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Sample records for acids detection folding

  1. Fold homology detection using sequence fragment composition profiles of proteins.

    Solis, Armando D; Rackovsky, Shalom R

    2010-10-01

    The effectiveness of sequence alignment in detecting structural homology among protein sequences decreases markedly when pairwise sequence identity is low (the so-called "twilight zone" problem of sequence alignment). Alternative sequence comparison strategies able to detect structural kinship among highly divergent sequences are necessary to address this need. Among them are alignment-free methods, which use global sequence properties (such as amino acid composition) to identify structural homology in a rapid and straightforward way. We explore the viability of using tetramer sequence fragment composition profiles in finding structural relationships that lie undetected by traditional alignment. We establish a strategy to recast any given protein sequence into a tetramer sequence fragment composition profile, using a series of amino acid clustering steps that have been optimized for mutual information. Our method has the effect of compressing the set of 160,000 unique tetramers (if using the 20-letter amino acid alphabet) into a more tractable number of reduced tetramers (approximately 15-30), so that a meaningful tetramer composition profile can be constructed. We test remote homology detection at the topology and fold superfamily levels using a comprehensive set of fold homologs, culled from the CATH database that share low pairwise sequence similarity. Using the receiver-operating characteristic measure, we demonstrate potentially significant improvement in using information-optimized reduced tetramer composition, over methods relying only on the raw amino acid composition or on traditional sequence alignment, in homology detection at or below the "twilight zone". PMID:20635424

  2. Folded Compact Range Development and Coherent Change Detection Measurement Project

    Sorensen, K.W.

    1995-03-01

    A novel, folded compact range configuration has been developed at the Sandia National Laboratories compact range antenna and radar cross section measurement facility, operated by the Radar/Antenna Department 2343, as a means of performing indoor, environmentally-controlled, far-field simulations of synthetic aperture radar (SAR) coherent change detection (CCD) measurements. This report describes the development of the folded compact range configuration, as well as the initial set of coherent change detection measurements made with the system. These measurements have been highly successful, and have demonstrated the viability of the folded compact range concept in simulating SAR CCD measurements. It is felt that follow-on measurements have the potential of contributing significantly to the body of knowledge available to the scientific community involved in CCD image generation and processing, and that this tool will be a significant aid in the research and development of change detection methodologies.

  3. Hyaluronic acid hydrogels for vocal fold wound healing

    Gaston, Joel; Thibeault, Susan L.

    2013-01-01

    The unique vibrational properties inherent to the human vocal fold have a significant detrimental impact on wound healing and scar formation. Hydrogels have taken prominence as a tissue engineered strategy to restore normal vocal structure and function as cellularity is low. The frequent vibrational and shear forces applied to, and present in this connective tissue make mechanical properties of such hydrogels a priority in this active area of research. Hyaluronic acid has been chemically modi...

  4. Nucleic acid detection kits

    Hall, Jeff G.; Lyamichev, Victor I.; Mast, Andrea L.; Brow, Mary Ann; Kwiatkowski, Robert W.; Vavra, Stephanie H.

    2005-03-29

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of nucleic acid from various viruses in a sample.

  5. Nucleic acid detection compositions

    Prudent, James R. (Madison, WI); Hall, Jeff G. (Madison, WI); Lyamichev, Victor I. (Madison, WI); Brow, Mary Ann (Madison, WI); Dahlberg, James L. (Madison, WI)

    2008-08-05

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  6. Nucleic acid detection assays

    Prudent, James R.; Hall, Jeff G.; Lyamichev, Victor I.; Brow, Mary Ann; Dahlberg, James E.

    2005-04-05

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof.

  7. Building multiclass classifiers for remote homology detection and fold recognition

    Karypis George

    2006-10-01

    Full Text Available Abstract Background Protein remote homology detection and fold recognition are central problems in computational biology. Supervised learning algorithms based on support vector machines are currently one of the most effective methods for solving these problems. These methods are primarily used to solve binary classification problems and they have not been extensively used to solve the more general multiclass remote homology prediction and fold recognition problems. Results We present a comprehensive evaluation of a number of methods for building SVM-based multiclass classification schemes in the context of the SCOP protein classification. These methods include schemes that directly build an SVM-based multiclass model, schemes that employ a second-level learning approach to combine the predictions generated by a set of binary SVM-based classifiers, and schemes that build and combine binary classifiers for various levels of the SCOP hierarchy beyond those defining the target classes. Conclusion Analyzing the performance achieved by the different approaches on four different datasets we show that most of the proposed multiclass SVM-based classification approaches are quite effective in solving the remote homology prediction and fold recognition problems and that the schemes that use predictions from binary models constructed for ancestral categories within the SCOP hierarchy tend to not only lead to lower error rates but also reduce the number of errors in which a superfamily is assigned to an entirely different fold and a fold is predicted as being from a different SCOP class. Our results also show that the limited size of the training data makes it hard to learn complex second-level models, and that models of moderate complexity lead to consistently better results.

  8. Direct observation of transition paths during the folding of proteins and nucleic acids.

    Neupane, Krishna; Foster, Daniel A N; Dee, Derek R; Yu, Hao; Wang, Feng; Woodside, Michael T

    2016-04-01

    Transition paths, the fleeting trajectories through the transition states that dominate the dynamics of biomolecular folding reactions, encapsulate the critical information about how structure forms. Owing to their brief duration, however, they have not previously been observed directly. We measured transition paths for both nucleic acid and protein folding, using optical tweezers to observe the microscopic diffusive motion of single molecules traversing energy barriers. The average transit times and the shapes of the transit-time distributions agreed well with theoretical expectations for motion over the one-dimensional energy landscapes reconstructed for the same molecules, validating the physical theory of folding reactions. These measurements provide a first look at the critical microscopic events that occur during folding, opening exciting new avenues for investigating folding phenomena. PMID:27124461

  9. Dependence of α-helical and β-sheet amino acid propensities on the overall protein fold type

    Fujiwara Kazuo

    2012-08-01

    Full Text Available Abstract Background A large number of studies have been carried out to obtain amino acid propensities for α-helices and β-sheets. The obtained propensities for α-helices are consistent with each other, and the pair-wise correlation coefficient is frequently high. On the other hand, the β-sheet propensities obtained by several studies differed significantly, indicating that the context significantly affects β-sheet propensity. Results We calculated amino acid propensities for α-helices and β-sheets for 39 and 24 protein folds, respectively, and addressed whether they correlate with the fold. The propensities were also calculated for exposed and buried sites, respectively. Results showed that α-helix propensities do not differ significantly by fold, but β-sheet propensities are diverse and depend on the fold. The propensities calculated for exposed sites and buried sites are similar for α-helix, but such is not the case for the β-sheet propensities. We also found some fold dependence on amino acid frequency in β-strands. Folds with a high Ser, Thr and Asn content at exposed sites in β-strands tend to have a low Leu, Ile, Glu, Lys and Arg content (correlation coefficient = −0.90 and to have flat β-sheets. At buried sites in β-strands, the content of Tyr, Trp, Gln and Ser correlates negatively with the content of Val, Ile and Leu (correlation coefficient = −0.93. "All-β" proteins tend to have a higher content of Tyr, Trp, Gln and Ser, whereas "α/β" proteins tend to have a higher content of Val, Ile and Leu. Conclusions The α-helix propensities are similar for all folds and for exposed and buried residues. However, β-sheet propensities calculated for exposed residues differ from those for buried residues, indicating that the exposed-residue fraction is one of the major factors governing amino acid composition in β-strands. Furthermore, the correlations we detected suggest that amino acid composition is related to folding

  10. Accuracy of sequence alignment and fold assessment using reduced amino acid alphabets.

    Melo, Francisco; Marti-Renom, Marc A

    2006-06-01

    Reduced or simplified amino acid alphabets group the 20 naturally occurring amino acids into a smaller number of representative protein residues. To date, several reduced amino acid alphabets have been proposed, which have been derived and optimized by a variety of methods. The resulting reduced amino acid alphabets have been applied to pattern recognition, generation of consensus sequences from multiple alignments, protein folding, and protein structure prediction. In this work, amino acid substitution matrices and statistical potentials were derived based on several reduced amino acid alphabets and their performance assessed in a large benchmark for the tasks of sequence alignment and fold assessment of protein structure models, using as a reference frame the standard alphabet of 20 amino acids. The results showed that a large reduction in the total number of residue types does not necessarily translate into a significant loss of discriminative power for sequence alignment and fold assessment. Therefore, some definitions of a few residue types are able to encode most of the relevant sequence/structure information that is present in the 20 standard amino acids. Based on these results, we suggest that the use of reduced amino acid alphabets may allow to increasing the accuracy of current substitution matrices and statistical potentials for the prediction of protein structure of remote homologs. PMID:16506243

  11. Improving the visualization and detection of tissue folds in whole slide images through color enhancement

    Pinky A Bautista

    2010-01-01

    Full Text Available Objective : The objective of this paper is to improve the visualization and detection of tissue folds, which are prominent among tissue slides, from the pre-scan image of a whole slide image by introducing a color enhancement method that enables the differentiation between fold and non-fold image pixels. Method: The weighted difference between the color saturation and luminance of the image pixels is used as shifting factor to the original RGB color of the image. Results: Application of the enhancement method to hematoxylin and eosin (H&E stained images improves the visualization of tissue folds regardless of the colorimetric variations in the images. Detection of tissue folds after application of the enhancement also improves but the presence of nuclei, which are also stained dark like the folds, was found to sometimes affect the detection accuracy. Conclusion: The presence of tissue artifacts could affect the quality of whole slide images, especially that whole slide scanners select the focus points from the pre-scan image wherein the artifacts are indistinguishable from real tissue area. We have a presented in this paper an enhancement scheme that improves the visualization and detection of tissue folds from pre-scan images. Since the method works on the simulated pre-scan images its integration to the actual whole slide imaging process should also be possible.

  12. Nucleic acid chaperons: a theory of an RNA-assisted protein folding

    Biro Jan C

    2005-09-01

    Full Text Available Summary Background Proteins are assumed to contain all the information necessary for unambiguous folding (Anfinsen's principle. However, ab initio structure prediction is often not successful because the amino acid sequence itself is not sufficient to guide between endless folding possibilities. It seems to be a logical to try to find the "missing" information in nucleic acids, in the redundant codon base. Results mRNA energy dot plots and protein residue contact maps were found to be rather similar. The structure of mRNA is also conserved if the protein structure is conserved, even if the sequence similarity is low. These observations led me to suppose that some similarity might exist between nucleic acid and protein folding. I found that amino acid pairs, which are co-located in the protein structure, are preferentially coded by complementary codons. This codon complementarity is not perfect; it is suboptimal where the 1st and 3rd codon residues are complementary to each other in reverse orientation, while the 2nd codon letters may be, but are not necessarily, complementary. Conclusion Partial complementary coding of co-locating amino acids in protein structures suggests that mRNA assists in protein folding and functions not only as a template but even as a chaperon during translation. This function explains the role of wobble bases and answers the mystery of why we have a redundant codon base.

  13. Use of a structural alphabet to find compatible folds for amino acid sequences.

    Mahajan, Swapnil; de Brevern, Alexandre G; Sanejouand, Yves-Henri; Srinivasan, Narayanaswamy; Offmann, Bernard

    2015-01-01

    The structural annotation of proteins with no detectable homologs of known 3D structure identified using sequence-search methods is a major challenge today. We propose an original method that computes the conditional probabilities for the amino-acid sequence of a protein to fit to known protein 3D structures using a structural alphabet, known as "Protein Blocks" (PBs). PBs constitute a library of 16 local structural prototypes that approximate every part of protein backbone structures. It is used to encode 3D protein structures into 1D PB sequences and to capture sequence to structure relationships. Our method relies on amino acid occurrence matrices, one for each PB, to score global and local threading of query amino acid sequences to protein folds encoded into PB sequences. It does not use any information from residue contacts or sequence-search methods or explicit incorporation of hydrophobic effect. The performance of the method was assessed with independent test datasets derived from SCOP 1.75A. With a Z-score cutoff that achieved 95% specificity (i.e., less than 5% false positives), global and local threading showed sensitivity of 64.1% and 34.2%, respectively. We further tested its performance on 57 difficult CASP10 targets that had no known homologs in PDB: 38 compatible templates were identified by our approach and 66% of these hits yielded correctly predicted structures. This method scales-up well and offers promising perspectives for structural annotations at genomic level. It has been implemented in the form of a web-server that is freely available at http://www.bo-protscience.fr/forsa. PMID:25297700

  14. Two methods of Haustral fold detection from computed tomographic virtual colonoscopy images

    Chowdhury, Ananda S.; Tan, Sovira; Yao, Jianhua; Linguraru, Marius G.; Summers, Ronald M.

    2009-02-01

    Virtual colonoscopy (VC) has gained popularity as a new colon diagnostic method over the last decade. VC is a new, less invasive alternative to the usually practiced optical colonoscopy for colorectal polyp and cancer screening, the second major cause of cancer related deaths in industrial nations. Haustral (colonic) folds serve as important landmarks for virtual endoscopic navigation in the existing computer-aided-diagnosis (CAD) system. In this paper, we propose and compare two different methods of haustral fold detection from volumetric computed tomographic virtual colonoscopy images. The colon lumen is segmented from the input using modified region growing and fuzzy connectedness. The first method for fold detection uses a level set that evolves on a mesh representation of the colon surface. The colon surface is obtained from the segmented colon lumen using the Marching Cubes algorithm. The second method for fold detection, based on a combination of heat diffusion and fuzzy c-means algorithm, is employed on the segmented colon volume. Folds obtained on the colon volume using this method are then transferred to the corresponding colon surface. After experimentation with different datasets, results are found to be promising. The results also demonstrate that the first method has a tendency of slight under-segmentation while the second method tends to slightly over-segment the folds.

  15. Structural analysis of Bacillus pumilus phenolic acid decarboxylase, a lipocalin-fold enzyme

    The crystal structure of phenolic acid decarboxylase from B. pumilus strain UI-670 has been determined and refined at 1.69 Å resolution. The enzyme is a dimer, with each subunit adopting a β-barrel structure belonging to the lipocalin fold. The decarboxylation of phenolic acids, including ferulic and p-coumaric acids, to their corresponding vinyl derivatives is of importance in the flavouring and polymer industries. Here, the crystal structure of phenolic acid decarboxylase (PAD) from Bacillus pumilus strain UI-670 is reported. The enzyme is a 161-residue polypeptide that forms dimers both in the crystal and in solution. The structure of PAD as determined by X-ray crystallography revealed a β-barrel structure and two α-helices, with a cleft formed at one edge of the barrel. The PAD structure resembles those of the lipocalin-fold proteins, which often bind hydrophobic ligands. Superposition of structurally related proteins bound to their cognate ligands shows that they and PAD bind their ligands in a conserved location within the β-barrel. Analysis of the residue-conservation pattern for PAD-related sequences mapped onto the PAD structure reveals that the conservation mainly includes residues found within the hydrophobic core of the protein, defining a common lipocalin-like fold for this enzyme family. A narrow cleft containing several conserved amino acids was observed as a structural feature and a potential ligand-binding site

  16. Amino acid alphabet reduction preserves fold information contained in contact interactions in proteins.

    Solis, Armando D

    2015-12-01

    To reduce complexity, understand generalized rules of protein folding, and facilitate de novo protein design, the 20-letter amino acid alphabet is commonly reduced to a smaller alphabet by clustering amino acids based on some measure of similarity. In this work, we seek the optimal alphabet that preserves as much of the structural information found in long-range (contact) interactions among amino acids in natively-folded proteins. We employ the Information Maximization Device, based on information theory, to partition the amino acids into well-defined clusters. Numbering from 2 to 19 groups, these optimal clusters of amino acids, while generated automatically, embody well-known properties of amino acids such as hydrophobicity/polarity, charge, size, and aromaticity, and are demonstrated to maintain the discriminative power of long-range interactions with minimal loss of mutual information. Our measurements suggest that reduced alphabets (of less than 10) are able to capture virtually all of the information residing in native contacts and may be sufficient for fold recognition, as demonstrated by extensive threading tests. In an expansive survey of the literature, we observe that alphabets derived from various approaches-including those derived from physicochemical intuition, local structure considerations, and sequence alignments of remote homologs-fare consistently well in preserving contact interaction information, highlighting a convergence in the various factors thought to be relevant to the folding code. Moreover, we find that alphabets commonly used in experimental protein design are nearly optimal and are largely coherent with observations that have arisen in this work. PMID:26407535

  17. Efficient fold-change detection based on protein-protein interactions

    Buijsman, Wouter

    2012-01-01

    Various biological sensory systems exhibit a response to the relative change of the stimulus, often reffered to as fold-change detection. Here, we present a mechanism consisting of two interacting proteins, able to detect a fold-change effectively. This mechanism, in contrast to other proposed mechanisms, does not consume chemical energy and is not subject to transcriptional and translational noise. We show by analytical and numerical calculations that the mechanism can have a fast, precise and efficient response for parameters that are relevant to eukaryotic cells.

  18. Nasal Alar Necrosis Following Hyaluronic Acid Injection into Nasolabial Folds: A Case Report

    Manafi, Ali; Barikbin, Behrooz; Manafi, Amir; Hamedi, Zahra Sadat; Ahmadi Moghadam, Shokoofeh

    2015-01-01

    Injection of synthetic fillers for soft tissue augmentation is increasing over the last decade. One of the most common materials used is hyaluronic acid (HA) that is safe and temporary filler for soft tissue augmentation. We present a case of 54-year-old female who experienced vascular occlusion and nasal alar necrosis following HA injection to the nasolabial folds. She suffered from pain, necrosis, infection, and alar loss that finally required a reconstructive surgery for cosmetic appearanc...

  19. Detecting Selection on Protein Stability through Statistical Mechanical Models of Folding and Evolution

    Ugo Bastolla

    2014-03-01

    Full Text Available The properties of biomolecules depend both on physics and on the evolutionary process that formed them. These two points of view produce a powerful synergism. Physics sets the stage and the constraints that molecular evolution has to obey, and evolutionary theory helps in rationalizing the physical properties of biomolecules, including protein folding thermodynamics. To complete the parallelism, protein thermodynamics is founded on the statistical mechanics in the space of protein structures, and molecular evolution can be viewed as statistical mechanics in the space of protein sequences. In this review, we will integrate both points of view, applying them to detecting selection on the stability of the folded state of proteins. We will start discussing positive design, which strengthens the stability of the folded against the unfolded state of proteins. Positive design justifies why statistical potentials for protein folding can be obtained from the frequencies of structural motifs. Stability against unfolding is easier to achieve for longer proteins. On the contrary, negative design, which consists in destabilizing frequently formed misfolded conformations, is more difficult to achieve for longer proteins. The folding rate can be enhanced by strengthening short-range native interactions, but this requirement contrasts with negative design, and evolution has to trade-off between them. Finally, selection can accelerate functional movements by favoring low frequency normal modes of the dynamics of the native state that strongly correlate with the functional conformation change.

  20. Injectable hyaluronic acid-dextran hydrogels and effects of implantation in ferret vocal fold.

    Luo, Ying; Kobler, James B; Heaton, James T; Jia, Xinqiao; Zeitels, Steven M; Langer, Robert

    2010-05-01

    Injectable hydrogels may potentially be used for augmentation/regeneration of the lamina propria of vocal fold tissue. In this study, hyaluronic acid (HA) and dextran were chemically modified and subsequently crosslinked via formation of hydrazone bonds in phosphate buffer. Swelling ratios, degradation, and compressive moduli of the resulting hydrogels were investigated. It was found that the properties of HA-dextran hydrogels were variable and the trend of variation could be correlated with the hydrogel composition. The biocompatibility of three injectable HA-dextran hydrogels with different crosslinking density was assessed in the vocal fold region using a ferret model. It was found that HA-dextran hydrogels implanted for three weeks stimulated mild foreign-body reactions. Distinct tissue-material interactions were also observed for hydrogels made from different formulations: the hydrogel with the lowest crosslinking density was completely degraded in vivo; while material residues were visible for other types of hydrogel injections, with or without cell penetration into the implantation depending on the hydrogel composition. The in vivo results suggest that the HA-dextran hydrogel matrices can be further developed for applications of vocal fold tissue restoration. PMID:20151459

  1. Evolution of the deaminase fold and multiple origins of eukaryotic editing and mutagenic nucleic acid deaminases from bacterial toxin systems

    Iyer, Lakshminarayan M; Zhang, Dapeng; Rogozin, Igor B.; Aravind, L.

    2011-01-01

    The deaminase-like fold includes, in addition to nucleic acid/nucleotide deaminases, several catalytic domains such as the JAB domain, and others involved in nucleotide and ADP-ribose metabolism. Using sensitive sequence and structural comparison methods, we develop a comprehensive natural classification of the deaminase-like fold and show that its ancestral version was likely to operate on nucleotides or nucleic acids. Consequently, we present evidence that a specific group of JAB domains ar...

  2. Improving protein fold recognition and structural class prediction accuracies using physicochemical properties of amino acids.

    Raicar, Gaurav; Saini, Harsh; Dehzangi, Abdollah; Lal, Sunil; Sharma, Alok

    2016-08-01

    Predicting the three-dimensional (3-D) structure of a protein is an important task in the field of bioinformatics and biological sciences. However, directly predicting the 3-D structure from the primary structure is hard to achieve. Therefore, predicting the fold or structural class of a protein sequence is generally used as an intermediate step in determining the protein's 3-D structure. For protein fold recognition (PFR) and structural class prediction (SCP), two steps are required - feature extraction step and classification step. Feature extraction techniques generally utilize syntactical-based information, evolutionary-based information and physicochemical-based information to extract features. In this study, we explore the importance of utilizing the physicochemical properties of amino acids for improving PFR and SCP accuracies. For this, we propose a Forward Consecutive Search (FCS) scheme which aims to strategically select physicochemical attributes that will supplement the existing feature extraction techniques for PFR and SCP. An exhaustive search is conducted on all the existing 544 physicochemical attributes using the proposed FCS scheme and a subset of physicochemical attributes is identified. Features extracted from these selected attributes are then combined with existing syntactical-based and evolutionary-based features, to show an improvement in the recognition and prediction performance on benchmark datasets. PMID:27164998

  3. Simultaneous detection of ascorbic acid, uric acid and homovanillic acid at copper modified electrode

    Selvaraju, T. [Centre for Photoelectrochemistry, School of Chemistry, Madurai Kamaraj University, Madurai 625021 (India); Ramaraj, R. [Centre for Photoelectrochemistry, School of Chemistry, Madurai Kamaraj University, Madurai 625021 (India)]. E-mail: ramarajr@yahoo.com

    2007-02-15

    The copper was deposited on glassy carbon (GC) and indium tin oxide (ITO) electrodes by electrochemical method. The copper structures on electrode were characterized by atomic force microscope, X-ray diffractometeric pattern and differential pulse voltammetric studies. Optimal conditions for uniform growth of copper structures on the electrode were established. Voltammetric sensor was fabricated using the copper deposited GC electrode for the simultaneous detection and determination of uric acid (UA) and homovanillic acid (HVA) in the presence of excess concentrations of ascorbic acid (AA). The voltammetric signals due to AA and UA oxidation were well separated with a potential difference of 400 mV and AA did not interfere with the measurement of UA and HVA at the GC/Cu electrode. Linear calibration curves were obtained in the concentration range 1-40 {mu}M for AA and 20-50 {mu}M for UA at physiological pH and a detection limit of 10 nM of UA in the presence of 10-fold excess concentrations of AA was achieved. The simultaneous detection of submicromolar concentrations of AA, UA and HVA was achieved at the GC/Cu electrode. The practical utility of the present GC/Cu modified electrode was demonstrated by measuring the AA content in Vitamin C tablet, UA content in human urine and blood serum samples with satisfactory results.

  4. Nucleic acid detection system and method for detecting influenza

    Cai, Hong; Song, Jian

    2015-03-17

    The invention provides a rapid, sensitive and specific nucleic acid detection system which utilizes isothermal nucleic acid amplification in combination with a lateral flow chromatographic device, or DNA dipstick, for DNA-hybridization detection. The system of the invention requires no complex instrumentation or electronic hardware, and provides a low cost nucleic acid detection system suitable for highly sensitive pathogen detection. Hybridization to single-stranded DNA amplification products using the system of the invention provides a sensitive and specific means by which assays can be multiplexed for the detection of multiple target sequences.

  5. Detection of Periodicity Based on Serial Dependence of Phase-Folded Data

    Zucker, Shay

    2015-01-01

    We introduce and test several novel approaches for periodicity detection in unevenly-spaced sparse datasets. Specifically, we examine five different kinds of periodicity metrics, which are based on non-parametric measures of serial dependence of the phase-folded data. We test the metrics through simulations in which we assess their performance in various situations, including various periodic signal shapes, different numbers of data points and different signal to noise ratios. One of the periodicity metrics we introduce seems to perform significantly better than the classical ones in some settings of interest to astronomers. We suggest that this periodicity metric - the Hoeffding-test periodicity metric - should be used in addition to the traditional methods, to increase periodicity detection probability.

  6. Single-molecule detection of folding and unfolding of the G-quadruplex aptamer in a nanopore nanocavity.

    Shim, Ji Wook; Tan, Qiulin; Gu, Li-Qun

    2009-02-01

    Guanine-rich nucleic acids can form G-quadruplexes that are important in gene regulation, biosensor design and nano-structure construction. In this article, we report on the development of a nanopore encapsulating single-molecule method for exploring how cations regulate the folding and unfolding of the G-quadruplex formed by the thrombin-binding aptamer (TBA, GGTTGGTGTGGTTGG). The signature blocks in the nanopore revealed that the G-quadruplex formation is cation-selective. The selectivity sequence is K(+) > NH(4)(+) approximately Ba(2+) > Cs(+) approximately Na(+) > Li(+), and G-quadruplex was not detected in Mg(2+) and Ca(2+). Ba(2+) can form a long-lived G-quadruplex with TBA. However, the capability is affected by the cation-DNA interaction. The cation-selective formation of the G-quadruplex is correlated with the G-quadruplex volume, which varies with cation species. The high formation capability of the K(+)-induced G-quadruplex is contributed largely by the slow unfolding reaction. Although the Na(+)- and Li(+)-quadruplexes feature similar equilibrium properties, they undergo radically different pathways. The Na(+)-quadruplex folds and unfolds most rapidly, while the Li(+)-quadruplex performs both reactions at the slowest rates. Understanding these ion-regulated properties of oligonucleotides is beneficial for constructing fine-tuned biosensors and nano-structures. The methodology in this work can be used for studying other quadruplexes and protein-aptamer interactions. PMID:19112078

  7. G-rich VEGF aptamer with locked and unlocked nucleic acid modifications exhibits a unique G-quadruplex fold

    Marusic, Maja; Veedu, Rakesh N; Wengel, Jesper;

    2013-01-01

    . Locked residues contribute to thermal stabilization of the adopted structure and formation of structurally pre-organized intermediates that facilitate folding into a single G-quadruplex. Understanding the impact of chemical modifications on folding, thermal stability and structural polymorphism of G......-quadruplexes provides means for the improvement of vascular endothelial growth factor aptamers and advances our insights into driving nucleic acid structure by locking or unlocking the conformation of sugar moieties of nucleotides in general....

  8. Effect of sodium dodecyl sulfate on folding and thermal stability of acid-denatured cytochrome c: A spectroscopic approach

    Xu, Qi; Keiderling, Timothy A

    2004-01-01

    The molten globule (MG) state can be an intermediate in the protein folding pathway; thus, its detailed description can help understanding protein folding. Sodium dodecyl sulfate (SDS), an anionic surfactant that is commonly used to mimic hydrophobic binding environments such as cell membranes, is known to denature some native state proteins, including horse cytochrome c (cyt c). In this article, refolding of acid denatured cyt c is studied under the influence of SDS to form MG-like states at...

  9. Stochastic adaptation and fold-change detection: from single-cell to population behavior

    Leier André

    2011-02-01

    Full Text Available Abstract Background In cell signaling terminology, adaptation refers to a system's capability of returning to its equilibrium upon a transient response. To achieve this, a network has to be both sensitive and precise. Namely, the system must display a significant output response upon stimulation, and later on return to pre-stimulation levels. If the system settles at the exact same equilibrium, adaptation is said to be 'perfect'. Examples of adaptation mechanisms include temperature regulation, calcium regulation and bacterial chemotaxis. Results We present models of the simplest adaptation architecture, a two-state protein system, in a stochastic setting. Furthermore, we consider differences between individual and collective adaptive behavior, and show how our system displays fold-change detection properties. Our analysis and simulations highlight why adaptation needs to be understood in terms of probability, and not in strict numbers of molecules. Most importantly, selection of appropriate parameters in this simple linear setting may yield populations of cells displaying adaptation, while single cells do not. Conclusions Single cell behavior cannot be inferred from population measurements and, sometimes, collective behavior cannot be determined from the individuals. By consequence, adaptation can many times be considered a purely emergent property of the collective system. This is a clear example where biological ergodicity cannot be assumed, just as is also the case when cell replication rates are not homogeneous, or depend on the cell state. Our analysis shows, for the first time, how ergodicity cannot be taken for granted in simple linear examples either. The latter holds even when cells are considered isolated and devoid of replication capabilities (cell-cycle arrested. We also show how a simple linear adaptation scheme displays fold-change detection properties, and how rupture of ergodicity prevails in scenarios where transitions between

  10. Cooperative folding near the downhill limit determined with amino acid resolution by hydrogen exchange.

    Yu, Wookyung; Baxa, Michael C; Gagnon, Isabelle; Freed, Karl F; Sosnick, Tobin R

    2016-04-26

    The relationship between folding cooperativity and downhill, or barrier-free, folding of proteins under highly stabilizing conditions remains an unresolved topic, especially for proteins such as λ-repressor that fold on the microsecond timescale. Under aqueous conditions where downhill folding is most likely to occur, we measure the stability of multiple H bonds, using hydrogen exchange (HX) in a λYA variant that is suggested to be an incipient downhill folder having an extrapolated folding rate constant of 2 × 10(5) s(-1) and a stability of 7.4 kcal·mol(-1) at 298 K. At least one H bond on each of the three largest helices (α1, α3, and α4) breaks during a common unfolding event that reflects global denaturation. The use of HX enables us to both examine folding under highly stabilizing, native-like conditions and probe the pretransition state region for stable species without the need to initiate the folding reaction. The equivalence of the stability determined at zero and high denaturant indicates that any residual denatured state structure minimally affects the stability even under native conditions. Using our ψ analysis method along with mutational ϕ analysis, we find that the three aforementioned helices are all present in the folding transition state. Hence, the free energy surface has a sufficiently high barrier separating the denatured and native states that folding appears cooperative even under extremely stable and fast folding conditions. PMID:27078098

  11. Logarithmic and power law input-output relations in sensory systems with fold-change detection.

    Adler, Miri; Mayo, Avi; Alon, Uri

    2014-08-01

    Two central biophysical laws describe sensory responses to input signals. One is a logarithmic relationship between input and output, and the other is a power law relationship. These laws are sometimes called the Weber-Fechner law and the Stevens power law, respectively. The two laws are found in a wide variety of human sensory systems including hearing, vision, taste, and weight perception; they also occur in the responses of cells to stimuli. However the mechanistic origin of these laws is not fully understood. To address this, we consider a class of biological circuits exhibiting a property called fold-change detection (FCD). In these circuits the response dynamics depend only on the relative change in input signal and not its absolute level, a property which applies to many physiological and cellular sensory systems. We show analytically that by changing a single parameter in the FCD circuits, both logarithmic and power-law relationships emerge; these laws are modified versions of the Weber-Fechner and Stevens laws. The parameter that determines which law is found is the steepness (effective Hill coefficient) of the effect of the internal variable on the output. This finding applies to major circuit architectures found in biological systems, including the incoherent feed-forward loop and nonlinear integral feedback loops. Therefore, if one measures the response to different fold changes in input signal and observes a logarithmic or power law, the present theory can be used to rule out certain FCD mechanisms, and to predict their cooperativity parameter. We demonstrate this approach using data from eukaryotic chemotaxis signaling. PMID:25121598

  12. Logarithmic and power law input-output relations in sensory systems with fold-change detection.

    Miri Adler

    2014-08-01

    Full Text Available Two central biophysical laws describe sensory responses to input signals. One is a logarithmic relationship between input and output, and the other is a power law relationship. These laws are sometimes called the Weber-Fechner law and the Stevens power law, respectively. The two laws are found in a wide variety of human sensory systems including hearing, vision, taste, and weight perception; they also occur in the responses of cells to stimuli. However the mechanistic origin of these laws is not fully understood. To address this, we consider a class of biological circuits exhibiting a property called fold-change detection (FCD. In these circuits the response dynamics depend only on the relative change in input signal and not its absolute level, a property which applies to many physiological and cellular sensory systems. We show analytically that by changing a single parameter in the FCD circuits, both logarithmic and power-law relationships emerge; these laws are modified versions of the Weber-Fechner and Stevens laws. The parameter that determines which law is found is the steepness (effective Hill coefficient of the effect of the internal variable on the output. This finding applies to major circuit architectures found in biological systems, including the incoherent feed-forward loop and nonlinear integral feedback loops. Therefore, if one measures the response to different fold changes in input signal and observes a logarithmic or power law, the present theory can be used to rule out certain FCD mechanisms, and to predict their cooperativity parameter. We demonstrate this approach using data from eukaryotic chemotaxis signaling.

  13. Transient dynamic phenotypes as criteria for model discrimination: fold-change detection in Rhodobacter sphaeroides chemotaxis.

    Hamadeh, Abdullah; Ingalls, Brian; Sontag, Eduardo

    2013-03-01

    The chemotaxis pathway of the bacterium Rhodobacter sphaeroides shares many similarities with that of Escherichia coli. It exhibits robust adaptation and has several homologues of the latter's chemotaxis proteins. Recent theoretical results have correctly predicted that the E. coli output behaviour is unchanged under scaling of its ligand input signal; this property is known as fold-change detection (FCD). In the light of recent experimental results suggesting that R. sphaeroides may also show FCD, we present theoretical assumptions on the R. sphaeroides chemosensory dynamics that can be shown to yield FCD behaviour. Furthermore, it is shown that these assumptions make FCD a property of this system that is robust to structural and parametric variations in the chemotaxis pathway, in agreement with experimental results. We construct and examine models of the full chemotaxis pathway that satisfy these assumptions and reproduce experimental time-series data from earlier studies. We then propose experiments in which models satisfying our theoretical assumptions predict robust FCD behaviour where earlier models do not. In this way, we illustrate how transient dynamic phenotypes such as FCD can be used for the purposes of discriminating between models that reproduce the same experimental time-series data. PMID:23293140

  14. Isolation, folding and structural investigations of the amino acid transporter OEP16

    Ni, Da Qun; Zook, James; Klewer, Douglas A.; Nieman, Ronald A.; Soll, J.; Fromme, Petra

    2011-12-01

    Membrane proteins compose more than 30% of all proteins in the living cell. However, many membrane proteins have low abundance in the cell and cannot be isolated from natural sources in concentrations suitable for structure analysis. The overexpression, reconstitution, and stabilization of membrane proteins are complex and remain a formidable challenge in membrane protein characterization. Here we describe a novel, in vitro folding procedure for a cation-selective channel protein, the outer envelope membrane protein 16 (OEP16) of pea chloroplast, overexpressed in Escherichia coli in the form of inclusion bodies. The protein is purified and then folded with detergent on a Ni-NTA affinity column. Final concentrations of reconstituted OEP16 of up to 24 mg/ml have been achieved, which provides samples that are sufficient for structural studies by NMR and crystallography. Reconstitution of OEP16 in detergent micelles was monitored by circular dichroism, fluorescence, and NMR spectroscopy. Tryptophan fluorescence spectra of heterologous expressed OEP16 in micelles are similar to spectra of functionally active OEP16 in liposomes, which indicates folding of the membrane protein in detergent micelles. CD spectroscopy studies demonstrate a folded protein consisting primarily of a-helices. 15N-HSQC NMR spectra also provide evidence for a folded protein. We present here a convenient, effective and quantitative method to screen large numbers of conditions for optimal protein stability by using microdialysis chambers in combination with fluorescence spectroscopy. Recent collection of multidimensional NMR data at 500, 600 and 800 MHz demonstrated that the protein is suitable for structure determination by NMR and stable for weeks during data collection.

  15. Swept-Source Optical Coherence Tomography Detecting Intraoperative Acute Descemet's Fold Formation

    Yu Ichioka

    2016-07-01

    Full Text Available Background: To present an intraoperative acute Descemet’s fold formation using swept-source optical coherence tomography (SS-OCT imaging. Case Report: A 67-year-old man complaining of reduced visual acuity in the left eye. A 25-gauge pars plana vitrectomy combined with phacoemulsification cataract surgery was performed to remove the vitreomacular traction. When hydro-sealing was performed, striae rapidly spread in the cornea. SS-OCT B-scan images performed on postoperative day 1 revealed a wavy Descemet’s membrane that might correspond to Descemet’s folds. Pairs of hypo- and hyperreflective narrow lesions running from the wavy Descemet’s membrane to almost half of the thickness of the whole cornea were observed. En face OCT imaging clearly showed the stromal fold, which continuously spread from the Descemet’s fold. Conclusion: The stromal fold might be due to the focal bulge of the stroma posteriorly caused by the rapid volume increase of the stroma which could push Descemet’s membrane posteriorly, thereby forming a wavy Descemet’s membrane layer.

  16. Single-molecule detection of folding and unfolding of the G-quadruplex aptamer in a nanopore nanocavity

    Shim, Ji Wook; Tan, Qiulin; Gu, Li-Qun

    2008-01-01

    Guanine-rich nucleic acids can form G-quadruplexes that are important in gene regulation, biosensor design and nano-structure construction. In this article, we report on the development of a nanopore encapsulating single-molecule method for exploring how cations regulate the folding and unfolding of the G-quadruplex formed by the thrombin-binding aptamer (TBA, GGTTGGTGTGGTTGG). The signature blocks in the nanopore revealed that the G-quadruplex formation is cation-selective. The selectivity s...

  17. Amino acid empirical contact energy definitions for fold recognition in the space of contact maps

    Fogolari Federico; Molinari Henriette; Berrera Marco

    2003-01-01

    Abstract Background Contradicting evidence has been presented in the literature concerning the effectiveness of empirical contact energies for fold recognition. Empirical contact energies are calculated on the basis of information available from selected protein structures, with respect to a defined reference state, according to the quasi-chemical approximation. Protein-solvent interactions are estimated from residue solvent accessibility. Results In the approach presented here, contact energ...

  18. Phosphatidic acid plays a special role in stabilizing and folding of the tetrameric potassium channel KcsA.

    Raja, Mobeen; Spelbrink, Robin E J; de Kruijff, Ben; Killian, J Antoinette

    2007-12-11

    In this study, we investigated how the presence of anionic lipids influenced the stability and folding properties of the potassium channel KcsA. By using a combination of gel electrophoresis, tryptophan fluorescence and acrylamide quenching experiments, we found that the presence of the anionic lipid phosphatidylglycerol (PG) in a phosphatidylcholine (PC) bilayer slightly stabilized the tetramer and protected it from trifluoroethanol-induced dissociation. Surprisingly, the presence of phosphatidic acid (PA) had a much larger effect on the stability of KcsA and this lipid, in addition, significantly influenced the folding properties of the protein. The data indicate that PA creates some specificity over PG, and that it most likely stabilizes the tetramer via both electrostatic and hydrogen bond interactions. PMID:18036565

  19. Safety and persistence of non-animal stabilized hyaluronic acid fillers for nasolabial folds correction in 30 Indian patients

    Shehnaz Z Arsiwala

    2010-01-01

    Full Text Available Background: Correction of nasolabial creases through minimally invasive procedures is increasingly being sought by patients. Injecting non-animal stabilized hyaluronic acid filler is a highly effective method to achieve an optimal and persistent cosmetic result. Aims: To evaluate the efficacy, persistence and safety of Restylane and Perlane (Q-Med, Sweden for correction of nasolabial folds in Indian patients. Materials and Methods: Thirty Indian patients with mild, moderate and severe nasolabial folds (based on Wrinkle Assessment Scale were recruited in the study after informed consent for correction of their folds with Restylane or Perlane or both. Injections were administered in a single sitting after global assessment of the patient′s face using Wrinkle assessment scale (WAS.Optimal filling was performed by using appropriate techniques and its safety and efficacy assessed independently by the investigator as well as by patients at immediately, 3, 6 and 9 months post-procedure. Any adverse reactions were noted. Results: Twenty two females and 8 males (age range 45-55 years, mean age 52 years were recruited in the study. An optimum cosmetic correction was obtained in all patients. The efficacy increased with time and was greatest at 3 months after the treatment. Grade 2 improvement was maintained at 9 months in mild and moderate folds, and grade 3 improvement for severe folds. Minor post injection side effects like erythema at puncture site, needle marks and bruising were seen. Conclusion: Restylane and Perlane are safe and effective dermal fillers for correction of nasolabial creases and offer immediate effect.

  20. Amplification-free in situ KRAS point mutation detection at 60 copies per mL in urine in a background of 1000-fold wild type.

    Kirimli, Ceyhun E; Shih, Wei-Heng; Shih, Wan Y

    2016-02-21

    We have examined the in situ detection of a single-nucleotide KRAS mutation in urine using a (Pb(Mg1/3Nb2/3)O3)0.65(PbTiO3)0.35 (PMN-PT) piezoelectric plate sensor (PEPS) coated with a 17-nucleotide (nt) locked nucleic acid (LNA) probe DNA complementary to the KRAS mutation. To enhance the in situ mutant (MT) DNA detection specificity against the wild type (WT), detection was carried out in a flow with a flow rate of 4 mL min(-1) and at 63 °C with the PEPS vertically situated at the center of the flow in which both the temperature and the flow impingement force discriminated the wild type. Under such conditions, PEPS was shown to specifically detect KRAS MT in situ with 60 copies per mL analytical sensitivity in a background of clinically-relevant 1000-fold more WT in 30 min without DNA isolation, amplification, or labeling. For validation, this detection was followed with detection in a mixture of blue MT fluorescent reporter microspheres (FRMs) (MT FRMs) that bound to only the captured MT and orange WT FRMs that bound to only the captured WT. Microscopic examinations showed that the captured blue MT FRMs still outnumbered the orange WT FRMs by a factor of 4 to 1 even though WT was 1000-fold of MT in urine. Finally, multiplexed specific mutation detection was demonstrated using a 6-PEPS array each with a probe DNA targeting one of the 6 codon-12 KRAS mutations. PMID:26783561

  1. Amplification-free In Situ KRAS Point Mutation Detection at 60 copies/mL in Urine in a Background of 1000-fold Wild Type

    KirimLi, Ceyhun E.; Shih, Wei-Heng; Shih, Wan Y.

    2016-01-01

    We have examined in situ detection of single-nucleotide KRAS mutation in urine using a (Pb(Mg1/3Nb2/3)O3)0.65(PbTiO3)0.35 (PMN-PT) piezoelectric plate sensor (PEPS) coated with a 17-nucleotide (nt) locked nucleic acid (LNA) probe DNA complementary to the KRAS mutation. To enhance in situ mutant (MT) DNA detection specificity against the wild type (WT), the detection was carried out in a flow with a flow rate of 4 mL/min and at 63°C with the PEPS vertically situated at the center of the flow in which both the temperature and the flow impingement force discriminated the wild type. Under such conditions, PEPS was shown to specifically detect KRAS MT in situ with 60 copies/mL analytical sensitivity in a background of clinically-relevant 1000-fold more WT in 30 min without DNA isolation, amplification, or labeling. For validation, the detection was followed with detection in a mixture of blue MT fluorescent reporter microspheres (FRMs) (MT FRMs) that bound to only the captured MT and orange WT FRMs that bound to only the captured WT. Microscopic examinations showed that the captured blue MT FRMs still outnumbered the orange WT FRMs by a factor of 4 to 1 even though WT was 1000-fold of MT in urine. Finally, multiplexed specific mutation detection was demonstrated using a 6-PEPS array each with a probe DNA targeting one of the 6 codon-12 KRAS mutations. PMID:26783561

  2. Scale-free behaviour of amino acid pair interactions in folded proteins

    Petersen, Steffen B.; Neves-Petersen, Maria Teresa; Mortensen, Rasmus J.;

    2012-01-01

    The protein structure is a cumulative result of interactions between amino acid residues interacting with each other through space and/or chemical bonds. Despite the large number of high resolution protein structures, the ‘‘protein structure code’’ has not been fully identified. Our manuscript...... presents a novel approach to protein structure analysis in order to identify rules for spatial packing of amino acid pairs in proteins. We have investigated 8706 high resolution non-redundant protein chains and quantified amino acid pair interactions in terms of solvent accessibility, spatial and sequence...... are in buried a-helices or b-strands, in a spatial distance of 3.8–4.3A° and in a sequence distance .4 residues. We speculate that the scale free organization of the amino acid pair interactions in the 8D protein structure combined with the clear dominance of pairs of Ala, Ile, Leu and Val is...

  3. Amino acid empirical contact energy definitions for fold recognition in the space of contact maps

    Fogolari Federico

    2003-02-01

    Full Text Available Abstract Background Contradicting evidence has been presented in the literature concerning the effectiveness of empirical contact energies for fold recognition. Empirical contact energies are calculated on the basis of information available from selected protein structures, with respect to a defined reference state, according to the quasi-chemical approximation. Protein-solvent interactions are estimated from residue solvent accessibility. Results In the approach presented here, contact energies are derived from the potential of mean force theory, several definitions of contact are examined and their performance in fold recognition is evaluated on sets of decoy structures. The best definition of contact is tested, on a more realistic scenario, on all predictions including sidechains accepted in the CASP4 experiment. In 30 out of 35 cases the native structure is correctly recognized and best predictions are usually found among the 10 lowest energy predictions. Conclusion The definition of contact based on van der Waals radii of alpha carbon and side chain heavy atoms is seen to perform better than other definitions involving only alpha carbons, only beta carbons, all heavy atoms or only backbone atoms. An important prerequisite for the applicability of the approach is that the protein structure under study should not exhibit anomalous solvent accessibility, compared to soluble proteins whose structure is deposited in the Protein Data Bank. The combined evaluation of a solvent accessibility parameter and contact energy allows for an effective gross screening of predictive models.

  4. An In Vivo Study of Composite Microgels Based on Hyaluronic Acid and Gelatin for the Reconstruction of Surgically Injured Rat Vocal Folds

    Coppoolse, Jiska M. S.; Van Kooten, T. G.; Heris, Hossein K.; Mongeau, Luc; Li, Nicole Y. K.; Thibeault, Susan L.; Pitaro, Jacob; Akinpelu, Olubunm; Daniel, Sam J.

    2014-01-01

    Purpose: The objective of this study was to investigate local injection with a hierarchically microstructured hyaluronic acid-gelatin (HA-Ge) hydrogel for the treatment of acute vocal fold injury using a rat model. Method: Vocal fold stripping was performed unilaterally in 108 Sprague-Dawley rats. A volume of 25 µl saline (placebo controls),…

  5. Prediction of the Occurrence of the ADP-binding βαβ-fold in Proteins, Using an Amino Acid Sequence Fingerprint

    Wierenga, Rik K.; Terpstra, Peter; Hol, Wim G.J.

    1986-01-01

    An amino acid sequence "fingerprint” has been derived that can be used to test if a particular sequence will fold into a βαβ-unit with ADP-binding properties. It was deduced from a careful analysis of the known three-dimensional structures of ADP-binding βαβ-folds. This fingerprint is in fact a set

  6. Device for the detection of acid vapors and particularly hydrofluoric acid vapors

    This device concerns the detection of acid vapors contained in a gaseous environment which have to be controlled. It uses a detector with a calorimetric material. It can be used to detect acid vapors, but it detects particularly hydrofluoric acid vapors. In nuclear industry, this device can detect hydrofluoric acid from UF6, even at high temperature. (TEC)

  7. Electrochemical Sensors for Detection of Acetylsalicylic Acid

    Rene Kizek

    2006-11-01

    Full Text Available Acetylsalicylic acid (AcSA, or aspirin, was introduced in the late 1890s and hasbeen used to treat a variety of inflammatory conditions. The aim of this work was to suggestelectrochemical sensor for acetylsalicylic detection. Primarily, we utilized square wavevoltammetry (SWV using both carbon paste electrode (CPE and of graphite pencilelectrode (GPE as working ones to indirect determination of AcSA. The principle ofindirect determination of AcSA bases in its hydrolysis on salicylic acid (SA, which isconsequently detected. Thus, we optimized both determination of SA and conditions forAcSA hydrolysis and found out that the most suitable frequency, amplitude, step potentialand the composition and pH of the supporting electrolyte for the determination of SA was260 Hz, 50 mV, 10 mV and Britton-Robinson buffer (pH 1.81, respectively. The detectionlimit (S/N = 3 of the SA was 1.3 ng/ml. After that, we aimed on indirect determination ofAcSA by SWV CPE. We tested the influence of pH of Britton-Robinson buffer andtemperature on yield of hydrolysis, and found out that 100% hydrolysis of AcSA wasreached after 80 minutes at pH 1.81 and 90°C. The method for indirect determination ofAcSA has been utilized to analyse pharmaceutical drug. The determined amount of AcSA in the pharmaceutical drug was in good agreement with the declared amounts. Moreover, weused GPE for determination of AcSA in a pharmaceutical drug. Base of the results obtainedfrom stationary electrochemical instrument we used flow injection analysis withelectrochemical detection to determine of salicylates (SA, AcSA, thiosalicylic acid, 3,5-dinitrosalicylic acid and 5-sulfosalicylic acid – SuSA. We found out that we are able todetermine all of detected salicylates directly without any pre-treatment, hydrolysis and so onat units of femtomoles per injection (5 μl.

  8. Distributional fold change test – a statistical approach for detecting differential expression in microarray experiments

    Farztdinov Vadim

    2012-11-01

    Full Text Available Abstract Background Because of the large volume of data and the intrinsic variation of data intensity observed in microarray experiments, different statistical methods have been used to systematically extract biological information and to quantify the associated uncertainty. The simplest method to identify differentially expressed genes is to evaluate the ratio of average intensities in two different conditions and consider all genes that differ by more than an arbitrary cut-off value to be differentially expressed. This filtering approach is not a statistical test and there is no associated value that can indicate the level of confidence in the designation of genes as differentially expressed or not differentially expressed. At the same time the fold change by itself provide valuable information and it is important to find unambiguous ways of using this information in expression data treatment. Results A new method of finding differentially expressed genes, called distributional fold change (DFC test is introduced. The method is based on an analysis of the intensity distribution of all microarray probe sets mapped to a three dimensional feature space composed of average expression level, average difference of gene expression and total variance. The proposed method allows one to rank each feature based on the signal-to-noise ratio and to ascertain for each feature the confidence level and power for being differentially expressed. The performance of the new method was evaluated using the total and partial area under receiver operating curves and tested on 11 data sets from Gene Omnibus Database with independently verified differentially expressed genes and compared with the t-test and shrinkage t-test. Overall the DFC test performed the best – on average it had higher sensitivity and partial AUC and its elevation was most prominent in the low range of differentially expressed features, typical for formalin-fixed paraffin-embedded sample sets

  9. Fabrication and characterization of cross-linkable hydrogel particles based on hyaluronic acid: potential application in vocal fold regeneration.

    Sahiner, Nurettin; Jha, Amit K; Nguyen, David; Jia, Xinqiao

    2008-01-01

    There is a critical need to engineer hyaluronic acid (HA)-based hydrogels with prolonged in vivo residence time, temporal release of therapeutics and matching viscoelasticity for use in vocal fold tissue engineering. We have previously demonstrated the synthesis and characterization of HA-based soft hydrogel particles (HGP) and particle cross-linked networks as injectable materials to treat vocal fold scarring. In this paper, we report a more versatile technique for preparing cross-linkable HA HGP with reduced sizes. HA HGP were synthesized via chemical cross-linking with divinyl sulfone using a sodium bis(2-ethylhexyl) sulfosuccinate (AOT)/isooctane reverse micelle system in the presence of 1-heptanol. These HGP were rendered cross-linkable by introducing aldehyde groups via sodium periodate oxidation (oxHGP). The presence of aldehyde groups was confirmed by multi-photon confocal microscope upon fluorescence staining using cascade blue hydrazide. The aldehyde groups were used as reactive handles for covalent cross-linking with HA that has been previously modified with adipic acid dihydrazide (HADH). The resulting doubly cross-linked networks (DXN) are highly pliable and do not break until approx. 200-300% strain. The measured elastic modulus of the DXN is around 500 Pa, while the dynamic viscosity decreases linearly with frequency in log- log scale. The mechanical characteristics of DXN are similar to that of vocal fold lamina propria. In vitro cell-proliferation assays showed that the cross-linkable HA HGP did not adversely affect the proliferation of the cultured fibroblasts as assessed by MTT assay. A low-molecular-weight model drug, rhodamine 6G (R6G), was loaded into oxHGP, and its release was monitored using UV-Vis spectroscopy. R6G-loaded oxHGP maintained their ability to form DXN when mixed with the HAADH solution. Approximately 84% of entrapped R6G was liberated from oxHGP at a rate of 0.24%/min in the first 6 h. When encapsulated in the DXN, R6G was

  10. Determination of acetylsalicylic acid and salicylic acid in foods, using HPLC with fluorescence detection.

    Venema, D.P.; Hollman, P.C.H.; Janssen, P.L.T.M.K.; Katan, M.B.

    1996-01-01

    We developed a specific and sensitive HPLC method with fluorescence detection for the determination of free acetylsalicylic acid, free salicylic acid, and free salicylic acid plus salicylic acid after alkaline hydrolysis (free-plus-bound) in foods. Acetylsalicylic acid was detected after postcolumn

  11. Acid denaturation of recombinant porcine growth hormone: formation and self-association of folding intermediates.

    Parkinson, E J; Morris, M B; Bastiras, S

    2000-10-10

    We have investigated the conformational changes incurred during the acid-induced unfolding and self-association of recombinant porcine growth hormone (pGH). Acidification (pH 8 to pH 2) of pGH resulted in intrinsic fluorescence, UV absorbance, and near-UV CD transitions centered at pH 4.10. At pH 2.0, a red shift in the fluorescence emission maximum of approximately 3 nm and a 15% loss of the far-UV CD signal at 222 nm imply that the protein did not become extensively unfolded. Acidification in the presence of 4 M urea resulted in similar pH-dependent transitions. However, these occurred at a higher pH (approximately 5.2). At pH 2.0 + 4 M urea, an 8 nm red shift in the fluorescence emission maximum suggests that unfolding was greater than in the absence of urea. The presence of a prominent peak centered at 298 nm in the near-UV CD spectrum, which is absent without urea, signifies further differences in the intermediates generated at pH 2. Sedimentation equilibrium experiments in the analytical ultracentrifuge showed that native pGH and the partially unfolded intermediates reversibly self-associate. Self-association was strongly promoted at pH 2 while urea reduced self-association at both pH 8 and pH 2. These results demonstrate that acidification of pGH in the absence or presence of 4 M urea induced the formation of molten globule-like states with measurable differences in conformation. Similarities and differences in these structural conformations with respect to other growth hormones are discussed. PMID:11015214

  12. Detection of substrate-dependent conformational changes in the P450 fold by nuclear magnetic resonance.

    Colthart, Allison M; Tietz, Drew R; Ni, Yuhua; Friedman, Jessica L; Dang, Marina; Pochapsky, Thomas C

    2016-01-01

    Cytochrome P450 monooxygenases typically catalyze the insertion of one atom of oxygen from O2 into unactivated carbon-hydrogen and carbon-carbon bonds, with concomitant reduction of the other oxygen atom to H2O by NAD(P)H. Comparison of the average structures of the camphor hydroxylase cytochrome P450(cam) (CYP101) obtained from residual dipolar coupling (RDC)-restrained molecular dynamics (MD) in the presence and absence of substrate camphor shows structural displacements resulting from the essential collapse of the active site upon substrate removal. This collapse has conformational consequences that extend across the protein structure, none of which were observed in analogous crystallographic structures. Mutations were made to test the involvement of the observed conformational changes in substrate binding and recognition. All of the mutations performed based upon the NMR-detected perturbations, even those remote from the active site, resulted in modified substrate selectivity, enzyme efficiency and/or haem iron spin state. The results demonstrate that solution NMR can provide insights into enzyme structure-function relationships that are difficult to obtain by other methods. PMID:26911901

  13. Tween 20-stabilized gold nanoparticles combined with adenosine triphosphate-BODIPY conjugates for the fluorescence detection of adenosine with more than 1000-fold selectivity

    Graphical abstract: A simple, enzyme-free, label-free, sensitive and selective system was developed for detecting adenosine based on the use of Tween 20-stabilized gold nanoparticles as an efficient quencher for boron dipyrromethene-conjugated adenosine 5′-triphosphate and as a recognition element for adenosine. - Highlights: • The proposed method can detect adenosine with more than 1000-fold selectivity. • The analysis of adenosine is rapid (∼6 min) using the proposed method. • This method provided better sensitivity for adenosine as compared to aptamer-based sensors. • This method can be applied for the determination of adenosine in urine. - Abstract: This study describes the development of a simple, enzyme-free, label-free, sensitive, and selective system for detecting adenosine based on the use of Tween 20-stabilized gold nanoparticles (Tween 20-AuNPs) as an efficient fluorescence quencher for boron dipyrromethene-conjugated adenosine 5′-triphosphate (BODIPY-ATP) and as a recognition element for adenosine. BODIPY-ATP can interact with Tween 20-AuNPs through the coordination between the adenine group of BODIPY-ATP and Au atoms on the NP surface, thereby causing the fluorescence quenching of BODIPY-ATP through the nanometal surface energy transfer (NSET) effect. When adenosine attaches to the NP surface, the attached adenosine exhibits additional electrostatic attraction to BODIPY-ATP. As a result, the presence of adenosine enhances the efficiency of AuNPs in fluorescence quenching of BODIPY-ATP. The AuNP-induced fluorescence quenching of BODIPY-ATP progressively increased with an increase in the concentration of adenosine; the detection limit at a signal-to-noise ratio of 3 for adenosine was determined to be 60 nM. The selectivity of the proposed system was more than 1000-fold for adenosine over any adenosine analogs and other nucleotides. The proposed system combined with a phenylboronic acid-containing column was successfully applied to the

  14. Detection of nucleic acids by multiple sequential invasive cleavages 02

    Hall, Jeff G. (Madison, WI); Lyamichev, Victor I. (Madison, WI); Mast, Andrea L. (Madison, WI); Brow, Mary Ann D. (Madison, WI)

    2002-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of human cytomegalovirus nucleic acid in a sample.

  15. Detection of nucleic acids by multiple sequential invasive cleavages

    Hall, Jeff G. (Madison, WI); Lyamichev, Victor I. (Madison, WI); Mast, Andrea L. (Madison, WI); Brow, Mary Ann D. (Madison, WI)

    1999-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of human cytomegalovirus nucleic acid in a sample.

  16. Detection of nucleic acids by multiple sequential invasive cleavages

    Hall, Jeff G; Lyamichev, Victor I; Mast, Andrea L; Brow, Mary Ann D

    2012-10-16

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The structure-specific nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based on charge. The present invention also provides methods for the detection of non-target cleavage products via the formation of a complete and activated protein binding region. The invention further provides sensitive and specific methods for the detection of human cytomegalovirus nucleic acid in a sample.

  17. Monitoring Homovanillic Acid and Vanillylmandelic Acid in Human Urine by Capillary Electrophoresis with Electrochemical Detection

    2002-01-01

    A simple, rapid and low-cost method of separation and determination of homovanillic acid and vanillylmandelic acid in human urine was developed based on capillary zone electrophoresis / amperometric detection with high sensitivity and good resolution.

  18. Advances in nucleic acid-based detection methods.

    Wolcott, M J

    1992-01-01

    Laboratory techniques based on nucleic acid methods have increased in popularity over the last decade with clinical microbiologists and other laboratory scientists who are concerned with the diagnosis of infectious agents. This increase in popularity is a result primarily of advances made in nucleic acid amplification and detection techniques. Polymerase chain reaction, the original nucleic acid amplification technique, changed the way many people viewed and used nucleic acid techniques in cl...

  19. Detection of nucleic acid sequences by invader-directed cleavage

    Brow, Mary Ann D. (Madison, WI); Hall, Jeff Steven Grotelueschen (Madison, WI); Lyamichev, Victor (Madison, WI); Olive, David Michael (Madison, WI); Prudent, James Robert (Madison, WI)

    1999-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The 5' nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based by charge.

  20. Detection of nucleic acid sequences by invader-directed cleavage

    Brow, Mary Ann D.; Hall, Jeff Steven Grotelueschen; Lyamichev, Victor; Olive, David Michael; Prudent, James Robert

    1999-01-01

    The present invention relates to means for the detection and characterization of nucleic acid sequences, as well as variations in nucleic acid sequences. The present invention also relates to methods for forming a nucleic acid cleavage structure on a target sequence and cleaving the nucleic acid cleavage structure in a site-specific manner. The 5' nuclease activity of a variety of enzymes is used to cleave the target-dependent cleavage structure, thereby indicating the presence of specific nucleic acid sequences or specific variations thereof. The present invention further relates to methods and devices for the separation of nucleic acid molecules based by charge.

  1. A combined single-molecule FRET and tryptophan fluorescence study of RNase H folding under acidic conditions

    Graphical abstract: A model-based global analysis of single-molecule FRET and tryptophan fluorescence spectroscopy data allows the folding of RNase H, pH 3.0, to be studied as a function of the concentration of the chemical denaturant GdmCl. Highlights: ► Global analysis of smFRET and tryptophan fluorescence data of RNase H. ► Analysis yields conclusive results despite strongly overlapping FRET distributions. ► RNase H’s folding intermediate is resolved under destabilizing conditions at pH 3 - Abstract: By using single-molecule Förster resonance energy transfer (smFRET), we have studied RNase H in the presence of the denaturant guanidinium chloride to explore its well-known folding intermediate. FRET efficiency histograms were determined from experiments on freely diffusing proteins at pH 3. Even with excellent data statistics, a folding intermediate is not obvious from the FRET efficiency histograms, owing to the broad and overlapping distributions associated with each of the three macroscopic (folded, intermediate, unfolded) states. We developed a global fitting procedure for the smFRET data based on a thermodynamic model that assumes the free energies of the three states to vary linearly with denaturant activity. In addition, we included tryptophan emission data measured on bulk samples, which place significant constraints on the relative populations of the three states. The analysis yields free energy differences between the states and, thus, fractional populations over the entire range of denaturant concentration.

  2. Methods for detecting formation mechanisms and determining a final strain value for different scales of folded structures

    Yakovlev, Fedor L.

    2012-03-01

    Linear folding, developing in fold and thrust belts, is treated as a hierarchic system, at each level of which objects are described by special kinematic models. Geometric parameters of natural folded structures are determined by a combination of various mechanisms incorporated in the model, and a value of finite strain. Several case studies demonstrate how such data enables one to solve structural and geodynamic problems for natural objects of different size. Shortening value of two morphological types of folds is determined based on the geometry of competent layers. Application of the method to analyze the folds of the Vorontsov nappe (Greater Caucasus) determines its gravitational origin. Structural cross-sections though several tectonic zones are subdivided into relatively small domains, the geometry of which, particularly in thin-bedded flysch deposits, making it possible to identify the mechanisms of formation of both local and large structures, and also to reconstruct the pre-folded state of each domain and of the entire cross-sections. By aggregation of tectonic domains into large modules and determination of the value of shortening, we have constructed for the first time a 3D model of the present-day structure of the northwestern Caucasus, which is balanced for the whole sedimentary cover. The geometry of large structures makes it possible to validate geodynamic models.

  3. Detecting coevolving amino acid sites using Bayesian mutational mapping

    Dimmic, Matthew W.; Hubisz, Melissa J.; Bustamente, Carlos D.;

    2005-01-01

    Motivation: The evolution of protein sequences is constrained by complex interactions between amino acid residues. Because harmful substitutions may be compensated for by other substitutions at neighboring sites, residues can coevolve. We describe a Bayesian phylogenetic approach to the detection...

  4. GANDivAWeb: A web server for detecting early folding units ("foldons" from protein 3D structures

    Krishnan Arun

    2008-03-01

    Full Text Available Abstract Background It has long been known that small regions of proteins tend to fold independently and are then stabilized by interactions between these distinct subunits or modules. Such units, also known as autonomous folding units (AFUs or"foldons" play a key role in protein folding. A knowledge of such early folding units has diverse applications in protein engineering as well as in developing an understanding of the protein folding process. Such AFUs can also be used as model systems in order to study the structural organization of proteins. Results In an earlier work, we had utilized a global network partitioning algorithm to identify modules in proteins. We had shown that these modules correlate well with AFUs. In this work, we have developed a webserver, GANDivAWeb, to identify early folding units or "foldons" in networks using the algorithm described earlier. The website has three functionalities: (a It is able to display information on the modularity of a database of 1420 proteins used in the original work, (b It can take as input an uploaded PDB file, identify the modules using the GANDivA algorithm and email the results back to the user and (c It can take as input an uploaded PDB file and a results file (obtained from functionality (b and display the results using the embedded viewer. The results include the module decomposition of the protein, plots of cartoon representations of the protein colored by module identity and connectivity as well as contour plots of the hydrophobicity and relative accessible surface area (RASA distributions. Conclusion We believe that the GANDivAWeb server, will be a useful tool for scientists interested in the phenomena of protein folding as well as in protein engineering. Our tool not only provides a knowledge of the AFUs through a natural graph partitioning approach but is also able to identify residues that are critical during folding. It is our intention to use this tool to study the topological

  5. Detection of formaldehyde in textiles by chromotropic acid method

    Rao Sanath; Shenoy Shruthakirthi; Davis Suraj; Nayak Sudhakar

    2004-01-01

    BACKGROUND: The common causes of textile dermatitis are formaldehyde resins and disperse dyes. There are various methods to detect the presence of formaldehyde in clothing. AIM: To detect the presence of formaldehyde in various types of textiles by the chromotropic acid method and to assess the effect of washing on the formaldehyde content. METHODS: Twenty randomly selected textiles from a local cloth store were tested for formaldehyde by the chromotropic acid method. A purple ring indicated ...

  6. Biomimetic folding particle chains

    Full text: The sequence of the amino acids in proteins dictates their folded 3-D structure. We have recently by simulations shown that this principle can be applied to flexible strings of isotropically interacting particles with at least one attractive patchy interaction, allowing the design of new materials and structures. Our goal is now to realize this directed self-folding on a colloidal size scale to study the folding in real time in real space. We discuss our use of polymer brushes, depletion interactions and liquid-interface scaffold chemistry to realize the goal. (author)

  7. Early kinetic intermediate in the folding of acyl-CoA binding protein detected by fluorescence labeling and ultrarapid mixing

    Teilum, Kaare; Maki, Kosuke; Kragelund, Birthe B;

    2002-01-01

    Early conformational events during folding of acyl-CoA binding protein (ACBP), an 86-residue alpha-helical protein, were explored by using a continuous-flow mixing apparatus with a dead time of 70 micros to measure changes in intrinsic tryptophan fluorescence and tryptophan-dansyl fluorescence...... energy transfer. Although the folding of ACBP was initially described as a concerted two-state process, the tryptophan fluorescence measurements revealed a previously unresolved phase with a time constant tau = 80 micros, indicating formation of an intermediate with only slightly enhanced fluorescence of...... partially collapsed states with approximately 1/3 of the solvent-accessible surface buried. The findings indicate that ultrafast mixing methods combined with sensitive conformational probes can reveal transient accumulation of intermediate states in proteins with apparent two-state folding mechanisms....

  8. Phosphatidic acid plays a special role in stabilizing and folding of the tetrameric potassium channel KcsA

    Raja, M.M.; Spelbrink, R E J; de Kruijff, B.; Killian, J A

    2007-01-01

    In this study, we investigated how the presence of anionic lipids influenced the stability and folding properties of the potassium channel KcsA. By using a combination of gel electrophoresis, tryptophan fluorescence and acrylamide quenching experiments, we found that the presence of the anionic lipid phosphatidylglycerol (PG) in a phosphatidylcholine (PC) bilayer slightly stabilized the tetramer and protected it from trifluoroethanol- induced dissociation. Surprisingly, the presence of phosph...

