Assay of hybrid ribonuclease using a membrane filter-immobilized synthetic hybrid: application to the human leukemic cell

International Nuclear Information System (INIS)

Papaphilis, A.D.; Kamper, E.F.

1985-01-01

A method for assaying hybrid ribonuclease has been devised which utilizes as substrate the synthetic hybrid [ 3 H]polyriboadenylic acid [poly(rA)]:polydeoxythymidylic acid [poly(dT)] immobilized on the solid matrix of nitrocellulose filters. The hybridization on filter of [ 3 H]poly(rA) to poly(dT) has been explored in terms of efficacy of the process and the response of the product to RNase H. A pulse of uv irradiation of poly(dT) while in dry state on the filter increased its firm binding to the filter in a concentration-dependent manner, resulting in a concomitant increase of the yield of hybrid formation. The filter-immobilized hybrid was 95% resistant to RNase A but sensitive to RNase H. When stored in toluene in the cold the hybrid maintained its stability for over 6 months, as judged by its resistance to RNase A. The method offers a number of advantages over assays that use solution hybrids as substrates and was readily applicable in the screening of leukemic patients, in the leukocytes of which it has demonstrated increased RNase H levels

A rapid and efficient branched DNA hybridization assay to titer lentiviral vectors.

Science.gov (United States)

Nair, Ayyappan; Xie, Jinger; Joshi, Sarasijam; Harden, Paul; Davies, Joan; Hermiston, Terry

2008-11-01

A robust assay to titer lentiviral vectors is imperative to qualifying their use in drug discovery, target validation and clinical applications. In this study, a novel branched DNA based hybridization assay was developed to titer lentiviral vectors by quantifying viral RNA genome copy numbers from viral lysates without having to purify viral RNA, and this approach was compared with other non-functional (p24 protein ELISA and viral RT-qPCR) and a functional method (reporter gene expression) used commonly. The RT-qPCR method requires purification of viral RNA and the accuracy of titration therefore depends on the efficiency of purification; this requirement is ameliorated in the hybridization assay as RNA is measured directly in viral lysates. The present study indicates that the hybridization based titration assay performed on viral lysates was more accurate and has additional advantages of being rapid, robust and not dependent on transduction efficiency in different cell types.

Detection of high-risk subtypes of human papillomavirus in cervical swabs: routine use of the Digene Hybrid Capture assay and polymerase chain reaction analysis.

LENUS (Irish Health Repository)

Brennan, M M

2012-02-03

Human papillomaviruses (HPVs) are major causative agents in the pathogenesis of cervical cancer, and more than twenty types are associated with its development. With the introduction of liquid-based preparation systems, it is envisaged that large-scale HPV testing will be established in the near future. Preliminary studies demonstrate the accessibility of these samples for DNA testing using both the Digene Hybrid Capture assay (DHCA) and polymerase chain reaction (PCR) techniques. This study aims to assess the validity and sensitivity of the DHCA system to detect high-risk HPV DNA, using two sets of HPV consensus primers (Gp5+\\/Gp6+ and MY09\\/MY11) in tandem with routine assessment of cervical smear and biopsy samples. Results indicate that the combination of DHCA and PCR detects more high-grade lesions than does the DHCA alone. DHCA-negative cases were categorised by subsequent PCR amplification into low-grade HPV-negative (12\\/16) cervical lesions and high-grade HPV-positive (7\\/9) cervical lesions. Gp5+\\/Gp6+ primers were less sensitive in detecting HPV-positive samples than was the MY09\\/MY11 primer set. These results support the use of high-risk HPV testing by DHCA, with subsequent analysis of DHCA-negative samples by PCR using the MY09\\/MY11 primers.

Appendix: a solution hybridization assay to detect radioactive globin messenger RNA nucleotide sequences

Energy Technology Data Exchange (ETDEWEB)

Ross, J

1976-09-15

In view of the sensitivity and specificity of the solution hybridization assay for unlabeled globin mRNA a similar technique has been devised to detect radioactive globin mRNA sequences with unlabeled globin cDNA. Several properties of the hybridization reaction are presented since RNA kinetic experiments reported recently depend on the validity of this assay. Data on hybridization analysis of (/sup 3/H)RNA from mouse fetal liver or erythroleukemia cell cytoplasm are presented. These data indicate that the excess cDNA solution assay for radioactive globin mRNA detection is specific for globin mRNA sequences. It can be performed rapidly and is highly reproducible from experiment. It is at least 500-fold less sensitive than the assay for unlabeled globin mRNA, due to the RNAase backgrounds of 0.05 to 0.15 %. However, this limitation has not affected kinetic experiments with non-dividing fetal liver erythroid cells, which synthesize relatively large quantities of globin mRNA.

Diagnosis of Fasciola gigantica infection in cattle using capture-ELISA assay for detecting antigen in faeces

Directory of Open Access Journals (Sweden)

Sarwitri Endah Estuningsih

2006-10-01

Full Text Available Capture-ELISA assay is a diagnosis for antigen detection in the serum or faeces using polyclonal or monoclonal antibodies. The purpose of this study was to determine the sensitivity and specificity of capture-ELISA assay using polyclonal antibody for diagnosing Fasciola gigantica infection in cattle by detecting antigen in the faeces. In this study, faecal samples and livers were collected from 141 cattle slaughtered in the abattoir in Jakarta. From each animal, liver was processed for liver flukes count and the corresponding faecal sample was analysed for coproantigen. The result of capture-ELISA assay for antigen detection showed that from 85 cattle infected with Fasciola gigantica, 83 had OD > 0.52 (range from 0.52-1.39 and 2 cattle had OD < 0.52 (range from 0.17-0.51. The sensitivity and specificity of the assay were 97.6% and 92.8% respectively. The assay also able to detect 50 ng/ml of antigen in faecal supernatant. It suggests that this assay will have the advantage over the other methods on its ability to detect the active infection. Collection of faeces, rather than serum, will allow a more cost-effective and adaptable method.

Continuously tunable nucleic acid hybridization probes.

Science.gov (United States)

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

2015-12-01

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

Referral population studies underestimate differences between human papillomavirus assays in primary cervical screening

DEFF Research Database (Denmark)

Rebolj, M.; Njor, S.; Lynge, E.

2017-01-01

with SurePath® cytology, and Hybrid Capture 2 (HC2), cobas, CLART and APTIMA HPV assays. Women with positive test results were offered a follow-up. For all detected HPV infections and HPV-positive high-grade cervical intraepithelial neoplasia (≥CIN2), we studied the distributions of assay-specific signal...

Use of neutron-capture plastic fibers for nondestructive assay

International Nuclear Information System (INIS)

Heger, A.S.; Grazioso, R.F.; Mayo, D.R.; Ensslin, N.; Miller, M.C.; Huang, H.Y.; Russo, P.A.

1998-01-01

Neutron-capture plastic fibers can be used as a nondestructive assay tool. The detectors consist of an active region assembled from ribbons of boron-( 10 B) loaded optical fibers. The mixture of the moderator and thermal neutron absorber in the fiber yields a detector with high efficiency (var-epsilon) and a short die-away time (τ). The deposited energy of the resultant charged particles is converted to light that is collected by photomultiplier tubes mounted at both ends of the fiber. Thermal neutron coincidence counters (TNCC) made of these fibers can serve to verify fissile materials generated from the nuclear fuel cycle. This type of detector may extend the range of materials now accessible to assay by 3 He detectors. Experiments with single fibers of diameters 0.25, 0.50, and 1.00 mm test their ability to distinguish between the signals generated from neutron interactions and those from gamma rays. These results are compared with those obtained from simulation analyses for the same purpose. Light output and attenuation, neutron detection efficiency, and the signal-to-noise ratios of these fibers have also been investigated. The experimental results for light attenuation and neutron detection efficiency are consistent with the values obtained from simulation studies. A comparison of the performance of various configurations of the plastic scintillating fibers with that of other neutron-capture devices such as 3 He detectors is also discussed

Human papillomavirus testing in primary cervical screening and the cut-off level for hybrid capture 2 tests

DEFF Research Database (Denmark)

Rebolj, Matejka; Bonde, Jesper; Njor, Sisse Helle

2011-01-01

To determine the trade-off between the sensitivity and the specificity for high grade cervical intraepithelial neoplasia at hybrid capture 2 cut-off values above the standard = 1 relative light units/cut-off level (rlu/co).......To determine the trade-off between the sensitivity and the specificity for high grade cervical intraepithelial neoplasia at hybrid capture 2 cut-off values above the standard = 1 relative light units/cut-off level (rlu/co)....

Oligonucleotide PIK3CA/Chromosome 3 Dual in Situ Hybridization Automated Assay with Improved Signals, One-Hour Hybridization, and No Use of Blocking DNA.

Science.gov (United States)

Zhang, Wenjun; Hubbard, Antony; Baca-Parkinson, Leslie; Stanislaw, Stacey; Vladich, Frank; Robida, Mark D; Grille, James G; Maxwell, Daniel; Tsao, Tsu-Shuen; Carroll, William; Gardner, Tracie; Clements, June; Singh, Shalini; Tang, Lei

2015-09-01

The PIK3CA gene at chromosome 3q26.32 was found to be amplified in up to 45% of patients with squamous cell carcinoma of the lung. The strong correlation between PIK3CA amplification and increased phosphatidylinositol 3-kinase (PI3K) pathway activities suggested that PIK3CA gene copy number is a potential predictive biomarker for PI3K inhibitors. Currently, all microscopic assessments of PIK3CA and chromosome 3 (CHR3) copy numbers use fluorescence in situ hybridization. PIK3CA probes are derived from bacterial artificial chromosomes whereas CHR3 probes are derived mainly from the plasmid pHS05. These manual fluorescence in situ hybridization assays mandate 12- to 18-hour hybridization and use of blocking DNA from human sources. Moreover, fluorescence in situ hybridization studies provide limited morphologic assessment and suffer from signal decay. We developed an oligonucleotide-based bright-field in situ hybridization assay that overcomes these shortcomings. This assay requires only a 1-hour hybridization with no need for blocking DNA followed by indirect chromogenic detection. Oligonucleotide probes produced discrete and uniform CHR3 stains superior to those from the pHS05 plasmid. This assay achieved successful staining in 100% of the 195 lung squamous cell carcinoma resections and in 94% of the 33 fine-needle aspirates. This robust automated bright-field dual in situ hybridization assay for the simultaneous detection of PIK3CA and CHR3 centromere provides a potential clinical diagnostic method to assess PIK3CA gene abnormality in lung tumors. Copyright © 2015 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

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

Science.gov (United States)

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

2013-02-05

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

A novel SERRS sandwich-hybridization assay to detect specific DNA target.

Directory of Open Access Journals (Sweden)

Cécile Feuillie

Full Text Available In this study, we have applied Surface Enhanced Resonance Raman Scattering (SERRS technology to the specific detection of DNA. We present an innovative SERRS sandwich-hybridization assay that allows specific DNA detection without any enzymatic amplification, such as is the case with Polymerase Chain Reaction (PCR. In some substrates, such as ancient or processed remains, enzymatic amplification fails due to DNA alteration (degradation, chemical modification or to the presence of inhibitors. Consequently, the development of a non-enzymatic method, allowing specific DNA detection, could avoid long, expensive and inconclusive amplification trials. Here, we report the proof of concept of a SERRS sandwich-hybridization assay that leads to the detection of a specific chamois DNA. This SERRS assay reveals its potential as a non-enzymatic alternative technology to DNA amplification methods (particularly the PCR method with several applications for species detection. As the amount and type of damage highly depend on the preservation conditions, the present SERRS assay would enlarge the range of samples suitable for DNA analysis and ultimately would provide exciting new opportunities for the investigation of ancient DNA in the fields of evolutionary biology and molecular ecology, and of altered DNA in food frauds detection and forensics.

Luminescence resonance energy transfer-based nucleic acid hybridization assay on cellulose paper with upconverting phosphor as donors.

Science.gov (United States)

Zhou, Feng; Noor, M Omair; Krull, Ulrich J

2014-03-04

A bioassay based on DNA hybridization on cellulose paper is a promising format for gene fragment detection that may be suited for in-field and rapid diagnostic applications. We demonstrate for the first time that luminescence resonance energy transfer (LRET) associated with upconverting phosphors (UCPs) can be used to develop a paper-based DNA hybridization assay with high sensitivity, selectivity and fast response. UCPs with strong green emission were synthesized and subsequently functionalized with streptavidin (UCP-strep). UCP-strep particles were immobilized on cellulose paper, and then biotinylated single-stranded oligonucleotide probes were conjugated onto the UCPs via streptavidin-biotin linkage. The UCPs served as donors that were LRET-paired with Cy3-labeled target DNA. Selective DNA hybridization enabled the proximity required for LRET-sensitized emission from Cy3, which was used as the detection signal. Hybridization was complete within 2 min, and the limit of detection of the method was 34 fmol, which is a significant improvement in comparison to an analogous fluorescence resonance energy transfer (FRET) assay based on quantum dots. The assay exhibited excellent resistance to nonspecific adsorption of noncomplementary short/long DNA and protein. The selectivity of the assay was further evaluated by one base pair mismatched (1BPM) DNA detection, where a maximum signal ratio of 3.1:1 was achieved between fully complementary and 1BPM samples. This work represents a preliminary but significant step for the development of paper-based UCP-LRET nucleic acid hybridization assays, which offer potential for lowering the limit of detection of luminescent hybridization assays due to the negligible background signal associated with optical excitation by near-infrared (NIR) light.

Comparison of analytical and clinical performance of CLART HPV2 genotyping assay to Linear Array and Hybrid Capture 2

DEFF Research Database (Denmark)

Ejegod, Ditte Møller; Rebolj, Matejka; Bonde, Jesper

2015-01-01

to the Danish nation-wide Pathology Data Bank. For comparison of CLART and LA in terms of genotype detection, we calculated κ-coefficients, and proportions of overall and positive agreement. For comparison of CIN detection between CLART, LA, and HC2, we calculated the relative sensitivity and specificity......), and Hybrid Capture 2 (HC2) using samples stored in SurePath. METHODS: Residual material from 401 routine samples from women with abnormal cytology was tested by CLART, LA, and HC2 (ClinicalTrial.gov: NCT01671462, Ethical Committee approval: H-2012-070). Histological outcomes were ascertained by linkage...... for high-grade CIN. RESULTS: The κ-coefficient for agreement in detection of genotypes 16, 18, 31, 33, 35, and 51 was ≥0.90 (overall agreement: 98-99%, positive agreement: 84-95%). The values were slightly lower, but still in the "substantial" range for genotypes 39, 45, 52, 56, 58, 59, and several low...

Use of Molecular Assays in Diagnosis and Monitoring of Cytomegalovirus Disease following Renal Transplantation

OpenAIRE

Aitken, Celia; Barrett-Muir, Winsome; Millar, Colin; Templeton, Kate; Thomas, Janice; Sheridan, Fran; Jeffries, Donald; Yaqoob, Magdi; Breuer, Judith

1999-01-01

We compared two commercial molecular assays (the Murex Hybrid Capture CMV DNA assay [HCA], version 2, and the Roche Amplicor plasma PCR assay) with a standard shell vial assay in detecting and predicting cytomegalovirus (CMV) disease in a group of renal transplant patients and assessed the role of viral load measurements (using the HCA) in their management. The sensitivity of the HCA and Amplicor assay in terms of disease detection was 100%, compared to 71% for the shell vial assay. Both the ...

Detecting West Nile virus in owls and raptors by an antigen-capture assay.

Science.gov (United States)

Gancz, Ady Y; Campbell, Douglas G; Barker, Ian K; Lindsay, Robbin; Hunter, Bruce

2004-12-01

We evaluated a rapid antigen-capture assay (VecTest) for detection of West Nile virus in oropharyngeal and cloacal swabs, collected at necropsy from owls (N = 93) and raptors (N = 27). Sensitivity was 93.5%-95.2% for northern owl species but raptors.

Detecting West Nile Virus in Owls and Raptors by an Antigen-capture Assay

OpenAIRE

Gancz, Ady Y.; Campbell, Douglas G.; Barker, Ian K.; Lindsay, Robbin; Hunter, Bruce

2004-01-01

We evaluated a rapid antigen-capture assay (VecTest) for detection of West Nile virus in oropharyngeal and cloacal swabs, collected at necropsy from owls (N = 93) and raptors (N = 27). Sensitivity was 93.5%–95.2% for northern owl species but

Conversion of a Capture ELISA to a Luminex xMAP Assay using a Multiplex Antibody Screening Method

Science.gov (United States)

Baker, Harold N.; Murphy, Robin; Lopez, Erica; Garcia, Carlos

2012-01-01

The enzyme-linked immunosorbent assay (ELISA) has long been the primary tool for detection of analytes of interest in biological samples for both life science research and clinical diagnostics. However, ELISA has limitations. It is typically performed in a 96-well microplate, and the wells are coated with capture antibody, requiring a relatively large amount of sample to capture an antigen of interest . The large surface area of the wells and the hydrophobic binding of capture antibody can also lead to non-specific binding and increased background. Additionally, most ELISAs rely upon enzyme-mediated amplification of signal in order to achieve reasonable sensitivity. Such amplification is not always linear and can thus skew results. In the past 15 years, a new technology has emerged that offers the benefits of the ELISA, but also enables higher throughput, increased flexibility, reduced sample volume, and lower cost, with a similar workflow 1, 2. Luminex xMAP Technology is a microsphere (bead) array platform enabling both monoplex and multiplex assays that can be applied to both protein and nucleic acid applications 3-5. The beads have the capture antibody covalently immobilized on a smaller surface area, requiring less capture antibody and smaller sample volumes, compared to ELISA, and non-specific binding is significantly reduced. Smaller sample volumes are important when working with limiting samples such as cerebrospinal fluid, synovial fluid, etc. 6. Multiplexing the assay further reduces sample volume requirements, enabling multiple results from a single sample. Recent improvements by Luminex include: the new MAGPIX system, a smaller, less expensive, easier-to-use analyzer; Low-Concentration Magnetic MagPlex Microspheres which eliminate the need for expensive filter plates and come in a working concentration better suited for assay development and low-throughput applications; and the xMAP Antibody Coupling (AbC) Kit, which includes a protocol, reagents, and

Evaluation of a reverse-hybridization StripAssay for the detection of genetic polymorphisms leading to acenocoumarol sensitivity.

Science.gov (United States)

Gialeraki, Argyri; Markatos, Christos; Grouzi, Elisabeth; Merkouri, Efrosyni; Travlou, Anthi; Politou, Marianna

2010-04-01

Acenocoumarol is mainly catabolized by CYP2C9 isoform of cytochrome P450 (CYP) liver complex and exerts its anticoagulant effect through the inhibition of Vitamin K Epoxide Reductase (VKOR). The most important genetic polymorphisms which lead to an impaired enzymatic activity and therefore predispose to acenocoumarol sensitivity, are considered to be CYP2C9*2 (Arg144Cys), CYP2C9*3 (Ile359Leu) and VKORC1-1639G>A, respectively. In this study we compared the results of the PGXThrombo StripAssay kit (ViennaLab Diagnostics,Vienna, Austria) with direct DNA sequencing and in house Restriction Fragment Length Polymorphisms (RFLP) for the detection of the aforementioned Single Nucleotide Polymorphisms (SNPs). The reverse hybridization StripAssay was found to be equally effective with RFLP and direct DNA sequencing for the detection of CYP2C9*2 and CYP2C9*3 polymorphisms, respectively. The comparison of the RFLP reference method with the reverse hybridization StripAssay for the detection of VKORC1-1639 G>A polymorphism showed that the reverse hybridization StripAsssay might misclassify some A/A homozygotes as heterozygotes. Optimization of the hybridization procedures may eliminate the extra low signal band observed in some samples at the reverse hybridization StripAssay and improve its diagnostic value.

Concordant testing results between various Human Papillomavirus assays in primary cervical cancer screening

DEFF Research Database (Denmark)

de Thurah, Lena; Bonde, Jesper; Hoa Lam, Janni Uyen

2018-01-01

OBJECTIVES: Human Papillomavirus (HPV) assays are increasingly used for primary cervical screening and HPV vaccination effect monitoring. We undertook a systematic literature review to determine the concordance in positive test results (i.e., detection of HPV infections) between Hybrid Capture 2 ...

A two-hybrid assay to study protein interactions within the secretory pathway.

Directory of Open Access Journals (Sweden)

Danielle H Dube

Full Text Available Interactions of transcriptional activators are difficult to study using transcription-based two-hybrid assays due to potent activation resulting in false positives. Here we report the development of the Golgi two-hybrid (G2H, a method that interrogates protein interactions within the Golgi, where transcriptional activators can be assayed with negligible background. The G2H relies on cell surface glycosylation to report extracellularly on protein-protein interactions occurring within the secretory pathway. In the G2H, protein pairs are fused to modular domains of the reporter glycosyltransferase, Och1p, and proper cell wall formation due to Och1p activity is observed only when a pair of proteins interacts. Cells containing interacting protein pairs are identified by selectable phenotypes associated with Och1p activity and proper cell wall formation: cells that have interacting proteins grow under selective conditions and display weak wheat germ agglutinin (WGA binding by flow cytometry, whereas cells that lack interacting proteins display stunted growth and strong WGA binding. Using this assay, we detected the interaction between transcription factor MyoD and its binding partner Id2. Interfering mutations along the MyoD:Id2 interaction interface ablated signal in the G2H assay. Furthermore, we used the G2H to detect interactions of the activation domain of Gal4p with a variety of binding partners. Finally, selective conditions were used to enrich for cells encoding interacting partners. The G2H detects protein-protein interactions that cannot be identified via traditional two-hybrid methods and should be broadly useful for probing previously inaccessible subsets of the interactome, including transcriptional activators and proteins that traffic through the secretory pathway.

[Correlation of hybrid II capture cytologic exam in diagnosis of cervical lesions related to HPV].

Science.gov (United States)

Saturnino, Ana Conceição Ribeiro Dantas; Diniz, Veruska Cassandra; Sisenando, Herbert Ary Arzabe Anteza Costa Nóbrega; de Souza, Eroisa Maria Conceição Oliveira; de Paiva, Magnus Sérgio Martins; de Brito, Maria Helena Marques Fonseca; Ramos, Eleni Souto Nóbrega

2005-01-01

The aim of the present study was confront the results of the cytological examination with hybrid capture II in the diagnosis of induced cervical intraepithelial lesion-HPV, correlating the cytological findings with biomoleculares. The research was carried through in a group of 160 sexually active women who had espontaneamente looked its gynecologists for consultation of routine, having been submitted to the collection of cervicovaginal material for cytology and for examination of hybrid capture II in the Centro de Patologia Clínica and the Hospital e Maternidade Promater, in the city of the Natal-RN. The results had shown to relatively high numbers of positive cases for HPV using hybrid capture II (41.87%) and the cytology (23.75%). The agreement between the two studied methods relatively was raised (59.38%). It was evident also that the viruses with high oncogênico potential had presented found in the compatible cytology with Lesion of low risk (11.88%), followed of Lesion of high risk (NIC II and III); already the viruses with low oncogênico potential were more associates the Lesion of low risk (6.25%), followed of Lesion of high risk. The cytology, exactly with its limitations, is an important method in the detention of attributable patologias to the HPV, emphasizing that the molecular method comes to complement it and to consolidate the cytological findings.

Preliminary carbon dioxide capture technical and economic feasibility study evaluation of carbon dioxide capture from existing fired plants by hybrid sorption using solid sorbents

Energy Technology Data Exchange (ETDEWEB)

Benson, Steven; Envergex, Srivats; Browers, Bruce; Thumbi, Charles

2013-01-01

Barr Engineering Co. was retained by the Institute for Energy Studies (IES) at University of North Dakota (UND) to conduct a technical and economic feasibility analysis of an innovative hybrid sorbent technology (CACHYS™) for carbon dioxide (CO2) capture and separation from coal combustion–derived flue gas. The project team for this effort consists of the University of North Dakota, Envergex LLC, Barr Engineering Co., and Solex Thermal Science, along with industrial support from Allete, BNI Coal, SaskPower, and the North Dakota Lignite Energy Council. An initial economic and feasibility study of the CACHYS™ concept, including definition of the process, development of process flow diagrams (PFDs), material and energy balances, equipment selection, sizing and costing, and estimation of overall capital and operating costs, is performed by Barr with information provided by UND and Envergex. The technology—Capture from Existing Coal-Fired Plants by Hybrid Sorption Using Solid Sorbents Capture (CACHYS™)—is a novel solid sorbent technology based on the following ideas: reduction of energy for sorbent regeneration, utilization of novel process chemistry, contactor conditions that minimize sorbent-CO2 heat of reaction and promote fast CO2 capture, and a low-cost method of heat management. The technology’s other key component is the use of a low-cost sorbent.

Sequence capture by hybridization to explore modern and ancient genomic diversity in model and nonmodel organisms.

Science.gov (United States)

Gasc, Cyrielle; Peyretaillade, Eric; Peyret, Pierre

2016-06-02

The recent expansion of next-generation sequencing has significantly improved biological research. Nevertheless, deep exploration of genomes or metagenomic samples remains difficult because of the sequencing depth and the associated costs required. Therefore, different partitioning strategies have been developed to sequence informative subsets of studied genomes. Among these strategies, hybridization capture has proven to be an innovative and efficient tool for targeting and enriching specific biomarkers in complex DNA mixtures. It has been successfully applied in numerous areas of biology, such as exome resequencing for the identification of mutations underlying Mendelian or complex diseases and cancers, and its usefulness has been demonstrated in the agronomic field through the linking of genetic variants to agricultural phenotypic traits of interest. Moreover, hybridization capture has provided access to underexplored, but relevant fractions of genomes through its ability to enrich defined targets and their flanking regions. Finally, on the basis of restricted genomic information, this method has also allowed the expansion of knowledge of nonreference species and ancient genomes and provided a better understanding of metagenomic samples. In this review, we present the major advances and discoveries permitted by hybridization capture and highlight the potency of this approach in all areas of biology. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Evaluation of Myc Gene Amplification in Prostate Cancer Using a Dual Color Chromogenic in-Situ Hybridization (Dual CISH) Assay

OpenAIRE

Daniel Lerda; Marta Cabrera; Jorge Flores; Luis Gutierrez; Armando Chierichetti; Martin Revol; Hernan Garcia Onto

2013-01-01

Objetive: The overall purpose of the study was to demonstrate applicability of the Dako dual-color chromogenic in situ hybridization (CISH) assay (DAKO Denmark, Glostrup) with respect to fluorescence in situ hybridization (FISH) probes MYC-C. Methods: MYC gene amplification by FISH and Dako dual-color CISH Results: The study showed that the dual-color CISH assay can convert Texas red and fluorescein isothiocyanate (FITC) signals into chromogenic signals. The dual –color CISH assay was p...

Standardization of a cytometric p24-capture bead-assay for the detection of main HIV-1 subtypes.

Science.gov (United States)

Merbah, Mélanie; Onkar, Sayali; Grivel, Jean-Charles; Vanpouille, Christophe; Biancotto, Angélique; Bonar, Lydia; Sanders-Buell, Eric; Kijak, Gustavo; Michael, Nelson; Robb, Merlin; Kim, Jerome H; Tovanabutra, Sodsai; Chenine, Agnès-Laurence

2016-04-01

The prevailing method to assess HIV-1 replication and infectivity is to measure the production of p24 Gag protein by enzyme-linked immunosorbent assay (ELISA). Since fluorescent bead-based technologies offer a broader dynamic range and higher sensitivity, this study describes a p24 capture Luminex assay capable of detecting HIV-1 subtypes A-D, circulating recombinant forms (CRF) CRF01_AE and CRF02_AG, which together are responsible for over 90% of HIV-1 infections worldwide. The success of the assay lies in the identification and selection of a cross-reactive capture antibody (clone 183-H12-5C). Fifty-six isolates that belonged to six HIV-1 subtypes and CRFs were successfully detected with p-values below 0.021; limits of detection ranging from 3.7 to 3 × 104 pg/ml. The intra- and inter-assay variation gave coefficient of variations below 6 and 14%, respectively. The 183-bead Luminex assay also displayed higher sensitivity of 91% and 98% compared to commercial p24 ELISA and a previously described Luminex assay. The p24 concentrations measured by the 183-bead Luminex assay showed a significant correlation (R=0.92, passay leverages the advantages of the Luminex platform, which include smaller sample volume and simultaneous detection of up to 500 analytes in a single sample, and delivers a valuable tool for the field. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparison of the cobas Human Papillomavirus (HPV) Test with the Hybrid Capture 2 and Linear Array HPV DNA Tests

Science.gov (United States)

Sadorra, Mark; LaMere, Brandon J.; Kail, Randi; Aldrich, Carrie; Kinney, Walter; Fetterman, Barbara; Lorey, Thomas; Schiffman, Mark; Castle, Philip E.

2012-01-01

The cobas human papillomavirus (HPV) test (cobas) was recently approved by the U.S. Food and Drug Administration (FDA) and identifies HPV16 and HPV18 separately as well as detecting a pool of 11 HR-HPV genotypes (HPV31, -33, -35, -39, -45, -51, -52, -56, -58, -59, -68) and also HPV66. We compared cobas, Linear Array (LA), and Hybrid Capture 2 (HC2) assays for detection of carcinogenic HPV DNA, and cobas and LA for detection of HPV16 and HPV18 DNA, among the first 1,852 women enrolled in the HPV Persistence and Progression Cohort (PaP Cohort) study. Specimens were tested by all 3 assays 1 year after an HC2-positive result. In 1,824 specimens with cobas results, cobas had an 85.9% agreement with HC2 and 91.0% agreement with LA for carcinogenic HPV detection. When results between cobas and HC2 disagreed, cobas tended to call more women HPV positive (P < 0.01). Categorizing cobas and LA results hierarchically according to cancer risk (HPV16, HPV18, other carcinogenic HPV genotypes, or carcinogen negative), there was a 90% agreement for all categories of HPV (n = 1,824). We found good agreement between the two U.S. FDA-approved HPV tests, with discrepancies between the two assays due to specific characteristics of the individual assays. Additional studies are needed to compare HC2 and cobas for detecting and predicting CIN3 to understand the clinical implications of the discrepant test results between the two tests. PMID:22075592

On-chip multiplexed solid-phase nucleic acid hybridization assay using spatial profiles of immobilized quantum dots and fluorescence resonance energy transfer.

Science.gov (United States)

Noor, M Omair; Tavares, Anthony J; Krull, Ulrich J

2013-07-25

A microfluidic based solid-phase assay for the multiplexed detection of nucleic acid hybridization using quantum dot (QD) mediated fluorescence resonance energy transfer (FRET) is described herein. The glass surface of hybrid glass-polydimethylsiloxane (PDMS) microfluidic channels was chemically modified to assemble the biorecognition interface. Multiplexing was demonstrated using a detection system that was comprised of two colors of immobilized semi-conductor QDs and two different oligonucleotide probe sequences. Green-emitting and red-emitting QDs were paired with Cy3 and Alexa Fluor 647 (A647) labeled oligonucleotides, respectively. The QDs served as energy donors for the transduction of dye labeled oligonucleotide targets. The in-channel assembly of the biorecognition interface and the subsequent introduction of oligonucleotide targets was accomplished within minutes using a combination of electroosmotic flow and electrophoretic force. The concurrent quantification of femtomole quantities of two target sequences was possible by measuring the spatial coverage of FRET sensitized emission along the length of the channel. In previous reports, multiplexed QD-FRET hybridization assays that employed a ratiometric method for quantification had challenges associated with lower analytical sensitivity arising from both donor and acceptor dilution that resulted in reduced energy transfer pathways as compared to single-color hybridization assays. Herein, a spatial method for quantification that is based on in-channel QD-FRET profiles provided higher analytical sensitivity in the multiplexed assay format as compared to single-color hybridization assays. The selectivity of the multiplexed hybridization assays was demonstrated by discrimination between a fully-complementary sequence and a 3 base pair sequence at a contrast ratio of 8 to 1. Copyright © 2013 Elsevier B.V. All rights reserved.

Comparison of kDNA PCR-hybridization assay with three PCR methods for canines visceral Leishmaniasis diagnosis

Energy Technology Data Exchange (ETDEWEB)

Pilatti, Marcia M.; Andrade, Antero S.R. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil)], e-mail: marciapilatti@yahoo.com.br, e-mail: antero@cdtn.br; Ferreira, Sidney A. [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Parasitologia], e-mail: saninoalmeida@gmail.com

2009-07-01

The sensitivity of the kDNA PCR-Hybridization assay, which uses radioactive DNA probes (labeled with {sup 32}P), was compared with three conventional PCR methods used for canine visceral leishmaniasis diagnosis. All PCR methods had two steps: a first amplification followed by hybridization or by a new amplification (nested or semi nested). Two methods (kDNA PCR-Hybridization and kDNA snPCR) used primers addressed to kinetoplast minicircles and the other two methods to the coding (LnPCR) and intergenic noncoding regions (ITS-1 nPCR) of the ribosomal rRNA genes. The comparison was accomplished in two groups of 23 infected dogs using samples collected by the conjunctival swab procedure. In the Group 1 the DNA was extracted from cotton swabs by phenol-chloroform and in Group 2 by boiling. The most efficient PCR methods in the Group 1 were those based on kDNA targets. The kDNA PCR-Hybridization was able to detect parasites in 22/23 dogs (95.6%) and in 40/46 samples (86.9%). The kDNA snPCR was positive for 21/23 dogs (91.3%) and for 40/46 samples (86.9%). The positivities of the kDNA based methods were significantly higher than the positivities verified for the methods based on ribosomal rRNA genes (p<0.05). In the Group 2 the kDNA PCR- Hybridization showed a better performance detecting parasites in 18/23 dogs (78.3%) and in 31/46 samples (67.4%), significantly higher than the other three methods (p<0.05). The higher sensitivity of the minicircle kDNA based assays reported by others was confirmed in this study and kDNA PCR-Hybridization showed the best sensitivity among the assays evaluated. (author)

Comparison of kDNA PCR-hybridization assay with three PCR methods for canines visceral Leishmaniasis diagnosis

International Nuclear Information System (INIS)

Pilatti, Marcia M.; Andrade, Antero S.R.; Ferreira, Sidney A.

2009-01-01

The sensitivity of the kDNA PCR-Hybridization assay, which uses radioactive DNA probes (labeled with 32 P), was compared with three conventional PCR methods used for canine visceral leishmaniasis diagnosis. All PCR methods had two steps: a first amplification followed by hybridization or by a new amplification (nested or semi nested). Two methods (kDNA PCR-Hybridization and kDNA snPCR) used primers addressed to kinetoplast minicircles and the other two methods to the coding (LnPCR) and intergenic noncoding regions (ITS-1 nPCR) of the ribosomal rRNA genes. The comparison was accomplished in two groups of 23 infected dogs using samples collected by the conjunctival swab procedure. In the Group 1 the DNA was extracted from cotton swabs by phenol-chloroform and in Group 2 by boiling. The most efficient PCR methods in the Group 1 were those based on kDNA targets. The kDNA PCR-Hybridization was able to detect parasites in 22/23 dogs (95.6%) and in 40/46 samples (86.9%). The kDNA snPCR was positive for 21/23 dogs (91.3%) and for 40/46 samples (86.9%). The positivities of the kDNA based methods were significantly higher than the positivities verified for the methods based on ribosomal rRNA genes (p<0.05). In the Group 2 the kDNA PCR- Hybridization showed a better performance detecting parasites in 18/23 dogs (78.3%) and in 31/46 samples (67.4%), significantly higher than the other three methods (p<0.05). The higher sensitivity of the minicircle kDNA based assays reported by others was confirmed in this study and kDNA PCR-Hybridization showed the best sensitivity among the assays evaluated. (author)

Clinical and analytical performance of the BD Onclarity™ HPV assay for detection of CIN2+ lesions on SurePath samples

DEFF Research Database (Denmark)

Ejegod, Ditte Møller; Junge, Jette; Franzmann, Maria

2016-01-01

BACKGROUND: The novel BD Onclarity(TM) HPV assay (Onclarity) on the BD Viper™ LT system (BD Diagnostics, Sparks, MD), detects E6/E7 DNA from 13 high-risk HPV genotypes and HPV66. We compared the analytical and clinical performance of the Onclarity Assay to that of Hybrid Capture 2 and LINEAR ARRAY...

Correlation between atomic negative muon capture and electron distribution in organic sp2-hybridization compounds CxHyClz

International Nuclear Information System (INIS)

Sakai, Yoichi; Tominaga, Takeshi; Ikuta, Shigeru

1986-01-01

The atomic negative muon capture ratios determined experimentally in organic sp 2 -hybridization compound, C x H y Cl z , were compared with the electron populations of carbon atomic orbitals obtained by an ab initio molecular orbital calculation in such systems. A clear positive correlation was found between the C 2s and C 2pz populations and the negative muon capture ratio A (C/Cl), suggesting the mesomolecular process in the initial stage of muon capture. (orig.)

Effect of SO 2 on CO 2 Capture Using Liquid-like Nanoparticle Organic Hybrid Materials

KAUST Repository

Lin, Kun-Yi Andrew

2013-08-15

Liquid-like nanoparticle organic hybrid materials (NOHMs), consisting of silica nanoparticles with a grafted polymeric canopy, were synthesized. Previous work on NOHMs has revealed that CO2 capture behaviors in these hybrid materials can be tuned by modifying the structure of the polymeric canopy. Because SO2, which is another acidic gas found in flue gas, would also interact with NOHMs, this study was designed to investigate its effect on CO2 capture in NOHMs. In particular, CO2 capture capacities as well as swelling and CO2 packing behaviors of NOHMs were analyzed using thermogravimetric analyses and Raman and attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopies before and after exposure of NOHMs to SO2. It was found that the SO2 absorption in NOHMs was only prominent at high SO2 levels (i.e., 3010 ppm; Ptot = 0.4 MPa) far exceeding the typical SO2 concentration in flue gas. As expected, the competitive absorption between SO2 and CO2 for the same absorption sites (i.e., ether and amine groups) resulted in a decreased CO2 capture capacity of NOHMs. The swelling of NOHMs was not notably affected by the presence of SO 2 within the given concentration range (Ptot = 0-0.68 MPa). On the other hand, SO2, owing to its Lewis acidic nature, interacted with the ether groups of the polymeric canopy and, thus, changed the CO2 packing behaviors in NOHMs. © 2013 American Chemical Society.

Effect of SO 2 on CO 2 Capture Using Liquid-like Nanoparticle Organic Hybrid Materials

KAUST Repository

Lin, Kun-Yi Andrew; Petit, Camille; Park, Ah-Hyung Alissa

2013-01-01

Liquid-like nanoparticle organic hybrid materials (NOHMs), consisting of silica nanoparticles with a grafted polymeric canopy, were synthesized. Previous work on NOHMs has revealed that CO2 capture behaviors in these hybrid materials can be tuned by modifying the structure of the polymeric canopy. Because SO2, which is another acidic gas found in flue gas, would also interact with NOHMs, this study was designed to investigate its effect on CO2 capture in NOHMs. In particular, CO2 capture capacities as well as swelling and CO2 packing behaviors of NOHMs were analyzed using thermogravimetric analyses and Raman and attenuated total reflectance (ATR) Fourier transform infrared (FTIR) spectroscopies before and after exposure of NOHMs to SO2. It was found that the SO2 absorption in NOHMs was only prominent at high SO2 levels (i.e., 3010 ppm; Ptot = 0.4 MPa) far exceeding the typical SO2 concentration in flue gas. As expected, the competitive absorption between SO2 and CO2 for the same absorption sites (i.e., ether and amine groups) resulted in a decreased CO2 capture capacity of NOHMs. The swelling of NOHMs was not notably affected by the presence of SO 2 within the given concentration range (Ptot = 0-0.68 MPa). On the other hand, SO2, owing to its Lewis acidic nature, interacted with the ether groups of the polymeric canopy and, thus, changed the CO2 packing behaviors in NOHMs. © 2013 American Chemical Society.

Analysis of hybrid membrane and chemical absorption systems for CO2 capture

International Nuclear Information System (INIS)

Binns, Michael; Oh, Se-Young; Kwak, Dong-Hun; Kim, Jin-Kuk

2015-01-01

Amine-based absorption of CO 2 is currently the industry standard technology for capturing CO 2 emitted from power plants, refineries and other large chemical plants. However, more recently there have been a number of competing technologies under consideration, including the use of membranes for CO 2 separation and purification. We constructed and analyzed two different hybrid configurations combining and connecting chemical absorption with membrane separation. For a particular flue gas which is currently treated with amine-based chemical absorption at a pilot plant we considered and tested how membranes could be integrated to improve the performance of the CO 2 capture. In particular we looked at the CO 2 removal efficiency and the energy requirements. Sensitivity analysis was performed varying the size of the membranes and the solvent flow rate

Identification and characterization of protein interactions in the mammalian mRNA processing body using a novel two-hybrid assay

Energy Technology Data Exchange (ETDEWEB)

Bloch, Donald B., E-mail: bloch@helix.mgh.harvard.edu; Nobre, Rita A.; Bernstein, Gillian A.; Yang, Wei-Hong

2011-09-10

Components of the mRNA processing body (P-body) regulate critical steps in mRNA storage, transport, translation and degradation. At the core of the P-body is the decapping complex, which removes the 5' cap from de-adenylated mRNAs and mediates an irreversible step in mRNA degradation. The assembly of P-bodies in Saccharomyces cerevisiae, Arabidopsis thaliana and Drosophila melanogaster has been previously described. Less is known about the assembly of mammalian P-bodies. To investigate the interactions that occur between components of mammalian P-bodies, we developed a fluorescence-based, two-hybrid assay system. The assay depends on the ability of one P-body component, fused to an exogenous nuclear localization sequence (NLS), to recruit other P-body components to the nucleus. The assay was used to investigate interactions between P-body components Ge-1, DCP2, DCP1, EDC3, RAP55, and RCK. The results of this study show that the modified two-hybrid assay can be used to identify protein interactions that occur in a macromolecular complex. The assay can also be used to efficiently detect protein interaction domains. The results provide important insights into mammalian P-body assembly and demonstrate similarities, and critical differences, between P-body assembly in mammalian cells compared with that of other species. -- Research highlights: {yields} A two-hybrid assay was developed to study interactions in macromolecular complexes. {yields} The assay was applied to interactions between components of mRNA P-bodies. {yields} The assay effectively and efficiently identified protein interaction domains. {yields} P-body assembly in mammalian cells differs from that in other species.

Identification and characterization of protein interactions in the mammalian mRNA processing body using a novel two-hybrid assay

International Nuclear Information System (INIS)

Bloch, Donald B.; Nobre, Rita A.; Bernstein, Gillian A.; Yang, Wei-Hong

2011-01-01

Components of the mRNA processing body (P-body) regulate critical steps in mRNA storage, transport, translation and degradation. At the core of the P-body is the decapping complex, which removes the 5' cap from de-adenylated mRNAs and mediates an irreversible step in mRNA degradation. The assembly of P-bodies in Saccharomyces cerevisiae, Arabidopsis thaliana and Drosophila melanogaster has been previously described. Less is known about the assembly of mammalian P-bodies. To investigate the interactions that occur between components of mammalian P-bodies, we developed a fluorescence-based, two-hybrid assay system. The assay depends on the ability of one P-body component, fused to an exogenous nuclear localization sequence (NLS), to recruit other P-body components to the nucleus. The assay was used to investigate interactions between P-body components Ge-1, DCP2, DCP1, EDC3, RAP55, and RCK. The results of this study show that the modified two-hybrid assay can be used to identify protein interactions that occur in a macromolecular complex. The assay can also be used to efficiently detect protein interaction domains. The results provide important insights into mammalian P-body assembly and demonstrate similarities, and critical differences, between P-body assembly in mammalian cells compared with that of other species. -- Research highlights: → A two-hybrid assay was developed to study interactions in macromolecular complexes. → The assay was applied to interactions between components of mRNA P-bodies. → The assay effectively and efficiently identified protein interaction domains. → P-body assembly in mammalian cells differs from that in other species.

Real-time PCR Detection of Brucella Abortus: A Comparative Study of SYBR Green I, 5'-exonuclease, and Hybridization Probe Assays

Energy Technology Data Exchange (ETDEWEB)

Newby, Deborah Trishelle; Hadfield, Ted; Roberto, Francisco Figueroa

2003-08-01

Real-time PCR provides a means of detecting and quantifying DNA targets by monitoring PCR product accumulation during cycling as indicated by increased fluorescence. A number of different approaches can be used to generate the fluorescence signal. Three approaches—SYBR Green I (a double-stranded DNA intercalating dye), 5'-exonuclease (enzymatically released fluors), and hybridization probes (fluorescence resonance energy transfer)—were evaluated for use in a real-time PCR assay to detect Brucella abortus. The three assays utilized the same amplification primers to produce an identical amplicon. This amplicon spans a region of the B. abortus genome that includes portions of the alkB gene and the IS711 insertion element. All three assays were of comparable sensitivity, providing a linear assay over 7 orders of magnitude (from 7.5 ng down to 7.5 fg). However, the greatest specificity was achieved with the hybridization probe assay.

Prevalence of Human Papillomavirus Infection in Unselected SurePath Samples Using the APTIMA HPV mRNA Assay

DEFF Research Database (Denmark)

Rebolj, Matejka; Preisler, Sarah; Ejegod, Ditte M

2013-01-01

The APTIMA Human Papillomavirus (HPV) Assay detects E6/E7 mRNA from 14 human papillomavirus genotypes. Horizon was a population-based split-sample study among well-screened women, with an aim to compare APTIMA, Hybrid Capture 2 (HC2), and liquid-based cytology (LBC) using SurePath samples. APTIMA...

EVALUATION OF CARBON DIOXIDE CAPTURE FROM EXISTING COAL FIRED PLANTS BY HYBRID SORPTION USING SOLID SORBENTS

Energy Technology Data Exchange (ETDEWEB)

Benson, Steven; Browers, Bruce; Srinivasachar, Srivats; Laudal, Daniel

2014-12-31

Under contract DE-FE0007603, the University of North Dakota conducted the project Evaluation of Carbon Dioxide Capture from Existing Coal Fired Plants by Hybrid Sorption Using Solid Sorbents. As an important element of this effort, a Technical and Economic Feasibility Study was conducted by Barr Engineering Co. (Barr) in association with the University of North Dakota. The assessment developed a process flow diagram, major equipment list, heat balances for the SCPC power plant, capital cost estimate, operating cost estimate, levelized cost of electricity, cost of CO2 capture ($/ton) and three sensitivity cases for the CACHYS™ process.

A Paper-Based Sandwich Format Hybridization Assay for Unlabeled Nucleic Acid Detection Using Upconversion Nanoparticles as Energy Donors in Luminescence Resonance Energy Transfer.

Science.gov (United States)

Zhou, Feng; Noor, M Omair; Krull, Ulrich J

2015-09-24

Bioassays based on cellulose paper substrates are gaining increasing popularity for the development of field portable and low-cost diagnostic applications. Herein, we report a paper-based nucleic acid hybridization assay using immobilized upconversion nanoparticles (UCNPs) as donors in luminescence resonance energy transfer (LRET). UCNPs with intense green emission served as donors with Cy3 dye as the acceptor. The avidin functionalized UCNPs were immobilized on cellulose paper and subsequently bioconjugated to biotinylated oligonucleotide probes. Introduction of unlabeled oligonucleotide targets resulted in a formation of probe-target duplexes. A subsequent hybridization of Cy3 labeled reporter with the remaining single stranded portion of target brought the Cy3 dye in close proximity to the UCNPs to trigger a LRET-sensitized emission from the acceptor dye. The hybridization assays provided a limit of detection (LOD) of 146.0 fmol and exhibited selectivity for one base pair mismatch discrimination. The assay was functional even in undiluted serum samples. This work embodies important progress in developing DNA hybridization assays on paper. Detection of unlabeled targets is achieved using UCNPs as LRET donors, with minimization of background signal from paper substrates owing to the implementation of low energy near-infrared (NIR) excitation.

A Techno-Economic Assessment of Hybrid Cooling Systems for Coal- and Natural-Gas-Fired Power Plants with and without Carbon Capture and Storage.

Science.gov (United States)

Zhai, Haibo; Rubin, Edward S

2016-04-05

Advanced cooling systems can be deployed to enhance the resilience of thermoelectric power generation systems. This study developed and applied a new power plant modeling option for a hybrid cooling system at coal- or natural-gas-fired power plants with and without amine-based carbon capture and storage (CCS) systems. The results of the plant-level analyses show that the performance and cost of hybrid cooling systems are affected by a range of environmental, technical, and economic parameters. In general, when hot periods last the entire summer, the wet unit of a hybrid cooling system needs to share about 30% of the total plant cooling load in order to minimize the overall system cost. CCS deployment can lead to a significant increase in the water use of hybrid cooling systems, depending on the level of CO2 capture. Compared to wet cooling systems, widespread applications of hybrid cooling systems can substantially reduce water use in the electric power sector with only a moderate increase in the plant-level cost of electricity generation.

Detection of target staphylococcal enterotoxin B antigen in orange juice and popular carbonated beverages using antibody-dependent antigen-capture assays.

Science.gov (United States)

Principato, MaryAnn; Njoroge, Joyce M; Perlloni, Andrei; O' Donnell, Michael; Boyle, Thomas; Jones, Robert L

2010-10-01

There is a critical need for qualitative and quantitative methodologies that provide the rapid and accurate detection of food contaminants in complex food matrices. However, the sensitivity of the assay can be affected when antigen-capture is applied to certain foods or beverages that are extremely acidic. This study was undertaken to assess the effects of orange juice and popular carbonated soft drink upon the fidelity of antibody-based antigen-capture assays and to develop simple approaches that could rescue assay performance without the introduction of additional or extensive extraction procedures. We examined the effects of orange juice and a variety of popular carbonated soft drink beverages upon a quantitative Interleukin-2 (IL-2) enzyme-linked immunosorbent assay (ELISA) assay system and a lateral flow device (LFD) adapted for the detection of staphylococcal enterotoxin B (SEB) in foods. Alterations in the performance and sensitivity of the assay were directly attributable to the food matrix, and alterations in pH were especially critical. The results demonstrate that approaches such as an alteration of pH and the use of milk as a blocking agent, either singly or in combination, will partially rescue ELISA performance. The same approaches permit lateral flow to efficiently detect antigen. Practical Application: The authors present ways to rescue an ELISA assay compromised by acidity in beverages and show that either the alteration of pH, or the use of milk as a blocking agent are not always capable of restoring the assay to its intended efficiency. However, the same methods, when employed with lateral flow technology, are rapid and extremely successful.

Paper-based solid-phase multiplexed nucleic acid hybridization assay with tunable dynamic range using immobilized quantum dots as donors in fluorescence resonance energy transfer.

Science.gov (United States)

Noor, M Omair; Krull, Ulrich J

2013-08-06

A multiplexed solid-phase nucleic acid hybridization assay on a paper-based platform is presented using multicolor immobilized quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET). The surface of paper was modified with imidazole groups to immobilize two types of QD-probe oligonucleotide conjugates that were assembled in solution. Green-emitting QDs (gQDs) and red-emitting QDs (rQDs) served as donors with Cy3 and Alexa Fluor 647 (A647) acceptors. The gQD/Cy3 FRET pair served as an internal standard, while the rQD/A647 FRET pair served as a detection channel, combining the control and analytical test zones in one physical location. Hybridization of dye-labeled oligonucleotide targets provided the proximity for FRET sensitized emission from the acceptor dyes, which served as an analytical signal. Hybridization assays in the multicolor format provided a limit of detection of 90 fmol and an upper limit of dynamic range of 3.5 pmol. The use of an array of detection zones was designed to provide improved analytical figures of merit compared to that which could be achieved on one type of array design in terms of relative concentration of multicolor QDs. The hybridization assays showed excellent resistance to nonspecific adsorption of oligonucleotides. Selectivity of the two-plex hybridization assay was demonstrated by single nucleotide polymorphism (SNP) detection at a contrast ratio of 50:1. Additionally, it is shown that the use of preformed QD-probe oligonucleotide conjugates and consideration of the relative number density of the two types of QD-probe conjugates in the two-color assay format is advantageous to maximize assay sensitivity and the upper limit of dynamic range.

Development of a quantitative bead capture assay for soluble IL-7 receptor alpha in human plasma.

Directory of Open Access Journals (Sweden)

Sylvie Faucher

Full Text Available BACKGROUND: IL-7 is an essential cytokine in T-cell development and homeostasis. It binds to the IL-7R receptor, a complex of the IL-7Ralpha (CD127 and common gamma (CD132 chains. There is significant interest in evaluating the expression of CD127 on human T-cells as it often decreased in medical conditions leading to lymphopenia. Previous reports showed the usefulness of CD127 as a prognostic marker in viral infections such as HIV, CMV, EBV and HCV. A soluble CD127 (sCD127 is released in plasma and may contribute to disease pathogenesis through its control on IL-7 activities. Measuring sCD127 is important to define its role and may complement existing markers used in lymphopenic disease management. We describe a new quantitative assay for the measurement of sCD127 in plasma and report sCD127 concentrations in healthy adults. METHODOLOGY/PRINCIPAL FINDINGS: We developed a quantitative bead-based sCD127 capture assay. Polyclonal CD127-specific antibodies were chosen for capture and a biotinylated monoclonal anti-CD127 antibody was selected for detection. The assay can detect native sCD127 and recombinant sCD127 which served as the calibrator. The analytical performance of the assay was characterized and the concentration and stability of plasma sCD127 in healthy adults was determined. The assay's range was 3.2-1000 ng/mL. The concentration of plasma sCD127 was 164+/-104 ng/mL with over a log variation between subjects. Individual sCD127 concentrations remained stable when measured serially during a period of up to one year. CONCLUSIONS/SIGNIFICANCE: This is the first report on the quantification of plasma sCD127 in a population of healthy adults. Soluble CD127 plasma concentrations remained stable over time in a given individual and sCD127 immunoreactivity was resistant to repeated freeze-thaw cycles. This quantitative sCD127 assay is a valuable tool for defining the potential role of sCD127 in lymphopenic diseases.

Evaluation of Myc Gene Amplification in Prostate Cancer Using a Dual Color Chromogenic in-Situ Hybridization (Dual CISH Assay

Directory of Open Access Journals (Sweden)

Daniel Lerda

2013-04-01

Full Text Available Objetive: The overall purpose of the study was to demonstrate applicability of the Dako dual-color chromogenic in situ hybridization (CISH assay (DAKO Denmark, Glostrup with respect to fluorescence in situ hybridization (FISH probes MYC-C. Methods: MYC gene amplification by FISH and Dako dual-color CISH Results: The study showed that the dual-color CISH assay can convert Texas red and fluorescein isothiocyanate (FITC signals into chromogenic signals. The dual –color CISH assay was performed on 40 cases of prostate cancer. Amplification was identified in 12 of 40 (30% tumors. No amplification was seen in 28 of 40 (70% tumors. FISH data were available in total of 40 tumors. All tumors showed concordant results between dual-color CISH and FISH for classifying a tumor as MYC amplified or not amplified. Conclusions: We conclude that dual-color Dako CISH assay is an accurate method for determining MYC gene amplification with added advantages that make it a more practically useful method. [J Interdiscipl Histopathol 2013; 1(2.000: 81-84

Double positive effect of adding hexaethyelene glycol when optimizing the hybridization efficiency of a microring DNA detection assay

Energy Technology Data Exchange (ETDEWEB)

Van Eeghem, Anabelle, E-mail: anabelle.vaneeghem@gmail.com [Polymer Chemistry and Biomaterials Research Group, Department of Organic and Macromolecular Chemistry, Ghent University (Belgium); Center for Nano- and Biophotonics, Ghent University (Belgium); Werquin, Sam [Center for Nano- and Biophotonics, Ghent University (Belgium); Photonics Research Group, Department of Information Technology, Ghent University – IMEC (Belgium); Hoste, Jan-Willem, E-mail: janwillem.hoste@ugent.be [Center for Nano- and Biophotonics, Ghent University (Belgium); Photonics Research Group, Department of Information Technology, Ghent University – IMEC (Belgium); Goes, Arne [Polymer Chemistry and Biomaterials Research Group, Department of Organic and Macromolecular Chemistry, Ghent University (Belgium); Agrosavfe NV, Technologiepark 4 (Bio-incubator), Zwijnaarde (Belgium); Vanderleyden, Els [Polymer Chemistry and Biomaterials Research Group, Department of Organic and Macromolecular Chemistry, Ghent University (Belgium); Center for Nano- and Biophotonics, Ghent University (Belgium); Bienstman, Peter [Center for Nano- and Biophotonics, Ghent University (Belgium); Photonics Research Group, Department of Information Technology, Ghent University – IMEC (Belgium); Dubruel, Peter [Polymer Chemistry and Biomaterials Research Group, Department of Organic and Macromolecular Chemistry, Ghent University (Belgium); Center for Nano- and Biophotonics, Ghent University (Belgium)

2017-05-31

Highlights: • The hybridization efficiency of a DNA assay was investigated based on SOI microring resonators. • A 4-fold increase in efficiency was obtained by using HEG as backfilling agent, as well as improving robustness. • The dual polarization microring technique shows that HEG reorients the DNA in an upright position. • Hybridizing at 35 °C and with a buffer containing 50 v/v% of formamide greatly improves the robustness. - Abstract: In this paper, a method for detection of DNA molecules using silicon-on-insulator (SOI) microring resonators is described. The influence of temperature and the use of formamide on the hybridization efficiency were studied. It was shown that 50 v/v% of formamide in the hybridization buffer can ensure hybridization when working close to physiological temperature. Furthermore, the use of hexaethylene glycol (HEG) as backfilling agent was studied in order to resolve issues of non-specific adsorption to the surface. The results indicated that not only non-specific binding was reduced significantly but also that HEG improves the orientation of the DNA probes on the surface. This led to a 4-fold increase in hybridization efficiency and thus in an equal decrease in the detection limit, compared to hybridization without the use of HEG. An improvement in robustness of the assay was also observed. This DNA reorientation hypothesis was confirmed by studying the thickness and density of the layers by using dual polarization microring sensing. Finally, the different steps in the sensing experiment were characterized in more detail by static contact angle (SCA) and X-ray photoelectron spectroscopy (XPS) analysis. The results showed quantitatively that the surface modifications were successful.

Carbon capture by hybrid separation processes

NARCIS (Netherlands)

van Benthum, R.J.; van Kemenade, H.P.; Brouwers, J.J.H.

2014-01-01

Even though there is an increasing development of carbon capture technology over the last decade, large-scale implementation is still far from common practice, mainly caused by the energy intensiveness of carbon capture processes and the lack of regulation. In absence of strict regulation, less

Investigation of parameters that affect the success rate of microarray-based allele-specific hybridization assays.

Directory of Open Access Journals (Sweden)

Lena Poulsen

Full Text Available BACKGROUND: The development of microarray-based genetic tests for diseases that are caused by known mutations is becoming increasingly important. The key obstacle to developing functional genotyping assays is that such mutations need to be genotyped regardless of their location in genomic regions. These regions include large variations in G+C content, and structural features like hairpins. METHODS/FINDINGS: We describe a rational, stable method for screening and combining assay conditions for the genetic analysis of 42 Phenylketonuria-associated mutations in the phenylalanine hydroxylase gene. The mutations are located in regions with large variations in G+C content (20-75%. Custom-made microarrays with different lengths of complementary probe sequences and spacers were hybridized with pooled PCR products of 12 exons from each of 38 individual patient DNA samples. The arrays were washed with eight buffers with different stringencies in a custom-made microfluidic system. The data were used to assess which parameters play significant roles in assay development. CONCLUSIONS: Several assay development methods found suitable probes and assay conditions for a functional test for all investigated mutation sites. Probe length, probe spacer length, and assay stringency sufficed as variable parameters in the search for a functional multiplex assay. We discuss the optimal assay development methods for several different scenarios.

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

Science.gov (United States)

Xu, Yao; Zheng, Zhi

2016-05-15

We have developed a convenient, robust and low-cost RNA detection system suitable for high-throughput applications. This system uses a highly specific sandwich hybridization to capture target RNA directly onto solid support, followed by on-site signal amplification via 2-dimensional, branched hybridizing chain polymerization through toehold-mediated strand displacement reaction. The assay uses SYBR Green to detect targets at concentrations as low as 1 pM, without involving nucleic acid purification or any enzymatic reaction, using ordinary oligonucleotides without modification or labeling. The system was demonstrated in the detection of malaria RNA in blood and GAPDH gene expression in cell lysate. Copyright © 2015 Elsevier B.V. All rights reserved.

Non-Enzymatic Detection of Bacterial Genomic DNA Using the Bio-Barcode Assay

Science.gov (United States)

Hill, Haley D.; Vega, Rafael A.; Mirkin, Chad A.

2011-01-01

The detection of bacterial genomic DNA through a non-enzymatic nanomaterials based amplification method, the bio-barcode assay, is reported. The assay utilizes oligonucleotide functionalized magnetic microparticles to capture the target of interest from the sample. A critical step in the new assay involves the use of blocking oligonucleotides during heat denaturation of the double stranded DNA. These blockers bind to specific regions of the target DNA upon cooling, and prevent the duplex DNA from re-hybridizing, which allows the particle probes to bind. Following target isolation using the magnetic particles, oligonucleotide functionalized gold nanoparticles act as target recognition agents. The oligonucleotides on the nanoparticle (barcodes) act as amplification surrogates. The barcodes are then detected using the Scanometric method. The limit of detection for this assay was determined to be 2.5 femtomolar, and this is the first demonstration of a barcode type assay for the detection of double stranded, genomic DNA. PMID:17927207

Hybridization Capture Reveals Evolution and Conservation across the Entire Koala Retrovirus Genome

Science.gov (United States)

Ishida, Yasuko; Cui, Pin; Vielgrader, Hanna; Helgen, Kristofer M.; Roca, Alfred L.; Greenwood, Alex D.

2014-01-01

The koala retrovirus (KoRV) is the only retrovirus known to be in the midst of invading the germ line of its host species. Hybridization capture and next generation sequencing were used on modern and museum DNA samples of koala (Phascolarctos cinereus) to examine ca. 130 years of evolution across the full KoRV genome. Overall, the entire proviral genome appeared to be conserved across time in sequence, protein structure and transcriptional binding sites. A total of 138 polymorphisms were detected, of which 72 were found in more than one individual. At every polymorphic site in the museum koalas, one of the character states matched that of modern KoRV. Among non-synonymous polymorphisms, radical substitutions involving large physiochemical differences between amino acids were elevated in env, potentially reflecting anti-viral immune pressure or avoidance of receptor interference. Polymorphisms were not detected within two functional regions believed to affect infectivity. Host sequences flanking proviral integration sites were also captured; with few proviral loci shared among koalas. Recently described variants of KoRV, designated KoRV-B and KoRV-J, were not detected in museum samples, suggesting that these variants may be of recent origin. PMID:24752422

Hybridization capture reveals evolution and conservation across the entire Koala retrovirus genome.

Directory of Open Access Journals (Sweden)

Kyriakos Tsangaras

Full Text Available The koala retrovirus (KoRV is the only retrovirus known to be in the midst of invading the germ line of its host species. Hybridization capture and next generation sequencing were used on modern and museum DNA samples of koala (Phascolarctos cinereus to examine ca. 130 years of evolution across the full KoRV genome. Overall, the entire proviral genome appeared to be conserved across time in sequence, protein structure and transcriptional binding sites. A total of 138 polymorphisms were detected, of which 72 were found in more than one individual. At every polymorphic site in the museum koalas, one of the character states matched that of modern KoRV. Among non-synonymous polymorphisms, radical substitutions involving large physiochemical differences between amino acids were elevated in env, potentially reflecting anti-viral immune pressure or avoidance of receptor interference. Polymorphisms were not detected within two functional regions believed to affect infectivity. Host sequences flanking proviral integration sites were also captured; with few proviral loci shared among koalas. Recently described variants of KoRV, designated KoRV-B and KoRV-J, were not detected in museum samples, suggesting that these variants may be of recent origin.

Efficient capture of CO2 over ordered micro-mesoporous hybrid carbon nanosphere

Science.gov (United States)

Chen, Changwei; Yu, Yanke; He, Chi; Wang, Li; Huang, Huang; Albilali, Reem; Cheng, Jie; Hao, Zhengping

2018-05-01

Four kinds of carbon-based adsorbents (micro-mesoporous hybrid carbon nanosphere and N-doped hollow carbon sphere with single-, double- or ruga-shell morphology) with different structural and textural properties were prepared and systematically studied in CO2 capture. All synthesized samples possess high specific surface area (828-910 m2 g-1), large pore volume (0.71-1.81 cm3 g-1), and different micropore contents varied from 2.1% to 46.4%. Amongst, the ordered micro-mesoporous carbon nanosphere (OM-CNS) exhibits the best adsorption performance with CO2 uptake as high as 3.01 mmol g-1 under conditions of 298 K and 1.0 bar, better than most of the reported CO2 adsorbents. The excellent CO2 adsorption capacity of OM-CNS can be reasonably attributed to the synergistic effect of ordered mesopore channels and abundant structural micropores which are beneficial for the diffusion and trapping of CO2 adsorbate. Moreover, the OM-CNS shows excellent CO2 trapping selectivity and superior stability and recyclability, which endow the OM-CNS as a promising and environmental-friendly adsorbent for CO2 capture and separation under practical conditions.

Antibody class capture assay (ACCA) for rubella-specific IgM antibody.

Science.gov (United States)

Isaac, M; Payne, R A

1982-01-01

Enzyme-linked immunosorbent assays for IgM antirubella were carried out on 1,546 sera, using an IgM capture method with a F (ab')2 conjugate (ACCA). Under the conditions described, sera containing IgM antirubella bound up to 15 times as much enzyme activity as negative specimens. Paired serum specimens from 27 patients, serial serum specimens from 6 patients, and single serum specimens from 15 patients who had had recent rubella were examined by the haemagglutination inhibition test (HAI) in the presence and absence of 2-mercaptoethanol following sucrose density gradient centrifugation (SDGC). ACCA confirmed all the results found with HAI following SDGC. Specimens were examined from ten patients with congenital rubella; ACCA confirmed the results found with both immunofluorescence following SDGC and radioimmunoassay. Pre- and post-vaccination specimens from 123 patients who had been vaccinated against rubella were examined. An IgM response could only be demonstrated in the 57 cases when IgG was absent in the first specimen. The specificity of the assay was confirmed by testing 31 serum specimens from rubella immune patients that also contained rheumatoid factor, 163 serum specimens from patients with acute infections other than rubella, and 12 serum specimens from infants with miscellaneous neonatal abnormalities other than congenital rubella. The ACCA proved a simple, sensitive, and specific test for IgM antirubella and the results compared favourably with those obtained by the SDGC technique.

Fluorescence in situ hybridization in combination with the comet assay and micronucleus test in genetic toxicology

Directory of Open Access Journals (Sweden)

Hovhannisyan Galina G

2010-09-01

Full Text Available Abstract Comet assay and micronucleus (MN test are widely applied in genotoxicity testing and biomonitoring. While comet assay permits to measure direct DNA-strand breaking capacity of a tested agent MN test allows estimating the induced amount of chromosome and/or genome mutations. The potential of these two methods can be enhanced by the combination with fluorescence in situ hybridization (FISH techniques. FISH plus comet assay allows the recognition of targets of DNA damage and repairing directly. FISH combined with MN test is able to characterize the occurrence of different chromosomes in MN and to identify potential chromosomal targets of mutagenic substances. Thus, combination of FISH with the comet assay or MN test proved to be promising techniques for evaluation of the distribution of DNA and chromosome damage in the entire genome of individual cells. FISH technique also permits to study comet and MN formation, necessary for correct application of these methods. This paper reviews the relevant literature on advantages and limitations of Comet-FISH and MN-FISH assays application in genetic toxicology.

