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Sample records for amplicon quantification maq

  1. Integrated Microfluidic Nucleic Acid Isolation, Isothermal Amplification, and Amplicon Quantification

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    Michael G. Mauk

    2015-10-01

    Full Text Available Microfluidic components and systems for rapid (<60 min, low-cost, convenient, field-deployable sequence-specific nucleic acid-based amplification tests (NAATs are described. A microfluidic point-of-care (POC diagnostics test to quantify HIV viral load from blood samples serves as a representative and instructive example to discuss the technical issues and capabilities of “lab on a chip” NAAT devices. A portable, miniaturized POC NAAT with performance comparable to conventional PCR (polymerase-chain reaction-based tests in clinical laboratories can be realized with a disposable, palm-sized, plastic microfluidic chip in which: (1 nucleic acids (NAs are extracted from relatively large (~mL volume sample lysates using an embedded porous silica glass fiber or cellulose binding phase (“membrane” to capture sample NAs in a flow-through, filtration mode; (2 NAs captured on the membrane are isothermally (~65 °C amplified; (3 amplicon production is monitored by real-time fluorescence detection, such as with a smartphone CCD camera serving as a low-cost detector; and (4 paraffin-encapsulated, lyophilized reagents for temperature-activated release are pre-stored in the chip. Limits of Detection (LOD better than 103 virons/sample can be achieved. A modified chip with conduits hosting a diffusion-mode amplification process provides a simple visual indicator to readily quantify sample NA template. In addition, a companion microfluidic device for extracting plasma from whole blood without a centrifuge, generating cell-free plasma for chip-based molecular diagnostics, is described. Extensions to a myriad of related applications including, for example, food testing, cancer screening, and insect genotyping are briefly surveyed.

  2. Amplicon sequencing for the quantification of spoilage microbiota in complex foods including bacterial spores

    NARCIS (Netherlands)

    Boer, de P.; Caspers, M.; Sanders, J.W.; Kemperman, R.; Wijman, J.; Lommerse, G.; Roeselers, G.; Montijn, R.; Abee, T.; Kort, R.

    2015-01-01

    Background
    Spoilage of food products is frequently caused by bacterial spores and lactic acid bacteria. Identification of these organisms by classic cultivation methods is limited by their ability to form colonies on nutrient agar plates. In this study, we adapted and optimized 16S rRNA amplicon

  3. The Quantification of Representative Sequences pipeline for amplicon sequencing: case study on within-population ITS1 sequence variation in a microparasite infecting Daphnia.

    Science.gov (United States)

    González-Tortuero, E; Rusek, J; Petrusek, A; Gießler, S; Lyras, D; Grath, S; Castro-Monzón, F; Wolinska, J

    2015-11-01

    Next generation sequencing (NGS) platforms are replacing traditional molecular biology protocols like cloning and Sanger sequencing. However, accuracy of NGS platforms has rarely been measured when quantifying relative frequencies of genotypes or taxa within populations. Here we developed a new bioinformatic pipeline (QRS) that pools similar sequence variants and estimates their frequencies in NGS data sets from populations or communities. We tested whether the estimated frequency of representative sequences, generated by 454 amplicon sequencing, differs significantly from that obtained by Sanger sequencing of cloned PCR products. This was performed by analysing sequence variation of the highly variable first internal transcribed spacer (ITS1) of the ichthyosporean Caullerya mesnili, a microparasite of cladocerans of the genus Daphnia. This analysis also serves as a case example of the usage of this pipeline to study within-population variation. Additionally, a public Illumina data set was used to validate the pipeline on community-level data. Overall, there was a good correspondence in absolute frequencies of C. mesnili ITS1 sequences obtained from Sanger and 454 platforms. Furthermore, analyses of molecular variance (amova) revealed that population structure of C. mesnili differs across lakes and years independently of the sequencing platform. Our results support not only the usefulness of amplicon sequencing data for studies of within-population structure but also the successful application of the QRS pipeline on Illumina-generated data. The QRS pipeline is freely available together with its documentation under GNU Public Licence version 3 at http://code.google.com/p/quantification-representative-sequences.

  4. HPC-MAQ : A PARALLEL SHORT-READ REFERENCE ASSEMBLER

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    Veeram Venkata Siva Prasad

    2011-07-01

    Full Text Available Bioinformatics and computational biology are rooted in life sciences as well as computer and information sciences and technologies. Bioinformatics applies principles of information sciences and technologies to make the vast, diverse, and complex life sciences data more understandable and useful. Computational biology uses mathematical and computational approaches to address theoretical and experimental questions in biology. Short read sequence assembly is one of the most important steps in the analysis of biological data. There are many open source software’s available for short read sequence assembly where MAQ is one such popularly used software by the research community. In general, biological data sets generated by next generation sequencers are very huge and massive which requires tremendous amount of computational resources. The algorithm used for the short read sequence assembly is NP Hard which is computationally expensive and time consuming. Also MAQ is single threaded software which doesn't use the power of multi core and distributed computing and it doesn't scale. In this paper we report HPC-MAQ which addresses the NP-Hard related challenges of genome reference assembly and enables MAQ parallel and scalable through Hadoop which is a software framework for distributed computing.

  5. Mobile Air Quality Studies (MAQS-an international project

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    Sudik Claudia

    2010-04-01

    Full Text Available Abstract Due to an increasing awareness of the potential hazardousness of air pollutants, new laws, rules and guidelines have recently been implemented globally. In this respect, numerous studies have addressed traffic-related exposure to particulate matter using stationary technology so far. By contrast, only few studies used the advanced technology of mobile exposure analysis. The Mobile Air Quality Study (MAQS addresses the issue of air pollutant exposure by combining advanced high-granularity spatial-temporal analysis with vehicle-mounted, person-mounted and roadside sensors. The MAQS-platform will be used by international collaborators in order 1 to assess air pollutant exposure in relation to road structure, 2 to assess air pollutant exposure in relation to traffic density, 3 to assess air pollutant exposure in relation to weather conditions, 4 to compare exposure within vehicles between front and back seat (children positions, and 5 to evaluate "traffic zone"-exposure in relation to non-"traffic zone"-exposure. Primarily, the MAQS-platform will focus on particulate matter. With the establishment of advanced mobile analysis tools, it is planed to extend the analysis to other pollutants including NO2, SO2, nanoparticles and ozone.