  9. A Hybrid of Genetic Algorithm and Support Vector Machine for Feature Reduction and Detection of Vocal Fold Pathology

    Vahid Majidnezhad

    2013-07-01

    Full Text Available Acoustic analysis is a proper method in vocal fold pathology diagnosis so that it can complement and in some cases replace the other invasive, based on direct vocal fold observation, methods. There are different approaches and algorithms for vocal fold pathology diagnosis. These algorithms usually have three stages which are Feature Extraction, Feature Reduction and Classification. While the third stage implies a choice of a variety of machine learning methods (Support Vector Machines, Artificial Neural Networks, etc, the first and second stages play a critical role in performance and accuracy of the classification system. In this paper we present initial study of feature extraction and feature reduction in the task of vocal fold pathology diagnosis. A new type of feature vector, based on wavelet packet decomposition and Mel-Frequency-Cepstral-Coefficients (MFCCs, is proposed. Also a new GA-based method for feature reduction stage is proposed and compared with conventional methods such as Principal Component Analysis (PCA. Support vector machine is used as a classifier for evaluating the performance of the proposed method. The results show the priority of the proposed method in comparison with the current methods.

  10. Developing nucleic acid-based electrical detection systems

    Gabig-Ciminska Magdalena

    2006-03-01

    Full Text Available Abstract Development of nucleic acid-based detection systems is the main focus of many research groups and high technology companies. The enormous work done in this field is particularly due to the broad versatility and variety of these sensing devices. From optical to electrical systems, from label-dependent to label-free approaches, from single to multi-analyte and array formats, this wide range of possibilities makes the research field very diversified and competitive. New challenges and requirements for an ideal detector suitable for nucleic acid analysis include high sensitivity and high specificity protocol that can be completed in a relatively short time offering at the same time low detection limit. Moreover, systems that can be miniaturized and automated present a significant advantage over conventional technology, especially if detection is needed in the field. Electrical system technology for nucleic acid-based detection is an enabling mode for making miniaturized to micro- and nanometer scale bio-monitoring devices via the fusion of modern micro- and nanofabrication technology and molecular biotechnology. The electrical biosensors that rely on the conversion of the Watson-Crick base-pair recognition event into a useful electrical signal are advancing rapidly, and recently are receiving much attention as a valuable tool for microbial pathogen detection. Pathogens may pose a serious threat to humans, animal and plants, thus their detection and analysis is a significant element of public health. Although different conventional methods for detection of pathogenic microorganisms and their toxins exist and are currently being applied, improvements of molecular-based detection methodologies have changed these traditional detection techniques and introduced a new era of rapid, miniaturized and automated electrical chip detection technologies into pathogen identification sector. In this review some developments and current directions in

  11. Acid catalysis of the formation of the slow-folding species of RNase A: Evidence that the reaction is proline isomerization

    Schmid, Franz X.; Baldwin, Robert L.

    1978-01-01

    Unfolded RNase A is known to contain an equilibrium mixture of two forms, a slow-folding form (U1) and a fast-folding form (U2). If U1 is produced after unfolding by the slow cis-trans isomerization of proline residues about X-Pro imide bonds, then the formation of U1 should be catalyzed by strong acids. Therefore, the rate of formation of U1 has been measured at different HClO4 concentrations. After rapid unfolding of the native protein in concentrated HClO4 at 0°, the slow formation of U1 was measured by use of refolding assays. Catalysis of its formation was found at HClO4 concentrations above 5 M. The uncatalyzed reaction follows apparent first-order kinetics but, in the acid-catalyzed range, two reactions are found. The faster reaction produces two-thirds of the slow-folding species and shows acid catalysis above 5 M HClO4. Catalysis of the slower reaction begins at 8 M HClO4. The faster reaction shows a 100-fold increase in rate at 10.6 M HClO4 over the rate of the uncatalyzed reaction of 5 M. The activation enthalpy of the uncatalyzed reaction has been measured in two sets of unfolding conditions: ΔH‡ is 21.5 kcal/mol (1 kcal = 4.2 × 103 J) in 3.3 M HClO4 and 21.0 kcal/mol in 5 M guanidine HCl, pH 2.5. Both acid catalysis of the formation of U1 and its high activation enthalpy are consistent with the rate-limiting step being cis-trans isomerization either of X-Pro imide bonds or of peptide bond. The rate of the uncatalyzed reaction is in the range expected for proline isomerization and is 0.1% of that of peptide bond isomerization; thus, the simplest explanation for the formation of U1 is proline isomerization. Earlier data, showing that the kinetic properties of the U1 ⇄ U2 reaction in refolding conditions differ from those of proline isomerization, can be explained if there is kinetic coupling between early steps in the folding of U1 and its conversion to U2. The existence of two acid-catalyzed reactions that are distinguished by the HClO4

  12. Detection of obstructive uropathy using 99m technetium diethylenetriaminepentaacetic acid

    A simple, accurate test with no morbidity and effective for the detection of urinary tract obstruction in patients with normal and impaired renal function would be of significant clinical use. The ability of 99m technetium diethylenetriaminepentaacetic acid to detect the presence or absence of obstruction is evaluated. Obstructions were defined as abnormal retention of activity in the collecting system persisting in the delayed images. Diethylentriaminepentaacetic acid identified correctly 13 of 13 patients proved to have obstruction and 17 of 18 without obstruction. These data indicate a sensitivity of 100 percent, specificity of 94 percent and accuracy of 97 percent

  13. Detection of obstructive uropathy using 99m technetium diethylenetriaminepentaacetic acid

    Powers, T.A.; Grove, R.B.; Bauriedel, J.K.; Orr, S.C.; Melton, R.E.; Bowenn, R.D.

    1980-11-01

    A simple, accurate test with no morbidity and effective for the detection of urinary tract obstruction in patients with normal and impaired renal function would be of significant clinical use. The ability of 99m technetium diethylenetriaminepentaacetic acid to detect the presence or absence of obstruction is evaluated. Obstructions were defined as abnormal retention of activity in the collecting system persisting in the delayed images. Diethylentriaminepentaacetic acid identified correctly 13 of 13 patients proved to have obstruction and 17 of 18 without obstruction. These data indicate a sensitivity of 100 percent, specificity of 94 percent and accuracy of 97 percent.

  14. Nucleic acid detection in the diagnosis and prevention of schistosomiasis.

    He, Ping; Song, Lan-Gui; Xie, Hui; Liang, Jin-Yi; Yuan, Dong-Ya; Wu, Zhong-Dao; Lv, Zhi-Yue

    2016-01-01

    Schistosomiasis is an important zoonotic parasitic disease that causes serious harms to humans and animals. Surveillance and diagnosis play key roles in schistosomiasis control, however, current techniques for surveillance and diagnosis of the disease have limitations. As genome data for parasites are increasing, novel techniques for detection incorporating nucleotide sequences are receiving widespread attention. These sensitive, specific, and rapid detection methods are particularly important in the diagnosis of low-grade and early infections, and may prove to have clinical significance. This paper reviews the progress of nucleic acid detection in the diagnosis and prevention of schistosomiasis, including such aspects as the selection of target genes, and development and application of nucleic acid detection methods. PMID:27025210

  15. Nucleic Acid-Based Approaches for Detection of Viral Hepatitis

    BEHZADI, Payam; Ranjbar, Reza; Alavian, Seyed Moayed

    2014-01-01

    Context: To determining suitable nucleic acid diagnostics for individual viral hepatitis agent, an extensive search using related keywords was done in major medical library and data were collected, categorized, and summarized in different sections. Results: Various types of molecular biology tools can be used to detect and quantify viral genomic elements and analyze the sequences. These molecular assays are proper technologies for rapidly detecting viral agents with high accuracy, high sensit...

  16. Graphdiyne as a promising material for detecting amino acids

    Chen, Xi; Gao, Pengfei; Guo, Lei; Zhang, Shengli

    2015-11-01

    The adsorption of glycine, glutamic acid, histidine and phenylalanine on single-layer graphdiyne/ graphene is investigated by ab initio calculations. The results show that for each amino acid molecule, the adsorption energy on graphdiyne is larger than the adsorption energy on graphene and dispersion interactions predominate in the adsorption. Molecular dynamics simulations reveal that at room temperature the amino acid molecules keep migrating and rotating on graphdiyne surface and induce fluctuation in graphdiyne bandgap. Additionally, the photon absorption spectra of graphdiyne-amino-acid systems are investigated. We uncover that the presence of amino acid molecules makes the photon absorption peaks of graphdiyne significantly depressed and shifted. Finally, quantum electronic transport properties of graphdiyne-amino-acid systems are compared with the transport properties of pure graphdiyne. We reveal that the amino acid molecules induce distinct changes in the electronic conductivity of graphdiyne. The results in this paper reveal that graphdiyne is a promising two-dimensional material for sensitively detecting amino acids and may potentially be used in biosensors.

  17. Chirality and Protein Folding

    Kwiecinska, Joanna I.; Cieplak, Marek

    2004-01-01

    There are several simple criteria of folding to a native state in model proteins. One of them involves crossing of a threshold value of the RMSD distance away from the native state. Another checks whether all native contacts are established, i.e. whether the interacting amino acids come closer than some characteristic distance. We use Go-like models of proteins and show that such simple criteria may prompt one to declare folding even though fragments of the resulting conformations have a wron...

  18. Liquid crystal based biosensors for bile acid detection

    He, Sihui; Liang, Wenlang; Tanner, Colleen; Fang, Jiyu; Wu, Shin-Tson

    2013-03-01

    The concentration level of bile acids is a useful indicator for early diagnosis of liver diseases. The prevalent measurement method in detecting bile acids is the chromatography coupled with mass spectrometry, which is precise yet expensive. Here we present a biosensor platform based on liquid crystal (LC) films for the detection of cholic acid (CA). This platform has the advantage of low cost, label-free, solution phase detection and simple analysis. In this platform, LC film of 4-Cyano-4'-pentylbiphenyl (5CB) was hosted by a copper grid supported with a polyimide-coated glass substrate. By immersing into sodium dodecyl sulfate (SDS) solution, the LC film was coated with SDS which induced a homeotropic anchoring of 5CB. Addition of CA introduced competitive adsorption between CA and SDS at the interface, triggering a transition from homeotropic to homogeneous anchoring. The detection limit can be tuned by changing the pH value of the solution from 12uM to 170uM.

  19. Ionizing radiation damage to the folded chromosome of Escherichia coli K-12: sedimentation properties of irradiated nucleoids and chromosomal deoxyribonucleic acid

    The structures of the membrane-free nucleoid of Escherichia coli K-12 and of unfolded chromosomal deoxyribonucleic acid (DNA) were investigated by low-speed sedimentation on neutral sucrose gradients after irradiation with 60Co gamma rays. Irradiation both in vivo and in vitro was used as a molecular probe of the constraints on DNA packaging in the bacterial chromosome. The number of domains of supercoiling was estimated to be approximately 180 per genome equivalent of DNA, based on measurements of relaxation caused by single-strand break formation in folded chromosomes gamma irradiated in vivo and in vitro. Similar estimates based on the target size of ribonucleic acid molecules responsible for maintaining the compact packaging of the nucleoid predicted negligible unfolding due to the formation of ribonucleic acid single-strand breaks at doses of up to 10 krad; this was born out by experimental measurements. Unfolding of the nucleoid in vitro by limit digestion with ribonuclease or by heating at 700C resulted in DNA complexes with sedimentation coefficients of 1,030 +- 59S and 625 +- 15S, respectively. The difference in these rates was apparently due to more complete deproteinization and thus less mass in the heated material. These structures are believed to represent intact, replicating genomes in the form of complex-theta structures containing two to three genome equivalents of DNA

  20. Optical detection of folded mini-zone-edge coherent acoustic modes in a doped GaAs/AlAs superlattice

    Beardsley, R.; Akimov, A. V.; Glavin, B. A.; Maryam, W.; Henini, M.; Kent, A. J.

    2010-07-01

    A coherent phonon mode with frequency corresponding to the first mini Brillouin-zone edge stop gap is observed in ultrafast pump-probe measurements on a doped semiconductor superlattice structure. It is proposed that the optical detection of the mode is facilitated by interactions with the free carriers present in the superlattice.

  1. Affinity sensor using 3-aminophenylboronic acid for bacteria detection.

    Wannapob, Rodtichoti; Kanatharana, Proespichaya; Limbut, Warakorn; Numnuam, Apon; Asawatreratanakul, Punnee; Thammakhet, Chongdee; Thavarungkul, Panote

    2010-10-15

    Boronic acid that can reversibly bind to diols was used to detect bacteria through its affinity binding reaction with diol-groups on bacterial cell walls. 3-aminophenylboronic acid (3-APBA) was immobilized on a gold electrode via a self-assembled monolayer. The change in capacitance of the sensing surface caused by the binding between 3-APBA and bacteria in a flow system was detected by a potentiostatic step method. Under optimal conditions the linear range of 1.5×10(2)-1.5×10(6) CFU ml(-1) and the detection limit of 1.0×10(2) CFU ml(-1) was obtained. The sensing surface can be regenerated and reused up to 58 times. The method was used for the analysis of bacteria in several types of water, i.e., bottled, well, tap, reservoir and wastewater. Compared with the standard plate count method, the results were within one standard deviation of each other. The proposed method can save both time and cost of analysis. The electrode modified with 3-APBA would also be applicable to the detection of other cis-diol-containing analytes. The concept could be extended to other chemoselective ligands, offering less expensive and more robust affinity sensors for a wide range of compounds. PMID:20801635

  2. Nucleic acid sequence detection using multiplexed oligonucleotide PCR

    Nolan, John P.; White, P. Scott

    2006-12-26

    Methods for rapidly detecting single or multiple sequence alleles in a sample nucleic acid are described. Provided are all of the oligonucleotide pairs capable of annealing specifically to a target allele and discriminating among possible sequences thereof, and ligating to each other to form an oligonucleotide complex when a particular sequence feature is present (or, alternatively, absent) in the sample nucleic acid. The design of each oligonucleotide pair permits the subsequent high-level PCR amplification of a specific amplicon when the oligonucleotide complex is formed, but not when the oligonucleotide complex is not formed. The presence or absence of the specific amplicon is used to detect the allele. Detection of the specific amplicon may be achieved using a variety of methods well known in the art, including without limitation, oligonucleotide capture onto DNA chips or microarrays, oligonucleotide capture onto beads or microspheres, electrophoresis, and mass spectrometry. Various labels and address-capture tags may be employed in the amplicon detection step of multiplexed assays, as further described herein.

  3. Complex folding and misfolding effects of deer-specific amino acid substitutions in the β2-α2 loop of murine prion protein

    Agarwal, Sonya; Döring, Kristina; Gierusz, Leszek A.; Iyer, Pooja; Lane, Fiona M.; Graham, James F.; Goldmann, Wilfred; Pinheiro, Teresa J. T.; Gill, Andrew C.

    2015-10-01

    The β2-α2 loop of PrPC is a key modulator of disease-associated prion protein misfolding. Amino acids that differentiate mouse (Ser169, Asn173) and deer (Asn169, Thr173) PrPC appear to confer dramatically different structural properties in this region and it has been suggested that amino acid sequences associated with structural rigidity of the loop also confer susceptibility to prion disease. Using mouse recombinant PrP, we show that mutating residue 173 from Asn to Thr alters protein stability and misfolding only subtly, whilst changing Ser to Asn at codon 169 causes instability in the protein, promotes oligomer formation and dramatically potentiates fibril formation. The doubly mutated protein exhibits more complex folding and misfolding behaviour than either single mutant, suggestive of differential effects of the β2-α2 loop sequence on both protein stability and on specific misfolding pathways. Molecular dynamics simulation of protein structure suggests a key role for the solvent accessibility of Tyr168 in promoting molecular interactions that may lead to prion protein misfolding. Thus, we conclude that ‘rigidity’ in the β2-α2 loop region of the normal conformer of PrP has less effect on misfolding than other sequence-related effects in this region.

  4. Clinical comparison between two hyaluronic acid-derived fillers in the treatment of nasolabial folds in Chinese subjects: BioHyalux versus Restylane.

    Wu, Yan; Sun, Nan; Xu, Yue; Liu, Huixian; Zhong, Shaomin; Chen, Liyang; Li, Dong

    2016-04-01

    Hyaluronic acid fillers are used to improve the appearance of nasolabial folds (NLF). This study aimed to compare the efficacy, safety, and durability of a new hyaluronic acid gel (BioHyalux) versus Restylane for the correction of NLF. This was a multicenter, double-blinded, randomized, controlled, non-inferiority clinical trial involving 88 subjects with moderate to severe NLF. Subjects were randomized to BioHyalux and Restylane on either sides of the NLF. NLF was assessed before and right after injection, and at 1 week, 1, 3, and 6 months. Patients were followed up for 13-15 months to evaluate the durability and long-term safety. A clinically meaningful response was predefined as at least one-point improvement on the Wrinkle Severity Rating Scale, which is a five-point scale. At 6 months, the response rate of BioHyalux was not inferior to that of Restylane (P  0.05) at all time points. At 6 months, 100 % reported improvements on both side; at 13-15 months, 60 % of subjects reported improvements with BioHyalux versus 64 % with Restylane. Adverse events were transient and predominantly mild or moderate in severity including injection site swelling, pain, itching, bruising, and tenderness. BioHyalux had reliable safety and tolerance, and could be an effective injectable filler for correcting NLF. PMID:26924549

  5. In-situ detection of tropospheric OH radicals by folded long-path laser absorption. Results from the POPCORN Field Campaign in August 1994

    Dorn, H.-P.; Brandenburger, U.; Brauers, T.; Hausmann, M.; Ehhalt, D. H.

    Ground based in-situ measurements of tropospheric hydroxyl radicals were conducted by folded long-path laser absorption as part of the field campaign POPCORN in August 1994. The OH instrument used an open optical multiple-reflection cell of 38.5 m base length through which the laser beam was passed up to 80 times. The broadband emission of a short-pulse UV laser together with a multichannel detection system allowed the simultaneous observation of six OH absorption lines in a spectral interval of Δλ≃0.24 nm at 308.1nm (A²Σ+,υ‧ = 0← X²Π,υ″ = 0 transition). Along with the OH radicals, the trace gases SO2, HCHO, and naphthalene were measured by this technique. The large spectral detection range covered a multitude of rotational absorption lines of these trace gases which were all used for multicomponent analysis, thus allowing a specific and sensitive detection of tropospheric OH radicals. An average 2σ detection limit of 1.5 × 106 OH/cm³ for an integration time of 200 seconds and an absorption light path length of 1848 m was determined from the field measurements. In total, 392 OH data were obtained by long-path absorption during 16 days of field measurements. The observed OH concentrations reached peak values of 13 × 106 cm-3 at noon.

  6. Preparation and detection of nonradioactive nucleic acid and oligonucleotide

    There is increasing interest worldwide in the development of nucleic acid probes which are detected by nonradioactive means. In the research laboratory, the use of 32P for detection is undoubtedly the method of choice and is likely to remain so for the forseeable future, in spite of the half life of only 14 days for 32P. In the diagnostic laboratory on the other hand, the use of nonradioactive probes has many potential advantages. Perhaps the major one is that nonradioactive probes are stable for at least 6 to 12 months, and probably much longer if properly stored, thus leading to a substantial reduction in cost by obviating the need to prepare them every 2 to 3 weeks. In addition, there is no radiation exposure from routine daily use and there are no storage and disposal problems. Numerous methods are described in this chapter for the preparation by enzymatic and chemical techniques of nonradioactive nucleic acid and oligonucleotide probes. In many cases, the resulting probes have yet to be fully tested under hybridization conditions. In others, initial results look very promising since some nonradioactive probes can provide a sensitivity of detection of target sequences similar to that provided by 32P-labeled probes

  7. Conformational Transition of Poly (Acrylic Acid) Detected by Microcantilever Sensing

    LI Kai; LIU Hong; ZHANG Qing-Chuan; XUE Chang-Guo; WU Xiao-Ping

    2007-01-01

    Poly (acrylic acid) (PAA) chains are grafted on one side of a microcantilever by the self-assembled method and the deflections of the microcantilever are detected as a function of medium pH from 3 to 11. It is found that when the pH varies, the microcantilever deflects because of the changing surface stress. By analysing the electrostatic repulsive effect, the surface stress change is related to the conformation transition of PAA from a collapse state to a swelling state. This method offers the interaction information among the polymer chains during the conformational transition and affords an alternative way to study conformational change of polymers.

  8. Methodology for detecting residual phosphoric acid in polybenzoxazole fibers.

    Park, Eun Su; Sieber, John; Guttman, Charles; Rice, Kirk; Flynn, Kathleen; Watson, Stephanie; Holmes, Gale

    2009-12-01

    Because of the premature failure of in-service soft-body armor containing the ballistic fiber poly[(benzo-[1,2-d:5,4-d']-benzoxazole-2,6-diyl)-1,4-phenylene] (PBO), the Office of Law Enforcement Standards (OLES) at the National Institute of Standards and Technology (NIST) initiated a research program to investigate the reasons for this failure and to develop testing methodologies and protocols to ensure that these types of failures do not reoccur. In a report that focused on the stability of the benzoxazole ring that is characteristic of PBO fibers, Holmes, G. A.; Rice, K.; Snyder, C. R. J. Mater. Sci. 2006, 41, 4105-4116, showed that the benzoxazole ring was susceptible to hydrolytic degradation under acid conditions. Because of the processing conditions for the fibers, it is suspected by many researchers that residual phosphoric acid may cause degradation of the benzoxazole ring resulting in a reduction of ballistic performance. Prior to this work, no definitive data have indicated the presence of phosphoric acid since the residual phosphorus is not easily extracted and the processed fibers are known to incorporate phosphorus containing processing aids. Methods to efficiently extract phosphorus from PBO are described in this article. Further, characterization determined that the majority of the extractable phosphorus in PBO was attributed to the octyldecyl phosphate processing aid with some phosphoric acid being detected. Analysis by matrix assisted laser desorption ionization of model PBO oligomers indicates that the nonextractable phosphorus is attached to the PBO polymer chain as a monoaryl phosphate ester. The response of model aryl phosphates to NaOH exposure indicates that monoaryl phosphate ester is stable to NaOH washes used in the manufacturing process to neutralize the phosphoric acid reaction medium and to extract residual phosphorus impurities. PMID:19899783

  9. The role of a conserved acidic residue in calcium-dependent protein folding for a low density lipoprotein (LDL)-A module: implications in structure and function for the LDL receptor superfamily.

    Guo, Ying; Yu, Xuemei; Rihani, Kayla; Wang, Qing-Yin; Rong, Lijun

    2004-04-16

    One common feature of the more than 1,000 complement-type repeats (or low density lipoprotein (LDL)-A modules) found in LDL receptor and the other members of the LDL receptor superfamily is a cluster of five highly conserved acidic residues in the C-terminal region, DXXXDXXDXXDE. However, the role of the third conserved aspartate of these LDL-A modules in protein folding and ligand recognition has not been elucidated. In this report, using a model LDL-A module and several experimental approaches, we demonstrate that this acidic residue, like the other four conserved acidic residues, is involved in calcium-dependent protein folding. These results suggest an alternative calcium coordination conformation for the LDL-A modules. The proposed model provides a plausible explanation for the conservation of this acidic residue among the LDL-A modules. Furthermore, the model can explain why mutations of this residue in human LDL receptor cause familial hypercholesterolemia. PMID:14749324

  10. Chirality and protein folding

    Kwiecinska, Joanna I; Cieplak, Marek [Institute of Physics, Polish Academy of Sciences, Aleja Lotnikow 32/46, 02-668 Warsaw (Poland)

    2005-05-11

    There are several simple criteria of folding to a native state in model proteins. One of them involves crossing of a threshold value of the root mean square deviation distance away from the native state. Another checks whether all native contacts are established, i.e. whether the interacting amino acids come closer than some characteristic distance. We use Go-like models of proteins and show that such simple criteria may prompt one to declare folding even though fragments of the resulting conformations have a wrong sense of chirality. We propose that a better condition of folding should augment the simple criteria with the requirement that most of the local values of the chirality should be nearly native. The kinetic discrepancy between the simple and compound criteria can be substantially reduced in the Go-like models by providing the Hamiltonian with a term which favours native values of the local chirality. We study the effects of this term as a function of its amplitude and compare it to other models such as ones with side groups and ones with angle-dependent potentials.

  11. Chirality and protein folding

    Kwiecinska, Joanna I.; Cieplak, Marek

    2005-05-01

    There are several simple criteria of folding to a native state in model proteins. One of them involves crossing of a threshold value of the root mean square deviation distance away from the native state. Another checks whether all native contacts are established, i.e. whether the interacting amino acids come closer than some characteristic distance. We use Go-like models of proteins and show that such simple criteria may prompt one to declare folding even though fragments of the resulting conformations have a wrong sense of chirality. We propose that a better condition of folding should augment the simple criteria with the requirement that most of the local values of the chirality should be nearly native. The kinetic discrepancy between the simple and compound criteria can be substantially reduced in the Go-like models by providing the Hamiltonian with a term which favours native values of the local chirality. We study the effects of this term as a function of its amplitude and compare it to other models such as ones with side groups and ones with angle-dependent potentials.

  12. Pretreatment of various feedstocks for lactic acid production: detection of sugars, organic acids and furanics in liquid fractions

    Harmsen, P.F.H.; Lips, S.J.J.; Bakker, R.R.C.

    2012-01-01

    Barley straw, sugarcane bagasse and empty fruit bunches were pretreated under acid- and alkaline conditions. Solid phase was separated from the liquid phase and the concentration of dissolved monomeric sugars, organic acids and furanics was determined. Acid hydrolysis yielded monomeric xylose concentrations (max 20 g/l) whereas for alkaline hydrolysis less than 1 g/l was found. Organic acids and furanics were detected with high concentrations when dried empty fruit bunches were used. Acetic a...

  13. Ionizing radiation damage to the folded chromosome of Escherichia coli K-12: repair of double-strand breaks in deoxyribonucleic acid

    The extremely gentle lysis and unfolding procedures that have been developed for the isolation of nucleoid deoxyribonucleic acid yield undamaged, replicating genomes, thus permitting direct measurement of the formation and repair of DNA double-strand breaks at biologically significant doses of ionizing radiation. Repair of ionizing radiation damage to folded chromosomes of Escherichia coli K-12 strain AB2497 was observed within 2 to 3 h of post-irradiation incubation in growth medium. Such behavior was not observed after post-irradiation incubation in growth medium of a recA13 strain (strain AB2487). A model based on recombinational repair is proposed to explain the formation of 2,200 to 2,300S material during early stages of incubation and to explain subsequent changes in the gradient profiles. Association of unrepaired DNA with the plasma membrane is proposed to explain the formation of a peak of rapidly sedimenting material (greater than 3,100S) during the later stage of repair. Direct evidence of repair of double-strand breaks during post-irradiation incubation in growth medium was obtained from gradient profiles of DNA from ribonuclease-digested chromosomes. The sedimentation coefficient of broken molecules was restored to the value of unirradiated DNA after 2 to 3 h of incubation, and the fraction of the DNA repaired in this fashion was equal to the fraction of cells that survived at the same dose. An average of 2.7 double-strand breaks per genome per lethal event was observed, suggesting that one to two double-strand breaks per genome are repairable in E. coli K-12 strain AB2497

  14. Detection and qualitative analysis of fatty acid amides in the urine of alcoholics using HPLC-QTOF-MS.

    Dabur, Rajesh; Mittal, Ashwani

    2016-05-01

    Fatty acid amides (FAAs) in alcoholism lead to liver diseases. These amides have been reported in plasma and in other organs of the body, while their detection or presence in the urine is still unknown. Therefore, the focus of the current study was to detect and analyze FAAs qualitatively in urine samples of alcoholics. Furthermore, the effects of Tinospora cordifolia (hepatoprotective medicinal plant) intervention on FAA levels in moderate alcoholics were also analyzed. In the study, asymptomatic chronic alcoholics (n = 22) without chronic liver disease and nonalcoholic healthy volunteers (n = 24) with a mean age of 39 ± 2.0 years were selected. The first-pass urine and fasting blood samples were collected in the morning on day 0 and day 14 after T. cordifolia water extract (TCE) treatment and analyzed using automated biochemistry analyzer and HPLC-QTOF-MS. Results indicated the increased levels of serum triglycerides, cholesterol, and liver function enzymes in alcoholic subjects, which were significantly down-regulated by TCE intervention. Multivariate discrimination analysis of QTOF-MS data showed increased urinary levels of oleoamide (2.55-fold), palmitamide (5.6-fold), and erucamide (1.6-fold) in alcoholics as compared to control subjects. Levels of oleamide (1.8-fold), palmitamide (1.7-fold), and linoleamide (1.5-fold) were found to be increased in plasma. Treatment with TCE in alcoholics (3.0 g lyophilized water extract/day) significantly decreased the plasma and urinary levels of all FAAs except linoleamide. The HPLC-QTOF-MS approach for FAAs analysis in both urinary and plasma samples of alcoholics worked very well. Moreover, findings (i.e., increased levels of FAAs in urine and in plasma) further support other findings that these amides play a very important role in alcoholism. Further, like our previous findings, TCE proved its hepatoprotective effect against alcoholism not only by lowering the levels of these detected FAAs, but also by decreasing the

  15. HMMerThread: detecting remote, functional conserved domains in entire genomes by combining relaxed sequence-database searches with fold recognition.

    Charles Richard Bradshaw

    Full Text Available Conserved domains in proteins are one of the major sources of functional information for experimental design and genome-level annotation. Though search tools for conserved domain databases such as Hidden Markov Models (HMMs are sensitive in detecting conserved domains in proteins when they share sufficient sequence similarity, they tend to miss more divergent family members, as they lack a reliable statistical framework for the detection of low sequence similarity. We have developed a greatly improved HMMerThread algorithm that can detect remotely conserved domains in highly divergent sequences. HMMerThread combines relaxed conserved domain searches with fold recognition to eliminate false positive, sequence-based identifications. With an accuracy of 90%, our software is able to automatically predict highly divergent members of conserved domain families with an associated 3-dimensional structure. We give additional confidence to our predictions by validation across species. We have run HMMerThread searches on eight proteomes including human and present a rich resource of remotely conserved domains, which adds significantly to the functional annotation of entire proteomes. We find ∼4500 cross-species validated, remotely conserved domain predictions in the human proteome alone. As an example, we find a DNA-binding domain in the C-terminal part of the A-kinase anchor protein 10 (AKAP10, a PKA adaptor that has been implicated in cardiac arrhythmias and premature cardiac death, which upon stress likely translocates from mitochondria to the nucleus/nucleolus. Based on our prediction, we propose that with this HLH-domain, AKAP10 is involved in the transcriptional control of stress response. Further remotely conserved domains we discuss are examples from areas such as sporulation, chromosome segregation and signalling during immune response. The HMMerThread algorithm is able to automatically detect the presence of remotely conserved domains in

  16. Flips for 3-folds and 4-folds

    Corti, Alessio

    2007-01-01

    This edited collection of chapters, authored by leading experts, provides a complete and essentially self-contained construction of 3-fold and 4-fold klt flips. A large part of the text is a digest of Shokurov's work in the field and a concise, complete and pedagogical proof of the existence of 3-fold flips is presented. The text includes a ten page glossary and is accessible to students and researchers in algebraic geometry.

  17. Detection of target DNA using fluorescent cationic polymer and peptide nucleic acid probes on solid support

    Leclerc Mario

    2005-04-01

    Full Text Available Abstract Background Nucleic acids detection using microarrays requires labelling of target nucleic acids with fluorophores or other reporter molecules prior to hybridization. Results Using surface-bound peptide nucleic acids (PNA probes and soluble fluorescent cationic polythiophenes, we show a simple and sensitive electrostatic approach to detect and identify unlabelled target nucleic acid on microarray. Conclusion This simple methodology opens exciting possibilities for applied genetic analysis for the diagnosis of infections, identification of genetic mutations, and forensic inquiries. This electrostatic strategy could also be used with other nucleic acid detection methods such as electrochemistry, silver staining, metallization, quantum dots, or electrochemical dyes.

  18. A novel nanocomposites sensor for epinephrine detection in the presence of uric acids and ascorbic acids

    Lu Xiaoquan, E-mail: luxq@nwnu.edu.cn [Key Laboratory of Bioelectrochemistry and Environmental Analysis of Gansu Province, College of Chemistry and Chemical Engineering, Northwest Normal University, LanZhou, 730070 (China); Li Yaya; Du Jie; Zhou Xibin; Xue Zhonghua; Liu Xiuhui; Wang Zhihua [Key Laboratory of Bioelectrochemistry and Environmental Analysis of Gansu Province, College of Chemistry and Chemical Engineering, Northwest Normal University, LanZhou, 730070 (China)

    2011-08-30

    Highlights: {center_dot} A novel PPy/AuNPs/SWCNTs nanomaterials biosensor was prepared to the selective determination of EP. {center_dot} The methods we employed to prepare PPy/AuNPs/SWCNTs nanomaterials are extremely simple. {center_dot} The PPy/AuNPs/SWCNTs nanocomposites biosensor we got from the results of experiments can totally eliminate the interference from AA and distinguish EP from UA. - Abstract: A novel nanocomposites film of conducting polymers including single-walled carbon nanotubes (SWCNTs), polypyrrole (PPy) and gold nanoparticles (AuNPs) modified electrode has been applied in voltammetric sensors to detect epinephrine (EP) sensitively when ascorbic acids (AA) and uric acids (UA) exist. The nanocomposites film of conducting polymers which show an excellent electrocatalystic activity for the oxidation of EP and UA was characterized by scanning electron microscopy (SEM) and electrochemical methods. The catalytic peak currents obtained from differential pulse voltammetry (DPV) increased linearly with increasing EP concentrations in the range of 4.0 x 10{sup -9}-1.0 x 10{sup -7} M with a detection limit of 2.0 x 10{sup -9} M (S/N = 3), respectively. The results showed that the nanocomposites of conducting polymers can selectively determine EP in the coexistence of a large amount of UA and AA. In addition, the sensor exhibited excellent sensitivity, selectivity and stability. The PPy/AuNPs/SWCNTs nanocomposites film can also be satisfactorily used for detecting EP in epinephrine hydrochloride injection when contain AA and UA, which also shows good recovery for determination of EP in some biological fluids.

  19. Simultaneous detection of seven phenolic acids in Danshen injection using HPLC with ultraviolet detector

    Jin-zhong XU; Jie SHEN; Yi-yu CHENG; Hai-bin QU

    2008-01-01

    A high-performance liquid chromatographic (HPLC) method with ultraviolet (UV) detector had been developed for simultaneous quantification of danshensu, protocatechuie aldehyde, caffeic acid, salvianolic acid D, rosmarinic acid, salvianolic acid B and salvianolic acid A in Danshen injection. According to the UV spectra of these components, three detection wavelengths have been selected as follows: 280 nm for danshensu and protocatechuic aldehyde, 326 nm for caffeic acid, salvianolic acid D and rosmarinic acid, 286 nm for salvianolic acid B and salvianolic acid A. The limit of detection (LOD) was improved to be in the range of 0.008~0.160 μg/ml. Moreover, excellent linear behavior over the investigated concentration range was observed, with R>0.999 for all the analytes.

  20. Development of ELISAs for detecting domoic acid, okadaic acid, and saxitoxin and their applicability for the detection of marine toxins in samples collected in Belgium

    Michel DUBOIS; Demoulin, Luc; Charlier, Caroline; Delahaut, Philippe; Campbell, Katrina; Elliott, Chris; Godefroy, Samuel; Singh, Gurmit

    2010-01-01

    Abstract Okadaic acid (OA), a diarrhetic shellfish poison (DSP), domoic acid (DA), an amnesic shellfish poison (ASP) and saxitoxin (SAX), a paralytic shellfish poison (PSP), are three of the best-known marine biotoxins. The mouse bioassay is the method most widely used to detect many of these toxins in shellfish samples, but animal welfare concerns have prompted researchers to seek alternative methods of detection. In this study, three direct competitive enzyme-linked immunosorbent...

  1. Development of a Method for Detection of Lactic Acid Bacteria Producing Exclusively the l-(+)- Isomer of Lactic Acid

    Jehanno, D.; Thuault, D; Bourgeois, C. M.

    1992-01-01

    A method was developed for the detection and isolation, within a population of lactic acid bacteria, of strains producing exclusively the l-(+)- isomer of lactic acid; the visual detection of colonies of these particular strains can be carried out directly on agar plates (50 to 70 colonies per plate). The method is based on an enzymatic stereospecific reaction involving d-(−)-lactate dehydrogenase and linked to a staining reaction; the diffusion area of the d-(−)- isomer stains red around the...

  2. Presence of a classical RRM-fold palm domain in Thg1-type 3'- 5'nucleic acid polymerases and the origin of the GGDEF and CRISPR polymerase domains

    Aravind L

    2010-06-01

    Full Text Available Abstract Background Almost all known nucleic acid polymerases catalyze 5'-3' polymerization by mediating the attack on an incoming nucleotide 5' triphosphate by the 3'OH from the growing polynucleotide chain in a template dependent or independent manner. The only known exception to this rule is the Thg1 RNA polymerase that catalyzes 3'-5' polymerization in vitro and also in vivo as a part of the maturation process of histidinyl tRNA. While the initial reaction catalyzed by Thg1 has been compared to adenylation catalyzed by the aminoacyl tRNA synthetases, the evolutionary relationships of Thg1 and the actual nature of the polymerase reaction catalyzed by it remain unclear. Results Using sensitive profile-profile comparison and structure prediction methods we show that the catalytic domain Thg1 contains a RRM (ferredoxin fold palm domain, just like the viral RNA-dependent RNA polymerases, reverse transcriptases, family A and B DNA polymerases, adenylyl cyclases, diguanylate cyclases (GGDEF domain and the predicted polymerase of the CRISPR system. We show just as in these polymerases, Thg1 possesses an active site with three acidic residues that chelate Mg++ cations. Based on this we predict that Thg1 catalyzes polymerization similarly to the 5'-3' polymerases, but uses the incoming 3' OH to attack the 5' triphosphate generated at the end of the elongating polynucleotide. In addition we identify a distinct set of residues unique to Thg1 that we predict as comprising a second active site, which catalyzes the initial adenylation reaction to prime 3'-5' polymerization. Based on contextual information from conserved gene neighborhoods we show that Thg1 might function in conjunction with a polynucleotide kinase that generates an initial 5' phosphate substrate for it at the end of a RNA molecule. In addition to histidinyl tRNA maturation, Thg1 might have other RNA repair roles in representatives from all the three superkingdoms of life as well as certain

  3. Determination of some hydroxybenzoic acids and catechins in white wine samples by liquid chromatography with luminescence detection.

    Rodríguez-Díaz, Rafael Carlos; Paz Aguilar-Caballos, María; Gómez-Hens, Agustina

    2006-12-01

    A liquid chromatographic method with luminescence detection for the determination of eight phenolic compounds is reported. The method involves postcolumn derivatization with terbium(III). This derivatization is based on the reaction between phenolics and terbium(III) to form luminescent chelates, which were determined at lamda ex 295 and lamda em 545 nm using the fluorescence mode. The long wavelength emission of lanthanide chelates can minimize interferences from background sample matrix, which usually emit at shorter wavelengths. Also, the chromatographic separation allows the individual determination of phenolics, which cannot be done using the direct measurement of the fluorescence of their corresponding terbium chelates. Dynamic ranges of the calibration graphs and detection limits, obtained with standard solutions of analytes were (microg/mL): gallic acid (0.9-40, 0.3), protocatechuic acid (0.05-7, 0.016), catechin (0.2-40, 0.07), vanillic acid (0.25-40, 0.08), p-hydroxybenzoic acid (0.8-40, 0.25), syringic acid (0.17-40, 0.05), epicatechin (0.3-40, 0.09) and salicylic acid (0.07-12, 0.02). The precision was established at two concentration levels of each analyte and expressed as the percentage of RSD with values ranging between 1.0 and 6.5%. The practical usefulness of the method was demonstrated by the analysis of white wine samples, which were diluted two-fold and directly injected into the chromatographic system. The recovery values obtained ranged between 93.3 and 108.0%. PMID:17305238

  4. Rapid detection method for fusaric acid-producing species of Fusarium by PCR

    Fusaric acid is a mycotoxin produced by species of the fungus Fusarium and can act synergistically with other Fusarium toxins. In order to develop a specific detection method for fusaric acid-producing fungus, PCR prim¬ers were designed to amplify FUB10, a transcription factor gene in fusaric acid ...

  5. Optimization of Chromatographic Conditions for Detecting Ellagic Acid in Pomegranate Peels Using HPLC Method

    2012-01-01

    [Objective] This study aimed to optimize the chromatographic conditions for detecting ellagic acid in pomegranate peels using HPLC method. [Method] By using 0.2 mg/ml ellagic acid standard solution, on the basis of single-factor experiment and orthogonal experiment, chromatographic conditions (mobile phase ratio, flow rate, col- umn temperature) for detecting ellagic acid using HPLC were optimized. Based on the optimal chromatographic conditions, the ellagic acid content in experimental pomegranate peels was determined. [Resull] The optimal chromatographic conditions for detecting ellagic acid in pomegranate peels using HPLC method are: 1.2% phos- phoric acid:acetonitrile=85:15, column temperature of 35 ℃, and flow rate of 1.0 ml/min. The linear regression equation of ellagic acid is: y=2.9e+0.6x+4.4e+5 (FF=9 999). Ac- cording to the standard addition recovery test, the average recovery rate of ellagic acid is 98.20%, and RSD is 0.60%. Under above optimized chromatographic condi- tions, ellagic acid can be well separated from other interfering components in pomegranate peels, with shorter peak time and ideal effect, which is convenient for the detection in production practices. [Conclusion] This study laid the foundation for detecting ellagic acid in pomegranate peels using HPLC method.

  6. Nucleic acid based fluorescent sensor for mercury detection

    Lu, Yi; Liu, Juewen

    2013-02-05

    A nucleic acid enzyme comprises an oligonucleotide containing thymine bases. The nucleic acid enzyme is dependent on both Hg.sup.2+and a second ion as cofactors, to produce a product from a substrate. The substrate comprises a ribonucleotide, a deoxyribonucleotide, or both.

  7. Reduced alphabet for protein folding prediction.

    Huang, Jitao T; Wang, Titi; Huang, Shanran R; Li, Xin

    2015-04-01

    What are the key building blocks that would have been needed to construct complex protein folds? This is an important issue for understanding protein folding mechanism and guiding de novo protein design. Twenty naturally occurring amino acids and eight secondary structures consist of a 28-letter alphabet to determine folding kinetics and mechanism. Here we predict folding kinetic rates of proteins from many reduced alphabets. We find that a reduced alphabet of 10 letters achieves good correlation with folding rates, close to the one achieved by full 28-letter alphabet. Many other reduced alphabets are not significantly correlated to folding rates. The finding suggests that not all amino acids and secondary structures are equally important for protein folding. The foldable sequence of a protein could be designed using at least 10 folding units, which can either promote or inhibit protein folding. Reducing alphabet cardinality without losing key folding kinetic information opens the door to potentially faster machine learning and data mining applications in protein structure prediction, sequence alignment and protein design. PMID:25641420

  8. Detection of COL III in Parchment by Amino Acid Analysis

    Vestergaard Poulsen Sommer, Dorte; Larsen, René

    2016-01-01

    Cultural heritage parchments made from the reticular dermis of animals have been subject to studies of deterioration and conservation by amino acid analysis. The reticular dermis contains a varying mixture of collagen I and III (COL I and III). When dealing with the results of the amino acid...... analyses, till now the COL III content has not been taken into account. Based on the available amino acid sequences we present a method for determining the amount of COL III in the reticular dermis of new and historical parchments calculated from the ratio of Ile/Val. We find COL III contents between 7 and...... 32 % in new parchments and between 0.2 and 40 % in the historical parchments. This is consistent with results in the literature. The varying content of COL III has a significant influence on the uncertainty of the amino acid analysis. Although we have not found a simple correlation between the COL...

  9. Visual detection of Ebola virus using reverse transcription loop-mediated isothermal amplification combined with nucleic acid strip detection.

    Xu, Changping; Wang, Hualei; Jin, Hongli; Feng, Na; Zheng, Xuexing; Cao, Zengguo; Li, Ling; Wang, Jianzhong; Yan, Feihu; Wang, Lina; Chi, Hang; Gai, Weiwei; Wang, Chong; Zhao, Yongkun; Feng, Yan; Wang, Tiecheng; Gao, Yuwei; Lu, Yiyu; Yang, Songtao; Xia, Xianzhu

    2016-05-01

    Ebola virus (species Zaire ebolavirus) (EBOV) is highly virulent in humans. The largest recorded outbreak of Ebola hemorrhagic fever in West Africa to date was caused by EBOV. Therefore, it is necessary to develop a detection method for this virus that can be easily distributed and implemented. In the current study, we developed a visual assay that can detect EBOV-associated nucleic acids. This assay combines reverse transcription loop-mediated isothermal amplification and nucleic acid strip detection (RT-LAMP-NAD). Nucleic acid amplification can be achieved in a one-step process at a constant temperature (58 °C, 35 min), and the amplified products can be visualized within 2-5 min using a nucleic acid strip detection device. The assay is capable of detecting 30 copies of artificial EBOV glycoprotein (GP) RNA and RNA encoding EBOV GP from 10(2) TCID50 recombinant viral particles per ml with high specificity. Overall, the RT-LAMP-NAD method is simple and has high sensitivity and specificity; therefore, it is especially suitable for the rapid detection of EBOV in African regions. PMID:26831931

  10. Nucleic acid hybridization techniques for the detection of bluetongue virus

    Virus isolation, antigen detection, and in situ hybridization were compared in their abilities to detect in cell culture, the five serotypes of bluetongue virus (BTV) occurring in the United States, serotypes 2, 10, 11, 13, and 17. For isolation, virus was propagated in baby hamster kidney (BHK-21) cell culture. For antigen detection, two techniques, indirect fluorescent-antibody (IFA) and enzyme immunocytoassay (EICA) were used. For in situ hybridization, a complementary DNA (cDNA) of the L3 RNA genome segment of BTV, serotype 17 (BTV-17) labeled with 35S was used as a group-specific probe. Virus isolation was the most sensitive technique, often detecting input virus and then detecting virus throughout the course of the study. IFA and EICA were of similar sensitivity and detected BTV antigen shortly after detection of virus by isolation. A direct-blot hybridization technique using a 32P-labeled, strand-specific RNA transcript probe was developed, optimized, and used to detect BTV in pools of infected Culicoides variipennis midges. The technique was able to detect as few as one infected Culicoides midge in a pool of 100 and as little as 3.5 log10 TCID50 per ml of virus. A sandwich hybridization technique was developed and used to detect BTV in pools of infected Culicoides variipennis midges. The sandwich hybridization technique used a single-stranded DNA catcher sequence bound to a solid support and a 32P-labeled, single-stranded RNA detector sequence. Sandwich hybridization was compared to direct blot hybridization using a strand-specific RNA transcript probe or a cDNA probe. Sandwich hybridization was able to detect as few as one infected Culicoides midge in a pool of 50; however, the technique was approximately tenfold less sensitive than direct blot hybridization

  11. Detecting to secret folded composite lamina package pairs in cores related slump dump structures and seismites with high resolution sampling of physical parameters

    Acar, Dursun; Cagatay, Namik; Feray Meydan, Aysegul; Eris, Kadir; Sari, Erol; Akcer, Sena; Makaroglu, Ozlem; Alkislar, Hakan; Biltekin, Demet; Nagehan Arslan, Tugce

    2016-04-01

    The core retrieved from Lake Van consists of seismites that were possibly deposited during the earthquakes around the Van region. Deformed parts of the core sediments display folded laminations that can be attributed to seismites. The problem arises that if the fold axis is deposited perpendicular to the liner and, if the hinge line is far enough, describing the true laminations might be impossible related to real age of basin evolution because extra laminae seem deposited to the area. Scientist must pay attention such problem that dating method like varve counting and basin evolution estimates can totally change due to extra laminae that explained before. For eliminate to wrong interpretations considering reversal reflected anomalies even with angularity effects to one package of pair can show significant difference than other symmetric one due to angle of the hinge line or soft sediment deformation. Considering the situation explained, p-wave is not enough to support the idea however; chemical analyses (x-ray florescence), ICP-MS (asdasd) analysis can provide appropriate results to identify laminae that appear on the limbs of the reversed micro folds. New easy designed extra U-Channel drive tray framework prepared by us. U-Channels are prepared well conditioned, saturated enough to well contact between sediment surface and plastic shield of u-channel samples from cores. Physical parameters are measured by Multi sensor core logger (MSCL) with high resolution step ratio fixed to 1mm. At the p- wave and gamma ray results, we observed together stair upwards form and reverse reflected downward data graphics, thus our interpretation of identifying the fold limbs are now visible. We understand that laminae packages are exactly the same. XRF and MSCL are totally supporting to origin of pairs generated after their sedimentation age with mechanical forces. For this reason, in this study, we attended to solve such problem to analyze deformed folded laminations that must be

  12. An Optical Test Strip for the Detection of Benzoic Acid in Food

    Fatimah Abu Bakar

    2011-07-01

    Full Text Available Fabrication of a test strip for detection of benzoic acid was successfully implemented by immobilizing tyrosinase, phenol and 3-methyl-2-benzothiazolinone hydrazone (MBTH onto filter paper using polystyrene as polymeric support. The sensing scheme was based on the decreasing intensity of the maroon colour of the test strip when introduced into benzoic acid solution. The test strip was characterized using optical fiber reflectance and has maximum reflectance at 375 nm. It has shown a highly reproducible measurement of benzoic acid with a calculated RSD of 0.47% (n = 10. The detection was optimized at pH 7. A linear response of the biosensor was obtained in 100 to 700 ppm of benzoic acid with a detection limit (LOD of 73.6 ppm. At 1:1 ratio of benzoic acid to interfering substances, the main interfering substance is boric acid. The kinetic analyses show that, the inhibition of benzoic is competitive inhibitor and the inhibition constant (Ki is 52.9 ppm. The activity of immobilized tyrosinase, phenol, and MBTH in the test strip was fairly sustained during 20 days when stored at 3 °C. The developed test strip was used for detection of benzoic acid in food samples and was observed to have comparable results to the HPLC method, hence the developed test strip can be used as an alternative to HPLC in detecting benzoic acid in food products.

  13. Clinical effect of nasolabial fold treated by hyaluronic acid with a multi-level injection%透明质酸多层次填充鼻唇沟凹陷的临床效果观察

    张晶晶; 林辉; 李红光; 文清; 王琳娜

    2016-01-01

    目的:探讨透明质酸多层次填充鼻唇沟凹陷的临床效果。方法:鼻唇沟凹陷明显的患者34例,给予多层次注射,注射后评价填充效果,记录不良反应。随访至治疗后6个月。结果:本组34例患者,鼻唇沟凹陷填充后的效果改善明显,患者对术后外观满意。注射后早期鼻唇沟处轻微疼痛、肿胀,一般在治疗后3d逐渐消退。未发现外形不对称、过敏、血管栓塞及皮肤坏死等并发症。填充效果可维持6个月以上。结论:透明质酸多层次填充鼻唇沟临床效果确切,操作简便,患者愿意接受,值得临床推广。%Objective To explore the clinical effect of nasolabial fold treated by hyaluronic acid with a multi-level injection. Methods 34 patients with varing degrees of nasolabial fold were treated by hyaluronic acid with a multi-level injection .The nasolabial fold iflling effect was evaluated after injection, adverse reactions were recorded,and follow-up was performed for 6 months.Results All nasolabial fold uplift in 34 patients was effective obviously.Patients were satisifed with their appearance. Maild pain and swelling after treatment disappeared at 3 days.No asymmetry, allergy, embolism or skin nerosis was observed. Augmention effect could last longer than 6 months.Conclusion Nasolabial fold augmention by hyaluronic acid with a multi-level injection is a safe and effective method, and it is easy to be accepted by patients.

  14. Bionic catalysis of porphyrin for electrochemical detection of nucleic acids

    Highlights: ► This is the first application of bionic catalysis of porphyrin as detection probe in bioanalysis. ► Porphyrin–DNA–gold nanoparticle probe is synthesized. ► Binding model between FeTMPyP and DNA is verified. ► The detection probe shows excellent electrocatalytic behaviors toward the reduction of O2. ► The biosensor exhibited good performance with wide linear range and high specificity. - Abstract: A novel electrochemical strategy was designed for the detection of DNA based on the bionic catalysis of porphyrin. The detection probe was prepared via the assembly of thiolated double strand DNA (dsDNA) with gold nanoparticles (AuNPs), and then interacted with cationic iron (III) meso-tetrakis (N-methylphyridinum-4-yl) porphyrin (FeTMPyP) via groove binding along the dsDNA surface. The resulting nanocomplex was characterized with transmission electron microscopy, UV–vis absorption and circular dichroism spectroscopy. The FeTMPyP–DNA–AuNPs probe on gold electrode demonstrated the excellent electrocatalytic behaviors toward the reduction of O2 due to the largely loading of FeTMPyP and good conductivity. Based on bionic catalysis of porphyrin for the reduction of O2, the resulting biosensor exhibited a good performance for the detection of DNA with a wide linear range from 1 × 10−12 to 1 × 10−8 mol L−1 and detection limit of 2.5 × 10−13 mol L−1 at the signal/noise of 3. More importantly, the biosensor presented excellent ability to discriminate the perfectly complementary target and the mismatched stand. This strategy could be conveniently extended for detection of other biomolecules. To the best of our knowledge, this is the first application of bionic catalysis of porphyrin as detection probe and opens new opportunities for sensitive detection of biorecognition events.

  15. STIS MAMA Fold Distribution

    Wheeler, Thomas

    2013-10-01

    The performance of MAMA microchannel plates can be monitored using a MAMA fold distribution procedure. The fold distribution provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of change in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the STIS MAMA Fold Distribution, Proposal 13149, as Cycle 20.

  16. Multiply Folded Graphene

    Kim, Kwanpyo; Lee, Zonghoon; Malone, Brad; Chan, Kevin T.; Alemán, Benjamín; Regan, William; Gannett, Will; Crommie, M. F.; Cohen, Marvin. L.; Zettl, A.

    2010-01-01

    The folding of paper, hide, and woven fabric has been used for millennia to achieve enhanced articulation, curvature, and visual appeal for intrinsically flat, two-dimensional materials. For graphene, an ideal two-dimensional material, folding may transform it to complex shapes with new and distinct properties. Here, we present experimental results that folded structures in graphene, termed grafold, exist, and their formations can be controlled by introducing anisotropic surface curvature dur...

  17. Nucleic Acid-based Detection of Bacterial Pathogens Using Integrated Microfluidic Platform Systems

    Carl A. Batt

    2009-05-01

    Full Text Available The advent of nucleic acid-based pathogen detection methods offers increased sensitivity and specificity over traditional microbiological techniques, driving the development of portable, integrated biosensors. The miniaturization and automation of integrated detection systems presents a significant advantage for rapid, portable field-based testing. In this review, we highlight current developments and directions in nucleic acid-based micro total analysis systems for the detection of bacterial pathogens. Recent progress in the miniaturization of microfluidic processing steps for cell capture, DNA extraction and purification, polymerase chain reaction, and product detection are detailed. Discussions include strategies and challenges for implementation of an integrated portable platform.

  18. CRISPR Spacer Arrays for Detection of Viral Signatures from Acidic Hot Springs

    Snyder, J. C.; Bateson, M. M.; Suciu, D.; Young, M. J.

    2010-04-01

    Viruses are the most abundant life-like entities on the planet Earth. Using CRISPR spacer sequences, we have developed a microarray-based approach to detecting viral signatures in the acidic hot springs of Yellowstone.

  19. A new method for quantitative determination of two uronic acids by CZE with direct UV detection.

    Xia, Yong-gang; Liang, Jun; Yang, Bing-you; Wang, Qiu-hong; Kuang, Hai-xue

    2011-09-01

    A new method using capillary zone electrophoresis was developed for the rapid quantification of two common uronic acids, galacturonic acid and glucuronic acid, based on utilization of an alkaline background electrolyte with reversed electroosmotic flow (EOF) within 16 min. The method relies on in-capillary reaction and direct UV detection at the wavelength 270 nm. The optimum electrolyte solution was prepared of 130 mm sodium hydroxide, 36 mm disodium hydrogen phosphate dihydrate and 0.5 mm cetyltrimethylammonium bromide. EOF was reversed to detect uronic acids and to improve the separation of neutral sugars. The established method was validated and the results showed good linearity, high precision and satisfactory sensitivity. The newly developed method was successfully applied to analyze galacturonic acid and glucuronic acid content in Forsythia suspensa polysaccharides. The method is fast since only sample hydrolysis and dilution are required in the sample preparation. PMID:21154888

  20. Detection of cancer cells using a peptide nanotube–folic acid modified graphene electrode

    Castillo, John J.; Svendsen, Winnie Edith; Rozlosnik, Noemi;

    2013-01-01

    This article describes the preparation of a graphene electrode modified with a new conjugate of peptide nanotubes and folic acid for the selective detection of human cervical cancer cells over-expressing folate receptors. The functionalization of peptide nanotubes with folic acid was confirmed by...... fluorescence microscopy and atomic force microscopy. The peptide nanotube–folic acid modified graphene electrode was characterized by scanning electron microscopy and cyclic voltammetry. The modification of the graphene electrode with peptide nanotube–folic acid led to an increase in the current signal. The......–folic acid modified electrode lowered the electron transfer resulting in a decrease in the measured current. A detection limit of 250 human cervical cancer cells per mL was obtained. Control experiments confirmed that the peptide nanotube–folic acid electrode specifically recognized folate receptors. The...

  1. Real-Time Detection of Deoxyribonucleic Acid Bases via their Negative Differential Conductance Signature

    Dragoman, Daniela; Dragoman, Mircea

    2009-01-01

    In this paper we present a method for the real-time detection of the bases of the deoxyribonucleic acid using their signatures in negative differential conductance measurements. The present methods of electronic detection of deoxyribonucleic acid bases are based on a statistical analysis because the electrical currents of the four bases are weak and do not differ significantly from one base to another. In contrast, we analyze a device that combines the accumulated knowledge in nanopore and sc...

  2. Detection of CIN by naked eye visualization after application of acetic acid.

    Londhe M; George S; Seshadri L

    1997-01-01

    A prospective study was undertaken to determine the sensitivity and specificity of acetic application to the cervix followed by naked eye visualization as a screening test for detection of cervical intraepithelial neoplasia. Three hundred and seventy two sexually active woman in the reproductive age group were studied. All the women underwent Papanicolaou test, acetic acid test and colposcopy. One hundred and seventy five woman were acetic acid test negative, 197 women were acetic acid test p...

  3. Cervical acid phosphatase detection: A guide to abnormal cells in cytology smear screening for cervical cancer

    Deb Prabal; Iyer Venkateswaran; Bhatla Neerja; Markovic O; Verma Kusum

    2008-01-01

    Background: Cervical acid phosphatase-Papanicolaou (CAP-PAP) test has recently been described for detection of acid phosphatase enzyme in abnormal squamous cells, and has been proposed as a biomarker-based technology for the screening of cervical cancer. Materials and Methods: Eighty-one consecutive cervical smears were subjected to routine Papanicolaou (Pap) staining as well as CAP-PAP, which combined cytochemical staining for acid phosphatase with modified Pap stain. Statistical evaluation ...

  4. Apparatus for point-of-care detection of nucleic acid in a sample

    Bearinger, Jane P.; Dugan, Lawrence C.

    2016-04-19

    Provided herein are methods and apparatus for detecting a target nucleic acid in a sample and related methods and apparatus for diagnosing a condition in an individual. The condition is associated with presence of nucleic acid produced by certain pathogens in the individual.

  5. Capillary electrophoresis of FITC labeled amino acids with laser-induced fluorescence detection

    党福全; 陈义

    1999-01-01

    FITC labeled amino acids have been separated using a home-huilt capillary electrophoresis with a laserinduced fluorescence detection (CE-LIF) system. Seventeen peaks can now be generated from the twenty common amino acids. The key conditions lie in the optimization of pH, buffer electrolytes and buffer additives.

  6. Interlaboratory comparison of ability to detect nucleic acid of Mycoplasma gallisepticum and Mycoplasma synoviae by PCR

    Hess, Michael; Neubauer, Claudia

    2007-01-01

    Abstract In recent years polymerase chain reaction (PCR) assays have become widely used as methods to confirm the presence of Mycoplasma gallisepticum and Mycoplasma synoviae in poultry flocks, but there has been limited standardization of the protocols used. Thirteen laboratories from 5 different countries participated in an interlaboratory comparison of detection of M. gallisepticum and M. synoviae DNA by PCR in samples that contained ten fold dilutions of these bacteria. The con...

  7. Quantitative detection of single amino acid polyrnorphisms by targeted proteornics

    Zhi-Duan Su; Jia-Rui Wu; Liang Sun; Dan-Xia Yu; Rong-Xia Li; Huai-Xing Li; Zhi-Jie Yu; Quan-Hu Sheng; Xu Lin; RongZeng

    2011-01-01

    Single-nucleotide polymorphisms (SNPs) are recognized as one kind of major genetic variants in population scale. However, polymorphisms at the proteome level in population scale remain elusive. In the present study, we named amino acid variances derived from SNPs within coding regions as single amino acid polymorphisms (SAPs) at the proteome level, and developed a pipeline of non-targeted and targeted proteomics to identify and quantify SAP peptides in human plasma. The absolute concentrations of three selected SAP-peptide pairs among 290 Asian individuals were measured by selected reaction monitoring (SRM) approach, and their associations with both obesity and diabetes were further analyzed. This work revealed that heterozygotes and homozygotes with various SAPs in a population could have different associations with particular traits. In addition, the SRM approach allows us for the first time to separately measure the absolute concentration of each SAP peptide in the heterozygotes, which also shows different associations with particular traits.%Single-nucleotide polymorphisms (SNPs) are recognized as one kind of major genetic variants in population scale.However,polymorphisms at the proteome level in population scale remain elusive.In the present study,we named amino acid variances derived from SNPs within coding regions as single amino acid polymorphisms (SAPs) at the proteome level,and developed a pipeline of non-targeted and targeted proteomics to identify and quantify SAP peptides in human plasma.The absolute concentrations of three selected SAP-peptide pairs among 290 Asian individuals were measured by selected reaction monitoring (SRM) approach,and their associations with both obesity and diabetes were further analyzed.This work revealed that heterozygotes and homozygotes with various SAPs in a population could have different associations with particular traits.In addition,the SRM approach allows us for the first time to separately measure the absolute

  8. Determination of free and total phenolic acids in plant-derived foods by HPLC with diode-array detection.

    Mattila, Pirjo; Kumpulainen, Jorma

    2002-06-19

    A high-performance liquid chromatographic (HPLC) method with diode-array detection (DAD) was used to identify and quantify free and total phenolic acids (m-hydroxybenzoic acid, p-hydroxybenzoic acid, protocatechuic acid, gallic acid, vanillic acid, syringic acid, o-coumaric acid, m-coumaric acid, p-coumaric acid, caffeic acid, ferulic acid, sinapic acid, chlorogenic acid, and ellagic acid) in plant foods. Free phenolic acids were extracted with a mixture of methanol and 10% acetic acid. Bound phenolic acids were liberated using first alkaline and then acid hydrolysis followed by extraction with diethyl ether/ethyl acetate (1:1). All fractions were quantified separately by HPLC. After HPLC quantification, results of alkali and acid hydrolysates were calculated to represent total phenolic acids. Ellagic acid was quantified separately after long (20 h) acid hydrolysis. The methods developed were effective for the determination of phenolic acids in plant foods. DAD response was linear for all phenolic acids within the ranges evaluated, with correlation coefficients exceeding 0.999. Coefficients of variation for 4-8 sample replicates were consistently below 10%. Recovery tests of phenolic acids were performed for every hydrolysis condition using several samples. Recoveries were generally good (mean >90%) with the exceptions of gallic acid and, in some cases, caffeic acid samples. PMID:12059140

  9. Design of acid-lead battery stage-of-charge detection system based on refractive index detection technology

    Chen, Junyao; Yang, Kecheng; Xia, Min; Li, Lei; Zeng, Xianjiang

    2015-10-01

    Based on optical total reflection critical Angle method, we have designed a refractive index measurement system. It adopted a divergent light source and a CCD camera as the occurrence and receiver of the signal. The divergent light source sent out a bunch of tapered beam, exposure to the interface of optical medium and sulfuric acid solution. Light intensity reflected from the interface could be detected by the CCD camera and then sent to the embedded system. In the DSP embedded system, we could obtain the critical edge position through the light intensity distribution curve and converted it to critical angle. Through experiment, we concluded the relation between liquid refractive index and the critical angle edge position. In this system, the detecting precision of the refractive index of sulfuric acid solution reached 10-4. Finally, through the conversion of the refractive index and density, we achieved high accuracy online measurement of electrolyte density in lead-acid battery.

  10. Understanding the folding process of synthetic polymers by small-molecule folding agents

    S G Ramkumar; S Ramakrishnan

    2008-01-01

    Two acceptor containing polyimides PDI and NDI carrying pyromellitic diimide units and 1,4,5,8-naphthalene tetracarboxy diimide units, respectively, along with hexa(oxyethylene) (EO6) segments as linkers, were prepared from the corresponding dianhydrides and diamines. These polyimides were made to fold by interaction with specifically designed folding agents containing a dialkoxynaphthalene (DAN) donor linked to a carboxylic acid group. The alkali-metal counter-ion of the donor carboxylic acid upon complexation with the EO6 segment brings the DAN unit in the right location to induce a charge-transfer complex formation with acceptor units in the polymer backbone. This two-point interaction between the folding agent and the polymer backbone leads to a folding of the polymer chain, which was readily monitored by NMR titrations. The effect of various parameters, such as structures of the folding agent and polymer, and the solvent composition, on the folding propensities of the polymer was studied.

  11. Structures of the first representatives of Pfam family PF06684 (DUF1185) reveal a novel variant of the Bacillus chorismate mutase fold and suggest a role in amino-acid metabolism

    Structures of the first representatives of PF06684 (DUF1185) reveal a Bacillus chorismate mutase-like fold with a potential role in amino-acid synthesis. The crystal structures of BB2672 and SPO0826 were determined to resolutions of 1.7 and 2.1 Å by single-wavelength anomalous dispersion and multiple-wavelength anomalous dispersion, respectively, using the semi-automated high-throughput pipeline of the Joint Center for Structural Genomics (JCSG) as part of the NIGMS Protein Structure Initiative (PSI). These proteins are the first structural representatives of the PF06684 (DUF1185) Pfam family. Structural analysis revealed that both structures adopt a variant of the Bacillus chorismate mutase fold (BCM). The biological unit of both proteins is a hexamer and analysis of homologs indicates that the oligomer interface residues are highly conserved. The conformation of the critical regions for oligomerization appears to be dependent on pH or salt concentration, suggesting that this protein might be subject to environmental regulation. Structural similarities to BCM and genome-context analysis suggest a function in amino-acid synthesis

  12. Stable dye-labelled oligonucleotide-nanoparticle conjugates for nucleic acid detection

    Barrett, Lee; Dougan, Jennifer A.; Faulds, Karen; Graham, Duncan

    2011-08-01

    Metallic nanoparticles functionalized with oligonucleotides are used for a number of nucleic acid detection strategies. However, oligonucleotide-nanoparticle conjugates suffer from a lack of stability when exposed to certain conditions associated with DNA detection assays. In this study, we report the synthesis of thiol and thioctic acid-modified oligonucleotide gold nanoparticle (OGNs) conjugates functionalized with a dye label and varying spacer groups. The thioctic acid-modified conjugates exhibit increased stability when treated with dithiothreitol (DTT) compared to the more commonly used thiol modification. When the dye labelled oligonucleotide nanoparticle conjugates are exposed to the same conditions there is a pronounced increase in the stability for both thioctic acid and thiol modified sequences. These results open up the possibility of simply using a dye label to enhance the stability of oligonucleotide-nanoparticle conjugates in DNA detection assays where the enhanced stability of the conjugate system can be advantageous in more complex biological environments.