Recent Advances in Anhydrous Solvents for CO2 Capture: Ionic Liquids, Switchable Solvents, and Nanoparticle Organic Hybrid Materials

International Nuclear Information System (INIS)

Park, Youngjune; Lin, Kun-Yi Andrew; Park, Ah-Hyung Alissa; Petit, Camille

2015-01-01

CO 2 capture by amine scrubbing, which has a high CO 2 capture capacity and a rapid reaction rate, is the most employed and investigated approach to date. There are a number of recent large-scale demonstrations including the Boundary Dam Carbon Capture Project by SaskPower in Canada that have reported successful implementations of aqueous amine solvent in CO 2 capture from flue gases. The findings from these demonstrations will significantly advance the field of CO 2 capture in the coming years. While the latest efforts in aqueous amine solvents are exciting and promising, there are still several drawbacks to amine-based CO 2 capture solvents including high volatility and corrosiveness of the amine solutions as well as the high parasitic energy penalty during the solvent regeneration step. Thus, in a parallel effort, alternative CO 2 capture solvents, which are often anhydrous, have been developed as the third-generation CO 2 capture solvents. These novel classes of liquid materials include ionic liquids, CO 2 -triggered switchable solvents (i.e., CO 2 -binding organic liquids, reversible ionic liquids), and nanoparticle organic hybrid materials. This paper provides a review of these various anhydrous solvents and their potential for CO 2 capture. Particular attention is given to the mechanisms of CO 2 absorption in these solvents, their regeneration and their processability – especially taking into account their viscosity. While not intended to provide a complete coverage of the existing literature, this review aims at pointing the major findings reported for these new classes of CO 2 capture media.

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

Science.gov (United States)

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

2013-07-08

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

Interfacial chemistry and the design of solid-phase nucleic acid hybridization assays using immobilized quantum dots as donors in fluorescence resonance energy transfer.

Science.gov (United States)

Algar, W Russ; Krull, Ulrich J

2011-01-01

The use of quantum dots (QDs) as donors in fluorescence resonance energy transfer (FRET) offer several advantages for the development of multiplexed solid-phase QD-FRET nucleic acid hybridization assays. Designs for multiplexing have been demonstrated, but important challenges remain in the optimization of these systems. In this work, we identify several strategies based on the design of interfacial chemistry for improving sensitivity, obtaining lower limits of detection (LOD) and enabling the regeneration and reuse of solid-phase QD-FRET hybridization assays. FRET-sensitized emission from acceptor dyes associated with hybridization events at immobilized QD donors provides the analytical signal in these assays. The minimization of active sensing area reduces background from QD donor PL and allows the resolution of smaller amounts of acceptor emission, thus lowering the LOD. The association of multiple acceptor dyes with each hybridization event can enhance FRET efficiency, thereby improving sensitivity. Many previous studies have used interfacial protein layers to generate selectivity; however, transient destabilization of these layers is shown to prevent efficient regeneration. To this end, we report a protein-free interfacial chemistry and demonstrate the specific detection of as little as 2 pmol of target, as well as an improved capacity for regeneration.

Effects of Bonding Types and Functional Groups on CO 2 Capture using Novel Multiphase Systems of Liquid-like Nanoparticle Organic Hybrid Materials

KAUST Repository

Lin, Kun-Yi Andrew; Park, Ah-Hyung Alissa

2011-01-01

Novel liquid-like nanoparticle organic hybrid materials (NOHMs) which possess unique features including negligible vapor pressure and a high degree of tunability were synthesized and their physical and chemical properties as well as CO 2 capture

A dual amplification strategy for DNA detection combining bio-barcode assay and metal-enhanced fluorescence modality.

Science.gov (United States)

Zhou, Zhenpeng; Li, Tian; Huang, Hongduan; Chen, Yang; Liu, Feng; Huang, Chengzhi; Li, Na

2014-11-11

Silver-enhanced fluorescence was coupled with a bio-barcode assay to facilitate a dual amplification assay to demonstrate a non-enzymatic approach for simple and sensitive detection of DNA. In the assay design, magnetic nanoparticles seeded with silver nanoparticles were modified with the capture DNA, and silver nanoparticles were modified with the binding of ssDNA and the fluorescently labeled barcode dsDNA. Upon introduction of the target DNA, a sandwich structure was formed because of the hybridization reaction. By simple magnetic separation, silver-enhanced fluorescence of barcode DNAs could be readily measured without the need of a further step to liberate barcode DNAs from silver nanoparticles, endowing the method with simplicity and high sensitivity with a detection limit of 1 pM.

A multiplex reverse transcription PCR and automated electronic microarray assay for detection and differentiation of seven viruses affecting swine.

Science.gov (United States)

Erickson, A; Fisher, M; Furukawa-Stoffer, T; Ambagala, A; Hodko, D; Pasick, J; King, D P; Nfon, C; Ortega Polo, R; Lung, O

2018-04-01

Microarray technology can be useful for pathogen detection as it allows simultaneous interrogation of the presence or absence of a large number of genetic signatures. However, most microarray assays are labour-intensive and time-consuming to perform. This study describes the development and initial evaluation of a multiplex reverse transcription (RT)-PCR and novel accompanying automated electronic microarray assay for simultaneous detection and differentiation of seven important viruses that affect swine (foot-and-mouth disease virus [FMDV], swine vesicular disease virus [SVDV], vesicular exanthema of swine virus [VESV], African swine fever virus [ASFV], classical swine fever virus [CSFV], porcine respiratory and reproductive syndrome virus [PRRSV] and porcine circovirus type 2 [PCV2]). The novel electronic microarray assay utilizes a single, user-friendly instrument that integrates and automates capture probe printing, hybridization, washing and reporting on a disposable electronic microarray cartridge with 400 features. This assay accurately detected and identified a total of 68 isolates of the seven targeted virus species including 23 samples of FMDV, representing all seven serotypes, and 10 CSFV strains, representing all three genotypes. The assay successfully detected viruses in clinical samples from the field, experimentally infected animals (as early as 1 day post-infection (dpi) for FMDV and SVDV, 4 dpi for ASFV, 5 dpi for CSFV), as well as in biological material that were spiked with target viruses. The limit of detection was 10 copies/μl for ASFV, PCV2 and PRRSV, 100 copies/μl for SVDV, CSFV, VESV and 1,000 copies/μl for FMDV. The electronic microarray component had reduced analytical sensitivity for several of the target viruses when compared with the multiplex RT-PCR. The integration of capture probe printing allows custom onsite array printing as needed, while electrophoretically driven hybridization generates results faster than conventional

Investigation of CO2 capture mechanisms of liquid-like nanoparticle organic hybrid materials via structural characterization

KAUST Repository

Park, Youngjune

2011-01-01

Nanoparticle organic hybrid materials (NOHMs) have been recently developed that comprise an oligomeric or polymeric canopy tethered to surface-modified nanoparticles via ionic or covalent bonds. It has already been shown that the tunable nature of the grafted polymeric canopy allows for enhanced CO 2 capture capacity and selectivity via the enthalpic intermolecular interactions between CO2 and the task-specific functional groups, such as amines. Interestingly, for the same amount of CO2 loading NOHMs have also exhibited significantly different swelling behavior compared to that of the corresponding polymers, indicating a potential structural effect during CO2 capture. If the frustrated canopy species favor spontaneous ordering due to steric and/or entropic effects, the inorganic cores of NOHMs could be organized into unusual structural arrangements. Likewise, the introduction of small gaseous molecules such as CO2 could reduce the free energy of the frustrated canopy. This entropic effect, the result of unique structural nature, could allow NOHMs to capture CO2 more effectively. In order to isolate the entropic effect, NOHMs were synthesized without the task-specific functional groups. The relationship between their structural conformation and the underlying mechanisms for the CO2 absorption behavior were investigated by employing NMR and ATR FT-IR spectroscopies. The results provide fundamental information needed for evaluating and developing novel liquid-like CO2 capture materials and give useful insights for designing and synthesizing NOHMs for more effective CO2 capture. © the Owner Societies 2011.

Investigation of CO2 capture mechanisms of liquid-like nanoparticle organic hybrid materials via structural characterization.

Science.gov (United States)

Park, Youngjune; Decatur, John; Lin, Kun-Yi Andrew; Park, Ah-Hyung Alissa

2011-10-28

Nanoparticle organic hybrid materials (NOHMs) have been recently developed that comprise an oligomeric or polymeric canopy tethered to surface-modified nanoparticles via ionic or covalent bonds. It has already been shown that the tunable nature of the grafted polymeric canopy allows for enhanced CO(2) capture capacity and selectivity via the enthalpic intermolecular interactions between CO(2) and the task-specific functional groups, such as amines. Interestingly, for the same amount of CO(2) loading NOHMs have also exhibited significantly different swelling behavior compared to that of the corresponding polymers, indicating a potential structural effect during CO(2) capture. If the frustrated canopy species favor spontaneous ordering due to steric and/or entropic effects, the inorganic cores of NOHMs could be organized into unusual structural arrangements. Likewise, the introduction of small gaseous molecules such as CO(2) could reduce the free energy of the frustrated canopy. This entropic effect, the result of unique structural nature, could allow NOHMs to capture CO(2) more effectively. In order to isolate the entropic effect, NOHMs were synthesized without the task-specific functional groups. The relationship between their structural conformation and the underlying mechanisms for the CO(2) absorption behavior were investigated by employing NMR and ATR FT-IR spectroscopies. The results provide fundamental information needed for evaluating and developing novel liquid-like CO(2) capture materials and give useful insights for designing and synthesizing NOHMs for more effective CO(2) capture. This journal is © the Owner Societies 2011

Recent Advances in Anhydrous Solvents for CO2 Capture: Ionic Liquids, Switchable Solvents, and Nanoparticle Organic Hybrid Materials

Directory of Open Access Journals (Sweden)

YOUNGJUNE ePARK

2015-10-01

Full Text Available CO2 capture by amine scrubbing, which has a high CO2 capture capacity and a rapid reaction rate, is the most employed and investigated approach to date. There are a number of recent large-scale demonstrations including the Boundary Dam Carbon Capture Project by SaskPower in Canada that have reported successful implementations of aqueous amine solvent in CO2 capture from flue gases. The findings from these demonstrations will significantly advance the field of CO2 capture in the coming years. While the latest efforts in aqueous amine solvents are exciting and promising, there are still several drawbacks to amine-based CO2 capture solvents including high volatility and corrosiveness of the amine solutions, as well as the high parasitic energy penalty during the solvent regeneration step. Thus, in a parallel effort, alternative CO2 capture solvents, which are often anhydrous, have been developed as the third-generation CO2 capture solvents. These novel classes of liquid materials include: Ionic Liquids (ILs, CO2-triggered switchable solvents (i.e., CO2 Binding Organic Liquids (CO2BOLs, Reversible Ionic Liquids (RevILs, and Nanoparticle Organic Hybrid Materials (NOHMs. This paper provides a review of these various anhydrous solvents and their potential for CO2 capture. Particular attention is given to the mechanisms of CO2 absorption in these solvents, their regeneration and their processability – especially taking into account their viscosity. While not intended to provide a complete coverage of the existing literature, this review aims at pointing the major findings reported for these new classes of CO2 capture media.

Electrochemical Branched-DNA Assay for Polymerase Chain Reaction-Free Detection and Quantification of Oncogenes in Messenger RNA

Energy Technology Data Exchange (ETDEWEB)

Lee, Ai Cheng; Dai, Ziyu; Chen, Baowei; Wu, Hong; Wang, Jun; Zhang, Aiguo; Zhang, Lurong; Lim, Tit-Meng; Lin, Yuehe

2008-12-01

We describe a novel electrochemical branched-DNA (bDNA) assay for polymerase chain reaction (PCR)-free detection and quantification of p185 BCR-ABL leukemia fusion transcript in the population of messenger RNA (mRNA) extracted from cell lines. The bDNA amplifier carrying high loading of alkaline phosphatase (ALP) tracers was used to amplify targets signal. The targets were captured on microplate well surfaces through cooperative sandwich hybridization prior to the labeling of bDNA. The activity of captured ALP was monitored by square-wave voltammetric (SWV) analysis of the electroactive enzymatic product in the presence of 1-napthyl-phosphate. The specificity and sensitivity of assay enabled direct detection of target transcript in as little as 4.6 ng mRNA without PCR amplification. In combination with the use of a well-quantified standard, the electrochemical bDNA assay was capable of direct use for a PCR-free quantitative analysis of target transcript in total mRNA population. The approach thus provides a simple, sensitive, accurate and quantitative tool alternate to the RQ-PCR for early disease diagnosis.

Comparison of clinical and analytical performance of the Abbott Realtime High Risk HPV test to the performance of hybrid capture 2 in population-based cervical cancer screening.

Science.gov (United States)

Poljak, Mario; Ostrbenk, Anja; Seme, Katja; Ucakar, Veronika; Hillemanns, Peter; Bokal, Eda Vrtacnik; Jancar, Nina; Klavs, Irena

2011-05-01

The clinical performance of the Abbott RealTime High Risk HPV (human papillomavirus) test (RealTime) and that of the Hybrid Capture 2 HPV DNA test (hc2) were prospectively compared in the population-based cervical cancer screening setting. In women >30 years old (n = 3,129), the clinical sensitivity of RealTime for detection of cervical intraepithelial neoplasia of grade 2 (CIN2) or worse (38 cases) and its clinical specificity for lesions of less than CIN2 (3,091 controls) were 100% and 93.3%, respectively, and those of hc2 were 97.4% and 91.8%, respectively. A noninferiority score test showed that the clinical specificity (P laboratories. RealTime can be considered to be a reliable and robust HPV assay clinically comparable to hc2 for the detection of CIN2+ lesions in a population-based cervical cancer screening setting.

Differential Detection of Human Papillomavirus Genotypes and Cervical Intraepithelial Neoplasia by Four Commercial Assays

DEFF Research Database (Denmark)

Rebolj, Matejka; Bonde, Jesper; Preisler, Sarah

2016-01-01

intraepithelial neoplasia (CIN) in 2.5 years after the baseline testing were determined from the national pathology register. HPV-positive women undergoing primary screening having concordant samples were more likely to harbor high-risk infections and less likely to harbor only low-risk infections than women......Laboratories can nowadays choose from >100 Human Papillomavirus (HPV) assays for cervical screening. Our previous analysis based on the data from the Danish Horizon study, however, showed that four widely used assays, Hybrid Capture 2 (HC2), cobas, CLART and APTIMA, frequently do not detect...... the same HPV infections. Here, we determined the characteristics of the concordant (all four assays returning a positive HPV test result) and discordant samples (all other HPV-positive samples) in primary cervical screening at 30-65 years (n=2859) and in a concurrent referral population from the same...

Competitive Reporter Monitored Amplification (CMA) - Quantification of Molecular Targets by Real Time Monitoring of Competitive Reporter Hybridization

Science.gov (United States)

Ullrich, Thomas; Ermantraut, Eugen; Schulz, Torsten; Steinmetzer, Katrin

2012-01-01

Background State of the art molecular diagnostic tests are based on the sensitive detection and quantification of nucleic acids. However, currently established diagnostic tests are characterized by elaborate and expensive technical solutions hindering the development of simple, affordable and compact point-of-care molecular tests. Methodology and Principal Findings The described competitive reporter monitored amplification allows the simultaneous amplification and quantification of multiple nucleic acid targets by polymerase chain reaction. Target quantification is accomplished by real-time detection of amplified nucleic acids utilizing a capture probe array and specific reporter probes. The reporter probes are fluorescently labeled oligonucleotides that are complementary to the respective capture probes on the array and to the respective sites of the target nucleic acids in solution. Capture probes and amplified target compete for reporter probes. Increasing amplicon concentration leads to decreased fluorescence signal at the respective capture probe position on the array which is measured after each cycle of amplification. In order to observe reporter probe hybridization in real-time without any additional washing steps, we have developed a mechanical fluorescence background displacement technique. Conclusions and Significance The system presented in this paper enables simultaneous detection and quantification of multiple targets. Moreover, the presented fluorescence background displacement technique provides a generic solution for real time monitoring of binding events of fluorescently labelled ligands to surface immobilized probes. With the model assay for the detection of human immunodeficiency virus type 1 and 2 (HIV 1/2), we have been able to observe the amplification kinetics of five targets simultaneously and accommodate two additional hybridization controls with a simple instrument set-up. The ability to accommodate multiple controls and targets into a

Competitive reporter monitored amplification (CMA--quantification of molecular targets by real time monitoring of competitive reporter hybridization.

Directory of Open Access Journals (Sweden)

Thomas Ullrich

Full Text Available BACKGROUND: State of the art molecular diagnostic tests are based on the sensitive detection and quantification of nucleic acids. However, currently established diagnostic tests are characterized by elaborate and expensive technical solutions hindering the development of simple, affordable and compact point-of-care molecular tests. METHODOLOGY AND PRINCIPAL FINDINGS: The described competitive reporter monitored amplification allows the simultaneous amplification and quantification of multiple nucleic acid targets by polymerase chain reaction. Target quantification is accomplished by real-time detection of amplified nucleic acids utilizing a capture probe array and specific reporter probes. The reporter probes are fluorescently labeled oligonucleotides that are complementary to the respective capture probes on the array and to the respective sites of the target nucleic acids in solution. Capture probes and amplified target compete for reporter probes. Increasing amplicon concentration leads to decreased fluorescence signal at the respective capture probe position on the array which is measured after each cycle of amplification. In order to observe reporter probe hybridization in real-time without any additional washing steps, we have developed a mechanical fluorescence background displacement technique. CONCLUSIONS AND SIGNIFICANCE: The system presented in this paper enables simultaneous detection and quantification of multiple targets. Moreover, the presented fluorescence background displacement technique provides a generic solution for real time monitoring of binding events of fluorescently labelled ligands to surface immobilized probes. With the model assay for the detection of human immunodeficiency virus type 1 and 2 (HIV 1/2, we have been able to observe the amplification kinetics of five targets simultaneously and accommodate two additional hybridization controls with a simple instrument set-up. The ability to accommodate multiple controls

Epidemiology of Babesia, Anaplasma and Trypanosoma species using a new expanded reverse line blot hybridization assay.

Science.gov (United States)

Paoletta, Martina Soledad; López Arias, Ludmila; de la Fournière, Sofía; Guillemi, Eliana Carolina; Luciani, Carlos; Sarmiento, Néstor Fabián; Mosqueda, Juan; Farber, Marisa Diana; Wilkowsky, Silvina Elizabeth

2018-02-01

Vector-borne hemoparasitic infections are a major problem that affects livestock industries worldwide, particularly in tropical and subtropical regions. In this work, a reverse line blot (RLB) hybridization assay was developed for the simultaneous detection and identification of Anaplasma, Babesia and bovine trypanosomes, encompassing in this way the most relevant hemoparasites that affect cattle. A total of 186 bovine blood samples collected from two different ecoepidemiological regions of northeast Argentina, with and without tick control, were analyzed with this new RLB. High diversity of parasites, such as Babesia bovis, B. bigemina, Anaplasma marginale and three different Trypanosoma species, was found. High rates of coinfections were also detected, and significant differences were observed not only in the prevalence of parasites but also in the level of coinfections between the two analyzed areas. Regarding the Trypanosoma genus, we provide molecular evidence of the presence of T. vivax and T. theileri for the first time in Argentina. Besides, since the RLB is a prospective tool, it allowed the identification of a yet unknown bovine trypanosome which could not be assigned to any of the bovine species known so far. In the present study we provide new insights on the prevalence of several pathogens that directly impact on livestock production in Argentina. The RLB assay developed here allows to identify simultaneously numerous pathogenic species which can also be easily expanded to detect other blood borne pathogens. These characteristics make the RLB hybridization assay an essential tool for epidemiological survey of all vector-borne pathogens. Copyright © 2017 Elsevier GmbH. All rights reserved.

Evaluation of a polymerase chain reaction reverse hybridization line probe assay for the detection and identification of medically important fungi in bronchoalveolar lavage fluids.

NARCIS (Netherlands)

Meletiadis, J.; Melchers, W.J.G.; Meis, J.F.G.M.; Hurk, P.J.J.C. van den; Jannes, G.; Verweij, P.E.

2003-01-01

An assay system in which polymerase chain reaction (PCR) amplification of the ITS-1 region of ribosomal DNA (rDNA) is combined with a reverse-hybridization line probe assay (LiPA) was used for the identification of six Candida species and four Aspergillus species in pure cultures of clinical

Recent Advances in Anhydrous Solvents for CO{sub 2} Capture: Ionic Liquids, Switchable Solvents, and Nanoparticle Organic Hybrid Materials

Energy Technology Data Exchange (ETDEWEB)

Park, Youngjune [School of Environmental Science and Engineering, Gwangju Institute of Science and Technology (GIST), Gwangju (Korea, Republic of); Lin, Kun-Yi Andrew [Department of Environmental Engineering, National Chung Hsing University, Taichung City (China); Park, Ah-Hyung Alissa, E-mail: ap2622@columbia.edu [Department of Earth and Environmental Engineering, Lenfest Center for Sustainable Energy, Columbia University, New York, NY (United States); Department of Chemical Engineering, Lenfest Center for Sustainable Energy, Columbia University, New York, NY (United States); Petit, Camille, E-mail: ap2622@columbia.edu [Department of Chemical Engineering, Imperial College London, London (United Kingdom)

2015-10-01

CO{sub 2} capture by amine scrubbing, which has a high CO{sub 2} capture capacity and a rapid reaction rate, is the most employed and investigated approach to date. There are a number of recent large-scale demonstrations including the Boundary Dam Carbon Capture Project by SaskPower in Canada that have reported successful implementations of aqueous amine solvent in CO{sub 2} capture from flue gases. The findings from these demonstrations will significantly advance the field of CO{sub 2} capture in the coming years. While the latest efforts in aqueous amine solvents are exciting and promising, there are still several drawbacks to amine-based CO{sub 2} capture solvents including high volatility and corrosiveness of the amine solutions as well as the high parasitic energy penalty during the solvent regeneration step. Thus, in a parallel effort, alternative CO{sub 2} capture solvents, which are often anhydrous, have been developed as the third-generation CO{sub 2} capture solvents. These novel classes of liquid materials include ionic liquids, CO{sub 2}-triggered switchable solvents (i.e., CO{sub 2}-binding organic liquids, reversible ionic liquids), and nanoparticle organic hybrid materials. This paper provides a review of these various anhydrous solvents and their potential for CO{sub 2} capture. Particular attention is given to the mechanisms of CO{sub 2} absorption in these solvents, their regeneration and their processability – especially taking into account their viscosity. While not intended to provide a complete coverage of the existing literature, this review aims at pointing the major findings reported for these new classes of CO{sub 2} capture media.

Branched-DNA signal amplification combined with paper chromatography hybridization assay and used in hepatitis B virus DNA detection

International Nuclear Information System (INIS)

Fu, F.Z.; Liu, L.X.; Wang, W.Q.; Sun, S. H.; Liu, L.B.

2002-01-01

Nucleic acids detection method is vital to the clinical pathogen diagnosis. The established method can be classified into target direct amplification and signal amplification format according to the target DNA or RNA being directly amplified or not. Those methods have advantages and disadvantages respectively in the clinical application. In the United States of American, branched-DNA as a strong signal amplifier is broadly used in the quantification of the nucleic acids. To gain satisfied sensitivity, some expensive label molecular and instruments should be adopted. Personnel should be special trained to perform. Hence, those can't be widely carried out in the Third World. To avoid those disadvantages, we used the branched-DNA amplifier in the paper chromatography hybridization assay. Methods: Branched DNA signal amplifier and series of probes complementary to the nucleic acid sequence of hepatitis B virus (HBV) have been synthesized. HBV-DNA or it's capture probe were immobilized on the high flow nitrocellulose strip. Having loaded at one end of the strip in turn, probes or HBV-DNA in the hybridization solution migrate to the opposite end of the strip by capillary forces and hybridizes to the immobilized DNA. The branched-DNA signal amplifier and probe labeled with biotin or 32P were then loaded. Through streptavidin-alkaline phosphatase (SA-AP) conjugate and NBT/BCIP ( the specific chromogenic substrate of AP) or autoradiography, the result can be visualized by color reaction or image production on the X-ray film. Results: The sensitivity of this HBV-DNA detection method used probe labeled with biotin and 32P are 1ng and 10pg. The method using the probe labeled with biotin is simple and rapid (2h) without depending on special instruments, it also avoids the pollution of EtBr which can lead to tumor. And the method using the probe labeled with 32P is simple and sensitive, with the exception of long time autoradiography and the inconvenient isotopic disposal

Accurate detection of male subclinical genital tract infection via cervical culture and DNA hybridization assay of the female partner

NARCIS (Netherlands)

Trum, J. W.; Pannekoek, Y.; Spanjaard, L.; Bleker, O. P.; van der Veen, F.

2000-01-01

The accuracy of the PACE2 DNA hybridization assay of the cervix and cervical culture in female partners for the diagnosis of male subclinical genital tract infection were assessed in a male infertility population. A total of 184 men were screened for the presence of Chlamydia trachomatis, Ureaplasma

Comparison of hybrid capture and reverse transcriptase polymerase chain reaction methods in terms of diagnosing human cytomegalovirus infection in patients following hematopoietic stem cell transplantation

International Nuclear Information System (INIS)

Orsal, Arif S.; Ozsan, M.; Dolapci, I.; Tekeli, A.; Becksac, M.

2006-01-01

Human cytomegalovirus (CMV) is a life threatening cause of infection among hematopoietic stem cell recipients. Developing reliable methods in detecting the CMV infection is important to identify the patients at risk of CMV infection and disease. The aim of this study was to compare the 2 tests- hybrid capture test, which is routinely used in the diagnosis of CMV infection among hematopoietic stem cell recipients, and reverse transcriptase polymerase chain reaction (RT-PCR) detecting UL21.5 mRNA transcripts of the active virus. In this prospective study, a total of 178 blood samples obtained 35 patients following allogeneic hematopoietic stem cell transplantation at the Bone Marrow Transplantation Unit of the Hematology Department, Ibn-i-Sina Hospital of Ankara University School of Medicine, Turkey between January 2003 and September 2003 were analyzed. Hybrid capture and RT-PCR using UL21.5 gene transcript method to investigate HCMV in blood samples were performed at the department of Microbiology and Clinic Microbiology, Ankara University School of Medicine, Turkey. When Hybrid capture test was accepted as the golden standard, the sensitivity of Rt-PCR was 3%, specificity 100%, false negativity 67%, false positivity 0%, positive predictive value 100%, negative predictive value 74%, and accuracy was 77%. Improving this test by quantification, and application of additional gene transcripts, primarily the late gene transcripts can help increase the sensitivity and feasibility. (author)

Population-based rare variant detection via pooled exome or custom hybridization capture with or without individual indexing.

Science.gov (United States)

Ramos, Enrique; Levinson, Benjamin T; Chasnoff, Sara; Hughes, Andrew; Young, Andrew L; Thornton, Katherine; Li, Allie; Vallania, Francesco L M; Province, Michael; Druley, Todd E

2012-12-06

Rare genetic variation in the human population is a major source of pathophysiological variability and has been implicated in a host of complex phenotypes and diseases. Finding disease-related genes harboring disparate functional rare variants requires sequencing of many individuals across many genomic regions and comparing against unaffected cohorts. However, despite persistent declines in sequencing costs, population-based rare variant detection across large genomic target regions remains cost prohibitive for most investigators. In addition, DNA samples are often precious and hybridization methods typically require large amounts of input DNA. Pooled sample DNA sequencing is a cost and time-efficient strategy for surveying populations of individuals for rare variants. We set out to 1) create a scalable, multiplexing method for custom capture with or without individual DNA indexing that was amenable to low amounts of input DNA and 2) expand the functionality of the SPLINTER algorithm for calling substitutions, insertions and deletions across either candidate genes or the entire exome by integrating the variant calling algorithm with the dynamic programming aligner, Novoalign. We report methodology for pooled hybridization capture with pre-enrichment, indexed multiplexing of up to 48 individuals or non-indexed pooled sequencing of up to 92 individuals with as little as 70 ng of DNA per person. Modified solid phase reversible immobilization bead purification strategies enable no sample transfers from sonication in 96-well plates through adapter ligation, resulting in 50% less library preparation reagent consumption. Custom Y-shaped adapters containing novel 7 base pair index sequences with a Hamming distance of ≥2 were directly ligated onto fragmented source DNA eliminating the need for PCR to incorporate indexes, and was followed by a custom blocking strategy using a single oligonucleotide regardless of index sequence. These results were obtained aligning raw

Population-based rare variant detection via pooled exome or custom hybridization capture with or without individual indexing

Directory of Open Access Journals (Sweden)

Ramos Enrique

2012-12-01

Full Text Available Abstract Background Rare genetic variation in the human population is a major source of pathophysiological variability and has been implicated in a host of complex phenotypes and diseases. Finding disease-related genes harboring disparate functional rare variants requires sequencing of many individuals across many genomic regions and comparing against unaffected cohorts. However, despite persistent declines in sequencing costs, population-based rare variant detection across large genomic target regions remains cost prohibitive for most investigators. In addition, DNA samples are often precious and hybridization methods typically require large amounts of input DNA. Pooled sample DNA sequencing is a cost and time-efficient strategy for surveying populations of individuals for rare variants. We set out to 1 create a scalable, multiplexing method for custom capture with or without individual DNA indexing that was amenable to low amounts of input DNA and 2 expand the functionality of the SPLINTER algorithm for calling substitutions, insertions and deletions across either candidate genes or the entire exome by integrating the variant calling algorithm with the dynamic programming aligner, Novoalign. Results We report methodology for pooled hybridization capture with pre-enrichment, indexed multiplexing of up to 48 individuals or non-indexed pooled sequencing of up to 92 individuals with as little as 70 ng of DNA per person. Modified solid phase reversible immobilization bead purification strategies enable no sample transfers from sonication in 96-well plates through adapter ligation, resulting in 50% less library preparation reagent consumption. Custom Y-shaped adapters containing novel 7 base pair index sequences with a Hamming distance of ≥2 were directly ligated onto fragmented source DNA eliminating the need for PCR to incorporate indexes, and was followed by a custom blocking strategy using a single oligonucleotide regardless of index sequence

Evaluation of Carbon Dioxide Capture From Existing Coal Fired Plants by Hybrid Sorption Using Solid Sorbents

Energy Technology Data Exchange (ETDEWEB)

Benson, Steven [Univ. of North Dakota, Grand Forks, ND (United States); Srinivasachar, Srivats [Envergex LLC, Sturbridge, MA (United States); Laudal, Daniel [Univ. of North Dakota, Grand Forks, ND (United States); Browers, Bruce [Barr Engineering, Minneapolis, MN (United States)

2014-12-31

A novel hybrid solid sorbent technology for CO₂ capture and separation from coal combustion-derived flue gas was evaluated. The technology – Capture of CO₂ by Hybrid Sorption (CACHYS™) – is a solid sorbent technology based on the following ideas: 1) reduction of energy for sorbent regeneration, 2) utilization of novel process chemistry, 3) contactor conditions that minimize sorbent-CO₂ heat of reaction and promote fast CO₂ capture, and 4) low-cost method of heat management. This report provides key information developed during the course of the project that includes sorbent performance, energy for sorbent regeneration, physical properties of the sorbent, the integration of process components, sizing of equipment, and overall capital and operational cost of the integrated CACHYS™ system. Seven sorbent formulations were prepared and evaluated at the lab-scale for energy requirements and CO₂ capture performance. Sorbent heat of regeneration ranged from 30-80 kJ/mol CO₂ and was found to be dependent on process conditions. Two sorbent formulations (designated HCK-4 & HCK-7) were down-selected for additional fixed-bed testing. Additional testing involved subjecting the sorbents to 100 continuous cycles in the fixed-bed reactor to determine performance as a function of time. The working capacity achieved for HCK-4 sorbent ranged from 5.5-8.0 g CO₂/100 g sorbent, while the HCK-7 typically ranged from 8.0-10.0 g CO₂/100 g sorbent. Overall, there was no deterioration in capacity with continuous cycling for either sorbent. The CACHYS™ bench-scale testing system designed and fabricated under this award consists of a dual circulating fluidized-bed adsorber and a moving-bed regenerator. The system takes a flue gas slipstream from the University of North Dakota’s coal-fired steam plant. Prior to being sent to the adsorber, the flue gas is scrubbed to remove SO₂ and particulate. During parametric testing of the adsorber, CO₂ capture achieved using

Detection of human papillomavirus by hybrid capture and real time PCR methods in patients with chronic cervicitis and cervical intraepithelial

Directory of Open Access Journals (Sweden)

Elisha Khandker

2016-07-01

Full Text Available Background and objectives:Cervical cancer due to Human papillomavirus (HPV is one of the leading causes of morbidity and mortality in women. Testing of HPV can identify women who are at risk of cervical cancer. Nowadays, molecular methods like real time polymerase chain reaction (PCR and hybrid capture technique are applied for detecting HPV in cervical specimens. The objective of the present study was to determine the rate of HPV infection in women with chronic cervicitis and cervical intraepithelial neoplasia (CIN by a commercial real time polymerase chain reaction test kit and by a hybrid capture HPV DNA test. Methods:Women aged between 20 to 55 years with chronic cervicitis and CIN were enrolled in the study after obtaining informed consent. Cervical specimen was collected by using cervical brush and stored in transport medium until used. HPV was detected by High Risk Screen Real-TM Quant 2x (Sacace, Biotechnologies SrI, Italy real time PCR kit (HR RT-PCR and by Hybrid Capture-2 High-Risk HPV DNA (Hc-2; Digene Corporation, USA test. Results: Total 72 women with chronic cervicitis and CIN of different grades were included in the study. Out of this, HPV infection detected by HR RT-PCR was 31 (43% and by Hc-2 was 14 (19.4%. Both the tests were able to detect HPV infection in all the CIN 3 cases and in most of the CIN 2 cases. However, HR RT-PCR detected higher number of HPV in chronic cervicitis and CIN1 cases. Conclusion:The study has shown that HR RT-PCR and Hc-2 tests are equally effective in detecting HPV infection in patients with CIN 2 and CIN 3 lesions. However, HR RT-PCR is more sensitive test for detecting HPV in chronic cervicitis and early CIN lesions and, therefore can be used in epidemiological study to detect presence of HPV in general population. IMC J Med Sci 2016; 10(2: 45-48

A fluorometric lateral flow assay for visual detection of nucleic acids using a digital camera readout.

Science.gov (United States)

Magiati, Maria; Sevastou, Areti; Kalogianni, Despina P

2018-06-04

A fluorometric lateral flow assay has been developed for the detection of nucleic acids. The fluorophores phycoerythrin (PE) and fluorescein isothiocyanate (FITC) were used as labels, while a common digital camera and a colored vinyl-sheet, acting as a cut-off optical filter, are used for fluorescence imaging. After DNA amplification by polymerase chain reaction (PCR), the biotinylated PCR product is hybridized to its complementary probe that carries a poly(dA) tail at 3΄ edge and then applied to the lateral flow strip. The hybrids are captured to the test zone of the strip by immobilized poly(dT) sequences and detected by streptavidin-fluorescein and streptavidin-phycoerythrin conjugates, through streptavidin-biotin interaction. The assay is widely applicable, simple, cost-effective, and offers a large multiplexing potential. Its performance is comparable to assays based on the use of streptavidin-gold nanoparticles conjugates. As low as 7.8 fmol of a ssDNA and 12.5 fmol of an amplified dsDNA target were detectable. Graphical abstract Schematic presentation of a fluorometric lateral flow assay based on fluorescein and phycoerythrin fluorescent labels for the detection of single-stranded (ssDNA) and double-stranded DNA (dsDNA) sequences and using a digital camera readout. SA: streptavidin, BSA: Bovine Serum Albumin, B: biotin, FITC: fluorescein isothiocyanate, PE: phycoerythrin, TZ: test zone, CZ: control zone.

Using a hybrid subtyping model to capture patterns and dimensionality of depressive and anxiety symptomatology in the general population.

Science.gov (United States)

Wardenaar, Klaas J; Wanders, Rob B K; Ten Have, Margreet; de Graaf, Ron; de Jonge, Peter

2017-06-01

Researchers have tried to identify more homogeneous subtypes of major depressive disorder (MDD) with latent class analyses (LCA). However, this approach does no justice to the dimensional nature of psychopathology. In addition, anxiety and functioning-levels have seldom been integrated in subtyping efforts. Therefore, this study used a hybrid discrete-dimensional approach to identify subgroups with shared patterns of depressive and anxiety symptomatology, while accounting for functioning-levels. The Comprehensive International Diagnostic Interview (CIDI) 1.1 was used to assess previous-year depressive and anxiety symptoms in the Netherlands Mental Health Survey and Incidence Study-1 (NEMESIS-1; n=5583). The data were analyzed with factor analyses, LCA and hybrid mixed-measurement item response theory (MM-IRT) with and without functioning covariates. Finally, the classes' predictors (measured one year earlier) and outcomes (measured two years later) were investigated. A 3-class MM-IRT model with functioning covariates best described the data and consisted of a 'healthy class' (74.2%) and two symptomatic classes ('sleep/energy' [13.4%]; 'mood/anhedonia' [12.4%]). Factors including older age, urbanicity, higher severity and presence of 1-year MDD predicted membership of either symptomatic class vs. the healthy class. Both symptomatic classes showed poorer 2-year outcomes (i.e. disorders, poor functioning) than the healthy class. The odds of MDD after two years were especially increased in the mood/anhedonia class. Symptoms were assessed for the past year whereas current functioning was assessed. Heterogeneity of depression and anxiety symptomatology are optimally captured by a hybrid discrete-dimensional subtyping model. Importantly, accounting for functioning-levels helps to capture clinically relevant interpersonal differences. Copyright © 2017 Elsevier B.V. All rights reserved.

Rapid detection of Cyprinid herpesvirus-3 (CyHV-3) using a gold nanoparticle-based hybridization assay.

Science.gov (United States)

Saleh, Mona; El-Matbouli, Mansour

2015-06-01

Cyprinid herpesvirus-3 (CyHV-3) is a highly infectious pathogen that causes fatal disease in common and koi carp Cyprinus carpio L. CyHV-3 detection is usually based on virus propagation or amplification of the viral DNA using the PCR or LAMP techniques. However, due to the limited susceptibility of cells used for propagation, it is not always possible to successfully isolate CyHV-3 even from tissue samples that have high virus titres. All previously described detection methods including PCR-based assays are time consuming, laborious and require specialized equipment. To overcome these limitations, gold nanoparticles (AuNPs) have been explored for direct and sensitive detection of DNA. In this study, a label-free colorimetric nanodiagnostic method for direct detection of unamplified CyHV-3 DNA using gold nanoparticles is introduced. Under appropriate conditions, DNA probes hybridize with their complementary target sequences in the sample DNA, which results in aggregation of the gold nanoparticles and a concomitant colour change from red to blue, whereas test samples with non complementary DNA sequences remain red. In this study, gold nanoparticles were used to develop and evaluate a specific and sensitive hybridization assay for direct and rapid detection of the highly infectious pathogen termed Cyprinid herpesvirus-3. Copyright © 2015 Elsevier B.V. All rights reserved.

Comparison of the accuracy of Hybrid Capture II and polymerase chain reaction in detecting clinically important cervical dysplasia: a systematic review and meta-analysis

International Nuclear Information System (INIS)

Luu, Hung N; Dahlstrom, Kristina R; Mullen, Patricia Dolan; VonVille, Helena M; Scheurer, Michael E

2013-01-01

The effectiveness of screening programs for cervical cancer has benefited from the inclusion of Human papillomavirus (HPV) DNA assays; which assay to choose, however, is not clear based on previous reviews. Our review addressed test accuracy of Hybrid Capture II (HCII) and polymerase chain reaction (PCR) assays based on studies with stronger designs and with more clinically relevant outcomes. We searched OvidMedline, PubMed, and the Cochrane Library for English language studies comparing both tests, published 1985–2012, with cervical dysplasia defined by the Bethesda classification. Meta-analysis provided pooled sensitivity, specificity, and 95% confidence intervals (CIs); meta-regression identified sources of heterogeneity. From 29 reports, we found that the pooled sensitivity and specificity to detect high-grade squamous intraepithelial lesion (HSIL) was higher for HCII than PCR (0.89 [CI: 0.89–0.90] and 0.85 [CI: 0.84–0.86] vs. 0.73 [CI: 0.73–0.74] and 0.62 [CI: 0.62–0.64]). Both assays had higher accuracy to detect cervical dysplasia in Europe than in Asia-Pacific or North America (diagnostic odd ratio – dOR = 4.08 [CI: 1.39–11.91] and 4.56 [CI: 1.86–11.17] for HCII vs. 2.66 [CI: 1.16–6.53] and 3.78 [CI: 1.50–9.51] for PCR) and accuracy to detect HSIL than atypical squamous cells of undetermined significance (ASCUS)/ low-grade squamous intraepithelial lesion (LSIL) (HCII-dOR = 9.04 [CI: 4.12–19.86] and PCR-dOR = 5.60 [CI: 2.87–10.94]). For HCII, using histology as a gold standard results in higher accuracy than using cytology (dOR = 2.87 [CI: 1.31–6.29]). Based on higher test accuracy, our results support the use of HCII in cervical cancer screening programs. The role of HPV type distribution should be explored to determine the worldwide comparability of HPV test accuracy

Optimization of AFP-radioimmunoassay using Antibody Capture Technique

International Nuclear Information System (INIS)

Moustafa, K.A.

2003-01-01

Alpha-fetoprotein (AFP) is a substance produced by the unborn baby. When the neural tube is not properly formed large amounts of AFP pass into the amniotic fluid and reach the mother's blood. By measuring AFP in the mother's blood and amniotic fluid, it is possible to tell whether or not there is a chance that the unborn baby has a neural tube defect. AFP also used as a tumor marker for hepatocellular carcinoma. There are many different techniques for measuring AFP in blood, but the most accurate one is the immunoassay technique. The immunoassays can be classified on the basis of methodology into three classes; (1) the antibody capture assays, (2) the antigen capture assay, (3)the two-antibody sandwich assays. In this present study, the antibody capture assay in which the antigen is attached to a solid support, and labeled antibody is allowed to bind, will be optimized

Development of a PCR/LDR/flow-through hybridization assay using a capillary tube, probe DNA-immobilized magnetic beads and chemiluminescence detection.

Science.gov (United States)

Hommatsu, Manami; Okahashi, Hisamitsu; Ohta, Keisuke; Tamai, Yusuke; Tsukagoshi, Kazuhiko; Hashimoto, Masahiko

2013-01-01

A polymerase chain reaction (PCR)/ligase detection reaction (LDR)/flow-through hybridization assay using chemiluminescence (CL) detection was developed for analyzing point mutations in gene fragments with high diagnostic value for colorectal cancers. A flow-through hybridization format using a capillary tube, in which probe DNA-immobilized magnetic beads were packed, provided accelerated hybridization kinetics of target DNA (i.e. LDR product) to the probe DNA. Simple fluid manipulations enabled both allele-specific hybridization and the removal of non-specifically bound DNA in the wash step. Furthermore, the use of CL detection greatly simplified the detection scheme, since CL does not require a light source for excitation of the fluorescent dye tags on the LDR products. Preliminary results demonstrated that this analytical system could detect both homozygous and heterozygous mutations, without the expensive instrumentation and cumbersome procedures required by conventional DNA microarray-based methods.

Development and characterization of a magnetic bead-quantum dot nanoparticles based assay capable of Escherichia coli O157:H7 quantification

Energy Technology Data Exchange (ETDEWEB)

Kim, Gha-Young [Department of Civil Engineering, Auburn University, Auburn, AL 36849 (United States); Son, Ahjeong, E-mail: ason@auburn.edu [Department of Civil Engineering, Auburn University, Auburn, AL 36849 (United States)

2010-09-10

The development and characterization of a magnetic bead (MB)-quantum dot (QD) nanoparticles based assay capable of quantifying pathogenic bacteria is presented here. The MB-QD assay operates by having a capturing probe DNA selectively linked to the signaling probe DNA via the target genomic DNA (gDNA) during DNA hybridization. The signaling probe DNA is labeled with fluorescent QD{sub 565} which serves as a reporter. The capturing probe DNA is conjugated simultaneously to a MB and another QD{sub 655}, which serve as a carrier and an internal standard, respectively. Successfully captured target gDNA is separated using a magnetic field and is quantified via a spectrofluorometer. The use of QDs (i.e., QD{sub 565}/QD{sub 655}) as both a fluorescence label and an internal standard increased the sensitivity of the assay. The passivation effect and the molar ratio between QD and DNA were optimized. The MB-QD assay demonstrated a detection limit of 890 zeptomolar (i.e., 10{sup -21} mol L{sup -1}) concentration for the linear single stranded DNA (ssDNA). It also demonstrated a detection limit of 87 gene copies for double stranded DNA (dsDNA) eaeA gene extracted from pure Escherichia coli (E. coli) O157:H7 culture. Its corresponding dynamic range, sensitivity, and selectivity were also presented. Finally, the bacterial gDNA of E. coli O157:H7 was used to highlight the MB-QD assay's ability to detect below the minimum infective dose (i.e., 100 organisms) of E. coli O157:H7 in water environment.

Establishment of 60Co dose calibration curve using fluorescent in situ hybridization assay technique: Result of preliminary study

International Nuclear Information System (INIS)

Rahimah Abdul Rahim; Noriah Jamal; Noraisyah Mohd Yusof; Juliana Mahamad Napiah; Nelly Bo Nai Lee

2010-01-01

This study aims at establishing an in-vitro 60 Co dose calibration curve using Fluorescent In-Situ Hybridization assay technique for the Malaysian National Bio dosimetry Laboratory. Blood samples collected from a female healthy donor were irradiated with several doses of 60 Co radiation. Following culturing of lymphocytes, microscopic slides are prepared, denatured and hybridized. The frequencies of translocation are estimated in the metaphases. A calibration curve was then generated using a regression technique. It shows a good fit to a linear-quadratic model. The results of this study might be useful in estimating absorbed dose for the individual exposed to ionizing radiation retrospectively. This information may be useful as a guide for medical treatment for the assessment of possible health consequences. (author)

An Aptamer Bio-barCode (ABC) assay using SPR, RNase H, and probes with RNA and gold-nanorods for anti-cancer drug screening.

Science.gov (United States)

Loo, Jacky Fong-Chuen; Yang, Chengbin; Tsang, Hing Lun; Lau, Pui Man; Yong, Ken-Tye; Ho, Ho Pui; Kong, Siu Kai

2017-10-07

With modifications to an ultra-sensitive bio-barcode (BBC) assay, we have developed a next generation aptamer-based bio-barcode (ABC) assay to detect cytochrome-c (Cyto-c), a cell death marker released from cancer cells, for anti-cancer drug screening. An aptamer is a short single-stranded DNA selected from a synthetic DNA library that is capable of binding to its target with high affinity and specificity based on its unique DNA sequence and 3D structure after folding. Similar to the BBC assay, Cyto-c is captured by a micro-magnetic particle (MMP) coated with capturing antibodies (Ab) and an aptamer specifically against Cyto-c to form sandwich structures ([MMP-Ab]-[Cyto-c]-[Aptamer]). After washing and melting, our aptamers, acting as a DNA bio-barcode, are released from the sandwiches and hybridized with the probes specially designed for RNase H for surface plasmon resonance (SPR) sensing. In an aptamer-probe duplex, RNase H digests the RNA in the probe and releases the intact aptamer for another round of hybridization and digestion. With signal enhancement effects from gold-nanorods (Au-NRs) on probes for SPR sensing, the detection limit was found to be 1 nM for the aptamer and 80 pM for Cyto-c. Without the time-consuming DNA amplification steps by PCR, the detection process of this new ABC assay can be completed within three hours. As a proof-of-concept, phenylarsine oxide was found to be a potent agent to kill liver cancer cells with multi-drug resistance at the nano-molar level. This approach thus provides a fast, sensitive and robust tool for anti-cancer drug screening.

Detection of H5 Avian Influenza Viruses by Antigen-Capture Enzyme-Linked Immunosorbent Assay Using H5-Specific Monoclonal Antibody▿

OpenAIRE

He, Qigai; Velumani, Sumathy; Du, Qingyun; Lim, Chee Wee; Ng, Fook Kheong; Donis, Ruben; Kwang, Jimmy

2007-01-01

The unprecedented spread of highly pathogenic avian influenza virus subtype H5N1 in Asia and Europe is threatening animals and public health systems. Effective diagnosis and control management are needed to control the disease. To this end, we developed a panel of monoclonal antibodies (MAbs) against the H5N1 avian influenza virus (AIV) and implemented an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) to detect the H5 viral antigen. Mice immunized with denatured hemagglutinin (H...

Performance of PCR-reverse blot hybridization assay for detection of rifampicin-resistant Mycobacterium leprae.

Science.gov (United States)

Wang, Hye-young; Kim, Hyunjung; Kim, Yeun; Bang, Hyeeun; Kim, Jong-Pill; Hwang, Joo Hwan; Cho, Sang-Nae; Kim, Tae Ue; Lee, Hyeyoung

2015-10-01

Drug resistance in Mycobacterium leprae is a significant problem in countries where leprosy is endemic. A sensitive, specific, and high-throughput reverse blot hybridization assay (REBA) for the detection of genotypic resistance to rifampicin (RIF) was designed and evaluated. It has been shown that resistance to RIF in M. leprae involves mutations in the rpoB gene encoding the -subunit of the RNA polymerase. The PCR-REBA simultaneously detects both 6 wild-type regions and 5 different mutations (507 AGC, 513 GTG, 516 TAT, 531 ATG, and 531 TTC) including the most prevalent mutations at positions 507 and 531. Thirty-one clinical isolates provided by Korea Institute of Hansen-s Disease were analyzed by PCR-REBA with RIF resistance of rpoB gene. As a result, missense mutations at codons 507 AGC and 531 ATG with 2-nucleotide substitutions were found in one sample, and a missense mutation at codon 516 TAT and ΔWT6 (deletion of 530-534) was found in another sample. These cases were confirmed by DNA sequence analysis. This rapid, simple, and highly sensitive assay provides a practical alternative to sequencing for genotypic evaluation of RIF resistance in M. leprae.

Detection of cystic fibrosis transmembrane conductance regulator ΔF508 gene mutation using a paper-based nucleic acid hybridization assay and a smartphone camera.

Science.gov (United States)

Malhotra, Karan; Noor, M Omair; Krull, Ulrich J

2018-05-29

Diagnostic technology that makes use of paper platforms in conjunction with the ubiquitous availability of digital cameras in cellular telephones and personal assistive devices offers opportunities for development of bioassays that are cost effective and widely distributed. Assays that operate effectively in aqueous solution require further development for implementation in paper substrates, overcoming issues associated with surface interactions on a matrix that offers a large surface-to-volume ratio and constraints on convective mixing. This report presents and compares two related methods for determination of oligonucleotides that serve as indicators of cystic fibrosis, differentiating between the normal wild-type sequence, and a mutant-type sequence that has a 3-base replacement. The transduction strategy operates by selective hybridization of oligonucleotide probes that are conjugated to fluorescent quantum dots, where hybridization of target sequences causes a molecular fluorophore to approach the quantum dot and become emissive through fluorescence resonance energy transfer. Detection can rely on hybridization of a target that is labelled with Cy3 fluorophore, or in the presence of an unlabelled target when a sandwich assay format is implemented with a labelled reporter oligonucleotide. Selectivity to determine the presence of mismatched sequences involves appropriate selection of nucleotide sequences to set melt temperatures, in conjunction with control of stringency conditions using formamide as a chaotrope. It was determined that both direct and sandwich assays on paper substrates are able to distinguish between wild-type and mutant-type samples.

Effects of Bonding Types and Functional Groups on CO 2 Capture using Novel Multiphase Systems of Liquid-like Nanoparticle Organic Hybrid Materials

KAUST Repository

Lin, Kun-Yi Andrew

2011-08-01

Novel liquid-like nanoparticle organic hybrid materials (NOHMs) which possess unique features including negligible vapor pressure and a high degree of tunability were synthesized and their physical and chemical properties as well as CO 2 capture capacities were investigated. NOHMs can be classified based on the synthesis methods involving different bonding types, the existence of linkers, and the addition of task-specific functional groups including amines for CO 2 capture. As a canopy of polymeric chains was grafted onto the nanoparticle cores, the thermal stability of the resulting NOHMs was improved. In order to isolate the entropy effect during CO 2 capture, NOHMs were first prepared using polymers that do not contain functional groups with strong chemical affinity toward CO 2. However, it was found that even ether groups on the polymeric canopy contributed to CO 2 capture in NOHMs via Lewis acid-base interactions, although this effect was insignificant compared to the effect of task-specific functional groups such as amine. In all cases, a higher partial pressure of CO 2 was more favorable for CO 2 capture, while a higher temperature caused an adverse effect. Multicyclic CO 2 capture tests confirmed superior recyclability of NOHMs and NOHMs also showed a higher selectivity toward CO 2 over N 2O, O 2 and N 2. © 2011 American Chemical Society.

Capturing multi-stage fuzzy uncertainties in hybrid system dynamics and agent-based models for enhancing policy implementation in health systems research.

Science.gov (United States)

Liu, Shiyong; Triantis, Konstantinos P; Zhao, Li; Wang, Youfa

2018-01-01

In practical research, it was found that most people made health-related decisions not based on numerical data but on perceptions. Examples include the perceptions and their corresponding linguistic values of health risks such as, smoking, syringe sharing, eating energy-dense food, drinking sugar-sweetened beverages etc. For the sake of understanding the mechanisms that affect the implementations of health-related interventions, we employ fuzzy variables to quantify linguistic variable in healthcare modeling where we employ an integrated system dynamics and agent-based model. In a nonlinear causal-driven simulation environment driven by feedback loops, we mathematically demonstrate how interventions at an aggregate level affect the dynamics of linguistic variables that are captured by fuzzy agents and how interactions among fuzzy agents, at the same time, affect the formation of different clusters(groups) that are targeted by specific interventions. In this paper, we provide an innovative framework to capture multi-stage fuzzy uncertainties manifested among interacting heterogeneous agents (individuals) and intervention decisions that affect homogeneous agents (groups of individuals) in a hybrid model that combines an agent-based simulation model (ABM) and a system dynamics models (SDM). Having built the platform to incorporate high-dimension data in a hybrid ABM/SDM model, this paper demonstrates how one can obtain the state variable behaviors in the SDM and the corresponding values of linguistic variables in the ABM. This research provides a way to incorporate high-dimension data in a hybrid ABM/SDM model. This research not only enriches the application of fuzzy set theory by capturing the dynamics of variables associated with interacting fuzzy agents that lead to aggregate behaviors but also informs implementation research by enabling the incorporation of linguistic variables at both individual and institutional levels, which makes unstructured linguistic data

Hydraulic Hybrid Fleet Vehicle Testing | Transportation Research | NREL

Science.gov (United States)

Hydraulic Hybrid Fleet Vehicle Evaluations Hydraulic Hybrid Fleet Vehicle Evaluations How Hydraulic Hybrid Vehicles Work Hydraulic hybrid systems can capture up to 70% of the kinetic energy that would -pressure reservoir to a high-pressure accumulator. When the vehicle accelerates, fluid in the high-pressure

CO 2 Capture Capacity and Swelling Measurements of Liquid-like Nanoparticle Organic Hybrid Materials via Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy

KAUST Repository

Park, Youngjune

2012-01-12

Novel nanoparticle organic hybrid materials (NOHMs), which are comprised of organic oligomers or polymers tethered to an inorganic nanosized cores of various sizes, have been synthesized, and their solvating property for CO 2 was investigated using attenuated total reflectance (ATR) Fourier transform infrared (FT-IR) spectroscopy. Simultaneous measurements of CO 2 capture capacity and swelling behaviors of polyetheramine (Jeffamine M-2070) and its corresponding NOHMs (NOHM-I-PE2070) were reported at temperatures of (298, 308, 323 and 353) K and CO 2 pressure conditions ranging from (0 to 5.5) MPa. The polymeric canopy, or polymer bound to the nanoparticle surface, showed significantly less swelling behavior with enhanced or comparable CO 2 capture capacity compared to pure unbound polyetheramine. © 2011 American Chemical Society.

Human Papillomavirus Assays and Cytology in Primary Cervical Screening of Women Aged 30 Years and Above

DEFF Research Database (Denmark)

Rebolj, Matejka; Bonde, Jesper; Preisler, Sarah

2016-01-01

In women aged ≥30 years, Human Papillomavirus testing will replace cytology for primary cervical screening. We compared Hybrid Capture 2 (HC2), cobas, CLART, and APTIMA HPV assays with cytology on 2869 SurePath samples from women undergoing routine screening at 30-65 years in Copenhagen, Denmark....... Women with cytological abnormalities were managed according to routine recommendations, with 92% completeness. Those with cytology-normal/HPV-positive samples (on any of the four assays) were invited for repeated cytology and HPV testing in 1.5 year, and 58% had additional testing. HPV testing detected...... more ≥CIN3 than cytology (HC2: 35, cobas, CLART: 37, APTIMA: 34, cytology: 31), although statistically the differences were not significant. Cobas and CLART detected significantly more ≥CIN2 than cytology (cobas, CLART: 49, cytology: 39). The proportion of women with false-positive test results...

Chromogenic in situ hybridization is a reliable assay for detection of ALK rearrangements in adenocarcinomas of the lung.

Science.gov (United States)

Schildhaus, Hans-Ulrich; Deml, Karl-Friedrich; Schmitz, Katja; Meiboom, Maren; Binot, Elke; Hauke, Sven; Merkelbach-Bruse, Sabine; Büttner, Reinhard

2013-11-01

Reliable detection of anaplastic lymphoma kinase (ALK) rearrangements is a prerequisite for personalized treatment of lung cancer patients, as ALK rearrangements represent a predictive biomarker for the therapy with specific tyrosine kinase inhibitors. Currently, fluorescent in situ hybridization (FISH) is considered to be the standard method for assessing formalin-fixed and paraffin-embedded tissue for ALK inversions and translocations. However, FISH requires a specialized equipment, the signals fade rapidly and it is difficult to detect overall morphology and tumor heterogeneity. Chromogenic in situ hybridization (CISH) has been successfully introduced as an alternative test for the detection of several genetic aberrations. This study validates a newly developed ALK CISH assay by comparing FISH and CISH signal patterns in lung cancer samples with and without ALK rearrangements. One hundred adenocarcinomas of the lung were included in this study, among them 17 with known ALK rearrangement. FISH and CISH were carried out and evaluated according to the manufacturers' recommendations. For both assays, tumors were considered positive if ≥15% of tumor cells showed either isolated 3' signals or break-apart patterns or a combination of both. A subset of tumors was exemplarily examined by using a novel EML4 (echinoderm microtubule-associated protein-like 4) CISH probe. Red, green and fusion CISH signals were clearcut and different signal patterns were easily recognized. The percentage of aberrant tumor cells was statistically highly correlated (PCISH. On the basis of 86 samples that were evaluable by ALK CISH, we found a 100% sensitivity and 100% specificity of this assay. Furthermore, EML4 rearrangements could be recognized by CISH. CISH is a highly reliable, sensitive and specific method for the detection of ALK gene rearrangements in pulmonary adenocarcinomas. Our results suggest that CISH might serve as a suitable alternative to FISH, which is the current gold

Rapid and Simple Detection of Hot Spot Point Mutations of Epidermal Growth Factor Receptor, BRAF, and NRAS in Cancers Using the Loop-Hybrid Mobility Shift Assay

Science.gov (United States)

Matsukuma, Shoichi; Yoshihara, Mitsuyo; Kasai, Fumio; Kato, Akinori; Yoshida, Akira; Akaike, Makoto; Kobayashi, Osamu; Nakayama, Haruhiko; Sakuma, Yuji; Yoshida, Tsutomu; Kameda, Yoichi; Tsuchiya, Eiju; Miyagi, Yohei

2006-01-01

A simple and rapid method to detect the epidermal growth factor receptor hot spot mutation L858R in lung adenocarcinoma was developed based on principles similar to the universal heteroduplex generator technology. A single-stranded oligonucleotide with an internal deletion was used to generate heteroduplexes (loop-hybrids) bearing a loop in the complementary strand derived from the polymerase chain reaction product of the normal or mutant allele. By placing deletion in the oligonucleotide adjacent to the mutational site, difference in electrophoretic mobility between loop-hybrids with normal and mutated DNA was distinguishable in a native polyacrylamide gel. The method was also modified to detect in-frame deletion mutations of epidermal growth factor receptor in lung adenocarcinomas. In addition, the method was adapted to detect hot spot mutations in the B-type Raf kinase (BRAF) at V600 and in a Ras-oncogene (NRAS) at Q61, the mutations commonly found in thyroid carcinomas. Our mutation detection system, designated the loop-hybrid mobility shift assay was sensitive enough to detect mutant DNA comprising 7.5% of the total DNA. As a simple and straightforward mutation detection technique, loop-hybrid mobility shift assay may be useful for the molecular diagnosis of certain types of clinical cancers. Other applications are also discussed. PMID:16931592

Hybrid Capture-Based Comprehensive Genomic Profiling Identifies Lung Cancer Patients with Well-Characterized Sensitizing Epidermal Growth Factor Receptor Point Mutations That Were Not Detected by Standard of Care Testing.

Science.gov (United States)

Suh, James H; Schrock, Alexa B; Johnson, Adrienne; Lipson, Doron; Gay, Laurie M; Ramkissoon, Shakti; Vergilio, Jo-Anne; Elvin, Julia A; Shakir, Abdur; Ruehlman, Peter; Reckamp, Karen L; Ou, Sai-Hong Ignatius; Ross, Jeffrey S; Stephens, Philip J; Miller, Vincent A; Ali, Siraj M

2018-03-14

In our recent study, of cases positive for epidermal growth factor receptor ( EGFR ) exon 19 deletions using comprehensive genomic profiling (CGP), 17/77 (22%) patients with prior standard of care (SOC) EGFR testing results available were previously negative for exon 19 deletion. Our aim was to compare the detection rates of CGP versus SOC testing for well-characterized sensitizing EGFR point mutations (pm) in our 6,832-patient cohort. DNA was extracted from 40 microns of formalin-fixed paraffin-embedded sections from 6,832 consecutive cases of non-small cell lung cancer (NSCLC) of various histologies (2012-2015). CGP was performed using a hybrid capture, adaptor ligation-based next-generation sequencing assay to a mean coverage depth of 576×. Genomic alterations (pm, small indels, copy number changes and rearrangements) involving EGFR were recorded for each case and compared with prior testing results if available. Overall, there were 482 instances of EGFR exon 21 L858R (359) and L861Q (20), exon 18 G719X (73) and exon 20 S768I (30) pm, of which 103 unique cases had prior EGFR testing results that were available for review. Of these 103 cases, CGP identified 22 patients (21%) with sensitizing EGFR pm that were not detected by SOC testing, including 9/75 (12%) patients with L858R, 4/7 (57%) patients with L861Q, 8/20 (40%) patients with G719X, and 4/7 (57%) patients with S768I pm (some patients had multiple EGFR pm). In cases with available clinical data, benefit from small molecule inhibitor therapy was observed. CGP, even when applied to low tumor purity clinical-grade specimens, can detect well-known EGFR pm in NSCLC patients that would otherwise not be detected by SOC testing. Taken together with EGFR exon 19 deletions, over 20% of patients who are positive for EGFR -activating mutations using CGP are previously negative by SOC EGFR mutation testing, suggesting that thousands of such patients per year in the U.S. alone could experience improved clinical

Chemosensitivity and radiosensitivity of small cell lung cancer cell lines studied by a newly developed 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) hybrid assay

International Nuclear Information System (INIS)

Hida, T.; Ueda, R.; Takahashi, T.; Watanabe, H.; Kato, T.; Suyama, M.; Sugiura, T.; Ariyoshi, Y.

1989-01-01

The 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) hybrid assay was developed by technically combining the human tumor clonogenic assay and the MTT assay to make the most of both assays. This assay was able to estimate the in vitro growth of cultured cell lines and of tumor cells in pleural effusion, suggesting the possibility of its use for assessment of chemosensitivity and radiosensitivity of fresh tumor samples. Multiple cell lines [including morphological and/or phenotypical in vitro converters and cisplatin (CDDP)-resistant lines] were established from three patients with small cell lung cancer at different stages of the disease. Chemosensitivity of these cell lines to four commonly used chemotherapeutic drugs was tested by the MTT hybrid assay. SK1 and SK3 lines were established from Patient S. K. before and after chemotherapy and radiotherapy, respectively. SK3/CDDP, a CDDP-resistant line derived from the SK3 line, was 30-fold more resistant to CDDP [50% inhibiting dose (IC50), 21.5 micrograms/ml] than the SK1 line. In Patient M. O., MOA2/CDDP, a CDDP-resistant line derived from MOA2 (an in vitro converter from the MO line), was 41-fold more resistant to CDDP (IC50, 37 micrograms/ml) than the parent MO line. From Patient T. M., TM1 and TM2 lines were established before and after chemotherapy, respectively. The latter showed 6-fold more resistance to CDDP than the former. Chemosensitivity of these lines to three other drugs, 4-hydroperoxycyclophosphamide, Adriamycin, and etoposide, suggested cross-resistance between CDDP and 4-hydroperoxycyclophosphamide. Radiosensitivity study was also carried out with the MTT hybrid assay. The MOA2 line was more resistant [Do, 3.0 Gy; extrapolation number (n), 4.0] than the parental MO line (Do, 1.6 Gy; n, 2.1). There was no clear difference in radiosensitivity between the cell lines established before and after radiation therapy in Patient S. K

A two-magnet strategy for improved mixing and capture from biofluids

Science.gov (United States)

Doyle, Andrew B.; Haselton, Frederick R.

2016-01-01

Magnetic beads are a popular method for concentrating biomolecules from solution and have been more recently used in multistep pre-arrayed microfluidic cartridges. Typical processing strategies rely on a single magnet, resulting in a tight cluster of beads and requiring long incubation times to achieve high capture efficiencies, especially in highly viscous patient samples. This report describes a two-magnet strategy to improve the interaction of the bead surface with the surrounding fluid inside of a pre-arrayed, self-contained assay-in-a-tube. In the two-magnet system, target biomarker capture occurs at a rate three times faster than the single-magnet system. In clinically relevant biomatrices, we find a 2.5-fold improvement in biomarker capture at lower sample viscosities with the two-magnet system. In addition, we observe a 20% increase in the amount of protein captured at high viscosity for the two-magnet configuration relative to the single magnet approach. The two-magnet approach offers a means to achieve higher biomolecule extraction yields and shorter assay times in magnetic capture assays and in self-contained processor designs. PMID:27158286

Diagnosis of visceral Leishmaniasis in asymptomatic dogs by the KDNA PCR-hybridization assay using noninvasive samples

International Nuclear Information System (INIS)

Leite, Rodrigo Souza; Andrade, Antero Silva Ribeiro de; Ferreira, Sydney de Almeida; Ituassu, Leonardo Trindade; Melo, Maria Norma de

2009-01-01

The visceral leishmaniasis (VL) in Brazil is caused by Leishmania (Leishmania) chagasi and the asymptomatic dogs may transmit the parasite to sand flies vectors. The VL epidemiological control in Brazil involves the elimination of seropositive dogs, insecticide treatment and systematic treatment of human cases. Therefore, the accurate diagnosis is important in order to avoid the disease transmission or unnecessary culling of dogs. Serological tests are used for screening of dogs. However, these techniques present limitations. The Polymerase Chain Reaction (PCR) is an attractive alternative to the diagnosis in this context; but non-invasive samplings have great importance because they are simpler, painless and less resisted by dog-owners. This study aimed at evaluating conjunctival swab (CS) for canine VL diagnosis. In this methodology a sterile cotton swab is used to sampling the dog conjunctiva in both eyes. Thirty asymptomatic seropositive dogs were used. The samples were analyzed by the kDNA PCR-hybridization procedure in which the PCR products are hybridized with cloned kDNA mini-circles labeled with 32 P[]dCTP. In addition, blood (B) was collected from each animal. L. chagasi was identified in 90% of CS samples and 13,6% of B samples. The high sensitivity obtained with asymptomatic dogs, in which the diagnosis is more difficult due the low number of parasites in the samples, allow concluding that the conjunctival swab associated to the kDNA PCR-hybridization assay provides a valuable alternative tool for the direct diagnosis of canine leishmaniasis. (author)

EVALUATION OF CARBON DIOXIDE CAPTURE FROM EXISTING COAL FIRED PLANTS BY HYBRID SORPTION USING SOLID SORBENTS

Energy Technology Data Exchange (ETDEWEB)

Benson, Steven; Palo, Daniel; Srinivasachar, Srivats; Laudal, Daniel

2014-12-01

Under contract DE-FE0007603, the University of North Dakota conducted the project Evaluation of Carbon Dioxide Capture from Existing Coal Fired Plants by Hybrid Sorption Using Solid Sorbents. As an important element of this effort, an Environmental Health and Safety (EH&S) Assessment was conducted by Barr Engineering Co. (Barr) in association with the University of North Dakota. The assessment addressed air and particulate emissions as well as solid and liquid waste streams. The magnitude of the emissions and waste streams was estimated for evaluation purposes. EH&S characteristics of materials used in the system are also described. This document contains data based on the mass balances from both the 40 kJ/mol CO2 and 80 kJ/mol CO2 desorption energy cases evaluated in the Final Technical and Economic Feasibility study also conducted by Barr Engineering.