  6. Application of Sharī‘ah contracts in contemporary Islamic finance: A maqāṣid perspective

    OpenAIRE

    Younes Soualhi

    2015-01-01

    This research exposes the underlying maqāṣid embedded in Sharī‘ah contracts as applied in Islamic banking and finance. It addresses the problem of not observing maqāṣid in nominated and combined Sharī‘ah contracts as well as the problem of not sufficiently imbuing maqāṣid in products developed by Islamic financial institutions. As a benchmark of the maqāṣid of wealth, the research adopts Ibn ‘Āshūr’s classification of maqāṣid to evaluate the conformity of Sharī‘ah contracts to Maqāṣid al-Shar...

  7. Measuring the performance of Islamic banks using maqāsid based model

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    Mustafa Omar Mohammed

    2015-12-01

    Full Text Available The vision and mission of Islamic banks were supposed to reflect the adherence of their activities and aspiration to Maqāṣid al-Sharī‘ah. However, there are contentions that Islamic banks have been converging towards conventional banking system. Efforts have been expended to reverse the tide and harmonise Islamic banking to its Sharī‘ah objectives. Hitherto, the existing conventional yardsticks have failed to measure the impact of the harmonisation exercise on Islamic banks’ performance. Therefore, using maqāṣid based yardstick to measure the performance of Islamic banks becomes imperative. This study has made use of al-Imām al-Ghazālī’s theory of Maqāṣid al-Sharī‘ah and Ibn ‘Āshūr’s reinterpretation, adopting content analysis and Sekaran (2000 behavioral science methods to develop a Maqāṣid Based Performance Evaluation Model (MPEM to measure the performance of Islamic banks. Experts’ opinions have validated the model and its acceptability. Suggestions are provided to policy makers and future research.

  8. Application of Sharī‘ah contracts in contemporary Islamic finance: A maqāṣid perspective

    Directory of Open Access Journals (Sweden)

    Younes Soualhi

    2015-12-01

    Full Text Available This research exposes the underlying maqāṣid embedded in Sharī‘ah contracts as applied in Islamic banking and finance. It addresses the problem of not observing maqāṣid in nominated and combined Sharī‘ah contracts as well as the problem of not sufficiently imbuing maqāṣid in products developed by Islamic financial institutions. As a benchmark of the maqāṣid of wealth, the research adopts Ibn ‘Āshūr’s classification of maqāṣid to evaluate the conformity of Sharī‘ah contracts to Maqāṣid al-Sharī‘ah namely, justice, circulation, transparency, and firmness. The study focuses on three markets related to the application of Sharī‘ah contracts, namely, banking, Islamic capital market, and takāful. The study concludes that, by and large, the application of Sharī‘ah contracts has observed Maqāṣid al-Sharī‘ah during its development and initial application stages of Islamic finance products; however, offering such products in the market has raised economic questions as to their viability and economic values. In addition, the malpractice of some Sharī‘ah contracts has long raised concerns as to the maqāṣid compliance of such products. The research recommends a de-sophistication of Islamic financial engineering to minimise the possibility of convergence with conventional finance. The research also emphasises product differentiation based on less complicated combined Sharī‘ah contracts.

  9. Maqâsid al-Qur’ân dan Deradikalisasi Penafsiran dalam Konteks Keindonesiaan

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    Ulya Fikriyati

    2015-09-01

    Full Text Available This article deals with the viewpoint that the reading of the Qur’ân by a certain generation is subject to criticism by the following generation. The article seeks to offer, as an example, the deradicalization of interpreting the so-called ‘radical’ verses of the Qur’ân in Indonesian context. Islam in Indonesia always interact with various races, ethnicities, religions and beliefs, and therefore requires a type of exegesis different from other regions such as the Middle East. For the radicalization of interpretation, this article offers what is called the maqâsid al-Qur’ân as its parameter. The maqâsid al-Qur’ân consists of seven points: 1 Hifz al-dîn wa tatwîr wasâilih, 2 Hifz al-nafs wa tatwîruhâ, 3 Hifz al-‘aql wa tatwîruh, 4 Hifz al-mâl wa tanmîyat wasâilih, 5 Hifz al-‘ird wa tatwîr al-wasâil li al-husûl ‘alayh, 6 Tahqîq al-huqûq al-insânîyah wa mâ yandarij tahtahâ, 7 Hifz al-‘âlam wa ‘imâratuhâ. As spirit and parameter, the maqâsid al-Qur’ân necessitates the dialectics of dynamic interpretation without any judgment of infidelity or heresy. If a certain reading of the Qur’anic verses deviates from these seven maqâsid al-Qur’ân above, it deserves to be examined further, but not to be immediately suppressed.

  10. Dispute management in Islamic financial services and products: A maqāṣid-based analysis

    OpenAIRE

    Umar A. Oseni

    2015-01-01

    The increasing expansion of the Islamic financial services industry beyond its original frontiers has not only come with success stories but has also been affected by the growing preference for litigation as the mode of dispute resolution. Exorbitant legal fees and cost of sustaining protracted litigation are two major challenges that require the attention of major stakeholders in the industry. This paper examines these challenges through a Maqāṣid al-Sharī‘ah focused prism considering the im...

  11. Removing Noise From Pyrosequenced Amplicons

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    Davenport Russell J

    2011-01-01

    Full Text Available Abstract Background In many environmental genomics applications a homologous region of DNA from a diverse sample is first amplified by PCR and then sequenced. The next generation sequencing technology, 454 pyrosequencing, has allowed much larger read numbers from PCR amplicons than ever before. This has revolutionised the study of microbial diversity as it is now possible to sequence a substantial fraction of the 16S rRNA genes in a community. However, there is a growing realisation that because of the large read numbers and the lack of consensus sequences it is vital to distinguish noise from true sequence diversity in this data. Otherwise this leads to inflated estimates of the number of types or operational taxonomic units (OTUs present. Three sources of error are important: sequencing error, PCR single base substitutions and PCR chimeras. We present AmpliconNoise, a development of the PyroNoise algorithm that is capable of separately removing 454 sequencing errors and PCR single base errors. We also introduce a novel chimera removal program, Perseus, that exploits the sequence abundances associated with pyrosequencing data. We use data sets where samples of known diversity have been amplified and sequenced to quantify the effect of each of the sources of error on OTU inflation and to validate these algorithms. Results AmpliconNoise outperforms alternative algorithms substantially reducing per base error rates for both the GS FLX and latest Titanium protocol. All three sources of error lead to inflation of diversity estimates. In particular, chimera formation has a hitherto unrealised importance which varies according to amplification protocol. We show that AmpliconNoise allows accurate estimates of OTU number. Just as importantly AmpliconNoise generates the right OTUs even at low sequence differences. We demonstrate that Perseus has very high sensitivity, able to find 99% of chimeras, which is critical when these are present at high

