to identify candidate genes to be knocked out for improving the production of succinic acid in Escherichia coli. Succinic acid is widely used as a precursor for many chemicals, for example production of antibiotics, therapeutic proteins and food. However, the chemical syntheses of succinic acid using the traditional methods usually result in the production that is far below their theoretical maximums. In silico gene knockout strategies are commonly implemented to delete the gene in E. coli to overcome this problem. In this paper, a hybrid of Ant Colony Optimization (ACO) and Minimization of Metabolic Adjustment (MoMA) is proposed to identify gene knockout strategies to improve the production of succinic acid in E. coli. As a result, the hybrid algorithm generated a list of knockout genes, succinic acid production rate and growth rate for E. coli after gene knockout. The results of the hybrid algorithm were compared with the previous methods, OptKnock and MOMAKnock. It was found that the hybrid algorithm performed better than OptKnock and MOMAKnock in terms of the production rate. The information from the results produced from the hybrid algorithm can be used in wet laboratory experiments to increase the production of succinic acid in E. coli. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Localization of glucocorticoid receptor messenger ribonucleic acid in hippocampus of rat brain using in situ hybridization

    SciTech Connect

    Yang, G.; Matocha, M.F.; Rapoport, S.I.

    1988-08-01

    An in situ hybridization procedure was applied to quantify glucocorticoid receptor (GR) mRNAs in the hippocampus of rat brain. Hybridization was carried out using a radiolabeled antisense probe complementary to the rat liver GR gene. The specificity of the method was validated by showing: 1) a high cellular grain density in sections hybridized with an antisense but not a sense probe; 2) agreement between the experimental and theoretical temperature at which 50% of the hybrids melted, and 3) a high signal distribution of GR mRNA in the hippocampus, a region of brain known to preferentially concentrate steroid hormones. Within the hippocampus, however, subregional differences in hybridization densities were observed. Quantitative autoradiography indicated that the average neuronal silver grain number was highest in the pyramidal cell layers of CA2 and CA4 and lowest in those of CA1 and CA3. Also, there was a significant difference in the average grain number between all of the cell fields except for that between CA2 and CA4. These results show that contiguous but neuroanatomically distinct cell fields of the hippocampus express different levels of GR transcripts, and indicate that differential regulation of GR expression occurs in subpopulations of hippocampal neurons.

  20. Identification of Dekkera bruxellensis (Brettanomyces) from wine by fluorescence in situ hybridization using peptide nucleic acid probes.

    PubMed

    Stender, H; Kurtzman, C; Hyldig-Nielsen, J J; Sørensen, D; Broomer, A; Oliveira, K; Perry-O'Keefe, H; Sage, A; Young, B; Coull, J

    2001-02-01

    A new fluorescence in situ hybridization method using peptide nucleic acid (PNA) probes for identification of Brettanomyces is described. The test is based on fluorescein-labeled PNA probes targeting a species-specific sequence of the rRNA of Dekkera bruxellensis. The PNA probes were applied to smears of colonies, and results were interpreted by fluorescence microscopy. The results obtained from testing 127 different yeast strains, including 78 Brettanomyces isolates from wine, show that the spoilage organism Brettanomyces belongs to the species D. bruxellensis and that the new method is able to identify Brettanomyces (D. bruxellensis) with 100% sensitivity and 100% specificity.

  1. Identification of Dekkera bruxellensis (Brettanomyces) from Wine by Fluorescence In Situ Hybridization Using Peptide Nucleic Acid Probes

    PubMed Central

    Stender, Henrik; Kurtzman, Cletus; Hyldig-Nielsen, Jens J.; Sørensen, Ditte; Broomer, Adam; Oliveira, Kenneth; Perry-O'Keefe, Heather; Sage, Andrew; Young, Barbara; Coull, James

    2001-01-01

    A new fluorescence in situ hybridization method using peptide nucleic acid (PNA) probes for identification of Brettanomyces is described. The test is based on fluorescein-labeled PNA probes targeting a species-specific sequence of the rRNA of Dekkera bruxellensis. The PNA probes were applied to smears of colonies, and results were interpreted by fluorescence microscopy. The results obtained from testing 127 different yeast strains, including 78 Brettanomyces isolates from wine, show that the spoilage organism Brettanomyces belongs to the species D. bruxellensis and that the new method is able to identify Brettanomyces (D. bruxellensis) with 100% sensitivity and 100% specificity. PMID:11157265

  2. Use of Peptide Nucleic Acid-Fluorescence In Situ Hybridization for Definitive, Rapid Identification of Five Common Candida Species▿

    PubMed Central

    Reller, Megan E.; Mallonee, Amanda B.; Kwiatkowski, Nicole P.; Merz, William G.

    2007-01-01

    We investigated a 2.5-h peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) assay with five Candida species-specific probes to identify Candida colonies and compared it to standard 2-h to 5-day phenotypic identification methods. Suspensions were made and slides were prepared and read for fluorescence per the manufacturer's instructions. Sensitivity was 99% (109/110), and specificity was 99% (129/130). PNA-FISH can rapidly identify those Candida species isolated most frequently. PMID:17804657

  3. Syntheses and structure characterization of ten acid-base hybrid crystals based on N-containing aromatic brønsted bases and mineral acids

    NASA Astrophysics Data System (ADS)

    Lin, Zhihao; Jin, Shouwen; Li, Xiaoliang; Xiao, Xiao; Hu, Kaikai; Guo, Ming; Chi, Xinchen; Liu, Hui; Wang, Daqi

    2017-10-01

    Cocrystallization of the aromatic brønsted bases with a series of mineral acids gave a total of ten hybrid salts with the compositions: (2-methylquinoline)2: (hydrochloride acid): 3H2O [(HL1)+. (L1)·· (Cl-) · (H2O)3] (1), (6-bromobenzo[d]thiazol-2-amine): (hydrochloride acid) [(HL2)+. (Cl-)] (2), (6-bromobenzo[d]thiazol-2-amine): (nitric acid) [(HL2)+. (NO3-)] (3), (6-bromobenzo[d]thiazol-2-amine): (sulfuric acid) [(HL2)+ · (HSO4)-] (4), (6-bromobenzo[d]thiazol-2-amine): (phosphoric acid) [(HL2)+ · (H2PO4)-] (5), (5,7-dimethyl-1,8-naphthyridine-2-amine): (hydrochloride acid): 3H2O [(HL3)+ · (Cl-) (H2O)3] (6), (5,7-dimethyl-1,8-naphthyridine-2-amine): (hydrobromic acid): CH3OH [(HL3)+ · (Br)- · CH3OH] (7), (5,7-dimethyl-1,8-naphthyridine-2-amine): (sulfuric acid): H2O [(HL3)+ · (HSO4)- · H2O] (8), (2-aminophenol): (phosphoric acid) [(HL4)+ · (H2PO4)-] (9), and (2-amino-4-chlorophenol): (phosphoric acid) [(HL5)+ · (H2PO4)-] (10). The ten salts have been characterized by X-ray diffraction analysis, IR, and elemental analysis, and the melting points of all the salts were also reported. And their structural and supramolecular aspects are fully analyzed. The result reveals that among the ten investigated crystals the ring N of the heterocycle or the NH2 in the aminophenol are protonated when the acids are deprotonated, and the crystal packing is interpreted in terms of the strong charge-assisted classical hydrogen bonds between the NH+/NH3+ and deprotonated acidic groups. Further analysis of the crystal packing of the salts indicated that a different family of additional CHsbnd O, CHsbnd Cl, CH3sbnd N, CH3sbnd O, CHsbnd Br, CH3sbnd Br, Brsbnd Cl, Clsbnd S, Osbnd S, Osbnd O, Brsbnd S, Hsbnd H, and π-π associations contribute to the stabilization and expansion of the total high-dimensional frameworks. For the coexistence of the various weak nonbonding interactions these structures adopted homo or hetero supramolecular synthons or both. Some classical

  4. Label-free amino acid detection based on nanocomposites of graphene oxide hybridized with gold nanoparticles.

    PubMed

    Zhang, Qian; Zhang, Diming; Lu, Yanli; Xu, Gang; Yao, Yao; Li, Shuang; Liu, Qingjun

    2016-03-15

    Nanocomposites of graphene oxide and gold nanoparticles (GO/GNPs) were synthesized for label-free detections of amino acids. Interactions between the composites and amino acids were investigated by both naked-eye observation and optical absorption spectroscopy. The GO/GNPs composites displayed apparent color changes and absorption spectra changes in presences of amino acids including glutamate, aspartate, and cysteine. The interaction mechanisms of the composites and amino acids were discussed and explored with sulfhydryl groups and non-α-carboxylic groups on the amino acids. Sensing properties of the composites were tested, while pure gold particles were used as the control. The results suggested that the GO/GNPs composites had better linearity and stability in dose-dependent responses to the amino acids than those of the particles, especially in detections for acidic amino acids. Therefore, the nanocomposites platform can provide a convenient and efficient approach for label-free optical detections of important molecules such as amino acids.

  5. Hybridization probe for femtomolar quantification of selected nucleic acid sequences on a disposable electrode.

    PubMed

    Jenkins, Daniel M; Chami, Bilal; Kreuzer, Matthias; Presting, Gernot; Alvarez, Anne M; Liaw, Bor Yann

    2006-04-01

    Mixed monolayers of electroactive hybridization probes on gold surfaces of a disposable electrode were investigated as a technology for simple, sensitive, selective, and rapid gene identification. Hybridization to the ferrocene-labeled hairpin probes reproducibly diminished cyclic redox currents, presumably due to a displacement of the label from the electrode. Observed peak current densities were roughly 1000x greater than those observed in previous studies, such that results could easily be interpreted without the use of algorithms to correct for background polarization currents. Probes were sensitive to hybridization with a number of oligonucleotide sequences with varying homology, but target oligonucleotides could be distinguished from competing nontarget sequences based on unique "melting" profiles from the probe. Detection limits were demonstrated down to nearly 100 fM, which may be low enough to identify certain genetic conditions or infections without amplification. This technology has rich potential for use in field devices for gene identification as well as in gene microarrays.

  6. Strong, Thermally Superinsulating Biopolymer-Silica Aerogel Hybrids by Cogelation of Silicic Acid with Pectin.

    PubMed

    Zhao, Shanyu; Malfait, Wim J; Demilecamps, Arnaud; Zhang, Yucheng; Brunner, Samuel; Huber, Lukas; Tingaut, Philippe; Rigacci, Arnaud; Budtova, Tatiana; Koebel, Matthias M

    2015-11-23

    Silica aerogels are excellent thermal insulators, but their brittle nature has prevented widespread application. To overcome these mechanical limitations, silica-biopolymer hybrids are a promising alternative. A one-pot process to monolithic, superinsulating pectin-silica hybrid aerogels is presented. Their structural and physical properties can be tuned by adjusting the gelation pH and pectin concentration. Hybrid aerogels made at pH 1.5 exhibit minimal dust release and vastly improved mechanical properties while remaining excellent thermal insulators. The change in the mechanical properties is directly linked to the observed "neck-free" nanoscale network structure with thicker struts. Such a design is superior to "neck-limited", classical inorganic aerogels. This new class of materials opens up new perspectives for novel silica-biopolymer nanocomposite aerogels.

  7. Multiplex Peptide Nucleic Acid Fluorescence In Situ Hybridization (PNA-FISH) for Diagnosis of Bacterial Vaginosis.

    PubMed

    Machado, Antonio; Cerca, Nuno

    2017-01-01

    Fluorescence in situ hybridization (FISH) is a molecular method used to identify and quantify microorganisms in a wide range of samples. This technique combines the simplicity of microscopic observation and the specificity of DNA/rRNA hybridization, allowing detection of selected bacterial species and morphologic visualization. Here, we describe a quantitative molecular diagnosis of bacterial vaginosis, based on the classical Nugent score. Our probes are able to differentiate Lactobacillus spp. and Gardnerella vaginalis from the other undefined bacterial species considered in the Nugent score.

  8. Hybridization-based detection of Helicobacter pylori at human body temperature using advanced locked nucleic acid (LNA) probes.

    PubMed

    Fontenete, Sílvia; Guimarães, Nuno; Leite, Marina; Figueiredo, Céu; Wengel, Jesper; Filipe Azevedo, Nuno

    2013-01-01

    The understanding of the human microbiome and its influence upon human life has long been a subject of study. Hence, methods that allow the direct detection and visualization of microorganisms and microbial consortia (e.g. biofilms) within the human body would be invaluable. In here, we assessed the possibility of developing a variant of fluorescence in situ hybridization (FISH), named fluorescence in vivo hybridization (FIVH), for the detection of Helicobacter pylori. Using oligonucleotide variations comprising locked nucleic acids (LNA) and 2'-O-methyl RNAs (2'OMe) with two types of backbone linkages (phosphate or phosphorothioate), we were able to successfully identify two probes that hybridize at 37 °C with high specificity and sensitivity for H. pylori, both in pure cultures and in gastric biopsies. Furthermore, the use of this type of probes implied that toxic compounds typically used in FISH were either found to be unnecessary or could be replaced by a non-toxic substitute. We show here for the first time that the use of advanced LNA probes in FIVH conditions provides an accurate, simple and fast method for H. pylori detection and location, which could be used in the future for potential in vivo applications either for this microorganism or for others.

  9. [Oligonucleotide derivatives in the nucleic acid hybridization analysis. II. Isothermal signal amplification in process of DNA analysis by minisequencing].

    PubMed

    Dmitrienko, E V; Khomiakova, E A; Pyshnaia; Bragin, A G; Vedernikov, V E; Pyshnyĭ, D V

    2010-01-01

    The isothermal amplification of reporter signal via limited probe extension (minisequencing) upon hybridization of nucleic acids has been studied. The intensity of reporter signal has been shown to increase due to enzymatic labeling of multiple probes upon consecutive hybridization with one DNA template both in homophase and heterophase assays using various kinds of detection signal: radioisotope label, fluorescent label, and enzyme-linked assay. The kinetic scheme of the process has been proposed and kinetic parameters for each step have been determined. The signal intensity has been shown to correlate with physicochemical characteristics of both complexes: probe/DNA and product/DNA. The maximum intensity has been observed at minimal difference between the thermodynamic stability of these complexes, provided the reaction temperature has been adjusted near their melting temperature values; rising or lowering the reaction temperature reduces the amount of reporting product. The signal intensity has been shown to decrease significantly upon hybridization with the DNA template containing single-nucleotide mismatches. Limited probe extension assay is useful not only for detection of DNA template but also for its quantitative characterization.

  10. Efficient boron abstraction using honeycomb-like porous magnetic hybrids: Assessment of techno-economic recovery of boric acid.

    PubMed

    Oladipo, Akeem Adeyemi; Gazi, Mustafa

    2016-12-01

    Porous magnetic hybrids were synthesized and functionalized with glycidol to produce boron-selective adsorbent. The magnetic hybrid (MH) comparatively out-performed the existing expensive adsorbents. MH had a saturation magnetisation of 63.48 emu/g and average pore diameter ranging from meso to macropores. The magnetic hybrids showed excellent selectivity towards boron and resulted in 79-93% boron removal even in the presence of competing metal ions (Na(+) and Cr(2+)). Experiments were performed in a column system, and breakthrough time was observed to increase with bed depths and decreased with flow rates. The batch experiments revealed that 60 min was enough to achieve equilibrium, and the level of boron sorption was 108.5 mg/g from a synthetic solution. Several adsorption-desorption cycles were performed using a simple acid-water treatment and evaluated using various kinetic models. The spent adsorbents could be separated easily from the mixture by an external magnetic field. The cost-benefit analysis was performed for the treatment of 72 m(3)/year boron effluent, including five years straight line depreciation charges of equipment. The net profit and standard percentage confirmed that the recovery process is economically feasible. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Hybridization-Based Detection of Helicobacter pylori at Human Body Temperature Using Advanced Locked Nucleic Acid (LNA) Probes

    PubMed Central

    Fontenete, Sílvia; Guimarães, Nuno; Leite, Marina; Figueiredo, Céu; Wengel, Jesper; Filipe Azevedo, Nuno

    2013-01-01

    The understanding of the human microbiome and its influence upon human life has long been a subject of study. Hence, methods that allow the direct detection and visualization of microorganisms and microbial consortia (e.g. biofilms) within the human body would be invaluable. In here, we assessed the possibility of developing a variant of fluorescence in situ hybridization (FISH), named fluorescence in vivo hybridization (FIVH), for the detection of Helicobacter pylori. Using oligonucleotide variations comprising locked nucleic acids (LNA) and 2’-O-methyl RNAs (2’OMe) with two types of backbone linkages (phosphate or phosphorothioate), we were able to successfully identify two probes that hybridize at 37 °C with high specificity and sensitivity for H. pylori, both in pure cultures and in gastric biopsies. Furthermore, the use of this type of probes implied that toxic compounds typically used in FISH were either found to be unnecessary or could be replaced by a non-toxic substitute. We show here for the first time that the use of advanced LNA probes in FIVH conditions provides an accurate, simple and fast method for H. pylori detection and location, which could be used in the future for potential in vivo applications either for this microorganism or for others. PMID:24278398

  12. Fabrication and structure analysis of poly(lactide-co-glycolic acid)/silk fibroin hybrid scaffold for wound dressing applications.

    PubMed

    Shahverdi, Sheida; Hajimiri, Mirhamed; Esfandiari, Mohammad Amin; Larijani, Bagher; Atyabi, Fatemeh; Rajabiani, Afsaneh; Dehpour, Ahmad Reza; Gharehaghaji, Ali Akbar; Dinarvand, Rassoul

    2014-10-01

    Silk fibroin (SF) and poly(lactide-co-glycolic acid) (PLGA) have been proved to be invaluable polymers in the field wound healing. This study aims at optimizing the electrospinning process of those polymers to make a hybrid membrane as a chronic wounds dressing. After characterizing the scaffolds, PLGA/SF (2:1), and PLGA scaffolds were selected for further study according to their superior tensile mechanical properties. The attachment and proliferation of mouse fibroblasts (L929) on scaffolds were measured using colorimetric assay and scanning electron microscopy. Furthermore, to evaluate the wound healing effect of the scaffolds in comparison with gauze and Comfeel(®) dressings, an excision wound model was conducted on diabetic rats. On the postoperative days of 3, 6, 9, 12, and 15, residual wound area was calculated using macroscopic data. In vitro results showed that the attachment and proliferation of L929 were significantly increased on PLGA/SF (2:1) hybrid scaffold. Animal study and histopathological evaluation outcomes confirmed the in vitro results as well. On day 15, the residual wound area in PLGA/SF (2:1) hybrid membrane group was significantly smaller than PLGA and control groups. This promising scaffold has the potential to be used for the upcoming development of wound dressings with or without biological drugs.

  13. 16S rRNA in situ Hybridization Followed by Flow Cytometry for Rapid Identification of Acetic Acid Bacteria Involved in Submerged Industrial Vinegar Production

    PubMed Central

    Lipoglavšek, Luka; Avguštin, Gorazd

    2016-01-01

    Summary Acetic acid bacteria are involved in many biotechnological processes such as vitamin C, gluconic acid, miglitol or acetic acid production, and others. For a technologist trying to control the industrial process, the ability to follow the microbiological development of the process is thus of importance. During the past few years hybridization in a combination with flow cytometry has often been used for this purpose. Since vinegar is a liquid, it is an ideal matrix for flow cytometry analysis. In this work we have constructed a specific probe for highly acetic acid-resistant species of the acetic acid bacteria and a protocol for in situ hybridization, which in combination with flow cytometry enables direct monitoring of bacteria producing vinegar with >10% of acetic acid. The approach was successfully applied for monitoring microbiota during industrial vinegar production. PMID:27904400

  14. 16S rRNA in situ Hybridization Followed by Flow Cytometry for Rapid Identification of Acetic Acid Bacteria Involved in Submerged Industrial Vinegar Production.

    PubMed

    Trček, Janja; Lipoglavšek, Luka; Avguštin, Gorazd

    2016-03-01

    Acetic acid bacteria are involved in many biotechnological processes such as vitamin C, gluconic acid, miglitol or acetic acid production, and others. For a technologist trying to control the industrial process, the ability to follow the microbiological development of the process is thus of importance. During the past few years hybridization in a combination with flow cytometry has often been used for this purpose. Since vinegar is a liquid, it is an ideal matrix for flow cytometry analysis. In this work we have constructed a specific probe for highly acetic acid-resistant species of the acetic acid bacteria and a protocol for in situ hybridization, which in combination with flow cytometry enables direct monitoring of bacteria producing vinegar with >10% of acetic acid. The approach was successfully applied for monitoring microbiota during industrial vinegar production.

  15. Novel Hybrid Catalyst for the Oxidation of Organic Acids: Pd Nanoparticles Supported on Mn-N-3D-Graphene Nanosheets

    DOE PAGES

    Perry, Albert; Kabir, Sadia; Matanovic, Ivana; ...

    2017-06-16

    This paper reports the fabrication and electrochemical performance of a hybrid catalyst composed of Pd nanoparticles and atomically dispersed Mn active centers integrated into the nitrogen-doped three-dimensional graphene nanosheets (Pd/Mn-N-3D-GNS). Our results show that the synergistic integration of both Pd nanoparticles and atomically dispersed Mn can be used to enhance the activity toward the electrochemical oxidation of organic acids at biologically relevant pH values. The hybrid catalyst (Pd/Mn-N-3D-GNS) showed increased maximum currents toward the oxidation of oxalic acid when compared to its individual catalysts, namely, Pd/3D-GNS and Mn N-3D-GNS catalysts. The hybrid also showed a decreased onset potential for oxidationmore » of mesoxalic acid as compared to Mn-N-3D-GNS and decreased onset potentials for the oxidation of glyoxalic acid when compared to both of its constituent catalysts. Oxidation of formic acid was also tested and the hybrid was shown to catalyze both dehydration and dehydrogenation mechanisms of formic acid electro-oxidation. Using density functional theory calculations, it was elucidated that a two-site catalysis most likely promotes dehydrogenation reaction for formic acid oxidation, which can explain the selectivity of Pd nanoparticles and atomically dispersed Mn towards the dehydrogenation/ dehydration pathway.« less

  16. Radical-generating coordination complexes as tools for rapid and effective fragmentation and fluorescent labeling of nucleic acids for microchip hybridization.

    SciTech Connect

    Kelly, J. J.; Chernov, B. K.; Tovstanovsky, I.; Mirzabekov, A. D.; Bavykin, S. G.; Biochip Technology Center; Northwestern Univ.; Engelhardt Inst. of Molecular Biology

    2002-12-15

    DNA microchip technology is a rapid, high-throughput method for nucleic acid hybridization reactions. This technology requires random fragmentation and fluorescent labeling of target nucleic acids prior to hybridization. Radical-generating coordination complexes, such as 1,10-phenanthroline-Cu(II) (OP-Cu) and Fe(II)-EDTA (Fe-EDTA), have been commonly used as sequence nonspecific 'chemical nucleases' to introduce single-strand breaks in nucleic acids. Here we describe a new method based on these radical-generating complexes for random fragmentation and labeling of both single- and double-stranded forms of RNA and DNA. Nucleic acids labeled with the OP-Cu and the Fe-EDTA protocols revealed high hybridization specificity in hybridization with DNA microchips containing oligonucleotide probes selected for identification of 16S rRNA sequences of the Bacillus group microorganisms.We also demonstrated cDNA- and cRNA-labeling and fragmentation with this method. Both the OP-Cu and Fe-EDTA fragmentation and labeling procedures are quick and inexpensive compared to other commonly used methods. A column-based version of the described method does not require centrifugation and therefore is promising for the automation of sample preparations in DNA microchip technology as well as in other nucleic acid hybridization studies.

  17. Synthesis and properties of poly(methyl methacrylate-2-acrylamido-2-methylpropane sulfonic acid)/PbS hybrid composite

    SciTech Connect

    Preda, N.; Rusen, E.; Musuc, A.; Enculescu, M.; Matei, E.; Marculescu, B.; Fruth, V.; Enculescu, I.

    2010-08-15

    The synthesis of a new hybrid composite based on PbS nanoparticles and poly(methyl methacrylate-2-acrylamido-2-methylpropane sulfonic acid) [P(MMA-AMPSA)] copolymer is reported. The chemical synthesis consists in two steps: (i) a surfactant-free emulsion copolymerization between methyl methacrylate and 2-acrylamido-2-methylpropane sulfonic acid and (ii) the generation of PbS particles in the presence of the P(MMA-AMPSA) latex, from the reaction between lead nitrate and thiourea. The composite was studied by scanning electron microscopy (SEM), X-ray diffraction, FTIR spectroscopy, thermogravimetric analysis and differential scanning calorimetry. The microstructure observed using SEM proves that the PbS nanoparticles are well dispersed in the copolymer matrix. The X-ray diffraction measurements demonstrate that the PbS nanoparticles have a cubic rock salt structure. It was also found that the inorganic semiconductor nanoparticles improve the thermal stability of the copolymer matrix.

  18. Carbon nanotube/raspberry hollow Pd nanosphere hybrids for methanol, ethanol, and formic acid electro-oxidation in alkaline media.

    PubMed

    Liu, Zhelin; Zhao, Bo; Guo, Cunlan; Sun, Yujing; Shi, Yan; Yang, Haibin; Li, Zhuang

    2010-11-01

    In this paper, raspberry hollow Pd nanospheres (HPNs)-decorated carbon nanotube (CNT) was developed for electro-oxidation of methanol, ethanol, and formic acid in alkaline media. The electrocatalyst was fabricated simply by attaching HPNs onto the surface of CNT which had been functionalized by polymer wrapping. The as-prepared HPN-CNTs (CHPNs) were characterized by transmission electron microscopy (TEM), scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), and X-ray photoelectron spectroscopy (XPS). The increasing interest and intensive research on fuel cell inspire us to investigate the electrocatalytic properties of the prepared nanostructures. Besides that, previous reports about alkaline other than acidic media could supply a more active environment guide us to examine the electrocatalytic properties in alkaline electrolyte. It is found that this novel hybrid electrocatalyst exhibits excellent electrocatalytic properties and can be further applied in fuel cells, catalysts, and sensors. Copyright 2010 Elsevier Inc. All rights reserved.

  19. Development and Application of Nucleic Acid Hybridization Techniques to Arbovirus Surveillance and Diagnosis.

    DTIC Science & Technology

    1987-08-14

    and was restricted to distinct peri-.uclear areas. By 12 hours, strong signal was detected in perinuclear spaces and signal was detectable in more...major problems or potential problems have been encountered which impede the use of hybridization in virus diagnosis and surveillan -e, especially in

  20. Unlocked nucleic acids with a pyrene-modified uracil: synthesis, hybridization studies, fluorescent properties and i-motif stability.

    PubMed

    Perlíková, Pavla; Karlsen, Kasper K; Pedersen, Erik B; Wengel, Jesper

    2014-01-03

    The synthesis of two new phosphoramidite building blocks for the incorporation of 5-(pyren-1-yl)uracilyl unlocked nucleic acid (UNA) monomers into oligonucleotides has been developed. Monomers containing a pyrene-modified nucleobase component were found to destabilize an i-motif structure at pH 5.2, both under molecular crowding and noncrowding conditions. The presence of the pyrene-modified UNA monomers in DNA strands led to decreases in the thermal stabilities of DNA*/DNA and DNA*/RNA duplexes, but these duplexes' thermal stabilities were better than those of duplexes containing unmodified UNA monomers. Pyrene-modified UNA monomers incorporated in bulges were able to stabilize DNA*/DNA duplexes due to intercalation of the pyrene moiety into the duplexes. Steady-state fluorescence emission studies of oligonucleotides containing pyrene-modified UNA monomers revealed decreases in fluorescence intensities upon hybridization to DNA or RNA. Efficient quenching of fluorescence of pyrene-modified UNA monomers was observed after formation of i-motif structures at pH 5.2. The stabilizing/destabilizing effect of pyrene-modified nucleic acids might be useful for designing antisense oligonucleotides and hybridization probes.

  1. A simple, rapid method of nucleic acid extraction without tissue homogenization for detecting viroids by hybridization and RT-PCR.

    PubMed

    Nakahara, K; Hataya, T; Uyeda, I

    1999-01-01

    A simple, rapid method of nucleic acid extraction on a microcentrifuge tube scale for detecting viroids is presented. Five distinct citrus viroids (CVds), chrysanthemum stunt viroid (CSVd), hop stunt viroid (HSVd), hop latent viroid (HLVd) and potato spindle tuber viroid (PSTVd) were detected in their natural host plants by hybridization using cRNA probes and reverse transcription-polymerase chain reaction (RT-PCR). Nucleic acids (NA) were liberated from tissues by incubation in a buffer containing potassium ethyl xanthogenate (PEX) without tissue homogenization, and then precipitated with ethanol (NA-PEX). All the viroids except CVd-IV could be detected clearly in NA-PEX by hybridization. HSVd, HLVd and PSTVd could also be detected in NA-PEX by RT-PCR. Although CVds and CSVd could not be detected in NA-PEX by RT-PCR, they were detected after further purification: differential precipitation with 2-butoxyethanol and HCl treatment followed by ethanol-precipitation. In addition, PCR in the presence of tetramethylammonium chloride specifically amplified the cDNA of all five distinct CVds under the same temperature and cycle conditions. Since all the viroids could be detected in NA liberated by PEX, the amount of NA extracted by the method described here is sufficient for detecting viroids, enabling the processing of a large number of samples.

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

    PubMed

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

    2013-02-05

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

  3. Identification of Bacillus strains isolated from milk and cream with classical and nucleic acid hybridization methods.

    PubMed

    Tatzel, R; Ludwig, W; Schleifer, K H; Wallnöfer, P R

    1994-11-01

    A total of 529 bacterial strains have been isolated from milk and cream sampled at different sites in a dairy production plant under conditions selective for aerobic sporeforming bacteria. Identification with classical methods based on morphological, physiological and biochemical criteria showed Bacillus licheniformis to be the most frequently occurring Bacillus sp. The investigation also revealed 62 unidentified strains. Classical identification methods were time consuming (3-7 d), lacked specificity and--because of their dependence on phenotypic gene expression--sometimes produced ambiguous results. Consequently, a colony hybridization method developed for the identification of B. licheniformis strains and using nonradioactive labelled 23S rRNA targeted oligonucleotide probes was also used. Identification of B. licheniformis with both classical and hybridization methods revealed diverging identification results for 70 strains.

  4. Hybrid poplar grows poorly on acid spoil banks at high elevations in West Virginia

    Treesearch

    George R., Jr. Trimble

    1963-01-01

    In the early 1950s, a region-wide series of hybrid poplar clonal tests was begun in the Northeast to evaluate the performance of 50 selected clones under a variety of site and climatic conditions. The basic test unit was a block of 50 randomized plots-1 plot for each of the 50 clones. In each plot, 16 cuttings were planted at 4-foot spacing.

  5. Structure and DNA Hybridization Properties of Mixed Nucleic Acid/Maleimide-ethylene glycol Monolayers

    PubMed Central

    Lee, Chi-Ying; Nguyen, Phuong-Cac T.; Grainger, David W.; Gamble, Lara J.; Castner, David G.

    2008-01-01

    The surface structure and DNA hybridization performance of thiolated single-strand DNA (HS-ssDNA) covalently attached to a maleimide-ethylene glycol disulfide (MEG) monolayer on gold have been investigated. Monolayer immobilization chemistry and surface coverage of reactive ssDNA probes were studied by X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS). Orientation of the ssDNA probes was determined by near edge X-ray absorption fine structure (NEXAFS). Target DNA hybridization on the DNA-MEG probe surfaces was measured by surface plasmon resonance (SPR) to demonstrate the utility of these probe surfaces for detection of DNA targets from both purified target DNA samples and complex biological mixtures such as blood serum. Data from complementary techniques showed that immobilized ssDNA density is strongly dependent on the spotted bulk DNA concentration and buffer ionic strength. Variation of the immobilized ssDNA density had a profound influence on the DNA probe orientation at the surface and subsequent target hybridization efficiency. With increasing surface probe density, NEXAFS polarization dependence results (followed by monitoring the N 1s → π* transition) indicate that the immobilized ssDNA molecules reorient towards a more upright position on the MEG monolayer. SPR assays of DNA targets from buffer and serum showed that DNA hybridization efficiency increased with decreasing surface probe density. However, target detection in serum was better on the “high density” probe surface than on the “high efficiency” probe surface. The amount of target detected for both ssDNA surfaces were several orders of magnitude poorer in serum than in purified DNA samples due to non-specific serum protein adsorption onto the sensing surface. PMID:17492838

  6. Development and Application of Nucleic Acid Hybridization Techniques to Arbovirus Surveillance and Diagnosis.

    DTIC Science & Technology

    1987-02-27

    Department of Microbiology and Enviromental Health College of Veterinary Medicine and Biomedical Sciences Colorado State University Fort Collins, C) 80523...1983). Detection of viral genomes in cultured cells and paraffin-embedded tissue sections using biotin-labeled hybridization probes. Virology 126: 32-50...sequence of the M RIA of snowshoe hare burzavirus reveals the presenoe of internal hydrophoic domains in the viral glycoprotein. Virology 137: 227-240. 15

  7. Ampholine-functionalized hybrid organic-inorganic silica material as sorbent for solid-phase extraction of acidic and basic compounds.

    PubMed

    Wang, Tingting; Chen, Yihui; Ma, Junfeng; Chen, Mingliang; Nie, Chenggang; Hu, Minjie; Li, Ying; Jia, Zhijian; Fang, Jianghua; Gao, Haoqi

    2013-09-20

    A novel sorbent for solid-phase extraction (SPE) was synthesized by chemical immobilization of ampholine on hybrid organic-inorganic silica material. The ampholine-functionalized hybrid organic-inorganic silica sorbent is consisted of aliphatic amine groups, carboxyl groups and long carbon chains, allowing for extraction of both acidic and basic compounds. The retention properties of the developed sorbent were evaluated for 1-hydroxy-2-naphthoic acid (HNA), 1-naphthoic acid (NA), 3-hydroxybenzoic acid (HBA), benzoic acid (BA), sorbic acid (SA), vanillic aldehyde (VA), butyl 4-hydroxybenzoate (BHB), propyl 4-hydroxybenzoate (PHB), ethyl 4-hydroxybenzoate (EHB), and methyl 4-hydroxybenzoate (MHB). The results show that such a sorbent has three types of interaction, i.e., electrostatic interaction, hydrophobic interaction, and hydrogen bonding, exhibiting high extraction efficiency towards the compounds tested. The adsorption capacities of the analytes ranged from 0.61 to 6.54μgmg(-1). The reproducibility of the sorbent preparation was evaluated at three spiking concentration levels, with relative standard deviations (RSDs) of 1.0-10.5%. The recoveries of ten acidic and basic compounds spiked in beverage Coca-Cola(®) sample ranged from 82.5% to 98.2% with RSDs less than 5.8%. Under optimum conditions, the ampholine-functionalized hybrid organic-inorganic silica sorbent rendered higher extraction efficiency for acidic compounds than that of the commercially available ampholine-functionalized silica particles, and was comparable to that of the commercial Oasis WAX and Oasis WCX.

  8. Self-assembly of nucleic acids, silk and hybrid materials thereof.

    PubMed

    Humenik, Martin; Scheibel, Thomas

    2014-12-17

    Top-down approaches based on etching techniques have almost reached their limits in terms of dimension. Therefore, novel assembly strategies and types of nanomaterials are required to allow technological advances. Self-assembly processes independent of external energy sources and unlimited in dimensional scaling have become a very promising approach. Here,we highlight recent developments in self-assembled DNA-polymer, silk-polymer and silk-DNA hybrids as promising materials with biotic and abiotic moieties for constructing complex hierarchical materials in ‘bottom-up’ approaches. DNA block copolymers assemble into nanostructures typically exposing a DNA corona which allows functionalization, labeling and higher levels of organization due to its specific addressable recognition properties. In contrast, self-assembly of natural silk proteins as well as their recombinant variants yields mechanically stable β-sheet rich nanostructures. The combination of silk with abiotic polymers gains hybrid materials with new functionalities. Together, the precision of DNA hybridization and robustness of silk fibrillar structures combine in novel conjugates enable processing of higher-order structures with nanoscale architecture and programmable functions.

  9. Self-assembly of nucleic acids, silk and hybrid materials thereof

    NASA Astrophysics Data System (ADS)

    Humenik, Martin; Scheibel, Thomas

    2014-12-01

    Top-down approaches based on etching techniques have almost reached their limits in terms of dimension. Therefore, novel assembly strategies and types of nanomaterials are required to allow technological advances. Self-assembly processes independent of external energy sources and unlimited in dimensional scaling have become a very promising approach. Here, we highlight recent developments in self-assembled DNA-polymer, silk-polymer and silk-DNA hybrids as promising materials with biotic and abiotic moieties for constructing complex hierarchical materials in ‘bottom-up’ approaches. DNA block copolymers assemble into nanostructures typically exposing a DNA corona which allows functionalization, labeling and higher levels of organization due to its specific addressable recognition properties. In contrast, self-assembly of natural silk proteins as well as their recombinant variants yields mechanically stable β-sheet rich nanostructures. The combination of silk with abiotic polymers gains hybrid materials with new functionalities. Together, the precision of DNA hybridization and robustness of silk fibrillar structures combine in novel conjugates enable processing of higher-order structures with nanoscale architecture and programmable functions.

  10. Mitochondrial proteomics of the acetic acid - induced programmed cell death response in a highly tolerant Zygosaccharomyces bailii - derived hybrid strain

    PubMed Central

    Guerreiro, Joana F.; Sampaio-Marques, Belém; Soares, Renata; Coelho, Ana V.; Leão, Cecília; Ludovico, Paula; Sá-Correia, Isabel

    2016-01-01

    Very high concentrations of acetic acid at low pH induce programmed cell death (PCD) in both the experimental model Saccharomyces cerevisiae and in Zygosaccharomyces bailii, the latter being considered the most problematic acidic food spoilage yeast due to its remarkable intrinsic resistance to this food preservative. However, while the mechanisms underlying S. cerevisiae PCD induced by acetic acid have been previously examined, the corresponding molecular players remain largely unknown in Z. bailii. Also, the reason why acetic acid concentrations known to be necrotic for S. cerevisiae induce PCD with an apoptotic phenotype in Z. bailii remains to be elucidated. In this study, a 2-DE-based expression mitochondrial proteomic analysis was explored to obtain new insights into the mechanisms involved in PCD in the Z. bailii derived hybrid strain ISA1307. This allowed the quantitative assessment of expression of protein species derived from each of the parental strains, with special emphasis on the processes taking place in the mitochondria known to play a key role in acetic acid - induced PCD. A marked decrease in the content of proteins involved in mitochondrial metabolism, in particular, in respiratory metabolism (Cor1, Rip1, Lpd1, Lat1 and Pdb1), with a concomitant increase in the abundance of proteins involved in fermentation (Pdc1, Ald4, Dld3) was registered. Other differentially expressed identified proteins also suggest the involvement of the oxidative stress response, protein translation, amino acid and nucleotide metabolism, among other processes, in the PCD response. Overall, the results strengthen the emerging concept of the importance of metabolic regulation of yeast PCD. PMID:28357336

  11. Alkyl substituent effects on gas-phase acidities - The influence of hybridization.

    NASA Technical Reports Server (NTRS)

    Brauman, J. I.; Blair, L. K.

    1971-01-01

    Exploration of the effect on acidity of alkyl groups bonded to trigonal and digonal carbon. Some results on the relative acidities of toluene and p-xylene, and acetylene and substitute acetylenes, as determined by ion cyclotron resonance (icr) spectroscopy, are described. Some limitations of the CNDO/2 calculation method are discussed.

  12. Application of a hybrid CFD-PBPK nasal dosimetry model in an inhalation risk assessment: an example with acrylic acid.

    PubMed

    Andersen, M; Sarangapani, R; Gentry, R; Clewell, H; Covington, T; Frederick, C B

    2000-10-01

    The available inhalation toxicity information for acrylic acid (AA) suggests that lesions to the nasal cavity, specifically olfactory degeneration, are the most sensitive end point for developing a reference concentration (RfC). Advances in physiologically based pharmacokinetic (PBPK) modeling, specifically the incorporation of computational fluid dynamic (CFD) models, now make it possible to estimate the flux of inhaled chemicals within the nasal cavity of experimental species, specifically rats. The focus of this investigation was to apply an existing CFD-PBPK hybrid model in the estimation of an RfC to determine the impact of incorporation of this new modeling technique into the risk assessment process. Information provided in the literature on the toxicity and mode of action for AA was used to determine the risk assessment approach. A comparison of the approach used for the current U.S. Environmental Protection Agency (U.S. EPA) RfC with the approach using the CFD-PBPK hybrid model was also conducted. The application of the CFD-PBPK hybrid model in a risk assessment for AA resulted in an RfC of 79 ppb, assuming a minute ventilation of 13.8 l/min (20 m(3)/day) in humans. This value differs substantially from the RfC of 0.37 ppb estimated for AA by the U.S. EPA before the PBPK modeling advances became available. The difference in these two RfCs arises from many factors, with the main difference being the species selected (mouse vs. rat). The choice to conduct the evaluation using the rat was based on the availability of dosimetry data in this species. Once these data are available in the mouse, an assessment should be conducted using this information. Additional differences included the methods used for estimating the target tissue concentration, the uncertainty factors (UFs) applied, and the application of duration and uncertainty adjustments to the internal target tissue dose rather than the external exposure concentration.

  13. Rapid identification, by use of the LTQ Orbitrap hybrid FT mass spectrometer, of antifungal compounds produced by lactic acid bacteria.

    PubMed

    Brosnan, Brid; Coffey, Aidan; Arendt, Elke K; Furey, Ambrose

    2012-07-01

    Fungal contamination of food causes health and economic concerns. Several species of lactic acid bacteria (LAB) have antifungal activity which may inhibit food spoilage fungi. LAB have GRAS (generally recognised as safe) status, allowing them to be safely integrated into food systems as natural food preservatives. A method is described herein that enables rapid screening of LAB cultures for 25 known antifungal compounds associated with LAB. This is the first chromatographic method developed which enables the rapid identification of a wide range of antifungal compounds by a single method with a short analysis time (23 min). Chromatographic separation was achieved on a Phenomenex Gemini C18 100A column (150 mm × 2.0 mm; 5 μm) by use of a mobile-phase gradient prepared from (A) water containing acetic acid (0.1%) and (B) acetonitrile containing acetic acid (0.1%), at a flow rate of 0.3 µL min(-1). The gradient involved a progressive ramp from 10-95% acetonitrile over 13 min. The LC was coupled to a hybrid LTQ Orbitrap XL fourier-transform mass spectrometer (FTMS) operated in negative ionisation mode. High mass accuracy data (<3 ppm) obtained by use of high resolution (30,000 K) enabled unequivocal identification of the target compounds. This method allows comprehensive profiling and comparison of different LAB strains and is also capable of the identification of additional compounds produced by these bacteria.

  14. A strategy for dual inhibition of the proteasome and fatty acid synthase with belactosin C-orlistat hybrids.

    PubMed

    Zhu, Mingzhao; Harshbarger, Wayne D; Robles, Omar; Krysiak, Joanna; Hull, Kenneth G; Cho, Sung Wook; Richardson, Robyn D; Yang, Yanyan; Garcia, Andres; Spiegelman, Lindsey; Ramirez, Bianca; Wilson, Christopher T; Yau, Ju Anne; Moore, James T; Walker, Caitlen B; Sacchettini, James C; Liu, Wenshe R; Sieber, Stephan A; Smith, Jeffrey W; Romo, Daniel

    2017-06-01

    The proteasome, a validated cellular target for cancer, is central for maintaining cellular homeostasis, while fatty acid synthase (FAS), a novel target for numerous cancers, is responsible for palmitic acid biosynthesis. Perturbation of either enzymatic machine results in decreased proliferation and ultimately cellular apoptosis. Based on structural similarities, we hypothesized that hybrid molecules of belactosin C, a known proteasome inhibitor, and orlistat, a known inhibitor of the thioesterase domain of FAS, could inhibit both enzymes. Herein, we describe proof-of-principle studies leading to the design, synthesis and enzymatic activity of several novel, β-lactone-based, dual inhibitors of these two enzymes. Validation of dual enzyme targeting through activity-based proteome profiling with an alkyne probe modeled after the most potent inhibitor, and preliminary serum stability studies of selected derivatives are also described. These results provide proof of concept for dual targeting of the proteasome and fatty acid synthase-thioesterase (FAS-TE) enabling a new approach for the development of drug-candidates with potential to overcome resistance. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Multidimensional Separation of Chiral Amino Acid Mixtures in a Multilayered Three Dimensional Hybrid Microfluidic/Nanofluidic Device

    PubMed Central

    Kim, Bo Young; Yang, Jing; Gong, Maojun; Flachsbart, Bruce R.; Shannon, Mark A.; Bohn, Paul W.; Sweedler, Jonathan V.

    2009-01-01

    Microscale total analysis systems (μTAS) allow high-throughput analyses by integrating multiple processes, parallelization and automation. Here we combine unit operations of μTAS to create a device that can perform multidimensional separations using a three-dimensional hybrid microfluidic/nanofluidic device composed of alternating layers of patterned polymethylmethacrylate and nanocapillary array membranes constructed from nuclear track-etched polycarbonate. Two consecutive electrophoretic separations are performed, the first being an achiral separation followed by a chiral separation of a selected analyte band. Separation conditions are optimized for a racemic mixture of fluorescein-labeled amino acids, serine and aspartic acid, chosen because there are endogenous D-forms of these amino acids in animals. The chiral separation is implemented using micellar electrokinetic chromatography using β–cyclodextrin as the chiral selector and sodium taurocholate as the micelle forming agent. Analyte separation is monitored by dual-beam laser-induced fluorescence detection. After separation in the first electrophoretic channel, the preselected analyte is sampled by the second-stage separation using an automated collection sequence with a zero-crossing algorithm for peak identification. The controlled fluidic environment inherent to the three-dimensional architecture enables a series of separations in varying fluidic environments and allows sample stacking via different background electrolyte pH conditions. The ability to interface sequential separations, selected analyte capture and other fluidic manipulations in the third dimension significantly improves the functionality of multilayer microfluidic devices. PMID:19271741

  16. Programmable release of multiple protein drugs from aptamer-functionalized hydrogels via nucleic acid hybridization.

    PubMed

    Battig, Mark R; Soontornworajit, Boonchoy; Wang, Yong

    2012-08-01

    Polymeric delivery systems have been extensively studied to achieve localized and controlled release of protein drugs. However, it is still challenging to control the release of multiple protein drugs in distinct stages according to the progress of disease or treatment. This study successfully demonstrates that multiple protein drugs can be released from aptamer-functionalized hydrogels with adjustable release rates at predetermined time points using complementary sequences (CSs) as biomolecular triggers. Because both aptamer-protein interactions and aptamer-CS hybridization are sequence-specific, aptamer-functionalized hydrogels constitute a promising polymeric delivery system for the programmable release of multiple protein drugs to treat complex human diseases.

  17. Peptide nucleic acid fluorescence in situ hybridization for identification of Listeria genus, Listeria monocytogenes and Listeria ivanovii.