Ultrasensitive Faraday cage-type electrochemiluminescence assay for femtomolar miRNA-141 via graphene oxide and hybridization chain reaction-assisted cascade amplification.

Science.gov (United States)

Lu, Jing; Wu, Lin; Hu, Yufang; Wang, Sui; Guo, Zhiyong

2018-06-30

In this study, a novel electrochemiluminescence (ECL) biosensor for sensitive detection of femtomolar miRNA-141 was constructed on the basis of Faraday cage-type strategy via graphene oxide (GO) and hybridization chain reaction (HCR)-assisted cascade amplification. A capture probe (CP) was immobilized on Fe 3 O 4 @SiO 2 @Au nanoparticles as capture unit, which could catch the miRNA-141, and the immobilization of the signal unit (Ru(phen) 3 2+ -HCR/GO) was allowed via nucleic acid hybridization. The prepared biosensor exhibited two advantages for signal amplification: firstly, GO could lap on the electrode surface directly, extending Outer Helmholtz Plane (OHP) of the sensor due to the large surface area and good electronic transport property; secondly, HCR-assisted cascade amplification was designed by anchoring all HCR products on the GO surface, then embedding Ru(phen) 3 2+ as a signal readout pathway. All these signal molecules could take part in electrochemical reactions, thus further enhancing the ECL signal drastically. Therefore, the proposed sensor constructed by integrating HCR with Faraday cage-type strategy displayed an ultrasensitive detection platform for the miRNA-141 with a low detection limit of 0.03 fM. In addition, this proposed biosensor provides a universal platform for analysis of other microRNAs. Copyright © 2018 Elsevier B.V. All rights reserved.

Calibration and LOD/LOQ estimation of a chemiluminescent hybridization assay for residual DNA in recombinant protein drugs expressed in E. coli using a four-parameter logistic model.

Science.gov (United States)

Lee, K R; Dipaolo, B; Ji, X

2000-06-01

Calibration is the process of fitting a model based on reference data points (x, y), then using the model to estimate an unknown x based on a new measured response, y. In DNA assay, x is the concentration, and y is the measured signal volume. A four-parameter logistic model was used frequently for calibration of immunoassay when the response is optical density for enzyme-linked immunosorbent assay (ELISA) or adjusted radioactivity count for radioimmunoassay (RIA). Here, it is shown that the same model or a linearized version of the curve are equally useful for the calibration of a chemiluminescent hybridization assay for residual DNA in recombinant protein drugs and calculation of performance measures of the assay.

Diagnosis of visceral Leishmaniasis in asymptomatic dogs by the KDNA PCR-hybridization assay using noninvasive samples

Energy Technology Data Exchange (ETDEWEB)

Leite, Rodrigo Souza; Andrade, Antero Silva Ribeiro de [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil). Lab. de Radiobiologia], e-mail: rleite2005@gmail.com; Ferreira, Sydney de Almeida; Ituassu, Leonardo Trindade; Melo, Maria Norma de [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Centro de Ciencias Biologicas. Dept. de Parasitologia], e-mail: saninoalmeida@gmail.com

2009-07-01

The visceral leishmaniasis (VL) in Brazil is caused by Leishmania (Leishmania) chagasi and the asymptomatic dogs may transmit the parasite to sand flies vectors. The VL epidemiological control in Brazil involves the elimination of seropositive dogs, insecticide treatment and systematic treatment of human cases. Therefore, the accurate diagnosis is important in order to avoid the disease transmission or unnecessary culling of dogs. Serological tests are used for screening of dogs. However, these techniques present limitations. The Polymerase Chain Reaction (PCR) is an attractive alternative to the diagnosis in this context; but non-invasive samplings have great importance because they are simpler, painless and less resisted by dog-owners. This study aimed at evaluating conjunctival swab (CS) for canine VL diagnosis. In this methodology a sterile cotton swab is used to sampling the dog conjunctiva in both eyes. Thirty asymptomatic seropositive dogs were used. The samples were analyzed by the kDNA PCR-hybridization procedure in which the PCR products are hybridized with cloned kDNA mini-circles labeled with {sup 32}P[]dCTP. In addition, blood (B) was collected from each animal. L. chagasi was identified in 90% of CS samples and 13,6% of B samples. The high sensitivity obtained with asymptomatic dogs, in which the diagnosis is more difficult due the low number of parasites in the samples, allow concluding that the conjunctival swab associated to the kDNA PCR-hybridization assay provides a valuable alternative tool for the direct diagnosis of canine leishmaniasis. (author)

Pregnancy does not affect HIV incidence test results obtained using the BED capture enzyme immunoassay or an antibody avidity assay.

Directory of Open Access Journals (Sweden)

Oliver Laeyendecker

2010-10-01

Full Text Available Accurate incidence estimates are needed for surveillance of the HIV epidemic. HIV surveillance occurs at maternal-child health clinics, but it is not known if pregnancy affects HIV incidence testing.We used the BED capture immunoassay (BED and an antibody avidity assay to test longitudinal samples from 51 HIV-infected Ugandan women infected with subtype A, C, D and intersubtype recombinant HIV who were enrolled in the HIVNET 012 trial (37 baseline samples collected near the time of delivery and 135 follow-up samples collected 3, 4 or 5 years later. Nineteen of 51 women were also pregnant at the time of one or more of the follow-up visits. The BED assay was performed according to the manufacturer's instructions. The avidity assay was performed using a Genetic Systems HIV-1/HIV-2 + O EIA using 0.1M diethylamine as the chaotropic agent.During the HIVNET 012 follow-up study, there was no difference in normalized optical density values (OD-n obtained with the BED assay or in the avidity test results (% when women were pregnant (n = 20 results compared to those obtained when women were not pregnant (n = 115; for BED: p = 0.9, generalized estimating equations model; for avidity: p = 0.7, Wilcoxon rank sum. In addition, BED and avidity results were almost exactly the same in longitudinal samples from the 18 women who were pregnant at only one study visit during the follow-up study (p = 0.6, paired t-test.These results from 51 Ugandan women suggest that any changes in the antibody response to HIV infection that occur during pregnancy are not sufficient to alter results obtained with the BED and avidity assays. Confirmation with larger studies and with other HIV subtypes is needed.

The use of Taka-diastase in a [3H]poly(A) hybridization assay of oligo(U) sequences in RNA

International Nuclear Information System (INIS)

De Herdt, E.; Kondo, M.; Slegers, H.

1981-01-01

A reliable assay for uridylate sequences longer than 10 is described. The procedure is based on the hybridization of [ 3 H]poly(A) with poly(U) or oligo(U) sequences in high ionic conditions and a subsequent degradation of single stranded polynucleotides with purified Taka-diastase. A 1:2 complex between poly(A) and poly(U) is formed on which one poly(U) strand is digested by Taka-diastase. The procedure is especially suitable for the detection and quantitation of Usub(n) (n > 10) in RNA preparations. (Auth.)

Carbon nanotube-polyamidoamine dendrimer hybrid-modified electrodes for highly sensitive electrochemical detection of microRNA24.

Science.gov (United States)

Li, Fengye; Peng, Jing; Zheng, Qiong; Guo, Xiang; Tang, Hao; Yao, Shouzhuo

2015-01-01

A simple and ultrasensitive microRNA (miRNA) electrochemical biosensor employing multiwalled carbon nanotube (MWCNT)-polyamidoamine (PAMAM) dendrimer and methylene blue (MB) redox indicator is reported in this work. The assay utilizes a glass carbon (GC) electrode modified with MWCNT-PAMAM, on which the oligonucleotide capture probes are immobilized. The electrochemical detection of miRNAs is completed by measuring the reduction signal change of MB before and after the probe hybridization with target miRNA (miRNA24 is used as a model case). The MWCNT-PAMAM/GC electrode shows greatly enhanced signal to MB reduction in contrast to bare GC electrode. The functionalization of MWCNT with PAMAM maintains the electrochemical property of MWCNT to MB reduction but minimizes the undesired adsorption of MB on the MWCNT surface. The effect of experimental variables on the miRNA detection is investigated and optimized. A detection limit of 0.5 fM and a linear peak current density-concentration relationship up to 100 nM are obtained following 60 min hybridization. The proposed assay is successfully used to detect miRNA24 in total RNA sample extracted from HeLa cells.

Cloth-based hybridization array system for expanded identification of the animal species origin of derived materials in feeds.

Science.gov (United States)

Murphy, Johanna; Armour, Jennifer; Blais, Burton W

2007-12-01

A cloth-based hybridization array system (CHAS) previously developed for the detection of animal species for which prohibited materials have been specified (cattle, sheep, goat, elk, and deer) has been expanded to include the detection of animal species for which there are no prohibitions (pig and horse) in Canadian and American animal feeds. Animal species were identified by amplification of mitochondrial DNA sequences by PCR and subsequent hybridization of the amplicons with an array of species-specific oligonucleotide capture probes immobilized on a polyester cloth support, followed by an immunoenzymatic assay of the bound PCR products. The CHAS permitted sensitive and specific detection of meat meals from different animal species blended in a grain-based feed and should provide a useful adjunct to microscopic examination for the identification of prohibited materials in animal feeds.

CO2 capture by Li-functionalized silicene

KAUST Repository

Zhu, Jiajie; Chroneos, Alexander; Schwingenschlö gl, Udo

2016-01-01

CO2 capture and storage technology is of key importance to reduce the greenhouse effect. By its large surface area and sp3 hybridization, Li-functionalized silicene is demonstrated to be a promising CO2 absorbent that is stable up to at least 500 K

Evaluación de las técnicas de detección del VPH en los programas de cribado para cáncer de cuello uterino Assessment of hpv detection assays for use in cervical cancer screening programs

Directory of Open Access Journals (Sweden)

M Paz Cañadas

2006-10-01

Full Text Available OBJETIVO: La identificación de la infección por tipos de alto riesgo del virus del papiloma humano (VPH es una herramienta útil para el cribado de cáncer del cuello uterino. Las distintas técnicas aplicadas para su detección deben contrastarse y validarse para su empleo en la tamización poblacional. MATERIAL Y MÉTODOS: Se evalúan tres técnicas para la detección del VPH en 166 muestras cervicales procedentes de mujeres atendidas en una clínica de dermatología en Oviedo (España: a PCR-EIA mediante consensos MY09/MY011; b PCR con line blot hybridization (PCR-LBH con consensos PGMY; y c hybrid capture 2. RESULTADOS: El ADN-VPH se reconoció en 29.5%, 25.3% y 24.7%, de acuerdo con el ensayo. La concordancia global entre PCR-EIA, PCR-LBH y HC2 fue de 73.5% con los valores de kappa superiores a 0.56 entre los ensayos (pOBJECTIVE: Detection of high-risk human papillomavirus types (HPV infection is an important tool in the screening of cervical cancer and triage of cytological abnormalities. The different techniques for detection of this cancer need to be contrasted and validated for use in population screening. MATERIAL AND METHODS: Cervical cell samples were collected from 166 women attending a dermatology clinic in Oviedo (Spain. We evaluated the performance of three different assays for VPH detection. The methods utilized were 1 In-house PCR-EIA using L1 consensus primers MY09/MY11, 2 A PCR-reverse line blot hybridization (PCR-LBH that uses L1 consensus PGMY primers. 3 Hybrid Capture 2. All assays were performed blinded. The kappa statistic was used to test for global agreement between assay pairs. RESULTS: HPV DNA was detected in 24,7%, 25,3% and 29,5% of the women, respective to the assay. The overall agreement between the in-house PCR, PCR-LBH and HC2 was (73.5% with all kappa values between assay pairs exceeding 0.56 (p<0.001. CONCLUSION: The three HPV assays were equally accurate in estimating high-risk HPV prevalence and HPV

Comparison of IgM Capture Enzyme- Linked Immunosorbent Assay (ELISA) using Inhouse method and commercially available MRL kit for serological confirmation of dengue infection

International Nuclear Information System (INIS)

Hapugoda, D.M.; De Silva R, Nilanthi; Abeywickreme, W.; Gunasena, Sunethra; Prithimala, L.D.; Jayawardene, S.L.G.J.; Kumari, Thamara

2003-01-01

Laboratory diagnosis of dengue infection is important for the management of the patients. In this study igM capture ELISA using an inhouse method and commercially available kit (MRL diagnostics,USA) was compared to detect diagnostic capability of Inhouse IgM ELISA for provision of diagnostic facilities to the public at an affordable cost. Eighty acute and convalescent serum samples were collected from serologically confirmed dengue patients. Serological confirmation of patients were performed by Haemagglutination Inhibition (HI) assay, gold standard assay for dengue on paired serum samples. All collected acute and convalescent sera were tested by IgM ELISA using the inhouse method and MRL kit. Antigen and conjugate for the inhouse IgM method were prepared in the laboratory. A cocktail of four dengue antigens containing 25 Antigen ELISA units of each type was prepared and used as the assay antigen. Conjugate was prepared using a serum sample with high dengue Anti flavi IgG antibody titre conjugated with Horseradish peroxidase. A prospective study of both IgM ELISA assays were performed using 113 acute sera collected from dengue suspected cases. Overall results showed that 46% and 52% acute sera collected from dengue confirmed patients were positive by inhouse ELISA assay and MRL kits respectively. In the prospective study done using acute sera collected from dengue suspected patients showed that 44% and 52% were positive by inhouse ELISA assay and MRL kits. There was no significant difference in positivity between these two assays. (P=0.18). Inhouse IgM ELISA can be used for provision of laboratory diagnosis of dengue virus infection more than 5 days. The assay is 10 times less costly than using MRL kits as assay antigen and conjugate can be prepared easily in the laboratory

Microbead agglutination based assays

KAUST Repository

Kodzius, Rimantas

2013-01-21

We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling the macroscopic observation. Such tests are most often used to explore antibody-antigen reactions. Agglutination has been used for protein assays using a biotin/streptavidin system as well as a hybridization based assay. The agglutination systems are prone to selftermination of the linking analyte, prone to active site saturation and loss of agglomeration at high analyte concentrations. We investigated the molecular target/ligand interaction, explaining the common agglutination problems related to analyte self-termination, linkage of the analyte to the same bead instead of different microbeads. We classified the agglutination process into three kinds of assays: a two- component assay, a three-component assay and a stepped three- component assay. Although we compared these three kinds of assays for recognizing DNA and protein molecules, the assay can be used for virtually any molecule, including ions and metabolites. In total, the optimized assay permits detecting analytes with high sensitivity in a short time, 5 min, at room temperature. Such a system is appropriate for POC testing.

Electrical potential-assisted DNA hybridization. How to mitigate electrostatics for surface DNA hybridization.

Science.gov (United States)

Tymoczko, Jakub; Schuhmann, Wolfgang; Gebala, Magdalena

2014-12-24

Surface-confined DNA hybridization reactions are sensitive to the number and identity of DNA capture probes and experimental conditions such as the nature and the ionic strength of the electrolyte solution. When the surface probe density is high or the concentration of bulk ions is much lower than the concentration of ions within the DNA layer, hybridization is significantly slowed down or does not proceed at all. However, high-density DNA monolayers are attractive for designing high-sensitivity DNA sensors. Thus, circumventing sluggish DNA hybridization on such interfaces allows a high surface concentration of target DNA and improved signal/noise ratio. We present potential-assisted hybridization as a strategy in which an external voltage is applied to the ssDNA-modified interface during the hybridization process. Results show that a significant enhancement of hybridization can be achieved using this approach.

Hybrid Membrane/Absorption Process for Post-combustion CO2 Capture

Energy Technology Data Exchange (ETDEWEB)

Li, Shiguang; Shou, S.; Pyrzynski, Travis; Makkuni, Ajay; Meyer, Howard

2013-12-31

This report summarizes scientific/technical progress made for bench-scale membrane contactor technology for post-combustion CO2 capture from DOE Contract No. DE-FE-0004787. Budget Period 1 (BP1) membrane absorber, Budget Period 2 (BP2) membrane desorber and Budget Period 3 (BP3) integrated system and field testing studies have been completed successfully and met or exceeded the technical targets (≥ 90% CO2 removal and CO2 purity of 97% in one membrane stage). Significant breakthroughs are summarized below: BP1 research: The feasibility of utilizing the poly (ether ether ketone), PEEK, based hollow fiber contractor (HFC) in combination with chemical solvents to separate and capture at least 90% of the CO2 from simulated flue gases has been successfully established. Excellent progress has been made as we have achieved the BP1 goal: ≥ 1,000 membrane intrinsic CO2 permeance, ≥ 90% CO2 removal in one stage, ≤ 2 psi gas side pressure drop, and ≥ 1 (sec)-1 mass transfer coefficient. Initial test results also show that the CO2 capture performance, using activated Methyl Diethanol Amine (aMDEA) solvent, was not affected by flue gas contaminants O2 (~3%), NO2 (66 ppmv), and SO2 (145 ppmv). BP2 research: The feasibility of utilizing the PEEK HFC for CO2-loaded solvent regeneration has been successfully established High CO2 stripping flux, one order of magnitude higher than CO2 absorption flux, have been achieved. Refined economic evaluation based on BP1 membrane absorber and BP2 membrane desorber laboratory test data indicate that the CO2 capture costs are 36% lower than DOE’s benchmark amine absorption technology. BP3 research: A bench-scale system utilizing a membrane absorber and desorber was integrated into a continuous CO2 capture process using contactors containing 10 to 20 ft2 of membrane area. The integrated process operation was stable through a 100-hour laboratory test, utilizing a simulated flue gas stream. Greater than 90% CO2 capture combined with 97

Spectroscopic Investigation of the Canopy Configurations in Nanoparticle Organic Hybrid Materials of Various Grafting Densities during CO 2 Capture

KAUST Repository

Petit, Camille; Park, Youngjune; Lin, Kun-Yi Andrew; Park, Ah-Hyung Alissa

2012-01-01

Novel liquid-like nanoparticle organic hybrid materials (NOHMs) made of polyetheramine chains tethered onto functionalized silica nanoparticles were synthesized and characterized before and after exposure to CO 2 using NMR, Raman, and ATR FT-IR spectroscopies in order to investigate the effect of the grafting densities on the NOHM canopy structure. Considering the promising tunable properties for CO 2 capture of NOHMs, this study was conducted to provide important structural information to better design NOHMs for CO 2 capture. In order to minimize the CO 2 absorption via enthalpic effect and provide a more accurate assessment of the structural effects, the NOHMs were synthesized without task-specific groups. A greater chain ordering and decreased intermolecular interactions were observed in NOHMs compared to the unbound polymer. This was attributed to the specific structural arrangement of the frustrated canopy. The distinct configuration of grafted polymer chains caused different CO 2 packing and CO 2-induced swelling behaviors between the NOHMs and the unbound polymer. The grafting density influenced the ordering and coupling of the polymer chains and CO 2-induced swelling. Its effect on the CO 2 packing behavior was less pronounced. © 2011 American Chemical Society.

Spectroscopic Investigation of the Canopy Configurations in Nanoparticle Organic Hybrid Materials of Various Grafting Densities during CO 2 Capture

KAUST Repository

Petit, Camille

2012-01-12

Novel liquid-like nanoparticle organic hybrid materials (NOHMs) made of polyetheramine chains tethered onto functionalized silica nanoparticles were synthesized and characterized before and after exposure to CO 2 using NMR, Raman, and ATR FT-IR spectroscopies in order to investigate the effect of the grafting densities on the NOHM canopy structure. Considering the promising tunable properties for CO 2 capture of NOHMs, this study was conducted to provide important structural information to better design NOHMs for CO 2 capture. In order to minimize the CO 2 absorption via enthalpic effect and provide a more accurate assessment of the structural effects, the NOHMs were synthesized without task-specific groups. A greater chain ordering and decreased intermolecular interactions were observed in NOHMs compared to the unbound polymer. This was attributed to the specific structural arrangement of the frustrated canopy. The distinct configuration of grafted polymer chains caused different CO 2 packing and CO 2-induced swelling behaviors between the NOHMs and the unbound polymer. The grafting density influenced the ordering and coupling of the polymer chains and CO 2-induced swelling. Its effect on the CO 2 packing behavior was less pronounced. © 2011 American Chemical Society.

Toronto hybrid taxi pilot

International Nuclear Information System (INIS)

Stevens, M.; Marans, B.

2009-10-01

This paper provided details of a hybrid taxi pilot program conducted to compare the on-road performance of Toyota Camry hybrid vehicles against conventional vehicles over a 1-year period in order to determine the business case and air emission reductions associated with the use of hybrid taxi cabs. Over 750,000 km worth of fuel consumption was captured from 10 Toyota Camry hybrids, a Toyota Prius, and 5 non-hybrid Camry vehicles over an 18-month period. The average real world fuel consumption for the taxis demonstrated that the Toyota Prius has the lowest cost of ownership, while the non-hybrid Camry has the highest cost of ownership. Carbon dioxide (CO 2 ) reductions associated with the 10 Camry hybrid taxis were calculated at 236 tonnes over a 7-year taxi service life. Results suggested that the conversion of Toronto's 5680 taxis would yield annual CO 2 emission reductions of over 19,000 tonnes. All hybrid purchasers identified themselves as highly likely to purchase a hybrid again. 5 tabs., 9 figs.

Toronto hybrid taxi pilot

Energy Technology Data Exchange (ETDEWEB)

Stevens, M. [CrossChasm Technologies, Cambridge, ON (Canada); Marans, B. [Toronto Atmospheric Fund, ON (Canada)

2009-10-15

This paper provided details of a hybrid taxi pilot program conducted to compare the on-road performance of Toyota Camry hybrid vehicles against conventional vehicles over a 1-year period in order to determine the business case and air emission reductions associated with the use of hybrid taxi cabs. Over 750,000 km worth of fuel consumption was captured from 10 Toyota Camry hybrids, a Toyota Prius, and 5 non-hybrid Camry vehicles over an 18-month period. The average real world fuel consumption for the taxis demonstrated that the Toyota Prius has the lowest cost of ownership, while the non-hybrid Camry has the highest cost of ownership. Carbon dioxide (CO{sub 2}) reductions associated with the 10 Camry hybrid taxis were calculated at 236 tonnes over a 7-year taxi service life. Results suggested that the conversion of Toronto's 5680 taxis would yield annual CO{sub 2} emission reductions of over 19,000 tonnes. All hybrid purchasers identified themselves as highly likely to purchase a hybrid again. 5 tabs., 9 figs.

Comparison of the clinical performances of the AdvanSure HPV Screening Real-Time PCR, the Abbott Real-Time High-Risk HPV Test, and the Hybrid Capture High-Risk HPV DNA Test for Cervical Cancer Screening.

Science.gov (United States)

Chung, Hae-Sun; Hahm, Chorong; Lee, Miae

2014-09-01

The clinical performance of three human papillomavirus (HPV) DNA commercial assays for cervical cancer screening was evaluated; the AdvanSure HPV Screening Real-Time PCR (AdvanSure PCR; LG Life Sciences) that was developed recently for the detection of both high-risk and low-risk genotypes, the Abbott RealTime High-Risk HPV Test (Abbott PCR; Abbott Molecular) and the Hybrid Capture High-Risk HPV DNA test (HC2; Qiagen). The three different HPV DNA tests were compared using cytology samples obtained from 619 women who underwent routine cervical cancer screening. The gold-standard assay was histopathological confirmation of cervical intraepithelial neoplasia of grade 2 or worse. The clinical sensitivities of the AdvanSure PCR, the Abbott PCR and the HC2 for the detection of cervical intraepithelial neoplasia of grade 2 or worse were 95.5%, 95.5% and 100%, respectively, while the clinical specificities were 61.6%, 86.4% and 83.3%, respectively. There were no significant differences in the clinical sensitivities of the Abbott PCR and the AdvanSure PCR compared to the HC2. The clinical specificities of the Abbott PCR and the AdvanSure PCR for the detection of HPV types 16/18 were 97.8% and 98.5%, respectively. For cervical cancer screening, all three tests showed relatively good clinical sensitivities, but the AdvanSure PCR had lower clinical specificity than the Abbott PCR and the HC2. The AdvanSure PCR and the Abbott PCR assays have the advantage of being automated and the ability to distinguish between HPV types 16/18 and other HPV types. The two real-time PCR assays could be useful tools in HPV testing for cervical cancer screening. Copyright © 2014 Elsevier B.V. All rights reserved.

Hybrid Calcium Phosphate-Polymeric Micelles Incorporating Gadolinium Chelates for Imaging-Guided Gadolinium Neutron Capture Tumor Therapy.

Science.gov (United States)

Mi, Peng; Dewi, Novriana; Yanagie, Hironobu; Kokuryo, Daisuke; Suzuki, Minoru; Sakurai, Yoshinori; Li, Yanmin; Aoki, Ichio; Ono, Koji; Takahashi, Hiroyuki; Cabral, Horacio; Nishiyama, Nobuhiro; Kataoka, Kazunori

2015-06-23

Gadolinium (Gd) chelates-loaded nanocarriers have high potential for achieving magnetic resonance imaging (MRI)-guided Gd neutron capture therapy (GdNCT) of tumors. Herein, we developed calcium phosphate micelles hybridized with PEG-polyanion block copolymers, and incorporated with the clinical MRI contrast agent Gd-diethylenetriaminepentaacetic acid (Gd-DTPA/CaP). The Gd-DTPA/CaP were nontoxic to cancer cells at the concentration of 100 μM based on Gd-DTPA, while over 50% of the cancer cells were killed by thermal neutron irradiation at this concentration. Moreover, the Gd-DTPA/CaP showed a dramatically increased accumulation of Gd-DTPA in tumors, leading to the selective contrast enhancement of tumor tissues for precise tumor location by MRI. The enhanced tumor-to-blood distribution ratio of Gd-DTPA/CaP resulted in the effective suppression of tumor growth without loss of body weight, indicating the potential of Gd-DTPA/CaP for safe cancer treatment.

Optimization of a magnetic capture RT-LAMP assay for fast and real-time detection of potato virus Y and differentiation of N and O serotypes.

Science.gov (United States)

Treder, Krzysztof; Chołuj, Joanna; Zacharzewska, Bogumiła; Babujee, Lavanya; Mielczarek, Mateusz; Burzyński, Adam; Rakotondrafara, Aurélie M

2018-02-01

Potato virus Y (PVY) infection has been a global challenge for potato production and the leading cause of downgrading and rejection of seed crops for certification. Accurate and timely diagnosis is a key for effective disease control. Here, we have optimized a reverse transcription loop-mediated amplification (RT-LAMP) assay to differentiate the PVY O and N serotypes. The RT-LAMP assay is based on isothermal autocyclic strand displacement during DNA synthesis. The high specificity of this method relies heavily on the primer sets designed for the amplification of the targeted regions. We designed specific primer sets targeting a region within the coat protein gene that contains nucleotide signatures typical for O and N coat protein types, and these primers differ in their annealing temperature. Combining this assay with total RNA extraction by magnetic capture, we have established a highly sensitive, simplified and shortened RT-LAMP procedure as an alternative to conventional nucleic acid assays for diagnosis. This optimized procedure for virus detection may be used as a preliminary test for identifying the viral serotype prior to investing time and effort in multiplex RT-PCR tests when a specific strain is needed.

Laser capture microdissection: Arcturus(XT) infrared capture and UV cutting methods.

Science.gov (United States)

Gallagher, Rosa I; Blakely, Steven R; Liotta, Lance A; Espina, Virginia

2012-01-01

Laser capture microdissection (LCM) is a technique that allows the precise procurement of enriched cell populations from a heterogeneous tissue under direct microscopic visualization. LCM can be used to harvest the cells of interest directly or can be used to isolate specific cells by ablating the unwanted cells, resulting in histologically enriched cell populations. The fundamental components of laser microdissection technology are (a) visualization of the cells of interest via microscopy, (b) transfer of laser energy to a thermolabile polymer with either the formation of a polymer-cell composite (capture method) or transfer of laser energy via an ultraviolet laser to photovolatize a region of tissue (cutting method), and (c) removal of cells of interest from the heterogeneous tissue section. Laser energy supplied by LCM instruments can be infrared (810 nm) or ultraviolet (355 nm). Infrared lasers melt thermolabile polymers for cell capture, whereas ultraviolet lasers ablate cells for either removal of unwanted cells or excision of a defined area of cells. LCM technology is applicable to an array of applications including mass spectrometry, DNA genotyping and loss-of-heterozygosity analysis, RNA transcript profiling, cDNA library generation, proteomics discovery, and signal kinase pathway profiling. This chapter describes the unique features of the Arcturus(XT) laser capture microdissection instrument, which incorporates both infrared capture and ultraviolet cutting technology in one instrument, using a proteomic downstream assay as a model.

Analytical applications of neutron capture gamma-rays

International Nuclear Information System (INIS)

Lindstrom, R.M.; Paul, R.L.; Anderson, D.L.; Paul, R.L.

1997-01-01

Field and industrial applications of neutron capture gamma-ray spectrometry with isotopic sources or neutron generators are economically important. Geochemical exploration in boreholes is done routinely with neutron probes. Coal and ores are assayed with analyzers adjacent to a conveyor belt in dozens of industrial facilities. The use of capture gamma rays for explosives detection has been described in the literature, both for scanning airline baggage and for characterizing obsolete munitions; a packaged system for the latter is available commercially. Generalizations are drawn from the history of the field, and predictions are made about the future usefulness of capture gamma rays. (author)

Effect of water on the physical properties and carbon dioxide capture capacities of liquid-like Nanoparticle Organic Hybrid Materials and their corresponding polymers

KAUST Repository

Petit, Camille; Bhatnagar, Sonali; Park, Ah-Hyung Alissa

2013-01-01

Binary systems composed of liquid-like Nanoparticle Organic Hybrid Materials (NOHMs) and the secondary fluid (i.e., water) were prepared, and their thermal stabilities, densities, viscosities, and CO2 absorption capacities were investigated. Recent work has suggested NOHMs as an alternative CO2 capture media with interesting chemical and physical tunability. Anhydrous CO2 capture solvents often degrade when they are exposed to water, while flue gas generally contains about 8-16% water. Thus, this study was conducted to investigate the effect of water on the NOHMs' properties relevant to CO2 capture as well as the chemical and thermal stabilities of H2O-loaded NOHMs. It was found that water acted as an antisolvent of NOHMs, and therefore, caused a decreased CO2 capture capacity. On the other hand, the results indicated that while water did not affect the NOHMs' thermal stability, it significantly helped lowering their density and viscosity. In order to investigate the effect of intermolecular interactions among two fluids on the density and viscosity, the excess volumes and viscosity deviations were calculated and correlated with Redlich-Kister equations. The trends revealed the existence of strong intermolecular interactions between water molecules and the poly(ethlyne glycol) component of NOHMs, which may have caused the drastic decrease in the NOHMs' viscosity with the addition of water. © 2013 Elsevier Inc.

Effect of water on the physical properties and carbon dioxide capture capacities of liquid-like Nanoparticle Organic Hybrid Materials and their corresponding polymers

KAUST Repository

Petit, Camille

2013-10-01

Binary systems composed of liquid-like Nanoparticle Organic Hybrid Materials (NOHMs) and the secondary fluid (i.e., water) were prepared, and their thermal stabilities, densities, viscosities, and CO2 absorption capacities were investigated. Recent work has suggested NOHMs as an alternative CO2 capture media with interesting chemical and physical tunability. Anhydrous CO2 capture solvents often degrade when they are exposed to water, while flue gas generally contains about 8-16% water. Thus, this study was conducted to investigate the effect of water on the NOHMs\\' properties relevant to CO2 capture as well as the chemical and thermal stabilities of H2O-loaded NOHMs. It was found that water acted as an antisolvent of NOHMs, and therefore, caused a decreased CO2 capture capacity. On the other hand, the results indicated that while water did not affect the NOHMs\\' thermal stability, it significantly helped lowering their density and viscosity. In order to investigate the effect of intermolecular interactions among two fluids on the density and viscosity, the excess volumes and viscosity deviations were calculated and correlated with Redlich-Kister equations. The trends revealed the existence of strong intermolecular interactions between water molecules and the poly(ethlyne glycol) component of NOHMs, which may have caused the drastic decrease in the NOHMs\\' viscosity with the addition of water. © 2013 Elsevier Inc.

Engineering cholesterol-based fibers for antibody immobilization and cell capture

Science.gov (United States)

Cohn, Celine

abilities of the CSS fibers were compared to that of hydrophobic polycaprolactone (PCL) fibers and hydrophilic plasma-treated PCL fibers. Electrospun CSS fibers were found to immobilize equivalent amounts of protein as hydrophobically immobilized proteins. However, these proteins captured 6 times more cells, indicative of retained protein function. The second key concept was the design and fabrication of a hybridized lipid fiber. Lipid fibers provide improved protein function but fabrication difficulties have limited their adoption. Thus, we sought to fabricate a lipid-polymer hybrid that is easily fabricated while maintaining protein function. The hybrid fiber consists of a PCL backbone with conjugated CSS. The hybrid lipid fibers showed improved protein function. In addition, higher lipid concentrations were directly correlated to higher cell capture efficiencies. The third key concept was on the development of dually functionalized lipid fibers and understanding the resulting cell capture efficiencies. Many platforms are unable to simultaneously search for heterogeneous populations of CTCs -- the ability to dually functionalize cell-capturing platforms would address this technological weakness. Studies indicated that dually functionalizing the lipid fibers did not compromise the platforms' abilities to capture the cells of interest. Such dually functionalized fibers allow for a single cell-capture platform to successfully detect heterogeneous populations of CTCs. The body of work encompassed herein describes the use of lipid fibers for antibody immobilization and cell capture. Data from various projects indicate that the use of cholesterol-based fibers produced from electrospun CSS are well suited for protein immobilization. The CSS fibers are able to immobilize equivalent amounts of protein as compared to other immobilization techniques. However, the benefit of these fibers is illustrated by the strong cell-capturing efficiencies, indicating that the immobilized

Hybridization Capture Using Short PCR Products Enriches Small Genomes by Capturing Flanking Sequences (CapFlank)

DEFF Research Database (Denmark)

Tsangaras, Kyriakos; Wales, Nathan; Sicheritz-Pontén, Thomas

2014-01-01

nucleotides) can result in enrichment across entire mitochondrial and bacterial genomes. Our findings suggest that some of the off-target sequences derived in capture experiments are non-randomly enriched, and that CapFlank will facilitate targeted enrichment of large contiguous sequences with minimal prior...

Combining hybrid cars and synthetic fuels with electricity generation and carbon capture and storage

International Nuclear Information System (INIS)

Vliet, Oscar van; Broek, Machteld van den; Turkenburg, Wim; Faaij, Andre

2011-01-01

We examined the co-evolution of the transportation, and electricity and heat generation sectors in the Netherlands until 2040 using a MARKAL bottom-up cost optimisation model. All scenario variants investigated indicate a switch away from crude oil-based diesel and petrol for transportation. Lowest overall CO 2 abatement cost is achieved by accommodating transportation first and using relatively expensive options for emissions reduction in electricity generation if needed. Biomass and carbon capture and storage (CCS) are used to full potential. Transportation CO 2 emissions are reduced by switching to ethanol or bio-based synthetic fuels combined with CCS, and series hybrid cars if needed. Depending on the availability of biomass and carbon storage capacity, electricity is produced from biomass, coal with CCS, or wind complemented with natural gas. Indirect greenhouse gas emissions rise to 34-54% of national emissions in 2040. The difference in annual investment required between the scenario variants with and without CO 2 emissions reductions of 68% by 2040 is 4-7 billion euro/year, or 0.5-1.2% of projected GDP. Investment costs are mostly determined by the cost of cars and electricity generation capacity. We observe competition for limited biomass supply and CO 2 storage capacity between the transportation and power sectors.

CO_2 capture by amine-functionalized nanoporous materials: A review

International Nuclear Information System (INIS)

Chen, Chao; Kim, Jun; Ahn, Wha-Seung

2014-01-01

Amine-functionalized nanoporous materials can be prepared by the incorporation of diverse organic amine moieties into the pore structures of a range of support materials, such as mesoporous silica and alumina, zeolite, carbon and metal organic frameworks (MOFs), either by direct functionalization or post-synthesis through physical impregnation or grafting. These hybrid materials have great potential for practical applications, such as dry adsorbents for postcombustion CO_2 capture, owing to their high CO_2 capture capacity, high capture selectivity towards CO_2 compared to other gases, and excellent stability. This paper summarizes the preparation methods and CO_2 capture performance based on the equilibrium CO_2 uptake of a range of amine-functionalized nanoporous materials

Hybrid male sterility is caused by mitochondrial DNA deletion.

Science.gov (United States)

Hayashida, Kenji; Kohno, Shigeru

2009-07-01

Although it is known that the hybrid male mouse is sterile just like any other animal's heterogametic sex, the reason why only the male germ cells are impaired has yet to be discovered. TdT-mediated dUTP nick end labeling assay using a confocal fluorescence microscope and DNA fragmentation assay of hybrid testis indicated destruction of the mitochondrial DNA (mtDNA) rather than the nuclear DNA. Previously we reported that maternal mtDNA inheritance is through selective sperm mtDNA elimination based on the sperm factor and two egg factors, and expression of these three factors was recognized in the hybrid testis. It was thereby assumed that mtDNA destruction caused by the expression of maternal mtDNA inheritance system in male germ cells is implicated in the hybrid male sterility of mice.

Sensitive electrochemical monitoring of nucleic acids coupling DNA nanostructures with hybridization chain reaction

International Nuclear Information System (INIS)

Zhuang, Junyang; Fu, Libing; Xu, Mingdi; Yang, Huanghao; Chen, Guonan; Tang, Dianping

2013-01-01

Graphical abstract: -- Highlights: •A new signal-on metallobioassay was developed for detection of nucleic acids. •Target-triggered long-range self-assembled DNA nanostructures are used for amplification of electronic signal. •Hybridization chain reaction is utilized for construction of long-range DNA nanostructures. -- Abstract: Methods based on metal nanotags have been developed for metallobioassay of nucleic acids, but most involve complicated labeling or stripping procedures and are unsuitable for routine use. Herein, we report the proof-of-concept of a novel and label-free metallobioassay for ultrasensitive electronic determination of human immunodeficiency virus (HIV)-related gene fragments at an ultralow concentration based on target-triggered long-range self-assembled DNA nanostructures and DNA-based hybridization chain reaction (HCR). The signal is amplified by silver nanotags on the DNA duplex. The assay mainly consists of capture probe, detection probe, and two different DNA hairpins. In the presence of target DNA, the capture probe immobilized on the sensor sandwiches target DNA with the 3′ end of detection probe. Another exposed part of detection probe at the 5′ end opens two alternating DNA hairpins in turn, and propagates a chain reaction of hybridization events to form a nicked double-helix. Finally, numerous silver nanotags are immobilized onto the long-range DNA nanostructures, each of which produces a strong electronic signal within the applied potentials. Under optimal conditions, the target-triggered long-range DNA nanostructures present good electrochemical behaviors for the detection of HIV DNA at a concentration as low as 0.5 fM. Importantly, the outstanding sensitivity can make this approach a promising scheme for development of next-generation DNA sensors without the need of enzyme labeling or fluorophore labeling

Genome-wide mapping in a house mouse hybrid zone reveals hybrid sterility loci and Dobzhansky-Muller interactions.

Science.gov (United States)

Turner, Leslie M; Harr, Bettina

2014-12-09

Mapping hybrid defects in contact zones between incipient species can identify genomic regions contributing to reproductive isolation and reveal genetic mechanisms of speciation. The house mouse features a rare combination of sophisticated genetic tools and natural hybrid zones between subspecies. Male hybrids often show reduced fertility, a common reproductive barrier between incipient species. Laboratory crosses have identified sterility loci, but each encompasses hundreds of genes. We map genetic determinants of testis weight and testis gene expression using offspring of mice captured in a hybrid zone between M. musculus musculus and M. m. domesticus. Many generations of admixture enables high-resolution mapping of loci contributing to these sterility-related phenotypes. We identify complex interactions among sterility loci, suggesting multiple, non-independent genetic incompatibilities contribute to barriers to gene flow in the hybrid zone.

FRET two-hybrid assay by linearly fitting FRET efficiency to concentration ratio between acceptor and donor

Science.gov (United States)

Du, Mengyan; Yang, Fangfang; Mai, Zihao; Qu, Wenfeng; Lin, Fangrui; Wei, Lichun; Chen, Tongsheng

2018-04-01

We here introduce a fluorescence resonance energy transfer (FRET) two-hybrid assay method to measure the maximal donor(D)- and acceptor(A)-centric FRET efficiency (ED,max and EA,max) of the D-A complex and its stoichiometry by linearly fitting the donor-centric FRET efficiency (ED) to the acceptor-to-donor concentration ratio (RC) and acceptor-centric FRET efficiency (EA) to 1/RC, respectively. We performed this method on a wide-field fluorescence microscope for living HepG2 cells co-expressing FRET tandem constructs and free donor/acceptor and obtained correct ED, EA, and stoichiometry values of those tandem constructs. Evaluation on the binding of Bad with Bcl-XL in Hela cells showed that Bad interacted strongly with Bcl-XL to form a Bad-Bcl-XL complex on mitochondria, and one Bad interacted mainly with one Bcl-XL molecule in healthy cells, while with multiple (maybe 2) Bcl-XL molecules in apoptotic cells.

11th GCC Closed Forum: cumulative stability; matrix stability; immunogenicity assays; laboratory manuals; biosimilars; chiral methods; hybrid LBA/LCMS assays; fit-for-purpose validation; China Food and Drug Administration bioanalytical method validation.

Science.gov (United States)

Islam, Rafiq; Briscoe, Chad; Bower, Joseph; Cape, Stephanie; Arnold, Mark; Hayes, Roger; Warren, Mark; Karnik, Shane; Stouffer, Bruce; Xiao, Yi Qun; van der Strate, Barry; Sikkema, Daniel; Fang, Xinping; Tudoroniu, Ariana; Tayyem, Rabab; Brant, Ashley; Spriggs, Franklin; Barry, Colin; Khan, Masood; Keyhani, Anahita; Zimmer, Jennifer; Caturla, Maria Cruz; Couerbe, Philippe; Khadang, Ardeshir; Bourdage, James; Datin, Jim; Zemo, Jennifer; Hughes, Nicola; Fatmi, Saadya; Sheldon, Curtis; Fountain, Scott; Satterwhite, Christina; Colletti, Kelly; Vija, Jenifer; Yu, Mathilde; Stamatopoulos, John; Lin, Jenny; Wilfahrt, Jim; Dinan, Andrew; Ohorodnik, Susan; Hulse, James; Patel, Vimal; Garofolo, Wei; Savoie, Natasha; Brown, Michael; Papac, Damon; Buonarati, Mike; Hristopoulos, George; Beaver, Chris; Boudreau, Nadine; Williard, Clark; Liu, Yansheng; Ray, Gene; Warrino, Dominic; Xu, Allan; Green, Rachel; Hayward-Sewell, Joanne; Marcelletti, John; Sanchez, Christina; Kennedy, Michael; Charles, Jessica St; Bouhajib, Mohammed; Nehls, Corey; Tabler, Edward; Tu, Jing; Joyce, Philip; Iordachescu, Adriana; DuBey, Ira; Lindsay, John; Yamashita, Jim; Wells, Edward

2018-04-01

The 11th Global CRO Council Closed Forum was held in Universal City, CA, USA on 3 April 2017. Representatives from international CRO members offering bioanalytical services were in attendance in order to discuss scientific and regulatory issues specific to bioanalysis. The second CRO-Pharma Scientific Interchange Meeting was held on 7 April 2017, which included Pharma representatives' sharing perspectives on the topics discussed earlier in the week with the CRO members. The issues discussed at the meetings included cumulative stability evaluations, matrix stability evaluations, the 2016 US FDA Immunogenicity Guidance and recent and unexpected FDA Form 483s on immunogenicity assays, the bioanalytical laboratory's role in writing PK sample collection instructions, biosimilars, CRO perspectives on the use of chiral versus achiral methods, hybrid LBA/LCMS assays, applications of fit-for-purpose validation and, at the Global CRO Council Closed Forum only, the status and trend of current regulated bioanalytical practice in China under CFDA's new BMV policy. Conclusions from discussions of these topics at both meetings are included in this report.

Toward a solid-phase nucleic acid hybridization assay within microfluidic channels using immobilized quantum dots as donors in fluorescence resonance energy transfer.

Science.gov (United States)

Chen, Lu; Algar, W Russ; Tavares, Anthony J; Krull, Ulrich J

2011-01-01

The optical properties and surface area of quantum dots (QDs) have made them an attractive platform for the development of nucleic acid biosensors based on fluorescence resonance energy transfer (FRET). Solid-phase assays based on FRET using mixtures of immobilized QD-oligonucleotide conjugates (QD biosensors) have been developed. The typical challenges associated with solid-phase detection strategies include non-specific adsorption, slow kinetics of hybridization, and sample manipulation. The new work herein has considered the immobilization of QD biosensors onto the surfaces of microfluidic channels in order to address these challenges. Microfluidic flow can be used to dynamically control stringency by adjustment of the potential in an electrokinetic-based microfluidics environment. The shearing force, Joule heating, and the competition between electroosmotic and electrophoretic mobilities allow the optimization of hybridization conditions, convective delivery of target to the channel surface to speed hybridization, amelioration of adsorption, and regeneration of the sensing surface. Microfluidic flow can also be used to deliver (for immobilization) and remove QD biosensors. QDs that were conjugated with two different oligonucleotide sequences were used to demonstrate feasibility. One oligonucleotide sequence on the QD was available as a linker for immobilization via hybridization with complementary oligonucleotides located on a glass surface within a microfluidic channel. A second oligonucleotide sequence on the QD served as a probe to transduce hybridization with target nucleic acid in a sample solution. A Cy3 label on the target was excited by FRET using green-emitting CdSe/ZnS QD donors and provided an analytical signal to explore this detection strategy. The immobilized QDs could be removed under denaturing conditions by disrupting the duplex that was used as the surface linker and thus allowed a new layer of QD biosensors to be re-coated within the channel

New Class of Hybrid Materials for Detection, Capture, and "On-Demand" Release of Carbon Monoxide.

Science.gov (United States)

Pitto-Barry, Anaïs; Lupan, Alexandru; Ellingford, Christopher; Attia, Amr A A; Barry, Nicolas P E

2018-04-25

Carbon monoxide (CO) is both a substance hazardous to health and a side product of a number of industrial processes, such as methanol steam reforming and large-scale oxidation reactions. The separation of CO from nitrogen (N 2 ) in industrial processes is considered to be difficult because of the similarities of their electronic structures, sizes, and physicochemical properties (e.g., boiling points). Carbon monoxide is also a major poison in fuel cells because of its adsorption onto the active sites of the catalysts. It is therefore of the utmost economic importance to discover new materials that enable effective CO capture and release under mild conditions. However, methods to specifically absorb and easily release CO in the presence of contaminants, such as water, nitrogen, carbon dioxide, and oxygen, at ambient temperature are not available. Here, we report the simple and versatile fabrication of a new class of hybrid materials that allows capture and release of carbon monoxide under mild conditions. We found that carborane-containing metal complexes encapsulated in networks made of poly(dimethylsiloxane) react with CO, even when immersed in water, leading to dramatic color and infrared signature changes. Furthermore, we found that the CO can be easily released from the materials by simply dipping the networks into an organic solvent for less than 1 min, at ambient temperature and pressure, which not only offers a straightforward recycling method, but also a new method for the "on-demand" release of carbon monoxide. We illustrated the utilization of the on-demand release of CO from the networks by carrying out a carbonylation reaction on an electron-deficient metal complex that led to the formation of the CO-adduct, with concomitant recycling of the gel. We anticipate that our sponge-like materials and scalable methodology will open up new avenues for the storage, transport, and controlled release of CO, the silent killer and a major industrial poison.

The Baseline Positron Production and Capture Scheme for CLIC

CERN Document Server

Dadoun, Olivier; Lepercq, Pierre; Poirier, Freddy; Variola, Alessandro; Chehab, Robert; Rinolfi, Louis; Vivoli, Alessandro; Strakhovenko, Vladimir; Xu, Chengai

2010-01-01

The CLIC study considers the hybrid source using channeling as the baseline for unpolarised positron production. The hybrid source uses a few GeV electron beam impinging on a tungsten crystal target. With the crystal oriented on its axis it results an intense relatively low energy photon beam. The later is then impinging on an amorphous tungsten target producing positrons by e+e− pair creation. Downstream the amorphous target, a capture section based on an adiabatic matching device followed by a 2 GHz Pre- Injector Linac focuses and accelerates the positron beam up to around 200 MeV

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

Science.gov (United States)

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

2008-11-04

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

Hybrid strategies in nanolithography

Energy Technology Data Exchange (ETDEWEB)

Saavedra, Hector M; Mullen, Thomas J; Zhang Pengpeng; Dewey, Daniel C; Claridge, Shelley A; Weiss, Paul S [Department of Chemistry, The Pennsylvania State University, University Park, PA 16802 (United States)], E-mail: psw@cnsi.ucla.edu

2010-03-15

Hybrid nanoscale patterning strategies combine the registration and addressability of conventional lithographic techniques with the chemical and physical functionality enabled by intermolecular, electrostatic and/or biological interactions. This review aims to highlight and to provide a comprehensive description of recent developments in hybrid nanoscale patterning strategies that enhance existing lithographic techniques or can be used to fabricate functional chemical patterns that interact with their environment. These functional structures create new capabilities, such as the fabrication of physicochemical surfaces that can recognize and capture analytes from complex liquid or gaseous mixtures. The nanolithographic techniques we describe can be classified into three general areas: traditional lithography, soft lithography and scanning-probe lithography. The strengths and limitations of each hybrid patterning technique will be discussed, along with the current and potential applications of the resulting patterned, functional surfaces.

Novel concepts for CO2 capture

International Nuclear Information System (INIS)

Dijkstra, J.W.; Jansen, D.

2004-01-01

This paper describes the possibilities for power generation with CO 2 capture using envisaged key technologies: gas turbines, membranes and solid oxide fuel cells (SOFCs). First, the underlying programs in the Netherlands and at ECN are introduced. Then the key technologies are introduced, and concepts using these technologies are discussed. A literature overview of systems for power generation with fuel cells in combination with CO 2 capture is presented. Then a novel concept is introduced. This concept uses a water gas shift membrane reactor to convert the CO and H 2 in the SOFC anode off-gas to gain a CO 2 rich stream, which can be used for sequestration without elaborate treatment. Several implementation schemes of the technique are discussed such as atmospheric systems and hybrid SOFC-GT systems

Developments in capture-γ libraries for nonproliferation applications

Science.gov (United States)

Hurst, A. M.; Firestone, R. B.; Sleaford, B. W.; Bleuel, D. L.; Basunia, M. S.; Bečvář, F.; Belgya, T.; Bernstein, L. A.; Carroll, J. J.; Detwiler, B.; Escher, J. E.; Genreith, C.; Goldblum, B. L.; Krtička, M.; Lerch, A. G.; Matters, D. A.; McClory, J. W.; McHale, S. R.; Révay, Zs.; Szentmiklosi, L.; Turkoglu, D.; Ureche, A.; Vujic, J.

2017-09-01

The neutron-capture reaction is fundamental for identifying and analyzing the γ-ray spectrum from an unknown assembly because it provides unambiguous information on the neutron-absorbing isotopes. Nondestructive-assay applications may exploit this phenomenon passively, for example, in the presence of spontaneous-fission neutrons, or actively where an external neutron source is used as a probe. There are known gaps in the Evaluated Nuclear Data File libraries corresponding to neutron-capture γ-ray data that otherwise limit transport-modeling applications. In this work, we describe how new thermal neutron-capture data are being used to improve information in the neutron-data libraries for isotopes relevant to nonproliferation applications. We address this problem by providing new experimentally-deduced partial and total neutron-capture reaction cross sections and then evaluate these data by comparison with statistical-model calculations.

Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay.

Science.gov (United States)

Black, W C; Gorrochotegui-Escalante, N; Duteau, N M

2006-03-01

Most single nucleotide polymorphism (SNP) detection requires expensive equipment and reagents. The oligonucleotide ligation assay (OLA) is an inexpensive SNP assay that detects ligation between a biotinylated "allele-specific detector" and a 3' fluorescein-labeled "reporter" oligonucleotide. No ligation occurs unless the 3' detector nucleotide is complementary to the SNP nucleotide. The original OLA used chemical denaturation and neutralization. Heated OLA (HOLA) instead uses a thermal stable ligase and cycles of denaturing and hybridization for ligation and SNP detection. The cost per genotype is approximately US$1.25 with two-allele SNPs or approximately US$1.75 with three-allele SNPs. We illustrate the development of HOLA for SNP detection in the Early Trypsin and Abundant Trypsin loci in the mosquito Aedes aegypti (L.) and at the a-glycerophosphate dehydrogenase locus in the mosquito Anopheles gambiae s.s.

Some options for the muon collider capture and decay solenoids

International Nuclear Information System (INIS)

Green, M.A.

1995-11-01

This report discusses some of the problems associated with using solenoid magnets to capture the secondary particles that are created when an intense beam of 8 to 10 GeV protons interacts with the target at the center of the capture region. Hybrid capture solenoids with inductions of 28 T and a 22T are described. The first 14 to 15 T of the solenoid induction will be generated by a superconducting magnet. The remainder of the field will be generated by a Bitter type of water cooled solenoid. The capture solenoids include a transition section from the high field solenoid to a 7 T decay channel where pions and kaons that come off of the target decay into muons. A short 7 T solenoidal decay channel between the capture solenoid system and the phase rotation system is described. A concept for separation of negative and positive pions and kaons is briefly discussed

Simultaneous Detection of CDC Category "A" DNA and RNA Bioterrorism Agents by Use of Multiplex PCR & RT-PCR Enzyme Hybridization Assays

Directory of Open Access Journals (Sweden)

Kelly J. Henrickson

2009-10-01

Full Text Available Assays to simultaneously detect multiple potential agents of bioterrorism are limited. Two multiplex PCR and RT-PCR enzyme hybridization assays (mPCR-EHA, mRT-PCR-EHA were developed to simultaneously detect many of the CDC category “A” bioterrorism agents. The “Bio T” DNA assay was developed to detect: Variola major (VM, Bacillus anthracis (BA, Yersinia pestis (YP, Francisella tularensis (FT and Varicella zoster virus (VZV. The “Bio T” RNA assay (mRT-PCR-EHA was developed to detect: Ebola virus (Ebola, Lassa fever virus (Lassa, Rift Valley fever (RVF, Hantavirus Sin Nombre species (HSN and dengue virus (serotypes 1-4. Sensitivity and specificity of the 2 assays were tested by using genomic DNA, recombinant plasmid positive controls, RNA transcripts controls, surrogate (spiked clinical samples and common respiratory pathogens. The analytical sensitivity (limit of detection (LOD of the DNA asssay for genomic DNA was 1×100~1×102 copies/mL for BA, FT and YP. The LOD for VZV whole organism was 1×10-2 TCID50/mL. The LOD for recombinant controls ranged from 1×102~1×103copies/mL for BA, FT, YP and VM. The RNA assay demonstrated LOD for RNA transcript controls of 1×104~1×106 copies/mL without extraction and 1×105~1×106 copies/mL with extraction for Ebola, RVF, Lassa and HSN. The LOD for dengue whole organisms was ~1×10-4 dilution for dengue 1 and 2, 1×104 LD50/mL and 1×102 LD50/mL for dengue 3 and 4. The LOD without extraction for recombinant plasmid DNA controls was ~1×103 copies/mL (1.5 input copies/reaction for Ebola, RVF, Lassa and HSN. No cross-reactivity of primers and probes used in both assays was detected with common respiratory pathogens or between targeted analytes. Clinical sensitivity was estimated using 264 surrogate clinical samples tested with the BioT DNA assay and 549 samples tested with the BioT RNA assay. The clinical specificity is 99.6% and 99.8% for BioT DNA assay and BioT RNA assay, respectively. The

Development and evaluation of a Sarcocystis neurona-specific IgM capture enzyme-linked immunosorbent assay.

Science.gov (United States)

Murphy, J E; Marsh, A E; Reed, S M; Meadows, C; Bolten, K; Saville, W J A

2006-01-01

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease of horses caused primarily by the protozoal parasite Sarcocystis neurona. Currently available antemortem diagnostic testing has low specificity. The hypothesis of this study was that serum and cerebrospinal fluid (CSF) of horses experimentally challenged with S neurona would have an increased S neurona-specific IgM (Sn-IgM) concentration after infection, as determined by an IgM capture enzyme linked immunoassay (ELISA). The ELISA was based on the S neurona low molecular weight protein SNUCD-1 antigen and the monoclonal antibody 2G5 labeled with horseradish peroxidase. The test was evaluated using serum and CSF from 12 horses experimentally infected with 1.5 million S neurona sporocysts and 16 horses experimentally infected with varying doses (100 to 100,000) of S neurona sporocysts, for which results of histopathologic examination of the central nervous system were available. For horses challenged with 1.5 million sporocysts, there was a significant increase in serum Sn-IgM concentrations compared with values before infection at weeks 2-6 after inoculation (P neurona, there were significant increases in serum Sn-IgM concentration at various points in time after inoculation, depending on the challenge dose (P < .01). In addition, there was a significant increase between the CSF Sn-IgM concentrations before and after inoculation (P < .0001). These results support further evaluation of the assay as a diagnostic test during the acute phase of EPM.

Evaluation of the Clinical Performance of the HPV-Risk Assay Using the VALGENT-3 Panel.

Science.gov (United States)

Polman, N J; Oštrbenk, A; Xu, L; Snijders, P J F; Meijer, C J L M; Poljak, M; Heideman, D A M; Arbyn, M

2017-12-01

Human papillomavirus (HPV) testing is increasingly being incorporated into cervical cancer screening. The Validation of HPV Genotyping Tests (VALGENT) is a framework designed to evaluate the clinical performance of various HPV tests relative to that of the validated and accepted comparator test in a formalized and uniform manner. The aim of this study was to evaluate the clinical performance of the HPV-Risk assay with samples from the VALGENT-3 panel and to compare its performance to that of the clinically validated Hybrid Capture 2 assay (HC2). The VALGENT-3 panel comprises 1,300 consecutive samples from women participating in routine cervical cancer screening and is enriched with 300 samples from women with abnormal cytology. DNA was extracted from original ThinPrep PreservCyt medium aliquots, and HPV testing was performed using the HPV-Risk assay by investigators blind to the clinical data. HPV prevalence was analyzed, and the clinical performance of the HPV-Risk assay for the detection of cervical intraepithelial neoplasia grade 3 or worse (CIN3+) and CIN2 or worse (CIN2+) relative to the performance of HC2 was assessed. The sensitivity of the HPV-Risk assay for the detection of CIN3+ was similar to that of HC2 (relative sensitivity, 1.00; 95% confidence interval [CI], 0.95 to 1.05; P = 1.000), but the specificity of the HPV-Risk assay was significantly higher than that of HC2 (relative specificity, 1.02; 95% CI, 1.01 to 1.04; P performance of the HPV-Risk assay for the detection of CIN3+ and CIN2+ was noninferior to that of HC2, with all P values being ≤0.006. In conclusion, the HPV-Risk assay demonstrated noninferiority to the clinically validated HC2 by the use of samples from the VALGENT-3 panel for test validation and comparison. Copyright © 2017 Polman et al.

Mixed DNA/Oligo(ethylene glycol) Functionalized Gold Surface Improve DNA Hybridization in Complex Media

International Nuclear Information System (INIS)

Lee, C.; Gamble, L.; Grainger, D.; Castner, D.

2006-01-01

Reliable, direct 'sample-to-answer' capture of nucleic acid targets from complex media would greatly improve existing capabilities of DNA microarrays and biosensors. This goal has proven elusive for many current nucleic acid detection technologies attempting to produce assay results directly from complex real-world samples, including food, tissue, and environmental materials. In this study, we have investigated mixed self-assembled thiolated single-strand DNA (ssDNA) monolayers containing a short thiolated oligo(ethylene glycol) (OEG) surface diluent on gold surfaces to improve the specific capture of DNA targets from complex media. Both surface composition and orientation of these mixed DNA monolayers were characterized with x-ray photoelectron spectroscopy (XPS) and near-edge x-ray absorption fine structure (NEXAFS). XPS results from sequentially adsorbed ssDNA/OEG monolayers on gold indicate that thiolated OEG diluent molecules first incorporate into the thiolated ssDNA monolayer and, upon longer OEG exposures, competitively displace adsorbed ssDNA molecules from the gold surface. NEXAFS polarization dependence results (followed by monitoring the N 1s→π* transition) indicate that adsorbed thiolated ssDNA nucleotide base-ring structures in the mixed ssDNA monolayers are oriented more parallel to the gold surface compared to DNA bases in pure ssDNA monolayers. This supports ssDNA oligomer reorientation towards a more upright position upon OEG mixed adlayer incorporation. DNA target hybridization on mixed ssDNA probe/OEG monolayers was monitored by surface plasmon resonance (SPR). Improvements in specific target capture for these ssDNA probe surfaces due to incorporation of the OEG diluent were demonstrated using two model biosensing assays, DNA target capture from complete bovine serum and from salmon genomic DNA mixtures. SPR results demonstrate that OEG incorporation into the ssDNA adlayer improves surface resistance to both nonspecific DNA and protein

Hybrid chemical and nondestructive-analysis technique

International Nuclear Information System (INIS)

Hsue, S.T.; Marsh, S.F.; Marks, T.

1982-01-01

A hybrid chemical/NDA technique has been applied at the Los Alamos National Laboratory to the assay of plutonium in ion-exchange effluents. Typical effluent solutions contain low concentrations of plutonium and high concentrations of americium. A simple trioctylphosphine oxide (TOPO) separation can remove 99.9% of the americium. The organic phase that contains the separated plutonium can be accurately assayed by monitoring the uranium L x-ray intensities

Hybridization chain reaction amplification for highly sensitive fluorescence detection of DNA with dextran coated microarrays.

Science.gov (United States)

Chao, Jie; Li, Zhenhua; Li, Jing; Peng, Hongzhen; Su, Shao; Li, Qian; Zhu, Changfeng; Zuo, Xiaolei; Song, Shiping; Wang, Lianhui; Wang, Lihua

2016-07-15

Microarrays of biomolecules hold great promise in the fields of genomics, proteomics, and clinical assays on account of their remarkably parallel and high-throughput assay capability. However, the fluorescence detection used in most conventional DNA microarrays is still limited by sensitivity. In this study, we have demonstrated a novel universal and highly sensitive platform for fluorescent detection of sequence specific DNA at the femtomolar level by combining dextran-coated microarrays with hybridization chain reaction (HCR) signal amplification. Three-dimensional dextran matrix was covalently coated on glass surface as the scaffold to immobilize DNA recognition probes to increase the surface binding capacity and accessibility. DNA nanowire tentacles were formed on the matrix surface for efficient signal amplification by capturing multiple fluorescent molecules in a highly ordered way. By quantifying microscopic fluorescent signals, the synergetic effects of dextran and HCR greatly improved sensitivity of DNA microarrays, with a detection limit of 10fM (1×10(5) molecules). This detection assay could recognize one-base mismatch with fluorescence signals dropped down to ~20%. This cost-effective microarray platform also worked well with samples in serum and thus shows great potential for clinical diagnosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Development of a genotype-by-sequencing immunogenetic assay as exemplified by screening for variation in red fox with and without endemic rabies exposure.

Science.gov (United States)

Donaldson, Michael E; Rico, Yessica; Hueffer, Karsten; Rando, Halie M; Kukekova, Anna V; Kyle, Christopher J

2018-01-01

Pathogens are recognized as major drivers of local adaptation in wildlife systems. By determining which gene variants are favored in local interactions among populations with and without disease, spatially explicit adaptive responses to pathogens can be elucidated. Much of our current understanding of host responses to disease comes from a small number of genes associated with an immune response. High-throughput sequencing (HTS) technologies, such as genotype-by-sequencing (GBS), facilitate expanded explorations of genomic variation among populations. Hybridization-based GBS techniques can be leveraged in systems not well characterized for specific variants associated with disease outcome to "capture" specific genes and regulatory regions known to influence expression and disease outcome. We developed a multiplexed, sequence capture assay for red foxes to simultaneously assess ~300-kbp of genomic sequence from 116 adaptive, intrinsic, and innate immunity genes of predicted adaptive significance and their putative upstream regulatory regions along with 23 neutral microsatellite regions to control for demographic effects. The assay was applied to 45 fox DNA samples from Alaska, where three arctic rabies strains are geographically restricted and endemic to coastal tundra regions, yet absent from the boreal interior. The assay provided 61.5% on-target enrichment with relatively even sequence coverage across all targeted loci and samples (mean = 50×), which allowed us to elucidate genetic variation across introns, exons, and potential regulatory regions (4,819 SNPs). Challenges remained in accurately describing microsatellite variation using this technique; however, longer-read HTS technologies should overcome these issues. We used these data to conduct preliminary analyses and detected genetic structure in a subset of red fox immune-related genes between regions with and without endemic arctic rabies. This assay provides a template to assess immunogenetic variation

Staphylococcus aureus DNA ligase: characterization of its kinetics of catalysis and development of a high-throughput screening compatible chemiluminescent hybridization protection assay.

Science.gov (United States)

Gul, Sheraz; Brown, Richard; May, Earl; Mazzulla, Marie; Smyth, Martin G; Berry, Colin; Morby, Andrew; Powell, David J

2004-11-01

DNA ligases are key enzymes involved in the repair and replication of DNA. Prokaryotic DNA ligases uniquely use NAD+ as the adenylate donor during catalysis, whereas eukaryotic enzymes use ATP. This difference in substrate specificity makes the bacterial enzymes potential targets for therapeutic intervention. We have developed a homogeneous chemiluminescence-based hybridization protection assay for Staphylococcus aureus DNA ligase that uses novel acridinium ester technology and demonstrate that it is an alternative to the commonly used radiometric assays for ligases. The assay has been used to determine a number of kinetic constants for S. aureus DNA ligase catalysis. These included the K(m) values for NAD+ (2.75+/-0.1 microM) and the acridinium-ester-labelled DNA substrate (2.5+/-0.2 nM). A study of the pH-dependencies of kcat, K(m) and kcat/K(m) has revealed values of kinetically influential ionizations within the enzyme-substrate complexes (kcat) and free enzyme (kcat/K(m)). In each case, the curves were shown to be composed of one kinetically influential ionization, for k(cat), pK(a)=6.6+/-0.1 and kcat/K(m), pK(a)=7.1+/-0.1. Inhibition characteristics of the enzyme against two Escherichia coli DNA ligase inhibitors have also been determined with IC50 values for these being 3.30+/-0.86 microM for doxorubicin and 1.40+/-0.07 microM for chloroquine diphosphate. The assay has also been successfully miniaturized to a sufficiently low volume to allow it to be utilized in a high-throughput screen (384-well format; 20 microl reaction volume), enabling the assay to be used in screening campaigns against libraries of compounds to discover leads for further drug development.