  12. Empirical Likelihood Inference for MA(q) Model%MA(q)模型的经验似然推断

    Institute of Scientific and Technical Information of China (English)

    陈燕红; 宋立新

    2009-01-01

    In this article we study the empirical likelihood inference for MA(q) model.We propose the moment restrictions,by which we get the empirical likelihood estimator of the model parameter,and we also propose an empirical log-likelihood ratio based on this estimator.Our result shows that the EL estimator is asymptotically normal,and the empirical log-likelihood ratio is proved to be asymptotical standard chi-square distribution.

  13. The validation of the Indonesian version of psychotic symptoms ratings scale (PSYRATS, the Indonesian version of cognitive bias questionnaire for psychosis (CBQP and metacognitive ability questionnaire (MAQ

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    Erna Erawati

    2014-10-01

    Full Text Available Aim: The present study was to validate the Indonesian version of Psychotic Symptoms Ratings Scales (PSYRATS, the Cognitive Bias Questionnaire for Psychosis (CBQp and the Metacognitive Ability Questionnaire (MAQ as a new scale to measure the ability of metacognition of schizophrenia. Background: The PSYRATS, CBQp, and MAQ have demonstrated their usefulness for the assessment of hallucinations and delusions, cognitive biases and metacognitive ability in schizophrenia. So far no validation of the Indonesian version has been carried out. Methods: The PSYRATS, CBQp and MAQ were administered to 155 subjects with a diagnosis of schizophrenia. Factor structure, reliability, test-retest stability, and convergent validity were analyzed. Findings: We found that the all psychometric were reliable and valid. Indonesian version of the PSYRATS and CBQp Indonesian version have high reliability. The reliability of new psychometric of MAQ was Cronbachs alpha=.759 and was checked in a subsample (n=32; r=.668; p<.01. Conclusions: Similar to the original PSYRATS and CBQp, the Indonesian version of PSYRATS and CBQp have good psychometric properties. The new psychometric of MAQ is a valid instrument for assessing metacognition. Implications for future research are discussed. Keywords: Schizophrenia; Hallucination Severity; Delusion Severity; Bias Cognitive for Psychosis; Metacognition: Instrumental Study.

  14. Dispute management in Islamic financial services and products: A maqāṣid-based analysis

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    Umar A. Oseni

    2015-12-01

    Full Text Available The increasing expansion of the Islamic financial services industry beyond its original frontiers has not only come with success stories but has also been affected by the growing preference for litigation as the mode of dispute resolution. Exorbitant legal fees and cost of sustaining protracted litigation are two major challenges that require the attention of major stakeholders in the industry. This paper examines these challenges through a Maqāṣid al-Sharī‘ah focused prism considering the importance of the sustainable dispute management framework in Islamic financial services and products. While singling out the important higher objective (maqṣad of ḥifẓ al-māl, this study argues that preservation of wealth and financial resources requires effective means of resolving increasingly diverse disputes in the Islamic financial services industry. It is further argued that an effective dispute management framework will consider the original value proposition of Islamic financial intermediation which promotes maṣlaḥah (benefits and prevents mafsadah (hardship and ḍarar (financial harm. This makes a case for the affirmative relevance, potential adoption, and systemic modernisation of Islamic dispute management mechanisms such as ṣulḥ, taḥkīm, and muḥtasib in order to fulfil the overarching objective of protection and preservation of wealth and financial resources as one of the core objectives of Sharī‘ah.

  15. The Discourse of Medicine in the Čahār Maqāla (Four Discourses) of Nezami Aruzi of Samarghand.

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    Afshar, Ahmadreza

    2015-09-01

    Nezami Aruzi prepared Čahār Maqāla (Four Discourses) as a guide and admonishment for the rulers and kings. The fourth discourse of Čahār Maqāla with 12 anecdotes is devoted to the science of medicine and the characteristics of the physicians. The discourse presents the name of the eminent scientists, physicians, as well as Farsi and Arabic medical books that had professional acceptance in the medieval in Persia. The author has described how medicine was studied in the medieval in Persia and has presented notes on the physiology of the nervous system, pulse, uroscopy, fever, spiritual affairs and medical ethics. The current essay is a brief review of the medical subjects in Čahār Maqāla. PMID:26317607

  16. Freedom, Sufism, maqâm hurrîyah, hâl hurrîyah.

    OpenAIRE

    Ah. Haris Fakhrudi

    2015-01-01

    This paper will discuss the meaning of freedom in the discourse of Sufi thought, especially of Ibn ‘Arabî. This is based on the consideration that Sufism before Ibn ‘Arabî’s more focused on ritualistic orientation for students and only revealed variant of Sufi’s expressions, both on maqâmât and ahwâl. The presence of Ibn ‘Arabî, therefore, became the turning point in the discourse of Sufism by expressing his beliefs in the theoretical formulation. The doctrine of Sufism—which previously only ...

  17. EKSEKUSI HUKUMAN MATI Tinjauan Maqāṣid al-Sharī’ah dan Keadilan

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    Imam Yahya

    2013-04-01

    Full Text Available The debate about death penalty, is still attracted attention of people. At least, there are, two mainstream firstly those who agrees and secondly who refuses the death penalty being imposed. For those who agrees reasoned that severe violations of the right to life, should be punished by death so that could provide a deterrent effect, while those who refuses argued that the death penalty is a denial of human rights, especially right to life. The essence of the death penalty is not a violation of the law, because the implementation the death penalty actually enforced in order to protect human rights itself. In the view of Islamic law, death penalty, can be done on four cases, namely that of adultery, killing intentionally, Hirabah and apostasy. Furthermore, the death penalty should be carried out in accordance with maqāṣid al-sharī'ah and justice. In maqāṣid al-sharī'ah perspective, the purpose of death penalty should refer to maintain religion (ḥifẓ al-dīn, maintain body or maintain the survival (ḥifẓ al-nafs, mind (ḥifẓ al-'aql, descent (ḥifẓ alnasl, and maintaining property (ḥifẓ al-māl. While in the perspective of justice, State, on behalf of the law must protect its citizens from legal events that harm society.