    PubMed

    Zhang, Xiaofeng; Wu, Shan; Li, Ke; Shuai, Jiangbing; Dong, Qiang; Fang, Weihuan

    2012-07-02

    A fluorescent in situ hybridization (FISH) method in conjunction with fluorescin-labeled peptide nucleic acid (PNA) probes (PNA-FISH) for detection of Listeria species was developed. In silico analysis showed that three PNA probes Lis-16S-1, Lm-16S-2 and Liv-16S-5 were suitable for specific identification of Listeria genus, Listeria monocytogenes and Listeria ivanovii, respectively. These probes were experimentally verified by their reactivity against 19 strains of six Listeria species (excluding newly described species Listeria marthii and Listeria rocourtiae) and eight other bacterial species. The PNA-FISH method was optimized as 30 min of hybridization with 0.2% Triton X-100 in the solution and used to identify 85 Listeria strains from individual putative Listeria colonies on PALCAM agar plates streaked from selectively enriched cultures of 780 food or food-related samples. Of the 85 Listeria strains, thirty-seven were identified as L. monocytogenes with the probe Lm-16S-2 and two as L. ivanovii with the probe Liv-16S-5 which was in agreement with the results obtained by the API LISTERIA method. Thus, the PNA-FISH protocol has the potential for identification of pathogenic Listeria spp. from food or food-related samples.

  18. Disaggregation of Amylin Aggregate by Novel Conformationally Restricted Aminobenzoic Acid containing α/β and α/γ Hybrid Peptidomimetics

    NASA Astrophysics Data System (ADS)

    Paul, Ashim; Kalita, Sourav; Kalita, Sujan; Sukumar, Piruthivi; Mandal, Bhubaneswar

    2017-01-01

    Diabetes has emerged as a threat to the current world. More than ninety five per cent of all the diabetic population has type 2 diabetes mellitus (T2DM). Aggregates of Amylin hormone, which is co-secreted with insulin from the pancreatic β-cells, inhibit the activities of insulin and glucagon and cause T2DM. Importance of the conformationally restricted peptides for drug design against T2DM has been invigorated by recent FDA approval of Symlin, which is a large conformationally restricted peptide. However, Symlin still has some issues including solubility, oral bioavailability and cost of preparation. Herein, we introduced a novel strategy for conformationally restricted peptide design adopting a minimalistic approach for cost reduction. We have demonstrated efficient inhibition of amyloid formation of Amylin and its disruption by a novel class of conformationally restricted β-sheet breaker hybrid peptidomimetics (BSBHps). We have inserted β, γ and δ -aminobenzoic acid separately into an amyloidogenic peptide sequence, synthesized α/β, α/γ and α/δ hybrid peptidomimetics, respectively. Interestingly, we observed the aggregation inhibitory efficacy of α/β and α/γ BSBHps, but not of α/δ analogues. They also disrupt existing amyloids into non-toxic forms. Results may be useful for newer drug design against T2DM as well as other amyloidoses and understanding amyloidogenesis.

  19. Modeling of the cranking and charging processes of conventional valve regulated lead acid (VRLA) batteries in micro-hybrid applications

    NASA Astrophysics Data System (ADS)

    Gou, Jun; Lee, Anson; Pyko, Jan

    2014-10-01

    The cranking and charging processes of a VRLA battery during stop-start cycling in micro-hybrid applications were simulated by one dimensional mathematical modeling, to study the formation and distribution of lead sulfate across the cell and analyze the resulting effect on battery aging. The battery focused on in this study represents a conventional VRLA battery without any carbon additives in the electrodes or carbon-based electrodes. The modeling results were validated against experimental data and used to analyze the "sulfation" of negative electrodes - the common failure mode of lead acid batteries under high-rate partial state of charge (HRPSoC) cycling. The analyses were based on two aging mechanisms proposed in previous studies and the predictions showed consistency with the previous teardown observations that the sulfate formed at the negative interface is more difficult to be converted back than anywhere else in the electrodes. The impact of cranking pulses during stop-start cycling on current density and the corresponding sulfate layer production was estimated. The effects of some critical design parameters on sulfate formation, distribution and aging over cycling were investigated, which provided guidelines for developing models and designing of VRLA batteries in micro-hybrid applications.

  20. Electrostatic nucleic acid nanoassembly enables hybridization chain reaction in living cells for ultrasensitive mRNA imaging.

    PubMed

    Wu, Zhan; Liu, Gao-Qin; Yang, Xiao-Li; Jiang, Jian-Hui

    2015-06-03

    Efficient approaches for intracellular delivery of nucleic acid reagents to achieve sensitive detection and regulation of gene and protein expressions are essential for chemistry and biology. We develop a novel electrostatic DNA nanoassembly that, for the first time, realizes hybridization chain reaction (HCR), a target-initiated alternating hybridization reaction between two hairpin probes, for signal amplification in living cells. The DNA nanoassembly has a designed structure with a core gold nanoparticle, a cationic peptide interlayer, and an electrostatically assembled outer layer of fluorophore-labeled hairpin DNA probes. It is shown to have high efficiency for cellular delivery of DNA probes via a unique endocytosis-independent mechanism that confers a significant advantage of overcoming endosomal entrapment. Moreover, electrostatic assembly of DNA probes enables target-initialized release of the probes from the nanoassembly via HCR. This intracellular HCR offers efficient signal amplification and enables ultrasensitive fluorescence activation imaging of mRNA expression with a picomolar detection limit. The results imply that the developed nanoassembly may provide an invaluable platform in low-abundance biomarker discovery and regulation for cell biology and theranostics.

  1. Toward a multiplexed solid-phase nucleic acid hybridization assay using quantum dots as donors in fluorescence resonance energy transfer.

    PubMed

    Algar, W Russ; Krull, Ulrich J

    2009-05-15

    Solid-phase assays using immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) have been developed for the selective detection of nucleic acids. QDs were immobilized on optical fibers and conjugated with probe oligonucleotides. Hybridization with acceptor labeled target oligonucleotides generated FRET-sensitized acceptor fluorescence that was used as the analytical signal. A sandwich assay was also introduced and avoided the need for target labeling. Green and red emitting CdSe/ZnS QDs were used as donors with Cy3 and Alexa Fluor 647 acceptors, respectively. Quantitative measurements were made via spectrofluorimetry or fluorescence microscopy. Detection limits as low as 1 nM were obtained, and the discrimination of single nucleotide polymorphisms (SNPs) with contrast ratios as high as 31:1 was possible. The assays retained their selectivity and at least 50% of their signal when tested in bovine serum and against a large background of noncomplementary genomic DNA. Mixed films of the two colors of QD and two probe oligonucleotide sequences were prepared for multiplexed solid-phase hybridization assays. It was possible to simultaneously detect two target sequences with retention of selectivity, including SNP discrimination. This research provides an important precedent and framework for the future development of QD-based bioassays and biosensors.

  2. Luminescent molecular hybrid system derived from 2-furancarboxylic acid and silylated monomer coordinated to rare earth ions

    NASA Astrophysics Data System (ADS)

    Sui, Yu-Long; Yan, Bing

    2006-04-01

    In this study, silica-based organic-inorganic hybrids were prepared by the sol-gel method. Tetraethoxysilane (abbreviated as TEOS) and a kind of monomer (abbreviated as FA-APES) derived from modified 2-furancarboxylic acid (abbreviated as FA) with (3-aminopropyl)triethoxysilane (abbreviated as APES) were used as the inorganic and organic fragments, respectively. Coordination reaction between lanthanides (europium and terbium ions) and sbnd C dbnd O group of the monomer happened simultaneously. And after days of aging process the resultant materials showed characteristic luminescence of lanthanides. The enhancement of luminescence can be seen by the comparison with simply doped lanthanide hybrid systems. And it can be explained by the coordination ability of the organic counterpart. IR, NMR, UV-vis absorption, low-temperature phosphorescence spectroscopy and fluorescence spectroscopy were applied to characterize and the above spectroscopic data revealed that the triplet state energy of organic ligand matches with the emissive energy level of lanthanides (especially of Tb 3+).

  3. Disaggregation of Amylin Aggregate by Novel Conformationally Restricted Aminobenzoic Acid containing α/β and α/γ Hybrid Peptidomimetics

    PubMed Central

    Paul, Ashim; Kalita, Sourav; Kalita, Sujan; Sukumar, Piruthivi; Mandal, Bhubaneswar

    2017-01-01

    Diabetes has emerged as a threat to the current world. More than ninety five per cent of all the diabetic population has type 2 diabetes mellitus (T2DM). Aggregates of Amylin hormone, which is co-secreted with insulin from the pancreatic β-cells, inhibit the activities of insulin and glucagon and cause T2DM. Importance of the conformationally restricted peptides for drug design against T2DM has been invigorated by recent FDA approval of Symlin, which is a large conformationally restricted peptide. However, Symlin still has some issues including solubility, oral bioavailability and cost of preparation. Herein, we introduced a novel strategy for conformationally restricted peptide design adopting a minimalistic approach for cost reduction. We have demonstrated efficient inhibition of amyloid formation of Amylin and its disruption by a novel class of conformationally restricted β-sheet breaker hybrid peptidomimetics (BSBHps). We have inserted β, γ and δ -aminobenzoic acid separately into an amyloidogenic peptide sequence, synthesized α/β, α/γ and α/δ hybrid peptidomimetics, respectively. Interestingly, we observed the aggregation inhibitory efficacy of α/β and α/γ BSBHps, but not of α/δ analogues. They also disrupt existing amyloids into non-toxic forms. Results may be useful for newer drug design against T2DM as well as other amyloidoses and understanding amyloidogenesis. PMID:28054630

  4. Hybrid therapy with locoregional steroid injection and polyglycolic acid sheets to prevent stricture after esophageal endoscopic submucosal dissection

    PubMed Central

    Nagami, Yasuaki; Shiba, Masatsugu; Tominaga, Kazunari; Ominami, Masaki; Fukunaga, Shusei; Sugimori, Satoshi; Tanaka, Fumio; Kamata, Noriko; Tanigawa, Tetsuya; Yamagami, Hirokazu; Watanabe, Toshio; Fujiwara, Yasuhiro; Arakawa, Tetsuo

    2016-01-01

    Background and study aim: The incidence of stricture formation caused by endoscopic submucosal dissection (ESD) for widespread lesions is high, and stricture formation can reduce quality of life. We evaluated the prophylactic efficacy of hybrid therapy using a locoregional steroid injection and polyglycolic acid (PGA) sheets with fibrin glue to prevent stricture formation after esophageal ESD in high risk patients in whom we predicted stricture formation would be difficult to prevent with a single prophylactic steroid injection. Methods: Ten patients who underwent esophageal ESD were enrolled (entire-circumference: n = 6; sub-circumference, more than 5/6 of the circumference: n = 4). A single locoregional steroid injection and PGA sheets with fibrin glue were used after ESD. We evaluated the incidence of stricture formation, the number of endoscopic balloon dilation (EBD) procedures needed to treat the stricture formation, and adverse events of the therapy. Results: Esophageal stricture formation occurred in 50.0 % of patients (5/10) (median EBD sessions 0.5, range 0 – 16). Subanalysis showed that stricture formation occurred in 37.5 % of patients (3/8) excluded the lesions located near a previous scar from ESD or surgical anastomosis site (median EBD sessions 0, range 0 – 4). Conclusion: Hybrid therapy using a locoregional steroid injection and PGA sheets with fibrin glue may have the potential to prevent esophageal stricture formation after esophageal ESD in high risk patients. PMID:27652294

  5. Fluorescence In Situ Hybridization with Peptide Nucleic Acid Probes for Rapid Identification of Candida albicans Directly from Blood Culture Bottles

    PubMed Central

    Rigby, Susan; Procop, Gary W.; Haase, Gerhard; Wilson, Deborah; Hall, Geraldine; Kurtzman, Cletus; Oliveira, Kenneth; Von Oy, Sabina; Hyldig-Nielsen, Jens J.; Coull, James; Stender, Henrik

    2002-01-01

    A new fluorescence in situ hybridization (FISH) method that uses peptide nucleic acid (PNA) probes for identification of Candida albicans directly from positive-blood-culture bottles in which yeast was observed by Gram staining (herein referred to as yeast-positive blood culture bottles) is described. The test (the C. albicans PNA FISH method) is based on a fluorescein-labeled PNA probe that targets C. albicans 26S rRNA. The PNA probe is added to smears made directly from the contents of the blood culture bottle and hybridized for 90 min at 55°C. Unhybridized PNA probe is removed by washing of the mixture (30 min), and the smears are examined by fluorescence microscopy. The specificity of the method was confirmed with 23 reference strains representing phylogenetically related yeast species and 148 clinical isolates covering the clinically most significant yeast species, including C. albicans (n = 72), C. dubliniensis (n = 58), C. glabrata (n = 5), C. krusei (n = 2), C. parapsilosis (n = 4), and C. tropicalis (n = 3). The performance of the C. albicans PNA FISH method as a diagnostic test was evaluated with 33 routine and 25 simulated yeast-positive blood culture bottles and showed 100% sensitivity and 100% specificity. It is concluded that this 2.5-h method for the definitive identification of C. albicans directly from yeast-positive blood culture bottles provides important information for optimal antifungal therapy and patient management. PMID:12037084

  6. Synthesis, Aqueous Reactivity, and Biological Evaluation of Carboxylic Acid Ester-Functionalized Platinum–Acridine Hybrid Anticancer Agents

    PubMed Central

    Graham, Leigh A.; Suryadi, Jimmy; West, Tiffany K.; Kucera, Gregory L.; Bierbach, Ulrich

    2012-01-01

    The synthesis of platinum–acridine hybrid agents containing carboxylic acid ester groups is described. The most active derivatives and the unmodified parent compounds showed up to 6-fold higher activity in ovarian cancer (OVCAR-3) and breast cancer (MCF-7, MDA-MB-23) cell lines than cisplatin. Inhibition of cell proliferation at nanomolar concentrations was observed in pancreatic (PANC-1) and non-small cell lung cancer cells (NSCLC, NCI-H460) of 80- and 150-fold, respectively. Introduction of the ester groups did not affect the cytotoxic properties of the hybrids, which form the same monofunctional–intercalative DNA adducts as the parent compounds, as demonstrated in a plasmid unwinding assay. In-line high-performance liquid chromatography and electrospray mass spectrometry (LC-ESMS) shows that the ester moieties undergo platinum-mediated hydrolysis in a chloride concentration-dependent manner to form carboxylate chelates. Potential applications of the chloride-sensitive ester hydrolysis as a self-immolative release mechanism for tumor-selective delivery of platinum–acridines are discussed. PMID:22871158

  7. Peptide nucleic acid fluorescence in-situ hybridization for identification of Vibrio spp. in aquatic products and environments.

    PubMed

    Zhang, Xiaofeng; Li, Ke; Wu, Shan; Shuai, Jiangbing; Fang, Weihuan

    2015-08-03

    A peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) method was developed for specific detection of the Vibrio genus. In silico analysis by BLAST and ProbeCheck showed that the designed PNA probe targeting the 16S rRNAs was suitable for specific identification of Vibrio. Specificity and sensitivity of the probe Vib-16S-1 were experimentally verified by its reactivity against 18 strains of 9 Vibrio species and 14 non-Vibrio strains of 14 representative species. The PNA-FISH assay was able to identify 47 Vibrio positive samples from selectively enriched cultures of 510 samples of aquatic products and environments, comparable with the results obtained by biochemical identification and real-time PCR. We conclude that PNA-FISH can be an alternative method for rapid identification of Vibrio species in a broad spectrum of seafood or related samples.

  8. Comparing hybrid hyaluronic acid with PRP in end career athletes with degenerative cartilage lesions of the knee.

    PubMed

    Papalia, R; Zampogna, B; Russo, F; Vasta, S; Tirindelli, M C; Nobile, C; Di Martino, A C; Vadalà, G; Denaro, V

    2016-01-01

    Cartilage lesions are very common causes of chronic knee pain in athletes. Current treatment options consist in conservative strategies, such as viscosupplementation and platelet-rich plasma injections. This randomized controlled trial aims to investigate the effect of intra-articular Hybrid Hyaluronic Acid injections compared to PRP for the treatment of cartilage lesions among athletes at the end of their career. Since March 2015, 48 professional soccer players were randomized into two groups: 24 patients received 3 injections of HHA and 23 patients received 3 intra-articular injections of PRP. All patients achieved a statistically significant clinical improvement from preoperative to postoperative time in both groups. Patients in the HHA group showed a significant superiority compared to PRP group at 3 and 6 months. Intergroup differences decrease gradually until loss of significance at 12 months follow-up. Athletes with chronic degenerative cartilage lesions of the knee responded positively both to HHA and PRP until last follow up.

  9. Hydrofluoric-nitric-sulphuric-acid surface treatment of tungsten for carbon fibre-reinforced composite hybrids in space applications

    NASA Astrophysics Data System (ADS)

    Kanerva, M.; Johansson, L.-S.; Campbell, J. M.; Revitzer, H.; Sarlin, E.; Brander, T.; Saarela, O.

    2015-02-01

    Hybrid material systems, such as combinations of tungsten foils and carbon fibre-reinforced plastic (CFRP), are replacing metal alloy concepts in spacecraft enclosures. However, a good adhesion between the tungsten oxide scale and the epoxy resin used is required. Here, the effects of a hydrofluoric-nitric-sulphuric-acid (HFNS) treatment on tungsten oxides and subsequent adhesion to CFRP are analysed using atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS) and fracture testing. The work shows that HFNS treatment results in decreased oxygen content, over 50% thinner tungsten trioxide (WO3) layer and increased nano-roughness on thin tungsten foils. Fracture testing established a 39% increase in the average critical strain for tungsten-CFRP specimens after HFNS treatment was carried out on tungsten. The effect of the oxide scale modification regarding the critical strain energy release rate was ΔGc≈ 8.4 J/m2.

  10. Structural and optical study of spin-coated camphorsulfonic acid-doped polyaniline/titanium-di-oxide nanoparticles hybrid thin films

    NASA Astrophysics Data System (ADS)

    Geethalakshmi, D.; Muthukumarasamy, N.; Balasundaraprabhu, R.

    2015-06-01

    Polyaniline (PANI) doped with Camphorsulfonic acid (CSA) has been prepared by chemical oxidative polymerization and blend with titanium-di-oxide (TiO2) nanoparticles prepared by sol-gel method to form CSA-doped PANI/TiO2 hybrid thin films. The properties of as-deposited and heat-treated (100 °C) hybrid thin films having different PANI:TiO2 weight ratios (1:0.5, 1:1, and 1:2) have been compared. FTIR study indicated that chemical bonding between CSA-doped PANI and TiO2 has been formed. XRD studies reveal that the as-deposited hybrid thin films are of amorphous nature and heat-treatment of such films initiates crystallization. SEM study shows that as-deposited hybrid films are rough; increase in TiO2 ratio and heat-treatment increased the roughness due to coalescing and agglomeration. UV-visible absorbance of hybrid films shows its characteristic peak in the visible region along with a peak in UV range and its intensity increased with TiO2 ratio and heat-treatment due to agglomeration of TiO2 particles. Photoluminescence spectra revealed that emission occurs in visible region (495 nm) for as-deposited hybrid thin film and this emission increased with TiO2 ratio and heat-treatment of hybrid films.

  11. Electrostatic surface plasmon resonance: Direct electric field-induced hybridization and denaturation in monolayer nucleic acid films and label-free discrimination of base mismatches

    PubMed Central

    Heaton, Richard J.; Peterson, Alexander W.; Georgiadis, Rosina M.

    2001-01-01

    We demonstrate that in situ optical surface plasmon resonance spectroscopy can be used to monitor hybridization kinetics for unlabeled DNA in tethered monolayer nucleic acid films on gold in the presence of an applied electrostatic field. The dc field can enhance or retard hybridization and can also denature surface-immobilized DNA duplexes. Discrimination between matched and mismatched hybrids is achieved by simple adjustment of the electrode potential. Although the electric field at the interface is extremely large, the tethered single-stranded DNA thiol probes remain bound and can be reused for subsequent hybridization reactions without loss of efficiency. Only capacitive charging currents are drawn; redox reactions are avoided by maintaining the gold electrode potential within the ideally polarizable region. Because of potential-induced changes in the shape of the surface plasmon resonance curve, we account for the full curve rather than simply the shift in the resonance minimum. PMID:11259682

  12. Babesia gibsoni: detection in blood smears and formalin-fixed, paraffin-embedded tissues using deoxyribonucleic acid in situ hybridization analysis.

    PubMed

    Yamasaki, Masahiro; Kobayashi, Yusuke; Nakamura, Kensuke; Sasaki, Noboru; Murakami, Masahiro; Rajapakshage, Bandula Kumara Wickramasekara; Ohta, Hiroshi; Yamato, Osamu; Maede, Yoshimitsu; Takiguchi, Mitsuyoshi

    2011-01-01

    In this study, we attempted to detect Babesia gibsoni in blood smears and formalin-fixed, paraffin-embedded tissues obtained from B. gibsoni-infected dogs using in situ hybridization. Using a digoxigenin-conjugated deoxyribonucleic acid (DNA) probe, both intraerythrocytic and exoerythrocytic parasites in the culture could be specifically stained in blood smears fixed with 4% phosphate-buffered paraformaldehyde. This indicated that genomic DNA extracted from the parasites could be detected using in situ hybridization. Moreover, the parasite could be specifically stained in paraffin-embedded spleen, lymph node, and kidney sections using in situ hybridization. Infected erythrocytes in blood vessels in the spleen and kidney, hemosiderin-laden macrophages in the spleen, and phagocytized erythrocytes, which seemed to be infected with the parasites, in lymph nodes were also specifically stained. This suggests that in situ hybridization can be utilized to investigate both the life cycle of B. gibsoni and the pathological condition of canine babesiosis.

  13. Removal of aliphatic amino acids by hybrid organic inorganic layered compounds

    NASA Astrophysics Data System (ADS)

    Silverio, Fabiano; dos Reis, Márcio José; Tronto, Jairo; Valim, João Barros

    2007-04-01

    Amino acids have been extensively used in several processes of the pharmaceutical and food industries. Treatments for the recovery and reuse of the wastewaters generated from these processes are few and little known. This work aims at studying the influence of variables like temperature, pH and ionic strength on the adsorption of the amino acids Asp and Glu, contained in aqueous solutions, on layered double hydroxides of the Mg-Al-CO 3-LDH system. The adsorption experiments were performed at two different temperatures (298 and 310 K), two different pH values (7.0 and 10.0), and two ionic strength conditions (with or without the addition of NaCl). The adsorption isotherms exhibited similar profiles under the various conditions studied: an increase in temperature as well as an increase in the pH value decreased the amount of adsorbed amino acid while an increase in the ionic strength increased Asp and Glu adsorption. The PXRD analysis showed that the diffractograms obtained before and after the adsorption of amino acids have a similar pattern. The FT-IR spectra of the adsorbed material presented specific bands, which are related to the amino acids. The concentration range varied up to the anion solubility product and the extraction rate lay between 2.7 and 23.4% at higher equilibrium concentrations, showing that Mg-Al-CO 3-LDH is efficient at removing the amino acids from the aqueous medium.

  14. Colorimetric detection of Ehrlichia canis via nucleic acid hybridization in gold nano-colloids.

    PubMed

    Muangchuen, Ajima; Chaumpluk, Piyasak; Suriyasomboon, Annop; Ekgasit, Sanong

    2014-08-08

    Canine monocytic ehrlichiosis (CME) is a major thick-bone disease of dog caused by Ehrlichia canis. Detection of this causal agent outside the laboratory using conventional methods is not effective enough. Thus an assay for E. canis detection based on the p30 outer membrane protein gene was developed. It was based on the p30 gene amplification using loop-mediated isothermal DNA amplification (LAMP). The primer set specific to six areas within the target gene were designed and tested for their sensitivity and specificity. Detection of DNA signals was based on modulation of gold nanoparticles' surface properties and performing DNA/DNA hybridization using an oligonucleotide probe. Presence of target DNA affected the gold colloid nanoparticles in terms of particle aggregation with a plasmonic color change of the gold colloids from ruby red to purple, visible by the naked eye. All the assay steps were completed within 90 min including DNA extraction without relying on standard laboratory facilities. This method was very specific to target bacteria. Its sensitivity with probe hybridization was sufficient to detect 50 copies of target DNA. This method should provide an alternative choice for point of care control and management of the disease.

  15. Colorimetric Detection of Ehrlichia Canis via Nucleic Acid Hybridization in Gold Nano-Colloids

    PubMed Central

    Muangchuen, Ajima; Chaumpluk, Piyasak; Suriyasomboon, Annop; Ekgasit, Sanong

    2014-01-01

    Canine monocytic ehrlichiosis (CME) is a major thick-bone disease of dog caused by Ehrlichia canis. Detection of this causal agent outside the laboratory using conventional methods is not effective enough. Thus an assay for E. canis detection based on the p30 outer membrane protein gene was developed. It was based on the p30 gene amplification using loop-mediated isothermal DNA amplification (LAMP). The primer set specific to six areas within the target gene were designed and tested for their sensitivity and specificity. Detection of DNA signals was based on modulation of gold nanoparticles' surface properties and performing DNA/DNA hybridization using an oligonucleotide probe. Presence of target DNA affected the gold colloid nanoparticles in terms of particle aggregation with a plasmonic color change of the gold colloids from ruby red to purple, visible by the naked eye. All the assay steps were completed within 90 min including DNA extraction without relying on standard laboratory facilities. This method was very specific to target bacteria. Its sensitivity with probe hybridization was sufficient to detect 50 copies of target DNA. This method should provide an alternative choice for point of care control and management of the disease. PMID:25111239

  16. Ternary Pd-Ni-P hybrid electrocatalysts derived from Pd-Ni core-shell nanoparticles with enhanced formic acid oxidation activity.

    PubMed

    Liang, Xin; Liu, Bo; Zhang, Juntao; Lu, Siqi; Zhuang, Zhongbin

    2016-09-25

    Ternary Pd-Ni-P hybrid electrocatalysts were synthesized through low temperature phosphidation of Pd-Ni core-shell nanoparticles. They show enhanced formic acid electro-oxidation activity compared to Pd, Pd-Ni and Pd-P nanoparticles, which is ascribed to the synergistic effect of the Ni and P components with Pd.

  17. Ozone sensitivity in hybrid poplar correlates with insensitivity to both salicylic acid and jasmonic acid. The role of programmed cell death in lesion formation.

    PubMed

    Koch, J R; Creelman, R A; Eshita, S M; Seskar, M; Mullet, J E; Davis, K R

    2000-06-01

    Our earlier studies demonstrated that the ozone-sensitive hybrid poplar clone NE-388 displays an attenuated level of ozone-, wound-, and phytopathogen-induced defense gene expression. To determine if this reduced gene activation involves signal transduction pathways dependent on salicylic acid (SA) and/or jasmonic acid (JA), we compared the responses of NE-388 and an ozone-tolerant clone, NE-245, to these signal molecules. JA levels increased in both clones in response to ozone, but only minimal increases in SA levels were measured for either clone. Treatment with SA and methyl jasmonate induced defense gene expression only in NE-245, indicating that NE-388 is insensitive to these signal molecules. DNA fragmentation, an indicator of programmed cell death (PCD), was detected in NE-245 treated with either ozone or an avirulent phytopathogen, but was not detected in NE-388. We conclude that these clones undergo two distinct mechanisms of ozone-induced lesion formation. In NE-388, lesions appear to be due to toxic cell death resulting from a limited ability to perceive and subsequently activate SA- and/or JA-mediated antioxidant defense responses. In NE-245, SA-dependent PCD precedes lesion formation via a process related to the PCD pathway activated by phytopathogenic bacteria. These results support the hypothesis that ozone triggers a hypersensitive response.

  18. Hybrid-DFT study on electronic structures of the active site of sweet potato purple acid phosphatase: the origin of stronger antiferromagnetic couplings than other purple acid phosphatases.

    PubMed

    Koizumi, Kenichi; Yamaguchi, Kizashi; Nakamura, Haruki; Takano, Yu

    2009-04-30

    The electronic structure and magnetic interactions of the active site of sweet potato purple acid phosphatase (PAP) were investigated by using UHF, pure DFT (UBLYP), and hybrid DFT methods (UB3LYP and UB2LYP). PAP catalyzes the hydrolysis of a phosphate ester under acidic conditions and contains a binuclear metal center. Sweet potato PAP provides stronger antiferromagnetic coupling than other PAPs. UB3LYP showed reasonably good agreement with the experimental magnetic coupling, indicating that this stronger antiferromagnetic coupling is caused by a mu-oxo bridge in the Fe(III)-Mn(II) binuclear metal center, which is the origin of the asymmetric spin delocalization. The type of bridging ligand is essential for the reaction mechanism, because the bridging ligand is suggested to function as a nucleophile in the reaction. Analyses of the natural orbital and spin density distributions implied the asymmetric spin delocalization on the bridging oxygen. The mechanism and the pathway of the antiferromagnetic coupling between Fe(III) and Mn(II) were discussed, using chemical indices introduced with the occupation numbers of singly occupied natural orbitals.

  19. Local sustained delivery of acetylsalicylic acid via hybrid stent with biodegradable nanofibers reduces adhesion of blood cells and promotes reendothelialization of the denuded artery.

    PubMed

    Lee, Cheng-Hung; Lin, Yu-Huang; Chang, Shang-Hung; Tai, Chun-Der; Liu, Shih-Jung; Chu, Yen; Wang, Chao-Jan; Hsu, Ming-Yi; Chang, Hung; Chang, Gwo-Jyh; Hung, Kuo-Chun; Hsieh, Ming-Jer; Lin, Fen-Chiung; Hsieh, I-Chang; Wen, Ming-Shien; Huang, Yenlin

    2014-01-01

    Incomplete endothelialization, blood cell adhesion to vascular stents, and inflammation of arteries can result in acute stent thromboses. The systemic administration of acetylsalicylic acid decreases endothelial dysfunction, potentially reducing thrombus, enhancing vasodilatation, and inhibiting the progression of atherosclerosis; but, this is weakened by upper gastrointestinal bleeding. This study proposes a hybrid stent with biodegradable nanofibers, for the local, sustained delivery of acetylsalicylic acid to injured artery walls. Biodegradable nanofibers are prepared by first dissolving poly(D,L)-lactide-co-glycolide and acetylsalicylic acid in 1,1,1,3,3,3-hexafluoro-2-propanol. The solution is then electrospun into nanofibrous tubes, which are then mounted onto commercially available bare-metal stents. In vitro release rates of pharmaceuticals from nanofibers are characterized using an elution method, and a highperformance liquid chromatography assay. The experimental results suggest that biodegradable nanofibers release high concentrations of acetylsalicylic acid for three weeks. The in vivo efficacy of local delivery of acetylsalicylic acid in reducing platelet and monocyte adhesion, and the minimum tissue inflammatory reaction caused by the hybrid stents in treating denuded rabbit arteries, are documented. The proposed hybrid stent, with biodegradable acetylsalicylic acid-loaded nanofibers, substantially contributed to local, sustained delivery of drugs to promote re-endothelialization and reduce thrombogenicity in the injured artery. The stents may have potential applications in the local delivery of cardiovascular drugs. Furthermore, the use of hybrid stents with acetylsalicylic acid-loaded nanofibers that have high drug loadings may provide insight into the treatment of patients with high risk of acute stent thromboses.

  20. A dual response organogel system based on an iridium complex and a Eu(iii) hybrid for volatile acid and organic amine vapors.

    PubMed

    Cao, Xinhua; Zhao, Na; Zou, Guodong; Gao, Aiping; Ding, Qianqian; Zeng, Guanjie; Wu, Yongquan

    2017-05-24

    A thiophene-based hybrid organogel system consisting of complex iridium (Ir) and EuCl3·6H2O was designed and synthesized to realize dual responses to volatile acids and organic amine vapors. The photophysical properties and self-assembly of compound 1 and the hybrid organogel were also studied. Compound 1 could gelate some organic solvents and self-assemble into 3D nanofibers in the gels. The stable hybrid organogel 1-Ir-Eu could be obtained after addition of complex Ir and EuCl3·6H2O. FTIR spectral results showed that the hydrogen bond still remained even upon addition of complex Ir, EuCl3·6H2O, NaOH and CF3COOH to organogel 1. Interestingly, the emission properties of the hybrid organogel 1-Ir-Eu could undergo interconversion between cyan light and red light via addition of NaOH and CF3COOH. The emission properties of xerogel film 1-Ir-Eu obtained in the presence of NaOH could also undergo fast and reversible transition in response to volatile acids such as CF3COOH, formic acid, acetic acid, propionic acid and organic amine vapors such as ammonium hydroxide, Et3N, tripropylamine, and ethylenediamine. The emission spectral change of Ir-Eu in the organogel or xerogel in the presence of base and acid demonstrated the formation of a new complex between complex Ir and EuCl3·6H2O. This dual-response process could be repeated many times. Contact angle experiment results further showed the morphology and internal components of the xerogel film surface in the process of response to gaseous CF3COOH and Et3N. This work provides a method for producing multifunctional supramolecular materials for sensing volatile acids and organic amine vapors.

  1. Local sustained delivery of acetylsalicylic acid via hybrid stent with biodegradable nanofibers reduces adhesion of blood cells and promotes reendothelialization of the denuded artery

    PubMed Central

    Lee, Cheng-Hung; Lin, Yu-Huang; Chang, Shang-Hung; Tai, Chun-Der; Liu, Shih-Jung; Chu, Yen; Wang, Chao-Jan; Hsu, Ming-Yi; Chang, Hung; Chang, Gwo-Jyh; Hung, Kuo-Chun; Hsieh, Ming-Jer; Lin, Fen-Chiung; Hsieh, I-Chang; Wen, Ming-Shien; Huang, Yenlin

    2014-01-01

    Incomplete endothelialization, blood cell adhesion to vascular stents, and inflammation of arteries can result in acute stent thromboses. The systemic administration of acetylsalicylic acid decreases endothelial dysfunction, potentially reducing thrombus, enhancing vasodilatation, and inhibiting the progression of atherosclerosis; but, this is weakened by upper gastrointestinal bleeding. This study proposes a hybrid stent with biodegradable nanofibers, for the local, sustained delivery of acetylsalicylic acid to injured artery walls. Biodegradable nanofibers are prepared by first dissolving poly(D,L)-lactide-co-glycolide and acetylsalicylic acid in 1,1,1,3,3,3-hexafluoro-2-propanol. The solution is then electrospun into nanofibrous tubes, which are then mounted onto commercially available bare-metal stents. In vitro release rates of pharmaceuticals from nanofibers are characterized using an elution method, and a highperformance liquid chromatography assay. The experimental results suggest that biodegradable nanofibers release high concentrations of acetylsalicylic acid for three weeks. The in vivo efficacy of local delivery of acetylsalicylic acid in reducing platelet and monocyte adhesion, and the minimum tissue inflammatory reaction caused by the hybrid stents in treating denuded rabbit arteries, are documented. The proposed hybrid stent, with biodegradable acetylsalicylic acid-loaded nanofibers, substantially contributed to local, sustained delivery of drugs to promote re-endothelialization and reduce thrombogenicity in the injured artery. The stents may have potential applications in the local delivery of cardiovascular drugs. Furthermore, the use of hybrid stents with acetylsalicylic acid-loaded nanofibers that have high drug loadings may provide insight into the treatment of patients with high risk of acute stent thromboses. PMID:24421640

  2. Amplified electrochemical detection of nucleic acid hybridization via selective preconcentration of unmodified gold nanoparticles.

    PubMed

    Li, Yuan; Tian, Rui; Zheng, Xingwang; Huang, Rongfu

    2016-08-31

    The common drawback of optical methods for rapid detection of nucleic acid by exploiting the differential affinity of single-/double-stranded nucleic acids for unmodified gold nanoparticles (AuNPs) is its relatively low sensitivity. In this article, on the basis of selective preconcentration of AuNPs unprotected by single-stranded DNA (ssDNA) binding, a novel electrochemical strategy for nucleic acid sequence identification assay has been developed. Through detecting the redox signal mediated by AuNPs on 1, 6-hexanedithiol blocked gold electrode, the proposed method is able to ensure substantial signal amplification and a low background current. This strategy is demonstrated for quantitative analysis of the target microRNA (let-7a) in human breast adenocarcinoma cells, and a detection limit of 16 fM is readily achieved with desirable specificity and sensitivity. These results indicate that the selective preconcentration of AuNPs for electrochemical signal readout can offer a promising platform for the detection of specific nucleic acid sequence.

  3. Oxygen-dependent chemical tumorigenesis in a Nicotiana hybrid: inhibition by ascorbic acid and dinitrophenol.

    PubMed

    Andersen, R A; Linney, T L

    1977-12-01

    Aqueous solutions of molecular oxygen, per se, or in combination with either pyrogallol or 6-azauracil increased tumorigenesis in Nicotiana suaveolens X Nicotiana langsdorffii seedlings relative to control seedlings. The biological activities of the organic chemicals were O2-dependent, because the substitution of N2 or O2 or the degassing of 0.1-1 mM solutions of the compounds eliminated or greatly reduced their tumorigenic effects. Rates of tumorigenesis exceeded 95% for 0.5 mM solutions of either pyrogallol or 6-azauracil solutions in the presence of l mM O2. Although tumors developed in 20% of seedlings in the presence of 1 mM O2, alone, 4-5 times more tumors were induced by the organic chemical--O2-H2O systems. Dinitrophenol and ascorbic acid, compounds which affect cellular respiration or redox systems, strongly inhibited the chemically-mediated tumorigenesis. Dinitrophenol was equally effective at one-tenth of the molar concentrations of ascorbic acid that were required for the suppressions of oncogenesis. Dehydroascorbic acid was much less inhibitory than ascorbic acid.

  4. Computer programs for analysis of nucleic acid hybridization, thermal denaturation, and gel electrophoresis data.

    PubMed Central

    Murphy, R F; Pearson, W R; Bonner, J

    1979-01-01

    Computer programs for the analysis of data from techniques frequently used in nucleic acids research are described. In addition to calculating non-linear, least-squares solutions to equations describing these systems, the programs allow for data editing, normalization, plotting and storage, and are flexible and simple to use. Typical applications of the programs are described. PMID:493129

  5. Hybrid Lead-Acid/Lithium-Ion Energy Storage System with Power-Mix Control for Light Electric Vehicles

    NASA Astrophysics Data System (ADS)

    Chung, Steven

    This work presents a hybrid energy storage system (HESS), using lithium-ion (LI) and lead-acid (PbA) batteries, for light electric vehicles (LEV) that is both cost and performance competitive with single energy storage system (SESS) configurations. A modular HESS architecture with a dc-dc converter and controller is proposed. The power-mix algorithm that minimizes the Peukert effect, and increases the usable energy of the lead-acid battery is presented and verified experimentally. A novel DLL and PLL based off-time control scheme is demonstrated for inductor ripple current interleaving in the multi-phase dc-dc converter. The cost and performance of the HESS are assessed side-by-side with PbA and LI SESS configurations of comparable total energy. The experimental HESS has a total projected cost midway between the SESS PbA cost and the SESS Li cost, while providing 17% range and 23% efficiency increase over the SESS PbA vehicle.

  6. Preparation and characterization of humic acid-carbon hybrid materials as adsorbents for organic micro-pollutants.

    PubMed

    Radwan, Emad K; Abdel Ghafar, Hany H; Moursy, Ahmed S; Langford, Cooper H; Bedair, Ahmed H; Achari, Gopal

    2015-08-01

    The present work involves the preparation of novel adsorbent materials by the insolubilization and hybridization of humic acid (HA) with carbon. The prepared materials were characterized by N2 adsorption, elemental analysis, Fourier transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy, solid-state (13)C cross polarization magic angle spinning nuclear magnetic resonance, and low-field nuclear magnetic resonance (NMR) relaxometry on wetted samples. The water solubility of these materials and the lack of effect of oxidants were also confirmed. With this background, the adsorption capacities toward phenol, 2,4,6-tricholrophenol, and atrazine were evaluated, using these as model compounds for organic micropollutants of concern in water. Experimental results show that the prepared materials are mesoporous and have a higher surface area than humic acid and even than the porous carbon in the case of carbon coating. They retain the basic features of the starting materials with lowered functional group content. Moreover, there are interesting new features. NMR relaxometry shows that equilibration of water uptake is very fast, making use in water simple. They have higher adsorption capacities than the pure materials, and they can be applied under a wide range of environmental conditions.

  7. [Oligonucleotide derivatives in the nucleic acid hybridization analysis. I. Covalent immobilization of oligonucleotide probes onto the nylon].

    PubMed

    Dmitrienko, E V; Pyshnaia, I A; Pyshnyĭ, D V

    2010-01-01

    The features of UV-induced immobilization of oligonucleotides on a nylon membranes and the effectiveness of enzymatic labeling of immobilized probes at heterophase detection of nucleic acids are studied. Short terminal oligothymidilate (up to 10 nt) sequences are suggested to attach to the probe via a flexible ethylene glycol based linker. The presence of such fragment enhances the intensity of immobilization and reduces UV-dependent degradation of the targeted (sequence-specific) part of the probe by reducing the dose needed for the immobilization of DNA. The optimum dose of UV-irradiation is determined to be ~0.4 J/cm(2) at the wavelength 254 nm. This dose provides high level of hybridization signal for immobilized probes with various nucleotide composition of the sequence specific moiety. The amide groups of the polyamide are shown to play the key role in the photoinduced immobilization of nucleic acids, whereas the primary amino groups in the structure of PA is not the center responsible for the covalent binding of DNA by UV-irradiation, as previously believed. Various additives in the soaking solution during the membrane of UV-dependent immobilization of probes are shown to influence its effectiveness. The use of alternative to UV-irradiation system of radical generation are shown to provide the immobilization of oligonucleotides onto the nylon membrane.

  8. A Strategy for Dual Inhibition of the Proteasome and Fatty Acid Synthase with Belactosin C-Orlistat Hybrids

    PubMed Central

    Zhu, Mingzhao; Harshbarger, Wayne D.; Robles, Omar; Krysiak, Joanna; Hull, Kenneth G.; Cho, Sung Wook; Richardson, Robyn D.; Yang, Yanyan; Garcia, Andres; Spiegelman, Lindsey; Ramirez, Bianca; Wilson, Christopher T.; Yau, Ju Anne; Moore, James T.; Walker, Caitlen B.; Sacchettini, James C.; Liu, Wenshe; Sieber, Stephan A.; Smith, Jeffrey W.; Romo, Daniel

    2017-01-01

    The proteasome, a validated cellular target for cancer, is central for maintaining cellular homeostasis, while fatty acid synthase (FAS), a novel target for numerous cancers, is responsible for palmitic acid biosynthesis. Perturbation of either enzymatic machine results in decreased proliferation and ultimately cellular apoptosis. Based on structural similarities, we hypothesized that hybrid molecules of belactosin C, a known proteasome inhibitor, and orlistat, a known inhibitor of the thioesterase domain of FAS, could inhibit both enzymes. Herein, we describe proof-of-principle studies leading to the design, synthesis and enzymatic activity of several novel, β-lactone-based, dual inhibitors of these two enzymes. Validation of dual enzyme targeting through activity-based proteome profiling with an alkyne probe modeled after the most potent inhibitor, and preliminary serum stability studies of selected derivatives are also described. These results provide proof of concept for dual targeting of the proteasome and FAS-TE enabling a new approach for the development of drug-candidates with potential to overcome resistance. PMID:28236510

  9. Hyaluronic Acid-Modified Cationic Lipid-PLGA Hybrid Nanoparticles as a Nanovaccine Induce Robust Humoral and Cellular Immune Responses.

    PubMed

    Liu, Lanxia; Cao, Fengqiang; Liu, Xiaoxuan; Wang, Hai; Zhang, Chao; Sun, Hongfan; Wang, Chun; Leng, Xigang; Song, Cunxian; Kong, Deling; Ma, Guilei

    2016-05-18

    Here, we investigated the use of hyaluronic acid (HA)-decorated cationic lipid-poly(lactide-co-glycolide) acid (PLGA) hybrid nanoparticles (HA-DOTAP-PLGA NPs) as vaccine delivery vehicles, which were originally developed for the cytosolic delivery of genes. Our results demonstrated that after the NPs uptake by dendritic cells (DCs), some of the antigens that were encapsulated in HA-DOTAP-PLGA NPs escaped to the cytosolic compartment, and whereas some of the antigens remained in the endosomal/lysosomal compartment, where both MHC-I and MHC-II antigen presentation occurred. Moreover, HA-DOTAP-PLGA NPs led to the up-regulation of MHC, costimulatory molecules, and cytokines. In vivo experiments further revealed that more powerful immune responses were induced from mice immunized with HA-DOTAP-PLGA NPs when compared with cationic lipid-PLGA nanoparticles and free ovalbumin (OVA); the responses included antigen-specific CD4(+) and CD8(+) T-cell responses, the production of antigen-specific IgG antibodies and the generation of memory CD4(+) and CD8(+) T cells. Overall, these data demonstrate the high potential of HA-DOTAP-PLGA NPs for use as vaccine delivery vehicles to elevate cellular and humoral immune responses.

  10. Multiplexed hybridizations of positively charge-tagged peptide nucleic acids detected by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    PubMed

    Bauer, Oliver; Guerasimova, Anna; Sauer, Sascha; Thamm, Sabine; Steinfath, Matthias; Herwig, Ralf; Janitz, Michal; Lehrach, Hans; Radelof, Uwe

    2004-01-01

    Peptide nucleic acid (PNA) is a novel class of DNA analogues in which the entire sugar-phosphate backbone is replaced by a pseudopeptide counterpart. Owing to its neutral character and the consequent lack of electrostatic repulsion, PNA exhibits very stable heteroduplex formation with complementary nucleic acid that is essentially ionic strength independent and enables hybridization under minimum salt conditions. This feature as well as its superior ion stability and easy ionization compared to DNA renders PNA very attractive for hybridization-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) applications. We have developed an approach to DNA characterization that takes advantage of multiplexed PNA hybridizations analyzed by MALDI-TOFMS. Our motivation was the further development of oligonucleotide fingerprinting, an efficient technique for cDNA and genomic DNA library characterization. Through positive 'charge-tagging' of PNA the efficiency of detection in MALDI-TOFMS was considerably enhanced permitting an unparalleled degree of multiplexing. Results from the simultaneous hybridization of 21 charge-tagged PNA hexamer oligonucleotides showed that genomic DNA and cDNA clones are successfully characterized on the basis of their hybridization profiles. The degree of multiplexing achieved may render a significant increase in throughput and hence efficiency of oligonucleotide fingerprinting possible.

  11. Packaging of an AAV vector encoding human acid alpha-glucosidase for gene therapy in glycogen storage disease type II with a modified hybrid adenovirus-AAV vector.

    PubMed

    Sun, Baodong; Chen, Y-T; Bird, Andrew; Xu, Fang; Hou, Yang-Xun; Amalfitano, Andrea; Koeberl, Dwight D

    2003-04-01

    We have developed an improved method for packaging adeno-associated virus (AAV) vectors with a replication-defective adenovirus-AAV (Ad-AAV) hybrid virus. The AAV vector encoding human acid alpha-glucosidase (hGAA) was cloned into an E1, polymerase/preterminal protein-deleted adenovirus, such that it is packaged as an Ad vector. Importantly, the Ad-AAV hybrid cannot replicate during AAV vector packaging in 293 cells, because of deletion of polymerase/preterminal protein. The residual Ad-AAV in the AAV vector stock was reduced to <1 infectious particle per 10(10) AAV vector particles. These modifications resulted in approximately 30-fold increased packaging of the AAV vector for the hybrid Ad-AAV vector method as compared with standard transfection-only methods. Similarly improved packaging was demonstrated for pseudotyping the AAV vector as AAV6, and for AAV vector packaging with a second Ad-AAV vector encoding canine glucose-6-phosphatase. Liver-targeted delivery of either the Ad-AAV hybrid or AAV vector particles in acid alpha-glucosidase-knockout (GAA-KO) mice revealed secretion of hGAA with the Ad-AAV vector, and sustained secretion of hGAA with an AAV vector in hGAA-tolerant GAA-KO mice. Further development of hybrid Ad-AAV vectors could offer distinct advantages for gene therapy in glycogen storage diseases.