Hybrid Droop Control Strategy Applied to Grid-Supporting Converters in DC Microgrids

DEFF Research Database (Denmark)

Han, Renke; Meng, Lexuan; Guerrero, Josep M.

2017-01-01

The paper proposes a hybrid droop control strategy to enhance the stability and increase maximum constant power loads (CPLs) capability of DC microgrids in a realistic scenario. By capturing the detailed model of inner control loops and hybrid droop control and general dc MG topology, a thorough...

DyHAP: Dynamic Hybrid ANFIS-PSO Approach for Predicting Mobile Malware.

Science.gov (United States)

Afifi, Firdaus; Anuar, Nor Badrul; Shamshirband, Shahaboddin; Choo, Kim-Kwang Raymond

2016-01-01

To deal with the large number of malicious mobile applications (e.g. mobile malware), a number of malware detection systems have been proposed in the literature. In this paper, we propose a hybrid method to find the optimum parameters that can be used to facilitate mobile malware identification. We also present a multi agent system architecture comprising three system agents (i.e. sniffer, extraction and selection agent) to capture and manage the pcap file for data preparation phase. In our hybrid approach, we combine an adaptive neuro fuzzy inference system (ANFIS) and particle swarm optimization (PSO). Evaluations using data captured on a real-world Android device and the MalGenome dataset demonstrate the effectiveness of our approach, in comparison to two hybrid optimization methods which are differential evolution (ANFIS-DE) and ant colony optimization (ANFIS-ACO).

DyHAP: Dynamic Hybrid ANFIS-PSO Approach for Predicting Mobile Malware.

Directory of Open Access Journals (Sweden)

Firdaus Afifi

Full Text Available To deal with the large number of malicious mobile applications (e.g. mobile malware, a number of malware detection systems have been proposed in the literature. In this paper, we propose a hybrid method to find the optimum parameters that can be used to facilitate mobile malware identification. We also present a multi agent system architecture comprising three system agents (i.e. sniffer, extraction and selection agent to capture and manage the pcap file for data preparation phase. In our hybrid approach, we combine an adaptive neuro fuzzy inference system (ANFIS and particle swarm optimization (PSO. Evaluations using data captured on a real-world Android device and the MalGenome dataset demonstrate the effectiveness of our approach, in comparison to two hybrid optimization methods which are differential evolution (ANFIS-DE and ant colony optimization (ANFIS-ACO.

DyHAP: Dynamic Hybrid ANFIS-PSO Approach for Predicting Mobile Malware

Science.gov (United States)

Afifi, Firdaus; Anuar, Nor Badrul; Shamshirband, Shahaboddin

2016-01-01

To deal with the large number of malicious mobile applications (e.g. mobile malware), a number of malware detection systems have been proposed in the literature. In this paper, we propose a hybrid method to find the optimum parameters that can be used to facilitate mobile malware identification. We also present a multi agent system architecture comprising three system agents (i.e. sniffer, extraction and selection agent) to capture and manage the pcap file for data preparation phase. In our hybrid approach, we combine an adaptive neuro fuzzy inference system (ANFIS) and particle swarm optimization (PSO). Evaluations using data captured on a real-world Android device and the MalGenome dataset demonstrate the effectiveness of our approach, in comparison to two hybrid optimization methods which are differential evolution (ANFIS-DE) and ant colony optimization (ANFIS-ACO). PMID:27611312

DETECTION OF DNA DAMAGE USING A FIBEROPTIC BIOSENSOR

Science.gov (United States)

A rapid and sensitive fiber optic biosensor assay for radiation-induced DNA damage is reported. For this assay, a biotin-labeled capture oligonucleotide (38 mer) was immobilized to an avidin-coated quartz fiber. Hybridization of a dye-labeled complementary sequence was observed...

Development of a lion-specific interferon-gamma assay.

Science.gov (United States)

Maas, M; van Kooten, P J S; Schreuder, J; Morar, D; Tijhaar, E; Michel, A L; Rutten, V P M G

2012-10-15

The ongoing spread of bovine tuberculosis (BTB) in African free-ranging lion populations, for example in the Kruger National Park, raises the need for diagnostic assays for BTB in lions. These, in addition, would be highly relevant for zoological gardens worldwide that want to determine the BTB status of their lions, e.g. for translocations. The present study concerns the development of a lion-specific IFN-γ assay, following the production and characterization of monoclonal antibodies specific for lion interferon-gamma (IFN-γ). Recombinant lion IFN-γ (rLIFN-γ) was produced in mammalian cells and used to immunize mice to establish hybridoma cell lines producing monoclonal antibodies. These were used to develop a sensitive, lion IFN-γ-specific capture ELISA, able to detect rLIFN-γ to the level of 160 pg/ml. Recognition of native lion IFN-γ was shown in an initial assessment of supernatants of mitogen stimulated whole blood cultures of 11 known BTB-negative lions. In conclusion, the capture ELISA shows potential as a diagnostic assay for bovine tuberculosis in lions. Preliminary results also indicate the possible use of the test for other (feline) species. Copyright © 2012 Elsevier B.V. All rights reserved.

Rapid Active Assay for the Detection of Antibodies to West Nile Virus in Chickens

National Research Council Canada - National Science Library

Groves, Stephanie S; Turell, Michael J; Bailey, Charles L; Morozov, Victor N

2008-01-01

... by detection of bound IgM molecules with functionalized magnetic beads as active labels. This assay takes only 15 minutes and has the same sensitivity as a commercially available human WNV IgM antibody-capture enzyme-linked immunosorbent assay...

Non-destructive assay system for uranium and plutonium in reprocessing input solutions. Hybrid K-edge/XRF Densitometer. JASPAS JC-11 final report

International Nuclear Information System (INIS)

Surugaya, N.; Abe, K.; Kurosawa, A.; Ikeda, H.; Kuno, Y.

1997-05-01

As a part of JASPAS programme, a non-radioactive assay system for the accountability of uranium and plutonium in input dissolver solutions of a spent fuel reprocessing plant, called Hybrid K-edge/XRF Densitometer, has been developed at the Tokai Reprocessing plant (TRP) since 1991. The instrument is the one of the hybrid type combined K-edge densitometry (KED) and X-ray fluorescence (XRF) analysis. The KED is used to determine the uranium concentration and the XRF is used to determine the U/Pu ratio. These results give the plutonium concentration in consequence. It is considered that the instrument has the capability of timely on-site verification for input accountancy. The instrument had been installed in the analytical hot cell at the TRP and the experiments comparing with Isotope Dilution Mass Spectrometry (IDMS) method have been carried out. As the results of measurements for the actual input solutions in the acceptance and performance tests, it was typically confirmed that the precision for determining uranium concentration by the KED was within 0.2%, whereas the XRF for plutonium performed within 0.7%. This final report summarizes the design information and performance data so as to end the JASPAS programme. (author)

Development of a VHH-Based Erythropoietin Quantification Assay

DEFF Research Database (Denmark)

Kol, Stefan; Beuchert Kallehauge, Thomas; Adema, Simon

2015-01-01

Erythropoietin (EPO) quantification during cell line selection and bioreactor cultivation has traditionally been performed with ELISA or HPLC. As these techniques suffer from several drawbacks, we developed a novel EPO quantification assay. A camelid single-domain antibody fragment directed against...... human EPO was evaluated as a capturing antibody in a label-free biolayer interferometry-based quantification assay. Human recombinant EPO can be specifically detected in Chinese hamster ovary cell supernatants in a sensitive and pH-dependent manner. This method enables rapid and robust quantification...

Sensitive electrochemical assaying of DNA methyltransferase activity based on mimic-hybridization chain reaction amplified strategy.

Science.gov (United States)

Zhang, Linqun; Liu, Yuanjian; Li, Ying; Zhao, Yuewu; Wei, Wei; Liu, Songqin

2016-08-24

A mimic-hybridization chain reaction (mimic-HCR) amplified strategy was proposed for sensitive electrochemically detection of DNA methylation and methyltransferase (MTase) activity In the presence of methylated DNA, DNA-gold nanoparticles (DNA-AuNPs) were captured on the electrode by sandwich-type assembly. It then triggered mimic-HCR of two hairpin probes to produce many long double-helix chains for numerous hexaammineruthenium (III) chloride ([Ru(NH3)6](3+), RuHex) inserting. As a result, the signal for electrochemically detection of DNA MTase activity could be amplified. If DNA was non-methylated, however, the sandwich-type assembly would not form because the short double-stranded DNAs (dsDNA) on the Au electrode could be cleaved and digested by restriction endonuclease HpaII (HapII) and exonuclease III (Exo III), resulting in the signal decrement. Based on this, an electrochemical approach for detection of M.SssI MTase activity with high sensitivity was developed. The linear range for M.SssI MTase activity was from 0.05 U mL(-1) to 10 U mL(-1), with a detection limit down to 0.03 U mL(-1). Moreover, this detecting strategy held great promise as an easy-to-use and highly sensitive method for other MTase activity and inhibition detection by exchanging the corresponding DNA sequence. Copyright © 2016 Elsevier B.V. All rights reserved.

AN ELISA ASSAY FOR HEME OXYGENASE (HO-1)

Science.gov (United States)

An ELISA assay for heme oxygenase (HO-l ) Abstract A double antibody capture ELISA for the HO-l protein has been developed to separately quantitate HO-I protein. The use of 2.5% NP40 detergent greatly assists in freeing HO-l protein from membranes and/or other cel...

Development and accuracy of quantitative real-time polymerase chain reaction assays for detection and quantification of enterotoxigenic Escherichia coli (ETEC) heat labile and heat stable toxin genes in travelers' diarrhea samples.

Science.gov (United States)

Youmans, Bonnie P; Ajami, Nadim J; Jiang, Zhi-Dong; Petrosino, Joseph F; DuPont, Herbert L; Highlander, Sarah K

2014-01-01

Enterotoxigenic Escherichia coli (ETEC), the leading bacterial pathogen of travelers' diarrhea, is routinely detected by an established DNA hybridization protocol that is neither sensitive nor quantitative. Quantitative real-time polymerase chain reaction (qPCR) assays that detect the ETEC toxin genes eltA, sta1, and sta2 in clinical stool samples were developed and tested using donor stool inoculated with known quantities of ETEC bacteria. The sensitivity of the qPCR assays is 89%, compared with 22% for the DNA hybridization assay, and the limits of detection are 10,000-fold lower than the DNA hybridization assays performed in parallel. Ninety-three clinical stool samples, previously characterized by DNA hybridization, were tested using the new ETEC qPCR assays. Discordant toxin profiles were observed for 22 samples, notably, four samples originally typed as ETEC negative were ETEC positive. The qPCR assays are unique in their sensitivity and ability to quantify the three toxin genes in clinical stool samples.

Specific RNP capture with antisense LNA/DNA mixmers.

Science.gov (United States)

Rogell, Birgit; Fischer, Bernd; Rettel, Mandy; Krijgsveld, Jeroen; Castello, Alfredo; Hentze, Matthias W

2017-08-01

RNA-binding proteins (RBPs) play essential roles in RNA biology, responding to cellular and environmental stimuli to regulate gene expression. Important advances have helped to determine the (near) complete repertoires of cellular RBPs. However, identification of RBPs associated with specific transcripts remains a challenge. Here, we describe "specific ribonucleoprotein (RNP) capture," a versatile method for the determination of the proteins bound to specific transcripts in vitro and in cellular systems. Specific RNP capture uses UV irradiation to covalently stabilize protein-RNA interactions taking place at "zero distance." Proteins bound to the target RNA are captured by hybridization with antisense locked nucleic acid (LNA)/DNA oligonucleotides covalently coupled to a magnetic resin. After stringent washing, interacting proteins are identified by quantitative mass spectrometry. Applied to in vitro extracts, specific RNP capture identifies the RBPs bound to a reporter mRNA containing the Sex-lethal (Sxl) binding motifs, revealing that the Sxl homolog sister of Sex lethal (Ssx) displays similar binding preferences. This method also revealed the repertoire of RBPs binding to 18S or 28S rRNAs in HeLa cells, including previously unknown rRNA-binding proteins. © 2017 Rogell et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Immunocapture loop-mediated isothermal amplification assays for the detection of canine parvovirus.

Science.gov (United States)

Sun, Yu-Ling; Yen, Chon-Ho; Tu, Ching-Fu

2017-11-01

A loop-mediated isothermal amplification (LAMP) assay was used for rapid canine parvovirus (CPV) diagnosis. To reduce the time required and increase the sensitivity of the assay, an immunocapture (IC) technique was developed in this study to exclude the DNA extraction step in molecular diagnostic procedures for CPV. A polyclonal rabbit anti-CPV serum was produced against VP2-EpC that was cloned via DNA recombination. The polyclonal anti-VP2-EpC serum was used for virus capture to prepare microtubes. IC-LAMP was performed to amplify a specific CPV target gene sequence from the CPV viral particles that were captured on the microtubes, and the amplicons were analyzed using agarose electrophoresis or enzyme-linked immunosorbent assay (IC-LAMP-ELISA) and lateral-flow dipstick (IC-LAMP-LFD). The detection sensitivities of IC-LAMP, IC-LAMP-ELISA, and IC-LAMP-LFD were 10 -1 , 10 -1 , and 10 -1 TCID 50 /mL, respectively. Using the IC-LAMP-ELISA and IC-LAMP-LFD assays, the complete CPV diagnostic process can be achieved within 1.5h. Both of the developed IC-LAMP-based assays are simple, direct visual and efficient techniques that are applicable to the detection of CPV. Copyright © 2017 Elsevier B.V. All rights reserved.

Grafted Cross-Linked Polyolefin Substrates for Peptide Synthesis and Assays

DEFF Research Database (Denmark)

1999-01-01

suited for use in solid-phase biosystems, notably bioassays, such as immunoassays, DNA hybridization assays or PCR amplification. The grafted chains may bear substituents which are such that the polymer-grafted cross-linked polyolefin substrate is swellable by water or aqueous media, in other words...

Cystatin capture elisa immunodiagnosis of human fasciolosis at Chupaca-Junin Province

International Nuclear Information System (INIS)

Cornejo, W.; Alva, P.; Sevilla, C.; Huiza, A.; Universidad Nacional Mayor de San Marcos, Lima

2003-01-01

To determine the prevalence of human fasciolosis in an endemic area by means of an Enzyme-Linked Immunosorbent Assay (ELISA) using cystatin as a capture agent for the detection of specific antibodies to fasciola hepatica cysteine proteinases. An ELISA plate was sensitized with cystatin, incubated with excretory-secretory products of adult flukes, and followed by standard ELISA procedures. Clinical applicability of the cystatin capture ELISA for the immunodiagnosis of fasciolosis was tested with 200 serum samples of children and adults from an endemic area in Chupaca province, Junin department. Serum samples from the endemic area tested by cystatin capture ELISA showed 27/200 (13,5%) of positive cases. Fasciolosis remains a major health problem at Chupaca province, Junin department

Femtomolar detection of single mismatches by discriminant analysis of DNA hybridization events using gold nanoparticles.

Science.gov (United States)

Ma, Xingyi; Sim, Sang Jun

2013-03-21

Even though DNA-based nanosensors have been demonstrated for quantitative detection of analytes and diseases, hybridization events have never been numerically investigated for further understanding of DNA mediated interactions. Here, we developed a nanoscale platform with well-designed capture and detection gold nanoprobes to precisely evaluate the hybridization events. The capture gold nanoprobes were mono-laid on glass and the detection probes were fabricated via a novel competitive conjugation method. The two kinds of probes combined in a suitable orientation following the hybridization with the target. We found that hybridization efficiency was markedly dependent on electrostatic interactions between DNA strands, which can be tailored by adjusting the salt concentration of the incubation solution. Due to the much lower stability of the double helix formed by mismatches, the hybridization efficiencies of single mismatched (MMT) and perfectly matched DNA (PMT) were different. Therefore, we obtained an optimized salt concentration that allowed for discrimination of MMT from PMT without stringent control of temperature or pH. The results indicated this to be an ultrasensitive and precise nanosensor for the diagnosis of genetic diseases.

High-throughput screening of metal-porphyrin-like graphenes for selective capture of carbon dioxide.

Science.gov (United States)

Bae, Hyeonhu; Park, Minwoo; Jang, Byungryul; Kang, Yura; Park, Jinwoo; Lee, Hosik; Chung, Haegeun; Chung, ChiHye; Hong, Suklyun; Kwon, Yongkyung; Yakobson, Boris I; Lee, Hoonkyung

2016-02-23

Nanostructured materials, such as zeolites and metal-organic frameworks, have been considered to capture CO2. However, their application has been limited largely because they exhibit poor selectivity for flue gases and low capture capacity under low pressures. We perform a high-throughput screening for selective CO2 capture from flue gases by using first principles thermodynamics. We find that elements with empty d orbitals selectively attract CO2 from gaseous mixtures under low CO2 pressures (~10(-3) bar) at 300 K and release it at ~450 K. CO2 binding to elements involves hybridization of the metal d orbitals with the CO2 π orbitals and CO2-transition metal complexes were observed in experiments. This result allows us to perform high-throughput screening to discover novel promising CO2 capture materials with empty d orbitals (e.g., Sc- or V-porphyrin-like graphene) and predict their capture performance under various conditions. Moreover, these findings provide physical insights into selective CO2 capture and open a new path to explore CO2 capture materials.

High-throughput screening of metal-porphyrin-like graphenes for selective capture of carbon dioxide

Science.gov (United States)

Bae, Hyeonhu; Park, Minwoo; Jang, Byungryul; Kang, Yura; Park, Jinwoo; Lee, Hosik; Chung, Haegeun; Chung, Chihye; Hong, Suklyun; Kwon, Yongkyung; Yakobson, Boris I.; Lee, Hoonkyung

2016-02-01

Nanostructured materials, such as zeolites and metal-organic frameworks, have been considered to capture CO2. However, their application has been limited largely because they exhibit poor selectivity for flue gases and low capture capacity under low pressures. We perform a high-throughput screening for selective CO2 capture from flue gases by using first principles thermodynamics. We find that elements with empty d orbitals selectively attract CO2 from gaseous mixtures under low CO2 pressures (~10-3 bar) at 300 K and release it at ~450 K. CO2 binding to elements involves hybridization of the metal d orbitals with the CO2 π orbitals and CO2-transition metal complexes were observed in experiments. This result allows us to perform high-throughput screening to discover novel promising CO2 capture materials with empty d orbitals (e.g., Sc- or V-porphyrin-like graphene) and predict their capture performance under various conditions. Moreover, these findings provide physical insights into selective CO2 capture and open a new path to explore CO2 capture materials.

Direct Competitive Enzyme-Linked Immunosorbent Assay (ELISA).

Science.gov (United States)

Kohl, Thomas O; Ascoli, Carl A

2017-07-05

The competitive enzyme-linked immunosorbent assay (ELISA) (cELISA; also called an inhibition ELISA) is designed so that purified antigen competes with antigen in the test sample for binding to an antibody that has been immobilized in microtiter plate wells. The same concept works if the immobilized molecule is antigen and the competing molecules are purified labeled antibody versus antibody in a test sample. Direct cELISAs incorporate labeled antigen or antibody, whereas indirect assay configurations use reporter-labeled secondary antibodies. The cELISA is very useful for determining the concentration of small-molecule antigens in complex sample mixtures. In the direct cELISA, antigen-specific capture antibody is adsorbed onto the microtiter plate before incubation with either known standards or unknown test samples. Enzyme-linked antigen (i.e., labeled antigen) is also added, which can bind to the capture antibody only when the antibody's binding site is not occupied by either the antigen standard or antigen in the test samples. Unbound labeled and unlabeled antigens are washed away and substrate is added. The amount of antigen in the standard or the test sample determines the amount of reporter-labeled antigen bound to antibody, yielding a signal that is inversely proportional to antigen concentration within the sample. Thus, the higher the antigen concentration in the test sample, the less labeled antigen is bound to the capture antibody, and hence the weaker is the resultant signal. © 2017 Cold Spring Harbor Laboratory Press.

A murine monoclonal antibody based enzyme-linked immunosorbent assay for almond (Prunus dulcis L.) detection.

Science.gov (United States)

Su, Mengna; Venkatachalam, Mahesh; Liu, Changqi; Zhang, Ying; Roux, Kenneth H; Sathe, Shridhar K

2013-11-13

A sandwich enzyme-linked immunosorbent assay (ELISA) using anti-almond soluble protein rabbit polyclonal antibodies as capture antibodies and murine monoclonal antibody 4C10 as the detection antibodies was developed. The assay is specific and sensitive (3-200 ng almond protein/mL) for almond detection. The standardized assay is accurate (assay variability assay did not register any cross-reactivity with the tested food matrices, suggesting the assay to be almond amandin specific. The assay could detect the presence of declared almond in the tested matched commercial samples. Further, the assay reliably detected the presence of almonds in the laboratory prepared food samples spiked with almond flour.

Serotype determination of Salmonella by xTAG assay.

Science.gov (United States)

Zheng, Zhibei; Zheng, Wei; Wang, Haoqiu; Pan, Jincao; Pu, Xiaoying

2017-10-01

Currently, no protocols or commercial kits are available to determine the serotypes of Salmonella by using Luminex MAGPIX®. In this study, an xTAG assay for serotype determination of Salmonella suitable for Luminex MAGPIX® is described and 228 Salmonella isolates were serotype determined by this xTAG assay. The xTAG assay consists of two steps: 1) Multiplex PCR to amplify simultaneously O, H and Vi antigen genes of Salmonella, and 2) Magplex-TAG™ microsphere hybridization to identify accurately the specific PCR products of different antigens. Compared with the serotyping results of traditional serum agglutination test, the sensitivity and specificity of the xTAG assay were 95.1% and 100%, respectively. The agreement rate of these two assays was 95.2%. Compared with Luminex xMAP® Salmonella Serotyping Assay (SSA) kit, the advantages of this xTAG assay are: First, the magnetic beads make it applicable to both the Luminex®100/200™ and MAGPIX® systems. Second, only primers rather than both primers and probes are needed in the xTAG assay, and the process of coupling antigen-specific oligonucleotide probes to beads is circumvented, which make the xTAG assay convenient to be utilized by other laboratories. The xTAG assay may serve as a rapid alternative or complementary method for traditional Salmonella serotyping tests, especially for laboratories that utilize the MAGPIX® systems. Copyright © 2017 Elsevier B.V. All rights reserved.

Detection of proteins using a colorimetric bio-barcode assay.

Science.gov (United States)

Nam, Jwa-Min; Jang, Kyung-Jin; Groves, Jay T

2007-01-01

The colorimetric bio-barcode assay is a red-to-blue color change-based protein detection method with ultrahigh sensitivity. This assay is based on both the bio-barcode amplification method that allows for detecting miniscule amount of targets with attomolar sensitivity and gold nanoparticle-based colorimetric DNA detection method that allows for a simple and straightforward detection of biomolecules of interest (here we detect interleukin-2, an important biomarker (cytokine) for many immunodeficiency-related diseases and cancers). The protocol is composed of the following steps: (i) conjugation of target capture molecules and barcode DNA strands onto silica microparticles, (ii) target capture with probes, (iii) separation and release of barcode DNA strands from the separated probes, (iv) detection of released barcode DNA using DNA-modified gold nanoparticle probes and (v) red-to-blue color change analysis with a graphic software. Actual target detection and quantification steps with premade probes take approximately 3 h (whole protocol including probe preparations takes approximately 3 days).

Amine-oxide hybrid materials for acid gas separations

KAUST Repository

Bollini, Praveen

2011-01-01

Organic-inorganic hybrid materials based on porous silica materials functionalized with amine-containing organic species are emerging as an important class of materials for the adsorptive separation of acid gases from dilute gas streams. In particular, these materials are being extensively studied for the adsorption of CO 2 from simulated flue gas streams, with an eye towards utilizing these materials as part of a post-combustion carbon capture process at large flue gas producing installations, such as coal-fired electricity-generating power plants. In this Application Article, the utilization of amine-modified organic-inorganic hybrid materials is discussed, focusing on important attributes of the materials, such as (i) CO 2 adsorption capacities, (ii) adsorption and desorption kinetics, and (iii) material stability, that will determine if these materials may one day be useful adsorbents in practical CO 2 capture applications. Specific research needs and limitations associated with the current body of work are identified. © 2011 The Royal Society of Chemistry.

Polyetheretherketone Hybrid Composites with Bioactive Nanohydroxyapatite and Multiwalled Carbon Nanotube Fillers

Directory of Open Access Journals (Sweden)

Chen Liu

2016-12-01

Full Text Available Polyetheretherketone (PEEK hybrid composites reinforced with inorganic nanohydroxyapatite (nHA and multiwalled carbon nanotube (MWNT were prepared by melt-compounding and injection molding processes. The additions of nHA and MWNT to PEEK were aimed to increase its elastic modulus, tensile strength, and biocompatibility, rendering the hybrids suitable for load-bearing implant applications. The structural behavior, mechanical property, wettability, osteoblastic cell adhesion, proliferation, differentiation, and mineralization of the PEEK/nHA-MWNT hybrids were studied. X-ray diffraction and SEM observation showed that both nHA and MWNT fillers are incorporated into the polymer matrix of PEEK-based hybrids. Tensile tests indicated that the elastic modulus of PEEK can be increased from 3.87 to 7.13 GPa by adding 15 vol % nHA and 1.88 vol % MWNT fillers. The tensile strength and elongation at break of the PEEK/(15% nHA-(1.88% MWNT hybrid were 64.48 MPa and 1.74%, respectively. Thus the tensile properties of this hybrid were superior to those of human cortical bones. Water contact angle measurements revealed that the PEEK/(15% nHA-(1.88% MWNT hybrid is hydrophilic due to the presence of nHA. Accordingly, hydrophilic PEEK/(15% nHA-(1.88% MWNT hybrid promoted the adhesion, proliferation, differentiation, and mineralization of murine MC3T3-E1 osteoblasts on its surface effectively on the basis of cell culture, fluorescence microscopy, MTT assay, WST-1 assay, alkaline phosphatase activity, and Alizarin red staining tests. Thus the PEEK/(15% nHA-(1.88% MWNT hybrid has the potential to be used for fabricating load-bearing bone implants.

Design of novel hybrid organic-inorganic nanostructured biomaterials for immunoassay applications

Energy Technology Data Exchange (ETDEWEB)

Andrade, G [Department of Microbiology, Institute of Biological Sciences, Federal University of Minas Gerais, PO Box 486, 31270.901, Belo Horizonte, MG (Brazil); Barbosa-Stancioli, E F [Department of Microbiology, Institute of Biological Sciences, Federal University of Minas Gerais, PO Box 486, 31270.901, Belo Horizonte, MG (Brazil); Piscitelli Mansur, A A [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil); Vasconcelos, W L [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil); Mansur, H S [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil)

2006-12-01

The purpose of this study was to develop novel hybrid organic-inorganic materials based on poly(vinyl alcohol) (PVA) polymer chemically crosslinked network to be tested as solid support on bovine herpesvirus immunoassay. Hybrids were synthesized by reacting PVA with three different alkoxysilanes modifying chemical groups: tetraethoxysilane (TEOS), 3-mercaptopropyltrimethoxysilane (MPTMS) and 3-glycidoxypropyltrimethoxysilane (GPTMS). PVA-derived hybrids were also modified by chemically crosslinking with glutaraldehyde (GA) during the synthesis reaction. In order to investigate the structure in the nanometer-scale, PVA-derived hybrids were characterized by using small-angle x-ray scattering synchrotron radiation (SAXS) and x-ray diffraction (XRD). PVA hybrids' chemical functionalities and their interaction with herpesviruses were also characterized by Fourier transform infrared spectroscopy (FTIR). The bioactivity assays were tested through enzyme linked immunosorbent assay (ELISA). SAXS results have indicated nano-ordered disperse domains for PVA hybrids with different x-ray scattering patterns for PVA polymer and PVA-derived hybrids. FTIR spectra have shown major vibration bands associated with organic-inorganic chemical groups present in the PVA, PVA-derived by silane modifier and PVA chemically crosslinked by GA. The immunoassay results have shown that PVA hybrids with chemically functionalized structures regulated to some extent the specific bioimmobilization of herpesvirus onto solid phase. We think that it is due to the overall balance of forces associated with van der Waals interaction, hydrophilic and hydrophobic forces and steric hindrance acting at the surface. PVA and PVA-derived hybrid materials were successfully produced with GA crosslinking in a nanometer-scale network. Also, such a PVA-based material could be advantageously used in immunoassays with enhanced specificity for diagnosis.

Design of novel hybrid organic-inorganic nanostructured biomaterials for immunoassay applications

Energy Technology Data Exchange (ETDEWEB)

Andrade, G [Department of Microbiology, Institute of Biological Sciences, Federal University of Minas Gerais, PO Box 486, 31270.901, Belo Horizonte, MG (Brazil); Barbosa-Stancioli, E F [Department of Microbiology, Institute of Biological Sciences, Federal University of Minas Gerais, PO Box 486, 31270.901, Belo Horizonte, MG (Brazil); Piscitelli Mansur, A A [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil); Vasconcelos, W L [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil); Mansur, H S [Department of Metallurgical and Materials Engineering, Biomaterials and Tissue Engineering Laboratory, Federal University of Minas Gerais, Belo Horizonte, MG (Brazil)

2006-12-01

The purpose of this study was to develop novel hybrid organic-inorganic materials based on poly(vinyl alcohol) (PVA) polymer chemically crosslinked network to be tested as solid support on bovine herpesvirus immunoassay. Hybrids were synthesized by reacting PVA with three different alkoxysilanes modifying chemical groups: tetraethoxysilane (TEOS), 3-mercaptopropyltrimethoxysilane (MPTMS) and 3-glycidoxypropyltrimethoxysilane (GPTMS). PVA-derived hybrids were also modified by chemically crosslinking with glutaraldehyde (GA) during the synthesis reaction. In order to investigate the structure in the nanometer-scale, PVA-derived hybrids were characterized by using small-angle x-ray scattering synchrotron radiation (SAXS) and x-ray diffraction (XRD). PVA hybrids' chemical functionalities and their interaction with herpesviruses were also characterized by Fourier transform infrared spectroscopy (FTIR). The bioactivity assays were tested through enzyme linked immunosorbent assay (ELISA). SAXS results have indicated nano-ordered disperse domains for PVA hybrids with different x-ray scattering patterns for PVA polymer and PVA-derived hybrids. FTIR spectra have shown major vibration bands associated with organic-inorganic chemical groups present in the PVA, PVA-derived by silane modifier and PVA chemically crosslinked by GA. The immunoassay results have shown that PVA hybrids with chemically functionalized structures regulated to some extent the specific bioimmobilization of herpesvirus onto solid phase. We think that it is due to the overall balance of forces associated with van der Waals interaction, hydrophilic and hydrophobic forces and steric hindrance acting at the surface. PVA and PVA-derived hybrid materials were successfully produced with GA crosslinking in a nanometer-scale network. Also, such a PVA-based material could be advantageously used in immunoassays with enhanced specificity for diagnosis.

Two types of nanoparticle-based bio-barcode amplification assays to detect HIV-1 p24 antigen

Directory of Open Access Journals (Sweden)

Dong Huahuang

2012-08-01

Full Text Available Abstract Background HIV-1 p24 antigen is a major viral component of human immunodeficiency virus type 1 (HIV-1 which can be used to identify persons in the early stage of infection and transmission of HIV-1 from infected mothers to infants. The detection of p24 is usually accomplished by using an enzyme-linked immunosorbent assay (ELISA with low detection sensitivity. Here we report the use of two bio-barcode amplification (BCA assays combined with polymerase chain reaction (PCR and gel electrophoresis to quantify HIV-1 p24 antigen. Method A pair of anti-p24 monoclonal antibodies (mAbs were used in BCA assays to capture HIV-1 p24 antigen in a sandwich format and allowed for the quantitative measurement of captured p24 using PCR and gel electrophoresis. The first 1 G12 mAb was coated on microplate wells or magnetic microparticles (MMPs to capture free p24 antigens. Captured p24 in turn captured 1D4 mAb coated gold nanoparticle probes (GNPs containing double-stranded DNA oligonucleotides. One strand of the oligonucleotides was covalently immobilized whereas the unbound complimentary bio-barcode DNA strand could be released upon heating. The released bio-barcode DNA was amplified by PCR, electrophoresed in agarose gel and quantified. Results The in-house ELISA assay was found to quantify p24 antigen with a limit of detection (LOD of 1,000 pg/ml and a linear range between 3,000 and 100,000 pg/ml. In contrast, the BCA-based microplate method yielded an LOD of 1 pg/ml and a linear detection range from 1 to 10,000 pg/ml. The BCA-based MMP method yielded an LOD of 0.1 pg/ml and a linear detection range from 0.1 to 1,000 pg/ml. Conclusions When combined with PCR and simple gel electrophoresis, BCA-based microplate and MMPs assays can be used to quantify HIV-1 p24 antigen. These methods are 3–4 orders of magnitude more sensitive than our in-house ELISA-based assay and may provide a useful approach to detect p24 in patients newly infected

What controls the hybridization thermodynamics of spherical nucleic acids?

Science.gov (United States)

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

2015-03-18

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

Early events in speciation: Polymorphism for hybrid male sterility in Drosophila

OpenAIRE

Reed, Laura K.; Markow, Therese A.

2004-01-01

Capturing the process of speciation early enough to determine the initial genetic causes of reproductive isolation remains a major challenge in evolutionary biology. We have found, to our knowledge, the first example of substantial intraspecific polymorphism for genetic factors contributing to hybrid male sterility. Specifically, we show that the occurrence of hybrid male sterility in crosses between Drosophila mojavensis and its sister species, Drosophila arizonae, is controlled by factors p...

Genomic Prediction of Barley Hybrid Performance

Directory of Open Access Journals (Sweden)

Norman Philipp

2016-07-01

Full Text Available Hybrid breeding in barley ( L. offers great opportunities to accelerate the rate of genetic improvement and to boost yield stability. A crucial requirement consists of the efficient selection of superior hybrid combinations. We used comprehensive phenotypic and genomic data from a commercial breeding program with the goal of examining the potential to predict the hybrid performances. The phenotypic data were comprised of replicated grain yield trials for 385 two-way and 408 three-way hybrids evaluated in up to 47 environments. The parental lines were genotyped using a 3k single nucleotide polymorphism (SNP array based on an Illumina Infinium assay. We implemented ridge regression best linear unbiased prediction modeling for additive and dominance effects and evaluated the prediction ability using five-fold cross validations. The prediction ability of hybrid performances based on general combining ability (GCA effects was moderate, amounting to 0.56 and 0.48 for two- and three-way hybrids, respectively. The potential of GCA-based hybrid prediction requires that both parental components have been evaluated in a hybrid background. This is not necessary for genomic prediction for which we also observed moderate cross-validated prediction abilities of 0.51 and 0.58 for two- and three-way hybrids, respectively. This exemplifies the potential of genomic prediction in hybrid barley. Interestingly, prediction ability using the two-way hybrids as training population and the three-way hybrids as test population or vice versa was low, presumably, because of the different genetic makeup of the parental source populations. Consequently, further research is needed to optimize genomic prediction approaches combining different source populations in barley.

Development of novel biocompatible hybrid nanocomposites based on polyurethane-silica prepared by sol gel process

Energy Technology Data Exchange (ETDEWEB)

Rashti, Ali [Department of Medical Nanotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Yahyaei, Hossein [Department of Polymer Engineering and Color Technology, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Firoozi, Saman [Department of Tissue Engineering & Regenerative Medicine, Faculty of Advanced Technologies in Medicine, Iran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Ramezani, Sara [Department of Neuroscience, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Rahiminejad, Ali [Department of Medical Nanotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Karimi, Roya [Department of Tissue Engineering and Applied Cell Science, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Farzaneh, Khadijeh [Tehran Heart Center, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Mohseni, Mohsen [Department of Polymer Engineering and Color Technology, Amirkabir University of Technology, Tehran (Iran, Islamic Republic of); Ghanbari, Hossein, E-mail: hghanbari@tums.ac.ir [Department of Medical Nanotechnology, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Tehran Heart Center, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Medical Biomaterials Research Center, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of)

2016-12-01

Due to high biocompatibility, polyurethane has found many applications, particularly in development of biomedical devices. A new nanocomposite based on thermoset polyurethane and silica nanoparticles was synthesized using sol-gel method. Sol-gel process was fulfilled in two acidic and basic conditions by using tetraethylorthosilicate (TEOS) and trimethoxyisocyanatesilane as precursors. The hybrid films characterized for mechanical and surface properties using tensile strength, contact angle, ATR-FTIR and scanning electron microscopy. Biocompatibility and cytotoxicity of the hybrids were assessed using standard MTT, LDH and TUNEL assays. The results revealed that incorporation of silica nanoparticles was significantly improved tensile strength and mechanical properties of the hybrids. Based on the contact angle results, silica nanoparticles increased hydrophilicity of the hybrids. Biocompatibility by using human lung epithelial cell line (MRC-5) demonstrated that the hybrids were significantly less cytotoxic compared to pristine polymer as tested by MTT and LDH assays. TUNEL assay revealed no signs of apoptosis in all tested samples. The results of this study demonstrated that incorporation of silica nanoparticles into polyurethane lead to the enhancement of biocompatibility, indicating that these hybrids could potentially be used in biomedical field in particular as a new coating for medical implants. - Highlights: • Nanocomposites based on polyurethane and nanosilica prepared by sol-gel method fabricated • Addition of inorganic phase improved mechanical properties. • Nanosilica prepared by sol-gel method increased hydrophilicity. • By adding nanosilica to polyurethane biocompatibility increased significantly.

Synovectomy by Neutron capture

International Nuclear Information System (INIS)

Vega C, H.R.; Torres M, C.

1998-01-01

The Synovectomy by Neutron capture has as purpose the treatment of the rheumatoid arthritis, illness which at present does not have a definitive curing. This therapy requires a neutron source for irradiating the articulation affected. The energy spectra and the intensity of these neutrons are fundamental since these neutrons induce nuclear reactions of capture with Boron-10 inside the articulation and the freely energy of these reactions is transferred at the productive tissue of synovial liquid, annihilating it. In this work it is presented the neutron spectra results obtained with moderator packings of spherical geometry which contains in its center a Pu 239 Be source. The calculations were realized through Monte Carlo method. The moderators assayed were light water, heavy water base and the both combination of them. The spectra obtained, the average energy, the neutron total number by neutron emitted by source, the thermal neutron percentage and the dose equivalent allow us to suggest that the moderator packing more adequate is what has a light water thickness 0.5 cm (radius 2 cm) and 24.5 cm heavy water (radius 26.5 cm). (Author)

Serological Assays Based on Recombinant Viral Proteins for the Diagnosis of Arenavirus Hemorrhagic Fevers

Directory of Open Access Journals (Sweden)

Masayuki Saijo

2012-10-01

Full Text Available The family Arenaviridae, genus Arenavirus, consists of two phylogenetically independent groups: Old World (OW and New World (NW complexes. The Lassa and Lujo viruses in the OW complex and the Guanarito, Junin, Machupo, Sabia, and Chapare viruses in the NW complex cause viral hemorrhagic fever (VHF in humans, leading to serious public health concerns. These viruses are also considered potential bioterrorism agents. Therefore, it is of great importance to detect these pathogens rapidly and specifically in order to minimize the risk and scale of arenavirus outbreaks. However, these arenaviruses are classified as BSL-4 pathogens, thus making it difficult to develop diagnostic techniques for these virus infections in institutes without BSL-4 facilities. To overcome these difficulties, antibody detection systems in the form of an enzyme-linked immunosorbent assay (ELISA and an indirect immunofluorescence assay were developed using recombinant nucleoproteins (rNPs derived from these viruses. Furthermore, several antigen-detection assays were developed. For example, novel monoclonal antibodies (mAbs to the rNPs of Lassa and Junin viruses were generated. Sandwich antigen-capture (Ag-capture ELISAs using these mAbs as capture antibodies were developed and confirmed to be sensitive and specific for detecting the respective arenavirus NPs. These rNP-based assays were proposed to be useful not only for an etiological diagnosis of VHFs, but also for seroepidemiological studies on VHFs. We recently developed arenavirus neutralization assays using vesicular stomatitis virus (VSV-based pseudotypes bearing arenavirus recombinant glycoproteins. The goal of this article is to review the recent advances in developing laboratory diagnostic assays based on recombinant viral proteins for the diagnosis of VHFs and epidemiological studies on the VHFs caused by arenaviruses.

Early events in speciation: polymorphism for hybrid male sterility in Drosophila.

Science.gov (United States)

Reed, Laura K; Markow, Therese A

2004-06-15

Capturing the process of speciation early enough to determine the initial genetic causes of reproductive isolation remains a major challenge in evolutionary biology. We have found, to our knowledge, the first example of substantial intraspecific polymorphism for genetic factors contributing to hybrid male sterility. Specifically, we show that the occurrence of hybrid male sterility in crosses between Drosophila mojavensis and its sister species, Drosophila arizonae, is controlled by factors present at different frequencies in different populations of D. mojavensis. In addition, we show that hybrid male sterility is a complex phenotype; some hybrid males with motile sperm still cannot sire offspring. Because male sterility factors in hybrids between these species are not yet fixed within D. mojavensis, this system provides an invaluable opportunity to characterize the genetics of reproductive isolation at an early stage.

Separation of magnetic beads in a hybrid continuous flow microfluidic device

Energy Technology Data Exchange (ETDEWEB)

Samanta, Abhishek [Haldia Institute of Technology, Production Engineering Department, Haldia (India); Ganguly, Ranjan; Datta, Amitava [Jadavpur University, Power Engineering Department (India); Modak, Nipu, E-mail: nmechju@gmail.com [Jadavpur University, Mechanical Engineering Department (India)

2017-04-01

Magnetic separation of biological entities in microfluidic environment is a key task for a large number of bio-analytical protocols. In magnetophoretic separation, biochemically functionalized magnetic beads are allowed to bind selectively to target analytes, which are then separated from the background stream using a suitably imposed magnetic field. Here we present a numerical study, characterizing the performance of a magnetophoretic hybrid microfluidic device having two inlets and three outlets for immunomagnetic isolation of three different species from a continuous flow. The hybrid device works on the principle of split-flow thin (SPLITT) fractionation and field flow fractionation (FFF) mechanisms. Transport of the magnetic particles in the microchannel has been predicted following an Eulerian-Lagrangian model and using an in-house numerical code. Influence of the salient geometrical parameters on the performance of the separator is studied by characterizing the particle trajectories and their capture and separation indices. Finally, optimum channel geometry is identified that yields the maximum capture efficiency and separation index. - Highlights: • Immunomagnetic separation in a hybrid microchannel design is investigated numerically. • Influence of salient geometric parameters on the device performance is analysed. • Optimum device dimension for best separation parameters are identified. • Optimized design of hybrid separator performs better than FFF or SPLITT devices.

An allele-specific polymerase chain reaction assay for the ...

Indian Academy of Sciences (India)

Unknown

designated as species A, B (Green and Miles 1980), C. (Subbarao et al 1983), ... 3 | September 2004. O P Singh et al. 276 able to distinguish species A from species B/C when sin- gle mosquito-extract was diluted to 1/200. However such hybridization assay .... in Rameshwaram Island and Sri Lanka only) are not pre- sent.

Synthetic wind speed scenarios generation for probabilistic analysis of hybrid energy systems

International Nuclear Information System (INIS)

Chen, Jun; Rabiti, Cristian

2017-01-01

Hybrid energy systems consisting of multiple energy inputs and multiple energy outputs have been proposed to be an effective element to enable ever increasing penetration of clean energy. In order to better understand the dynamic and probabilistic behavior of hybrid energy systems, this paper proposes a model combining Fourier series and autoregressive moving average (ARMA) to characterize historical weather measurements and to generate synthetic weather (e.g., wind speed) data. In particular, Fourier series is used to characterize the seasonal trend in historical data, while ARMA is applied to capture the autocorrelation in residue time series (e.g., measurements with seasonal trends subtracted). The generated synthetic wind speed data is then utilized to perform probabilistic analysis of a particular hybrid energy system configuration, which consists of nuclear power plant, wind farm, battery storage, natural gas boiler, and chemical plant. Requirements on component ramping rate, economic and environmental impacts of hybrid energy systems, and the effects of deploying different sizes of batteries in smoothing renewable variability, are all investigated. - Highlights: • Computational model to synthesize artificial wind speed data with consistent characteristics with database. • Fourier series to capture seasonal trends in the database. • Monte Carlo simulation and probabilistic analysis of hybrid energy systems. • Investigation of the effect of battery in smoothing variability of wind power generation.

Fast and automated DNA assays on a compact disc (CD)-based microfluidic platform

Science.gov (United States)

Jia, Guangyao

Nucleic acid-based molecular diagnostics offers enormous potential for the rapid and accurate diagnosis of infectious diseases. However, most of the existing commercial tests are time-consuming and technically complicated, and are thus incompatible with the need for rapid identification of infectious agents. We have successfully developed a CD-based microfluidic platform for fast and automated DNA array hybridization and a low cost, disposable plastic microfluidic platform for polymerase chain reaction (PCR). These platforms have proved to be a promising approach to meet the requirements in terms of detection speed and operational convenience in diagnosis of infectious diseases. In the CD-based microfluidic platform for DNA hybridization, convection is introduced to the system to enhance mass transport so as to accelerate the hybridization rate since DNA hybridization is a diffusion limited reaction. Centrifugal force is utilized for sample propulsion and surface force is used for liquid gating. Standard microscope glass slides are used as the substrates for capture probes owing to their compatibility with commercially available instrumentation (e.g. laser scanners) for detection. Microfabricated polydimethylsiloxane (PDMS) structures are used to accomplish the fluidic functions required by the protocols for DNA hybridization. The assembly of the PDMS structure and the glass slide forms a flow-through hybridization unit that can be accommodated onto the CD platform for reagent manipulation. The above scheme has been validated with oligonucleotides as the targets using commercially available enzyme-labeled fluorescence (ELF 97) for detection of the hybridization events, and tested with amplicons of genomic staphylococcus DNA labeled with Cy dye. In both experiments, significantly higher fluorescence intensities were observed in the flow-through hybridization unit compared to the passive assays. The CD fluidic scheme was also adapted to the immobilization of

DNA hybridization sensing for cytogenetic analysis

DEFF Research Database (Denmark)

Kwasny, Dorota; Dapra, Johannes; Brøgger, Anna Line

2013-01-01

are rearrangements between two chromosome arms that results in two derivative chromosomes having a mixed DNA sequence. The current detection method is a Fluorescent In situ Hybridization, which requires a use of expensive, fluorescently labeled probes that target the DNA sequences of two chromosomes involved...... in the translocation (Kwasny et al., 2012). We have developed a new double hybridization assay that allows for sorting of the DNA chromosomal fragments into separate compartment, moreover allowing for detection of the translocation. To detect the translocation it is necessary to determine that the two DNA sequences...... forming a derivative chromosome are connected, which is achieved by two subsequent hybridization steps. The first example of the translocation detection was presented on lab-on-a-disc using fluorescently labeled DNA fragments, representing the derivative chromosome (Brøgger et al., 2012). To allow...

A UK NEQAS ISH multicenter ring study using the Ventana HER2 dual-color ISH assay.

LENUS (Irish Health Repository)

Bartlett, J M S

2011-01-01

We performed a multicenter assessment of a new HER2 dual-color chromogenic in situ hybridization (CISH) test and herein report on concordance of CISH data with fluorescence in situ hybridization (FISH) data and intraobserver and interlaboratory scoring consistency. HER2 results were evaluated using duplicate cores from 30 breast cancers in 5 laboratories using the Ventana HER2 dual-color ISH assay (Ventana Medical Systems, Cambridgeshire, England) and in 1 central laboratory using a standard FISH assay. Overall 93.3% of cases were successfully analyzed by CISH across the 5 participating laboratories. There was excellent concordance (98.0% overall) for diagnosis of HER2 amplification by CISH compared with FISH. Intraobserver variability (7.7%) and intersite variability (9.1%) of absolute HER2\\/chromosome enumeration probe 17 ratios were tightly controlled across all participating laboratories. The Ventana HER2 dual-color ISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national and international guidelines for performance of ISH-based diagnostic tests.

Gene protein detection platform--a comparison of a new human epidermal growth factor receptor 2 assay with conventional immunohistochemistry and fluorescence in situ hybridization platforms.

Science.gov (United States)

Stålhammar, Gustav; Farrajota, Pedro; Olsson, Ann; Silva, Cristina; Hartman, Johan; Elmberger, Göran

2015-08-01

Human epidermal growth factor receptor 2 (HER2) immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) are widely used semiquantitative assays for selecting breast cancer patients for HER2 antibody therapy. However, both techniques have been shown to have disadvantages. Our aim was to test a recent automated technique of combined IHC and brightfield dual in situ hybridization-gene protein detection platform (GPDP)-in breast cancer HER2 protein, gene, and chromosome 17 centromere status evaluations, comparing the results in accordance to the American Society of Clinical Oncology/College of American Pathologists recommendations for HER2 testing in breast cancer from both 2007 and 2013. The GPDP technique performance was evaluated on 52 consecutive whole slide invasive breast cancer cases with HER2 IHC 2/3+ scoring results. Applying in turns the American Society of Clinical Oncology/College of American Pathologists recommendations for HER2 testing in breast cancer from 2007 and 2013 to both FISH and GPDP DISH assays, the HER2 gene amplification results showed 100% concordance among amplified/nonamplified cases, but there was a shift in 4 cases toward positive from equivocal results and toward equivocal from negative results. This might be related to the emphasis on the average HER2 copy number in the 2013 criteria. HER2 expression by IVD market IHC kit (Pathway®) has a strong correlation with GPDP HER2 protein, including a full concordance for all cases scored as 3+ and a reduction from 2+ to 1+ in 7 cases corresponding to nonamplified cases. Gene protein detection platform HER2 protein "solo" could have spared the need for 7 FISH studies. In addition, the platform offered advantages on interpretation reassurance including selecting areas for counting gene signals paralleled with protein IHC expression, on heterogeneity detection, interpretation time, technical time, and tissue expense. Copyright © 2015 Elsevier Inc. All rights reserved.

Optimal Threshold for a Positive Hybrid Capture 2 Test for Detection of Human Papillomavirus: Data from the ARTISTIC Trial▿

Science.gov (United States)

Sargent, A.; Bailey, A.; Turner, A.; Almonte, M.; Gilham, C.; Baysson, H.; Peto, J.; Roberts, C.; Thomson, C.; Desai, M.; Mather, J.; Kitchener, H.

2010-01-01

We present data on the use of the Hybrid Capture 2 (HC2) test for the detection of high-risk human papillomavirus (HR HPV) with different thresholds for positivity within a primary screening setting and as a method of triage for low-grade cytology. In the ARTISTIC population-based trial, 18,386 women were screened by cytology and for HPV. Cervical intraepithelial neoplasia lesions of grade two and higher (CIN2+ lesions) were identified for 453 women within 30 months of an abnormal baseline sample. When a relative light unit/cutoff (RLU/Co) ratio of ≥1 was used as the threshold for considering an HC2 result positive, 15.6% of results were positive, and the proportion of CIN2+ lesions in this group was 14.7%. The relative sensitivity for CIN2+ lesion detection was 93.4%. When an RLU/Co ratio of ≥2 was used as the threshold, there was a 2.5% reduction in positivity, with an increase in the proportion of CIN2+ lesions detected. The relative sensitivity decreased slightly, to 90.3%. Among women with low-grade cytology, HPV prevalences were 43.7% and 40.3% at RLU/Co ratios of ≥1 and ≥2, respectively. The proportions of CIN2+ lesions detected were 17.3% and 18.0%, with relative sensitivities of 87.7% at an RLU/Co ratio of ≥1 and 84.2% at an RLU/Co ratio of ≥2. At an RLU/Co ratio of ≥1, 68.3% of HC2-positive results were confirmed by the Roche line blot assay, compared to 77.2% of those at an RLU/Co ratio of ≥2. Fewer HC2-positive results were confirmed for 35- to 64-year-olds (50.3% at an RLU/Co ratio of ≥1 and 63.2% at an RLU/Co ratio of >2) than for 20- to 34-year-olds (78.7% at an RLU/Co ratio of ≥1 and 83.7% at an RLU/Co ratio of >2). If the HC2 test is used for routine screening as an initial test or as a method of triage for low-grade cytology, we would suggest increasing the threshold for positivity from the RLU/Co ratio of ≥1, recommended by the manufacturer, to an RLU/Co ratio of ≥2, since this study has shown that a beneficial balance

Establishment of hybridized focus measure functions as a universal method for autofocusing

Science.gov (United States)

Shah, Mohammad Imran; Mishra, Smriti; Rout, Chittaranjan

2017-12-01

Exact focusing is essential for any automatic image capturing system. Performances of focus measure functions (FMFs) used for autofocusing are sensitive to image contents and imaging systems. Therefore, identification of universal FMF assumes a lot of significance. Eight FMFs were hybridized in pairs of two and implemented simultaneously on a single stack to calculate the hybrid focus measure. In total, 28 hybrid FMFs (HFMFs) and eight FMFs were implemented on stacks of images from three different imaging modalities. Performance of FMFs was found to the best at 50% region sampling. Accuracy, focus error, and false maxima were calculated to evaluate the performance of each FMF. Nineteen HFMFs provided >90% accuracy. Image distortion (noise, contrast, saturation, illumination, etc.) was performed to evaluate robustness of HFMFs. Hybrid of tenengrad variance and steerable filter-based (VGRnSFB) FMFs was identified as the most robust and accurate function with an accuracy of ≥90% and a relatively lower focus error and false maxima rate. Sharpness of focus curve of VGRnSFB along with eight individual FMFs was also computed to determine the efficacy of HFMF for the optimization process. VGRnSFB HFMF may be implemented for automated capturing of an image for any imaging system.

Evaluation of PCR and DNA hybridization protocols for detection of viable enterotoxigenic Clostridium perfringens in irradiated beef

International Nuclear Information System (INIS)

Baez, L.A.; Juneja, V.K.; Thayer, D.W.; Sackitey, S.

1997-01-01

The sensitivity of DNA hybridization and polymerase chain reaction (PCR), was evaluated in irradiated cooked and raw beef samples. A membrane-based colony hybridization assay and a PCR protocol, both with specificity for the enterotoxin A gene of Clostridium perfringens, were compared with viable plate counts. The results of the colony hybridization procedure were in agreement with viable plate counts for detection and enumeration of enterotoxigenic C. perfringens. The PCR procedure combined a 4 h enrichment followed by a nucleic acid extraction step and assessed the amplification of 183 and 750 base pair enterotoxin gene targets. Detection of C. perfringens by PCR did not show a reliable correlation with viable plate counts or the colony hybridization assay. C. perfringens killed by irradiation were not detected by the plate count or colony hybridization methods; however, killed cells were detected with the PCR technique. By relying on the growth of viable cells for detection and/or enumeration, the colony hybridization and plate count methods provided a direct correlation with the presence of viable bacteria

An efficient algorithm for the stochastic simulation of the hybridization of DNA to microarrays

Directory of Open Access Journals (Sweden)

Laurenzi Ian J

2009-12-01

Full Text Available Abstract Background Although oligonucleotide microarray technology is ubiquitous in genomic research, reproducibility and standardization of expression measurements still concern many researchers. Cross-hybridization between microarray probes and non-target ssDNA has been implicated as a primary factor in sensitivity and selectivity loss. Since hybridization is a chemical process, it may be modeled at a population-level using a combination of material balance equations and thermodynamics. However, the hybridization reaction network may be exceptionally large for commercial arrays, which often possess at least one reporter per transcript. Quantification of the kinetics and equilibrium of exceptionally large chemical systems of this type is numerically infeasible with customary approaches. Results In this paper, we present a robust and computationally efficient algorithm for the simulation of hybridization processes underlying microarray assays. Our method may be utilized to identify the extent to which nucleic acid targets (e.g. cDNA will cross-hybridize with probes, and by extension, characterize probe robustnessusing the information specified by MAGE-TAB. Using this algorithm, we characterize cross-hybridization in a modified commercial microarray assay. Conclusions By integrating stochastic simulation with thermodynamic prediction tools for DNA hybridization, one may robustly and rapidly characterize of the selectivity of a proposed microarray design at the probe and "system" levels. Our code is available at http://www.laurenzi.net.

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

Science.gov (United States)

Algar, W Russ; Krull, Ulrich J

2009-01-06

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

Hybrid Predictive Control for Dynamic Transport Problems

CERN Document Server

Núñez, Alfredo A; Cortés, Cristián E

2013-01-01

Hybrid Predictive Control for Dynamic Transport Problems develops methods for the design of predictive control strategies for nonlinear-dynamic hybrid discrete-/continuous-variable systems. The methodology is designed for real-time applications, particularly the study of dynamic transport systems. Operational and service policies are considered, as well as cost reduction. The control structure is based on a sound definition of the key variables and their evolution. A flexible objective function able to capture the predictive behaviour of the system variables is described. Coupled with efficient algorithms, mainly drawn from the area of computational intelligence, this is shown to optimize performance indices for real-time applications. The framework of the proposed predictive control methodology is generic and, being able to solve nonlinear mixed-integer optimization problems dynamically, is readily extendable to other industrial processes. The main topics of this book are: ●hybrid predictive control (HPC) ...

Enzyme activity assay of glycoprotein enzymes based on a boronate affinity molecularly imprinted 96-well microplate.

Science.gov (United States)

Bi, Xiaodong; Liu, Zhen

2014-12-16

Enzyme activity assay is an important method in clinical diagnostics. However, conventional enzyme activity assay suffers from apparent interference from the sample matrix. Herein, we present a new format of enzyme activity assay that can effectively eliminate the effects of the sample matrix. The key is a 96-well microplate modified with molecularly imprinted polymer (MIP) prepared according to a newly proposed method called boronate affinity-based oriented surface imprinting. Alkaline phosphatase (ALP), a glycoprotein enzyme that has been routinely used as an indicator for several diseases in clinical tests, was taken as a representative target enzyme. The prepared MIP exhibited strong affinity toward the template enzyme (with a dissociation constant of 10(-10) M) as well as superb tolerance for interference. Thus, the enzyme molecules in a complicated sample matrix could be specifically captured and cleaned up for enzyme activity assay, which eliminated the interference from the sample matrix. On the other hand, because the boronate affinity MIP could well retain the enzymatic activity of glycoprotein enzymes, the enzyme captured by the MIP was directly used for activity assay. Thus, additional assay time and possible enzyme or activity loss due to an enzyme release step required by other methods were avoided. Assay of ALP in human serum was successfully demonstrated, suggesting a promising prospect of the proposed method in real-world applications.

Multiplex real-time PCR assay for Legionella species.

Science.gov (United States)

Kim, Seung Min; Jeong, Yoojung; Sohn, Jang Wook; Kim, Min Ja

2015-12-01

Legionella pneumophila serogroup 1 (sg1) accounts for the majority of infections in humans, but other Legionella species are also associated with human disease. In this study, a new SYBR Green I-based multiplex real-time PCR assay in a single reaction was developed to allow the rapid detection and differentiation of Legionella species by targeting specific gene sequences. Candidate target genes were selected, and primer sets were designed by referring to comparative genomic hybridization data of Legionella species. The Legionella species-specific groES primer set successfully detected all 30 Legionella strains tested. The xcpX and rfbA primers specifically detected L. pneumophila sg1-15 and L. pneumophila sg1, respectively. In addition, this assay was validated by testing clinical samples and isolates. In conclusion, this novel multiplex real-time PCR assay might be a useful diagnostic tool for the rapid detection and differentiation of Legionella species in both clinical and epidemiological studies. Copyright © 2015 Elsevier Ltd. All rights reserved.

Hybrid Optimization for Wind Turbine Thick Airfoils

Energy Technology Data Exchange (ETDEWEB)

Grasso, F. [ECN Wind Energy, Petten (Netherlands)

2012-06-15

One important element in aerodynamic design of wind turbines is the use of specially tailored airfoils to increase the ratio of energy capture and reduce cost of energy. This work is focused on the design of thick airfoils for wind turbines by using numerical optimization. A hybrid scheme is proposed in which genetic and gradient based algorithms are combined together to improve the accuracy and the reliability of the design. Firstly, the requirements and the constraints for this class of airfoils are described; then, the hybrid approach is presented. The final part of this work is dedicated to illustrate a numerical example regarding the design of a new thick airfoil. The results are discussed and compared to existing airfoils.

Evaluation of the BeTha gene 1 kit for the qualitative detection of the eight most common Mediterranean beta-thalassemia mutations.

Science.gov (United States)

Ugozzoli, L A; Lowery, J D; Reyes, A A; Lin, C I; Re, A; Locati, F; Galanello, R; Macioni, L; Maggio, A; Giambona, A; Loutradi, A; Boussiou, M; Wallace, R B

1998-11-01

We describe the evaluation of the Bio-Rad BeTha Gene 1 kit (Bio-Rad Laboratories, Hercules, CA), a DNA-probe assay designed for the qualitative determination of the eight most common Mediterranean beta-thalassemia mutations. The kit utilizes the principle of allele-specific oligonucleotide (ASO) hybridization. Following sample preparation and in vitro DNA amplification by the polymerase chain reaction (PCR), an allele-specific detection of the amplified products by a nonradioactive enzymatic assay is performed. Genomic DNA is prepared from an individual's whole blood with a DNA purification matrix. In a second step, the beta-globin gene is amplified in a multiplex PCR reaction containing four 5' biotinylated oligonucleotide primers. In a final step, an aliquot of the PCR reaction is first chemically denatured and then captured in two eight-well strips of a 96-well enzyme-linked immunosorbent assay (ELISA) plate by hybridization to an immobilized ASO probe. Each DNA sequence at each of the eight mutation sites is represented by one normal and one mutant ASO. During this capture/hybridization step, which is performed at 37 degrees C, only perfectly matched PCR products will be captured by an ASO. Subsequently, the allele-specific captured biotin-labeled PCR products are detected by a colorimetric enzymatic reaction. The system permits the detection of 16 beta-thalassemia alleles using a high-throughput format that can be automated easily. A clinical feasibility study was performed to evaluate the functionality (method comparison study, assay validity using samples previously collected and stored at various temperatures for different periods of time, interference on kit performance, and assay validity for prenatal diagnosis) and the usability (ease of use, sample throughput) of the kit. The analysis of 110 samples previously studied with reference methods showed 100% clinical sensitivity and specificity. We demonstrate here that the procedure not only increases the

Hydrogen fluoride capture by imidazolium acetate ionic liquid

Science.gov (United States)

Chaban, Vitaly

2015-04-01

Extraction of hydrofluoric acid (HF) from oils is a drastically important problem in petroleum industry, since HF causes quick corrosion of pipe lines and brings severe health problems to humanity. Some ionic liquids (ILs) constitute promising scavenger agents thanks to strong binding to polar compounds and tunability. PM7-MD simulations and hybrid density functional theory are employed here to consider HF capture ability of ILs. Discussing the effects and impacts of the cation and the anion separately and together, we evaluate performance of imidazolium acetate and outline systematic search guidelines for efficient adsorption and extraction of HF.

Membrane Separation Processes for Post-Combustion Carbon Dioxide Capture: State of the Art and Critical Overview

Directory of Open Access Journals (Sweden)

Belaissaoui Bouchra

2014-11-01

Full Text Available Membrane processes have been initially seldom considered within a post-combustion carbon dioxide capture framework. More traditional processes, particularly gas-liquid absorption in chemical solvents, are often considered as the most appropriate solution for the first generation of technologies. In this paper, a critical state of the art of gas separation membranes for CO2 capture is proposed. In a first step, the key performances (selectivity, permeability of different membrane materials such as polymers, inorganic membranes, hybrid matrices and liquid membranes, including recently reported results, are reviewed. In a second step, the process design characteristics of a single stage membrane unit are studied. Purity and energy constraints are analysed as a function of operating conditions and membrane materials performances. The interest of multistage and hybrid systems, two domains which have not sufficiently investigated up to now, are finally discussed. The importance of technico-economical analyses is highlighted in order to better estimate the optimal role of membranes for CCS applications.

Validation of an immunodiagnostic assay for detection of 13 Streptococcus pneumoniae serotype-specific polysaccharides in human urine.

Science.gov (United States)

Pride, Michael W; Huijts, Susanne M; Wu, Kangjian; Souza, Victor; Passador, Sherry; Tinder, Chunyan; Song, Esther; Elfassy, Arik; McNeil, Lisa; Menton, Ronald; French, Roger; Callahan, Janice; Webber, Chris; Gruber, William C; Bonten, Marc J M; Jansen, Kathrin U

2012-08-01

To improve the clinical diagnosis of pneumococcal infection in bacteremic and nonbacteremic community-acquired pneumonia (CAP), a Luminex technology-based multiplex urinary antigen detection (UAD) diagnostic assay was developed and validated. The UAD assay can simultaneously detect 13 different serotypes of Streptococcus pneumoniae by capturing serotype-specific S. pneumoniae polysaccharides (PnPSs) secreted in human urine. Assay specificity is achieved by capturing the polysaccharides with serotype-specific monoclonal antibodies (MAbs) on spectrally unique microspheres. Positivity for each serotype was based on positivity cutoff values calculated from a standard curve run on each assay plate together with positive- and negative-control urine samples. The assay is highly specific, since significant signals are detected only when each PnPS was paired with its homologous MAb-coated microspheres. Validation experiments demonstrated excellent accuracy and precision. The UAD assay and corresponding positivity cutoff values were clinically validated by assessing 776 urine specimens obtained from patients with X-ray-confirmed CAP. The UAD assay demonstrated 97% sensitivity and 100% specificity using samples obtained from patients with bacteremic, blood culture-positive CAP. Importantly, the UAD assay identified Streptococcus pneumoniae (13 serotypes) in a proportion of individuals with nonbacteremic CAP, a patient population for which the pneumococcal etiology of CAP was previously difficult to assess. Therefore, the UAD assay provides a specific, noninvasive, sensitive, and reproducible tool to support vaccine efficacy as well as epidemiological evaluation of pneumococcal disease, including CAP, in adults.

Hybrid approaches to nanometer-scale patterning: Exploiting tailored intermolecular interactions

International Nuclear Information System (INIS)

Mullen, Thomas J.; Srinivasan, Charan; Shuster, Mitchell J.; Horn, Mark W.; Andrews, Anne M.; Weiss, Paul S.

2008-01-01

In this perspective, we explore hybrid approaches to nanometer-scale patterning, where the precision of molecular self-assembly is combined with the sophistication and fidelity of lithography. Two areas - improving existing lithographic techniques through self-assembly and fabricating chemically patterned surfaces - will be discussed in terms of their advantages, limitations, applications, and future outlook. The creation of such chemical patterns enables new capabilities, including the assembly of biospecific surfaces to be recognized by, and to capture analytes from, complex mixtures. Finally, we speculate on the potential impact and upcoming challenges of these hybrid strategies.

Automated 5 ' nuclease assay for detection of virulence factors in porcine Escherichia coli

DEFF Research Database (Denmark)

Frydendahl, K.; Imberechts, H.; Lehmann, S.