  13. Determination of aliphatic organic acids by high-performance liquid chromatography with pulsed electrochemical detection.

    Casella, Innocenzo G; Gatta, Maria

    2002-01-01

    A new ion exclusion HPLC procedure accomplished with a pulsed electrochemical detection for the determination of several common aliphatic acids is described. A triple-step waveform of the applied potentials, based on the formation/inhibition of PtOH species on the electrode surface, is successfully used for sensitive detection of several aliphatic acids in flowing systems avoiding pre- or postcolumn derivatization and/or cleanup procedures. Under optimal chromatographic conditions (i.e., 50 mM HClO(4)) the proposed method allowed detection limits between 0.5 and 7 microM for all investigated acids, and the dynamic linear range spanned generally over 1 or 2 orders of magnitude. Determination of citric, malic, tartaric, lactic, formic, and acetic acids in several foods and beverages was performed, in approximately 15 min, without the necessity of any sample pretreatment. PMID:11754537

  14. JAWS: Just Add Water System - A device for detection of nucleic acids in Martian ice caps

    Hansen, Anders J.; Willerslev, Eske; Mørk, Søren;

    2002-01-01

    The design of a device for nucleic acid detection in the Martian ice caps is presented; the Just Add Water System (JAWS). It is based on fiber-optic PNA (peptide nucleic acid) light up probe random microsphere universal array technology. JAWS is designed to be part of a larger system with a...... regulation of pH and salt concentrations e.g. the MOD systems and could be installed on a planetary probe melting its way down the Martian ice caps e.g. the NASA Cryobot. JAWS can be used for detection of remains of ancient life preserved in the Martian ice as well as for detection of contamination brought...

  15. Real-Time PCR for detection of herpes simplex virus without nucleic acid extraction

    Klausner Jeffrey

    2006-06-01

    Full Text Available Abstract Background The speed and sensitivity of real-time polymerase chain reaction (PCR have made it a popular method for the detection of microbiological agents in both research and clinical specimens. For the detection and genotyping of herpes simplex virus (HSV in clinical specimens, real-time PCR has proven to be faster, more sensitive and safer than earlier methods which included isolation of the virus in cell culture followed by immunofluorescence microscopy. While PCR-based assays for HSV detection posses clear advantages over these earlier techniques, certain aspects of the PCR method remain onerous. The process of extraction and purification of nucleic acid from clinical specimens prior to PCR is particularly cumbersome. Nucleic acid extraction is expensive, time-consuming and provides a step whereby specimens can become contaminated prior to their analysis. Herein, we investigate the necessity of nucleic acid extraction from swab-based clinical specimens for HSV detection by real-time PCR. We find that nucleic acid extraction is unnecessary for specific and sensitive detection of HSV in clinical specimens using real-time PCR. Methods Prospective (n = 36 and retrospective (n = 21 clinical specimens from various anatomical sites were analyzed for the presence of herpes simplex virus 1 or 2 by real-time PCR using the RealArt HSV 1/2 LC PCR Kit. Specimens were analyzed by PCR both before and following automated nucleic acid extraction. PCR using extracted and unextracted specimens was also compared to cell culture as a means of detecting HSV. Results Detection of HSV 1/2 DNA in clinical specimens by real-time PCR did not require that the specimen be subjected to nucleic acid extraction/purification prior to analysis. Each specimen that was detectable by real-time PCR when analyzed in the extracted form was also detectable when analyzed in the unextracted form using the methods herein. The limit of detection of HSV-1 and HSV-2 particles

  16. Detection of exogenous citric acid in fruit juices by stable isotope ratio analysis.

    Jamin, Eric; Martin, Frédérique; Santamaria-Fernandez, Rebeca; Lees, Michèle

    2005-06-29

    A new method has been developed for measuring the D/H ratio of the nonexchangeable sites of citric acid by isotope ratio mass spectrometry (IRMS). Pure citric acid is transformed into its calcium salt and subsequently analyzed by pyrolysis-IRMS. The citric acid isolated from authentic fruit juices (citrus, pineapple, and red fruits) systematically shows higher D/H values than its nonfruit counterpart produced by fermentation of various sugar sources. The discrimination obtained with this simplified method is similar to that obtained previously by applying site specific isotopic fractionation-nuclear magnetic resonance (SNIF-NMR) to an ester derivative of citric acid. The combination of carbon 13 and deuterium measurements of extracted citric acid is proposed as a routine method for an optimum detection of exogenous citric acid in all kinds of fruit juices. PMID:15969486

  17. Ion-exclusion chromatography with conductimetric detection of aliphatic carboxylic acids on a weakly acidic cation-exchange resin by elution with benzoic acid-beta-cyclodextrin.

    Tanaka, Kazuhiko; Mori, Masanobu; Xu, Qun; Helaleh, Murad I H; Ikedo, Mikaru; Taoda, Hiroshi; Hu, Wenzhi; Hasebe, Kiyoshi; Fritz, James S; Haddad, Paul R

    2003-05-16

    In this study, an aqueous solution consisting of benzoic acid with low background conductivity and beta-cyclodextrin (beta-CD) of hydrophilic nature and the inclusion effect to benzoic acid were used as eluent for the ion-exclusion chromatographic separation of aliphatic carboxylic acids with different pKa values and hydrophobicity on a polymethacrylate-based weakly acidic cation-exchange resin in the H+ form. With increasing concentration of beta-cyclodextrin in the eluent, the retention times of the carboxylic acids decreased due to the increased hydrophilicity of the polymethacrylate-based cation-exchange resin surface from the adsorption of OH groups of beta-cyclodextrin. Moreover, the eluent background conductivity decreased with increasing concentration of beta-cyclodextrin in 1 mM benzoic acid, which could result in higher sensitivity for conductimetric detection. The ion-exclusion chromatographic separation of carboxylic acids with high resolution and sensitivity was accomplished successfully by elution with a 1 mM benzoic acid-10 mM cyclodextrin solution without chemical suppression. PMID:12830884

  18. Determination of Monochloroacetic Acid in Swimming Pool Water by Ion Chromatography-Conductivity Detection

    Maria Pythias B. Espino

    2013-02-01

    Full Text Available In this study, an analytical method involving ion chromatography with conductivity detection was developed and optimized for the determination of monochloroacetic acid in swimming pool water. The ion chromatographic method has a detection limit of 0.02 mg L-1 and linear range of 0.05 to 1.0 mg L-1 with correlation coeff icient of 0.9992. The method is reproducible with percent RSD of 0.052% (n=10. The recovery of monochloroacetic acid spiked in different water types (bottled, tap and swimming pool water ranged from 28 to 122%. In dilute solutions, chloride and bromide were simultaneously analyzed along with monochloroacetic acid using the optimized method. Chloride and bromide have detection limits of 0.01 to 0.05 mg L-1, respectively. The usefulness of the ion chromatographic method was demonstrated in the analysis of monochloroacetic acid in swimming pool water samples. In such highly-chlorinated samples, an Ag/H cartridge was used prior to the ion chromatographic determination so as to minimize the signal due to chloride ion. Monochloroacetic acid was detected in concentrations between 0.020 and 0.093 mg L-1 in three of the six swimming pool water samples studied. The presence of monochloroacetic acid in the swimming pool water samples suggests the possible occurrence of other disinfection by-products in these waters.

  19. Development of a bacterial bioassay for atrazine and cyanuric acid detection

    Anna eHUA

    2015-03-01

    Full Text Available The s-triazine herbicides are compounds which can disseminate into soils and water. Due to their toxic effects on living organisms, their concentrations in drinking water are legislated by WHO recommendations. Here we have developed for the first time, to the best of our knowledge, an alternative method for physicochemical quantification using two bioluminescent bacterial biosensors: E. coli SM003 for cyanuric acid detection and E. coli SM004 for both atrazine and cyanuric acid detection. The concentration of cyanuric acid detection for E. coli SM003 ranges from 7.83 µM to 2.89 mM, and for E. coli SM004 ranges from 0.22 µM to 15 µM. Moreover, atrazine detection by E. coli SM004 ranges from 1.08 µM to 15 µM. According to WHO recommendations, the cyanuric acid detection range is sensitive enough to discriminate between polluted and drinking water. Nevertheless, the detection of atrazine by E. coli SM004 is only applicable for high concentrations of contaminants.

  20. Protein folding, protein homeostasis, and cancer

    John H. Van Drie

    2011-01-01

    Proteins fold into their functional 3-dimensional structures from a linear amino acid sequence. In vitro this process is spontaneous; while in vivo it is orchestrated by a specialized set of proteins, called chaperones. Protein folding is an ongoing cellular process, as cellular proteins constantly undergo synthesis and degradation. Here emerging links between this process and cancer are reviewed. This perspective both yields insights into the current struggle to develop novel cancer chemotherapeutics and has implications for future chemotherapy discovery.

  1. The Folding of de Novo Designed Protein DS119 via Molecular Dynamics Simulations.

    Wang, Moye; Hu, Jie; Zhang, Zhuqing

    2016-01-01

    As they are not subjected to natural selection process, de novo designed proteins usually fold in a manner different from natural proteins. Recently, a de novo designed mini-protein DS119, with a βαβ motif and 36 amino acids, has folded unusually slowly in experiments, and transient dimers have been detected in the folding process. Here, by means of all-atom replica exchange molecular dynamics (REMD) simulations, several comparably stable intermediate states were observed on the folding free-energy landscape of DS119. Conventional molecular dynamics (CMD) simulations showed that when two unfolded DS119 proteins bound together, most binding sites of dimeric aggregates were located at the N-terminal segment, especially residues 5-10, which were supposed to form β-sheet with its own C-terminal segment. Furthermore, a large percentage of individual proteins in the dimeric aggregates adopted conformations similar to those in the intermediate states observed in REMD simulations. These results indicate that, during the folding process, DS119 can easily become trapped in intermediate states. Then, with diffusion, a transient dimer would be formed and stabilized with the binding interface located at N-terminals. This means that it could not quickly fold to the native structure. The complicated folding manner of DS119 implies the important influence of natural selection on protein-folding kinetics, and more improvement should be achieved in rational protein design. PMID:27128902

  2. The Folding of de Novo Designed Protein DS119 via Molecular Dynamics Simulations

    Moye Wang

    2016-04-01

    Full Text Available As they are not subjected to natural selection process, de novo designed proteins usually fold in a manner different from natural proteins. Recently, a de novo designed mini-protein DS119, with a βαβ motif and 36 amino acids, has folded unusually slowly in experiments, and transient dimers have been detected in the folding process. Here, by means of all-atom replica exchange molecular dynamics (REMD simulations, several comparably stable intermediate states were observed on the folding free-energy landscape of DS119. Conventional molecular dynamics (CMD simulations showed that when two unfolded DS119 proteins bound together, most binding sites of dimeric aggregates were located at the N-terminal segment, especially residues 5–10, which were supposed to form β-sheet with its own C-terminal segment. Furthermore, a large percentage of individual proteins in the dimeric aggregates adopted conformations similar to those in the intermediate states observed in REMD simulations. These results indicate that, during the folding process, DS119 can easily become trapped in intermediate states. Then, with diffusion, a transient dimer would be formed and stabilized with the binding interface located at N-terminals. This means that it could not quickly fold to the native structure. The complicated folding manner of DS119 implies the important influence of natural selection on protein-folding kinetics, and more improvement should be achieved in rational protein design.

  3. Water soluble and efficient amino acid Schiff base receptor for reversible fluorescence turn-on detection of Zn2+ ions: Quantum chemical calculations and detection of bacteria

    Subha, L.; Balakrishnan, C.; Natarajan, Satheesh; Theetharappan, M.; Subramanian, Balanehru; Neelakantan, M. A.

    2016-01-01

    An amino acid Schiff base (R) capable of recognizing Zn2+ ions selectively and sensitively in an aqueous medium was prepared and characterized. Upon addition of Zn2+ ions, the receptor exhibits fluorescence intensity enhancements (~ 40 fold) at 460 nm (quantum yield, Φ = 0.05 for R and Φ = 0.18 for R-Zn2+) and can be detected by naked eye under UV light. The receptor can recognize the Zn2+ (1.04 × 10- 8 M) selectively for other metal ions in the pH range of 7.5-11. The Zn2+ chelation with R decreases the loss of energy through non-radiative transition and leads to fluorescence enhancement. The binding mode of the receptor with Zn2+ was investigated by 1H NMR titration and further validated by ESI-MS. The elemental color mapping and SEM/EDS analysis were also used to study the binding of R with Zn2+. Density functional theory calculations were carried out to understand the binding mechanism. The receptor was applied as a microbial sensor for Escherichia coli and Staphylococcus aureus.

  4. Specific in situ hepatitis B viral double mutation (HBVDM) detection in urine with 60 copies ml(-1) analytical sensitivity in a background of 250-fold wild type without DNA isolation and amplification.

    Kirimli, Ceyhun E; Shih, Wei-Heng; Shih, Wan Y

    2015-03-01

    We have examined in situ detection of hepatitis B virus 1762T/1764A double mutation (HBVDM) in urine using a (Pb(Mg(1/3)Nb(2/3))O3)(0.65)(PbTiO3)(0.35) (PMN-PT) piezoelectric plate sensor (PEPS) coated with a 16-nucleotide (nt) probe DNA (pDNA) complementary to the HBVDM. The in situ mutation (MT) detection was carried out in a flow with the PEPS vertically situated at the center of the flow in a background of wild type (WT). For validation, this detection was followed by detection in the mixture of MT fluorescent reporter microspheres (FRMs) (MT FRMs) and WT FRMs that emitted different fluorescence colours and were designed to specifically bind to MT and WT, respectively. At 30 °C and 4 ml min(-1), a PEPS was shown to specifically detect HBVDM in situ with 60 copies ml(-1) analytical sensitivity in a background of clinically-relevant 250-fold more WT in 30 min without DNA isolation, amplification, or labelling as validated by the visualization of the captured MT FRMs and WT FRMs following FRM detection where the captured MT FRMs outnumbered the WT FRMs by a factor of 5 to 1. PMID:25599103

  5. Synthesis of structurally defined scaffolds for bivalent ligand display based on glucuronic acid anilides. The degree of tertiary amide isomerism and folding depends on the configuration of a glycosyl azide.

    Tosin, Manuela; Murphy, Paul V

    2005-05-13

    [structures: see text] Syntheses and structural analyses of bivalent carbohydrates based on anilides of glucuronic acid are described. Secondary anilides predominantly adopted the Z-anti structure; there is also evidence for population of the Z-syn isomer. Bivalent tertiary anilides displayed two signal sets in their NMR spectra, consistent with the presence of (i) a major isomer where both amides have E configurations (EE) and (ii) a minor isomer where one amide is E and the other Z (EZ). Qualitative NOE/ROE spectroscopic studies in solution support the proposal that the anti conformation is preferred for E amides. The crystal structure of one bivalent tertiary anilide showed E-anti and E-syn structural isomers; intramolecular carbohydrate-carbohydrate stacking was observed and mediated by carbonyl-pyranose, azide-azide, and pyranose-aromatic interactions. The EE to EZ isomer ratio, or the degree of folding, for tertiary amides, was greatest for a bivalent compound containing two alpha-glycosyl azide groups; this was enhanced in water, suggesting that hydrophobic interactions are partially but not wholly responsible. Computational methods predicted azide-aromatic (N...H-C interaction) and azide-azide interactions for folded isomers. The close contact of the azide and aromatic protons (N...H-C interaction) was observed upon examination of the close packing in the crystal structure of a related monomer. It is proposed that the alpha-azide group is more optimally aligned, compared to the beta-azide, to facilitate interaction and minimize the surface area of the hydrophobic groups exposed to water, and this leads to the increased folding. The alkylation of bivalent secondary anilides induces a switch from Z to E amide that alters the scaffold orientation. The synthesis of a bivalent mannoside, based on a secondary anilide scaffold, for investigation of mannose-binding receptor cross-linking and lattice formation is described. PMID:15876103

  6. Screen printing of nucleic acid detecting carbon electrodes.

    Dequaire, Murielle; Heller, Adam

    2002-09-01

    A large fraction of the presently mass-manufactured (> 10(8) units/year) electrochemical biosensors, used mostly by diabetic people to monitor their blood glucose levels, have screen-printed carbon working electrodes. An earlier study (Campbell, C. N., et al. Anal. Chem. 2002, 74, 158-162) showed that nucleic acids can be assayed at 1 nM concentrations by a sandwich-type amperometric method. The assay was performed with vitreous carbon working electrodes on which an electron-conducting polycationic redox polymer and avidin were coelectrodeposited. Because the rate of the electrodeposition increases with the surface density of the polycationic redox polymer, its practicality depends on pretreatment of the surface, which adds anionic functions. (Gao, Z., et al. Angew. Chem. Int. Ed. 2002, 41, 810-813). Here it is shown that the required conducting redox polymer films can be electrodeposited on potentially mass manufacturable electrodes made by screen-printing hydrophilic carbon inks on polyester sheets. The modified electrodes are made in two steps. First a polycationic electron-conducting redox polymer is cross-linked and electrodeposited by applying a negative potential. Next, an amine-terminated 20-base single-stranded oligonucleotide is electrodeposited by ligand-exchange. Both steps involve exchange of a labile inner sphere chloride ligand of the polymer-bound osmium-complex: Cross-linking and electrodeposition of the redox polymer result when inner-sphere chloride anions of the osmium complexes are exchanged by imidazole functions of neighboring chains. Incorporation of the oligonucleotide in the redox polymer results in the formation of a coordinative bond between the terminal amine (attached through a spacer to the oligonucleotide) and the osmium complex. In testing for the presence of a 38-base oligonucleotide, the analyte, in a 15- or 25-microL droplet of hybridization solution, is hybridized with and captured by the 20-base electrode-bound sequence; then

  7. α-Lipoic Acid Induced Elevated S-adenosylhomocysteine and Depleted S-adenosylmethionine

    Stabler, Sally P.; Sekhar, Jeevan; Allen, Robert H.; O'Neill, Heidi C.; White, Carl W.

    2009-01-01

    Lipoic Acid is a disulfhydryl-containing compound used in clinical medicine and in experimental models as an antioxidant. We developed a stable isotope dilution capillary gas chromatography/mass spectrometry assay for lipoic acid. We assayed a panel of the metabolites of transmethylation and transsulfuration 30 minutes after injecting lipoic acid 100 mg/kg in a rat model. Lipoic acid values rose 1000-fold in serum and 10-fold in liver. A methylated metabolite of lipoic acid was also detected ...

  8. Association of amino acids embedded in helium droplets detected by mass spectrometry

    Lalanne, Matthieu R.; Achazi, Georg; Reichwald, Sebastian; Lindinger, Albrecht

    2015-12-01

    Amino acids were embedded in helium droplets. The electron impact ionization allows for detecting positively charged glycine, valine, histidine, tryptophan and their principal fragments. Monomers and polymers with up to four amino acids are reported. Heterodimers of tryptophan and valine or histidine are observed as well as heterodimers of included fragments. The ability of these associations of molecules to form complexes with water is examined.

  9. Nucleic acid tests for the detection of alpha human papillomaviruses.

    Poljak, Mario; Cuzick, Jack; Kocjan, Boštjan J; Iftner, Thomas; Dillner, Joakim; Arbyn, Marc

    2012-11-20

    Testing for high-risk types of alpha human papillomaviruses (HPV) is an invaluable part of clinical guidelines for cervical carcinoma screening, management and treatment. In this comprehensive inventory of commercial tests for detection of alpha-HPV, we identified at least 125 distinct HPV tests and at least 84 variants of the original tests. However, only a small subset of HPV tests has documented clinical performance for any of the standard HPV testing indications. For more than 75% of HPV tests currently on the market, no single publication in peer-reviewed literature can be identified. HPV tests that have not been validated and lack proof of reliability, reproducibility and accuracy should not be used in clinical management. Once incorporated in the lab, it is essential that the whole procedure of HPV testing is subject to continuous and rigorous quality assurance to avoid sub-optimal, potentially harmful practices. Manufacturers of HPV tests are urged to put more effort into evaluating their current and future products analytically, using international standards, and for clinical applications, using clinically validated endpoints. To assist with analytical validation, the World Health Organization is developing international standards for HPV types other than HPV16 and HPV18 and is planning development of external quality control panels specifically designed to be used for performance evaluation of current and future HPV tests. There is a need for more competitively priced HPV tests, especially for resource-poor countries, and uniform test validation criteria based on international standards should enable issuing more competitive and fair tender notices for purchasing. Automation systems allowing large-scale testing, as well as further increases in clinical performance, are the main needs in the further improvement of HPV tests. This article forms part of a special supplement entitled "Comprehensive Control of HPV Infections and Related Diseases" Vaccine

  10. Programmable matter by folding

    Hawkes, E.; An, B.; Benbernou, N. M.; Tanaka, H.; Kim, S.; Demaine, E. D.; Rus, D.; Wood, R. J.

    2010-01-01

    Programmable matter is a material whose properties can be programmed to achieve specific shapes or stiffnesses upon command. This concept requires constituent elements to interact and rearrange intelligently in order to meet the goal. This paper considers achieving programmable sheets that can form themselves in different shapes autonomously by folding. Past approaches to creating transforming machines have been limited by the small feature sizes, the large number of components, and the associated complexity of communication among the units. We seek to mitigate these difficulties through the unique concept of self-folding origami with universal crease patterns. This approach exploits a single sheet composed of interconnected triangular sections. The sheet is able to fold into a set of predetermined shapes using embedded actuation. To implement this self-folding origami concept, we have developed a scalable end-to-end planning and fabrication process. Given a set of desired objects, the system computes an optimized design for a single sheet and multiple controllers to achieve each of the desired objects. The material, called programmable matter by folding, is an example of a system capable of achieving multiple shapes for multiple functions. PMID:20616049

  11. Detection of North American eastern and western equine encephalitis viruses by nucleic acid amplification assays.

    Lambert, Amy J; Martin, Denise A; Lanciotti, Robert S

    2003-01-01

    We have developed nucleic acid sequence-based amplification (NASBA), standard reverse transcription PCR (RT-PCR), and TaqMan nucleic acid amplification assays for the rapid detection of North American eastern equine encephalitis (EEE) and western equine encephalitis (WEE) viral RNAs from samples collected in the field and clinical samples. The sensitivities of these assays have been compared to that of virus isolation. While all three types of nucleic acid amplification assays provide rapid detection of viral RNAs comparable to the isolation of viruses in Vero cells, the TaqMan assays for North American EEE and WEE viral RNAs are the most sensitive. We have shown these assays to be specific for North American EEE and WEE viral RNAs by testing geographically and temporally distinct strains of EEE and WEE viruses along with a battery of related and unrelated arthropodborne viruses. In addition, all three types of nucleic acid amplification assays have been used to detect North American EEE and WEE viral RNAs from mosquito and vertebrate tissue samples. The sensitivity, specificity, and rapidity of nucleic acid amplification demonstrate the usefulness of NASBA, standard RT-PCR, and TaqMan assays, in both research and diagnostic settings, to detect North American EEE and WEE viral RNAs. PMID:12517876

  12. Acetic Acid Detection Threshold in Synthetic Wine Samples of a Portable Electronic Nose

    Miguel Macías Macías

    2012-12-01

    Full Text Available Wine quality is related to its intrinsic visual, taste, or aroma characteristics and is reflected in the price paid for that wine. One of the most important wine faults is the excessive concentration of acetic acid which can cause a wine to take on vinegar aromas and reduce its varietal character. Thereby it is very important for the wine industry to have methods, like electronic noses, for real-time monitoring the excessive concentration of acetic acid in wines. However, aroma characterization of alcoholic beverages with sensor array electronic noses is a difficult challenge due to the masking effect of ethanol. In this work, in order to detect the presence of acetic acid in synthetic wine samples (aqueous ethanol solution at 10% v/v we use a detection unit which consists of a commercial electronic nose and a HSS32 auto sampler, in combination with a neural network classifier (MLP. To find the characteristic vector representative of the sample that we want to classify, first we select the sensors, and the section of the sensors response curves, where the probability of detecting the presence of acetic acid will be higher, and then we apply Principal Component Analysis (PCA such that each sensor response curve is represented by the coefficients of its first principal components. Results show that the PEN3 electronic nose is able to detect and discriminate wine samples doped with acetic acid in concentrations equal or greater than 2 g/L.

  13. Detection of CIN by naked eye visualization after application of acetic acid.

    Londhe, M; George, S S; Seshadri, L

    1997-06-01

    A prospective study was undertaken to determine the sensitivity and specificity of acetic application to the cervix followed by naked eye visualization as a screening test for detection of cervical intraepithelial neoplasia. Three hundred and seventy two sexually active woman in the reproductive age group were studied. All the women underwent Papanicolaou test, acetic acid test and colposcopy. One hundred and seventy five woman were acetic acid test negative, 197 women were acetic acid test positive. The sensitivity of acetic acid test was 72.4%, specificity 54% and false negative rate 15.2%, as compared to papanicolaou test which had a sensitivity of 13.2%, specificity of 96.3% and false negative rate of 24.4%. The advantage of the acetic acid test lies in its easy technique, low cost and high sensitivity which are important factors for determining the efficacy of any screening programme in developing countries. PMID:9491668

  14. Isothermal amplification detection of nucleic acids by a double-nicked beacon.

    Shi, Chao; Zhou, Meiling; Pan, Mei; Zhong, Guilin; Ma, Cuiping

    2016-03-01

    Isothermal and rapid amplification detection of nucleic acids is an important technology in environmental monitoring, foodborne pathogen detection, and point-of-care clinical diagnostics. Here we have developed a novel method of isothermal signal amplification for single-stranded DNA (ssDNA) detection. The ssDNA target could be used as an initiator, coupled with a double-nicked molecular beacon, to originate amplification cycles, achieving cascade signal amplification. In addition, the method showed good specificity and strong anti-jamming capability. Overall, it is a one-pot and isothermal strand displacement amplification method without the requirement of a stepwise procedure, which greatly simplifies the experimental procedure and decreases the probability of contamination of samples. With its advantages, the method would be very useful to detect nucleic acids in point-of-care or field use. PMID:26706801

  15. DNA tetrahedron and star trigon nanostructures for target recycling detection of nucleic acid.

    Li, Yueran; Chen, Xifeng; Wang, Bidou; Liu, Guangxing; Tang, Yuguo; Miao, Peng

    2016-06-01

    Human immunodeficiency virus (HIV) is a retrovirus which attacks the human body's immune system and further leads to acquired immunodeficiency syndrome (AIDS). Nucleic acid detection is of great importance in the medical diagnosis of such diseases. Herein, we develop a simple and enzyme-free electrochemical method for the target recycling detection of nuclei acid. DNA tetrahedron and star trigon nanostructures are designed and constructed on the electrode interface for target capture and signal enrichment. This strategy is convenient and sensitive, with a limit of detection as low as 1 fM, and can also successfully distinguish single-base mismatched DNA. Therefore, the proposed method has a promising potential application for HIV DNA detection. PMID:27170090

  16. Vocal Fold Collision Modeling

    Granados, Alba; Brunskog, Jonas; Misztal, M. K.

    2015-01-01

    When vocal folds vibrate at normal speaking frequencies, collisions occurs. The numerics and formulations behind a position-based continuum model of contact is an active field of research in the contact mechanics community. In this paper, a frictionless three-dimensional finite element model of the...... vocal fold collision is proposed, which incorporates different procedures used in contact mechanics and mathematical optimization theories. The penalty approach and the Lagrange multiplier method are investigated. The contact force solution obtained by the penalty formulation is highly dependent on the...

  17. Simulations of Protein Folding

    Cahill, M; Cahill, K E; Cahill, Michael; Fleharty, Mark; Cahill, Kevin

    2000-01-01

    We have developed a simple, phenomenological, Monte-Carlo code that predicts the three-dimensional structure of globular proteins from the DNA sequences that define them. We have applied this code to two small proteins, the villin headpiece (1VII) and cole1 rop (1ROP). Our code folded the 36-residue villin headpiece to a mean rms distance of less than 5 A from its native structure as revealed by NMR; it folded a 56-residue fragment of the protein cole1 rop to within 11 A of its native structure. The denatured starting configurations of these two proteins were, respectively, 29 A and 55 A distant from their native structures.

  18. Folding Detonation Waves

    V. P. Singh

    1983-01-01

    Full Text Available Propagation of converging detonation waves in solid explosive is discussed. Whitham's method modified for solid explosives is used. Using folding coordinates, it is found that the strength of detonation waves increases as it moves towards the centre of implosion.

  19. Folding worlds between pages

    Meier, Matthias

    2010-01-01

    "We all remember pop-up books form our childhood. As fascinated as we were back then, we probably never imagined how much engineering know-how went into these books. Pop-up engineer Anton Radevsky has even managed to fold a 27-kilometre particle accelerator into a book" (4 pages)

  20. Simulations of protein folding

    We have developed a simple, phenomenological, Monte-Carlo code that predicts the three-dimensional structure of globular proteins from the DNA sequences that define them. We have applied this code to two small proteins, the villin headpiece (1VII) and colel rop (1ROP). Our code folds both proteins to within 5 A rms of their native structures

  1. Application of RP-HPLC in Detecting Content of Ferulic Acid in Fuke Qianjin Capsule

    Ming-zhi WANG

    2015-09-01

    Full Text Available Objective: To establish a method for the determination of ferulic acid content in Fuke Qianjin Capsule. Methods: Reversed phase high performance liquid chromatography (RP-HPLC was applied in this study, with the detection conditions as follows: chromatographic column: Boston green ODS·min-1, detection wavelength: 316 nm, and column temperature: 30℃. C18 (250 mm × 4.6 mm, 5 μm, mobile phase: methanol-0.1% phosphoric acid (25:75, flow velocity: 1.0 mLResults: Ferulic acid, whose sample size was 0.017760-0.10656 μg, was in favorable linear relationship with the integral value of peak area, with correlation coefficient r=0.9998. The average sample-injecting recovery rate and degree of precision (RSD were 97.6% (n=6 and 1.6%, respectively. The results of this study also showed that the specificity of RP-HPLC in this study was excellent; negative samples had no interference on chromatographic peak of target substance (ferulic acid; and RSD of accuracy, repeatability, stability and recovery rate were 1.1%, 1.4%, 1.2% and 1.6%, respectively.Conclusion: RP-HPLC is accurate, rapid, stable and convenient, so it can be used as an optimal method for the detection of ferulic acid content in Fuke Qianjin Capsule.

  2. Application of RP-HPLC in Detecting Content of Ferulic Acid in Fuke Qianjin Capsule

    WANG Ming-zhi

    2015-01-01

    Objective:To establish a method for the determination of ferulic acid content in Fuke Qianjin Capsule. Methods: Reversed phase high performance liquid chromatography (RP-HPLC) was applied in this study, with the detection conditions as follows: chromatographic column: Boston green ODS C18 (250 mm×4.6 mm, 5 μm), mobile phase: methanol-0.1% phosphoric acid (25:75), lfow velocity: 1.0 mL·min-1, detection wavelength: 316 nm, and column temperature: 30℃. Results:Ferulic acid, whose sample size was 0.017760-0.10656 μg, was in favorable linear relationship with the integral value of peak area, with correlation coefifcient r=0.9998. The average sample-injecting recovery rate and degree of precision (RSD) were 97.6% (n=6) and 1.6%, respectively. The results of this study also showed that the speciifcity of RP-HPLC in this study was excellent; negative samples had no interference on chromatographic peak of target substance (ferulic acid); and RSD of accuracy, repeatability, stability and recovery rate were 1.1%, 1.4%, 1.2% and 1.6%, respectively. Conclusion: RP-HPLC is accurate, rapid, stable and convenient, so it can be used as an optimal method for the detection of ferulic acid content in Fuke Qianjin Capsule.

  3. Capillary electrophoresis method with UV-detection for analysis of free amino acids concentrations in food.

    Omar, Mei Musa Ali; Elbashir, Abdalla Ahmed; Schmitz, Oliver J

    2017-01-01

    Simple and inexpensive capillary electrophoresis with UV-detection method (CE-UV) was optimized and validated for determination of six amino acids namely (alanine, asparagine, glutamine, proline, serine and valine) for Sudanese food. Amino acids in the samples were derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) prior to CE-UV analysis. Labeling reaction conditions (100mM borate buffer at pH 8.5, labeling reaction time 60min, temperature 70°C and NBD-Cl concentration 40mM) were systematically investigated. The optimal conditions for the separation were 100mM borate buffer at pH 9.7 and detected at 475nm. The method was validated in terms of linearity, limit of detection (LOD), limit of quantification (LOQ), precision (repeatability) (RSD%) and accuracy (recovery). Good linearity was achieved for all amino acids (r(2)>0.9981) in the concentration range of 2.5-40mg/L. The LODs in the range of 0.32-0.56mg/L were obtained. Recoveries of amino acids ranging from 85% to 108%, (n=3) were obtained. The validated method was successfully applied for the determination of amino acids for Sudanese food samples. PMID:27507479

  4. Robert Feulgen Prize Lecture 1995. New approaches to in situ detection of nucleic acids.

    Thiry, M

    1995-08-01

    The present paper reviews recent results obtained by different molecular biology-based, immunocytological approaches to the localization and identification of nucleic acids in sections of biological material. Examples of sensitive, high-resolution detection methods for RNA, DNA or specialized DNA regions are presented. Special emphasis is placed on the potential values and limitations of these new methods. PMID:8536076

  5. Monitoring the Hydrolysis of Olive Oil Catalyzed by Lipase via Acid Value Detection

    2007-01-01

    Hydrolysis of olive oil catalyzed by Candida lipolytica lipase was investigated. The relative concentration of the components in the product was determined by using high performance liquid chromatography(HPLC). Furthermore, a novel rapid method to detect the hydrolytic process of olive oil was developed based on the relationship between the acid value and the relative concentration of the different components.

  6. Detection of mitochondrial deoxyribonucleic acid alterations in urine from urothelial cell carcinoma patients.

    Dasgupta, S.; Shao, C.; Keane, T.E.; Duberow, D.P.; Mathies, R.A.; Fisher, P.B.; Kiemeney, L.A.L.M.; Sidransky, D.

    2012-01-01

    Our study aims at understanding the timing and nature of mitochondrial deoxyribonucleic acid (mtDNA) alterations in urothelial cell carcinoma (UCC) and their detection in urine sediments. The entire 16.5 kb mitochondrial genome was sequenced in matched normal lymphocytes, tumor and urine sediments f

  7. Analysis of Ascorbic Acid in Single Human Neutrophils by Electrochemical Detection

    2002-01-01

    Ascorbic acid in individual human neutrophils was determined by capillary zone electrophoresis with electrochemical detection. In order to overcome the influence of the adsorption of the substances in cells on the inner surface wall of the capillary on the migration time and the number of theoretical plates, a procedure for treating capillaries has been described.

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

    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. PMID:27506344

  9. Single and double stranded DNA detection using locked nucleic acid (LNA) functionalized nanoparticles

    McKenzie, Fiona; Stokes, Robert; Faulds, Karen; Graham, Duncan

    2008-08-01

    Gold and silver nanoparticles functionalized with oligonucleotides can be used for the detection of specific sequences of DNA. We show that gold nanoparticles modified with locked nucleic acid (LNA) form stronger duplexes with a single stranded DNA target and offer better discrimination against single base pair mismatches than analogous DNA probes. Our LNA nanoparticle probes have also been used to detect double stranded DNA through triplex formation, whilst still maintaining selectivity for only complementary targets. Nanoparticle conjugates embedded with suitable surface enhanced resonance Raman scattering (SERRS) labels have been synthesized enabling simultaneous detection and identification of multiple DNA targets.

  10. A paper based microfluidic device for easy detection of uric acid using positively charged gold nanoparticles.

    Kumar, Anand; Hens, Abhiram; Arun, Ravi Kumar; Chatterjee, Monosree; Mahato, Kuldeep; Layek, Keya; Chanda, Nripen

    2015-03-21

    A paper based microfluidic device is fabricated that can rapidly detect very low concentrations of uric acid (UA) using 3,5,3',5'-tetramethyl benzidine (TMB), H2O2 and positively charged gold nanoparticles ((+)AuNPs). In the presence of (+)AuNPs, H2O2 reacts with TMB to produce a bluish-green colour which becomes colourless on reaction with UA. This colorimetric method can detect as low as 8.1 ppm of UA within <20 minutes on white filter paper. This technique provides an alternative way for UA detection. PMID:25655365

  11. Modelling of lateral fold growth and fold linkage: Applications to fold-and-thrust belt tectonics

    Grasemann, Bernhard; Schmalholz, Stefan

    2013-04-01

    We use a finite element model to investigate the three-dimensional fold growth and interference of two initially isolated fold segments. The most critical parameter, which controls the fold linkage mode, is the phase difference between the laterally growing fold hinge lines: 1) "Linear-linkage" yields a sub-cylindrical fold with a saddle at the location where the two initial folds linked. 2) "Oblique-linkage" produces a curved fold resembling a Type II refold structure. 3) "Oblique-no-linkage" results in two curved folds with fold axes plunging in opposite directions. 4) "Linear-no-linkage" yields a fold train of two separate sub-cylindrical folds with fold axes plunging in opposite directions. The transition from linkage to no-linkage occurs when the fold separation between the initially isolated folds is slightly larger than one half of the low-amplitude fold wavelength. The model results compare well with previously published plasticine analogue models and can be directly applied to the investigation of fold growth history in fold-and-thust belts. An excellent natural example of lateral fold linkage is described from the Zagros fold-and-thrust belt in the Kurdistan Region of Iraq. The fold growth in this region is not controlled by major thrust faults but the shortening of the Paleozoic to Cenozoic passive margin sediments of the Arabian plate occurred mainly by detachment folding. The sub-cylindrical anticlines with hinge-parallel lengths of more than 50 km have not developed from single sub-cylindrical embryonic folds but they have merged from different fold segments that joined laterally during fold amplification and lateral fold growth. Linkage points are marked by geomorphological saddle points which are structurally the lowermost points of antiforms and points of principal curvatures with opposite sign. Linkage points can significantly influence the migration of mineral-rich fluids and hydrocarbons and are therefore of great economic importance.

  12. Estimation of vocal fold plane in 3D CT images for diagnosis of vocal fold abnormalities.

    Hewavitharanage, Sajini; Gubbi, Jayavardhana; Thyagarajan, Dominic; Lau, Ken; Palaniswami, Marimuthu

    2015-01-01

    Vocal folds are the key body structures that are responsible for phonation and regulating air movement into and out of lungs. Various vocal fold disorders may seriously impact the quality of life. When diagnosing vocal fold disorders, CT of the neck is the commonly used imaging method. However, vocal folds do not align with the normal axial plane of a neck and the plane containing vocal cords and arytenoids does vary during phonation. It is therefore important to generate an algorithm for detecting the actual plane containing vocal folds. In this paper, we propose a method to automatically estimate the vocal fold plane using vertebral column and anterior commissure localization. Gray-level thresholding, connected component analysis, rule based segmentation and unsupervised k-means clustering were used in the proposed algorithm. The anterior commissure segmentation method achieved an accuracy of 85%, a good estimate of the expert assessment. PMID:26736949

  13. Self-assembled monolayer based electrochemical nucleic acid sensor for Vibrio cholerae detection

    Nucleic acid sensor has been fabricated by immobilization of thiolated (5' end) single stranded deoxyribonucleic acid probe (ssDNA-SH) onto gold (Au) coated glass electrode for Vibriocholerae detection. This ssDNA-SH/Au bioelectrode characterized using atomic force microscopy (AFM),Fourier transforms infrared spectroscopy (FT-IR) and electrochemical technique, has been used for hybridization detection of genomic DNA (dsDNA/Au). This ssDNA-SH/Au bioelectrode can specifically detect up to 100- 500 ng/μL genomic DNA of Vibriocholeare within 60 s of hybridization time at 25°C by cyclic voltammetry (CV) using methylene blue (MB) as electro-active DNA hybridization indicator. The value of sensitivity of the dsDNA/Au electrode has been determined as 0.027μA/ng cm−2 with regression coefficient as 0.978. This DNA bioelectrode is stable for about 4 months when stored at 4°C.

  14. A micro E-DNA sensor for selective detection of dopamine in presence of ascorbic acid

    朱丹; 李敏; 王丽华; 左小磊

    2015-01-01

    In this paper, a novel method for selectively detection of dopamine (DA) in the interference of ascorbic acid (AA) is described. A nanometer-sized gold flower microelectrode (NGFME) is prepared by flame-etching and electrochemical deposition. The electrode tip was characterized by scanning electron microscope (SEM). The NGFME is sized at about 100 µm and dimensions of thorns of the electrode were in nanometers. By modifying with DA aptamer on the surface, the prepared aptasensor can selectively detect DA even in the presence of high concentration AA. Experimental results show that this NGFME has no response to AA. As a comparison, the carbon fiber electrode without DA aptamer modification is unable to effectively detect DA in the presence of AA. The NGFME is easy-to-prepare, selective and sensitive for DA detection down to 25 µM. The electrode can be expected to detect DA in vivo and in real biological samples.

  15. Application of a Mass Spectrometric Approach to Detect the Presence of Fatty Acid Biosynthetic Phosphopeptides.

    Lau, Benjamin Yii Chung; Clerens, Stefan; Morton, James D; Dyer, Jolon M; Deb-Choudhury, Santanu; Ramli, Umi Salamah

    2016-04-01

    The details of plant lipid metabolism are relatively well known but the regulation of fatty acid production at the protein level is still not understood. Hence this study explores the importance of phosphorylation as a mechanism to control the activity of fatty acid biosynthetic enzymes using low and high oleic acid mesocarps of oil palm fruit (Elaeis guineensis variety of Tenera). Adaptation of neutral loss-triggered tandem mass spectrometry and selected reaction monitoring to detect the neutral loss of phosphoric acid successfully found several phosphoamino acid-containing peptides. These peptides corresponded to the peptides from acetyl-CoA carboxylase and 3-enoyl-acyl carrier protein reductase as identified by their precursor ion masses. These findings suggest that these enzymes were phosphorylated at 20th week after anthesis. Phosphorylation could have reduce their activities towards the end of fatty acid biosynthesis at ripening stage. Implication of phosphorylation in the regulation of fatty acid biosynthesis at protein level has never been reported. PMID:26993480

  16. Protein folding and cosmology

    González-Diáz, P F

    1997-01-01

    Protein denaturing induced by supercooling is interpreted as a process where some or all internal symmetries of the native protein are spontaneously broken. Hence, the free-energy potential corresponding to a folding-funnel landscape becomes temperature-dependent and describes a phase transition. The idea that deformed vortices could be produced in the transition induced by temperature quenching, from native proteins to unfolded conformations is discussed in terms of the Zurek mechanism that implements the analogy between vortices, created in the laboratory at low energy, and the cosmic strings which are thought to have been left after symmetry breaking phase transitions in the early universe. An experiment is proposed to test the above idea which generalizes the cosmological analogy to also encompass biological systems and push a step ahead the view that protein folding is a biological equivalent of the big bang.

  17. Ab initio RNA folding

    RNA molecules are essential cellular machines performing a wide variety of functions for which a specific three-dimensional structure is required. Over the last several years, the experimental determination of RNA structures through x-ray crystallography and NMR seems to have reached a plateau in the number of structures resolved each year, but as more and more RNA sequences are being discovered, the need for structure prediction tools to complement experimental data is strong. Theoretical approaches to RNA folding have been developed since the late nineties, when the first algorithms for secondary structure prediction appeared. Over the last 10 years a number of prediction methods for 3D structures have been developed, first based on bioinformatics and data-mining, and more recently based on a coarse-grained physical representation of the systems. In this review we are going to present the challenges of RNA structure prediction and the main ideas behind bioinformatic approaches and physics-based approaches. We will focus on the description of the more recent physics-based phenomenological models and on how they are built to include the specificity of the interactions of RNA bases, whose role is critical in folding. Through examples from different models, we will point out the strengths of physics-based approaches, which are able not only to predict equilibrium structures, but also to investigate dynamical and thermodynamical behavior, and the open challenges to include more key interactions ruling RNA folding. (topical review)

  18. Ab initio RNA folding

    Cragnolini, Tristan; Derreumaux, Philippe; Pasquali, Samuela

    2015-06-01

    RNA molecules are essential cellular machines performing a wide variety of functions for which a specific three-dimensional structure is required. Over the last several years, the experimental determination of RNA structures through x-ray crystallography and NMR seems to have reached a plateau in the number of structures resolved each year, but as more and more RNA sequences are being discovered, the need for structure prediction tools to complement experimental data is strong. Theoretical approaches to RNA folding have been developed since the late nineties, when the first algorithms for secondary structure prediction appeared. Over the last 10 years a number of prediction methods for 3D structures have been developed, first based on bioinformatics and data-mining, and more recently based on a coarse-grained physical representation of the systems. In this review we are going to present the challenges of RNA structure prediction and the main ideas behind bioinformatic approaches and physics-based approaches. We will focus on the description of the more recent physics-based phenomenological models and on how they are built to include the specificity of the interactions of RNA bases, whose role is critical in folding. Through examples from different models, we will point out the strengths of physics-based approaches, which are able not only to predict equilibrium structures, but also to investigate dynamical and thermodynamical behavior, and the open challenges to include more key interactions ruling RNA folding.

  19. Determination of acrylamide and acrylic acid by isocratic liquid chromatography with pulsed electrochemical detection.

    Casella, Innocenzo G; Pierri, Marianna; Contursi, Michela

    2006-02-24

    The electrochemical behaviour of the polycrystalline platinum electrode towards the oxidation/reduction of short-chain unsaturated aliphatic molecules such as acrylamide and acrylic acid was investigated in acidic solutions. Analytes were separated by reverse phase liquid chromatographic and quantified using a pulsed amperometric detection. A new two-step waveform, is introduced for detection of acrylamide and acrylic acid. Detection limits (LOD) of 20 nM (1. 4 microg/kg) and 45 nM (3.2 microg/kg) were determined in water solutions containing acrylamide and acrylic acid, respectively. Compared to the classical three-step waveform, the proposed two-step waveform shows favourable analytical performance in terms of LOD, linear range, precision and improved long-term reproducibility. The proposed analytical method combined with clean-up procedure accomplished by Carrez clearing reagent and subsequent extraction with a strong cation exchanger cartridges (SPE), was successfully used for the quantification of low concentrations of acrylamide in foodstuffs such as coffee and potato fries. PMID:16426623

  20. Separation and detection of amino acid metabolites of Escherichia coli in microbial fuel cell with CE.

    Wang, Wei; Ma, Lihong; Lin, Ping; Xu, Kaixuan

    2016-07-01

    In this work, CE-LIF was employed to investigate the amino acid metabolites produced by Escherichia coli (E. coli) in microbial fuel cell (MFC). Two peptides, l-carnosine and l-alanyl-glycine, together with six amino acids, cystine, alanine, lysine, methionine, tyrosine, arginine were separated and detected in advance by a CE-LIF system coupled with a homemade spontaneous injection device. The injection device was devised to alleviate the effect of electrical discrimination for analytes during sample injection. All analytes could be completely separated within 8 min with detection limits of 20-300 nmol/L. Then this method was applied to analyze the substrate solution containing amino acid metabolites produced by E. coli. l-carnosine, l-alanyl-glycine, and cystine were used as the carbon, nitrogen, and sulfur source for the E. coli culture in the MFC to investigate the amino acid metabolites during metabolism. Two MFCs were used to compare the activity of metabolism of the bacteria. In the sample collected at the running time 200 h of MFC, the amino acid methionine was discovered as the metabolite with the concentrations 23.3 μg/L. PMID:27121957

  1. Selective extraction by dissolvable (nitriloacetic acid-nickel)-layered double hydroxide coupled with reaction with potassium thiocyanate for sensitive detection of iron(III).

    Tang, Sheng; Chang, Yuepeng; Shen, Wei; Lee, Hian Kee

    2016-07-01

    A highly selective method has been proposed for the determination of iron cation (Fe(3+)). (Nitriloacetic acid-nickel)-layered double hydroxide ((NTA-Ni)-LDH) was successfully synthesized and used as dissolvable sorbent in dispersive solid-phase extraction to pre-concentrate and separate Fe(3+) from aqueous phase. Since Fe(3+) has a larger formation constant with NTA compared to Ni(2+), subsequently ion exchange occurred when (NTA-Ni)-LDH was added to the sample solution. The resultant (NTA-Fe)-LDH sol was isolated and transferred in an acidic medium containing potassium thiocyanate (KSCN). Since (NTA-Fe)-LDH could be dissolved in acidic conditions, Fe(3+)was released and reacted with SCN(-) to form an Fe-SCN complex. The resulting product was measured by ultraviolet-visible spectrometry for quantitative detection of Fe(3+). Extraction factors, including sample pH, reaction pH, extraction temperature, extraction time, reaction time and concentration of KSCN were optimized. This method achieved a low limit of detection of 15.2nM and a good linear range from 0.05 to 50μM (r(2)=0.9937). A nearly 18-fold enhancement of signal intensity was achieved after selective extraction. The optimized conditions were validated by applying the method to determine Fe(3+) in seawater samples. PMID:27154694

  2. Use of radiolabeled nucleic acid probes for the detection and identification of infectious organisms

    Nucleic acid probes can be used to detect and identify a wide variety of infectious organisms from bacteria to helminthic worms. Commercially-produced kits are now available for the direct detection of Legionella species in clinical samples, for identification of Mycobacterium species growing on solid media or in broth, and for detection of a variety of other bacteria and viruses directly in clinical samples or tissues. 32P and 125I, which are used to label probes to high specific activity, continue to be used in commercially-prepared kits. 32P-labeling remains as one of the most sensitive means of labeling nucleic acids probes. The advantages of using probes to detect and identify infectious agents include the rapid generation of results, the ability to broaden the spectrum of organisms identified by a clinical laboratory, and the high specificity of probe-based tests. The disadvantages of using probes include the high cost of the assays, the additional equipment required, and the inability to perform ancillary tests, such as antimicrobial susceptibility testing, which require having a pure culture of the organism available. None-the-less, nucleic acid probes already are having a very positive impact on the diagnosis of infectious diseases both in the industrialized and developing nations. (author). 11 refs

  3. Electrochemical detection of uric acid via uricase-immobilized graphene oxide.

    Omar, Muhamad Nadzmi; Salleh, Abu Bakar; Lim, Hong Ngee; Ahmad Tajudin, Asilah

    2016-09-15

    Measurement of the uric acid level in the body can be improved by biosensing with respect to the accuracy, sensitivity and time consumption. This study has reported the immobilization of uricase onto graphene oxide (GO) and its function for electrochemical detection of uric acid. Through chemical modification of GO using 1-ethyl-3-(dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide (NHS) as cross-linking reagents, the enzyme activity of the immobilized uricase was much comparable to the free enzyme with 88% of the activity retained. The modified GO-uricase (GOU) was then subjected to electrocatalytic detection of uric acid (UA) via cyclic voltammetry (CV). For that reason, a glassy carbon electrode (GCE) was modified by adhering the GO along with the immobilized uricase to facilitate the redox reaction between the enzyme and the substrate. The modified GOU/GCE outperformed a bare electrode through the electrocatalytic activity with an amplified electrical signal for the detection of UA. The electrocatalytic response showed a linear dependence on the UA concentration ranging from 0.02 to 0.49 mM with a detection limit of 3.45 μM at 3σ/m. The resulting biosensor also exhibited a high selectivity towards UA in the presence of other interference as well as good reproducibility. PMID:27402177

  4. Fluorescent probes for detection of picric acid explosive: A greener approach

    Green materials with advantages of low cost and high sensitivity are important from the perspective of human health, environment and homeland security. Herein, we have reported two cost effective modified biomaterials as fluorophores for detection of Picric acid in aqueous state. The biomaterials Scutellarin–Hispiduloside and Curcumin have been modified with green solvent glycerol for Picric acid detection in aqueous solution. The limit of detection for Picric acid by Scutellarin–Hispiduloside–glycerol and Curcumin–glycerol are 9.1×10−8 M and 6.03×10−8 M respectively. These luminescence based sensors have also been able to detect Picric acid in real samples with high efficiency. The fluorescence quenching efficiency of Scutellarin–Hispiduloside–glycerol has been found to be 99% while that for Curcumin–glycerol, it is 88.9% for 0.5 µM Picric acid in aqueous state. In both the cases, the quenching is governed by FRET between the fluorophore and the quencher and the FRET efficiency has been found to be 0.968 and 0.792 respectively. In addition, both the systems show excellent selectivity towards PA in presence of other nitroaromatic compounds and are also statistically accessible. The utilization of readily available cheap biomaterials without using multistep protocol for synthesis and devoid of any kind of sophisticated equipments for the processs further enhances the utility of the method. - Highlights: • Environmentally benign systems – Scutellarin, Hispiduloside and curcumin with green solvent glycerol – have been used for Picric acid sensing. • The method is simple and cost effective with a detection limit for CIG and CG found to be 9.1×10−8 M and 6.03×10−8 M of PA respectively. • Both the sensing systems were found to be highly selective for Picric acid in the presence of structurally similar compounds. • The quenching occurs by FRET between the fluorophore and the quencher and the FRET efficiency is determined. • The

  5. Fluorescent probes for detection of picric acid explosive: A greener approach

    Chakravarty, Sudesna; Gogoi, Bedanta; Sen Sarma, Neelotpal, E-mail: neelot@iasst.gov.in

    2015-09-15

    Green materials with advantages of low cost and high sensitivity are important from the perspective of human health, environment and homeland security. Herein, we have reported two cost effective modified biomaterials as fluorophores for detection of Picric acid in aqueous state. The biomaterials Scutellarin–Hispiduloside and Curcumin have been modified with green solvent glycerol for Picric acid detection in aqueous solution. The limit of detection for Picric acid by Scutellarin–Hispiduloside–glycerol and Curcumin–glycerol are 9.1×10{sup −8} M and 6.03×10{sup −8} M respectively. These luminescence based sensors have also been able to detect Picric acid in real samples with high efficiency. The fluorescence quenching efficiency of Scutellarin–Hispiduloside–glycerol has been found to be 99% while that for Curcumin–glycerol, it is 88.9% for 0.5 µM Picric acid in aqueous state. In both the cases, the quenching is governed by FRET between the fluorophore and the quencher and the FRET efficiency has been found to be 0.968 and 0.792 respectively. In addition, both the systems show excellent selectivity towards PA in presence of other nitroaromatic compounds and are also statistically accessible. The utilization of readily available cheap biomaterials without using multistep protocol for synthesis and devoid of any kind of sophisticated equipments for the processs further enhances the utility of the method. - Highlights: • Environmentally benign systems – Scutellarin, Hispiduloside and curcumin with green solvent glycerol – have been used for Picric acid sensing. • The method is simple and cost effective with a detection limit for CIG and CG found to be 9.1×10−8 M and 6.03×10−8 M of PA respectively. • Both the sensing systems were found to be highly selective for Picric acid in the presence of structurally similar compounds. • The quenching occurs by FRET between the fluorophore and the quencher and the FRET efficiency is determined

  6. Sequence-specific nucleic acid detection from binary pore conductance measurement

    Esfandiari, Leyla; Monbouquette, Harold G.; Jacob J. Schmidt

    2012-01-01

    We describe a platform for sequence-specific nucleic acid (NA) detection utilizing a micropipette tapered to a 2 μm diameter pore and 3 μm diameter polystyrene beads to which uncharged peptide nucleic acid (PNA) probe molecules have been conjugated. As the target NAs hybridize to the complementary PNA-beads, the beads acquire negative charge and become electrophoretically mobile. An applied electric field guides these NA-PNA-beads toward the pipette tip, which they obstruct, leading to an ind...

  7. Non Invasive Microwave Sensor for the Detection of Lactic Acid in Cerebrospinal Fluid (CSF)

    This research involves the use of a low power microwave sensor for analysis of lactic acid in cerebrospinal fluid (CSF), an indicator of neurological impairment during aortic aneurysm surgery which could provide the basis for improved treatment regimes and better quality of care with more efficient use of resources. This paper presents initial work using standard lactate curves in water followed by lactate in synthetic CSF. A multi-modal spectral signature has been defined for lactate, forming the basis for subsequent development of microwave sensor platform that is able to detect concentrations of lactic acid in CSF of volumes less than 1ml.

  8. Nucleic acid detection based on the use of microbeads: a review

    Microbead-based technologies represent elegant and versatile approaches for highly parallelized quantitative multiparameter assays. They also form the basis of various techniques for detection and quantification of nucleic acids and proteins. Nucleic acid-based methods include hybridization assays, solid-phase PCR, sequencing, and trapping assays. Microbead assays have been improved in the past decades and are now important tools in routine and point-of-care diagnostics as well as in life science. Its advances include low costs, low workload, high speed and high-throughput automation. The potential of microbead-based assays therefore is apparent, and commercial applications can be found in the detection and discrimination of single nucleotide polymorphism, of pathogens, and in trapping assays. This review provides an overview on microbead-based platforms for biosensing with a main focus on nucleic acid detection (including amplification strategies and on selected probe systems using fluorescent labeling). Specific sections cover chemical properties of microbeads, the coupling of targets onto solid surfaces, microbead probe systems (mainly oligonucleotide probes), microbead detection schemes (with subsections on suspension arrays, microfluidic devices, and immobilized microbeads), quantification of nucleic acids, PCR in solution and the detection of amplicons, and methods for solid-phase amplification. We discuss selected trends such as microbead-coupled amplification, heterogeneous and homogenous DNA hybridization assays, real-time assays, melting curve analysis, and digital microbead assays. We finally discuss the relevance and trends of the methods in terms of high-level multiplexed analysis and their potential in diagnosis and personalized medicine. (author)

  9. Determination of some aliphatic carboxylic acids in anaerobic digestion process waters by ion-exclusion chromatography with conductimetric detection on a weakly acidic cation-exchange resin column.

    Ito, Kazuaki; Takayama, Yohichi; Ikedo, Mikaru; Mori, Masanobu; Taoda, Hiroshi; Xu, Qun; Hu, Wenzhi; Sunahara, Hiroshi; Hayashi, Tsuneo; Sato, Shinji; Hirokawa, Takeshi; Tanaka, Kazuhiko

    2004-06-11

    The determination of seven aliphatic carboxylic acids, formic, acetic, propionic, isobutyric, n-butyric, isovaleric and n-valeric acids in anaerobic digestion process waters was examined using ion-exclusion chromatography with conductimetric detection. The analysis of these biologically important carboxylic acids is necessary as a measure for evaluating and controlling the process. The ion-exclusion chromatography system employed consisted of polymethacrylate-based weakly acidic cation-exchange resin columns (TSKgel OApak-A or TSKgel Super IC-A/C). weakly acidic eluent (benzoic acid), and conductimetric detection. Particle size and cation-exchange capacity were 5 microm and 0.1 meq./ml for TSKgel OApak-A and 3 microm and 0.2 meq./ml for TSKgel Super IC-A/C, respectively. A dilute eluent (1.0-2.0 mM) of benzoic acid was effective for the high resolution and highly conductimetric detection of the carboxylic acids. The good separation of isobutyric and n-butyric acids was performed using the TSKgel Super IC-A/C column (150 mm x 6.0 mm i.d. x 2). The simple and good chromatograms were obtained by the optimized ion-exclusion chromatography conditions for real samples from mesophilic anaerobic digestors, thus the aliphatic carboxylic acids were successfully determined without any interferences. PMID:15250416

  10. Paper-based fluorescence resonance energy transfer assay for directly detecting nucleic acids and proteins.

    Li, Hua; Fang, Xueen; Cao, Hongmei; Kong, Jilie

    2016-06-15

    Paper-based fluorescence resonance energy transfer assay (FRET) is gaining great interest in detecting macro-biological molecule. It is difficult to achieve conveniently and fast detection for macro-biological molecule. Herein, a graphene oxide (GO)-based paper chip (glass fiber) integrated with fluorescence labeled single-stranded DNA (ssDNA) for fast, inexpensive and direct detection of biological macromolecules (proteins and nucleic acids) has been developed. In this paper, we employed the Cy3/FAM-labeled ssDNA as the reporter and the GO as quencher and the original glass fiber paper as data acquisition substrates. The chip which was designed and fabricated by a cutting machine is a miniature biosensor that monitors fluorescence recovery from resonance energy transfer. The hybridization assays and fluorescence detection were all simplified, and the surface of the chip did not require immobilization or washing. A Nikon Eclipse was employed as excited resource and a commercial digital camera was employed for capturing digital images. This paper-based microfluidics chip has been applied in the detection of proteins and nucleic acids. The biosensing capability meets many potential requirements for disease diagnosis and biological analysis. PMID:26807518

  11. [Detection of sorbic acid in food by homemade micro-spectrometer analytical system].

    Chuan, Na; Xu, Yi; Chen, Gang; Wen, Zhong-quan; He, Li; Wen, Zhi-yu

    2012-08-01

    A homemade micro-spectrometer analytical system was developed for the quantitative determination of the sorbic acid in the food based on the photometric principle. And with the standard addition method it was applied to eliminate the interference coming from the food substrate. The detecting result illustrated a good relativity in the range of 0-10.0 mg x L(-1) with the linear correlation coefficient of 0.9989, and the sample recovery was 99.2%-99.5% with RSD of 0.58%. The micro-spectrometer analysis system has shown potential prospective application in the fields of rapid and high performance detection for food additives. PMID:23156802

  12. A Single-Tube Nucleic Acid Extraction, Amplification, and Detection Method Using Aluminum Oxide

    Dames, Shale; Bromley, L. Kathryn; Herrmann, Mark; Elgort, Marc; Erali, Maria; Smith, Roger; Voelkerding, Karl V.

    2006-01-01

    A disposable 0.2-ml polymerase chain reaction (PCR) tube modified with an aluminum oxide membrane (AOM) has been developed for the extraction, amplification, and detection of nucleic acids. To assess the dynamic range of AOM tubes for real-time PCR, quantified herpes simplex virus (HSV) DNA was used to compare AOM tubes to standard PCR tubes. AOM PCR tubes used for amplification and detection of quantified HSV-1 displayed a crossing threshold (CT) shift 0.1 cycles greater than PCR tube contro...

  13. Detection of Thiobacillus ferrooxidans in acid mine environments by indirect fluorescent antibody staining.

    Apel, W A; Dugan, P R; Filppi, J A; Rheins, M S

    1976-07-01

    An indirect fluorescent antibody (FA) staining technique was developed for the rapid detection of Thiobacillus ferrooxidans. The specificity of the FA stain for T. ferrooxidans was demonstrated with both laboratory and environmental samples. Coal refuse examined by scanning electron microscopy exhibited a rough, porous surface, which was characteristically covered by water-soluble crystals. Significant numbers of T. ferrooxidans were detected in the refuse pores. A positive correlation between numbers of T. ferrooxidans and acid production in coal refuse in the laboratory was demonstrated with the FA technique. PMID:61736

  14. Detection of anhydrous hydrochloric acid, HCl, in IRC+10216 with the Herschel SPIRE and PACS spectrometers

    Cernicharo, J.; Decin, L.; Barlow, M.J.; Agundez, M.; Royer, P.; Wesson, R.; Polehampton, E.T.; Beck, E; Blommaert, J. A. D. L.; Daniel, F; De Meester, W.; Exter, K. M.; Feuchtgruber, H.; Gear, W. K.; Goicoechea, J. R.

    2010-01-01

    We report on the detection of anhydrous hydrochloric acid (hydrogen chlorine, HCl) in the carbon-rich star IRC+10216 using the spectroscopic facilities onboard the Herschel satellite. Lines from J=1-0 up to J=7-6 have been detected. From the observed intensities, we conclude that HCl is produced in the innermost layers of the circumstellar envelope with an abundance relative to H2 of 5x10^-8 and extends until the molecules reach its photodissociation zone. Upper limits to the column densities...

  15. Autoinducer-2 detection among commensal oral streptococci is dependent on pH and boric acid.

    Cuadra, Giancarlo A; Frantellizzi, Ashley J; Gaesser, Kimberly M; Tammariello, Steven P; Ahmed, Anika

    2016-07-01

    Autoinducer-2, considered a universal signaling molecule, is produced by many species of bacteria; including oral strains. Structurally, autoinducer-2 can exist bound to boron (borated autoinducer-2). Functionally, autoinducer-2 has been linked to important bacterial processes such as virulence and biofilm formation. In order to test production of autoinducer-2 by a given bacterial strain, a bioassay using marine bioluminescent bacteria Vibrio harveyi as a reporter for autoinducer-2 has been designed. We hypothesize that pH adjustment and addition of boron are required for optimal bioluminescence and accurate autoinducer-2 detection. Using this reporter strain we tested autoinducer-2 activity from two oral commensal species, Streptococcus gordonii DL1 and Streptococcus oralis 34. Spent broth was collected and adjusted to pH 7.5 and supplemented with boric acid prior to measuring autoinducer- 2 activity. Results show that low pH inhibits bioluminescence of the reporter strain, but pH 7.5 allows for bioluminescence induction and proper readings of autoinducer-2 activity. Addition of boric acid also has a positive effect on bioluminescence allowing for a more sensitive detection of autoinducer-2 activity. Our data suggests that although autoinducer-2 is present in spent broth, low pH and/or low levels of boric acid become an obstacle for proper autoinducer-2 detection. For proper autoinducer-2 detection, we propose a protocol using this bioassay to include pH adjustment and boric acid addition to spent broth. Studies on autoinducer-2 activity in several bacteria species represent an important area of study as this universal signaling molecule is involved in critical bacterial phenotypes such as virulence and biofilm formation. PMID:27350615

  16. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy

    Ramla Gary; Giovani Carbone; Gia Petriashvili; Maria Penelope De Santo; Riccardo Barberi

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra ...

  17. Monoclonal antibodies targeted to alpha-oligonucleotides. Characterisation and application in nucleic acid detection.

    Cros, P.; Kurfürst, R; Allibert, P; Battail, N; Piga, N; Roig, V; Thuong, N T; Mandrand, B; Hélène, C

    1994-01-01

    The aim of the present study was to test the antigenicity of alpha-deoxyribonucleotides in order to develop a new tool for the detection of nucleic acid sequences for use in diagnostic applications. We describe four monoclonal antibodies (Mabs) which recognize alpha-deoxyribonucleotides. Two were raised against a poly(alpha-dT) sequence and specifically recognized the alpha-dT nucleotide. Two were raised against a sequence containing all four common nucleotides as alpha-nucleotides and, surpr...

  18. Amino acid biosignatures : implications for the detection of extinct or extant microbial communities on Mars

    Aubrey, Andrew D.

    2008-01-01

    Investigations of Mars have recently found strong geochemical evidence for the presence of standing bodies of water early in the planet’s history. It still remains to be discovered whether organic compounds exist on Mars, a question which concurrent international scientific efforts are focused on for future in situ planetary missions. Amino acids are at the core of terrestrial biochemistry, ubiquitous in terrestrial life, and are easily detectable via highly advanced instrumentation with pa...