  12. Optimization of a peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) method for the detection of bacteria and disclosure of a formamide effect.

    PubMed

    Santos, Rita S; Guimarães, Nuno; Madureira, Pedro; Azevedo, Nuno F

    2014-10-10

    Despite the fact that fluorescence in situ hybridization (FISH) is a well-established technique to identify microorganisms, there is a lack of understanding concerning the interaction of the different factors affecting the obtained fluorescence. In here, we used flow cytometry to study the influence of three essential factors in hybridization - temperature, time and formamide concentration - in an effort to optimize the performance of a Peptide Nucleic Acid (PNA) probe targeting bacteria (EUB338). The PNA-FISH optimization was performed with bacteria representing different families employing response surface methodology. Surprisingly, the optimum concentration of formamide varied according to the bacterium tested. While hybridization on the bacteria possessing the thickest peptidoglycan was more successful at nearly 50% (v/v) formamide, hybridization on all other microorganisms appeared to improve with much lower formamide concentrations. Gram staining and transmission electron microscopy allowed us to confirm that the overall effect of formamide concentration on the fluorescence intensity is a balance between a harmful effect on the bacterial cell envelope, affecting cellular integrity, and the beneficial denaturant effect in the hybridization process. We also conclude that microorganisms belonging to different families will require different hybridization parameters for the same FISH probe, meaning that an optimum universal PNA-FISH procedure is non-existent for these situations.

  13. Multiplexed interfacial transduction of nucleic acid hybridization using a single color of immobilized quantum dot donor and two acceptors in fluorescence resonance energy transfer.

    PubMed

    Algar, W Russ; Krull, Ulrich J

    2010-01-01

    A multiplexed solid-phase assay for the detection of nucleic acid hybridization was developed on the basis of a single color of immobilized CdSe/ZnS quantum dot (QD) as a donor in fluorescence resonance energy transfer (FRET). This work demonstrated that two channels of detection did not necessitate two different QD donors. Two probe oligonucleotides were coimmobilized on optical fibers modified with QDs, and a sandwich assay was used to associate the acceptor dyes with interfacial hybridization events without target labeling. FRET-sensitized acceptor emission provided an analytical signal that was concentration dependent down to 10 nM. Changes in the ratio of coimmobilized probe oligonucleotides were found to yield linear changes in the relative amounts of acceptor emission. These changes were compared to previous studies that used mixed films of two QD donors for two detection channels. The analysis indicated that probe dilution effects were primarily driven by changes in acceptor number density and that QD dilution effects or changes in mean donor-acceptor distance were secondary. Hybridization kinetics were found to be consistent between different ratios of coimmobilized probes, suggesting that hybridization in this type of system occurred via the accepted model for solid-phase hybridization, where adsorption and then diffusion at the solid interface drove hybridization.

  14. Hybrid striped bass feeds based on fish oil, beef tallow, and eicosapentaenoic acid/docosahexaenoic acid supplements: Insight regarding fish oil sparing and demand for -3 long-chain polyunsaturated fatty acids.

    PubMed

    Bowzer, J; Jackson, C; Trushenski, J

    2016-03-01

    Previous research suggests that saturated (SFA) and monounsaturated fatty acid (MUFA) rich lipids, including beef tallow, can make utilization or diet-to-tissue transfer of long-chain polyunsaturated fatty acids (LC-PUFA) more efficient. We hypothesized that using beef tallow as an alternative to fish oil may effectively reduce the LC-PUFA demand of hybrid striped bass × and allow for greater fish oil sparing. Accordingly, we evaluated growth performance and tissue fatty acid profiles of juvenile fish (23.7 ± 0.3 g) fed diets containing menhaden fish oil (considered an ideal source of LC-PUFA for this taxon), beef tallow (BEEF ONLY), or beef tallow amended with purified sources of eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) to achieve levels corresponding to 50 or 100% of those observed in the FISH ONLY feed. Diets were randomly assigned to quadruplicate tanks of fish ( = 4; 10 fish/tank), and fish were fed assigned diets to apparent satiation once daily for 10 wk. Survival (98-100%) was equivalent among treatments, but weight gain (117-180%), specific growth rate (1.1-1.5% BW/d), feed intake (1.4-1.8% BW/d), thermal growth coefficient (0.50-0.70), and feed conversion ratio (FCR; 1.1-1.4, DM basis) varied. Except for FCR, no differences were observed between the FISH ONLY and BEEF ONLY treatments, but performance was generally numerically superior among fish fed the diets containing beef tallow supplemented with DHA at the 100% or both EPA and DHA at the 50% or 100% level. Tissue fatty acid composition was significantly distorted in favor among fish fed the beef tallow-based feeds; however, profile distortion was most overt in peripheral tissues. Results suggest that beef tallow may be used as a primary lipid source in practical diets for hybrid striped bass, but performance may be improved by supplementation with LC-PUFA, particularly DHA. Furthermore, our results suggest that -3 LC-PUFA requirements reported for hybrid striped bass may not be

  15. Design and synthesis of palladium/graphitic carbon nitride/carbon black hybrids as high-performance catalysts for formic acid and methanol electrooxidation

    NASA Astrophysics Data System (ADS)

    Qian, Huayu; Huang, Huajie; Wang, Xin

    2015-02-01

    Here we report a facile two-step method to synthesize high-performance palladium/graphitic carbon nitride/carbon black (Pd/g-C3N4/carbon black) hybrids for electrooxidizing formic acid and methanol. The coating of g-C3N4 on carbon black surface is realized by a low-temperature heating treatment, followed by the uniform deposition of palladium nanoparticles (Pd NPs) via a wet chemistry route. Owning to the significant synergistic effects of the individual components, the preferred Pd/g-C3N4/carbon black electrocatalyst exhibits exceptional forward peak current densities as high as 2155 and 1720 mA mg-1Pd for formic acid oxidation in acid media and methanol oxidation in alkaline media, respectively, far outperforming the commercial Pd-C catalyst. The catalyst also shows reliable stability, demonstrating that the newly-designed hybrids have great promise in constructing high-performance portable fuel cell systems.

  16. Remote control of lipophilic nucleic acids domain partitioning by DNA hybridization and enzymatic cleavage.

    PubMed

    Schade, Matthias; Knoll, Andrea; Vogel, Alexander; Seitz, Oliver; Liebscher, Jürgen; Huster, Daniel; Herrmann, Andreas; Arbuzova, Anna

    2012-12-19

    Lateral partitioning of lipid-modified molecules between liquid-disordered (ld) and liquid-ordered (lo) domains depends on the type of lipid modification, presence of a spacer, membrane composition, and temperature. Here, we show that the lo domain partitioning of the palmitoylated peptide nucleic acid (PNA) can be influenced by formation of a four-component complex with the ld domain partitioning tocopherol-modified DNA: the PNA-DNA complex partitioned into the ld domains. Enzymatic cleavage of the DNA linker led to the disruption of the complex and restored the initial distribution of the lipophilic nucleic acids into the respective domains. This modular system offers strategies for dynamic functionalization of biomimetic surfaces, for example, in nanostructuring and regulation of enzyme catalysis, and it provides a tool to study the molecular basis of controlled reorganization of lipid-modified proteins in membranes, for example, during signal transduction.

  17. Electric and hybrid vehicles charge efficiency tests of ESB EV-106 lead-acid batteries

    SciTech Connect

    Rowlette, J.J.

    1981-01-15

    Charge efficiencies were determined for ESB EV-106 lead-acid batteries by measurements made under widely differing conditions of temperature, charge procedure, and battery age. The measurements were used to optimize charge procedures and to evaluate the concept of a modified, coulometric state-of-charge indicator. Charge efficiency determinations were made by measuring gassing rates and oxygen fractions. A novel, positive displacement gas flow meter which proved to be both simple and highly accurate is described and illustrated.

  18. Enhanced proton conductivity of Nafion hybrid membrane under different humidities by incorporating metal-organic frameworks with high phytic acid loading.

    PubMed

    Li, Zhen; He, Guangwei; Zhang, Bei; Cao, Ying; Wu, Hong; Jiang, Zhongyi; Tiantian, Zhou

    2014-06-25

    In this study, phytic acid (myo-inositol hexaphosphonic acid) was first immobilized by MIL101 via vacuum-assisted impregnation method. The obtained phytic@MIL101 was then utilized as a novel filler to incorporate into Nafion to fabricate hybrid proton exchange membrane for application in PEMFC under different relative humidities (RHs), especially under low RHs. High loading and uniform dispersion of phytic acid in MIL 101(Cr) were achieved as demonstrated by ICP, FT-IR, XPS, and EDS-mapping. The phytic@MIL101 was dispersed homogeneously in the Nafion matrix when the filler content was less than 12%. Hybrid membranes were evaluated by proton conductivity, mechanical property, thermal stability, and so forth. Remarkably, the Nafion/phytic@MIL hybrid membranes showed high proton conductivity at different RHs, especially under low RHs, which was up to 0.0608 S cm(-1) and 7.63 × 10(-4) S cm(-1) at 57.4% RH and 10.5% RH (2.8 and 11.0 times higher than that of pristine membrane), respectively. Moreover, the mechanical property of Nafion/phtic@MIL hybrid membranes was substantially enhanced and the thermal stability of membranes was well preserved.

  19. Synthesis, Characterization, and Environmental Applications of Hybrid Materials Based on Humic Acid Obtained by the Sol-Gel Route.

    PubMed

    Oliveira, Lílian Karla de; Molina, Eduardo Ferreira; Moura, André L A; de Faria, Emerson Henrique; Ciuffi, Katia Jorge

    2016-01-20

    Humic acids (HAs) are ubiquitous macromolecules in the environment. Due to their high contents of oxygenated functional groups, they can interact with contaminants present in the natural environment and therefore influence the behavior of pollutants. However, a pH of 2 or lower is required to maintain HAs in the solid form. To increase the stability of HAs and their capacity to bind to contaminants, this work proposes the development of new hybrid materials based on alkoxysilanes and HAs for environmental applications such as dye adsorption. Three different materials with new functional groups were prepared by employing the following alkoxysilanes: tetraethyl orthosilicate, (3-aminopropyl)triethoxysilane, and N-[3-(trimethoxylsilyl)propyl]ethylenediamine. The final materials were denoted HWA, HOA, and HTA, respectively, and they were characterized by elemental analysis, diffuse reflectance Fourier-transform infrared spectroscopy (DRIFT), small-angle X-ray scattering (SAXS), scanning electron microscopy (SEM), and N2 gas-volumetric adsorption. The point of zero charge (pzc) and stability of these materials were also determined. Their selectivity was evaluated in adsorption experiments performed with two different charged dyes in aqueous medium, namely anionic rose bengal (RB) and cationic methylene blue (MB). The elemental, DRIFT, SAXS, SEM, and textural analyses confirmed the presence of a combination of the features of HAs and alkoxysilanes. The pzc results showed that the new materials displayed different characteristics and affinities. All the materials were stable in aqueous solution up to pH 10. For MB, the percentage removal values obtained by using HWA, HOA, and HTA were 98, 85, and 67%, respectively. As for RB, the percentage removal values were 19, 18, and 44% for HWA, HOA, and HTA, respectively. These hybrid materials have potential use as adsorbents for the removal of cationic or anionic species and could be viable alternatives to remove various

  20. Microbial populations identified by fluorescence in situ hybridization in a constructed wetland treating acid coal mine drainage.

    PubMed

    Nicomrat, Duongruitai; Dick, Warren A; Tuovinen, Olli H

    2006-01-01

    Microorganisms are an integral part of the biogeochemical processes in wetlands, yet microbial communities in sediments within constructed wetlands receiving acid mine drainage (AMD) are only poorly understood. The purpose of this study was to characterize the microbial diversity and abundance in a wetland receiving AMD using fluorescence in situ hybridization (FISH) analysis. Seasonal samples of oxic surface sediments, comprised of Fe(III) precipitates, were collected from two treatment cells of the constructed wetland system. The pH of the bulk samples ranged between pH 2.1 and 3.9. Viable counts of acidophilic Fe and S oxidizers and heterotrophs were determined with a most probable number (MPN) method. The MPN counts were only a fraction of the corresponding FISH counts. The sediment samples contained microorganisms in the Bacteria (including the subgroups of acidophilic Fe- and S-oxidizing bacteria and Acidiphilium spp.) and Eukarya domains. Archaea were present in the sediment surface samples at < 0.01% of the total microbial community. The most numerous bacterial species in this wetland system was Acidithiobacillus ferrooxidans, comprising up to 37% of the bacterial population. Acidithiobacillus thiooxidans was also abundant. Heterotrophs in the Acidiphilium genus totaled 20% of the bacterial population. Leptospirillum ferrooxidans was below the level of detection in the bacterial community. The results from the FISH technique from this field study are consistent with results from other experiments involving enumeration by most probable number, dot-blot hybridization, and denaturing gradient gel electrophoresis analyses and with the geochemistry of the site.

  1. Preparation of hybrid molecularly imprinted polymer with double-templates for rapid simultaneous purification of theophylline and chlorogenic acid in green tea.

    PubMed

    Tang, Weiyang; Li, Guizhen; Row, Kyung Ho; Zhu, Tao

    2016-05-15

    A novel double-templates technique was adopted for solid-phase extraction packing agent, and the obtained hybrid molecularly imprinted polymers with double-templates (theophylline and chlorogenic acid) were characterized by fourier transform infrared and field emission scanning electron microscope. The molecular recognition ability and binding capability for theophylline and chlorogenic acid of polymers was evaluated by static absorption and dynamic adsorption curves. A rapid and accurate approach was established for simultaneous purification of theophylline and chlorogenic acid in green tea by coupling hybrid molecularly imprinted solid-phase extraction with high performance liquid chromatography. With optimization of SPE procedure, a reliable analytical method was developed for highly recognition towards theophylline and chlorogenic acid in green tea with satisfactory extraction recoveries (theophylline: 96.7% and chlorogenic acid: 95.8%). The limit of detection and limit of quantity of the method were 0.01 μg/mL and 0.03 μg/mL for theophylline, 0.05 μg/mL and 0.17 μg/mL for chlorogenic acid, respectively. The recoveries of proposed method at three spiked levels analysis were 98.7-100.8% and 98.3-100.2%, respectively, with the relative standard deviation less than 1.9%. Hybrid molecularly imprinted polymers with double-templates showed good performance for two kinds of targets, and the proposed approach with high affinity of hybrid molecularly imprinted polymers might offer a novel method for the purification of complex samples. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Growth, body fatty acid composition, immune response and resistance to Streptococcus iniae of hybrid tilapia, Oreochromis niloticus X O. aureus, fed diets containing various levels of linoleic and linolenic acids

    USDA-ARS?s Scientific Manuscript database

    The effects of dietary linoleic (LA) and linolenic acids (LN) on growth and immunity of all-male hybrid tilapia, Oreochromis niloticus × O. aureus, were evaluated for 10 weeks. Fish fed 0.12% LA + 0% LN had the lowest weight gain (WG) but was not significantly different from diets containing 0.5% LA...

  3. Effect of acid or alkaline catalyst and of different capping agents on the optical properties of CdS nanoparticles incorporated within a diureasil hybrid matrix

    NASA Astrophysics Data System (ADS)

    Gonçalves, Luis F. F. F.; Silva, Carlos J. R.; Kanodarwala, Fehmida K.; Stride, John A.; Pereira, Mario R.

    2015-11-01

    CdS nanoparticles (NPs) were synthesized using colloidal methods and incorporated within a diureasil hybrid matrix. The surface capping of the CdS NPs by 3-mercaptopropyltrimethoxysilane (MPTMS) and 3-aminopropyltrimethoxysilane (APTMS) organic ligands during the incorporation of the NPs within the hybrid matrix has been investigated. The matrix is based on poly(ethylene oxide)/poly(propylene oxide) chains grafted to a siliceous skeleton through urea bonds and was produced by sol-gel process. Both alkaline and acidic catalysis of the sol-gel reaction were used to evaluate the effect of each organic ligand on the optical properties of the CdS NPs. The hybrid materials were characterized by absorption, steady-state and time-resolved photoluminescence spectroscopy and High Resolution Transmission Electron Microscopy (HR-TEM). The preservation of the optical properties of the CdS NPs within the diureasil hybrids was dependent on the experimental conditions used. Both organic ligands (APTMS and MPTMS) demonstrated to be crucial in avoiding the increase of size distribution and clustering of the NPs within the hybrid matrix. The use of organic ligands was also shown to influence the level of interaction between the hybrid host and the CdS NPs. The CdS NPs showed large Stokes shifts and long average lifetimes, both in colloidal solution and in the xerogels, due to the origin of the PL emission in surface states. The CdS NPs capped with MPTMS have lower PL lifetimes compared to the other xerogel samples but still larger than the CdS NPs in the original colloidal solution. An increase in PL lifetimes of the NPs after their incorporation within the hybrid matrix is related to interaction between the NPs and the hybrid host matrix.

  4. Lead-acid batteries for micro- and mild-hybrid applications

    NASA Astrophysics Data System (ADS)

    Valenciano, J.; Fernández, M.; Trinidad, F.; Sanz, L.

    Car manufactures have announced the launch in coming months of vehicles with reduced emissions due to the introduction of new functions like stop-start and regenerative braking. Initial performance request of automotive lead-acid batteries are becoming more and more demanding and, in addition to this, cycle life with new accelerated ageing profiles are being proposed in order to determine the influence of the new functions on the expected battery life. This paper will show how different lead-acid battery technologies comply with these new demands, from an improved version of the conventional flooded SLI battery to the high performance of spiral wound valve-regulated lead-acid (VRLA) battery. Different approaches have been studied for improving conventional flooded batteries, i.e., either by the addition of new additives for reducing electrolyte stratification or by optimisation of the battery design to extend cycling life in partial state of charge conditions. With respect to VRLA technology, two different battery designs have been compared. Spiral wound design combines excellent power capability and cycle life under different depth of discharge (DoD) cycling conditions, but flat plate design outperform the latter in energy density due to better utilization of the space available in a prismatic enclosure. This latter design is more adequate for high end class vehicles with high electrical energy demand, whereas spiral wound is better suited for high power/long life demand of commercial vehicle. High temperature behaviour (75 °C) is rather poor for both designs due to water loss, and then VRLA batteries should preferably be located out of the engine compartment.

  5. Sialyl Lewis(x) hybridized complement receptor type 1 moderates acid aspiration injury.

    PubMed

    Kyriakides, C; Wang, Y; Austen, W G; Favuzza, J; Kobzik, L; Moore, F D; Hechtman, H B

    2001-12-01

    The potentially enhanced anti-inflammatory effects of the sialyl Lewis(x) (sLe(x))-decorated version of soluble complement receptor type 1 (sCR1) in moderating acid aspiration injury are examined. HCl was instilled in tracheostomy tubes placed in mice, and extravasation of (125)I-labeled albumin in bronchoalveolar lavage (BAL) fluid was used to calculate the vascular permeability index (PI). Neutrophil counts in BAL fluid and immunohistochemistry were performed. PI was moderated by 82% after treatment with sCR1sLe(x) compared with 54% in sCR1-untreated mice (P < 0.05). Respective reductions in PI in mice treated 0.5 and 1 h after acid aspiration with sCR1sLe(x) of 70 and 57% were greater than the decreases in PI of 45 and 38% observed in respective sCR1-treated groups (P < 0.05). BAL fluid neutrophil counts in sCR1sLe(x)-treated mice were significantly less than those in sCR1-treated animals, which were similar to those in untreated mice. Immunohistochemistry stained for sCR1 only on the pulmonary vascular endothelium of sCR1sLe(x)- but not sCR1-treated mice. In conclusion, sCR1sLe(x) moderates permeability by antagonizing complement activation and neutrophil adhesion. Delayed complement and neutrophil antagonism significantly reduces injury.

  6. Unique Honey Bee (Apis mellifera) Hive Component-Based Communities as Detected by a Hybrid of Phospholipid Fatty-Acid and Fatty-Acid Methyl Ester Analyses

    PubMed Central

    2015-01-01

    Microbial communities (microbiomes) are associated with almost all metazoans, including the honey bee Apis mellifera. Honey bees are social insects, maintaining complex hive systems composed of a variety of integral components including bees, comb, propolis, honey, and stored pollen. Given that the different components within hives can be physically separated and are nutritionally variable, we hypothesize that unique microbial communities may occur within the different microenvironments of honey bee colonies. To explore this hypothesis and to provide further insights into the microbiome of honey bees, we use a hybrid of fatty acid methyl ester (FAME) and phospholipid-derived fatty acid (PLFA) analysis to produce broad, lipid-based microbial community profiles of stored pollen, adults, pupae, honey, empty comb, and propolis for 11 honey bee hives. Averaging component lipid profiles by hive, we show that, in decreasing order, lipid markers representing fungi, Gram-negative bacteria, and Gram-positive bacteria have the highest relative abundances within honey bee colonies. Our lipid profiles reveal the presence of viable microbial communities in each of the six hive components sampled, with overall microbial community richness varying from lowest to highest in honey, comb, pupae, pollen, adults and propolis, respectively. Finally, microbial community lipid profiles were more similar when compared by component than by hive, location, or sampling year. Specifically, we found that individual hive components typically exhibited several dominant lipids and that these dominant lipids differ between components. Principal component and two-way clustering analyses both support significant grouping of lipids by hive component. Our findings indicate that in addition to the microbial communities present in individual workers, honey bee hives have resident microbial communities associated with different colony components. PMID:25849080

  7. Superabsorbent hydrogels via graft polymerization of acrylic acid from chitosan-cellulose hybrid and their potential in controlled release of soil nutrients.

    PubMed

    Essawy, Hisham A; Ghazy, Mohamed B M; El-Hai, Farag Abd; Mohamed, Magdy F

    2016-08-01

    Superabsorbent polymers fabricated via grafting polymerization of acrylic acid from chitosan (CTS) yields materials that suffer from poor mechanical strength. Hybridization of chitosan with cellulose (Cell) via chemical bonding using thiourea formaldehyde resin increases the flexibility of the produced hybrid (CTS/Cell). The hybridization process and post graft polymerization of acrylic acid was followed using Fourier transform infrared (FTIR). Also, the obtained structures were homogeneous and exhibited uniform surface as could be shown from imaging with scanning electron microscopy (SEM). Thus, the polymers derived from the grafting of polyacrylic acid from (CTS/Cell) gave rise to much more mechanically robust structures ((CTS/Cell)-g-PAA) that bear wide range of pH response due to presence of chitosan and polyacrylic acid in one homogeneous entity. Additionally, the obtained structures possessed greater water absorbency 390, 39.5g/g in distilled water and saline (0.9wt.% NaCl solution), respectively, and enhanced retention potential even at elevated temperatures as revealed by thermogravimetric analysis (TGA). This could be explained by the high grafting efficiency (GE%), 86.4%, and grafting yield (GY%), 750%. The new superabsorbent polymers proved to be very efficient devices for controlled release of fertilizers into the soil which expands their use in agriculture and horticultural applications. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Artificial mismatch hybridization

    DOEpatents

    Guo, Zhen; Smith, Lloyd M.

    1998-01-01

    An improved nucleic acid hybridization process is provided which employs a modified oligonucleotide and improves the ability to discriminate a control nucleic acid target from a variant nucleic acid target containing a sequence variation. The modified probe contains at least one artificial mismatch relative to the control nucleic acid target in addition to any mismatch(es) arising from the sequence variation. The invention has direct and advantageous application to numerous existing hybridization methods, including, applications that employ, for example, the Polymerase Chain Reaction, allele-specific nucleic acid sequencing methods, and diagnostic hybridization methods.

  9. A Radial Concentration Gradient of Indole-3-Acetic Acid Is Related to Secondary Xylem Development in Hybrid Aspen.

    PubMed Central

    Tuominen, H.; Puech, L.; Fink, S.; Sundberg, B.

    1997-01-01

    The radial distribution pattern of indole-3-acetic acid (IAA) was determined across the developing tissues of the cambial region in the stem of hybrid aspen (Populus tremula L. x Populus tremuloides Michx). IAA content was measured in consecutive tangential cryo-sections using a microscale mass spectrometry technique. Analysis was performed with wild-type and transgenic trees with an ectopic expression of Agrobacterium tumefaciens IAA-biosynthetic genes. In all tested trees IAA was distributed as a steep concentration gradient across the developing tissues of the cambial region. The peak level of IAA was within the cambial zone, where cell division takes place. Low levels were reached in the region where secondary wall formation was initiated. The transgenic trees displayed a lower peak level and a wider radial gradient of IAA compared with the wild type. This alteration was related to a lower rate of cambial cell division and a longer duration of xylem cell expansion in the transgenic trees, resulting in a decreased xylem production and a larger fiber lumen area. The results indicate that IAA has a role in regulating not only the rate of physiological processes such as cell division, but also the duration of developmental processes such as xylem fiber expansion, suggesting that IAA functions as a morphogen, conveying positional information during xylem development. PMID:12223825

  10. Microbial populations identified by fluorescence in situ hybridization in a constructed wetland treating acid coal mine drainage

    SciTech Connect

    Nicomrat, D.; Dick, W.A.; Tuovinen, O.H.

    2006-07-15

    Microorganisms are an integral part of the biogeochemical processes in wetlands, yet microbial communities in sediments within constructed wetlands receiving acid mine drainage (AMD) are only poorly understood. The purpose of this study was to characterize the microbial diversity and abundance in a wetland receiving AMD using fluorescence in situ hybridization (FISH) analysis. Seasonal samples of oxic surface sediments, comprised of Fe(III) precipitates, were collected from two treatment cells of the constructed wetland system. The pH of the bulk samples ranged between pH 2.1 and 3.9. Viable counts of acidophilic Fe and S oxidizers and heterotrophs were determined with a most probable number (MPN) method. The MPN counts were only a fraction of the corresponding FISH counts. The sediment samples contained microorganisms in the Bacteria (including the subgroups of acidophilic Fe- and S-oxidizing bacteria and Acidiphilium spp.) and Eukarya domains. Archaea were present in the sediment surface samples at < 0.01% of the total microbial community. The most numerous bacterial species in this wetland system was Acidithiobacillus ferrooxidans, comprising up to 37% of the bacterial population. Acidithiobacillus thiooxidans was also abundant.

  11. Hybrid Cross-Linked Lipase Aggregates with Magnetic Nanoparticles: A Robust and Recyclable Biocatalysis for the Epoxidation of Oleic Acid.

    PubMed

    Cui, Jiandong; Cui, Lili; Jia, Shiru; Su, Zhiguo; Zhang, Songping

    2016-09-28

    Highly stable and easily recyclable hybrid magnetic cross-linked lipase aggregates (HM-CSL-CLEAs) were prepared by coaggregation of lipase aggregates with nonfunctionalized magnetic nanoparticles and subsequent chemical cross-linking with glutaraldehyde. Analysis by SEM and CLSM indicated that the CLEAs were embedded in nanoparticle aggregates instead of covalently immobilized. The resulting HM-CSL-CLEAs exhibited higher thermostability, storage stability, and reusability than standard CLEAs. For example, HM-CSL-CLEAs maintained >60% of their initial activity after 40 min of incubation at 60 °C, whereas standard CLEAs lost most of their activities. The HM-CSL-CLEAs can be easily recovered from the reaction mixture by an external magnetic field. Moreover, the H2O2 tolerance of the lipase in HM-CSL-CLEAs was also enhanced, which could relieve the inhibitory effect on lipase activity. A high conversion yield (55%) for the epoxidation of oleic acid using H2O2 as oxidizing agent was achieved by HM-CSL-CLEAs.

  12. Hybrid organic/inorganic copolymers with strongly hydrogen-bond acidic properties for acoustic wave and optical sensors

    SciTech Connect

    Grate, J.W.; Kaganove, S.N.; Patrash, S.J.

    1997-05-01

    Hybrid organic/inorganic polymers have been prepared incorporating fluoroalkyl-substituted bisphenol groups linked using oligosiloxane spacers. These hydrogen-bond acidic materials have glass-to-rubber transition temperatures below room temperature and are excellent sorbents for basic vapors. The physical properties such as viscosity and refractive index can be tuned by varying the length of the oligosiloxane spacers and the molecular weight. In addition, the materials are easily cross-linked to yield solid elastomers. The potential use of these materials for chemical sensing has been demonstrated by applying them to surface acoustic wave devices as thin films and detecting the hydrogen-bond basic vapor dimethyl methylphosphonate with high sensitivity. It has also been demonstrated that one of these materials with suitable viscosity and refractive index can be used to clad silica optical fibers; the cladding was applied to freshly drawn fiber using a fiber drawing tower. These fibers have potential as evanescent wave optical fiber sensors. 38 refs., 2 figs.

  13. Anti-melanogenic effects of resveratryl triglycolate, a novel hybrid compound derived by esterification of resveratrol with glycolic acid.

    PubMed

    Park, Soojin; Seok, Jin Kyung; Kwak, Jun Yup; Choi, Yun-Hyeok; Hong, Seong Su; Suh, Hwa-Jin; Park, Woncheol; Boo, Yong Chool

    2016-07-01

    Resveratrol is known to inhibit cellular melanin synthesis by multiple mechanisms. Glycolic acid (GA) is used in skin care products for its excellent skin penetration. The purpose of this study was to examine the anti-melanogenic effects of resveratryl triglycolate (RTG), a novel hybrid compound of resveratrol and GA, in comparison with resveratrol, GA, resveratryl triacetate (RTA) and arbutin. Resveratrol, RTG, and RTA inhibited the catalytic activity human tyrosinase (TYR) more potently than arbutin or GA did. Their cytotoxic and anti-melanogenic effects were examined using murine melanoma B16/F10 cells and human epidermal melanocytes (HEMs). The cytotoxicity of RTG was similar to that of resveratrol and RTA. RTG at 3-10 μM decreased melanin levels and cellular TYR activities in α-melanocyte-stimulating hormone-stimulated B16/F10 cells, and L-tyrosine-stimulated HEMs. RTG also suppressed mRNA and protein expression of TYR, tyrosinase-related protein 1, L-3,4-dihydroxyphenylalanine chrome tautomerase, and microphthalmia-associated transcription factor (MITF) in HEMs stimulated with L-tyrosine. This study suggests that, like resveratrol and RTA, RTG can attenuate cellular melanin synthesis effectively through the suppression of MITF-dependent expression of melanogenic enzymes and the inhibition of catalytic activity of TYR enzyme. RTG therefore has potential for use as a cosmeceutical ingredient for skin whitening.

  14. Identification of antisense nucleic acid hybridization sites in mRNA molecules with self-quenching fluorescent reporter molecules.

    PubMed

    Gifford, Lida K; Opalinska, Joanna B; Jordan, David; Pattanayak, Vikram; Greenham, Paul; Kalota, Anna; Robbins, Michelle; Vernovsky, Kathy; Rodriguez, Lesbeth C; Do, Bao T; Lu, Ponzy; Gewirtz, Alan M

    2005-02-17

    We describe a physical mRNA mapping strategy employing fluorescent self-quenching reporter molecules (SQRMs) that facilitates the identification of mRNA sequence accessible for hybridization with antisense nucleic acids in vitro and in vivo, real time. SQRMs are 20-30 base oligodeoxynucleotides with 5-6 bp complementary ends to which a 5' fluorophore and 3' quenching group are attached. Alone, the SQRM complementary ends form a stem that holds the fluorophore and quencher in contact. When the SQRM forms base pairs with its target, the structure separates the fluorophore from the quencher. This event can be reported by fluorescence emission when the fluorophore is excited. The stem-loop of the SQRM suggests that SQRM be made to target natural stem-loop structures formed during mRNA synthesis. The general utility of this method is demonstrated by SQRM identification of targetable sequence within c-myb and bcl-6 mRNA. Corresponding antisense oligonucleotides reduce these gene products in cells.

  15. Locked Nucleic Acid and Flow Cytometry-Fluorescence In Situ Hybridization for the Detection of Bacterial Small Noncoding RNAs

    PubMed Central

    Robertson, Kelly L.

    2012-01-01

    We describe the development and testing of a high-throughput method that enables the detection of small noncoding RNAs (ncRNAs) from single bacterial cells using locked nucleic acid probes (LNA) and flow cytometry-fluorescence in situ hybridization (flow-FISH). The LNA flow-FISH method and quantitative reverse transcription-PCR (qRT-PCR) were used to monitor the expression of three ncRNAs (6S, CsrB, and TPP-2) in Vibrio campbellii ATCC BAA-1116 cultures during lag phase, mid-log phase, and stationary phase. Both LNA flow-FISH and qRT-PCR revealed that CsrB and TPP-2 were highly expressed during lag phase but markedly reduced in mid-log phase and stationary phase, whereas 6S demonstrated no to little expression during lag phase but increased thereafter. Importantly, while LNA flow-FISH and qRT-PCR demonstrated similar overall expression trends, only LNA flow-FISH, which enabled the detection of ncRNAs in individual cells as opposed to the lysate-based ensemble measurements generated by qRT-PCR, was able to capture the cell-to-cell heterogeneity in ncRNA expression. As such, this study demonstrates a new method that simultaneously enables the in situ detection of ncRNAs and the determination of gene expression heterogeneity within an isogenic bacterial population. PMID:22057868

  16. A potential non-invasive glioblastoma treatment: Nose-to-brain delivery of farnesylthiosalicylic acid incorporated hybrid nanoparticles.

    PubMed

    Sekerdag, Emine; Lüle, Sevda; Bozdağ Pehlivan, Sibel; Öztürk, Naile; Kara, Aslı; Kaffashi, Abbas; Vural, Imran; Işıkay, Ilkay; Yavuz, Burҫin; Oguz, Kader Karlı; Söylemezoğlu, Figen; Gürsoy-Özdemir, Yasemin; Mut, Melike

    2017-09-10

    New drug delivery systems are highly needed in research and clinical area to effectively treat gliomas by reaching a high antineoplastic drug concentration at the target site without damaging healthy tissues. Intranasal (IN) administration, an alternative route for non-invasive drug delivery to the brain, bypasses the blood-brain-barrier (BBB) and eliminates systemic side effects. This study evaluated the antitumor efficacy of farnesylthiosalicylic acid (FTA) loaded (lipid-cationic) lipid-PEG-PLGA hybrid nanoparticles (HNPs) after IN application in rats. FTA loaded HNPs were prepared, characterized and evaluated for cytotoxicity. Rat glioma 2 (RG2) cells were implanted unilaterally into the right striatum of female Wistar rats. 10days later, glioma bearing rats received either no treatment, or 5 repeated doses of 500μM freshly prepared FTA loaded HNPs via IN or intravenous (IV) application. Pre-treatment and post-treatment tumor sizes were determined with MRI. After a treatment period of 5days, IN applied FTA loaded HNPs achieved a significant decrease of 55.7% in tumor area, equal to IV applied FTA loaded HNPs. Herewith, we showed the potential utility of IN application of FTA loaded HNPs as a non-invasive approach in glioblastoma treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Identification of antisense nucleic acid hybridization sites in mRNA molecules with self-quenching fluorescent reporter molecules

    PubMed Central

    Gifford, Lida K.; Opalinska, Joanna B.; Jordan, David; Pattanayak, Vikram; Greenham, Paul; Kalota, Anna; Robbins, Michelle; Vernovsky, Kathy; Rodriguez, Lesbeth C.; Do, Bao T.; Lu, Ponzy; Gewirtz, Alan M.

    2005-01-01

    We describe a physical mRNA mapping strategy employing fluorescent self-quenching reporter molecules (SQRMs) that facilitates the identification of mRNA sequence accessible for hybridization with antisense nucleic acids in vitro and in vivo, real time. SQRMs are 20–30 base oligodeoxynucleotides with 5–6 bp complementary ends to which a 5′ fluorophore and 3′ quenching group are attached. Alone, the SQRM complementary ends form a stem that holds the fluorophore and quencher in contact. When the SQRM forms base pairs with its target, the structure separates the fluorophore from the quencher. This event can be reported by fluorescence emission when the fluorophore is excited. The stem–loop of the SQRM suggests that SQRM be made to target natural stem–loop structures formed during mRNA synthesis. The general utility of this method is demonstrated by SQRM identification of targetable sequence within c-myb and bcl-6 mRNA. Corresponding antisense oligonucleotides reduce these gene products in cells. PMID:15718294

  18. Sulfonic acid-functionalized hybrid organic-inorganic proton exchange membranes synthesized by sol-gel using 3-mercaptopropyl trimethoxysilane (MPTMS)

    NASA Astrophysics Data System (ADS)

    Mosa, J.; Durán, A.; Aparicio, M.

    2015-11-01

    Organic/inorganic hybrid membranes based on (3-glycidoxypropyl) trimethoxysilane (GPTMS) and 3-mercaptopropyl trimethoxysilane (MPTMS) have been prepared by sol-gel method and organic polymerisation, as candidate materials for proton exchange membranes in direct alcohol fuel cell (DMFC) applications. The -SH groups of MPTMS are oxidized to sulfonic acid groups, which are attributed to enhance the proton conductivity of hybrid membranes. FTIR, XPS and contact angle were used to characterize and confirm the hybrid structure and oxidation reaction progress. Membranes characterization also includes ion exchange capacity, water uptake, methanol permeability and proton conductivity to confirm their applicability in fuel cells. All the membranes were homogeneous and thermally and chemically resistant. In particular, the hybrid membranes demonstrated proton conductivities as high as 0.16 S cm-1 at high temperature, while exhibiting a low methanol permeability as compared to Nafion®. These results are associated with proton conducting paths through the silica pseudo-PEO network in which sulfonic acid groups work as proton donor.

  19. High-quality reduced graphene oxide-nanocrystalline platinum hybrid materials prepared by simultaneous co-reduction of graphene oxide and chloroplatinic acid.

    PubMed

    Wang, Yinjie; Liu, Jincheng; Liu, Lei; Sun, Darren Delai

    2011-03-21

    Reduced graphene oxide-nanocrystalline platinum (RGO-Pt) hybrid materials were synthesized by simultaneous co-reduction of graphene oxide (GO) and chloroplatinic acid with sodium citrate in water at 80°C, of pH 7 and 10. The resultant RGO-Pt hybrid materials were characterized using transmission electron microscopy (TEM), powder X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), Fourier-transform infrared spectroscopy, and thermogravimetric analysis. Platinum (Pt) nanoparticles were anchored randomly onto the reduced GO (RGO) sheets with average mean diameters of 1.76 (pH 7) and 1.93 nm (pH 10). The significant Pt diffraction peaks and the decreased intensity of (002) peak in the XRD patterns of RGO-Pt hybrid materials confirmed that the Pt nanoparticles were anchored onto the RGO sheets and intercalated into the stacked RGO layers at these two pH values. The Pt loadings for the hybrid materials were determined as 36.83 (pH 7) and 49.18% (pH 10) by mass using XPS analysis. With the assistance of oleylamine, the resultant RGO-Pt hybrid materials were soluble in the nonpolar organic solvents, and the dispersion could remain stable for several months.

  20. High-quality reduced graphene oxide-nanocrystalline platinum hybrid materials prepared by simultaneous co-reduction of graphene oxide and chloroplatinic acid

    PubMed Central

    2011-01-01

    Reduced graphene oxide-nanocrystalline platinum (RGO-Pt) hybrid materials were synthesized by simultaneous co-reduction of graphene oxide (GO) and chloroplatinic acid with sodium citrate in water at 80°C, of pH 7 and 10. The resultant RGO-Pt hybrid materials were characterized using transmission electron microscopy (TEM), powder X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), Fourier-transform infrared spectroscopy, and thermogravimetric analysis. Platinum (Pt) nanoparticles were anchored randomly onto the reduced GO (RGO) sheets with average mean diameters of 1.76 (pH 7) and 1.93 nm (pH 10). The significant Pt diffraction peaks and the decreased intensity of (002) peak in the XRD patterns of RGO-Pt hybrid materials confirmed that the Pt nanoparticles were anchored onto the RGO sheets and intercalated into the stacked RGO layers at these two pH values. The Pt loadings for the hybrid materials were determined as 36.83 (pH 7) and 49.18% (pH 10) by mass using XPS analysis. With the assistance of oleylamine, the resultant RGO-Pt hybrid materials were soluble in the nonpolar organic solvents, and the dispersion could remain stable for several months. PMID:21711745

  1. Immobilization of CdSe/ZnS quantum dots on glass beads for the detection of nucleic acid hybridization using fluorescence resonance energy transfer

    NASA Astrophysics Data System (ADS)

    Algar, W. Russ; Krull, Ulrich J.

    2011-03-01

    The photoluminescence (PL) properties of quantum dots (QD) are of significant interest in the development of new methods for bioanalysis. Multiplexed solid-phase nucleic acid hybridization assays that use immobilized QDs as donors in fluorescence resonance energy transfer (FRET) are one such example, and offer several unique advantages over other methods. In this work, new interfacial chemistry is described for the immobilization of red-emitting CdSe/ZnS QDs on glass beads for use in hybridization assays. The beads were chemically modified with a dithiolate surface ligand and the QDs immobilized via self-assembly. Further derivatization of the QDs with dithiolate-terminated probe oligonucleotides enabled a hybridization assay that could detect unlabeled target down to nanomolar levels with discrimination of single base-pair mismatches. The use of beads as an immobilization platform afforded shorter analysis times and superior reusability compared to previous studies using optical fibers. Hybridization between probe, target, and Alexa Fluor 647 (A647) labeled reporter oligonucleotides in a sandwich format generated a spectroscopic signal by introducing the proximity needed for FRET between the QDs and A647. The results indicate clear directions for the optimization of solid-phase hybridization assays, and are important for the future development of true multiplexed biosensors based on QDs and FRET.

  2. Poly(lactic acid)/chitosan hybrid nanoparticles for controlled release of anticancer drug.

    PubMed

    Wang, Wenlong; Chen, Shu; Zhang, Liang; Wu, Xi; Wang, Jiexin; Chen, Jian-Feng; Le, Yuan

    2015-01-01

    Poly(lactic acid) (PLA) is a kind of non-toxic biological materials with excellent absorbability, biocompatibility and biodegradability, which can be used for drug release, tissue engineering and surgical treatment applications. In this study, we prepared chitosan modified PLA nanoparticles as carriers for encapsulation of docetaxel by anti-solvent precipitation method. The morphology, particle size, zeta potential and composition of the PLA/chitosan were characterized by SEM, DLS, FTIR and XPS. As-prepared PLA/chitosan particles exhibited average size of 250 nm and showed very narrow distribution with polydispersity index of 0.098. Their large surface charge-ability was confirmed by zeta potential value of 53.9 mV. Docetaxel was released from PLA/chitosan nanoparticles with 40% initial burst release in 5 h and 70% cumulative release within 24 h, while from PLA nanoparticles 65% of docetaxel was released in 5h. In vitro drug release study demonstrated that PLA/chitosan nanoparticles prolonged drug release and decreased the burst release over the unmodified PLA nanoparticles. These results illustrated high potential of chitosan modified PLA nanoparticles for usage as anticancer drug carriers.

  3. Hybridization assay of insect antifreezing protein gene by novel multilayered porous silicon nucleic acid biosensor.

    PubMed

    Lv, Xiaoyi; Chen, Liangliang; Zhang, Hongyan; Mo, Jiaqing; Zhong, Furu; Lv, Changwu; Ma, Ji; Jia, Zhenhong

    2013-01-15

    A fabrication of a novel simple porous silicon polybasic photonic crystal with symmetrical structure has been reported as a nucleic acid biosensor for detecting antifreeze protein gene in insects (Microdera puntipennis dzhungarica), which would be helpful in the development of some new transgenic plants with tolerance of freezing stress. Compared to various porous silicon-based photonic configurations, porous silicon polytype layered structure is quite easy to prepare and shows more stability; moreover, polybasic photonic crystals with symmetrical structure exhibit interesting optical properties with a sharp resonance in the reflectance spectrum, giving a higher Q factor which causes higher sensitivity for sensing performance. In this experiment, DNA oligonucleotides were immobilized into the porous silicon pores using a standard crosslink chemistry method. The porous silicon polybasic symmetrical structure sensor possesses high specificity in performing controlled experiments with non-complementary DNA. The detection limit was found to be 21.3nM for DNA oligonucleotides. The fabricated multilayered porous silicon-based DNA biosensor has potential commercial applications in clinical chemistry for determination of an antifreeze protein gene or other genes.

  4. Morphology and properties of hybrid composites based on polypropylene/polylactic acid blend and bamboo fiber.

    PubMed

    Ying-Chen, Zhang; Hong-Yan, Wu; Yi-Ping, Qiu

    2010-10-01

    The design of new composites based on a polypropylene (PP)/polylactic acid (PLA) matrix and filler bamboo fiber (BF) leads to changes in process ability, morphology, and rheological properties of the raw thermoplastic. We have designed more environmentally friendly composites blended with PLA, filled with bamboo fiber. To refine dispersion of PLA and improve composite toughness, MAH-g-PP was included in the filler-matrix interface to enhance interface strength for PP, PLA and BF. The incorporation of MAH-g-PP into composites brought about beneficial changes in morphology and rheological, related with improved dispersion of PLA and increased bamboo fiber-matrix interactions. The SEM results showed the particle size of the dispersed PLA and BF phase significantly reduced with adding a litter MAH-g-PP. The XRD and DSC results showed that spread and phases in the composites preserved crystallization. The good rheological, morphological and thermal properties obtained when the ratio of PP/PLA/BF/MAH-g-PP was 48.75/13/35/3.25. Copyright © 2010 Elsevier Ltd. All rights reserved.

  5. Weathering and Biodegradation Study on Graft Copolymer Compatibilized Hybrid Bionanocomposites of Poly(Lactic Acid)

    NASA Astrophysics Data System (ADS)

    Sajna, VP; Nayak, Sanjay K.; Mohanty, Smita

    2016-07-01

    This work reports on the influence of moisture absorption and accelerated weathering on the properties of graft copolymer compatibilized bionanocomposites of poly(lactic acid) (PLA). Moisture absorption tests were conducted for 30 days by immersing the samples in a distilled water bath at room temperature, and the amount of moisture absorbed in each time interval was measured. The rate of moisture uptake decreased by incorporation of C30B nanoclay and graft copolymer into fiber-reinforced PLA composites. Changes in the mechanical properties of composites in each time interval of moisture absorption were investigated using tensile and impact tests. Exposure to moisture caused significant drops in the mechanical properties. The morphological characterization of biocomposites during the aforementioned tests has been made using SEM, while bionanocomposites were analyzed by TEM. Further, this paper also reported the effect of accelerated weathering on the mechanical properties and the results are confirmed through SEM analysis. Biodegradation behaviors of PLA biocomposites and bionanocomposites have also been studied.

  6. Preparation of hydroxyapatite/poly(lactic acid) hybrid microparticles for local drug delivery

    NASA Astrophysics Data System (ADS)

    Loca, D.; Locs, J.; Berzina-Cimdina, L.