2001-01-01

(STa, STb, EAST1) and heat labile LT) enterotoxins and the verocytotoxin variant 2e (VT2e). To correctly identify false negative results, an endogenous internal control targeting the E. coil 16S rRNA gene was incorporated in each test tube. The assay was evaluated using a collection of E. coil...... reference strains which have previously been examined with phenotypical assays or DNA hybridization. Furthermore, the assay was evaluated by testing porcine E. coil field strains, previously characterized. The 5' nuclease assay correctly detected the presence of virulence genes in all reference strains....... When testing field strains there was generally excellent agreement with results obtained by laboratories in Belgium and Germany. In conclusion, the 5' nuclease assay developed is a fast and specific tool for detection of E. coli virulence genes in the veterinary diagnostic laboratory....

Spacecraft Hybrid (Mixed-Actuator) Attitude Control Experiences on NASA Science Missions

Science.gov (United States)

Dennehy, Cornelius J.

2014-01-01

There is a heightened interest within NASA for the design, development, and flight implementation of mixed-actuator hybrid attitude control systems for science spacecraft that have less than three functional reaction wheel actuators. This interest is driven by a number of recent reaction wheel failures on aging, but what could be still scientifically productive, NASA spacecraft if a successful hybrid attitude control mode can be implemented. Over the years, hybrid (mixed-actuator) control has been employed for contingency attitude control purposes on several NASA science mission spacecraft. This paper provides a historical perspective of NASA's previous engineering work on spacecraft mixed-actuator hybrid control approaches. An update of the current situation will also be provided emphasizing why NASA is now so interested in hybrid control. The results of the NASA Spacecraft Hybrid Attitude Control Workshop, held in April of 2013, will be highlighted. In particular, the lessons learned captured from that workshop will be shared in this paper. An update on the most recent experiences with hybrid control on the Kepler spacecraft will also be provided. This paper will close with some future considerations for hybrid spacecraft control.

[Effectiveness of FTA Elute® indicating cartridge in combination with hybrid capture 2 for cervical cancer screening].

Science.gov (United States)

Chen, Feng; Zhang, Xi; Wang, Shaoming; Hu, Shangying; Chen, Wen; Zhao, Fanghui; He, Wei; Zhang, Yuqing; Qiao, Youlin

2015-02-01

To evaluate the effectiveness of FTA Elute® Cartridge (GE healthcare, Kent, UK) in combination with hybrid capture 2 (HC2) testing for cervical cancer screening. From May to June 2012, 412 women aged 25 to 65 years in Jiangxi Tonggu were enrolled in the study. We used pathological outcome as the gold standard, and the accuracy of the FTA card in combination with HC2 testing was investigated from both physician- and self-sampling, respectively. Physician sampling using the FTA card in combination with HC2 testing showed a comparable sensitivity (12/13) with the liquid based medium, but a higher specificity 69.5% (266/383) vs (77.8%, 298/383) (P FTA card in combination with HC2 testing with liquid based medium was 10/13 vs 8/13(P = 0.625) and (62.3%, 238/382) vs (75.7%, 289/382) (P FTA and liquid-based sampling medium was 86.1% (340/395) and 79.5% (314/395). For physician-collected samples used for HC2 testing to detect CIN2+, the accuracy of the FTA card was superior to that of the liquid-based medium (area under the receiver operating characteristic curve (AUC) = 0.898, 95%CI:0.838-0.958). FTA Elute® cartridge in combination with HC2 testing is a promising method of specimen transport for cervical cancer screening programs with a good precision.With further optimization, it could become an effective method for cervical cancer screening in various economic levels of areas.

BioAssay templates for the semantic web

Directory of Open Access Journals (Sweden)

Alex M. Clark

2016-05-01

Full Text Available Annotation of bioassay protocols using semantic web vocabulary is a way to make experiment descriptions machine-readable. Protocols are communicated using concise scientific English, which precludes most kinds of analysis by software algorithms. Given the availability of a sufficiently expressive ontology, some or all of the pertinent information can be captured by asserting a series of facts, expressed as semantic web triples (subject, predicate, object. With appropriate annotation, assays can be searched, clustered, tagged and evaluated in a multitude of ways, analogous to other segments of drug discovery informatics. The BioAssay Ontology (BAO has been previously designed for this express purpose, and provides a layered hierarchy of meaningful terms which can be linked to. Currently the biggest challenge is the issue of content creation: scientists cannot be expected to use the BAO effectively without having access to software tools that make it straightforward to use the vocabulary in a canonical way. We have sought to remove this barrier by: (1 defining a BioAssay Template (BAT data model; (2 creating a software tool for experts to create or modify templates to suit their needs; and (3 designing a common assay template (CAT to leverage the most value from the BAO terms. The CAT was carefully assembled by biologists in order to find a balance between the maximum amount of information captured vs. low degrees of freedom in order to keep the user experience as simple as possible. The data format that we use for describing templates and corresponding annotations is the native format of the semantic web (RDF triples, and we demonstrate some of the ways that generated content can be meaningfully queried using the SPARQL language. We have made all of these materials available as open source (http://github.com/cdd/bioassay-template, in order to encourage community input and use within diverse projects, including but not limited to our own

Selective functional activity measurement of a PEGylated protein with a modification-dependent activity assay.

Science.gov (United States)

Weber, Alfred; Engelmaier, Andrea; Mohr, Gabriele; Haindl, Sonja; Schwarz, Hans Peter; Turecek, Peter L

2017-01-05

BAX 855 (ADYNOVATE) is a PEGylated recombinant factor VIII (rFVIII) that showed prolonged circulatory half-life compared to unmodified rFVIII in hemophilic patients. Here, the development and validation of a novel assay is described that selectively measures the activity of BAX 855 as cofactor for the serine protease factor IX, which actives factor X. This method type, termed modification-dependent activity assay, is based on PEG-specific capture of BAX 855 by an anti-PEG IgG preparation, followed by a chromogenic FVIII activity assay. The assay principle enabled sensitive measurement of the FVIII cofactor activity of BAX 855 down to the pM-range without interference by non-PEGylated FVIII. The selectivity of the capture step, shown by competition studies to primarily target the terminal methoxy group of PEG, also allowed assessment of the intactness of the attached PEG chains. Altogether, the modification-dependent activity not only enriches, but complements the group of methods to selectively, accurately, and precisely measure a PEGylated drug in complex biological matrices. In contrast to all other methods described so far, it allows measurement of the biological activity of the PEGylated protein. Data obtained demonstrate that this new method principle can be extended to protein modifications other than PEGylation and to a variety of functional activity assays. Copyright © 2016 Elsevier B.V. All rights reserved.

Modeling and control of a hybrid wind-tidal turbine with hydraulic accumulator

International Nuclear Information System (INIS)

Fan, YaJun; Mu, AnLe; Ma, Tao

2016-01-01

This paper presents the modeling and control of a hybrid wind-tidal turbine with hydraulic accumulator. The hybrid turbine captures the offshore wind energy and tidal current energy simultaneously and stores the excess energy in hydraulic accumulator prior to electricity generation. Two hydraulic pumps installed respectively in wind and tidal turbine nacelles are used to transform the captured mechanical energy into hydraulic energy. To extract the maximal power from wind and tidal current, standard torque controls are achieved by regulating the displacements of the hydraulic pumps. To meet the output power demand, a Proportion Integration Differentiation (PID) controller is designed to distribute the hydraulic energy between the accumulator and the Pelton turbine. A simulation case study based on combining a 5 MW offshore wind turbine and a 1 MW tidal current turbine is undertaken. Case study demonstrates that the hybrid generation system not only captures all the available wind and tidal energy and also delivers the desired generator power precisely through the accumulator damping out all the power fluctuations from the wind and tidal speed disturbances. Energy and exergy analyses show that the energy efficiency can exceed 100% as the small input speeds are considered, and the exergy efficiency has the consistent change trends with demand power. Further more parametric sensitivity study on hydraulic accumulator shows that there is an inversely proportional relationship between accumulator and hydraulic equipments including the pump and nozzle in terms of dimensions. - Highlights: • A hybrid wind-tidal turbine is presented. • Hydraulic accumulator stores/releases the surplus energy. • Standard torque controls extract the maximal power from wind and tidal. • Generator outputs meet the electricity demand precisely. • Parametric sensitivity study on accumulator is implemented.

Performance comparison of two low-CO2 emission solar/methanol hybrid combined cycle power systems

International Nuclear Information System (INIS)

Li, Yuanyuan; Zhang, Na; Lior, Noam

2015-01-01

Highlights: • Two novel solar hybrid combined cycle systems have been proposed and analyzed. • The power systems integrate solar-driven thermo-chemical conversion and CO 2 capture. • Exergy efficiency of about 55% and specific CO 2 emissions of 34 g/kW h are predicted. • Systems CO 2 emissions are 36.8% lower compared to a combined cycle with CO 2 capture. • The fossil fuel demand is ∼30% lower with a solar share of ∼20%. - Abstract: Two novel hybrid combined cycle power systems that use solar heat and methanol, and integrate CO 2 capture, are proposed and analyzed, one based on solar-driven methanol decomposition and the other on solar-driven methanol reforming. The high methanol conversion rates at relatively low temperatures offer the advantage of using the solar heat at only 200–300 °C to drive the syngas production by endothermic methanol conversions and its conversion to chemical energy. Pre-combustion decarbonization is employed to produce CO 2 -free fuel from the fully converted syngas, which is then burned to produce heat at the high temperature for power generation in the proposed advanced combined cycle systems. To improve efficiency, the systems’ configurations were based on the principle of cascade use of multiple heat sources of different temperatures. The thermodynamic performance of the hybrid power systems at its design point is simulated and evaluated. The results show that the hybrid systems can attain an exergy efficiency of about 55%, and specific CO 2 emissions as low as 34 g/kW h. Compared to a gas/steam combined cycle with flue gas CO 2 capture, the proposed solar-assisted system CO 2 emissions are 36.8% lower, and a fossil fuel saving ratio of ∼30% is achievable with a solar thermal share of ∼20%. The system integration predicts high efficiency conversion of solar heat and low-energy-penalty CO 2 capture, with the additional advantage that solar heat is at relatively low temperature where its collection is cheaper and

Translating tumor biology into personalized treatment planning: analytical performance characteristics of the Oncotype DX? Colon Cancer Assay

OpenAIRE

Clark-Langone, Kim M; Sangli, Chithra; Krishnakumar, Jayadevi; Watson, Drew

2010-01-01

Abstract Background The Oncotype DX® Colon Cancer Assay is a new diagnostic test for determining the likelihood of recurrence in stage II colon cancer patients after surgical resection using fixed paraffin embedded (FPE) primary colon tumor tissue. Like the Oncotype DX Breast Cancer Assay, this is a high complexity, multi-analyte, reverse transcription (RT) polymerase chain reaction (PCR) assay that measures the expression levels of specific cancer-related genes. By capturing the biology unde...

Expression of endogenous proteins in maize hybrids in a multi-location field trial in India.

Science.gov (United States)

Gutha, Linga R; Purushottam, Divakar; Veeramachaneni, Aruna; Tigulla, Sarita; Kodappully, Vikas; Enjala, Chandana; Rajput, Hitendrasinh; Anderson, Jennifer; Hong, Bonnie; Schmidt, Jean; Bagga, Shveta

2018-05-17

Genetically modified (GM) crops undergo large scale multi-location field trials to characterize agronomics, composition, and the concentration of newly expressed protein(s) [herein referred to as transgenic protein(s)]. The concentration of transgenic proteins in different plant tissues and across the developmental stages of the plant is considered in the safety assessment of GM crops. Reference or housekeeping proteins are expected to maintain a relatively stable expression pattern in healthy plants given their role in cellular functions. Understanding the effects of genotype, growth stage and location on the concentration of endogenous housekeeping proteins may provide insight into the contribution these factors could have on transgenic protein concentrations in GM crops. The concentrations of three endogenous proteins (actin, elongation factor 1-alpha, and glyceraldehyde 3-phosphate dehydrogenase) were measured in several different maize hybrids grown across multiple field locations over 2 years. Leaf samples were collected from healthy plants at three developmental stages across the growing seasons, and protein concentrations were quantified by indirect enzyme-linked immunosorbent assay (ELISA) for each protein. In general, the concentrations of these three endogenous proteins were relatively consistent across hybrid backgrounds, when compared within one growth stage and location (2-26%CV), whereas the concentrations of proteins in the same hybrid and growth stage across different locations were more variable (12-64%CV). In general, the protein concentrations in 2013 and 2014 show similar trends in variability. Some degree of variability in protein concentrations should be expected for both transgenic and endogenous plant-expressed proteins. In the case of GM crops, the potential variation in protein concentrations due to location effects is captured in the current model of multi-location field testing.

Electrochemical DNA sandwich assay with a lipase label for attomole detection of DNA

DEFF Research Database (Denmark)

Ferapontova, Elena; Hansen, Majken Nørgaard; Saunders, Aaron Marc

2010-01-01

A fast and sensitive electrochemical lipase-based sandwich hybridization assay for detection of attomole levels of DNA has been developed. A combination of magnetic beads, used for pre-concentration and bioseparation of the analyte with a lipase catalyst label allowed detection of DNA with a limi...

Hybrid nuclear reactors and muon catalysis

International Nuclear Information System (INIS)

Petrov, Yu.

1983-01-01

Three methods are described of the conversion of isotope 238 U to 239 Pu by neutron capture in fast breeder reactors, in the breeding blanket of hybrid thermonuclear reactors using neutrons generated by fusion and electronuclear breeding in which the target is bombarded with 1 GeV protons. Their possible use in power production is discussed. Another prospective energy source is the use of muon catalysis in the fusion of deuterium and tritium nuclei. (J.P.)

A multiplex branched DNA assay for parallel quantitative gene expression profiling.

Science.gov (United States)

Flagella, Michael; Bui, Son; Zheng, Zhi; Nguyen, Cung Tuong; Zhang, Aiguo; Pastor, Larry; Ma, Yunqing; Yang, Wen; Crawford, Kimberly L; McMaster, Gary K; Witney, Frank; Luo, Yuling

2006-05-01

We describe a novel method to quantitatively measure messenger RNA (mRNA) expression of multiple genes directly from crude cell lysates and tissue homogenates without the need for RNA purification or target amplification. The multiplex branched DNA (bDNA) assay adapts the bDNA technology to the Luminex fluorescent bead-based platform through the use of cooperative hybridization, which ensures an exceptionally high degree of assay specificity. Using in vitro transcribed RNA as reference standards, we demonstrated that the assay is highly specific, with cross-reactivity less than 0.2%. We also determined that the assay detection sensitivity is 25,000 RNA transcripts with intra- and interplate coefficients of variance of less than 10% and less than 15%, respectively. Using three 10-gene panels designed to measure proinflammatory and apoptosis responses, we demonstrated sensitive and specific multiplex gene expression profiling directly from cell lysates. The gene expression change data demonstrate a high correlation coefficient (R(2)=0.94) compared with measurements obtained using the single-plex bDNA assay. Thus, the multiplex bDNA assay provides a powerful means to quantify the gene expression profile of a defined set of target genes in large sample populations.

A new gaseous imaging detector for the assay of lymphocyte cultures

International Nuclear Information System (INIS)

Bateman, J.E.; Joyce, A.; Knight, S.C.; Bedford, P.

1991-01-01

Tritium-labelled cell cultures used in studies of lymphocyte proliferation at the Clinical Research Centre are blotted in arrays of 10x6 spots spaced at 6 mm. An imaging detector based on the differential induction signals produced at a central amplifying electrode has been developed for the imaging and assay of these blots. A spatial resolution ≅ 2.5 mm FWHM attained over the aperture of 60 mmx36mm enables the individual spots to be reliably counted. Data is captured in a PC/AT at rates which permit an assay to be completed in typically 30-60 min. The simplicity of both the detector and the readout electronics leads to a low cost system. Images and assay results are presented. (orig.)

Restriction Cascade Exponential Amplification (RCEA) assay with an attomolar detection limit: a novel, highly specific, isothermal alternative to qPCR.

Science.gov (United States)

Ghindilis, Andrey L; Smith, Maria W; Simon, Holly M; Seoudi, Ihab A; Yazvenko, Nina S; Murray, Iain A; Fu, Xiaoqing; Smith, Kenneth; Jen-Jacobson, Linda; Xu, Shuang-Yong

2015-01-13

An alternative to qPCR was developed for nucleic acid assays, involving signal rather than target amplification. The new technology, Restriction Cascade Exponential Amplification (RCEA), relies on specific cleavage of probe-target hybrids by restriction endonucleases (REase). Two mutant REases for amplification (Ramp), S17C BamHI and K249C EcoRI, were conjugated to oligonucleotides, and immobilized on a solid surface. The signal generation was based on: (i) hybridization of a target DNA to a Ramp-oligonucleotide probe conjugate, followed by (ii) specific cleavage of the probe-target hybrid using a non-immobilized recognition REase. The amount of Ramp released into solution upon cleavage was proportionate to the DNA target amount. Signal amplification was achieved through catalysis, by the free Ramp, of a restriction cascade containing additional oligonucleotide-conjugated Ramp and horseradish peroxidase (HRP). Colorimetric quantification of free HRP indicated that the RCEA achieved a detection limit of 10 aM (10(-17) M) target concentration, or approximately 200 molecules, comparable to the sensitivity of qPCR-based assays. The RCEA assay had high specificity, it was insensitive to non-specific binding, and detected target sequences in the presence of foreign DNA. RCEA is an inexpensive isothermal assay that allows coupling of the restriction cascade signal amplification with any DNA target of interest.

Advances in hybrid optics physical sensors for extreme environments

Science.gov (United States)

Riza, Nabeel A.

2010-04-01

Highlighted are novel innovations in hybrid optical design physical sensors for extreme environments. Various hybrid design compositions are proposed that are suited for a particular sensor application. Examples includes combining freespace (wireless) and fiber-optics (wired) for gas turbine sensing and combining single crystal and sintered Silicon Carbide (SiC) materials for robust extreme environment Coefficent of Thermal Expansion (CTE) matched frontend probe design. Sensor signal processing also includes the hybrid theme where for example Black-Body radiation thermometry (pyrometry) is combined with laser interferometry to provide extreme temperature measurements. The hybrid theme also operates on the optical device level where a digital optical device such as a Digital Micromirror Device (DMD) is combined with an analog optical device such as an Electronically Controlled Variable Focal Length Lens (ECVFL) to deliver a smart and compressive Three Dimensional (3-D) imaging sensor for remote scene and object shape capture including both ambient light (passive) mode and active laser targeting and receive processing. Within a device level, the hybrid theme also operates via combined analog and digital control such as within a wavelength-coded variable optical delay line. These powerful hybrid design optical sensors have numerous applications in engineering and science applications from the military to the commercial/industrial sectors.

Destination bonding: Hybrid cognition using Instagram

Directory of Open Access Journals (Sweden)

Arup Kumar Baksi

2015-01-01

Full Text Available Empirical research has identified the phenomenon of destination bonding as a result of summated physical and emotional values associated with the destination. Physical values, namely natural landscape & other physical settings and emotional values, namely the enculturation processes, have a significant role to play in portraying visitors’ cognitive framework for destination preference. The physical values seemed to be the stimulator for bonding that embodies action or behavior tendencies in imagery. The emotional values were the conditions that lead to affective bonding and are reflected in attitudes for a place which were evident in text narratives. Social networking on virtual platforms offers the scope for hybrid cognitive expression using imagery and text to the visitors. Instagram has emerged as an application-window to capture these hybrid cognitions of visitors. This study focuses on assessing the relationship between hybrid cognition of visitors expressed via Instagram and their bond with the destination. Further to this, the study attempts to examine the impact of hybrid cognition of visitors on the behavioral pattern of prospective visitors to the destination. The study revealed that sharing of visual imageries and related text by the visitors is an expression of the physico-emotional bonding with the destination. It was further established that hybrid cognition strongly asserts destination bonding and has been also found to have moderating impact on the link between destination bonding and electronic-word-of-mouth.

The narrow therapeutic window of glycated hemoglobin and assay variability.

Science.gov (United States)

Hosseini, S S; Bibler, I; Charles, M A

1999-12-01

Glycated hemoglobin is measured by a variety of assays, each of which has a unique normal level. Our purpose is to show that among the different assays available in the United States, using the same patient's blood sample, assay results may vary widely and may more or less easily achieve a glycated hemoglobin value within the normal range. The following assays were compared using the same patient's blood sample for each pair of assays: glycohemoglobin affinity assay (GHB Reader; Isolab, Akron, OH) versus gel electrophoresis assay (n = 76); Isolab versus ion capture assay (IMX; Abbott Laboratories, Irving, TX) (n = 57); monoclonal antibody assay (DCA2000; Bayer Diagnostics, Pittsburgh, PA) versus IMX (n = 100); and high-performance liquid chromatography (HPLC) assay (Bio-Rad Variant A1c; Bio-Rad Laboratories, Richmond, CA) versus IMX assay (n = 55). Our analyses indicate that a relative ranking can be established for the ease of achieving a normal glycated hemoglobin level. The ranking indicates that the most stringent or difficult assays for achieving a normal level are the Isolab and DCA2000 assays. The intermediate assays are the IMX and Bio-Rad Variant, and the easiest method for achieving a normal value is the gel electrophoresis assay. Our results indicate that various glycated hemoglobin assays vary widely and are associated with more or less difficulty for an individual patient to achieve a glycated hemoglobin level within the normal range. These results are especially significant with respect to (1) the clinically narrow therapeutic window of glycated hemoglobin values in type 1 diabetes to avoid rapidly advancing severe hypoglycemia rates and chronic microvascular complication rates, and (2) the glycated hemoglobin threshold for rapidly advancing macrovascular disease in both type 1 and type 2 patients.

Reproductive isolation and the expansion of an invasive hybrid swarm

Science.gov (United States)

Blum, Michael J.; Walters, David M.; Burkhead, Noel M.; Freeman, Byron J.; Porter, Brady A.

2010-01-01

Biological invasions involving hybridization proceed according to prezygotic and postzygotic reproductive isolating mechanisms. Yet few comparisons of reproductive isolation have been carried out to understand how different mechanisms prevent or promote invasions involving hybridization. Here we present a study of prezygotic and postzygotic isolation between non-native red shiner (Cyprinella lutrensis) and native blacktail shiner (C. venusta stigmatura) from the Coosa River basin (USA) to better understand the formation and expansion of invasive hybrid swarms. We conducted spawning trials to measure mating preferences and raised broods from crosses to assay hybrid viability through early juvenile development. Females of both species were more responsive to conspecific mates, although blacktail shiner females responded more often to heterospecific mates than did red shiner females. Fecundity of red shiner females was also higher than blacktail shiner females. Heterospecific crosses resulted in lower fertilization and egg hatching rates, but we found no other evidence of inviability. Rather, we found comparatively low larval mortality of F1 hybrids, which is suggestive of heterosis. These findings support prior inferences of assortative mating from genetic descriptions of hybridization, and that the invasion in the Coosa River is likely proceeding due to interspecific competition and intrinsic hybrid viability.

Validation of a next-generation sequencing assay for clinical molecular oncology.

Science.gov (United States)

Cottrell, Catherine E; Al-Kateb, Hussam; Bredemeyer, Andrew J; Duncavage, Eric J; Spencer, David H; Abel, Haley J; Lockwood, Christina M; Hagemann, Ian S; O'Guin, Stephanie M; Burcea, Lauren C; Sawyer, Christopher S; Oschwald, Dayna M; Stratman, Jennifer L; Sher, Dorie A; Johnson, Mark R; Brown, Justin T; Cliften, Paul F; George, Bijoy; McIntosh, Leslie D; Shrivastava, Savita; Nguyen, Tudung T; Payton, Jacqueline E; Watson, Mark A; Crosby, Seth D; Head, Richard D; Mitra, Robi D; Nagarajan, Rakesh; Kulkarni, Shashikant; Seibert, Karen; Virgin, Herbert W; Milbrandt, Jeffrey; Pfeifer, John D

2014-01-01

Currently, oncology testing includes molecular studies and cytogenetic analysis to detect genetic aberrations of clinical significance. Next-generation sequencing (NGS) allows rapid analysis of multiple genes for clinically actionable somatic variants. The WUCaMP assay uses targeted capture for NGS analysis of 25 cancer-associated genes to detect mutations at actionable loci. We present clinical validation of the assay and a detailed framework for design and validation of similar clinical assays. Deep sequencing of 78 tumor specimens (≥ 1000× average unique coverage across the capture region) achieved high sensitivity for detecting somatic variants at low allele fraction (AF). Validation revealed sensitivities and specificities of 100% for detection of single-nucleotide variants (SNVs) within coding regions, compared with SNP array sequence data (95% CI = 83.4-100.0 for sensitivity and 94.2-100.0 for specificity) or whole-genome sequencing (95% CI = 89.1-100.0 for sensitivity and 99.9-100.0 for specificity) of HapMap samples. Sensitivity for detecting variants at an observed 10% AF was 100% (95% CI = 93.2-100.0) in HapMap mixes. Analysis of 15 masked specimens harboring clinically reported variants yielded concordant calls for 13/13 variants at AF of ≥ 15%. The WUCaMP assay is a robust and sensitive method to detect somatic variants of clinical significance in molecular oncology laboratories, with reduced time and cost of genetic analysis allowing for strategic patient management. Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

Innovative hybrid pile oscillator technique in the Minerve reactor: open loop vs. closed loop

Science.gov (United States)

Geslot, Benoit; Gruel, Adrien; Bréaud, Stéphane; Leconte, Pierre; Blaise, Patrick

2018-01-01

Pile oscillator techniques are powerful methods to measure small reactivity worth of isotopes of interest for nuclear data improvement. This kind of experiments has long been implemented in the Mineve experimental reactor, operated by CEA Cadarache. A hybrid technique, mixing reactivity worth estimation and measurement of small changes around test samples is presented here. It was made possible after the development of high sensitivity miniature fission chambers introduced next to the irradiation channel. A test campaign, called MAESTRO-SL, took place in 2015. Its objective was to assess the feasibility of the hybrid method and investigate the possibility to separate mixed neutron effects, such as fission/capture or scattering/capture. Experimental results are presented and discussed in this paper, which focus on comparing two measurements setups, one using a power control system (closed loop) and another one where the power is free to drift (open loop). First, it is demonstrated that open loop is equivalent to closed loop. Uncertainty management and methods reproducibility are discussed. Second, results show that measuring the flux depression around oscillated samples provides valuable information regarding partial neutron cross sections. The technique is found to be very sensitive to the capture cross section at the expense of scattering, making it very useful to measure small capture effects of highly scattering samples.

Development of high-throughput SNP-based genotyping in Acacia auriculiformis x A. mangium hybrids using short-read transcriptome data

Directory of Open Access Journals (Sweden)

Wong Melissa ML

2012-12-01

Full Text Available Abstract Background Next Generation Sequencing has provided comprehensive, affordable and high-throughput DNA sequences for Single Nucleotide Polymorphism (SNP discovery in Acacia auriculiformis and Acacia mangium. Like other non-model species, SNP detection and genotyping in Acacia are challenging due to lack of genome sequences. The main objective of this study is to develop the first high-throughput SNP genotyping assay for linkage map construction of A. auriculiformis x A. mangium hybrids. Results We identified a total of 37,786 putative SNPs by aligning short read transcriptome data from four parents of two Acacia hybrid mapping populations using Bowtie against 7,839 de novo transcriptome contigs. Given a set of 10 validated SNPs from two lignin genes, our in silico SNP detection approach is highly accurate (100% compared to the traditional in vitro approach (44%. Further validation of 96 SNPs using Illumina GoldenGate Assay gave an overall assay success rate of 89.6% and conversion rate of 37.5%. We explored possible factors lowering assay success rate by predicting exon-intron boundaries and paralogous genes of Acacia contigs using Medicago truncatula genome as reference. This assessment revealed that presence of exon-intron boundary is the main cause (50% of assay failure. Subsequent SNPs filtering and improved assay design resulted in assay success and conversion rate of 92.4% and 57.4%, respectively based on 768 SNPs genotyping. Analysis of clustering patterns revealed that 27.6% of the assays were not reproducible and flanking sequence might play a role in determining cluster compression. In addition, we identified a total of 258 and 319 polymorphic SNPs in A. auriculiformis and A. mangium natural germplasms, respectively. Conclusion We have successfully discovered a large number of SNP markers in A. auriculiformis x A. mangium hybrids using next generation transcriptome sequencing. By using a reference genome from the most closely

Future Potential of Hybrid and Diesel Powertrains in the U.S. Light-duty Vehicle Market

Energy Technology Data Exchange (ETDEWEB)

Greene, D.L.

2004-08-23

Diesel and hybrid technologies each have the potential to increase light-duty vehicle fuel economy by a third or more without loss of performance, yet these technologies have typically been excluded from technical assessments of fuel economy potential on the grounds that hybrids are too expensive and diesels cannot meet Tier 2 emissions standards. Recently, hybrid costs have come down and the few hybrid makes available are selling well. Diesels have made great strides in reducing particulate and nitrogen oxide emissions, and are likely though not certain to meet future standards. In light of these developments, this study takes a detailed look at the market potential of these two powertrain technologies and their possible impacts on light-duty vehicle fuel economy. A nested multinomial logit model of vehicle choice was calibrated to 2002 model year sales of 930 makes, models and engine-transmission configurations. Based on an assessment of the status and outlook for the two technologies, market shares were predicted for 2008, 2012 and beyond, assuming no additional increase in fuel economy standards or other new policy initiatives. Current tax incentives for hybrids are assumed to be phased out by 2008. Given announced and likely introductions by 2008, hybrids could capture 4-7% and diesels 2-4% of the light-duty market. Based on our best guesses for further introductions, these shares could increase to 10-15% for hybrids and 4-7% for diesels by 2012. The resulting impacts on fleet average fuel economy would be about +2% in 2008 and +4% in 2012. If diesels and hybrids were widely available across vehicle classes, makes, and models, they could capture 40% or more of the light-duty vehicle market.

An immunoenzymatic assay for the diagnosis of hepatitis A utilising immunoglobulin Y

Directory of Open Access Journals (Sweden)

Alexandre dos Santos da Silva

2012-11-01

Full Text Available The detection of anti-hepatitis A virus (HAV antibody levels by diagnostic kits in the convalescent period of disease generally use immunoglobulin G (IgG, which is expensive. An alternative to IgG is immunoglobulin Y (IgY, an immunoglobulin antibody encountered in birds and reptiles. The aim of this study was to develop a competitive immunoenzymatic assay to measure total anti-HAV antibody levels using anti-HAV IgY as the capture and conjugated immunoglobulins. For this purpose, anti-HAV IgY was conjugated to horseradish peroxidase (HRP and the optimal dilution of HRP-conjugated antibodies was evaluated to establish the competitive immuneenzymatic assay. The results obtained from our "in-house" assay were plotted on a receiver operator curve, which showed a sensitivity of 95% and a specificity of 98.8%, demonstrating that a competitive anti-HAV IgY immunoenzymatic assay developed "in house" could be used as an alternative to commercial assays that utilise IgG.

HYBRID CONTINUUM-DISCONTINUUM MODELLING OF ROCK FRACUTRE PROCESS IN BRAZILIAN TENSILE STRENGTH TEST

Directory of Open Access Journals (Sweden)

Huaming An

2017-10-01

Full Text Available A hybrid continuum-discontinuum method is introduced to model the rock failure process in Brazilian tensile strength (BTS test. The key component of the hybrid continuum-discontinuum method, i.e. transition from continuum to discontinuum through fracture and fragmentation, is introduced in detail. A laboratory test is conducted first to capture the rock fracture pattern in the BTS test while the tensile strength is calculated according to the peak value of the loading forces. Then the proposed method is used to model the rock behaviour during BTS test. The stress propagation is modelled and compared with those modelled by finite element method in literatures. In addition, the crack initiation and propagation are captured and compared with the facture patter in laboratory test. Moreover, the force-loading displacement curve is obtained which represents a typical brittle material failure process. Furthermore, the stress distributions along the vertical direction are compared with the theoretical solution. It is concluded that the hybrid continuum-discontinuum method can model the stress propagation process and the entire rock failure process in BTS test. The proposed method is a valuable numerical tool for studying the rock behaviour involving the fracture and fragmentation processes.

High-throughput screening assay of hepatitis C virus helicase inhibitors using fluorescence-quenching phenomenon

International Nuclear Information System (INIS)

Tani, Hidenori; Akimitsu, Nobuyoshi; Fujita, Osamu; Matsuda, Yasuyoshi; Miyata, Ryo; Tsuneda, Satoshi; Igarashi, Masayuki; Sekiguchi, Yuji; Noda, Naohiro

2009-01-01

We have developed a novel high-throughput screening assay of hepatitis C virus (HCV) nonstructural protein 3 (NS3) helicase inhibitors using the fluorescence-quenching phenomenon via photoinduced electron transfer between fluorescent dyes and guanine bases. We prepared double-stranded DNA (dsDNA) with a 5'-fluorescent-dye (BODIPY FL)-labeled strand hybridized with a complementary strand, the 3'-end of which has guanine bases. When dsDNA is unwound by helicase, the dye emits fluorescence owing to its release from the guanine bases. Our results demonstrate that this assay is suitable for quantitative assay of HCV NS3 helicase activity and useful for high-throughput screening for inhibitors. Furthermore, we applied this assay to the screening for NS3 helicase inhibitors from cell extracts of microorganisms, and found several cell extracts containing potential inhibitors.

Thermal modeling of a hydraulic hybrid vehicle transmission based on thermodynamic analysis

International Nuclear Information System (INIS)

Kwon, Hyukjoon; Sprengel, Michael; Ivantysynova, Monika

2016-01-01

Hybrid vehicles have become a popular alternative to conventional powertrain architectures by offering improved fuel efficiency along with a range of environmental benefits. Hydraulic Hybrid Vehicles (HHV) offer one approach to hybridization with many benefits over competing technologies. Among these benefits are lower component costs, more environmentally friendly construction materials, and the ability to recover a greater quantity of energy during regenerative braking which make HHVs partially well suited to urban environments. In order to further the knowledge base regarding HHVs, this paper explores the thermodynamic characteristics of such a system. A system model is detailed for both the hydraulic and thermal components of a closed circuit hydraulic hybrid transmission following the FTP-72 driving cycle. Among the new techniques proposed in this paper is a novel method for capturing rapid thermal transients. This paper concludes by comparing the results of this model with experimental data gathered on a Hardware-in-the-Loop (HIL) transmission dynamometer possessing the same architecture, components, and driving cycle used within the simulation model. This approach can be used for several applications such as thermal stability analysis of HHVs, optimal thermal management, and analysis of the system's thermodynamic efficiency. - Highlights: • Thermal modeling for HHVs is introduced. • A model for the hydraulic and thermal system is developed for HHVs. • A novel method for capturing rapid thermal transients is proposed. • The thermodynamic system diagram of a series HHV is predicted.

Co-ordinated research program on development of kits for radioimmunometric assays of tumor markers

International Nuclear Information System (INIS)

Acevedo Castro, B.E.; Perera Negrin, Y.; Pichardo Diaz, D.; Murugiah, R.; Ayala Avila, M.; Gavilondo Cowley, J.; Ruiz Pena, M.; Caso Pena, R.; Hernandez Pagarizabal, M.

1999-01-01

Mice were immunized with semen and natural PSA, following three different schemes. Splenocytes from two hyper immune animals were fused with the P3/x63.Ag8.653 myeloma under conventional hybridoma procedures. Stable hybrid cell cultures secreting antibodies specific to natural PSA were obtained by cloning dilutions procedures. With a group of stable hybridoma cultures we developed epitope characterization assays to determine whether the antibodies were capable of recognizing PSA. According to the recognition level in ELISA, the hybridomas were classified in different groups,. We select the best pairs of Mabs for developing a total and free PSA assays based on ELISA or IRMA format. Our total-PSA based on IRMA format presented a good correlation in comparison with CIS bio total PSA assay. We recommend our anti-PSA monoclonal antibodies to develop an IRMA assay for total PSA. Cuban free-PSA assay is under evaluation at present. (author)

A highly scalable peptide-based assay system for proteomics.

Directory of Open Access Journals (Sweden)

Igor A Kozlov

Full Text Available We report a scalable and cost-effective technology for generating and screening high-complexity customizable peptide sets. The peptides are made as peptide-cDNA fusions by in vitro transcription/translation from pools of DNA templates generated by microarray-based synthesis. This approach enables large custom sets of peptides to be designed in silico, manufactured cost-effectively in parallel, and assayed efficiently in a multiplexed fashion. The utility of our peptide-cDNA fusion pools was demonstrated in two activity-based assays designed to discover protease and kinase substrates. In the protease assay, cleaved peptide substrates were separated from uncleaved and identified by digital sequencing of their cognate cDNAs. We screened the 3,011 amino acid HCV proteome for susceptibility to cleavage by the HCV NS3/4A protease and identified all 3 known trans cleavage sites with high specificity. In the kinase assay, peptide substrates phosphorylated by tyrosine kinases were captured and identified by sequencing of their cDNAs. We screened a pool of 3,243 peptides against Abl kinase and showed that phosphorylation events detected were specific and consistent with the known substrate preferences of Abl kinase. Our approach is scalable and adaptable to other protein-based assays.

Rapid detection of the CYP2A6*12 hybrid allele by Pyrosequencing® technology

Directory of Open Access Journals (Sweden)

Gallagher Margaret L

2009-08-01

Full Text Available Abstract Background Identification of CYP2A6 alleles associated with reduced enzyme activity is important in the study of inter-individual differences in drug metabolism. CYP2A6*12 is a hybrid allele that results from unequal crossover between CYP2A6 and CYP2A7 genes. The 5' regulatory region and exons 1–2 are derived from CYP2A7, and exons 3–9 are derived from CYP2A6. Conventional methods for detection of CYP2A6*12 consist of two-step PCR protocols that are laborious and unsuitable for high-throughput genotyping. We developed a rapid and accurate method to detect the CYP2A6*12 allele by Pyrosequencing technology. Methods A single set of PCR primers was designed to specifically amplify both the CYP2A6*1 wild-type allele and the CYP2A6*12 hybrid allele. An internal Pyrosequencing primer was used to generate allele-specific sequence information, which detected homozygous wild-type, heterozygous hybrid, and homozygous hybrid alleles. We first validated the assay on 104 DNA samples that were also genotyped by conventional two-step PCR and by cycle sequencing. CYP2A6*12 allele frequencies were then determined using the Pyrosequencing assay on 181 multi-ethnic DNA samples from subjects of African American, European Caucasian, Pacific Rim, and Hispanic descent. Finally, we streamlined the Pyrosequencing assay by integrating liquid handling robotics into the workflow. Results Pyrosequencing results demonstrated 100% concordance with conventional two-step PCR and cycle sequencing methods. Allele frequency data showed slightly higher prevalence of the CYP2A6*12 allele in European Caucasians and Hispanics. Conclusion This Pyrosequencing assay proved to be a simple, rapid, and accurate alternative to conventional methods, which can be easily adapted to the needs of higher-throughput studies.

Polymerase chain reaction-hybridization method using urease gene sequences for high-throughput Ureaplasma urealyticum and Ureaplasma parvum detection and differentiation.

Science.gov (United States)

Xu, Chen; Zhang, Nan; Huo, Qianyu; Chen, Minghui; Wang, Rengfeng; Liu, Zhili; Li, Xue; Liu, Yunde; Bao, Huijing

2016-04-15

In this article, we discuss the polymerase chain reaction (PCR)-hybridization assay that we developed for high-throughput simultaneous detection and differentiation of Ureaplasma urealyticum and Ureaplasma parvum using one set of primers and two specific DNA probes based on urease gene nucleotide sequence differences. First, U. urealyticum and U. parvum DNA samples were specifically amplified using one set of biotin-labeled primers. Furthermore, amine-modified DNA probes, which can specifically react with U. urealyticum or U. parvum DNA, were covalently immobilized to a DNA-BIND plate surface. The plate was then incubated with the PCR products to facilitate sequence-specific DNA binding. Horseradish peroxidase-streptavidin conjugation and a colorimetric assay were used. Based on the results, the PCR-hybridization assay we developed can specifically differentiate U. urealyticum and U. parvum with high sensitivity (95%) compared with cultivation (72.5%). Hence, this study demonstrates a new method for high-throughput simultaneous differentiation and detection of U. urealyticum and U. parvum with high sensitivity. Based on these observations, the PCR-hybridization assay developed in this study is ideal for detecting and discriminating U. urealyticum and U. parvum in clinical applications. Copyright © 2016 Elsevier Inc. All rights reserved.

CO2 capture by Li-functionalized silicene

KAUST Repository

Zhu, Jiajie

2016-05-18

CO2 capture and storage technology is of key importance to reduce the greenhouse effect. By its large surface area and sp3 hybridization, Li-functionalized silicene is demonstrated to be a promising CO2 absorbent that is stable up to at least 500 K and has a very high storage capacity of 28.6 mol/kg (55.7 wt%). The adsorption energy of CO2 on Li-functionalized silicene is enhanced as compared to pristine silicene, to attain an almost ideal value that still facilitates easy release. In addition, the band gap is found to change sensitively with the CO2 coverage. (© 2016 WILEY-VCH Verlag GmbH &Co. KGaA, Weinheim). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Misregulation of Gene Expression and Sterility in Interspecies Hybrids: Causal Links and Alternative Hypotheses.

Science.gov (United States)

Civetta, Alberto

2016-05-01

Understanding the origin of species is of interest to biologist in general and evolutionary biologist in particular. Hybrid male sterility (HMS) has been a focus in studies of speciation because sterility imposes a barrier to free gene flow between organisms, thus effectively isolating them as distinct species. In this review, I focus on the role of differential gene expression in HMS and speciation. Microarray and qPCR assays have established associations between misregulation of gene expression and sterility in hybrids between closely related species. These studies originally proposed disrupted expression of spermatogenesis genes as a causative of sterility. Alternatively, rapid genetic divergence of regulatory elements, particularly as they relate to the male sex (fast-male evolution), can drive the misregulation of sperm developmental genes in the absence of sterility. The use of fertile hybrids (both backcross and F1 progeny) as controls has lent support to this alternative explanation. Differences in gene expression between fertile and sterile hybrids can also be influenced by a pattern of faster evolution of the sex chromosome (fast-X evolution) than autosomes. In particular, it would be desirable to establish whether known X-chromosome sterility factors can act as trans-regulatory drivers of genome-wide patterns of misregulation. Genome-wide expression studies coupled with assays of proxies of sterility in F1 and BC progeny have identified candidate HMS genes but functional assays, and a better phenotypic characterization of sterility phenotypes, are needed to rigorously test how these genes might contribute to HMS.

Construction and characterisation of a modular microfluidic system: coupling magnetic capture and electrochemical detection

DEFF Research Database (Denmark)

Godino, N.; Snakenborg, Detlef; Kutter, Jörg Peter

2010-01-01

, and a polycarbonate base where permanent magnets are hosted; these parts are designed to fit so that wire bonding and encapsulation are avoided. This system can perform bioassays over the surface of magnetic beads and uses only 50 mu L of bead suspension per assay. Following detection, captured beads are released...

Ultrasensitive electrochemical detection of DNA based on Zn²⁺ assistant DNA recycling followed with hybridization chain reaction dual amplification.

Science.gov (United States)

Qian, Yong; Wang, Chunyan; Gao, Fenglei

2015-01-15

A new strategy to combine Zn(2+) assistant DNA recycling followed with hybridization chain reaction dual amplification was designed for highly sensitive electrochemical detection of target DNA. A gold electrode was used to immobilize molecular beacon (MB) as the recognition probe and perform the amplification procedure. In the presence of the target DNA, the hairpin probe 1 was opened, and the DNAzyme was liberated from the caged structure. The activated DNAzyme hybridized with the MB and catalyzed its cleavage in the presence of Zn(2+) cofactor and resulting in a free DNAzyme strand. Finally, each target-induced activated DNAzyme underwent many cycles triggering the cleavage of MB, thus forming numerous MB fragments. The MB fragments triggered the HCR and formed a long double-helix DNA structure. Because both H1 and H2 were labeled by biotin, a lot of SA-ALP was captured on the electrode surface, thus catalyzing a silver deposition process for electrochemical stripping analysis. This novel cascade signal amplification strategy can detect target DNA down to the attomolar level with a dynamic range spanning 6 orders of magnitude. This highly sensitive and specific assay has a great potential to become a promising DNA quantification method in biomedical research and clinical diagnosis. Copyright © 2014 Elsevier B.V. All rights reserved.

Phylogenetic tests of a Cercopithecus monkey hybrid reveal X ...

African Journals Online (AJOL)

A captive Cercopithecus nictitans × C. cephus male was examined at loci on the X- and Y-chromosomes as a test of previously described phylogenetic methods for identifying hybrid Cercopithecus monkeys. The results confirm the reliability of such assays, indicating that they can be of immediate utility for studies of wild ...

Gene expression disruptions of organism versus organ in Drosophila species hybrids.

Directory of Open Access Journals (Sweden)

Daniel J Catron

2008-08-01

Full Text Available Hybrid dysfunctions, such as sterility, may result in part from disruptions in the regulation of gene expression. Studies of hybrids within the Drosophila simulans clade have reported genes expressed above or below the expression observed in their parent species, and such misexpression is associated with male sterility in multigenerational backcross hybrids. However, these studies often examined whole bodies rather than testes or had limited replication using less-sensitive but global techniques. Here, we use a new RNA isolation technique to re-examine hybrid gene expression disruptions in both testes and whole bodies from single Drosophila males by real-time quantitative RT-PCR. We find two early-spermatogenesis transcripts are underexpressed in hybrid whole-bodies but not in assays of testes alone, while two late-spermatogenesis transcripts seem to be underexpressed in both whole-bodies and testes alone. Although the number of transcripts surveyed is limited, these results provide some support for a previous hypothesis that the spermatogenesis pathway in these sterile hybrids may be disrupted sometime after the expression of the early meiotic arrest genes.

Comparison of central HER2 testing with quantitative total HER2 expression and HER2 homodimer measurements using a novel proximity-based assay.

Science.gov (United States)

Huang, Weidong; Reinholz, Monica; Weidler, Jodi; Yolanda, Lie; Paquet, Agnes; Whitcomb, Jeannette; Lingle, Wilma; Jenkins, Robert B; Chen, Beiyun; Larson, Jeffrey S; Tan, Yuping; Sherwood, Thomas; Bates, Michael; Perez, Edith A

2010-08-01

The accuracy and reliability of immunohistochemical analysis and in situ hybridization for the assessment of HER2 status remains a subject of debate. We developed a novel assay (HERmark Breast Cancer Assay, Monogram Biosciences, South San Francisco, CA) that provides precise quantification of total HER2 protein expression (H2T) and HER2 homodimers (H2D) in formalin-fixed, paraffin-embedded tissue specimens. H2T and H2D results of 237 breast cancers were compared with those of immunohistochemical studies and fluorescence in situ hybridization (FISH) centrally performed at the Mayo Clinic, Rochester, MN. H2T described a continuum across a wide dynamic range ( approximately 2.5 log). Excluding the equivocal cases, HERmark showed 98% concordance with immunohistochemical studies for positive and negative assay values. For the 94 immunohistochemically equivocal cases, 67% and 39% concordance values were observed between HERmark and FISH for positive and negative assay values, respectively. Polysomy 17 in the absence of HER2 gene amplification did not result in HER2 overexpression as evaluated quantitatively using the HERmark assay.

Hybrid Semantics of Stochastic Programs with Dynamic Reconfiguration

Directory of Open Access Journals (Sweden)

Alberto Policriti

2009-10-01

Full Text Available We begin by reviewing a technique to approximate the dynamics of stochastic programs --written in a stochastic process algebra-- by a hybrid system, suitable to capture a mixed discrete/continuous evolution. In a nutshell, the discrete dynamics is kept stochastic while the continuous evolution is given in terms of ODEs, and the overall technique, therefore, naturally associates a Piecewise Deterministic Markov Process with a stochastic program. The specific contribution in this work consists in an increase of the flexibility of the translation scheme, obtained by allowing a dynamic reconfiguration of the degree of discreteness/continuity of the semantics. We also discuss the relationships of this approach with other hybrid simulation strategies for biochemical systems.

Biological evaluation of zirconia/PEG hybrid materials synthesized via sol–gel technique

Energy Technology Data Exchange (ETDEWEB)

Catauro, M., E-mail: michelina.catauro@unina2.it [Department of Industrial and Information Engineering, Second University of Naples, Via Roma 29, 81031 Aversa (Italy); Papale, F.; Bollino, F. [Department of Industrial and Information Engineering, Second University of Naples, Via Roma 29, 81031 Aversa (Italy); Gallicchio, M.; Pacifico, S. [Department Environmental, Biological and Pharmaceutical Sciences and Technologies, Second University of Naples, Via Vivaldi 43, 81100 Caserta (Italy)

2014-07-01

The objective of the following study has been the synthesis via sol–gel and the characterization of novel organic–inorganic hybrid materials to be used in biomedical field. The prepared materials consist of an inorganic zirconia matrix containing as organic component the polyethylene glycol (PEG), a water-soluble polymer used in medical and pharmaceutical fields. Various hybrids have been synthesized changing the molar ratio between the organic and inorganic parts. Fourier transform spectroscopy suggests that the structure of the interpenetrating network is realized by hydrogen bonds between the Zr-OH group in the sol–gel intermediate species and both the terminal alcoholic group and ethereal oxygen atoms in the repeating units of polymer The amorphous nature of the gels has been ascertained by X-ray diffraction analysis. The morphology observation has been carried out by using the Scanning Electron Microscope and has confirmed that the obtained materials are nanostructurated hybrids. The bioactivity of the synthesized system has been shown by the formation of a hydroxyapatite layer on the surface of samples soaked in a fluid simulating the human blood plasma. The potential biocompatibility of hybrids has been assessed as performing indirect MTT cytotoxicity assay towards 3T3 cell line at 24, 48, and 72 h exposure times. - Highlights: • ZrO{sub 2}/PEG amorphous class I organic–inorganic hybrid synthesis via sol–gel • Bioactivity evaluation of materials by the formation of apatite on surface in SBF • Biocompatibility test with indirect MTT cytotoxicity assay on NHI 3T3 cell line.

Biological evaluation of zirconia/PEG hybrid materials synthesized via sol–gel technique

International Nuclear Information System (INIS)

Catauro, M.; Papale, F.; Bollino, F.; Gallicchio, M.; Pacifico, S.

2014-01-01

The objective of the following study has been the synthesis via sol–gel and the characterization of novel organic–inorganic hybrid materials to be used in biomedical field. The prepared materials consist of an inorganic zirconia matrix containing as organic component the polyethylene glycol (PEG), a water-soluble polymer used in medical and pharmaceutical fields. Various hybrids have been synthesized changing the molar ratio between the organic and inorganic parts. Fourier transform spectroscopy suggests that the structure of the interpenetrating network is realized by hydrogen bonds between the Zr-OH group in the sol–gel intermediate species and both the terminal alcoholic group and ethereal oxygen atoms in the repeating units of polymer The amorphous nature of the gels has been ascertained by X-ray diffraction analysis. The morphology observation has been carried out by using the Scanning Electron Microscope and has confirmed that the obtained materials are nanostructurated hybrids. The bioactivity of the synthesized system has been shown by the formation of a hydroxyapatite layer on the surface of samples soaked in a fluid simulating the human blood plasma. The potential biocompatibility of hybrids has been assessed as performing indirect MTT cytotoxicity assay towards 3T3 cell line at 24, 48, and 72 h exposure times. - Highlights: • ZrO 2 /PEG amorphous class I organic–inorganic hybrid synthesis via sol–gel • Bioactivity evaluation of materials by the formation of apatite on surface in SBF • Biocompatibility test with indirect MTT cytotoxicity assay on NHI 3T3 cell line

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

Science.gov (United States)

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

2010-01-01

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

Enhanced CO_2 capture on graphene via N, S dual-doping

International Nuclear Information System (INIS)

Li, Jieyuan; Hou, Meiling; Chen, Yanqiu; Cen, Wanglai; Chu, Yinghao; Yin, Shi

2017-01-01

Highlights: • Sluggish conjugated π bonds of graphene should be weakened to promote adsorption activity. • A charge delivery channel along S → N → CO_2 path should be prior responsible for the enhancement of CO_2 capture on graphene. • Applicative temperature range of graphene-based adsorbents for CO_2 capture is extend to about 100 °C via N, S dual-doping. - Abstract: N, S doped graphene-based materials have been recently recognized as promising adsorbents for CO_2 capture, but understanding of the adsorption mechanism at the atomic level is still limited. Herein, the local structures and promotion mechanism of CO_2 capture by N, S doped graphene were investigated by combining density functional theory and ab initio thermodynamics. A single vacancy defected graphene involving N, S dual-doping was found to be a superior adsorbent for CO_2 capture under mild conditions (<100 °C, 1 atm). The enhanced CO_2 adsorption performance should be ascribed to a charge delivery channel along the S → N → CO_2 path, leading to extra charge transfer from graphene to CO_2. It is worth mentioning that the extra charge transfer was stimulated by the unique sp"2 hybridization of pyridine N and further enhanced by S in N, S dual-doped graphene. A possible mechanism has been proposed to explain the high adsorption performance of CO_2 by N, S dual-doped graphene, which offers insights for the design of new graphene-based adsorbents.

Yield stability and adaptability of maize hybrids based on GGE biplot analysis characteristics

Directory of Open Access Journals (Sweden)

Marcio Balestre

2009-01-01

Full Text Available The objective of this study was to evaluate stability and adaptability of the grain yield of commercial intervarietalmaize hybrids by the GGE (Genotype and Genotype by Environment Interaction biplot and AMMI (Additive Main Effects andMultiplicative Interaction analyses. Two intervarietal hybrids (BIO 2 and BIO4 were evaluated together with single, doubleand three-way cross hybrids. The performance of the intervarietal hybrid BIO 4 was superior to all double and three-waycross hybrids and outmatched the single-cross hybrids by 43%. In terms of stability, BIO 2 was more stable than BIO4, whichis desirable, but biological stability, which is not necessarily desirable, was also observed, since the yield was below theenvironmental mean. The graphical GGE biplot analysis was superior to the AMMI1 since a greater portion of the sum ofsquares of GE and G+GE was captured and the predictive accuracy was higher. On the other hand, the AMMI2 graphoutperformed the GGE biplot in predictive accuracy and explanation of G + GE and GE, although the difference in accuracywas smaller than between GGE2 and AMMI1.

Innovative hybrid pile oscillator technique in the Minerve reactor: open loop vs. closed loop

Directory of Open Access Journals (Sweden)

Geslot Benoit

2018-01-01

Full Text Available Pile oscillator techniques are powerful methods to measure small reactivity worth of isotopes of interest for nuclear data improvement. This kind of experiments has long been implemented in the Mineve experimental reactor, operated by CEA Cadarache. A hybrid technique, mixing reactivity worth estimation and measurement of small changes around test samples is presented here. It was made possible after the development of high sensitivity miniature fission chambers introduced next to the irradiation channel. A test campaign, called MAESTRO-SL, took place in 2015. Its objective was to assess the feasibility of the hybrid method and investigate the possibility to separate mixed neutron effects, such as fission/capture or scattering/capture. Experimental results are presented and discussed in this paper, which focus on comparing two measurements setups, one using a power control system (closed loop and another one where the power is free to drift (open loop. First, it is demonstrated that open loop is equivalent to closed loop. Uncertainty management and methods reproducibility are discussed. Second, results show that measuring the flux depression around oscillated samples provides valuable information regarding partial neutron cross sections. The technique is found to be very sensitive to the capture cross section at the expense of scattering, making it very useful to measure small capture effects of highly scattering samples.

A daunting challenge

DEFF Research Database (Denmark)

Rebolj, Matejka; Bonde, Jesper; Ejegod, Ditte

2015-01-01

We compared cytology with Hybrid Capture 2 (HC2), cobas, CLART and APTIMA Human Papillomavirus (HPV) assays in primary cervical screening at age 23-29 years based on data from the Danish Horizon study. SurePath samples were collected from 1278 women undergoing routine cytology-based screening...

PCR associated with hybridization with DNA radioactive probes for diagnosis of asymptomatic infection caused by Leishmania Chagasi

International Nuclear Information System (INIS)

Andrade, Antero Silva Ribeiro de; Moreno, Elizabeth Castro; Gomes, Rosangela Fatima; Melo, Maria Norma de; Carneiro, Mariangela; Fernandes, Octavio

2002-01-01

Detection systems for diagnosis of leishmaniasis based on PCR are very promising due to their sensitivity and specificity. Secondary detection by specific radioactive DNA probes, able to type the PCR amplified products, increase the specificity and raise about tem-fold the sensitivity of the assay. The aim of this work was evaluate PCR and hybridization as a tool to identify Leishmania (Leishmania) chagasi (the specie that cause the visceral leishmaniasis in Brazil) infection in asymptomatic persons living in a endemic area. Material and Methods: A group of 226 asymptomatic individuals, living in General Carneiro (MG), was selected. Blood samples were harvested and the DNA extracted from the mononucleate cells. PCR was performed using primers addressed to the kinetoplast DNA minicircles. This protocol gives a positive reaction for all Leishmania species. The amplified products were further hybridized with cloned L.chagasi minicircles labeled with 32 P. Results: were identified 111 samples PCR positive, 2 of them hybridization negative and 133 samples hybridization positive, 24 of them PCR negative. The occurrence of samples with hybridization positive and PCR negative was expected since hybridization, with DNA probes labeled with 32 P, increase the sensitivity of the assay. The samples that presented positive PCR and negative hybridization were probably due the presence of other Leishmania species, likely L. (V.) braziliensis (that produce tegumentary leishmaniasis in the region), since L. (L.) chagasi cloned minicircles were used as hybridization probe. We conclude that this procedure is a valuable tool to access subclinical L. (L.) chagasi infections in epidemiological studies. (author)

Evaluation of environmental genotoxicity by comet assay in Columba livia.

Science.gov (United States)

González-Acevedo, Anahi; García-Salas, Juan A; Gosálvez, Jaime; Fernández, José Luis; Dávila-Rodríguez, Martha I; Cerda-Flores, Ricardo M; Méndez-López, Luis F; Cortés-Gutiérrez, Elva I

2016-01-01

The concentrations of recognized or suspected genotoxic and carcinogenic agents found in the air of large cities and, in particular, developing countries, have raised concerns about the potential for chronic health effects in the populations exposed to them. The biomonitoring of environmental genotoxicity requires the selection of representative organisms as "sentinels," as well as the development of suitable and sensitive assays, such as those aimed at assessing DNA damage. The aim of this study was to evaluate DNA damage levels in erythrocytes from Columba livia living in the metropolitan area of Monterrey, Mexico, compared with control animals via comet assay, and to confirm the results via Micronuclei test (MN) and DNA breakage detection-fluorescence in situ hybridization (DBD-FISH). Our results showed a significant increase in DNA migration in animals from the area assayed compared with that observed in control animals sampled in non-contaminated areas. These results were confirmed by MN test and DBD-FISH. In conclusion, these observations confirm that the examination of erythrocytes from Columba livia via alkaline comet assay provides a sensitive and reliable end point for the detection of environmental genotoxicants.

PCR-free quantitative detection of genetically modified organism from raw materials – A novel electrochemiluminescence-based bio-barcode method

Science.gov (United States)

Zhu, Debin; Tang, Yabing; Xing, Da; Chen, Wei R.

2018-01-01

Bio-barcode assay based on oligonucleotide-modified gold nanoparticles (Au-NPs) provides a PCR-free method for quantitative detection of nucleic acid targets. However, the current bio-barcode assay requires lengthy experimental procedures including the preparation and release of barcode DNA probes from the target-nanoparticle complex, and immobilization and hybridization of the probes for quantification. Herein, we report a novel PCR-free electrochemiluminescence (ECL)-based bio-barcode assay for the quantitative detection of genetically modified organism (GMO) from raw materials. It consists of tris-(2’2’-bipyridyl) ruthenium (TBR)-labele barcode DNA, nucleic acid hybridization using Au-NPs and biotin-labeled probes, and selective capture of the hybridization complex by streptavidin-coated paramagnetic beads. The detection of target DNA is realized by direct measurement of ECL emission of TBR. It can quantitatively detect target nucleic acids with high speed and sensitivity. This method can be used to quantitatively detect GMO fragments from real GMO products. PMID:18386909

PCR-free quantitative detection of genetically modified organism from raw materials. An electrochemiluminescence-based bio bar code method.

Science.gov (United States)

Zhu, Debin; Tang, Yabing; Xing, Da; Chen, Wei R

2008-05-15

A bio bar code assay based on oligonucleotide-modified gold nanoparticles (Au-NPs) provides a PCR-free method for quantitative detection of nucleic acid targets. However, the current bio bar code assay requires lengthy experimental procedures including the preparation and release of bar code DNA probes from the target-nanoparticle complex and immobilization and hybridization of the probes for quantification. Herein, we report a novel PCR-free electrochemiluminescence (ECL)-based bio bar code assay for the quantitative detection of genetically modified organism (GMO) from raw materials. It consists of tris-(2,2'-bipyridyl) ruthenium (TBR)-labeled bar code DNA, nucleic acid hybridization using Au-NPs and biotin-labeled probes, and selective capture of the hybridization complex by streptavidin-coated paramagnetic beads. The detection of target DNA is realized by direct measurement of ECL emission of TBR. It can quantitatively detect target nucleic acids with high speed and sensitivity. This method can be used to quantitatively detect GMO fragments from real GMO products.

Comparison of clinical samples for visceral Leishmaniasis diagnosis in asymptomatic dogs by PCR hybridization

International Nuclear Information System (INIS)

Ferreira, Sidney A.; Ituassu, Leonardo T.; Melo, Maria N.

2009-01-01

The canine visceral leishmaniasis (CVL) diagnosis still represents a challenge because of complexity of this disease. The aim of present study was to compare different clinical samples for diagnosis of CVL by Polymerase Chain Reaction (PCR) combined with hybridization of 32 P labeled probes. Bone marrow (BM), skin biopsy (SB), peripheral blood (PB) and conjunctival swab (CS) were used in this work. With this purpose 40 asymptomatic dogs, all positive by parasitological test, were obtained. From each animal were collected SB with sterile punches from ear internal surface, 1.0 mL of PB, BM aspirates from sternum and CS from both lower eyelid. Each clinical sample was submitted to suitable DNA purification process and PCR-hybridization assays. The positive results obtained with PCR were 55%, 25%, 30% and 22.5% for CS, BM, SB and PB respectively while the PCR followed by hybridization showed a positivity of 87.5%, 50%, 45% and 27.5% respectively. The hybridization assay was able to increase the PCR positivity in all kinds of clinical samples. The best performance was obtained using CS samples. We concluded that the PCR associated with DNA radioactive probes was a very sensitive tool for diagnosis of CVL in asymptomatic dogs and the CS has an important potential for regular screening of dogs. (author)

Hybrid capture vs. PCR screening of cervical human papilloma virus infections. Cytological and histological associations in 1270 women

International Nuclear Information System (INIS)

Tsiodras, Sotirios; Georgoulakis, John; Chranioti, Aikaterini; Voulgaris, Zanis; Psyrri, Amanda; Tsivilika, Angeliki; Panayiotides, John; Karakitsos, Petros

2010-01-01

We evaluated two molecular methods of HPV detection and their correlation with cytological and histological diagnosis in a large sample of Greek women. All women with liquid-based cytology performed at a University Hospital between 2000 and 2003 were included. The Hybrid Capture 2 (HC2) kit and in house Polymerase Chain Reaction (PCR) were used for HPV DNA detection. Cervical biopsy was performed for women with ASCUS+ cytology, HPV detection, or abnormal colposcopy. Positive (PLR) and negative (NLR) likelihood ratios were calculated for cytology and HPV molecular testing for the prediction of CIN2 and greater histology. Of the 1270 women evaluated 241 (18.5%) had abnormal cytology. Cytology diagnosed high-grade squamous intraepithelial lesion (HSIL) or invasive carcinoma in 21(1.7%) cases whereas 26 (2%) women had CIN2+ or greater histology. PCR detected HPV in 397/1270 (31.3%) and HC2 in 260/1270 (20.4%) samples. Both molecular tests exhibited high reproducibility (Cohen's kappa value 0.691, 95% CI: 0.664 - 0.718). Positive likelihood ratios (PLR) of 9.4, 3.8 and 3.4 and negative likelihood ratios of 0.13, 0.21, and 0 were noted for ≥ LSIL, any positive HC2 or any positive PCR-HPV testing, for predicting CIN2+ histology, respectively. All CIN 3+ lesions harbored high risk oncogenic HPV type infections. HPV infection was found in a large proportion of this population and was associated with CIN 2/3 lesions and infiltrating carcinomas. Thin prep testing and HPV detection by HC2 or PCR performed very well with regards to identifying high grade lesions in an environment with experienced examiners

Structured oligonucleotides for target indexing to allow single-vessel PCR amplification and solid support microarray hybridization.

Science.gov (United States)

Girard, Laurie D; Boissinot, Karel; Peytavi, Régis; Boissinot, Maurice; Bergeron, Michel G

2015-02-07

The combination of molecular diagnostic technologies is increasingly used to overcome limitations on sensitivity, specificity or multiplexing capabilities, and provide efficient lab-on-chip devices. Two such techniques, PCR amplification and microarray hybridization are used serially to take advantage of the high sensitivity and specificity of the former combined with high multiplexing capacities of the latter. These methods are usually performed in different buffers and reaction chambers. However, these elaborate methods have high complexity and cost related to reagent requirements, liquid storage and the number of reaction chambers to integrate into automated devices. Furthermore, microarray hybridizations have a sequence dependent efficiency not always predictable. In this work, we have developed the concept of a structured oligonucleotide probe which is activated by cleavage from polymerase exonuclease activity. This technology is called SCISSOHR for Structured Cleavage Induced Single-Stranded Oligonucleotide Hybridization Reaction. The SCISSOHR probes enable indexing the target sequence to a tag sequence. The SCISSOHR technology also allows the combination of nucleic acid amplification and microarray hybridization in a single vessel in presence of the PCR buffer only. The SCISSOHR technology uses an amplification probe that is irreversibly modified in presence of the target, releasing a single-stranded DNA tag for microarray hybridization. Each tag is composed of a 3-nucleotide sequence-dependent segment and a unique "target sequence-independent" 14-nucleotide segment allowing for optimal hybridization with minimal cross-hybridization. We evaluated the performance of five (5) PCR buffers to support microarray hybridization, compared to a conventional hybridization buffer. Finally, as a proof of concept, we developed a multiplexed assay for the amplification, detection, and identification of three (3) DNA targets. This new technology will facilitate the design

Tumorigenic hybrids between mesenchymal stem cells and gastric cancer cells enhanced cancer proliferation, migration and stemness

International Nuclear Information System (INIS)

Xue, Jianguo; Zhu, Yuan; Sun, Zixuan; Ji, Runbi; Zhang, Xu; Xu, Wenrong; Yuan, Xiao; Zhang, Bin; Yan, Yongmin; Yin, Lei; Xu, Huijuan; Zhang, Leilei; Zhu, Wei; Qian, Hui

2015-01-01

Emerging evidence indicates that inappropriate cell-cell fusion might contribute to cancer progression. Similarly, mesenchymal stem cells (MSCs) can also fuse with other cells spontaneously and capable of adopting the phenotype of other cells. The aim of our study was to investigate the role of MSCs participated cell fusion in the tumorigenesis of gastric cancer. We fused human umbilical cord mesenchymal stem cells (hucMSCs) with gastric cancer cells in vitro by polyethylene glycol (PEG), the hybrid cells were sorted by flow cytometer. The growth and migration of hybrids were assessed by cell counting, cell colony formation and transwell assays. The proteins and genes related to epithelial-mesenchymal transition and stemness were tested by western blot, immunocytochemistry and real-time RT-PCR. The expression of CD44 and CD133 was examined by immunocytochemistry and flow cytometry. The xenograft assay was used to evaluation the tumorigenesis of the hybrids. The obtained hybrids exhibited epithelial- mesenchymal transition (EMT) change with down-regulation of E-cadherin and up-regulation of Vimentin, N-cadherin, α-smooth muscle actin (α-SMA), and fibroblast activation protein (FAP). The hybrids also increased expression of stemness factors Oct4, Nanog, Sox2 and Lin28. The expression of CD44 and CD133 on hybrid cells was stronger than parental gastric cancer cells. Moreover, the migration and proliferation of heterotypic hybrids were enhanced. In addition, the heterotypic hybrids promoted the growth abilities of gastric xenograft tumor in vivo. Taken together, our results suggest that cell fusion between hucMSCs and gastric cancer cells could contribute to tumorigenic hybrids with EMT and stem cell-like properties, which may provide a flexible tool for investigating the roles of MSCs in gastric cancer. The online version of this article (doi:10.1186/s12885-015-1780-1) contains supplementary material, which is available to authorized users

Hybrid electrospun chitosan-phospholipids nanofibers for transdermal drug delivery.