  19. A Randomized, Evaluator-Blinded, Split-Face Comparison Study of the Efficacy and Safety of a Novel Mannitol Containing Monophasic Hyaluronic Acid Dermal Filler for the Treatment of Moderate to Severe Nasolabial Folds

    Kim, Byung Wook; Moon, Ik Jun; Yun, Woo Jin; Chung, Bo Young; Kim, Sang Duck

    2016-01-01

    Background Mannitol containing monophasic filler with higher crosslinking has not been well studied for moderate and severe nasolabial fold (NLF) correction. Objective To compare the efficacy and safety of a novel mannitol containing hyaluronic acid (HA) filler (HA-G) with biphasic HA filler (HA-P) for moderate and severe NLF correction. Methods Thirteen subjects with symmetric moderate to severe NLF received HA-G (in one NLF) and HA-P (in other NLF) and were evaluated for 24 weeks. Results At both 12 and 24 weeks, the mean improvement in Genzyme 6-point grading scale from baseline was significantly greater in the side of face that was treated with HA-G than HA-P (1.96±0.91 vs. 1.54±0.73 at week 12; p=0.044, 1.88±0.78 vs. 1.3±0.79 at week 24; p=0.027, respectively). At 12 weeks, the mean Global Aesthetic Improvement Scale score was 2.92±0.93 for HA-G and 2.31±0.95 for HA-P (p=0.008). Both fillers were well tolerated. Conclusion The HA filler HA-G provides better efficacy and similar local tolerability compared with HA-P in 6 months following treatment for moderate and severe NLF. PMID:27274627

  20. Low-ppm-Level colorimetric acid detection using gold nanoparticles with electro-steric stabilization.

    Bae, Doo Ri; Lee, You-Jin; Lee, Sung Woo; Han, Young-Kyu; Yoon, Jae-Sik; Lee, Ji-Hyun; Lee, Sang-Gil; Chang, Ki Soo; Yi, Gi-Ra; Lee, Gaehang

    2014-12-01

    Electro-sterically stabilized gold suspensions were employed in a colorimetric system for the detection of strong acid in water. Using oleyamine and oleic acid as steric stabilizer in 1,2-dichlorobenzene, hydrophobic gold nanoparticles were first synthesized by a reduction reaction of gold salts and were then transferred into water with a cationic surfactant. When the hydrochlo- ric acid solution higher than critical concentration was injected, particles were quickly aggregated and precipitated, creating a clear solution from the colored suspension. The particles were stable against chemical etching by corrosive ion such as chloride. Critical concentration was dependent of the size and concentration of the particles. The minimum concentration of dramatic color change was at 5 ppm level of hydrochloric acid, in which the largest colloidal gold nanoparticles (54 nm) were used. Furthermore, because of their steric repulsive soft layer on particles, particles could be reused for further detection experiments after regeneration by the simple pH-neutralization and washing process. PMID:25971086

  1. Highly-sensitive Detection of Salvianolic Acid B using Alumina Microfibers-modified Electrode

    Alumina microfibers with porous structures were prepared through hydrothermal reaction, and then used to modify the surface of carbon paste electrode (CPE). After modification with alumina microfibers, the electrochemical activity of CPE was found to be greatly improved. On the surface of alumina microfibers-modified CPE, the oxidation peak current of salvianolic acid B, a main bioactive compound in Danshen with anti-oxidative and anti-inflammatory effects, was remarkably increased compared with that on the bare CPE surface. The influences of pH value, amount of alumina microfibers and accumulation time were studied. Based on the strong signal amplification effects of alumina microfibers, a novel electrochemical method was developed for the detection of salvianolic acid B. The linear range was from 5 μg L-1 to 0.3 mg L-1, and the detection limit was 2 μg L-1 (2.78 nM) after 1-min accumulation. The new method was successfully used to detect salvianolic acid B in ShuangDan oral liquid samples, and the recovery was over the range from 97.4% to 102.9%

  2. BMAA detected as neither free nor protein bound amino acid in blue mussels.

    Rosén, Johan; Westerberg, Erik; Schmiedt, Sebastian; Hellenäs, Karl-Erik

    2016-01-01

    The results of this study imply that β-methylamino-alanine (BMAA) obtained from extracts of blue mussels from the Swedish west coast is neither free nor protein bound. The results were obtained by separation (precipitation and ultrafiltration) of low and high molecular weight compounds from neutral extracts of blue mussels, and treatment of these extracts with low and high concentrations of acids, varying time and temperature. The main portion of BMAA was obtained from the low molecular weight fraction, released or formed at 95 °C in dilute acids. The measured amount of BMAA did not increase by strong acid treatment. Lysine was used as reference and was only released at significant amounts when treating the high molecular weight fraction with concentrated acid. The results also indicated that breakage of peptide bonds was not involved in the formation/release of BMAA in these extracts unless any BMAA peptide bond would be significantly more susceptible to dilute acid than e.g. the monitored lysine peptide bond. BMAA was measured using isotope dilution and detection of the underivatized compound by HILIC-UHPLC-MS/MS (Hydrophilic Interaction Liquid Chromatography, Ultra-High Performance Liquid Chromatography, tandem Mass Spectrometry). The findings might add to the understanding of conflicting data in the literature regarding the occurrence of BMAA, and have implications for studies on possible biomagnification of BMAA in the food chain and bioavailability from food. PMID:26577502

  3. Carbonic acid revisited: Vibrational spectra, energetics and the possibility of detecting an elusive molecule

    Stefan E. Huber

    2012-09-01

    Full Text Available We calculate harmonic frequencies of the three most abundant carbonic acid conformers. For this, different model chemistries are investigated with respect to their benefits and shortcomings. Based on these results we use perturbation theory to calculate anharmonic corrections at the ωB97XD/aug-cc-pVXZ, X = D, T, Q, level of theory and compare them with recent experimental data and theoretical predictions. A discrete variable representation method is used to predict the large anharmonic contributions to the frequencies of the stretching vibrations in the hydrogen bonds in the carbonic acid dimer. Moreover, we re-investigate the energetics of the formation of the carbonic acid dimer from its constituents water and carbon dioxide using a high-level extrapolation method. We find that the ωB97XD functional performs well in estimating the fundamental frequencies of the carbonic acid conformers. Concerning the reaction energetics, the accuracy of ωB97XD is even comparable to the high-level extrapolation method. We discuss possibilities to detect carbonic acid in various natural environments such as Earth's and Martian atmospheres.

  4. Detection of amino acid neurotransmitters by surface enhanced Raman scattering and hollow core photonic crystal fiber

    Tiwari, Vidhu S.; Khetani, Altaf; Monfared, Ali Momenpour T.; Smith, Brett; Anis, Hanan; Trudeau, Vance L.

    2012-03-01

    The present work explores the feasibility of using surface enhanced Raman scattering (SERS) for detecting the neurotransmitters such as glutamate (GLU) and gamma-amino butyric acid (GABA). These amino acid neurotransmitters that respectively mediate fast excitatory and inhibitory neurotransmission in the brain, are important for neuroendocrine control, and upsets in their synthesis are also linked to epilepsy. Our SERS-based detection scheme enabled the detection of low amounts of GLU (10-7 M) and GABA (10-4 M). It may complement existing techniques for characterizing such kinds of neurotransmitters that include high-performance liquid chromatography (HPLC) or mass spectrography (MS). This is mainly because SERS has other advantages such as ease of sample preparation, molecular specificity and sensitivity, thus making it potentially applicable to characterization of experimental brain extracts or clinical diagnostic samples of cerebrospinal fluid and saliva. Using hollow core photonic crystal fiber (HC-PCF) further enhanced the Raman signal relative to that in a standard cuvette providing sensitive detection of GLU and GABA in micro-litre volume of aqueous solutions.

  5. Antisera production to detect indoleacetic acid in cultures of plant-growth promoting bacteria

    Rabbit polyclonal antisera against indoleacetic acid (IAA) bound to nitrocellulose membrane were obtained, which exhibited a high titer and specificity. The dot immunobinding technique with colloidal gold was used to detect auxin production by several strains belonging to Gluconacetobacter, Herbaspirillum, Azospirillum, Pseudomonas, Burkholderia and Bacillus genera, using culture supernatants as antigens. Moreover, auxin production was quantified by the Salkowski's method to corroborate the previous results. It was found that that all the studied microorganisms produce IAA and the feasibility of using these antisera to detect the metabolite was confirmed. Taking into account the potentialities of plant growth promoting bacteria as biofertilizers, the use of these antisera for a rapid and easy detection of IAA in bacteria associated with important crops is thus recommended.

  6. Improved detection of coastal acid sulfate soil hotspots through biomonitoring of metal(loid) accumulation in water lilies (Nymphaea capensis).

    Stroud, Jacqueline L; Collins, Richard N

    2014-07-15

    Anthropogenically disturbed coastal acid sulfate soils along the east coast of Australia, and worldwide, periodically result in the discharge of acid waters containing high concentrations of metals. Identifying priority sites (hotspots) within a catchment for acid sulfate soil remediation activities typically involves long-term monitoring of drainwater chemistry, including the capture of data on unpredictable rain-induced groundwater discharge events. To improve upon this monitoring approach, this study investigated using the water lily (Nymphaea capensis) as a biomonitor of drainage waters to identify hotspots in three acid sulfate soil impacted catchments (83 km(2)) in north-eastern New South Wales, Australia. In one catchment where the location of hotspots was known, water lily lamina concentrations of a suite of metal(loid)s were significantly (pcatchment-scale water lily sampling program undertaken in catchments with unidentified hotspots revealed within catchment variation of plant metal concentrations up to 70-fold. High resolution maps produced from these results, therefore, provided strong evidence for the location of potential hotspots which were confirmed with measurements of drainwater chemistry during rain-induced groundwater discharge events. Median catchment lily accumulation was ca. 160 mg Al kg(-1) and 1,300 mg Fe kg(-1), with hotspots containing up to 6- and 10-fold higher Al and Fe concentrations. These findings suggest that biomonitoring with N. capensis can be an important tool to rapidly identify priority sites for remediation in acid sulfate soil impacted landscapes. PMID:24805963

  7. Folding Gravitational-Wave Interferometers

    Sanders, J R

    2016-01-01

    The sensitivity of kilometer-scale terrestrial gravitational wave interferometers is limited by mirror coating thermal noise. We explore the effect of folding the arm cavities of such interferometers. While simple folding alone does not reduce the mirror coating thermal noise, it makes the folding mirror the critical mirror, opening up a variety of design and upgrade options.

  8. Novel derivatization technique for the detection of phenoxyacetic acid herbicides by gas chromatography-mass spectrometry

    Complete text of publication follows. The analytical detection of phenoxyacetic acid type herbicides (2,4-D, dichloprop, MCPA, etc.) from environmental samples is often a problem in instrumental analysis, as these compounds containing free carboxylic groups require chemical derivatisation prior to gas chromatographic (GC) methods. Commonly used derivatising agents include diazomethane and pentachlorobenzyl bromide, the former being toxic and explosive, the latter is being most suitable for GC with electron capture detector. In order to detect these compounds in GC coupled with mass spectrometry (GC-MS), a novel method for the formation of stable chemical derivatives from the acidic herbicides prior to GC analysis was developed. Earlier we have applied silylcarbamates for the derivatisation of chlorophenols (A. Kovacs et al., J. Chromat. A, 1194 (1) (2008) 139-142.), and this procedure was extended now to the silylation of phenoxyacetic acid type compounds. We studied the reactions between commercially available trimethylsilyl N,N-dimethylcarbamate and mecoprop, MCPA, dichlorprop, 2,4-D, MCPB 2,4,5-T, 2,4,5-TP and 2,4-DB, respectively. Beside the trimethylsilylated derivatives of the target compounds the corresponding tert-butyldimethylsilyl derivatives were also prepared in order to enhance hydrolytic stability and improve mass spectrometric properties. For that purpose the corresponding tert-butyldimethylsilyl N,N-dimethylcarbamate agent was prepared, allowing detection limits at ppb level. The reactions proved to be fast and the reagent excess and by-products were proven not to interfere with the target compounds. In accordance with the expectations tert-butyldimethylsilyl esters of phenoxyacetic acids surpassed the corresponding trimethylsilyl analogues both in stability and detectability. The procedure was also compared with the results obtained by commercial silylating agent BSTFA and certain alkylating agents (TMAH, Me3SOH, trimethylsilyldiazomethane

  9. Novel bioluminescent quantitative detection of nucleic acid amplification in real-time.

    Olga A Gandelman

    Full Text Available BACKGROUND: The real-time monitoring of polynucleotide amplification is at the core of most molecular assays. This conventionally relies on fluorescent detection of the amplicon produced, requiring complex and costly hardware, often restricting it to specialised laboratories. PRINCIPAL FINDINGS: Here we report the first real-time, closed-tube luminescent reporter system for nucleic acid amplification technologies (NAATs enabling the progress of amplification to be continuously monitored using simple light measuring equipment. The Bioluminescent Assay in Real-Time (BART continuously reports through bioluminescent output the exponential increase of inorganic pyrophosphate (PPi produced during the isothermal amplification of a specific nucleic acid target. BART relies on the coupled conversion of inorganic pyrophosphate (PPi produced stoichiometrically during nucleic acid synthesis to ATP by the enzyme ATP sulfurylase, and can therefore be coupled to a wide range of isothermal NAATs. During nucleic acid amplification, enzymatic conversion of PPi released during DNA synthesis into ATP is continuously monitored through the bioluminescence generated by thermostable firefly luciferase. The assay shows a unique kinetic signature for nucleic acid amplifications with a readily identifiable light output peak, whose timing is proportional to the concentration of original target nucleic acid. This allows qualitative and quantitative analysis of specific targets, and readily differentiates between negative and positive samples. Since quantitation in BART is based on determination of time-to-peak rather than absolute intensity of light emission, complex or highly sensitive light detectors are not required. CONCLUSIONS: The combined chemistries of the BART reporter and amplification require only a constant temperature maintained by a heating block and are shown to be robust in the analysis of clinical samples. Since monitoring the BART reaction requires only a

  10. A liquid chromatography-mass spectrometric method for the detection of cyclic β-amino fatty acid lipopeptides.

    Urajová, Petra; Hájek, Jan; Wahlsten, Matti; Jokela, Jouni; Galica, Tomáš; Fewer, David P; Kust, Andreja; Zapomělová-Kozlíková, Eliška; Delawská, Kateřina; Sivonen, Kaarina; Kopecký, Jiří; Hrouzek, Pavel

    2016-03-18

    Bacterial lipopeptides, which contain β-amino fatty acids, are an abundant group of bacterial secondary metabolites exhibiting antifungal and/or cytotoxic properties. Here we have developed an LC-HRMS/MS method for the selective detection of β-amino fatty acid containing cyclic lipopeptides. The method was optimized using the lipopeptides iturin A and puwainaphycin F, which contain fatty acids of similar length but differ in the amino acid composition of the peptide cycle. Fragmentation energies of 10-55eV were used to obtain the amino acid composition of the peptide macrocycle. However, fragmentation energies of 90-130eV were used to obtain an intense fragment specific for the β-amino fatty acid (CnH2n+2N(+)). The method allowed the number of carbons and consequently the length of the β-amino fatty acid to be estimated. We identified 21 puwainaphycin variants differing in fatty acid chain in the crude extract of cyanobacterium Cylindrospermum alatosporum using this method. Analogously 11 iturin A variants were detected. The retention time of the lipopeptide variants showed a near perfect linear dependence (R(2)=0.9995) on the length of the fatty acid chain in linear separation gradient which simplified the detection of minor variants. We used the method to screen 240 cyanobacterial strains and identified lipopeptides from 8 strains. The HPLC-HRMS/MS method developed here provides a rapid and easy way to detecting novel variants of cyclic lipopeptides. PMID:26893022

  11. A novel silicon based mags-biosensor for nucleic acid detection by magnetoelectronic transduction

    Maria Eloisa Castagna

    2015-12-01

    Full Text Available We developed a novel silicon biosensor based on magnetoelectronic transduction (MAGS for nucleic acid detection. The mags-biosensor is a planar device composed by a primary micro-coil, and two secondary coils which produce a differential voltage due to the induced magnetic field. The presence of magnetic material over one of the secondary coils causes variations of induced magnetic field density that in turn results in a total output voltage different from zero. The voltage variation, therefore, is a measure of the amount of magnetic material present in the active zone. A device sensitivity of 5.1 mV/ng and a resolution of 0.008 ng have been observed. The biosensor also presents a micro-heater and a thermal sensor respectively to set and read-out the chip temperature: this aspect enables the device to be used for several biochemical applications that need temperature control and activation such for example nucleic acid amplification (real-time PCR, antigen- antibody detection (immune-assay and SNP detection.

  12. Enzyme-free detection and quantification of double-stranded nucleic acids.

    Feuillie, Cécile; Merheb, Maxime Mohamad; Gillet, Benjamin; Montagnac, Gilles; Hänni, Catherine; Daniel, Isabelle

    2012-08-01

    We have developed a fully enzyme-free SERRS hybridization assay for specific detection of double-stranded DNA sequences. Although all DNA detection methods ranging from PCR to high-throughput sequencing rely on enzymes, this method is unique for being totally non-enzymatic. The efficiency of enzymatic processes is affected by alterations, modifications, and/or quality of DNA. For instance, a limitation of most DNA polymerases is their inability to process DNA damaged by blocking lesions. As a result, enzymatic amplification and sequencing of degraded DNA often fail. In this study we succeeded in detecting and quantifying, within a mixture, relative amounts of closely related double-stranded DNA sequences from Rupicapra rupicapra (chamois) and Capra hircus (goat). The non-enzymatic SERRS assay presented here is the corner stone of a promising approach to overcome the failure of DNA polymerase when DNA is too degraded or when the concentration of polymerase inhibitors is too high. It is the first time double-stranded DNA has been detected with a truly non-enzymatic SERRS-based method. This non-enzymatic, inexpensive, rapid assay is therefore a breakthrough in nucleic acid detection. PMID:22695500

  13. Extramedullary plasmacytoma of the true vocal fold.

    De Zoysa, Nilantha; Sandler, Belinda; Amonoo-Kuofi, Kwame; Swamy, Rajiv; Kothari, Prasad; Mochloulis, George

    2012-08-01

    We report a rare case of extramedullary plasmacytoma (EMP) of the true vocal fold. Our patient, a 62-year-old woman, presented with dysphonia. On workup, fiberoptic laryngoscopy detected a lesion arising from the anterior half of her left true vocal fold. No evidence of other pathology was noted. The patient underwent radical radiotherapy, and the lesion resolved. Follow-up revealed no sign of recurrence. A type of myeloma, EMP is rare, especially in the larynx. To the best of our knowledge, our patient represents the sixth case of glottic EMP to be reported in the literature. PMID:22930090

  14. Evaluation of automated nucleic acid extraction methods for virus detection in a multicenter comparative trial

    Rasmussen, Thomas Bruun; Uttenthal, Åse; Hakhverdyan, M.; Belak, S.; Wakeley, P. R.; Reid, S. M.; Ebert, K.; King, D. P.

    2009-01-01

    Five European veterinary laboratories participated in an exercise to compare the performance of nucleic acid extraction robots. Identical sets of coded samples were prepared using serial dilutions of bovine viral diarrhoea virus (BVDV) from serum and cell culture propagated material. Each...... laboratory extracted nucleic acid from this panel using available robotic equipment (12 separate instruments, comprising 8 different models), after which the processed samples were frozen and sent to a single laboratory for subsequent testing by real-time RT-PCR. In general, there was good concordance...... between the results obtained for the different automated extraction platforms. In particular, the limit of detection was identical for 9/12 and 8/12 best performing robots (using dilutions of BVDV infected-serum and cell culture material, respectively), which was similar to a manual extraction method used...

  15. Simultaneous separation and quantitative determination of monosaccharides, uronic acids, and aldonic acids by high performance anion-exchange chromatography coupled with pulsed amperometric detection in corn stover prehydrolysates

    Xing Wang

    2012-11-01

    Full Text Available A method for simultaneous separation and quantitative determination of arabinose, galactose, glucose, xylose, xylonic acid, gluconic acid, galacturonic acid, and glucuronic acid was developed by using high performance anion-exchange chromatography coupled with pulsed amperometric detection (HPAEC-PAD. The separation was performed on a CarboPacTM PA-10 column (250 mm × 2 mm with a various gradient elution of NaOH-NaOAc solution as the mobile phase. The calibration curves showed good linearity (R2 ≥ 0.9993 for the monosaccharides, uronic acids, and aldonic acids in the range of 0.1 to 12.5 mg/L. The detection limits (LODs and the quantification limits (LOQs were 4.91 to 18.75 μg/L and 16.36 to 62.50 μg/L, respectively. Relative standard deviations (RSDs of the retention times and peak areas for the seven consecutive determinations of an unknown amount of mixture were 0.15% to 0.44% and 0.22% to 2.31%, respectively. The established method was used to separate and determine four monosaccharides, two uronic acids, and two aldonic acids in the prehydrolysate from dilute acid steam-exploded corn stover within 21 min. The spiked recoveries of monosaccharides, uronic acids, and aldonic acids ranged from 91.25% to 108.81%, with RSDs (n=3 of 0.04% ~ 6.07%. This method was applied to evaluate the quantitative variation of sugar and sugar acid content in biomass prehydrolysates.

  16. Comparison of the boronic acid disk potentiation test and cefepime-clavulanic acid method for the detection of ESBL among AmpC-producing Enterobacteriaceae

    R M Shoorashetty

    2011-01-01

    Full Text Available Purpose: Extended spectrum β-lactamase (ESBL and AmpC β-lactamase are important mechanisms of betalactam resistance among Enterobacteriaceae . The ESBL confirmation test described by Clinical Laboratory Standards Institute (CLSI is in routine use. This method fails to detect ESBL in the presence of AmpC. Therefore, we compared two different ESBL detection methods against the CLSI confirmatory test. Materials and Methods: A total 200 consecutive clinical isolates of Enterobacteriaceae from various clinical samples were tested for ESBL production using (i CLSI described phenotypic confirmatory test (PCT, (ii boronic acid disk potentiation test and (iii cefepime-CA disk potentiation method. AmpC confirmation was done by a modified three-dimensional test. Results: Among total 200 Enterobacteriaceae isolates, 82 were only ESBL producers, 12 were only AmpC producers, 55 were combined ESBL and AmpC producers, 14 were inducible AmpC producers and 37 isolates did not harboured any enzymes. The CLSI described PCT detected ESBL-producing organisms correctly but failed to detect 36.3% of ESBLs among combined enzyme producers. The boronic acid disk potentiation test reliably detected all ESBL, AmpC, and combined enzyme producers correctly. The cefepime-CA method detected all ESBLs correctly but another method of AmpC detection has to be adopted. Conclusion: The use of boronic acid in disk diffusion testing along with the CLSI described PCT enhances ESBL detection in the presence of AmpC betalactamases.

  17. Determination of Amino Acids in Single Human Lymphocytes after On-capillary Derivatization by Capillary Zone Electrophoresis with Electrochemical Detection

    2002-01-01

    Amino acids in individual human lymphocytes were determined by capillary zone electrophoresis with electrochemical detection after on-capillary derivatization. In order to inject cells easily, a cell injector was designed. Four amino acids (serine, alanine, taurine, and glycine) in single human lymphocytes have been identified. Quantitation has been accomplished through the use of calibration curves.

  18. Preparation of a novel colorimetric luminescence sensor strip for the detection of indole-3-acetic acid.

    Liu, Yan; Dong, Haitao; Zhang, Wenzhu; Ye, Zhiqiang; Wang, Guilan; Yuan, Jingli

    2010-06-15

    A novel colorimetric luminescence sensor strip for the detection of indole-3-acetic acid (IAA) has been fabricated by using green emissive quantum dots of cadmium telluride (CdTe QDs) as a background layer and a red emissive europium chelate, [4'-(9-anthryl)-2,2':6',2''-terpyridine-6,6''-diyl]bis(methylenenitrilo) tetrakis(acetate)-Eu(3+) (ATTA-Eu(3+)), as a specific sensing layer coated on the surface of glass slide, respectively. The luminescence response of the sensor strip is given by the dramatic changes in emission colors from green to red at different IAA concentrations. This approach provides a simple, rapid, sensitive and accurate method for the detection of IAA without using any special scientific instruments. PMID:20353890

  19. Detection of anhydrous hydrochloric acid, HCl, in IRC+10216 with the Herschel SPIRE and PACS spectrometers

    Cernicharo, J; Barlow, M J; Agundez, M; Royer, P; Vandenbussche, B; Wesson, R; Polehampton, E T; De Beck, E; Blommaert, J A D L; Daniel, F; De Meester, W; Exter, K M; Feuchtgruber, H; Gear, W K; Goicoechea, J R; Gomez, H L; Groenewegen, M A T; Hargrave, P C; Huygen, R; Imhof, P; Ivison, R J; Jean, C; Kerschbaum, F; Leeks, S J; Lim, T L; Matsuura, M; Olofsson, G; Posch, T; Regibo, S; Savini, G; Sibthorpe, B; Swinyard, B M; Vandenbussche, B; Waelkens, C

    2010-01-01

    We report on the detection of anhydrous hydrochloric acid (hydrogen chlorine, HCl) in the carbon-rich star IRC+10216 using the spectroscopic facilities onboard the Herschel satellite. Lines from J=1-0 up to J=7-6 have been detected. From the observed intensities, we conclude that HCl is produced in the innermost layers of the circumstellar envelope with an abundance relative to H2 of 5x10^-8 and extends until the molecules reach its photodissociation zone. Upper limits to the column densities of AlH, MgH, CaH, CuH, KH, NaH, FeH, and other diatomic hydrides have also been obtained.

  20. The comparative detection of renal scarring by intravenous pyelography versus dimercaptosuccinic acid renal scan

    Yamaguchi, Takanori; Itoi, Tatsunori; Nagata, Toyoharu; Ohfuji, Tetsuro; Kitada, Shinichiro; Osada, Yukio (Miyazaki Medical Coll., Kiyotake (Japan))

    1993-06-01

    Intravenous pyelography (IVP) and dimercaptosuccinic acid (DMSA) renal scan were performed in 82 kidneys of 52 patients with vesicoureteral reflux (VUR). Thirty-three kidneys (40%) were found to be scarred on IVP whereas 61 kidneys (74%) were found to be scarred on DMSA renal scan. In one of the 82 refluxing kidneys, IVP demonstrated a scar not detectable on DMSA study. The concordance rate was 40 of 82 kidneys (49%). When both studies were positive, there was an excellent correlation on a site by site basis. The incidence of positive DMSA increased with high grade VUR and decreased with low grade reflux. When IVP identifies renal scarring, there is usually a more significant area of scars demonstrated on DMSA renal scan. Our findings indicate that DMSA renal scan is more sensitive than IVP regarding the early detection of renal scarring in patients with VUR. (author).

  1. Detection of Glutamate and γ-aminobutyric Acid in Vitreous of Patients with Proliferative Diabetic Retinopathy

    Juan Deng; De-Zheng Wu; Rulong Gao

    2000-01-01

    Purpose: To study the levels of glutamate and γ-aminobutyric acid (GABA) in vitreous of patients with proliferative diabetic retinopathy(PDR) and to investigate their roles in retinal ischemia.Method: Vitreous samples were collected from 25 patients (27 eyes) with PDR and 14patients ( 14 eyes) with idiopathic macular hole. Glutamate and GABA detection were performed by high-performance liquid chromatography (HPLC).Results: Patients with PDR had significantly higher concentrations of glutamate and GABA than the control group. The glutamate level has a significantly positive correlation with GABA level.Conclusion: Detection of glutamate and GABA in vitreous provides biochemical support for the mechanism and treatment of ischemic retinal damage in patients with PDR.

  2. A FRET-enabled molecular peptide beacon with a significant red shift for the ratiometric detection of nucleic acids.

    Maity, Debabrata; Jiang, Juanjuan; Ehlers, Martin; Wu, Junchen; Schmuck, Carsten

    2016-05-01

    A cationic molecular peptide beacon NAP1 functionalized with a fluorescence resonance energy transfer-pair at its ends allows the ratiometric detection of ds-DNA with a preference for AT rich sequences. NAP1 most likely binds in a folded form into the minor groove of ds-DNA, which results in a remarkable change in its fluorescence properties. As NAP1 exhibits quite low cytotoxicity, it can also be used for imaging of nuclear DNA in cells. PMID:27071707

  3. Analysis of High-Fold Gamma Data

    Historically, γ-γ and γ-γ-γ coincidence spectra were utilized to build nuclear level schemes. With the development of large detector arrays, it has became possible to analyze higher fold coincidence data sets. This paper briefly reports on software to analyze 4-fold coincidence data sets that allows creation of 4-fold histograms (hypercubes) of at least 1024 channels per side (corresponding to a 43 gigachannel data space) that will fit onto a few gigabytes of disk space, and extraction of triple-gated spectra in a few seconds. Future detector arrays may have even higher efficiencies, and detect an many as 15 or 20 γ rays simultaneously; such data will require very different algorithms for storage and analysis. Difficulties inherent in the analysis of such data are discussed, and two possible new solutions are presented, namely adaptive list-mode systems and list-list-mode storage

  4. In vivo detection of prostatic carcinoma with antibodies against prostatic acid phosphatase

    Serum prostatic acid phosphates (PAP) immunoassay is used to evaluate patients with prostatic carcinoma; however, as with other tumor markers, the enzyme levels do not necessarily reflect the presence or extent of tumor. The authors investigated the use of radiolabeled PAP antibodies for the in vivo detection of prostatic carcinoma by external scintillation imaging. Nine patients with prostatic carcinoma were entered into the study. Each received from 2.0 to 2.5 mCi of I-131 labeled antibody to PAP, administered i.v. The immunogen (PAP) was purified from normal human seminal fluid. Antiserum was prepared in rabbits by injecting the purified PAP. The antibodies were labeled with I-131 by chloramine-T method (10 to 20 Ci/g of IgG). Total body images were obtained at 24 and 48 hrs following administration of the labeled antibody. Nontarget I-131 activity was diminished by computer processing. Tumor sites detected by I-131 antibodies were correlated with other diagnostic procedures. In 7 of 9 patients primary and metastatic sites of cancer were detected by antibody imaging, however, no bone lesions were detected (6 cases). In 3 patients with concomitant pulmonary tumors, one was identified as of prostate origin. The serum PAP was normal in 4 patients; however, the primary tumor was identified in 3 of these. These findings suggest that the localization of prostatic carcinoma by means of in-vivo imaging of labeled antibodies to PAP is feasible and offers diagnostic opportunities based upon the functional characteristics

  5. Simultaneous separation and detection of actinides in acidic solutions using an extractive scintillating resin.

    Roane, J E; DeVol, T A

    2002-11-01

    An extractive scintillating resin was evaluated for the simultaneous separation and detection of actinides in acidic solutions. The transuranic extractive scintillating (TRU-ES) resin is composed of an inert macroporous polystyrene core impregnated with organic fluors (diphenyloxazole and 1,4-bis-(4-methyl-5-phenyl-2-oxazolyl)benzene) and an extractant (octyl(phenyl)-N,N-diisobutylcarbamoylmethylphosphine oxide in tributyl phosphate). The TRU-ES resin was packed into FEP Teflon tubing to produce a flow cell (0.2-mL free column volume), which is placed into a scintillation detection system to obtain pulse height spectra and time series data during loading and elution of actinides onto/from the resin. The alpha-particle absolute detection efficiencies ranged from 77% to 96.5%, depending on the alpha energy and quench. In addition to the on-line analyses, off-line analyses of the effluent can be conducted using conventional detection methods. The TRU-ES resin was applied to the quantification of a mixed radionuclide solution and two actual waste samples. The on-line characterization of the mixed radionuclide solution was within 10% of the reported activities whereas the agreement with the waste samples was not as good due to sorption onto the sample container walls and the oxidation state of plutonium. Agreement between the on-line and off-line analyses was within 35% of one another for both waste samples. PMID:12433098

  6. Vocal fold and ventricular fold vibration in period-doubling phonation: physiological description and aerodynamic modeling.

    Bailly, Lucie; Henrich, Nathalie; Pelorson, Xavier

    2010-05-01

    Occurrences of period-doubling are found in human phonation, in particular for pathological and some singing phonations such as Sardinian A Tenore Bassu vocal performance. The combined vibration of the vocal folds and the ventricular folds has been observed during the production of such low pitch bass-type sound. The present study aims to characterize the physiological correlates of this acoustical production and to provide a better understanding of the physical interaction between ventricular fold vibration and vocal fold self-sustained oscillation. The vibratory properties of the vocal folds and the ventricular folds during phonation produced by a professional singer are analyzed by means of acoustical and electroglottographic signals and by synchronized glottal images obtained by high-speed cinematography. The periodic variation in glottal cycle duration and the effect of ventricular fold closing on glottal closing time are demonstrated. Using the detected glottal and ventricular areas, the aerodynamic behavior of the laryngeal system is simulated using a simplified physical modeling previously validated in vitro using a larynx replica. An estimate of the ventricular aperture extracted from the in vivo data allows a theoretical prediction of the glottal aperture. The in vivo measurements of the glottal aperture are then compared to the simulated estimations. PMID:21117769

  7. Direct folding simulation of a long helix in explicit water

    Gao, Ya; Lu, Xiaoliang; Duan, Lili; Zhang, Dawei; Mei, Ye; Zhang, John Z. H.

    2013-05-01

    A recently proposed Polarizable Hydrogen Bond (PHB) method has been employed to simulate the folding of a 53 amino acid helix (PDB ID 2KHK) in explicit water. Under PHB simulation, starting from a fully extended structure, the peptide folds into the native state as confirmed by measured time evolutions of radius of gyration, root mean square deviation (RMSD), and native hydrogen bond. Free energy and cluster analysis show that the folded helix is thermally stable under the PHB model. Comparison of simulation results under, respectively, PHB and standard nonpolarizable force field demonstrates that polarization is critical for stable folding of this long α-helix.

  8. High-performance liquid chromatography with conductimetric detection of perfluorocarboxylic acids and perfluorosulfonates.

    Hori, Hisao; Hayakawa, Etsuko; Yamashita, Nobuyoshi; Taniyasu, Sachi; Nakata, Fumiya; Kobayashi, Yoshimi

    2004-10-01

    A rapid and simple method for separating and determining various environmentally harmful perfluorocarboxylic acids and perfluorosulfonates was successfully developed using high- performance liquid chromatography with conductimetric detection, for product and waste management of these compounds at manufacturing and processing sites. Compounds having C(3)-C(8) perfluoroalkyl groups were separated using a Tosoh TSKgel Super-ODS column and a mobile phase consisting of a mixture of methanol and aqueous NaH(2)PO(4) at several mixing ratios. The best detection limits for the compounds ranged from 0.12 to 0.66 mg l(-1) (ppm), and linear calibration graphs were obtained up to 87-109 mg l(-1). The combination of this method with concentration of the sample by solid-phase extraction with cartridges based on styrene-divinylbenzene-copolymer enabled the determination of approximately 50 microg l(-1) (ppb) for compounds with C(4)-C(8) perfluoroalkyl groups. This method was successfully used to monitor the artificial decomposition of the perfluorocarboxylic acid n-C(4)F(9)COOH induced by a photocatalyst. PMID:15312725

  9. Enhanced Sensitivity for Hydrogen Peroxide Detection: Polydiacetylene Vesicles with Phenylboronic Acid Head Group.

    Jia, Chen; Tang, Jie; Lu, Shengguo; Han, Yuwang; Huang, He

    2016-01-01

    It was recently reported that, besides UV irradiated polymerization, polymerization of diacetylene compounds could also been initiated by radicals generated from enzyme catalyzed hydrogen peroxide (H2O2) decomposition. A new optical sensing method for H2O2 was proposed based on this phenomenon. However, the sensitivity of this method is relatively lower than existed ones. In the present work, phenylboronic acid (PBA) functionalized 10, 12-pentacosadiynoic acid (PDA-PBA) was synthesized and its vesicles were formed successfully as colorimetric sensor for H2O2 detection. It was found that color change during the polymerization of vesicles composed of the PBA modified monomer is much stronger than that of the non-modified one. The response of PDA-PBA vesicles to H2O2 is 16 times more sensitive than that of the PDA. The absorption of PDA-PBA at 650 nm is linearly related to the concentration of H2O2 and a detection limit of ~5 μM could be achieved. PMID:26511954

  10. Uptake of gamma-valerolactone--detection of gamma-hydroxyvaleric acid in human urine samples.

    Andresen-Streichert, H; Jungen, H; Gehl, A; Müller, A; Iwersen-Bergmann, S

    2013-05-01

    Gamma-valerolactone (GVL) is reported to be a substance that can be used as a legal substitute for gamma-hydroxybutyric acid (GHB), which is currently a controlled substance in several countries. Unlike gamma-butyrolactone and 1,4-butanediol, GVL is not metabolized to GHB, which causes the effects after uptake of these two chemicals. In the case of GVL, the lactone ring is split to gamma-hydroxyvaleric acid (GHV or 4-methyl-GHB) by a lactonase. Because of its affinity for the GHB receptor, GHV reveals similar effects to GHB, although it is less potent. Intoxications with GVL, or its use as a date rape drug, are conceivable. Despite these facts, there are no publications in the literature regarding detections of GHV in human samples. This study reports three cases, including five urine samples, in which GHV could be detected in concentrations between 3 and 5.8 mg/L. In one of these cases, a drug-facilitated sexual assault (DFSA) was assumed; four of these samples were from two people suspected of abusing GHB. The results indicate that GVL is used as an alternative to GHB and its precursors and should be taken seriously. GVL or GHV should be included in toxicological analysis, particularly in DFSA cases. More information is needed regarding the pharmacokinetics of GVL/GHV for the meaningful interpretation of positive or negative results. PMID:23486087

  11. Study on the Spectrophotometric Detection of Free Fatty Acids in Palm Oil Utilizing Enzymatic Reactions

    Nur Hidayah Azeman

    2015-07-01

    Full Text Available In this paper, a comprehensive study has been made on the detection of free fatty acids (FFAs in palm oil via an optical technique based on enzymatic aminolysis reactions. FFAs in crude palm oil (CPO were converted into fatty hydroxamic acids (FHAs in a biphasic lipid/aqueous medium in the presence of immobilized lipase. The colored compound formed after complexation between FHA and vanadium (V ion solution was proportional to the FFA content in the CPO samples and was analyzed using a spectrophotometric method. In order to develop a rapid detection system, the parameters involved in the aminolysis process were studied. The utilization of immobilized lipase as catalyst during the aminolysis process offers simplicity in the product isolation and the possibility of conducting the process under extreme reaction conditions. A good agreement was found between the developed method using immobilized Thermomyces lanuginose lipase as catalyst for the aminolysis process and the Malaysian Palm Oil Board (MPOB standard titration method (R2 = 0.9453.

  12. Immobilization of Tyrosinase from Avocado Crude Extract in Polypyrrole Films for Inhibitive Detection of Benzoic Acid

    André Brisolari

    2014-07-01

    Full Text Available Inhibition-based biosensors were developed by immobilizing tyrosinase (Tyr, polyphenol oxidase from the crude extract of avocado fruit on electrochemically prepared polypyrrole (PPy films. The biosensors were prepared during the electropolymerization of pyrrole in a solution containing a fixed volume of the crude extract of avocado. The dependence of the biosensor responses on the volume used from the crude extract, values of pH and temperature was studied, and a substrate, catechol, at different concentrations, was amperometrically detected by these biosensors. Benzoic acid, a competitive inhibitor of Try, was added to the catechol solutions at specific concentrations aimed at obtaining the inhibition constant, K’m, which ranged from 1.7 to 4.6 mmol∙L−1 for 0.0 and 60 µmol∙L−1 of benzoic acid, respectively. Studies on the inhibition caused by benzoic acid by using PPy/Try films, and catechol as a substrate, allowed us propose how to develop, under optimized conditions, simple and low-cost biosensors based on the use of avocado fruit.

  13. Colorimetric detection of Cr{sup 3+} using gold nanoparticles functionalized with 4-amino hippuric acid

    Jin, Weiwei; Huang, Pengcheng; Chen, Yueji; Wu, Fangying, E-mail: fywu@ncu.edu.cn; Wan, Yiqun [Nanchang University, College of Chemistry (China)

    2015-09-15

    A facile and effective technique for monitoring Cr{sup 3+} concentration based on 4-amino hippuric acid (PAH) decorated Au nanoparticles (PAH-AuNPs) is introduced. The modified AuNPs were easily aggregated in the presence of Cr{sup 3+}, resulting in the color change from red to violet or blue, which is in response to the surface plasmon absorption of dispersed or aggregated nanoparticles. Under the optimized conditions, a good linear relationship (correlation coefficient r = 0.998) was obtained between the ratio of the absorbance at 635 nm to that at 520 nm (A{sub 635 nm}/A{sub 520 nm}), and the concentration of Cr{sup 3+} was over the range of 5.0–120 µM with detection limit of 1.17 µM. This method exhibited excellent selectivity for Cr{sup 3+} over other tested heavy metal ions. Furthermore, there was no significant difference for the parameters of calibration equation between the presence and absence of ethylenediamine tetraacetic acid (EDTA), which suggests that the method can be applied in various real samples owing to the strong masking ability of EDTA. The assay was used to detect the concentrations of Cr{sup 3+} in liquid milk, milk power, and lake water samples with recoveries ranging from 93.5 to 114 %, indicating that the method could be used for extensive practical application. Graphical Abstract: A facile and effective technique for monitoring Cr{sup 3+} based on 4-amino hippuric acid decorated Au nanoparticles is introduced. The modified AuNPs were aggregated in the presence of Cr{sup 3+} resulting in the color change from red to violet or blue, which is in response to the surface plasmon absorption of dispersed or aggregated nanoparticles.

  14. Colorimetric detection of Cr3+ using gold nanoparticles functionalized with 4-amino hippuric acid

    A facile and effective technique for monitoring Cr3+ concentration based on 4-amino hippuric acid (PAH) decorated Au nanoparticles (PAH-AuNPs) is introduced. The modified AuNPs were easily aggregated in the presence of Cr3+, resulting in the color change from red to violet or blue, which is in response to the surface plasmon absorption of dispersed or aggregated nanoparticles. Under the optimized conditions, a good linear relationship (correlation coefficient r = 0.998) was obtained between the ratio of the absorbance at 635 nm to that at 520 nm (A635 nm/A520 nm), and the concentration of Cr3+ was over the range of 5.0–120 µM with detection limit of 1.17 µM. This method exhibited excellent selectivity for Cr3+ over other tested heavy metal ions. Furthermore, there was no significant difference for the parameters of calibration equation between the presence and absence of ethylenediamine tetraacetic acid (EDTA), which suggests that the method can be applied in various real samples owing to the strong masking ability of EDTA. The assay was used to detect the concentrations of Cr3+ in liquid milk, milk power, and lake water samples with recoveries ranging from 93.5 to 114 %, indicating that the method could be used for extensive practical application. Graphical Abstract: A facile and effective technique for monitoring Cr3+ based on 4-amino hippuric acid decorated Au nanoparticles is introduced. The modified AuNPs were aggregated in the presence of Cr3+ resulting in the color change from red to violet or blue, which is in response to the surface plasmon absorption of dispersed or aggregated nanoparticles

  15. Copper- or manganese-doped ZnS quantum dots as fluorescent probes for detecting folic acid in aqueous media

    3-Mercaptopropionic acid-capped core/shell ZnS:Cu/ZnS and ZnS:Mn/ZnS doped quantum dots (QDs) prepared through hydrothermal methods exhibit high photoluminescence intensity as well as good photostability. These water-dispersible nanoparticles exhibit high fluorescence sensitivity to folic acid due to the high affinity of the carboxylate groups and nitrogen atoms of folic acid towards the Zn surface atoms of the doped dots. Quenching of the fluorescence intensity of the QDs allows the detection of folic acid concentrations as low as 11 μM, thus affording a very sensitive system for the sensing of this biologically active molecule in aqueous solution. The possible quenching mechanism is discussed. - Graphical abstract: A sensitive method for the detection of folic acid based on the fluorescence quenching of Mn- or Cu-doped ZnS quantum dots was developed. Highlights: ► Quenching of the fluorescence intensity of doped ZnS QDs in the presence of folic acid. ► New fluorescent sensors for folic acid. ► Detection of folic acid concentrations as low as 11 μM in aqueous solution. ► The Perrin model and fluorescence lifetimes of ZnS:Mn QDs demonstrate a static quenching mechanism. ► Quenching efficiency of ZnS:Cu QDs correlates with the Stern-Volmer model.

  16. Natural selection of more designable folds: A mechanism for thermophilic adaptation

    England, Jeremy L.; Boris E Shakhnovich; Eugene I Shakhnovich

    2003-01-01

    An open question of great interest in biophysics is whether variations in structure cause protein folds to differ in the number of amino acid sequences that can fold to them stably, i.e., in their designability. Recently, we have shown that a novel quantitative measure of a fold's tertiary topology, called its contact trace, strongly correlates with the fold's designability. Here, we investigate the relationship between a fold's contact trace and its relative frequency...

  17. Synthesis of Au/Graphene Oxide Composites for Selective and Sensitive Electrochemical Detection of Ascorbic Acid

    Song, Jian; Xu, Lin; Xing, Ruiqing; Li, Qingling; Zhou, Chunyang; Liu, Dali; Song, Hongwei

    2014-12-01

    In this work, we present a novel ascorbic acid (AA) sensor applied to the detection of AA in human sera and pharmaceuticals. A series of Au nanoparticles (NPs) and graphene oxide sheets (Au NP/GO) composites were successfully synthesized by reduction of gold (III) using sodium citrate. Then the Au NP/GO composites were used to construct nonenzymatic electrodes in practical AA measurement. The electrode that has the best performance presents attractive analytical features, such as a low working potential of +0.15 V, a high sensitivity of 101.86 μA mM-1 cm-2 to AA, a low detection limit of 100 nM, good reproducibility and excellent selectivity. And more,it was also employed to accurately and practically detect AA in human serum and clinical vitamin C tablet with the existence of some food additive. The enhanced AA electrochemical properties of the Au NP/GO modified electrode in our work can be attributed to the improvement of electroactive surface area of Au NPs and the synergistic effect from the combination of Au NPs and GO sheets. This work shows that the Au NP/GO/GCEs hold the prospect for sensitive and selective determination of AA in practical clinical application.

  18. Excretion and detection of SARS coronavirus and its nucleic acid from digestive system

    Xin-Wei Wang; Xiao-Ming Wu; Wen-Jun Xiao; Xiu-Mei Zhu; Chang-Qing Gu; Jing Yin; Wei Wei; Wei Yao; Chao Liu; Jian-Feng Li; Guo-Rong Ou; Jin-Song Li; Min-Nian Wang; Tong-Yu Fang; Gui-Jie Wang; Yao-Hui Qiu; Huai-Huan Wu; Fu-Huan Chao; Jun-Wen Li; Ting-Kai Guo; Bei Zhen; Qing-Xin Kong; Bin Yi; Zhong Li; Nong Song; Min Jin

    2005-01-01

    AIM: To study whether severe acute respiratory syndrome coronavirus (SARS-CoV) could be excreted from digestive system.METHODS: Cell culture and semi-nested RT-PCR were used to detect SARS-CoV and its RNA from 21 stool and urine samples, and a kind of electropositive filter media particles was used to concentrate the virus in 10 sewage samples from two hospitals receiving SAPS patients in Beijing in China.RESULTS: It was demonstrated that there was no live SARS-CoV in all samples collected, but the RNA of SARS-CoV could be detected in seven stool samples from SARS patients with any one of the symptoms of fever, malaise,cough, or dyspnea, in 10 sewage samples before disinfection and 3 samples after disinfection from the two hospitals.The RNA could not be detected in urine and stool samples from patients recovered from SARS.CONCLUSION: Nucleic acid of SARS-CoV can be excreted through the stool of patients into sewage system, and the possibility of SARS-CoV transmitting through digestive system cannot be excluded.

  19. Porphyrin-functionalized gold nanoparticles for selective electrochemical detection of peroxyacetic acid

    Li Jie; Tu Wenwen [Key Laboratory of Analytical Chemistry for Life Science (Ministry of Education of China), Department of Chemistry, Nanjing University, Nanjing 210093 (China); Lei Jianping, E-mail: jpl@nju.edu.c [Key Laboratory of Analytical Chemistry for Life Science (Ministry of Education of China), Department of Chemistry, Nanjing University, Nanjing 210093 (China); Tang Sheng [Key Laboratory of Analytical Chemistry for Life Science (Ministry of Education of China), Department of Chemistry, Nanjing University, Nanjing 210093 (China); Ju Huangxian, E-mail: hxju@nju.edu.c [Key Laboratory of Analytical Chemistry for Life Science (Ministry of Education of China), Department of Chemistry, Nanjing University, Nanjing 210093 (China)

    2011-03-30

    Two layers of cationic iron(III) meso-tetrakis (N-methylpyridinum-4-yl)porphyrin (FeTMPyP) and anionic gold nanoparticles (GNPs) were alternately assembled on a poly(diallyldimethylammonium chloride)-wrapped carbon nanotube (PDDA-CNT)-modified electrode via electrostatic interactions. The porphyrin-functionalized gold nanoparticles were characterized by scanning electron microscopy and UV-vis absorption spectrometry. The (FeTMPyP-GNP){sub 2}/PDDA-CNT modified electrode showed two stable and well-defined peaks at -0.112 V and -0.154 V, which were attributed to the GNP-accelerated redox process of Fe(III)TMPyP/Fe(II)TMPyP. The modified electrode possessed excellent electrocatalytic behavior for the reduction of peroxyacetic acid (PAA). The resulting biosensor exhibited a fast amperometric response to PAA ({approx}3 s), with a wide linear range from 2.5 x 10{sup -6} M to 1.05 x 10{sup -3} M and a detection limit of 0.5 {mu}M at a signal-to-noise ratio of 3. More importantly, H{sub 2}O{sub 2} did not interfere with the detection. Thus, this biosensor enabled highly sensitive detection of PAA without removing H{sub 2}O{sub 2} and showed a promising potential in practical applications.

  20. Cell-free nucleic acids as noninvasive biomarkers for colorectal cancer detection

    Mansour, Hicham

    2014-08-27

    Cell-free nucleic acids (CFNA) have been reported by several authors in blood, stool, and urine of patients with colorectal cancer (CRC). These genetic biomarkers can be an indication of neoplastic colorectal epithelial cells, and can thus potentially be used as noninvasive tests for the detection of the disease in CRC patients and monitor their staging, without the need to use heavier and invasive tools. In a number of test-trials, these genetic tests have shown the advantage of non-invasiveness, making them well accepted by most of the patients, without major side effects. They have also shown a promising sensitivity and specificity in the detection of malignant and premalignant neoplasms. Moreover, costs for performing such tests are very low. Several studies reported and confirmed the proof of the principle for these genetic tests for screening, diagnosis, and prognosis; the main challenge of translating this approach from research to clinical laboratory is the validation from large and long-term randomized trials to prove sustainable high sensitivity and specificity. In this paper, we present a review on the noninvasive genetics biomarkers for CRC detection described in the literature and the challenges that can be encountered for validation processes.

  1. Supersensitive and selective detection of picric acid explosive by fluorescent Ag nanoclusters.

    Zhang, Jian Rong; Yue, Yuan Yuan; Luo, Hong Qun; Li, Nian Bing

    2016-02-01

    Picric acid (PA) explosive is a hazard to public safety and health, so the sensitive and selective detection of PA is very important. In the present work, polyethyleneimine stabilized Ag nanoclusters were successfully used for the sensitive and selective quantification of PA on the basis of fluorescence quenching. The quenching efficiency of Ag nanoclusters is proportional to the concentration of PA and the logarithm of PA concentration over two different concentration ranges (1.0 nM-1 μM for the former and 0.25-20 μM for the latter), thus the proposed quantitative strategy for PA provides a wide linear range of 1.0 nM-20 μM. The detection limit based on 3σ/K is 0.1 nM. The quenching mechanism of Ag nanoclusters by PA is discussed in detail. The results indicate that the selective detection of PA over other nitroaromatics including 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (2,4-DNT), p-nitrotoluene (p-NT), m-dinitrobenzene (m-DNB), and nitrobenzene (NB), is due to the electron transfer and energy transfer between PA and polyethyleneimine-capped Ag nanoclusters. In addition, the experimental data obtained for the analysis of artificial samples show that the proposed PA sensor is potentially applicable in the determination of trace PA explosive in real samples. PMID:26661456

  2. Field Effect Sensors for Nucleic Acid Detection: Recent Advances and Future Perspectives

    Bruno Veigas

    2015-05-01

    Full Text Available In the last decade the use of field-effect-based devices has become a basic structural element in a new generation of biosensors that allow label-free DNA analysis. In particular, ion sensitive field effect transistors (FET are the basis for the development of radical new approaches for the specific detection and characterization of DNA due to FETs’ greater signal-to-noise ratio, fast measurement capabilities, and possibility to be included in portable instrumentation. Reliable molecular characterization of DNA and/or RNA is vital for disease diagnostics and to follow up alterations in gene expression profiles. FET biosensors may become a relevant tool for molecular diagnostics and at point-of-care. The development of these devices and strategies should be carefully designed, as biomolecular recognition and detection events must occur within the Debye length. This limitation is sometimes considered to be fundamental for FET devices and considerable efforts have been made to develop better architectures. Herein we review the use of field effect sensors for nucleic acid detection strategies—from production and functionalization to integration in molecular diagnostics platforms, with special focus on those that have made their way into the diagnostics lab.

  3. Determination of catecholamines by ion chromatography coupled to acidic potassium permanganate chemiluminescence detection

    Hong Wei Wu; Mei Lan Chen; Dan Shou; Yan Zhu

    2012-01-01

    A simple,fast,sensitive,highly selective and eco-friendly analytical method for the determination of catecholamines in human urine by ion chromatography (IC) with chemiluminescence (CL) detection was described in this paper.Using 12 mmoi/L H2SO4 without any organic additive as eluent,three catecholamines including epinephrine (EP),norepinephrine (NE) and dopamine (DA)were well separated on a cation-exchange column.The CL detection was based on the reaction of analytes with acidic potassium permanganate in the presence of formaldehyde as an enhancer.The absence of methanol and acetonitrile in eluent made the proposed method more sensitive and eco-friendly.Under the optimal conditions,the linear range of the proposed method was in the range of 0.02-0.5 μg/mL.The limit of detection (LOD) was in the range of 0.6 and 5.1 μg/L.The relative standard deviations (RSD) for 0.1 μg/mL mixed standard solution were in the range of 0.8-1.9% (n =11).The method has been applied to the determination of catecholamines in human urine successfully.Excellent spiked recoveries were achieved for catecholamines ranged from 91.2% to 112.7%.

  4. Two-fold symmetric singularity

    Jacquemard, Alain; Teixeira, Marco Antonio; Tonon, Durval Jose

    2011-01-01

    We explore some qualitative dynamics in the neighborhood of the $3-dimensional$ two-fold symmetric singularity. We study the existence of an one-parameter family of regular (pseudo) periodic orbits of such systems near a reversible two-fold singularity.

  5. Problem Solving through Paper Folding

    Wares, Arsalan

    2014-01-01

    The purpose of this article is to describe a couple of challenging mathematical problems that involve paper folding. These problem-solving tasks can be used to foster geometric and algebraic thinking among students. The context of paper folding makes some of the abstract mathematical ideas involved relatively concrete. When implemented…

  6. COS NUV MAMA Fold Distribution

    Wheeler, Thomas

    2013-10-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as Cycle 20 proposal 13128.

  7. Separation and conductimetric detection of C1-C7 aliphatic monocarboxylic acids and C1-C7 aliphatic monoamines on unfunctionized polymethacrylate resin columns.

    Ohta, Kazutoku; Towata, Atsuya; Ohashi, Masayoshi; Takeuchi, Toyohide

    2004-06-11

    The application of unfunctionized polymethacrylate resin (TSKgel G3000PWXL) as a stationary phase in liquid chromatography with conductimetric detection for C1-C7 aliphatic monocarboxylic acids (formic acid, acetic acid, propionic acid, butyric acid, isovaleric acid, valeric acid, 3,3-dimethylbutyric acid, 4-methylvaleric acid, hexanoic acid, 2-methylhexanoic acid, 5-methylhexanoic acid and heptanoic acid) and C1-C7 aliphatic monoamines (methylamine, ethylamine, propylamine, isobutylamine, butylamine, isoamylamine, amylamine, 1,3-dimethylbutylamine, hexylamine, 2-heptylamine and heptylamine) was attempted with C8 aliphatic monocarboxylic acids (2-propylvaleric acid, 2-ethylhexanoic acid, 2-methylheptanoic acid and octanoic acid) and C8 aliphatic monoamines (1,5-dimethylhexylamine, 2-ethylhexylamine, 1-methylheptylamine and octylamine) as eluents, respectively. Using 1 mM 2-methylheptanoic acid at pH 4.0 as the eluent, excellent separation and relatively high sensitive detection for these C1-C7 carboxylic acids were achieved on a TSKgel G3000PWXL column (150 mm x 6 mm i.d.) in 60 min. Using 2 mM octylamine at pH 11.0 as the eluent, excellent separation and relatively high sensitive detection for these C1-C7 amines were also achieved on the TSKgel G3000PWXL column in 60 min. PMID:15250420

  8. Equi-Gaussian Curvature Folding

    E M El-Kholy; El-Said R Lashin; Salama N Daoud

    2007-08-01

    In this paper we introduce a new type of folding called equi-Gaussian curvature folding of connected Riemannian 2-manifolds. We prove that the composition and the cartesian product of such foldings is again an equi-Gaussian curvature folding. In case of equi-Gaussian curvature foldings, $f:M→ P_n$, of an orientable surface onto a polygon $P_n$ we prove that (i) $f\\in\\mathcal{F}_{EG}(S^2)\\Leftrightarrow n=3$ (ii) $f\\in\\mathcal{F}_{EG}(T^2)\\Rightarrow n=4$ (iii) $f\\in\\mathcal{F}_{EG}(\\# 2T^2)\\Rightarrow n=5, 6$ and we generalize (iii) for $\\# nT^2$.

  9. Protein folding and protein metallocluster studies using synchrotron small angler X-ray scattering

    Proteins, biological macromolecules composed of amino-acid building blocks, possess unique three dimensional shapes or conformations which are intimately related to their biological function. All of the information necessary to determine this conformation is stored in a protein's amino acid sequence. The problem of understanding the process by which nature maps protein amino-acid sequences to three-dimensional conformations is known as the protein folding problem, and is one of the central unsolved problems in biophysics today. The possible applications of a solution are broad, ranging from the elucidation of thousands of protein structures to the rational modification and design of protein-based drugs. The scattering of X-rays by matter has long been useful as a tool for the characterization of physical properties of materials, including biological samples. The high photon flux available at synchrotron X-ray sources allows for the measurement of scattering cross-sections of dilute and/or disordered samples. Such measurements do not yield the detailed geometrical information available from crystalline samples, but do allow for lower resolution studies of dynamical processes not observable in the crystalline state. The main focus of the work described here has been the study of the protein folding process using time-resolved small-angle x-ray scattering measurements. The original intention was to observe the decrease in overall size which must accompany the folding of a protein from an extended conformation to its compact native state. Although this process proved too fast for the current time-resolution of the technique, upper bounds were set on the probable compaction times of several small proteins. In addition, an interesting and unexpected process was detected, in which the folding protein passes through an intermediate state which shows a tendency to associate. This state is proposed to be a kinetic molten globule folding intermediate

  10. Protein folding and protein metallocluster studies using synchrotron small angler X-ray scattering

    Eliezer, D.

    1994-06-01

    Proteins, biological macromolecules composed of amino-acid building blocks, possess unique three dimensional shapes or conformations which are intimately related to their biological function. All of the information necessary to determine this conformation is stored in a protein`s amino acid sequence. The problem of understanding the process by which nature maps protein amino-acid sequences to three-dimensional conformations is known as the protein folding problem, and is one of the central unsolved problems in biophysics today. The possible applications of a solution are broad, ranging from the elucidation of thousands of protein structures to the rational modification and design of protein-based drugs. The scattering of X-rays by matter has long been useful as a tool for the characterization of physical properties of materials, including biological samples. The high photon flux available at synchrotron X-ray sources allows for the measurement of scattering cross-sections of dilute and/or disordered samples. Such measurements do not yield the detailed geometrical information available from crystalline samples, but do allow for lower resolution studies of dynamical processes not observable in the crystalline state. The main focus of the work described here has been the study of the protein folding process using time-resolved small-angle x-ray scattering measurements. The original intention was to observe the decrease in overall size which must accompany the folding of a protein from an extended conformation to its compact native state. Although this process proved too fast for the current time-resolution of the technique, upper bounds were set on the probable compaction times of several small proteins. In addition, an interesting and unexpected process was detected, in which the folding protein passes through an intermediate state which shows a tendency to associate. This state is proposed to be a kinetic molten globule folding intermediate.

  11. Direct determination of seleno-amino acids in biological tissues by anion-exchange separation and electrochemical detection.

    Cavalli, S; Cardellicchio, N

    1995-07-01

    Several studies have described the determination of selenium in protein extracts from tissues of marine or terrestrial animals, but have not identified the different chemical forms of selenium that are present. Selenium may be present as seleno-amino acids. Selenocysteine, for example, is a normal component of glutathione peroxidase, an antioxidant enzyme which may behave like other antioxidants, such as vitamin E, protecting tissues against methylmercury toxicity. The present study illustrates a method for the characterization of seleno-amino acids, such as selenocysteine and selenomethionine, in proteins extracted from the liver of marine mammals. The mechanism of detoxification of methylmercury, which involves seleno-compounds, is identified. The analytical determination was carried out using high-performance anion-exchange chromatography coupled with integrated pulsed amperometric detection (HPAEC-IPAD). This method allows the direct determination of underivatized amino acids, eliminating the procedure of pre- or postcolumn derivatization. The chromatographic separation was carried out on an anion-exchange column using a quaternary gradient elution. In order to optimize this method, interferences of amino acids and the influence of pH and ionic strength on the separation and electrochemical detection were studied. The IPAD response for the direct detection of amino acids is optimum at pH > 11. The detection limit (S/N = 3) for selenocysteine was found to be 450 micrograms/l. The application of this method for the identification of seleno-amino acids in protein hydrolysates is also shown. PMID:7640774

  12. A comparison of fluorescamine and naphthalene-2,3-dicarboxaldehyde fluorogenic reagents for microplate-based detection of amino acids.

    Bantan-Polak, T; Kassai, M; Grant, K B

    2001-10-15

    The use of appropriate fluorometric derivatization procedures is of considerable importance for accurate determination of amino acids in biological samples and in metal-assisted peptide hydrolysis reactions. It is especially critical for the relative fluorescence intensities (RFI) of equal amounts of amino acids to be as similar as possible. While fluorescamine and naphthalene-2,3-dicarboxaldehyde (NDA) have proven to be excellent fluorogenic reagents for amino acid detection, the effects of various factors such as organic solvent, buffer, and pH have never been rigorously evaluated with respect to normalizing the relative fluorescence intensities of individual amino acids. To this end, here we describe optimized fluorescamine and NDA derivatization reactions that enhance the accuracy of microplate-based detection of amino acids. For both fluorescamine and NDA, we have shown that the RFI values of 16 of 19 amino acids are greater than 70%. Although determination of tryptophan is problematic, this difficulty is overcome by the addition of beta-cyclodextrin to the NDA reaction. In principle, the optimized fluorescamine and NDA microplate procedures reported here can be utilized as complementary techniques for the detection of 19 of 20 naturally occurring amino acids. PMID:11673879

  13. Standardization of natural mycolic acid antigen composition and production for use in biomarker antibody detection to diagnose active tuberculosis.

    Ndlandla, F L; Ejoh, V; Stoltz, A C; Naicker, B; Cromarty, A D; van Wyngaardt, S; Khati, M; Rotherham, L S; Lemmer, Y; Niebuhr, J; Baumeister, C R; Al Dulayymi, J R; Swai, H; Baird, M S; Verschoor, J A

    2016-08-01

    Mycobacterium tuberculosis, the causative agent of tuberculosis, is characterized by the abundance of species specific, antigenic cell wall lipids called mycolic acids. These wax-like molecules all share an identical, amphiphilic mycolic motif, but have different functional groups in a long hydrophobic hydrocarbon mero-chain that divide them into three main classes: alpha-, keto- and methoxy-mycolic acids. Whereas alpha-mycolic acids constitutively maintain an abundance of around 50%, the ratio of methoxy- to keto-mycolic acid types may vary depending on, among other things, the growth stage of M. tuberculosis. In human patients, antibodies to mycolic acids have shown potential as diagnostic serum biomarkers for active TB. Variations in mycolic acid composition affect the antigenic properties and can potentially compromise the precision of detection of anti-mycolic acids antibodies in patient sera to natural mixtures. We demonstrate this here with combinations of synthetic mycolic acid antigens, tested against TB patient and control sera. Combinations of methoxy- and α-mycolic acids are more antigenic than combinations of keto- and α-mycolic acids, showing the former to give a more sensitive test for TB biomarker antibodies. Natural mixtures of mycolic acids isolated from mature cultures of M. tuberculosis H37Rv give the same sensitivity as that with synthetic methoxy- and α-mycolic acids in combination, in a surface plasmon resonance inhibition biosensor test. To ensure that the antigenic activity of isolates of natural mycolic acids is reproducible, we cultured M. tuberculosis H37Rv on Middlebrook 7H10 solid agar plates to stationary growth phase in a standardized, optimal way. The proportions of mycolic acid classes in various batches of the isolates prepared from these cultures were compared to a commercially available natural mycolic acid isolate. LC-MS/MS and NMR data for quantitation of mycolic acids class compositions show that the variation in batches

  14. Detection of Lymph Node Metastases in Human Colorectal Cancer by Using 5-Aminolevulinic Acid-Induced Protoporphyrin IX Fluorescence with Spectral Unmixing

    Kenichi Harada

    2013-11-01

    Full Text Available Accurate evaluation of metastatic lymph nodes (LNs is indispensable for adequate treatment of colorectal cancer (CRC patients. Here, we demonstrate detection of metastases of human CRC in removed fresh LNs using 5-aminolevulinic acid (ALA-induced protoporphyrin IX (PpIX fluorescence. A spectral unmixing method was employed to reduce the overlap of collagen autofluorescence on PpIX fluorescence. A total of 17 surgery patients with advanced CRC were included in this study. After 5-ALA at a dose of 15 mg/kg of body weight was applied orally 2 h prior to surgery, 87 LNs were subjected to spectral fluorescence imaging and histopathological diagnosis, and statistical analysis was performed. No apparent side effect was observed to be associated with 5-ALA administration. The spectral unmixing fluorescence intensity of PpIX in metastatic LNs was 10.2-fold greater than that in nonmetastaic LNs. The receiver-operating-characteristic (ROC analysis showed that the area under the curve (AUC was calculated as 0.95. Our results show the potential of 5-ALA-induced PpIX fluorescence processed by spectral unmixing for detecting metastases in excised fresh LNs from patients with CRC, suggesting that this rapid and feasible method is applicable to gross evaluation of resected LN samples in pathology laboratories.

  15. Production and detection of neutral molecular beams : from single amino acids to biomolecular complexes

    This thesis presents a laser desorption source for neutral organic molecules and clusters as well as the first exploration of a superconducting single photon detector for the detection of massive neutral particles. Whereas the source can produce beams of biomolecules for various gas-phase applications, the detector can be used to overcome the current post- ionization detection mass limit of neutral molecules. The aim of our work is to produce and detect neutral molecular gas-phase beams, ranging from small amino acids overlarge polypeptides to massive complexes. The purpose of creating these beams is to use them for quantum optics experiments, like near field matter wave interference and its applications in metrology. Standard effusive sources usually lack the ability to cool the evaporated organic molecules fast enough to prevent fragmentation. In contrast to that, the presented laser desorption source cools the initially evaporated molecules by embedding them into a supersonic seed gas beam. The mixing of the seed gas and the desorbed molecules is implemented both in free expansion as well as inside a closed mixing channel. The desorbed neutral molecules are detected by photo-ionization using UV (266 nm) and FUV (157 nm) light followed by time-of-flight mass spectrometry. For the amino acid tryptophan (204amu) and for the antibiotic polypeptide gramicidin (1884amu) the ion yields for both photo-ionization wavelengths are examined and the ionization cross sections for the UV wavelength are measured. In case of tryptophan the ionization yield is comparable for both wavelengths, whereas gramicidin is detected fifteen times more efficiently under VUV ionization than for UV ionization at equal intensity. Desorption of heavier molecules than gramicidin never resulted in a detectable ion yield, which confirms the known inefficiency for the post-ionization of isolated large organic molecules [1-3]. The desorption source is also used for the formation of large neutral

  16. Cascade enzymatic catalysis in poly(acrylic acid) brushes-nanospherical silica for glucose detection.

    Zhao, Yan; Wang, Ying; Zhang, Xiaobin; Kong, Rongmei; Xia, Lian; Qu, Fengli

    2016-08-01

    The ultrasensitive monitoring of glucose with a fast and accurate method is significant in potential therapeutics and optimizes protein biosynthesis. Incorporation of enzyme into matrix is considered as promising candidates for constructing highly sensitive glucose-responsive systems. In this study, three-dimensional poly(acrylic acid) brushes-nanospherical silica (PAA-nano silica) with high amplification capability and stability were used to covalently immobilize bienzymes for cascade enzymatic catalysis. The major advantages of PAA-nano silica-bienzyme co-incorporation is that the enzymes are proximity distribution, and such close confinement both minimized the diffusion of intermediates among the enzymes in the consecutive reaction and improve the utilization efficiency of enzymes, thereby enhancing the overall reaction efficiency and specificity. Thus, this present bienzymatic biosensor shows robust signal amplification and ultrasensitivity of glucose-responsive properties with a detection limit of 0.04μM. PMID:27216683

  17. Amperometric Low-Potential Detection of Malic Acid Using Single-Wall Carbon Nanotubes Based Electrodes

    Camelia Bala

    2008-03-01

    Full Text Available The electrocatalytical property of single-wall carbon nanotube (SWNTmodified electrode toward NADH detection was explored by cyclic voltammetry andamperometry techniques. The experimental results show that SWNT decrease theovervoltage required for oxidation of NADH (to 300 mV vs. Ag/AgCl and this propertymake them suitable for dehydrogenases based biosensors. The behavior of the SWNTmodified biosensor for L-malic acid was studied as an example for dehydrogenasesbiosensor. The amperometric measurements indicate that malate dehydrogenase (MDHcan be strongly adsorbed on the surface of the SWNT-modified electrode to form anapproximate monolayer film. Enzyme immobilization in Nafion membrane can increasethe biosensor stability. A linear calibration curve was obtained for L-malic acidconcentrations between 0.2 and 1mM.