    2013-12-01

    Calcium phosphate (CaP) bioceramic is well known as bioactive and biocompatible material in bone tissue regeneration applications. Apatitic CaP, especially nano sized hydroxyapatite (NHAp), is more similar to the natural apatite presented in the bone tissue than CaP bioceramics. In the current research NHAp was modified using biodegradable polymer - poly(lactic acid) (PLA) to develop composites providing bone regeneration and local drug delivery. NHAp/PLA microcapsules were prepared using solid-in-water-in-oil-in-water (s/w1/o/w2) encapsulation technology. The impact of primary and secondary emulsion stability on the emulsion droplet and microparticle properties was evaluated. The stability of final emulsion can be increased by varying the process parameters. Stable s/w1/o/w2 emulsion using 3ml of NHAp suspension, not less than 100ml of 4% PVA water solution and 10ml of 10% PLA solution in dichloromethane can be obtained. S/w1/o/w2 microencapuslation method can be effectively used for the preparation of multi-domain microcapsules achieving high NHAp encapsulation efficacy (93%).

  7. Hybrid porous tin(IV) phosphonate: an efficient catalyst for adipic acid synthesis and a very good adsorbent for CO2 uptake.

    PubMed

    Dutta, Arghya; Pramanik, Malay; Patra, Astam K; Nandi, Mahasweta; Uyama, Hiroshi; Bhaumik, Asim

    2012-07-07

    A new porous organic-inorganic hybrid tin phosphonate material has been synthesized hydrothermally, which shows a Brunauer-Emmett-Teller surface area of 723 m(2) g(-1) and it adsorbs 4.8 mmol g(-1) CO(2) at 273 K and 5 bar pressure. The material also shows remarkable catalytic activity in one-pot liquid phase oxidation of cyclohexanone to adipic acid under eco-friendly conditions.

  8. PDMS microchip coated with polydopamine/gold nanoparticles hybrid for efficient electrophoresis separation of amino acids.

    PubMed

    Liang, Ru-Ping; Meng, Xiang-Ying; Liu, Chun-Ming; Qiu, Jian-Ding

    2011-11-01

    In this paper, a novel, simple, economical and environmentally friendly method based on in situ chemically induced synthesis strategy was designed and developed for the modification of a poly(dimethylsiloxane) (PDMS) microchip channel with polydopamine/gold nanoparticles (PDA/Au NPs) to create a hydrophilic and biofouling resistant surface. Dopamine as a reductant and a monomer, and HAuCl(4) as an oxidant to trigger dopamine polymerization and the source of metallic nanoparticles, were filled into the PDMS microchannel to yield in situ a well-distributed and robust PDA/Au NP coating. Au NPs were highly and uniformly dispersed in/on the PDA matrix with a narrow size distribution, as verified by scanning electron microscopy and UV-vis spectra. Compared with the native PDMS microchannel, the modified surfaces exhibited much better wettability, high stability and suppressed electroosmotic mobility, and less nonspecific adsorption towards biomolecules. The water contact angle and EOF of PDA/Au NP-coated PDMS microchip were measured to be 13° and 4.17×10(-4) cm(2)/V s, compared to those of 111° and 5.33×10(-4) cm(2)/V s from the native one, respectively. Fast and efficient separations of five amino acids such as arginine, proline, histidine, valine and threonine suggested greatly improved electrophoretic performance of the PDA/Au NP-functionalized PDMS microchips. This one-step procedure offers an effective approach for a biomimetic surface design on microfluidic chips, which is promising in high-throughput and complex biological analysis.

  9. Hybridization between periodic mesoporous organosilica and a Ru(II) polypyridyl complex with phosphonic acid anchor groups.

    PubMed

    Yui, Tatsuto; Takeda, Hiroyuki; Ueda, Yutaro; Sekizawa, Keita; Koike, Kazuhide; Inagaki, Shinji; Ishitani, Osamu

    2014-02-12

    A new method for the hybridization of a ruthenium(II) polypyridyl complex ([Ru(bpy)2((CH2PO3H2)2-bpy)](2+) (RuP2(2+): bpy =2,2'-bipyridine; (CH2PO3H2)2-bpy =2,2'-bipyridine-4,4'di(metylphosphonic acid)) with biphenylene-bearing periodic mesoporous organosilica (Bp-PMO made from 4,4'bis(triethoxysilyl)biphenyl [(C2H5O)3Si-(C6H4)2-Si(OC2H5)3]) was developed. Efficient and secure fixation of the ruthenium(II) complex with methylphosphonic acid groups (RuP2(2+)) in the mesopores of Bp-PMO occurred. This method introduced up to 660 μmol of RuP2(2+) in 1 g of Bp-PMO. Two modes of adsorption of RuP2(2+) in the mesopores of Bp-PMO were observed: one is caused by the chemical interaction between the methylphosphonic acid groups of RuP2(2+) and the silicate moieties of Bp-PMO and the other is attributed to aggregation of the RuP2(2+) complexes. In the case of the former mode, adsorbed RuP2(2+) (up to 80-100 μmol g(-1)) did not detach from Bp-PMO after washing with acetonitrile, dimethylformamide, or even water. Emission from the excited biphenylene (Bp) units was quantitatively quenched by the adsorbed RuP2(2+) molecules in cases where more than 60 μmol g(-1) of RuP2(2+) was adsorbed, and emission from RuP2(2+) was observed. Quantitative emission measurements indicated that emission from approximately 100 Bp units can be completely quenched by only one RuP2(2+) molecule in the mesopore, and photons absorbed by approximately 400 Bp units are potentially accumulated in one RuP2(2+) molecule.

  10. Open L-lactic acid fermentation of food refuse using thermophilic Bacillus coagulans and fluorescence in situ hybridization analysis of microflora.

    PubMed

    Sakai, Kenji; Ezaki, Yutaka

    2006-06-01

    In the production of commercially useful poly-L-lactic acid plastic from biomass wastes, a feasible fermentation process to produce optically active L-lactic acid would be required. Here, model kitchen refuse (MKR) was inoculated with Bacillus coagulans NBRC12583 under nonsterilized openculture conditions. At temperatures below 45 degrees C, a racemic mixture of D- and L-lactic acids was accumulated, whereas only L-lactic acid was selectively accumulated by incubation at 50-65 degrees C. At 45 degrees C, the results of fermentation could not be consistently reproduced. To analyze microflora in this type of mixed culture system, whole-cell fluorescence in situ hybridization (FISH) using 16S rRNA-targeted oligonucleotide probes for B. coagulans, Bcoa191, and LAC722(L), a group-specific probe for a wide range of mesophilic lactic acid bacteria was applied. The dominancy of mesophilic lactic acid bacteria at lower temperatures, and that of B. coagulans at higher temperatures were confirmed. By using a saccharified liquid of collected kitchen refuse, 86 g/l of L-lactic acid was accumulated under nonsterile conditions by a 5-d incubation at 55 degrees C, pH 6.5, with 53% carbon yield and 97% optical purity. To conclude, high temperature open lactic acid fermentation is a simple and promising method for producing high-grade L-lactic acid from biomass waste, and FISH analysis of such mixed-culture systems is helpful for monitoring the microflora in these cultures.

  11. Corrosion resistance of siloxane-poly(methyl methacrylate) hybrid films modified with acetic acid on tin plate substrates: Influence of tetraethoxysilane addition

    NASA Astrophysics Data System (ADS)

    Kunst, S. R.; Cardoso, H. R. P.; Oliveira, C. T.; Santana, J. A.; Sarmento, V. H. V.; Muller, I. L.; Malfatti, C. F.

    2014-04-01

    The aim of this paper is to study the corrosion resistance of hybrid films. Tin plate was coated with a siloxane-poly (methyl methacrylate) (PMMA) hybrid film prepared by sol-gel route with covalent bonds between the organic (PMMA) and inorganic (siloxane) phases obtained by hydrolysis and polycondensation of 3-(trimethoxysilylpropyl) methacrylate (TMSM) and polymerization of methyl methacrylate (MMA) using benzoyl peroxide (BPO) as a thermic initiator. Hydrolysis reactions were catalyzed by acetic acid solution avoiding the use of chlorine or stronger acids in the film preparation. The effect of the addition of tetraethoxysilane (TEOS) on the protective properties of the film was evaluated. The hydrophobicity of the film was determined by contact angle measurements, and the morphology was evaluated by scanning electron microscopy (SEM) and profilometry. The local nanostructure was investigated by Fourier transform infrared spectroscopy (FT-IR). The electrochemical behavior of the films was assessed by open circuit potential monitoring, potentiodynamic polarization and electrochemical impedance spectroscopy (EIS) measurements in a 0.05 M NaCl solution. The mechanical behavior was evaluated by tribology. The results highlighted that the siloxane-PMMA hybrid films modified with acetic acid are promising anti-corrosive coatings that acts as an efficient diffusion barrier, protecting tin plates against corrosion. However, the coating properties were affected by the TEOS addition, which contributed for the thickness increase and irregular surface coverage.

  12. Preparation of organic-inorganic hybrid silica monolith with octyl and sulfonic acid groups for capillary electrochromatograhpy and application in determination of theophylline and caffeine in beverage.

    PubMed

    Chen, Ming-Luan; Zheng, Ming-Ming; Feng, Yu-Qi

    2010-05-21

    An organic-inorganic hybrid silica monolithic column with octyl and sulfonic acid groups has been prepared by sol-gel technique for capillary electrochromatograhpy. The structure of hybrid monolith was optimized by changing the composition of tetraethoxysilane (TEOS), octyltriethoxysilane (C(8)-TEOS) and 3-mercaptopropyltrimethoxysilane (MPTMS) in the mixture of precursors. Then, the obtained hybrid monolith was oxidized using hydrogen peroxide (30%, w/w) to yield sulfonic acid groups. The sulfonic acid group, which served as strong cation-exchanger, dominated the charge on the surface of the capillary column and generated stable electroosmotic flow (EOF) in a wide range of pH. The monolithic column was characterized by scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy and elemental analysis (EA), and the performance of column was evaluated in detail by separating different kinds of compounds with column efficiency up to 155,000 plates/m for thiourea. In addition, this monolithic column was also applied in the analysis of theophylline (TP) and caffeine (CA) in beverages. The detection limits were 0.39 and 0.48 microg/mL for theophylline and caffeine, respectively. The method reproducibility was tested by evaluating the intra- and inter-day precisions, and relative standard deviations of less than 3.9 and 8.4%, respectively, were obtained. Recoveries of compounds from spiked beverage samples ranged from 87.2 to 105.2%. 2010 Elsevier B.V. All rights reserved.

  13. Helices with additional H-bonds: crystallographic conformations of α,γ-hybrid peptides helices composed of β-hydroxy γ-amino acids (statines).

    PubMed

    Malik, Ankita; Kumar, Mothukuri Ganesh; Bandyopadhyay, Anupam; Gopi, Hosahudya N

    2017-01-01

    β-Hydroxy-γ-amino acids (Statines) are a class of naturally occurring non-ribosomal amino acids frequently found in many peptide natural products. Peptidomimetics constituted with statines have been used as inhibitors for various aspartic acid proteases. In contrast to the synthetic γ-amino acids, very little is known about the folding behavior of these naturally occurring β-hydroxy γ-amino acids. To understand the folding behavior of statines, three α,γ-hybrid peptides P1 (Ac-Aib-γPhe-Aib-(R, S)Phesta-Aib-γPhe-Aib-CONH2 ), P2 (Ac-Aib-γPhe-Aib-(S, S)Phesta-Aib-γPhe-Aib-CONH2 ), and P3 (Ac-Aib-γPhe-Aib-(S, S)Phesta-Aib-(S, S)Phesta-Aib-CONH2 ) were synthesized on solid phase and their helical conformations in single crystals were studied. Results suggest that both syn and anti diastereoisomers of statines can be accommodated into the helix without deviating overall helical conformation of α,γ-hybrid peptides. In comparison with syn diastereoisomer, the anti diastereoisomer was found to be directly involved in the intramolecular H-bonding with the backbone carbonyl groups (i to i + 3) similar to the backbone amide NHs in the helix.

  14. Metal–organic hybrid materials built with tetrachlorophthalate acid and different N-donor coligands: Structure diversity and photoluminescence

    SciTech Connect

    Xiao, Zhenyu; Yang, Xiao; Zhao, Siwei; Wang, Debao Yang, Yu; Wang, Lei

    2016-02-15

    Eight new metal–organic hybrid materials, namely {Cd(Tcph)(4,4′-bipy)_1_/_2} (1), {[Cd_2(Tcph)_2(1,4-bimb)_1_/_2(H_2O)_4]·H_2O} (2), {Cd_2(Tcph)_2(1,4-bmimb)_1_/_2(H_2O)_4} (3), {Cd(Tcph)(1,2-bmimb)} (4), {Cu(Tcph)(1,4-bimb)(H_2O)} (5), {[Co(Tcph)(1,4-bimb)_1_/_2(H_2O)_3]·(H_2O)} (6), {Zn(Tcph)(1,2-bimb)} (7), {Cu_2(Tcph)_2(1,2-bimb)(H_2O)_4} (8), where Tcph=tetrachlorophthalate acid, 4,4′-bipy=4,4′-bipyridine, 1,4-bimb=1,4-bis(imidazol-1-ylmethyl)benzene, 1,4-bmimb=1,4-bis(2-methylimidazol-1-ylmethyl)benzene, 1,2-bimb=1,2-bis(imidazol-1-ylmethyl)-benzene, 1,2-bmimb=1,2-bis(2-methylimidazol-1-ylmethyl)benzene, have been synthesized and characterized. Their structures are determined by single crystal X-ray diffraction and further characterized by infrared spectra (IR) and thermogravimetric (TG) analyses. Complex 1, 4 and 7 display 2D layer structures. 1 possesses two-dimensional sheet containing an unusual [Cd(Tcph)] chains linked by 4,4′-bipy co-ligand, while 4 and 7 hold the similar 4-connected 4{sup 4}-sql nets. Complex 2 and 3 feature a similar three dimensional (3D) internal compensation structure with a topology of {4"2·6"3·8}{sub 2}{6"3}. 5 is a novel 2-fold self-penetrating 3D network with 4-coordinated 6{sup 5}·8–CdSO{sub 4} subnets. The ladder-like chains of 6 are further connected through O–H···O interactions to yield a 3D supramolecular structure. 8 is a discrete tetranuclear complex. The thermal stabilities of 1–8 and the luminescent properties of 1–4 and 7 in the solid state are also discussed. - Graphical abstract: Structure diversity and photoluminescence of eight new metal–organic hybrid materials constructed by Tetrachlorophthalate acid and different N-donor coligands are discussed in the context. - Highlights: • Eight new coordination polymers were synthesized based on mix-ligand strategy. • Complex 2 and 3 feature a similar 3D structure with {4"2·6"3·8}{sub 2}{6"3} topology. • Seven coordination modes of

  15. Excellent anti-corrosive pretreatment layer on iron substrate based on three-dimensional porous phytic acid/silane hybrid

    NASA Astrophysics Data System (ADS)

    Gao, Xiang; Lu, Ke; Xu, Lei; Xu, Hua; Lu, Haifeng; Gao, Feng; Hou, Shifeng; Ma, Houyi

    2016-01-01

    A novel, highly effective and environmentally friendly film-forming material, phytic acid (PA)/silane (denoted as PAS) hybrid with a three-dimensional (3D) network structure, was prepared through a condensation reaction of PA with methyltrihydroxysilane generated from the hydrolysis of methyltriethoxysilane (MTES). Two kinds of PAS-based pretreatment layers, namely NaBrO3-free and NaBrO3-doped PAS layers, were fabricated on iron substrates using the dip-coating method. SEM and AFM observations showed that the as-fabricated PAS-based layers possessed a 3D porous microstructure at the nanoscale and a rough surface morphology. X-ray photoelectron spectroscopic (XPS) and attenuated total reflection infrared (ATR-IR) spectroscopic characterization demonstrated that the above PAS layers bound to the iron surface via the -P-O- bond. Moreover, analyses of steady-state polarization curves and electrochemical impedance spectroscopic (EIS) data indicated that the corrosion rates of the iron substrates decreased considerably in the presence of the two PAS-based pretreatment layers. In particular, the NaBrO3-dosed PAS layer displayed the better corrosion resistance ability as well as maintaining the original microstructure and surface morphology. The PAS-based pretreatment layers are expected to act as substitutes for chromate and phosphate conversion layers and will find widespread application in the surface pretreatment of iron and steel materials due to the advantages of being environmentally friendly, the rapid film-forming process, and, especially, the nanoporous microstructure and rough surface morphology.A novel, highly effective and environmentally friendly film-forming material, phytic acid (PA)/silane (denoted as PAS) hybrid with a three-dimensional (3D) network structure, was prepared through a condensation reaction of PA with methyltrihydroxysilane generated from the hydrolysis of methyltriethoxysilane (MTES). Two kinds of PAS-based pretreatment layers, namely Na

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

    PubMed

    Noor, M Omair; Tavares, Anthony J; Krull, Ulrich J

    2013-07-25

    A microfluidic based solid-phase assay for the multiplexed detection of nucleic acid hybridization using quantum dot (QD) mediated fluorescence resonance energy transfer (FRET) is described herein. The glass surface of hybrid glass-polydimethylsiloxane (PDMS) microfluidic channels was chemically modified to assemble the biorecognition interface. Multiplexing was demonstrated using a detection system that was comprised of two colors of immobilized semi-conductor QDs and two different oligonucleotide probe sequences. Green-emitting and red-emitting QDs were paired with Cy3 and Alexa Fluor 647 (A647) labeled oligonucleotides, respectively. The QDs served as energy donors for the transduction of dye labeled oligonucleotide targets. The in-channel assembly of the biorecognition interface and the subsequent introduction of oligonucleotide targets was accomplished within minutes using a combination of electroosmotic flow and electrophoretic force. The concurrent quantification of femtomole quantities of two target sequences was possible by measuring the spatial coverage of FRET sensitized emission along the length of the channel. In previous reports, multiplexed QD-FRET hybridization assays that employed a ratiometric method for quantification had challenges associated with lower analytical sensitivity arising from both donor and acceptor dilution that resulted in reduced energy transfer pathways as compared to single-color hybridization assays. Herein, a spatial method for quantification that is based on in-channel QD-FRET profiles provided higher analytical sensitivity in the multiplexed assay format as compared to single-color hybridization assays. The selectivity of the multiplexed hybridization assays was demonstrated by discrimination between a fully-complementary sequence and a 3 base pair sequence at a contrast ratio of 8 to 1.

  17. Mechanism of thermal renaturation and hybridization of nucleic acids: Kramers' process and universality in Watson-Crick base pairing.

    PubMed

    Sikorav, Jean-Louis; Orland, Henri; Braslau, Alan

    2009-03-26

    Renaturation and hybridization reactions lead to the pairing of complementary single-stranded nucleic acids. We present here a theoretical investigation of the mechanism of these reactions in vitro under thermal conditions (dilute solutions of single-stranded chains, in the presence of molar concentrations of monovalent salts and at elevated temperatures). The mechanism follows a Kramers' process, whereby the complementary chains overcome a potential barrier through Brownian motion. The barrier originates from a single rate-limiting nucleation event in which the first complementary base pairs are formed. The reaction then proceeds through a fast growth of the double helix. For the DNA of bacteriophages T7, T4, and phiX174, as well as for Escherichia coli DNA, the bimolecular rate k2 of the reaction increases as a power law of the average degree of polymerization of the reacting single-strands: k2 is proportional to alpha. This relationship holds for 100 < or = < or = 50,000 with an experimentally determined exponent alpha = 0.51 +/- 0.01. The length dependence results from a thermodynamic excluded-volume effect. The reacting single-stranded chains are predicted to be in universal good solvent conditions, and the scaling law is determined by the relevant equilibrium monomer contact probability. The value theoretically predicted for the exponent is alpha = 1 - nutheta2, where nu is Flory's swelling exponent (nu approximately equal 0.588), and theta2 is a critical exponent introduced by des Cloizeaux (theta2 approximately equal 0.82), yielding alpha = 0.52 +/- 0.01, in agreement with the experimental results.

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

    PubMed

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

    2017-01-01

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

  19. Identification of differentially expressed genes in SHSY5Y cells exposed to okadaic acid by suppression subtractive hybridization

    PubMed Central

    2012-01-01

    Background Okadaic acid (OA), a toxin produced by several dinoflagellate species is responsible for frequent food poisonings associated to shellfish consumption. Although several studies have documented the OA effects on different processes such as cell transformation, apoptosis, DNA repair or embryogenesis, the molecular mechanistic basis for these and other effects is not completely understood and the number of controversial data on OA is increasing in the literature. Results In this study, we used suppression subtractive hybridization in SHSY5Y cells to identify genes that are differentially expressed after OA exposure for different times (3, 24 and 48 h). A total of 247 subtracted clones which shared high homology with known genes were isolated. Among these, 5 specific genes associated with cytoskeleton and neurotransmission processes (NEFM, TUBB, SEPT7, SYT4 and NPY) were selected to confirm their expression levels by real-time PCR. Significant down-regulation of these genes was obtained at the short term (3 and 24 h OA exposure), excepting for NEFM, but their expression was similar to the controls at 48 h. Conclusions From all the obtained genes, 114 genes were up-regulated and 133 were down-regulated. Based on the NCBI GenBank and Gene Ontology databases, most of these genes are involved in relevant cell functions such as metabolism, transport, translation, signal transduction and cell cycle. After quantitative PCR analysis, the observed underexpression of the selected genes could underlie the previously reported OA-induced cytoskeleton disruption, neurotransmission alterations and in vivo neurotoxic effects. The basal expression levels obtained at 48 h suggested that surviving cells were able to recover from OA-caused gene expression alterations. PMID:22284234

  20. Impact of a rapid peptide nucleic acid fluorescence in situ hybridization assay on treatment of Candida infections.

    PubMed

    Heil, Emily L; Daniels, Lindsay M; Long, Dustin M; Rodino, Kyle G; Weber, David J; Miller, Melissa B

    2012-11-01

    The impact of a rapid peptide nucleic acid fluorescence in situ hybridization (PNA FISH) assay with an antimicrobial stewardship intervention on the treatment of Candida infections was studied. The utility of implementing the PNA FISH assay with an antimicrobial stewardship intervention in hospitalized patients with candidemia was evaluated by measuring the median time to Candida species identification, time to targeted therapy, and clinical outcomes, including time to culture clearance, hospital length of stay, and hospital mortality. Secondary objectives included determining the cost-effectiveness of the PNA FISH assay by assessing estimated antifungal drug costs (as average wholesale price) before (June 26, 2009-September 19, 2010) and after (September 20, 2010-June 13, 2011) test implementation and confirming test accuracy. For both groups, laboratory personnel notified the physician of the results of Gram's stain from blood culture. Time to targeted therapy significantly decreased after the implementation of the PNA FISH assay (p = 0.0016). The postimplementation group had a higher rate of culture clearance (p = 0.01). Median time to species identification was 0.2 day with the PNA FISH test versus 4 days with routine methods (p < 0.001). Accounting for the cost of the test itself and the cases in which patients were switched to more-expensive therapy on the basis of the test, we estimated that the PNA FISH test resulted in savings of approximately $415 per patient. Implementing a PNA FISH test to identify Candida species from yeast-positive blood cultures in conjunction with a pharmacy-driven antimicrobial stewardship protocol decreased the time to targeted antifungal therapy and the time to culture clearance.

  1. Identification of differentially expressed genes in SHSY5Y cells exposed to okadaic acid by suppression subtractive hybridization.

    PubMed

    Valdiglesias, Vanessa; Fernández-Tajes, Juan; Pásaro, Eduardo; Méndez, Josefina; Laffon, Blanca

    2012-01-27

    Okadaic acid (OA), a toxin produced by several dinoflagellate species is responsible for frequent food poisonings associated to shellfish consumption. Although several studies have documented the OA effects on different processes such as cell transformation, apoptosis, DNA repair or embryogenesis, the molecular mechanistic basis for these and other effects is not completely understood and the number of controversial data on OA is increasing in the literature. In this study, we used suppression subtractive hybridization in SHSY5Y cells to identify genes that are differentially expressed after OA exposure for different times (3, 24 and 48 h). A total of 247 subtracted clones which shared high homology with known genes were isolated. Among these, 5 specific genes associated with cytoskeleton and neurotransmission processes (NEFM, TUBB, SEPT7, SYT4 and NPY) were selected to confirm their expression levels by real-time PCR. Significant down-regulation of these genes was obtained at the short term (3 and 24 h OA exposure), excepting for NEFM, but their expression was similar to the controls at 48 h. From all the obtained genes, 114 genes were up-regulated and 133 were down-regulated. Based on the NCBI GenBank and Gene Ontology databases, most of these genes are involved in relevant cell functions such as metabolism, transport, translation, signal transduction and cell cycle. After quantitative PCR analysis, the observed underexpression of the selected genes could underlie the previously reported OA-induced cytoskeleton disruption, neurotransmission alterations and in vivo neurotoxic effects. The basal expression levels obtained at 48 h suggested that surviving cells were able to recover from OA-caused gene expression alterations.

  2. Fluorescence in situ Hybridization method using Peptide Nucleic Acid probes for rapid detection of Lactobacillus and Gardnerella spp.

    PubMed Central

    2013-01-01

    Background Bacterial vaginosis (BV) is a common vaginal infection occurring in women of reproductive age. It is widely accepted that the microbial switch from normal microflora to BV is characterized by a decrease in vaginal colonization by Lactobacillus species together with an increase of Gardnerella vaginalis and other anaerobes. Our goal was to develop and optimize a novel Peptide Nucleic Acid (PNA) Fluorescence in situ Hybridization assay (PNA FISH) for the detection of Lactobacillus spp. and G. vaginalis in mixed samples. Results Therefore, we evaluated and validated two specific PNA probes by using 36 representative Lactobacillus strains, 22 representative G. vaginalis strains and 27 other taxonomically related or pathogenic bacterial strains commonly found in vaginal samples. The probes were also tested at different concentrations of G. vaginalis and Lactobacillus species in vitro, in the presence of a HeLa cell line. Specificity and sensitivity of the PNA probes were found to be 98.0% (95% confidence interval (CI), from 87.8 to 99.9%) and 100% (95% CI, from 88.0 to 100.0%), for Lactobacillus spp.; and 100% (95% CI, from 92.8 to 100%) and 100% (95% CI, from 81.5 to 100.0%) for G. vaginalis. Moreover, the probes were evaluated in mixed samples mimicking women with BV or normal vaginal microflora, demonstrating efficiency and applicability of our PNA FISH. Conclusions This quick method accurately detects Lactobacillus spp. and G. vaginalis species in mixed samples, thus enabling efficient evaluation of the two bacterial groups, most frequently encountered in the vagina. PMID:23586331

  3. A universal colorimetry for nucleic acids and aptamer-specific ligands detection based on DNA hybridization amplification.

    PubMed

    Li, Shuang; Shang, Xinxin; Liu, Jia; Wang, Yujie; Guo, Yingshu; You, Jinmao

    2017-07-01

    We present a universal amplified-colorimetric for detecting nucleic acid targets or aptamer-specific ligand targets based on gold nanoparticle-DNA (GNP-DNA) hybridization chain reaction (HCR). The universal arrays consisted of capture probe and hairpin DNA-GNP. First, capture probe recognized target specificity and released the initiator sequence. Then dispersed hairpin DNA modified GNPs were cross-linked to form aggregates through HCR events triggered by initiator sequence. As the aggregates accumulate, a significant red-to purple color change can be easily visualized by the naked eye. We used miRNA target sequence (miRNA-203) and aptamer-specific ligand (ATP) as target molecules for this proof-of-concept experiment. Initiator sequence (DNA2) was released from the capture probe (MNP/DNA1/2 conjugates) under the strong competitiveness of miRNA-203. Hairpin DNA (H1 and H2) can be complementary with the help of initiator DNA2 to form GNP-H1/GNP-H2 aggregates. The absorption ratio (A620/A520) values of solutions were a sensitive function of miRNA-203 concentration covering from 1.0 × 10(-11) M to 9.0 × 10(-10) M, and as low as 1.0 × 10(-11) M could be detected. At the same time, the color changed from light wine red to purple and then to light blue have occurred in the solution. For ATP, initiator sequence (5'-end of DNA3) was released from the capture probe (DNA3) under the strong combination of aptamer-ATP. The present colorimetric for specific detection of ATP exhibited good sensitivity and 1.0 × 10(-8) M ATP could be detected. The proposed strategy also showed good performances for qualitative analysis and quantitative analysis of intracellular nucleic acids and aptamer-specific ligands. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Sensitive electrochemical detection of telomerase activity using spherical nucleic acids gold nanoparticles triggered mimic-hybridization chain reaction enzyme-free dual signal amplification.

    PubMed

    Wang, Wen-Jing; Li, Jing-Jing; Rui, Kai; Gai, Pan-Pan; Zhang, Jian-Rong; Zhu, Jun-Jie

    2015-03-03

    We report an electrochemical sensor for telomerase activity detection based on spherical nucleic acids gold nanoparticles (SNAs AuNPs) triggered mimic-hybridization chain reaction (mimic-HCR) enzyme-free dual signal amplification. In the detection strategy, SNAs AuNPs and two hairpin probes were employed. SNAs AuNPs as the primary amplification element, not only hybridized with the telomeric repeats on the electrode to amplify signal but also initiated the subsequent secondary amplification, mimic-hybridization chain reaction of two hairpin probes. If the cells' extracts were positive for telomerase activity, SNAs AuNPs could be captured on the electrode. The carried initiators could trigger an alternative hybridization reaction of two hairpin probes that yielded nicked double helices. The signal was further amplified enzyme-free by numerous hexaammineruthenium(III) chloride ([Ru(NH3)6](3+), RuHex) inserting into double-helix DNA long chain by electrostatic interaction, each of which could generate an electrochemical signal at appropriate potential. With this method, a detection limit of down to 2 HeLa cells and a dynamic range of 10-10,000 cells were achieved. Telomerase activities of different cell lines were also successfully evaluated.

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

    PubMed Central

    Algar, W. Russ; Krull, Ulrich J.

    2011-01-01

    The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling the regeneration and reuse of solid-phase QD-FRET hybridization assays. FRET-sensitized emission from acceptor dyes associated with hybridization events at immobilized QD donors provides the analytical signal in these assays. The minimization of active sensing area reduces background from QD donor PL and allows the resolution of smaller amounts of acceptor emission, thus lowering the LOD. The association of multiple acceptor dyes with each hybridization event can enhance FRET efficiency, thereby improving sensitivity. Many previous studies have used interfacial protein layers to generate selectivity; however, transient destabilization of these layers is shown to prevent efficient regeneration. To this end, we report a protein-free interfacial chemistry and demonstrate the specific detection of as little as 2 pmol of target, as well as an improved capacity for regeneration. PMID:22163951

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

    PubMed Central

    Zhou, Feng; Noor, M. Omair; Krull, Ulrich J.

    2015-01-01

    Bioassays based on cellulose paper substrates are gaining increasing popularity for the development of field portable and low-cost diagnostic applications. Herein, we report a paper-based nucleic acid hybridization assay using immobilized upconversion nanoparticles (UCNPs) as donors in luminescence resonance energy transfer (LRET). UCNPs with intense green emission served as donors with Cy3 dye as the acceptor. The avidin functionalized UCNPs were immobilized on cellulose paper and subsequently bioconjugated to biotinylated oligonucleotide probes. Introduction of unlabeled oligonucleotide targets resulted in a formation of probe-target duplexes. A subsequent hybridization of Cy3 labeled reporter with the remaining single stranded portion of target brought the Cy3 dye in close proximity to the UCNPs to trigger a LRET-sensitized emission from the acceptor dye. The hybridization assays provided a limit of detection (LOD) of 146.0 fmol and exhibited selectivity for one base pair mismatch discrimination. The assay was functional even in undiluted serum samples. This work embodies important progress in developing DNA hybridization assays on paper. Detection of unlabeled targets is achieved using UCNPs as LRET donors, with minimization of background signal from paper substrates owing to the implementation of low energy near-infrared (NIR) excitation. PMID:28347081

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

    PubMed

    Algar, W Russ; Krull, Ulrich J

    2011-01-01

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

  8. N-Terminal Fatty Acid Substitution Increases the Leishmanicidal Activity of CA(1-7)M(2-9), a Cecropin-Melittin Hybrid Peptide

    PubMed Central

    Chicharro, Cristina; Granata, Cesare; Lozano, Rosario; Andreu, David; Rivas, Luis

    2001-01-01

    In order to improve the leishmanicidal activity of the synthetic cecropin A-melittin hybrid peptide CA(1-7)M(2-9) (KWKLFKKIGAVLKVL-NH2), a systematic study of its acylation with saturated linear fatty acids was carried out. Acylation of the Nɛ-7 lysine residue led to a drastic decrease in leishmanicidal activity, whereas acylation at lysine 1, in either the α or the ɛ NH2 group, increased up to 3 times the activity of the peptide against promastigotes and increased up to 15 times the activity of the peptide against amastigotes. Leishmanicidal activity increased with the length of the fatty acid chain, reaching a maximum for the lauroyl analogue (12 carbons). According to the fast kinetics, dissipation of membrane potential, and parasite membrane permeability to the nucleic acid binding probe SYTOX green, the lethal mechanism was directly related to plasma membrane permeabilization. PMID:11502512

  9. A new microcolumn-type microchip for examining the expression of chimeric fusion genes using a nucleic acid sandwich hybridization technique.

    PubMed

    Ohnishi, Michihiro; Sasaki, Naoyuki; Kishimoto, Takuya; Watanabe, Hidetoshi; Takagi, Masatoshi; Mizutani, Shuki; Kishii, Noriyuki; Yasuda, Akio

    2014-11-01

    We report a new type of microcolumn installed in a microchip. The architecture allows use of a nucleic acid sandwich hybridization technique to detect a messenger RNA (mRNA) chain as a target. Data are presented that demonstrate that the expression of a chimeric fusion gene can be detected. The microcolumn was filled with semi-transparent microbeads made of agarose gel that acted as carriers, allowing increased efficiency of the optical detection of fluorescence from the microcolumn. The hybrid between the target trapped on the microbeads and a probe DNA labeled with a fluorescent dye was detected by measuring the intensity of the fluorescence from the microcolumn directly. These results demonstrate an easy and simple method for determining the expression of chimeric fusion genes with no preamplification. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Hybrid polymeric hydrogels for ocular drug delivery: nanoparticulate systems from copolymers of acrylic acid-functionalized chitosan and N-isopropylacrylamide or 2-hydroxyethyl methacrylate.

    PubMed

    Barbu, Eugen; Verestiuc, Liliana; Iancu, Mihaela; Jatariu, Anca; Lungu, Adriana; Tsibouklis, John

    2009-06-03

    Nanoparticulate hybrid polymeric hydrogels (10-70 nm) have been obtained via the radical-induced co-polymerization of acrylic acid-functionalized chitosan with either N-isopropylacrylamide or 2-hydroxyethyl methacrylate, and the materials have been investigated for their ability to act as controlled release vehicles in ophthalmic drug delivery. Studies on the effects of network structure upon swelling properties, adhesiveness to substrates that mimic mucosal surfaces and biodegradability, coupled with in vitro drug release investigations employing ophthalmic drugs with differing aqueous solubilities, have identified nanoparticle compositions for each of the candidate drug molecules. The hybrid nanoparticles combine the temperature sensitivity of N-isopropylacrylamide or the good swelling characteristics of 2-hydroxyethyl methacrylate with the susceptibility of chitosan to lysozyme-induced biodegradation.

  11. Hybrid polymeric hydrogels for ocular drug delivery: nanoparticulate systems from copolymers of acrylic acid-functionalized chitosan and N-isopropylacrylamide or 2-hydroxyethyl methacrylate

    NASA Astrophysics Data System (ADS)

    Barbu, Eugen; Verestiuc, Liliana; Iancu, Mihaela; Jatariu, Anca; Lungu, Adriana; Tsibouklis, John

    2009-06-01

    Nanoparticulate hybrid polymeric hydrogels (10-70 nm) have been obtained via the radical-induced co-polymerization of acrylic acid-functionalized chitosan with either N-isopropylacrylamide or 2-hydroxyethyl methacrylate, and the materials have been investigated for their ability to act as controlled release vehicles in ophthalmic drug delivery. Studies on the effects of network structure upon swelling properties, adhesiveness to substrates that mimic mucosal surfaces and biodegradability, coupled with in vitro drug release investigations employing ophthalmic drugs with differing aqueous solubilities, have identified nanoparticle compositions for each of the candidate drug molecules. The hybrid nanoparticles combine the temperature sensitivity of N-isopropylacrylamide or the good swelling characteristics of 2-hydroxyethyl methacrylate with the susceptibility of chitosan to lysozyme-induced biodegradation.

  12. Ionic Liquid-Hybrid Molecularly Imprinted Material-Filter Solid-Phase Extraction Coupled with HPLC for Determination of 6-Benzyladenine and 4-Chlorophenoxyacetic Acid in Bean Sprouts.

    PubMed

    Han, Yehong; Yang, Chunliu; Zhou, Yang; Han, Dandan; Yan, Hongyuan

    2017-03-01

    A new method involving ionic liquid-hybrid molecularly imprinted material-filter solid-phase extraction coupled to high-performance liquid chromatography (IL-HIM-FSPE-HPLC) was developed for the simultaneous isolation and determination of 6-benzyladenine (6-BA) and 4-chlorophenoxyacetic acid (4-CPA) in bean sprouts. Sample preconcentration was performed using a modified filter, with the new IL-HIM as the adsorbent, which shows double adsorption. The first adsorption involves special recognition of molecular imprinting, and the second involves ion exchange and electrostatic attraction caused by the ionic liquid. This method combines the advantages of ionic liquids, hybrid materials, and molecularly imprinted polymers and was successfully applied to determine 6-BA and 4-CPA in bean sprouts. The adsorption of 6-BA to IL-HIM is based on selective imprinted recognition, whereas the adsorption of 4-CPA is mainly dependent on ion-exchange interactions.

  13. Polycarboxylic acids as network modifiers for water durability improvement of inorganic-organic hybrid tin-silico-phosphate low-melting glasses

    SciTech Connect

    Menaa, Bouzid . E-mail: bouzidmenaa@noncry.kuicr.kyoto-u.ac.jp; Mizuno, Megumi; Takahashi, Masahide . E-mail: masahide@noncry.kuicr.kyoto-u.ac.jp; Tokuda, Yomei; Yoko, Toshinobu

    2006-02-15

    We investigated the water durability of the inorganic-organic hybrid tin-silico-phosphate glasses Me{sub 2}SiO-SnO-P{sub 2}O{sub 5} (Me designs the organic methyl group) doped with organic acids (salicylic acid (SA), tartaric acid (TA), citric acid (Canada) and butane tetracarboxylic acid (BTCA)) containing one or more of carboxylic groups per molecule. The structure, thermal properties and durability of the final glasses obtained via a non-aqueous acid-base reaction were discussed owing to the nature and the concentration of the acid added. {sup 29}Si magic angle spinning (MAS) NMR and {sup 31}P MAS NMR spectra, respectively, showed clearly a modification of the network in the host glass matrix of the Me{sub 2}SiO-SnO-P{sub 2}O{sub 5} system. The polycondensation enhancement to form -P-O-Si-O-P- linkages (PSP) and the increase of the Q {sup 2} unit (two bridging oxygens per phosphorus atom) over the Q {sup 3} unit (three bridging oxygens per phosphorus atom) as a function of the acid in the order SAacids containing a large number of carboxylic groups per molecule. The presence of carboxylic groups of the acid acting as network modifier may retard the movement of water molecules through the glasses due to the steric hindrance strengthening the PSP connections in a chain-like structure.

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

    PubMed

    Noor, M Omair; Krull, Ulrich J

    2013-08-06

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

  15. Metal-organic hybrid materials built with tetrachlorophthalate acid and different N-donor coligands: Structure diversity and photoluminescence

    NASA Astrophysics Data System (ADS)

    Xiao, Zhenyu; Yang, Xiao; Zhao, Siwei; Wang, Debao; Yang, Yu; Wang, Lei

    2016-02-01

    Eight new metal-organic hybrid materials, namely {Cd(Tcph)(4,4‧-bipy)1/2} (1), {[Cd2(Tcph)2(1,4-bimb)1/2(H2O)4]·H2O} (2), {Cd2(Tcph)2(1,4-bmimb)1/2(H2O)4} (3), {Cd(Tcph)(1,2-bmimb)} (4), {Cu(Tcph)(1,4-bimb)(H2O)} (5), {[Co(Tcph)(1,4-bimb)1/2(H2O)3]·(H2O)} (6), {Zn(Tcph)(1,2-bimb)} (7), {Cu2(Tcph)2(1,2-bimb)(H2O)4} (8), where Tcph=tetrachlorophthalate acid, 4,4‧-bipy=4,4‧-bipyridine, 1,4-bimb=1,4-bis(imidazol-1-ylmethyl)benzene, 1,4-bmimb=1,4-bis(2-methylimidazol-1-ylmethyl)benzene, 1,2-bimb=1,2-bis(imidazol-1-ylmethyl)-benzene, 1,2-bmimb=1,2-bis(2-methylimidazol-1-ylmethyl)benzene, have been synthesized and characterized. Their structures are determined by single crystal X-ray diffraction and further characterized by infrared spectra (IR) and thermogravimetric (TG) analyses. Complex 1, 4 and 7 display 2D layer structures. 1 possesses two-dimensional sheet containing an unusual [Cd(Tcph)] chains linked by 4,4‧-bipy co-ligand, while 4 and 7 hold the similar 4-connected 44-sql nets. Complex 2 and 3 feature a similar three dimensional (3D) internal compensation structure with a topology of {42·63·8}2{63}. 5 is a novel 2-fold self-penetrating 3D network with 4-coordinated 65·8-CdSO4 subnets. The ladder-like chains of 6 are further connected through O-H···O interactions to yield a 3D supramolecular structure. 8 is a discrete tetranuclear complex. The thermal stabilities of 1-8 and the luminescent properties of 1-4 and 7 in the solid state are also discussed.

  16. Discriminating Multi-Species Populations in Biofilms with Peptide Nucleic Acid Fluorescence In Situ Hybridization (PNA FISH)

    PubMed Central

    Almeida, Carina; Azevedo, Nuno F.; Santos, Sílvio; Keevil, Charles W.; Vieira, Maria J.

    2011-01-01

    Background Our current understanding of biofilms indicates that these structures are typically composed of many different microbial species. However, the lack of reliable techniques for the discrimination of each population has meant that studies focusing on multi-species biofilms are scarce and typically generate qualitative rather than quantitative data. Methodology/Principal Findings We employ peptide nucleic acid fluorescence in situ hybridization (PNA FISH) methods to quantify and visualize mixed biofilm populations. As a case study, we present the characterization of Salmonella enterica/Listeria monocytogenes/Escherichia coli single, dual and tri-species biofilms in seven different support materials. Ex-situ, we were able to monitor quantitatively the populations of ∼56 mixed species biofilms up to 48 h, regardless of the support material. In situ, a correct quantification remained more elusive, but a qualitative understanding of biofilm structure and composition is clearly possible by confocal laser scanning microscopy (CLSM) at least up to 192 h. Combining the data obtained from PNA FISH/CLSM with data from other established techniques and from calculated microbial parameters, we were able to develop a model for this tri-species biofilm. The higher growth rate and exopolymer production ability of E. coli probably led this microorganism to outcompete the other two [average cell numbers (cells/cm2) for 48 h biofilm: E. coli 2,1×108 (±2,4×107); L. monocytogenes 6,8×107 (±9,4×106); and S. enterica 1,4×106 (±4,1×105)]. This overgrowth was confirmed by CSLM, with two well-defined layers being easily identified: the top one with E. coli, and the bottom one with mixed regions of L. monocytogenes and S. enterica. Significance While PNA FISH has been described previously for the qualitative study of biofilm populations, the present investigation demonstrates that it can also be used for the accurate quantification and spatial distribution of species in

  17. Luminescent hybrid lanthanide sulfates and lanthanide sulfonate-carboxylates with 1,10-phenanthroline involving in-situ oxidation of 2-mercaptonbenzoic acid

    SciTech Connect

    Zhong, Jie-Cen; Wan, Fang; Sun, Yan-Qiong; Chen, Yi-Ping

    2015-01-15

    A series of lanthanide sulfates and lanthanide sulfonate-carboxylates, [Ln{sub 2}(phen){sub 2}(SO{sub 4}){sub 3}(H{sub 2}O){sub 2}]{sub n} (I:Ln=Nd(1a), Sm(1b), Eu(1c), phen=1,10-phenanthroline) and [Ln(phen)(2-SBA)(BZA)]{sub n} (II: Ln=Sm(2a), Eu(2b), Dy(2c), 2-SBA=2-sulfobenzoate, BZA=benzoate) have been hydrothermally synthesized from lanthanide oxide, 2-mercaptonbenzoic acid with phen as auxiliary ligand and characterized by single-crystal X-ray diffraction, elemental analyses, IR spectra, TG analyses and luminescence spectroscopy. Interestingly, SO{sub 4}{sup 2−} anions in I came from the in situ deep oxidation of thiol groups of 2-mercaptonbenzoic acid while 2-sulfobenzoate and benzoate ligands in II from the middle oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. Compounds I are organic–inorganic hybrid lanthanide sulfates, which have rare one-dimensional column-like structures. Complexes II are binuclear lanthanide sulfonate-carboxylates with 2-sulfobenzoate and benzoate as bridges and 1,10-phenanthroline as terminal. Photoluminescence studies reveal that complexes I and II exhibit strong lanthanide characteristic emission bands in the solid state at room temperature. - Graphical abstract: Lanthanide sulfates and lanthanide sulfonate-carboxylates have been hydrothermally synthesized. Interestingly, sulfate anions, 2-sulfobenzoate and benzoate ligands came from the in situ oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. - Highlights: • In situ oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. • The organic–inorganic hybrid lanthanide sulfates with one-dimensional column-like structure. • The dinuclear lanthanide sulfonate-carboxylates. • The emission spectra exhibit the characteristic transition of {sup 5}D{sub 0}→{sup 7}F{sub J} (J=0–4) of the Eu(III)

  18. Design, synthesis and biological evaluation of hybrids of β-carboline and salicylic acid as potential anticancer and apoptosis inducing agents

    PubMed Central

    Xu, Qi-Bing; Chen, Xiang-Fan; Feng, Jiao; Miao, Jie-Fei; Liu, Ji; Liu, Feng-Tao; Niu, Bi-Xi; Cai, Jin-Yang; Huang, Chao; Zhang, Yanan; Ling, Yong

    2016-01-01

    A novel series of hybrids (7a-l, 8a-l) from β-carboline and salicylic acid (SA) were designed and synthesized, and their in vitro biological activities were evaluated. Most of the hybrids displayed potent antiproliferative activity against five cancer cell lines in vitro, showing potencies superior to 5-FU and harmine. In particular, compound 8h selectively inhibited proliferation of liver cancer SMMC-7721 cells but not normal liver LO2 cells, and displayed greater inhibitory selectivity than intermediate 5h and SA. 8h also induced cancer cell apoptosis in an Annexin V-FITC/propidium iodide flow cytometry assay, and triggered the mitochondrial/caspase apoptosis by decreasing mitochondrial membrane potential which was associated with up-regulation of Bax, down-regulation of Bcl-2 and activation levels of the caspase cascade in a concentration-dependent manner. Our findings suggest that the β-carboline/SA hybrids may hold greater promise as therapeutic agents for the intervention of human cancers. PMID:27824091

  19. Impedance measurements on lead-acid batteries for state-of-charge, state-of-health and cranking capability prognosis in electric and hybrid electric vehicles

    NASA Astrophysics Data System (ADS)

    Blanke, Holger; Bohlen, Oliver; Buller, Stephan; De Doncker, Rik W.; Fricke, Birger; Hammouche, Abderrezak; Linzen, Dirk; Thele, Marc; Sauer, Dirk Uwe

    Various attempts have been made to use impedance measurements for online analysis and offline modelling of lead-acid batteries. This presentation gives an overview on the latest and successful approaches based on impedance measurements to assess state-of-charge (SoC), state-of-health (SoH) and cranking capability of lead-acid batteries. Furthermore, it is shown that impedance data can serve as a basis for dynamic battery models for the simulation of vehicle power-supply systems. The methods and procedures aim for a reliable prediction of battery performance in electric vehicles, hybrid cars and classical automotive applications. Although, it will become obvious that impedance measurements give valuable information on the battery state, typically the information needs to be combined with other conventional algorithms or self-learning tools to achieve reliable and stable results for real-world applications.