  18. Freedom, Sufism, maqâm hurrîyah, hâl hurrîyah.

    Directory of Open Access Journals (Sweden)

    Ah. Haris Fakhrudi

    2015-09-01

    Full Text Available This paper will discuss the meaning of freedom in the discourse of Sufi thought, especially of Ibn ‘Arabî. This is based on the consideration that Sufism before Ibn ‘Arabî’s more focused on ritualistic orientation for students and only revealed variant of Sufi’s expressions, both on maqâmât and ahwâl. The presence of Ibn ‘Arabî, therefore, became the turning point in the discourse of Sufism by expressing his beliefs in the theoretical formulation. The doctrine of Sufism—which previously only implicitly contained in the words of the Sufi shaykh—in the hands of Ibn ‘Arabî flashed into an open, theoretical, and obvios and thus opened the door for anyone who has a high intelligence in reflecting at once and realizing the metaphysical theories through operational forms. Therefore, this article will discuss some of the key concepts in the thought of Ibn ‘Arabî including the meaning of freedom (al-hurrîyah in Sufism, maqâm hurrîyah, and hâl hurrîyah obtained by the Sufis during their spiritual journey.

  19. MaqFACS (Macaque Facial Action Coding System) can be used to document facial movements in Barbary macaques (Macaca sylvanus).

    Science.gov (United States)

    Julle-Danière, Églantine; Micheletta, Jérôme; Whitehouse, Jamie; Joly, Marine; Gass, Carolin; Burrows, Anne M; Waller, Bridget M

    2015-01-01

    Human and non-human primates exhibit facial movements or displays to communicate with one another. The evolution of form and function of those displays could be better understood through multispecies comparisons. Anatomically based coding systems (Facial Action Coding Systems: FACS) are developed to enable such comparisons because they are standardized and systematic and aid identification of homologous expressions underpinned by similar muscle contractions. To date, FACS has been developed for humans, and subsequently modified for chimpanzees, rhesus macaques, orangutans, hylobatids, dogs, and cats. Here, we wanted to test whether the MaqFACS system developed in rhesus macaques (Macaca mulatta) could be used to code facial movements in Barbary macaques (M. sylvanus), a species phylogenetically close to the rhesus macaques. The findings show that the facial movement capacity of Barbary macaques can be reliably coded using the MaqFACS. We found differences in use and form of some movements, most likely due to specializations in the communicative repertoire of each species, rather than morphological differences.

  20. Pendekatan Maqāṣid Syarī’ah: Konstruksi Terhadap Pengembangan Ilmu Ekonomi dan Keuangan Islam

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    Ayief Fathurrahman

    2014-12-01

    Full Text Available This article examines the problems that occur in the development process of Islamic economic and finance, both in theory and practice, especially in the Islamic banking practice in various parts of the world. To cope with the problems, the maqāṣid al-syarī’ah approach can be employed to understand the verses of the Qur’an and the sunnah, to reconsolidate the conflicting arguments, and to decide the legal standing to a case of which legal sanding is not mentioned in the Qur’an and the sunnah. The purpose is to make Islamic economic and finance relevant to the change of time and to reduce the turbulence taking place in the practice of Islamic economy and finance themselves.

  1. Metabarcoding Marine Sediments: Preparation of Amplicon Libraries.

    Science.gov (United States)

    Fonseca, Vera G; Lallias, Delphine

    2016-01-01

    The accurate assessment of community composition and ultimately species identification is of utmost importance in any ecological and evolutionary study. Advances in sequencing technologies have allowed the unraveling of levels of biodiversity never imagined before when applied to large-scale environmental DNA studies (also termed metabarcoding/metagenetics/metasystematics/environmental barcoding). Here, we describe a detailed protocol to assess eukaryotic biodiversity in marine sediments, identifying key steps that should not be neglected when preparing Next-Generation Sequencing (NGS) amplicon libraries: DNA extraction, multiple PCR amplification of DNA barcode markers with index/ tag-primers, and final Illumina MiSeq sequencing library preparation. PMID:27460378

  2. Sample Preparation for Fungal Community Analysis by High-Throughput Sequencing of Barcode Amplicons.

    Science.gov (United States)

    Clemmensen, Karina Engelbrecht; Ihrmark, Katarina; Durling, Mikael Brandström; Lindahl, Björn D

    2016-01-01

    Fungal species participate in vast numbers of processes in the landscape around us. However, their often cryptic growth, inside various substrates and in highly diverse species assemblages, has been a major obstacle to thorough analysis of fungal communities, hampering exhaustive description of the fungal kingdom. Recent technological developments allowing rapid, high-throughput sequencing of mixed communities from many samples at once are currently having a tremendous impact in fungal community ecology. Universal DNA extraction followed by amplification and sequencing of fungal species-level barcodes such as the nuclear internal transcribed spacer (ITS) region now enable identification and relative quantification of fungal community members across well-replicated experimental settings. Here, we present the sample preparation procedure presently used in our laboratory for fungal community analysis by high-throughput sequencing of amplified ITS2 markers. We focus on the procedure optimized for studies of total fungal communities in humus-rich soils, wood, and litter. However, this procedure can be applied to other sample types and markers. We focus on the laboratory-based part of sample preparation, that is, the procedure from the point where samples enter the laboratory until amplicons are submitted for sequencing. Our procedure comprises four main parts: (1) universal DNA extraction, (2) optimization of PCR conditions, (3) production of tagged ITS amplicons, and (4) preparation of the multiplexed amplicon mix to be sequenced. The presented procedure is independent of the specific high-throughput sequencing technology used, which makes it highly versatile. PMID:26791497

  3. A Maqāṣidī approach to contemporary application of the Sharī‘ah

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    Jasser Auda

    2011-12-01

    Full Text Available Normal 0 false false false EN-US KO AR-SA /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-parent:""; mso-padding-alt:0in 5.4pt 0in 5.4pt; mso-para-margin-top:0in; mso-para-margin-right:0in; mso-para-margin-bottom:10.0pt; mso-para-margin-left:0in; line-height:115%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:Arial; mso-bidi-theme-font:minor-bidi; mso-ansi-language:EN-US; mso-fareast-language:EN-US;} This paper explores how Maqāṣid al-Sharī‘ah (higher purposes and intents of Islamic law could contribute to the application of the Sharī‘ah itself in contemporary Muslim societies and to making the appropriate related juridical policies. The soundness of the application of the Shar‘īah and related policies is subject to the degree of universality and flexibility of the Islamic rulings with changing circumstances, are discussed from various viewpoints in this paper. After a survey of the system of values that Maqāṣid al-Sharī‘ah represent, three methods are explored: (1 differentiating between scripts that are means (wasā’il to higher ends and scripts that are ends (ahdāf in their own right, (2 preferring a multi-dimensional understanding for the conciliation of opposing juridical evidence, instead of reductionist methods such as abrogation (naskh and elimination (tarjīḥ, and (3 achieving a universality of Sharī‘ah across cultures via the consideration of customs (al-‘urf. A number of examples are provided throughout the paper in order to explain the impact of the proposed methods on contemporary Islamic rulings and juridical policies related to them.