  18. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy

    Gary, Ramla; Carbone, Giovani; Petriashvili, Gia; De Santo, Maria Penelope; Barberi, Riccardo

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA. PMID:26907286

  19. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy.

    Gary, Ramla; Carbone, Giovani; Petriashvili, Gia; De Santo, Maria Penelope; Barberi, Riccardo

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA. PMID:26907286

  20. Ultramild protein-mediated click chemistry creates efficient oligonucleotide probes for targeting and detecting nucleic acids

    Nåbo, Lina J.; Madsen, Charlotte Stahl; Jensen, Knud Jørgen;

    2015-01-01

    Functionalized synthetic oligonucleotides are finding growing applications in research, clinical studies, and therapy. However, it is not easy to prepare them in a biocompatible and highly efficient manner. We report a new strategy to synthesize oligonucleotides with promising nucleic acid...... targeting and detection properties. We focus in particular on the pH sensitivity of these new probes and their high target specificity. For the first time, human copper(I)-binding chaperon Cox17 was applied to effectively catalyze click labeling of oligonucleotides. This was performed under ultramild...... conditions with fluorophore, peptide, and carbohydrate azide derivatives. In thermal denaturation studies, the modified probes showed specific binding to complementary DNA and RNA targets. Finally, we demonstrated the pH sensitivity of the new rhodamine-based fluorescent probes in vitro and rationalize our...

  1. Carbon nanotube-based labels for highly sensitive colorimetric and aggregation-based visual detection of nucleic acids

    A novel carbon nanotube (CNT) derived label capable of dramatic signal amplification of nucleic acid detection and direct visual detection of target hybridization has been developed. Highly sensitive colorimetric detection of human acute lymphocytic leukemia (ALL) related oncogene sequences amplified by the novel CNT-based label was demonstrated. Atomic force microscope (AFM) images confirmed that a monolayer of horseradish peroxidase and detection probe molecules was immobilized along the carboxylated CNT carrier. The resulting CNT labels significantly enhanced the nucleic acid assay sensitivity by at least 1000 times compared to that of conventional labels used in enzyme-linked oligosorbent assay (ELOSA). An excellent detection limit of 1 x 10-12 M (60 x 10-18 mol in 60 μl) and a four-order wide dynamic range of target concentration were achieved. Hybridizations using these labels were coupled to a concentration-dependent formation of visible dark aggregates. Targets can thus be detected simply with visual inspection, eliminating the need for expensive and sophisticated detection systems. The approach holds promise for ultrasensitive and low cost visual inspection and colorimetric nucleic acid detection in point-of-care and early disease diagnostic application

  2. Electrogenerated chemiluminescence detection for deoxyribonucleic acid hybridization based on gold nanoparticles carrying multiple probes

    A novel sensitive electrogenerated chemiluminescence (ECL) method for the detection deoxyribonucleic acid (DNA) hybridization based on gold nanoparticles carrying multiple probes was developed. Ruthenium bis(2,2'-bipyridine)(2,2'-bipyridine-4,4'-dicarboxylic acid)-N-hydroxysuccinimide ester (Ru(bpy)2(dcbpy)NHS) was used as a ECL label and gold nanoparticle as a carrier. Probe single strand DNA (ss-DNA) was self-assembled at the 3'-terminal with a thiol group to the surface of gold nanoparticle and covalently labeled at the 5'-terminal of a phosphate group with Ru(bpy)2(dcbpy)NHS and the resulting conjugate (Ru(bpy)2(dcbpy)NHS)-ss-DNA-Au, was taken as a ECL probe. When target analyte ss-DNA was immobilized on a gold electrode by self-assembled monolayer technique and then hybridized with the ECL probe to form a double-stranded DNA (ds-DNA), a strong ECL response was electrochemically generated. The ECL intensity was linearly related to the concentration of the complementary sequence (target ss-DNA) in the range from 1.0 x 10-11 to 1.0 x 10-8 mol L-1, and the linear regression equation was S = 57301 + 4579.6 lg C (unit of C is mol L-1). A detection limit of 5.0 x 10-12 mol L-1 for target ss-DNA was achieved. The ECL signal generated from many reporters of ECL probe prepared is greatly amplified, compared to the convention scheme which is based on one reporter per hybridization event

  3. Determination of Vitamin C (Ascorbic Acid Using High Performance Liquid Chromatography Coupled with Electrochemical Detection

    Ondrej Zitka

    2008-11-01

    Full Text Available Vitamin C (ascorbic acid, ascorbate, AA is a water soluble organic compound that participates in many biological processes. The main aim of this paper was to utilize two electrochemical detectors (amperometric – Coulouchem III and coulometric – CoulArray coupled with flow injection analysis for the detection of ascorbic acid. Primarily, we optimized the experimental conditions. The optimized conditions were as follows: detector potential 100 mV, temperature 25 °C, mobile phase 0.09% TFA:ACN, 3:97 (v/v and flow rate 0.13 mL·min-1. The tangents of the calibration curves were 0.3788 for the coulometric method and 0.0136 for the amperometric one. The tangent of the calibration curve measured by the coulometric detector was almost 30 times higher than the tangent measured by the amperometric detector. Consequently, we coupled a CoulArray electrochemical detector with high performance liquid chromatography and estimated the detection limit for AA as 90 nM (450 fmol per 5 μL injection. The method was used for the determination of vitamin C in a pharmaceutical preparations (98 ± 2 mg per tablet, in oranges (Citrus aurantium (varied from 30 to 56 mg/100 g fresh weight, in apples (Malus sp. (varied from 11 to 19 mg/100 g fresh weight, and in human blood serum (varied from 38 to 78 μM. The recoveries were also determined.

  4. A combinatorial approach to detect coevolved amino acid networks in protein families of variable divergence.

    Julie Baussand

    2009-09-01

    Full Text Available Communication between distant sites often defines the biological role of a protein: amino acid long-range interactions are as important in binding specificity, allosteric regulation and conformational change as residues directly contacting the substrate. The maintaining of functional and structural coupling of long-range interacting residues requires coevolution of these residues. Networks of interaction between coevolved residues can be reconstructed, and from the networks, one can possibly derive insights into functional mechanisms for the protein family. We propose a combinatorial method for mapping conserved networks of amino acid interactions in a protein which is based on the analysis of a set of aligned sequences, the associated distance tree and the combinatorics of its subtrees. The degree of coevolution of all pairs of coevolved residues is identified numerically, and networks are reconstructed with a dedicated clustering algorithm. The method drops the constraints on high sequence divergence limiting the range of applicability of the statistical approaches previously proposed. We apply the method to four protein families where we show an accurate detection of functional networks and the possibility to treat sets of protein sequences of variable divergence.

  5. Mutational Effects on the Folding Dynamics of a Minimized Hairpin‡

    Scian, Michele; Shu, Irene; Olsen, Katherine A.; Hassam, Khalil; Andersen, Niels H.

    2013-01-01

    The fold stabilities and folding dynamics of a series of mutants of a model hairpin, KTW-NPATGK-WTE (HP7), are reported. The parent system and the corresponding DPATGK loop species display sub-μs folding time constants. The mutational studies revealed that ultrafast folding requires both some pre-structuring of the loop and a favorable interaction between the chain termini at the transition state. In the case of YY-DPETGT-WY, another sub-μs folding species [Davis, C. M.; Xiao, S.; Raleigh, D. P.; Dyer, R. B. (2012) J. Am. Chem. Soc. 134, 14476–14482], a hydrophobic cluster provides the latter. In the case of HP7, the Coulombic interaction between the terminal NH3+ and CO2− units provides this; a C-terminal Glu to amidated Ala mutation results in a 5-fold folding rate retardation. The effects of mutations within the reversing loop indicate the balance between loop flexibility (favoring fast conformational searching) and turn-formation in the unfolded state is a major factor in determining the folding dynamics. The –NAAAKX- loops examined display no detectable turn formation propensity in other hairpin constructs, but do result in stable analogs of HP7. Peptide KTW-NAAAKK-WTE displays the same fold stability as HP7 but both the folding and unfolding time constants are greater by a factor of 20. PMID:23521619

  6. Mutational effects on the folding dynamics of a minimized hairpin.

    Scian, Michele; Shu, Irene; Olsen, Katherine A; Hassam, Khalil; Andersen, Niels H

    2013-04-16

    The fold stabilities and folding dynamics of a series of mutants of a model hairpin, KTW-NPATGK-WTE (HP7), are reported. The parent system and the corresponding DPATGK loop species display submicrosecond folding time constants. The mutational studies revealed that ultrafast folding requires both some prestructuring of the loop and a favorable interaction between the chain termini in the transition state. In the case of YY-DPETGT-WY, another submicrosecond folding species [Davis, C. M., Xiao, S., Raleigh, D. P., and Dyer, R. B. (2012) J. Am. Chem. Soc. 134, 14476-14482], a hydrophobic cluster provides the latter. In the case of HP7, the Coulombic interaction between the terminal NH3(+) and CO2(-) units provides this; a C-terminal Glu to amidated Ala mutation results in a 5-fold retardation of the folding rate. The effects of mutations within the reversing loop indicate the balance between loop flexibility (favoring fast conformational searching) and turn formation in the unfolded state is a major factor in determining the folding dynamics. The -NAAAKX- loops examined display no detectable turn formation propensity in other hairpin constructs but do result in stable analogues of HP7. Peptide KTW-NAAAKK-WTE displays the same fold stability as HP7, but both the folding and unfolding time constants are greater by a factor of 20. PMID:23521619

  7. Detection of Acid Rain Stress Effect on Plant Using Hyperspectral Data in Three Gorges Region,China

    SONG Xiaodong; JIANG Hong; YU Shuquan; ZHOU Guomo

    2008-01-01

    This paper aims to use hyperspectral data to detect the spectral change caused by acid stress to a native forest type in the Three Gorges region of China.For this purpose,a ground-based hyperspectral experiment was conducted at the Three Gorges region to detect acid deposition that caused Masson pine (Pinus massoniana) forest degradation.Continuum removal method was used to isolate wavebands more responsive to stress in wavelengths 450-750nm.The differences in chlorophyll concentrations and needle thickness caused by acidic stress are found to be explicable to the different spectral reflectance patterns in the visible and near-infrared wavelengths.Two new chlorotic indices were utilized to explain the stress-caused leaf chiorosis.The comparison of simulated vegetation indices and principal component analysis (PCA) results suggests that it would be possible to monitor acid rain stress effect on forest ecosystem from some wider spectral regions.

  8. Self Contained Encrypted Image Folding

    Rebollo-Neira, Laura; Constantinides, Anthony; Plastino, Angel

    2012-01-01

    The recently introduced approach for Encrypted Image Folding is generalized to make it Self Contained. The goal is achieved by enlarging the folded image so as to embed all the necessary information for the image recovery. The need for extra size is somewhat compensated by considering a transformation with higher folding capacity. Numerical examples show that the size of the resulting cipher image may be significantly smaller than the plain text one. The implementation of the approach is further extended to deal also with color images.

  9. Teaching computers to fold proteins

    Winther, Ole; Krogh, Anders Stærmose

    2004-01-01

    A new general algorithm for optimization of potential functions for protein folding is introduced. It is based upon gradient optimization of the thermodynamic stability of native folds of a training set of proteins with known structure. The iterative update rule contains two thermodynamic averages...... which are estimated by (generalized ensemble) Monte Carlo. We test the learning algorithm on a Lennard-Jones (LJ) force field with a torsional angle degrees-of-freedom and a single-atom side-chain. In a test with 24 peptides of known structure, none folded correctly with the initial potential functions...

  10. Detection of acid moisture in photovoltaic modules using a dual wavelength pH-sensitive fluorescent dye

    Asaka, Takashi; Iwami, Kentaro; Taguchi, Atsushi; Umeda, Norihiro; Masuda, Atsushi

    2014-01-01

    The formation of acetic acid via the penetration of moisture into ethylene vinyl acetate (EVA) in photovoltaic (PV) modules is cited as the main reason for PV modules’ degradation. Currently, there is no effective method for detecting acetic moisture in PV modules. We proposed a simple method for detecting acid moisture in PV modules using a dual-wavelength pH-sensitive dye that measures pH by the ratio of the intensities of two peaks in the fluorescence spectra of the dye. We detected the pH change caused by acetic acid with the change in the intensity ratio of the fluorescence spectra of the dried dye. Furthermore, we observed that the dry fluorescent dye is heat resistant to withstand the lamination process for the manufacturing of PV modules, and has good long-term durability.

  11. Polyoxometalate-Graphene Nanocomposite Modified Electrode for Electrocatalytic Detection of Ascorbic Acid

    Zhang, Weiying; Du, Dan; Gunaratne, Don; Colby, Robert; Lin, Yuehe; Laskin, Julia

    2013-11-15

    Phosphomolybdate functionalized graphene nanocomposite (PMo12-GS) has been successfully formed on a glassy carbon electrode (GCE) for the detection of ascorbic acid (AA). The obtained PMo12-GS modified GCE, was characterized by cyclic voltammetry, electrochemical impedance spectroscopy, scanning electron microscopy (SEM) and Fourier transform infrared (FT-IR) spectroscopy and compared with GCE, GS modified GCE, and PMo12 modified GCE. It shows an increased current and a decrease in over-potential of ~210 mV. The amperometric signals are linearly proportional to the AA concentration in a wide concentration range from 1×10-6 M to 8×10-3 M, with a detection limit of 0.5×10-6 M. Finally, the PMo12-GS modified electrode was employed for the determination of the AA level in vitamin C tablets, with recoveries between 96.3 and 100.8 %.

  12. Molecularly imprinted polyaniline-polyvinyl sulphonic acid composite based sensor for para-nitrophenol detection

    Graphical abstract: -- Highlights: •Biomimetic para-nitrophenol (PNP) imprinted sensor is used to detect PNP. •The PVSA doped polyaniline shows high conductivity. •The sensor shows excellent sensitivity, 18 times reusability and higher stability. •PANI-PVSA composite shows increased stability of the sensor. -- Abstract: We report results of the studies relating to the fabrication and characterization of a conducting polymer based molecularly imprinted para-nitrophenol (PNP) sensor. A water pollutant, para-nitrophenol is electrochemically imprinted with polyvinyl sulphonic acid (PVSA) doped polyaniline onto indium tin oxide (ITO) glass substrate. This PNP imprinted electrode (PNPI-PANI-PVSA/ITO) prepared via chronopotentiometric polymerization and over-oxidation is characterized by Fourier transform infra-red spectroscopy (FT-IR), UV–visible (UV–vis) spectroscopy, contact angle (CA), scanning electron microscopy (SEM), cyclic voltammetry (CV) and differential pulse voltammetry (DPV) studies. The response studies of PNPI-PANI-PVSA/ITO electrode carried out using DPV reveal a lower detection limit of 1 × 10−3 mM, improved sensitivity as 1.5 × 10−3 A mM−1 and stability of 45 days. The PNPI-PANI-PVSA/ITO electrode shows good precision with relative standard deviation of 2.1% and good reproducibility with standard deviation of 3.78%

  13. Improved microscopical detection of acid-fast bacilli by the modified bleach method in lymphnode aspirates

    Annam Vamseedhar

    2009-07-01

    Full Text Available Objectives: To improve the smear microscopy for detection of acid-fast bacilli (AFB in fine needle aspiration cytology (FNAC of lymph node using the bleach method and also to compare this with cytological diagnosis and the conventional Ziehl-Neelsen (ZN method. Study Design: In 99 consecutive patients with clinical suspicion of tuberculosis (TB presenting with lymphadenopathy, FNACs were performed. Smears from the aspirates were processed for routine cytology and the conventional ZN method. The remaining material in the needle hub and/or the syringe was used for the bleach method. The significance of the bleach method over the conventional ZN method and cytology was analyzed using the χ2 test. Results: Of 99 aspirates, 93 were studied and the remaining six were excluded from the study due to diagnosis of malignancy in 4.04% (4/6 and inadequate aspiration in 2.02% (2/6. Among the 93 aspirates, 33.33% (31/93 were positive for AFB on conventional ZN method, 41.94% (39/93 were indicative of TB on cytology and the smear positivity increased to 63.44% (59/93 on bleach method. Conclusion: The bleach method is simple, inexpensive and potent disinfectant, also limiting the risk of laboratory-acquired infections. The implementation of the bleach method clearly improves microscopic detection and can be a useful contribution to routine cytology.

  14. Capillary electrophoresis based on nucleic acid detection for diagnosing human infectious disease.

    Lian, Dong-Sheng; Zhao, Shu-Jin

    2016-04-01

    Rapid transmission, high morbidity, and mortality are the features of human infectious diseases caused by microorganisms, such as bacteria, fungi, and viruses. These diseases may lead within a short period of time to great personal and property losses, especially in regions where sanitation is poor. Thus, rapid diagnoses are vital for the prevention and therapeutic intervention of human infectious diseases. Several conventional methods are often used to diagnose infectious diseases, e.g. methods based on cultures or morphology, or biochemical tests based on metabonomics. Although traditional methods are considered gold standards and are used most frequently, they are laborious, time consuming, and tedious and cannot meet the demand for rapid diagnoses. Disease diagnosis using capillary electrophoresis methods has the advantages of high efficiency, high throughput, and high speed, and coupled with the different nucleic acid detection strategies overcomes the drawbacks of traditional identification methods, precluding many types of false positive and negative results. Therefore, this review focuses on the application of capillary electrophoresis based on nucleic detection to the diagnosis of human infectious diseases, and offers an introduction to the limitations, advantages, and future developments of this approach. PMID:26352354

  15. PREFACE Protein folding: lessons learned and new frontiers Protein folding: lessons learned and new frontiers

    Pappu, Rohit V.; Nussinov, Ruth

    2009-03-01

    In appropriate physiological milieux proteins spontaneously fold into their functional three-dimensional structures. The amino acid sequences of functional proteins contain all the information necessary to specify the folds. This remarkable observation has spawned research aimed at answering two major questions. (1) Of all the conceivable structures that a protein can adopt, why is the ensemble of native-like structures the most favorable? (2) What are the paths by which proteins manage to robustly and reproducibly fold into their native structures? Anfinsen's thermodynamic hypothesis has guided the pursuit of answers to the first question whereas Levinthal's paradox has influenced the development of models for protein folding dynamics. Decades of work have led to significant advances in the folding problem. Mean-field models have been developed to capture our current, coarse grain understanding of the driving forces for protein folding. These models are being used to predict three-dimensional protein structures from sequence and stability profiles as a function of thermodynamic and chemical perturbations. Impressive strides have also been made in the field of protein design, also known as the inverse folding problem, thereby testing our understanding of the determinants of the fold specificities of different sequences. Early work on protein folding pathways focused on the specific sequence of events that could lead to a simplification of the search process. However, unifying principles proved to be elusive. Proteins that show reversible two-state folding-unfolding transitions turned out to be a gift of natural selection. Focusing on these simple systems helped researchers to uncover general principles regarding the origins of cooperativity in protein folding thermodynamics and kinetics. On the theoretical front, concepts borrowed from polymer physics and the physics of spin glasses led to the development of a framework based on energy landscape theories. These

  16. Anion-exchange high-performance liquid chromatography with conductivity detection for the analysis of phytic acid in food

    Talamond, Pascale; Doulbeau, Sylvie; Rochette, Isabelle; Guyot, Jean-Pierre; Trèche, Serge

    1999-01-01

    A sensitive method for the accurate determination of phytic acid in food samples is described. The proposed procedure involves the anion-exchange liquid chromatography with conductivity detection. Initially, two methods of determination of phytic acid were compared : absorptiometry and high-performance ion chromatography (HPIC) with chemically suppressed conductivity detector. Unlike most conventional methods involving precipitation by FeCl3, the simpler and more reliable HPIC assay avoids th...

  17. Adipose-Derived Mesenchymal Stem Cells in the Regeneration of Vocal Folds: A Study on a Chronic Vocal Fold Scar

    Angelou Valerie

    2016-01-01

    Full Text Available Background. The aim of the study was to assess the histological effects of autologous infusion of adipose-derived stem cells (ADSC on a chronic vocal fold scar in a rabbit model as compared to an untreated scar as well as in injection of hyaluronic acid. Study Design. Animal experiment. Method. We used 74 New Zealand rabbits. Sixteen of them were used as control/normal group. We created a bilateral vocal fold wound in the remaining 58 rabbits. After 18 months we separated our population into three groups. The first group served as control/scarred group. The second one was injected with hyaluronic acid in the vocal folds, and the third received an autologous adipose-derived stem cell infusion in the scarred vocal folds (ADSC group. We measured the variation of thickness of the lamina propria of the vocal folds and analyzed histopathologic changes in each group after three months. Results. The thickness of the lamina propria was significantly reduced in the group that received the ADSC injection, as compared to the normal/scarred group. The collagen deposition, the hyaluronic acid, the elastin levels, and the organization of elastic fibers tend to return to normal after the injection of ADSC. Conclusions. Autologous injection of adipose-derived stem cells on a vocal fold chronic scar enhanced the healing of the vocal folds and the reduction of the scar tissue, even when compared to other treatments.

  18. Analysis of olfactory sensitivity in rainbow trout (Oncorhynchus mykiss) reveals their ability to detect lactic acid, pyruvic acid and four B vitamins.

    Valdés, Joaquín; Olivares, Jesús; Ponce, Daniela; Schmachtenberg, Oliver

    2015-08-01

    Salmonid fishes like the rainbow trout Oncorhynchus mykiss have a highly developed olfactory sense that allows them to perceive some odorants at very low concentrations, such as certain amino acids and bile salts. Previous behavioral and electrophysiological studies in salmonids have shown strong responses to human skin odor. Because this stimulus represents a complex and heterogeneous mixture of components, we sought to determine which odorants contribute to the sensitive detection of human skin odor by salmonids. In vivo electroolfactogram recordings in O. mykiss revealed lactic acid, pyruvic acid and two B vitamins, thiamine and riboflavin, as novel, potent odorants which triggered responses at nanomolar concentrations. Two more B vitamins, nicotinic and pantothenic acid, were detected at micromolar concentrations. These compounds share important roles in cellular energy metabolism, supporting an original role in food search and feeding behavior of this species and most likely other fishes. The olfactory detection of B vitamins by salmonids represents a new paradigm in chemosensation, warranting further investigation in other teleosts. PMID:25864178

  19. Fog spontaneously folds mosquito wings

    Dickerson, Andrew K.; Liu, Xing; Zhu, Ting; Hu, David L.

    2015-02-01

    The flexibility of insect wings confers aerodynamic benefits, but can also present a hazard if exposed to fog or dew. Fog can cause water to accumulate on wings, bending them into tight taco shapes and rendering them useless for flight. In this combined experimental and theoretical study, we use high-speed video to film the spontaneous folding of isolated mosquito wings due to the evaporation of a water drop. We predict shapes of the deformed wing using two-dimensional elastica theory, considering both surface tension and Laplace pressure. We also recommend fold-resistant geometries for the wings of flapping micro-aerial vehicles. Our work reveals the mechanism of insect wing folding and provides a framework for further study of capillarity-driven folding in both natural and biomimetic systems at small scales.

  20. A naphthalene-based two-photon fluorescent probe for selective and sensitive detection of endogenous hypochlorous acid.

    Zhou, Xiao-Hong; Jiang, Yu-Ren; Zhao, Xiong-Jie; Guo, Dong

    2016-11-01

    An efficient naphthalene-based two-photon fluorescent probe for endogenous HClO has been reported in the present study, which consists of a 6-(2-benzothiazolyl)-2-naphthalenol fluorophore connected with a 4-aminophenol (the fluorescence quenching and response group). This probe exhibits a high selectivity and excellent sensitivity with a detection limit of 7.6nM over other reactive oxygen species and analyte species, and the fluorescence intensity enhanced 103-fold when responsed. Furthermore, it was successfully used for two-photon imaging of endogenous HClO in live cells with high-resolution. PMID:27591640

  1. Rapid diagnosis of rotavirus infection by direct detection of viral nucleic acid in silver-stained polyacrylamide gels.

    Herring, A J; Inglis, N F; Ojeh, C K; Snodgrass, D R; Menzies, J D

    1982-01-01

    A rapid simple technique for the diagnosis of rotavirus has been developed based on the sensitive detection of rotavirus double-stranded RNA genome segments separated in polyacrylamide gels. The method utilizes a recently described ultrasensitive silver stain for polypeptides, which can also detect subnanogram amounts of nucleic acid. The sensitivity of the technique is comparable with that of electron microscopy or enzyme-linked immunosorbent assay.

  2. Real-Time Nucleic Acid Sequence-Based Amplification Using Molecular Beacons for Detection of Enterovirus RNA in Clinical Specimens

    Landry, Marie L.; Garner, Robin; Ferguson, David

    2005-01-01

    Real-time nucleic acid sequence-based amplification (NASBA) using molecular beacon technology (NASBA-beacon) was compared to standard NASBA with postamplification hybridization using electrochemiluminescently labeled probes (NASBA-ECL) for detection of enteroviruses (EV) in 133 cerebrospinal fluid and 27 stool samples. NASBA-ECL and NASBA-beacon were similar in sensitivity, detecting 55 (100%) and 52 (94.5%) EV-positive samples, respectively. There were no false positives. Both NASBA assays w...

  3. Simultaneous detection of cyclopiazonic acid and aflatoxin B1 by HPLC in methanol/water mobile phase

    Soares, Célia Maria Gonçalves; Freitas-Silva, O.; Abrunhosa, Luís; Venâncio, Armando

    2009-01-01

    A simple procedure for the simultaneous detection of cyclopiazonic acid (CPA) and aflatoxin B1 from fungal extracts is presented, using a methanol and water mobile phase and fluorescence detection. This methodology has been tested with standard solutions of both mycotoxins CPA and Aflatoxin B1 and with methanolic extracts of Aspergillus section Flavi strains, previously characterized for their mycotoxin production profile. Previously available methodology required the use of two different c...

  4. Qualitative analysis of some carboxylic acids by ion-exclusion chromatography with atmospheric pressure chemical ionization mass spectrometric detection.

    Helale, Murad I H; Tanaka, Kazuhiko; Taoda, Hiroshi; Hu, Wenzhi; Hasebe, Kiyoshi; Haddad, Paul R

    2002-05-17

    A simple, selective and sensitive method for the determination of carboxylic acids has been developed. A mixture of formic, acetic, propionic, valeric, isovaleric, isobutyric, and isocaproic acids has been separated on a polymethacrylate-based weak acidic cation-exchange resin (TSK gel OA pak-A) based on an ion-exclusion chromatographic mechanism with detection using UV-photodiode array, conductivity and atmospheric pressure chemical ionization mass spectrometry (APCI-MS). A mobile phase consisting of 0.85 mM benzoic acid in 10% aqueous methanol (pH 3.89) was used to separate the above carboxylic acids in about 40 min. For LC-MS, the APCI interface was used in the negative ionization mode. Linear plots of peak area versus concentration were obtained over the range 1-30 mM (r2=0.9982) and 1-30 mM (r2=0.9958) for conductimetric and MS detection, respectively. The detection limits of the target carboxylic acids calculated at S/N=3 ranged from 0.078 to 2.3 microM for conductimetric and photometric detection and from 0.66 to 3.82 microM for ion-exclusion chromatography-APCI-MS. The reproducibility of retention times was 0.12-0.16% relative standard deviation for ion-exclusion chromatography and 1.21-2.5% for ion-exclusion chromatography-APCI-MS. The method was applied to the determination of carboxylic acids in red wine, white wine, apple vinegar, and Japanese rice wine. PMID:12108651

  5. NMR and protein folding: equilibrium and stopped-flow studies.

    Frieden, C.; Hoeltzli, S D; Ropson, I. J.

    1993-01-01

    NMR studies are now unraveling the structure of intermediates of protein folding using hydrogen-deuterium exchange methodologies. These studies provide information about the time dependence of formation of secondary structure. They require the ability to assign specific resonances in the NMR spectra to specific amide protons of a protein followed by experiments involving competition between folding and exchange reactions. Another approach is to use 19F-substituted amino acids to follow change...

  6. Abnormal myocardial fatty acid metabolism in dilated cardiomyopathy detected by iodine-123 phenylpentadecanoic acid and tomographic imaging

    The radioidinated synthetic fatty acid iodine-123 phenylpentadecanoic acid (IPPA) has proven useful in the identification of regional abnormalities of cardiac metabolism in patients with myocardial ischemia. The present study was performed to test the hypothesis that the myocardial distribution and turnover of fatty acids, assessed noninvasively with IPPA, are altered in patients with cardiomyopathy. Nine normal volunteers and 19 patients with dilated cardiomyopathy of various etiologies underwent cardiac imaging with single-photon emission computed tomography (SPECT) after intravenous injection of IPPA. Apical short-axis and basal short-axis sections were reconstructed and quantitatively analyzed for relative IPPA activity distribution and washout. Patients with congestive cardiomyopathy demonstrated significantly greater heterogeneity of IPPA uptake than normal subjects (maximal percent variation of activity 27 +/- 11 vs 18 +/- 4, p less than 0.01). They also demonstrated a more rapid percent washout rate than control subjects (24 +/- 8 vs 17 +/- 6 for the apical short-axis section, p less than 0.05; 26 +/- 7 vs 18 +/- 5 for the basal short-axis section, p less than 0.01). These abnormalities of fatty acid distribution and turnover were independent of the etiology of the cardiomyopathy. The degree of heterogeneity of IPPA uptake was significantly related to the patients' New York Heart Association functional class (r = 0.64, p less than 0.01). Thus, compared with normal myocardium, the myocardium of patients with congestive cardiomyopathy demonstrates a more heterogeneous distribution of fatty acid uptake, which parallels the clinical severity of the disease. Furthermore, patients with congestive cardiomyopathy demonstrate a more rapid myocardial clearance of the labeled fatty acid, as assessed with SPECT imaging

  7. Co-transcriptional folding is encoded within RNA genes

    Miklós István

    2004-08-01

    Full Text Available Abstract Background Most of the existing RNA structure prediction programs fold a completely synthesized RNA molecule. However, within the cell, RNA molecules emerge sequentially during the directed process of transcription. Dedicated experiments with individual RNA molecules have shown that RNA folds while it is being transcribed and that its correct folding can also depend on the proper speed of transcription. Methods The main aim of this work is to study if and how co-transcriptional folding is encoded within the primary and secondary structure of RNA genes. In order to achieve this, we study the known primary and secondary structures of a comprehensive data set of 361 RNA genes as well as a set of 48 RNA sequences that are known to differ from the originally transcribed sequence units. We detect co-transcriptional folding by defining two measures of directedness which quantify the extend of asymmetry between alternative helices that lie 5' and those that lie 3' of the known helices with which they compete. Results We show with statistical significance that co-transcriptional folding strongly influences RNA sequences in two ways: (1 alternative helices that would compete with the formation of the functional structure during co-transcriptional folding are suppressed and (2 the formation of transient structures which may serve as guidelines for the co-transcriptional folding pathway is encouraged. Conclusions These findings have a number of implications for RNA secondary structure prediction methods and the detection of RNA genes.

  8. Thermostability in endoglucanases is fold-specific

    Wolt Jeffrey D

    2011-02-01

    Full Text Available Abstract Background Endoglucanases are usually considered to be synergistically involved in the initial stages of cellulose breakdown-an essential step in the bioprocessing of lignocellulosic plant materials into bioethanol. Despite their economic importance, we currently lack a basic understanding of how some endoglucanases can sustain their ability to function at elevated temperatures required for bioprocessing, while others cannot. In this study, we present a detailed comparative analysis of both thermophilic and mesophilic endoglucanases in order to gain insights into origins of thermostability. We analyzed the sequences and structures for sets of endoglucanase proteins drawn from the Carbohydrate-Active enZymes (CAZy database. Results Our results demonstrate that thermophilic endoglucanases and their mesophilic counterparts differ significantly in their amino acid compositions. Strikingly, these compositional differences are specific to protein folds and enzyme families, and lead to differences in intramolecular interactions in a fold-dependent fashion. Conclusions Here, we provide fold-specific guidelines to control thermostability in endoglucanases that will aid in making production of biofuels from plant biomass more efficient.

  9. Analysis of protein folds using protein contact networks

    Pankaj Barah; Somdatta Sinha

    2008-08-01

    Proteins are important biomolecules, which perform diverse structural and functional roles in living systems. Starting from a linear chain of amino acids, proteins fold to different secondary structures, which then fold through short- and long-range interactions to give rise to the final three-dimensional shapes useful to carry out the biophysical and biochemical functions. Proteins are defined as having a common `fold' if they have major secondary structural elements with same topological connections. It is known that folding mechanisms are largely determined by a protein's topology rather than its interatomic interactions. The native state protein structures can, thus, be modelled, using a graph-theoretical approach, as coarse-grained networks of amino acid residues as `nodes' and the inter-residue interactions/contacts as `links'. Using the network representation of protein structures and their 2D contact maps, we have identified the conserved contact patterns (groups of contacts) representing two typical folds – the EF-hand and the ubiquitin-like folds. Our results suggest that this direct and computationally simple methodology can be used to infer about the presence of specific folds from the protein's contact map alone.

  10. Quantitative detection of uric acid by electrochemical-surface enhanced Raman spectroscopy using a multilayered Au/Ag substrate.

    Zhao, Lili; Blackburn, Jonathan; Brosseau, Christa L

    2015-01-01

    Uric acid is a potential important biomarker in urine and serum samples for early diagnosis of preeclampsia, a life-threatening hypertensive disorder that occurs during pregnancy. Preeclampsia is a leading cause of maternal death, especially in developing nation settings. Quantitative detection of uric acid for rapid and routine diagnosis of early preeclampsia using electrochemical-surface enhanced Raman spectroscopy (EC-SERS) is presented herein. A uniform EC-SERS active Au/Ag substrate was developed by depositing nearly monodisperse gold and silver nanoparticles on the carbon working electrode surface of screen printed electrodes. The multilayered Au/Ag substrates were characterized by electron microscopy and used for quantitative detection of uric acid in 0.1 M NaF and synthetic urine at clinically relevant concentrations. These results showed a linear relationship between the EC-SERS signal intensity and the uric acid concentration. Relative errors calculated for selected concentrations were all within the Clinical Laboratory Improvement Amendments (CLIA) criterion for uric acid analysis (±17%). It is believed that routine and early diagnosis of disease could be possible through such quantitative detection of biomarkers in patient samples using this EC-SERS method. PMID:25483146

  11. Simultaneous detection of ascorbic acid, dopamine, uric acid and tryptophan with Azure A-interlinked multi-walled carbon nanotube/gold nanoparticles composite modified electrode

    Hayati Filik

    2016-05-01

    Full Text Available In this paper, multi-walled carbon nanotube/Azure A/gold nanoparticle composites (Nafion/AuNPs/AzA/MWCNTs were prepared by binding gold nanoparticles to the surfaces of Azure A-coated carbon nanotubes. Nafion/AuNPs/AzA/MWCNTs based electrochemical sensor was fabricated for the simultaneous determination of ascorbic acid, dopamine, uric acid, and tryptophan. Cyclic voltammetry and electrochemical impedance spectroscopy were used to characterize the electrochemical properties of the modified electrodes. The modified electrode showed excellent electrocatalytic activity toward ascorbic acid, dopamine, uric acid, and tryptophan (pH 7.0. The experiment results showed that the linear response range for simultaneous detection of AA, DA, UA and Trp were 300–10,000 μM, 0.5–50 μM, 0.5–50 μM and 1.0–100 μM, respectively, and the detection limits were 16 μM, 0.014 μM, 0.028 μM and 0.56 μM (S/N = 3. The proposed method offers promise for simple, rapid, selective and cost-effective analysis of small biomolecules. The procedure was also applied to the determination of tryptophan in spiked milk samples.

  12. First improvements in the detection and quantification of label-free nucleic acids by laser-induced breakdown spectroscopy: Application to the deoxyribonucleic acid micro-array technology

    The accurate quantification of nucleic acids is essential in many fields of modern biology and industry, and in some cases requires the use of fluorescence labeling. Yet, in addition to standardization problems and quantification reproducibility, labeling can modify the physicochemical properties of molecules or affect their stability. To address these limitations, we have developed a novel method to detect and quantify label-free nucleic acids. This method is based on stoichiometric proportioning of phosphorus in the nucleic acid skeleton, using laser-induced breakdown spectroscopy, and a specific statistical analysis, which indicates the error probability for each measurement. The results obtained appear to be quantitative, with a limit of detection of 105 nucleotides/μm2 (i.e. 2 x 1013 phosphorus atoms/cm2). Initial micro-array analysis has given very encouraging results, which point to new ways of quantifying hybridized nucleic acids. This is essential when comparing molecules of different sequences, which is presently very difficult with fluorescence labeling

  13. Phylogenetic group- and species-specific oligonucleotide probes for single-cell detection of lactic acid bacteria in oral biofilms

    Quevedo, Beatrice; Giertsen, Elin; Zijnge, Vincent; Luethi-Schaller, Helga; Guggenheim, Bernhard; Thurnheer, Thomas; Gmuer, Rudolf

    2011-01-01

    Background: The purpose of this study was to design and evaluate fluorescent in situ hybridization (FISH) probes for the single-cell detection and enumeration of lactic acid bacteria, in particular organisms belonging to the major phylogenetic groups and species of oral lactobacilli and to Abiotroph

  14. Quantification of fold growth of frontal antiforms in the Zagros fold and thrust belt (Kurdistan, NE Iraq)

    Bretis, Bernhard; Bartl, Nikolaus; Graseman, Bernhard; Lockhart, Duncan

    2010-05-01

    The Zagros fold and thrust belt is a seismically active orogen, where actual kinematic models based on GPS networks suggest a north-south shortening between Arabian and Eurasian in the order of 1.5-2.5 cm/yr. Most of this deformation is partitioned in south-southwest oriented folding and thrusting with northwest-southeast to north-south trending dextral strike slip faults. The Zagros fold and thrust belt is of great economic interest because it has been estimated that this area contains about 15% of the global recoverable hydrocarbons. Whereas the SE parts of the Zagros have been investigated by detailed geological studies, the NW extent being part of the Republic of Iraq have experienced considerably less attention. In this study we combine field work and remote sensing techniques in order to investigate the interaction of erosion and fold growth in the area NE of Erbil (Kurdistan, Iraq). In particular we focus on the interaction of the transient development of drainage patterns along growing antiforms, which directly reflects the kinematics of progressive fold growth. Detailed geomorphological studies of the Bana Bawi-, Permam- and Safeen fold trains show that these anticlines have not developed from subcylindrical embryonic folds but they have merged from different fold segments that joined laterally during fold amplification. This fold segments with length between 5 and 25 km have been detected by mapping ancient and modern river courses that initially cut the nose of growing folds and eventually got defeated leaving behind a wind gap. Fold segments, propagating in different directions force rivers to join resulting in steep gorges, which dissect the merging fold noses. Along rapidly lateral growing folds (e.g. at the SE end of the Bana Bawi Anticline) we observed "curved wind gaps", a new type of abandoned river course, where form of the wind gap mimics a formed nose of a growing antiform. The inherited curved segments of uplifted curved river courses strongly

  15. THE APPLICATION OF PEPTIDE NUCLEIC ACID PROBES FOR RAPID DETECTION AND ENUMERATION OF EUBACTERIA, STAPHYLOCOCCUS AUREUS AND PSEUDOMONAS AERUGINOSA IN RECREATIONAL BEACHES OF S. FLORIDA. (R828830)

    A novel chemiluminescent in situ hybridization technique using peptide nucleic acids (PNA) was adapted for the detection of bacteria in beach sand and recreational waters in South Florida. The simultaneous detection and enumeration of eubacteria and the novel indicators, S...

  16. Detection of Serum Hyaluronic Acid and Laminin in Patients with Bladder Tumors

    李令勋; 丁国富

    2003-01-01

    In order to investigate the changes of serum hyaluronic acid (HA) and laminin (LN) levels and their clinical implication in the patients with bladder tumors, the serum HA and LN levels in 34 patients with bladder tumor and 30 cases of control group were detected by radioimmunoassay before and after operation. The results showed that the serum HA and LN levels in the patients with bladder tumors were significantly higher than those in control group (P<0. 01) before operation, and decreased significantly after operation (P<0. 05). The serum levels of HA and LN in infiltration tumors were higher than those in superficial tumors (P<0.05). The serum HA and LN levels in patients with lymph node metastasis were higher than those without lymph node metastasis (P<0.01 ). The investigation revealed that HA and LN might be involved in the malignant biology behavior of bladder tumors and could be used as important markers of assistant diagnosis and condition monitoring.

  17. Colorimetric detection of lead (II) based on silver nanoparticles capped with iminodiacetic acid

    We report on a colorimetric probe for the determination of Pb(II). It is based on the use of silver nanoparticles that have been functionalized with iminodiacetic acid (IDA-Ag NPs). The absorption spectrum and solution color of IDA-Ag NPs undergo dramatic changes on exposure to Pb(II) with a new absorption peak appearing at 650 nm and a concomitant color change from yellow to green. This is assumed to result from the aggregation of IDA-Ag NPs induced by Pb(II). Under optimum conditions, there is a linear relationship between the ratio of the absorbances at 650 and 396 nm, respectively, and the concentration of Pb(II) in the 0.4 to 8.0 μM concentration range, with a detection limit of 13 nM. The method was applied to the determination of Pb(II) in tap water and urea samples, and recoveries ranged from 93.7 % to 98.6 %. (author)

  18. Detection of diaphragmatic disruptions by peritoneoscintigraphy using technetium-99M diethylene-triamine pentacetic acid

    Intraperitoneal injection of a selected radiopharmaceutical results in the diffusion of radioactive material throughout the peritoneum. A diaphragmatic injury should theoretically result in the diffusion of the radioactive material into the chest. To test this hypothesis, Technetium-99m diethylene-triamine pentacetic acid (Tc-99m DTPA) was administered intraperitoneally by either direct needle injection or catheter into 18 rabbits. Four of the rabbits served as controls and did not have any diaphragmatic injury. Fourteen rabbits had surgically induced diaphragmatic tears of varying size (1/4 to 1 cm) after thoracotomy. Four of the 14 rabbits were dropped from the study because they had inadequate peritoneal injections of the radiopharmaceutical. The remaining ten rabbits showed peritoneoscintigraphic evidence of diaphragmatic injury either by showing passage of the radiotracer into the chest, demonstrating the site of injury as a focal region of increased radiotracer uptake, or showing both of these features. Peritoneoscintigraphy appears to be a potentially useful modality in the detection of diaphragmatic injury

  19. Cytotoxicity detection of poly(lactic-co-glycolic acid/tricalcium phosphate

    Meng SUN

    2011-12-01

    Full Text Available Objective To detecte the cytotoxicity of the PLGA/TCP(poly(lactic-co-glycolic acid/Tricalcium phosphate composite that based on the precedent experiments conducted in Tsinghua University.Methods Compared with the PLGA scaffold material,observated the surface and interior structure of the PLGA/TCP scaffold material by SEM(scanning electron microscope,the surface and interior of PLGA/TCP scaffold material appeared to be homogeneous porous under SEM,with fairly even porosity distribution.The pore diameter was approximately 400μm.The interpenetrative micro-pores were scattered over bigger pores’ periphery with approximately circular contour and 3~5 μm in diameter.These pores were interpenetrative,the average factor of porosity was 89.6%.And which selected rat L929 cell strain,and detected the cytotoxicity of the PLGA/TCP composite in vitro by MTT method.Results The surface and interior of PLGA/TCP scaffold material appeared to be homogeneous porous under SEM,with fairly even porosity distribution.The pore diameter was approximately 400μm.The interpenetrative micro-pores were scattered over bigger pores’ periphery with approximately circular contour and 3~5 μm in diameter.These pores were interpenetrative,the average factor of porosity was 89.6%.On rat L929 cell strain,used MTT Method to detect the cytotoxicity of the composite PLGA/ TCP in vitro,the result showed that the cytotoxicity of the PLGA/TCP composite was level I,according to the criterion,it can be considered as non cytotoxic.Conclusion This research has proved that the PLGA/TCP compound scaffold material has a more homogeneous structure,with the vesicular interior and the structure of PLGA/TCP composite is similar to natural bone trabecula,PLGA/TCP is non cytotoxicity,which satisfy the basic requirement of biological material application and provides a good experimental foundation for repairing autologous bone defect in the near future.

  20. Folding and escape of nascent proteins at ribosomal exit tunnel

    Bui, Phuong Thuy; Hoang, Trinh Xuan

    2016-03-01

    We investigate the interplay between post-translational folding and escape of two small single-domain proteins at the ribosomal exit tunnel by using Langevin dynamics with coarse-grained models. It is shown that at temperatures lower or near the temperature of the fastest folding, folding proceeds concomitantly with the escape process, resulting in vectorial folding and enhancement of foldability of nascent proteins. The concomitance between the two processes, however, deteriorates as temperature increases. Our folding simulations as well as free energy calculation by using umbrella sampling show that, at low temperatures, folding at the tunnel follows one or two specific pathways without kinetic traps. It is shown that the escape time can be mapped to a one-dimensional diffusion model with two different regimes for temperatures above and below the folding transition temperature. Attractive interactions between amino acids and attractive sites on the tunnel wall lead to a free energy barrier along the escape route of the protein. It is suggested that this barrier slows down the escape process and consequently promotes correct folding of the released nascent protein.

  1. Water soluble and efficient amino acid Schiff base receptor for reversible fluorescence turn-on detection of Zn²⁺ ions: Quantum chemical calculations and detection of bacteria.

    Subha, L; Balakrishnan, C; Natarajan, Satheesh; Theetharappan, M; Subramanian, Balanehru; Neelakantan, M A

    2016-01-15

    An amino acid Schiff base (R) capable of recognizing Zn(2+) ions selectively and sensitively in an aqueous medium was prepared and characterized. Upon addition of Zn(2+) ions, the receptor exhibits fluorescence intensity enhancements (~40 fold) at 460 nm (quantum yield, Φ=0.05 for R and Φ=0.18 for R-Zn(2+)) and can be detected by naked eye under UV light. The receptor can recognize the Zn(2+) (1.04×10(-8) M) selectively for other metal ions in the pH range of 7.5-11. The Zn(2+) chelation with R decreases the loss of energy through non-radiative transition and leads to fluorescence enhancement. The binding mode of the receptor with Zn(2+) was investigated by (1)H NMR titration and further validated by ESI-MS. The elemental color mapping and SEM/EDS analysis were also used to study the binding of R with Zn(2+). Density functional theory calculations were carried out to understand the binding mechanism. The receptor was applied as a microbial sensor for Escherichia coli and Staphylococcus aureus. PMID:26318699

  2. Evaluation of [1-11C]-α-aminoisobutyric acid for tumor detection and amino acid transport measurement: Spontaneous canine tumor studies

    Alpha-aminoisobutyric acid (AIB) or α-methyl alanine, is a nonmetabolized amino acid treansported into cells particularly malignant cells, predominantly by the ''A'' amino acid transport system. Since it is not metabolized, [1-11C]-AIB can be used to quantify A-type amino acid transport into cells using a relatively simple compartmental model and quantitative imaging procedures (e.g. positron tomography). The tissue distribution of [1-11C]-AIB was determined in six dogs bearing spontaneous tumors, including lymphosarcoma, osteogenic sarcoma, mammary carcinoma, and adenocarcinoma. Quantitative imaging with tissue radioassay confirmation at necropsy showed poor to excellent tumor localization. However, in all cases the concentrations achieved appear adequate for amino acid transport measurement at known tumor locations. The observed low normal brain (due to blood-brain barrier exclusion) and high (relative to brain) tumor concentrations of [1-11C]-AIB suggest that this agent may prove effective for the early detection of human brain tumors. (orig.)

  3. Wheat gluten amino acid analysis by high-performance anion-exchange chromatography with integrated pulsed amperometric detection.

    Rombouts, Ine; Lagrain, Bert; Lamberts, Lieve; Celus, Inge; Brijs, Kristof; Delcour, Jan A

    2012-01-01

    This chapter describes an accurate and user-friendly method for determining amino acid composition of wheat gluten proteins and their gliadin and glutenin fractions. The method consists of hydrolysis of the peptide bonds in 6.0 M hydrochloric acid solution at 110°C for 24 h, followed by evaporation of the acid and separation of the free amino acids by high-performance anion-exchange chromatography with integrated pulsed amperometric detection. In contrast to conventional methods, the analysis requires neither pre- or postcolumn derivatization, nor a time-consuming oxidation or derivatization step prior to hydrolysis. Correction factors account for incomplete release of Val and Ile even after hydrolysis for 24 h, and for losses of Ser during evaporation. Gradient conditions including an extra eluent allow multiple sequential sample analyses without risk of Glu accumulation on the anion-exchange column which otherwise would result from high Gln levels in gluten proteins. PMID:22125156

  4. A highly sensitive and stable electrochemical sensor for simultaneous detection towards ascorbic acid, dopamine, and uric acid based on the hierarchical nanoporous PtTi alloy.

    Zhao, Dianyun; Yu, Guolong; Tian, Kunlong; Xu, Caixia

    2016-08-15

    In current work highly sensitive and stable electrochemical sensor for simultaneous detection of ascorbic acid (AA), dopamine (DA), and uric acid (UA) is constructed based on the hierarchical nanoporous (HNP) PtTi alloy. The HNP-PtTi alloy is simply fabricated by two-step dealloying process, characterized by the bimodal ligament/pore size distributions and interconnected hollow channels. The HNP structure with the advantages of large surface area, excellent structure stability, and rich pore channels is used for facilitating the electron conductivity and the mass transfer. Combined with the dual effects of the bimodal nanoporous architecture and the excellent electrocatalytic activity of PtTi alloy, the constructed sensor exhibits high electrochemical sensing activity with wide linear responses from 0.2 to 1mM, 0.004 to 0.5mM, and 0.1 to 1mM for simultaneous detection of AA, DA, and UA, respectively. In addition, HNP-PtTi alloy also shows long-term sensing stability towards the AA, DA, and UA detection and behaves as a good anti-interference towards NaCl, KCl, FeCl3, CuCl2, AlCl3, glucose, and H2O2. The HNP-PtTi alloy manifests intriguing application potential as the candidate for the application of the electrochemical sensor for simultaneous detection of AA, DA, and UA. PMID:27058442

  5. Detection and formation scenario of citric acid, pyruvic acid, and other possible metabolism precursors in carbonaceous meteorites

    Cooper, George; Reed, Chris; Nguyen,Dang; Carter, Malika; Wang, Yi

    2011-01-01

    Carbonaceous meteorites deliver a variety of organic compounds to Earth that may have played a role in the origin and/or evolution of biochemical pathways. Some apparently ancient and critical metabolic processes require several compounds, some of which are relatively labile such as keto acids. Therefore, a prebiotic setting for any such individual process would have required either a continuous distant source for the entire suite of intact precursor molecules and/or an energetic and compact ...

  6. Combined measurement of thyroid and plasma homocysteic acid for detecting the severity of vascular dementia

    Bin Zhao; Junjian Zhang; Shifeng Wang; Guanghui Chen; Fayun Hu

    2006-01-01

    and content of homocysteic acid was measured with high performance liquid chromatogram (HPLC) electrochemical detection.MAIN OUTCOME MEASURES: Levels of homocysteic acid and thyroxine among patients with vascular dementia and non-dementia cerebral infarction.RESULTS: A total of 38 patients with vascular dementia and 40 patients with non-dementia cerebral infarction were involved in the final analysis. ① Levels of triiodothyronine (T3), thyroxine (T4) and free T3 (FT3) were (0.9±0.4) μg/L, (92.9±26.4) μg/L and (3.9±1.8) pmol/L in vascular dementia group respectively, which were higher than those in control group [(1.3±0.3) μg/L, (110.2±28.7) μg/L, (7.2±2.1) pmol/L, t=2.766 6-7.433 6, P<0.01]; while, level of homocysteic acid was (29.57±7.12) μmol/L in vascular dementia group, which was higher than that in control group [(24.53±4.98) μmol/L, t =3.637 7, P < 0.01]. There were no significant differences of free T4 (FT4) and thyrotropic-stimulating hormone (TSH) between the two groups (P> 0.05). ② Levels of FT3of patients with mild, moderate and severe vascular dementia were (1.0±0.2), (0.9±0.1) and (0.8±0.1) μg/L,respectively; levels of homocysteic acid were (26.52±4.84), (29.59±5.56) and (32.71±6.17) μmol/L,respectively. There were significant differences among patients at the three degrees of vascular dementia (F=3.59-32.4, P < 0.01). However, there were no significant differences of T4, FT4 and TSH among the three kinds of patients (P> 0.05).CONCLUSION: Levels of thyroxine of patients with vascular dementia decrease; however, levels of homocysteic acid increase. Therefore, the results can indirectly reflect severities of vascular dementia.

  7. Hidden Markov models that use predicted local structure for fold recognition: alphabets of backbone geometry.

    Karchin, Rachel; Cline, Melissa; Mandel-Gutfreund, Yael; Karplus, Kevin

    2003-06-01

    An important problem in computational biology is predicting the structure of the large number of putative proteins discovered by genome sequencing projects. Fold-recognition methods attempt to solve the problem by relating the target proteins to known structures, searching for template proteins homologous to the target. Remote homologs that may have significant structural similarity are often not detectable by sequence similarities alone. To address this, we incorporated predicted local structure, a generalization of secondary structure, into two-track profile hidden Markov models (HMMs). We did not rely on a simple helix-strand-coil definition of secondary structure, but experimented with a variety of local structure descriptions, following a principled protocol to establish which descriptions are most useful for improving fold recognition and alignment quality. On a test set of 1298 nonhomologous proteins, HMMs incorporating a 3-letter STRIDE alphabet improved fold recognition accuracy by 15% over amino-acid-only HMMs and 23% over PSI-BLAST, measured by ROC-65 numbers. We compared two-track HMMs to amino-acid-only HMMs on a difficult alignment test set of 200 protein pairs (structurally similar with 3-24% sequence identity). HMMs with a 6-letter STRIDE secondary track improved alignment quality by 62%, relative to DALI structural alignments, while HMMs with an STR track (an expanded DSSP alphabet that subdivides strands into six states) improved by 40% relative to CE. PMID:12784210

  8. Investigation of the parallel tempering method for protein folding

    Schug, Alexander; Herges, Thomas; Verma, Abhinav; Wenzel, Wolfgang

    2005-05-01

    We investigate the suitability and efficiency of an adapted version of the parallel tempering method for all-atom protein folding. We have recently developed an all-atom free energy force field (PFF01) for protein structure prediction with stochastic optimization methods. Here we report reproducible folding of the 20-amino-acid trp-cage protein and the conserved 40-amino-acid three-helix HIV accessory protein with an adapted parallel tempering method. We find that the native state, for both proteins, is correctly predicted to 2 Å backbone root mean square deviation and analyse the efficiency of the simulation approach.

  9. Metabotropic glutamate receptors are involved in the detection of IMP and L-amino acids by mouse taste sensory cells.

    Pal Choudhuri, S; Delay, R J; Delay, E R

    2016-03-01

    G-protein-coupled receptors are thought to be involved in the detection of umami and L-amino acid taste. These include the heterodimer taste receptor type 1 member 1 (T1r1)+taste receptor type 1 member 3 (T1r3), taste and brain variants of mGluR4 and mGluR1, and calcium sensors. While several studies suggest T1r1+T1r3 is a broadly tuned lLamino acid receptor, little is known about the function of metabotropic glutamate receptors (mGluRs) in L-amino acid taste transduction. Calcium imaging of isolated taste sensory cells (TSCs) of T1r3-GFP and T1r3 knock-out (T1r3 KO) mice was performed using the ratiometric dye Fura 2 AM to investigate the role of different mGluRs in detecting various L-amino acids and inosine 5' monophosphate (IMP). Using agonists selective for various mGluRs such as (RS)-3,5-dihydroxyphenylglycine (DHPG) (an mGluR1 agonist) and L-(+)-2-amino-4-phosphonobutyric acid (l-AP4) (an mGluR4 agonist), we evaluated TSCs to determine if they might respond to these agonists, IMP, and three L-amino acids (monopotassium L-glutamate, L-serine and L-arginine). Additionally, we used selective antagonists against different mGluRs such as (RS)-L-aminoindan-1,5-dicarboxylic acid (AIDA) (an mGluR1 antagonist), and (RS)-α-methylserine-O-phosphate (MSOP) (an mGluR4 antagonist) to determine if they can block responses elicited by these L-amino acids and IMP. We found that L-amino acid- and IMP-responsive cells also responded to each agonist. Antagonists for mGluR4 and mGluR1 significantly blocked the responses elicited by IMP and each of the L-amino acids. Collectively, these data provide evidence for the involvement of taste and brain variants of mGluR1 and mGluR4 in L-amino acid and IMP taste responses in mice, and support the concept that multiple receptors contribute to IMP and L-amino acid taste. PMID:26701297

  10. Influência do envelhecimento na concentração de ácido hialurônico nas pregas vocais de ratas fêmeas Influence of aging on hyaluronic acid concentration in the vocal folds of female rats

    Hugo Valter Lisboa Ramos

    2012-06-01

    Full Text Available A vibração das pregas vocais é um importante fator envolvido na produção vocal e o envelhecimento pode alterar a quantidade de ácido hialurônico da prega vocal levando a disfonia. OBJETIVO: Este estudo compara a concentração de ácido hialurônico nas pregas vocais de ratas fêmeas idosas e jovens. Desenho do estudo: estudo experimental. MATERIAL E MÉTODO: Foram utilizadas pregas vocais de 13 ratas fêmeas divididas em dois grupos: cinco ratas idosas e oito ratas jovens. A concentração tecidual do ácido hialurônico foi determinada por meio de método fluorimétrico utilizando a proteína de ligação ao ácido hialurônico imobilizada em placas de enzyme-linked immunosorbent assay (ELISA e também conjugada à biotina. Estreptavidina marcada com európio foi adicionada e, depois de európio ter sido liberado com o uso de solução de enhancement; a fluorescência final foi medida em um fluorímetro. RESULTADOS: Foram encontradas as seguintes concentrações de ácido hialurônico nas pregas vocais de acordo com os grupos: 581,7 ng/mg em ratas idosas e 1275,6 ng/mg em ratas jovens. A análise estatística mostrou diferença entre os grupos. CONCLUSÃO: A prega vocal de ratas idosas tem uma menor concentração de ácido hialurônico do que a concentração da prega vocal de ratas jovens.The vibration of the vocal fold lamina propria is an important factor involved in vocal production and aging may change the amount of hyaluronic acid in the vocal fold leading to dysphonia. AIMS: This study compares the concentration of hyaluronic acid in vocal folds of aged and young female rats. Study design: experimental. MATERIALS AND METHODS: We used the vocal cords of 13 female rats divided into two groups: five aged rats and eight young ones. The tissue concentration of hyaluronic acid was determined using the fluorimetric method with the hyaluronic acid binding-protein coated on plates of enzyme-linked immunosorbent assay (ELISA, conjugated

  11. Protein folding and wring resonances

    Bohr, Jakob; Bohr, Henrik; Brunak, Søren

    1997-01-01

    protein folding takes place when the amplitude of a wring excitation becomes so large that it is energetically favorable to bend the protein backbone. The condition under which such structural transformations can occur is found, and it is shown that both cold and hot denaturation (the unfolding of...

  12. Direct Determination of a Small-Molecule Drug, Valproic Acid, by an Electrically-Detected Microcantilever Biosensor for Personalized Diagnostics

    Long-Sun Huang

    2015-01-01

    Full Text Available Direct, small-molecule determination of the antiepileptic drug, valproic acid, was investigated by a label-free, nanomechanical biosensor. Valproic acid has long been used as an antiepileptic medication, which is administered through therapeutic drug monitoring and has a narrow therapeutic dosage range of 50–100 μg·mL−1 in blood or serum. Unlike labeled and clinically-used measurement techniques, the label-free, electrical detection microcantilever biosensor can be miniaturized and simplified for use in portable or hand-held point-of-care platforms or personal diagnostic tools. A micromachined microcantilever sensor was packaged into the micro-channel of a fluidic system. The measurement of the antiepileptic drug, valproic acid, in phosphate-buffered saline and serum used a single free-standing, piezoresistive microcantilever biosensor in a thermally-controlled system. The measured surface stresses showed a profile over a concentration range of 50–500 μg·mL−1, which covered the clinically therapeutic range of 50–100 μg·mL−1. The estimated limit of detection (LOD was calculated to be 45 μg·mL−1, and the binding affinity between the drug and the antibody was measured at around 90 ± 21 μg·mL−1. Lastly, the results of the proposed device showed a similar profile in valproic acid drug detection with those of the clinically-used fluorescence polarization immunoassay.

  13. Metalophthalocyanine complexes as ion-carriers in membrane-selective electrodes for detection of thiosalicylic acid

    Shahrokhian, Saeed; Souri, Ali

    2004-08-02

    The potentiometric response properties of several PVC-based membrane electrodes using phthalocyanine complexes of aluminum (AlPc), nickel (NiPc) and copper (CuPc) as anion carriers, toward thiosalicylic acid (TSA) were investigated. The influences of lipophilic ionic additives (cationic and anionic) and the pH of the buffered solutions were used for the interpretation of the mechanism of the potentiometric response of sensors. The sensitivity, linear range, detection limit, and potentiometric selectivity of the membrane sensors show a considerable dependence on the nature of central metal of the ionophore. The membrane electrodes based on AlPc demonstrate sub-Nernstian responses toward TSA over the range of 0.01 to 1x10{sup -5} M. In the case of NiPc and CuPc as ionophores and in the presence of trioctylmethyl ammonium (TOMA{sup +}) as a cationic additive, a Nernstian response could be established in a range of 4 orders of magnitudes of TSA concentration (0.01 to 1x10{sup -6} M). The results of potentiometric investigations revealed that from thermodynamic point of view, the axial coordination of thiosalicylate with the central metal of NiPc and CuPc is more efficient with respect to AlPc. This preference in response to TSA was discussed on the basis of the softness nature of NiPc and CuPc and more affinity for coordination with the thiolate group of thiosalicylate as a soft anion. These potentiometric sensors manifest prominent advantages of high selectivity for TSA over the various inorganic and organic anions, fast response times and micromolar detection limits and can be used over a wide pH range of 4.0-8.0. The prepared electrodes based on NiPc and CuPc were successfully applied in the potentiometric titration of sub-milimolar quantities of Hg{sup 2+} in aqueous solutions and very good recovery results were obtained in these measurements. The results of complexometric studies between Hg{sup 2+} and TSA using electrodes based on NiPc and CuPc as indicator

  14. Development of magnetic resonance imaging based detection methods for beta amyloids via sialic acid-functionalized magnetic nanoparticles

    Kouyoumdjian, Hovig

    The development of a non-invasive method for the detection of Alzheimer's disease is of high current interest, which can be critical in early diagnosis and in guiding preventive treatment of the disease. The aggregates of beta amyloids are a pathological hallmark of Alzheimer's disease. Carbohydrates such as sialic acid terminated gangliosides have been shown to play significant roles in initiation of amyloid aggregation. Herein, we report a biomimetic approach using sialic acid coated iron oxide superparamagnetic nanoparticles for in vitro detection in addition to the assessment of the in vivo mouse-BBB (Blood brain barrier) crossing of the BSA (bovine serum albumin)-modified ones. The sialic acid functionalized dextran nanoparticles were shown to bind with beta amyloids through several techniques including ELISA (enzyme linked immunosorbent assay), MRI (magnetic resonance imaging), TEM (transmission electron microscopy), gel electrophoresis and tyrosine fluorescence assay. The superparamagnetic nature of the nanoparticles allowed easy detection of the beta amyloids in mouse brains in both in vitro and ex vivo model by magnetic resonance imaging. Furthermore, the sialic acid nanoparticles greatly reduced beta amyloid induced cytotoxicity to SH-SY5Y neuroblastoma cells, highlighting the potential of the glyconanoparticles for detection and imaging of beta amyloids. Sialic acid functionalized BSA (bovine serum albumin) nanoparticles also showed significant binding to beta amyloids, through ELISA and ex vivo mouse brain MRI experiments. Alternatively, the BBB crossing was demonstrated by several techniques such as confocal microscopy, endocytosis, exocytosis assays and were affirmed by nanoparticles transcytosis assays through bEnd.3 endothelial cells. Finally, the BBB crossing was confirmed by analyzing the MRI signal of nanoparticle-injected CD-1 mice.

  15. Sensitive and specific detection of the non-human sialic Acid N-glycolylneuraminic acid in human tissues and biotherapeutic products.

    Sandra L Diaz

    Full Text Available BACKGROUND: Humans are genetically defective in synthesizing the common mammalian sialic acid N-glycolylneuraminic acid (Neu5Gc, but can metabolically incorporate it from dietary sources (particularly red meat and milk into glycoproteins and glycolipids of human tumors, fetuses and some normal tissues. Metabolic incorporation of Neu5Gc from animal-derived cells and medium components also results in variable contamination of molecules and cells intended for human therapies. These Neu5Gc-incorporation phenomena are practically significant, because normal humans can have high levels of circulating anti-Neu5Gc antibodies. Thus, there is need for the sensitive and specific detection of Neu5Gc in human tissues and biotherapeutic products. Unlike monoclonal antibodies that recognize Neu5Gc only in the context of underlying structures, chicken immunoglobulin Y (IgY polyclonal antibodies can recognize Neu5Gc in broader contexts. However, prior preparations of such antibodies (including our own suffered from some non-specificity, as well as some cross-reactivity with the human sialic acid N-acetylneuraminic acid (Neu5Ac. METHODOLOGY/PRINCIPAL FINDINGS: We have developed a novel affinity method utilizing sequential columns of immobilized human and chimpanzee serum sialoglycoproteins, followed by specific elution from the latter column by free Neu5Gc. The resulting mono-specific antibody shows no staining in tissues or cells from mice with a human-like defect in Neu5Gc production. It allows sensitive and specific detection of Neu5Gc in all underlying glycan structural contexts studied, and is applicable to immunohistochemical, enzyme-linked immunosorbent assay (ELISA, Western blot and flow cytometry analyses. Non-immune chicken IgY is used as a reliable negative control. We show that these approaches allow sensitive detection of Neu5Gc in human tissue samples and in some biotherapeutic products, and finally show an example of how Neu5Gc might be eliminated

  16. Quantification of fold curvature and fracturing using terrestrial laser scanning

    Pearce, M. A.; eospatial Research Ltd., Department of Earth Sciences, University of Durham, Durham, DH1 3LE, United Kingdom;; Jones, R. R.; Geospatial Research Ltd., Department of Earth Sciences, University of Durham, Durham, DH1 3LE, United Kingdom;; Smith, S. A. F.; Istituto Nazionale di Geofisica e Vulcanologia, Sezione Roma1, Roma, Italia; McCaffrey, K .J. W.; Geospatial Research Ltd., Department of Earth Sciences, University of Durham, Durham, DH1 3LE, United Kingdom;

    2011-01-01

    Terrestrial laser scanning is used to capture the geometry of three single folded bedding surfaces. The resulting light detection and ranging (LIDAR) point clouds are filtered and smoothed to enable meshing and calculation of principal curvatures. Fracture traces, picked from the LIDAR data, are used to calculate fracture densities. The rich data sets produced by this method provide statistically robust estimates of spatial variations in fracture density across the fold surface. The digital n...