  20. MoS2/reduced graphene oxide as active hybrid material for the electrochemical detection of folic acid in human serum.

    PubMed

    Chekin, Fereshteh; Teodorescu, Florina; Coffinier, Yannick; Pan, Guo-Hui; Barras, Alexandre; Boukherroub, Rabah; Szunerits, Sabine

    2016-11-15

    In this study, a new matrix based on a molybdenum disulfide-reduced graphene oxide hybrid (MoS2-rGO) was prepared and characterized. Modification of a glassy carbon electrode (GCE) with MoS2-rGO (MG) using drop casting allowed for the selective analysis of folic acid in the presence of a variety of interference species with a limit of detection of 10nM, a linear range between 0.01μM and 100μM with a sensitivity of 14µAµM(-1). In addition, the analytical performance of the proposed sensor was successfully conducted for the determination of folic acid in human serum samples, making MG-GC electrodes promising interfaces for bio-electrochemical applications.

  1. Au/ZnO hybrid nanocatalysts impregnated in N-doped graphene for simultaneous determination of ascorbic acid, acetaminophen and dopamine.

    PubMed

    Chen, Xianlan; Zhang, Guowei; Shi, Ling; Pan, Shanqing; Liu, Wei; Pan, Hiabo

    2016-08-01

    The formation of nitrogen-doped (N-doped) graphene uses hydrothermal method with urea as reducing agent and nitrogen source. The surface elemental composition of the catalyst was analyzed through XPS, which showed a high content of a total N species (7.12at.%), indicative of the effective N-doping, present in the form of pyridinic N, pyrrolic N and graphitic N groups. Moreover, Au nanoparticles deposited on ZnO nanocrystals surface, forming Au/ZnO hybrid nanocatalysts, undergo a super-hydrophobic to super-hydrophilic conversion. Herein, we present Au/ZnO hybrid nanocatalysts impregnated in N-doped graphene sheets through sonication technique of the Au/ZnO/N-doped graphene hybrid nanostructures. The as-prepared Au/ZnO/N-doped graphene hybrid nanostructure modified glassy carbon electrode (Au/ZnO/N-doped graphene/GCE) was first employed for the simultaneous determination of ascorbic acid (AA), dopamine (DA) and acetaminophen (AC). The oxidation over-potentials of AA, DA and AC decreased dramatically, and their oxidation peak currents increased significantly at Au/ZnO/N-doped graphene/GCE compared to those obtained at the N-doped graphene/GCE and bare CCE. The peak separations between AA and DA, DA and AC, and AC and AA are large up to 195, 198 and 393mV, respectively. The calibration curves for AA, DA and AC were obtained in the range of 30.00-13.00×10(3), 2.00-0.18×10(3) and 5.00-3.10×10(3)μM, respectively. The detection limits (S/N=3) were 5.00, 0.40 and 0.80μM for AA, DA and AC, respectively. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Differential Effects of Elevated Ozone on Two Hybrid Aspen Genotypes Predisposed to Chronic Ozone Fumigation. Role of Ethylene and Salicylic Acid1

    PubMed Central

    Vahala, Jorma; Keinänen, Markku; Schützendübel, Andres; Polle, Andrea; Kangasjärvi, Jaakko

    2003-01-01

    The role of ethylene (ET) signaling in the responses of two hybrid aspen (Populus tremula L. × P. tremuloides Michx.) clones to chronic ozone (O3; 75 nL L−1) was investigated. The hormonal responses differed between the clones; the O3-sensitive clone 51 had higher ET evolution than the tolerant clone 200 during the exposure, whereas the free salicylic acid concentration in clone 200 was higher than in clone 51. The cellular redox status, measured as glutathione redox balance, did not differ between the clones suggesting that the O3 lesions were not a result of deficient antioxidative capacity. The buildup of salicylic acid during chronic O3 exposure might have prevented the up-regulation of ET biosynthesis in clone 200. Blocking of ET perception with 1-methylcyclopropene protected both clones from the decrease in net photosynthesis during chronic exposure to O3. After a pretreatment with low O3 for 9 d, an acute 1.5-fold O3 elevation caused necrosis in the O3-sensitive clone 51, which increased substantially when ET perception was blocked. The results suggest that in hybrid aspen, ET signaling had a dual role depending on the severity of the stress. ET accelerated leaf senescence under low O3, but under acute O3 elevation, ET signaling seemed to be required for protection from necrotic cell death. PMID:12746525

  3. Micro-hybrid electric vehicle application of valve-regulated lead-acid batteries in absorbent glass mat technology: Testing a partial-state-of-charge operation strategy

    NASA Astrophysics Data System (ADS)

    Schaeck, S.; Stoermer, A. O.; Hockgeiger, E.

    The BMW Group has launched two micro-hybrid functions in high volume models in order to contribute to reduction of fuel consumption in modern passenger cars. Both the brake energy regeneration (BER) and the auto-start-stop function (ASSF) are based on the conventional 14 V vehicle electrical system and current series components with only little modifications. An intelligent control algorithm of the alternator enables recuperative charging in braking and coasting phases, known as BER. By switching off the internal combustion engine at a vehicle standstill the idling fuel consumption is effectively reduced by ASSF. By reason of economy and package a lead-acid battery is used as electrochemical energy storage device. The BMW Group assembles valve-regulated lead-acid (VRLA) batteries in absorbent glass mat (AGM) technology in the micro-hybrid electrical power system since special challenges arise for the batteries. By field data analysis a lower average state-of-charge (SOC) due to partial state-of-charge (PSOC) operation and a higher cycling rate due to BER and ASSF are confirmed in this article. Similar to a design of experiment (DOE) like method we present a long-term lab investigation. Two types of 90 Ah VRLA AGM batteries are operated with a test bench profile that simulates the micro-hybrid vehicle electrical system under varying conditions. The main attention of this lab testing is focused on capacity loss and charge acceptance over cycle life. These effects are put into context with periodically refresh charging the batteries in order to prevent accelerated battery aging due to hard sulfation. We demonstrate the positive effect of refresh chargings concerning preservation of battery charge acceptance. Furthermore, we observe moderate capacity loss over 90 full cycles both at 25 °C and at 3 °C battery temperature.

  4. Application of a hybrid computational fluid dynamics and physiologically based inhalation model for interspecies dosimetry extrapolation of acidic vapors in the upper airways.

    PubMed

    Frederick, C B; Bush, M L; Lomax, L G; Black, K A; Finch, L; Kimbell, J S; Morgan, K T; Subramaniam, R P; Morris, J B; Ultman, J S

    1998-09-01

    This study provides a scientific basis for interspecies extrapolation of nasal olfactory irritants from rodents to humans. By using a series of short-term in vivo studies, in vitro studies with nasal explants, and computer modeling, regional nasal tissue dose estimates were made and comparisons of tissue doses between species were conducted. To make these comparisons, this study assumes that human and rodent olfactory epithelium have similar susceptibility to the cytotoxic effects of organic acids based on similar histological structure and common mode of action considerations. Interspecies differences in susceptibility to the toxic effects of acidic vapors are therefore assumed to be driven primarily by differences in nasal tissue concentrations that result from regional differences in nasal air flow patterns relative to the species-specific distribution of olfactory epithelium in the nasal cavity. The acute, subchronic, and in vitro studies have demonstrated that the nasal olfactory epithelium is the most sensitive tissue to the effects of inhalation exposure to organic acids and that the sustentacular cells are the most sensitive cell type of this epithelium. A hybrid computational fluid dynamics (CFD) and physiologically based pharmacokinetic (PBPK) dosimetry model was constructed to estimate the regional tissue dose of organic acids in the rodent and human nasal cavity. The CFD-PBPK model simulations indicate that the olfactory epithelium of the human nasal cavity is exposed to two- to threefold lower tissue concentrations of a representative inhaled organic acid vapor, acrylic acid, than the olfactory epithelium of the rodent nasal cavity when the exposure conditions are the same. The magnitude of this difference varies somewhat with the specific exposure scenario that is simulated. The increased olfactory tissue dose in rats relative to humans may be attributed to the large rodent olfactory surface area (greater than 50% of the nasal cavity) and its highly

  5. Oxidative degradation of different chlorinated phenoxyalkanoic acid herbicides by a hybrid ZrO2 gel-derived catalyst without light irradiation.

    PubMed

    Sannino, Filomena; Pernice, Pasquale; Minieri, Luciana; Camandona, Gaia Aurora; Aronne, Antonio; Pirozzi, Domenico

    2015-01-14

    The oxidative degradation of 2-methyl-4-chlorophenoxyacetic acid (MCPA), 4-(4-chloro-2-methylphenoxy)butanoic acid (MCPB), 4-chlorophenoxyacetic acid (4-CPA) and 2,4-dichlorophenoxyacetic acid (2,4 D) by ZrO2-acetylacetonate hybrid catalyst (HSGZ) without light irradiation was assessed. The thermal stability of the catalyst was investigated by thermogravimetry, differential thermal analysis, and Fourier transform infrared spectroscopy. For each herbicide, a virtually complete removal in about 3 days without light irradiation at room temperature was achieved. The removal kinetics of the herbicides has been satisfactorily characterized by a double-stage physico-mathematical model, in the hypothesis that a first-order adsorption on HSGZ surface is followed by the herbicide degradation, catalytically driven by HSGZ surface groups. The long-term use of the HSGZ catalyst was assessed by repeated-batch tests. The specific cost for unit-volume removal of herbicide was evaluated by a detailed cost analysis showing that it is comparable with those pertaining to alternative methods.

  6. Fermentative production of poly (γ-glutamic acid) from renewable carbon source and downstream purification through a continuous membrane-integrated hybrid process.

    PubMed

    Kumar, Ramesh; Pal, Parimal

    2015-02-01

    Experimental investigations were carried out on continuous and direct production of poly-(γ-glutamic acid) in a hybrid reactor system that integrated conventional fermentative production step with membrane-based downstream separation and purification. Novelty of the integrated system lies in high degree of purity, conversion, yield and productivity of poly-(γ-glutamic acid) through elimination of substrate-product inhibitions of traditional batch production system. This new system is compact, flexible, eco-friendly and largely fouling-free ensuring steady and continuous production of poly-(γ-glutamic acid) directly from a renewable carbon source at the rate of 0.91 g/L/h. Cross-flow microfiltration membrane modules ensured almost complete separation and recycle of cells without much fouling problem. Well-screened ultrafiltration membrane module helped to concentrate poly-(γ-glutamic acid) while ensuring recovery and recycle of 96% unconverted carbon source resulting in yield of 0.6g/g along with high product purity.

  7. Simultaneous optimization of monolayer formation factors, including temperature, to significantly improve nucleic acid hybridization efficiency on gold substrates.

    PubMed

    Pris, Andrew D; Ostrowski, Sara G; Garaas, Sarah D

    2010-04-20

    Past literature investigations have optimized various single factors used in the formation of thiolated, single stranded DNA (ss-DNA) monolayers on gold. In this study a more comprehensive approach is taken, where a design of experiment (DOE) is employed to simultaneously optimize all of the factors involved in construction of the capture monolayer used in a fluorescence-based hybridization assay. Statistical analysis of the fluorescent intensities resulting from the DOE provides empirical evidence for the importance and the optimal levels of traditional and novel factors included in this investigation. We report on the statistical importance of a novel factor, temperature of the system during monolayer formation of the capture molecule and lateral spacer molecule, and how proper usage of this temperature factor increased the hybridization signal 50%. An initial theory of how the physical factor of heat is mechanistically supplementing the function of the lateral spacer molecule is provided.

  8. Comparison of nucleic acid targets prepared from total RNA or poly(A) RNA for DNA oligonucleotide microarray hybridization.

    PubMed

    Petersen, Kjell; Oyan, Anne Margrete; Rostad, Kari; Olsen, Sue; Bø, Trond Hellem; Salvesen, Helga B; Gjertsen, Bjørn Tore; Bruserud, Oystein; Halvorsen, Ole Johan; Akslen, Lars Andreas; Steen, Vidar M; Jonassen, Inge; Kalland, Karl-Henning

    2007-07-01

    The aim of this work was to compare DNA microarray results using either total RNA or affinity-purified poly(A) RNA from the same biological sample for target preparation. The high-density oligonucleotide microarrays of both Agilent Technologies (based on two-color detection) and Applied Biosystems (based on single-color detection) were evaluated. Real-time quantitative PCR was used to quantify messenger RNA (mRNA) and ribosomal RNA (rRNA) at different stages of target preparations. Poly(A) RNA versus total RNA target hybridizations exhibited slightly lower correlation coefficients than did self versus self hybridizations (i.e., poly(A) RNA targets vs. poly(A) RNA targets or total RNA targets vs. total RNA targets). Only a small fraction of all transcripts appeared to be significantly over- or underrepresented when total RNA targets or poly(A) RNA targets from the same biological sample were compared. Therefore, the conclusion is that poly(A) affinity purification from total RNA can be omitted during target preparation for routine mRNA expression analysis using high-density oligonucleotide microarrays. Among consistently overrepresented transcripts in total RNA targets were histone mRNAs known to lack poly(A) tails. Therefore, structurally exceptional RNA species can be identified by comparing targets derived from either poly(A) RNA or total RNA using microarray hybridization.

  9. Subcellular localization of low-abundance human immunodeficiency virus nucleic acid sequences visualized by fluorescence in situ hybridization

    SciTech Connect

    Lawrence, J.B.; Marselle, L.M.; Byron, K.S.; Johnson, C.V.; Sullivan, J.L.; Singer, R.H. )

    1990-07-01

    Detection and subcellular localization of human immunodeficiency virus (HIV) were investigated using sensitive high-resolution in situ hybridization methodology. Lymphocytes infected with HIV in vitro or in vivo were detected by fluorescence after hybridization with either biotin or digoxigenin-labeled probes. At 12 hr after infection in vitro, a single intense signal appeared in the nuclei of individual cells. Later in infection, when cytoplasmic fluorescence became intense, multiple nuclear foci frequently appeared. The nuclear focus consisted of newly synthesized HIV RNA as shown by hybridization in the absence of denaturation and by susceptibility to RNase and actinomycin D. Virus was detected in patient lymphocytes and it was shown that a singular nuclear focus also characterizes cells infected in vivo. The cell line 8E5/LAV containing one defective integrated provirus revealed a similar focus of nuclear RNA, and the single integrated HIV genome was unequivocally visualized on a D-group chromosome. This demonstrates an extremely sensitive single-cell assay for the presence of a single site of HIV transcription in vitro and in vivo and suggests that it derives from one (or very few) viral genomes per cell. In contrast, productive Epstein-Barr virus infection exhibited many foci of nuclear RNA per cell.

  10. Camera-based ratiometric fluorescence transduction of nucleic acid hybridization with reagentless signal amplification on a paper-based platform using immobilized quantum dots as donors.

    PubMed

    Noor, M Omair; Krull, Ulrich J

    2014-10-21

    Paper-based diagnostic assays are gaining increasing popularity for their potential application in resource-limited settings and for point-of-care screening. Achievement of high sensitivity with precision and accuracy can be challenging when using paper substrates. Herein, we implement the red-green-blue color palette of a digital camera for quantitative ratiometric transduction of nucleic acid hybridization on a paper-based platform using immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). A nonenzymatic and reagentless means of signal enhancement for QD-FRET assays on paper substrates is based on the use of dry paper substrates for data acquisition. This approach offered at least a 10-fold higher assay sensitivity and at least a 10-fold lower limit of detection (LOD) as compared to hydrated paper substrates. The surface of paper was modified with imidazole groups to assemble a transduction interface that consisted of immobilized QD-probe oligonucleotide conjugates. Green-emitting QDs (gQDs) served as donors with Cy3 as an acceptor. A hybridization event that brought the Cy3 acceptor dye in close proximity to the surface of immobilized gQDs was responsible for a FRET-sensitized emission from the acceptor dye, which served as an analytical signal. A hand-held UV lamp was used as an excitation source and ratiometric analysis using an iPad camera was possible by a relative intensity analysis of the red (Cy3 photoluminescence (PL)) and green (gQD PL) color channels of the digital camera. For digital imaging using an iPad camera, the LOD of the assay in a sandwich format was 450 fmol with a dynamic range spanning 2 orders of magnitude, while an epifluorescence microscope detection platform offered a LOD of 30 fmol and a dynamic range spanning 3 orders of magnitude. The selectivity of the hybridization assay was demonstrated by detection of a single nucleotide polymorphism at a contrast ratio of 60:1. This work provides an

  11. Design, synthesis, and biological evaluation of novel hybrid dicaffeoyltartaric/diketo acid and tetrazole-substituted L-chicoric acid analogue inhibitors of human immunodeficiency virus type 1 integrase.

    PubMed

    Crosby, David C; Lei, Xiangyang; Gibbs, Charles G; McDougall, Brenda R; Robinson, W Edward; Reinecke, Manfred G

    2010-11-25

    Fourteen analogues of the anti-HIV-1 integrase (IN) inhibitor L-chicoric acid (L-CA) were prepared. Their IC(50) values for 3'-end processing and strand transfer against recombinant HIV-1 IN were determined in vitro, and their cell toxicities and EC(50) against HIV-1 were measured in cells (ex vivo). Compounds 1-6 are catechol/β-diketoacid hybrids, the majority of which exhibit submicromolar potency against 3'-end processing and strand transfer, though only with modest antiviral activities. Compounds 7-10 are L-CA/p-fluorobenzylpyrroloyl hybrids, several of which were more potent against strand transfer than 3'-end processing, a phenomenon previously attributed to the β-diketo acid pharmacophore. Compounds 11-14 are tetrazole bioisosteres of L-CA and its analogues, whose in vitro potencies were comparable to L-CA but with enhanced antiviral potency. The trihydroxyphenyl analogue 14 was 30-fold more potent than L-CA at relatively nontoxic concentrations. These data indicate that L-CA analogues are attractive candidates for development into clinically relevant inhibitors of HIV-1 IN.

  12. Comparing the effects of different dietary organic acids on the growth, intestinal short-chain fatty acids, and liver histopathology of red hybrid tilapia (Oreochromis sp.) and potential use of these as preservatives.

    PubMed

    Ebrahimi, Mahdi; Daeman, Nor Hafizah; Chong, Chou Min; Karami, Ali; Kumar, Vikas; Hoseinifar, Seyed Hossein; Romano, Nicholas

    2017-08-01

    Dietary organic acids are increasingly being investigated as a potential means of improving growth and nutrient utilization in aquatic animals. A 9-week study was performed to compare equal amounts (2%) of different organic acids (sodium butyrate, acetate, propionate, or formate) on the growth, muscle proximate composition, fatty acid composition, cholesterol and lipid peroxidation, differential cell counts, plasma biochemistry, intestinal short-chain fatty acid (SCFA) level, and liver histopathology to red hybrid tilapia (Oreochromis sp.) (initial mean weight of 2.87 g). A second experiment was performed to determine their effects on lipid peroxidation and trimethylamine (TMA) when added at 1% to tilapia meat and left out for 24 h. The results of the first experiment showed no treatment effect to growth, feeding efficiencies, or muscle fatty acid composition, but all dietary organic acids significantly decreased intestinal SCFA. Dietary butyrate and propionate significantly decreased muscle lipid peroxidation compared to the control group, but the dietary formate treatment had the lowest lipid peroxidation compared to all treatments. Muscle crude protein and lipid in tilapia fed the formate diet were significantly lower and higher, respectively, and showed evidence of stress based on the differential cell counts, significantly higher plasma glucose and liver glycogen, as well as inflammatory responses in the liver. Although a potential benefit of dietary organic acids was a reduction to lipid peroxidation, this could be accomplished post-harvest by direct additions to the meat. In addition, inclusions of butyrate and propionate to tilapia meat significantly decreased TMA, which might be a more cost-effective option to improve the shelf life of tilapia products.

  13. Import of desired nucleic acid sequences using addressing motif of mitochondrial ribosomal 5S-rRNA for fluorescent in vivo hybridization of mitochondrial DNA and RNA.

    PubMed

    Zelenka, Jaroslav; Alán, Lukáš; Jabůrek, Martin; Ježek, Petr

    2014-04-01

    Based on the matrix-addressing sequence of mitochondrial ribosomal 5S-rRNA (termed MAM), which is naturally imported into mitochondria, we have constructed an import system for in vivo targeting of mitochondrial DNA (mtDNA) or mt-mRNA, in order to provide fluorescence hybridization of the desired sequences. Thus DNA oligonucleotides were constructed, containing the 5'-flanked T7 RNA polymerase promoter. After in vitro transcription and fluorescent labeling with Alexa Fluor(®) 488 or 647 dye, we obtained the fluorescent "L-ND5 probe" containing MAM and exemplar cargo, i.e., annealing sequence to a short portion of ND5 mRNA and to the light-strand mtDNA complementary to the heavy strand nd5 mt gene (5'-end 21 base pair sequence). For mitochondrial in vivo fluorescent hybridization, HepG2 cells were treated with dequalinium micelles, containing the fluorescent probes, bringing the probes proximally to the mitochondrial outer membrane and to the natural import system. A verification of import into the mitochondrial matrix of cultured HepG2 cells was provided by confocal microscopy colocalizations. Transfections using lipofectamine or probes without 5S-rRNA addressing MAM sequence or with MAM only were ineffective. Alternatively, the same DNA oligonucleotides with 5'-CACC overhang (substituting T7 promoter) were transcribed from the tetracycline-inducible pENTRH1/TO vector in human embryonic kidney T-REx®-293 cells, while mitochondrial matrix localization after import of the resulting unlabeled RNA was detected by PCR. The MAM-containing probe was then enriched by three-order of magnitude over the natural ND5 mRNA in the mitochondrial matrix. In conclusion, we present a proof-of-principle for mitochondrial in vivo hybridization and mitochondrial nucleic acid import.

  14. New hybrid nanocomposite of copper terephthalate MOF-graphene oxide: synthesis, characterization and application as adsorbents for toxic metal ion removal from Sungun acid mine drainage.

    PubMed

    Rahimi, Esmaeil; Mohaghegh, Neda

    2017-08-11

    The application of a hybrid Cu(tpa).GO (Cu(tpa) copper terephthalate metal organic framework, GO graphene oxide) composite as a new adsorbent for the removal of toxic metal ions was reported. New hybrid nanocomposite with excellent dispersibility and stability was successfully fabricated by the simple and effective ultrasonication method. The synthesized composite was characterized by scanning electron microscopy (SEM), UV-Vis and Fourier-transform infrared (FT-IR) techniques. The characterization results concluded that the binding mechanism of the Cu(tpa) and GO was related to both π-π packing and hydrogen bonding. For scrutinizing the sorption activity, the prepared adsorbents were assessed for the removal of Mn(2+), Cu(2+), Zn(2+), Cd(2+), Pb(2+) and Fe(3+) metal ions from aqueous synthetic solution and also acid mine drainage (AMD) wastewater. The sorption experiments demonstrated that the removal efficiency was significantly improved by modified hybrid Cu(tpa).GO composite, owing to the significant number of active binding sites and unique structure formed based on π-conjugated networks. Also, it was shown that the adsorption reaction was mainly attributed to the chemical interactions between metal ions and the surface functional groups. Moreover, kinetic and adsorption studies clarified that the adsorption process onto the Cu(tpa).GO follows a pseudo-second-order kinetics and fits the Langmuir and Freundlich adsorption models. Holistically, the results of this research represent that applying Cu(tpa).GO can be remarked as an effective adsorbent with high possibility at conventional water treatment.

  15. Acetic acid recovery from a hybrid biological-hydrothermal treatment process of sewage sludge - a pilot plant study.

    PubMed

    Andrews, J; Dare, P; Estcourt, G; Gapes, D; Lei, R; McDonald, B; Wijaya, N

    2015-01-01

    A two-stage process consisting of anaerobic fermentation followed by sub-critical wet oxidation was used to generate acetic acid from sewage sludge at pilot scale. Volatile fatty acids, dominated by propionic acid, were produced over 4-6 days in the 2,000 L fermentation reactor, which also achieved 31% solids reduction. Approximately 96% of the carbon was retained in solution over the fermentation stage. Using a 200 L wet oxidation reactor operating in batch mode, the second stage achieved 98% volatile suspended solids (VSS) destruction and 67% total chemical oxygen demand (tCOD) destruction. Acetic acid produced in this stage was recalcitrant to further degradation and was retained in solution. The gross yield from VSS was 16% for acetic acid and 21% for volatile fatty acids across the process, higher than reported yields for wet oxidation alone. The pilot plant results showed that 72% of the incoming phosphorus was retained in the solids, 94% of the nitrogen became concentrated in solution and 41% of the carbon was converted to a soluble state, in a more degradable form. Acetic acid produced from the process has the potential to be used to offset ethanol requirements in biological nutrient removal plants.

  16. Positively charged polymer brush-functionalized filter paper for DNA sequence determination following Dot blot hybridization employing a pyrrolidinyl peptide nucleic acid probe.

    PubMed

    Laopa, Praethong S; Vilaivan, Tirayut; Hoven, Voravee P

    2013-01-07

    As inspired by the Dot blot analysis, a well known technique in molecular biology and genetics for detecting biomolecules, a new paper-based platform for colorimetric detection of specific DNA sequences employing peptide nucleic acid (PNA) as a probe has been developed. In this particular study, a pyrrolidinyl PNA bearing a conformationally rigid d-prolyl-2-aminocyclopentanecarboxylic acid backbone (acpcPNA) was used as a probe. The filter paper was modified to be positively charged with grafted polymer brushes of quaternized poly(dimethylamino)ethyl methacrylate (QPDMAEMA) prepared by surface-initiated polymerization of 2-(dimethylamino)ethyl methacrylate from the filter paper via ARGET ATRP followed by quaternization with methyl iodide. Following the Dot blot format, a DNA target was first immobilized via electrostatic interactions between the positive charges of the QPDMAEMA brushes and negative charges of the phosphate backbone of DNA. Upon hybridization with the biotinylated pyrrolidinyl peptide nucleic acid (b-PNA) probe, the immobilized DNA can be detected by naked eye observation of the yellow product generated by the enzymatic reaction employing HRP-labeled streptavidin. It has been demonstrated that this newly developed assay was capable of discriminating between complementary and single base mismatch targets at a detection limit of at least 10 fmol. In addition, the QPDMAEMA-grafted filter paper exhibited a superior performance to the commercial membranes, namely Nylon 66 and nitrocellulose.

  17. Luminescent hybrid lanthanide sulfates and lanthanide sulfonate-carboxylates with 1,10-phenanthroline involving in-situ oxidation of 2-mercaptonbenzoic acid

    NASA Astrophysics Data System (ADS)

    Zhong, Jie-Cen; Wan, Fang; Sun, Yan-Qiong; Chen, Yi-Ping

    2015-01-01

    A series of lanthanide sulfates and lanthanide sulfonate-carboxylates, [Ln2(phen)2(SO4)3(H2O)2]n (I:Ln=Nd(1a), Sm(1b), Eu(1c), phen=1,10-phenanthroline) and [Ln(phen)(2-SBA)(BZA)]n (II: Ln=Sm(2a), Eu(2b), Dy(2c), 2-SBA=2-sulfobenzoate, BZA=benzoate) have been hydrothermally synthesized from lanthanide oxide, 2-mercaptonbenzoic acid with phen as auxiliary ligand and characterized by single-crystal X-ray diffraction, elemental analyses, IR spectra, TG analyses and luminescence spectroscopy. Interestingly, SO4 2 - anions in I came from the in situ deep oxidation of thiol groups of 2-mercaptonbenzoic acid while 2-sulfobenzoate and benzoate ligands in II from the middle oxidation and desulfuration reactions of 2-mercaptonbenzoic acid. Compounds I are organic-inorganic hybrid lanthanide sulfates, which have rare one-dimensional column-like structures. Complexes II are binuclear lanthanide sulfonate-carboxylates with 2-sulfobenzoate and benzoate as bridges and 1,10-phenanthroline as terminal. Photoluminescence studies reveal that complexes I and II exhibit strong lanthanide characteristic emission bands in the solid state at room temperature.

  18. Toward an on-chip multiplexed nucleic acid hybridization assay using immobilized quantum dot-oligonucleotide conjugates and fluorescence resonance energy transfer

    NASA Astrophysics Data System (ADS)

    Tavares, Anthony J.; Noor, M. Omair; Algar, W. Russ; Vannoy, Charles H.; Chen, Lu; Krull, Ulrich J.

    2011-03-01

    Semiconductor quantum dots (QD) are a class of NP with photophysical properties that are ideally suited for optical multiplexing and use as donors in fluorescence resonance energy transfer (FRET). A new strategy is presented for the development of multiplexed DNA hybridization assays using immobilized QDs in a microfluidic system. Green- or red-emitting QDs were immobilized via self-assembly with a multidentate-thiol-derivatized glass slide, and subsequently conjugated with amine-terminated probe oligonucleotides using carbodiimide activation. Immobilized QD-probe conjugates were then passivated with adsorbed non-complementary oligonucleotides to achieve selectivity in microfluidic assays. Target nucleic acid sequences hybridized with QD-probe conjugates and were labeled with Cy3 or Alexa Fluor 647 as acceptor dyes for the QD donors, where FRET-sensitized dye emission provided a signal for the detection of picomolar quantities of target. The simultaneous immobilization of green- and red-emitting QDs at different ratios within a microfluidic channel was demonstrated as a step toward multiplexed assays.

  19. Fischer carbene mediated covalent grafting of a peptide nucleic acid on gold surfaces and IR optical detection of DNA hybridization with a transition metalcarbonyl label

    NASA Astrophysics Data System (ADS)

    Srivastava, Pratima; Ghasemi, Mahsa; Ray, Namrata; Sarkar, Amitabha; Kocabova, Jana; Lachmanova, Stepanka; Hromadova, Magdalena; Boujday, Souhir; Cauteruccio, Silvia; Thakare, Pramod; Licandro, Emanuela; Fosse, Céline; Salmain, Michèle

    2016-11-01

    Amine-reactive surfaces comprising N-hydroxysuccinimide ester groups as well as much more unusual Fischer alkoxymetallocarbene groups were generated on gold-coated surfaces via self-assembled monolayers of carboxy- and azido-terminated thiolates, respectively. These functions were further used to immobilize homothymine peptide nucleic acid (PNA) decamer in a covalent fashion involving the primary amine located at its N-terminus. These stepwise processes were monitored by polarization modulation reflection - absorption infrared spectroscopy (PM-RAIRS) that gave useful information on the molecular composition of the organic layers. PNA grafting and hybridization with complementary DNA strand were successfully transduced by quartz crystal microbalance (QCM) measurements. Unfortunately, attempts to transduce the hybridization optically by IR in a label-free fashion were inconclusive. Therefore we undertook to introduce an IR reporter group, namely a transition metalcarbonyl (TMC) entity at the 5‧ terminus of complementary DNA. Evidence for the formation of PNA-DNA heteroduplex was brought by the presence of ν(Ctbnd O) bands in the 2000 cm-1 region of the IR spectrum of the gold surface owing to the metalcarbonyl label.

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

    PubMed

    Doughan, Samer; Uddayasankar, Uvaraj; Krull, Ulrich J

    2015-06-09

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

  1. Peptide Nucleic Acid-Fluorescence In Situ Hybridization for Detection of Staphylococci From Endophthalmitis Isolates: A Proof-of-Concept Study.

    PubMed

    Patel, Nimesh; Miller, Darlene; Relhan, Nidhi; Flynn, Harry W

    2017-08-01

    Rapid identification of pathogens causing endophthalmitis may improve treatment outcomes through early administration of species-specific medication. The current study reports a new molecular application of peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) with Staphylococcus-specific molecular PNA probes for the potential rapid detection of common pathogens causing endophthalmitis. An experimental study was designed to evaluate the proof of concept at the microbiology laboratory of the Bascom Palmer Eye Institute. Stored culture-positive staphylococci endophthalmitis isolates obtained from prior vitreous samples (n = 15), along with broth as negative controls (n = 5) were used. Inoculum was prepared to a final concentration of 1 × 105 colony-forming units/mL to ensure that the isolates were viable. Smears of samples were fixed and hybridized using QuickFISH protocol with probes for Staphylococcus. With PNA-FISH technique, Staphylococcus aureus was identified in 9 of 10 samples and coagulase-negative staphylococci were identified in 10 of 10 samples. Detection time was 20 minutes. This study serves a proof of concept using a new microbial detection system with FISH probes, and may have the potential for clinical use in the rapid and accurate identification of isolates from patients with endophthalmitis.

  2. Identification of the Mtv-2 gene responsible for the early appearance of mammary tumors in the GR mouse by nucleic acid hybridization

    PubMed Central

    Michalides, R.; Deemter, L. Van; Nusse, R.; Nie, R. Van

    1978-01-01

    In the mouse strain GR, the Mtv-2 gene controls the expression of large amounts of mammary tumor virus (MTV) antigens in the milk at first lactation. It also controls the early appearance of mammary tumors. We have investigated the number of MTV proviral sequences associated with this Mtv-2 gene by nucleic acid hybridization between MTV [3H]cDNA and DNA from GR, B10, and GR-Mtv-2- mice. B10 and GR-Mtv-2- mice lack Mtv-2 gene expression. The molecular hybridizations revealed that the DNA of GR mice contains 12 copies of MTV proviral sequences, whereas only 4 copies are present in the DNA of B10 and GR-Mtv-2- mice. We therefore conclude that the Mtv-2 gene in the GR mouse strain is associated with eight additional MTV proviral sequences. The four Mtv proviral sequences in the GR-Mtv-2- DNA might represent another Mtv gene in the GR mouse. Different amounts of MTV RNA are detected in mammary glands at first lactation of B10 and GR-Mtv-2- mice, even though both contain four copies of MTV proviral sequences. This indicates a difference between these two mouse strains either in the regulation of expression of these MTV proviral sequences or in the location of these sequences in the murine genome. PMID:209461

  3. Highly Sensitive Detection of Deoxyribonucleic Acid Hybridization Using Au-Gated AlInN/GaN High Electron Mobility Transistor-Based Sensors

    NASA Astrophysics Data System (ADS)

    Zhan, Xiang-Mi; Hao, Mei-Lan; Wang, Quan; Li, Wei; Xiao, Hong-Ling; Feng, Chun; Jiang, Li-Juan; Wang, Cui-Mei; Wang, Xiao-Liang; Wang, Zhan-Guo

    2017-03-01

    Gallium nitride- (GaN) based high electron mobility transistors (HEMTs) provide a good platform for biological detection. In this work, both Au-gated AlInN/GaN HEMT and AlGaN/GaN HEMT biosensors are fabricated for the detection of deoxyribonucleic acid (DNA) hybridization. The Au-gated AlInN/GaN HEMT biosensor exhibits higher sensitivity in comparison with the AlGaN/GaN HEMT biosensor. For the former, the drain-source current ( {V}{DS}=0.5 V) shows a clear decrease of 69 μA upon the introduction of 1 μmolL {}-1 (μM) complimentary DNA to the probe DNA at the sensor area, while for the latter it is only 38 μA. This current reduction is a notable indication of the hybridization. The high sensitivity can be attributed to the thinner barrier of the AlInN/GaN heterostructure, which makes the two-dimensional electron gas channel more susceptible to a slight change of the surface charge. Supported by the National Key Research and Development Program of China under Grant Nos 2016YFB0400104 and 2016YFB0400301, the National Natural Sciences Foundation of China under Grant No 61334002, and the National Science and Technology Major Project.

  4. Alternative hybrid electrolytes based on a series of bis(trialkoxysilyl)alkanes and 3-(trihydroxysilyl)-1-propane sulfonic acid applied in gas diffusion electrodes of proton exchange membrane fuel cells

    NASA Astrophysics Data System (ADS)

    Lin, C. W.; Chung, L. C.; Veerapur, R. S.; Yang, F. C.

    This study demonstrates a method for improving the electrolyte distribution in catalyst layers and enhancing the utilization of catalyst existing in primary pores. Bis(trialkoxysilyl)alkanes (BTAS-alkanes) and 3-(trihydroxysilyl)-1-propane sulfonic acid (THS)Pro-SO 3H) precursors have been used to prepare a series of hybrid electrolytes with various organic segment lengths of BTAS-alkanes and ratios of organic moiety and sulfonic acid groups. Investigations of BTAS-alkanes series includes bis(triethoxysilyl)octane (BTES-Oct), bis(trimethoxysilyl)hexane (BTMS-Hex), and bis(triethoxysilyl)ethane (BTES-Eth). Small angle X-ray spectroscopy (SAXS) identifies morphological phase separation in BTES-Oct and BTMS-Hex hybrid electrolytes. The results of mercury porosimetry and BET porosimetry show that the hybrid electrolytes have better capability than Nafion ionomer to penetrate into primary pores of the catalyst layers. Electrochemical measurements including electrode polarization, electrochemical active surface (EAS) and electrochemical impedance spectroscopy (EIS) are discussed. The BTES-Oct or BTMS-Hex hybrid electrolytes with higher ratio of organic moiety and sulfonic acid group have achieved better electrode performance. Oxygen benefit current (OBC) results indicate that higher ratios of BTES-Oct/(THS)Pro-SO 3H provides higher hydrophobicity with better gas transport properties. However, the hybrid electrodes exhibit lower cathode performance than Nafion ®-based electrodes due to excessive electrolyte incorporated in the catalyst layer.

  5. Hybrid Quantum Mechanics/Molecular Mechanics-Based Molecular Dynamics Simulation of Acid-Catalyzed Dehydration of Polyols in Liquid Water

    SciTech Connect

    Caratzoulas, Stavros; Courtney, Timothy; Vlachos, Dionisios G.

    2011-01-01

    We use the conversion of protonated glycerol to acrolein for a case study of the mechanism of acid-catalyzed dehydration of polyols in aqueous environments. We employ hybrid Quamtum Mechanics/Molecular Mechanics Molecular Dynamics (QM/MM MD) simulations with biased sampling and perform free energy calculations for the elementary steps of the reaction. We investigate the effects of solvent dynamics and in particular the role of quantum mechanical water in the dehydration mechanism. We present results supporting a mechanism that proceeds via water-mediated proton transfers and thus through an enol intermediate. We find that the first dehydration may take place by two, low-energy pathways requiring, respectively, 20.9 and 18.8 kcal/mol of activation free energy. The second dehydration requires 19.9 kcal/mol of activation free energy while for the overall reaction we compute a free energy change of -8 kcal/mol.

  6. Template-free synthesis of a porous organic-inorganic hybrid tin(IV) phosphonate and its high catalytic activity for esterification of free fatty acids.

    PubMed

    Dutta, Arghya; Patra, Astam K; Uyama, Hiroshi; Bhaumik, Asim

    2013-10-23

    Here we have synthesized an organic-inorganic hybrid mesoporous tin phosphonate monolith (MLSnP-1) with crystalline pore walls by a template-free sol-gel route. N2 sorption analysis shows Brunauer-Emmett-Teller (BET) surface area of 347 m2 g(-1). Wide-angle powder X-ray diffraction (PXRD) pattern shows few broad diffraction peaks indicating crystalline pore wall of the material. High-resolution transmission electron microscopic (HR TEM) image further reveals the crystal fringes on the pore wall. Framework bonding and local environment around phosphorus and carbon were examined by Fourier transform infrared (FT IR) spectroscopy and solid-state MAS NMR spectroscopy. The material exhibits remarkable catalytic activity for esterification of long chain fatty acids under mild reaction conditions at room temperature.

  7. The genome sequence of the highly acetic acid-tolerant Zygosaccharomyces bailii-derived interspecies hybrid strain ISA1307, isolated from a sparkling wine plant.

    PubMed

    Mira, Nuno P; Münsterkötter, Martin; Dias-Valada, Filipa; Santos, Júlia; Palma, Margarida; Roque, Filipa C; Guerreiro, Joana F; Rodrigues, Fernando; Sousa, Maria João; Leão, Cecília; Güldener, Ulrich; Sá-Correia, Isabel

    2014-06-01

    In this work, it is described the sequencing and annotation of the genome of the yeast strain ISA1307, isolated from a sparkling wine continuous production plant. This strain, formerly considered of the Zygosaccharomyces bailii species, has been used to study Z. bailii physiology, in particular, its extreme tolerance to acetic acid stress at low pH. The analysis of the genome sequence described in this work indicates that strain ISA1307 is an interspecies hybrid between Z. bailii and a closely related species. The genome sequence of ISA1307 is distributed through 154 scaffolds and has a size of around 21.2 Mb, corresponding to 96% of the genome size estimated by flow cytometry. Annotation of ISA1307 genome includes 4385 duplicated genes (∼ 90% of the total number of predicted genes) and 1155 predicted single-copy genes. The functional categories including a higher number of genes are 'Metabolism and generation of energy', 'Protein folding, modification and targeting' and 'Biogenesis of cellular components'. The knowledge of the genome sequence of the ISA1307 strain is expected to contribute to accelerate systems-level understanding of stress resistance mechanisms in Z. bailii and to inspire and guide novel biotechnological applications of this yeast species/strain in fermentation processes, given its high resilience to acidic stress. The availability of the ISA1307 genome sequence also paves the way to a better understanding of the genetic mechanisms underlying the generation and selection of more robust hybrid yeast strains in the stressful environment of wine fermentations.

  8. The Genome Sequence of the Highly Acetic Acid-Tolerant Zygosaccharomyces bailii-Derived Interspecies Hybrid Strain ISA1307, Isolated From a Sparkling Wine Plant

    PubMed Central

    Mira, Nuno P.; Münsterkötter, Martin; Dias-Valada, Filipa; Santos, Júlia; Palma, Margarida; Roque, Filipa C.; Guerreiro, Joana F.; Rodrigues, Fernando; Sousa, Maria João; Leão, Cecília; Güldener, Ulrich; Sá-Correia, Isabel

    2014-01-01

    In this work, it is described the sequencing and annotation of the genome of the yeast strain ISA1307, isolated from a sparkling wine continuous production plant. This strain, formerly considered of the Zygosaccharomyces bailii species, has been used to study Z. bailii physiology, in particular, its extreme tolerance to acetic acid stress at low pH. The analysis of the genome sequence described in this work indicates that strain ISA1307 is an interspecies hybrid between Z. bailii and a closely related species. The genome sequence of ISA1307 is distributed through 154 scaffolds and has a size of around 21.2 Mb, corresponding to 96% of the genome size estimated by flow cytometry. Annotation of ISA1307 genome includes 4385 duplicated genes (∼90% of the total number of predicted genes) and 1155 predicted single-copy genes. The functional categories including a higher number of genes are ‘Metabolism and generation of energy’, ‘Protein folding, modification and targeting’ and ‘Biogenesis of cellular components’. The knowledge of the genome sequence of the ISA1307 strain is expected to contribute to accelerate systems-level understanding of stress resistance mechanisms in Z. bailii and to inspire and guide novel biotechnological applications of this yeast species/strain in fermentation processes, given its high resilience to acidic stress. The availability of the ISA1307 genome sequence also paves the way to a better understanding of the genetic mechanisms underlying the generation and selection of more robust hybrid yeast strains in the stressful environment of wine fermentations. PMID:24453040

  9. Improved selective cholesterol adsorption by molecularly imprinted poly(methacrylic acid)/silica (PMAA-SiO₂) hybrid material synthesized with different molar ratios.

    PubMed

    Clausen, Débora Nobile; Pires, Igor Matheus Ruiz; Tarley, César Ricardo Teixeira

    2014-11-01

    The present paper describes the synthesis of molecularly imprinted polymer - poly(methacrylic acid)/silica and reports its performance feasibility with desired adsorption capacity and selectivity for cholesterol extraction. Two imprinted hybrid materials were synthesized at different methacrylic acid (MAA)/tetraethoxysilane (TEOS) molar ratios (6:1 and 1:5) and characterized by FT-IR, TGA, SEM and textural data. Cholesterol adsorption on hybrid materials took place preferably in apolar solvent medium, especially in chloroform. From the kinetic data, the equilibrium time was reached quickly, being 12 and 20 min for the polymers synthesized at MAA/TEOS molar ratio of 6:1 and 1:5, respectively. The pseudo-second-order model provided the best fit for cholesterol adsorption on polymers, confirming the chemical nature of the adsorption process, while the dual-site Langmuir-Freundlich equation presented the best fit to the experimental data, suggesting the existence of two kinds of adsorption sites on both polymers. The maximum adsorption capacities obtained for the polymers synthesized at MAA/TEOS molar ratios of 6:1 and 1:5 were found to be 214.8 and 166.4 mg g(-1), respectively. The results from isotherm data also indicated higher adsorption capacity for both imprinted polymers regarding to corresponding non-imprinted polymers. Nevertheless, taking into account the retention parameters and selectivity of cholesterol in the presence of structurally analogue compounds (5-α-cholestane and 7-dehydrocholesterol), it was observed that the polymer synthesized at the MAA/TEOS molar ratio of 6:1 was much more selective for cholesterol than the one prepared at the ratio of 1:5, thus suggesting that selective binding sites ascribed to the carboxyl group from MAA play a central role in the imprinting effect created on MIP.

  10. Hybrid polypeptides having cellobiohydrolase activity and polynucleotides encoding same

    SciTech Connect

    Liu, Ye; Shaghasi, Tarana

    2016-11-01

    The present invention provides hybrid polypeptides having cellobiohydrolase activity. The present invention also provides polynucleotides encoding the hybrid polypeptides; nucleic acid constructs, vectors and host cells comprising the polynucleotides; and processes of using the hybrid polypeptides.

  11. Production of hybrid diesel fuel precursors from carbohydrates and petrochemicals using formic acid as a reactive solvent.

    PubMed

    Zhou, Xiaoyuan; Rauchfuss, Thomas B

    2013-02-01

    We report the one-pot alkylation of mesitylene with carbohydrate-derived 5-(hydroxymethyl)furfural (HMF) as a step toward diesel-range liquids. Using FeCl(3) as a catalyst, HMF is shown to alkylate toluene, xylene, and mesitylene in high yields in CH(2)Cl(2) and MeNO(2) solvents. Efforts to extend this reaction to greener or safer solvents showed that most ether-based solvents are unsatisfactory. Acid catalysts (e.g, p-TsOH) also proved to be ineffective. Using formic acid as a reactive solvent, mesitylene could be alkylated to give mesitylmethylfurfural (MMF) starting from fructose with yields up to approximately 70 %. The reaction of fructose with formic acid in the absence of mesitylene gave rise to low yields of the formate ester of HMF, which indicates the stabilizing effect of replacing the hydroxyl substituent with mesityl. The arene also serves as a second phase into which the product is extracted. Even by using formic acid, the mesitylation of less expensive precursors such as glucose and cellulose proceeded only in modest yields (ca. 20 %). These simpler substrates were found to undergo mesitylation by using hydrogen chloride/formic acid via the intermediate chloromethylfurfural. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Nucleic acid spot hybridization based species-specific detection of Sclerotium rolfsii associated with collar rot disease of Amorphophallus paeoniifolius.