Science.gov (United States)

Mendes, Ana C; Gorzelanny, Christian; Halter, Natalia; Schneider, Stefan W; Chronakis, Ioannis S

2016-08-20

Chitosan (Ch) polysaccharide was mixed with phospholipids (P) to generate electrospun hybrid nanofibers intended to be used as platforms for transdermal drug delivery. Ch/P nanofibers exibithed average diameters ranging from 248±94nm to 600±201nm, depending on the amount of phospholipids used. Fourier Transformed Infra-Red (FTIR) spectroscopy and Dynamic Light Scattering (DLS) data suggested the occurrence of electrostatic interactions between amine groups of chitosan with the phospholipid counterparts. The nanofibers were shown to be stable for at least 7days in Phosphate Buffer Saline (PBS) solution. Cytotoxicity studies (WST-1 and LDH assays) demonstrated that the hybrid nanofibers have suitable biocompatibility. Fluorescence microscopy, also suggested that L929 cells seeded on top of the CH/P hybrid have similar metabolic activity comparatively to the cells seeded on tissue culture plate (control). The release of curcumin, diclofenac and vitamin B12, as model drugs, from Ch/P hybrid nanofibers was investigated, demonstrating their potential utilization as a transdermal drug delivery system. Copyright © 2016 Elsevier B.V. All rights reserved.

A Gaussian process regression based hybrid approach for short-term wind speed prediction

International Nuclear Information System (INIS)

Zhang, Chi; Wei, Haikun; Zhao, Xin; Liu, Tianhong; Zhang, Kanjian

2016-01-01

Highlights: • A novel hybrid approach is proposed for short-term wind speed prediction. • This method combines the parametric AR model with the non-parametric GPR model. • The relative importance of different inputs is considered. • Different types of covariance functions are considered and combined. • It can provide both accurate point forecasts and satisfactory prediction intervals. - Abstract: This paper proposes a hybrid model based on autoregressive (AR) model and Gaussian process regression (GPR) for probabilistic wind speed forecasting. In the proposed approach, the AR model is employed to capture the overall structure from wind speed series, and the GPR is adopted to extract the local structure. Additionally, automatic relevance determination (ARD) is used to take into account the relative importance of different inputs, and different types of covariance functions are combined to capture the characteristics of the data. The proposed hybrid model is compared with the persistence model, artificial neural network (ANN), and support vector machine (SVM) for one-step ahead forecasting, using wind speed data collected from three wind farms in China. The forecasting results indicate that the proposed method can not only improve point forecasts compared with other methods, but also generate satisfactory prediction intervals.

Programmable display of DNA-protein chimeras for controlling cell-hydrogel interactions via reversible intermolecular hybridization.

Science.gov (United States)

Zhang, Zhaoyang; Li, Shihui; Chen, Niancao; Yang, Cheng; Wang, Yong

2013-04-08

Extensive studies have been recently carried out to achieve dynamic control of cell-material interactions primarily through physicochemical stimulation. The purpose of this study was to apply reversible intermolecular hybridization to program cell-hydrogel interactions in physiological conditions based on DNA-antibody chimeras and complementary oligonucleotides. The results showed that DNA oligonucleotides could be captured to and released from the immobilizing DNA-functionalized hydrogels with high specificity via DNA hybridization. Accordingly, DNA-antibody chimeras were captured to the hydrogels, successfully inducing specific cell attachment. The cell attachment to the hydrogels reached the plateau at approximately half an hour after the functionalized hydrogels and the cells were incubated together. The attached cells were rapidly released from the bound hydrogels when triggering complementary oligonucleotides were introduced to the system. However, the capability of the triggering complementary oligonucleotides in releasing cells was affected by the length of intermolecular hybridization. The length needed to be at least more than 20 base pairs in the current experimental setting. Notably, because the procedure of intermolecular hybridization did not involve any harsh condition, the released cells maintained the same viability as that of the cultured cells. The functionalized hydrogels also exhibited the potential to catch and release cells repeatedly. Therefore, this study demonstrates that it is promising to regulate cell-material interactions dynamically through the DNA-programmed display of DNA-protein chimeras.

Investigation of CO2 capture mechanisms of liquid-like nanoparticle organic hybrid materials via structural characterization

KAUST Repository

Park, Youngjune; Decatur, John; Lin, Kun-Yi Andrew; Park, Ah-Hyung Alissa

2011-01-01

Nanoparticle organic hybrid materials (NOHMs) have been recently developed that comprise an oligomeric or polymeric canopy tethered to surface-modified nanoparticles via ionic or covalent bonds. It has already been shown that the tunable nature

ANALYSIS OF SOIL AND DUST SAMPLES FOR POLYCHLORINATED BIPHENYLS BY ENZYME LINKED IMMUNOSORBENT ASSAY (ELISA)

Science.gov (United States)

An inhibition enzyme-linked immunosorbent assay (ELISA) was used to determine polychlorinated biphenyls (PCBs) in house dust and soil. Soil and house dust samples were analyzed for PCB by both gas chromatography/electron capture detection (GC/ECD) and ELISA methods. A correlati...

Development and field evaluation of the sentinel mosquito arbovirus capture kit (SMACK)

OpenAIRE

Johnson, Brian J.; Kerlin, Tim; Hall-Mendelin, Sonja; van den Hurk, Andrew F.; Cortis, Giles; Doggett, Stephen L.; Toi, Cheryl; Fall, Ken; McMahon, Jamie L.; Townsend, Michael; Ritchie, Scott A.

2015-01-01

Background Although sentinel animals are used successfully throughout the world to monitor arbovirus activity, ethical considerations and cross-reactions in serological assays highlight the importance of developing viable alternatives. Here we outline the development of a passive sentinel mosquito arbovirus capture kit (SMACK) that allows for the detection of arboviruses on honey-baited nucleic acid preservation cards (Flinders Technology Associates; FTA®) and has a similar trap efficacy as s...

Localized surface plasmon resonance (LSPR) study of DNA hybridization at single nanoparticle transducers

International Nuclear Information System (INIS)

Schneider, T.; Jahr, N.; Jatschka, J.; Csaki, A.; Stranik, O.; Fritzsche, W.

2013-01-01

The effect of DNA–DNA interaction on the localized surface plasmon resonance of single 80 nm gold nanoparticles is studied. Therefore, both the attachment of the capture DNA strands at the particle surface and the sequence-specific DNA binding (hybridization) of analyte DNA to the immobilized capture DNA is subject of investigations. The influence of substrate attachment chemistry, the packing density of DNA as controlled by an assisting layer of smaller molecules, and the distance as increased by a linker on the LSPR efficiency is investigated. The resulting changes in signal can be related to a higher hybridization efficiency of the analyte DNA to the immobilized capture DNA. The subsequent attachment of additional DNA strands to this system is studied, which allows for a multiple step detection of binding and an elucidation of the resulting resonance shifts. The detection limit was determined for the utilized DNA system by incubation with various concentration of analyte DNA. Although the method allows for a marker-free detection, we show that additional markers such as 20 nm gold particle labels increase the signal and thereby the sensitivity significantly. The study of resonance shift for various DNA lengths revealed that the resonance shift per base is stronger for shorter DNA molecules (20 bases) as compared to longer ones (46 bases).

The indicating FTA elute cartridge a solid sample carrier to detect high-risk HPV and high-grade cervical lesions

NARCIS (Netherlands)

Bie, R.P. de; Schmeink, C.E.; Bakkers, J.M.J.E.; Snijders, P.J.L.M.; Quint, W.G.V.; Massuger, L.F.A.G.; Bekkers, R.L.M.; Melchers, W.J.G.

2011-01-01

The clinically validated high-risk human papillomavirus (hrHPV) Hybrid Capture 2 (HC2) and GP5+/6+-PCR assays were analyzed on an Indicating FTA Elute cartridge (FTA cartridge). The FTA cartridge is a solid dry carrier that allows safe transport of cervical samples. FTA cartridge samples were

Simultaneous detection of viruses and Toxoplasma gondii in cerebrospinal fluid specimens by multiplex polymerase chain reaction-based reverse hybridization assay.

Science.gov (United States)

Del Prete, Raffaele; Di Taranto, Anna Maria; Lipsi, Maria Rosaria; Natalicchio, Maria Iole; Antonetti, Raffaele; Miragliotta, Giuseppe

2009-04-01

The lack of rapidity and the low sensitivity and specificity of traditional laboratory methods limits their usefulness in the laboratory diagnosis of viral central nervous system (CNS) infections. This study describes the use of a commercially available multiplex polymerase chain reaction (mPCR)-based reverse hybridization assay (RHA) for the simultaneous detection of the genomes of 8 viruses and Toxoplasma gondii in cerebrospinal fluids (CSF) from 181 patients suspected of having viral meningitis. Twenty-two/181 (12.15%) CSF samples resulted positive by mPCR. Eighteen/22 were positive for 1 viral pathogen, whereas a dual infection was detected in 4/22 samples. Epstein-Barr virus (EBV) was the most commonly detected virus (6/22), followed by herpes simplex virus type-1 (HSV-1) (5/22) and -2 (HSV-2) (4/22). Cytomegalovirus (CMV), human herpesvirus-6 (HHV-6), and Epstein-Barr virus (EBV) were detected in 1 specimen each. Two CSF samples were co-infected by HSV-1/HSV-2, 1 sample by HHV-6/T. gondii, and 1 sample by EBV/EV, respectively. Our data support the usefulness of mPCR as a rapid molecular method for the simultaneous detection of major viral pathogens and T. gondii in aseptic meningitis also to allow the earlier application of specific antiviral therapy.

A flow cytometric assay technology based on quantum dots-encoded beads

International Nuclear Information System (INIS)

Wang Haiqiao; Liu Tiancai; Cao Yuancheng; Huang Zhenli; Wang Jianhao; Li Xiuqing; Zhao Yuandi

2006-01-01

A flow cytometric detecting technology based on quantum dots (QDs)-encoded beads has been described. Using this technology, several QDs-encoded beads with different code were identified effectively, and the target molecule (DNA sequence) in solution was also detected accurately by coupling to its complementary sequence probed on QDs-encoded beads through DNA hybridization assay. The resolution of this technology for encoded beads is resulted from two longer wavelength fluorescence identification signals (yellow and red fluorescent signals of QDs), and the third shorter wavelength fluorescence signal (green reporting signal of fluorescein isothiocyanate (FITC)) for the determination of reaction between probe and target. In experiment, because of QDs' unique optical character, only one excitation light source was needed to excite the QDs and probe dye FITC synchronously comparing with other flow cytometric assay technology. The results show that this technology has present excellent repeatability and good accuracy. It will become a promising multiple assay platform in various application fields after further improvement

Forecasting Inflow and Outflow of Money Currency in East Java Using a Hybrid Exponential Smoothing and Calendar Variation Model

Science.gov (United States)

Susanti, Ana; Suhartono; Jati Setyadi, Hario; Taruk, Medi; Haviluddin; Pamilih Widagdo, Putut

2018-03-01

Money currency availability in Bank Indonesia can be examined by inflow and outflow of money currency. The objective of this research is to forecast the inflow and outflow of money currency in each Representative Office (RO) of BI in East Java by using a hybrid exponential smoothing based on state space approach and calendar variation model. Hybrid model is expected to generate more accurate forecast. There are two studies that will be discussed in this research. The first studies about hybrid model using simulation data that contain pattern of trends, seasonal and calendar variation. The second studies about the application of a hybrid model for forecasting the inflow and outflow of money currency in each RO of BI in East Java. The first of results indicate that exponential smoothing model can not capture the pattern calendar variation. It results RMSE values 10 times standard deviation of error. The second of results indicate that hybrid model can capture the pattern of trends, seasonal and calendar variation. It results RMSE values approaching the standard deviation of error. In the applied study, the hybrid model give more accurate forecast for five variables : the inflow of money currency in Surabaya, Malang, Jember and outflow of money currency in Surabaya and Kediri. Otherwise, the time series regression model yields better for three variables : outflow of money currency in Malang, Jember and inflow of money currency in Kediri.

Cost-effective multiplexing before capture allows screening of 25 000 clinically relevant SNPs in childhood acute lymphoblastic leukemia

DEFF Research Database (Denmark)

Wesolowska, Agata; Dalgaard, M. D.; Borst, L.

2011-01-01

designed a cost-effective, high-throughput capture assay of â¼25â000 clinically relevant SNPs, and demonstrated that multiple samples can be tagged and pooled before genome capture in targeted enrichment with a sufficient sequencing depth for genotyping. This multiplexed, targeted sequencing method allows...... exploration of the impact of pharmacogenetics on efficacy and toxicity in childhood ALL treatment, which will be of importance for personalized chemotherapy.Leukemia advance online publication, 18 March 2011; doi:10.1038/leu.2011.32....

OpenComet: An automated tool for comet assay image analysis

Directory of Open Access Journals (Sweden)

Benjamin M. Gyori

2014-01-01

Full Text Available Reactive species such as free radicals are constantly generated in vivo and DNA is the most important target of oxidative stress. Oxidative DNA damage is used as a predictive biomarker to monitor the risk of development of many diseases. The comet assay is widely used for measuring oxidative DNA damage at a single cell level. The analysis of comet assay output images, however, poses considerable challenges. Commercial software is costly and restrictive, while free software generally requires laborious manual tagging of cells. This paper presents OpenComet, an open-source software tool providing automated analysis of comet assay images. It uses a novel and robust method for finding comets based on geometric shape attributes and segmenting the comet heads through image intensity profile analysis. Due to automation, OpenComet is more accurate, less prone to human bias, and faster than manual analysis. A live analysis functionality also allows users to analyze images captured directly from a microscope. We have validated OpenComet on both alkaline and neutral comet assay images as well as sample images from existing software packages. Our results show that OpenComet achieves high accuracy with significantly reduced analysis time.

Optimized mtDNA Control Region Primer Extension Capture Analysis for Forensically Relevant Samples and Highly Compromised mtDNA of Different Age and Origin

Directory of Open Access Journals (Sweden)

Mayra Eduardoff

2017-09-01

Full Text Available The analysis of mitochondrial DNA (mtDNA has proven useful in forensic genetics and ancient DNA (aDNA studies, where specimens are often highly compromised and DNA quality and quantity are low. In forensic genetics, the mtDNA control region (CR is commonly sequenced using established Sanger-type Sequencing (STS protocols involving fragment sizes down to approximately 150 base pairs (bp. Recent developments include Massively Parallel Sequencing (MPS of (multiplex PCR-generated libraries using the same amplicon sizes. Molecular genetic studies on archaeological remains that harbor more degraded aDNA have pioneered alternative approaches to target mtDNA, such as capture hybridization and primer extension capture (PEC methods followed by MPS. These assays target smaller mtDNA fragment sizes (down to 50 bp or less, and have proven to be substantially more successful in obtaining useful mtDNA sequences from these samples compared to electrophoretic methods. Here, we present the modification and optimization of a PEC method, earlier developed for sequencing the Neanderthal mitochondrial genome, with forensic applications in mind. Our approach was designed for a more sensitive enrichment of the mtDNA CR in a single tube assay and short laboratory turnaround times, thus complying with forensic practices. We characterized the method using sheared, high quantity mtDNA (six samples, and tested challenging forensic samples (n = 2 as well as compromised solid tissue samples (n = 15 up to 8 kyrs of age. The PEC MPS method produced reliable and plausible mtDNA haplotypes that were useful in the forensic context. It yielded plausible data in samples that did not provide results with STS and other MPS techniques. We addressed the issue of contamination by including four generations of negative controls, and discuss the results in the forensic context. We finally offer perspectives for future research to enable the validation and accreditation of the PEC MPS

Identification and Differentiation of Verticillium Species and V. longisporum Lineages by Simplex and Multiplex PCR Assays

Science.gov (United States)

Inderbitzin, Patrik; Davis, R. Michael; Bostock, Richard M.; Subbarao, Krishna V.

2013-01-01

Accurate species identification is essential for effective plant disease management, but is challenging in fungi including Verticillium sensu stricto (Ascomycota, Sordariomycetes, Plectosphaerellaceae), a small genus of ten species that includes important plant pathogens. Here we present fifteen PCR assays for the identification of all recognized Verticillium species and the three lineages of the diploid hybrid V. longisporum. The assays were based on DNA sequence data from the ribosomal internal transcribed spacer region, and coding and non-coding regions of actin, elongation factor 1-alpha, glyceraldehyde-3-phosphate dehydrogenase and tryptophan synthase genes. The eleven single target (simplex) PCR assays resulted in amplicons of diagnostic size for V. alfalfae, V. albo-atrum, V. dahliae including V. longisporum lineage A1/D3, V. isaacii, V. klebahnii, V. nonalfalfae, V. nubilum, V. tricorpus, V. zaregamsianum, and Species A1 and Species D1, the two undescribed ancestors of V. longisporum. The four multiple target (multiplex) PCR assays simultaneously differentiated the species or lineages within the following four groups: Verticillium albo-atrum, V. alfalfae and V. nonalfalfae; Verticillium dahliae and V. longisporum lineages A1/D1, A1/D2 and A1/D3; Verticillium dahliae including V. longisporum lineage A1/D3, V. isaacii, V. klebahnii and V. tricorpus; Verticillium isaacii, V. klebahnii and V. tricorpus. Since V. dahliae is a parent of two of the three lineages of the diploid hybrid V. longisporum, no simplex PCR assay is able to differentiate V. dahliae from all V. longisporum lineages. PCR assays were tested with fungal DNA extracts from pure cultures, and were not evaluated for detection and quantification of Verticillium species from plant or soil samples. The DNA sequence alignments are provided and can be used for the design of additional primers. PMID:23823707

A Fully Automated High-Throughput Zebrafish Behavioral Ototoxicity Assay.

Science.gov (United States)

Todd, Douglas W; Philip, Rohit C; Niihori, Maki; Ringle, Ryan A; Coyle, Kelsey R; Zehri, Sobia F; Zabala, Leanne; Mudery, Jordan A; Francis, Ross H; Rodriguez, Jeffrey J; Jacob, Abraham

2017-08-01

Zebrafish animal models lend themselves to behavioral assays that can facilitate rapid screening of ototoxic, otoprotective, and otoregenerative drugs. Structurally similar to human inner ear hair cells, the mechanosensory hair cells on their lateral line allow the zebrafish to sense water flow and orient head-to-current in a behavior called rheotaxis. This rheotaxis behavior deteriorates in a dose-dependent manner with increased exposure to the ototoxin cisplatin, thereby establishing itself as an excellent biomarker for anatomic damage to lateral line hair cells. Building on work by our group and others, we have built a new, fully automated high-throughput behavioral assay system that uses automated image analysis techniques to quantify rheotaxis behavior. This novel system consists of a custom-designed swimming apparatus and imaging system consisting of network-controlled Raspberry Pi microcomputers capturing infrared video. Automated analysis techniques detect individual zebrafish, compute their orientation, and quantify the rheotaxis behavior of a zebrafish test population, producing a powerful, high-throughput behavioral assay. Using our fully automated biological assay to test a standardized ototoxic dose of cisplatin against varying doses of compounds that protect or regenerate hair cells may facilitate rapid translation of candidate drugs into preclinical mammalian models of hearing loss.

Enhanced CO{sub 2} capture on graphene via N, S dual-doping

Energy Technology Data Exchange (ETDEWEB)

Li, Jieyuan; Hou, Meiling [College of Architecture and Environment, Sichuan University (China); Chen, Yanqiu [Institute of New Energy and Low Carbon Technology, Sichuan University (China); Cen, Wanglai [Institute of New Energy and Low Carbon Technology, Sichuan University (China); National Engineering Research Center for Flue Gas Desulfurization (China); Chu, Yinghao, E-mail: chuyinghao@scu.edu.cn [College of Architecture and Environment, Sichuan University (China); National Engineering Research Center for Flue Gas Desulfurization (China); Yin, Shi, E-mail: yinshi_scu@foxmail.com [College of Architecture and Environment, Sichuan University (China)

2017-03-31

Highlights: • Sluggish conjugated π bonds of graphene should be weakened to promote adsorption activity. • A charge delivery channel along S → N → CO{sub 2} path should be prior responsible for the enhancement of CO{sub 2} capture on graphene. • Applicative temperature range of graphene-based adsorbents for CO{sub 2} capture is extend to about 100 °C via N, S dual-doping. - Abstract: N, S doped graphene-based materials have been recently recognized as promising adsorbents for CO{sub 2} capture, but understanding of the adsorption mechanism at the atomic level is still limited. Herein, the local structures and promotion mechanism of CO{sub 2} capture by N, S doped graphene were investigated by combining density functional theory and ab initio thermodynamics. A single vacancy defected graphene involving N, S dual-doping was found to be a superior adsorbent for CO{sub 2} capture under mild conditions (<100 °C, 1 atm). The enhanced CO{sub 2} adsorption performance should be ascribed to a charge delivery channel along the S → N → CO{sub 2} path, leading to extra charge transfer from graphene to CO{sub 2}. It is worth mentioning that the extra charge transfer was stimulated by the unique sp{sup 2} hybridization of pyridine N and further enhanced by S in N, S dual-doped graphene. A possible mechanism has been proposed to explain the high adsorption performance of CO{sub 2} by N, S dual-doped graphene, which offers insights for the design of new graphene-based adsorbents.

Rationalizing and advancing the 3-MPBA SERS sandwich assay for rapid detection of bacteria in environmental and food matrices.

Science.gov (United States)

Pearson, Brooke; Mills, Alexander; Tucker, Madeline; Gao, Siyue; McLandsborough, Lynne; He, Lili

2018-06-01

Bacterial foodborne illness continues to be a pressing issue in our food supply. Rapid detection methods are needed for perishable foods due to their short shelf lives and significant contribution to foodborne illness. Previously, a sensitive and reliable surface-enhanced Raman spectroscopy (SERS) sandwich assay based on 3-mercaptophenylboronic acid (3-MBPA) as a capturer and indicator molecule was developed for rapid bacteria detection. In this study, we explored the advantages and constraints of this assay over the conventional aerobic plate count (APC) method and further developed methods for detection in real environmental and food matrices. The SERS sandwich assay was able to detect environmental bacteria in pond water and on spinach leaves at higher levels than the APC method. In addition, the SERS assay appeared to have higher sensitivity to quantify bacteria in the stationary phase. On the other hand, the APC method was more sensitive to cell viability. Finally, a method to detect bacteria in a challenging high-sugar juice matrix was developed to enhance bacteria capture. This study advanced the SERS technique for real applications in environment and food matrices. Copyright © 2017 Elsevier Ltd. All rights reserved.

Comparison of Fluorescence In Situ Hybridization and Chromogenic In Situ Hybridization for Low and High Throughput HER2 Genetic Testing

Science.gov (United States)

Poulsen, Tim S.; Espersen, Maiken L. M.; Kofoed, Vibeke; Dabetic, Tanja; Høgdall, Estrid; Balslev, Eva

2013-01-01

The purpose was to evaluate and compare 5 different HER2 genetic assays with different characteristics that could affect the performance to analyze the human epidermal growth factor 2 (HER2) gene copy number under low and high throughput conditions. The study included 108 tissue samples from breast cancer patients with HER2 immunohistochemistry (IHC) results scored as 0/1+, 2+, and 3+. HER2 genetic status was analysed using chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH). Scoring results were documented through digital image analysis. The cancer region of interest was identified from a serial H&E stained slide following tissue cores were transferred to a tissue microarrays (TMA). When using TMA in a routine flow, all patients will be tested for HER2 status with IHC followed by CISH or FISH, thereby providing individual HER2 results. In conclusion, our results show that the differences between the HER2 genetic assays do not have an effect on the analytic performance and the CISH technology is superior to high throughput HER2 genetic testing due to scanning speed, while the IQ-FISH may still be a choice for fast low throughput HER2 genetic testing. PMID:24383005

Cloned polynucleotide and synthetic oligonucleotide probes used in colony hybridization are equally efficient in the identification of enterotoxigenic Escherichia coli

International Nuclear Information System (INIS)

Sommerfelt, H.; Kalland, K.H.; Raj, P.; Moseley, S.L.; Bhan, M.K.; Bjorvatn, B.

1988-01-01

Restriction endonuclease-generated polynucleotide and synthetically produced oligonucleotide gene probes used in colony hybridization assays proved to be efficient for the detection and differentiation of enterotoxigenic Escherichia coli. To compare their relative efficiencies, these two sets of probes were radiolabeled with 32 P and were applied to 74 strains of E. coli with known enterotoxin profiles and to 156 previously unexamined E. coli isolates. The enterotoxigenic bacteria Vibrio cholerae O1, Vibrio cholerae non-O1 (NAG), Yersinia enterocolitica, and E. coli harboring the plasmid vectors of the polynucleotide gene probes were examined for further evaluation of probe specificity. The two classes of probes showed a perfect concordance in their specific detection and differentiation of enterotoxigenic E. coli. In the analysis of six strains, the signal strength on autoradiography after hybridization with oligonucleotides was weaker than that obtained after hybridization with polynucleotide probes. The probes did not hybridize with DNA from V. cholerae O1, V. cholerae non-O1 (NAG), or Y. enterocolitica. The strains of E. coli harboring the plasmid vectors of the polynucleotide gene probes were, likewise, negative in the hybridization assays

Cloned polynucleotide and synthetic oligonucleotide probes used in colony hybridization are equally efficient in the identification of enterotoxigenic Escherichia coli

Energy Technology Data Exchange (ETDEWEB)

Sommerfelt, H.; Kalland, K.H.; Raj, P.; Moseley, S.L.; Bhan, M.K.; Bjorvatn, B.

1988-11-01

Restriction endonuclease-generated polynucleotide and synthetically produced oligonucleotide gene probes used in colony hybridization assays proved to be efficient for the detection and differentiation of enterotoxigenic Escherichia coli. To compare their relative efficiencies, these two sets of probes were radiolabeled with /sup 32/P and were applied to 74 strains of E. coli with known enterotoxin profiles and to 156 previously unexamined E. coli isolates. The enterotoxigenic bacteria Vibrio cholerae O1, Vibrio cholerae non-O1 (NAG), Yersinia enterocolitica, and E. coli harboring the plasmid vectors of the polynucleotide gene probes were examined for further evaluation of probe specificity. The two classes of probes showed a perfect concordance in their specific detection and differentiation of enterotoxigenic E. coli. In the analysis of six strains, the signal strength on autoradiography after hybridization with oligonucleotides was weaker than that obtained after hybridization with polynucleotide probes. The probes did not hybridize with DNA from V. cholerae O1, V. cholerae non-O1 (NAG), or Y. enterocolitica. The strains of E. coli harboring the plasmid vectors of the polynucleotide gene probes were, likewise, negative in the hybridization assays.

Multifunctional Nanotechnology-Enabled Sensors for Rapid Capture and Detection of Pathogens.

Science.gov (United States)

Mustafa, Fatima; Hassan, Rabeay Y A; Andreescu, Silvana

2017-09-15

Nanomaterial-based sensing approaches that incorporate different types of nanoparticles (NPs) and nanostructures in conjunction with natural or synthetic receptors as molecular recognition elements provide opportunities for the design of sensitive and selective assays for rapid detection of contaminants. This review summarizes recent advancements over the past ten years in the development of nanotechnology-enabled sensors and systems for capture and detection of pathogens. The most common types of nanostructures and NPs, their modification with receptor molecules and integration to produce viable sensing systems with biorecognition, amplification and signal readout are discussed. Examples of all-in-one systems that combine multifunctional properties for capture, separation, inactivation and detection are also provided. Current trends in the development of low-cost instrumentation for rapid assessment of food contamination are discussed as well as challenges for practical implementation and directions for future research.

Identification of hybrid node and link communities in complex networks.

Science.gov (United States)

He, Dongxiao; Jin, Di; Chen, Zheng; Zhang, Weixiong

2015-03-02

Identifying communities in complex networks is an effective means for analyzing complex systems, with applications in diverse areas such as social science, engineering, biology and medicine. Finding communities of nodes and finding communities of links are two popular schemes for network analysis. These schemes, however, have inherent drawbacks and are inadequate to capture complex organizational structures in real networks. We introduce a new scheme and an effective approach for identifying complex mixture structures of node and link communities, called hybrid node-link communities. A central piece of our approach is a probabilistic model that accommodates node, link and hybrid node-link communities. Our extensive experiments on various real-world networks, including a large protein-protein interaction network and a large network of semantically associated words, illustrated that the scheme for hybrid communities is superior in revealing network characteristics. Moreover, the new approach outperformed the existing methods for finding node or link communities separately.

Hybrid Vehicle Technologies and their potential for reducing oil use

Science.gov (United States)

German, John

2006-04-01

Vehicles with hybrid gasoline-electric powertrains are starting to gain market share. Current hybrid vehicles add an electric motor, battery pack, and power electronics to the conventional powertrain. A variety of engine/motor configurations are possible, each with advantages and disadvantages. In general, efficiency is improved due to engine shut-off at idle, capture of energy during deceleration that is normally lost as heat in the brakes, downsizing of the conventional engine, and, in some cases, propulsion on the electric motor alone. Ongoing increases in hybrid market share are dependent on cost reduction, especially the battery pack, efficiency synergies with other vehicle technologies, use of the high electric power to provide features desired by customers, and future fuel price and availability. Potential barriers include historically low fuel prices, high discounting of the fuel savings by new vehicle purchasers, competing technologies, and tradeoffs with other factors desired by customers, such as performance, utility, safety, and luxury features.

Identification of hybrid node and link communities in complex networks

Science.gov (United States)

He, Dongxiao; Jin, Di; Chen, Zheng; Zhang, Weixiong

2015-03-01

Identifying communities in complex networks is an effective means for analyzing complex systems, with applications in diverse areas such as social science, engineering, biology and medicine. Finding communities of nodes and finding communities of links are two popular schemes for network analysis. These schemes, however, have inherent drawbacks and are inadequate to capture complex organizational structures in real networks. We introduce a new scheme and an effective approach for identifying complex mixture structures of node and link communities, called hybrid node-link communities. A central piece of our approach is a probabilistic model that accommodates node, link and hybrid node-link communities. Our extensive experiments on various real-world networks, including a large protein-protein interaction network and a large network of semantically associated words, illustrated that the scheme for hybrid communities is superior in revealing network characteristics. Moreover, the new approach outperformed the existing methods for finding node or link communities separately.

Rapid detection of EBOLA VP40 in microchip immunofiltration assay

Science.gov (United States)

Miethe, Peter; Gary, Dominik; Hlawatsch, Nadine; Gad, Anne-Marie

2015-05-01

In the spring of 2014, the Ebola virus (EBOV) strain Zaire caused a dramatic outbreak in several regions of West Africa. The RT-PCR and antigen capture diagnostic proved to be effective for detecting EBOV in blood and serum. In this paper, we present data of a rapid antigen capture test for the detection of VP40. The test was performed in a microfluidic chip for immunofiltration analysis. The chip integrates all necessary assay components. The analytical sensitivity of the rapid test was 8 ng/ml for recombinant VP40. In serum and whole blood samples spiked with virus culture material, the detection limit was 2.2 x 102 PFU/ml. The performance data of the rapid test (15 min) are comparable to that of the VP40 laboratory ELISA.

4-Aminoquinoline-pyrimidine hybrids: synthesis, antimalarial activity, heme binding and docking studies.

Science.gov (United States)

Kumar, Deepak; Khan, Shabana I; Tekwani, Babu L; Ponnan, Prija; Rawat, Diwan S

2015-01-07

A series of novel 4-aminoquinoline-pyrimidine hybrids has been synthesized and evaluated for their antimalarial activity. Several compounds showed promising in vitro antimalarial activity against both CQ-sensitive and CQ-resistant strains with high selectivity index. All the compounds were found to be non-toxic to the mammalian cell lines. Selected compound 7g exhibited significant suppression of parasitemia in the in vivo assay. The heme binding studies were conducted to determine the mode of action of these hybrid molecules. These compounds form a stable 1:1 complex with hematin suggesting that heme may be one of the possible targets of these hybrids. The interaction of these conjugate hybrids was also investigated by the molecular docking studies in the binding site of PfDHFR. The pharmacokinetic property analysis of best active compounds was also studied using ADMET prediction. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Hybrid CFD/CAA Modeling for Liftoff Acoustic Predictions

Science.gov (United States)

Strutzenberg, Louise L.; Liever, Peter A.

2011-01-01

This paper presents development efforts at the NASA Marshall Space flight Center to establish a hybrid Computational Fluid Dynamics and Computational Aero-Acoustics (CFD/CAA) simulation system for launch vehicle liftoff acoustics environment analysis. Acoustic prediction engineering tools based on empirical jet acoustic strength and directivity models or scaled historical measurements are of limited value in efforts to proactively design and optimize launch vehicles and launch facility configurations for liftoff acoustics. CFD based modeling approaches are now able to capture the important details of vehicle specific plume flow environment, identifY the noise generation sources, and allow assessment of the influence of launch pad geometric details and sound mitigation measures such as water injection. However, CFD methodologies are numerically too dissipative to accurately capture the propagation of the acoustic waves in the large CFD models. The hybrid CFD/CAA approach combines the high-fidelity CFD analysis capable of identifYing the acoustic sources with a fast and efficient Boundary Element Method (BEM) that accurately propagates the acoustic field from the source locations. The BEM approach was chosen for its ability to properly account for reflections and scattering of acoustic waves from launch pad structures. The paper will present an overview of the technology components of the CFD/CAA framework and discuss plans for demonstration and validation against test data.

Comparison of clinical samples for visceral Leishmaniasis diagnosis in asymptomatic dogs by PCR hybridization

Energy Technology Data Exchange (ETDEWEB)

Ferreira, Sidney A.; Ituassu, Leonardo T.; Melo, Maria N. [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Parasitologia], e-mail: saninoalmeida@gmail.com, e-mail: Itituassu@yahoo.com.br, e-mail: melo@icb.ufmg.br; Leite, Rodrigo S.; Andrade, Antero S.R. [Centro de Desenvolvimento da Tecnologia Nuclear (CDTN-CNEN/MG), Belo Horizonte, MG (Brazil)], e-mail: rleite2005@gmail.com, e-mail: antero@cdtn.br

2009-07-01

The canine visceral leishmaniasis (CVL) diagnosis still represents a challenge because of complexity of this disease. The aim of present study was to compare different clinical samples for diagnosis of CVL by Polymerase Chain Reaction (PCR) combined with hybridization of {sup 32}P labeled probes. Bone marrow (BM), skin biopsy (SB), peripheral blood (PB) and conjunctival swab (CS) were used in this work. With this purpose 40 asymptomatic dogs, all positive by parasitological test, were obtained. From each animal were collected SB with sterile punches from ear internal surface, 1.0 mL of PB, BM aspirates from sternum and CS from both lower eyelid. Each clinical sample was submitted to suitable DNA purification process and PCR-hybridization assays. The positive results obtained with PCR were 55%, 25%, 30% and 22.5% for CS, BM, SB and PB respectively while the PCR followed by hybridization showed a positivity of 87.5%, 50%, 45% and 27.5% respectively. The hybridization assay was able to increase the PCR positivity in all kinds of clinical samples. The best performance was obtained using CS samples. We concluded that the PCR associated with DNA radioactive probes was a very sensitive tool for diagnosis of CVL in asymptomatic dogs and the CS has an important potential for regular screening of dogs. (author)

Assessment of a New Lower-Cost Real-Time PCR Assay for Detection of High-Risk Human Papillomavirus: Useful for Cervical Screening in Limited-Resource Settings?

Science.gov (United States)

Fokom Domgue, Joel; Schiffman, Mark; Wentzensen, Nicolas H; Gage, Julia C; Castle, Philip E; Raine-Bennett, Tina R; Fetterman, Barbara; Lorey, Thomas; Poitras, Nancy E; Befano, Brian; Xie, Yi; Miachon, Lais S; Dean, Michael

2017-08-01

Inexpensive and easy-to-perform human papillomavirus (HPV) tests are needed for primary cervical cancer screening in lower-resource regions. In a convenience sample of 516 residual exfoliative cervical specimens from the Kaiser Permanente Northern California and U.S. National Cancer Institute Persistence and Progression Study, we assessed the agreement and clinical performance of a simple, inexpensive real-time PCR assay for the detection of 13 carcinogenic HPV types (the H13 assay; Hybribio, Hong Kong) that is marketed in limited-resource settings compared to previous testing by the Hybrid Capture 2 assay (HC2; Qiagen, Germantown, MD) and the Onclarity assay (BD Diagnostics, Sparks, MD). The test set was chosen to include many HPV-positive specimens. The reference standard was a combination of HC2 and Onclarity results for HPV detection and histologic diagnosis of controls (less than cervical intraepithelial neoplasia grade 2 [

Enzyme-linked electrochemical DNA ligation assay using magnetic beads.

Science.gov (United States)

Stejskalová, Eva; Horáková, Petra; Vacek, Jan; Bowater, Richard P; Fojta, Miroslav

2014-07-01

DNA ligases are essential enzymes in all cells and have been proposed as targets for novel antibiotics. Efficient DNA ligase activity assays are thus required for applications in biomedical research. Here we present an enzyme-linked electrochemical assay based on two terminally tagged probes forming a nicked junction upon hybridization with a template DNA. Nicked DNA bearing a 5' biotin tag is immobilized on the surface of streptavidin-coated magnetic beads, and ligated product is detected via a 3' digoxigenin tag recognized by monoclonal antibody-alkaline phosphatase conjugate. Enzymatic conversion of napht-1-yl phosphate to napht-1-ol enables sensitive detection of the voltammetric signal on a pyrolytic graphite electrode. The technique was tested under optimal conditions and various situations limiting or precluding the ligation reaction (such as DNA substrates lacking 5'-phosphate or containing a base mismatch at the nick junction, or application of incompatible cofactor), and utilized for the analysis of the nick-joining activity of a range of recombinant Escherichia coli DNA ligase constructs. The novel technique provides a fast, versatile, specific, and sensitive electrochemical assay of DNA ligase activity.

Modeling, control, and simulation of grid connected intelligent hybrid battery/photovoltaic system using new hybrid fuzzy-neural method.

Science.gov (United States)

Rezvani, Alireza; Khalili, Abbas; Mazareie, Alireza; Gandomkar, Majid

2016-07-01

Nowadays, photovoltaic (PV) generation is growing increasingly fast as a renewable energy source. Nevertheless, the drawback of the PV system is its dependence on weather conditions. Therefore, battery energy storage (BES) can be considered to assist for a stable and reliable output from PV generation system for loads and improve the dynamic performance of the whole generation system in grid connected mode. In this paper, a novel topology of intelligent hybrid generation systems with PV and BES in a DC-coupled structure is presented. Each photovoltaic cell has a specific point named maximum power point on its operational curve (i.e. current-voltage or power-voltage curve) in which it can generate maximum power. Irradiance and temperature changes affect these operational curves. Therefore, the nonlinear characteristic of maximum power point to environment has caused to development of different maximum power point tracking techniques. In order to capture the maximum power point (MPP), a hybrid fuzzy-neural maximum power point tracking (MPPT) method is applied in the PV system. Obtained results represent the effectiveness and superiority of the proposed method, and the average tracking efficiency of the hybrid fuzzy-neural is incremented by approximately two percentage points in comparison to the conventional methods. It has the advantages of robustness, fast response and good performance. A detailed mathematical model and a control approach of a three-phase grid-connected intelligent hybrid system have been proposed using Matlab/Simulink. Copyright © 2016 ISA. Published by Elsevier Ltd. All rights reserved.

Comparison of RT-PCR-Dot blot hybridization based on radioisotope 32P with conventional RT-PCR and commercial ELISA Assays for blood screening of HIV-1

International Nuclear Information System (INIS)

Maria Lina R; Andi Yasmon

2011-01-01

There are many commercial ELISA and rapid test kits that have been used for blood screening; however, the kits can give false positive and negative results. Therefore, RT-PCR (Reverse Transcription Polymerase Chain Reaction) - Dot Blot Hybridization based on radioisotope 32 P (RDBR) method was developed in this research, to compare the method with the conventional RT-PCR and commercial ELISA Enzyme-Linked lmmunosorbent Assay) kit. This method is efficient for screening of large blood specimens and surveillance study. Eighty seven samples were used and serum of the samples were tested by ELISA to detect HIV-1. The HIV-l RNA genome was extracted from plasma samples and tested using the RT-PCR and RDBR methods. Of 87 samples that were tested, the rates of positive testing of the RT-PCR, the RDBR, and the ELISA were 71.26%, 74.71%, and 80.46%, respectively. The RDBR (a combination of RTPCR and dot blot hybridization) was more sensitive than conventional RT-PCR by showing 3.45% in increase number of positive specimens. The results showed that of 9 samples (10.34%) were negative RDBR and positive ELISA, while 4 samples (4.60%) were negative ELISA and positive RDBR. The two methods showed slightly difference in the results but further validation is still needed. However, RDBR has high potential as an alternative method for screening of blood in large quantities when compared to method of conventional RT-PCR and ELISA. (author)

The Lesser Antillean Iguana (Iguana delicatissima) on St. Eustatius: genetically depauperate and threatened by ongoing hybridization.

Science.gov (United States)

van den Burg, Matthijs P; Meirmans, Patrick G; Wagensveld, Tim van; Kluskens, Bart; Madden, Hannah; Welch, Mark E; Breeuwer, Johannes A J

2018-02-19

The Lesser Antillean Iguana (Iguana delicatissima) is an endangered species threatened by habitat loss and hybridization with non-native Green Iguanas (Iguana iguana). Iguanadelicatissima has been extirpated on several islands, and the Green Iguana has invaded most islands with extant populations. Information is essential to protect this species from extinction. We collected data on 293 iguanas including 17 juveniles from St. Eustasius, one of the few remaining I. delicatissima strongholds. Genetic data was leveraged to test for hybridization presence with the Green Iguana using both mitochondrial and nuclear genes, including 16 microsatellite loci. The microsatellites were also analysed to estimate genetic diversity, population structure and effective population size. Using molecular and morphological data we identified 286 I. delicatissima individuals captured during our first fieldwork effort, and seven non-native iguanas captured during a second effort, showing hybridization occurs within this population. Comparing homologous microsatellites used in studies on Dominica and Chancel, the I. delicatissima population on St. Eustatius has extremely low genetic diversity (HO=0.051; HE=0.057), suggesting this population is genetically depauperate. Furthermore, there is significant evidence for inbreeding (FIS=0.12) and weak spatial genetic structure (FST=0.021, p=0.002) within this population. Besides immediate threats including hybridization, this population's low genetic diversity, presence of physiological abnormalities and low recruitment could indicate presence of inbreeding depression that threatens its long-term survival. We conclude there is a continued region-wide threat to I. delicatissima and highlight the need for immediate conservation action to stop the continuing spread of Green Iguanas and to eliminate hybridization from St. Eustatius.

High-sensitivity HLA typing by Saturated Tiling Capture Sequencing (STC-Seq).

Science.gov (United States)

Jiao, Yang; Li, Ran; Wu, Chao; Ding, Yibin; Liu, Yanning; Jia, Danmei; Wang, Lifeng; Xu, Xiang; Zhu, Jing; Zheng, Min; Jia, Junling

2018-01-15

Highly polymorphic human leukocyte antigen (HLA) genes are responsible for fine-tuning the adaptive immune system. High-resolution HLA typing is important for the treatment of autoimmune and infectious diseases. Additionally, it is routinely performed for identifying matched donors in transplantation medicine. Although many HLA typing approaches have been developed, the complexity, low-efficiency and high-cost of current HLA-typing assays limit their application in population-based high-throughput HLA typing for donors, which is required for creating large-scale databases for transplantation and precision medicine. Here, we present a cost-efficient Saturated Tiling Capture Sequencing (STC-Seq) approach to capturing 14 HLA class I and II genes. The highly efficient capture (an approximately 23,000-fold enrichment) of these genes allows for simplified allele calling. Tests on five genes (HLA-A/B/C/DRB1/DQB1) from 31 human samples and 351 datasets using STC-Seq showed results that were 98% consistent with the known two sets of digitals (field1 and field2) genotypes. Additionally, STC can capture genomic DNA fragments longer than 3 kb from HLA loci, making the library compatible with the third-generation sequencing. STC-Seq is a highly accurate and cost-efficient method for HLA typing which can be used to facilitate the establishment of population-based HLA databases for the precision and transplantation medicine.

A high-throughput in vitro ring assay for vasoactivity using magnetic 3D bioprinting

Science.gov (United States)

Tseng, Hubert; Gage, Jacob A.; Haisler, William L.; Neeley, Shane K.; Shen, Tsaiwei; Hebel, Chris; Barthlow, Herbert G.; Wagoner, Matthew; Souza, Glauco R.

2016-01-01

Vasoactive liabilities are typically assayed using wire myography, which is limited by its high cost and low throughput. To meet the demand for higher throughput in vitro alternatives, this study introduces a magnetic 3D bioprinting-based vasoactivity assay. The principle behind this assay is the magnetic printing of vascular smooth muscle cells into 3D rings that functionally represent blood vessel segments, whose contraction can be altered by vasodilators and vasoconstrictors. A cost-effective imaging modality employing a mobile device is used to capture contraction with high throughput. The goal of this study was to validate ring contraction as a measure of vasoactivity, using a small panel of known vasoactive drugs. In vitro responses of the rings matched outcomes predicted by in vivo pharmacology, and were supported by immunohistochemistry. Altogether, this ring assay robustly models vasoactivity, which could meet the need for higher throughput in vitro alternatives. PMID:27477945

Evaluation of a competitive enzyme-linked immunosorbent assay for measurements of soluble HLA-G protein

DEFF Research Database (Denmark)

Rasmussen, M; Dahl, M; Buus, S

2014-01-01

. We report a novel method, a competitive immunoassay, for measuring HLA-G5/sHLA-G1 in biological fluids. The sHLA-G immunoassay is based upon a competitive enzyme-linked immunosorbent assay (ELISA) principle. It includes a recombinant sHLA-G1 protein in complex with β2-microglobulin and a peptide...... as a standard, biotinylated recombinant sHLA-G1 as an indicator, and the MEM-G/9 anti-HLA-G monoclonal antibody (mAb) as the capture antibody. The specificity and sensitivity of the assay were evaluated. Testing with different recombinant HLA class I proteins and different anti-HLA class I mAbs showed....../ml. An intra-assay coefficient of variation (CV) of 15.5% at 88 ng/ml and an inter-assay CV of 23.1% at 39 ng/ml were determined. An assay based on the competitive sHLA-G ELISA may be important for measurements of sHLA-G proteins in several conditions: assisted reproduction, organ transplantation, cancer...

An Improved Iris Recognition Algorithm Based on Hybrid Feature and ELM

Science.gov (United States)

Wang, Juan

2018-03-01

The iris image is easily polluted by noise and uneven light. This paper proposed an improved extreme learning machine (ELM) based iris recognition algorithm with hybrid feature. 2D-Gabor filters and GLCM is employed to generate a multi-granularity hybrid feature vector. 2D-Gabor filter and GLCM feature work for capturing low-intermediate frequency and high frequency texture information, respectively. Finally, we utilize extreme learning machine for iris recognition. Experimental results reveal our proposed ELM based multi-granularity iris recognition algorithm (ELM-MGIR) has higher accuracy of 99.86%, and lower EER of 0.12% under the premise of real-time performance. The proposed ELM-MGIR algorithm outperforms other mainstream iris recognition algorithms.

Phenolic content and antioxidant capacity of hybrid variety cocoa beans.

Science.gov (United States)

Jonfia-Essien, W A; West, G; Alderson, P G; Tucker, G

2008-06-01

Cocoa (Theobroma cacao L.) is a major, economically important, international crop and has been associated with several nutritional benefits including high antioxidant capacity. New cocoa hybrids have been developed in Ghana that exhibit resistance to pest damage during storage. The aim of this work was to assess the phenolic content and antioxidant capacity of these new hybrids in comparison to more traditional cocoa varieties. Total extractable phenolics were similar in all the four hybrids tested ranging from 69.9 to 81.6FAEg(-1). These levels were very similar to that extracted from traditional beans (73.8±2.5FAEg(-1)). The "phenolic profile" was determined by HPLC. A total of 25 peaks was observed but there were only minor differences in this profile between traditional and hybrid bean extracts. Antioxidant capacity was determined using the FRAP assay and traditional beans were found to possess 12.4μmolTEg(-1). In comparison the hybrid beans had antioxidant capacities ranging from 21.6 to 45.5μmolTEg(-1), and these were significantly higher than in the traditional beans for three out of the four hybrids. Since the phenolic and antioxidant levels and in these hybrid varieties were either similar to, or higher than, that obtained from traditional beans, the introduction of these new varieties would be unlikely to impact detrimentally on these nutritional components of the beans. Copyright © 2007 Elsevier Ltd. All rights reserved.

Simple Objective Detection of Human Lyme Disease Infection Using Immuno-PCR and a Single Recombinant Hybrid Antigen

Science.gov (United States)

Halpern, Micah D.; Molins, Claudia R.; Schriefer, Martin

2014-01-01

A serology-based tiered approach has, to date, provided the most effective means of laboratory confirmation of clinically suspected cases of Lyme disease, but it lacks sensitivity in the early stages of disease and is often dependent on subjectively scored immunoblots. We recently demonstrated the use of immuno-PCR (iPCR) for detecting Borrelia burgdorferi antibodies in patient serum samples that were positive for Lyme disease. To better understand the performance of the Lyme disease iPCR assay, the repeatability and variability of the background of the assay across samples from a healthy population (n = 36) were analyzed. Both of these parameters were found to have coefficients of variation of Lyme disease patient serum samples (n = 12) demonstrated a strong correlation with that of 2-tier testing. Furthermore, a simplified iPCR approach using a single hybrid antigen and detecting only IgG antibodies confirmed the 2-tier diagnosis in the Lyme disease patient serum samples (n = 12). Validation of the hybrid antigen IgG iPCR assay using a blinded panel of Lyme disease and non-Lyme disease patient serum samples (n = 92) resulted in a sensitivity of 69% (95% confidence interval [CI], 50% to 84%), compared to that of the 2-tier analysis at 59% (95% CI, 41% to 76%), and a specificity of 98% (95% CI, 91% to 100%) compared to that of the 2-tier analysis at 97% (95% CI, 88% to 100%). A single-tier hybrid antigen iPCR assay has the potential to be an improved method for detecting host-generated antibodies against B. burgdorferi. PMID:24899074

Hybridization Capture Using RAD Probes (hyRAD, a New Tool for Performing Genomic Analyses on Collection Specimens.

Directory of Open Access Journals (Sweden)

Tomasz Suchan

Full Text Available In the recent years, many protocols aimed at reproducibly sequencing reduced-genome subsets in non-model organisms have been published. Among them, RAD-sequencing is one of the most widely used. It relies on digesting DNA with specific restriction enzymes and performing size selection on the resulting fragments. Despite its acknowledged utility, this method is of limited use with degraded DNA samples, such as those isolated from museum specimens, as these samples are less likely to harbor fragments long enough to comprise two restriction sites making possible ligation of the adapter sequences (in the case of double-digest RAD or performing size selection of the resulting fragments (in the case of single-digest RAD. Here, we address these limitations by presenting a novel method called hybridization RAD (hyRAD. In this approach, biotinylated RAD fragments, covering a random fraction of the genome, are used as baits for capturing homologous fragments from genomic shotgun sequencing libraries. This simple and cost-effective approach allows sequencing of orthologous loci even from highly degraded DNA samples, opening new avenues of research in the field of museum genomics. Not relying on the restriction site presence, it improves among-sample loci coverage. In a trial study, hyRAD allowed us to obtain a large set of orthologous loci from fresh and museum samples from a non-model butterfly species, with a high proportion of single nucleotide polymorphisms present in all eight analyzed specimens, including 58-year-old museum samples. The utility of the method was further validated using 49 museum and fresh samples of a Palearctic grasshopper species for which the spatial genetic structure was previously assessed using mtDNA amplicons. The application of the method is eventually discussed in a wider context. As it does not rely on the restriction site presence, it is therefore not sensitive to among-sample loci polymorphisms in the restriction sites

Selection of matched pair of monoclonal antibodies for development of immunoradiometric assay (IRMA) : our experience with IRMA of TSH

International Nuclear Information System (INIS)

Kadwad, V.B.; Jyotsna, N.; Sivaprasad, N.

1998-01-01

Full text: In immunoradiometricassay (IRMA) two antibodies raised against two different epitopes of the same antigen are used, one bound to a solid phase (capture antibody) and the other labelled with 125 I (detector antibody). The development of any IRMA thus involves proper selection of the capture and detector antibody, preparation of solid phase, labelling of the antibody and assay optimization. Extensive studies have been carried out on these aspects in our laboratory with greater emphasis on the behavior of different pairs of antibodies as sandwich partners : monoclonal-monoclonal and monoclonal-polyclonal antibodies. The parameters studied include the ease of radio-iodination of different monoclonal antibodies, the effect of interchange of capture and detector antibody etc. Keeping TSH antibody as a model, two different monoclonal antibodies, a polyclonal antibody and a tracer from a commercial TSH IRMA kit were used in this study. Based on our studies an assay procedure for in-house IRMA of TSH has been developed with a sensitivity of 0.1 μIU/ml and validated

Hybridization State Detection of DNA-Functionalized Gold Nanoparticles Using Hyperspectral Imaging

Directory of Open Access Journals (Sweden)

Richard C. Murdock

2017-01-01

Full Text Available Hyperspectral imaging has the unique ability of capturing spectral data for multiple wavelengths at each pixel in an image. This gives the ability to distinguish, with certainty, different nanomaterials and/or distinguish nanomaterials from biological materials. In this study, 4 nm and 13 nm gold nanoparticles (Au NPs were synthesized, functionalized with complimentary oligonucleotides, and hybridized to form large networks of NPs. Scattering spectra were collected from each sample (unfunctionalized, functionalized, and hybridized and evaluated. The spectra showed unique peaks for each size of Au NP sample and also exhibited narrowing and intensifying of the spectra as the NPs were functionalized and then subsequently hybridized. These spectra are different from normal aggregation effects where the LSPR and reflected spectrum broaden and are red-shifted. Rather, this appears to be dependent on the ability to control the interparticle distance through oligonucleotide length, which is also investigated through the incorporation of a poly-A spacer. Also, hybridized Au NPs were exposed to cells with no adverse effects and retained their unique spectral signatures. With the ability to distinguish between hybridization states at nearly individual NP levels, this could provide a new method of tracking the intracellular actions of nanomaterials as well as extracellular biosensing applications.

Optional carbon capture

Energy Technology Data Exchange (ETDEWEB)

Alderson, T.; Scott, S.; Griffiths, J. [Jacobs Engineering, London (United Kingdom)

2007-07-01

In the case of IGCC power plants, carbon capture can be carried out before combustion. The carbon monoxide in the syngas is catalytically shifted to carbon dioxide and then captured in a standard gas absorption system. However, the insertion of a shift converter into an existing IGCC plant with no shift would mean a near total rebuild of the gasification waste heat recovery, gas treatment system and HRSG, with only the gasifier and gas turbine retaining most of their original features. To reduce the extent, cost and time taken for the revamping, the original plant could incorporate the shift, and the plant would then be operated without capture to advantage, and converted to capture mode of operation when commercially appropriate. This paper examines this concept of placing a shift converter into an IGCC plant before capture is required, and operating the same plant first without and then later with CO{sub 2} capture in a European context. The advantages and disadvantages of this 'capture ready' option are discussed. 6 refs., 2 figs., 4 tabs.

Miniaturized Aptamer-Based Assays for Protein Detection

Directory of Open Access Journals (Sweden)

Alessandro Bosco

2016-09-01

Full Text Available The availability of devices for cancer biomarker detection at early stages of the disease is one of the most critical issues in biomedicine. Towards this goal, to increase the assay sensitivity, device miniaturization strategies empowered by the employment of high affinity protein binders constitute a valuable approach. In this work we propose two different surface-based miniaturized platforms for biomarker detection in body fluids: the first platform is an atomic force microscopy (AFM-based nanoarray, where AFM is used to generate functional nanoscale areas and to detect biorecognition through careful topographic measurements; the second platform consists of a miniaturized electrochemical cell to detect biomarkers through electrochemical impedance spectroscopy (EIS analysis. Both devices rely on robust and highly-specific protein binders as aptamers, and were tested for thrombin detection. An active layer of DNA-aptamer conjugates was immobilized via DNA directed immobilization on complementary single-stranded DNA self-assembled monolayers confined on a nano/micro area of a gold surface. Results obtained with these devices were compared with the output of surface plasmon resonance (SPR assays used as reference. We succeeded in capturing antigens in concentrations as low as a few nM. We put forward ideas to push the sensitivity further to the pM range, assuring low biosample volume (μL range assay conditions.

Hybrid ligand-alkylating agents targeting telomeric G-quadruplex structures.

Science.gov (United States)

Doria, Filippo; Nadai, Matteo; Folini, Marco; Di Antonio, Marco; Germani, Luca; Percivalle, Claudia; Sissi, Claudia; Zaffaroni, Nadia; Alcaro, Stefano; Artese, Anna; Richter, Sara N; Freccero, Mauro

2012-04-14

The synthesis, physico-chemical properties and biological effects of a new class of naphthalene diimides (NDIs) capable of reversibly binding telomeric DNA and alkylate it through an electrophilic quinone methide moiety (QM), are reported. FRET and circular dichroism assays showed a marked stabilization and selectivity towards telomeric G4 DNA folded in a hybrid topology. NDI-QMs' alkylating properties revealed a good reactivity on single nucleosides and selectivity towards telomeric G4. A selected NDI was able to significantly impair the growth of melanoma cells by causing telomere dysfunction and down-regulation of telomerase expression. These findings points to our hybrid ligand-alkylating NDIs as possible tools for the development of novel targeted anticancer therapies. This journal is © The Royal Society of Chemistry 2012

Resistance of solanum species to phytophthora infestans evaluated in the detached-leaf and whole-plant assays

International Nuclear Information System (INIS)

Akhtar, K.P.; Saleem, M.Y.; Asghar, M.

2012-01-01

The reaction of 82 tomato genotypes belonging to 8 Solanum and a Lycopersicon species against Phytophthora infestans causing late blight was determined using detached-leaf and whole-plant assays. None of the test genotypes was immune or highly resistant. Of the 82 commercial and wild genotypes only TMS-2 (male-sterile and characterized by indeterminate growth) belonging to Lycopersicon esculentum was resistant with severity index of 2.4 in the detached-leaf assay on 0-5 scale (where 5 was highly susceptible) and percent disease index (%DI) of 23.3% under the whole-plant assay. Among the remaining genotypes, 41 were susceptible and 40 were highly susceptible under the detached-leaf assay, while 18 were susceptible and 63 were highly susceptible under the whole-plant assay. However, there was a significant difference in %DI for genotypes under the whole-plant assay. The response of whole-plants to inoculation with P. infestans in the detached-leaf assay was similar in all cases. The overall screening results indicate that TMS-2 is a good source of resistance and it can be useful for the development of tomato hybrid cultivars resistant to late blight. (author)

Multiplex PCR-based assay for detection of Bordetella pertussis in nasopharyngeal swab specimens.

Science.gov (United States)

Wadowsky, R M; Michaels, R H; Libert, T; Kingsley, L A; Ehrlich, G D

1996-11-01

A multiplex PCR-based assay was developed for the detection of Bordetella pertussis in nasopharyngeal swab specimens. The assay simultaneously amplified two separate DNA targets (153 and 203 bp) within a B. pertussis repetitive element and a 438-bp target within the beta-actin gene of human DNA (PCR amplification control). PCR products were detected by a sensitive and specific liquid hybridization gel retardation assay. A total of 496 paired nasopharyngeal swab specimens were tested by both the PCR-based assay and culture. Although 30 (6%) of the specimens inhibited the amplification of the beta-actin target, in all 29 specimens studied, the inhibition disappeared on repeat testing or was easily overcome with a 1:8 dilution or less of specimen digest. Of the 495 specimen pairs yielding a final evaluable result by the PCR-based assay, 19.0% were positive by the PCR-based assay, whereas 13.9% were positive by culture (P < 0.0001). After resolving the PCR-positive, culture-negative results by testing an additional aliquot from these specimens by the multiplex PCR-based assay, the PCR-based assay had a sensitivity and specificity of 98.9 and 99.7%, respectively, compared with values of 73.4 and 100%, respectively, for culture. In comparison with patients with culture-confirmed pertussis, those with PCR-positive, culture-negative results were older and more likely to have had prolonged cough, immunization with pertussis vaccine, or treatment with erythromycin. This multiplex PCR-based assay is substantially more sensitive than culture and identifies specimens that contain inhibitors of PCR.

Gravitational capture

International Nuclear Information System (INIS)

Bondi, H.

1979-01-01

In spite of the strength of gravitational focres between celestial bodies, gravitational capture is not a simple concept. The principles of conservation of linear momentum and of conservation of angular momentum, always impose severe constraints, while conservation of energy and the vital distinction between dissipative and non-dissipative systems allows one to rule out capture in a wide variety of cases. In complex systems especially those without dissipation, long dwell time is a more significant concept than permanent capture. (author)

Ratiometric fluorescence transduction by hybridization after isothermal amplification for determination of zeptomole quantities of oligonucleotide biomarkers with a paper-based platform and camera-based detection

Energy Technology Data Exchange (ETDEWEB)

Noor, M. Omair; Hrovat, David [Chemical Sensors Group, Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Road, Mississauga, ON L5L 1C6 (Canada); Moazami-Goudarzi, Maryam [Department of Cell and Systems Biology, University of Toronto Mississauga, 3359 Mississauga Road, Mississauga, ON L5L 1C6 (Canada); Espie, George S. [Department of Cell and Systems Biology, University of Toronto Mississauga, 3359 Mississauga Road, Mississauga, ON L5L 1C6 (Canada); Department of Biology, University of Toronto Mississauga, 3359 Mississauga Road, Mississauga, ON L5L 1C6 (Canada); Krull, Ulrich J., E-mail: ulrich.krull@utoronto.ca [Chemical Sensors Group, Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Road, Mississauga, ON L5L 1C6 (Canada)

2015-07-23

Highlights: • Solid-phase QD-FRET transduction of isothermal tHDA amplicons on paper substrates. • Ratiometric QD-FRET transduction improves assay precision and lowers the detection limit. • Zeptomole detection limit by an iPad camera after isothermal amplification. • Tunable assay sensitivity by immobilizing different amounts of QD–probe bioconjugates. - Abstract: Paper is a promising platform for the development of decentralized diagnostic assays owing to the low cost and ease of use of paper-based analytical devices (PADs). It can be challenging to detect on PADs very low concentrations of nucleic acid biomarkers of lengths as used in clinical assays. Herein we report the use of thermophilic helicase-dependent amplification (tHDA) in combination with a paper-based platform for fluorescence detection of probe-target hybridization. Paper substrates were patterned using wax printing. The cellulosic fibers were chemically derivatized with imidazole groups for the assembly of the transduction interface that consisted of immobilized quantum dot (QD)–probe oligonucleotide conjugates. Green-emitting QDs (gQDs) served as donors with Cy3 as the acceptor dye in a fluorescence resonance energy transfer (FRET)-based transduction method. After probe-target hybridization, a further hybridization event with a reporter sequence brought the Cy3 acceptor dye in close proximity to the surface of immobilized gQDs, triggering a FRET sensitized emission that served as an analytical signal. Ratiometric detection was evaluated using both an epifluorescence microscope and a low-cost iPad camera as detectors. Addition of the tHDA method for target amplification to produce sequences of ∼100 base length allowed for the detection of zmol quantities of nucleic acid targets using the two detection platforms. The ratiometric QD-FRET transduction method not only offered improved assay precision, but also lowered the limit of detection of the assay when compared with the non

Ratiometric fluorescence transduction by hybridization after isothermal amplification for determination of zeptomole quantities of oligonucleotide biomarkers with a paper-based platform and camera-based detection

International Nuclear Information System (INIS)

Noor, M. Omair; Hrovat, David; Moazami-Goudarzi, Maryam; Espie, George S.; Krull, Ulrich J.

2015-01-01

Highlights: • Solid-phase QD-FRET transduction of isothermal tHDA amplicons on paper substrates. • Ratiometric QD-FRET transduction improves assay precision and lowers the detection limit. • Zeptomole detection limit by an iPad camera after isothermal amplification. • Tunable assay sensitivity by immobilizing different amounts of QD–probe bioconjugates. - Abstract: Paper is a promising platform for the development of decentralized diagnostic assays owing to the low cost and ease of use of paper-based analytical devices (PADs). It can be challenging to detect on PADs very low concentrations of nucleic acid biomarkers of lengths as used in clinical assays. Herein we report the use of thermophilic helicase-dependent amplification (tHDA) in combination with a paper-based platform for fluorescence detection of probe-target hybridization. Paper substrates were patterned using wax printing. The cellulosic fibers were chemically derivatized with imidazole groups for the assembly of the transduction interface that consisted of immobilized quantum dot (QD)–probe oligonucleotide conjugates. Green-emitting QDs (gQDs) served as donors with Cy3 as the acceptor dye in a fluorescence resonance energy transfer (FRET)-based transduction method. After probe-target hybridization, a further hybridization event with a reporter sequence brought the Cy3 acceptor dye in close proximity to the surface of immobilized gQDs, triggering a FRET sensitized emission that served as an analytical signal. Ratiometric detection was evaluated using both an epifluorescence microscope and a low-cost iPad camera as detectors. Addition of the tHDA method for target amplification to produce sequences of ∼100 base length allowed for the detection of zmol quantities of nucleic acid targets using the two detection platforms. The ratiometric QD-FRET transduction method not only offered improved assay precision, but also lowered the limit of detection of the assay when compared with the non

Full wave simulations of fast wave mode conversion and lower hybrid wave propagation in tokamaks

DEFF Research Database (Denmark)

Wright, J.C.; Bonoli, P.T.; Brambilla, M.

2004-01-01

Fast wave (FW) studies of mode conversion (MC) processes at the ion-ion hybrid layer in toroidal plasmas must capture the disparate scales of the FW and mode converted ion Bernstein and ion cyclotron waves. Correct modeling of the MC layer requires resolving wavelengths on the order of k...

Capture ready study

Energy Technology Data Exchange (ETDEWEB)

Minchener, A.