  17. Nucleic Acid-Based Detection and Identification of Bacterial and Fungal Plant Pathogens - Final Report

    Kingsley, Mark T.

    2001-03-13

    The threat to American interests from terrorists is not limited to attacks against humans. Terrorists might seek to inflict damage to the U.S. economy by attacking our agricultural sector. Infection of commodity crops by bacterial or fungal crop pathogens could adversely impact U.S. agriculture, either directly from damage to crops or indirectly from damage to our ability to export crops suspected of contamination. Recognizing a terrorist attack against U.S. agriculture, to be able to prosecute the terrorists, is among the responsibilities of the members of Hazardous Material Response Unit (HMRU) of the Federal Bureau of Investigation (FBI). Nucleic acid analysis of plant pathogen strains by the use of polymerase chain reaction (PCR) amplification techniques is a powerful method for determining the exact identity of pathogens, as well as their possible region of origin. This type of analysis, however, requires that PCR assays be developed specific to each particular pathogen strain, and analysis protocols developed that are specific to the particular instrument used for detection. The objectives of the work described here were threefold: 1) to assess the potential terrorist threat to U.S. agricultural crops, 2) to determine whether suitable assays exist to monitor that threat, and 3) where assays are needed for priority plant pathogen threats, to modify or develop those assays for use by specialists at the HMRU. The assessment of potential threat to U.S. commodity crops and the availability of assays for those threats were described in detail in the Technical Requirements Document (9) and will be summarized in this report. This report addresses development of specific assays identified in the Technical Requirements Document, and offers recommendations for future development to ensure that HMRU specialists will be prepared with the PCR assays they need to protect against the threat of economic terrorism.

  18. Reliability of nucleic acid amplification for detection of Mycobacterium tuberculosis: an international collaborative quality control study among 30 laboratories.

    Noordhoek, G T; van Embden, J D; Kolk, A H

    1996-01-01

    Nucleic acid amplification to detect Mycobacterium tuberculosis in clinical specimens is increasingly used as a laboratory tool for the diagnosis of tuberculosis. However, the specificity and sensitivity of these tests may be questioned, and no standardized reagents for quality control assessment are available. To estimate the performance of amplification tests for routine diagnosis, we initiated an interlaboratory study involving 30 laboratories in 18 countries. We prepared blinded panels of 20 sputum samples containing no, 100, or 1,000 mycobacterial cells. Each laboratory was asked to detect M. tuberculosis by their routine method of nucleic acid amplification. Only five laboratories correctly identified the presence or absence of mycobacterial DNA in all 20 samples. Seven laboratories detected mycobacterial DNA in all positive samples, and 13 laboratories correctly reported the absence of DNA in the negative samples. Lack of specificity was more of a problem than lack of sensitivity. Reliability was not found to be associated with the use of any particular method. Reliable detection of M. tuberculosis in clinical samples by nucleic acid amplification techniques is possible, but many laboratories do not use adequate quality controls. This study underlines the need for good laboratory practice and reference reagents to monitor the performance of the whole assay, including pretreatment of clinical samples. PMID:8880513

  19. Determination of Amino Acids in Panax notoginseng by Microwave Hydrolysis and Derivatization Coupled with Capillary Zone Electrophoresis Detection

    LI Xiao-tian; ZHAO Ya-jing; JIANG Cheng-fei; ZHANG Han-qi; YU Ai-min

    2013-01-01

    The microwave hydrolysis and derivatization coupled with capillary electrophoresis detection were developed for the separation and determination of the amino acids in Panax notoginseng.The experimental conditions for the microwave hydrolysis and derivatization were examined and optimized.Several parameters of capillary electrophoresis,such as pH value of background electrolyte,borate concentration and applied voltage were optimized.Under the selected conditions,11 amino acids were completely separated.The real sample was analyzed and the results were satisfactory.Compared with that of conventional heat hydrolysis and derivatization,the analytical time of this method was significantly shortened.

  20. Probing folding free energy landscape of small proteins through minimalistic models: Folding of HP-36 and -amyloid

    Arnab Mukherjee; Biman Bagchi

    2003-10-01

    Folding dynamics and energy landscape picture of protein conformations of HP-36 and -amyloid (A) are investigated by extensive Brownian dynamics simulations, where the inter amino acid interactions are given by a minimalistic model (MM) we recently introduced [J. Chem. Phys. 118 4733 (2003)]. In this model, a protein is constructed by taking two atoms for each amino acid. One atom represents the backbone C atom, while the other mimics the whole side chain residue. Sizes and interactions of the side residues are all different and specific to a particular amino acid. The effect of water-mediated folding is mapped into the MM by suitable choice of interaction parameters of the side residues obtained from the amino acid hydropathy scale. A new non-local helix potential is incorporated to generate helices at the appropriate positions in a protein. Simulations have been done by equilibrating the protein at high temperature followed by a sudden quench. The subsequent folding is monitored to observe the dynamics of topological contacts (topo), relative contact order parameter (RCO), and the root mean square deviation (RMSD) from the realprotein native structure. The folded structures of different model proteins (HP-36 and ) resemble their respective real native state rather well. The dynamics of folding shows multistage decay, with an initial hydrophobic collapse followed by a long plateau. Analysis of topo and RCO correlates the late stage folding with rearrangement of the side chain residues, particularly those far apart in the sequence. The long plateau also signifies large entropic free energy barrier near the native state, as predicted from theories of protein folding.

  1. Intestinal involvement in progressive systemic sclerosis detected by increased unconjugated serum bile acids.

    Stellaard, F.; Sauerbruch, T; Luderschmidt, C H; Leisner, B.; Paumgartner, G

    1987-01-01

    In patients with progressive systemic sclerosis, impaired motor function of the small intestine may lead to bacterial overgrowth causing diarrhoea, steatorrhoea and malabsorption. As unconjugated serum bile acids have been proposed as markers for small bowel bacterial overgrowth, we studied individual unconjugated serum bile acids in 36 patients with progressive systemic sclerosis. These patients had significantly higher serum concentrations of unconjugated cholic acid (median 0.18; range 0.0...

  2. Ultra-sensitive detection of zinc oxide nanowires using a quartz crystal microbalance and phosphoric acid DNA

    Jang, Kuewhan; You, Juneseok; Park, Chanhoo; Park, Hyunjun; Choi, Jaeyeong; Choi, Chang-Hwan; Park, Jinsung; Lee, Howon; Na, Sungsoo

    2016-09-01

    Recent advancements of nanomaterials have inspired numerous scientific and industrial applications. Zinc oxide nanowires (ZnO NWs) is one of the most important nanomaterials due to their extraordinary properties. However, studies performed over the past decade have reported toxicity of ZnO NWs. Therefore, there has been increasing demand for effective detection of ZnO NWs. In this study, we propose a method for the detection of ZnO NW using a quartz crystal microbalance (QCM) and DNA probes. The detection method is based on the covalent interaction between ZnO NWs and the phosphoric acid group of single-stranded DNA (i.e., linker DNA), and DNA hybridization between the linker DNA and the probe DNA strand on the QCM electrode. Rapid, high sensitivity, in situ detection of ZnO NWs was demonstrated for the first time. The limit of detection was 10‑4 μg ml‑1 in deionized water, which represents a sensitivity that is 100000 times higher than the toxic ZnO NW concentration level. Moreover, the selectivity of the ZnO NW detection method was demonstrated by comparison with other types of nanowires and the method was able to detect ZnO NWs in tap water sensitively even after stored for 14 d in a refrigerator. The performance of our proposed method was sufficient to achieve detection of ZnO NW in the ‘real-world’ environment.

  3. Microfluidic Chip-based Nucleic Acid Testing using Gingival Crevicular Fluid as a New Technique for Detecting HIV-1 Infection

    Alex Willyandre

    2013-05-01

    Full Text Available Transmission of HIV-1 infection by individuals in window period who are tested negative in conventional HIV-1 detection would pose the community with serious problems. Several diagnostic tools require specific labora-tory equipment, perfect timing of diagnosis, antibody to HIV-1, and invasive technique to get sample for examination, until high amount of time to process the sample as well as accessibility of remote areas. Many attempts have been made to solve those problems to come to a new detection technique. This review aims to give information about the current development technique for detection of HIV infection. Microfluidic Chip-based Nucleic Acid Testing is currently introduced for detection of HIV-1 infection. This review also cover the possible usage of gingival crevicular fluid as sample specimen that could be taken noninvasively from the individual.DOI: 10.14693/jdi.v18i2.63

  4. Slow alpha helix formation during folding of a membrane protein.

    Riley, M L; Wallace, B A; Flitsch, S L; Booth, P J

    1997-01-01

    Very little is known about the folding of proteins within biological membranes. A "two-stage" model has been proposed on thermodynamic grounds for the folding of alpha helical, integral membrane proteins, the first stage of which involves formation of transmembrane alpha helices that are proposed to behave as autonomous folding domains. Here, we investigate alpha helix formation in bacteriorhodopsin and present a time-resolved circular dichroism study of the slow in vitro folding of this protein. We show that, although some of the protein's alpha helices form early, a significant part of the protein's secondary structure appears to form late in the folding process. Over 30 amino acids, equivalent to at least one of bacteriorhodopsin's seven transmembrane segments, slowly fold from disordered structures to alpha helices with an apparent rate constant of about 0.012 s-1 at pH 6 or 0.0077 s-1 at pH 8. This is a rate-limiting step in protein folding, which is dependent on the pH and the composition of the lipid bilayer. PMID:8993333

  5. Amino acid detection using fluoroquinolone–Cu2+ complex as a switch-on fluorescent probe by competitive complexation without derivatization

    In this work, we describe the use of fluoroquinolone–Cu2+ complex as a competitive switch-on fluorescence probe for amino acid determination without derivatization. The fluorescence intensity of this probe, which has been reduced due to effective quenching by Cu2+ ion, increases drastically by an addition of amino acid (glycine, phenylalanine, sarcosine, aspargine, alanine, proline, arginine, aspartic acid, glutamic acid, lysine, leucine and isoleucine). The overall stability constants of Cu2+ ion complexes with amino acids were determined by fluorometric titration of fluoroquinolone-Cu2+ complex with the amino acid solution. Furthermore, the probe shows high calibration sensitivity toward aspartic acid. The fluorescence signal depends linearly on the amino acid concentration within the range of concentration from 1.2×10−7 to 1.1×10−5 mol L−1 for aspartic acid. The detection limit was found 2.7×10−8 mol L−1 with the relative standard deviation (RSD%) about 2.1% (five replicate). -- Highlights: • Amino acids are detected by using fluoroquinolone–Cu2+ complex as fluorescent probe. • Amino acids were detected based on a competitive complexation reaction. • Probe has been able to recognize amino acids through switch-on fluorescence behavior. • Ultra-trace level of aspartic and glutamic acid is determined without derivatization

  6. Amino acid detection using fluoroquinolone–Cu{sup 2+} complex as a switch-on fluorescent probe by competitive complexation without derivatization

    Farokhcheh, Alireza; Alizadeh, Naader, E-mail: alizaden@modares.ac.ir

    2014-01-15

    In this work, we describe the use of fluoroquinolone–Cu{sup 2+} complex as a competitive switch-on fluorescence probe for amino acid determination without derivatization. The fluorescence intensity of this probe, which has been reduced due to effective quenching by Cu{sup 2+} ion, increases drastically by an addition of amino acid (glycine, phenylalanine, sarcosine, aspargine, alanine, proline, arginine, aspartic acid, glutamic acid, lysine, leucine and isoleucine). The overall stability constants of Cu{sup 2+} ion complexes with amino acids were determined by fluorometric titration of fluoroquinolone-Cu{sup 2+} complex with the amino acid solution. Furthermore, the probe shows high calibration sensitivity toward aspartic acid. The fluorescence signal depends linearly on the amino acid concentration within the range of concentration from 1.2×10{sup −7} to 1.1×10{sup −5} mol L{sup −1} for aspartic acid. The detection limit was found 2.7×10{sup −8} mol L{sup −1} with the relative standard deviation (RSD%) about 2.1% (five replicate). -- Highlights: • Amino acids are detected by using fluoroquinolone–Cu{sup 2+} complex as fluorescent probe. • Amino acids were detected based on a competitive complexation reaction. • Probe has been able to recognize amino acids through switch-on fluorescence behavior. • Ultra-trace level of aspartic and glutamic acid is determined without derivatization.

  7. Selective colorimetric detection of Cr(iii) and Cr(vi) using gallic acid capped gold nanoparticles.

    Dong, Chen; Wu, Genhua; Wang, Zhuqing; Ren, Wenzhi; Zhang, Yujie; Shen, Zheyu; Li, Tianhua; Wu, Aiguo

    2016-05-28

    A colorimetric assay is proposed for the selective detection of Cr(iii) and Cr(vi) via the aggregation-induced color change of gallic acid capped gold nanoparticles (GA-AuNPs). The AuNPs are characterized using UV-vis spectroscopy, transmission electron microscopy (TEM), dynamic light scattering (DLS) and Fourier-transform infrared spectrometry (FT-IR). To detect Cr(iii) and Cr(vi) coexisting in a sample, citrate and thiosulfate were applied to mask Cr(vi) for the detection of Cr(iii), and ethylenediaminetetraacetic acid disodium salt (EDTA) was applied to mask Cr(iii) for the detection of Cr(vi). At optimized experimental conditions, the selectivity of these AuNPs-based detection systems is excellent for Cr(iii) and/or Cr(vi) compared with other types of metal ions. The limit of detections (LODs) of a mixture of Cr(iii) and Cr(vi), Cr(iii) and Cr(vi) by eye vision are 1.5, 1.5 and 2 μM, respectively, and those by UV-vis spectroscopy are 0.05, 0.1 and 0.1 μM, respectively. The minimum detectable concentrations for Cr(iii) or Cr(vi) are all below the guideline value set by the US Environmental Protection Agency (EPA). The applicability of the AuNPs-based colorimetric sensor is also validated by the detection of Cr(iii) and Cr(vi) in electroplating wastewater and real water samples with high recoveries. PMID:26606324

  8. A fully integrated paperfluidic molecular diagnostic chip for the extraction, amplification, and detection of nucleic acids from clinical samples.

    Rodriguez, Natalia M; Wong, Winnie S; Liu, Lena; Dewar, Rajan; Klapperich, Catherine M

    2016-02-21

    Paper diagnostics have successfully been employed to detect the presence of antigens or small molecules in clinical samples through immunoassays; however, the detection of many disease targets relies on the much higher sensitivity and specificity achieved via nucleic acid amplification tests (NAAT). The steps involved in NAAT have recently begun to be explored in paper matrices, and our group, among others, has reported on paper-based extraction, amplification, and detection of DNA and RNA targets. Here, we integrate these paper-based NAAT steps into a single paperfluidic chip in a modular, foldable system that allows for fully integrated fluidic handling from sample to result. We showcase the functionality of the chip by combining nucleic acid isolation, isothermal amplification, and lateral flow detection of human papillomavirus (HPV) 16 DNA directly from crude cervical specimens in less than 1 hour for rapid, early detection of cervical cancer. The chip is made entirely of paper and adhesive sheets, making it low-cost, portable, and disposable, and offering the potential for a point-of-care molecular diagnostic platform even in remote and resource-limited settings. PMID:26785636

  9. Quantitative Morphology of Epithelial Folds.

    Štorgel, Nick; Krajnc, Matej; Mrak, Polona; Štrus, Jasna; Ziherl, Primož

    2016-01-01

    The shape of spatially modulated epithelial morphologies such as villi and crypts is usually associated with the epithelium-stroma area mismatch leading to buckling. We propose an alternative mechanical model based on intraepithelial stresses generated by differential tensions of apical, lateral, and basal sides of cells as well as on the elasticity of the basement membrane. We use it to theoretically study longitudinal folds in simple epithelia and we identify four types of corrugated morphologies: compact, invaginated, evaginated, and wavy. The obtained tissue contours and thickness profiles are compared to epithelial folds observed in invertebrates and vertebrates, and for most samples, the agreement is within the estimated experimental error. Our model establishes the groove-crest modulation of tissue thickness as a morphometric parameter that can, together with the curvature profile, be used to estimate the relative differential apicobasal tension in the epithelium. PMID:26745429

  10. Protein folding and wring resonances

    Bohr, Jakob; Bohr, Henrik; Brunak, Søren

    1997-01-01

    protein folding takes place when the amplitude of a wring excitation becomes so large that it is energetically favorable to bend the protein backbone. The condition under which such structural transformations can occur is found, and it is shown that both cold and hot denaturation (the unfolding of......The polypeptide chain of a protein is shown to obey topological contraints which enable long range excitations in the form of wring modes of the protein backbone. Wring modes of proteins of specific lengths can therefore resonate with molecular modes present in the cell. It is suggested that...... proteins) are natural consequences of the suggested wring mode model. Native (folded) proteins are found to possess an intrinsic standing wring mode....

  11. Folded supersymmetry with a twist

    Cohen, Timothy; Craig, Nathaniel; Lou, Hou Keong; Pinner, David

    2016-03-01

    Folded supersymmetry ( f-SUSY) stabilizes the weak scale against radiative corrections from the top sector via scalar partners whose gauge quantum numbers differ from their Standard Model counterparts. This non-trivial pairing of states can be realized in extra-dimensional theories with appropriate supersymmetry-breaking boundary conditions. We present a class of calculable f-SUSY models that are parametrized by a non-trivial twist in 5D boundary conditions and can accommodate the observed Higgs mass and couplings. Although the distinctive phenomenology associated with the novel folded states should provide strong evidence for this mechanism, the most stringent constraints are currently placed by conventional supersymmetry searches. These models remain minimally fine-tuned in light of LHC8 data and provide a range of both standard and exotic signatures accessible at LHC13.

  12. Adulteration of Argentinean milk fats with animal fats: Detection by fatty acids analysis and multivariate regression techniques.

    Rebechi, S R; Vélez, M A; Vaira, S; Perotti, M C

    2016-02-01

    The aims of the present study were to test the accuracy of the fatty acid ratios established by the Argentinean Legislation to detect adulterations of milk fat with animal fats and to propose a regression model suitable to evaluate these adulterations. For this purpose, 70 milk fat, 10 tallow and 7 lard fat samples were collected and analyzed by gas chromatography. Data was utilized to simulate arithmetically adulterated milk fat samples at 0%, 2%, 5%, 10% and 15%, for both animal fats. The fatty acids ratios failed to distinguish adulterated milk fats containing less than 15% of tallow or lard. For each adulterant, Multiple Linear Regression (MLR) was applied, and a model was chosen and validated. For that, calibration and validation matrices were constructed employing genuine and adulterated milk fat samples. The models were able to detect adulterations of milk fat at levels greater than 10% for tallow and 5% for lard. PMID:26304443

  13. GPU accelerated Rna folding algorithm

    Rizk, Guillaume; Lavenier, Dominique

    2009-01-01

    Many bioinformatics studies require the analysis of RNA or DNA structures. More specifically, extensive work is done to elaborate efficient algorithms able to predict the 2-D folding structures of RNA or DNA sequences. However, the high computational complexity of the algorithms, combined with the rapid increase of genomic data, triggers the need of faster methods. Current approaches focus on parallelizing these algorithms on multiprocessor systems or on clusters, yielding to good performance...

  14. Hydrogen Bonds in Polymer Folding

    Borg, J; Jensen, M. H.; K. Sneppen; Tiana, G.

    2000-01-01

    The thermodynamics of a homopolymeric chain with both Van der Waals and highly-directional hydrogen bond interaction is studied. The effect of hydrogen bonds is to reduce dramatically the entropy of low-lying states and to give raise to long-range order and to conformations displaying secondary structures. For compact polymers a transition is found between helix-rich states and low-entropy sheet-dominated states. The consequences of this transition for protein folding and, in particular, for ...

  15. Hydrodynamic Interactions in Protein Folding

    Cieplak, Marek; Niewieczerzał, Szymon

    2008-01-01

    We incorporate hydrodynamic interactions (HI) in a coarse-grained and structure-based model of proteins by employing the Rotne-Prager hydrodynamic tensor. We study several small proteins and demonstrate that HI facilitate folding. We also study HIV-1 protease and show that HI make the flap closing dynamics faster. The HI are found to affect time correlation functions in the vicinity of the native state even though they have no impact on same time characteristics of the structure fluctuations ...

  16. Solitons and protein folding: An In Silico experiment

    Protein folding [1] is the process of formation of a functional 3D structure from a random coil — the shape in which amino-acid chains leave the ribosome. Anfinsen’s dogma states that the native 3D shape of a protein is completely determined by protein’s amino acid sequence. Despite the progress in understanding the process rate and the success in folding prediction for some small proteins, with presently available physics-based methods it is not yet possible to reliably deduce the shape of a biologically active protein from its amino acid sequence. The protein-folding problem endures as one of the most important unresolved problems in science; it addresses the origin of life itself. Furthermore, a wrong fold is a common cause for a protein to lose its function or even endanger the living organism. Soliton solutions of a generalized discrete non-linear Schrödinger equation (GDNLSE) obtained from the energy function in terms of bond and torsion angles κ and τ provide a constructive theoretical framework for describing protein folds and folding patterns [2]. Here we study the dynamics of this process by means of molecular-dynamics simulations. The soliton manifestation is the pattern helix–loop–helix in the secondary structure of the protein, which explains the importance of understanding loop formation in helical proteins. We performed in silico experiments for unfolding one subunit of the core structure of gp41 from the HIV envelope glycoprotein (PDB ID: 1AIK [3]) by molecular-dynamics simulations with the MD package GROMACS. We analyzed 80 ns trajectories, obtained with one united-atom and two different all-atom force fields, to justify the side-chain orientation quantification scheme adopted in the studies and to eliminate force-field based artifacts. Our results are compatible with the soliton model of protein folding and provide first insight into soliton-formation dynamics

  17. Solitons and protein folding: An In Silico experiment

    Ilieva, N.; Dai, J.; Sieradzan, A.; Niemi, A.

    2015-10-01

    Protein folding [1] is the process of formation of a functional 3D structure from a random coil — the shape in which amino-acid chains leave the ribosome. Anfinsen's dogma states that the native 3D shape of a protein is completely determined by protein's amino acid sequence. Despite the progress in understanding the process rate and the success in folding prediction for some small proteins, with presently available physics-based methods it is not yet possible to reliably deduce the shape of a biologically active protein from its amino acid sequence. The protein-folding problem endures as one of the most important unresolved problems in science; it addresses the origin of life itself. Furthermore, a wrong fold is a common cause for a protein to lose its function or even endanger the living organism. Soliton solutions of a generalized discrete non-linear Schrödinger equation (GDNLSE) obtained from the energy function in terms of bond and torsion angles κ and τ provide a constructive theoretical framework for describing protein folds and folding patterns [2]. Here we study the dynamics of this process by means of molecular-dynamics simulations. The soliton manifestation is the pattern helix-loop-helix in the secondary structure of the protein, which explains the importance of understanding loop formation in helical proteins. We performed in silico experiments for unfolding one subunit of the core structure of gp41 from the HIV envelope glycoprotein (PDB ID: 1AIK [3]) by molecular-dynamics simulations with the MD package GROMACS. We analyzed 80 ns trajectories, obtained with one united-atom and two different all-atom force fields, to justify the side-chain orientation quantification scheme adopted in the studies and to eliminate force-field based artifacts. Our results are compatible with the soliton model of protein folding and provide first insight into soliton-formation dynamics.

  18. Solitons and protein folding: An In Silico experiment

    Ilieva, N., E-mail: nevena.ilieva@parallel.bas.bg [Institute of Information and Communication Technologies, Bulgarian Aacademy of Sciences, Sofia (Bulgaria); Dai, J., E-mail: daijing491@gmail.com [School of Physics, Beijing Institute of Technology, Beijing (China); Sieradzan, A., E-mail: adams86@wp.pl [Faculty of Chemistry, University of Gdańsk, Gdańsk (Poland); Niemi, A., E-mail: Antti.Niemi@physics.uu.se [Department of Physics and Astronomy, Uppsala University, Uppsala (Sweden); LMPT–CNRS, Université de Tours, Tours (France)

    2015-10-28

    Protein folding [1] is the process of formation of a functional 3D structure from a random coil — the shape in which amino-acid chains leave the ribosome. Anfinsen’s dogma states that the native 3D shape of a protein is completely determined by protein’s amino acid sequence. Despite the progress in understanding the process rate and the success in folding prediction for some small proteins, with presently available physics-based methods it is not yet possible to reliably deduce the shape of a biologically active protein from its amino acid sequence. The protein-folding problem endures as one of the most important unresolved problems in science; it addresses the origin of life itself. Furthermore, a wrong fold is a common cause for a protein to lose its function or even endanger the living organism. Soliton solutions of a generalized discrete non-linear Schrödinger equation (GDNLSE) obtained from the energy function in terms of bond and torsion angles κ and τ provide a constructive theoretical framework for describing protein folds and folding patterns [2]. Here we study the dynamics of this process by means of molecular-dynamics simulations. The soliton manifestation is the pattern helix–loop–helix in the secondary structure of the protein, which explains the importance of understanding loop formation in helical proteins. We performed in silico experiments for unfolding one subunit of the core structure of gp41 from the HIV envelope glycoprotein (PDB ID: 1AIK [3]) by molecular-dynamics simulations with the MD package GROMACS. We analyzed 80 ns trajectories, obtained with one united-atom and two different all-atom force fields, to justify the side-chain orientation quantification scheme adopted in the studies and to eliminate force-field based artifacts. Our results are compatible with the soliton model of protein folding and provide first insight into soliton-formation dynamics.

  19. Detection and Quantification of Valerenic Acid in Commercially Available Valerian Products

    Douglas, Ruth H.; Muldowney, Ciaran A.; Mohamed, Rabab; Keohane, Fiona; Shanahan, Catherine; Walsh, John J.; Kavanagh, Pierce V.

    2007-01-01

    Several valerian-containing products sold in pharmacies were evaluated to verify the presence of Valeriana officinalis by identifying the presence of valerenic acid found only in species of Valeriana. The content of valerenic acid was found to vary considerably in the products analyzed, thus emphasizing the importance of standardizing herbal…

  20. Detection of plasmid deoxyribonucleic acid in an isolate of Lactobacillus acidophilus.

    Klaenhammer, T R; Sutherland, S M

    1980-01-01

    Eight strains of Lactobacillus acidophilus were examined for the presence of plasmid deoxyribonucleic acid, and one, a pig intestinal isolate, showed the presence of a 13.7- and a 6.3-megadalton plasmid. This is the first reported evidence for plasmid deoxyribonucleic acid in Lactobacillus acidophilus. The functions of these plasmids are presently unknown.

  1. Folding of proteins in presculpted free energy landscape

    Recent studies of the tube model of protein have indicated that the free energy landscape of proteins is presculpted by symmetry of the protein backbone and geometrical constraints played by the hydrogen bonds. In this study, we investigate the role of amino acid sequences in the folding of proteins. We consider two models that are differed by sequence specificity: the tube HP model with hydrophobic (H) and polar (P) sequences, and the tube Go model with native-centric contact potentials. Monte Carlo simulations are carried out for two sequences of length of 48 amino acids, whose ground states are a three-helix bundle and a GB1-like structure. The results show that folding in the Go model is more cooperative than in the HP model. In the HP model the collapse transition and the folding transition are separated, whereas in the Go model the two transitions coincide. (author)

  2. A sensor of a polyoxometalate and Au–Pd alloy for simultaneously detection of dopamine and ascorbic acid

    A composite film based on K8P2W16V2O62·18H2O decorated by Au–Pd nanoparticles was prepared by the layer-by-layer self-assembly method. This composite film exhibits enhanced electrocatalytic performance, repeatability and long-term stability for the simultaneous determination of dopamine and ascorbic acid at biological pH (pH 7.0). The proposed electrochemical sensing film for simultaneous detecting dopamine and ascorbic acid shows rather low detection limit of 8.3 × 10−7 and 4.3 × 10−7 M and a linear response range from 2.1 × 10−6 to 2.06 × 10−3 M and 1.2 × 10−6 to 1.61 × 10−3 M, as well as no interference from the common interfering species at an applied potential. -- Abstract: A novel composite film based on Dawson-type phosphovanadotungstate K8P2W16V2O62·18H2O (P2W16V2) decorated by Au–Pd alloy nanoparticles (Au–Pd) was fabricated on quartz, silicon and ITO using the layer-by-layer self-assembly method. The composite film was characterized by UV–vis spectroscopy, X-ray photoelectron spectra, atomic force microscopy, Scanning electronic microscope, cyclic voltammetry and differential pulse voltammetry. The composite film can be employed for sensitive and simultaneous determination of dopamine and ascorbic acid at biological pH (pH 7.0). Linear analytical curves were obtained in the ranges from 2.1 × 10−6 to 2.06 × 10−3 M and 1.2 × 10−6 to 1.61 × 10−3 M for dopamine and ascorbic acid by DPV methods, respectively. The low detection limit for dopamine and ascorbic acid were 8.3 × 10−7 and 4.3 × 10−7 M, as well as no interference was observed from the common interfering species such as glucose, uric acid, L-cysteine, CH3OH, CH3CH2OH and H2O2. The composite film was used for dopamine and ascorbic acid determinations in real samples with satisfactory results. With high sensitivity and selectivity, the proposed electrochemical sensor would provide a simple method for simultaneous determination of dopamine and ascorbic acid

  3. Peptide nucleic acid (PNA) probe, kit and procedure for specific detection of Helicobacter pylori and applications thereof

    Azevedo, N. F.; N. Guimarães; Figueiredo, C.; Keevil, C. W.; Vieira, M. J.

    2008-01-01

    The current invention relates to the development of a peptide nucleic acid (PNA) probe for the specific detection of Helicobacter pylori. These probes are used in a process based on molecular biology techniques, namely the fluorescence in situ hybridization (FISH), and can be applied in a variety of samples such as biopsies, blood, air, food, water and other environmental samples. Due to the physical and chemical characteristics inherent to their structure, these probes are able to provide a ...

  4. Design and Synthesis of Immunoconjugates and Development of an Indirect ELISA for Rapid Detection of 3, 5-Dinitrosalicyclic Acid Hydrazide

    Yuan-Ming Sun; Yue-Ming Jiang; Zhi-Li Xiao; Hong Wang; Hong-Tao Lei; Shi-Wei Zhang; Yu-Dong Shen

    2008-01-01

    In this study novel immunoconjugates were designed, synthesized and then used to develop a rapid, specific and sensitive indirect ELISA method to directly detect residues of 3,5-dinitrosalicyclic acid hydrazide (DNSH), a toxic metabolite of nifursol present in chicken tissues. The hapten DNSHA was first designed and used to covalently couple to BSA to form an immunogen which was immunized to rabbits to produce a polyclonal antibody against DNSH. Furthermore, a novel 3,5-dinitrosalicylic acido...

  5. Simultaneous determination of furfural, acetic acid, and 5-hydroxymethylfurfural in corncob hydrolysates using liquid chromatography with ultraviolet detection.

    Dong, Bo-Yu; Chen, Ye-Fu; Zhao, Chang-Chun; Zhang, Shi-Jie; Guo, Xue-Wu; Xiao, Dong-Guang

    2013-01-01

    A single-laboratory validation study was conducted using HPLC for detecting and quantifying acetic acid, furfural, and 5-hydroxymethylfurfural (HMF) in corncob hydrolysates. A pretreatment procedure using dilute sulfuric acid was optimized for corncob hydrolysis. The final hydrolysates were analyzed by HPLC using a C18 RP column with aqueous 0.01% (v/v) H2SO4-CH3OH (95 + 5) as the mobile phase at a flow rate of 1 mL/min. The wavelengths for detecting the three compounds were changed to their optimal UV detection wavelengths at the time of elution. The wavelength detection adjustments were as follow: 205 nm (0 to 4 min); 284 nm (4 to 7 min); and 276 nm (7 to 10 min). Separation was achieved with a chromatographic run time of 10 min. The calibration curves for the three compounds had correlation coefficients (r2) > or = 99.8%. The analytical range, as defined by the calibration curves, was 0.5-10 mg/L for acetic acid, 0.4-22 mg/L for furfural, and 0.1-18 mg/L for HMF. The LODs for acetic acid, furfural, and HMF were estimated to be 0.05, 0.03, and 0.02 mg/L, respectively; the LOQs were 0.196, 0.135, and 0.074 mg/L, respectively. The RSD values for the intraday precision study ranged from 0.31 to 2.22%, and from 0.57 to 2.43% for the interday study. The mean recovery rates in all compounds were between 100.08 and 101.49%. PMID:24645500

  6. Application of functionalized lanthanide-based nanoparticles for the detection of okadaic acid-specific immunoglobulin G

    Stipić, Filip; Pletikapić, Galja; Jakšić, Željko; Frkanec, Leo; Zgrablić, Goran; Burić, Petra; Lyons, Daniel Mark

    2015-01-01

    Marine biotoxins are widespread in the environment and impact human health via contaminated shellfish, causing diarrhetic, amnesic, paralytic, or neurotoxic poisoning. In spite of this, methods for determining if poisoning has occurred are limited. We show the development of a simple and sensitive luminescence resonance energy transfer (LRET)-based concept which allows the detection of anti-okadaic acid rabbit polyclonal IgG (mouse monoclonal IgG1) using functionalized lanthanide-based nanopa...

  7. Incorporation and excision of 5-fluorouracil from deoxyribonucleic acid in Escherichia coli.

    Warner, H. R.; Rockstroh, P A

    1980-01-01

    When Escherichia coli are grown in the presence of 5-fluorouracil, the 5-fluorouracil is incorporated almost exclusively into ribonucleic acid as fluorouridylate. In this study, small but detectable amounts were incorporated into ribonucleic acid as fluorocytidylate and into deoxyribonucleic acid as fluorodeoxyuridylate and fluorodeoxycytidylate. The amount of 5-fluorouracil found in deoxyribonucleic acid as fluorodeoxyuridylate increased 50-fold when the cells were deficient in both deoxyuri...

  8. Gadoxetic acid-enhanced MRI and diffusion-weighted imaging for the detection of colorectal liver metastases after neoadjuvant chemotherapy

    Yu, Mi Hye [Konkuk University Medical Center, Department of Radiology, Seoul (Korea, Republic of); Lee, Jeong Min; Han, Joon Koo; Choi, Byung-Ihn [Seoul National University Hospital, Department of Radiology, Seoul (Korea, Republic of); Hur, Bo Yun [Boramae Medical Center, Department of Radiology, Seoul (Korea, Republic of); Kim, Tae-You [Seoul National University Hospital, Department of Internal Medicine, Seoul (Korea, Republic of); Jeong, Seung-Yong; Yi, Nam-Joon; Suh, Kyung-Suk [Seoul National University Hospital, Department of Surgery, Seoul (Korea, Republic of)

    2015-08-15

    To investigate the diagnostic performance of gadoxetic acid-enhanced MRI including diffusion-weighted imaging (DWI) for the detection of colorectal liver metastases (CRLMs) after neoadjuvant chemotherapy (NAC). Our study population comprised 77 patients with 140 CRLMs who underwent gadoxetic acid-enhanced MRI within 1 month prior to surgery: group A (without NAC, n = 38) and group B (with NAC, n = 39). Two radiologists independently assessed all MR images and graded their diagnostic confidence for CRLM on a 5-point scale. Diagnostic accuracy, sensitivity and positive predictive values (PPV) were calculated and compared between the two groups. Diagnostic accuracy of gadoxetic acid-enhanced MRI in group B was slightly lower than in group A, but a statistically significant difference was not observed (observer 1: A{sub z}, 0.926 in group A, 0.905 in group B; observer 2: A{sub z}, 0.944 in group A, 0.885 in group B; p > 0.05). Sensitivity and PPV of group B were comparable to those of group A (observer 1: sensitivity = 93.5 % vs. 93.6 %, PPV = 95.1 % vs. 86.9 %; observer 2: sensitivity = 96.8 % vs. 91.0 %; PPV = 90.0 % vs. 89.7 %; all p > 0.05). Gadoxetic acid-enhanced MRI including DWI provided good diagnostic performance with high sensitivity (>90 %) for the detection of CRLMs, regardless of the influence of NAC. (orig.)

  9. Protein folding in a force clamp

    Cieplak, Marek; Szymczak, P.

    2006-05-01

    Kinetics of folding of a protein held in a force clamp are compared to an unconstrained folding. The comparison is made within a simple topology-based dynamical model of ubiquitin. We demonstrate that the experimentally observed variations in the end-to-end distance reflect microscopic events during folding. However, the folding scenarios in and out of the force clamp are distinct.

  10. Mycolic acids – biological role and potential application in Mycobacterium detection and differentiation

    Konrad Kowalski

    2014-04-01

    Full Text Available Mycolic acids are one of the basic structural elements of the cell wall of bacteria from Corynebacterineae suborder. These compounds are long-chain α-hydroxy β-alkyl fatty acids with two hydrocarbon chains: longer meromycolic and shorter α-chain meromycolic α-chain. The genus Mycobacterium is characterized by the presence of mycolic acids in length from 60 to 90 carbon atoms having a fully saturated α-chain with a defined length of 22, 24 or 26 carbon atoms. Current research indicates that not only the presence of mycolic acids in the cell wall of mycobacteria is essential for the virulence of mycobacteria. It is proved that the relationship between different types of mycolic acids, their length and the degree of cyclopropanation may vary depending on the stage of infection and mycobacterial culture conditions. At the same time it has been shown that some mycolic acid types are crucial for biofilm formation, antimycobacterial drug resistance or interactions with the immune system. Recent studies also indicate that analysis of mycolic acid profiles could be an alternative to conventional methods of diagnosis of diseases such as tuberculosis, leprosy or mycobacteriosis.

  11. Amplified voltammetric detection of glycoproteins using 4-mercaptophenylboronic acid/biotin-modified multifunctional gold nanoparticles as labels

    Liu L

    2014-05-01

    Full Text Available Lin Liu,1,2 Yun Xing,1 Hui Zhang,1 Ruili Liu,1 Huijing Liu,1 Ning Xia1,21College of Chemistry and Chemical Engineering, Anyang Normal University, Anyang, Henan, People’s Republic of China; 2College of Chemistry and Chemical Engineering, Central South University, Changsha, Hunan, People’s Republic of ChinaAbstract: Ultrasensitive detection of protein biomarkers is essential for early diagnosis and therapy of many diseases. Glycoproteins, differing from other types of proteins, contain carbohydrate moieties in the oligosaccharide chains. Boronic acid can form boronate ester covalent bonds with diol-containing species. Herein, we present a sensitive and cost-effective electrochemical method for glycoprotein detection using 4-mercaptophenylboronic acid (MBA/biotin-modified gold nanoparticles (AuNPs (MBA-biotin-AuNPs as labels. To demonstrate the feasibility and sensitivity of this method, recombinant human erythropoietin (rHuEPO was tested as a model analyte. Specifically, rHuEPO was captured by the anti-rHuEPO aptamer-covered electrode and then derivatized with MBA-biotin-AuNPs through the boronic acid–carbohydrate interaction. The MBA-biotin-AuNPs facilitated the attachment of streptavidin-conjugated alkaline phosphatase for the production of electroactive p-aminophenol from p-aminophenyl phosphate substrate. A detection limit of 8 fmol L-1 for rHuEPO detection was achieved. Other glycosylated and non-glycosylated proteins, such as horseradish peroxidase, prostate specific antigen, metallothionein, streptavidin, and thrombin showed no interference in the detection assay.Keywords: electrochemical biosensor, boronic acid, signal amplification, alkaline phosphatase

  12. Mycolic acids – biological role and potential application in Mycobacterium detection and differentiation

    Konrad Kowalski; Przemysław Trzepiński; Magdalena Druszczyńska; Janusz Boratyński

    2014-01-01

    Mycolic acids are one of the basic structural elements of the cell wall of bacteria from Corynebacterineae suborder. These compounds are long-chain α-hydroxy β-alkyl fatty acids with two hydrocarbon chains: longer meromycolic and shorter α-chain meromycolic α-chain. The genus Mycobacterium is characterized by the presence of mycolic acids in length from 60 to 90 carbon atoms having a fully saturated α-chain with a defined length of 22, 24 or 26 carbon atoms. Current research indicates that no...

  13. Quantifying the similarities within fold space.

    Harrison, Andrew; Pearl, Frances; Mott, Richard; Thornton, Janet; Orengo, Christine

    2002-11-01

    We have used GRATH, a graph-based structure comparison algorithm, to map the similarities between the different folds observed in the CATH domain structure database. Statistical analysis of the distributions of the fold similarities has allowed us to assess the significance for any similarity. Therefore we have examined whether it is best to represent folds as discrete entities or whether, in fact, a more accurate model would be a continuum wherein folds overlap via common motifs. To do this we have introduced a new statistical measure of fold similarity, termed gregariousness. For a particular fold, gregariousness measures how many other folds have a significant structural overlap with that fold, typically comprising 40% or more of the larger structure. Gregarious folds often contain commonly occurring super-secondary structural motifs, such as beta-meanders, greek keys, alpha-beta plait motifs or alpha-hairpins, which are matching similar motifs in other folds. Apart from one example, all the most gregarious folds matching 20% or more of the other folds in the database, are alpha-beta proteins. They also occur in highly populated architectural regions of fold space, adopting sandwich-like arrangements containing two or more layers of alpha-helices and beta-strands.Domains that exhibit a low gregariousness, are those that have very distinctive folds, with few common motifs or motifs that are packed in unusual arrangements. Most of the superhelices exhibit low gregariousness despite containing some commonly occurring super-secondary structural motifs. In these folds, these common motifs are combined in an unusual way and represent a small proportion of the fold (<10%). Our results suggest that fold space may be considered as continuous for some architectural arrangements (e.g. alpha-beta sandwiches), in that super-secondary motifs can be used to link neighbouring fold groups. However, in other regions of fold space much more discrete topologies are observed with

  14. Detection system of acid rain pollution using light-induced delayed fluorescence of plant leaf in vivo

    Zeng, Lizhang; Xing, Da

    2006-09-01

    Photosynthetic apparatus is susceptible to environmental stress. Light-induced delayed fluorescence (DF) in plant is an intrinsic label of the efficiency of charge separation at P680 in photosystem II (PS II). In this investigation, we have developed a biosensor that can accurately inspect acid rain pollution by means of DF in vivo. Compared with traditional methods, the proposed technique can continuously monitor environmental changes, making fast, real-time and noninvasive inspection possible. The biosensor is an all-weather measuring instrument; it has its own illumination power and utilizes intrinsic DF as the measurement marker. With soybean (Glycine max (L.) Merr.) seedling as a testing model, which is sensitive to acid rain pollution, the relationship that delayed fluorescence properties and capability of photosynthetic apparatus after being affected by simulated acid rain with different pH value was studied. The current investigation has revealed that the changes of delayed fluorescence (equation available in paper) can probably characterize the pollution degree of simulated acid rain, Inspecting the changes in DF characteristics (φ i) of plant leaf in vivo may be a new approach for the detection of acid rain pollution and its impact on the ecosystem.

  15. Nucleic Acid, Antibody, and Virus Culture Methods to Detect Xenotropic MLV-Related Virus in Human Blood Samples

    M. F. Kearney

    2011-01-01

    Full Text Available The MLV-related retrovirus, XMRV, was recently identified and reported to be associated with both prostate cancer and chronic fatigue syndrome. At the National Cancer Institute-Frederick, MD (NCI-Frederick, we developed highly sensitive methods to detect XMRV nucleic acids, antibodies, and replication competent virus. Analysis of XMRV-spiked samples and/or specimens from two pigtail macaques experimentally inoculated with 22Rv1 cell-derived XMRV confirmed the ability of the assays used to detect XMRV RNA and DNA, and culture isolatable virus when present, along with XMRV reactive antibody responses. Using these assays, we did not detect evidence of XMRV in blood samples ( or prostate specimens ( from two independent cohorts of patients with prostate cancer. Previous studies detected XMRV in prostate tissues. In the present study, we primarily investigated the levels of XMRV in blood plasma samples collected from patients with prostate cancer. These results demonstrate that while XMRV-related assays developed at the NCI-Frederick can readily measure XMRV nucleic acids, antibodies, and replication competent virus, no evidence of XMRV was found in the blood of patients with prostate cancer.

  16. Folded MEMS approach to NMRG

    Gundeti, Venu Madhav

    Atomic gyroscopes have a potential for good performance advantages and several attempts are being made to miniaturize them. This thesis describes the efforts made in implementing a Folded MEMS based NMRG. The micro implementations of all the essential components for NMRG (Nuclear Magnetic Resonance Gyroscope) are described in detail in regards to their design, fabrication, and characterization. A set of micro-scale Helmholtz coils are described and the homogeneity of the generated magnetic field is analyzed for different designs of heaters. The dielectric mirrors and metallic mirrors are compared in terms of reflectivity and polarization change up on reflection. A pyramid shaped folded backbone structure is designed, fabricated, and assembled along with all the required components. A novel double-folded structure 1/4th the size of original version is fabricated and assembled. Design and modeling details of a 5 layered shield with shielding factor > 106 and total volume of around 90 cc are also presented. A table top setup for characterization of atomic vapor cell is described in detail. A micro vapor cell based Rb magnetometer with a sensitivity of 108 pT/√Hz is demonstrated. The challenges due to DC heating are addressed and mitigated using an AC heater. Several experiments related to measuring the relaxation time of Xe are provided along with results. For Xe131, relaxation times of T1 = 23.78 sec, T2 = 18.06 sec and for Xe129, T1 = 21.65 sec and T2 = 20.45 sec are reported.

  17. Hydrodynamic interactions in protein folding

    Cieplak, Marek; Niewieczerzał, Szymon

    2009-03-01

    We incorporate hydrodynamic interactions (HIs) in a coarse-grained and structure-based model of proteins by employing the Rotne-Prager hydrodynamic tensor. We study several small proteins and demonstrate that HIs facilitate folding. We also study HIV-1 protease and show that HIs make the flap closing dynamics faster. The HIs are found to affect time correlation functions in the vicinity of the native state even though they have no impact on same time characteristics of the structure fluctuations around the native state.

  18. Direct detection of circulating free DNA extracted from serum samples of breast cancer using locked nucleic acid molecular beacon.

    Gui, Zhen; Wang, Quanbo; Li, Jinchang; Zhu, Mingchen; Yu, Lili; Xun, Tang; Yan, Feng; Ju, Huangxian

    2016-07-01

    As an emerging noninvasive blood biomarker, circulating free DNA (cfDNA) can be utilized to assess diagnosis, progression and evaluate prognosis of cancer. However, cfDNAs are not "naked", they can be part of complexes, or are bound to the surface of the cells via proteins, which make the detection more challenging. Here, a simple method for the detection of Ubiquitin-like with PHD and ring finger domains 1 (UHRF1) DNA exacted from serum of breast cancer (BC) has been developed using a novel locked nucleic acid molecular beacon (LNA-MB). In order to enhance the stability and detection efficiency of the probe in biofluids, we design a shared-stem molecular beacon containing a 27-mer loop and a 4-mer stem with DNA/LNA alternating bases. The fluorescence is released in the presence of target. The detection procedure is simple and can be completed within 1h. This method shows a sensitive response to UHRF1 DNA with a dynamic range of 3 orders of magnitude. The limit of detection is 11nM (S/N=3) with excellent selectivity. It can discriminate UHRF1 DNA from three-base mismatched DNA with a high specificity. More importantly, this method can distinguish the expression of serum UHRF1 DNA among 5 breast cancer patients and 5 healthy controls. The mentioned superiority may suggest that this assay can be served as a promising noninvasive detection tool for early BC diagnosis and monitoring. PMID:27154709

  19. 100-fold but not 50-fold dystrophin overexpression aggravates electrocardiographic defects in the mdx model of Duchenne muscular dystrophy.

    Yue, Yongping; Wasala, Nalinda B; Bostick, Brian; Duan, Dongsheng

    2016-01-01

    Dystrophin gene replacement holds the promise of treating Duchenne muscular dystrophy. Supraphysiological expression is a concern for all gene therapy studies. In the case of Duchenne muscular dystrophy, Chamberlain and colleagues found that 50-fold overexpression did not cause deleterious side effect in skeletal muscle. To determine whether excessive dystrophin expression in the heart is safe, we studied two lines of transgenic mdx mice that selectively expressed a therapeutic minidystrophin gene in the heart at 50-fold and 100-fold of the normal levels. In the line with 50-fold overexpression, minidystrophin showed sarcolemmal localization and electrocardiogram abnormalities were corrected. However, in the line with 100-fold overexpression, we not only detected sarcolemmal minidystrophin expression but also observed accumulation of minidystrophin vesicles in the sarcoplasm. Excessive minidystrophin expression did not correct tachycardia, a characteristic feature of Duchenne muscular dystrophy. Importantly, several electrocardiogram parameters (QT interval, QRS duration and the cardiomyopathy index) became worse than that of mdx mice. Our data suggests that the mouse heart can tolerate 50-fold minidystrophin overexpression, but 100-fold overexpression leads to cardiac toxicity. PMID:27419194

  20. 100-fold but not 50-fold dystrophin overexpression aggravates electrocardiographic defects in the mdx model of Duchenne muscular dystrophy

    Yue, Yongping; Wasala, Nalinda B; Bostick, Brian; Duan, Dongsheng

    2016-01-01

    Dystrophin gene replacement holds the promise of treating Duchenne muscular dystrophy. Supraphysiological expression is a concern for all gene therapy studies. In the case of Duchenne muscular dystrophy, Chamberlain and colleagues found that 50-fold overexpression did not cause deleterious side effect in skeletal muscle. To determine whether excessive dystrophin expression in the heart is safe, we studied two lines of transgenic mdx mice that selectively expressed a therapeutic minidystrophin gene in the heart at 50-fold and 100-fold of the normal levels. In the line with 50-fold overexpression, minidystrophin showed sarcolemmal localization and electrocardiogram abnormalities were corrected. However, in the line with 100-fold overexpression, we not only detected sarcolemmal minidystrophin expression but also observed accumulation of minidystrophin vesicles in the sarcoplasm. Excessive minidystrophin expression did not correct tachycardia, a characteristic feature of Duchenne muscular dystrophy. Importantly, several electrocardiogram parameters (QT interval, QRS duration and the cardiomyopathy index) became worse than that of mdx mice. Our data suggests that the mouse heart can tolerate 50-fold minidystrophin overexpression, but 100-fold overexpression leads to cardiac toxicity. PMID:27419194

  1. Electrochemical Co-Reduction Synthesis of AuPt Bimetallic Nanoparticles-Graphene Nanocomposites for Selective Detection of Dopamine in the Presence of Ascorbic Acid and Uric Acid

    Zongya Zhao

    2015-07-01

    Full Text Available In this paper, AuPt bimetallic nanoparticles-graphene nanocomposites were obtained by electrochemical co-reduction of graphene oxide (GO, HAuCl4 and H2PtCl6. The as-prepared AuPt bimetallic nanoparticles-graphene nanocomposites were characterized by scanning electron microscopy (SEM, electrochemical impedance spectroscopy (EIS and other electrochemical methods. The morphology and composition of the nanocomposite could be easily controlled by adjusting the HAuCl4/H2PtCl6 concentration ratio. The electrochemical experiments showed that when the concentration ratio of HAuCl4/H2PtCl6 was 1:1, the obtained AuPt bimetallic nanoparticles-graphene nanocomposite (denoted as Au1Pt1NPs-GR possessed the highest electrocatalytic activity toward dopamine (DA. As such, Au1Pt1NPs-GR nanocomposites were used to detect DA in the presence of ascorbic acid (AA and uric acid (UA using the differential pulse voltammetry (DPV technique and on the modified electrode, there were three separate DPV oxidation peaks with the peak potential separations of 177 mV, 130 mV and 307 mV for DA and AA, DA and UA, AA and UA, respectively. The linear range of the constructed DA sensor was from 1.6 μM to 39.7 μM with a detection limit of 0.1 μM (S/N = 3. The obtained DA sensor with good stability, high reproducibility and excellent selectivity made it possible to detect DA in human urine samples.

  2. Electrodes Modification Based on Metal-Free Phthalocyanine: Example of Electrochemical Sensors for the Detection of Acetic Acid

    Amadou L. Ndiaye

    2015-01-01

    Full Text Available Electroanalytical properties of tetra-tert-butyl phthalocyanine (PcH2-tBu modified electrodes are studied by cyclic voltammetry (CV. The modified electrodes are obtained by CV deposition techniques on gold (Au and glassy carbon (C screen-printed electrodes (SPEs and used for the electrochemical detection of acetic acid (AA. Based on the CV experiments, the electrodeposition mechanism is detailed. The modified PcH2-tBu electrodes reveal one oxidation and one reduction peak within the potential window of the working electrodes. In the presence of the analyte (acetic acid, the modified electrodes show sensitivity in the range of 10 mM to 400 mM. For the PcH2-tBu modified Au electrode, a limit of detection (LOD of 5.89 mM (based on the +0.06 V peak was obtained while for the PcH2-tBu modified C electrode a LOD of 17.76 mM (based on the +0.07 V peak was achieved. A signal decay of 17%, based on 20 experiments, is obtained when gold is used as working electrode. If carbon is used as working electrode a value of 7% is attained. A signal decay is observed after more than 50 cycles of experiments and is more pronounced when higher concentrations of acetic acid are used. A mechanism of sensing is proposed at the end.

  3. Detection of Elevated Signaling Amino Acids in Human Diabetic Vitreous by Rapid Capillary Electrophoresis

    Miao-Jen Lu

    2007-01-01

    Full Text Available Elevated glutamate is implicated in the pathology of PDR. The ability to rapidly assess the glutamate and amino acid content of vitreous provides a more complete picture of the chemical changes occurring at the diabetic retina and may lead to a better understanding of the pathology of PDR. Vitreous humor was collected following vitrectomies of patients with PDR and control conditions of macular hole or epiretinal membrane. A capillary electrophoresis method was developed to quantify glutamate and arginine. The analysis is relatively fast (<6 minutes and utilizes a poly(ethyleneoxide and sodium dodecylsulfate run buffer. Both amino acid levels show significant increases in PDR patients versus controls and are comparable to other reports. The levels of vitreal glutamate vary inversely with the degree of observed hemorrhage. The results demonstrate a rapid method for assessment of a number of amino acids to characterize the chemical changes at the diabetic retina to better understand tissue changes and potentially identify new treatments.

  4. Topology, Geometry, and Stability: Protein Folding and Evolution

    Simmons, Walter

    2015-01-01

    The protein folding problem must ultimately be solved on all length scales from the atomic up through a hierarchy of complicated structures. By analyzing the stability of the folding process using physics and mathematics, this paper shows that features without length scales, i.e. topological features, are potentially of central importance. Topology is a natural mathematical tool for the study of shape and we avail ourselves of that tool to examine the relationship between the amino acid sequence and the shapes of protein molecules. We apply what we learn to conjectures about their biological evolution.

  5. Sensitive detection of nucleic acids by PNA hybridization directed co-localization of fluorescent beads

    Shiraishi, Takehiko; Deborggraeve, Stijn; Büscher, Philippe; Nielsen, Peter E

    2011-01-01

    We have designed a pair of biotinylated peptide nucleic acid (PNA) probes targeting two sequences in 18S rRNA (from the parasite Trypanosoma brucei) at a distance of 191 nt (corresponding to maximum distance of ca. 60 nm) from each other. The PNA probes were individually bound to (strept)avidin-c......We have designed a pair of biotinylated peptide nucleic acid (PNA) probes targeting two sequences in 18S rRNA (from the parasite Trypanosoma brucei) at a distance of 191 nt (corresponding to maximum distance of ca. 60 nm) from each other. The PNA probes were individually bound to (strept...

  6. Direct Colorimetric Detection of Hydrogen Peroxide Using 4-Nitrophenyl Boronic Acid or Its Pinacol Ester

    Gregory Su; Yibin Wei; Maolin Guo

    2011-01-01

    A colorimetric method for the direct determination of hydrogen peroxide in aqueous solution is described. H2O2 stoichiometrically converts 4-nitrophenyl boronic acid or 4-nitrophenyl boronic acid pinacol ester into 4-nitrophenol, which can be quantified by measuring the absorption at 400 nm in neutral or basic media. The reactions proceed fast under basic conditions and complete in 2 minutes to at pH 11 and 80?C. The linear range for the colorimetric method extends beyond 1.0 to 40 µM H2O2, a...

  7. Poly(3,4-ethylenedioxythiophene-co-(5-amino-2-naphthalenesulfonic acid)) (PEDOT-PANS) film modified glassy carbon electrode for selective detection of dopamine in the presence of ascorbic acid and uric acid

    Poly(3,4-ethylenedioxythiophene-co-(5-amino-2-naphthalenesulfonic acid)) (PEDOT-PANS) film modified glassy carbon electrode was prepared by electrochemical polymerization technique. The properties of modified electrode was studied. It was found that the electrochemical properties of modified electrode was very much dependent on the experimental conditions, such as monomer oxidation potential and pH. The modified electrode surface was characterized by scanning electron microscopy (SEM). The PEDOT-PANS film modified electrode shows electrocatalytic activity toward oxidation of dopamine (DA) in acetate buffer solution (pH 5.0) and results in a marked enhancement of the current response. The linear sweep voltammetric (LSV) peak heights are linear with DA concentration from 2 x 10-6 to 1 x 10-5 M. The detection limit is 5 x 10-7 M. More over, the interferences of ascorbic acid (AA) and uric acid (UA) were effectively diminished. This work provides a simple and easy approach for selective determination of dopamine in the presence of ascorbic acid and uric acid

  8. Detection of Staphylococcus epidermidis by a Quartz Crystal Microbalance Nucleic Acid Biosensor Array Using Au Nanoparticle Signal Amplification

    Weiling Fu

    2008-10-01

    Full Text Available Staphylococcus epidermidis is a critical pathogen of nosocomial blood infections, resulting in significant morbidity and mortality. A piezoelectric quartz crystal microbalance (QCM nucleic acid biosensor array using Au nanoparticle signal amplification was developed to rapidly detect S. epidermidis in clinical samples. The synthesized thiolated probes specific targeting S. epidermidis 16S rRNA gene were immobilized on the surface of QCM nucleic acid biosensor arrays. Hybridization was induced by exposing the immobilized probes to the PCR amplified fragments of S. epidermidis, resulting in a mass change and a consequent frequency shift of the QCM biosensor. To further enhance frequency shift results from above described hybridizations, streptavidin coated Au nanoparticles were conjugated to the PCR amplified fragments. The results showed that the lowest detection limit of current QCM system was 1.3×103 CFU/mL. A linear correlation was found when the concentration of S. epidermidis varied from 1.3×103 to 1.3×107 CFU/mL. In addition, 55 clinical samples were detected with both current QCM biosensor system and conventional clinical microbiological method, and the sensitivity and specificity of current QCM biosensor system were 97.14% and 100%, respectively. In conclusion, the current QCM system is a rapid, low-cost and sensitive method that can be used to identify infection of S. epidermidis in clinical samples.

  9. Automated flow-through amperometric immunosensor for highly sensitive and on-line detection of okadaic acid in mussel sample.

    Dominguez, Rocio B; Hayat, Akhtar; Sassolas, Audrey; Alonso, Gustavo A; Munoz, Roberto; Marty, Jean-Louis

    2012-09-15

    An electrochemical immunosensor for okadaic acid (OA) detection has been developed, and used in an indirect competitive immunoassay format under automated flow conditions. The biosensor was fabricated by injecting OA modified magnetic beads onto screen printed carbon electrode (SPCE) in the flow system. The OA present in the sample competed with the immobilized OA to bind with anti-okadaic acid monoclonal antibody (anti-OA-MAb). The secondary alkaline phosphatase labeled antibody was used to perform electrochemical detection. The current response obtained from the labeled alkaline phosphatase to 1-naphthyl phosphate decreased proportionally to the concentration of free OA in the sample. The calculated limit of detection (LOD) was 0.15 μg/L with a linear range of 0.19-25 μg/L. The good recoveries percentages validated the immunosensor application for real mussel samples. The developed system automatically controlled the incubation, washing and current measurement steps, showing its potential use for OA determination in field analysis. PMID:22967546

  10. Two-stage sample-to-answer system based on nucleic acid amplification approach for detection of malaria parasites.

    Liu, Qing; Nam, Jeonghun; Kim, Sangho; Lim, Chwee Teck; Park, Mi Kyoung; Shin, Yong

    2016-08-15

    Rapid, early, and accurate diagnosis of malaria is essential for effective disease management and surveillance, and can reduce morbidity and mortality associated with the disease. Although significant advances have been achieved for the diagnosis of malaria, these technologies are still far from ideal, being time consuming, complex and poorly sensitive as well as requiring separate assays for sample processing and detection. Therefore, the development of a fast and sensitive method that can integrate sample processing with detection of malarial infection is desirable. Here, we report a two-stage sample-to-answer system based on nucleic acid amplification approach for detection of malaria parasites. It combines the Dimethyl adipimidate (DMA)/Thin film Sample processing (DTS) technique as a first stage and the Mach-Zehnder Interferometer-Isothermal solid-phase DNA Amplification (MZI-IDA) sensing technique as a second stage. The system can extract DNA from malarial parasites using DTS technique in a closed system, not only reducing sample loss and contamination, but also facilitating the multiplexed malarial DNA detection using the fast and accurate MZI-IDA technique. Here, we demonstrated that this system can deliver results within 60min (including sample processing, amplification and detection) with high sensitivity (malaria in low-resource settings. PMID:27031184

  11. Sensitive Detection of Capsaicinoids Using a Surface Plasmon Resonance Sensor with Anti-Homovanillic Acid Polyclonal Antibodies

    Kiyoshi Toko

    2013-11-01

    Full Text Available Recently, highly functional biosensors have been developed in preparation for possible large-scale terrorist attacks using chemical warfare agents. Practically applicable sensors are required to have various abilities, such as high portability and operability, the capability of performing rapid and continuous measurement, as well as high sensitivity and selectivity. We developed the detection method of capsaicinoids, the main component of some lachrymators, using a surface plasmon resonance (SPR immunosensor as an on-site detection sensor. Homovanillic acid, which has a vanillyl group similar to capsaicinoids such as capsaicin and dihydrocapsaicin, was bound to Concholepas concholepas hemocyanin (CCH for use as an immunogen to generate polyclonal antibodies. An indirect competitive assay was carried out to detect capsaicinoids using SPR sensor chips on which different capsaicin analogues were immobilized. For the sensor chip on which 4-hydroxy-3-methoxybenzylamine hydrochloride was immobilized, a detection limit of 150 ppb was achieved. We found that the incubation time was not required and the detection can be completed in five minutes.

  12. Sensitive detection of capsaicinoids using a surface plasmon resonance sensor with anti-homovanillic Acid polyclonal antibodies.

    Nakamura, Shingo; Yatabe, Rui; Onodera, Takeshi; Toko, Kiyoshi

    2013-01-01

    Recently, highly functional biosensors have been developed in preparation for possible large-scale terrorist attacks using chemical warfare agents. Practically applicable sensors are required to have various abilities, such as high portability and operability, the capability of performing rapid and continuous measurement, as well as high sensitivity and selectivity. We developed the detection method of capsaicinoids, the main component of some lachrymators, using a surface plasmon resonance (SPR) immunosensor as an on-site detection sensor. Homovanillic acid, which has a vanillyl group similar to capsaicinoids such as capsaicin and dihydrocapsaicin, was bound to Concholepas concholepas hemocyanin (CCH) for use as an immunogen to generate polyclonal antibodies. An indirect competitive assay was carried out to detect capsaicinoids using SPR sensor chips on which different capsaicin analogues were immobilized. For the sensor chip on which 4-hydroxy-3-methoxybenzylamine hydrochloride was immobilized, a detection limit of 150 ppb was achieved. We found that the incubation time was not required and the detection can be completed in five minutes. PMID:25586413

  13. Conductimetric Biosensor for the Detection of Uric Acid by Immobilization Uricase on Nata de Coco Membrane—Pt Electrode

    Mulyasuryani, Ani; Srihardiastutie, Arie

    2011-01-01

    A conductimetric enzyme biosensor for uric acid detection has been developed. The uricase, as enzyme, is isolated from Candida utilis and immobilized on a nata de coco membrane-Pt electrode. The biosensor demonstrates a linear response to urate over the concentration range 1–6 ppm and has good selectivity properties. The response is affected by the membrane thickness and pH change in the range 7.5–9.5. The response time is three minutes in aqueous solutions and in human serum samples. Application of the biosensor to the determination of uric acid in human serum gave results that compared favourably with those obtained by medical laboratory. The operational stability of the biosensor was not less than three days and the relative error is smaller than 10%. PMID:21792276

  14. Conductimetric biosensor for the detection of uric Acid by immobilization uricase on nata de coco membrane-pt electrode.

    Mulyasuryani, Ani; Srihardiastutie, Arie

    2011-01-01

    A conductimetric enzyme biosensor for uric acid detection has been developed. The uricase, as enzyme, is isolated from Candida utilis and immobilized on a nata de coco membrane-Pt electrode. The biosensor demonstrates a linear response to urate over the concentration range 1-6 ppm and has good selectivity properties. The response is affected by the membrane thickness and pH change in the range 7.5-9.5. The response time is three minutes in aqueous solutions and in human serum samples. Application of the biosensor to the determination of uric acid in human serum gave results that compared favourably with those obtained by medical laboratory. The operational stability of the biosensor was not less than three days and the relative error is smaller than 10%. PMID:21792276

  15. Near-infrared Spectral Detection of the Content of Soybean Fat Acids Based on Genetic Multilayer Feed forward Neural Network

    CHAI Yu-hua; PAN Wei; NING Hai-long

    2005-01-01

    In the paper, a method of building mathematic model employing genetic multilayer feed forward neural network is presented, and the quantitative relationship of chemical measured values and near-infrared spectral data is established. In the paper, quantitative mathematic model related chemical assayed values and near-infrared spectral data is established by means of genetic multilayer feed forward neural network, acquired near-infrared spectral data are taken as input of network with the content of five kinds of fat acids tested from chemical method as output,weight values of multilayer feed forward neural network are trained by genetic algorithms and detection model of neural network of soybean is built. A kind of multilayer feed forward neural network trained by genetic algorithms is designed in the paper. Through experiments, all the related coefficients of five fat acids can approach 0.9 which satisfies the preliminary test of soybean breeding.

  16. Carbonic Acid Revisited: Critical Remarks on Theoretical Predictions and the Possibility of Detecting an Elusive Molecule

    Huber, Stefan E; Kausch, Wolfgang; Kimeswenger, Stefan; Probst, Michael

    2012-01-01

    We calculate harmonic frequencies of the three most abundant carbonic acid conformers. For this, different model chemistries are investigated with respect to their benefits and shortcomings. Based on these results we use perturbation theory to calculate anharmonic corrections at the {\\omega}B97XD/aug-cc-pVXZ, X=D, T, Q, level of theory and compare them with recent experimental data and theoretical predictions. A discrete variable representation method is used to predict the large anharmonic contributions to the frequencies of the stretching vibrations in the hydrogen bonds in the carbonic acid dimer. Moreover, we re-investigate the energetics of the formation of the carbonic acid dimer from its constituents water and carbon dioxide using a high-level extrapolation method. We find that the {\\omega}B97XD functional performs well in estimating the fundamental frequencies of the carbonic acid conformers. Concerning the reaction energetics, the accuracy of {\\omega}B97XD is even comparable to the high-level extrapo...