  4. Brief communication: MaqFACS: A muscle-based facial movement coding system for the rhesus macaque.

    Science.gov (United States)

    Parr, L A; Waller, B M; Burrows, A M; Gothard, K M; Vick, S J

    2010-12-01

    Over 125 years ago, Charles Darwin (1872) suggested that the only way to fully understand the form and function of human facial expression was to make comparisons with other species. Nevertheless, it has been only recently that facial expressions in humans and related primate species have been compared using systematic, anatomically based techniques. Through this approach, large-scale evolutionary and phylogenetic analyses of facial expressions, including their homology, can now be addressed. Here, the development of a muscular-based system for measuring facial movement in rhesus macaques (Macaca mulatta) is described based on the well-known FACS (Facial Action Coding System) and ChimpFACS. These systems describe facial movement according to the action of the underlying facial musculature, which is highly conserved across primates. The coding systems are standardized; thus, their use is comparable across laboratories and study populations. In the development of MaqFACS, several species differences in the facial movement repertoire of rhesus macaques were observed in comparison with chimpanzees and humans, particularly with regard to brow movements, puckering of the lips, and ear movements. These differences do not seem to be the result of constraints imposed by morphological differences in the facial structure of these three species. It is more likely that they reflect unique specializations in the communicative repertoire of each species.

  5. Harmonising legality with morality in Islamic banking and finance: A quest for Maqāṣid al-Sharī‘ah paradigm

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    Luqman Zakariyah

    2015-12-01

    Full Text Available Scholars in Islamic Finance Industry (IFI have been calling for the integration of Islamic morality with legal theories in the industry. Among the reasons for this call is an unethical trend in product innovation. Implementing Islamic banking and financial practices would require adopting their undergirding Islamic legal and moral frameworks. Departing from these foundations of Islamic law could render the activities conducted under its name religiously unacceptable. Many approaches have been put forward to achieve this cause. One of the most complex yet subjective approaches is the quest for Maqāṣid al-Sharī‘ah. This paper critically examines the feasibility of harmonising morality with legality in Islamic finance. In doing so, it will reveal what constitutes morality and legality in Islamic legal theory, and critically examine the approaches of Muslim classical scholars in fusing the two elements together for the realisation and actualisation of the very objectives of Sharī‘ah. Questions of the relationship between morality and legality are raised, and samples of Islamic finance products are evaluated to expose their moral and legal dimensions. Lastly, the role of Maqāṣid al-Sharī‘ah in the process of harmonisation is discussed with some observations and reservations on the practicality of their implementation.

  6. Systems consequences of amplicon formation in human breast cancer.

    Science.gov (United States)

    Inaki, Koichiro; Menghi, Francesca; Woo, Xing Yi; Wagner, Joel P; Jacques, Pierre-Étienne; Lee, Yi Fang; Shreckengast, Phung Trang; Soon, Wendy WeiJia; Malhotra, Ankit; Teo, Audrey S M; Hillmer, Axel M; Khng, Alexis Jiaying; Ruan, Xiaoan; Ong, Swee Hoe; Bertrand, Denis; Nagarajan, Niranjan; Karuturi, R Krishna Murthy; Miranda, Alfredo Hidalgo; Liu, Edison T

    2014-10-01

    Chromosomal structural variations play an important role in determining the transcriptional landscape of human breast cancers. To assess the nature of these structural variations, we analyzed eight breast tumor samples with a focus on regions of gene amplification using mate-pair sequencing of long-insert genomic DNA with matched transcriptome profiling. We found that tandem duplications appear to be early events in tumor evolution, especially in the genesis of amplicons. In a detailed reconstruction of events on chromosome 17, we found large unpaired inversions and deletions connect a tandemly duplicated ERBB2 with neighboring 17q21.3 amplicons while simultaneously deleting the intervening BRCA1 tumor suppressor locus. This series of events appeared to be unusually common when examined in larger genomic data sets of breast cancers albeit using approaches with lesser resolution. Using siRNAs in breast cancer cell lines, we showed that the 17q21.3 amplicon harbored a significant number of weak oncogenes that appeared consistently coamplified in primary tumors. Down-regulation of BRCA1 expression augmented the cell proliferation in ERBB2-transfected human normal mammary epithelial cells. Coamplification of other functionally tested oncogenic elements in other breast tumors examined, such as RIPK2 and MYC on chromosome 8, also parallel these findings. Our analyses suggest that structural variations efficiently orchestrate the gain and loss of cancer gene cassettes that engage many oncogenic pathways simultaneously and that such oncogenic cassettes are favored during the evolution of a cancer. PMID:25186909

  7. Multiplex PCR, amplicon size and hybridization efficiency on the NanoChip electronic microarray

    DEFF Research Database (Denmark)

    Børsting, Claus; Sanchez, Juan J; Morling, Niels

    2004-01-01

    . Hybridization to individual amplicons in multiplexes was less efficient suggesting that intramolecular and intermolecular interactions may block access to the target sequence on the NanoChip array. We observed a high risk of contamination with amplicons shorter than 60 bp and therefore, we recommend the use......We tested the SNP typing protocol developed for the NanoChip electronic microarray by analyzing the four Y chromosome loci SRY1532, SRY8299, TAT, and 92R7. Amplicons of different lengths containing the same locus were purified and addressed to the NanoChip array and fluorescently labelled reporter...... probes were hybridized to the amplicons. We demonstrated that as little as 10-30 fmol of 50 bp DNA amplicons was sufficient to obtain strong and reproducible results. The hybridization to 50 bp amplicons was up to 10 times more efficient than the hybridization to 200 bp amplicons containing the same SNP...