2007-07-15

There are a large number of ways in which the capture of carbon as carbon dioxide (CO{sub 2}) can be integrated into fossil fuel power stations, most being applicable for both gas and coal feedstocks. To add to the choice of technology is the question of whether an existing plant should be retrofitted for capture, or whether it is more attractive to build totally new. This miscellany of choices adds considerably to the commercial risk of investing in a large power station. An intermediate stage between the non-capture and full capture state would be advantageous in helping to determine the best way forward and hence reduce those risks. In recent years the term 'carbon capture ready' or 'capture ready' has been coined to describe such an intermediate stage plant and is now widely used. However a detailed and all-encompassing definition of this term has never been published. All fossil fuel consuming plant produce a carbon dioxide gas byproduct. There is a possibility of scrubbing it with an appropriate CO{sub 2} solvent. Hence it could be said that all fossil fuel plant is in a condition for removal of its CO{sub 2} effluent and therefore already in a 'capture ready' state. Evidently, the practical reality of solvent scrubbing could cost more than the rewards offered by such as the ETS (European Trading Scheme). In which case, it can be said that although the possibility exists of capturing CO{sub 2}, it is not a commercially viable option and therefore the plant could not be described as ready for CO{sub 2} capture. The boundary between a capture ready and a non-capture ready condition using this definition cannot be determined in an objective and therefore universally acceptable way and criteria must be found which are less onerous and less potentially contentious to assess. 16 refs., 2 annexes.

Multiply osmium-labeled reporter probes for electrochemical DNA hybridization assays: detection of trinucleotide repeats

Czech Academy of Sciences Publication Activity Database

Fojta, Miroslav; Havran, Luděk; Kizek, René; Paleček, Emil

2004-01-01

Roč. 20, č. 5 (2004), s. 985-994 ISSN 0956-5663 R&D Projects: GA MPO 1H-PK/42; GA AV ČR IAA4004108; GA AV ČR IBS5004355; GA AV ČR KJB4004302; GA AV ČR KSK4055109 Institutional research plan: CEZ:AV0Z5004920 Keywords : electrochemical sensors * DNA hybridization * DNA labeling Subject RIV: BO - Biophysics Impact factor: 3.251, year: 2004

Prey capture behaviour evoked by simple visual stimuli in larval zebrafish

Directory of Open Access Journals (Sweden)

Isaac Henry Bianco

2011-12-01

Full Text Available Understanding how the nervous system recognises salient stimuli in the environ- ment and selects and executes the appropriate behavioural responses is a fundamen- tal question in systems neuroscience. To facilitate the neuroethological study of visually-guided behaviour in larval zebrafish, we developed virtual reality assays in which precisely controlled visual cues can be presented to larvae whilst their behaviour is automatically monitored using machine-vision algorithms. Freely swimming larvae responded to moving stimuli in a size-dependent manner: they directed multiple low amplitude orienting turns (∼ 20◦ towards small moving spots (1◦ but reacted to larger spots (10◦ with high-amplitude aversive turns (∼ 60◦. The tracking of small spots led us to examine how larvae respond to prey during hunting routines. By analysing movie sequences of larvae hunting parame- cia, we discovered that all prey capture routines commence with eye convergence and larvae maintain their eyes in a highly converged position for the duration of the prey-tracking and capture swim phases. We adapted our virtual reality assay to deliver artificial visual cues to partially restrained larvae and found that small moving spots evoked convergent eye movements and J-turns of the tail, which are defining features of natural hunting. We propose that eye convergence represents the engagement of a predatory mode of behaviour in larval fish and serves to increase the region of binocular visual space to enable stereoscopic targeting of prey.

Measurement of cetuximab and panitumumab-unbound serum EGFR extracellular domain using an assay based on slow off-rate modified aptamer (SOMAmer reagents.

Directory of Open Access Journals (Sweden)

Noh Jin Park

Full Text Available Response to cetuximab (Erbitux® and panitumumab (Vectibix® varies among individuals, and even those who show response ultimately gain drug resistance. One possible etiologic factor is differential interaction between the drug and target. We describe the development of an assay based on Slow Off-rate Modified Aptamer (SOMAmer(™ reagents that can distinguish drug-bound from unbound epidermal growth factor receptor (EGFR.This quantitative assay uses a SOMAmer reagent specific for EGFR extracellular domain (ECD as a capturing reagent. Captured SOMAmer is quantitated using PCR. Linearity and accuracy (recovery of the assay were assessed using normal sera and purified EGFR ECD.This EGFR ECD assay showed linearity between 2.5 and 600 ng/mL. Average recovery was 101%. The assay detected EGFR but showed little cross-reactivity to other ErbB proteins: 0.4% for ErbB2, 6.9% for ErbB3, and 1.3% for ErbB4. Preincubation of normal serum with either cetuximab or panitumumab resulted in a dose-dependent decrease in EGFR ECD levels measured using the SOMAmer assay; preincubation did not affect measurement with an ELISA.This SOMAmer-based serum EGFR ECD assay accurately and specifically measures EGFR in serum. Detection of significant amounts of drug-unbound EGFR in patients undergoing cetuximab or panitumumab treatment could be an indicator of poor drug response. Further studies are needed to evaluate the utility of the assay as an indicator of drug efficacy or as a guide to dosing.

Ultra-sensitive DNA assay based on single-molecule detection coupled with fluorescent quantum dot-labeling and its application to determination of messenger RNA

International Nuclear Information System (INIS)

Li Li; Li Xincang; Li Lu; Wang Jinxing; Jin Wenrui

2011-01-01

An ultra-sensitive single-molecule detection (SMD) method for quantification of DNA using total internal reflection fluorescence microscopy (TIRFM) coupled with fluorescent quantum dot (QD)-labeling was developed. In this method, the target DNA (tDNA) was captured by the capture DNA immobilized on the silanized coverslip blocked with ethanolamine and bovine serum albumin. Then, the QD-labeled probe DNA was hybridized to the tDNA. Ten fluorescent images of the QD-labeled sandwich DNA hybrids on the coverslip were taken by a high-sensitive CCD. The tDNA was quantified by counting the bright spots on the images using a calibration curve. The LOD of the method was 1 x 10 -14 mol L -1 . Several key factors, including image acquirement, fluorescence probe, substrate preparation, noise elimination from solutions and glass coverslips, and nonspecific adsorption and binding of solution-phase detection probes were discussed in detail. The method could be applied to quantify messenger RNA (mRNA) in cells. In order to determine mRNA, double-stranded RNA-DNA hybrids consisting of mRNA and corresponding cDNA were synthesized from the cellular mRNA template using reverse transcription in the presence of reverse transcriptase. After removing the mRNA in the double-stranded hybrids using ribonuclease, cDNA was quantified using the SMD-based TIRFM. Osteopontin mRNA in decidual stromal cells was chosen as the model analyte.

Rapid hybridization of nucleic acids using isotachophoresis

Science.gov (United States)

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

2012-01-01

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

Increasing the analytical sensitivity by oligonucleotides modified with para- and ortho-twisted intercalating nucleic acids--TINA.

Directory of Open Access Journals (Sweden)

Uffe V Schneider

Full Text Available The sensitivity and specificity of clinical diagnostic assays using DNA hybridization techniques are limited by the dissociation of double-stranded DNA (dsDNA antiparallel duplex helices. This situation can be improved by addition of DNA stabilizing molecules such as nucleic acid intercalators. Here, we report the synthesis of a novel ortho-Twisted Intercalating Nucleic Acid (TINA amidite utilizing the phosphoramidite approach, and examine the stabilizing effect of ortho- and para-TINA molecules in antiparallel DNA duplex formation. In a thermal stability assay, ortho- and para-TINA molecules increased the melting point (Tm of Watson-Crick based antiparallel DNA duplexes. The increase in Tm was greatest when the intercalators were placed at the 5' and 3' termini (preferable or, if placed internally, for each half or whole helix turn. Terminally positioned TINA molecules improved analytical sensitivity in a DNA hybridization capture assay targeting the Escherichia coli rrs gene. The corresponding sequence from the Pseudomonas aeruginosa rrs gene was used as cross-reactivity control. At 150 mM ionic strength, analytical sensitivity was improved 27-fold by addition of ortho-TINA molecules and 7-fold by addition of para-TINA molecules (versus the unmodified DNA oligonucleotide, with a 4-fold increase retained at 1 M ionic strength. Both intercalators sustained the discrimination of mismatches in the dsDNA (indicated by ΔTm, unless placed directly adjacent to the mismatch--in which case they partly concealed ΔTm (most pronounced for para-TINA molecules. We anticipate that the presented rules for placement of TINA molecules will be broadly applicable in hybridization capture assays and target amplification systems.

Highly sensitive antibody-aptamer sensor for vascular endothelial growth factor based on hybridization chain reaction and pH meter/indicator.

Science.gov (United States)

Xu, Huifeng; Kou, Fangxia; Ye, Hongzhi; Wang, Zongwen; Huang, Suixin; Liu, Xianxiang; Zhu, Xi; Lin, Zhenyu; Chen, Guonan

2017-12-01

Vascular endothelial growth factor (VEGF) is a crucial signaling protein for the tumor growth and metastasis, which is also acted as the biomarkers for various diseases. In this research, we fabricate an aptamer-antibody sensor for point-of-care test of VEGF. Firstly, target VEGF is captured by antibody immobilized on the microplate, and then binds with aptamer to form the sandwich structure. Next, with the assist of glucose oxidase (GOx)-functionalized ssDNAs, hybridization chain reaction occurs using the aptamer as the primer. Thus, GOx are greatly gathered on the microplate, which catalyzes the oxidization of glucose, leading to the pH change. As a result, the detect limit at a signal-to-noise was estimated to be 0.5pg/mL of target by pH meter, and 1.6pg/mL of VEGF was able to be distinguished by naked eyes. Meanwhile, this method has been used assay VEGF in the serum with the satisfactory results. Copyright © 2017. Published by Elsevier B.V.

A global hybrid coupled model based on atmosphere-SST feedbacks

Energy Technology Data Exchange (ETDEWEB)

Cimatoribus, Andrea A.; Drijfhout, Sybren S. [Royal Netherlands Meteorological Institute, De Bilt (Netherlands); Dijkstra, Henk A. [Utrecht University, Institute for Marine and Atmospheric Research Utrecht, Utrecht (Netherlands)

2012-02-15

A global hybrid coupled model is developed, with the aim of studying the effects of ocean-atmosphere feedbacks on the stability of the Atlantic meridional overturning circulation. The model includes a global ocean general circulation model and a statistical atmosphere model. The statistical atmosphere model is based on linear regressions of data from a fully coupled climate model on sea surface temperature both locally and hemispherically averaged, being the footprint of Atlantic meridional overturning variability. It provides dynamic boundary conditions to the ocean model for heat, freshwater and wind-stress. A basic but consistent representation of ocean-atmosphere feedbacks is captured in the hybrid coupled model and it is more than 10 times faster than the fully coupled climate model. The hybrid coupled model reaches a steady state with a climate close to the one of the fully coupled climate model, and the two models also have a similar response (collapse) of the Atlantic meridional overturning circulation to a freshwater hosing applied in the northern North Atlantic. (orig.)

In vivo immuno-reactivity analysis of the porous three-dimensional chitosan/SiO2 and chitosan/SiO2 /hydroxyapatite hybrids.

Science.gov (United States)

Guo, Mengxia; Dong, Yifan; Xiao, Jiangwei; Gu, Ruicai; Ding, Maochao; Huang, Tao; Li, Junhua; Zhao, Naru; Liao, Hua

2018-05-01

Inorganic/organic hybrid silica-chitosan (CS) scaffolds have promising potential for bone defect repair, due to the controllable mechanical properties, degradation behavior, and scaffold morphology. However, the precise in vivo immuno-reactivity of silica-CS hybrids with various compositions is still poorly defined. In this study, we fabricated the three-dimensional (3D) interconnected porous chitosan-silica (CS/SiO 2 ) and chitosan-silica-hydroxyapatite (CS/SiO 2 /HA) hybrids, through sol-gel process and 3D plotting skill, followed by the naturally or freeze drying separately. Scanning electron microscopy demonstrated the hybrids possessed the uniform geometric structure, while, transmission electron microscopy displayed nanoscale silica, or HA nanoparticles dispersed homogeneously in the CS matrix, or CS/silica hybrids. After intramuscular implantation, CS/SiO 2 and CS/SiO 2 /HA hybrids triggered a local and limited monocyte/macrophage infiltration and myofiber degeneration. Naturally dried CS/SiO 2 hybrid provoked a more severe inflammation than the freeze-dried ones. Dendritic cells were attracted to invade into the implants embedded-muscle, but not be activated to prime the adaptive immunity, because the absence of cytotoxic T cells and B cells in muscle received the implants. Fluorescence-activated cell sorting (FACS) analysis indicated the implanted hybrids were incapable to initiate splenocytes activation. Plasma complement C3 enzyme linked immunosorbent assay (ELISA) assay showed the hybrids induced C3 levels increase in early implanting phase, and the subsequent striking decrease. Thus, the present results suggest that, in vivo, 3D plotted porous CS/SiO 2 and CS/SiO 2 /HA hybrids are relatively biocompatible in vivo, which initiate a localized inflammatory procedure, instead of a systematic immune response. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1223-1235, 2018. © 2018 Wiley Periodicals, Inc.

Hybrid automata models of cardiac ventricular electrophysiology for real-time computational applications.

Science.gov (United States)

Andalam, Sidharta; Ramanna, Harshavardhan; Malik, Avinash; Roop, Parthasarathi; Patel, Nitish; Trew, Mark L

2016-08-01

Virtual heart models have been proposed for closed loop validation of safety-critical embedded medical devices, such as pacemakers. These models must react in real-time to off-the-shelf medical devices. Real-time performance can be obtained by implementing models in computer hardware, and methods of compiling classes of Hybrid Automata (HA) onto FPGA have been developed. Models of ventricular cardiac cell electrophysiology have been described using HA which capture the complex nonlinear behavior of biological systems. However, many models that have been used for closed-loop validation of pacemakers are highly abstract and do not capture important characteristics of the dynamic rate response. We developed a new HA model of cardiac cells which captures dynamic behavior and we implemented the model in hardware. This potentially enables modeling the heart with over 1 million dynamic cells, making the approach ideal for closed loop testing of medical devices.

[Super sweet corn hybrids adaptability for industrial processing. I freezing].

Science.gov (United States)

Alfonzo, Braunnier; Camacho, Candelario; Ortiz de Bertorelli, Ligia; De Venanzi, Frank

2002-09-01

With the purpose of evaluating adaptability to the freezing process of super sweet corn sh2 hybrids Krispy King, Victor and 324, 100 cobs of each type were frozen at -18 degrees C. After 120 days of storage, their chemical, microbiological and sensorial characteristics were compared with a sweet corn su. Industrial quality of the process of freezing and length and number of rows in cobs were also determined. Results revealed yields above 60% in frozen corns. Length and number of rows in cobs were acceptable. Most of the chemical characteristics of super sweet hybrids were not different from the sweet corn assayed at the 5% significance level. Moisture content and soluble solids of hybrid Victor, as well as total sugars of hybrid 324 were statistically different. All sh2 corns had higher pH values. During freezing, soluble solids concentration, sugars and acids decreased whereas pH increased. Frozen cobs exhibited acceptable microbiological rank, with low activities of mesophiles and total coliforms, absence of psychrophiles and fecal coliforms, and an appreciable amount of molds. In conclusion, sh2 hybrids adapted with no problems to the freezing process, they had lower contents of soluble solids and higher contents of total sugars, which almost doubled the amount of su corn; flavor, texture, sweetness and appearance of kernels were also better. Hybrid Victor was preferred by the evaluating panel and had an outstanding performance due to its yield and sensorial characteristics.

Radiative electron capture

International Nuclear Information System (INIS)

Biggerstaff, J.A.; Appleton, B.R.; Datz, S.; Moak, C.D.; Neelavathi, V.N.; Noggle, T.S.; Ritchie, R.H.; VerBeek, H.

1975-01-01

Some data are presented for radiative electron capture by fast moving ions. The radiative electron capture spectrum is shown for O 8+ in Ag, along with the energy dependence of the capture cross-section. A discrepancy between earlier data, theoretical prediction, and the present data is pointed out. (3 figs) (U.S.)

Novel Platinum (Pt)-Vandetanib Hybrid Compounds: Design, Synthesis and Investigation of Anti-cancer Activity and Mechanism of Action

Science.gov (United States)

Fei, Rong

Purpose: Lung cancer is one of the most common cancers and non-small cell lung cancer (NSCLC) accounts for 80-85% of lung cancers. 70% of individuals with NSCLC harboring somatic mutations in exons of the epidermal growth factor receptor (EGFR) gene that encode tyrosine kinase domain. EGFR tyrosine kinase inhibitors (TKIs) are promising molecular targeted therapy for NSCLC with sensitizing EGFR mutations. However, secondary mutation of EGFR after treatment of TKIs develops resistance. Vandetanib is introduced to overcome erlotinib resistance as a multi-targeted TKI. However, its anticancer effect is still compromised by EGFR T790M mutation. Therefore, new molecular anticancer strategies are necessarily needed. In this study, vandetanib is incorporated with Pt-based anticancer agents as hybrid compounds, aiming to circumvent TKI resistance. Furthermore, hybrid compounds are investigated in cisplatin resistant problem to expect to overcome resistance by introduction of vandetanib. Methods: Three novel Pt-vandetanib hybrid compounds were synthesized and its physicochemical properties were characterized. Anticancer activity and cytotoxicity were evaluated by sulforhodamine B assay and lactate dehydrogenase release. Docking simulation was performed to investigate the interaction of compounds with EGFR harboring different mutations. Inhibition efficacy of hybrids to kinases was evaluated by kinase inhibition profiling service and cell-free kinase inhibition assay. Mechanistic studies on cytotoxicity activity of the hybrid compounds were carried out. DNA damage response of hybrid compounds was further investigated in KB cells. The cytotoxicity of hybrids was tested in cisplatin resistant KB CP20 cells. Mechanistic of anticancer activity was studied to test inhibition on oncoprotein CIP2Aand DNA damage. Results: Platinum-vandetanib hybrid compounds were synthesized and test to be stable under extracellular condition. Hybrids reacted with 5'-GMP2- and glutathione, and both

About how to capture and exploit the CO2 surplus that nature, per se, is not capable of fixing.

Science.gov (United States)

Godoy, Manuel S; Mongili, Beatrice; Fino, Debora; Prieto, M Auxiliadora

2017-09-01

Human activity has been altering many ecological cycles for decades, disturbing the natural mechanisms which are responsible for re-establishing the normal environmental balances. Probably, the most disrupted of these cycles is the cycle of carbon. In this context, many technologies have been developed for an efficient CO 2 removal from the atmosphere. Once captured, it could be stored in large geological formations and other reservoirs like oceans. This strategy could present some environmental and economic problems. Alternately, CO 2 can be transformed into carbonates or different added-value products, such as biofuels and bioplastics, recycling CO 2 from fossil fuel. Currently different methods are being studied in this field. We classified them into biological, inorganic and hybrid systems for CO 2 transformation. To be environmentally compatible, they should be powered by renewable energy sources. Although hybrid systems are still incipient technologies, they have made great advances in the recent years. In this scenario, biotechnology is the spearhead of ambitious strategies to capture CO 2 and reduce global warming. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Fuel economy and life-cycle cost analysis of a fuel cell hybrid vehicle

Science.gov (United States)

Jeong, Kwi Seong; Oh, Byeong Soo

The most promising vehicle engine that can overcome the problem of present internal combustion is the hydrogen fuel cell. Fuel cells are devices that change chemical energy directly into electrical energy without combustion. Pure fuel cell vehicles and fuel cell hybrid vehicles (i.e. a combination of fuel cell and battery) as energy sources are studied. Considerations of efficiency, fuel economy, and the characteristics of power output in hybridization of fuel cell vehicle are necessary. In the case of Federal Urban Driving Schedule (FUDS) cycle simulation, hybridization is more efficient than a pure fuel cell vehicle. The reason is that it is possible to capture regenerative braking energy and to operate the fuel cell system within a more efficient range by using battery. Life-cycle cost is largely affected by the fuel cell size, fuel cell cost, and hydrogen cost. When the cost of fuel cell is high, hybridization is profitable, but when the cost of fuel cell is less than 400 US$/kW, a pure fuel cell vehicle is more profitable.

Application of DNA hybridization techniques in the assessment of diarrheal disease among refugess in Thailand. [Shigella; Escherichia coli; Campylobacter; Cryptosporidium

Energy Technology Data Exchange (ETDEWEB)

Taylor, D.N.; Echeverria, P.; Pitarangsi, C.; Seriwatana, J.; Sethabutr, O.; Bodhidatta, L.; Brown, C.; Herrmann, J.E.; Blacklow, N.R.

1988-01-01

The epidemiology and etiology of acute diarrheal disease were determined in a Hmong refugee camp on the Thai-Laotian border from April 11 to May 14, 1985. DNA hybridization techniques were used to detect Shigella species, enteroinvasive Escherichia coli, and enterotoxigenic E. coli. A monoclonal enzyme-linked immunosorbent assay was used to detect rotavirus, and standard microbiology was used to detect other enteropathogens. The age-specific diarrheal disease rates were 47 episodes per month per 1000 children less than five years old and 113 episodes per month per 1000 children less than one year old. Rotavirus, enterotoxigenic E. coli, Campylobacter, and Cryptosporidium were the predominant pathogens in children less than two years old. The DNA probe hybridized with 94% of 31 specimens identified as enterotoxigenic E. coli by the standard assays and with none of the specimens in which the standard assays were negative. The probe for Shigella and enteroinvasive E. coli hybridized in eight of 10 stools that contained Shigella and four of 314 stools from which Shigella and enteroinvasive E. coli were not isolated. The use of DNA probes allows specimens to be collected in remote areas with a minimum amount of equipment and technical expertise so that they can be easily transported to a central laboratory for further processing.

A homogeneous fluorometric assay platform based on novel synthetic proteins

International Nuclear Information System (INIS)

Vardar-Schara, Goenuel; Krab, Ivo M.; Yi, Guohua; Su, Wei Wen

2007-01-01

Novel synthetic recombinant sensor proteins have been created to detect analytes in solution, in a rapid single-step 'mix and read' noncompetitive homogeneous assay process, based on modulating the Foerster resonance energy transfer (FRET) property of the sensor proteins upon binding to their targets. The sensor proteins comprise a protein scaffold that incorporates a specific target-capturing element, sandwiched by genetic fusion between two molecules that form a FRET pair. The utility of the sensor proteins was demonstrated via three examples, for detecting an anti-biotin Fab antibody, a His-tagged recombinant protein, and an anti-FLAG peptide antibody, respectively, all done directly in solution. The diversity of sensor-target interactions that we have demonstrated in this study points to a potentially universal applicability of the biosensing concept. The possibilities for integrating a variety of target-capturing elements with a common sensor scaffold predict a broad range of practical applications

Verification of a novel innovative blade root design for wind turbines using a hybrid numerical method

DEFF Research Database (Denmark)

Zhu, Wei Jun; Shen, Wen Zhong; Sørensen, Jens Nørkær

2017-01-01

captured at the outer part of the blades, where the relative wind speed is high. To assess the impact of this novel design idea, a hybrid numerical technique, based on solving the Reynolds-averaged Navier-Stokes equations, is utilized to determine the aerodynamic performance. The in-house developed Ellip...

Species delineation and hybrid identification using diagnostic nuclear markers for Plectropomus leopardus and Plectropomus maculatus

KAUST Repository

He, Song

2018-06-01

Diagnostic molecular markers are an essential tool in the study of species’ ecology and evolution, particularly in closely related and sympatric species. Furthermore, the increasing awareness of wild-hybrids has led to a renewed interest in rapid diagnostic assays. Here, we test the ability of two mitochondrial (Cytb and COI) and two nuclear markers (ETS2 and TMO-4c4) to confidently discriminate purebred P. leopardus and P. maculatus and their first-generation hybrids. A sample of 48 purebred individuals and 91 interspecific hybrids were used in this study and their delineation confirmed using a set of microsatellite markers. Our results indicate mitochondrial markers could not distinguish even between species but both nuclear markers confidently identified species and first-generation hybrids. However, later-generation hybrids could not always be confidently identified due to on-going introgression between species. Our findings provide a robust tool to distinguish purebred individuals and interspecific hybrids in a pair of species with an unexpectedly high incidence of hybridization. The quick species discrimination abilities provided by these diagnostic markers are important for stock assessment and recruitment studies of these important fishery species.

Species delineation and hybrid identification using diagnostic nuclear markers for Plectropomus leopardus and Plectropomus maculatus

KAUST Repository

He, Song; Harrison, Hugo B.; Berumen, Michael L.

2018-01-01

Diagnostic molecular markers are an essential tool in the study of species’ ecology and evolution, particularly in closely related and sympatric species. Furthermore, the increasing awareness of wild-hybrids has led to a renewed interest in rapid diagnostic assays. Here, we test the ability of two mitochondrial (Cytb and COI) and two nuclear markers (ETS2 and TMO-4c4) to confidently discriminate purebred P. leopardus and P. maculatus and their first-generation hybrids. A sample of 48 purebred individuals and 91 interspecific hybrids were used in this study and their delineation confirmed using a set of microsatellite markers. Our results indicate mitochondrial markers could not distinguish even between species but both nuclear markers confidently identified species and first-generation hybrids. However, later-generation hybrids could not always be confidently identified due to on-going introgression between species. Our findings provide a robust tool to distinguish purebred individuals and interspecific hybrids in a pair of species with an unexpectedly high incidence of hybridization. The quick species discrimination abilities provided by these diagnostic markers are important for stock assessment and recruitment studies of these important fishery species.

A hybrid nanoantenna for highly enhanced directional spontaneous emission

Energy Technology Data Exchange (ETDEWEB)

Chou, R. Yuanying; Lu, Guowei, E-mail: guowei.lu@pku.edu.cn; Shen, Hongming; He, Yingbo; Cheng, Yuqing [State Key Laboratory for Mesoscopic Physics, Department of Physics, Peking University, Beijing 100871 (China); Perriat, Pascal [MATEIS, UMR 5510 CNRS, INSA-Lyon, Université de Lyon, Villeurbanne Cedex 69621 (France); Martini, Matteo; Tillement, Olivier [ILM, UMR 5306 CNRS, Université de Lyon, Villeurbanne Cedex 69622 (France); Gong, Qihuang [State Key Laboratory for Mesoscopic Physics, Department of Physics, Peking University, Beijing 100871 (China); Collaborative Innovation Center of Quantum Matter, Beijing 100871 (China)

2014-06-28

Spontaneous emission modulated by a hybrid plasmonic nanoantenna has been investigated by employing finite-difference time-domain method. The hybrid nanoantenna configurations constituted by a gap hot-spot and of a plasmonic corrugated grating and a metal reflector sandwiching a SiO{sub 2} thin layer which appears promising for high spontaneous emission enhancement devices. Simulation assays show that the coupling between the gap-antenna and plasmonic corrugations reaches an ultra-high near-field enhancement factor in the excitation process. Moreover, concerning the emission process, the corrugations concentrate the far-field radiated power within a tiny angular volume, offering unprecedented collection efficiency. In the past decades, many kinds of optical antennas have been proposed and optimized to enhance single molecule detection. However, the excitation enhancement effect for single individual or dimmer plasmonic nanostructure is limited due to intrinsic nonradiative decay of the nanoparticle plasmon and quantum tunneling effect. The proposed hybrid configuration overwhelms the enhancement limit of single individual plasmonic structure. The findings provide an insight into spontaneous emission high enhancement through integrating the functions of different metallic nanostructures.

A hybrid nanoantenna for highly enhanced directional spontaneous emission

International Nuclear Information System (INIS)

Chou, R. Yuanying; Lu, Guowei; Shen, Hongming; He, Yingbo; Cheng, Yuqing; Perriat, Pascal; Martini, Matteo; Tillement, Olivier; Gong, Qihuang

2014-01-01

Spontaneous emission modulated by a hybrid plasmonic nanoantenna has been investigated by employing finite-difference time-domain method. The hybrid nanoantenna configurations constituted by a gap hot-spot and of a plasmonic corrugated grating and a metal reflector sandwiching a SiO 2 thin layer which appears promising for high spontaneous emission enhancement devices. Simulation assays show that the coupling between the gap-antenna and plasmonic corrugations reaches an ultra-high near-field enhancement factor in the excitation process. Moreover, concerning the emission process, the corrugations concentrate the far-field radiated power within a tiny angular volume, offering unprecedented collection efficiency. In the past decades, many kinds of optical antennas have been proposed and optimized to enhance single molecule detection. However, the excitation enhancement effect for single individual or dimmer plasmonic nanostructure is limited due to intrinsic nonradiative decay of the nanoparticle plasmon and quantum tunneling effect. The proposed hybrid configuration overwhelms the enhancement limit of single individual plasmonic structure. The findings provide an insight into spontaneous emission high enhancement through integrating the functions of different metallic nanostructures.

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

Science.gov (United States)

Algar, W Russ; Krull, Ulrich J

2010-01-01

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

Hybrid Cascading Outage Analysis of Extreme Events with Optimized Corrective Actions

Energy Technology Data Exchange (ETDEWEB)

Vallem, Mallikarjuna R.; Vyakaranam, Bharat GNVSR; Holzer, Jesse T.; Samaan, Nader A.; Makarov, Yuri V.; Diao, Ruisheng; Huang, Qiuhua; Ke, Xinda

2017-10-19

Power system are vulnerable to extreme contingencies (like an outage of a major generating substation) that can cause significant generation and load loss and can lead to further cascading outages of other transmission facilities and generators in the system. Some cascading outages are seen within minutes following a major contingency, which may not be captured exclusively using the dynamic simulation of the power system. The utilities plan for contingencies either based on dynamic or steady state analysis separately which may not accurately capture the impact of one process on the other. We address this gap in cascading outage analysis by developing Dynamic Contingency Analysis Tool (DCAT) that can analyze hybrid dynamic and steady state behavior of the power system, including protection system models in dynamic simulations, and simulating corrective actions in post-transient steady state conditions. One of the important implemented steady state processes is to mimic operator corrective actions to mitigate aggravated states caused by dynamic cascading. This paper presents an Optimal Power Flow (OPF) based formulation for selecting corrective actions that utility operators can take during major contingency and thus automate the hybrid dynamic-steady state cascading outage process. The improved DCAT framework with OPF based corrective actions is demonstrated on IEEE 300 bus test system.

Effect of insulin analogues on insulin/IGF1 hybrid receptors: increased activation by glargine but not by its metabolites M1 and M2.

Directory of Open Access Journals (Sweden)

Cécile Pierre-Eugene

Full Text Available BACKGROUND: In diabetic patients, the pharmacokinetics of injected human insulin does not permit optimal control of glycemia. Fast and slow acting insulin analogues have been developed, but they may have adverse properties, such as increased mitogenic or anti-apoptotic signaling. Insulin/IGF1 hybrid receptors (IR/IGF1R, present in most tissues, have been proposed to transmit biological effects close to those of IGF1R. However, the study of hybrid receptors is difficult because of the presence of IR and IGF1R homodimers. Our objective was to perform the first study on the pharmacological properties of the five marketed insulin analogues towards IR/IGF1R hybrids. METHODOLOGY: To study the effect of insulin analogues on IR/IGF1R hybrids, we used our previously developed Bioluminescence Resonance Energy Transfer (BRET assay that permits specific analysis of the pharmacological properties of hybrid receptors. Moreover, we have developed a new, highly sensitive BRET-based assay to monitor phophatidylinositol-3 phosphate (PIP(3 production in living cells. Using this assay, we performed a detailed pharmacological analysis of PIP(3 production induced by IGF1, insulin and insulin analogues in living breast cancer-derived MCF-7 and MDA-MB231 cells. RESULTS: Among the five insulin analogues tested, only glargine stimulated IR/IGF1R hybrids with an EC50 that was significantly lower than insulin and close to that of IGF1. Glargine more efficiently stimulated PIP(3 production in MCF-7 cells but not in MDA-MB231 cells as compared to insulin. In contrast, glargine metabolites M1 and M2 showed lower potency for hybrid receptors stimulation, PIP(3 production, Akt and Erk1/2 phosphorylation and DNA synthesis in MCF-7 cells, compared to insulin. CONCLUSION: Glargine, possibly acting through IR/IGF1R hybrids, displays higher potency, whereas its metabolites M1 and M2 display lower potency than insulin for the stimulation of proliferative/anti-apoptotic pathways in

The hybrid MPC-MINLP algorithm for optimal operation of coal-fired power plants with solvent based post-combustion CO2 capture

Directory of Open Access Journals (Sweden)

Norhuda Abdul Manaf

2017-03-01

Full Text Available This paper presents an algorithm that combines model predictive control (MPC with MINLP optimization and demonstrates its application for coal-fired power plants retrofitted with solvent based post-combustion CO2 capture (PCC plant. The objective function of the optimization algorithm works at a primary level to maximize plant economic revenue while considering an optimal carbon capture profile. At a secondary level, the MPC algorithm is used to control the performance of the PCC plant. Two techno-economic scenarios based on fixed (capture rate is constant and flexible (capture rate is variable operation modes are developed using actual electricity prices (2011 with fixed carbon prices ($AUD 5, 25, 50/tonne-CO2 for 24 h periods. Results show that fixed operation mode can bring about a ratio of net operating revenue deficit at an average of 6% against the superior flexible operation mode.

Phenotypic and Genotypic Analysis of Newly Obtained Interspecific Hybrids in the Campanula Genus.

Directory of Open Access Journals (Sweden)

Anna-Catharina Röper

Full Text Available Interspecific hybridisation creates new phenotypes within several ornamental plant species including the Campanula genus. We have employed phenotypic and genotypic methods to analyse and evaluate interspecific hybridisation among cultivars of four Campanula species, i.e. C. cochleariifolia, C. isophylla, C. medium and C. formanekiana. Hybrids were analysed using amplified fragment length polymorphism (AFLP, flow cytometry and biometrical measurements. Results of correlation matrices demonstrated heterogeneous phenotypes for the parental species, which confirmed our basic premise for new phenotypes of interspecific hybrids. AFLP assays confirmed the hybridity and identified self-pollinated plants. Limitation of flow cytometry analysis detection was observed while detecting the hybridity status of two closely related parents, e.g. C. cochleariiafolia × C. isophylla. Phenotypic characteristics such as shoot habitus and flower colour were strongly influenced by one of the parental species in most crosses. Rooting analysis revealed that inferior rooting quality occurred more often in interspecific hybrids than in the parental species. Only interspecific hybrid lines of C. formanekiana 'White' × C. medium 'Pink' showed a high rooting level. Phenotype analyses demonstrated a separation from the interspecific hybrid lines of C. formanekiana 'White' × C. medium 'Pink' to the other clustered hybrids of C. formanekiana and C. medium. In our study we demonstrated that the use of correlation matrices is a suitable tool for identifying suitable cross material. This study presents a comprehensive overview for analysing newly obtained interspecific hybrids. The chosen methods can be used as guidance for analyses for further interspecific hybrids in Campanula, as well as in other ornamental species.

Optimization of ultrasonic array inspections using an efficient hybrid model and real crack shapes

Energy Technology Data Exchange (ETDEWEB)

Felice, Maria V., E-mail: maria.felice@bristol.ac.uk [Department of Mechanical Engineering, University of Bristol, Bristol, U.K. and NDE Laboratory, Rolls-Royce plc., Bristol (United Kingdom); Velichko, Alexander, E-mail: p.wilcox@bristol.ac.uk; Wilcox, Paul D., E-mail: p.wilcox@bristol.ac.uk [Department of Mechanical Engineering, University of Bristol, Bristol (United Kingdom); Barden, Tim; Dunhill, Tony [NDE Laboratory, Rolls-Royce plc., Bristol (United Kingdom)

2015-03-31

Models which simulate the interaction of ultrasound with cracks can be used to optimize ultrasonic array inspections, but this approach can be time-consuming. To overcome this issue an efficient hybrid model is implemented which includes a finite element method that requires only a single layer of elements around the crack shape. Scattering Matrices are used to capture the scattering behavior of the individual cracks and a discussion on the angular degrees of freedom of elastodynamic scatterers is included. Real crack shapes are obtained from X-ray Computed Tomography images of cracked parts and these shapes are inputted into the hybrid model. The effect of using real crack shapes instead of straight notch shapes is demonstrated. An array optimization methodology which incorporates the hybrid model, an approximate single-scattering relative noise model and the real crack shapes is then described.

Fluorescence in situ hybridization for phytoplasma and endophytic bacteria localization in plant tissues.

Science.gov (United States)

Bulgari, Daniela; Casati, Paola; Faoro, Franco

2011-11-01

In the present study, we developed a rapid and efficient fluorescence in situ hybridization assay (FISH) in non-embedded tissues of the model plant Catharanthus roseus for co-localizing phytoplasmas and endophytic bacteria, opening new perspectives for studying the interaction between these microorganisms. Copyright © 2011 Elsevier B.V. All rights reserved.

ZyFISH: a simple, rapid and reliable zygosity assay for transgenic mice.

Directory of Open Access Journals (Sweden)

Donal McHugh

Full Text Available Microinjection of DNA constructs into fertilized mouse oocytes typically results in random transgene integration at a single genomic locus. The resulting transgenic founders can be used to establish hemizygous transgenic mouse lines. However, practical and experimental reasons often require that such lines be bred to homozygosity. Transgene zygosity can be determined by progeny testing assays which are expensive and time-consuming, by quantitative Southern blotting which is labor-intensive, or by quantitative PCR (qPCR which requires transgene-specific design. Here, we describe a zygosity assessment procedure based on fluorescent in situ hybridization (zyFISH. The zyFISH protocol entails the detection of transgenic loci by FISH and the concomitant assignment of homozygosity using a concise and unbiased scoring system. The method requires small volumes of blood, is scalable to at least 40 determinations per assay, and produces results entirely consistent with the progeny testing assay. This combination of reliability, simplicity and cost-effectiveness makes zyFISH a method of choice for transgenic mouse zygosity determinations.

Single-Step Incubation Determination of miRNAs in Cancer Cells Using an Amperometric Biosensor Based on Competitive Hybridization onto Magnetic Beads

Directory of Open Access Journals (Sweden)

Eva Vargas

2018-03-01

Full Text Available This work reports an amperometric biosensor for the determination of miRNA-21, a relevant oncogene. The methodology involves a competitive DNA-target miRNA hybridization assay performed on the surface of magnetic microbeads (MBs and amperometric transduction at screen-printed carbon electrodes (SPCEs. The target miRNA competes with a synthetic fluorescein isothiocyanate (FITC-modified miRNA with an identical sequence for hybridization with a biotinylated and complementary DNA probe (b-Cp immobilized on the surface of streptavidin-modified MBs (b-Cp-MBs. Upon labeling, the FITC-modified miRNA attached to the MBs with horseradish peroxidase (HRP-conjugated anti-FITC Fab fragments and magnetic capturing of the MBs onto the working electrode surface of SPCEs. The cathodic current measured at −0.20 V (versus the Ag pseudo-reference electrode was demonstrated to be inversely proportional to the concentration of the target miRNA. This convenient biosensing method provided a linear range between 0.7 and 10.0 nM and a limit of detection (LOD of 0.2 nM (5 fmol in 25 μL of sample for the synthetic target miRNA without any amplification step. An acceptable selectivity towards single-base mismatched oligonucleotides, a high storage stability of the b-Cp-MBs, and usefulness for the accurate determination of miRNA-21 in raw total RNA (RNAt extracted from breast cancer cells (MCF-7 were demonstrated.

Virologic versus cytologic triage of women with equivocal Pap smears: a meta-analysis of the accuracy to detect high-grade intraepithelial neoplasia.

Science.gov (United States)

Arbyn, Marc; Buntinx, Frank; Van Ranst, Marc; Paraskevaidis, Evangelos; Martin-Hirsch, Pierre; Dillner, Joakim

2004-02-18

The appropriate management of women with minor cytologic lesions in their cervix is unclear. We performed a meta-analysis to assess the accuracy of human papillomavirus (HPV) DNA testing as an alternative to repeat cytology in women who had equivocal results on a previous Pap smear. Data were extracted from articles published between 1992 and 2002 that contained results of virologic and cytologic testing followed by colposcopically directed biopsy in women with an index smear showing atypical cells of undetermined significance (ASCUS). Fifteen studies were identified in which HPV triage and the histologic outcome (presence or absence of a cervical intraepithelial neoplasia of grade II or worse [CIN2+]) was documented. Nine, seven, and two studies also documented the accuracy of repeat cytology when the cutoff for abnormal cytology was set at a threshold of ASCUS or worse, low-grade squamous intraepithelial lesion (LSIL) or worse, or high-grade squamous intraepithelial lesion (HSIL) or worse, respectively. Random-effects models were used for pooling of accuracy parameters in case of interstudy heterogeneity. Differences in accuracy were assessed by pooling the ratio of the sensitivity (or specificity) of HPV testing to that of repeat cytology. The sensitivity and specificity were 84.4% (95% confidence interval [CI] = 77.6% to 91.1%) and 72.9% (95% CI = 62.5% to 83.3%), respectively, for HPV testing overall and 94.8% (95% CI = 92.7% to 96.9%) and 67.3% (95% CI = 58.2% to 76.4%), respectively, for HPV testing in the eight studies that used the Hybrid Capture II assay. Sensitivity and specificity of repeat cytology at a threshold for abnormal cytology of ASCUS or worse was 81.8% (95% CI = 73.5% to 84.3%) and 57.6% (95% CI = 49.5% to 65.7%), respectively. Repeat cytology that used higher cytologic thresholds yielded substantially lower sensitivity but higher specificity than triage with the Hybrid Capture II assay. The ratio of the sensitivity of the Hybrid Capture II

Poly(ethyleneimine) infused and functionalized Torlon®-silica hollow fiber sorbents for post-combustion CO2 capture

KAUST Repository

Li, Fuyue Stephanie

2014-03-01

Organic-inorganic hybrid materials functionalized with amine-containing reagents are emerging as an important class of materials for capturing carbon dioxide from flue gas. Polymeric silica hollow fiber sorbents are fabricated through the proven dry-jet/wet-quench spinning process. In our study, a new technique for functionalizing polymeric silica hollow fiber sorbents with poly(ethyleneimine), followed by a post-spinning infusion step was studied. This two step process introduces a sufficient amount of poly(ethyleneimine) to the polymeric silica hybrid material support to improve the CO2 sorption capacity due to the added amine groups. The poly(ethyleneimine) infused and functionalized hollow fiber sorbents are also characterized by a thermal gravimetric analyzer (TGA) to assess their CO2 sorption capacities. © 2014 Elsevier Ltd. All rights reserved.

Hybrid mimics and hybrid vigor in Arabidopsis

Science.gov (United States)

Wang, Li; Greaves, Ian K.; Groszmann, Michael; Wu, Li Min; Dennis, Elizabeth S.; Peacock, W. James

2015-01-01

F1 hybrids can outperform their parents in yield and vegetative biomass, features of hybrid vigor that form the basis of the hybrid seed industry. The yield advantage of the F1 is lost in the F2 and subsequent generations. In Arabidopsis, from F2 plants that have a F1-like phenotype, we have by recurrent selection produced pure breeding F5/F6 lines, hybrid mimics, in which the characteristics of the F1 hybrid are stabilized. These hybrid mimic lines, like the F1 hybrid, have larger leaves than the parent plant, and the leaves have increased photosynthetic cell numbers, and in some lines, increased size of cells, suggesting an increased supply of photosynthate. A comparison of the differentially expressed genes in the F1 hybrid with those of eight hybrid mimic lines identified metabolic pathways altered in both; these pathways include down-regulation of defense response pathways and altered abiotic response pathways. F6 hybrid mimic lines are mostly homozygous at each locus in the genome and yet retain the large F1-like phenotype. Many alleles in the F6 plants, when they are homozygous, have expression levels different to the level in the parent. We consider this altered expression to be a consequence of transregulation of genes from one parent by genes from the other parent. Transregulation could also arise from epigenetic modifications in the F1. The pure breeding hybrid mimics have been valuable in probing the mechanisms of hybrid vigor and may also prove to be useful hybrid vigor equivalents in agriculture. PMID:26283378

Effect of secondary structure on the thermodynamics and kinetics of PNA hybridization to DNA hairpins

DEFF Research Database (Denmark)

Kushon, S A; Jordan, J P; Seifert, J L

2001-01-01

The binding of a series of PNA and DNA probes to a group of unusually stable DNA hairpins of the tetraloop motif has been observed using absorbance hypochromicity (ABS), circular dichroism (CD), and a colorimetric assay for PNA/DNA duplex detection. These results indicate that both stable PNA...... structures in both target and probe molecules are shown to depress the melting temperatures and free energies of the probe-target duplexes. Kinetic analysis of hybridization yields reaction rates that are up to 160-fold slower than hybridization between two unstructured strands. The thermodynamic and kinetic...

Detection and quantification of Epstein-Barr virus EBER1 in EBV-infected cells by fluorescent in situ hybridization and flow cytometry

Science.gov (United States)

Stowe, R. P.; Cubbage, M. L.; Sams, C. F.; Pierson, D. L.; Barrett, A. D.

1998-01-01

A rapid and highly sensitive fluorescent in situ hybridization (FISH) assay was developed to detect Epstein Barr virus (EBV)-infected cells in peripheral blood. Multiple fluorescein-labeled antisense oligonucleotide probes were designed to hybridize to the EBER1 transcript, which is highly expressed in latently infected cells. After a rapid (30 min) hybridization, the cells were analyzed by flow cytometry. EBER1 was detected in several positive control cell lines that have variable numbers of EBV genome copies. No EBER1 was detected in two known EBV-negative cell lines. Northern blot analyses confirmed the presence and quantity of EBER1 transcripts in each cell line. This method was used to quantify the number of EBV-infected cells in peripheral blood from a patient with chronic mononucleosis. These results indicate that EBV-infected cells can be detected at the single cell level, and that this assay can be used to quantify the number of EBV-infected cells in clinical samples.

A Low Cost Air Hybrid Concept Un concept hybride à air et à bas prix

Directory of Open Access Journals (Sweden)

Lee C. Y.

2010-02-01

Full Text Available The air hybrid engine absorbs the vehicle kinetic energy during braking, stores it in an air tank in the form of compressed air, and reuses it to propel a vehicle during cruising and acceleration. Capturing, storing and reusing this braking energy to give additional power can therefore improve fuel economy, particularly in cities and urban areas where the traffic conditions involve many stops and starts. In order to reuse the residual kinetic energy, the vehicle operation consists of 3 basic modes, i.e. Compression Mode (CM, Expander Mode (EM and normal firing mode. Unlike previous works, a low cost air hybrid engine has been proposed and studied. The hybrid engine operation can be realized by means of production technologies, such as VVT and valve deactivation. In this work, systematic investigation has been carried out on the performance of the hybrid engine concept through detailed gas dynamic modelling using Ricardo WAVE software. Valve timing optimization has been done for the more efficient operation of air hybrid operation and obtaining higher braking and motoring mean effective pressure for CM and EM respectively. Le moteur hybride à air absorbe l’énergie cinétique du véhicule en cas de freinage, la stocke sous forme d’air comprimé, puis la réutilise pour faire avancer le véhicule en circulation ou en accélération. Capter, stocker et réutiliser cette énergie et créer ainsi une puissance plus importante peut donc permettre de plus grandes économies de carburant, surtout en ville et zone urbaine, lorsque les conditions de circulation imposent de nombreux arrêts et démarrages. Pour pouvoir réutiliser l’énergie cinétique, on distingue trois modes de base d’utilisation du véhicule : les modes compression (MC, expansion (ME et standard. Un moteur hybride à air et à bas prix a été proposé et étudié. Un tel moteur peut être opéré par le biais de technologies de production, comme le VVT et la d

Synovectomy by Neutron capture; Sinovectomia por captura de neutrones

Energy Technology Data Exchange (ETDEWEB)

Vega C, H.R.; Torres M, C. [Centro Regional de Estudios Nucleares, Universidad Autonoma de Zacatecas, C. Cipres 10, Fracc. La Penuela, 98000 Zacatecas (Mexico)

1998-12-31

The Synovectomy by Neutron capture has as purpose the treatment of the rheumatoid arthritis, illness which at present does not have a definitive curing. This therapy requires a neutron source for irradiating the articulation affected. The energy spectra and the intensity of these neutrons are fundamental since these neutrons induce nuclear reactions of capture with Boron-10 inside the articulation and the freely energy of these reactions is transferred at the productive tissue of synovial liquid, annihilating it. In this work it is presented the neutron spectra results obtained with moderator packings of spherical geometry which contains in its center a Pu{sup 239} Be source. The calculations were realized through Monte Carlo method. The moderators assayed were light water, heavy water base and the both combination of them. The spectra obtained, the average energy, the neutron total number by neutron emitted by source, the thermal neutron percentage and the dose equivalent allow us to suggest that the moderator packing more adequate is what has a light water thickness 0.5 cm (radius 2 cm) and 24.5 cm heavy water (radius 26.5 cm). (Author)

On the Mechanism of Berberine-INF55 (5-Nitro-2-phenylindole) Hybrid Antibacterials.

Science.gov (United States)

Dolla, Naveen K; Chen, Chao; Larkins-Ford, Jonah; Rajamuthiah, Rajmohan; Jagadeesan, Sakthimala; Conery, Annie L; Ausubel, Frederick M; Mylonakis, Eleftherios; Bremner, John B; Lewis, Kim; Kelso, Michael J

Berberine-INF55 hybrids are a promising class of antibacterials that combine berberine and the NorA multidrug resistance pump inhibitor INF55 (5-nitro-2-phenylindole) together in one molecule via a chemically stable linkage. Previous studies demonstrated the potential of these compounds for countering efflux-mediated antibacterial drug resistance but they didn't establish whether the compounds function as originally intended, i.e. with the berberine moiety providing antibacterial activity and the attached INF55 component independently blocking multidrug resistance pumps, thereby enhancing the activity of berberine by reducing its efflux. We hypothesised that if the proposed mechanism is correct, then hybrids carrying more potent INF55 pump inhibitor structures should show enhanced antibacterial effects relative to those bearing weaker inhibitors. Two INF55 analogues showing graded reductions in NorA inhibitory activity compared with INF55 were identified and their corresponding berberine-INF55 hybrids carrying equivalent INF55 inhibitor structures synthesised. Multiple assays comparing the antibacterial effects of the hybrids and their corresponding berberine-INF55 analogue combinations showed that the three hybrids all show very similar activities, leading us to conclude that the antibacterial mechanism(s) of berberine-INF55 hybrids is different from berberine-INF55 combinations.

Multiplex bio-assay with inductively coupled plasma mass spectrometry: Towards a massively multivariate single-cell technology

International Nuclear Information System (INIS)

Tanner, Scott D.; Ornatsky, Olga; Bandura, Dmitry R.; Baranov, Vladimir I.

2007-01-01

Recent progress in the development of massively multiplexed bioanalytical assays using element tags with inductively coupled plasma mass spectrometry detection is reviewed. Feasibility results using commercially available secondary immunolabeling reagents for leukemic cell lines are presented. Multiplex analysis of higher order is shown with first generation tag reagents based on functionalized carriers that bind lanthanide ions. DNA quantification using metallointercalation allows for cell enumeration or mitotic state differentiation. In situ hybridization permits the determination of cellular RNA. The results provide a feasibility basis for the development of a multivariate assay tool for individual cell analysis based on inductively coupled plasma mass spectrometry in a cytometer configuration

Multiplex bio-assay with inductively coupled plasma mass spectrometry: Towards a massively multivariate single-cell technology

Energy Technology Data Exchange (ETDEWEB)

Tanner, Scott D. [Institute of Biomaterials and Biomedical Engineering, University of Toronto, Room 407, 164 College Street, Toronto, Ontario, M5S 3G9 (Canada)], E-mail: sd.tanner@utoronto.ca; Ornatsky, Olga; Bandura, Dmitry R.; Baranov, Vladimir I. [Institute of Biomaterials and Biomedical Engineering, University of Toronto, Room 407, 164 College Street, Toronto, Ontario, M5S 3G9 (Canada)

2007-03-15

Recent progress in the development of massively multiplexed bioanalytical assays using element tags with inductively coupled plasma mass spectrometry detection is reviewed. Feasibility results using commercially available secondary immunolabeling reagents for leukemic cell lines are presented. Multiplex analysis of higher order is shown with first generation tag reagents based on functionalized carriers that bind lanthanide ions. DNA quantification using metallointercalation allows for cell enumeration or mitotic state differentiation. In situ hybridization permits the determination of cellular RNA. The results provide a feasibility basis for the development of a multivariate assay tool for individual cell analysis based on inductively coupled plasma mass spectrometry in a cytometer configuration.

Capturing Thoughts, Capturing Minds?

DEFF Research Database (Denmark)

Nielsen, Janni

2004-01-01

Think Aloud is cost effective, promises access to the user's mind and is the applied usability technique. But 'keep talking' is difficult, besides, the multimodal interface is visual not verbal. Eye-tracking seems to get around the verbalisation problem. It captures the visual focus of attention...

Ultra-sensitive DNA assay based on single-molecule detection coupled with fluorescent quantum dot-labeling and its application to determination of messenger RNA

Energy Technology Data Exchange (ETDEWEB)

Li Li [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Li Xincang [School of Life Sciences, Shandong University, Jinan 250100 (China); Li Lu [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Wang Jinxing [School of Life Sciences, Shandong University, Jinan 250100 (China); Jin Wenrui, E-mail: jwr@sdu.edu.cn [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

2011-01-24

An ultra-sensitive single-molecule detection (SMD) method for quantification of DNA using total internal reflection fluorescence microscopy (TIRFM) coupled with fluorescent quantum dot (QD)-labeling was developed. In this method, the target DNA (tDNA) was captured by the capture DNA immobilized on the silanized coverslip blocked with ethanolamine and bovine serum albumin. Then, the QD-labeled probe DNA was hybridized to the tDNA. Ten fluorescent images of the QD-labeled sandwich DNA hybrids on the coverslip were taken by a high-sensitive CCD. The tDNA was quantified by counting the bright spots on the images using a calibration curve. The LOD of the method was 1 x 10{sup -14} mol L{sup -1}. Several key factors, including image acquirement, fluorescence probe, substrate preparation, noise elimination from solutions and glass coverslips, and nonspecific adsorption and binding of solution-phase detection probes were discussed in detail. The method could be applied to quantify messenger RNA (mRNA) in cells. In order to determine mRNA, double-stranded RNA-DNA hybrids consisting of mRNA and corresponding cDNA were synthesized from the cellular mRNA template using reverse transcription in the presence of reverse transcriptase. After removing the mRNA in the double-stranded hybrids using ribonuclease, cDNA was quantified using the SMD-based TIRFM. Osteopontin mRNA in decidual stromal cells was chosen as the model analyte.

Fuel economy of hybrid fuel-cell vehicles

Science.gov (United States)

Ahluwalia, Rajesh K.; Wang, X.; Rousseau, A.

The potential improvement in fuel economy of a mid-size fuel-cell vehicle by combining it with an energy storage system has been assessed. An energy management strategy is developed and used to operate the direct hydrogen, pressurized fuel-cell system in a load-following mode and the energy storage system in a charge-sustaining mode. The strategy places highest priority on maintaining the energy storage system in a state where it can supply unanticipated boost power when the fuel-cell system alone cannot meet the power demand. It is found that downsizing a fuel-cell system decreases its efficiency on a drive cycle which is compensated by partial regenerative capture of braking energy. On a highway cycle with limited braking energy the increase in fuel economy with hybridization is small but on the stop-and-go urban cycle the fuel economy can improve by 27%. On the combined highway and urban drive cycles the fuel economy of the fuel-cell vehicle is estimated to increase by up to 15% by hybridizing it with an energy storage system.

Acetohydroxyacid synthase (AHAS) in vivo assay for screening imidazolinone-resistance in sunflower (Helianthus annuus L.).

Science.gov (United States)

Vega, T; Breccia, G; Gil, M; Zorzoli, R; Picardi, L; Nestares, G

2012-12-01

The objective of this work was to evaluate the in vivo acetohydroxyacid synthase (AHAS) activity response to imidazolinones and its possible use as a selection method for evaluating AHAS inhibitor resistance. In vivo AHAS assay and the comparison of parameters from dose-response curves have been used as a valid tool for comparing sunflower lines and hybrids differing in imidazolinone resistance. The sunflower resistant genotypes evaluated here were 100-fold and 20-fold more resistant compared with the susceptible line for imazethapyr and imazapyr, respectively. This assay also allowed discrimination of homozygous from heterozygous genotypes for I(mr1) locus that codify for the catalytic subunit of AHAS. The in vivo AHAS assay described in this study was useful for the selection of sunflower genotypes differing in herbicide resistance and could be a useful tool when breeding for imidazolinone resistance in sunflower. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Performance Evaluation of the VIDAS® Measles IgG Assay and Its Diagnostic Value for Measuring IgG Antibody Avidity in Measles Virus Infection

Science.gov (United States)

Dina, Julia; Creveuil, Christian; Gouarin, Stephanie; Viron, Florent; Hebert, Amelie; Freymuth, Francois; Vabret, Astrid

2016-01-01

The objective of this study is primarily to compare the performance of the VIDAS® Measles immunoglobulin (Ig)G assay to that of two other serological assays using an immunoassay technique, Enzygnost® Anti-measles Virus/IgG (Siemens) and Measles IgG CAPTURE EIA® (Microimmune). The sensitivity and the agreement of the VIDAS® Measles IgG assay compared to the Enzygnost® Anti-measles Virus/IgG assay and the Measles IgG CAPTURE EIA® assay are 100%, 97.2% and 99.0%, 98.4%, respectively. The very low number of negative sera for IgG antibodies does not allow calculation of specificity. As a secondary objective, we have evaluated the ability of the VIDAS® Measles IgG assay to measure anti-measles virus IgG antibody avidity with the help of the VIDAS® CMV IgG Avidity reagent, using 76 sera from subjects with measles and 238 other sera. Different groups of populations were analyzed. In the primary infection measles group, the mean IgG avidity index was 0.16 (range of 0.07 to 0.93) compared to 0.79 (range of 0.25 to 1) in the serum group positive for IgG antibodies and negative for IgM. These data allow to define a weak anti-measles virus IgG antibody avidity as an avidity index (AI) 0.6. The VIDAS® Measles IgG assay has a performance equivalent to that of other available products. Its use, individual and quick, is well adapted to testing for anti-measles immunity in exposed subjects. PMID:27556477

Thermodynamic Modeling and Dispatch of Distributed Energy Technologies including Fuel Cell -- Gas Turbine Hybrids

Science.gov (United States)

McLarty, Dustin Fogle

Distributed energy systems are a promising means by which to reduce both emissions and costs. Continuous generators must be responsive and highly efficiency to support building dynamics and intermittent on-site renewable power. Fuel cell -- gas turbine hybrids (FC/GT) are fuel-flexible generators capable of ultra-high efficiency, ultra-low emissions, and rapid power response. This work undertakes a detailed study of the electrochemistry, chemistry and mechanical dynamics governing the complex interaction between the individual systems in such a highly coupled hybrid arrangement. The mechanisms leading to the compressor stall/surge phenomena are studied for the increased risk posed to particular hybrid configurations. A novel fuel cell modeling method introduced captures various spatial resolutions, flow geometries, stack configurations and novel heat transfer pathways. Several promising hybrid configurations are analyzed throughout the work and a sensitivity analysis of seven design parameters is conducted. A simple estimating method is introduced for the combined system efficiency of a fuel cell and a turbine using component performance specifications. Existing solid oxide fuel cell technology is capable of hybrid efficiencies greater than 75% (LHV) operating on natural gas, and existing molten carbonate systems greater than 70% (LHV). A dynamic model is calibrated to accurately capture the physical coupling of a FC/GT demonstrator tested at UC Irvine. The 2900 hour experiment highlighted the sensitivity to small perturbations and a need for additional control development. Further sensitivity studies outlined the responsiveness and limits of different control approaches. The capability for substantial turn-down and load following through speed control and flow bypass with minimal impact on internal fuel cell thermal distribution is particularly promising to meet local demands or provide dispatchable support for renewable power. Advanced control and dispatch

Colorimetric microwell plate reverse-hybridization assay for Mycobacterium tuberculosis detection

Directory of Open Access Journals (Sweden)

Candice Tosi Michelon

2011-03-01

Full Text Available Direct smear examination using Ziehl-Neelsen staining for pulmonary tuberculosis (PTB diagnosis is inexpensive and easy to use, but has the major limitation of low sensitivity. Rapid molecular methods are becoming more widely available in centralized laboratories, but they depend on timely reporting of results and strict quality assurance obtainable only from costly commercial kits available in high burden nations. This study describes a pre-commercial colorimetric method, Detect-TB, for detecting Mycobacterium tuberculosis DNA in which an oligonucleotide probe is fixed onto wells of microwell plates and hybridized with biotinylated polymerase chain reaction amplification products derived from clinical samples. The probe is capable of hybridising with the IS6110 insertion element and was used to specifically recognise the M. tuberculosis complex. When combined with an improved silica-based DNA extraction method, the sensitivity of the test was 50 colony-forming units of the M. tuberculosis reference strain H37Rv. The results that were in agreement with reference detection methods were observed in 95.2% (453/476 of samples included in the analysis. Sensitivity and specificity for 301 induced sputum samples and 175 spontaneous sputum samples were 85% and 98%, and 94% and 100%, respectively. This colorimetric method showed similar specificity to that described for commercially available kits and may provide an important contribution for PTB diagnosis.

A hybrid agent-based approach for modeling microbiological systems.

Science.gov (United States)

Guo, Zaiyi; Sloot, Peter M A; Tay, Joc Cing

2008-11-21

Models for systems biology commonly adopt Differential Equations or Agent-Based modeling approaches for simulating the processes as a whole. Models based on differential equations presuppose phenomenological intracellular behavioral mechanisms, while models based on Multi-Agent approach often use directly translated, and quantitatively less precise if-then logical rule constructs. We propose an extendible systems model based on a hybrid agent-based approach where biological cells are modeled as individuals (agents) while molecules are represented by quantities. This hybridization in entity representation entails a combined modeling strategy with agent-based behavioral rules and differential equations, thereby balancing the requirements of extendible model granularity with computational tractability. We demonstrate the efficacy of this approach with models of chemotaxis involving an assay of 10(3) cells and 1.2x10(6) molecules. The model produces cell migration patterns that are comparable to laboratory observations.

Lack of detection of human papillomavirus infection by hybridization test in prostatic biopsies

International Nuclear Information System (INIS)

Gazzaz, Faten S; Mosli, Hisham A

2009-01-01

To explore the possibility of finding human papillomavirus (HPV) infection in the prostate tissue of a cohort of Saudi men presenting with benign prostatic hyperplasia (BPH) or prostate cancer. A cohort study on prospectively collected tissue samples was conducted at King Abdulaziz University Hospital (KAUH), Jeddah, Kingdom of Saudi Arabia from March 2007 to December 2008 on a total of 56 male patients, age range 50-93 years (average 68), diagnosed as having BPH or prostate cancer. The HPV DNA hybridization by hybrid capture 2 technology was performed on prostate biopsies of these patients to detect 18 types of HPV infection, and differentiate between 2 HPV DNA groups, the low-risk types, and the high/intermediate risk types.The tissues of all the prostatic biopsies were negative for HPV DNA. Our results, using the hybridization test, indicate that it is unlikely that HPV-16 or HPV-18, or the other tested subtypes, enhance the risk of prostate cancer. (author)

Micropatterned comet assay enables high throughput and sensitive DNA damage quantification.

Science.gov (United States)

Ge, Jing; Chow, Danielle N; Fessler, Jessica L; Weingeist, David M; Wood, David K; Engelward, Bevin P

2015-01-01

The single cell gel electrophoresis assay, also known as the comet assay, is a versatile method for measuring many classes of DNA damage, including base damage, abasic sites, single strand breaks and double strand breaks. However, limited throughput and difficulties with reproducibility have limited its utility, particularly for clinical and epidemiological studies. To address these limitations, we created a microarray comet assay. The use of a micrometer scale array of cells increases the number of analysable comets per square centimetre and enables automated imaging and analysis. In addition, the platform is compatible with standard 24- and 96-well plate formats. Here, we have assessed the consistency and sensitivity of the microarray comet assay. We showed that the linear detection range for H2O2-induced DNA damage in human lymphoblastoid cells is between 30 and 100 μM, and that within this range, inter-sample coefficient of variance was between 5 and 10%. Importantly, only 20 comets were required to detect a statistically significant induction of DNA damage for doses within the linear range. We also evaluated sample-to-sample and experiment-to-experiment variation and found that for both conditions, the coefficient of variation was lower than what has been reported for the traditional comet assay. Finally, we also show that the assay can be performed using a 4× objective (rather than the standard 10× objective for the traditional assay). This adjustment combined with the microarray format makes it possible to capture more than 50 analysable comets in a single image, which can then be automatically analysed using in-house software. Overall, throughput is increased more than 100-fold compared to the traditional assay. Together, the results presented here demonstrate key advances in comet assay technology that improve the throughput, sensitivity, and robustness, thus enabling larger scale clinical and epidemiological studies. © The Author 2014. Published by

Improvement of non-destructive fissile mass assays in α low-level waste drums: A matrix correction method based on neutron capture gamma-rays and a neutron generator

Science.gov (United States)

Jallu, F.; Loche, F.

2008-08-01

Within the framework of radioactive waste control, non-destructive assay (NDA) methods may be employed. The active neutron interrogation (ANI) method is now well-known and effective in quantifying low α-activity fissile masses (mainly 235U, 239Pu, 241Pu) with low densities, i.e. less than about 0.4, in radioactive waste drums of volumes up to 200 l. The PROMpt Epithermal and THErmal interrogation Experiment (PROMETHEE [F. Jallu, A. Mariani, C. Passard, A.-C. Raoux, H. Toubon, Alpha low level waste control: improvement of the PROMETHEE 6 assay system performances. Nucl. Technol. 153 (January) (2006); C. Passard, A. Mariani, F. Jallu, J. Romeyer-Dherber, H. Recroix, M. Rodriguez, J. Loridon, C. Denis, PROMETHEE: an alpha low level waste assay system using passive and active neutron measurement methods. Nucl. Technol. 140 (December) (2002) 303-314]) based on ANI has been under development since 1996 to reach the incinerating α low level waste (LLW) criterion of about 50 Bq[α] per gram of crude waste (≈50 μg Pu) in 118 l drums on the date the drums are conditioned. Difficulties arise when dealing with matrices containing neutron energy moderators such as H and neutron absorbents such as Cl. These components may have a great influence on the fissile mass deduced from the neutron signal measured by ANI. For example, the calibration coefficient measured in a 118 l drum containing a cellulose matrix (density d = 0.144 g cm -3) may be 50 times higher than that obtained in a poly-vinyl-chloride matrix ( d = 0.253 g cm -3). Without any information on the matrix, the fissile mass is often overestimated due to safety procedures and by considering the most disadvantageous calibration coefficient corresponding to the most absorbing and moderating calibration matrix. The work discussed in this paper was performed at the CEA Nuclear Measurement Laboratory in France. It concerns the development of a matrix effect correction method, which consists in identifying and quantifying

Improvement of non-destructive fissile mass assays in α low-level waste drums: A matrix correction method based on neutron capture gamma-rays and a neutron generator

International Nuclear Information System (INIS)

Jallu, F.; Loche, F.