    PubMed

    Pravi, V; Jeeva, M L; Archana, P V

    2015-02-01

    Collar rot is one of the most destructive and prevalent disease of Amorphophallus paeoniifolius, resulting in heavy yield losses. The causative organism, Sclerotium rolfsii is a soil-borne polyphagous fungus characterized by prolific growth and ability to produce persistent sclerotia. The pathogen propagules surviving in soil and planting material are the major sources of inoculum. This study presents the suitability of DNA hybridization technique for species specific detection of S. rolfsii in soil and planting material. The detection limit of the probe was 10-15 pg of pure pathogen DNA. The developed probe was found to be highly specific and could be used for accurate identification of pathogen up to the species level. The protocol was standardized for detection of the pathogen in naturally infected field samples.

  13. Efficient conversion of furfuryl alcohol into alkyl levulinates catalyzed by an organic-inorganic hybrid solid acid catalyst.

    PubMed

    Zhang, Zehui; Dong, Kun; Zhao, Zongbao Kent

    2011-01-17

    A clean, facile, and environment-friendly catalytic method has been developed for the conversion of furfuryl alcohol into alkyl levulinates making use of the novel solid catalyst methylimidazolebutylsulfate phosphotungstate ([MIMBS]₃PW₁₂O₄₀). The solid catalyst is an organic-inorganic hybrid material, which consists of an organic cation and an inorganic anion. A study for optimizing the reaction conditions such as the reaction time, the temperature and the catalyst loading has been performed. Under optimal conditions, a high n-butyl levulinate yield of up to 93 % is obtained. Furthermore, the kinetics of the reaction pathways and the mechanism for the alcoholysis of furfuryl alcohol are discussed. This method is environmentally benign and economical for the conversion of biomass-based derivatives into fine chemicals.

  14. Arginine-Glycine-Aspartic Acid-Modified Lipid-Polymer Hybrid Nanoparticles for Docetaxel Delivery in Glioblastoma Multiforme.

    PubMed

    Shi, Kairong; Zhou, Jin; Zhang, Qianyu; Gao, Huile; Liu, Yayuan; Zong, Taili; He, Qin

    2015-03-01

    Hybrid nanoparticles consisting of lipids and the biodegradable polymer, poly (D,L-lactide-co-glycolide) (PLGA), were developed for the targeted delivery of the anticancer drug, docetaxel. Transmission electron microscopic observations confirmed the presence of a lipid coating over the polymeric core. Using coumarin-6 as a fluorescent probe, the uptake efficacy of RGD conjugated lipid coated nanoparticles (RGD-L-P) by C6 cells was increased significantly, compared with that of lipid-polymer hybrid nanoparticles (L-P; 2.5-fold higher) or PLGA-nanoparticles (PLGA-P; 1.76-fold higher). The superior tumor spheroid penetration of RGD-L-P indicated that RGD-L-P could target effectively and specifically to C6 cells overexpressing integrin α(v)β3. The anti-proliferative activity of docetaxel-loaded RGD-L-P against C6 cells was increased 2.69- and 4.13-fold compared with L-P and PLGA-P, respectively. Regarding biodistribution, the strongest brain-localized fluorescence signals were detected in glioblastoma multiforme (GBM)-bearing rats treated with 1,10-Dioctadecyl-3,3,30,30-tetramethylindotricarb-ocyanine iodide (DiR)-loaded RGD-L-P, compared to rats treated with DiR-loaded L-P or PLGA-P. The median survival time of GBM-bearing rats treated with docetaxel-loaded RGD-L-P was 57 days, a fold increase of 1.43, 1.78, 3.35, and 3.56 compared with animals given L-P (P < 0.05), PLGA-P (P < 0.05), Taxotere (P < 0.01) and saline (P < 0.01), respectively. Collectively, these results support RGD-L-P as a promising drug delivery system for the specific targeting and the treatment of GBM.

  15. Recognition of RNA by amide modified backbone nucleic acids: molecular dynamics simulations of DNA-RNA hybrids in aqueous solution.

    PubMed

    Nina, Mafalda; Fonné-Pfister, Raymonde; Beaudegnies, Renaud; Chekatt, Habiba; Jung, Pierre M J; Murphy-Kessabi, Fiona; De Mesmaeker, Alain; Wendeborn, Sebastian

    2005-04-27

    Thermodynamic and structural properties of a chemically modified DNA-RNA hybrid in which a phosphodiester linkage is replaced by a neutral amide-3 linkage (3'-CH(2)-CONH-5') were investigated using UV melting experiments, molecular dynamics simulations in explicit water, and continuum solvent models. van't Hoff analysis of the experimental UV melting curves suggests that the significant increase of the thermodynamic stability of a 15-mer DNA-RNA with seven alternated amide-3 modifications (+11 degrees C) is mainly due to an increased binding enthalpy. To further evaluate the origin in the observed affinities differences, the electrostatic contribution to the binding free energy was calculated by solving the Poisson-Boltzmann equation numerically. The nonelectrostatic contribution was estimated as the product of a hydrophobic surface tension coefficient and the surface area that is buried upon double strand formation. Structures were taken from 10 ns molecular dynamics simulations computed in a consistent fashion using explicit solvent, counterions, and the particle-mesh Ewald procedure. The present preliminary thermodynamic study suggests that the favorable binding free energy of the amide-3 DNA single strand to the complementary RNA is equally driven by electrostatic and nonpolar contributions to the binding compared to their natural analogues. In addition, molecular dynamics simulations in explicit water were performed on an amide-3 DNA single strand and the corresponding natural DNA. Results from the conformations cluster analysis of the simulated amide-3 DNA single strand ensembles suggest that the 25% of the population sampled within 10 ns has a pre-organized conformation where the sugar C3' endo pucker is favored at the 3'-flanking nucleotides. These structural and thermodynamic features contribute to the understanding of the observed increased affinities of the amide-3 DNA-RNA hybrids at the microscopic level.

  16. Effect of tannic acid-fish scale gelatin hydrolysate hybrid nanoparticles on intestinal barrier function and α-amylase activity.

    PubMed

    Wu, Shao-Jung; Ho, Yi-Cheng; Jiang, Shun-Zhou; Mi, Fwu-Long

    2015-07-01

    Practical application of tannic acid is limited because it readily binds proteins to form insoluble aggregates. In this study, tannic acid was self-assembled with fish scale gelatin hydrolysates (FSGH) to form stable colloidal complex nanoparticles. The nanoparticles prepared from 4 mg ml(-1) tannic acid and 4 mg ml(-1) FSGH had a mean particle size of 260.8 ± 3.6 nm, and showed a positive zeta potential (20.4 ± 0.4 mV). The nanoparticles acted as effective nano-biochelators and free radical scavengers because they provided a large number of adsorption sites for interaction with heavy metal ions and scavenging free radicals. The maximum adsorption capacity for Cu(2+) ions was 123.5 mg g(-1) and EC50 of DPPH radical scavenging activity was 21.6 ± 1.2 μg ml(-1). Hydroxyl radical scavenging effects of the nanoparticles were investigated by electron spin resonance spectroscopy. The copper-chelating capacity and free radical scavenging activity of the nanoparticles were associated with their capacity to inhibit Cu(2+) ion-induced barrier impairment and hyperpermeability of Caco-2 intestinal epithelial tight junction (TJ). However, α-amylase inhibitory activity of the nanoparticles was significantly lower than that of free tannic acid. The results suggest that the nanoparticles can ameliorate Cu(2+) ion induced intestinal epithelial TJ dysfunction without severely inhibiting the activity of the digestive enzymes.

  17. Synthesis and characterization of conformationally preorganized, (R)-diethylene glycol-containing γ-peptide nucleic acids with superior hybridization properties and water solubility.

    PubMed

    Sahu, Bichismita; Sacui, Iulia; Rapireddy, Srinivas; Zanotti, Kimberly J; Bahal, Raman; Armitage, Bruce A; Ly, Danith H

    2011-07-15

    Developed in the early 1990s, peptide nucleic acid (PNA) has emerged as a promising class of nucleic acid mimic because of its strong binding affinity and sequence selectivity toward DNA and RNA and resistance to enzymatic degradation by proteases and nucleases; however, the main drawbacks, as compared to other classes of oligonucleotides, are water solubility and biocompatibility. Herein we show that installation of a relatively small, hydrophilic (R)-diethylene glycol ("miniPEG", R-MP) unit at the γ-backbone transforms a randomly folded PNA into a right-handed helix. Synthesis of optically pure (R-MP)γPNA monomers is described, which can be accomplished in a few simple steps from a commercially available and relatively cheap Boc-l-serine. Once synthesized, (R-MP)γPNA oligomers are preorganized into a right-handed helix, hybridize to DNA and RNA with greater affinity and sequence selectivity, and are more water soluble and less aggregating than the parental PNA oligomers. The results presented herein have important implications for the future design and application of PNA in biology, biotechnology, and medicine, as well as in other disciplines, including drug discovery and molecular engineering.

  18. The perils of pathogen discovery: origin of a novel parvovirus-like hybrid genome traced to nucleic acid extraction spin columns.

    PubMed

    Naccache, Samia N; Greninger, Alexander L; Lee, Deanna; Coffey, Lark L; Phan, Tung; Rein-Weston, Annie; Aronsohn, Andrew; Hackett, John; Delwart, Eric L; Chiu, Charles Y

    2013-11-01

    Next-generation sequencing was used for discovery and de novo assembly of a novel, highly divergent DNA virus at the interface between the Parvoviridae and Circoviridae. The virus, provisionally named parvovirus-like hybrid virus (PHV), is nearly identical by sequence to another DNA virus, NIH-CQV, previously detected in Chinese patients with seronegative (non-A-E) hepatitis. Although we initially detected PHV in a wide range of clinical samples, with all strains sharing ∼99% nucleotide and amino acid identity with each other and with NIH-CQV, the exact origin of the virus was eventually traced to contaminated silica-binding spin columns used for nucleic acid extraction. Definitive confirmation of the origin of PHV, and presumably NIH-CQV, was obtained by in-depth analyses of water eluted through contaminated spin columns. Analysis of environmental metagenome libraries detected PHV sequences in coastal marine waters of North America, suggesting that a potential association between PHV and diatoms (algae) that generate the silica matrix used in the spin columns may have resulted in inadvertent viral contamination during manufacture. The confirmation of PHV/NIH-CQV as laboratory reagent contaminants and not bona fide infectious agents of humans underscores the rigorous approach needed to establish the validity of new viral genomes discovered by next-generation sequencing.

  19. Facile "one-pot" synthesis of poly(methacrylic acid)-based hybrid monolith via thiol-ene click reaction for hydrophilic interaction chromatography.

    PubMed

    Lv, Xumei; Tan, Wangming; Chen, Ye; Chen, Yingzhuang; Ma, Ming; Chen, Bo; Yao, Shouzhuo

    2016-07-08

    A novel sol-gel "one-pot" approach in tandem with a radical-mediated thiol-ene reaction for the synthesis of a methacrylic acid-based hybrid monolith was developed. The polymerization monomers, tetramethoxysilane (TMOS) and 3-mercaptopropyl trimethoxysilane (MPTS), were hydrolyzed in high-concentration methacrylic acid solution that also served as a hydrophilic functional monomer. The resulting solution was then mixed with initiator (2, 2'-azobis (2-methylpropionamide) dihydrochloride) and porogen (urea, polyethylene glycol 20,000) in a capillary column and polymerized in water bath. The column had a uniform porous structure and a good permeability. The evaluation of the monolith was performed by separation of small molecules including nucleosides, phenols, amides, bases and Triton X-100. The calibration curves for uridine, inosine, adenosine and cytidine were determined. All the calibration curves exhibited good linear regressions (R(2)≥0.995) within the test ranges of 0.5-40μg/mL for four nucleosides. Additionaliy, atypical hydrophilic mechanism was proved by elution order from low to high according to polarity retention time increased with increases in the content of the organic solvent in the mobile phase. Further studies indicated that hydrogen bond and electrostatic interactions existed between the polar analytes and the stationary phase. This was the mechanism of retention. The excellent separation of the BSA digest showed good hydrophility of the column and indicated the potential in separation of complex biological samples. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Organic inorganic hybrid materials based on polyaniline/TiO2 nanocomposites for ascorbic acid fuel cell systems

    NASA Astrophysics Data System (ADS)

    Ganesan, Raman; Gedanken, Aharon

    2008-10-01

    Polyaniline was grafted onto a mixture of rutile and anatase TiO2 nanoparticles by in situ chemical oxidative polymerization. These nanocomposites were characterized by carbon, hydrogen and nitrogen (CHN) analysis, x-ray diffraction (XRD), Fourier transform infrared (FTIR), ultraviolet-visible (UV-vis), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analysis. FTIR and UV-vis confirm the formation of polyaniline on TiO2 nanoparticles. The TEM shows that the composites consist of PANI and TiO2 nanoparticles. Compared to the neat polyaniline, PANI/TiO2 composites show a higher capacitance and also a higher activity per mass of polyaniline. Since the PANI/TiO2 composites are stable during the electrooxidation of ascorbic acid, they can be used as an alternative catalyst for direct ascorbic acid fuel cells.

  1. Organic-inorganic hybrid materials based on polyaniline/TiO(2) nanocomposites for ascorbic acid fuel cell systems.

    PubMed

    Ganesan, Raman; Gedanken, Aharon

    2008-10-29

    Polyaniline was grafted onto a mixture of rutile and anatase TiO(2) nanoparticles by in situ chemical oxidative polymerization. These nanocomposites were characterized by carbon, hydrogen and nitrogen (CHN) analysis, x-ray diffraction (XRD), Fourier transform infrared (FTIR), ultraviolet-visible (UV-vis), scanning electron microscopy (SEM) and transmission electron microscopy (TEM) analysis. FTIR and UV-vis confirm the formation of polyaniline on TiO(2) nanoparticles. The TEM shows that the composites consist of PANI and TiO(2) nanoparticles. Compared to the neat polyaniline, PANI/TiO(2) composites show a higher capacitance and also a higher activity per mass of polyaniline. Since the PANI/TiO(2) composites are stable during the electrooxidation of ascorbic acid, they can be used as an alternative catalyst for direct ascorbic acid fuel cells.

  2. Poly (N-isopropylacrylamide)-co-(acrylic acid) microgel/Ag nanoparticle hybrids for the colorimetric sensing of H2O2

    NASA Astrophysics Data System (ADS)

    Han, De-Man; Matthew Zhang, Qiang; Serpe, Michael J.

    2015-01-01

    Poly (N-isopropylacrylamide)-co-(acrylic acid) (pNIPAm-co-AAc) microgels composed of Ag nanoparticles (Ag NPs) have been synthesized and employed for the colorimetric sensing of H2O2. Each pNIPAm-co-AAc microgel, which exhibited a diameter of ~800 nm, contained multiple Ag NPs (diameter of ~5 nm), and solutions of these hybrid materials showed a UV-vis absorption band at ~400 nm. This is due to the excitation of the Ag NP surface plasmon. We go on to show that the intensity of this absorption band is dependent on the concentration of H2O2 in solution. Specifically, in the presence of H2O2 the magnitude of the absorption peak dramatically decreases in a linear fashion over the concentration range of 0.30 to 3.00 μM H2O2 (r2 = 0.9918). We go on to show that the response is selective for H2O2 and can still function in complex mixtures, e.g., we showed that the response is still robust in milk samples. While Ag NPs themselves can exhibit similar responses, this system has many benefits including sample processing and long term stability - i.e., Ag NPs are destabilized in solutions of a certain pH, and aggregate readily. Our microgel/Ag NP hybrids have been shown to be extremely stable and are easily purified prior to use by simple centrifugation/washing protocols. This system is simple and straightforward to use, is low cost, and can be used in complex media, which makes it practical for analyzing complex biological and environmental samples.Poly (N-isopropylacrylamide)-co-(acrylic acid) (pNIPAm-co-AAc) microgels composed of Ag nanoparticles (Ag NPs) have been synthesized and employed for the colorimetric sensing of H2O2. Each pNIPAm-co-AAc microgel, which exhibited a diameter of ~800 nm, contained multiple Ag NPs (diameter of ~5 nm), and solutions of these hybrid materials showed a UV-vis absorption band at ~400 nm. This is due to the excitation of the Ag NP surface plasmon. We go on to show that the intensity of this absorption band is dependent on the concentration

  3. Focused upon hybridization: rapid and high sensitivity detection of DNA using isotachophoresis and peptide nucleic acid probes.

    PubMed

    Ostromohov, Nadya; Schwartz, Ortal; Bercovici, Moran

    2015-09-15

    We present a novel assay for rapid and high sensitivity detection of nucleic acids without amplification. Utilizing the neutral backbone of peptide nucleic acids (PNA), our method is based on the design of low electrophoretic mobility PNA probes, which do not focus under isotachophoresis (ITP) unless bound to their target sequence. Thus, background noise associated with free probes is entirely eliminated, significantly improving the signal-to-noise ratio while maintaining a simple single-step assay requiring no amplification steps. We provide a detailed analytical model and experimentally demonstrate the ability to detect targets as short as 17 nucleotides (nt) and a limit of detection of 100 fM with a dynamic range of 5 decades. We also demonstrate that the assay can be successfully implemented for detection of DNA in human serum without loss of signal. The assay requires 15 min to complete, and it could potentially be used in applications where rapid and highly sensitive amplification-free detection of nucleic acids is desired.

  4. Synthesis, hybridization characteristics, and fluorescence properties of oligonucleotides modified with nucleobase-functionalized locked nucleic acid adenosine and cytidine monomers.

    PubMed

    Kaura, Mamta; Kumar, Pawan; Hrdlicka, Patrick J

    2014-07-03

    Conformationally restricted nucleotides such as locked nucleic acid (LNA) are very popular as affinity-, specificity-, and stability-enhancing modifications in oligonucleotide chemistry to produce probes for nucleic acid targeting applications in molecular biology, biotechnology, and medicinal chemistry. Considerable efforts have been devoted in recent years to optimize the biophysical properties of LNA through additional modification of the sugar skeleton. We recently introduced C5-functionalization of LNA uridines as an alternative and synthetically more straightforward approach to improve the biophysical properties of LNA. In the present work, we set out to test the generality of this concept by studying the characteristics of oligonucleotides modified with four different C5-functionalized LNA cytidine and C8-functionalized LNA adenosine monomers. The results strongly suggest that C5-functionalization of LNA pyrimidines is indeed a viable approach for improving the binding affinity, target specificity, and/or enzymatic stability of LNA-modified ONs, whereas C8-functionalization of LNA adenosines is detrimental to binding affinity and specificity. These insights will impact the future design of conformationally restricted nucleotides for nucleic acid targeting applications.

  5. Poly (N-isopropylacrylamide)-co-(acrylic acid) microgel/Ag nanoparticle hybrids for the colorimetric sensing of H2O2.

    PubMed

    Han, De-Man; Zhang, Qiang Matthew; Serpe, Michael J

    2015-02-14

    Poly (N-isopropylacrylamide)-co-(acrylic acid) (pNIPAm-co-AAc) microgels composed of Ag nanoparticles (Ag NPs) have been synthesized and employed for the colorimetric sensing of H2O2. Each pNIPAm-co-AAc microgel, which exhibited a diameter of ∼800 nm, contained multiple Ag NPs (diameter of ∼5 nm), and solutions of these hybrid materials showed a UV-vis absorption band at ∼400 nm. This is due to the excitation of the Ag NP surface plasmon. We go on to show that the intensity of this absorption band is dependent on the concentration of H2O2 in solution. Specifically, in the presence of H2O2 the magnitude of the absorption peak dramatically decreases in a linear fashion over the concentration range of 0.30 to 3.00 μM H2O2 (r(2) = 0.9918). We go on to show that the response is selective for H2O2 and can still function in complex mixtures, e.g., we showed that the response is still robust in milk samples. While Ag NPs themselves can exhibit similar responses, this system has many benefits including sample processing and long term stability - i.e., Ag NPs are destabilized in solutions of a certain pH, and aggregate readily. Our microgel/Ag NP hybrids have been shown to be extremely stable and are easily purified prior to use by simple centrifugation/washing protocols. This system is simple and straightforward to use, is low cost, and can be used in complex media, which makes it practical for analyzing complex biological and environmental samples.

  6. Cellular delivery of quantum dot-bound hybridization probe for detection of intracellular pre-microRNA using chitosan/poly(γ-glutamic acid) complex as a carrier.

    PubMed

    Geng, Yao; Lin, Dajie; Shao, Lijia; Yan, Feng; Ju, Huangxian

    2013-01-01

    A quantum dot (QD)-bound hybridization probe was designed for detection of intracellular pre-miRNA using chitosan (CS)/poly(γ-glutamic acid) (γ-PGA) complex as a gene vector. The probe was prepared by assembling thiolated RNA to gold nanoparticle (Au NP) via Au-S bond and then binding 3'-end amine of the RNA to the carboxy group capped on quantum dot surface. The QD-RNA-Au NP probe was assembled on the vector by mixing with aqueous γ-PGA solution and then CS solution to construct a gene delivery system for highly effective cellular uptake and delivery. After the probe was released from CS/γ-PGA complex to the cytoplasm by electrostatic repulsion at intracellular pH, it hybridized with pre-miRNA precursor as target. The formed product was then cleaved by RNase III Dicer, leading to the separation of QDs from Au NPs and fluorescence emission of QDs, which could be detected by confocal microscopic imaging to monitor the amount of the intracellular pre-miRNA precursor. The in vitro assays revealed that the QD-RNA-Au NP was a robust, sensitive and selective probe for quantitative detection of target pre-miRNA. Using MDA-MB231 and MCF-7 breast cancer cells as models, the relative amount of pre-miRNA let-7a could be successfully compared. Since the amount of miRNA is related to the progress and prognosis of cancer, this strategy could be expected to hold promising application potential in medical research and clinical diagnostics.

  7. Optimization of peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) for the detection of bacteria: The effect of pH, dextran sulfate and probe concentration.

    PubMed

    Rocha, Rui; Santos, Rita S; Madureira, Pedro; Almeida, Carina; Azevedo, Nuno F

    2016-05-20

    Fluorescence in situ hybridization (FISH) is a molecular technique widely used for the detection and characterization of microbial populations. FISH is affected by a wide variety of abiotic and biotic variables and the way they interact with each other. This is translated into a wide variability of FISH procedures found in the literature. The aim of this work is to systematically study the effects of pH, dextran sulfate and probe concentration in the FISH protocol, using a general peptide nucleic acid (PNA) probe for the Eubacteria domain. For this, response surface methodology was used to optimize these 3 PNA-FISH parameters for Gram-negative (Escherichia coli and Pseudomonas fluorescens) and Gram-positive species (Listeria innocua, Staphylococcus epidermidis and Bacillus cereus). The obtained results show that a probe concentration higher than 300nM is favorable for both groups. Interestingly, a clear distinction between the two groups regarding the optimal pH and dextran sulfate concentration was found: a high pH (approx. 10), combined with lower dextran sulfate concentration (approx. 2% [w/v]) for Gram-negative species and near-neutral pH (approx. 8), together with higher dextran sulfate concentrations (approx. 10% [w/v]) for Gram-positive species. This behavior seems to result from an interplay between pH and dextran sulfate and their ability to influence probe concentration and diffusion towards the rRNA target. This study shows that, for an optimum hybridization protocol, dextran sulfate and pH should be adjusted according to the target bacteria. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Preparation of high efficiency and low carry-over immobilized enzymatic reactor with methacrylic acid-silica hybrid monolith as matrix for on-line protein digestion.

    PubMed

    Yuan, Huiming; Zhang, Lihua; Zhang, Yukui

    2014-12-05

    In this work, a novel kind of organic-silica hybrid monolith based immobilized enzymatic reactor (IMER) was developed. The monolithic support was prepared by a single step "one-pot" strategy via the polycondensation of tetramethoxysilane and vinyltrimethoxysilane and in situ copolymerization of methacrylic acid and vinyl group on the precondensed siloxanes with ammonium persulfate as the thermal initiator. Subsequently, the monolith was activated by N-(3-dimethylaminopropyl) - N'-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS), followed by the modification of branched polyethylenimine (PEI) to improve the hydrophilicity. Finally, after activated by EDC and NHS, trypsin was covalently immobilized onto the monolithic support. The performance of such a microreactor was evaluated by the in sequence digestion of bovine serum albumin (BSA) and myoglobin, followed by MALDI-TOF-MS analysis. Compared to those obtained by traditional in-solution digestion, not only higher sequence coverages for BSA (74±1.4% vs. 59.5±2.7%, n=6) and myoglobin (93±3% vs. 81±4.5%, n=6) were obtained, but also the digestion time was shortened from 24h to 2.5 min, demonstrating the high digestion efficiency of such an IMER. The carry-over of these two proteins on the IMER was investigated, and peptides from BSA could not be found in mass spectrum of myoglobin digests, attributed to the good hydrophilicity of our developed monolithic support. Moreover, the dynamic concentration range for protein digestion was proved to be four orders of magnitude, and the IMER could endure at least 7-day consecutive usage. Furthermore, such an IMER was coupled with nano-RPLC-ESI/MS/MS for the analysis of extracted proteins from Escherichia coli. Compared to formerly reported silica hybrid monolith based IMER and the traditional in-solution counterpart, by our developed IMER, although the identified protein number was similar, the identified distinct peptide number was improved by 7% and 25% respectively

  9. The use of activated carbon and graphite for the development of lead-acid batteries for hybrid vehicle applications

    NASA Astrophysics Data System (ADS)

    Fernández, M.; Valenciano, J.; Trinidad, F.; Muñoz, N.

    Future vehicle applications require the development of reliable and long life batteries operating under high-rate partial-state-of-charge (HRPSoC) working conditions. This paper updates work carried out to develop spiral wound valve-regulated batteries for vehicles with different hybridisation degrees, ranging from stop-start to mild hybrid applications. In order to develop a battery that can withstand the hard operating conditions that the work at High Rate Partial-State-of-Charge (HRPSoC) implies, it is necessary to modify the negative AM formulation by using special, additives like carbon and graphite that reduce lead sulphate accumulation during HRPSoC cycling within in the negative plate. Several batches of negative active material (NAM) with the addition of graphites of different types, as well as combinations of graphite and activated carbons, have been made on 6 V 24 Ah Spiral wound modules. Electrical results show a dramatic increase of the charge acceptance at different SoC's that for some combinations approach 200%. On the other hand, on cycle life according to EUCAR Power Assist cycling, values in the range 200,000-220,000 cycles have been obtain in most part of the batch. This represents a capacity turnover of 5000-5500 times the nominal capacity. The paper is divided into three parts. The first part is devoted to identify the cause of failure of the negative plate on Power Assist Cycle Life, that turned to be the development of high amounts of lead sulphate and its accumulation on the surface of the plate. The second part covers the addition of carbon and graphite of low SSA to NAM and finally the third part is dedicated to the test of additions of medium/high SSA carbon to NAM with the specific objective of trying to implement the supercapacitor effect inside the battery.

  10. Array comparative genomic hybridization profiling analysis reveals deoxyribonucleic acid copy number variations associated with premature ovarian failure.

    PubMed

    Aboura, Azzedine; Dupas, Claire; Tachdjian, Gérard; Portnoï, Marie-France; Bourcigaux, Nathalie; Dewailly, Didier; Frydman, René; Fauser, Bart; Ronci-Chaix, Nathalie; Donadille, Bruno; Bouchard, Philippe; Christin-Maitre, Sophie

    2009-11-01

    Premature ovarian failure (POF) is defined by amenorrhea of at least 4- to 6-month duration, occurring before 40 yr of age, with two FSH levels in the postmenopausal range. Its etiology remains unknown in more than 80% of cases. Standard karyotypes, having a resolution of 5-10 Mb, have identified critical chromosomal regions, mainly located on the long arm of the X chromosome. Array comparative genomic hybridization (a-CGH) analysis is able to detect submicroscopic chromosomal rearrangements with a higher genomic resolution. We searched for copy number variations (CNVs), using a-CGH analysis with a resolution of approximately 0.7 Mb, in a cohort of patients with POF. We prospectively included 99 women. Our study included a conventional karyotype and DNA microarrays comprising 4500 bacterial artificial chromosome clones spread on the entire genome. Thirty-one CNVs have been observed, three on the X chromosome and 28 on autosomal chromosomes. Data have been compared to control populations obtained from the Database of Genomic Variants (http://projects.tcag.ca/variation). Eight statistically significantly different CNVs have been identified in chromosomal regions 1p21.1, 5p14.3, 5q13.2, 6p25.3, 14q32.33, 16p11.2, 17q12, and Xq28. We report the first study of CNV analysis in a large cohort of Caucasian POF patients. In the eight statistically significant CNVs we report, we found five genes involved in reproduction, thus representing potential candidate genes in POF. The current study along with emerging information regarding CNVs, as well as data on their potential association with human diseases, emphasizes the importance of assessing CNVs in cohorts of POF women.

  11. Synthesis and DNA-binding properties of novel DNA cyclo-intercalators containing purine-glucuronic acid hybrids.

    PubMed

    Zhang, Renshuai; Chen, Shaopeng; Wang, Xueting; Yu, Rilei; Li, Mingjing; Ren, Sumei; Jiang, Tao

    2016-06-24

    Novel DNA cyclo-intercalators, which incorporated two intercalator subunits linked by two bridges, were synthesized. Binding of the compounds to calf-thymus DNA was studied by fluorescence spectroscopy, and docking simulations were used to predict the binding modes of these cyclic compounds. The spectral data demonstrated that all of these compounds can interact with CT-DNA. The sugar moiety played an important role in the process of binding between the intercalators containing glucuronic acid and DNA. The length and flexibility of the connecting bridges affected the binding affinity of the resultant cyclo-intercalators. Docking simulations showed that compounds 7 and 8 interact with DNA as mono-intercalators.

  12. Poly(citric acid)-block-poly(ethylene glycol) copolymers--new biocompatible hybrid materials for nanomedicine.

    PubMed

    Naeini, Ashkan Tavakoli; Adeli, Mohsen; Vossoughi, Manouchehr

    2010-08-01

    Linear-dendritic ABA triblock copolymers containing poly(ethylene glycol) (PEG) as B block and hyperbranched poly(citric acid) (PCA) as A blocks were synthesized through polycondensation. The molecular self-assembly of synthesized PCA-PEG-PCA copolymers in water led to formation of nanoparticles and fibers in different sizes and shapes depending on the time and size of PCA blocks. Ten days after dissolving PCA-PEG-PCA copolymers in water, the size of fibers had reached several millimeters. Mixing a water solution of fluorescein as a small guest molecule and PCA-PEG-PCA copolymers led to the encapsulation of fluorescein by products of molecular self-assembly. To investigate their potential application in nanomedicine and to understand the limitations and capabilities of these materials as nanoexcipients in biological systems, different types of short-term in vitro cytotoxicity experiments on the HT1080 cell line (human fibrosarcoma) and hemocompatibility tests were performed. From the clinical editor: This manuscript investigates the potentials of linear-dendritic ABA triblock copolymers containing poly(ethylene glycol) (PEG) as B block and hyperbranched poly(citric acid) (PCA) as A blocks for future applications in nanomedicine.

  13. A label-free electrochemical immunosensor for hepatitis B based on hyaluronic acid-carbon nanotube hybrid film.

    PubMed

    Cabral, Diego G A; Lima, Erika C S; Moura, Patrícia; Dutra, Rosa F

    2016-01-01

    An electrochemical immunosensor developed for detection of antibodies to hepatitis B core protein (anti-HBc) is described. Anti-HBc is the earliest serological marker from hepatitis B virus (HBV) infection, remaining all life after contact with virus, being considered the most important marker for uses in screening of blood bank. A nanohybrid surface assembled onto a glassy carbon electrode consisting of amino carbon nanotubes recovered by hyaluronic acid was used as sensing platform to detect the anti-HBc. All the steps of electrode surface modification were characterized by Scanning Electronic Microscopy and extensively evaluated by electrochemical techniques. The electrode response was measured by direct anti-HBc antigen interactions by square wave voltammetry, dispensing uses of label or chemical mediators. Under optimal conditions, the anodic peak current which was proportional to the anti-HBs concentration. The immunosensor response was linear toward anti-HBc in concentrations up to 6 ng mL(-1), with a detection limit of 0.03 ng mL(-1). The linear range achieved was according to clinical level, indicating the immunosensor as promising tool for use as a criterion for blood bag disposal. The enhancement of the hyaluronic acid by carbon nanotube promoted an increase of charge electron transfer, besides a stable platform for HBc. Copyright © 2015 Elsevier B.V. All rights reserved.

  14. Comparative analysis of poly-glycolic acid-based hybrid polymer starter matrices for in vitro tissue engineering.

    PubMed

    Generali, Melanie; Kehl, Debora; Capulli, Andrew K; Parker, Kevin K; Hoerstrup, Simon P; Weber, Benedikt

    2017-07-01

    Biodegradable scaffold matrixes form the basis of any in vitro tissue engineering approach by acting as a temporary matrix for cell proliferation and extracellular matrix deposition until the scaffold is replaced by neo-tissue. In this context several synthetic polymers have been investigated, however a concise systematic comparative analyses is missing. Therefore, the present study systematically compares three frequently used polymers for the in vitro engineering of extracellular matrix based on poly-glycolic acid (PGA) under static as well as dynamic conditions. Ultra-structural analysis was used to examine the polymers structure. For tissue engineering (TE) three human fibroblast cell lines were seeded on either PGA-poly-4-hydroxybutyrate (P4HB), PGA-poly-lactic acid (PLA) or PGA-poly-caprolactone (PCL) patches. These patches were analyzed after 21days of culture qualitative by histology and quantitative by determining the amount of DNA, glycosaminoglycan and hydroxyproline. We found that PGA-P4HB and PGA-PLA scaffolds enhance tissue formation significantly higher than PGA-PCL scaffolds (p<0.05). Polymer remnants were visualized by polarization microscopy. In addition, biomechanical properties of the tissue engineered patches were determined in comparison to native tissue. This study may allow future studies to specifically select certain polymer starter matrices aiming at specific tissue properties of the bioengineered constructs in vitro. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Synthesis, structural studies, and cytotoxic evaluation of novel ursolic acid hybrids with capabilities to arrest breast cancer cells in mitosis.

    PubMed

    Pattnaik, Banita; Lakshmi, Jerripothula K; Kavitha, Rachineni; Jagadeesh, Bharatam; Bhattacharjee, Debanjan; Jain, Nishant; Mallavadhani, Uppuluri V

    2017-03-01

    Some novel chemically modified frameworks of ursolic acid have been designed and synthesized. The key step was the cycloaddition of azidopropyl-3β-hydroxy-urs-12-en-28-oate with the appropriate C28 propargyl esters of ursolic, corosolic, asiatic, oleanolic, and betulinic acid under Click reaction conditions, and the products were obtained in 74-84% yields. In view of their intriguing structural diversity, they have been subjected to detailed 1D and 2D NMR studies and their structures are thoroughly assigned. The synthesized compounds were screened for their anticancer potential against two human breast cancer cell lines (MCF-7 & MDA-MB-231) using sulforhodamine B cell proliferation assay. The GI50 data revealed that the synthesized compounds exhibit highly potent activities against the two tested cell lines. Interestingly, the synthesized compounds showed selectivity and higher activity against MDA-MB-231 cell line than MCF-7. Among the tested compounds, compound 17 is the most potent one with GI50 value of 1.4 ± 0.1 μM and showed 2.9 times more activity than the standard doxorubicin against MDA-MB-231. In addition, 17 arrests cells in mitotic phase of cell cycle, resulting in a change in cell phenotype. In view of the selective and highly promising activity against breast cancer cell lines, these compounds can serve as promising leads for further development.

  16. Label-free fluorometric detection of chymotrypsin activity using graphene oxide/nucleic-acid-stabilized silver nanoclusters hybrid materials.

    PubMed

    Li, Shuangqin; Fu, Yuewei; Ma, Xuejuan; Zhang, Yaodong

    2017-02-15

    Pancreatic function tests are used to determine the presence of chronic pancreatitis, particularly in the early stage of the disease. Chymotrypsin is an indicator of pancreatic function and is thus related to pancreatic diseases. A new fluorescent biosensing method for assay of chymotrypsin activity was developed using DNA (dC12)-templated silver nanoclusters and graphene oxide (GO). A peptide probe was also designed using chymotrypsin-cleavable amino acid sequence and a cysteine terminus. The peptide probe formed Ag-S bond to dC12-AgNCs to enhance the fluorescence of dC12-AgNCs. After the addition of GO, the peptide was adsorbed to the negative GO surface and the fluorescence of dC12-AgNCs was quenched by FRET. The peptide was then degraded into amino acid fragments upon addition of chymotrypsin; these fragments were released from the GO surface, and the FRET was terminated. The developed label-free method features lower cost and higher sensitivity to chymotrypsin activity assay compared with conventional fluorescence analysis. The method can be used to analyze chymotrypsin (as low as 3ng/mL, signal/noise =3) across a dynamic range of 0.0-50.0ng/mL. The proposed biosensing strategy can also be extended to other proteases by using different peptide substrates.

  17. Graphene nanosheets-polypyrrole hybrid material as a highly active catalyst support for formic acid electro-oxidation.

    PubMed

    Yang, Sudong; Shen, Chengmin; Liang, Yanyu; Tong, Hao; He, Wei; Shi, Xuezhao; Zhang, Xiaogang; Gao, Hong-jun

    2011-08-01

    A novel electrode material based on graphene oxide (GO)-polypyrrole (PPy) composites was synthesized by in situ chemical oxidation polymerization. Palladium nanoparticles (NPs) with a diameter of 4.0 nm were loaded on the reduced graphene oxide(RGO)-PPy composites by a microwave-assisted polyol process. Microstructure analysis showed that a layer of coated PPy film with monodisperse Pd NPs is present on the RGO surface. The Pd/RGO-PPy catalysts exhibit excellent catalytic activity and stability for formic acid electro-oxidation when the weight feed ratio of GO to pyrrole monomer is 2:1. The superior performance of Pd/RGO-PPy catalysts may arise from utilization of heterogeneous nucleation sites for NPs and the greatly increased electronic conductivity of the supports. This journal is © The Royal Society of Chemistry 2011

  18. Graphene nanosheets-polypyrrole hybrid material as a highly active catalyst support for formic acid electro-oxidation

    NASA Astrophysics Data System (ADS)

    Yang, Sudong; Shen, Chengmin; Liang, Yanyu; Tong, Hao; He, Wei; Shi, Xuezhao; Zhang, Xiaogang; Gao, Hong-Jun

    2011-08-01

    A novel electrode material based on graphene oxide (GO)-polypyrrole (PPy) composites was synthesized by in situ chemical oxidation polymerization. Palladium nanoparticles (NPs) with a diameter of 4.0 nm were loaded on the reduced graphene oxide(RGO)-PPy composites by a microwave-assisted polyol process. Microstructure analysis showed that a layer of coated PPy film with monodisperse Pd NPs is present on the RGO surface. The Pd/RGO-PPy catalysts exhibit excellent catalytic activity and stability for formic acid electro-oxidation when the weight feed ratio of GO to pyrrole monomer is 2 : 1. The superior performance of Pd/RGO-PPy catalysts may arise from utilization of heterogeneous nucleation sites for NPs and the greatly increased electronic conductivity of the supports.

  19. Improved Work Function of Poly(3,4-ethylenedioxythiophene): Poly(styrenesulfonic acid) and its Effect on Hybrid Silicon/Organic Heterojunction Solar Cells.

    PubMed

    Shen, Xiaojuan; Chen, Ling; Pan, Jianmei; Hu, Yue; Li, Songjun; Zhao, Jie

    2016-12-01

    Hybrid silicon/organic solar cells have been recently extensively investigated due to their simple structure and low-cost fabrication process. However, the efficiency of the solar cells is greatly limited by the barrier height as well as the carrier recombination at the silicon/organic interface. In this work, hydrochloroplatinic acid (H2PtCl6) is employed into the poly(3,4-ethlenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) solution, and the work function (WF) of the PEDOT:PSS layer has been successfully improved. Based on the Pt-modified PEDOT:PSS layer, the efficiency of the silicon/PEDOT:PSS cell can be increased to 11.46%, corresponding to ~20% enhancement to the one without platinum (Pt) modification. Theoretical and experimental results show that, when increasing the WF of the PEDO:PSS layer, the barrier height between the silicon/PEDOT:PSS interface can be effectively enhanced. Meanwhile, the carrier recombination at the interface is significantly reduced. These results can contribute to better understanding of the interfacial mechanism of silicon/PEDOT:PSS interface, and further improving the device performance of silicon/organic solar cells.

  20. Improved Work Function of Poly(3,4-ethylenedioxythiophene): Poly(styrenesulfonic acid) and its Effect on Hybrid Silicon/Organic Heterojunction Solar Cells

    NASA Astrophysics Data System (ADS)

    Shen, Xiaojuan; Chen, Ling; Pan, Jianmei; Hu, Yue; Li, Songjun; Zhao, Jie

    2016-11-01

    Hybrid silicon/organic solar cells have been recently extensively investigated due to their simple structure and low-cost fabrication process. However, the efficiency of the solar cells is greatly limited by the barrier height as well as the carrier recombination at the silicon/organic interface. In this work, hydrochloroplatinic acid (H2PtCl6) is employed into the poly(3,4-ethlenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) solution, and the work function (WF) of the PEDOT:PSS layer has been successfully improved. Based on the Pt-modified PEDOT:PSS layer, the efficiency of the silicon/PEDOT:PSS cell can be increased to 11.46%, corresponding to 20% enhancement to the one without platinum (Pt) modification. Theoretical and experimental results show that, when increasing the WF of the PEDO:PSS layer, the barrier height between the silicon/PEDOT:PSS interface can be effectively enhanced. Meanwhile, the carrier recombination at the interface is significantly reduced. These results can contribute to better understanding of the interfacial mechanism of silicon/PEDOT:PSS interface, and further improving the device performance of silicon/organic solar cells.

  1. Zirconium-carbon hybrid sorbent for removal of fluoride from water: oxalic acid mediated Zr(IV) assembly and adsorption mechanism.

    PubMed

    Velazquez-Jimenez, Litza Halla; Hurt, Robert H; Matos, Juan; Rangel-Mendez, Jose Rene

    2014-01-21

    When activated carbon (AC) is modified with zirconium(IV) by impregnation or precipitation, the fluoride adsorption capacity is typically improved. There is significant potential to improve these hybrid sorbents by controlling the impregnation conditions, which determine the assembly and dispersion of the Zr phases on carbon surfaces. Here, commercial activated carbon was modified with Zr(IV) together with oxalic acid (OA) used to maximize the zirconium dispersion and enhance fluoride adsorption. Adsorption experiments were carried out at pH 7 and 25 °C with a fluoride concentration of 40 mg L(-1). The OA/Zr ratio was varied to determine the optimal conditions for subsequent fluoride adsorption. The data was analyzed using the Langmuir and Freundlich isotherm models. FTIR, XPS, and the surface charge distribution were performed to elucidate the adsorption mechanism. Potentiometric titrations showed that the modified activated carbon (ZrOx-AC) possesses positive charge at pH lower than 7, and FTIR analysis demonstrated that zirconium ions interact mainly with carboxylic groups on the activated carbon surfaces. Moreover, XPS analysis demonstrated that Zr(IV) interacts with oxalate ions, and the fluoride adsorption mechanism is likely to involve -OH(-) exchange from zirconyl oxalate complexes.

  2. Quantitative determination of isoquinoline alkaloids and chlorogenic acid in Berberis species using ultra high performance liquid chromatography with hybrid triple quadrupole linear ion trap mass spectrometry.

    PubMed

    Singh, Awantika; Bajpai, Vikas; Kumar, Sunil; Arya, Kamal Ram; Sharma, Kulwant Rai; Kumar, Brijesh

    2015-06-01

    Berberis species are well known and used extensively as medicinal plants in traditional medicine. They have many medicinal values attributable to the presence of alkaloids having different pharmacological activities. In this study, a method was developed and validated as per international conference on harmonization guidelines using ultra high performance liquid chromatography with hybrid triple quadrupole-linear ion trap mass spectrometry operated in the multiple reaction monitoring mode for nine bioactive compounds, including protoberberine alkaloids, aporphine alkaloids and chlorogenic acid. This method was applied in different plant parts of eight Berberis species to determine variations in content of nine bioactive compounds. The separation was achieved on an ACQUITY UPLC CSH™ C18 column using a gradient mobile phase at flow rate 0.3 mL/min. Calibration curves for all the nine analytes provided optimum linear detector response (with R(2) ≥0.9989) over the concentration range of 0.5-1000 ng/mL. The precision and accuracy were within RSDs ≤2.4 and ≤2.3%, respectively. The results indicated significant variation in the total contents of the nine compounds in Berberis species.

  3. Identification of genes involved in indole-3-butyric acid-induced adventitious root formation in nodal cuttings of Camellia sinensis (L.) by suppression subtractive hybridization.

    PubMed

    Wei, Kang; Wang, Liyuan; Cheng, Hao; Zhang, Chengcai; Ma, Chunlei; Zhang, Liqun; Gong, Wuyun; Wu, Liyun

    2013-02-10

    The plant hormone auxin plays a key role in adventitious rooting. To increase our understanding of genes involved in adventitious root formation, we identified transcripts differentially expressed in single nodal cuttings of Camellia sinensis treated with or without indole-3-butyric acid (IBA) by suppressive subtractive hybridization (SSH). A total of 77 differentially expressed transcripts, including 70 up-regulated and 7 down-regulated sequences, were identified in tea cuttings under IBA treatment. Seven candidate transcripts were selected and analyzed for their response to IBA, and IAA by real time RT-PCR. All these transcripts were up regulated by at least two folds one day after IBA treatment. Meanwhile, IAA showed less positive effects on the expression of candidate transcripts. The full-length cDNA of a F-box/kelch gene was also isolated and found to be similar to a group of At1g23390 like genes. These unigenes provided a new source for mining genes related to adventitious root formation, which facilitate our understanding of relative fundamental metabolism.

  4. pH-responsive drug delivery system based on luminescent CaF(2):Ce(3+)/Tb(3+)-poly(acrylic acid) hybrid microspheres.

    PubMed

    Dai, Yunlu; Zhang, Cuimiao; Cheng, Ziyong; Ma, Ping'an; Li, Chunxia; Kang, Xiaojiao; Yang, Dongmei; Lin, Jun

    2012-03-01

    In this study, we design a controlled release system based on CaF(2):Ce(3+)/Tb(3+)-poly(acrylic acid) (PAA) composite microspheres, which were fabricated by filling the pH-responsive PAA inside CaF(2):Ce(3+)/Tb(3+) hollow spheres via photopolymerization route. The CaF(2):Ce(3+)/Tb(3+) hollow spheres prepared by hydrothermal route possess mesoporous structure and show strong green fluorescence from Tb(3+) under UV excitation. Doxorubicin hydrochloride (DOX), a widely used anti-cancer drug, was used as a model drug to evaluate the loading and controlled release behaviors of the composite microspheres due to the good biocompatibility of the samples using MTT assay. The composite carriers provide a strongly pH-dependent drug release behavior owing to the intrinsic property of PAA and its interactions with DOX. The endocytosis process of drug-loaded microspheres was observed using confocal laser scanning microscopy (CLSM) and the in vitro cytotoxic effect against SKOV3 ovarian cancer cells of the DOX-loaded carriers was investigated. In addition, the extent of drug release could be monitored by the altering of photoluminescence (PL) intensity of CaF(2):Ce(3+)/Tb(3+). Considering the good biocompatibility, high drug loading content and pH-dependent drug release of the materials, these hybrid luminescent microspheres have potential applications in drug controlled release and disease therapy.