  17. Direct determination of amino acids by hydrophilic interaction liquid chromatography with charged aerosol detection.

    Socia, Adam; Foley, Joe P

    2016-05-13

    A chromatographic analytical method for the direct determination of amino acids by hydrophilic interaction liquid chromatography (HILIC) was developed. A dual gradient simultaneously varying the pH 3.2 ammonium formate buffer concentration and level of acetonitrile (ACN) in the mobile phase was employed. Using a charged aerosol detector (CAD) and a 2(nd) order regression analysis, the fit of the calibration curve showed R(2) values between 0.9997 and 0.9985 from 1.5mg/mL to 50μg/mL (600ng to 20ng on column). Analyte chromatographic parameters such as the sensitivity of retention to the water fraction in the mobile phase values (mHILIC) were determined as part of method development. A degradation product of glutamine (5-pyrrolidone-2-carboxylic acid; pGlu) was observed and resolved chromatographically with no method modifications. The separation was used to quantitate amino acid content in acid hydrolysates of various protein samples. PMID:27059400

  18. Detecting relationships between amylose content and amino acid contents of indica rice with conditional approach

    Chun Hai Shi; Qiong Qiong Zhu; Ke Ming Wang; Guo Ke Ge; Jian Guo Wu; Zhen Ghao Xu

    2010-04-01

    The relationship between the genetic effects of endosperm, cytoplasm and maternal plant on amylose content (AC) and amino acid contents of indica rice was studied using unconditional and conditional analysis methods. The results indicated that the protein content (PC) and brown rice weight (WBR) could significantly affect the relationships between AC and amino acid contents of rice. The phenotypic and genotypic covariances between AC and amino acid contents were most significantly negative under the interference of PC or WBR, but most of the relationships for the paired traits were not significant after excluding the influence of PC or WBR on AC. For the conditional genetic relationship analysis of different genetic systems including endosperm, cytoplasm and maternal plant, visible changes were found in many genetic correlation components between AC and amino acid content after eliminating the influences of PC, especially, for the endosperm or maternal additive effects, endosperm additive or dominance interaction effects and maternal additive interaction effects. The relationships of the paired traits conditioned on WBR were mainly controlled by the endosperm dominance or additive interaction effects.

  19. BIENZYME REACTOR FOR ELECTROCHEMICAL DETECTION OF LOW CONCENTRATIONS OF URIC-ACID AND GLUCOSE

    ELEKES, O; MOSCONE, D; VENEMA, K; KORF, J

    1995-01-01

    An enzyme-based flow-injection amperometric analysis system (FIA) for monitoring of uric acid and glucose is described, The oxidase and peroxidase enzymes are physically co-immobilised in a sandwich-type reactor and ferrocene serves as a mediator. The assays are based on the measurement of a reducti

  20. 78 FR 16513 - Application of Advances in Nucleic Acid and Protein Based Detection Methods to Multiplex...

    2013-03-15

    ... workshop are to review the status of multiplex platforms and the technological advances in gene based and... academic institutions, blood establishments, industry, and government agencies. Date and Time: The public... HUMAN SERVICES Food and Drug Administration Application of Advances in Nucleic Acid and Protein...

  1. Stretching Folding Instability and Nanoemulsions

    Chan, Chon U

    2009-01-01

    Here we show a folding-stretching instability in a microfluidic flow focusing device using silicon oil (100cSt) and water. The fluid dynamics video demonstrates an oscillating thread of oil focused by two co-flowing streams of water. We show several high-speed sequences of these oscillations with 30,000 frames/s. Once the thread is decelerated in a slower moving pool downstream an instability sets in and water-in-oil droplets are formed. We reveal the details of the pinch-off with 500,000 frames/s. The pinch-off is so repeatable that complex droplet patterns emerge. Some of droplets are below the resolution limit, thus smaller than 1 micrometer in diameter.

  2. Motives of uniruled 3-folds

    Angel, P L; Angel, Pedro Luis del; Mueller-Stach, Stefan

    1996-01-01

    We construct projectors in the ring of correspondences of a complex uniruled 3-fold $X$ which lift the Kuenneth components of the diagonal in singular cohomology and have other properties which were conjectured by J. Murre. Such Murre decompositions have been already obtained for curves, surfaces, abelian varieties and varieties with cell decompositions by the work of Manin, Shermenev, Beauville, Murre et.al.. In particular they define a natural filtration on the Chow groups of $X$ which was conjectured by Bloch and Beilinson. To do this we use Mori theory and construct projectors in the situation of a fiber space over a surface. These projectors may also be used in more general situations.

  3. A Study on Tannic Acid-doped Polypyrrole Films on Gold Electrodes for Selective Electrochemical Detection of Dopamine

    Shouzhuo Yao

    2005-04-01

    Full Text Available Tannic acid-doped polypyrrole (PPY/TA films have been grown on goldelectrodes for selective electrochemical detection of dopamine (DA. Electrochemicalquartz crystal microbalance (EQCM studies revealed that, in vivid contrast toperchlorate-doped polypyrrole films (PPY/ClO4-, the redox switching of PPY/TA filmsin aqueous solutions involved only cation transport if the solution pH was greater than3~4. The PPY/TA Au electrodes also exhibited attractive permselectivity forelectroactive cations, namely, effectively blocking the electrochemical reactions ofanionic ferricyanide and ascorbic acid (AA while well retaining the electrochemicalactivities of hexaammineruthenium (III and dopamine as cationic species. A 500 HzPPY/TA film could effectively block the redox current of up to 5.0 mM AA. Thecoexistence of ascorbic acid in the measurement solution notably enhanced the currentsignal for dopamine oxidation, due probably to the chemical regeneration of dopaminethrough an ascorbic acid-catalyzed reduction of the electro-oxidation product ofdopamine (EC’ mechanism, and the greatest amplification was found at an ascorbic acidconcentration of 1.0 mM. The differential pulse voltammetry peak current for DAoxidation was linear with DA concentration in the range of 0 to 10 μM, with sensitivityof 0.125 and 0.268 μA/μM, as well as lower detection limit of 2.0 and 0.3 μM in a PBSsolution without AA and with 1.0 mM coexisting AA, respectively.

  4. Highly Sensitive and Selective Detection of Dopamine at Poly(chromotrope 2B)-Modified Glassy Carbon Electrode in the Presence of Uric Acid and Ascorbic Acid

    A highly sensitive and selective electrochemical method based on a poly(chromotrope 2B)-modified anodized glassy carbon electrode (PCHAGCE) was developed for the determination of dopamine (DA) in the presence of uric acid (UA) and ascorbic acid (AA). The PCHAGCE sensor exhibited excellent electron-mediating behavior towards the oxidation of DA in 0.1 M phosphate buffer solution (PBS) (pH 7.0). It was found that the electrocatalytic activity was significantly dependent on the charge status and molecular structure of the target molecules. Differential pulse voltammetry (DPV) measurements revealed oxidation signals for DA, UA, and AA that were well-resolved into three distinct peaks with AA–DA, DA–UA, and AA–UA peak potential separations (ΔEp) of 172, 132, and 304 mV, respectively. A detection limit of 0.04 ± 0.001 μM (S/N = 3) and a quantification limit (S/N = 10) of 0.149 ± 0.03 μM were obtained for DA sensing in a linear range of 1 to 40 μM in PBS (pH 7.0) with a very high sensitivity of 1.522 ± 0.032 μA·μM−1. The DA concentrations in human urine samples were also successfully determined with recoveries of 94.0–98.0%. This approach provides a simple, easy, sensitive, and selective method to detect DA in the presence of AA and UA

  5. Conductimetric Biosensor for the Detection of Uric Acid by Immobilization Uricase on Nata de Coco Membrane—Pt Electrode

    Ani Mulyasuryani; Arie Srihardiastutie

    2011-01-01

    A conductimetric enzyme biosensor for uric acid detection has been developed. The uricase, as enzyme, is isolated from Candida utilis and immobilized on a nata de coco membrane-Pt electrode. The biosensor demonstrates a linear response to urate over the concentration range 1–6 ppm and has good selectivity properties. The response is affected by the membrane thickness and pH change in the range 7.5–9.5. The response time is three minutes in aqueous solutions and in human serum samples. Applica...

  6. In situ detection of denitrifying bacteria by mRNA-targeted nucleic acid probes and catalyzed reporter deposition

    Kofoed, Michael Vedel; Stief, Peter; Poulsen, Morten; Schramm, Andreas

    In situ detection of denitrifying bacteria by mRNA-targeted nucleic acid probes and catalyzed reporter deposition   Michael V.W. Kofoed, Peter Stief, Morten Poulsen, and Andreas Schramm Department of Biological Sciences, Microbiology, University of Aarhus, Denmark Denitrification, the sequential...... and catalyzed fluorescent reporter deposition (CARD-FISH). The general feasibility of the approach was first tested with pure cultures of Pseudomonas stutzeri and various denitrifying and nitrate-reducing isolates. Detailed studies of probe specificity and hybridization conditions using Clone-FISH of...

  7. Energy driven cascade recognition for selective detection of nucleic acids with high discrimination factor at room temperature.

    Zhang, Zhang; Li, Jun Long; Yao, Juan; Wang, Ting; Yin, Dan; Xiang, Yu; Chen, Zhongping; Xie, Guoming

    2016-05-15

    In this article, we demonstrated a cascade recognition strategy for the detection of single strand nucleic acid with high discrimination factor at room temperature. The cascade recognition strategy contains a toehold mediated strand displacement and a double-toehold mediated double strand displacement reaction, thus enable the high ability to discern point mutation of target. The discrimination factor of the model target is between 45 and 109, with the medium of 70. This strategy is homogeneous, easy operation, enzyme-free, isothermal, and can be easily adapted to high-throughput devices without the need of designing complicated instruments. PMID:26745796

  8. A spatio-temporal mining approach towards summarizing and analyzing protein folding trajectories

    Ucar Duygu

    2007-04-01

    Full Text Available Abstract Understanding the protein folding mechanism remains a grand challenge in structural biology. In the past several years, computational theories in molecular dynamics have been employed to shed light on the folding process. Coupled with high computing power and large scale storage, researchers now can computationally simulate the protein folding process in atomistic details at femtosecond temporal resolution. Such simulation often produces a large number of folding trajectories, each consisting of a series of 3D conformations of the protein under study. As a result, effectively managing and analyzing such trajectories is becoming increasingly important. In this article, we present a spatio-temporal mining approach to analyze protein folding trajectories. It exploits the simplicity of contact maps, while also integrating 3D structural information in the analysis. It characterizes the dynamic folding process by first identifying spatio-temporal association patterns in contact maps, then studying how such patterns evolve along a folding trajectory. We demonstrate that such patterns can be leveraged to summarize folding trajectories, and to facilitate the detection and ordering of important folding events along a folding path. We also show that such patterns can be used to identify a consensus partial folding pathway across multiple folding trajectories. Furthermore, we argue that such patterns can capture both local and global structural topology in a 3D protein conformation, thereby facilitating effective structural comparison amongst conformations. We apply this approach to analyze the folding trajectories of two small synthetic proteins-BBA5 and GSGS (or Beta3S. We show that this approach is promising towards addressing the above issues, namely, folding trajectory summarization, folding events detection and ordering, and consensus partial folding pathway identification across trajectories.

  9. Simultaneous/Selective Detection of Dopamine and Ascorbic Acid at Synthetic Zeolite-Modified/Graphite-Epoxy Composite Macro/Quasi-Microelectrodes

    Rodica Pode; Elida Cristina Ilinoiu; Pier Andrea Serra; Florica Manea

    2013-01-01

    The present paper aims to miniaturize a graphite-epoxy and synthetic zeolite-modified graphite-epoxy composite macroelectrode as a quasi-microelectrode aiming in vitro and also, envisaging in vivo simultaneous electrochemical detection of dopamine (DA) and ascorbic acid (AA) neurotransmitters, or DA detection in the presence of AA. The electrochemical behavior and the response of the designed materials to the presence of dopamine and ascorbic acid without any protective membranes were studied...

  10. Determination of Amino Acids in Cell Culture and Fermentation Broth Media Using Anion-Exchange Chromatography with Integrated Pulsed Amperometric Detection

    Hanko, Valoran P.; Heckenberg, Andrea; Rohrer, Jeffrey S.

    2004-01-01

    Anion-exchange chromatography with integrated pulsed amperometric detection (AE-IPAD) separates and directly detects amino acids, carbohydrates, alditols, and glycols in the same injection without pre- or post-column derivatization. These separations use a combination of NaOH and NaOH/sodium acetate eluents. We previously published the successful use of this technique, also known as AAA-Direct, to determine free amino acids in cell culture and fermentation broth media. We showed that retentio...

  11. Accelerated molecular dynamics simulations of protein folding

    Miao, Y.; Feixas, F; Eun, C; McCammon, JA

    2015-01-01

    © 2015 Wiley Periodicals, Inc. Folding of four fast-folding proteins, including chignolin, Trp-cage, villin headpiece and WW domain, was simulated via accelerated molecular dynamics (aMD). In comparison with hundred-of-microsecond timescale conventional molecular dynamics (cMD) simulations performed on the Anton supercomputer, aMD captured complete folding of the four proteins in significantly shorter simulation time. The folded protein conformations were found within 0.2-2.1 Å of the native ...

  12. Protein folding in a force-clamp

    Cieplak, Marek; Szymczak, Piotr

    2006-03-01

    Kinetics of folding of a protein held in a force-clamp are compared to an unconstrained folding. The comparison is made within a simple topology-based dynamical model of ubiquitin. We demonstrate that the experimentally observed rapid changes in the end-to-end distance mirror microscopic events during folding. However, the folding scenarios in and out of the force-clamp are distinct.

  13. Protein folding in a force-clamp

    Cieplak, Marek; Szymczak, Piotr

    2006-01-01

    Kinetics of folding of a protein held in a force-clamp are compared to an unconstrained folding. The comparison is made within a simple topology-based dynamical model of ubiquitin. We demonstrate that the experimentally observed variations in the end-to-end distance reflect microscopic events during folding. However, the folding scenarios in and out of the force-clamp are distinct.

  14. Electrocatalytic detection of dopamine in the presence of ascorbic acid and uric acid using single-walled carbon nanotubes modified electrode.

    Li, Yaya; Du, Jie; Yang, Jiandong; Liu, Dong; Lu, Xiaoquan

    2012-09-01

    Single-walled carbon nanotubes (SWCNTs) fabricated by sodium dodecyl sulfate (SDS) (f-SWCNTs) modified glassy carbon electrodes (f-SWCNTs/GCE) for the simultaneous determination of ascorbic acid (AA), dopamine (DA) and uric acid (UA). The f-SWCNTs/GCE displayed very good electrochemical catalytic activities with respect to GCE. The oxidation over-potentials of DA and UA decreased dramatically, and their oxidation peak currents increased significantly at f-SWCNTs/GCE compared to those obtained at the bare GCE. Simultaneously, the oxidation peak currents of AA decreased accordingly. The f-SWCNTs/GCE not only divide the overlapping voltammetric responses of them into individual voltammetric peaks, but also totally eliminate the interference from AA and distinguish DA from UA. The catalytic peak currents obtained from square-wave voltammetry increased linearly with increasing DA concentrations in the range of 5.0×10(-6) to 1.0×10(-4)M with a detection limit of 2.0×10(-8)M (S/N=3). The method was also successfully applied for determination of DA and showed good recovery in some biological fluids. PMID:22580482

  15. Determination of Amino Acids in an Individual Erythrocyteby Capillary Electrophoresis with Intracellular FITC-derivatization and Laser-induced Fluorescence Detection

    Hua ZHANG; Wen Rui JIN

    2003-01-01

    A novel approach for analysis of amino acids in individual erythrocytes was established.In this method, the derivatization reagent was introduced into the living cells by electroporation.After derivatization, the amino acids in a single cell were determined by capillary electrophoresiswith laser-induced fluorescence detection.

  16. Analysis of Amino Acids in a Single Human Red Blood Cell by Capillary Zone Electrophoresis with Intracellular NDA—derivatization and Electrochemical Detection

    QianDONG; XiaoLeiWANG; 等

    2002-01-01

    A novel method for determination of amino acids in individual red blood cells has been developed. In this method, the derivatization reagents (NDA and CN-) are introduced into living cells by electroporation. After completion of derivatization,the amino acids in a single cell is determined by capillary zone electrophoresis with end-column amperometric detection.

  17. Analysis of Amino Acids in a Single Human Red Blood Cell by Capillary Zone Electrophoresis with Intracellular NDA derivatization and Electrochemical Detection

    2002-01-01

    A novel method for detcrmination of amino acids in individual human red blood cells has been dcveloped. In this method, the derivatization reagents (NDA and CN-) are introduced into living cells by clcctroporation. After completion of derivatization, the amino acids in a single cell is determined by capillary zone electrophoresis with end-column ampcrometric detection.

  18. Functional stimuli responsive hydrogel devices by self-folding

    We describe a photolithographic approach to create functional stimuli responsive, self-folding, microscale hydrogel devices using thin, gradient cross-linked hinges and thick, fully cross-linked panels. The hydrogels are composed of poly (N-isopropylacrylamide-co-acrylic acid) (pNIPAM-AAc) with reversible stimuli responsive properties just below physiological temperatures. We show that a variety of three-dimensional structures can be formed and reversibly actuated by temperature or pH. We experimentally characterized the swelling and mechanical properties of pNIPAM-AAc and developed a finite element model to rationalize self-folding and its variation with hinge thickness and swelling ratio. Finally, we highlight applications of this approach in the creation of functional devices such as self-folding polymeric micro-capsules, untethered micro-grippers and thermally steered micro-mirror systems. (paper)

  19. Characterization of Non-Trivial Neighborhood Fold Constraints from Protein Sequences using Generalized Topohydrophobicity

    Guillaume Fourty

    2008-01-01

    Full Text Available Prediction of key features of protein structures, such as secondary structure, solvent accessibility and number of contacts between residues, provides useful structural constraints for comparative modeling, fold recognition, ab-initio fold prediction and detection of remote relationships. In this study, we aim at characterizing the number of non-trivial close neighbors, or long-range contacts of a residue, as a function of its “topohydrophobic” index deduced from multiple sequence alignments and of the secondary structure in which it is embedded. The “topohydrophobic” index is calculated using a twoclass distribution of amino acids, based on their mean atom depths. From a large set of structural alignments processed from the FSSP database, we selected 1485 structural sub-families including at least 8 members, with accurate alignments and limited redundancy. We show that residues within helices, even when deeply buried, have few non-trivial neighbors (0–2, whereas β-strand residues clearly exhibit a multimodal behavior, dominated by the local geometry of the tetrahedron (3 non-trivial close neighbors associated with one tetrahedron; 6 with two tetrahedra. This observed behavior allows the distinction, from sequence profiles, between edge and central β-strands within β-sheets. Useful topological constraints on the immediate neighborhood of an amino acid, but also on its correlated solvent accessibility, can thus be derived using this approach, from the simple knowledge of multiple sequence alignments.

  20. Phylogenetic group- and species-specific oligonucleotide probes for single-cell detection of lactic acid bacteria in oral biofilms

    Thurnheer Thomas

    2011-01-01

    Full Text Available Abstract Background The purpose of this study was to design and evaluate fluorescent in situ hybridization (FISH probes for the single-cell detection and enumeration of lactic acid bacteria, in particular organisms belonging to the major phylogenetic groups and species of oral lactobacilli and to Abiotrophia/Granulicatella. Results As lactobacilli are known for notorious resistance to probe penetration, probe-specific assay protocols were experimentally developed to provide maximum cell wall permeability, probe accessibility, hybridization stringency, and fluorescence intensity. The new assays were then applied in a pilot study to three biofilm samples harvested from variably demineralized bovine enamel discs that had been carried in situ for 10 days by different volunteers. Best probe penetration and fluorescent labeling of reference strains were obtained after combined lysozyme and achromopeptidase treatment followed by exposure to lipase. Hybridization stringency had to be established strictly for each probe. Thereafter all probes showed the expected specificity with reference strains and labeled the anticipated morphotypes in dental plaques. Applied to in situ grown biofilms the set of probes detected only Lactobacillus fermentum and bacteria of the Lactobacillus casei group. The most cariogenic biofilm contained two orders of magnitude higher L. fermentum cell numbers than the other biofilms. Abiotrophia/Granulicatella and streptococci from the mitis group were found in all samples at high levels, whereas Streptococcus mutans was detected in only one sample in very low numbers. Conclusions Application of these new group- and species-specific FISH probes to oral biofilm-forming lactic acid bacteria will allow a clearer understanding of the supragingival biome, its spatial architecture and of structure-function relationships implicated during plaque homeostasis and caries development. The probes should prove of value far beyond the field of

  1. Lactic acid bacteria and their antimicrobial peptides : Induction,detection,partial characterization, and potential applications

    Abbas Hilmi, Hanan

    2010-01-01

    Bacteriocin-producing lactic acid bacteria and their isolated peptide bacteriocins are of value to control pathogens and spoiling microorganisms in foods and feed. Nisin is the only bacteriocin that is commonly accepted as a food preservative and has a broad spectrum of activity against Gram-positive organisms including spore forming bacteria. In this study nisin induction was studied from two perspectives, induction from inside of the cell and selection of nisin inducible strains with increa...

  2. Detection of irradiation history for health foods. Calcium salt of organic acid and its basic ingredient

    Calcium carbonate and calcium salt of organic acid are well-known food additives used for the improvement of the shelf life and eating quality of health food. Calcium carbonate is a precursor in the synthesis of calcium salts of organic acid. Certain calcium carbonates made of natural limestone mined from very old stratum have silicate minerals exposed to a low level of natural radiation over a long period of time and food additives derived from calcium carbonates contained of such silicate minerals are possible to classify as irradiated foods by PSL and TL analysis in spite of non-irradiation. The study of calcium carbonates and calcium salts of organic acid obtained from different producers were allow to provided appropriate decisions by using the information of both the TL response (Glow1 peak temperature and TL ratio) and PSL ratio. ESR measurements of radicals in such food additives caused by gamma- irradiation were effective tool for correctly determining for irradiation history of those because the measurements were not affected by silicate minerals contained in those. (author)

  3. Electrochemical detection of nanomolar dopamine in the presence of neurophysiological concentration of ascorbic acid and uric acid using charge-coated carbon nanotubes via facile and green preparation.

    Oh, Jeong-Wook; Yoon, Yeo Woon; Heo, Jihye; Yu, Joonhee; Kim, Hasuck; Kim, Tae Hyun

    2016-01-15

    Negatively charged multi-walled carbon nanotubes (MWCNTs) were prepared using simple sonication technique with non-toxic citric acid (CA) for the electrochemical detection of dopamine (DA). CA/MWCNTs were placed on glassy carbon (GC) electrodes by drop-casting method and then electrochemical determinations of DA were performed in the presence of highly concentrated ascorbic acid (AA). For the comparison of the charge effect on MWCNTs surface, positively charged polyethyleneimine (PEI)/MWCNT/GC electrode and pristine MWCNT/GC electrode were also prepared. Contrary to conventional GC electrode, all three types of MWCNT modified electrodes (CA/MWCNT/GC, PEI/MWCNT/GC, and pristine MWCNT/GC) can discriminate ~μM of DA from 1mM AA using cyclic voltammetry (CV) and differential pulse voltammetry (DPV) due to the inherent electrocatalytic effect of MWCNTs. Compared to positively charged PEI/MWCNT/GC and pristine MWCNT/GC electrodes, negatively charged CA/MWCNT/GC electrode remarkably enhanced the electrochemical sensitivity and selectivity of DA, showing the linear relationship between DPV signal and DA concentration in the range of 10-1000nM even in the presence of ~10(5) times concentrated AA, which is attributed to the synergistic effect of the electrostatic interaction between cationic DA molecules and negatively charged MWCNTs and the inherent electrocatalytic property of MWCNT. As a result, the limit of detection (LOD) of DA for CA/MWCNT/GC electrode was 4.2nM, which is 5.2 and 16.5 times better than those for MWCNT/GC electrode and PEI/MWCNT/GC electrode even in the presence of 1mM AA. This LOD value for DA at CA/MWCNT/GC electrode is one of the lowest values compared to the previous reports and is low enough for the early diagnosis of neurological disorder in the presence of physiological AA concentration (~0.5mM). In addition, the high selectivity and sensitivity of DA at CA/MWCNT/GC electrode were well kept even in the presence of both 1mM AA and 10μM uric acid

  4. Proficient Detection of Multi-Drug-Resistant Mycobacterium tuberculosis by Padlock Probes and Lateral Flow Nucleic Acid Biosensors.

    Pavankumar, Asalapuram R; Engström, Anna; Liu, Jie; Herthnek, David; Nilsson, Mats

    2016-04-19

    Tuberculosis is a major communicable disease. Its causative agent, Mycobacterium tuberculosis, becomes resistant to antibiotics by acquisition of point mutations in the chromosome. Multi-drug-resistant tuberculosis (MDR-TB) is an increasing public health threat, and prompt detection of such strains is of critical importance. As rolling circle amplification of padlock probes can be used to robustly distinguish single-nucleotide variants, we combined this technique with a sensitive lateral flow nucleic acid biosensor to develop a rapid molecular diagnostic test for MDR-TB. A proof-of-concept test was established for detection of the most common mutations [rpoB 531 (TCG/TTG) and katG 315 (AGC/ACC)] causing MDR-TB and verification of loss of the respective wild type. The molecular diagnostic test produces visual signals corresponding to the respective genotypes on lateral flow strips in approximately 75 min. By detecting only two mutations, the test can detect about 60% of all MDR-TB cases. The padlock probe-lateral flow (PLP-LF) test is the first of its kind and can ideally be performed at resource-limited clinical laboratories. Rapid information about the drug-susceptibility pattern can assist clinicians to choose suitable treatment regimens and take appropriate infection control actions rather than prescribing empirical treatment, thereby helping to control the spread of MDR-TB in the community. PMID:26985774

  5. The quality control of glycoprotein folding in the endoplasmic reticulum, a trip from trypanosomes to mammals

    A.J. Parodi

    1998-05-01

    Full Text Available The present review deals with the stages of synthesis and processing of asparagine-linked oligosaccharides occurring in the lumen of the endoplasmic reticulum and their relationship to the acquisition by glycoproteins of their proper tertiary structures. Special emphasis is placed on reactions taking place in trypanosomatid protozoa since their study has allowed the detection of the transient glucosylation of glycoproteins catalyzed by UDP-Glc:glycoprotein glucosyltransferase and glucosidase II. The former enzyme has the unique property of covalently tagging improperly folded conformations by catalyzing the formation of protein-linked Glc1Man7GlcNAc2, Glc1Man8GlcNac2 and Glc1Man9GlcNAc2 from the unglucosylated proteins. Glucosyltransferase is a soluble protein of the endoplasmic reticulum that recognizes protein domains exposed in denatured but not in native conformations (probably hydrophobic amino acids and the innermost N-acetylglucosamine unit that is hidden from macromolecular probes in most native glycoproteins. In vivo, the glucose units are removed by glucosidase II. The influence of oligosaccharides in glycoprotein folding is reviewed as well as the participation of endoplasmic reticulum chaperones (calnexin and calreticulin that recognize monoglucosylated species in the same process. A model for the quality control of glycoprotein folding in the endoplasmic reticulum, i.e., the mechanism by which cells recognize the tertiary structure of glycoproteins and only allow transit to the Golgi apparatus of properly folded species, is discussed. The main elements of this control are calnexin and calreticulin as retaining components, the UDP-Glc:glycoprotein glucosyltransferase as a sensor of tertiary structures and glucosidase II as the releasing agent.

  6. Detection of toxins in single molecule level using deoxyribonucleic acid aptamers

    Toxins in foodstuffs are always a threat to food safety Among many toxins related to food, ricin (category B toxin) from castor beans has been mentioned in some poisoning cases happened. Atomic Force Microscopy (AFM) is a widely used nanotechnology to detect biospecies in vitro and in situ. The AFM...

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

    Ajima Muangchuen

    2014-08-01

    Full Text Available 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.

  8. A New Heuristic Algorithm for Protein Folding in the HP Model.

    Traykov, Metodi; Angelov, Slav; Yanev, Nicola

    2016-08-01

    This article presents an efficient heuristic for protein folding. The protein folding problem is to predict the compact three-dimensional structure of a protein based on its amino acid sequence. The focus is on an original integer programming model derived from a platform used for Contact Map Overlap problem. PMID:27153764

  9. Multivariate detection limits of on-line NIR model for extraction process of chlorogenic acid from Lonicera japonica.

    Wu, Zhisheng; Sui, Chenglin; Xu, Bing; Ai, Lu; Ma, Qun; Shi, Xinyuan; Qiao, Yanjiang

    2013-04-15

    A methodology is proposed to estimate the multivariate detection limits (MDL) of on-line near-infrared (NIR) model in Chinese Herbal Medicines (CHM) system. In this paper, Lonicera japonica was used as an example, and its extraction process was monitored by on-line NIR spectroscopy. Spectra of on-line NIR could be collected by two fiber optic probes designed to transmit NIR radiation by a 2mm-flange. High performance liquid chromatography (HPLC) was used as a reference method to determine the content of chlorogenic acid in the extract solution. Multivariate calibration models were carried out including partial least squares regression (PLS) and interval partial least-squares (iPLS). The result showed improvement of model performance: compared with PLS model, the root mean square errors of prediction (RMSEP) of iPLS model decreased from 0.111mg to 0.068mg, and the R(2) parameter increased from 0.9434 to 0.9801. Furthermore, MDL values were determined by a multivariate method using the type of errors and concentration ranges. The MDL of iPLS model was about 14ppm, which confirmed that on-line NIR spectroscopy had the ability to detect trace amounts of chlorogenic acid in L. japonica. As a result, the application of on-line NIR spectroscopy for monitoring extraction process in CHM could be very encouraging and reliable. PMID:23376723

  10. Colorimetric detection of Bi (III) in water and drug samples using pyridine-2,6-dicarboxylic acid modified silver nanoparticles

    Mohammadi, Somayeh; Khayatian, Gholamreza

    2015-09-01

    A new selective, simple, fast and sensitive method is developed for sensing assay of Bi (III) using pyridine-2,6-dicarboxylic acid or dipicolinic acid (DPA) modified silver nanoparticles (DPA-AgNPs). Silver nanoparticles (AgNPs) were synthesized by reducing silver nitrate (AgNO3) with sodium borohydride (NaBH4) in the presence of DPA. Bismuth detection is based on color change of nanoparticle solution from yellow to red that is induced in the presence of Bi (III). Aggregation of DPA-AgNPs has been confirmed with UV-vis absorption spectra and transmission electron microscopy (TEM) images. Under the optimized conditions, a good linear relationship (correlation coefficient r = 0.995) is obtained between the absorbance ratio (A525/A390) and the concentration of Bi (III) in the 0.40-8.00 μM range. This colorimetric probe allows Bi (III) to be rapidly quantified with a 0.01 μM limit of detection. The present method successfully applied to determine bismuth in real water and drug samples. Recoveries of water samples were in the range of 91.2-99.6%.

  11. Enzyme-based microfluidic chip coupled to graphene electrodes for the detection of D-amino acid enantiomer-biomarkers.

    Batalla, Pilar; Martín, Aída; López, Miguel Ángel; González, María Cristina; Escarpa, Alberto

    2015-01-01

    An electrochemical microfluidic strategy for the separation and enantiomeric detection of D-methionine (D-Met) and D-leucine (D-Leu) is presented. These D-amino acids (D-AAs) act as biomarkers involved in relevant diseases caused by Vibrio cholerae. On a single layout microfluidic chip (MC), highly compatible with extremely low biological sample consumption, the strategy allowed the controlled microfluidic D-AA separation and the specific reaction between D-amino acid oxidase (DAAO) and each D-AA biomarker avoiding the use of additives (i.e., cyclodextrins) for enantiomeric separation as well as any covalent immobilization of the enzyme into the wall channels or on the electrode surface such as in the biosensor-based approaches. Hybrid polymer/graphene-based electrodes were end-channel coupled to the microfluidic system to improve the analytical performance. D-Met and D-Leu were successfully detected becoming this proof-of-the-concept a promising principle for the development of point-of-care (POC) devices for in situ screening of V. cholerae related diseases. PMID:25870911

  12. Molecular assays for the detection of prostate tumor derived nucleic acids in peripheral blood

    Kinnunen Martin

    2010-07-01

    Full Text Available Abstract Background Prostate cancer is the second leading cause of cancer mortality in American men. Although serum PSA testing is widely used for early detection, more specific prognostic tests are needed to guide treatment decisions. Recently, the enumeration of circulating prostate epithelial cells has been shown to correlate with disease recurrence and metastasis following definitive treatment. The purpose of our study was to investigate an immunomagnetic fractionation procedure to enrich circulating prostate tumor cells (CTCs from peripheral blood specimens, and to apply amplified molecular assays for the detection of prostate-specific markers (PSA, PCA3 and TMPRSS2:ERG gene fusion mRNAs. Results As few as five prostate cancer cells were detected per 5 mL of whole blood in model system experiments using anti-EpCAM magnetic particles alone or in combination with anti-PSMA magnetic particles. In our experiments, anti-EpCAM magnetic particles alone exhibited equivalent or better analytical performance with patient samples compared to a combination of anti-EpCAM + anti-PSMA magnetic particles. Up to 39% of men with advanced prostate cancer tested positive with one or more of the molecular assays tested, whereas control samples from men with benign prostate hyperplasia gave consistently negative results as expected. Interestingly, for the vast majority of men who tested positive for PSA mRNA following CTC enrichment, their matched plasma samples also tested positive, although CTC enrichment gave higher overall mRNA copy numbers. Conclusion CTCs were successfully enriched and detected in men with advanced prostate cancer using an immunomagnetic enrichment procedure coupled with amplified molecular assays for PSA, PCA3, and TMPRSS2:ERG gene fusion mRNAs. Our results indicate that men who test positive following CTC enrichment also exhibit higher detectable levels of non-cellular, circulating prostate-specific mRNAs.

  13. A high-resolution mitochondria-targeting ratiometric fluorescent probe for detection of the endogenous hypochlorous acid

    Zhou, Liyi; Lu, Dan-Qing; Wang, Qianqian; Hu, Shunqin; Wang, Haifei; Sun, Hongyan; Zhang, Xiaobing

    2016-09-01

    Hypochlorite anion, one of the biologically important reactive oxygen species, plays an essential role in diverse normal biochemical functions and abnormal pathological processes. Herein, an efficient high-resolution mitochondria-targeting ratiometric fluorescent probe for hypochlorous acid detection has been designed, synthesized and characterized. It is easily synthesized by the condensation reaction (Cdbnd C) of a 2-(2-hydroxyphenyl) quinazolin-4(3H)-one fluorophore and a cyanine group (mitochondria-targeting), which made the whole molecular a large Stokes shift (210 nm) and the two well-resolved emission peaks separated by 140 nm. As a result, it is considered as a good candidate for high resolution hypochlorous acid imaging in live cells. The ratiometric fluorescent probe exhibited outstanding features of high sensitivity, high selectivity, rapid response time (within 50 s), and excellent mitochondria-targeting ability. Moreover, the probe can also be successfully applied to imaging endogenously hypochlorous acid in the mitochondria of living cells with low cytotoxicity, and high resolution.

  14. A high-resolution mitochondria-targeting ratiometric fluorescent probe for detection of the endogenous hypochlorous acid.

    Zhou, Liyi; Lu, Dan-Qing; Wang, Qianqian; Hu, Shunqin; Wang, Haifei; Sun, Hongyan; Zhang, Xiaobing

    2016-09-01

    Hypochlorite anion, one of the biologically important reactive oxygen species, plays an essential role in diverse normal biochemical functions and abnormal pathological processes. Herein, an efficient high-resolution mitochondria-targeting ratiometric fluorescent probe for hypochlorous acid detection has been designed, synthesized and characterized. It is easily synthesized by the condensation reaction (CC) of a 2-(2-hydroxyphenyl) quinazolin-4(3H)-one fluorophore and a cyanine group (mitochondria-targeting), which made the whole molecular a large Stokes shift (210nm) and the two well-resolved emission peaks separated by 140nm. As a result, it is considered as a good candidate for high resolution hypochlorous acid imaging in live cells. The ratiometric fluorescent probe exhibited outstanding features of high sensitivity, high selectivity, rapid response time (within 50s), and excellent mitochondria-targeting ability. Moreover, the probe can also be successfully applied to imaging endogenously hypochlorous acid in the mitochondria of living cells with low cytotoxicity, and high resolution. PMID:27236136

  15. Protein Collapse is Encoded in the Folded State Architecture

    Samanta, Himadri S; Hinczewski, Michael; Hori, Naoto; Chakrabarti, Shaon; Thirumalai, D

    2016-01-01

    Natural protein sequences that self-assemble to form globular structures are compact with high packing densities in the folded states. It is known that proteins unfold upon addition of denaturants, adopting random coil structures. The dependence of the radii of gyration on protein size in the folded and unfolded states obeys the same scaling laws as synthetic polymers. Thus, one might surmise that the mechanism of collapse in proteins and polymers ought to be similar. However, because the number of amino acids in single domain proteins is not significantly greater than about two hundred, it has not been resolved if the unfolded states of proteins are compact under conditions that favor the folded states - a problem at the heart of how proteins fold. By adopting a theory used to derive polymer-scaling laws, we find that the propensity for the unfolded state of a protein to be compact is universal and is encoded in the contact map of the folded state. Remarkably, analysis of over 2000 proteins shows that protei...

  16. k-fold coloring of planar graphs

    2010-01-01

    A k-fold n-coloring of G is a mapping φ: V (G) → Zk(n) where Zk(n) is the collection of all ksubsets of {1,2,...,n} such that φ(u) ∩φ(v) = φ if uv ∈ E(G).If G has a k-fold n-coloring,i.e.,G is k-fold n-colorable.Let the smallest integer n such that G is k-fold n-colorable be the k-th chromatic number,denoted by χk(G).In this paper,we show that any outerplanar graph is k-fold 2k-colorable or k-fold χk(C*)-colorable,where C* is a shortest odd cycle of G.Moreover,we investigate that every planar graph with odd girth at least 10k-9(k 3) can be k-fold (2k + 1)-colorable.

  17. Identification and Antimicrobial Activity Detection of Lactic Acid Bacteria Isolated from Corn Stover Silage

    Li, Dongxia; Ni, Kuikui; Pang, Huili; Wang, Yanping; Cai, Yimin; Jin, Qingsheng

    2015-01-01

    A total of 59 lactic acid bacteria (LAB) strains were isolated from corn stover silage. According to phenotypic and chemotaxonomic characteristics, 16S ribosomal DNA (rDNA) sequences and recA gene polymerase chain reaction amplification, these LAB isolates were identified as five species: Lactobacillus (L.) plantarum subsp. plantarum, Pediococcus pentosaceus, Enterococcus mundtii, Weissella cibaria and Leuconostoc pseudomesenteroides, respectively. Those strains were also screened for antimicrobial activity using a dual-culture agar plate assay. Based on excluding the effects of organic acids and hydrogen peroxide, two L. plantarum subsp. plantarum strains ZZU 203 and 204, which strongly inhibited Salmonella enterica ATCC 43971T, Micrococcus luteus ATCC 4698T and Escherichia coli ATCC 11775T were selected for further research on sensitivity of the antimicrobial substance to heat, pH and protease. Cell-free culture supernatants of the two strains exhibited strong heat stability (60 min at 100°C), but the antimicrobial activity was eliminated after treatment at 121°C for 15 min. The antimicrobial substance remained active under acidic condition (pH 2.0 to 6.0), but became inactive under neutral and alkaline condition (pH 7.0 to 9.0). In addition, the antimicrobial activities of these two strains decreased remarkably after digestion by protease K. These results preliminarily suggest that the desirable antimicrobial activity of strains ZZU 203 and 204 is the result of the production of a bacteriocin-like substance, and these two strains with antimicrobial activity could be used as silage additives to inhibit proliferation of unwanted microorganism during ensiling and preserve nutrients of silage. The nature of the antimicrobial substances is being investigated in our laboratory. PMID:25924957

  18. Detection of Cryptosporidium sp infection by PCR and modified acid fast staining from potassium dichromate preserved stool

    Agnes Kurniawan

    2009-09-01

    Full Text Available Aim To identify the frequency of Cryptosporidium infection in children below 3 years old by examining concentrated long term preserved stool using PCR detection of 18S rRNA gene and compared with modified acid fast staining technique.Methods Hundred eighty eight stools from children ≤ 3 years old were stored for 13 months in 2.5% K2Cr2O7 solution at 40C. Cryptosporidium oocysts were isolated by water-ether concentration technique. The concentrates were smeared onto object glass and stained with modified acid fast staining, and the rest of the concentrates were DNA extracted by freezing and thawing cycles and proteinase K digestion, then direct PCR was done to detect 18S rRNA gene.Result The proportion of positive stools for Cryptosporidium sp by acid fast staining from concentrated stools and 18S rRNA PCR were 4.8% and 34.6% respectively, which showed statistically significant difference.Conclusion The frequency of Cryptosporidium infection among children ≤ 3 years old was very high and stool storage in K2Cr2O7 for 13 months did not affect the PCR result. High prevalence of Cryptosporidium infection indicated high transmission in that area and the potential to be transmitted to other individuals such as the immunocompromised. (Med J Indones 2009;18:147-52Key words: 18S rRNA, cryptosporidiosis

  19. Selecting fast folding proteins by their rate of convergence

    Gridnev, Dmitry K; Garcia, Martin E

    2012-01-01

    We propose a general method for predicting potentially good folders from a given number of amino acid sequences. Our approach is based on the calculation of the rate of convergence of each amino acid chain towards the native structure using only the very initial parts of the dynamical trajectories. It does not require any preliminary knowledge of the native state and can be applied to different kinds of models, including atomistic descriptions. We tested the method within both the lattice and off-lattice model frameworks and obtained several so far unknown good folders. The unbiased algorithm also allows to determine the optimal folding temperature and takes by 3--4 orders of magnitude less time steps than those needed to compute folding times.

  20. Detecting Early Biomechanical Effects of Zoledronic Acid on Femurs of Osteoporotic Female Rats

    Evandro Pereira Palacio

    2012-01-01

    Full Text Available Aim. To investigate the biomechanical effects of zoledronic acid (ZA on femurs of female osteoporotic rats after follow-up periods of 9 and 12 months. Methods. Eighty female Wistar rats were prospectively assessed. At 60 days of age, the animals were randomly divided into two groups: bilateral oophorectomy (O (n=40 and sham surgery (S (n=40. At 90 days of age, groups O and S were randomly subdivided into four groups, according to whether 0.1 mg/kg of ZA or distilled water (DW was intraperitoneally administered: OZA (n=20, ODW (n=20, SZA (n=20, and SDW (n=20. The animals were sacrificed at 9 and 12 months after the administration of the substances, and then their right femurs were removed and analyzed biomechanically. Axial compression tests that focused on determining the maximum load (N, yield point (N, and stiffness coefficient (N/mm of the proximal femur were performed in the biomechanical study. Results. ZA significantly increased the maximum load and yield point, reducing the stiffness coefficient concerning the oophorectomy status and follow-up period. Conclusion. Zoledronic acid, at a dose of 0.1 mg/kg, significantly increased the maximum loads and yield points and reduced the stiffness coefficients in the femurs of female rats with osteoporosis caused by bilateral oophorectomy.

  1. A highly sensitive dual-readout assay based on gold nanoclusters for folic acid detection

    We describe a sensitive fluorometric and colorimetric dual-readout probe for folic acid (FA). It is based on the use of the gold nanoclusters (AuNCs) and cysteamine–modified gold nanoparticles (cyst-AuNPs). The bovine serum albumin stabilized AuNCs exhibit strong fluorescence emission at 652 nm. Upon addition of cyst-AuNPs, the fluorescence intensity of the AuNCs showed dramatic decrease due to the surface plasmon enhanced energy transfer process. This is due to an FA-induced aggregation of the cyst-AuNPs which shifts the absorption peaks from 530 to 670 nm. Thus, the surface plasmon enhanced energy transfer between cyst-AuNPs and AuNCs is weakened and the fluorescence intensity of AuNCs is recovered. The fluorescence intensity of the AuNCs/cyst-AuNPs system is proportional to the concentration of FA in the range from 0.11 to 2.27 μmol L−1. The dual-readout probe reported here was successfully applied to the determination of FA in spiked serum samples and folic acid tablets. (author)

  2. Detection of hyaluronidase activity using fluorescein labeled hyaluronic acid and fluorescence correlation spectroscopy

    Rich, Ryan M.; Mummert, Mark; Foldes-Papp, Zeno; Gryczynski, Zygmunt; Borejdo, Julian; Gryczynski, Ignacy; Fudala, Rafal

    2012-01-01

    The over-expression of hyaluronidase has been observed in many types of cancer, suggesting that it may have utility for diagnosis. Here we present a technique for the detection of hyaluronidase using Fluorescence Correlation Spectroscopy (FCS). Hyaluronan macromolecules (HAs) have been heavily labeled with fluorescein amine resulting in strong self-quenching. In the presence of hyaluronidase, HA is cleaved into smaller, fluorescein-labeled fragments and the self-quenching is released. Such cl...

  3. Amino acid precursors for the detection of transketolase activity in Escherichia coli auxotrophs.

    Simon, Grégory; Bouzon, Madeleine; Charmantray, Franck; Hélaine, Virgil; Légeret, Bertrand; Marlière, Philippe; Hecquet, Laurence

    2009-07-15

    Probes were developed for the in vivo detection of transketolase activity by the use of a complementation assay in Escherichia coli auxotrophs They combine the d-threo ketose moiety recognised by transketolase and the side chain of leucine or methionine. These compounds were donor substrates of yeast transketolase leading to the release of the corresponding alpha-hydroxyaldehydes which could be converted in E. coli by a cascade of reactions into leucine or methionine required for cellular growth. PMID:19535247

  4. Peroxynitrous-acid-induced chemiluminescence detection of nitrite based on Microfluidic chip.

    Wu, Jing; Wang, Xiong; Lin, Yitong; Zheng, Yongzan; Lin, Jin-Ming

    2016-07-01

    A chemiluminescent method for nitrite detection was developed on microfluidic chip. Carbon dots-NaNO2(-) acidified H2O2 system was adopted. Chemiluminescence (CL) spectrum of this system was detected. The radiative recombination of hole-injected and electron-injected carbon dots explained their CL property. Spiral microchannels were designed on the microfluidic chip to allow enough reaction time for the carbon dots-NaNO2-acidified H2O2 system. Carbon dots and NaNO2 were premixed in the branch microchannel, then, the mixture reacted with acidified H2O2 in spiral microchannels. Concentrations of H2SO4 and H2O2, dilution ratio of carbon dots in H2O and flow rate were optimized to obtain the best CL signals. The approach presented satisfactory linear relationship between NaNO2 concentration and CL intensity. The tolerance of metal ions in determination of 1×10(-5)M nitrite was analyzed. The nitrites in water and beverage samples were successfully analyzed on the microfluidic chip with good repeatability. The data were well accordance with the results obtained from GB 5009.33(-) 2010. This microfluidic CL detection method is believed to be a simple, automatic and agent-save approach for inorganic ion analysis. PMID:27154650

  5. Label-free nucleic acids detection based on DNA templated silver nanoclusters fluorescent probe.

    Zhao, Haiyan; Wang, Lei; Zhu, Jing; Wei, Haiping; Jiang, Wei

    2015-06-01

    Based on DNA templated Ag NCs (DNA/Ag NCs) fluorescent probe, a label-free fluorescent method was developed for the detection of clinical significant DNA fragments from human immunodeficiency virus type 1 (HIV-1) DNA. Firstly, a hairpin probe, containing target DNA recognition sequence and guanine-rich sequence, was designed to hybridize with the target DNA and form a blunt 3'-terminus DNA duplex. Then, exonuclease III (Exo III) was employed to stepwise hydrolyze the mononucleotides from formed blunt 3'-terminus DNA duplex, releasing the target DNA and guanine-rich sequence. Finally, DNA/Ag NCs fluorescent probe was introduced to hybridize with the guanine-rich sequence, leading to an enhanced fluorescence signal for detection. The proposed method could detect as low as 2.9×10(-10) mol L(-1) HIV-1 DNA and exhibited excellent selectivity against mismatched target DNA. Furthermore, the method possessed perfect recoveries in cells lysate and human serum, showing potential to be used in biological samples. PMID:25863386

  6. The role of low-level magnification in visual inspection with acetic acid for the early detection of cervical neoplasia.

    Sankaranarayanan, Rengaswamy; Shastri, Surendra S; Basu, Parthasarathi; Mahé, Cédric; Mandal, Ranajit; Amin, Geethanjali; Roy, Chinmayi; Muwonge, Richard; Goswami, Smriti; Das, Pradip; Chinoy, Roshini; Frappart, Lucien; Patil, Sharmila; Choudhury, Devjani; Mukherjee, Titha; Dinshaw, Ketayun

    2004-01-01

    Several studies have investigated the accuracy of naked eye visual inspection with acetic acid (VIA) in the early detection of cervical neoplasia. It is not clear whether low-level (2-4x) magnification (VIAM) can improve the sensitivity and specificity of VIA. The accuracy of both VIA and VIAM, provided by independent health workers, were evaluated in three cross-sectional studies involving 18,675 women aged 25-65 years in Kolkata and Mumbai in India. All screened women were investigated with colposcopy and biopsies were obtained based on colposcopy findings. The final disease status was based on the reference standard of histology (if biopsies had been taken) or colposcopy. Data from the studies were pooled to calculate the test characteristics for the detection of high-grade squamous intraepithelial lesions (HSIL). 14.1% and 14.2% were positive on testing with VIA and VIAM respectively. Two hundred twenty-nine were diagnosed with HSIL and 68 with invasive cancer. The pooled sensitivity, specificity, positive and negative predictive values for VIA in detecting high-grade squamous intraepithelial lesions (HSIL) were 60.3% (95% CI: 53.6-66.7), 86.8% (95% CI: 86.3-87.3), 5.9% (95% CI: 5.0-7.0), and 99.4% (95% CI: 99.2-99.5), respectively. The values were 64.2% (95% CI: 57.6-70.4), 86.8% (95% CI: 86.2-87.3), 6.3% (95% CI: 5.3-7.3) and 99.4% (95% CI: 99.3-99.6), respectively, for VIAM. Low-level magnification did not improve the test performance of naked eye visualization of acetic acid impregnated uterine cervix. PMID:15542259

  7. Stress and strain evolution of folding rocks

    Llorens, Maria-Gema; Griera, Albert; Bons, Paul; Gomez-Rivas, Enrique; Weikusat, Ilka

    2015-04-01

    One of the main objectives of structural geology is to unravel rock deformation histories. Fold shapes can be used to estimate the orientation and amount of strain associated with folding. However, much more information on rheology and kinematics can potentially be extracted from fold geometries (Llorens et al., 2013a). We can study the development of folds, quantify the relationships between the different parameters that determine their geometries and estimate their mechanical evolution. This approach allows us to better understand and predict not only rock but also ice deformation. One of the main parameters in fold development is the viscosity contrast between the folding layer and the matrix in which it is embedded (m), since it determines the initial fold wavelength and the amplification rate of the developing folds. Moreover, non-linear viscous rheology influences fold geometry too (Llorens et al., 2013b). We present a series of 2-dimensional simulations of folding of viscous single layers in pure and simple shear. We vary different parameters in order to compare and determine their influence on the resulting fold patterns and the associated mechanical response of the material. To perform these simulations we use the software platform ELLE (www.elle.ws) with the non-linear viscous finite element code BASIL. The results show that layers thicken at the beginning of deformation in all simulations, and visible folds start earlier or later depending on the viscosity contrast. When folds start to nucleate the layer maximum shear strain decreases, moving away from the theoretical trend for homogeneous strain (no folding). This allows the accurate determination of the onset of folding. Maximum deviatoric stresses are higher in power-law than in linear-viscosity materials, and it is initially double in pure shear than in simple shear conditions. Therefore, folding a competent layer requires less work in simple than in pure shear. The maximum deviatoric stress

  8. Viscoelastic properties of the false vocal fold

    Chan, Roger W.

    2001-05-01

    The biomechanical properties of vocal fold tissues have been the focus of many previous studies, as vocal fold viscoelasticity critically dictates the acoustics and biomechanics of phonation. However, not much is known about the viscoelastic response of the ventricular fold or false vocal fold. It has been shown both clinically and in computer simulations that the false vocal fold may contribute significantly to the aerodynamics and sound generation processes of human voice production, with or without flow-induced oscillation of the false fold. To better understand the potential role of the false fold in phonation, this paper reports some preliminary measurements on the linear and nonlinear viscoelastic behavior of false vocal fold tissues. Linear viscoelastic shear properties of human false fold tissue samples were measured by a high-frequency controlled-strain rheometer as a function of frequency, and passive uniaxial tensile stress-strain response of the tissue samples was measured by a muscle lever system as a function of strain and loading rate. Elastic moduli (Young's modulus and shear modulus) of the false fold tissues were calculated from the measured data. [Work supported by NIH.

  9. Kinetic partitioning mechanism of HDV ribozyme folding

    Chen, Jiawen; Gong, Sha; Wang, Yujie; Zhang, Wenbing, E-mail: wbzhang@whu.edu.cn [Department of Physics, Wuhan University, Wuhan, Hubei 430072 (China)

    2014-01-14

    RNA folding kinetics is directly tied to RNA biological functions. We introduce here a new approach for predicting the folding kinetics of RNA secondary structure with pseudoknots. This approach is based on our previous established helix-based method for predicting the folding kinetics of RNA secondary structure. In this approach, the transition rates for an elementary step: (1) formation, (2) disruption of a helix stem, and (3) helix formation with concomitant partial melting of an incompatible helix, are calculated with the free energy landscape. The folding kinetics of the Hepatitis delta virus (HDV) ribozyme and the mutated sequences are studied with this method. The folding pathways are identified by recursive searching the states with high net flux-in(out) population starting from the native state. The theory results are in good agreement with that of the experiments. The results indicate that the bi-phasic folding kinetics for the wt HDV sequence is ascribed to the kinetic partitioning mechanism: Part of the population will quickly fold to the native state along the fast pathway, while another part of the population will fold along the slow pathway, in which the population is trapped in a non-native state. Single mutation not only changes the folding rate but also the folding pathway.

  10. Detection of hyaluronic acid and laminin in serum of patients with urinary bladder tumors

    Objective: To investigate the serum Hyaluronic Acid (HA) and laminin (LN) level and its clinical significance in bladder tumors. Methods: Serum samples from 34 patients with bladder tumor and 30 cases as control group were analyzed by radioimmunoassay. Results: The results showed that the HA and LN levels of serum in patients with bladder tumors were significantly higher than that in control group (P<0.01) before operation, and decreased significantly after operation. The serum levels of HA and LN in infiltrated tumors were higher than that in superficial tumors (P<0.05). The markers in patients with lymph node metastasis were higher than that without lymph node metastasis (P<0.01). Conclusion: HA and LN take part in the malignant biology behavior of bladder tumors. They may be the important markers in assisting diagnosis and disease monitoring

  11. Isothermal cycling and cascade signal amplification strategy for ultrasensitive colorimetric detection of nucleic acids

    We have designed a novel isothermal cascade signal-amplification strategy for ultrasensitive colorimetric determination of nucleic acids. It is based on double-cycling amplification with formation of DNAzyme via a polymerase-induced strand-displacement reaction and nicking endonuclease-assisted recycling. The assay makes use of a hairpin DNA, a short primer, KF-polymerase, and nicking endonuclease. The presence of a target DNA triggers the strand-displacement and polymerization reaction with the formation of numerous DNAzyme molecules. Upon addition of H2O2 to the resulting mixture, the H2O2 reacts with 2,2′-azino-bis (3-ethylbenzothiozoline)-6-sulfonate to form a colored product in the aid of DNAzyme, which is quantified by photometry at 415 nm. Under optimal conditions, the assay allows target DNA to be determined at concentration as low as 0.6 aM. (author)

  12. Aptamer- and nucleic acid enzyme-based systems for simultaneous detection of multiple analytes

    Lu, Yi; Liu, Juewen

    2011-11-15

    The present invention provides aptamer- and nucleic acid enzyme-based systems for simultaneously determining the presence and optionally the concentration of multiple analytes in a sample. Methods of utilizing the system and kits that include the sensor components are also provided. The system includes a first reactive polynucleotide that reacts to a first analyte; a second reactive polynucleotide that reacts to a second analyte; a third polynucleotide; a fourth polynucleotide; a first particle, coupled to the third polynucleotide; a second particle, coupled to the fourth polynucleotide; and at least one quencher, for quenching emissions of the first and second quantum dots, coupled to the first and second reactive polynucleotides. The first particle includes a quantum dot having a first emission wavelength. The second particle includes a second quantum dot having a second emission wavelength different from the first emission wavelength. The third polynucleotide and the fourth polynucleotide are different.

  13. Detection and quantification of genetically modified organisms using very short, locked nucleic acid TaqMan probes.

    Salvi, Sergio; D'Orso, Fabio; Morelli, Giorgio

    2008-06-25

    Many countries have introduced mandatory labeling requirements on foods derived from genetically modified organisms (GMOs). Real-time quantitative polymerase chain reaction (PCR) based upon the TaqMan probe chemistry has become the method mostly used to support these regulations; moreover, event-specific PCR is the preferred method in GMO detection because of its high specificity based on the flanking sequence of the exogenous integrant. The aim of this study was to evaluate the use of very short (eight-nucleotide long), locked nucleic acid (LNA) TaqMan probes in 5'-nuclease PCR assays for the detection and quantification of GMOs. Classic TaqMan and LNA TaqMan probes were compared for the analysis of the maize MON810 transgene. The performance of the two types of probes was tested on the maize endogenous reference gene hmga, the CaMV 35S promoter, and the hsp70/cryIA(b) construct as well as for the event-specific 5'-integration junction of MON810, using plasmids as standard reference molecules. The results of our study demonstrate that the LNA 5'-nuclease PCR assays represent a valid and reliable analytical system for the detection and quantification of transgenes. Application of very short LNA TaqMan probes to GMO quantification can simplify the design of 5'-nuclease assays. PMID:18494480

  14. The significance and robustness of a plasma free amino acid (PFAA) profile-based multiplex function for detecting lung cancer

    We have recently reported on the changes in plasma free amino acid (PFAA) profiles in lung cancer patients and the efficacy of a PFAA-based, multivariate discrimination index for the early detection of lung cancer. In this study, we aimed to verify the usefulness and robustness of PFAA profiling for detecting lung cancer using new test samples. Plasma samples were collected from 171 lung cancer patients and 3849 controls without apparent cancer. PFAA levels were measured by high-performance liquid chromatography (HPLC)–electrospray ionization (ESI)–mass spectrometry (MS). High reproducibility was observed for both the change in the PFAA profiles in the lung cancer patients and the discriminating performance for lung cancer patients compared to previously reported results. Furthermore, multivariate discriminating functions obtained in previous studies clearly distinguished the lung cancer patients from the controls based on the area under the receiver-operator characteristics curve (AUC of ROC = 0.731 ~ 0.806), strongly suggesting the robustness of the methodology for clinical use. Moreover, the results suggested that the combinatorial use of this classifier and tumor markers improves the clinical performance of tumor markers. These findings suggest that PFAA profiling, which involves a relatively simple plasma assay and imposes a low physical burden on subjects, has great potential for improving early detection of lung cancer

  15. Folic acid conjugated magnetic iron oxide nanoparticles for nondestructive separation and detection of ovarian cancer cells from whole blood.

    Liu, Wenting; Nie, Liju; Li, Fulai; Aguilar, Zoraida P; Xu, Hong; Xiong, Yonghua; Fu, Fen; Xu, Hengyi

    2016-01-01

    Because of the lack of early screening strategies, ovarian cancer is the most deadly cause of gynecologic malignancies. This paper describes an effective method for the separation and detection of ovarian cancer cells from female whole blood, using folic acid (FA) conjugated magnetic iron oxide nanoparticles (IO-FA nanoparticles). The IO nanoparticles were synthesized by thermal decomposition and then covalently conjugated with FA. The IO-FA nanoparticles were stably attached to the surface of ovarian cancer cells by coupling to the over-expressed folate receptor (FR), thereby making the cells magnetic. These "magnetic cells" were separated from the complex blood matrix without destruction under a magnetic field. The separation efficiency was as high as 61.3% when the abundance of spiked ovarian cancer SKOV3 cells was as low as 5 × 10(-5)%. We also successfully detected five (5) out of ten (10) metastatic ovarian cancer patients' whole blood. This study suggested the feasibility of early detecting of metastatic ovarian cancer cells, which may potentially improve the ovarian cancers patients' overall survival rate for clinical applications. PMID:26478922

  16. Use of pooled sodium acetate acetic acid formalin-preserved fecal specimens for the detection of intestinal parasites.

    Gaafar, Maha R

    2011-01-01

    This study aimed at comparing detection of intestinal parasites from single unpreserved stool sample vs. sodium acetate acetic acid formalin (SAF)-preserved pooled samples, and stained with chlorazol black dye in routine practice. Unpreserved samples were collected from 120 patients and represented as Group I. Other three SAF-preserved samples were collected from the same patients over a 6-day period and represented as Groups IIa, IIb, and IIc. The latter groups were equally subdivided into two subgroups. The first subgroup of each of the three samples was examined individually, whereas the second subgroup of each were pooled and examined as a single specimen. All groups were examined by the routine diagnostic techniques; however, in group II when the diagnosis was uncertain, the chlorazol black dye staining procedure was carried out. Results demonstrated that out of 74 patients who continued the study, 12 cases (16%) were positive in group I, compared with 29 (39%) in the subgroups examined individually, and 27 (36%) in the pooled subgroups. Therefore, pooling of preserved fecal samples is an efficient and economical procedure for the detection of parasites. Furthermore, the chlorazol black dye was simple and effective in detecting the nuclear details of different parasites. PMID:21567472

  17. On-Chip Microfluidic Components for In Situ Analysis, Separation, and Detection of Amino Acids

    Zheng, Yun; Getty, Stephanie; Dworkin, Jason; Balvin, Manuel; Kotecki, Carl

    2013-01-01

    The Astrobiology Analytical Laboratory at GSFC has identified amino acids in meteorites and returned cometary samples by using liquid chromatography-electrospray ionization time-of-flight mass spectrometry (LCMS). These organic species are key markers for life, having the property of chirality that can be used to distinguish biological from non-biological amino acids. One of the critical components in the benchtop instrument is liquid chromatography (LC) analytical column. The commercial LC analytical column is an over- 250-mm-long and 4.6-mm-diameter stainless steel tube filled with functionized microbeads as stationary phase to separate the molecular species based on their chemistry. Miniaturization of this technique for spaceflight is compelling for future payloads for landed missions targeting astrobiology objectives. A commercial liquid chromatography analytical column consists of an inert cylindrical tube filled with a stationary phase, i.e., microbeads, that has been functionalized with a targeted chemistry. When analyte is sent through the column by a pressurized carrier fluid (typically a methanol/ water mixture), compounds are separated in time due to differences in chemical interactions with the stationary phase. Different species of analyte molecules will interact more strongly with the column chemistry, and will therefore take longer to traverse the column. In this way, the column will separate molecular species based on their chemistry. A lab-on-chip liquid analysis tool was developed. The microfluidic analytical column is capable of chromatographically separating biologically relevant classes of molecules based on their chemistry. For this analytical column, fabrication, low leak rate, and stationary phase incorporation of a serpentine microchannel were demonstrated that mimic the dimensions of a commercial LC column within a 5 10 1 mm chip. The microchannel in the chip has a 75- micrometer-diameter oval-shaped cross section. The serpentine

  18. Chip-based device for parallel sorting, amplification, detection, and identification of nucleic acid subsequences

    Beer, Neil Reginald; Colston, Jr, Billy W.

    2016-08-09

    An apparatus for chip-based sorting, amplification, detection, and identification of a sample having a planar substrate. The planar substrate is divided into cells. The cells are arranged on the planar substrate in rows and columns. Electrodes are located in the cells. A micro-reactor maker produces micro-reactors containing the sample. The micro-reactor maker is positioned to deliver the micro-reactors to the planar substrate. A microprocessor is connected to the electrodes for manipulating the micro-reactors on the planar substrate. A detector is positioned to interrogate the sample contained in the micro-reactors.

  19. 4-(Dimethylamino)butyric acid labeling for electrochemiluminescence detection of biological substances by increasing sensitivity with gold nanoparticle amplification.

    Yin, Xue-Bo; Qi, Bin; Sun, Xuping; Yang, Xiurong; Wang, Erkang

    2005-06-01

    4-(Dimethylamino)butyric acid (DMBA) labeling combined with gold nanoparticle amplification for electrochemiluminescence (ECL) determination of a biological substance (bovine serum albumin (BSA) and immunoglobulin G (IgG) as models) was presented. After DMBA, an analogue of tripropylamine, was tagged on the (anti)analytes, an ECL signal related to the content of the analytes was generated when the analyte tagged with DMBA was in contact with tris(2,2'-bipyridine)ruthenium (Ru(bpy)(3)2+) solution and a potential was applied. To improve the adsorption capacity, a gold nanoparticle layer was first combined into the surface of the 2-mm-diameter gold electrode. For the determination of BSA, avidin was covalently conjugated to a self-assembled monolayer of 3-mercaptopropanoic acid on the gold nanoparticle layer. Biotinylated BSA-DMBA was then immobilized on the gold nanoparticle layer of the gold electrode via the avidin-biotin reaction. IgG was tested via a typical sandwich-type immobilization method. ECL signals were generated from the electrodes immobilized with BSA or IgG by immersing them in a 1 mmol L-1 Ru(bpy)(3)2+ solution and scanning from 0.5 to 1.3 V versus Ag/AgCl. With gold nanoparticle amplification, the ECL peak intensity was proportional to the concentration over the range 1-80 and 5-100 microg/mL for BSA and IgG consuming 50 microL of sample, respectively. A 10- and 6-fold sensitivity enhancement was obtained for BSA and IgG over their direct immobilization on an electrode using DMBA labeling. The relative standard deviations of five replicate determinations of 10 microg/mL BSA and 20 microg/mL IgG were 8.4 and 10.2%, respectively. High biocompatibility and low cost were the main advantages of the present DMBA labeling technique over the traditional Ru(bpy)(3)2+ labeling. PMID:15924384

  20. Analysis of hyaluronic acid concentration in rat vocal folds during estral and gravidic puerperal cycles Análise da concentração do ácido hialurônico nas pregas vocais de ratas durante o ciclo estral e ciclo gravídico-puerperal

    José Eduardo de Sá Pedroso

    2009-10-01

    Full Text Available Hormone plays an important role in the larynx. Among other substances, vocal folds contain hyaluronic acid, which tissue concentration may vary according to hormone action. AIM: the objective of this study is to analyze hyaluronic acid concentration in the vocal folds during estral and gravidic-puerperal cycles. MATERIALS AND METHODS: Experimental study. 40 adult rats were divided into two groups. In the first group we used 20 rats to establish the concentration of hyaluronic acid during the estral cycle and in the second group, 20 animals were submitted to the same procedure but during the gravidic-puerperal cycle. RESULTS: Variations in hyaluronic acid concentration was not observed during the estral cycle. In the gravidic puerperal cycle group, an increase in hyaluronic acid concentration was observed in the puerperal subgroup. Comparing the two groups of estral and gravidic-puerperal cycles, no difference was observed. CONCLUSIONS: In comparing all subgroups of estral and gravidic-puerperal cycles, an increase in hyaluronic acid concentration was noticed only in the puerperal phase.Os hormônios exercem importante influência sobre a laringe. A prega vocal contém, entre outras substâncias, o ácido hialurônico, cuja concentração nos tecidos pode variar com a ação dos hormônios. OBJETIVO: O objetivo deste trabalho é analisar comparativamente a concentração do ácido hialurônico nas pregas vocais de ratas durante o ciclo estral e ciclo gravídico-puerperal. FORMA DE ESTUDO: Experimental. MATERIAL E MÉTODO: Foram utilizadas 40 ratas adultas, divididas em dois grupos, no primeiro grupo utilizamos 20 ratas para determinação da concentração do ácido hialurônico no ciclo estral, no segundo grupo, também de 20 animais, foi realizado o mesmo experimento no ciclo gravídico-puerperal. RESULTADOS: No grupo do ciclo estral não observou-se variação da concentração do ácido hialurônico. No grupo do ciclo grav

  1. Chemical ionization mass spectrometry of indol-3yl-acetic acid and cis-abscisic acid: evaluation of negative ion detection and quantification of cis-abscisic acid in growing maize roots

    Mass spectra of the derivatives of indol-3yl-acetic acid and cis-abscisic acid were obtained in electron impact and chemical ionization positive ion and negative ion modes. The respective merits of methane, isobutane, and ammonia as reagent gases for structure determination and sensitive detection were compared using the methyl esters. From one to 10 fluorine atoms were attached to IAA to improve the electron-capturing properties of the molecule. The best qualitative information was obtained when using positive ion chemical ionization with methane. However, the most sensitive detection, with at least two ions per molecule, was achieved by electron impact on the IAA-HFB-ME derivative and by negative ion chemical ionization with NH3 on the ABA-methyl ester derivative. p ]Quantitative analyses of ABA in different parts of maize (Zea mays cv. LG 11) root tips were performed by the latter technique. It was found that the cap and apex contained less ABA than the physiologically older parts of the root such as the elongation zone and the more differentiated tissues. This technique was also used to show a relation between maize root growth and the endogenous ABA level of the elongation zone and root tip: there is more ABA in the slowly growing roots than in the rapidly growing ones. (author)

  2. In-capillary derivatization with o-phthalaldehyde in the presence of 3-mercaptopropionic acid for the simultaneous determination of monosodium glutamate, benzoic acid, and sorbic acid in food samples via capillary electrophoresis with ultraviolet detection.