2008-01-01

Within the framework of radioactive waste control, non-destructive assay (NDA) methods may be employed. The active neutron interrogation (ANI) method is now well-known and effective in quantifying low α-activity fissile masses (mainly 235 U, 239 Pu, 241 Pu) with low densities, i.e. less than about 0.4, in radioactive waste drums of volumes up to 200 l. The PROMpt Epithermal and THErmal interrogation Experiment (PROMETHEE [F. Jallu, A. Mariani, C. Passard, A.-C. Raoux, H. Toubon, Alpha low level waste control: improvement of the PROMETHEE 6 assay system performances. Nucl. Technol. 153 (January) (2006); C. Passard, A. Mariani, F. Jallu, J. Romeyer-Dherber, H. Recroix, M. Rodriguez, J. Loridon, C. Denis, PROMETHEE: an alpha low level waste assay system using passive and active neutron measurement methods. Nucl. Technol. 140 (December) (2002) 303-314]) based on ANI has been under development since 1996 to reach the incinerating α low level waste (LLW) criterion of about 50 Bq[α] per gram of crude waste (∼50 μg Pu) in 118 l drums on the date the drums are conditioned. Difficulties arise when dealing with matrices containing neutron energy moderators such as H and neutron absorbents such as Cl. These components may have a great influence on the fissile mass deduced from the neutron signal measured by ANI. For example, the calibration coefficient measured in a 118 l drum containing a cellulose matrix (density d = 0.144 g cm -3 ) may be 50 times higher than that obtained in a poly-vinyl-chloride matrix (d = 0.253 g cm -3 ). Without any information on the matrix, the fissile mass is often overestimated due to safety procedures and by considering the most disadvantageous calibration coefficient corresponding to the most absorbing and moderating calibration matrix. The work discussed in this paper was performed at the CEA Nuclear Measurement Laboratory in France. It concerns the development of a matrix effect correction method, which consists in identifying and

Improvement of non-destructive fissile mass assays in {alpha} low-level waste drums: A matrix correction method based on neutron capture gamma-rays and a neutron generator

Energy Technology Data Exchange (ETDEWEB)

Jallu, F. [Commissariat a l' Energie Atomique, CEA, DEN, Nuclear Measurement Laboratory, Bat. 224, 13108 Saint Paul lez Durance (France)], E-mail: fanny.jallu@cea.fr; Loche, F. [Commissariat a l' Energie Atomique, CEA, DEN, Nuclear Measurement Laboratory, Bat. 224, 13108 Saint Paul lez Durance (France)

2008-08-15

Within the framework of radioactive waste control, non-destructive assay (NDA) methods may be employed. The active neutron interrogation (ANI) method is now well-known and effective in quantifying low {alpha}-activity fissile masses (mainly {sup 235}U, {sup 239}Pu, {sup 241}Pu) with low densities, i.e. less than about 0.4, in radioactive waste drums of volumes up to 200 l. The PROMpt Epithermal and THErmal interrogation Experiment (PROMETHEE [F. Jallu, A. Mariani, C. Passard, A.-C. Raoux, H. Toubon, Alpha low level waste control: improvement of the PROMETHEE 6 assay system performances. Nucl. Technol. 153 (January) (2006); C. Passard, A. Mariani, F. Jallu, J. Romeyer-Dherber, H. Recroix, M. Rodriguez, J. Loridon, C. Denis, PROMETHEE: an alpha low level waste assay system using passive and active neutron measurement methods. Nucl. Technol. 140 (December) (2002) 303-314]) based on ANI has been under development since 1996 to reach the incinerating {alpha} low level waste (LLW) criterion of about 50 Bq[{alpha}] per gram of crude waste ({approx}50 {mu}g Pu) in 118 l drums on the date the drums are conditioned. Difficulties arise when dealing with matrices containing neutron energy moderators such as H and neutron absorbents such as Cl. These components may have a great influence on the fissile mass deduced from the neutron signal measured by ANI. For example, the calibration coefficient measured in a 118 l drum containing a cellulose matrix (density d = 0.144 g cm{sup -3}) may be 50 times higher than that obtained in a poly-vinyl-chloride matrix (d = 0.253 g cm{sup -3}). Without any information on the matrix, the fissile mass is often overestimated due to safety procedures and by considering the most disadvantageous calibration coefficient corresponding to the most absorbing and moderating calibration matrix. The work discussed in this paper was performed at the CEA Nuclear Measurement Laboratory in France. It concerns the development of a matrix effect correction

Carbon captured from the air

International Nuclear Information System (INIS)

Keith, D.

2008-01-01

This article presented an innovative way to achieve the efficient capture of atmospheric carbon. A team of scientists from the University of Calgary's Institute for Sustainable Energy, Environment and Economy have shown that it is possible to reduce carbon dioxide (CO 2 ) using a simple machine that can capture the trace amount of CO 2 present in ambient air at any place on the planet. The thermodynamics of capturing the small concentrations of CO 2 from the air is only slightly more difficult than capturing much larger concentrations of CO 2 from power plants. The research is significant because it offers a way to capture CO 2 emissions from transportation sources such as vehicles and airplanes, which represent more than half of the greenhouse gases emitted on Earth. The energy efficient and cost effective air capture technology could complement other approaches for reducing emissions from the transportation sector, such as biofuels and electric vehicles. Air capture differs from carbon capture and storage (CCS) technology used at coal-fired power plants where CO 2 is captured and pipelined for permanent storage underground. Air capture can capture the CO 2 that is present in ambient air and store it wherever it is cheapest. The team at the University of Calgary showed that CO 2 could be captured directly from the air with less than 100 kWhrs of electricity per tonne of CO 2 . A custom-built tower was able to capture the equivalent of 20 tonnes per year of CO 2 on a single square meter of scrubbing material. The team devised a way to use a chemical process from the pulp and paper industry to cut the energy cost of air capture in half. Although the technology is only in its early stage, it appears that CO 2 could be captured from the air with an energy demand comparable to that needed for CO 2 capture from conventional power plants, but costs will be higher. The simple, reliable and scalable technology offers an opportunity to build a commercial-scale plant. 1 fig

Degradation of Akt using protein-catalyzed capture agents.

Science.gov (United States)

Henning, Ryan K; Varghese, Joseph O; Das, Samir; Nag, Arundhati; Tang, Grace; Tang, Kevin; Sutherland, Alexander M; Heath, James R

2016-04-01

Abnormal signaling of the protein kinase Akt has been shown to contribute to human diseases such as diabetes and cancer, but Akt has proven to be a challenging target for drugging. Using iterative in situ click chemistry, we recently developed multiple protein-catalyzed capture (PCC) agents that allosterically modulate Akt enzymatic activity in a protein-based assay. Here, we utilize similar PCCs to exploit endogenous protein degradation pathways. We use the modularity of the anti-Akt PCCs to prepare proteolysis targeting chimeric molecules that are shown to promote the rapid degradation of Akt in live cancer cells. These novel proteolysis targeting chimeric molecules demonstrate that the epitope targeting selectivity of PCCs can be coupled with non-traditional drugging moieties to inhibit challenging targets. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.

Identification of a novel interspecific hybrid yeast from a metagenomic spontaneously inoculated beer sample using Hi-C.

Science.gov (United States)

Smukowski Heil, Caiti; Burton, Joshua N; Liachko, Ivan; Friedrich, Anne; Hanson, Noah A; Morris, Cody L; Schacherer, Joseph; Shendure, Jay; Thomas, James H; Dunham, Maitreya J

2018-01-01

Interspecific hybridization is a common mechanism enabling genetic diversification and adaptation; however, the detection of hybrid species has been quite difficult. The identification of microbial hybrids is made even more complicated, as most environmental microbes are resistant to culturing and must be studied in their native mixed communities. We have previously adapted the chromosome conformation capture method Hi-C to the assembly of genomes from mixed populations. Here, we show the method's application in assembling genomes directly from an uncultured, mixed population from a spontaneously inoculated beer sample. Our assembly method has enabled us to de-convolute four bacterial and four yeast genomes from this sample, including a putative yeast hybrid. Downstream isolation and analysis of this hybrid confirmed its genome to consist of Pichia membranifaciens and that of another related, but undescribed, yeast. Our work shows that Hi-C-based metagenomic methods can overcome the limitation of traditional sequencing methods in studying complex mixtures of genomes. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

A hybrid approach to device integration on a genetic analysis platform

International Nuclear Information System (INIS)

Brennan, Des; Justice, John; Aherne, Margaret; Galvin, Paul; Jary, Dorothee; Kurg, Ants; Berik, Evgeny; Macek, Milan

2012-01-01

Point-of-care (POC) systems require significant component integration to implement biochemical protocols associated with molecular diagnostic assays. Hybrid platforms where discrete components are combined in a single platform are a suitable approach to integration, where combining multiple device fabrication steps on a single substrate is not possible due to incompatible or costly fabrication steps. We integrate three devices each with a specific system functionality: (i) a silicon electro-wetting-on-dielectric (EWOD) device to move and mix sample and reagent droplets in an oil phase, (ii) a polymer microfluidic chip containing channels and reservoirs and (iii) an aqueous phase glass microarray for fluorescence microarray hybridization detection. The EWOD device offers the possibility of fully integrating on-chip sample preparation using nanolitre sample and reagent volumes. A key challenge is sample transfer from the oil phase EWOD device to the aqueous phase microarray for hybridization detection. The EWOD device, waveguide performance and functionality are maintained during the integration process. An on-chip biochemical protocol for arrayed primer extension (APEX) was implemented for single nucleotide polymorphism (SNiP) analysis. The prepared sample is aspirated from the EWOD oil phase to the aqueous phase microarray for hybridization. A bench-top instrumentation system was also developed around the integrated platform to drive the EWOD electrodes, implement APEX sample heating and image the microarray after hybridization. (paper)

Capture by colour: evidence for dimension-specific singleton capture.

Science.gov (United States)

Harris, Anthony M; Becker, Stefanie I; Remington, Roger W

2015-10-01

Previous work on attentional capture has shown the attentional system to be quite flexible in the stimulus properties it can be set to respond to. Several different attentional "modes" have been identified. Feature search mode allows attention to be set for specific features of a target (e.g., red). Singleton detection mode sets attention to respond to any discrepant item ("singleton") in the display. Relational search sets attention for the relative properties of the target in relation to the distractors (e.g., redder, larger). Recently, a new attentional mode was proposed that sets attention to respond to any singleton within a particular feature dimension (e.g., colour; Folk & Anderson, 2010). We tested this proposal against the predictions of previously established attentional modes. In a spatial cueing paradigm, participants searched for a colour target that was randomly either red or green. The nature of the attentional control setting was probed by presenting an irrelevant singleton cue prior to the target display and assessing whether it attracted attention. In all experiments, the cues were red, green, blue, or a white stimulus rapidly rotated (motion cue). The results of three experiments support the existence of a "colour singleton set," finding that all colour cues captured attention strongly, while motion cues captured attention only weakly or not at all. Notably, we also found that capture by motion cues in search for colour targets was moderated by their frequency; rare motion cues captured attention (weakly), while frequent motion cues did not.

Materials For Gas Capture, Methods Of Making Materials For Gas Capture, And Methods Of Capturing Gas

KAUST Repository

Polshettiwar, Vivek

2013-06-20

In accordance with the purpose(s) of the present disclosure, as embodied and broadly described herein, embodiments of the present disclosure, in one aspect, relate to materials that can be used for gas (e.g., CO.sub.2) capture, methods of making materials, methods of capturing gas (e.g., CO.sub.2), and the like, and the like.

Determination for Enterobacter cloacae based on a europium ternary complex labeled DNA probe

Science.gov (United States)

He, Hui; Niu, Cheng-Gang; Zeng, Guang-Ming; Ruan, Min; Qin, Pin-Zhu; Liu, Jing

2011-11-01

The fast detection and accurate diagnosis of the prevalent pathogenic bacteria is very important for the treatment of disease. Nowadays, fluorescence techniques are important tools for diagnosis. A two-probe tandem DNA hybridization assay was designed for the detection of Enterobacter cloacae based on time-resolved fluorescence. In this work, the authors synthesized a novel europium ternary complex Eu(TTA) 3(5-NH 2-phen) with intense luminescence, high fluorescence quantum yield and long lifetime before. We developed a method based on this europium complex for the specific detection of original extracted DNA from E. cloacae. In the hybridization assay format, the reporter probe was labeled with Eu(TTA) 3(5-NH 2-phen) on the 5'-terminus, and the capture probe capture probe was covalent immobilized on the surface of the glutaraldehyde treated glass slides. The original extracted DNA of samples was directly used without any DNA purification and amplification. The detection was conducted by monitoring the fluorescence intensity from the glass surface after DNA hybridization. The detection limit of the DNA was 5 × 10 -10 mol L -1. The results of the present work proved that this new approach was easy to operate with high sensitivity and specificity. It could be conducted as a powerful tool for the detection of pathogen microorganisms in the environment.

Hybrid epidemics--a case study on computer worm conficker.

Directory of Open Access Journals (Sweden)

Changwang Zhang

Full Text Available Conficker is a computer worm that erupted on the Internet in 2008. It is unique in combining three different spreading strategies: local probing, neighbourhood probing, and global probing. We propose a mathematical model that combines three modes of spreading: local, neighbourhood, and global, to capture the worm's spreading behaviour. The parameters of the model are inferred directly from network data obtained during the first day of the Conficker epidemic. The model is then used to explore the tradeoff between spreading modes in determining the worm's effectiveness. Our results show that the Conficker epidemic is an example of a critically hybrid epidemic, in which the different modes of spreading in isolation do not lead to successful epidemics. Such hybrid spreading strategies may be used beneficially to provide the most effective strategies for promulgating information across a large population. When used maliciously, however, they can present a dangerous challenge to current internet security protocols.

Hybrid epidemics--a case study on computer worm conficker.

Science.gov (United States)

Zhang, Changwang; Zhou, Shi; Chain, Benjamin M

2015-01-01

Conficker is a computer worm that erupted on the Internet in 2008. It is unique in combining three different spreading strategies: local probing, neighbourhood probing, and global probing. We propose a mathematical model that combines three modes of spreading: local, neighbourhood, and global, to capture the worm's spreading behaviour. The parameters of the model are inferred directly from network data obtained during the first day of the Conficker epidemic. The model is then used to explore the tradeoff between spreading modes in determining the worm's effectiveness. Our results show that the Conficker epidemic is an example of a critically hybrid epidemic, in which the different modes of spreading in isolation do not lead to successful epidemics. Such hybrid spreading strategies may be used beneficially to provide the most effective strategies for promulgating information across a large population. When used maliciously, however, they can present a dangerous challenge to current internet security protocols.

Hybrid Epidemics—A Case Study on Computer Worm Conficker

Science.gov (United States)

Zhang, Changwang; Zhou, Shi; Chain, Benjamin M.

2015-01-01

Conficker is a computer worm that erupted on the Internet in 2008. It is unique in combining three different spreading strategies: local probing, neighbourhood probing, and global probing. We propose a mathematical model that combines three modes of spreading: local, neighbourhood, and global, to capture the worm’s spreading behaviour. The parameters of the model are inferred directly from network data obtained during the first day of the Conficker epidemic. The model is then used to explore the tradeoff between spreading modes in determining the worm’s effectiveness. Our results show that the Conficker epidemic is an example of a critically hybrid epidemic, in which the different modes of spreading in isolation do not lead to successful epidemics. Such hybrid spreading strategies may be used beneficially to provide the most effective strategies for promulgating information across a large population. When used maliciously, however, they can present a dangerous challenge to current internet security protocols. PMID:25978309

Bioinspired near-infrared-excited sensing platform for in vitro antioxidant capacity assay based on upconversion nanoparticles and a dopamine-melanin hybrid system.

Science.gov (United States)

Wang, Dong; Chen, Chuan; Ke, Xuebin; Kang, Ning; Shen, Yuqing; Liu, Yongliang; Zhou, Xi; Wang, Hongjun; Chen, Changqing; Ren, Lei

2015-02-11

A novel core-shell structure based on upconversion fluorescent nanoparticles (UCNPs) and dopamine-melanin has been developed for evaluation of the antioxidant capacity of biological fluids. In this approach, dopamine-melanin nanoshells facilely formed on the surface of UCNPs act as ultraefficient quenchers for upconversion fluorescence, contributing to a photoinduced electron-transfer mechanism. This spontaneous oxidative polymerization of the dopamine-induced quenching effect could be effectively prevented by the presence of various antioxidants (typically biothiols, ascorbic acid (Vitamin C), and Trolox). The chemical response of the UCNPs@dopamine-melanin hybrid system exhibited great selectivity and sensitivity toward antioxidants relative to other compounds at 100-fold higher concentration. A satisfactory correlation was established between the ratio of the "anti-quenching" fluorescence intensity and the concentration of antioxidants. Besides the response of the upconversion fluorescence signal, a specific evaluation process for antioxidants could be visualized by the color change from colorless to dark gray accompanied by the spontaneous oxidation of dopamine. The near-infrared (NIR)-excited UCNP-based antioxidant capacity assay platform was further used to evaluate the antioxidant capacity of cell extracts and human plasma, and satisfactory sensitivity, repeatability, and recovery rate were obtained. This approach features easy preparation, fluorescence/visual dual mode detection, high specificity to antioxidants, and enhanced sensitivity with NIR excitation, showing great potential for screening and quantitative evaluation of antioxidants in biological systems.

Interatomic Coulombic electron capture

International Nuclear Information System (INIS)

Gokhberg, K.; Cederbaum, L. S.

2010-01-01

In a previous publication [K. Gokhberg and L. S. Cederbaum, J. Phys. B 42, 231001 (2009)] we presented the interatomic Coulombic electron capture process--an efficient electron capture mechanism by atoms and ions in the presence of an environment. In the present work we derive and discuss the mechanism in detail. We demonstrate thereby that this mechanism belongs to a family of interatomic electron capture processes driven by electron correlation. In these processes the excess energy released in the capture event is transferred to the environment and used to ionize (or to excite) it. This family includes the processes where the capture is into the lowest or into an excited unoccupied orbital of an atom or ion and proceeds in step with the ionization (or excitation) of the environment, as well as the process where an intermediate autoionizing excited resonance state is formed in the capturing center which subsequently deexcites to a stable state transferring its excess energy to the environment. Detailed derivation of the asymptotic cross sections of these processes is presented. The derived expressions make clear that the environment assisted capture processes can be important for many systems. Illustrative examples are presented for a number of model systems for which the data needed to construct the various capture cross sections are available in the literature.

Streptococcus pneumoniae Supragenome Hybridization Arrays for Profiling of Genetic Content and Gene Expression.

Science.gov (United States)

Kadam, Anagha; Janto, Benjamin; Eutsey, Rory; Earl, Joshua P; Powell, Evan; Dahlgren, Margaret E; Hu, Fen Z; Ehrlich, Garth D; Hiller, N Luisa

2015-02-02

There is extensive genomic diversity among Streptococcus pneumoniae isolates. Approximately half of the comprehensive set of genes in the species (the supragenome or pangenome) is present in all the isolates (core set), and the remaining is unevenly distributed among strains (distributed set). The Streptococcus pneumoniae Supragenome Hybridization (SpSGH) array provides coverage for an extensive set of genes and polymorphisms encountered within this species, capturing this genomic diversity. Further, the capture is quantitative. In this manner, the SpSGH array allows for both genomic and transcriptomic analyses of diverse S. pneumoniae isolates on a single platform. In this unit, we present the SpSGH array, and describe in detail its design and implementation for both genomic and transcriptomic analyses. The methodology can be applied to construction and modification of SpSGH array platforms, as well to other bacterial species as long as multiple whole-genome sequences are available that collectively capture the vast majority of the species supragenome. Copyright © 2015 John Wiley & Sons, Inc.

Highly efficient solid-state neutron scintillators based on hybrid sol-gel nanocomposite materials

International Nuclear Information System (INIS)

Kesanli, Banu; Hong, Kunlun; Meyer, Kent; Im, Hee-Jung; Dai, Sheng

2006-01-01

This research highlights opportunities in the formulation of neutron scintillators that not only have high scintillation efficiencies but also can be readily cast into two-dimensional detectors. Series of transparent, crack-free monoliths were prepared from hybrid polystyrene-silica nanocomposites in the presence of arene-containing alkoxide precursor through room temperature sol-gel processing. The monoliths also contain lithium-6 salicylate as a target material for neutron-capture reactions and amphiphilic scintillator solution as a fluorescent sensitizer. Polystyrene was functionalized by trimethoxysilyl group in order to enable the covalent incorporation of aromatic functional groups into the inorganic sol-gel matrices for minimizing macroscopic phase segregation and facilitating lithium-6 doping in the sol-gel samples. Neutron and alpha responses of these hybrid polystyrene-silica monoliths were explored

Rapid Diagnostic Assay for Intact Influenza Virus Using a High Affinity Hemagglutinin Binding Protein.

Science.gov (United States)

Anderson, Caitlin E; Holstein, Carly A; Strauch, Eva-Maria; Bennett, Steven; Chevalier, Aaron; Nelson, Jorgen; Fu, Elain; Baker, David; Yager, Paul

2017-06-20

Influenza is a ubiquitous and recurring infection that results in approximately 500 000 deaths globally each year. Commercially available rapid diagnostic tests are based upon detection of the influenza nucleoprotein, which are limited in that they are unable to differentiate by species and require an additional viral lysis step. Sample preprocessing can be minimized or eliminated by targeting the intact influenza virus, thereby reducing assay complexity and leveraging the large number of hemagglutinin proteins on the surface of each virus. Here, we report the development of a paper-based influenza assay that targets the hemagglutinin protein; the assay employs a combination of antibodies and novel computationally designed, recombinant affinity proteins as the capture and detection agents. This system leverages the customizability of recombinant protein design to target the conserved receptor-binding pocket of the hemagglutinin protein and to match the trimeric nature of hemagglutinin for improved avidity. Using this assay, we demonstrate the first instance of intact influenza virus detection using a combination of antibody and affinity proteins within a porous network. The recombinant head region binder based assays yield superior analytical sensitivity as compared to the antibody based assay, with lower limits of detection of 3.54 × 10 7 and 1.34 × 10 7 CEID 50 /mL for the mixed and all binder stacks, respectively. Not only does this work describe the development of a novel influenza assay, it also demonstrates the power of recombinant affinity proteins for use in rapid diagnostic assays.

Hybrid RANS/LES method for wind flow over complex terrain

DEFF Research Database (Denmark)

Bechmann, Andreas; Sørensen, Niels N.

2010-01-01

for flows at high Reynolds numbers. To reduce the computational cost of traditional LES, a hybrid method is proposed in which the near-wall eddies are modelled in a Reynolds-averaged sense. Close to walls, the flow is treated with the Reynolds-averaged Navier-Stokes (RANS) equations (unsteady RANS...... rough walls. Previous attempts of combining RANS and LES has resulted in unphysical transition regions between the two layers, but the present work improves this region by using a stochastic backscatter model. To demonstrate the ability of the proposed hybrid method, simulations are presented for wind...... the turbulent kinetic energy, whereas the new method captures the high turbulence levels well but underestimates the mean velocity. The presented results are for a relative mild configuration of complex terrain, but the proposed method can also be used for highly complex terrain where the benefits of the new...

Investigation of Parameters that Affect the Success Rate of Microarray-Based Allele-Specific Hybridization Assays

DEFF Research Database (Denmark)

Poulsen, Lena; Søe, Martin Jensen; Moller, Lisbeth Birk

2011-01-01

Background: The development of microarray-based genetic tests for diseases that are caused by known mutations is becoming increasingly important. The key obstacle to developing functional genotyping assays is that such mutations need to be genotyped regardless of their location in genomic regions...

Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

DEFF Research Database (Denmark)

Schmidt, Gunilla Veslemøy; Mellerup, Anders; Christiansen, Lasse Engbo

2015-01-01

The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays...... for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined...... in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes...

Novel RNA hybridization method for the in situ detection of ETV1, ETV4, and ETV5 gene fusions in prostate cancer.

Science.gov (United States)

Kunju, Lakshmi P; Carskadon, Shannon; Siddiqui, Javed; Tomlins, Scott A; Chinnaiyan, Arul M; Palanisamy, Nallasivam

2014-09-01

The genetic basis of 50% to 60% of prostate cancer (PCa) is attributable to rearrangements in E26 transformation-specific (ETS) (ERG, ETV1, ETV4, and ETV5), BRAF, and RAF1 genes and overexpression of SPINK1. The development and validation of reliable detection methods are warranted to classify various molecular subtypes of PCa for diagnostic and prognostic purposes. ETS gene rearrangements are typically detected by fluorescence in situ hybridization and reverse-transcription polymerase chain reaction methods. Recently, monoclonal antibodies against ERG have been developed that detect the truncated ERG protein in immunohistochemical assays where staining levels are strongly correlated with ERG rearrangement status by fluorescence in situ hybridization. However, specific antibodies for ETV1, ETV4, and ETV5 are unavailable, challenging their clinical use. We developed a novel RNA in situ hybridization-based assay for the in situ detection of ETV1, ETV4, and ETV5 in formalin-fixed paraffin-embedded tissues from prostate needle biopsies, prostatectomy, and metastatic PCa specimens using RNA probes. Further, with combined RNA in situ hybridization and immunohistochemistry we identified a rare subset of PCa with dual ETS gene rearrangements in collisions of independent tumor foci. The high specificity and sensitivity of RNA in situ hybridization provides an alternate method enabling bright-field in situ detection of ETS gene aberrations in routine clinically available PCa specimens.

Feasibility and accuracy evaluation of three human papillomavirus assays for FTA card-based sampling: a pilot study in cervical cancer screening.

Science.gov (United States)

Wang, Shao-Ming; Hu, Shang-Ying; Chen, Wen; Chen, Feng; Zhao, Fang-Hui; He, Wei; Ma, Xin-Ming; Zhang, Yu-Qing; Wang, Jian; Sivasubramaniam, Priya; Qiao, You-Lin

2015-11-04

Liquid-state specimen carriers are inadequate for sample transportation in large-scale screening projects in low-resource settings, which necessitates the exploration of novel non-hazardous solid-state alternatives. Studies investigating the feasibility and accuracy of a solid-state human papillomavirus (HPV) sampling medium in combination with different down-stream HPV DNA assays for cervical cancer screening are needed. We collected two cervical specimens from 396 women, aged 25-65 years, who were enrolled in a cervical cancer screening trial. One sample was stored using DCM preservative solution and the other was applied to a Whatman Indicating FTA Elute® card (FTA card). All specimens were processed using three HPV testing methods, including Hybrid capture 2 (HC2), careHPV™, and Cobas®4800 tests. All the women underwent a rigorous colposcopic evaluation that included using a microbiopsy protocol. Compared to the liquid-based carrier, the FTA card demonstrated comparable sensitivity for detecting high grade Cervical Intraepithelial Neoplasia (CIN) using HC2 (91.7 %), careHPV™ (83.3 %), and Cobas®4800 (91.7 %) tests. Moreover, the FTA card showed a higher specificity compared to a liquid-based carrier for HC2 (79.5 % vs. 71.6 %, P = 0.015), comparable specificity for careHPV™ (78.1 % vs. 73.0 %, P > 0.05), but lower specificity for the Cobas®4800 test (62.4 % vs. 69.9 %, P = 0.032). Generally, the FTA card-based sampling medium's accuracy was comparable with that of liquid-based medium for the three HPV testing assays. FTA cards are a promising sample carrier for cervical cancer screening. With further optimization, it can be utilized for HPV testing in areas of varying economic development.

Hybrid intelligent monironing systems for thermal power plant trips

Science.gov (United States)

Barsoum, Nader; Ismail, Firas Basim

2012-11-01

Steam boiler is one of the main equipment in thermal power plants. If the steam boiler trips it may lead to entire shutdown of the plant, which is economically burdensome. Early boiler trips monitoring is crucial to maintain normal and safe operational conditions. In the present work two artificial intelligent monitoring systems specialized in boiler trips have been proposed and coded within the MATLAB environment. The training and validation of the two systems has been performed using real operational data captured from the plant control system of selected power plant. An integrated plant data preparation framework for seven boiler trips with related operational variables has been proposed for IMSs data analysis. The first IMS represents the use of pure Artificial Neural Network system for boiler trip detection. All seven boiler trips under consideration have been detected by IMSs before or at the same time of the plant control system. The second IMS represents the use of Genetic Algorithms and Artificial Neural Networks as a hybrid intelligent system. A slightly lower root mean square error was observed in the second system which reveals that the hybrid intelligent system performed better than the pure neural network system. Also, the optimal selection of the most influencing variables performed successfully by the hybrid intelligent system.

Evaluation of a competitive enzyme-linked immunosorbent assay for measurements of soluble HLA-G protein.

Science.gov (United States)

Rasmussen, M; Dahl, M; Buus, S; Djurisic, S; Ohlsson, J; Hviid, T V F

2014-08-01

The human leukocyte antigen (HLA) class Ib molecule, HLA-G, has gained increased attention because of its assumed important role in immune regulation. The HLA-G protein exists in several soluble isoforms. Most important are the actively secreted HLA-G5 full-length isoform generated by alternative splicing retaining intron 4 with a premature stop codon, and the cleavage of full-length membrane-bound HLA-G1 from the cell surface, so-called soluble HLA-G1 (sHLA-G1). A specific and sensitive immunoassay for measurements of soluble HLA-G is mandatory for conceivable routine testing and research projects. We report a novel method, a competitive immunoassay, for measuring HLA-G5/sHLA-G1 in biological fluids. The sHLA-G immunoassay is based upon a competitive enzyme-linked immunosorbent assay (ELISA) principle. It includes a recombinant sHLA-G1 protein in complex with β2-microglobulin and a peptide as a standard, biotinylated recombinant sHLA-G1 as an indicator, and the MEM-G/9 anti-HLA-G monoclonal antibody (mAb) as the capture antibody. The specificity and sensitivity of the assay were evaluated. Testing with different recombinant HLA class I proteins and different anti-HLA class I mAbs showed that the sHLA-G immunoassay was highly specific. Optimal combinations of competitor sHLA-G1 and capture mAb concentrations were determined. Two versions of the assay were tested. One with a relatively wide dynamic range from 3.1 to 100.0 ng/ml, and another more sensitive version ranging from 1.6 to 12.5 ng/ml. An intra-assay coefficient of variation (CV) of 15.5% at 88 ng/ml and an inter-assay CV of 23.1% at 39 ng/ml were determined. An assay based on the competitive sHLA-G ELISA may be important for measurements of sHLA-G proteins in several conditions: assisted reproduction, organ transplantation, cancer, and certain pregnancy complications, both in research studies and possibly in the future also for clinical routine use. © 2014 John Wiley & Sons A/S. Published by John Wiley

Electrochemical immunosensor assay (EIA) for sensitive detection of E. coli O157:H7 with signal amplification on a SG-PEDOT-AuNPs electrode interface.

Science.gov (United States)

Guo, Yuna; Wang, Yu; Liu, Su; Yu, Jinghua; Wang, Hongzhi; Cui, Min; Huang, Jiadong

2015-01-21

A novel electrochemical immunosensor assay (EIA) for highly sensitive and specific detection of Escherichia coli O157:H7 has been developed. This immunosensor is constructed by the assembly of capture antibody on SG-PEDOT-AuNPs composites modified glass carbon electrode. In the presence of target E. coli O157:H7, horseradish peroxidase (HRP)-labeled antibody is captured on the electrode surface to form a sandwich-type system via the specific identification. As a result, E. coli O157:H7 detection is realized by outputting a redox current from electro-reduction of hydrogen peroxide reaction catalyzed by HRP. In our assay, the combination of the unique properties of sulfonated graphene (SG) and gold nanoparticles (AuNPs) can not only accelerate electron transfer on electrode interface, but also provide an excellent scaffold for the conjugation of capture antibody that significantly improves the target capture efficiency and enhances the sensitivity of the biosensor. The results reveal the calibration plot obtained for E. coli O157:H7 is approximately linear from 7.8 × 10-7.8 × 10(6) colony-forming unit (cfu) mL(-1) with the limit of detection of 3.4 × 10 cfu mL(-1). In addition, the biosensor has been successfully applied to the quantitative assay of E. coli O157:H7 in synthetic samples (spring water and milk). Hence, the developed electrochemical-based immunosensor might provide a useful and practical tool for E. coli O157:H7 determination and related food safety analysis and clinical diagnosis.

Dynamical analysis of Parkinsonian state emulated by hybrid Izhikevich neuron models

Science.gov (United States)

Liu, Chen; Wang, Jiang; Yu, Haitao; Deng, Bin; Wei, Xile; Li, Huiyan; Loparo, Kenneth A.; Fietkiewicz, Chris

2015-11-01

Computational models play a significant role in exploring novel theories to complement the findings of physiological experiments. Various computational models have been developed to reveal the mechanisms underlying brain functions. Particularly, in the development of therapies to modulate behavioral and pathological abnormalities, computational models provide the basic foundations to exhibit transitions between physiological and pathological conditions. Considering the significant roles of the intrinsic properties of the globus pallidus and the coupling connections between neurons in determining the firing patterns and the dynamical activities of the basal ganglia neuronal network, we propose a hypothesis that pathological behaviors under the Parkinsonian state may originate from combined effects of intrinsic properties of globus pallidus neurons and synaptic conductances in the whole neuronal network. In order to establish a computational efficient network model, hybrid Izhikevich neuron model is used due to its capacity of capturing the dynamical characteristics of the biological neuronal activities. Detailed analysis of the individual Izhikevich neuron model can assist in understanding the roles of model parameters, which then facilitates the establishment of the basal ganglia-thalamic network model, and contributes to a further exploration of the underlying mechanisms of the Parkinsonian state. Simulation results show that the hybrid Izhikevich neuron model is capable of capturing many of the dynamical properties of the basal ganglia-thalamic neuronal network, such as variations of the firing rates and emergence of synchronous oscillations under the Parkinsonian condition, despite the simplicity of the two-dimensional neuronal model. It may suggest that the computational efficient hybrid Izhikevich neuron model can be used to explore basal ganglia normal and abnormal functions. Especially it provides an efficient way of emulating the large-scale neuron network

Carbon captured from the air

Energy Technology Data Exchange (ETDEWEB)

Keith, D. [Calgary Univ., AB (Canada)

2008-10-15

This article presented an innovative way to achieve the efficient capture of atmospheric carbon. A team of scientists from the University of Calgary's Institute for Sustainable Energy, Environment and Economy have shown that it is possible to reduce carbon dioxide (CO{sub 2}) using a simple machine that can capture the trace amount of CO{sub 2} present in ambient air at any place on the planet. The thermodynamics of capturing the small concentrations of CO{sub 2} from the air is only slightly more difficult than capturing much larger concentrations of CO{sub 2} from power plants. The research is significant because it offers a way to capture CO{sub 2} emissions from transportation sources such as vehicles and airplanes, which represent more than half of the greenhouse gases emitted on Earth. The energy efficient and cost effective air capture technology could complement other approaches for reducing emissions from the transportation sector, such as biofuels and electric vehicles. Air capture differs from carbon capture and storage (CCS) technology used at coal-fired power plants where CO{sub 2} is captured and pipelined for permanent storage underground. Air capture can capture the CO{sub 2} that is present in ambient air and store it wherever it is cheapest. The team at the University of Calgary showed that CO{sub 2} could be captured directly from the air with less than 100 kWhrs of electricity per tonne of CO{sub 2}. A custom-built tower was able to capture the equivalent of 20 tonnes per year of CO{sub 2} on a single square meter of scrubbing material. The team devised a way to use a chemical process from the pulp and paper industry to cut the energy cost of air capture in half. Although the technology is only in its early stage, it appears that CO{sub 2} could be captured from the air with an energy demand comparable to that needed for CO{sub 2} capture from conventional power plants, but costs will be higher. The simple, reliable and scalable technology

Hybrid Propulsion Demonstration Program 250K Hybrid Motor

Science.gov (United States)

Story, George; Zoladz, Tom; Arves, Joe; Kearney, Darren; Abel, Terry; Park, O.

2003-01-01

The Hybrid Propulsion Demonstration Program (HPDP) program was formed to mature hybrid propulsion technology to a readiness level sufficient to enable commercialization for various space launch applications. The goal of the HPDP was to develop and test a 250,000 pound vacuum thrust hybrid booster in order to demonstrate hybrid propulsion technology and enable manufacturing of large hybrid boosters for current and future space launch vehicles. The HPDP has successfully conducted four tests of the 250,000 pound thrust hybrid rocket motor at NASA's Stennis Space Center. This paper documents the test series.

Novel nuclear localization and potential function of insulin-like growth factor-1 receptor/insulin receptor hybrid in corneal epithelial cells.

Directory of Open Access Journals (Sweden)

Yu-Chieh Wu

Full Text Available BACKGROUND: Type I insulin-like growth factor receptor (IGF-1R and insulin receptor (INSR are highly homologous molecules, which can heterodimerize to form an IGF-1R/INSR hybrid (Hybrid-R. The presence and biological significance of the Hybrid-R in human corneal epithelium has not yet been established. In addition, while nuclear localization of IGF-1R was recently reported in cancer cells and human corneal epithelial cells, the function and profile of nuclear IGF-1R is unknown. In this study, we characterized the nuclear localization and function of the Hybrid-R and the role of IGF-1/IGF-1R and Hybrid-R signaling in the human corneal epithelium. METHODOLOGY/PRINCIPLE FINDINGS: IGF-1-mediated signaling and cell growth were examined in a human telomerized corneal epithelial (hTCEpi cell line using co-immunoprecipitation, immunoblotting and cell proliferation assays. The presence of Hybrid-R in hTCEpi and primary cultured human corneal epithelial cells was confirmed by immunofluorescence and reciprocal immunoprecipitation of whole cell lysates. We found that IGF-1 stimulated Akt and promoted cell growth through IGF-1R activation, which was independent of the Hybrid-R. The presence of Hybrid-R, but not IGF-1R/IGF-1R, was detected in nuclear extracts. Knockdown of INSR by small interfering RNA resulted in depletion of the INSR/INSR and preferential formation of Hybrid-R. Chromatin-immunoprecipitation sequencing assay with anti-IGF-1R or anti-INSR was subsequently performed to identify potential genomic targets responsible for critical homeostatic regulatory pathways. CONCLUSION/SIGNIFICANCE: In contrast to previous reports on nuclear localized IGF-1R, this is the first report identifying the nuclear localization of Hybrid-R in an epithelial cell line. The identification of a nuclear Hybrid-R and novel genomic targets suggests that IGF-1R traffics to the nucleus as an IGF-1R/INSR heterotetrameric complex to regulate corneal epithelial homeostatic

A signal amplification assay for HSV type 1 viral DNA detection using nanoparticles and direct acoustic profiling

Directory of Open Access Journals (Sweden)

Hammond Richard

2010-02-01

Full Text Available Abstract Background Nucleic acid based recognition of viral sequences can be used together with label-free biosensors to provide rapid, accurate confirmation of viral infection. To enhance detection sensitivity, gold nanoparticles can be employed with mass-sensitive acoustic biosensors (such as a quartz crystal microbalance by either hybridising nanoparticle-oligonucleotide conjugates to complimentary surface-immobilised ssDNA probes on the sensor, or by using biotin-tagged target oligonucleotides bound to avidin-modified nanoparticles on the sensor. We have evaluated and refined these signal amplification assays for the detection from specific DNA sequences of Herpes Simplex Virus (HSV type 1 and defined detection limits with a 16.5 MHz fundamental frequency thickness shear mode acoustic biosensor. Results In the study the performance of semi-homogeneous and homogeneous assay formats (suited to rapid, single step tests were evaluated utilising different diameter gold nanoparticles at varying DNA concentrations. Mathematical models were built to understand the effects of mass transport in the flow cell, the binding kinetics of targets to nanoparticles in solution, the packing geometries of targets on the nanoparticle, the packing of nanoparticles on the sensor surface and the effect of surface shear stiffness on the response of the acoustic sensor. This lead to the selection of optimised 15 nm nanoparticles that could be used with a 6 minute total assay time to achieve a limit of detection sensitivity of 5.2 × 10-12 M. Larger diameter nanoparticles gave poorer limits of detection than smaller particles. The limit of detection was three orders of magnitude lower than that observed using a hybridisation assay without nanoparticle signal amplification. Conclusions An analytical model was developed to determine optimal nanoparticle diameter, concentration and probe density, which allowed efficient and rapid optimisation of assay parameters

Enhancing Hybrid Perovskite Detectability in the Deep Ultraviolet Region with Down-Conversion Dual-Phase (CsPbBr3-Cs4PbBr6) Films.

Science.gov (United States)

Tong, Guoqing; Li, Huan; Zhu, Zhifeng; Zhang, Yan; Yu, Linwei; Xu, Jun; Jiang, Yang

2018-04-05

Hybrid perovskite photodetectors (PDs) exhibit outstanding performance in the ultraviolet-visible (UV-vis) spectrum but have poor detectability in the deep ultraviolet (DUV) region (200-350 nm). In this work, a novel inorganic-hybrid architecture that incorporates a dual-phase (CsPbBr 3 -Cs 4 PbBr 6 ) inorganic perovskite material as a down-conversion window layer and a hybrid perovskite as a light capture layer was prepared to achieve faster, highly sensitive photodetection in the DUV spectrum. A dual-phase inorganic perovskite film coated on the back surface of the photodetector enables strong light absorption and tunes the incident energy into emission bands that are optimized for the perovskite photodetector. The presence of Cs 4 PbBr 6 enhances the capture and down-conversion of the incident DUV light. Due to the down-conversion and transport of the DUV photons, a self-driven perovskite photodetector with this composite structure exhibits a fast response time of 7.8/33.6 μs and a high responsivity of 49.4 mA W -1 at 254 nm without extra power supply.

Hybrid Large-Eddy/Reynolds-Averaged Simulation of a Supersonic Cavity Using VULCAN

Science.gov (United States)

Quinlan, Jesse; McDaniel, James; Baurle, Robert A.

2013-01-01

Simulations of a supersonic recessed-cavity flow are performed using a hybrid large-eddy/Reynolds-averaged simulation approach utilizing an inflow turbulence recycling procedure and hybridized inviscid flux scheme. Calorically perfect air enters a three-dimensional domain at a free stream Mach number of 2.92. Simulations are performed to assess grid sensitivity of the solution, efficacy of the turbulence recycling, and the effect of the shock sensor used with the hybridized inviscid flux scheme. Analysis of the turbulent boundary layer upstream of the rearward-facing step for each case indicates excellent agreement with theoretical predictions. Mean velocity and pressure results are compared to Reynolds-averaged simulations and experimental data for each case and indicate good agreement on the finest grid. Simulations are repeated on a coarsened grid, and results indicate strong grid density sensitivity. Simulations are performed with and without inflow turbulence recycling on the coarse grid to isolate the effect of the recycling procedure, which is demonstrably critical to capturing the relevant shear layer dynamics. Shock sensor formulations of Ducros and Larsson are found to predict mean flow statistics equally well.

Modelling and experimentation on air hybrid engine concepts for automotive applications

OpenAIRE

Psanis, Christodoulos

2007-01-01

This thesis was submitted for the degree of Doctor of Philosophy and awarded by Brunel University. Hybrid powertrains that use compressed air to help power a vehicle could dramatically improve the fuel economy, particularly in cities and urban areas where the traffic conditions involve a lot of starts and stops. In such conditions, a large amount of fuel is needed to accelerate the vehicle, and much of this is converted to heat in brake friction during decelerations. Capturing, storing and...

A hybrid approach to the surface biofunctionalization of nanostructured porous alumina

Energy Technology Data Exchange (ETDEWEB)

Silvan, Miguel Manso; Ruiz, Josefa Predestinacion Garcia [Departamento de Fisica Aplicada y Departamento de Biologia Molecular, Facultad de Ciencias, Universidad Autonoma de Madrid, Unidad Asociada GMNF (ICMM-CSIC), 28049 Madrid (Spain); Centro de Investigaciones Biomedicas en Red, Bioingenieria Biomateriales y Nanomedicina (CIBERbbn) (Spain); Gonzalez, Ruy Sanz [Instituto de Ciencia de Materiales de Madrid, Consejo Superior de Investigaciones Cientificas, 28049 Madrid (Spain); Velez, Manuel Hernandez [Departamento de Fisica Aplicada y Departamento de Biologia Molecular, Facultad de Ciencias, Universidad Autonoma de Madrid, Unidad Asociada GMNF (ICMM-CSIC), 28049 Madrid (Spain)

2010-02-15

The application of nanostructured porous alumina templates as a solid support in biomedical assays requires a surface biofunctionalization process that has been addressed in this work by an hybrid aminopropyl-triethoxysilane/tetraisopropyl-orthotitanate (APTS/ TIPT) self assembled film. The nanostructured porous alumina templates are activated in a peroxide solution before immersion in the biofunctionalizing APTS/TIPT solution. The biofunctionalization process was followed up by UV-vis spectroscopy, which confirmed the modification of the dielectric structure of the alumina surface. The influence of the biofunctionalization step in an immunological assay was carried out by fluorescence microscopy. Results confirm the gain in activity after the immobilization of an FITC labelled mouse Igg. Specific biological recognition in a bovine serum albumin (BSA)-antiBSA assay is proved afterwards by shifts observed in the reflectance interferograms thus providing a fast biosensing transducer platform. (copyright 2010 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

A CO-FISH assay to assess sister chromatid segregation patterns in mitosis of mouse embryonic stem cells.

Science.gov (United States)

Sauer, Stephan; Burkett, Sandra S; Lewandoski, Mark; Klar, Amar J S

2013-05-01

Sister chromatids contain identical DNA sequence but are chiral with respect to both their helical handedness and their replication history. Emerging evidence from various model organisms suggests that certain stem cells segregate sister chromatids nonrandomly to either maintain genome integrity or to bias cellular differentiation in asymmetric cell divisions. Conventional methods for tracing of old vs. newly synthesized DNA strands generally lack resolution for individual chromosomes and employ halogenated thymidine analogs with profound cytotoxic effects on rapidly dividing cells. Here, we present a modified chromosome orientation fluorescence in situ hybridization (CO-FISH) assay, where identification of individual chromosomes and their replication history is achieved in subsequent hybridization steps with chromosome-specific DNA probes and PNA telomere probes. Importantly, we tackle the issue of BrdU cytotoxicity and show that our method is compatible with normal mouse ES cell biology, unlike a recently published related protocol. Results from our CO-FISH assay show that mitotic segregation of mouse chromosome 7 is random in ES cells, which contrasts previously published results from our laboratory and settles a controversy. Our straightforward protocol represents a useful resource for future studies on chromatid segregation patterns of in vitro-cultured cells from distinct model organisms.

Determination of nitrosourea compounds in brain tissue by gas chromatography and electron capture detection.

Science.gov (United States)

Hassenbusch, S J; Colvin, O M; Anderson, J H

1995-07-01

A relatively simple, high-sensitivity gas chromatographic assay is described for nitrosourea compounds, such as BCNU [1,3-bis(2-chloroethyl)-1-nitrosourea] and MeCCNU [1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea], in small biopsy samples of brain and other tissues. After extraction with ethyl acetate, secondary amines in BCNU and MeCCNU are derivatized with trifluoroacetic anhydride. Compounds are separated and quantitated by gas chromatography using a capillary column with temperature programming and an electron capture detector. Standard curves of BCNU indicate a coefficient of variance of 0.066 +/- 0.018, a correlation coefficient of 0.929, and an extraction efficiency from whole brain of 68% with a minimum detectable amount of 20 ng in 5-10 mg samples. The assay has been facile and sensitive in over 1000 brain biopsy specimens after intravenous and intraarterial infusions of BCNU.

Simultaneous capture and sequential detection of two malarial biomarkers on magnetic microparticles.

Science.gov (United States)

Markwalter, Christine F; Ricks, Keersten M; Bitting, Anna L; Mudenda, Lwiindi; Wright, David W

2016-12-01

We have developed a rapid magnetic microparticle-based detection strategy for malarial biomarkers Plasmodium lactate dehydrogenase (pLDH) and Plasmodium falciparum histidine-rich protein II (PfHRPII). In this assay, magnetic particles functionalized with antibodies specific for pLDH and PfHRPII as well as detection antibodies with distinct enzymes for each biomarker are added to parasitized lysed blood samples. Sandwich complexes for pLDH and PfHRPII form on the surface of the magnetic beads, which are washed and sequentially re-suspended in detection enzyme substrate for each antigen. The developed simultaneous capture and sequential detection (SCSD) assay detects both biomarkers in samples as low as 2.0parasites/µl, an order of magnitude below commercially available ELISA kits, has a total incubation time of 35min, and was found to be reproducible between users over time. This assay provides a simple and efficient alternative to traditional 96-well plate ELISAs, which take 5-8h to complete and are limited to one analyte. Further, the modularity of the magnetic bead-based SCSD ELISA format could serve as a platform for application to other diseases for which multi-biomarker detection is advantageous. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Detection of Antibodies to U.S. Isolates of Avian Pneumovirus by a Recombinant Nucleocapsid Protein-Based Sandwich Enzyme-Linked Immunosorbent Assay

OpenAIRE

Gulati, Baldev R.; Munir, Shirin; Patnayak, Devi P.; Goyal, Sagar M.; Kapur, Vivek

2001-01-01

The nucleocapsid (N) protein of subgroup C (United States-specific) avian pneumovirus (APV/US) was expressed in Escherichia coli, and antibodies to the recombinant N protein were shown to specifically recognize the ≈47-kDa N protein of APV/US by Western immunoblot analysis. The recombinant APV/US N protein was used in a sandwich-capture enzyme-linked immunosorbent assay (ELISA), and the resulting assay was found to be more sensitive and specific than the routine indirect ELISA for the detecti...

ASTRO Research Fellow Presentation - A comparison of the comet assay and pulsed-field gel electrophoresis as a predictive assay for radiosensitivity in human fibroblasts

International Nuclear Information System (INIS)

Sarkaria, Jann N.; Eady, John J.; Peacock, John H.; Steel, G. Gordon

1996-01-01

Purpose/Objective: To determine whether neutral lysis single-cell gel electrophoresis (comet assay) or pulsed-field gel electrophoresis (PFGE) can be used as a predictive assay for tissue response to radiotherapy as an alternative to clonogenic survival measurements. Materials and Methods: The comet assay has been widely used to measure DNA double strand breaks (dsb) in individual cells, and it has been suggested that it could be used as an alternative to clonogenic assays to measure radiosensitivity. Previous studies in this lab have demonstrated the ability of pulsed-field gel electrophoresis, which also measures DNA dsb, to accurately predict the radiosensitivity of a panel of fibroblasts based on determination of residual DNA dsb. As part of an ongoing study examining the relationship between fibroblast radiosensitivity and normal-tissue radiation reactions, we have compared the sensitivity and accuracy of the comet assay and PFGE on a different panel of non-transformed fibroblasts derived from breast cancer patients who developed severe radiation late effects and from case-matched controls. For the measurement of initial damage, cells were suspended in PBS and irradiated on ice for the comet assay and irradiated in agarose plugs on ice for pFGE. Residual damage was measured following irradiation of confluent cultures at 37 degree sign C and subsequent incubation for four hours prior to preparation of agarose slides and plugs. All irradiations were performed with a 59 TBq 60 Co source at a dose rate of 1.7 Gy/min. Electrophoresis was performed following neutral pH cell lysis. Comet images were captured and analyzed using Optimas software with DNA damage quantitated by the comet moment. PFGE gels were analyzed using a phosphor-image analysis system and damage was quantitated based on the percent of activity released from the well. Results: The comet assay was able to detect initial DNA damage at a threshold of 5 Gy and exhibited a linear dose

A real time chemotaxis assay unveils unique migratory profiles amongst different primary murine macrophages.

Directory of Open Access Journals (Sweden)

Asif J Iqbal

Full Text Available Chemotaxis assays are an invaluable tool for studying the biological activity of inflammatory mediators such as CC chemokines, which have been implicated in a wide range of chronic inflammatory diseases. Conventional chemotaxis systems such as the modified Boyden chamber are limited in terms of the data captured given that the assays are analysed at a single time-point. We report the optimisation and validation of a label-free, real-time cell migration assay based on electrical cell impedance to measure chemotaxis of different primary murine macrophage populations in response to a range of CC chemokines and other chemoattractant signalling molecules. We clearly demonstrate key differences in the migratory behavior of different murine macrophage populations and show that this dynamic system measures true macrophage chemotaxis rather than chemokinesis or fugetaxis. We highlight an absolute requirement for Gαi signaling and actin cytoskeletal rearrangement as demonstrated by Pertussis toxin and cytochalasin D inhibition. We also studied the chemotaxis of CD14(+ human monocytes and demonstrate distinct chemotactic profiles amongst different monocyte donors to CCL2. This real-time chemotaxis assay will allow a detailed analysis of factors that regulate macrophage responses to chemoattractant cytokines and inflammatory mediators.

A Real Time Chemotaxis Assay Unveils Unique Migratory Profiles amongst Different Primary Murine Macrophages

Science.gov (United States)

Iqbal, Asif J.; Regan-Komito, Daniel; Christou, Ivy; White, Gemma E.; McNeill, Eileen; Kenyon, Amy; Taylor, Lewis; Kapellos, Theodore S.; Fisher, Edward A.; Channon, Keith M.; Greaves, David R.

2013-01-01

Chemotaxis assays are an invaluable tool for studying the biological activity of inflammatory mediators such as CC chemokines, which have been implicated in a wide range of chronic inflammatory diseases. Conventional chemotaxis systems such as the modified Boyden chamber are limited in terms of the data captured given that the assays are analysed at a single time-point. We report the optimisation and validation of a label-free, real-time cell migration assay based on electrical cell impedance to measure chemotaxis of different primary murine macrophage populations in response to a range of CC chemokines and other chemoattractant signalling molecules. We clearly demonstrate key differences in the migratory behavior of different murine macrophage populations and show that this dynamic system measures true macrophage chemotaxis rather than chemokinesis or fugetaxis. We highlight an absolute requirement for Gαi signaling and actin cytoskeletal rearrangement as demonstrated by Pertussis toxin and cytochalasin D inhibition. We also studied the chemotaxis of CD14+ human monocytes and demonstrate distinct chemotactic profiles amongst different monocyte donors to CCL2. This real-time chemotaxis assay will allow a detailed analysis of factors that regulate macrophage responses to chemoattractant cytokines and inflammatory mediators. PMID:23516549

Attenuation correction for the collimated gamma ray assay of cylindrical samples

International Nuclear Information System (INIS)

Patra, Sabyasachi; Agarwal, Chhavi; Goswami, A.; Gathibandhe, M.

2015-01-01

The Hybrid Monte Carlo (HMC) method developed earlier for attenuation correction of non-collimated samples [Agarwal et al., 2008, Nucl. Instrum. Methods A 597, 198], has been extended to the segmented gamma ray assay of cylindrical samples. The method has been validated both experimentally and theoretically. For experimental validation, the results of HMC calculation have been compared with the experimentally obtained attenuation correction factors. The HMC attenuation correction factors have also been compared with the results obtained from literature available near-field and far-field formulae at two sample-to-detector distances (10.3 cm and 20.4 cm). The method has been found to be valid at all sample-to-detector distances over a wide range of transmittance. On the other hand, the literature available near-field and far-field formulae have been found to work over a limited range of sample-to detector distances and transmittances. The HMC method has been further extended to circular collimated geometries where analytical formula for attenuation correction does not exist. - Highlights: • Hybrid Monte Carlo method for attenuation correction developed for SGA system. • Method found to work for all sample-detector geometries for all transmittances. • The near-field formula applicable only after certain sample-detector distance. • The far-field formula applicable only for higher transmittances (>18%). • Hybrid Monte Carlo method further extended to circular collimated geometry

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

Science.gov (United States)

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

2014-08-08

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

Quantitative analyses reveal distinct sensitivities of the capture of HIV-1 primary viruses and pseudoviruses to broadly neutralizing antibodies.

Science.gov (United States)

Kim, Jiae; Jobe, Ousman; Peachman, Kristina K; Michael, Nelson L; Robb, Merlin L; Rao, Mangala; Rao, Venigalla B

2017-08-01

Development of vaccines capable of eliciting broadly neutralizing antibodies (bNAbs) is a key goal to controlling the global AIDS epidemic. To be effective, bNAbs must block the capture of HIV-1 to prevent viral acquisition and establishment of reservoirs. However, the role of bNAbs, particularly during initial exposure of primary viruses to host cells, has not been fully examined. Using a sensitive, quantitative, and high-throughput qRT-PCR assay, we found that primary viruses were captured by host cells and converted into a trypsin-resistant form in less than five minutes. We discovered, unexpectedly, that bNAbs did not block primary virus capture, although they inhibited the capture of pseudoviruses/IMCs and production of progeny viruses at 48h. Further, viruses escaped bNAb inhibition unless the bNAbs were present in the initial minutes of exposure of virus to host cells. These findings will have important implications for HIV-1 vaccine design and determination of vaccine efficacy. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

The KRAS Strip Assay for detection of KRAS mutation in Egyptian patients with colorectal cancer (CRC): A pilot study

International Nuclear Information System (INIS)

Abd El Kader, Y.; Safwat, E.; Kassem, H.A.; Kassem, N.M.; Emera, G.

2013-01-01

Background: Epidermal growth factor receptor (EGFR) and its downstream factors KRAS and BRAF are mutated in several types of cancer, affecting the clinical response to EGFR inhibitors. Mutations in the EGFR kinase domain predict sensitivity to the tyrosine kinase inhibitors gefltinib and erlotinib in lung adenocarcinoma, while activating point mutations in KRAS and BRAF confer resistance to the anti-EGFR monoclonal antibody cetuximab in colorectal cancer. The development of new generation methods for systematic mutation screening of these genes will allow more appropriate therapeutic choices. Purpose: Detection of KRAS mutation in Egyptian colorectal cancer (CRC) patients by the KRAS Strip Assay. Methods: Examination of 20 colorectal cancer (CRC) patients is done to detect KRAS mutations by KRAS Strip Assay. For the Strip Assay, a mutant-enriched PCR was followed by hybridization to KRAS-specific probes bound to a nitrocellulose strip. Results: Among 20 patients, KRAS mutations were identified in 80% of patients by the KRAS Strip Assay. Conclusions: Our preliminary results suggest that KRAS Strip Assay is an alternative to protocols currently in use for KRAS mutation detection

Hybrid Analytical and Data-Driven Modeling for Feed-Forward Robot Control.

Science.gov (United States)

Reinhart, René Felix; Shareef, Zeeshan; Steil, Jochen Jakob

2017-02-08

Feed-forward model-based control relies on models of the controlled plant, e.g., in robotics on accurate knowledge of manipulator kinematics or dynamics. However, mechanical and analytical models do not capture all aspects of a plant's intrinsic properties and there remain unmodeled dynamics due to varying parameters, unmodeled friction or soft materials. In this context, machine learning is an alternative suitable technique to extract non-linear plant models from data. However, fully data-based models suffer from inaccuracies as well and are inefficient if they include learning of well known analytical models. This paper thus argues that feed-forward control based on hybrid models comprising an analytical model and a learned error model can significantly improve modeling accuracy. Hybrid modeling here serves the purpose to combine the best of the two modeling worlds. The hybrid modeling methodology is described and the approach is demonstrated for two typical problems in robotics, i.e., inverse kinematics control and computed torque control. The former is performed for a redundant soft robot and the latter for a rigid industrial robot with redundant degrees of freedom, where a complete analytical model is not available for any of the platforms.

Effects of a hybrid micro/nanorod topography-modified titanium implant on adhesion and osteogenic differentiation in rat bone marrow mesenchymal stem cells.

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Zhang, Wenjie; Li, Zihui; Huang, Qingfeng; Xu, Ling; Li, Jinhua; Jin, Yuqin; Wang, Guifang; Liu, Xuanyong; Jiang, Xinquan

2013-01-01

Various methods have been used to modify titanium implant surfaces with the aim of achieving better osseointegration. In this study, we fabricated a clustered nanorod structure on an acid-etched, microstructured titanium plate surface using hydrogen peroxide. We also evaluated biofunctionalization of the hybrid micro/nanorod topography on rat bone marrow mesenchymal stem cells. Scanning electron microscopy and x-ray diffraction were used to investigate the surface topography and phase composition of the modified titanium plate. Rat bone marrow mesenchymal stem cells were cultured and seeded on the plate. The adhesion ability of the cells was then assayed by cell counting at one, 4, and 24 hours after cell seeding, and expression of adhesion-related protein integrin β1 was detected by immunofluorescence. In addition, a polymerase chain reaction assay, alkaline phosphatase and Alizarin Red S staining assays, and osteopontin and osteocalcin immunofluorescence analyses were used to evaluate the osteogenic differentiation behavior of the cells. The hybrid micro/nanoscale texture formed on the titanium surface enhanced the initial adhesion activity of the rat bone marrow mesenchymal stem cells. Importantly, the hierarchical structure promoted osteogenic differentiation of these cells. This study suggests that a hybrid micro/nanorod topography on a titanium surface fabricated by treatment with hydrogen peroxide followed by acid etching might facilitate osseointegration of a titanium implant in vivo.

A Dual Electrochemical Sensor Based on a Test-strip Assay for the Quantitative Determination of Albumin and Creatinine.

Science.gov (United States)

Yasukawa, Tomoyuki; Kiba, Yuya; Mizutani, Fumio

2015-01-01

A dual-electrochemical sensor based on a test-strip assay with immunochemistry and enzyme reactions has been developed for the determination of albumin and creatinine. Each nitrocellulose membrane with an immobilization area of an anti-albumin antibody or three enzymes was prepared in the device with three working electrodes for measuring albumin, creatinine, and ascorbic acid, as well as an Ag/AgCl electrode used as a counter/pseudo-reference electrode. The reactions of three enzymes were initiated by flowing a solution containing creatinine to detect an oxidation current of hydrogen peroxide. A sandwich-type immunocomplex was formed by albumin and antibody labeled with glucose oxidase (GOx). Captured GOx catalyzed the reduction of Fe(CN)6(3-) to Fe(CN)6(4-), which was oxidized electrochemically to determine the captured albumin. The responses for creatinine and albumin increased with the concentrations in millimolar order and over the range 18.75 - 150 μg mL(-1), respectively. The present sensor would be a distinct demonstration for producing quantitative dual-assays for various biomolecules used for clinical diagnoses.

Automation of the ELISpot assay for high-throughput detection of antigen-specific T-cell responses.

Science.gov (United States)

Almeida, Coral-Ann M; Roberts, Steven G; Laird, Rebecca; McKinnon, Elizabeth; Ahmed, Imran; Pfafferott, Katja; Turley, Joanne; Keane, Niamh M; Lucas, Andrew; Rushton, Ben; Chopra, Abha; Mallal, Simon; John, Mina

2009-05-15

The enzyme linked immunospot (ELISpot) assay is a fundamental tool in cellular immunology, providing both quantitative and qualitative information on cellular cytokine responses to defined antigens. It enables the comprehensive screening of patient derived peripheral blood mononuclear cells to reveal the antigenic restriction of T-cell responses and is an emerging technique in clinical laboratory investigation of certain infectious diseases. As with all cellular-based assays, the final results of the assay are dependent on a number of technical variables that may impact precision if not highly standardised between operators. When studies that are large scale or using multiple antigens are set up manually, these assays may be labour intensive, have many manual handling steps, are subject to data and sample integrity failure and may show large inter-operator variability. Here we describe the successful automated performance of the interferon (IFN)-gamma ELISpot assay from cell counting through to electronic capture of cytokine quantitation and present the results of a comparison between automated and manual performance of the ELISpot assay. The mean number of spot forming units enumerated by both methods for limiting dilutions of CMV, EBV and influenza (CEF)-derived peptides in six healthy individuals were highly correlated (r>0.83, pautomated system compared favourably with the manual ELISpot and further ensured electronic tracking, increased through-put and reduced turnaround time.

Enzymatic production of single-molecule FISH and RNA capture probes.

Science.gov (United States)

Gaspar, Imre; Wippich, Frank; Ephrussi, Anne

2017-10-01

Arrays of singly labeled short oligonucleotides that hybridize to a specific target revolutionized RNA biology, enabling quantitative, single-molecule microscopy analysis and high-efficiency RNA/RNP capture. Here, we describe a simple and efficient method that allows flexible functionalization of inexpensive DNA oligonucleotides by different fluorescent dyes or biotin using terminal deoxynucleotidyl transferase and custom-made functional group conjugated dideoxy-UTP. We show that (i) all steps of the oligonucleotide labeling-including conjugation, enzymatic synthesis, and product purification-can be performed in a standard biology laboratory, (ii) the process yields >90%, often >95% labeled product with minimal carryover of impurities, and (iii) the oligonucleotides can be labeled with different dyes or biotin, allowing single-molecule FISH, RNA affinity purification, and Northern blot analysis to be performed. © 2017 Gaspar et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Flow cytometric immunobead assay for quantitative detection of platelet autoantibodies in immune thrombocytopenia patients.

Science.gov (United States)

Zhai, Juping; Ding, Mengyuan; Yang, Tianjie; Zuo, Bin; Weng, Zhen; Zhao, Yunxiao; He, Jun; Wu, Qingyu; Ruan, Changgeng; He, Yang

2017-10-23

Platelet autoantibody detection is critical for immune thrombocytopenia (ITP) diagnosis and prognosis. Therefore, we aimed to establish a quantitative flow cytometric immunobead assay (FCIA) for ITP platelet autoantibodies evaluation. Capture microbeads coupled with anti-GPIX, -GPIb, -GPIIb, -GPIIIa and P-selectin antibodies were used to bind the platelet-bound autoantibodies complex generated from plasma samples of 250 ITP patients, 163 non-ITP patients and 243 healthy controls, a fluorescein isothiocyanate (FITC)-conjugated secondary antibody was the detector reagent and mean fluorescence intensity (MFI) signals were recorded by flow cytometry. Intra- and inter-assay variations of the quantitative FCIA assay were assessed. Comparisons of the specificity, sensitivity and accuracy between quantitative and qualitative FCIA or monoclonal antibody immobilization of platelet antigen (MAIPA) assay were performed. Finally, treatment process was monitored by our quantitative FCIA in 8 newly diagnosed ITPs. The coefficient of variations (CV) of the quantitative FCIA assay were respectively 9.4, 3.8, 5.4, 5.1 and 5.8% for anti-GPIX, -GPIb, -GPIIIa, -GPIIb and -P-selectin autoantibodies. Elevated levels of autoantibodies against platelet glycoproteins GPIX, GPIb, GPIIIa, GPIIb and P-selectin were detected by our quantitative FCIA in ITP patients compared to non-ITP patients or healthy controls. The sensitivity, specificity and accuracy of our quantitative assay were respectively 73.13, 81.98 and 78.65% when combining all 5 autoantibodies, while the sensitivity, specificity and accuracy of MAIPA assay were respectively 41.46, 90.41 and 72.81%. A quantitative FCIA assay was established. Reduced levels of platelet autoantibodies could be confirmed by our quantitative FCIA in ITP patients after corticosteroid treatment. Our quantitative assay is not only good for ITP diagnosis but also for ITP treatment monitoring.

On-chip transduction of nucleic acid hybridization using spatial profiles of immobilized quantum dots and fluorescence resonance energy transfer.

Science.gov (United States)

Tavares, Anthony J; Noor, M Omair; Vannoy, Charles H; Algar, W Russ; Krull, Ulrich J

2012-01-03

The glass surface of a glass-polydimethylsiloxane (PDMS) microfluidic channel was modified to develop a solid-phase assay for quantitative determination of nucleic acids. Electroosmotic flow (EOF) within channels was used to deliver and immobilize semiconductor quantum dots (QDs), and electrophoresis was used to decorate the QDs with oligonucleotide probe sequences. These processes took only minutes to complete. The QDs served as energy donors in fluorescence resonance energy transfer (FRET) for transduction of nucleic acid hybridization. Electrokinetic injection of fluorescent dye (Cy3) labeled oligonucleotide target into a microfluidic channel and subsequent hybridization (within minutes) provided the proximity for FRET, with emission from Cy3 being the analytical signal. The quantification of target concentration was achieved by measurement of the spatial length of coverage by target along a channel. Detection of femtomole quantities of target was possible with a dynamic range spanning an order of magnitude. The assay provided excellent resistance to nonspecific interactions of DNA. Further selectivity of the assay was achieved using 20% formamide, which allowed discrimination between a fully complementary target and a 3 base pair mismatch target at a contrast ratio of 4:1. © 2011 American Chemical Society

Characterization and evaluation of rice blast resistance of Chinese indica hybrid rice parental lines