  5. Fabrication of microfibrous and nano-/microfibrous scaffolds: melt and hybrid electrospinning and surface modification of poly(L-lactic acid) with plasticizer.

    PubMed

    Yoon, Young Il; Park, Ko Eun; Lee, Seung Jin; Park, Won Ho

    2013-01-01

    Biodegradable poly(L-lactic acid) (PLA) fibrous scaffolds were prepared by electrospinning from a PLA melt containing poly(ethylene glycol) (PEG) as a plasticizer to obtain thinner fibers. The effects of PEG on the melt electrospinning of PLA were examined in terms of the melt viscosity and fiber diameter. Among the parameters, the content of PEG had a more significant effect on the average fiber diameter and its distribution than those of the spinning temperature. Furthermore, nano-/microfibrous silk fibroin (SF)/PLA and PLA/PLA composite scaffolds were fabricated by hybrid electrospinning, which involved a combination of solution electrospinning and melt electrospinning. The SF/PLA (20/80) scaffolds consisted of a randomly oriented structure of PLA microfibers (average fiber diameter = 8.9 µm) and SF nanofibers (average fiber diameter = 820 nm). The PLA nano-/microfiber (20/80) scaffolds were found to have similar pore parameters to the PLA microfiber scaffolds. The PLA scaffolds were treated with plasma in the presence of either oxygen or ammonia gas to modify the surface of the fibers. This approach of controlling the surface properties and diameter of fibers could be useful in the design and tailoring of novel scaffolds for tissue engineering.

  6. Double-labeled donor probe can enhance the signal of fluorescence resonance energy transfer (FRET) in detection of nucleic acid hybridization

    PubMed Central

    Okamura, Yukio; Kondo, Satoshi; Sase, Ichiro; Suga, Takayuki; Mise, Kazuyuki; Furusawa, Iwao; Kawakami, Shigeki; Watanabe, Yuichiro

    2000-01-01

    A set of fluorescently-labeled DNA probes that hybridize with the target RNA and produce fluorescence resonance energy transfer (FRET) signals can be utilized for the detection of specific RNA. We have developed probe sets to detect and discriminate single-strand RNA molecules of plant viral genome, and sought a method to improve the FRET signals to handle in vivo applications. Consequently, we found that a double-labeled donor probe labeled with Bodipy dye yielded a remarkable increase in fluorescence intensity compared to a single-labeled donor probe used in an ordinary FRET. This double-labeled donor system can be easily applied to improve various FRET probes since the dependence upon sequence and label position in enhancement is not as strict. Furthermore this method could be applied to other nucleic acid substances, such as oligo RNA and phosphorothioate oligonucleotides (S-oligos) to enhance FRET signal. Although the double-labeled donor probes labeled with a variety of fluorophores had unexpected properties (strange UV-visible absorption spectra, decrease of intensity and decay of donor fluorescence) compared with single-labeled ones, they had no relation to FRET enhancement. This signal amplification mechanism cannot be explained simply based on our current results and knowledge of FRET. Yet it is possible to utilize this double-labeled donor system in various applications of FRET as a simple signal-enhancement method. PMID:11121494

  7. Click chemistry-based synthesis and anticancer activity evaluation of novel C-14 1,2,3-triazole dehydroabietic acid hybrids.

    PubMed

    Hou, Wei; Luo, Zhi; Zhang, Guanjun; Cao, Danhui; Li, Di; Ruan, Haoqiang; Ruan, Benfang Helen; Su, Lin; Xu, Hongtao

    2017-09-29

    A concise and efficient synthetic approach has been established to readily access a series of novel C-14 1,2,3-triazole-tethered dehydroabietic acid derivatives in moderate to high yields. In vitro antiproliferative activity evaluation indicated that most of the hybrids exhibited potent inhibitory activities in a variety of cancer cell lines with low micromolar to submicromolar IC50 values. Further studies demonstrated that some of these analogues such as 20, 21, and 24 were also effective against adriamycin-resistant MCF-7 clone at low concentrations in a dose-dependent manner. Notably, the most potent compound 24, which possesses a 3-(tert-butoxycarbonylamino)phenyl-substituted triazole moiety, not only exhibited obviously improved IC50 values ranging from 0.7 to 1.2 μM against a panel of tested cancer cells, but also showed very weak cytotoxicity on normal cells. Preliminary mechanism studies indicated that compound 24 could induce apoptosis in MDA-MB-231 cells and was worth developing into a novel natural product-like anticancer lead by proper structure modification. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  8. Zirconium-carbon hybrid sorbent for removal of fluoride from water: oxalic acid mediated Zr(IV) assembly and adsorption mechanism

    PubMed Central

    Halla, Velazquez-Jimenez Litza; Hurt Robert, H; Juan, Matos; Rene, Rangel-Mendez Jose

    2014-01-01

    When activated carbon (AC) is modified with zirconium(IV) by impregnation or precipitation, the fluoride adsorption capacity is typically improved. There is significant potential to improve these hybrid sorbent by controlling the impregnation conditions, which determine the assembly and dispersion of the Zr phases on carbon surfaces. Here, commercial activated carbon was modified with Zr(IV) together with oxalic acid (OA) used to maximize the zirconium dispersion and enhance fluoride adsorption. Adsorption experiments were carried out at pH 7 and 25 °C with a fluoride concentration of 40 mg L−1. The OA/Zr ratio was varied to determine the optimal conditions for subsequent fluoride adsorption. The data was analyzed using the Langmuir and Freundlich isotherm models. FTIR, XPS and the surface charge distribution were performed to elucidate the adsorption mechanism. Potentiometric titrations showed that the modified activated carbon (ZrOx-AC) possesses positive charge at pH lower than 7, and FTIR analysis demonstrated that zirconium ions interact mainly with carboxylic groups on the activated carbon surfaces. Moreover, XPS analysis demonstrated that Zr(IV) interacts with oxalate ions, and the fluoride adsorption mechanism is likely to involve –OH− exchange from zirconyl oxalate complexes. PMID:24359079

  9. Hybrid calcium phosphate coatings with the addition of trace elements and polyaspartic acid by a low-thermal process.

    PubMed

    Xu, Sanzhong; Yang, Xianyan; Chen, Xiaoyi; Lin, Xiangjin; Zhang, Lei; Yang, Guojing; Gao, Changyou; Gou, Zhongru

    2011-06-01

    Research in the field of orthopedic implantology is currently focused on developing methodologies to potentiate osseointegration and to expedite the reestablishment of full functionality. We have developed a simple biomimetic approach for preparing trace elements-codoped calcium phosphate (teCaP) coatings on a titanium substrate. The reaction proceeded via low-thermal incubation in trace elements (TEs)-added simulated body fluid (teSBF) at 90 and 120 °C. The x-ray photoelectron spectroscopy, x-ray diffraction and energy-dispersive x-ray analyses demonstrated that the teCaP coating was the composite of hydroxyapatite and whitlockite, simultaneously doped with magnesium, strontium, zinc and silicon. The addition of polyaspartic acid and TEs into SBF significantly densified the coating. The incubation temperature is another important factor controlling the coating precipitation rate and bonding strength. An incubation temperature of 120 °C could accelerate the coating precipitation and improve the interface bonding strength. The in vitro cell culture investigation indicated that the teCaP coating supported the adhesion and spreading of ovariectomized rat mesenchymal stem cells (rMSCs) and particularly, promoted rMSCs proliferation compared to the CaP coating prepared in SBF. Collectively, from such a biomimetic route there potentially arises a general procedure to prepare a wide range of bioactive teCaP coatings of different composition for osteoporotic osteogenic cells activation response.

  10. Development of a peptide nucleic acid probe to Trichosporon species and identification of trichosporonosis by use of in situ hybridization in formalin-fixed and paraffin-embedded (FFPE) sections.

    PubMed

    Shinozaki, Minoru; Okubo, Yoichiro; Sasai, Daisuke; Nakayama, Haruo; Murayama, Somay Yamagata; Ide, Tadashi; Wakayama, Megumi; Ishiwatari, Takao; Tochigi, Naobumi; Nemoto, Tetsuo; Shibuya, Kazutoshi

    2013-01-01

    In order to identify Trichosporon species in formalin-fixed and paraffin-embedded sections from which visual discrimination of non-glabrata Candida species is mostly ineffective but critical for the choice of antifungals, we tested the usefulness of a newly designed peptide nucleic acid probe (PNA) for in situ hybridization (ISH). Results confirmed the usefulness of ISH with our PNA probe in identifying Trichosporon species from Candida albicans.

  11. Amino acids and peptides. XXX. Preparation of Arg-Gly-Asp (RGD) hybrids with poly(ethylene glycol) analogs and their antimetastatic effect.

    PubMed

    Maeda, M; Izuno, Y; Kawasaki, K; Kaneda, Y; Mu, Y; Tsutsumi, Y; Nakagawa, S; Mayumi, T

    1997-11-01

    Hybrids of a fibronectin-related peptide[Arg-Gly-Asp (RGD)] with poly(ethylene glycol) (PEG) analogs were prepared by a simple and easy procedure. Two amino-PEG analogs were used as carriers for hybrid formation of the RGD. One was poly(oxyethylene)dipropylamine and the other was Jeffamine ED type, which has branched chains. RGD peptides were formed stepwise on PEG analogs by the diisopropylcarbodiimide method. The synthetic intermediates were easily purified by molecular-sieve gel chromatography and the final products were purified by molecular-sieve gel chromatography, followed by HPLC. This simple and easy preparation procedure using molecular-sieve gel chromatography for purification of synthetic intermediates is advantageous for the preparation of peptide-polymer hybrids. We found that PEG is stable to HF treatment at 0 degree C for 1 h. The inhibitory effect of the RGD hybrids on experimental metastasis of B16-BL6 was examined in mice. The Jeffamine type hybrid showed no inhibitory effect at the dose of 1 mg/mouse, but poly(oxyethylene)dipropylamine type hybrid was inhibitory at the same dose. The effect of the latter hybrid was about the same as that of 1 mg of RGD. One mg of the hybrid contains 0.18 mumol of RGD and 1 mg of RGD is 2.38 mumol. Thus it can be said that the inhibitory effect of RGD was potentiated by hybrid formation with poly(oxyethylene)diisopropylamine.

  12. Triplex in-situ hybridization

    SciTech Connect

    Fresco, Jacques R.; Johnson, Marion D.

    2002-01-01

    Disclosed are methods for detecting in situ the presence of a target sequence in a substantially double-stranded nucleic acid segment, which comprises: a) contacting in situ under conditions suitable for hybridization a substantially double-stranded nucleic acid segment with a detectable third strand, said third strand being capable of hybridizing to at least a portion of the target sequence to form a triple-stranded structure, if said target sequence is present; and b) detecting whether hybridization between the third strand and the target sequence has occured.

  13. Nucleic acid hybridization in plasma: method for the quantitation of poly(I).poly(C12, U) in plasma of cancer patients.

    PubMed

    Krueger, L J; Andryuk, P J; Borigini, M J

    1986-12-01

    Using a new method for the direct measurement of the double-stranded RNA (dsRNA) molecule poly(I).poly(C12, U) in plasma, levels of 100 X 10(-9) g of drug were routinely quantified. The samples were digested by proteinase K in a buffered solution containing 0.1% of Brij-35 and deoxycholate detergents. The digestions were terminated after 1 h by the addition of Brij-58 and boiling saturated NaI (1.67 g/ml). Serially diluted samples were filtered onto nitrocellulose and the filters washed and hybridized. Levels of the hybridized-radioactive probe, synthesized de novo in an RNA dependent DNA transcription system, were determined by liquid scintillation spectrophotometry and quantified by comparison to a standard curve. The efficiency of hybridization declined when the plasma concentration in the reaction fell below 1.0 mg/ml. Incubation and denaturation temperatures significantly altered the amount of radioactive probe hybridized; results varied in the extent of hybridization and in the concentration range of dsRNA showing a linear response. Elevated temperature during proteinase K digestion showed reduced hybridization efficiencies: 100% at 25, 80% at 37, 35% at 45, and 25% at 55 degrees C. Incubation at elevated temperatures, prior to the addition of NaI, caused a decline in the amount of radioactivity hybridized, but did not have an effect during hybridization.

  14. Bifunctional sensor of pentachlorophenol and copper ions based on nanostructured hybrid films of humic acid and exfoliated layered double hydroxide via a facile layer-by-layer assembly.

    PubMed

    Yuan, Shuang; Peng, Dinghua; Hu, Xianluo; Gong, Jingming

    2013-06-27

    A new, highly sensitive bifunctional electrochemical sensor for the simultaneous determination of pentachlorophenol (PCP) and copper ions (Cu(2+)) has been developed, where organic-inorganic hybrid ultrathin films were fabricated by alternate assembly of humic acid (HA) and exfoliated Mg-Al-layered double hydroxide (LDH) nanosheets onto ITO substrates via a layer-by-layer (LBL) approach. The multilayer films were then characterized by means of UV-vis spectrometry, scanning electron microscopy (SEM), and atomic force microscope (AFM). These films were found to have a relatively smooth surface with almost equal amounts of HA incorporated in each cycle. Its electrochemical performance was systematically investigated. Our results demonstrate that such a newly designed (LDH/HA)n multilayer films, combining the individual properties of HA (dual recognition ability for organic herbicides and metal ions) together with LDH nanosheets (a rigid inorganic matrix), can be applied to the simultaneous analysis of PCP and Cu(II) without interference from each other. The LBL assembled nanoarchitectures were further investigated by X-ray photoelectron spectroscopy (XPS) and infrared spectroscopy (IR), which provides insight for bifunctional sensing behavior. Under the optimized conditions, the detection limit was found to be as low as 0.4 nM PCP, well below the guideline value of PCP in drinking water (3.7 nM) set by the United States Environmental Protection Agency (U.S. EPA), and 2.0 nM Cu(2+), much below the guideline value (2.0 mg L(-1), ~31.2 nM) from the World Health Organization (WHO), respectively. Toward the goal for practical applications, this simple and cost-effective probe was further evaluated by monitoring PCP and Cu(II) in water samples. Copyright © 2013 Elsevier B.V. All rights reserved.

  15. Suppression Subtractive Hybridization Analysis of Genes Regulated by Application of Exogenous Abscisic Acid in Pepper Plant (Capsicum annuum L.) Leaves under Chilling Stress

    PubMed Central

    Gong, Zhen-Hui; Yin, Yan-Xu; Li, Da-Wei

    2013-01-01

    Low temperature is one of the major factors limiting pepper (Capsicum annuum L.) production during winter and early spring in non-tropical regions. Application of exogenous abscisic acid (ABA) effectively alleviates the symptoms of chilling injury, such as wilting and formation of necrotic lesions on pepper leaves; however, the underlying molecular mechanism is not understood. The aim of this study was to identify genes that are differentially up- or downregulated in ABA-pretreated hot pepper seedlings incubated at 6°C for 48 h, using a suppression subtractive hybridization (SSH) method. A total of 235 high-quality ESTs were isolated, clustered and assembled into a collection of 73 unigenes including 18 contigs and 55 singletons. A total of 37 unigenes (50.68%) showed similarities to genes with known functions in the non-redundant database; the other 36 unigenes (49.32%) showed low similarities or unknown functions. Gene ontology analysis revealed that the 37 unigenes could be classified into nine functional categories. The expression profiles of 18 selected genes were analyzed using quantitative RT-PCR; the expression levels of 10 of these genes were at least two-fold higher in the ABA-pretreated seedlings under chilling stress than water-pretreated (control) plants under chilling stress. In contrast, the other eight genes were downregulated in ABA-pretreated seedlings under chilling stress, with expression levels that were one-third or less of the levels observed in control seedlings under chilling stress. These results suggest that ABA can positively and negatively regulate genes in pepper plants under chilling stress. PMID:23825555

  16. Comparison of MALDI-TOF MS, nucleic acid hybridization and the MPT64 immunochromatographic test for the identification of M. tuberculosis and non-tuberculosis Mycobacterium species.

    PubMed

    Şamlı, Asuman; İlki, Arzu

    2016-10-01

    Mycobacteria are an important cause of morbidity in humans. Rapid and accurate mycobacterial identification is important for improving patient outcomes. However, identification of Mycobacterium species is not easy, due to the slow and fastidious growth of mycobacteria. Recently, biochemical, sequencing, and probing methods have come to be used for identification. This study compared the performance of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the identification of M.tuberculosis and non-tuberculosis Mycobacteria (NTM) to those of nucleic acid hybridization (NAH) and the MPT64 immunochromatographic test. A total of 69 isolates from Marmara University Hospital, Microbiology Laboratory obtained between 2012 and 2013 were included in our study. All strains were grown on Lowenstein-Jensen and Middlebrook 7H9 medium. Among the 69 isolates, 56 (81%) were isolated as Mycobacterium tuberculosis complex (MTC), and 13 (19%) were isolated as NTM by the MPT64 ICT. NAH was able to identify all isolates to the species level. The isolated NTM included M. intracellulare (n:5), M. lentiflavum (n:3), M. xenopi (n:2), M. malmoense (n:1), M. abscessus (n:1), and M. avium (n:1). MALDI-TOF MS identified 88% of the mycobacterial isolates. All M. tuberculosis strains were identified correctly, but the ratio was 38.5% for NTM. Mycobacterial identification using MALDI-TOF MS takes 45 minutes and costs 3 Euro/test, whereas mycobacterial identification using NAH takes 6-7 hours and costs 30 Euro/test. In conclusion, MALDI-TOF MS has the potential to identify mycobacteria in the clinical laboratory setting by reducing identification turnaround time and laboratory costs for isolate referral.

  17. Suppression Subtractive Hybridization Analysis of Genes Regulated by Application of Exogenous Abscisic Acid in Pepper Plant (Capsicum annuum L.) Leaves under Chilling Stress.

    PubMed

    Guo, Wei-Li; Chen, Ru-Gang; Gong, Zhen-Hui; Yin, Yan-Xu; Li, Da-Wei

    2013-01-01

    Low temperature is one of the major factors limiting pepper (Capsicum annuum L.) production during winter and early spring in non-tropical regions. Application of exogenous abscisic acid (ABA) effectively alleviates the symptoms of chilling injury, such as wilting and formation of necrotic lesions on pepper leaves; however, the underlying molecular mechanism is not understood. The aim of this study was to identify genes that are differentially up- or downregulated in ABA-pretreated hot pepper seedlings incubated at 6°C for 48 h, using a suppression subtractive hybridization (SSH) method. A total of 235 high-quality ESTs were isolated, clustered and assembled into a collection of 73 unigenes including 18 contigs and 55 singletons. A total of 37 unigenes (50.68%) showed similarities to genes with known functions in the non-redundant database; the other 36 unigenes (49.32%) showed low similarities or unknown functions. Gene ontology analysis revealed that the 37 unigenes could be classified into nine functional categories. The expression profiles of 18 selected genes were analyzed using quantitative RT-PCR; the expression levels of 10 of these genes were at least two-fold higher in the ABA-pretreated seedlings under chilling stress than water-pretreated (control) plants under chilling stress. In contrast, the other eight genes were downregulated in ABA-pretreated seedlings under chilling stress, with expression levels that were one-third or less of the levels observed in control seedlings under chilling stress. These results suggest that ABA can positively and negatively regulate genes in pepper plants under chilling stress.

  18. Quantitative Use of Fluorescent In Situ Hybridization To Examine Relationships between Mycolic Acid-Containing Actinomycetes and Foaming in Activated Sludge Plants

    PubMed Central

    Davenport, Russell J.; Curtis, Thomas P.; Goodfellow, Michael; Stainsby, Fiona M.; Bingley, Marc

    2000-01-01

    The formation of viscous foams on aeration basins and secondary clarifiers of activated sludge plants is a common and widespread problem. Foam formation is often attributed to the presence of mycolic acid-containing actinomycetes (mycolata). In order to examine the relationship between the number of mycolata and foam, we developed a group-specific probe targeting the 16S rRNA of the mycolata, a protocol to permeabilize mycolata, and a statistically robust quantification method. Statistical analyses showed that a lipase-based permeabilization method was quantitatively superior to previously described methods (P << 0.05). When mixed liquor and foam samples were examined, most of the mycolata present were rods or cocci, although filamentous mycolata were also observed. A nested analysis of variance showed that virtually all of the measured variance occurred between fields of view and not between samples. On this basis we determined that as few as five fields of view could be used to give a statistically meaningful sample. Quantitative fluorescent in situ hybridization (FISH) was used to examine the relationship between foaming and the concentration of mycolata in a 20-m3 completely mixed activated sludge plant. Foaming occurred when the number of mycolata exceeded a certain threshold value. Baffling of the plant affected foaming without affecting the number of mycolata. We tentatively estimated that the threshold foaming concentration of mycolata was about 2 × 106 cells ml−1 or 4 × 1012 cells m−2. We concluded that quantitative use of FISH is feasible and that quantification is a prerequisite for rational investigation of foaming in activated sludge. PMID:10698786

  19. Quantitative use of fluorescent in situ hybridization to examine relationships between mycolic acid-containing actinomycetes and foaming in activated sludge plants.

    PubMed

    Davenport, R J; Curtis, T P; Goodfellow, M; Stainsby, F M; Bingley, M

    2000-03-01

    The formation of viscous foams on aeration basins and secondary clarifiers of activated sludge plants is a common and widespread problem. Foam formation is often attributed to the presence of mycolic acid-containing actinomycetes (mycolata). In order to examine the relationship between the number of mycolata and foam, we developed a group-specific probe targeting the 16S rRNA of the mycolata, a protocol to permeabilize mycolata, and a statistically robust quantification method. Statistical analyses showed that a lipase-based permeabilization method was quantitatively superior to previously described methods (P < 0.05). When mixed liquor and foam samples were examined, most of the mycolata present were rods or cocci, although filamentous mycolata were also observed. A nested analysis of variance showed that virtually all of the measured variance occurred between fields of view and not between samples. On this basis we determined that as few as five fields of view could be used to give a statistically meaningful sample. Quantitative fluorescent in situ hybridization (FISH) was used to examine the relationship between foaming and the concentration of mycolata in a 20-m(3) completely mixed activated sludge plant. Foaming occurred when the number of mycolata exceeded a certain threshold value. Baffling of the plant affected foaming without affecting the number of mycolata. We tentatively estimated that the threshold foaming concentration of mycolata was about 2 x 10(6) cells ml(-1) or 4 x 10(12) cells m(-2). We concluded that quantitative use of FISH is feasible and that quantification is a prerequisite for rational investigation of foaming in activated sludge.

  20. Improved enumeration of lactic acid bacteria in mesophilic dairy starter cultures by using multiplex quantitative real-time PCR and flow cytometry-fluorescence in situ hybridization.

    PubMed

    Friedrich, Udo; Lenke, Jan

    2006-06-01

    Nucleic acid-based assays were developed to enumerate members of the three taxa Lactococcus lactis subsp. cremoris, L. lactis subsp. lactis, and Leuconostoc spp. in mesophilic starter cultures. To our knowledge the present is the first study to present a multiplex quantitative PCR (qPCR) strategy for the relative enumeration of bacteria. The multiplex qPCR strategy was designed to quantify the target DNA simultaneously relative to total bacterial DNA. The assay has a high discriminatory power and resolves concentration changes as low as 1.3-fold. The methodology was compared with flow cytometric fluorescence in situ hybridization (FLOW-FISH) and 5-bromo-4-chloro-3-indolyl-beta-d-galactopyranoside (X-Gal)-calcium citrate agar-based plate counting. For enumeration by FLOW-FISH, three new probes having the same specificity as the qPCR assay were designed and established. A combination with flow cytometry greatly reduced the time consumed compared to manual enumeration. Both qPCR and FLOW-FISH yielded similar community compositions for 10 complex starter cultures, with all detected subpopulations being highly significantly correlated (P < 0.001). Correlations between X-Gal-calcium citrate agar-based CFU and qPCR-derived counts were highly significant (P < 0.01 and P < 0.001, respectively) for the number of acidifiers versus L. lactis subsp. cremoris and for Leuconostoc spp. as quantified by the two techniques, respectively. This confirmed that most acidifiers in the studied PROBAT cultures are members of L. lactis subsp. cremoris. Quantitative real-time PCR and FLOW-FISH were found to be effective and accurate tools for the bacterial community analysis of complex starter cultures.

  1. Interobserver agreement in the reporting of cervical biopsy specimens obtained from women screened by visual inspection with acetic acid and hybrid capture 2.

    PubMed

    Basu, Partha; Kamal, Meherbano; Ray, Chinmayi; Bhat, Dharitri; Ghosh, Ishita; Mittal, Srabani; Chatterjee, Simi; Samaddar, Anushree; Biswas, Jaydip

    2013-09-01

    Visual inspection with acetic acid (VIA) and human papillomavirus detection have sensitivity higher than cytology but lower specificity. The high false-positive rate of either test poses a challenge to the colposcopists who obtain biopsies from the innocuous changes and to the pathologists who have to interpret large numbers of specimens that are either normal or have low-grade abnormalities. Interobserver variation in histopathologic interpretations of cervical punch biopsy specimens is high, specially for the lower-grade abnormalities. Use of the modified Bethesda system to report histology in place of the cervical intraepithelial neoplasia (CIN) system has the potential to reduce such variability as there are fewer categories. The present study aimed to assess the interobserver agreement to interpret cervical punch biopsies when both pathologists followed the modified Bethesda classification system and also when one pathologist followed the modified Bethesda classification system and the other followed the CIN classification system. Colposcopy-directed punch biopsies were obtained from VIA and/or Hybrid Capture 2-positive women. The Institute pathologist interpreted the slides using the CIN system. Blinded review was done by 2 external pathologists who independently interpreted cervical punch biopsies using the Bethesda system. The Institute pathologist's diagnoses based on CIN system were converted post hoc into categories belonging to the Bethesda system for comparison. The overall agreement was poor (κ=0.36). The lowest agreement was observed in the low-grade squamous intraepithelial lesion category (κ=0.23) and the highest in the squamous cell carcinoma category (κ=0.76). The agreement between the reviewers, both of whom used the Bethesda system, was substantial.

  2. AIDS-related lymphoma. Histopathology, immunophenotype, and association with Epstein-Barr virus as demonstrated by in situ nucleic acid hybridization.

    PubMed Central

    Hamilton-Dutoit, S. J.; Pallesen, G.; Franzmann, M. B.; Karkov, J.; Black, F.; Skinhøj, P.; Pedersen, C.

    1991-01-01

    To investigate the range of pathology shown by acquired immune deficiency syndrome (AIDS)-related lymphomas arising in an epidemiologically well-defined group of patients, all cases of lymphoma recognized in Danish human immunodeficiency virus (HIV)-infected individuals up to the end of 1988 were studied. Twenty-seven cases (26 high-grade non-Hodgkin's lymphoma [NHL], 1 Hodgkin's disease) were found, to give a cumulative incidence rate of 8% among Danish AIDS patients. Morphologically most NHL patients were classified into two groups: 1) high-grade tumors with a predominant population of immunoblasts, either monomorphic or more often polymorphic with plasmacytic differentiation; 2) Burkitt-type. Of 26 NHLs, 22 had a B-cell paraffin-section immunophenotype and 4 were non-B, non-T. Epstein-Barr virus (EBV) DNA was demonstrated in tumor cells of 12 of 24 cases (50%) using in situ nucleic acid hybridization with a 35S-labeled probe in paraffin sections. Epstein-Barr virus DNA was found in 65% of group 1 and 20% of group 2 tumors. This study suggests the existence of two main groups of AIDS-related lymphoma with different pathogeneses. First there are immunoblast-rich lesions, which usually are associated with EBV and morphologically resemble lymphomas described in immunosuppressed organ-transplantation patients. Second there are Burkitt-type tumors in which EBV sequences are less common and that may be pathogenetically analogous to sporadic Burkitt's lymphoma. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:1846263

  3. GPCR-2L: predicting G protein-coupled receptors and their types by hybridizing two different modes of pseudo amino acid compositions.

    PubMed

    Xiao, Xuan; Wang, Pu; Chou, Kuo-Chen

    2011-03-01

    G protein-coupled receptors (GPCRs) are among the most frequent targets of therapeutic drugs. With the avalanche of newly generated protein sequences in the post genomic age, to expedite the process of drug discovery, it is highly desirable to develop an automated method to rapidly identify GPCRs and their types. A new predictor was developed by hybridizing two different modes of pseudo-amino acid composition (PseAAC): the functional domain PseAAC and the low-frequency Fourier spectrum PseAAC. The new predictor is called GPCR-2L, where "2L" means that it is a two-layer predictor: the 1st layer prediction engine is to identify a query protein as GPCR or not; if it is, the prediction will be automatically continued to further identify it as belonging to one of the following six types: (1) rhodopsin-like (Class A), (2) secretin-like (Class B), (3) metabotropic glutamate/pheromone (Class C), (4) fungal pheromone (Class D), (5) cAMP receptor (Class E), or (6) frizzled/smoothened family (Class F). The overall success rate of GPCR-2L in identifying proteins as GPCRs or non-GPCRs is over 97.2%, while identifying GPCRs among their six types is over 97.8%. Such high success rates were derived by the rigorous jackknife cross-validation on a stringent benchmark dataset, in which none of the included proteins had ≥40% pairwise sequence identity to any other protein in a same subset. As a user-friendly web-server, GPCR-2L is freely accessible to the public at http://icpr.jci.edu.cn/, by which one can obtain the 2-level results in about 20 s for a query protein sequence of 500 amino acids. The longer the sequence is, the more time it may usually need. The high success rates reported here indicate that it is a quite effective approach to identify GPCRs and their types with the functional domain information and the low-frequency Fourier spectrum analysis. It is anticipated that GPCR-2L may become a useful tool for both basic research and drug development in the areas related to

  4. Systems for hybrid cars

    NASA Astrophysics Data System (ADS)

    Bitsche, Otmar; Gutmann, Guenter

    Not only sharp competition but also legislation are pushing development of hybrid drive trains. Based on conventional internal combustion engine (ICE) vehicles, these drive trains offer a wide range of benefits from reduced fuel consumption and emission to multifaceted performance improvements. Hybrid electric drive trains may also facilitate the introduction of fuel cells (FC). The battery is the key component for all hybrid drive trains, as it dominates cost and performance issues. The selection of the right battery technology for the specific automotive application is an important task with an impact on costs of development and use. Safety, power, and high cycle life are a must for all hybrid applications. The greatest pressure to reduce cost is in soft hybrids, where lead-acid embedded in a considerate management presents the cheapest solution, with a considerable improvement in performance needed. From mild to full hybridization, an improvement in specific power makes higher costs more acceptable, provided that the battery's service life is equivalent to the vehicle's lifetime. Today, this is proven for the nickel-metal hydride system. Lithium ion batteries, which make use of a multiple safety concept, and with some development anticipated, provide even better prospects in terms of performance and costs. Also, their scalability permits their application in battery electric vehicles—the basis for better performance and enhanced user acceptance. Development targets for the batteries are discussed with a focus on system aspects such as electrical and thermal management and safety.

  5. Signal enhancement for gene detection based on a redox reaction of [Fe(CN)(6)](4-) mediated by ferrocene at the terminal of a peptide nucleic acid as a probe with hybridization-amenable conformational flexibility.

    PubMed

    Aoki, Hiroshi; Tao, Hiroaki

    2008-07-01

    Electrochemically enhanced DNA detection was demonstrated by utilizing the couple of a synthesized ferrocene-terminated peptide nucleic acid (PNA) with a cysteine anchor and a sacrificial electron donor [Fe(CN)(6)](4-). DNA detection sensors were prepared by modifying a gold electrode surface with a mixed monolayer of the probe PNA and 11-hydroxy-1-undecanethiol (11-HUT), protecting [Fe(CN)(6)](4-) from any unexpected redox reaction. Before hybridization, the terminal ferrocene moiety of the probe was subject to a redox reaction due to the flexible probe structure and, in the presence of [Fe(CN)(6)](4-), the observed current was amplified based on regeneration of the ferrocene moiety. Hybridization decreased the redox current of the ferrocene. This occurred because hybridization rigidified the probe structure: the ferrocene moiety was then removed from the electrode surface, and the redox reaction of [Fe(CN)(6)](4-) was again prevented. The change in the anodic current before and after hybridization was enhanced 1.75-fold by using the electron donor [Fe(CN)(6)](4-). Sequence-specific detection of the complementary target DNA was also demonstrated.

  6. Locked Nucleic Acid Flow Cytometry-fluorescence in situ Hybridization (LNA flow-FISH): A Method for Bacterial Small RNA Detection

    DTIC Science & Technology

    2012-01-10

    LNA oligonucleotide. The biotinylation is necessary for post-hybridization staining with a streptavidin-dye conjugate. It is possible to add this...which the LNA probe is hybridized to the target sRNA but the hybridization event is not detected due to the absence of the fluorescent stain , and (iii) a... peptidoglycan present in bacterial cell walls. 2. Prepare a 3.0 μg/mL solution of proteinase K in TE buffer. This solution should be prepared fresh from

  7. Hybrid Vehicles

    DTIC Science & Technology

    2008-12-08

    hybrid electric vehicles typically contain potentially hazardous levels of electrical voltage or current. It is important to protect the operators...60740. ITOP 2-2-607(1)41 is used for tracked vehicles. 13 TOP 2-1-003 08 December 2008 Hybrid electric vehicles often employ much more

  8. Hexagonal cobalt oxyhydroxide-carbon dots hybridized surface: high sensitive fluorescence turn-on probe for monitoring of ascorbic acid in rat brain following brain ischemia.

    PubMed

    Li, Linbo; Wang, Chao; Liu, Kangyu; Wang, Yuhan; Liu, Kun; Lin, Yuqing

    2015-03-17

    In this study, we report a novel and efficient fluorescence probe synthesized by Tris(hydroxymethyl)aminomethane-derived carbon dots (CDs)-modified hexagonal cobalt oxyhydroxide(CoOOH) nanoflakes (Tris-derived CDs-CoOOH) for monitoring of cerebral ascorbic acid (AA) in brain microdialysate. The as-prepared Tris-derived CDs with the fluorescence quantum yield of 7.3% are prepared by a one-step pyrolysis strategy of the sole precursor and used as the signal output. After being hybridized with CoOOH nanoflakes to form Tris-derived CDs-CoOOH, the luminescence of the Tris-derived CDs can be efficiently quenched by CoOOH via fluorescence resonance energy transfer (FRET). Due to the specific redox reaction between the enediol group of AA and hexagonal CoOOH nanoflakes, AA can reduce the hexagonal CoOOH nanoflakes in the Tris-derived CDs-CoOOH and lead to collapse of the hybrized structure, then the release of Tris-derived CDs, and thus finally the fluorescence recovery. Moreover, cobalt ions (II), generated by CoOOH nanoflakes oxidizing AA, almost have no obvious interference on the fluorescence probe, i.e., Tris-derived CDs, which could be ascribed to the surface of Tris-derived CDs containing a few strong chelation groups such as amino/carboxyl/thiol groups, instead of plenty of -OH groups with weak chelation with Co(2+). On the basis of this feature, the Tris-derived CDs-CoOOH fluorescent probe demonstrates a linear range from 100 nM to 20 μM with the detection limit of ∼50 nM, i.e., with an improved sensitivity toward AA detection. Compared with other turn-on fluorescent methods using convenient fluorophore-nitroxide fluorescent probes for detection of AA, the method demonstrated here possesses a facial synthesis route, lower limit of detection, and wider linear range, which validates sensing of AA in the cerebral systems during the calm/ischemia process. This study provides a fluorescence assay for the simple yet facial detection of AA in the cerebral systems and

  9. Strategies for optimizing DNA hybridization on surfaces.

    PubMed

    Ravan, Hadi; Kashanian, Soheila; Sanadgol, Nima; Badoei-Dalfard, Arastoo; Karami, Zahra

    2014-01-01

    Specific and predictable hybridization of the polynucleotide sequences to their complementary counterparts plays a fundamental role in the rational design of new nucleic acid nanodevices. Generally, nucleic acid hybridization can be performed using two major strategies, namely hybridization of DNA or RNA targets to surface-tethered oligonucleotide probes (solid-phase hybridization) and hybridization of the target nucleic acids to randomly distributed probes in solution (solution-phase hybridization). Investigations into thermodynamic and kinetic parameters of these two strategies showed that hybridization on surfaces is less favorable than that of the same sequence in solution. Indeed, the efficiency of DNA hybridization on surfaces suffers from three constraints: (1) electrostatic repulsion between DNA strands on the surface, (2) steric hindrance between tethered DNA probes, and (3) nonspecific adsorption of the attached oligonucleotides to the solid surface. During recent years, several strategies have been developed to overcome the problems associated with DNA hybridization on surfaces. Optimizing the probe surface density, application of a linker between the solid surface and the DNA-recognizing sequence, optimizing the pH of DNA hybridization solutions, application of thiol reagents, and incorporation of a polyadenine block into the terminal end of the recognizing sequence are among the most important strategies for enhancing DNA hybridization on surfaces.

  10. Anatomic viral detection is automated: the application of a robotic molecular pathology system for the detection of DNA viruses in anatomic pathology substrates, using immunocytochemical and nucleic acid hybridization techniques.

    PubMed Central

    Montone, K. T.; Brigati, D. J.; Budgeon, L. R.

    1989-01-01

    This paper presents the first automated system for simultaneously detecting human papilloma, herpes simplex, adenovirus, or cytomegalovirus viral antigens and gene sequences in standard formalin-fixed, paraffin-embedded tissue substrates and tissue culture. These viruses can be detected by colorimetric in situ nucleic acid hybridization, using biotinylated DNA probes, or by indirect immunoperoxidase techniques, using polyclonal or monoclonal antibodies, in a 2.0-hour assay performed at a single automated robotic workstation. Images FIG. 1 FIG. 4 FIG. 5 FIG. 6 FIG. 7 FIG. 8 FIG. 9 FIG. 10 FIG. 11 PMID:2773514

  11. A nucleic acid probe labeled with desmethyl thiazole orange: a new type of hybridization-sensitive fluorescent oligonucleotide for live-cell RNA imaging.

    PubMed

    Okamoto, Akimitsu; Sugizaki, Kaori; Yuki, Mizue; Yanagisawa, Hiroyuki; Ikeda, Shuji; Sueoka, Takuma; Hayashi, Gosuke; Wang, Dan Ohtan

    2013-01-14

    A new fluorescent nucleotide with desmethyl thiazole orange dyes, D'(505), has been developed for expansion of the function of fluorescent probes for live-cell RNA imaging. The nucleoside unit of D'(505) for DNA autosynthesis was soluble in organic solvents, which made the preparation of nucleoside units and the reactions in the cycles of DNA synthesis more efficient. The dyes of D'(505)-containing oligodeoxynucleotide were protonated below pH 7 and the oligodeoxynucleotide exhibited hybridization-sensitive fluorescence emission through the control of excitonic interactions of the dyes of D'(505). The simplified procedure and effective hybridization-sensitive fluorescence emission produced multicolored hybridization-sensitive fluorescent probes, which were useful for live-cell RNA imaging. The acceptor-bleaching method gave us information on RNA in a specific cell among many living cells.

  12. DNA/DNA in situ hybridization with enzyme linked probes

    SciTech Connect

    Grillo, S.; Mosher, M.; Charles, P.; Henry, S.; Taub, F.

    1987-05-01

    A non-radioactive in situ nucleic acid hybridization method which requires no antibodies, haptens, avidin or biotin intermediateries is presented. Horseradish peroxidase (HRP) labeled nucleic acid probes are hybridized in situ for 2 hours or less, followed by brief washing of hybridized cells and the direct detection of in situ hybrids with diaminobenzidine (DAB). Application of this method to the detection of Human Papilloma Virus (HPV) in human cells is shown.

  13. A hybrid fluorous monolithic capillary column with integrated nanoelectrospray ionization emitter for determination of perfluoroalkyl acids by nano-liquid chromatography-nanoelectrospray ionization-mass spectrometry/mass spectrometry.

    PubMed

    Zhang, Haiyang; Ou, Junjie; Wei, Yinmao; Wang, Hongwei; Liu, Zhongshan; Zou, Hanfa

    2016-04-01

    A hybrid fluorous monolithic column was simply prepared via photo-initiated free radical polymerization of an acrylopropyl polyhedral oligomeric silsesquioxane (acryl-POSS) and a perfluorous monomer (2,2,3,3,4,4,5,5,6,6,7,7-dodecafluoroheptyl acrylate) in UV-transparent fused-silica capillaries within 5min. The physical characterization of hybrid fluorous monolith, including scanning electron microscopy (SEM), Fourier transform infrared (FT-IR) spectroscopy, mercury intrusion porosimetry (MIP) and nitrogen adsorption/desorption measurement was performed. Chromatographic performance was also evaluated by capillary liquid chromatography (cLC). Due to the fluorous-fluorous interaction between fluorous monolith and analytes, fluorobenzenes could well be separated, and the column efficiencies reached 86,600-92,500plates/m at the velocity of 0.87mm/s for alkylbenzenes and 51,900-76,000plates/m at the velocity of 1.10mm/s for fluorobenzenes. Meanwhile, an approach to integrate nanoelectrospray ionization (ESI) emitter with hybrid fluorous monolithic column was developed for quantitative determination of perfluoroalkyl acids by nanoHPLC-ESI-MS/MS. The integration design could minimize extracolumn volume, thus excluding undesirable peak broadening and improving separation performance. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Stellarator hybrids

    SciTech Connect

    Furth, H.P.; Ludescher, C.

    1984-08-01

    The present paper briefly reviews the subject of tokamak-stellarator and pinch-stellarator hybrids, and points to two interesting new possibilities: compact-torus-stellarators and mirror-stellarators.

  15. Effect of winter feeding frequency on growth, survival, and fatty acid metabolism of juvenile bluegill (Lepomis macrochirus) and hybrid bluegill (L.cyanellus ×L. macrochirus)

    USDA-ARS?s Scientific Manuscript database

    There has been an increasing global demand for large bluegill (Lepomis macrochirus) and its hybrid (female Lepomis cyanellus x male L. macrochirus) to supply the food-fish market. However, production of market-size bluegill requires producers to overwinter them in temperate regions. Winter fish mort...

  16. [Evaluation of peptide nucleic acid fluorescent in situ hybridization (PNA FISH) method in the identifi cation of Candida species isolated from blood cultures].

    PubMed

    Aydemir, Gonca; Koç, Ayşe Nedret; Atalay, Mustafa Altay

    2016-04-01

    In recent years, increased number of patients who are hospitalized in intensive care units, received immunosuppressive therapy and treated with broad-spectrum antibiotics that can lead an increase in the incidence of systemic candidiasis. In these patients, the most common clinical manifestation is candidemia. Since the identification of Candida species isolated from blood cultures is time consuming by conventional (morphological and biochemical) methods, rapid, reliable and accurate methods are needed. For this purpose novel systems have been developed to identify the agent directly. The aim of this study was to evaluate the peptide nucleic acid fluorescent in situ hybridization (PNA FISH) method for the identification of Candida species by comparing with the conventional methods. A total of 50 patients who were admitted to Erciyes University Medical Faculty Hospital clinics and followed with prediagnosis of systemic fungal infections whose blood cultures were positive for the yeasts between July 2011 and July 2012 were included in the study. The conventional identification of Candida isolates was performed by considering macroscopic and microscopic morphology, germ tube test, cycloheximide sensitivity, urease activity and carbohydrate assimilation patterns with API 20C AUX (bioMerieux, France) test. PNA FISH method was conducted by the use of a commercial kit namely Yeast Traffic Light(®) PNA FISH (AdvanDx, USA). According to morphological and biochemical characteristics (conventional methods), 19 (38%) out of 50 Candida isolates were identified as C.albicans, 12 (24%) as C.glabrata, five (10%) as C.parapsilosis, five (10%) as C.kefyr, four (8%) as C.krusei, two (4%) as C.guilliermondii, two (4%) as C.tropicalis and one (2%) as C.lusitaniae. On the other hand, 24 (48%) of the isolates were identified as C.albicans/C.parapsilosis (with green fluorescence), 16 (32%) as C.glabrata/C.krusei (with red fluorescence) and one (%2) as C.tropicalis (with yellow

  17. Mass spectral study of hybrid peptides derived from (R)-aminoxy ester and [beta]-amino acids: The influence of aminoxy peptide bond (CO-NH-O) on peptide fragmentation under electrospray ionization conditions

    NASA Astrophysics Data System (ADS)

    Ramesh, V.; Ramesh, M.; Srinivas, R.; Sharma, G. V. M.; Manohar, V.

    2009-04-01

    A new class of Boc-protected aminoxy hybrid peptides containing repeats of [beta]-hAla-(R)-Ama-, and [beta]-Caa-(R)-Ama- ([beta]-hAla = [beta]3-(S)-hAlanine, (R)-Ama = (R)-aminoxy ester, and [beta]-Caa = (R)-C-linked carbo-[beta]3-amino acid) have been studied by electrospray ionization (ESI) ion-trap and quadrupole time-of-flight tandem mass spectrometry (Q-TOF MS/MS) of their protonated, cationized, and negative ions. MS3 CID of protonated aminoxy peptides of [beta]-hAla-(R)-Ama- yield intense [beta]-amino acid characteristic retro-Mannich fragmentation. The bn+ and [bn-methyl imine]+ (n = 3, 5) ions formed by cleavage of aminoxy peptide bond (CO-NH-O) are more intense than bn+ (n = 2, 4) formed by that of peptide bond (CO-NH-C) cleavage. Another characteristic ion observed is due to loss of H3NO from yn+ ions. The cationized (Li+, and Na+) peptides dissociate differently compared to protonated peptides. Intense cationized cn and zn ions are formed due to the cleavage of N-O bond. The deprotonated peptides also show abundant cn- and zn- ions (n = 1, 3, 5) and do not form any yn- ions. All these results clearly indicate the influence of aminoxy peptide bond on fragmentation of these hybrid peptides.

  18. DNA-LCEB: a high-capacity and mutation-resistant DNA data-hiding approach by employing encryption, error correcting codes, and hybrid twofold and fourfold codon-based strategy for synonymous substitution in amino acids.

    PubMed

    Hafeez, Ibbad; Khan, Asifullah; Qadir, Abdul

    2014-11-01

    Data-hiding in deoxyribonucleic acid (DNA) sequences can be used to develop an organic memory and to track parent genes in an offspring as well as in genetically modified organism. However, the main concerns regarding data-hiding in DNA sequences are the survival of organism and successful extraction of watermark from DNA. This implies that the organism should live and reproduce without any functional disorder even in the presence of the embedded data. Consequently, performing synonymous substitution in amino acids for watermarking becomes a primary option. In this regard, a hybrid watermark embedding strategy that employs synonymous substitution in both twofold and fourfold codons of amino acids is proposed. This work thus presents a high-capacity and mutation-resistant watermarking technique, DNA-LCEB, for hiding secret information in DNA of living organisms. By employing the different types of synonymous codons of amino acids, the data storage capacity has been significantly increased. It is further observed that the proposed DNA-LCEB employing a combination of synonymous substitution, lossless compression, encryption, and Bose-Chaudary-Hocquenghem coding is secure and performs better in terms of both capacity and robustness compared to existing DNA data-hiding schemes. The proposed DNA-LCEB is tested against different mutations, including silent, miss-sense, and non-sense mutations, and provides substantial improvement in terms of mutation detection/correction rate and bits per nucleotide. A web application for DNA-LCEB is available at http://111.68.99.218/DNA-LCEB.