  8. Innovación y ruptura en la métrica de la poesía en al-Maqāmāt al-Luzūmiyya de al-Saraqusṭī (s. VI/XII

    Directory of Open Access Journals (Sweden)

    Ferrando, Ignacio

    2016-06-01

    Full Text Available Al-Maqāmāt al-Luzūmiyya, a collection of 50 picaresque-like stories couched in rhymed prose by Abū l-Ṭāhir al-Saraqusṭī in al-Andalus in the VI/XIIth century, were considered by Western scholars, up until the eighties of XXth century, as highly rhetorical pieces with a negligible and irrelevant content. However, some contemporary scholars argued that they contain some degree of social and political criticism, as they constitute a fictional domain in which subversion and irony play a fundamental role. In this paper, we cast some light on the particular use of metrical patterns in Al-Maqāmāt al-Luzūmiyya, which should be considered another subversion field: a poetical rhythm that avoids the most usual Arabic metrical patterns, contributing to provide the maqāma a flavor of the countergenre.Al-Maqāmāt al-Luzūmiyya, la colección de 50 relatos de corte picaresco escritos en una exigente prosa rimada por Abū l-Ṭāhir al-Saraqusṭī en al-Andalus en el siglo VI/XII, han sido consideradas por los críticos occidentales como piezas de alta retórica y de virtuosismo lingüístico en las que el contenido es prácticamente irrelevante. Sin embargo, a partir de los años 80 del siglo pasado, algunos investigadores han tratado de incidir en el hecho de que esta obra contiene cierto grado de crítica social y política, puesto que constituye un espacio ficticio en el que la subversión y la ironía desempeñan un papel fundamental. En este trabajo vamos a tratar de arrojar luz sobre el uso particular de los esquemas métricos por parte de al-Saraqusṭī, tratando de mostrar que los metros constituyen otro más de los ámbitos de subversión tan caros al autor. De hecho, el uso de ritmos “invertidos” o contrarios a los más habituales de la poesía clásica contribuye a darle a la maqāma ese aire de anti género que la puebla.

  9. ExploMaq, software para la evaluación energética y económica de la maquinaria agrícola

    Directory of Open Access Journals (Sweden)

    Carlos A. Pereira Marín

    2015-01-01

    Full Text Available El presente trabajo se realizó con el objetivo de desarrollar un software para viabilizar la evaluación energética y económica del conjunto tractor-implemento, con un diseño de aplicación sobre ventanas y sistema portable. El software ExploMaq, determina la evaluación energética del conjunto tractor-máquina agrícola a partir del cálculo de las fuerzas que actúan en una única labor, también evalúa el balance de potencia, calculado a partir de las pérdidas de diferentes tipos de potencia y otros factores de resistencia. Es posible además, la evaluación de los gastos directos de explotación. El software, en su primera versión de prueba y puesta a punto, está a disposición de usuarios con objetivos académicos e investigativos sobre la evaluación energética de máquinas agrícolas.

  10. Mobile air quality studies (MAQS in inner cities: particulate matter PM10 levels related to different vehicle driving modes and integration of data into a geographical information program

    Directory of Open Access Journals (Sweden)

    Uibel Stefanie

    2012-10-01

    Full Text Available Abstract Background Particulate matter (PM is assumed to exert a major burden on public health. Most studies that address levels of PM use stationary measure systems. By contrast, only few studies measure PM concentrations under mobile conditions to analyze individual exposure situations. Methods By combining spatial-temporal analysis with a novel vehicle-mounted sensor system, the present Mobile Air Quality Study (MAQS aimed to analyse effects of different driving conditions in a convertible vehicle. PM10 was continuously monitored in a convertible car, driven with roof open, roof closed, but windows open, or windows closed. Results PM10 values inside the car were nearly always higher with open roof than with roof and windows closed, whereas no difference was seen with open or closed windows. During the day PM10 values varied with high values before noon, and occasional high median values or standard deviation values due to individual factors. Vehicle speed in itself did not influence the mean value of PM10; however, at traffic speed (10 – 50 km/h the standard deviation was large. No systematic difference was seen between PM10 values in stationary and mobile cars, nor was any PM10 difference observed between driving within or outside an environmental (low emission zone. Conclusions The present study has shown the feasibility of mobile PM analysis in vehicles. Individual exposure of the occupants varies depending on factors like time of day as well as ventilation of the car; other specific factors are clearly identifiably and may relate to specific PM10 sources. This system may be used to monitor individual exposure ranges and provide recommendations for preventive measurements. Although differences in PM10 levels were found under certain ventilation conditions, these differences are likely not of concern for the safety and health of passengers.

  11. Deep amplicon sequencing reveals mixed phytoplasma infection within single grapevine plants

    DEFF Research Database (Denmark)

    Nicolaisen, Mogens; Contaldo, Nicoletta; Makarova, Olga;

    2011-01-01

    The diversity of phytoplasmas within single plants has not yet been fully investigated. In this project, deep amplicon sequencing was used to generate 50,926 phytoplasma sequences from 11 phytoplasma-infected grapevine samples from a PCR amplicon in the 5' end of the 16S region. After clustering ...

  12. High throughput 16S rRNA gene amplicon sequencing

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Larsen, Poul; Jørgensen, Mads Koustrup;

    A reliable and reproducible method for identification and quantification of microorganisms is important for the studies of microbial communities in activated sludge and for the demonstration of their significance for plant operation and stability. DNA based identification of microorganisms using 16...... was correlated with the bacterial species composition in 25 Danish full-scale WWTPs with nutrient removal. Examples of properties were SVI, filament index, floc size, floc strength, content of cations and amount of extracellular polymeric substances. Multivariate statistics provided several important insights...

  13. JRC GMO-Amplicons, a collection of nucleic acid sequences related to genetically modified organisms

    OpenAIRE

    PETRILLO MAURO; ANGERS ALEXANDRE; HENRIKSSON PETER; Bonfini, Laura; PATAK DENNSTEDT Alexandre; KREYSA JOACHIM

    2015-01-01

    The DNA target sequence is the key element in designing detection methods for genetically modified organisms (GMOs). Unfortunately this information is frequently lacking, especially for unauthorized GMOs. In addition, patent sequences are generally poorly annotated, buried in complex and extensive documentation and hard to link to the corresponding GM event. Here, we present the JRC GMO-Amplicons, a database of amplicons collected by screening public nucleotide sequence databanks by in silico...