    Aung, Hnin-Pwint; Pyell, Ute

    2016-06-01

    For the rapid simultaneous determination of monosodium glutamate (MSG), benzoic acid (BA), and sorbic acid (SA) in canned food and other processed food samples, we developed a method that combines in-capillary derivatization with separation by capillary electrophoresis. This method employs the rapid derivatization of MSG with o-phthalaldehyde (OPA) in the presence of 3-mercaptopropionic acid (3-MPA) and enables the detection of the resulting OPA-MSG derivative and of non-derivatized BA and SA at 230nm. The composition of the background electrolyte and the parameters of derivatization and separation are as follows: 25mM borax containing 5mM OPA and 6mM 3-MPA, separation voltage 25mV, injection at 30mbar for 20s, and column temperature 25°C. Because of the high reaction rate and suitably adapted effective electrophoretic mobilities, band broadening due to the derivatization reaction at the start of the separation process is kept to a minimum. The optimized method is validated with respect to LOD, LOQ, linearity, recovery, and precision. This method can be applied to real samples such as soy, fish, oyster and sweet and sour chili sauces after application of appropriate clean-up steps. Mechanisms of zone broadening and zone focusing are discussed showing the validity of the employed theoretical approach regarding the dependence of the peak shape for OPA-MSG on the concentration of MSG in the sample. PMID:27156753

  3. Hydration of the folding transition-state ensemble of a protein

    Brun, Ludovic; Isom, Daniel G.; Velu, Priya; García-Moreno, Bertrand; Royer, Catherine

    2006-01-01

    A complete description of the mechanisms of protein folding requires knowledge of the structural and physical character of the folding transition state ensembles (TSE). A key question remains, concerning the role of hydration of the hydrophobic core in determining folding mechanisms. To address this we probed the state of hydration of the TSE of staphylococcal nuclease (SNase) by examining the fluorescence-detected pressure-jump relaxation behavior of six SNase variants in which a residue in ...

  4. Protein folding guides disulfide bond formation

    Qin, Meng; Wang, Wei; Thirumalai, D.

    2015-01-01

    Anfinsen inferred the principles of protein folding by studying a protein containing four disulfide bonds in the native state. However, how protein folding drives disulfide bond formation is poorly understood despite the role such proteins play in variety of extracellular and intracellular functions. We developed a method to mimic the complex chemistry of disulfide bond formation in molecular simulations, which is used to decipher the mechanism of folding of bovine pancreatic trypsin inhibito...

  5. [Design of a medical folding fridge].

    Sun, Jianjun; Wei, Jiancang; Wu, Taihu; Meng, Xingju

    2011-07-01

    This article introduces a design of a medical folding fridge, which consists of three major components, base, folding frame and insulated cover. The base has a cooling system. The frame and cover are expanded during normal use and folded during storage or transportation. The device is compact, durable, transportable and well environmental adaptable. The system design is proved proper and the temperature inside is reliable. It is very suitable for temperature sensitive supplies stored in the medical emergency field. PMID:22097750

  6. Acquired retinal folds in the cat.

    MacMillan, A D

    1976-06-01

    Retinal folds were found in 5 cats. The apparent cause of the folding was varied: in 1 cat the folds appeared after a localized retinal detachment; in 2 cats the condition accompanied other intraocular abnormalities associated with feline infectious peritonitis; 1 cat had active keratitis, and the retinal changes were thought to have been injury related; and 1 cat, bilaterally affected, had chronic glomerulonephritis. PMID:945253

  7. Direct estimation of sialic acid in milk and milk products by fluorimetry and its application in detection of sweet whey adulteration in milk.

    Neelima; Rao, Priyanka Singh; Sharma, Rajan; Rajput, Yudhishthir S

    2012-11-01

    Sialic acid, being a biologically active compound, is recognised as an important component of milk and milk products. Almost all the sialic acid estimation protocols in milk require prior hydrolysis step to release the bound sialic acid followed by its estimation. The objective of this work was to estimate sialic acid in milk and milk products by fluorimetric assay which does not require a prior hydrolysis step thus decreasing the estimation time. The recovery of added sialic acid in milk was 91·6 to 95·8%. Sialic acid in milk was found to be dependent on cattle breed and was in the range of 1·68-3·93 g/kg (dry matter basis). The assay was further extended to detect adulteration of milk with sweet whey which is based on the detection of glycomacropeptide (GMP) bound sialic acid in adulterated milk. GMP is the C-terminal part of κ-casein which is released into the whey during cheese making. For detection of adulteration, selective precipitation of GMP was done using trichloroacetic acid (TCA). TCA concentration in milk was first raised to 5% to precipitate milk proteins, especially κ-casein, followed by raising the TCA concentration to 14% to precipitate out GMP. In the precipitates GMP bound sialic acid was estimated using fluorimetric method and the fluorescence intensity was found to be directly proportional to the level of sweet whey in adulterated milk samples. The method was found to detect the presence of 5% sweet whey in milk. PMID:23089266

  8. Nanosensors having dipicolinic acid imprinted nanoshell for Bacillus cereus spores detection

    Molecular imprinted polymers (MIPs) as a recognition element for sensors are increasingly of interest and MIP nanoclusters have started to appear in the literature. In this study, we have proposed a novel thiol ligand-capping method with polymerizable methacryloylamido-cysteine (MAC) attached to gold-silver nanoclusters, reminiscent of a self-assembled monolayer and have reconstructed surface shell by synthetic host polymers based on molecular imprinting method for recognition. In this method, methacryloylamidoantipyrine-terbium ((MAAP)2-Tb(III)) has been used as a new metal-chelating monomer via metal coordination-chelation interactions and dipicolinic acid (DPA) which is main participant of Bacillus cereus spores used as a model. Nanoshell sensors with templates give a cavity that is selective for DPA. The DPA can simultaneously chelate to Tb(III) metal ion and fit into the shape-selective cavity. Thus, the interaction between Tb(III) ion and free coordination spheres has an effect on the binding ability of the gold-silver nanoclusters nanosensor. The binding affinity of the DPA imprinted nanoclusters has been investigated by using the Langmuir and Scatchard methods, and the respective affinity constants (Kaffinity) determined were found to be 1.43 x 104 and 9.1 x 106 mol L-1.

  9. Nanosensors having dipicolinic acid imprinted nanoshell for Bacillus cereus spores detection

    Gueltekin, Aytac [Trakya University, Department of Chemistry (Turkey); Ersoez, Arzu [Anadolu University, Department of Chemistry, Faculty of Science, Yunusemre Campus (Turkey); Sarioezlue, Nalan Yilmaz [Anadolu University, Department of Biology (Turkey); Denizli, Adil [Hacettepe University, Department of Chemistry (Turkey); Say, Ridvan, E-mail: rsay@anadolu.edu.t [Anadolu University, Department of Chemistry, Faculty of Science, Yunusemre Campus (Turkey)

    2010-08-15

    Molecular imprinted polymers (MIPs) as a recognition element for sensors are increasingly of interest and MIP nanoclusters have started to appear in the literature. In this study, we have proposed a novel thiol ligand-capping method with polymerizable methacryloylamido-cysteine (MAC) attached to gold-silver nanoclusters, reminiscent of a self-assembled monolayer and have reconstructed surface shell by synthetic host polymers based on molecular imprinting method for recognition. In this method, methacryloylamidoantipyrine-terbium ((MAAP){sub 2}-Tb(III)) has been used as a new metal-chelating monomer via metal coordination-chelation interactions and dipicolinic acid (DPA) which is main participant of Bacillus cereus spores used as a model. Nanoshell sensors with templates give a cavity that is selective for DPA. The DPA can simultaneously chelate to Tb(III) metal ion and fit into the shape-selective cavity. Thus, the interaction between Tb(III) ion and free coordination spheres has an effect on the binding ability of the gold-silver nanoclusters nanosensor. The binding affinity of the DPA imprinted nanoclusters has been investigated by using the Langmuir and Scatchard methods, and the respective affinity constants (K{sub affinity}) determined were found to be 1.43 x 10{sup 4} and 9.1 x 10{sup 6} mol L{sup -1}.

  10. Protein Folding and Self-Organized Criticality

    Bajracharya, Arun; Murray, Joelle

    Proteins are known to fold into tertiary structures that determine their functionality in living organisms. However, the complex dynamics of protein folding and the way they consistently fold into the same structures is not fully understood. Self-organized criticality (SOC) has provided a framework for understanding complex systems in various systems (earthquakes, forest fires, financial markets, and epidemics) through scale invariance and the associated power law behavior. In this research, we use a simple hydrophobic-polar lattice-bound computational model to investigate self-organized criticality as a possible mechanism for generating complexity in protein folding.

  11. COS Side 2 NUV MAMA Fold Test

    Bacinski, John

    2013-10-01

    The performance of the MAMA microchannel plate can be monitored using a MAMA fold analysis procedure. The fold analysis provides a measurement of the distribution of charge cloud sizes incident upon the anode giving some measure of changes in the pulse-height distribution of the MCP and, therefore, MCP gain. This proposal executes the same steps as the COS MAMA Fold Analysis {13128} during Cycle 20.This proposal is an exact duplication of nominal COS MAMA Fold Analysis {proposal 13128, Cycle 20}. Any changes 13128 or subsequent cycle submissions should be reflected in this proposal and vice versa.

  12. Dependence of Internal Friction on Folding Mechanism

    2016-01-01

    An outstanding challenge in protein folding is understanding the origin of “internal friction” in folding dynamics, experimentally identified from the dependence of folding rates on solvent viscosity. A possible origin suggested by simulation is the crossing of local torsion barriers. However, it was unclear why internal friction varied from protein to protein or for different folding barriers of the same protein. Using all-atom simulations with variable solvent viscosity, in conjunction with transition-path sampling to obtain reaction rates and analysis via Markov state models, we are able to determine the internal friction in the folding of several peptides and miniproteins. In agreement with experiment, we find that the folding events with greatest internal friction are those that mainly involve helix formation, while hairpin formation exhibits little or no evidence of friction. Via a careful analysis of folding transition paths, we show that internal friction arises when torsion angle changes are an important part of the folding mechanism near the folding free energy barrier. These results suggest an explanation for the variation of internal friction effects from protein to protein and across the energy landscape of the same protein. PMID:25721133

  13. Direct detection of fatty acid ethyl esters using low temperature plasma (LTP) ambient ionization mass spectrometry for rapid bacterial differentiation.

    Zhang, J Isabella; Costa, Anthony B; Tao, W Andy; Cooks, R Graham

    2011-08-01

    Low temperature plasma mass spectrometry (LTP-MS) was employed to detect fatty acid ethyl esters (FAEE) from bacterial samples directly. Positive ion mode FAEE mass spectrometric profiles of sixteen different bacterial samples were obtained without extraction or other sample preparation. In the range m/z 200-300, LTP mass spectra show highly reproducible and characteristic patterns. To identify the FAEE's associated with the characteristic peaks, accurate masses were recorded in the full scan mode using an LTQ/Orbitrap instrument, and tandem mass spectrometry was performed. Data were examined by principal component analysis (PCA) to determine the degree of differentiation possible amongst different bacterial species. Gram-positive and gram-negative bacteria are readily distinguished, and 11 out of 13 Salmonella strains show distinctive patterns. Growth media effects are observed but do not interfere with species recognition based on the PCA results. PMID:21706093

  14. New high-performance liquid chromatography assay for glycosyltransferases based on derivatization with anthranilic acid and fluorescence detection.

    Anumula, Kalyan Rao

    2012-07-01

    Assays were developed using the unique labeling chemistry of 2-aminobenzoic acid (2AA; anthranilic acid, AA) for measuring activities of both β1-4 galactosyltransferase (GalT-1) and α2-6 sialyltransferase (ST-6) by high-performance liquid chromatography (HPLC) with fluorescence detection (Anumula KR. 2006. Advances in fluorescence derivatization methods for high-performance liquid chromatographic analysis of glycoprotein carbohydrates. Anal Biochem. 350:1-23). N-Acetylglucosamine (GlcNAc) and N-acetyllactosamine were used as acceptors and uridine diphosphate (UDP)-galactose and cytidine monophosphate (CMP)-N-acetylneuraminic acid (NANA) as donors for GalT-1 and ST-6, respectively. Enzymatic products were labeled in situ with AA and were separated from the substrates on TSKgel Amide 80 column using normal-phase conditions. Enzyme units were determined from the peak areas by comparison with the concomitantly derivatized standards Gal-β1-4GlcNAc and NANA-α2-6 Gal-β1-4GlcNAc. Linearity (time and enzyme concentration), precision (intra- and interassay) and reproducibility for the assays were established. The assays were found to be useful in monitoring the enzyme activities during isolation and purification. The assays were highly sensitive and performed equal to or better than the traditional radioactive sugar-based measurements. The assay format can also be used for measuring the activity of other transferases, provided that the carbohydrate acceptors contain a reducing end for labeling. An assay for glycoprotein acceptors was developed using IgG. A short HPLC profiling method was developed for the separation of IgG glycans (biantennary G0, G1, G2, mono- and disialylated), which facilitated the determination of GalT-1 and ST-6 activities in a rapid manner. Furthermore, this profiling method should prove useful for monitoring the changes in IgG glycans in clinical settings. PMID:22459802

  15. A novel glucose biosensor based on phosphonic acid-functionalized silica nanoparticles for sensitive detection of glucose in real samples

    An effective strategy for preparation amperometric biosensor by using the phosphonic acid-functionalized silica nanoparticles (PFSi NPs) as special modified materials is proposed. In such a strategy, glucose oxidase (GOD) was selected as model protein to fabricate glucose biosensor in the presence of phosphonic acid-functionalized silica nanoparticles (PFSi NPs). The PFSi NPs were first modified on the surface of glassy carbon (GC) electrode, then, GOD was adsorbed onto the PFSi NPs film by drop-coating. The PFSi NPs were characterized by transmission electron microscopy (TEM) and nuclear magnetic resonance (NMR) spectra. The interaction of PFSi NPs with GOD was investigated by the circular dicroism spectroscopy (CD). The results showed PFSi NPs could essentially maintain the native conformation of GOD. The direct electron transfer of GOD on (PFSi NPs)/GCE electrode exhibited excellent electrocatalytic activity for the oxidation of glucose. The proposed biosensor modified with PFSi NPs displayed a fast amperometric response (5 s) to glucose, a good linear current–time relation over a wide range of glucose concentrations from 5.00 × 10−4 to 1.87 × 10−1 M, and a low detection limit of 2.44 × 10−5 M (S/N = 3). Moreover, the biosensor can be used for assessment of the concentration of glucose in many real samples (relative error < 3%). The GOD biosensor modified with PFSi NPs will have essential meaning and practical application in future that attributed to the simple method of fabrication and good performance

  16. Colorimetric detection of fluoride ion by 5-arylidenebarbituric acids: dual interaction mode for fluoride ion with single receptor.

    Saravanan, Chinnusamy; Easwaramoorthi, Shanmugam; Wang, Leeyih

    2014-04-01

    Two 5-arylidenebarbituric acid derivatives (IH and IM) have been synthesized by the Knoevenagel condensation of barbituric acid with 4-N,N-dimethylamino benzaldehyde and studied for anion sensing activities. Both receptors sense fluoride ion with high selectivity and sensitivity and the sensing action has been demonstrated by naked eye detection, UV-visible absorption, and fluorescence spectral changes in the presence of F(-). Indeed, the F(-) sensing mechanism for receptor IH depends on F(-) ion concentration. While at higher concentrations F(-) forms strong hydrogen bonding interaction with the N-H proton of the receptor, at lower concentrations sensing is influenced by the deprotonation of the methylene proton, followed by the chemical reaction, which is also confirmed by the (1)H-NMR technique. On the other hand, when replacing the N-H proton with a methyl group, IM does not show any concentration dependent behaviour with F(-). The F(-) concentration dependent sensing is attributed to the changes in the receptor-anion interaction equilibrium, where at higher F(-) concentrations, F(-) interacts with the receptor through hydrogen bonding and at lower concentrations it induces a chemical reaction. PMID:24500374

  17. Poly-glutamic acid modified carbon nanotube-doped carbon paste electrode for sensitive detection of L-tryptophan.

    Liu, Xiao; Luo, Liqiang; Ding, Yaping; Ye, Daixin

    2011-08-01

    A novel poly-glutamic acid (PGA) film modified carbon paste electrode (CPE) incorporating carbon nanotubes (CNTs) was first prepared for the determination of l-tryptophan (l-Trp). Scanning electron microscopy and Fourier transform infrared spectroscopy were applied for characterization of the surface morphology of the modified electrodes and cyclic voltammetry was used to investigate the electrochemical properties of the proposed electrode towards the oxidation of l-Trp. Optimization of the experimental parameters was performed with regard to pH, ratio of CNTs, concentration of glutamic acid, electro-polymerization cycles, accumulation time and concentration of sodium dodecylbenzene sulfonate. The linearity between the oxidation peak current and the l-Trp concentration was obtained in the range of 5.0×10(-8) to 1.0×10(-4)M with a detection limit of 1.0×10(-8)M (S/N=3) and the sensitivity was calculated to be 1143.79μA∙mM(-1)∙cm(-2). In addition, the PGA modified CPE incorporating CNTs displayed high selectivity, good stability and reproducibility, making it suitable for the routine analysis of l-Trp in clinical use. PMID:21640670

  18. Centrifugal Microfluidic System for Nucleic Acid Amplification and Detection

    Baogang Miao

    2015-11-01

    Full Text Available We report here the development of a rapid PCR microfluidic system comprising a double-shaft turntable and centrifugal-based disc that rapidly drives the PCR mixture between chambers set at different temperatures, and the bidirectional flow improved the space utilization of the disc. Three heating resistors and thermistors maintained uniform, specific temperatures for the denaturation, annealing, and extension steps of the PCR. Infrared imaging showed that there was little thermal interference between reaction chambers; the system enabled the cycle number and reaction time of each step to be independently adjusted. To validate the function and efficiency of the centrifugal microfluidic system, a 350-base pair target gene from the hepatitis B virus was amplified and quantitated by fluorescence detection. By optimizing the cycling parameters, the reaction time was reduced to 32 min as compared to 120 min for a commercial PCR machine. DNA samples with concentrations ranging from 10 to 106 copies/mL could be quantitatively analyzed using this system. This centrifugal-based microfluidic platform is a useful system and possesses industrialization potential that can be used for portable diagnostics.

  19. Detection of biologically active isomers of conjugated linoleic acid in kaymak

    Ökten, Sevtap

    2005-12-01

    Full Text Available Numerous physiological effects are attributed to conjugated linoleic acids (CLA. Biologically active isomers of CLA ( cis -9, trans -11 (C18:2 and trans- 10, cis- 12 (C18:2 have been reported to have anticarcinogenic, antioxidative and antiatherosclerotic properties. Relatively rich sources of CLA include milk fat-containing foods such as kaymak. Kaymak is a kind of concentrated cream which is traditionally manufactured from buffalo or cow's milk mainly in Turkey . The objective of this study was to determine CLA concentrations during kaymak production. Kaymak was manufactured from cow's milk which was enriched with unfermented cream. Biologically active isomers of CLA in raw milk, cream and kaymak were analyzed using gas chromatography. The method was quick, repeatable and sensitive for the CLA determination of samples. Significant differences were found among the concentrations of both isomer and total CLA during the production process (pNumerosos efectos fisiológicos se atribuyen a los ácidos linoleico conjugados (CLA. Así los isómeros biológicamente activos ( cis -9, trans -11 (C18:2 y trans- 10, cis del ácido linoleico han sido descritos con propiedades anticarcinogénicas, antioxidantes y antiarterioscleróticas. Fuentes relativamente ricas de CLA incluyen alimentos con grasas lácteas tales como el kaymak. El kaymak es una crema concentrada elaborada de leche de búfalo o vaca principalmente en Turquía. El objetivo de este estudio fue la determinación de la concentración de CLA durante la producción de kaymak. El kaymak objeto de estudio fue elaborado a partir de leche de vaca que fue enriquecida con crema no fermentada. Los isómeros biológicamente activos del CLA fueron analizados por cromatografía gaseosa en leche cruda, crema y kaymak. El método empleado fue rápido, reproducible y sensible. Se encontraron diferencias significativas en las concentraciones de ambos isómeros y de CLA total durante el proceso de producci

  20. Determination of aromatic and sulfur-containing amino acids, peptides, and proteins using high-performance liquid chromatography with photolytic electrochemical detection

    Dou, Lin; Krull, I.S. (Northeastern Univ., Boston, MA (USA))

    1990-12-01

    Aromatic amino acids, sulfur-containing amino acids, peptides containing such constituents, and proteins can now be detected in high-performance liquid chromatography by the use of on-line, postcolumn, continuous photolytic derivatization with electrochemical (HPLC-h{nu}-EC) detection. The overall approach is a very simple, reproducible, rapid, and fully automatable approach for the determination of certain amino acids, peptides, and proteins with excellent selectivity, sensitivity, and linearities of response. Dual-electrode response ratios, lamp-on/lamp-off behavior, and chromatographic capacity factors all contribute to the enhanced selectivity of the overall HPLC-h{nu}-EC determination for these particular classes of bioorganics and biopolymers. The analytical figures of merit, chromatography detection, and method validation approaches have all be optimally derived and demonstrated reproducible. Applications of the basic methodology to real-world samples are demonstrated and validated.

  1. Impaired folding and subunit assembly as disease mechanism

    Bross, P; Andresen, B S; Gregersen, N

    1998-01-01

    Rapid progress in DNA technology has entailed the possibility of readily detecting mutations in disease genes. In contrast to this, techniques to characterize the effects of mutations are still very time consuming. It has turned out that many of the mutations detected in disease genes are missense...... mutations. Characterization of the effect of these mutations is particularly important in order to establish that they are disease causing and to estimate their severity. We use the experiences with investigation of medium-chain acyl-CoA dehydrogenase deficiency as an example to illustrate that (i) impaired...... folding is a common effect of missense mutations occurring in genetic diseases, (ii) increasing the level of available chaperones may augment the level of functional mutant protein in vivo, and (iii) one mutation may have multiple effects. The interplay between the chaperones assisting folding and...

  2. Simple synthesis of thioglycolic acid-coated CdTe quantum dots as probes for Norfloxacin lactate detection

    Wei, Xiao; Zhou, Zhiping; Hao, Tongfan [School of Material Science and Engineering, Jiangsu University, Zhenjiang 212013 (China); Li, Hongji [School of Chemistry and Chemical Engineering, Jiangsu University, Zhenjiang 212013 (China); Dai, Jiangdong [School of Material Science and Engineering, Jiangsu University, Zhenjiang 212013 (China); Gao, Lin; Zheng, Xudong; Wang, Jixiang [School of Chemistry and Chemical Engineering, Jiangsu University, Zhenjiang 212013 (China); Yan, Yongsheng, E-mail: weixiaokeyan@163.com [School of Chemistry and Chemical Engineering, Jiangsu University, Zhenjiang 212013 (China)

    2015-05-15

    In this study, a simple and effective fluorometry method has been developed and used for the determination of Norfloxacin lactate (NOR-L) by the fluorescence quenching of thioglycolic acid (TGA)-coated CdTe quantum dots (QDs). The TGA-CdTe QDs were obtained in a simple way without precursor preparation, heating, pH adjustment and N{sub 2} protection. The CdTe QDs were characterized by TEM, UV–vis spectrophotometer and spectrofluorometer. Meanwhile, spectrofluorometer was used to evaluation of simple, convenient and highly sensitive determination of NOR-L. After the experimental conditions were optimized, a good linear relationship was obtained from 0.1–100 μg/mL with the coefficient of determination (0.99342) and the limit of detection (LOD) was 0.031 μg/mL. Moreover, a possible quenching mechanism was investigated and the CdTe QDs were also successfully used to confirm the NOR-L in pharmaceutical formations. The proposed method is rapid, simple, and applied. - Highlights: • The synthesis procedures were very simple. • The CdTe QDs were used to detect Norfloxacin lactate. • The mechanism of the proposed reaction was discussed.

  3. Simple synthesis of thioglycolic acid-coated CdTe quantum dots as probes for Norfloxacin lactate detection

    In this study, a simple and effective fluorometry method has been developed and used for the determination of Norfloxacin lactate (NOR-L) by the fluorescence quenching of thioglycolic acid (TGA)-coated CdTe quantum dots (QDs). The TGA-CdTe QDs were obtained in a simple way without precursor preparation, heating, pH adjustment and N2 protection. The CdTe QDs were characterized by TEM, UV–vis spectrophotometer and spectrofluorometer. Meanwhile, spectrofluorometer was used to evaluation of simple, convenient and highly sensitive determination of NOR-L. After the experimental conditions were optimized, a good linear relationship was obtained from 0.1–100 μg/mL with the coefficient of determination (0.99342) and the limit of detection (LOD) was 0.031 μg/mL. Moreover, a possible quenching mechanism was investigated and the CdTe QDs were also successfully used to confirm the NOR-L in pharmaceutical formations. The proposed method is rapid, simple, and applied. - Highlights: • The synthesis procedures were very simple. • The CdTe QDs were used to detect Norfloxacin lactate. • The mechanism of the proposed reaction was discussed

  4. Label-free surface-enhanced Raman scattering strategy for rapid detection of penicilloic acid in milk products.

    Qi, Meihui; Huang, Xiaoyan; Zhou, Yujie; Zhang, Liying; Jin, Yang; Peng, Yan; Jiang, Huijun; Du, Shuhu

    2016-04-15

    A label-free surface-enhanced Raman scattering (SERS) strategy based on silver-coated gold nanoparticles (Au@Ag NPs) was developed for rapid detection of penicilloic acid (PA) in milk products. It has been demonstrated that core size and shell thickness of Au@Ag NPs are two critical variants affecting enhancement of Raman signals by coupling of two plasma resonance absorption. The Au@Ag NPs with 26-nm core and 9-nm Ag shell exhibit excellent Raman enhancement, in particular, upon the formation of hot spots through NPs aggregation induced by interaction between target molecules and Au@Ag NPs. Compared to the early studies limited to laboratory settings, our analytical approach is simple (without sample pretreatment), less time-consuming (within ∼3 min) and inexpensive. The limit of detection of PA is 3.00 ppm, 3.00 ppm and 4.00 ppm in liquid milk, yogurt and milk powder, respectively. The label-free SERS technique offers a potential for the on-site monitoring of chemical contaminants in milk products. PMID:26617009

  5. The value of peri-interventional procedure serum bile acid (TBA) detection in patients with primary liver cancer

    Objective: To investigate the clinical value of peri-interventional procedure serum bile acid (TBA) detection in patients with primary liver cancer. Methods: The serum TBA was examined peri-operatively in 160 patients with primary liver cancer for testing the correlations between TBA, liver function, the degree of hepatocirrhosis, interventional therapy method and hepatic failure. Results: The preoperative mean value of serum TBA increased significantly in comparing with that of the control group (P<0.01). The preoperative value of serum TBA in different Child grading patients with primary liver cancer were different significantly (P<0.01), Child A< Child B< Child C, the increased degree of serum TBA corresponded with Child grading of the liver function and the cirrhotic degree of liver. In patients with liver function of Child B and C, the postoperative mean values of serum TBA in different interventional therapy methods were different significantly (P<0.01). Comparing with that of the patients without hepatic failure, the postoperative value of serum TBA in the patients with hepatic failure increased significantly (P<0.01). Conclusions: The value of serum TBA can sensitively and accurately reflect liver reserve ability and damage degree of peri-interventional procedure liver function. Hepatic failure can be detected in time and the prognosis of the patients with primary liver cancer can be predicted by testing the value of serum TBA continually. (authors)

  6. Fast quantification of α-lipoic acid in biological samples and dietary supplements using batch injection analysis with amperometric detection.

    Santos Pereira, Laise Nayra Dos; da Silva, Iranaldo Santos; Araújo, Thaylan Pinheiro; Tanaka, Auro Atsushi; Angnes, Lúcio

    2016-07-01

    Batch injection analysis (BIA) with amperometric detection, using a pyrolytic graphite electrode modified with cobalt phthalocyanine (PG/CoPc), was employed for determination of α-lipoic acid (ALA) in pharmaceutical product and in synthetic urine samples. The proposed BIA method is based on the application of a potential of +0.9V vs. Ag/AgCl, KCl sat, enabling quantification of ALA over a concentration range from 1.3×10(-6) to 1.0×10(-4)molL(-1), with a detection limit of 1.5×10(-8)molL(-1). A sampling rate of 180 injections per hour was attained and measurements of the reproducibility of successive injections (100µmolL(-1) ALA on the same electrode) showed a RSD of 2.11% for 40 successive injections. The new sensor was utilised for ALA quantification in a dietary pharmaceutical supplement and in synthetic urine and the results obtained for both samples were compared with parallel analysis using high performance liquid chromatography (HPLC), the method recommended by the United States Pharmacopeia. The results obtained were similar (at a 95% confidence level) and in the case of the synthetic urine sample (prepared with a known amount of ALA) the recovery was situated between 98.0% and 102.6%. PMID:27154671

  7. A screening lateral flow immunochromatographic assay for on-site detection of okadaic acid in shellfish products.

    Lu, Shi-Ying; Lin, Chao; Li, Yan-Song; Zhou, Yu; Meng, Xian-Mei; Yu, Shi-Yu; Li, Zhao-Hui; Li, Le; Ren, Hong-Lin; Liu, Zeng-Shan

    2012-03-15

    A lateral flow immunochromatographic (LFIC) test strip based on a colloidal gold-monoclonal antibody (McAb) conjugate was developed for on-site rapid detection of okadaic acid (OA) in shellfish. It applies a competitive format using an immobilized toxin conjugate and free toxin present in samples. The McAb against OA was conjugated with 20-nm colloidal gold as detector reagent. The toxin in the sample competed with the immobilized toxin to bind to the gold conjugated with McAb. The colloidal gold/McAb/toxin mobile complex was not captured by OA-bovine serum albumin (BSA) on the test line, but it was captured by goat anti-mouse immunoglobulin G (IgG) on the control line. The color density of the test line correlated with the concentration of toxin in the range of 10-50 ng ml(-1). The qualitative detection limit of 150 μg kg(-1) sample was close to the European Union (EU) regulatory limit (160 μg kg(-1)). Therefore, these strips were able to directly and qualitatively estimate the consuming safety of shellfish. They require no equipment because of available visual results, and they screened numerous samples within 10 min. The results were further confirmed by high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). As a food safety screening tool, the test strips are convenient and useful to rapidly on-site test the presence of OA in shellfish products. PMID:22266294

  8. Nonenzymatic L-lysine amino acid detection using titanium oxide nanoparticles/multi wall carbon nanotube composite electrodes

    Graphical abstract: - Abstract: For the first time, a nonenzymatic electrochemical sensor for the detection of lysine was proposed based on immobilizing Multi wall carbon nanotube (MWCNT) and Titanium oxide nanoparticles (TiO2NPs) on glassy carbon (GC) electrode. Scaning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) were used to confirm the successful stepwise assembly procedure of the sensor. The electrocatalytical behaviors of the sensor were also investigated by cyclic voltammetry (CV) and differential pulse voltammetry (DPV). The results showed that MWCNT- TiO2NPs exhibited a remarkable electrocatalytic activity for the oxidation of lysine. Under optimal conditions, the DPV response of the sensor was proportional to the lysine concentration in the range of 500 to 5500 nanomolar with a detection limit and sensitivity of 390 nM and 0.1795 μAμM−1. This electrode show many advantages such as simple preparation without using any enzyme special electron transfer mediator or specific reagent, excellent catalytic activity at physiological pH values and antifouling property toward lysine and its oxidation product. Furthermore, the selectivity of the proposed sensor was tested in the presence of some amino acids and the response of the sensor was encountered with interferences of proline and tryptophan at equimolar concentrations

  9. Folding and Stabilization of Native-Sequence-Reversed Proteins

    Zhang, Yuanzhao; Zhou, Ruhong

    2016-01-01

    Though the problem of sequence-reversed protein folding is largely unexplored, one might speculate that reversed native protein sequences should be significantly more foldable than purely random heteropolymer sequences. In this article, we investigate how the reverse-sequences of native proteins might fold by examining a series of small proteins of increasing structural complexity ({\\alpha}-helix, \\b{eta}-hairpin, {\\alpha}-helix bundle, and {\\alpha}/\\b{eta}-protein). Employing a tandem protein structure prediction algorithmic and molecular dynamics simulation approach, we find that the ability of reverse sequences to adopt native-like folds is strongly in influenced by protein size and the flexibility of the native hydrophobic core. For \\b{eta}-hairpins with reverse-sequences that fail to fold, we employ a simple mutational strategy for guiding stable hairpin formation that involves the insertion of amino acids into the \\b{eta}-turn region. This systematic look at reverse sequence duality sheds new light on t...

  10. How does a simplified-sequence protein fold?

    Guarnera, Enrico; Pellarin, Riccardo; Caflisch, Amedeo

    2009-09-16

    To investigate a putatively primordial protein we have simplified the sequence of a 56-residue alpha/beta fold (the immunoglobulin-binding domain of protein G) by replacing it with polyalanine, polythreonine, and diglycine segments at regions of the sequence that in the folded structure are alpha-helical, beta-strand, and turns, respectively. Remarkably, multiple folding and unfolding events are observed in a 15-micros molecular dynamics simulation at 330 K. The most stable state (populated at approximately 20%) of the simplified-sequence variant of protein G has the same alpha/beta topology as the wild-type but shows the characteristics of a molten globule, i.e., loose contacts among side chains and lack of a specific hydrophobic core. The unfolded state is heterogeneous and includes a variety of alpha/beta topologies but also fully alpha-helical and fully beta-sheet structures. Transitions within the denatured state are very fast, and the molten-globule state is reached in structure of the wild-type is more rigid than the molten-globule conformation of the simplified-sequence variant. The difference in structural stability and the very fast folding of the simplified protein suggest that evolution has enriched the primordial alphabet of amino acids mainly to optimize protein function by stabilization of a unique structure with specific tertiary interactions. PMID:19751679

  11. Anion-exchange high-performance liquid chromatography with post-column detection for the analysis of phytic acid and other inositol phosphates

    Rounds, M. A.; Nielsen, S. S.; Mitchell, C. A. (Principal Investigator)

    1993-01-01

    The use of gradient anion-exchange HPLC, with a simple post-column detection system, is described for the separation of myo-inositol phosphates, including "phytic acid" (myo-inositol hexaphosphate). Hexa-, penta-, tetra-, tri- and diphosphate members of this homologous series are clearly resolved within 30 min. This method should facilitate analysis and quantitation of "phytic acid" and other inositol phosphates in plant, food, and soil samples.

  12. A comparison of RNA folding measures

    Gardner Paul P

    2005-10-01

    Full Text Available Abstract Background In the last few decades there has been a great deal of discussion concerning whether or not noncoding RNA sequences (ncRNAs fold in a more well-defined manner than random sequences. In this paper, we investigate several existing measures for how well an RNA sequence folds, and compare the behaviour of these measures over a large range of Rfam ncRNA families. Such measures can be useful in, for example, identifying novel ncRNAs, and indicating the presence of alternate RNA foldings. Results Our analysis shows that ncRNAs, but not mRNAs, in general have lower minimal free energy (MFE than random sequences with the same dinucleotide frequency. Moreover, even when the MFE is significant, many ncRNAs appear to not have a unique fold, but rather several alternative folds, at least when folded in silico. Furthermore, we find that the six investigated measures are correlated to varying degrees. Conclusion Due to the correlations between the different measures we find that it is sufficient to use only two of them in RNA folding studies, one to test if the sequence in question has lower energy than a random sequence with the same dinucleotide frequency (the Z-score and the other to see if the sequence has a unique fold (the average base-pair distance, D.

  13. A comparison of RNA folding measures

    Freyhult, E.; Gardner, P. P.; Moulton, V.

    2005-01-01

    Background In the last few decades there has been a great deal of discussion concerning whether or not noncoding RNA sequences (ncRNAs) fold in a more well-defined manner than random sequences. In this paper, we investigate several existing measures for how well an RNA sequence folds, and compare...

  14. Stochastic Resonance in Protein Folding Dynamics.

    Davtyan, Aram; Platkov, Max; Gruebele, Martin; Papoian, Garegin A

    2016-05-01

    Although protein folding reactions are usually studied under static external conditions, it is likely that proteins fold in a locally fluctuating cellular environment in vivo. To mimic such behavior in in vitro experiments, the local temperature of the solvent can be modulated either harmonically or using correlated noise. In this study, coarse-grained molecular simulations are used to investigate these possibilities, and it is found that both periodic and correlated random fluctuations of the environment can indeed accelerate folding kinetics if the characteristic frequencies of the applied fluctuations are commensurate with the internal timescale of the folding reaction; this is consistent with the phenomenon of stochastic resonance observed in many other condensed-matter processes. To test this theoretical prediction, the folding dynamics of phosphoglycerate kinase under harmonic temperature fluctuations are experimentally probed using Förster resonance energy transfer fluorescence measurements. To analyze these experiments, a combination of theoretical approaches is developed, including stochastic simulations of folding kinetics and an analytical mean-field kinetic theory. The experimental observations are consistent with the theoretical predictions of stochastic resonance in phosphoglycerate kinase folding. When combined with an alternative experiment on the protein VlsE using a power spectrum analysis, elaborated in Dave et al., ChemPhysChem 2016, 10.1002/cphc.201501041, the overall data overwhelmingly point to the experimental confirmation of stochastic resonance in protein folding dynamics. PMID:26992148

  15. Enzyme immunoassay for carminic acid in foods.

    Yoshida, A; Takagaki, Y; Nishimune, T

    1995-01-01

    A competitive enzyme immunoassay (EIA) for carminic acid was investigated. Monoclonal anticarminic acid antibody was obtained from A/J mice immunized with carminic acid-human immunoglobulin G (IgG) conjugate. Carminic acid was extracted with distilled water from beverage, jelly, candy, pasta sauce, yogurt, or ice cream samples. Ham or fish paste samples were digested with pronase, then carminic acid was extracted from samples with sodium hydroxide solution. The extract was diluted more than 10-fold with 1% gelatin in borate buffer solution. Microtiter plates were coated with carminic acid-bovine serum albumin (BSA) conjugate or just BSA. Goat anti-mouse IgG(H+L)-peroxidase complex was used as a second antibody, and 3,3',5,5'-tetramethylbenzidine was used as a substrate for the peroxidase. The working range for quantitative analysis was 0.3-10 ng/mL, and the detection limit was 0.2 micrograms/g original sample. Recoveries of carminic acid by this assay were > 95% for milk beverage and jelly, and > 85% for yogurt and fish paste. Carminic acid was detected in 7 of 26 red-colored commercial food products and ranged from 3.5 to 356 micrograms/g. This EIA system also responded to the structural analogue of carminic acid, laccaic acid. PMID:7756895

  16. Development of a multiplex real time PCR to detect thermophilic lactic acid bacteria in natural whey starters.

    Bottari, Benedetta; Agrimonti, Caterina; Gatti, Monica; Neviani, Erasmo; Marmiroli, Nelson

    2013-01-01

    A multiplex real time PCR (mRealT-PCR) useful to rapidly screen microbial composition of thermophilic starter cultures for hard cooked cheeses and to compare samples with potentially different technological properties was developed. Novel primers directed toward pheS gene were designed and optimized for multiple detection of Lactobacillus helveticus, Lactobacillus delbrueckii, Streptococcus thermophilus and Lactobacillus fermentum. The assay was based on SYBR Green chemistry followed by melting curves analysis. The method was then evaluated for applications in the specific detection of the 4 lactic acid bacteria (LAB) in 29 different natural whey starters for Parmigiano Reggiano cheese production. The results obtained by mRealT-PCR were also compared with those obtained on the same samples by Fluorescence in Situ Hybridization (FISH) and Length-Heterogeneity PCR (LH-PCR). The mRealT-PCR developed in this study, was found to be effective for analyzing species present in the samples with an average sensitivity down to less than 600 copies of DNA and therefore sensitive enough to detect even minor LAB community members of thermophilic starter cultures. The assay was able to describe the microbial population of all the different natural whey starter samples analyzed, despite their natural variability. A higher number of whey starter samples with S. thermophilus and L. fermentum present in their microbial community were revealed, suggesting that these species could be more frequent in Parmigiano Reggiano natural whey starter samples than previously shown. The method was more effective than LH-PCR and FISH and, considering that these two techniques have to be used in combination to detect the less abundant species, the mRealT-PCR was also faster. Providing a single step sensitive detection of L. helveticus, L. delbrueckii, S. thermophilus and L. fermentum, the developed mRealT-PCR could be used for screening thermophilic starter cultures and to follow the presence of

  17. Fabrication of GNPs/CDSH-Fc/nafion modified electrode for the detection of dopamine in the presence of ascorbic acid

    A novel dopamine sensor was fabricated by forming the inclusion complex between mono-6-thio-β-cyclodextrin (CD-SH) and ferrocene (Fc) functionalized gold nanoparticles (GNPs) films on a platinum electrode. The properties of the GNPs/CDSH-Fc nanocomposite were characterized by Fourier transform infrared spectra, UV-visible absorption spectroscopy, transmission electron microscopy and cyclic voltammetry. The electrochemistry of dopamine (DA) was investigated by cyclic voltammetry (CV) and differential pulse voltammograms (DPV). The electrooxidation of dopamine could be catalyzed by Fc/Fc+ couple as a mediator and had a higher electrochemical response due to the unique performance of GNPs/CDSH-Fc. The anodic peaks of DA and ascorbic acid (AA) in their mixture can be well separated by the prepared electrode. Under optimum conditions linear calibration graphs were obtained over the DA concentration range 2.0 x 10-6 to 5.0 x 10-5 M with a correlation coefficient of 0.998 and a detection limit of 9.0 x 10-8 M (S/N = 3). The modified electrode had been effectively applied for the assay of DA in dopamine hydrochloride injections. This work provides a simple and easy approach to selectively detect DA in the presence of AA. - Research highlights: → The sensor of DA was constructed by using GNPs/CDSH-Fc as the building block. → Inclusion complex on the surface of GNPs decreased the leakage of mediator. → The electro-oxidation of DA could be catalyzed by Fc/Fc+ couple as a mediator. → This work provides a simple approach to selectively detect DA in the presence of AA.

  18. Fabrication of GNPs/CDSH-Fc/nafion modified electrode for the detection of dopamine in the presence of ascorbic acid

    Chen Ming; Wei Xiujuan; Qian Hui; Diao Guowang, E-mail: gwdiao@yzu.edu.cn

    2011-10-10

    A novel dopamine sensor was fabricated by forming the inclusion complex between mono-6-thio-{beta}-cyclodextrin (CD-SH) and ferrocene (Fc) functionalized gold nanoparticles (GNPs) films on a platinum electrode. The properties of the GNPs/CDSH-Fc nanocomposite were characterized by Fourier transform infrared spectra, UV-visible absorption spectroscopy, transmission electron microscopy and cyclic voltammetry. The electrochemistry of dopamine (DA) was investigated by cyclic voltammetry (CV) and differential pulse voltammograms (DPV). The electrooxidation of dopamine could be catalyzed by Fc/Fc{sup +} couple as a mediator and had a higher electrochemical response due to the unique performance of GNPs/CDSH-Fc. The anodic peaks of DA and ascorbic acid (AA) in their mixture can be well separated by the prepared electrode. Under optimum conditions linear calibration graphs were obtained over the DA concentration range 2.0 x 10{sup -6} to 5.0 x 10{sup -5} M with a correlation coefficient of 0.998 and a detection limit of 9.0 x 10{sup -8} M (S/N = 3). The modified electrode had been effectively applied for the assay of DA in dopamine hydrochloride injections. This work provides a simple and easy approach to selectively detect DA in the presence of AA. - Research highlights: {yields} The sensor of DA was constructed by using GNPs/CDSH-Fc as the building block. {yields} Inclusion complex on the surface of GNPs decreased the leakage of mediator. {yields} The electro-oxidation of DA could be catalyzed by Fc/Fc{sup +} couple as a mediator. {yields} This work provides a simple approach to selectively detect DA in the presence of AA.

  19. Cooperative Tertiary Interaction Network Guides RNA Folding

    Behrouzi, Reza; Roh, Joon Ho; Kilburn, Duncan; Briber, R.M.; Woodson, Sarah A. (JHU); (Maryland)

    2013-04-08

    Noncoding RNAs form unique 3D structures, which perform many regulatory functions. To understand how RNAs fold uniquely despite a small number of tertiary interaction motifs, we mutated the major tertiary interactions in a group I ribozyme by single-base substitutions. The resulting perturbations to the folding energy landscape were measured using SAXS, ribozyme activity, hydroxyl radical footprinting, and native PAGE. Double- and triple-mutant cycles show that most tertiary interactions have a small effect on the stability of the native state. Instead, the formation of core and peripheral structural motifs is cooperatively linked in near-native folding intermediates, and this cooperativity depends on the native helix orientation. The emergence of a cooperative interaction network at an early stage of folding suppresses nonnative structures and guides the search for the native state. We suggest that cooperativity in noncoding RNAs arose from natural selection of architectures conducive to forming a unique, stable fold.

  20. The geometry and wetting of capillary folding

    Péraud, Jean-Philippe

    2014-01-01

    Capillary forces are involved in a variety of natural phenomena, ranging from droplet breakup to the physics of clouds. The forces from surface tension can also be exploited in industrial application provided the length scales involved are small enough. Recent experimental investigations showed how to take advantage of capillarity to fold planar structures into three-dimensional configurations by selectively melting polymeric hinges joining otherwise rigid shapes. In this paper we use theoretical calculations to quantify the role of geometry and fluid wetting on the final folded state. Considering folding in two and three dimensions, studying both hydrophilic and hydrophobic situations with possible contact angle hysteresis, and addressing the shapes to be folded to be successively infinite, finite, curved, kinked, elastic, we are able to derive an overview of the geometrical parameter space available for capillary folding.

  1. Detection of the HIV-1 nucleic acid in spermatozoa of HIV/AIDS patients%HIV/AIDS患者精子中病毒核酸的检测

    王典; 李练兵; 侯志伟; 谢庆东; 康祥锦; 黄天华

    2011-01-01

    Objective To detect the presence of HIV - 1 nucleic acid in spermatozoa of HIV/AIDS patients.Methods Spermatozoon samples of 33 HIV/AIDS patients were prepared. Fluorescence in situ hybridization ( FISH ) ,in which biotinylated HIV gag and HIV pol were used as probes, was performed to detect HIV nucleic acid in spermatozoa. Results HIV nucleic acid was detected in spermatozoa of 9 patients( 27. 3% ). Conclusion These results further confirm the presence of HIV nucleic acid in sperm of HIV/AIDS patients and indicates that FISH is a sensitive and reliable method to detect the nucleic acid in sperm.%目的 检测HIV/AIDS患者精子中HIV核酸.方法 制备33例HIV/AIDS患者精子样本,采用荧光原位杂交(FISH),以生物素标记的HIV gag及 HIV pol作为双探针对这些患者精子进行HIV核酸检测.结果 在9例(27.3%)患者中检测到阳性荧光信号.结论 HIV/AIDS患者精子存有HIV核酸;FISH是一种检测HIV/AIDS患者精子病毒核酸灵敏而可靠的方法.

  2. Environmental Fluctuations and Stochastic Resonance in Protein Folding.

    Dave, Kapil; Davtyan, Aram; Papoian, Garegin A; Gruebele, Martin; Platkov, Max

    2016-05-01

    Stochastic resonance is a mechanism whereby a weak signal becomes detectable through the addition of noise. It is common in many macroscopic biological phenomena, but here we ask whether it can be observed in a microscopic biological phenomenon, protein folding. We investigate the folding kinetics of the protein VlsE, with a folding relaxation time of about 0.7 seconds at 38 °C in vitro. First we show that the VlsE unfolding/refolding reaction can be driven by a periodic thermal excitation above the reaction threshold. We detect the reaction by fluorescence from FRET labels on VlSE and show that accurate rate coefficients and activation barriers can be obtained from modulated kinetics. Then we weaken the periodic temperature modulation below the reaction threshold, and show that addition of artificial thermal noise speeds up the reaction from an undetectable to a detectable rate. We observe a maximum in the recovered signal as a function of thermal noise, a stochastic resonance. Simulation of a small model-protein, analysis in an accompanying theory paper, and our experimental result here all show that correlated noise is a physically and chemically plausible mechanism by which cells could modulate biomolecular dynamics during threshold processes such as signaling. PMID:26711088

  3. Fabrication of Palladium Nanoparticles on Porous Aromatic Frameworks as a Sensing Platform to Detect Vanillin.

    Vilian, A T Ezhil; Puthiaraj, Pillaiyar; Kwak, Cheol Hwan; Hwang, Seung-Kyu; Huh, Yun Suk; Ahn, Wha-Seung; Han, Young-Kyu

    2016-05-25

    Here, we report the fabrication of palladium nanoparticles on porous aromatic frameworks (Pd/PAF-6) using a facile chemical approach, which was characterized by various spectro- and electrochemical techniques. The differential pulse voltammetry (DPV) response of Pd/PAF-6 toward the vanillin (VA) sensor shows a linear relationship over concentrations (10-820 pM) and a low detection limit (2 pM). Pd/PAF-6 also exhibited good anti-interference performance toward 2-fold excess of ascorbic acid, nitrophenol, glutathione, glucose, uric acid, dopamine, ascorbic acid, 4-nitrophenol, glutathione, glucose, uric acid, dopamine, and 100-fold excess of Na(+), Mg(2+), and K(+) during the detection of VA. The developed electrochemical sensor based on Pd/PAF-6 had good reproducibility, as well as high selectivity and stability. The established sensor revealed that Pd/PAF-6 could be used to detect VA in biscuit and ice cream samples with satisfactory results. PMID:27149292

  4. Improving detection sensitivity of amino acids in thyroid tissues by using phthalic acid as a mobile phase additive in hydrophilic interaction chromatography-electrospray ionization-tandem mass spectrometry

    Highlights: • HILIC–ESI-MS/MS method was used to quantify 24 free AAs in human thyroid tissues. • Addition of 0.08 mM of phthalic acid to the eluent enhanced the sensitivity of AAs. • Narrowed peak shapes of AAs were achieved with phthalic acid in the mobile phase. • The mechanism for the signal intensity enhancement by phthalic acid was investigated. - Abstract: In this work, 0.08 mmol L−1 of phthalic acid was introduced as a mobile phase additive to quantify free amino acids (AAs) by hydrophilic interaction liquid chromatography (HILIC) coupled with electrospray ionization tandem mass spectrometry (ESI-MS/MS). The addition of phthalic acid significantly increased the signal intensity of protonated AA ions, resulting from the decrease of the relative abundance of AA sodium adducts. Meanwhile, the chromatographic peak shapes of AAs were optimized. As a consequence, there was a noticeable increase in the sensitivity of detection for AAs. The limits of detection (LOD) and quantification (LOQ) of the AAs ranged from 0.0500 to 20.0 ng mL−1 and from 0.100 to 50.0 ng mL−1, respectively, which were 4–50 times lower compared to the values measured without the addition of phthalic acid. The enhanced detection and separation of AAs were obtained by merely adding phthalic acid to the mobile phase without changing other conditions. Eventually, this simple method was validated and successfully applied to the analysis of twenty-four kinds of free AAs in human thyroid carcinoma and para-carcinoma tissues, demonstrating a significant increase of most AAs in thyroid carcinoma tissues (p < 0.05)

  5. Improving detection sensitivity of amino acids in thyroid tissues by using phthalic acid as a mobile phase additive in hydrophilic interaction chromatography-electrospray ionization-tandem mass spectrometry

    Qi, Wanshu [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China); Guan, Qing [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center (FUSCC), Shanghai 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Sun, Tuanqi, E-mail: tuanqisun@163.com [Department of Head and Neck Surgery, Fudan University Shanghai Cancer Center (FUSCC), Shanghai 200032 (China); Department of Oncology, Shanghai Medical College, Fudan University, Shanghai 200032 (China); Cao, Yanjing [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China); Zhang, Li, E-mail: zhangli7488@sioc.ac.cn [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China); Guo, Yinlong, E-mail: ylguo@sioc.ac.cn [National Center for Organic Mass Spectrometry in Shanghai, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai 200032 (China)

    2015-04-22

    Highlights: • HILIC–ESI-MS/MS method was used to quantify 24 free AAs in human thyroid tissues. • Addition of 0.08 mM of phthalic acid to the eluent enhanced the sensitivity of AAs. • Narrowed peak shapes of AAs were achieved with phthalic acid in the mobile phase. • The mechanism for the signal intensity enhancement by phthalic acid was investigated. - Abstract: In this work, 0.08 mmol L{sup −1} of phthalic acid was introduced as a mobile phase additive to quantify free amino acids (AAs) by hydrophilic interaction liquid chromatography (HILIC) coupled with electrospray ionization tandem mass spectrometry (ESI-MS/MS). The addition of phthalic acid significantly increased the signal intensity of protonated AA ions, resulting from the decrease of the relative abundance of AA sodium adducts. Meanwhile, the chromatographic peak shapes of AAs were optimized. As a consequence, there was a noticeable increase in the sensitivity of detection for AAs. The limits of detection (LOD) and quantification (LOQ) of the AAs ranged from 0.0500 to 20.0 ng mL{sup −1} and from 0.100 to 50.0 ng mL{sup −1}, respectively, which were 4–50 times lower compared to the values measured without the addition of phthalic acid. The enhanced detection and separation of AAs were obtained by merely adding phthalic acid to the mobile phase without changing other conditions. Eventually, this simple method was validated and successfully applied to the analysis of twenty-four kinds of free AAs in human thyroid carcinoma and para-carcinoma tissues, demonstrating a significant increase of most AAs in thyroid carcinoma tissues (p < 0.05)

  6. The human PDI family: Versatility packed into a single fold

    Appenzeller-Herzog, Christian; Ellgaard, Lars

    2007-01-01

    in promoting oxidative protein folding in the ER has been extended in recent years to include roles in other processes such as ER-associated degradation (ERAD), trafficking, calcium homeostasis, antigen presentation and virus entry. Some of these functions are performed by non-catalytic members of...... the family that lack the active-site cysteines. Regardless of their function, all human PDIs contain at least one domain of approximately 100 amino acid residues with structural homology to thioredoxin. As we learn more about the individual proteins of the family, a complex picture is emerging that...... emphasizes as much their differences as their similarities, and underlines the versatility of the thioredoxin fold. Here, we primarily explore the diversity of cellular functions described for the human PDIs. Udgivelsesdato: 2007-Dec-3...

  7. Internally folded expanded metal electrode for battery construction

    Korinek, Paul D. (Inventor); Morgan, Maurice C. (Inventor); Pierce, Doug C. (Inventor)

    1993-01-01

    A battery system is disclosed which includes folded grids of expanded metal inserted through non-conductive substrates and pasted with electrochemically active materials. In the most preferred embodiment, a frame is provided with a plastic insert, and slots are provided in the latter to receive the expanded metal grid. After suitable coinage of the grid and insertion through the plastic film, the grid is sealed and pasted on opposite sides with positive and negative active material. A battery is assembled using one or a plurality of the resulting electrode elements, with separators, to produce a high-power, lead-acid battery. The folded grid provides many of the design benefits of standard bipolar construction.

  8. Energy landscape exploration of the folding processes of biological molecules

    Engel, Megan Clare

    For decades, scientists from every discipline have struggled to understand the mechanism of biological self-assembly, which allows proteins and nucleic acids to fold reliably into functional three-dimensional structures. Such an understanding may hold the key to eliminating diseases such as Alzheimer's and Parkinson's and to effective protein engineering. The current best framework for describing biological folding processes is that of statistical mechanical energy landscape theory, and one of the most promising experimental techniques for exploring molecular energy landscapes is single molecule force spectroscopy (SMFS), in which molecules are mechanically denatured. Theoretical advances have enabled the extraction of complete energy landscape profiles from SMFS data. Here, SMFS experiments performed using laser optical tweezers are analyzed to yield the first ever full landscape profile for an RNA pseudoknot. Further, a promising novel landscape reconstruction technique is validated for the first time using experimental data from a DNA hairpin.

  9. Evaluation of Peptide Nucleic Acid Probe-based Real-time PCR for Detection of Mycobacterium tuberculosis Complex and Nontuberculous Mycobacteria in Respiratory Specimens

    Choi, Young Jin; Kim, Hwi Jun; Shin, Hee Bong; Nam, Hae Seon; Lee, Sang Han; Park, Joon Soo; Park, Kwi Sung; Baek, Kyoung Ah

    2012-01-01

    Background A peptide nucleic acid (PNA) probe-based real-time PCR (PNAqPCR™ TB/NTM detection kit; PANAGENE, Korea) assay has been recently developed for the simultaneous detection of Mycobacterium tuberculosis complex (MTBC) and nontuberculous mycobacteria (NTM) in clinical specimens. The study was aimed at evaluation of the performance of PNA probe-based real-time PCR in respiratory specimens. Methods To evaluate potential cross-reactivity, the extracted DNA specimens from Mycobacterium spec...

  10. Real-Time Detection of Noroviruses in Surface Water by Use of a Broadly Reactive Nucleic Acid Sequence-Based Amplification Assay

    Rutjes, Saskia A.; van den Berg, Harold H. J. L.; Lodder, Willemijn J.; Roda Husman, Ana Maria de

    2006-01-01

    Noroviruses are the most common agents causing outbreaks of viral gastroenteritis. Outbreaks originating from contaminated drinking water and from recreational waters have been described. Due to a lack of cell culture systems, noroviruses are detected mostly by molecular methods. Molecular detection assays for viruses in water are often repressed by inhibitory factors present in the environment, like humic acids and heavy metals. To study the effect of environmental inhibitors on the performa...

  11. Real-time detection of noroviruses in surface water by use of a broadly reactive nucleic acid sequence-based amplification assay.

    Rutjes, Saskia A.; Berg, Harold H J L van den; Lodder, Willemijn J.; Roda Husman, Ana Maria de

    2006-01-01

    Noroviruses are the most common agents causing outbreaks of viral gastroenteritis. Outbreaks originating from contaminated drinking water and from recreational waters have been described. Due to a lack of cell culture systems, noroviruses are detected mostly by molecular methods. Molecular detection assays for viruses in water are often repressed by inhibitory factors present in the environment, like humic acids and heavy metals. To study the effect of environmental inhibitors on the performa...

  12. Detection of Chlamydia trachomatis in urine samples by nucleic acid tests: comparison with culture and enzyme immunoassay of genital swab specimens.

    Schepetiuk, S.; Kok, T.; Martin, L; Waddell, R; Higgins, G

    1997-01-01

    Two commercially available nucleic acid-based tests, ligase chain reaction (LCR; Abbott Laboratories) and PCR (Roche Diagnostics), for the detection of Chlamydia trachomatis in male and female urine samples were compared with culture and enzyme immunoassay (EIA) (Microtrak; Syva) for C. trachomatis detection in genital samples. The samples were collected from 1,005 patients who attended a sexually transmitted disease clinic. In this study population, the prevalence of the infection was 4%. Sp...

  13. Detection of Extended-Spectrum β-Lactamase and Klebsiella pneumoniae Carbapenemase Genes Directly from Blood Cultures by Use of a Nucleic Acid Microarray

    Fishbain, Joel T.; Sinyavskiy, Oleg; Riederer, Kathleen; Hujer, Andrea M.; Robert A Bonomo

    2012-01-01

    The growing crisis of multidrug-resistant (MDR) Gram-negative bacteria requires that current technologies permit the rapid detection of extended-spectrum β-lactamase (blaESBL) and Klebsiella pneumoniae carbapenemase (blaKPC) genes. In the present study, we assessed the performance characteristics of a commercially available nucleic acid microarray system for the detection of blaESBL and blaKPC genes directly from positive blood cultures. Using blood cultures (BCs) that contained Gram-negative...

  14. Comparison of the Folding Mechanism of Highly Homologous Proteins in the Lipid-binding Protein Family

    The folding mechanism of two closely related proteins in the intracellular lipid binding protein family, human bile acid binding protein (hBABP) and rat bile acid binding protein (rBABP) were examined. These proteins are 77% identical (93% similar) in sequence Both of these singl...

  15. Reliable allele detection using SNP-based PCR primers containing Locked Nucleic Acid: application in genetic mapping

    Trognitz Friederike

    2007-02-01

    Full Text Available Abstract Background The diploid, Solanum caripense, a wild relative of potato and tomato, possesses valuable resistance to potato late blight and we are interested in the genetic base of this resistance. Due to extremely low levels of genetic variation within the S. caripense genome it proved impossible to generate a dense genetic map and to assign individual Solanum chromosomes through the use of conventional chromosome-specific SSR, RFLP, AFLP, as well as gene- or locus-specific markers. The ease of detection of DNA polymorphisms depends on both frequency and form of sequence variation. The narrow genetic background of close relatives and inbreds complicates the detection of persisting, reduced polymorphism and is a challenge to the development of reliable molecular markers. Nonetheless, monomorphic DNA fragments representing not directly usable conventional markers can contain considerable variation at the level of single nucleotide polymorphisms (SNPs. This can be used for the design of allele-specific molecular markers. The reproducible detection of allele-specific markers based on SNPs has been a technical challenge. Results We present a fast and cost-effective protocol for the detection of allele-specific SNPs by applying Sequence Polymorphism-Derived (SPD markers. These markers proved highly efficient for fingerprinting of individuals possessing a homogeneous genetic background. SPD markers are obtained from within non-informative, conventional molecular marker fragments that are screened for SNPs to design allele-specific PCR primers. The method makes use of primers containing a single, 3'-terminal Locked Nucleic Acid (LNA base. We demonstrate the applicability of the technique by successful genetic mapping of allele-specific SNP markers derived from monomorphic Conserved Ortholog Set II (COSII markers mapped to Solanum chromosomes, in S. caripense. By using SPD markers it was possible for the first time to map the S. caripense alleles

  16. Reducing the dimensionality of the protein-folding search problem.

    Chellapa, George D; Rose, George D

    2012-08-01

    How does a folding protein negotiate a vast, featureless conformational landscape and adopt its native structure in biological real time? Motivated by this search problem, we developed a novel algorithm to compare protein structures. Procedures to identify structural analogs are typically conducted in three-dimensional space: the tertiary structure of a target protein is matched against each candidate in a database of structures, and goodness of fit is evaluated by a distance-based measure, such as the root-mean-square distance between target and candidate. This is an expensive approach because three-dimensional space is complex. Here, we transform the problem into a simpler one-dimensional procedure. Specifically, we identify and label the 11 most populated residue basins in a database of high-resolution protein structures. Using this 11-letter alphabet, any protein's three-dimensional structure can be transformed into a one-dimensional string by mapping each residue onto its corresponding basin. Similarity between the resultant basin strings can then be evaluated by conventional sequence-based comparison. The disorder → order folding transition is abridged on both sides. At the onset, folding conditions necessitate formation of hydrogen-bonded scaffold elements on which proteins are assembled, severely restricting the magnitude of accessible conformational space. Near the end, chain topology is established prior to emergence of the close-packed native state. At this latter stage of folding, the chain remains molten, and residues populate natural basins that are approximated by the 11 basins derived here. In essence, our algorithm reduces the protein-folding search problem to mapping the amino acid sequence onto a restricted basin string. PMID:22692765

  17. Periodic and stochastic thermal modulation of protein folding kinetics.

    Platkov, Max; Gruebele, Martin

    2014-07-21

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude. PMID:25053342

  18. Periodic and stochastic thermal modulation of protein folding kinetics

    Chemical reactions are usually observed either by relaxation of a bulk sample after applying a sudden external perturbation, or by intrinsic fluctuations of a few molecules. Here we show that the two ideas can be combined to measure protein folding kinetics, either by periodic thermal modulation, or by creating artificial thermal noise that greatly exceeds natural thermal fluctuations. We study the folding reaction of the enzyme phosphoglycerate kinase driven by periodic temperature waveforms. As the temperature waveform unfolds and refolds the protein, its fluorescence color changes due to FRET (Förster resonant Energy Transfer) of two donor/acceptor fluorophores labeling the protein. We adapt a simple model of periodically driven kinetics that nicely fits the data at all temperatures and driving frequencies: The phase shifts of the periodic donor and acceptor fluorescence signals as a function of driving frequency reveal reaction rates. We also drive the reaction with stochastic temperature waveforms that produce thermal fluctuations much greater than natural fluctuations in the bulk. Such artificial thermal noise allows the recovery of weak underlying signals due to protein folding kinetics. This opens up the possibility for future detection of a stochastic resonance for protein folding subject to noise with controllable amplitude

  19. Development of a Microfluidic-Based Optical Sensing Device for Label-Free Detection of Circulating Tumor Cells (CTCs Through Their Lactic Acid Metabolism

    Tzu-Keng Chiu

    2015-03-01

    Full Text Available This study reports a microfluidic-based optical sensing device for label-free detection of circulating tumor cells (CTCs, a rare cell species in blood circulation. Based on the metabolic features of cancer cells, live CTCs can be quantified indirectly through their lactic acid production. Compared with the conventional schemes for CTC detection, this label-free approach could prevent the biological bias due to the heterogeneity of the surface antigens on cancer cells. In this study, a microfluidic device was proposed to generate uniform water-in-oil cell-encapsulating micro-droplets, followed by the fluorescence-based optical detection of lactic acid produced within the micro-droplets. To test its feasibility to quantify cancer cells, experiments were carried out. Results showed that the detection signals were proportional to the number of cancer cells within the micro-droplets, whereas such signals were insensitive to the existence and number of leukocytes within. To further demonstrate its feasibility for cancer cell detection, the cancer cells with known cell number in a cell suspension was detected based on the method. Results revealed that there was no significant difference between the detected number and the real number of cancer cells. As a whole, the proposed method opens up a new route to detect live CTCs in a label-free manner.

  20. Mechanical Models of Fault-Related Folding

    Johnson, A. M.

    2003-01-09

    The subject of the proposed research is fault-related folding and ground deformation. The results are relevant to oil-producing structures throughout the world, to understanding of damage that has been observed along and near earthquake ruptures, and to earthquake-producing structures in California and other tectonically-active areas. The objectives of the proposed research were to provide both a unified, mechanical infrastructure for studies of fault-related foldings and to present the results in computer programs that have graphical users interfaces (GUIs) so that structural geologists and geophysicists can model a wide variety of fault-related folds (FaRFs).