  19. Naturally occurring amino acid substitutions at Arg1174 in the human insulin receptor result in differential effects on receptor biosynthesis and hybrid formation, leading to discordant clinical phenotypes.

    PubMed

    Rau, H; Kocova, M; O'Rahilly, S; Whitehead, J P

    2000-07-01

    Missense mutations in the tyrosine kinase domain of the human insulin receptor frequently result in a dominantly inherited form of insulin resistance. We noted a marked disparity in the clinical phenotypes of our study subjects with different missense mutations at the same residue (Arg1174) of the insulin receptor. Subjects with a tryptophan substitution (W) were only moderately hyperinsulinemic, whereas those with a glutamine substitution (Q) had severe clinical and biochemical insulin resistance. Studies were undertaken to explore the molecular mechanisms underlying these differences. Both W and Q mutant receptors bound insulin normally but were kinase inactive. The W mutation resulted in more rapid degradation of newly synthesized mutant receptor, which contrasted with the near-normal biosynthesis of the Q receptor. The propensity of the W receptor to form hybrids with the cotransfected wild-type (WT) receptor was also markedly impaired compared with the Q receptor, to an extent greater than could be explained by lower steady-state expression. Thus, the more clinically benign consequences of the heterozygous W mutant receptor are likely to relate to its impaired biosynthesis and/or reduced capacity to form hybrids with WT receptors. In addition to providing an explanation for the milder phenotype of 1174W versus 1174Q carriers, these studies provide further support for the notion that the dominant-negative effect of insulin receptor tyrosine kinase mutations involves the competition between inactive mutant homodimers and WT/mutant hybrids with active WT homodimers for both ligands and intracellular substrates.

  20. A field operational test on valve-regulated lead-acid absorbent-glass-mat batteries in micro-hybrid electric vehicles. Part I. Results based on kernel density estimation

    NASA Astrophysics Data System (ADS)

    Schaeck, S.; Karspeck, T.; Ott, C.; Weckler, M.; Stoermer, A. O.

    2011-03-01

    In March 2007 the BMW Group has launched the micro-hybrid functions brake energy regeneration (BER) and automatic start and stop function (ASSF). Valve-regulated lead-acid (VRLA) batteries in absorbent glass mat (AGM) technology are applied in vehicles with micro-hybrid power system (MHPS). In both part I and part II of this publication vehicles with MHPS and AGM batteries are subject to a field operational test (FOT). Test vehicles with conventional power system (CPS) and flooded batteries were used as a reference. In the FOT sample batteries were mounted several times and electrically tested in the laboratory intermediately. Vehicle- and battery-related diagnosis data were read out for each test run and were matched with laboratory data in a data base. The FOT data were analyzed by the use of two-dimensional, nonparametric kernel estimation for clear data presentation. The data show that capacity loss in the MHPS is comparable to the CPS. However, the influence of mileage performance, which cannot be separated, suggests that battery stress is enhanced in the MHPS although a battery refresh function is applied. Anyway, the FOT demonstrates the unsuitability of flooded batteries for the MHPS because of high early capacity loss due to acid stratification and because of vanishing cranking performance due to increasing internal resistance. Furthermore, the lack of dynamic charge acceptance for high energy regeneration efficiency is illustrated. Under the presented FOT conditions charge acceptance of lead-acid (LA) batteries decreases to less than one third for about half of the sample batteries compared to new battery condition. In part II of this publication FOT data are presented by multiple regression analysis (Schaeck et al., submitted for publication [1]).

  1. Anthocyanins and Phenolic Acids of Hybrid and Native Blue Maize (Zea mays L.) Extracts and Their Antiproliferative Activity in Mammary (MCF7), Liver (HepG2), Colon (Caco2 and HT29) and Prostate (PC3) Cancer Cells.

    PubMed

    Urias-Lugo, D A; Heredia, J B; Muy-Rangel, M D; Valdez-Torres, J B; Serna-Saldívar, S O; Gutiérrez-Uribe, J A

    2015-06-01

    Blue maize is an excellent source of bioactive components such as phenolic acids and anthocyanins but when it is processed for human consumption, these compounds decrease considerably. Therefore, blue maize could be directed to produce nutraceutical extracts. The aim of this study was to evaluate the relation between anthocyanins composition of acidified and non-acidified extracts from native and hybrid blue maize genotypes and their antiproliferative effect in mammary (MCF7), liver (HepG2), colon (Caco2 and HT29) and prostate (PC3) cancer cells. The most abundant phenolic acid was ferulic acid. Nine anthocyanins were quantified in the extracts, being Cy3-Glu the most abundant. Acylated forms were also obtained in high abundance depending of the extraction method. An extract concentration range of 4.31 to 7.23 mg/mL inhibited by 50% the growth of untransformed cells NIH3T3. Antiproliferative effect on PC3, Caco2, HepG2 and MCF7 cancer cells of acidified extracts from hybrid blue maize was larger than the observed using non-acidified extracts. Among the nine compounds that were quantified in the extracts tested, CyMalGlu I showed the strongest correlation with the reduction of cell viability in Caco2 (-0.876), HepG2 (-0.813), MCF7 (-0.765) and PC3 (-0.894). No significant correlation or differences in antiproliferative effect on HT29 was found among the extracts. The method of extraction of maize anthocyanins must be selected to obtain a high yield of CyMalGlu I more than only Cy3-Glu since acylation affects the inhibition of cancer cell growth.

  2. A hybrid computational fluid dynamics and physiologically based pharmacokinetic model for comparison of predicted tissue concentrations of acrylic acid and other vapors in the rat and human nasal cavities following inhalation exposure.

    PubMed

    Frederick, C B; Gentry, P R; Bush, M L; Lomax, L G; Black, K A; Finch, L; Kimbell, J S; Morgan, K T; Subramaniam, R P; Morris, J B; Ultman, J S

    2001-05-01

    To assist in interspecies dosimetry comparisons for risk assessment of the nasal effects of organic acids, a hybrid computational fluid dynamics (CFD) and physiologically based pharmacokinetic (PBPK) dosimetry model was constructed to estimate the regional tissue dose of inhaled vapors in the rat and human nasal cavity. Application to a specific vapor would involve the incorporation of the chemical-specific reactivity, metabolism, partition coefficients, and diffusivity (in both air and tissue phases) of the vapor. This report describes the structure of the CFD-PBPK model and its application to a representative acidic vapor, acrylic acid, for interspecies tissue concentration comparisons to assist in risk assessment. By using the results from a series of short-term in vivo studies combined with computer modeling, regional nasal tissue dose estimates were developed and comparisons of tissue doses between species were conducted. To make these comparisons, the assumption was made that the susceptibilities of human and rat olfactory epithelium to the cytotoxic effects of organic acids were similar, based on similar histological structure and common mode of action considerations. Interspecies differences in response were therefore assumed to be driven primarily by differences in nasal tissue concentrations that result from regional differences in nasal air flow patterns relative to the species-specific distribution of olfactory epithelium in the nasal cavity. The results of simulations with the seven-compartment CFD-PBPK model suggested that the olfactory epithelium of the human nasal cavity would be exposed to tissue concentrations of acrylic acid similar to that of the rat nasal cavity when the exposure conditions are the same. Similar analysis of CFD data and CFD-PBPK model simulations with a simpler one-compartment model of the whole nasal cavities of rats and humans provides comparable results to averaging over the compartments of the seven-compartment model. These

  3. [Oligonucleotide derivatives in the nucleic acid hybridization analysis. III. Synthesis and investigation of properties of oligonucleotides, bearing bifunctional non-nucleotide insert].

    PubMed

    Kupriushkin, M S; Pyshnyĭ, D V

    2012-01-01

    Non-nucleotide phosporamidites were synthetized, having branched backbone with different position of functional groups. Obtained phosphoramidite monomers contain intercalator moiety--6-chloro-2-methoxyacridine, and additional hydroxyl residue protected with dimethoxytrityl group or with tert-butyldimethylsilyl group for post-synthetic modification. Synthesized oligothymidilates contain one or more modified units in different positions of sequence. Melting temperature and thermodynamic parameters of formation of complementary duplexes formed by modified oligonucleotides was defined (change in enthalpy and entropy). The introduction of intercalating residue causes a significant stabilization of DNA duplexes. It is shown that the efficiency of the fluorescence of acridine residue in the oligonucleotide conjugate significantly changes upon hybridization with DNA.

  4. A field operational test on valve-regulated lead-acid absorbent-glass-mat batteries in micro-hybrid electric vehicles. Part II. Results based on multiple regression analysis and tear-down analysis

    NASA Astrophysics Data System (ADS)

    Schaeck, S.; Karspeck, T.; Ott, C.; Weirather-Koestner, D.; Stoermer, A. O.

    2011-03-01

    In the first part of this work [1] a field operational test (FOT) on micro-HEVs (hybrid electric vehicles) and conventional vehicles was introduced. Valve-regulated lead-acid (VRLA) batteries in absorbent glass mat (AGM) technology and flooded batteries were applied. The FOT data were analyzed by kernel density estimation. In this publication multiple regression analysis is applied to the same data. Square regression models without interdependencies are used. Hereby, capacity loss serves as dependent parameter and several battery-related and vehicle-related parameters as independent variables. Battery temperature is found to be the most critical parameter. It is proven that flooded batteries operated in the conventional power system (CPS) degrade faster than VRLA-AGM batteries in the micro-hybrid power system (MHPS). A smaller number of FOT batteries were applied in a vehicle-assigned test design where the test battery is repeatedly mounted in a unique test vehicle. Thus, vehicle category and specific driving profiles can be taken into account in multiple regression. Both parameters have only secondary influence on battery degradation, instead, extended vehicle rest time linked to low mileage performance is more serious. A tear-down analysis was accomplished for selected VRLA-AGM batteries operated in the MHPS. Clear indications are found that pSoC-operation with periodically fully charging the battery (refresh charging) does not result in sulphation of the negative electrode. Instead, the batteries show corrosion of the positive grids and weak adhesion of the positive active mass.

  5. Characterization of Hybrid Polyhedral Oligomeric Silsesquioxane (POSS)-Polybenzimidazole (PBI)-Phosphoric Acid (PA) Materials Intended for Proton Exchange Membranes (PEM)

    NASA Astrophysics Data System (ADS)

    Bubeck, Robert; Stark, Edmund; Decker, Berryinne; Hartmann-Thompson, Claire

    2013-03-01

    Isophthalic acid and 3,3'-diaminobenzidine (DAB) were polymerized in the presence of polyphosphoric acid (PPA) and various additives, degree of polymerization was monitored by viscosity and torque change measurements, and membranes were prepared by casting the reaction solution and allowing PPA to hydrolyze to PA under ambient conditions. As a function of relative humidity, the membranes were characterized for (1) acid content, (2) in-plane conductivity and (3) complex shear modulus G* obtained via oscillatory parallel plate dynamic mechanical spectroscopy. The addition of sulfonated octaphenyl polyhedral oligomeric silsesquixane (S-POSS) to m-polybenzimidazole (PBI)-phosphoric acid (PA) membranes resulted in increased in-plane proton conductivity at high temperatures (120-150 °C) and increased G* relative to a m-PBI control membrane and to m-PBI control membranes carrying comparable weight loadings of non-proton conducting octaphenyl-POSS nanoadditive or silica.

  6. Foldamers to nanotubes: influence of amino acid side chains in the hierarchical assembly of α,γ(4)-hybrid peptide helices.

    PubMed

    Jadhav, Sandip V; Misra, Rajkumar; Gopi, Hosahudya N

    2014-12-08

    Supramolecular assembly of various artificially folded 12-helical architectures composed of γ(4) -Val, γ(4) -Leu and γ(4) -Phe residues is investigated. In contrast to the 12-helices composed of γ(4) -Val and γ(4) -Leu residues, the helices with γ(4) -Phe residues displayed unique elongated nanotubular architectures. The elongated nanotube assembly was further explored as a template for biomineralization of silver ions to silver nanowires. A comparative study using an analogous α-peptide helix reveals the importance of the spatial arrangement of aromatic side chains along the helical cylinder in a 12-helix. These results suggested that the proteolytically and structurally stable α,γ(4) -hybrid peptide 12-helices may serve as a new generation of potential templates in the design of functional biomaterials.

  7. Dietary fish oil supplements increase tissue n-3 fatty acid composition and expression of delta-6 desaturase and elongase-2 in Jade Tiger hybrid abalone.

    PubMed

    Mateos, Hintsa T; Lewandowski, Paul A; Su, Xiao Q

    2011-08-01

    This study was conducted to investigate the effects of fish oil (FO) supplements on fatty acid composition and the expression of ∆6 desaturase and elongase 2 genes in Jade Tiger abalone. Five test diets were formulated to contain 0.5, 1.0, 1.5, 2.0 and 2.5% of FO respectively, and the control diet was the normal commercial abalone diet with no additional FO supplement. The muscle, gonad and digestive glands (DG) of abalone fed with all of the five test diets showed significantly high levels of total n-3 polyunsaturated fatty acid (PUFA), eicosapentaenoic acid (EPA), docosapentaenoic acid n-3 (DPAn-3), and docosahexaenoic acid (DHA) than the control group. In all three types of tissue, abalone fed diet supplemented with 1.5% FO showed the highest level of these fatty acids (P < 0.05). For DPAn-3 the higher level was also found in muscle and gonad of abalone fed diet supplemented with 2% FO (P < 0.05). Elongase 2 expression was markedly higher in the muscle of abalone fed diet supplemented with 1.5% FO (P < 0.05), followed by the diet containing 2% FO supplement. For ∆6 desaturase, significantly higher expression was observed in muscle of abalone fed with diet containing 0.5% FO supplement (P < 0.05). Supplementation with FO in the normal commercial diet can significantly improve long chain n-3 PUFA level in cultured abalone, with 1.5% being the most effective supplementation level.

  8. A Nucleic Acid Probe and Method for the Rapid Detection of Typhoid Fever Bacteria.

    DTIC Science & Technology

    This invention relates to a nucleic acid probe and method for the rapid detection of typhoid fever bacteria by use of a nucleic acid hybridization...a nucleic acid hybridization reaction with a clinical specimen containing typhoid fever bacteria.

  9. Inhibition of AAC(6')-Ib-mediated resistance to amikacin in Acinetobacter baumannii by an antisense peptide-conjugated 2',4'-bridged nucleic acid-NC-DNA hybrid oligomer.

    PubMed

    Lopez, Christina; Arivett, Brock A; Actis, Luis A; Tolmasky, Marcelo E

    2015-09-01

    Multiresistant Acinetobacter baumannii, a common etiologic agent of severe nosocomial infections in compromised hosts, usually harbors aac(6')-Ib. This gene specifies resistance to amikacin and other aminoglycosides, seriously limiting the effectiveness of these antibiotics. An antisense oligodeoxynucleotide (ODN4) that binds to a duplicated sequence on the aac(6')-Ib mRNA, one of the copies overlapping the initiation codon, efficiently inhibited translation in vitro. An isosequential nuclease-resistant hybrid oligomer composed of 2',4'-bridged nucleic acid-NC (BNA(NC)) residues and deoxynucleotides (BNA(NC)-DNA) conjugated to the permeabilizing peptide (RXR)4XB ("X" and "B" stand for 6-aminohexanoic acid and β-alanine, respectively) (CPPBD4) inhibited translation in vitro at the same levels observed in testing ODN4. Furthermore, CPPBD4 in combination with amikacin inhibited growth of a clinical A. baumannii strain harboring aac(6')-Ib in liquid cultures, and when both compounds were used as combination therapy to treat infected Galleria mellonella organisms, survival was comparable to that seen with uninfected controls. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  10. Peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) assay for specific detection of Mycobacterium immunogenum and DNA-FISH assay for analysis of pseudomonads in metalworking fluids and sputum.

    PubMed

    Selvaraju, Suresh B; Kapoor, Renuka; Yadav, Jagjit S

    2008-01-01

    Specific and rapid detection and quantification of mycobacteria in contaminated metalworking fluid (MWF) are problematic due to complexity of the matrix and heavy background co-occurring microflora. Furthermore, cross-reactivity among neighboring species of Mycobacterium makes species differentiation difficult for this genus. Here, we report for the first time a species-specific peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) method for Mycobacterium immunogenum, a non-tuberculous Mycobacterium species prevalent in MWF and implicated in occupational lung disease hypersensitivity pneumonitis and pseudo-outbreaks. A novel species-specific 14-bp PNA probe was designed for M. immunogenum based on its 16S rRNA gene sequence and was validated for specificity, by testing against a panel of other phylogenetically closely related rapidly growing mycobacteria and representative species of gram-positive, gram-negative, and acid fast organisms. In addition, a DNA-FISH protocol was optimized for co-detection of Pseudomonas, the most predominantly co-occurring genus in contaminated MWF. Reliable quantification for both the test organisms was achieved at or above a cell density of 10(3)cellsml(-1), a recognized minimum limit for microscopic quantification. The mycobacterial PNA-FISH assay was successfully adapted to human sputum demonstrating its potential for clinical diagnostic applications in addition to industrial MWF monitoring, to assess MWF-associated exposures and pseudo-outbreaks.

  11. Development of a Simple Adjustable Zinc Acid/Base Hybrid Catalyst for C-C and C-O Bond-Forming and C-C Bond-Cleavage Reactions.

    PubMed

    Yamashita, Yasuhiro; Minami, Kodai; Saito, Yuki; Kobayashi, Shū

    2016-09-06

    A newly designed zinc Lewis acid/base hybrid catalyst was developed. By adjusting the Lewis acidity of the zinc center, aldol-type additions of 2-picolylamine Schiff base to aldehydes proceeded smoothly to afford syn-aldol adduct equivalents, trans-N,O-acetal adducts, in high yields with high selectivities. NMR experiments, including microchanneled cell for synthesis monitoring (MICCS) NMR analysis, revealed that anti-aldol adducts were formed at the initial stage of the reactions under kinetic control, but the final products were the trans-(syn)-N,O-acetal adducts that were produced through a retro-aldol process under thermodynamic control. In the whole reaction process, the zinc catalyst played three important roles: i) promotion of the aldol process (C-C bond formation), ii) cyclization process to the N,O-acetal product (C-O bond formation), and iii) retro-aldol process from the anti-aldol adduct to the syn-aldol adduct (C-C bond cleavage and C-C bond formation).

  12. Hybrid Gear

    NASA Technical Reports Server (NTRS)

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

    2016-01-01

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

  13. Covalently bonded sulfonic acid magnetic graphene oxide: Fe3O4@GO-Pr-SO3H as a powerful hybrid catalyst for synthesis of indazolophthalazinetriones.

    PubMed

    Doustkhah, Esmail; Rostamnia, Sadegh

    2016-09-15

    Multistep synthesis of covalently sulfonated magnetic graphene oxide was achieved by starting from Hummer's method to produce graphene oxide (GO) from chemical oxidation of graphite. Then, GO nanosheets were applied to support Fe3O4 nanoparticles (Fe3O4@GO) using co-precipitation method in the presence of GO sheets. This strategy led to formation of uniform particles of Fe3O4 on the surface of GO sheets. Then, it was sulfonated (Fe3O4@GO-Pr-SO3H) through modification with 3-mercaptopropyltrimethoxysilane (MPTMS) and subsequent oxidation with hydrogen peroxide (H2O2). In comparison, the covalently bonded propyl sulfonic acid groups were more prevailing rather to sulfonic acids of GO itself. The proposed catalyst was more active and recyclable at least for 11 runs. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Selective detection of dopamine in the presence of uric acid using a gold nanoparticles-poly(luminol) hybrid film and multi-walled carbon nanotubes with incorporated β-cyclodextrin modified glassy carbon electrode.

    PubMed

    Jia, Dong; Dai, Jianyuan; Yuan, Hongyan; Lei, Ling; Xiao, Dan

    2011-10-15

    Gold nanoparticles-poly(luminol) (Plu-AuNPs) hybrid film and multi-walled carbon nanotubes with incorporated β-cyclodextrin modified glassy carbon electrode (β-CD-MWCNTs/Plu-AuNPs/GCE) was successfully prepared for simultaneous determination of dopamine (DA) and uric acid (UA). The surface of the modified electrode has been characterized by X-ray photo-electron spectroscopy (XPS), energy dispersive X-ray spectroscopy (EDS), field-emission scanning electron microscope (SEM) and transmission electron microscope (TEM). Cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS) and differential pulse voltammetry (DPV) have been used to investigate the β-CD-MWCNTs/Plu-AuNPs composite film. Gold nanoparticles anchored into poly(luminol) film exhibited catalytic activity for DA. MWCNTs with incorporated β-CD can greatly promote the direct electron transfer. In 0.10 M phosphate buffer solution (PBS, pH 7.0), the DPV response of the β-CD-MWCNTs/Plu-AuNPs/GCE sensor to DA is about 8-fold as compared with the Plu-AuNPs/GCE sensor, and the detection limit for DA is about one order of magnitude lower than the Plu-AuNPs/GCE sensor. The steady-state current response increases linearly with DA concentration from 1.0 × 10(-6) to 5.6 × 10(-5)M with a low detection limit (S/N=3) of 1.9 × 10(-7)M. Moreover, the interferences of ascorbic acid (AA) and uric acid (UA) are effectively diminished. The applicability of the prepared electrode has been demonstrated by measuring DA contents in dopamine hydrochloride injection.

  15. Post-Translational Modification and Secretion of Azelaic Acid Induced 1 (AZI1), a Hybrid Proline-Rich Protein from Arabidopsis.

    PubMed

    Pitzschke, Andrea; Xue, Hui; Persak, Helene; Datta, Sneha; Seifert, Georg J

    2016-01-12

    Arabidopsis EARLI-type hybrid proline-rich proteins (HyPRPs) consist of a putative N-terminal secretion signal, a proline-rich domain (PRD), and a characteristic eight-cysteine-motif (8-CM). They have been implicated in biotic and abiotic stress responses. AZI1 is required for systemic acquired resistance and it has recently been identified as a target of the stress-induced mitogen-activated protein kinase MPK3. AZI1 gel migration properties strongly indicate AZI1 to undergo major post-translational modifications. These occur in a stress-independent manner and are unrelated to phosphorylation by MAPKs. As revealed by transient expression of AZI1 in Nicotiana benthamiana and Tropaeolum majus, the Arabidopsis protein is similarly modified in heterologous plant species. Proline-rich regions, resembling arabinogalactan proteins point to a possible proline hydroxylation and subsequent O-glycosylation of AZI1. Consistently, inhibition of prolyl hydroxylase reduces its apparent protein size. AZI1 secretion was examined using Arabidopsis protoplasts and seedling exudates. Employing Agrobacterium-mediated leaf infiltration of N. benthamiana, we attempted to assess long-distance movement of AZI1. In summary, the data point to AZI1 being a partially secreted protein and a likely new member of the group of hydroxyproline-rich glycoproteins. Its dual location suggests AZI1 to exert both intra- and extracellular functions.

  16. Locked nucleic acid (LNA) induced effect on the hybridization and fluorescence properties of oligodeoxyribonucleotides modified with nucleobase-functionalized DNA monomers.

    PubMed

    Kaura, Mamta; Hrdlicka, Patrick J

    2015-07-14

    LNA and nucleobase-modified DNA monomers are two types of building blocks that are used extensively in oligonucleotide chemistry. However, there are only very few reports in which these two monomer families are used alongside each other. In the present study we set out to characterize the biophysical properties of oligodeoxyribonucleotides in which C5-modified 2'-deoxyuridine or C8-modified 2'-deoxyadenosine monomers are flanked by LNA nucleotides. We hypothesized that the LNA monomers would alter the sugar rings of the modified DNA monomers toward more RNA-like North-type conformations for maximal DNA/RNA affinity and specificity. Indeed, the incorporation of LNA monomers almost invariably results in increased target affinity and specificity relative to the corresponding LNA-free ONs, but the magnitude of the stabilization varies greatly. Introduction of LNA nucleotides as direct neighbors into C5-pyrene-functionalized pyrimidine DNA monomers yields oligonucleotide probes with more desirable photophysical properties as compared to the corresponding LNA-free probes, including more intense fluorescence emission upon target binding and improved discrimination of single nucleotide polymorphisms (SNPs). These hybrid oligonucleotides are therefore promising probes for diagnostic applications.

  17. Epitope mapping and key amino acid identification of anti-CD22 immunotoxin CAT-8015 using hybrid β-lactamase display

    PubMed Central

    Bannister, D.; Popovic, B.; Sridharan, S.; Giannotta, F.; Filée, P.; Yilmaz, N.; Minter, R.

    2011-01-01

    Monoclonal antibodies are a commercially successful class of drug molecules and there are now a growing number of antibodies coupled to toxic payloads, which demonstrate clinical efficacy. Determining the precise epitope of therapeutic antibodies is beneficial in understanding the structure–activity relationship of the drug, but in many cases is not done due to the structural complexity of, in particular, conformational protein epitopes. Using the immunotoxin CAT-8015 as a test case, this study demonstrates that a new methodology, hybrid β-lactamase display, can be employed to elucidate a complex epitope on CD22. Following insertion of random CD22 gene fragments into a permissive site within β-lactamase, proteins expressed in Escherichia coli were first screened for correct folding by resistance to ampicillin and then selected by phage display for affinity to CAT-8015. The optimal protein region recognised by CAT-8015 could then be used as a tool for fine epitope mapping, using alanine-scanning analysis, demonstrating that this technology is well suited to the rapid characterisation of antibody epitopes. PMID:21159620

  18. Epitope mapping and key amino acid identification of anti-CD22 immunotoxin CAT-8015 using hybrid β-lactamase display.

    PubMed

    Bannister, D; Popovic, B; Sridharan, S; Giannotta, F; Filée, P; Yilmaz, N; Minter, R

    2011-04-01

    Monoclonal antibodies are a commercially successful class of drug molecules and there are now a growing number of antibodies coupled to toxic payloads, which demonstrate clinical efficacy. Determining the precise epitope of therapeutic antibodies is beneficial in understanding the structure-activity relationship of the drug, but in many cases is not done due to the structural complexity of, in particular, conformational protein epitopes. Using the immunotoxin CAT-8015 as a test case, this study demonstrates that a new methodology, hybrid β-lactamase display, can be employed to elucidate a complex epitope on CD22. Following insertion of random CD22 gene fragments into a permissive site within β-lactamase, proteins expressed in Escherichia coli were first screened for correct folding by resistance to ampicillin and then selected by phage display for affinity to CAT-8015. The optimal protein region recognised by CAT-8015 could then be used as a tool for fine epitope mapping, using alanine-scanning analysis, demonstrating that this technology is well suited to the rapid characterisation of antibody epitopes.

  19. Post-Translational Modification and Secretion of Azelaic Acid Induced 1 (AZI1), a Hybrid Proline-Rich Protein from Arabidopsis

    PubMed Central

    Pitzschke, Andrea; Xue, Hui; Persak, Helene; Datta, Sneha; Seifert, Georg J.

    2016-01-01

    Arabidopsis EARLI-type hybrid proline-rich proteins (HyPRPs) consist of a putative N-terminal secretion signal, a proline-rich domain (PRD), and a characteristic eight-cysteine-motif (8-CM). They have been implicated in biotic and abiotic stress responses. AZI1 is required for systemic acquired resistance and it has recently been identified as a target of the stress-induced mitogen-activated protein kinase MPK3. AZI1 gel migration properties strongly indicate AZI1 to undergo major post-translational modifications. These occur in a stress-independent manner and are unrelated to phosphorylation by MAPKs. As revealed by transient expression of AZI1 in Nicotiana benthamiana and Tropaeolum majus, the Arabidopsis protein is similarly modified in heterologous plant species. Proline-rich regions, resembling arabinogalactan proteins point to a possible proline hydroxylation and subsequent O-glycosylation of AZI1. Consistently, inhibition of prolyl hydroxylase reduces its apparent protein size. AZI1 secretion was examined using Arabidopsis protoplasts and seedling exudates. Employing Agrobacterium-mediated leaf infiltration of N. benthamiana, we attempted to assess long-distance movement of AZI1. In summary, the data point to AZI1 being a partially secreted protein and a likely new member of the group of hydroxyproline-rich glycoproteins. Its dual location suggests AZI1 to exert both intra- and extracellular functions. PMID:26771603

  20. Understanding the effect of locked nucleic acid and 2'-O-methyl modification on the hybridization thermodynamics of a miRNA-mRNA pair in the presence and absence of AfPiwi protein.

    PubMed

    Kumar, Santosh; Mapa, Koyeli; Maiti, Souvik

    2014-03-18

    miRNAs are some of the key epigenetic regulators of gene expression. They act through hybridization with their target mRNA and modulate the level of respective proteins via different mechanisms. Various cancer conditions are known to be associated with up- and downregulation of the oncogenic and tumor suppressor miRNAs, respectively. The levels of aberrantly expressed oncogenic miRNAs can be downregulated in different ways. Similarly, restoration of tumor suppressor miRNAs to their normal levels can be achieved using miRNA mimics. However, the use of miRNA mimics is limited by their reduced biostability and function. We have studied the hybridization thermodynamics of the miRNA 26a (11-mer, including the seed sequence) guide strand with the mRNA (11-mer) target strand in the absence and presence of AfPiwi protein. We have also inserted locked nucleic acids (LNAs) and 2'-O-methyl-modified nucleotides into the guide strand, in a walk-through manner, to assess their effect on the binding efficiency between guide and target RNA. Insertion of LNA and 2'-O-methyl-modified nucleotides into the guide strand helped to strengthen the binding affinity irrespective of the position of insertion. However, in the presence of AfPiwi protein, these modifications reduced the binding affinity to different extents depending on the position of insertion. Insertion of a modification leads to an increase in the enthalpic contribution with an increased unfavorable entropic contribution, which negatively compensates for the higher favorable enthalpy.

  1. Efficacy and tolerance of an injectable medical device containing stable hybrid cooperative complexes of high- and low-molecular-weight hyaluronic acid: a monocentric 16 weeks open-label evaluation

    PubMed Central

    Sparavigna, Adele; Tenconi, Beatrice

    2016-01-01

    Background An injectable medical device containing stable hybrid cooperative complexes of high- and low-molecular-weight hyaluronic acid (HA) has been developed with characteristics suited for a global improvement of facial esthetics. Objective To evaluate the HA product performance in improving some key facial esthetic features. The study employed clinical scales, subjective evaluations, and facial skin objective measurements. Methods A single Italian site treated 64 female subjects aged 38–60 years, with injections at five predetermined points, on each side of the face, with a 4-week time lapse between the first and the second product administration. Subjects were evaluated after 4, 8, 12, and 16 weeks, using validated clinical scales, subjective evaluation, and objective quantitative outcome measures. Assessment of esthetic results included photographic documentation. Results Both the clinical and subjective assessments, and the majority of objective instrumental parameters indicated an improvement throughout the study and were already significant at week 4 or 8 and were still significant at week 16 (3 months after the second treatment). Minor and temporary local skin reactions were observed in 23% of subjects at the site of the injections, and the global judgment on tolerability was good or excellent, both in the investigators’ opinion and volunteers’ self-evaluation. Conclusion Both subjective and objective improvement of the facial parameters was consistent with the bio-remodeling purpose, and persistent and still statistically significant at the end of the study. The tolerability and safety profile of the product were judged good or excellent both by investigators and volunteers. This study supports the claim for bio-remodeling of these stable hybrid cooperative complexes of low- and high-molecular-weight HA. PMID:27713647

  2. Developing mixed films of immobilized oligonucleotides and quantum dots for the multiplexed detection of nucleic acid hybridization using a combination of fluorescence resonance energy transfer and direct excitation of fluorescence.

    PubMed

    Algar, W Russ; Krull, Ulrich J

    2010-04-20

    Methods have been developed for the simultaneous and selective detection of three target nucleic acid sequences based on mixed films of immobilized quantum dots (QDs) and oligonucleotide probes. CdSe/ZnS QDs were immobilized on optical fibers and conjugated with mixtures of different probe oligonucleotides. Hybridization events were detected using a combination of fluorescence from direct excitation and fluorescence sensitized by resonance energy transfer (FRET). A sandwich assay format was used to associate dye labeled reporter oligonucleotides with probe-target hybrids formed at the surface of the optical fiber. One detection channel utilized direct excitation of Pacific Blue and the two other detection channels were based on FRET. In one strategy, green emitting QDs were used as donors with Cy3 and Rhodamine Red-X acceptors. In a second strategy, green and red emitting QDs were coimmobilized and used as donors with Cy3 and Alexa Fluor 647 acceptors, respectively. Selective three-plex detection was demonstrated with both strategies. Several key design criteria that were explored to optimize the relative signal magnitude between channels included: the ratio of probe associated with direct excitation versus probes associated with FRET; the relative amounts of each FRET probe and corresponding spectral overlap; and the photoluminescence ratio between immobilized green and red emitting QDs (where applicable). Careful selection of probe sequences and lengths were important for the discrimination of single nucleotide polymorphisms in one channel without suppressing binding of target in the other two channels. This work provides a basis for the development of multiplexed biosensors that are ensemble compatible and do not require discrete sensor elements, spatial registration, sorting technology, or single molecule spectroscopy.

  3. Hybrid Simulator

    SciTech Connect

    Trujillo, David J.; Sridharan, Srikesh; Weinstock, Irvin

    2005-10-15

    HybSim (short for Hybrid Simulator) is a flexible, easy to use screening tool that allows the user to quanti the technical and economic benefits of installing a village hybrid generating system and simulates systems with any combination of —Diesel generator sets —Photovoltaic arrays -Wind Turbines and -Battery energy storage systems Most village systems (or small population sites such as villages, remote military bases, small communities, independent or isolated buildings or centers) depend on diesel generation systems for their source of energy. HybSim allows the user to determine other "sources" of energy that can greatly reduce the dollar to kilo-watt hour ratio. Supported by the DOE, Energy Storage Program, HybSim was initially developed to help analyze the benefits of energy storage systems in Alaskan villages. Soon after its development, other sources of energy were added providing the user with a greater range of analysis opportunities and providing the village with potentially added savings. In addition to village systems, HybSim has generated interest for use from military institutions in energy provisions and USAID for international village analysis.

  4. Organic/inorganic hybrid coatings for anticorrosion

    NASA Astrophysics Data System (ADS)

    He, Zhouying

    Compared to organic coatings, organic-inorganic hybrid coatings can potentially improve the anticorrosion performance. The organic phase provides the excellent mechaincal and barrier properties while the inorganic phase acts as an adhesion promoter and corrosion inhibitor. Despite that many studies on alkoxylsilane-based hybrid coatings have been developed and studied, their weatherability and anticorrosion performance has been rarely evaluated. On the other hand, organic-inorganic hybrid coatings based on mixed sol-gel precursors have received much less attention compared to alkoxylsilane-based hybrid coatings. In the first part, polyurethane hybrid coatings with a unique hybrid crosslinked structure as an improved unicoat were successfully prepared. The effect of polyesters on physical properties of the hybrid coatings was studied. Polyurethane coatings derived from cycloaliphatic polyester show comparable properties than those derived from the commercially viable aromatic polyester. Introducing the polysiloxane part into the polyurethane coatings enhanced the crosslinking density, Tg, mechanical properties, and general coating properties. The increased adhesion between the hybrid coating and the substrate make the hybrid coating a good candidate for anticorrosion application, which is shown by electrochemical impedance spectroscopy (EIS). The degradation mechanism of the polyurethane/polysiloxane hybrid coatings under various weathering conditions was shown to be the scission of the urethane and ester groups in the organic phase along with reorganizing and rearranging of the inorganic phase. The anticorrosion performance of the cycloaliphatic hybrid was much better than that of aromatic based hybrid under outdoor weathering based on visual observation and EIS analysis. Acid undercutting is an issue for TEOS based hybrid coating. In the second part, design of experiments (DOEs) was used to statistically investigate on the effect of sol-gel precursors. The

  5. Hybridization parameters revisited: solutions containing SDS.

    PubMed

    Rose, Ken; Mason, John O; Lathe, Richard

    2002-07-01

    Salt concentration governs nucleic acid hybridization according to the Schildkraut-Lifson equation. High concentrations of SDS are used in some common protocols, but the effects of SDS on hybridization stringency have not been reported. We investigated hybridization parameters in solutions containing SDS. With targets immobilized on nylon membranes and PCR- or transcription-generated probes, we report that the 50% dissociation temperature (Tm*) in the absence of SDS was 15 degrees C-17degrees C lower than the calculated Tm. SDS had only modest effects on Tm* [1% (w/v) equating to 8 mM NaCl]. RNA/DNA hybrids were approximately 11 degrees C more stable than DNA/DNA hybrids. Incomplete homology (69%) significantly reduced the Tm* for DNA/DNA hybrids (approximately /4degrees C; 0.45 degrees C/% nonhomology) but far less so for RNA/DNA hybrids (approximately 2.3 degrees C; approximately 0.07 degrees C/% non-homology); incomplete homology also markedly reduced the extent of hybridization. On these nylonfilters, SDS had a major effect on nonspecific binding. Buffers lacking SDS, or with low salt concentration, gave high hybridization backgrounds; buffers containing SDS, or high-salt buffers, gave reproducibly low backgrounds.

  6. Hybrid optofluidic biosensors

    NASA Astrophysics Data System (ADS)

    Parks, Joshua W.

    Optofluidics, born of the desire to create a system containing microfluidic environments with integrated optical elements, has seen dramatic increases in popularity over the last 10 years. In particular, the application of this technology towards chip based molecular sensors has undergone significant development. The most sensitive of these biosensors interface liquid- and solid-core antiresonant reflecting optical waveguides (ARROWs). These sensor chips are created using conventional silicon microfabrication. As such, ARROW technology has previously been unable to utilize state-of-the-art microfluidic developments because the technology used--soft polydimethyl siloxane (PDMS) micromolded chips--is unamenable to the silicon microfabrication workflows implemented in the creation of ARROW detection chips. The original goal of this thesis was to employ hybrid integration, or the connection of independently designed and fabricated optofluidic and microfluidic chips, to create enhanced biosensors with the capability of processing and detecting biological samples on a single hybrid system. After successful demonstration of this paradigm, this work expanded into a new direction--direct integration of sensing and detection technologies on a new platform with dynamic, multi-dimensional photonic re-configurability. This thesis reports a number of firsts, including: • 1,000 fold optical transmission enhancement of ARROW optofluidic detection chips through thermal annealing, • Detection of single nucleic acids on a silicon-based ARROW chip, • Hybrid optofluidic integration of ARROW detection chips and passive PDMS microfluidic chips, • Hybrid optofluidic integration of ARROW detection chips and actively controllable PDMS microfluidic chips with integrated microvalves, • On-chip concentration and detection of clinical Ebola nucleic acids, • Multimode interference (MMI) waveguide based wavelength division multiplexing for detection of single influenza virions,

  7. Hybridization-Modulated Ion Fluxes through Peptide-Nucleic-Acid-Functionalized Gold Nanotubules. A New Approach to Quantitative Label-Free DNA Analysis

    PubMed Central

    Jágerszki, Gyula; Gyurcsányi, Róbert E.; Höfler, Lajos; Pretsch, Ernő

    2008-01-01

    The inner walls of gold nanotubules, prepared by template synthesis in the nanopores of polycarbonate track etch membranes, have been chemically modified with peptide nucleic acid (PNA) and used for label-free quantification of complementary DNA sequences. Selective binding of DNA to the PNA modified nanotubules are shown to decrease the flux of optically detected anionic markers through the nanotubules in a concentration-dependent manner. The strong dependence of the biorecognition-modulated ion transport through the nanopores on the ionic strength suggests a dominantly electrostatic exclusion mechanism of the ion flux decrease as a result of DNA binding to the PNA-modified nanopores. PMID:17488052

  8. Synthesis and hybridization property of a boat-shaped pyranosyl nucleic acid containing an exocyclic methylene group in the sugar moiety.

    PubMed

    Mori, Kazuto; Kodama, Tetsuya; Obika, Satoshi

    2015-01-01

    A boat-shaped pyranosyl nucleic acid (BsNA) having an exocyclic methylene group in the sugar moiety was synthesized to investigate the possibility that the axial H3' of original BsNA is the cause of its duplex destabilization. The synthesized BsNA analog was chemically stable against various nucleophiles. From the thermal stability of duplex oligonucleotides including the BsNA analog, it was found that the duplex-forming ability can be sensitive to the size of functional groups at the 3'-position.

  9. Hybridization-modulated ion fluxes through peptide-nucleic-acid- functionalized gold nanotubes. A new approach to quantitative label-free DNA analysis.

    PubMed

    Jágerszki, Gyula; Gyurcsányi, Róbert E; Höfler, Lajos; Pretsch, Ernö

    2007-06-01

    The inner walls of gold nanotubes, prepared by template synthesis in the nanopores of polycarbonate track etch membranes, have been chemically modified with peptide nucleic acid (PNA) and used for label-free quantification of complementary DNA sequences. Selective binding of DNA to the PNA-modified nanotubes is shown to decrease the flux of optically detected anionic markers through the nanotubes in a concentration-dependent manner. The strong dependence of the biorecognition-modulated ion transport through the nanopores on the ionic strength suggests a dominantly electrostatic exclusion mechanism of the ion flux decrease as a result of DNA binding to the PNA-modified nanopores.

  10. Using a combined hydrolysis factor to balance enzymatic saccharification and the structural characteristics of lignin during pretreatment of Hybrid poplar with a fully recyclable solid acid.

    PubMed

    Ji, Hairui; Song, Yanliang; Zhang, Xu; Tan, Tianwei

    2017-08-01

    In this study, a new pretreatment strategy for lignocellulosic was developed using a fully recyclable solid acid, Toluenesulfonic acid (p-TsOH). A combined hydrolysis factor (CHF) as a pretreatment severity was used to balance enzymatic saccharification and the structural characteristics of lignin. The results from degradation of carbohydrates, enzymatic hydrolysis of cellulose and characterization of lignin by FT-IR, (31)P NMR, GPC, 2D-HSQC NMR indicated that a CHF of approximately 3.90 was the optimal pretreatment severity to facilitate enzymatic saccharification and the potential serviceability of lignin. Then approximately 90% of the xylan was removed to result in a reasonable sugar yield of 76%. Residual lignin showed low molecular weight (Mw, 5783g/mol), narrow polydispersities (Mw/Mn, 1.10) and high content of phenolic hydroxyl groups (3.702mmol/g); it may be a potential feedstock for phenol monomer and polymeric materials production. In short, this process was regarded as a promising approach to achieve an efficient conversion of lignocellulosic biomass to sugar products and lignin-based materials. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Identification of low molecular weight organic acids by ion chromatography/hybrid quadrupole time-of-flight mass spectrometry during Uniblu-A ozonation.

    PubMed

    Amorisco, Apollonia; Locaputo, Vito; Pastore, Carlo; Mascolo, Giuseppe

    2013-01-15

    The balance of organic nitrogen and sulfur during ozonation of organic pollutants often shows a lack of complete mineralization. It follows that polar and ionic by-products are likely to be present that are difficult to identify by liquid chromatography/mass spectrometry (LC/MS). The structural elucidation of low molecular weight organic acids arising from Uniblu-OH ozonation has been investigated by ion chromatography/electrospray tandem mass spectrometry (IC/ESI-MS/MS) employing a quadrupole time-of-flight mass spectrometer. Unequivocal elemental composition of the by-products was determined by a combination of mass accuracy and high spectral accuracy. The employed identification strategy was demonstrated to be a powerful method of unequivocally assigning a single chemical composition to each identified compound. The exact mass measurements of [M-H](-) ions allowed the elemental formulae and related structures of eighteen by-products to be determined confidently. The main degradation pathways were found to be decarboxylation and oxidation. The experimental procedure allowed the identification of both nitrogen- and sulfur-containing organic acid by-products arising from Uniblu-OH ozonation. The obtained results are of environmental relevance for the balance of organic nitrogen and sulfur during the ozonation of organic pollutants due to the lack of complete mineralization of the compounds containing these atoms. Copyright © 2012 John Wiley & Sons, Ltd.

  12. Hybridized tetraquarks

    DOE PAGES

    Esposito, Angelo; Pilloni, Alessadro; Polosa, A. D.

    2016-05-12

    In this study, we propose a new interpretation of the neutral and charged X,Z exotic hadron resonances. Hybridized-tetraquarks are neither purely compact tetraquark states nor bound or loosely bound molecules. The latter would require a negative or zero binding energy whose counterpart in h-tetraquarks is a positive quantity. The formation mechanism of this new class of hadrons is inspired by that of Feshbach metastable states in atomic physics. The recent claim of an exotic resonance in the B0s π± channel by the D0 collaboration and the negative result presented subsequently by the LHCb collaboration are understood in this scheme, togethermore » with a considerable portion of available data on X, Z particles. Considerations on a state with the same quantum numbers as the X(5568) are also made.« less

  13. Regulation of progesterone receptor messenger ribonucleic acid in the rat medial preoptic nucleus by estrogenic and antiestrogenic compounds: an in situ hybridization study.

    PubMed

    Shughrue, P J; Lane, M V; Merchenthaler, I

    1997-12-01

    Progesterone receptor (PR) messenger RNA (mRNA) is concentrated in neurons of the preoptic area and other regions of the rat hypothalamus where it is colocalized with the estrogen receptor and regulated by changes in the steroid hormonal milieu. To date, little is known about the regulation of PR mRNA by estrogens and whether antiestrogenic compounds are capable of modulating its expression. The present studies used in situ hybridization to ascertain the time course of PR mRNA regulation in the medial preoptic nucleus by 17beta-estradiol, determine the effective dose required to elicit a response, and compare the efficacy of 17beta-estradiol with a variety of estrogenic or antiestrogenic compounds. The first series of studies revealed that the treatment of ovariectomized rats with 17beta-estradiol resulted in an increase in PR expression within 2 h, after which it remained elevated until 10 h postinjection and then returned to baseline levels. When ovariectomized rats were injected with 25-1000 ng/kg of 17beta-estradiol and euthanized 6 h later, a dose-dependent increase in the level of PR mRNA was observed, with a maximal response at 1000 ng/kg and an EC50 of 93.5 ng/kg. Subsequent studies evaluated the efficacy of a variety of estrogenic and antiestrogenic compounds in the rat preoptic nucleus. 17Beta-estradiol, diethylstilbestrol, and 17alpha-estradiol all significantly increased the level of PR mRNA, although the degree of induction varied with each compound. The injection of tamoxifen, raloxifene, toremifene, droloxifene, clomiphene, GW 5638, or ICI 182,780 had no significant estrogenic effect on PR gene expression at the dose evaluated. In contrast, when tamoxifen or raloxifene, but not ICI 182,780, was administered in the antagonist mode, a significant dose-related decrease in the estradiol-induced level of PR mRNA was seen in the preoptic area. The results of these studies clearly demonstrate that PR mRNA expression in the rat preoptic area is rapidly

  14. Identifying a base in a nucleic acid