  14. Efficient error correction for next-generation sequencing of viral amplicons

    Directory of Open Access Journals (Sweden)

    Skums Pavel

    2012-06-01

    Full Text Available Abstract Background Next-generation sequencing allows the analysis of an unprecedented number of viral sequence variants from infected patients, presenting a novel opportunity for understanding virus evolution, drug resistance and immune escape. However, sequencing in bulk is error prone. Thus, the generated data require error identification and correction. Most error-correction methods to date are not optimized for amplicon analysis and assume that the error rate is randomly distributed. Recent quality assessment of amplicon sequences obtained using 454-sequencing showed that the error rate is strongly linked to the presence and size of homopolymers, position in the sequence and length of the amplicon. All these parameters are strongly sequence specific and should be incorporated into the calibration of error-correction algorithms designed for amplicon sequencing. Results In this paper, we present two new efficient error correction algorithms optimized for viral amplicons: (i k-mer-based error correction (KEC and (ii empirical frequency threshold (ET. Both were compared to a previously published clustering algorithm (SHORAH, in order to evaluate their relative performance on 24 experimental datasets obtained by 454-sequencing of amplicons with known sequences. All three algorithms show similar accuracy in finding true haplotypes. However, KEC and ET were significantly more efficient than SHORAH in removing false haplotypes and estimating the frequency of true ones. Conclusions Both algorithms, KEC and ET, are highly suitable for rapid recovery of error-free haplotypes obtained by 454-sequencing of amplicons from heterogeneous viruses. The implementations of the algorithms and data sets used for their testing are available at: http://alan.cs.gsu.edu/NGS/?q=content/pyrosequencing-error-correction-algorithm

  15. JRC GMO-Amplicons: a collection of nucleic acid sequences related to genetically modified organisms.

    Science.gov (United States)

    Petrillo, Mauro; Angers-Loustau, Alexandre; Henriksson, Peter; Bonfini, Laura; Patak, Alex; Kreysa, Joachim

    2015-01-01

    The DNA target sequence is the key element in designing detection methods for genetically modified organisms (GMOs). Unfortunately this information is frequently lacking, especially for unauthorized GMOs. In addition, patent sequences are generally poorly annotated, buried in complex and extensive documentation and hard to link to the corresponding GM event. Here, we present the JRC GMO-Amplicons, a database of amplicons collected by screening public nucleotide sequence databanks by in silico determination of PCR amplification with reference methods for GMO analysis. The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF) provides these methods in the GMOMETHODS database to support enforcement of EU legislation and GM food/feed control. The JRC GMO-Amplicons database is composed of more than 240 000 amplicons, which can be easily accessed and screened through a web interface. To our knowledge, this is the first attempt at pooling and collecting publicly available sequences related to GMOs in food and feed. The JRC GMO-Amplicons supports control laboratories in the design and assessment of GMO methods, providing inter-alia in silico prediction of primers specificity and GM targets coverage. The new tool can assist the laboratories in the analysis of complex issues, such as the detection and identification of unauthorized GMOs. Notably, the JRC GMO-Amplicons database allows the retrieval and characterization of GMO-related sequences included in patents documentation. Finally, it can help annotating poorly described GM sequences and identifying new relevant GMO-related sequences in public databases. The JRC GMO-Amplicons is freely accessible through a web-based portal that is hosted on the EU-RL GMFF website. Database URL: http://gmo-crl.jrc.ec.europa.eu/jrcgmoamplicons/.

  16. JRC GMO-Amplicons: a collection of nucleic acid sequences related to genetically modified organisms.

    Science.gov (United States)

    Petrillo, Mauro; Angers-Loustau, Alexandre; Henriksson, Peter; Bonfini, Laura; Patak, Alex; Kreysa, Joachim

    2015-01-01

    The DNA target sequence is the key element in designing detection methods for genetically modified organisms (GMOs). Unfortunately this information is frequently lacking, especially for unauthorized GMOs. In addition, patent sequences are generally poorly annotated, buried in complex and extensive documentation and hard to link to the corresponding GM event. Here, we present the JRC GMO-Amplicons, a database of amplicons collected by screening public nucleotide sequence databanks by in silico determination of PCR amplification with reference methods for GMO analysis. The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF) provides these methods in the GMOMETHODS database to support enforcement of EU legislation and GM food/feed control. The JRC GMO-Amplicons database is composed of more than 240 000 amplicons, which can be easily accessed and screened through a web interface. To our knowledge, this is the first attempt at pooling and collecting publicly available sequences related to GMOs in food and feed. The JRC GMO-Amplicons supports control laboratories in the design and assessment of GMO methods, providing inter-alia in silico prediction of primers specificity and GM targets coverage. The new tool can assist the laboratories in the analysis of complex issues, such as the detection and identification of unauthorized GMOs. Notably, the JRC GMO-Amplicons database allows the retrieval and characterization of GMO-related sequences included in patents documentation. Finally, it can help annotating poorly described GM sequences and identifying new relevant GMO-related sequences in public databases. The JRC GMO-Amplicons is freely accessible through a web-based portal that is hosted on the EU-RL GMFF website. Database URL: http://gmo-crl.jrc.ec.europa.eu/jrcgmoamplicons/. PMID:26424080

  17. Formation of linear amplicons with inverted duplications in Leishmania requires the MRE11 nuclease.

    Directory of Open Access Journals (Sweden)

    Marie-Claude N Laffitte

    2014-12-01

    Full Text Available Extrachromosomal DNA amplification is frequent in the protozoan parasite Leishmania selected for drug resistance. The extrachromosomal amplified DNA is either circular or linear, and is formed at the level of direct or inverted homologous repeated sequences that abound in the Leishmania genome. The RAD51 recombinase plays an important role in circular amplicons formation, but the mechanism by which linear amplicons are formed is unknown. We hypothesized that the Leishmania infantum DNA repair protein MRE11 is required for linear amplicons following rearrangements at the level of inverted repeats. The purified LiMRE11 protein showed both DNA binding and exonuclease activities. Inactivation of the LiMRE11 gene led to parasites with enhanced sensitivity to DNA damaging agents. The MRE11(-/- parasites had a reduced capacity to form linear amplicons after drug selection, and the reintroduction of an MRE11 allele led to parasites regaining their capacity to generate linear amplicons, but only when MRE11 had an active nuclease activity. These results highlight a novel MRE11-dependent pathway used by Leishmania to amplify portions of its genome to respond to a changing environment.

  18. A flexible and economical barcoding approach for highly multiplexed amplicon sequencing of diverse target genes

    Directory of Open Access Journals (Sweden)

    Craig W. Herbold

    2015-07-01

    Full Text Available High throughput sequencing of phylogenetic and functional gene amplicons provides tremendous insight into the structure and functional potential of complex microbial communities. Here, we introduce a highly adaptable and economical PCR approach to barcoding and pooling libraries of numerous target genes. In this approach, we replace gene- and sequencing platform-specific fusion primers with general, interchangeable barcoding primers, enabling nearly limitless customized barcode-primer combinations. Compared to barcoding with long fusion primers, our multiple-target gene approach is more economical because it overall requires lower number of primers and is based on short primers with generally lower synthesis and purification costs. To highlight our approach, we pooled over 900 different small-subunit rRNA and functional gene amplicon libraries obtained from various environmental or host-associated microbial community samples into a single, paired-end Illumina MiSeq run. Although the amplicon regions ranged in size from approximately 290 to 720 bp, we found no significant systematic sequencing bias related to amplicon length or gene target. Our results indicate that this flexible multiplexing approach produces large, diverse and high quality sets of amplicon sequence data for modern studies in microbial ecology.

  19. Silencing Status Epilepticus-Induced BDNF Expression with Herpes Simplex Virus Type-1 Based Amplicon Vectors.

    Directory of Open Access Journals (Sweden)

    Chiara Falcicchia

    Full Text Available Brain-derived neurotrophic factor (BDNF has been found to produce pro- but also anti-epileptic effects. Thus, its validity as a therapeutic target must be verified using advanced tools designed to block or to enhance its signal. The aim of this study was to develop tools to silence the BDNF signal. We generated Herpes simplex virus type 1 (HSV-1 derived amplicon vectors, i.e. viral particles containing a genome of 152 kb constituted of concatameric repetitions of an expression cassette, enabling the expression of the gene of interest in multiple copies. HSV-1 based amplicon vectors are non-pathogenic and have been successfully employed in the past for gene delivery into the brain of living animals. Therefore, amplicon vectors should represent a logical choice for expressing a silencing cassette, which, in multiple copies, is expected to lead to an efficient knock-down of the target gene expression. Here, we employed two amplicon-based BDNF silencing strategies. The first, antisense, has been chosen to target and degrade the cytoplasmic mRNA pool of BDNF, whereas the second, based on the convergent transcription technology, has been chosen to repress transcription at the BDNF gene. Both these amplicon vectors proved to be effective in down-regulating BDNF expression in vitro, in BDNF-expressing mesoangioblast cells. However, only the antisense strategy was effective in vivo, after inoculation in the hippocampus in a model of status epilepticus in which BDNF mRNA levels are strongly increased. Interestingly, the knocking down of BDNF levels induced with BDNF-antisense was sufficient to produce significant behavioral effects, in spite of the fact that it was produced only in a part of a single hippocampus. In conclusion, this study demonstrates a reliable effect of amplicon vectors in knocking down gene expression in vitro and in vivo. Therefore, this approach may find broad applications in neurobiological studies.

  20. Silencing Status Epilepticus-Induced BDNF Expression with Herpes Simplex Virus Type-1 Based Amplicon Vectors.

    Science.gov (United States)

    Falcicchia, Chiara; Trempat, Pascal; Binaschi, Anna; Perrier-Biollay, Coline; Roncon, Paolo; Soukupova, Marie; Berthommé, Hervé; Simonato, Michele

    2016-01-01

    Brain-derived neurotrophic factor (BDNF) has been found to produce pro- but also anti-epileptic effects. Thus, its validity as a therapeutic target must be verified using advanced tools designed to block or to enhance its signal. The aim of this study was to develop tools to silence the BDNF signal. We generated Herpes simplex virus type 1 (HSV-1) derived amplicon vectors, i.e. viral particles containing a genome of 152 kb constituted of concatameric repetitions of an expression cassette, enabling the expression of the gene of interest in multiple copies. HSV-1 based amplicon vectors are non-pathogenic and have been successfully employed in the past for gene delivery into the brain of living animals. Therefore, amplicon vectors should represent a logical choice for expressing a silencing cassette, which, in multiple copies, is expected to lead to an efficient knock-down of the target gene expression. Here, we employed two amplicon-based BDNF silencing strategies. The first, antisense, has been chosen to target and degrade the cytoplasmic mRNA pool of BDNF, whereas the second, based on the convergent transcription technology, has been chosen to repress transcription at the BDNF gene. Both these amplicon vectors proved to be effective in down-regulating BDNF expression in vitro, in BDNF-expressing mesoangioblast cells. However, only the antisense strategy was effective in vivo, after inoculation in the hippocampus in a model of status epilepticus in which BDNF mRNA levels are strongly increased. Interestingly, the knocking down of BDNF levels induced with BDNF-antisense was sufficient to produce significant behavioral effects, in spite of the fact that it was produced only in a part of a single hippocampus. In conclusion, this study demonstrates a reliable effect of amplicon vectors in knocking down gene expression in vitro and in vivo. Therefore, this approach may find broad applications in neurobiological studies.

  1. Use of AmpliWax to optimize amplicon sterilization by isopsoralen.

    Science.gov (United States)

    De la Viuda, M; Fille, M; Ruiz, J; Aslanzadeh, J

    1996-12-01

    The photochemical inactivation of amplicons by isopsoralen (IP-10) has been suggested as a possible means to prevent PCR carryover contamination. To evaluate the technique, serial dilutions of amplicons (10(11) to 10(3)) from the Borrelia burgdorferi OSP A gene were amplified in the presence of 0, 25, 50, and 100 micrograms of IP-10 per ml for 45 cycles. The PCR products were exposed to UV light for 15 min to activate IP-10 and sterilize the amplicons. One microliter of each sterilized sample was reamplified for an additional 45 cycles. The PCR products were then resolved in an agarose gel, blotted onto a nylon membrane, and probed with an alkaline phosphatase-conjugated chemiluminescent probe. Although IP-10 at concentrations of 50 and 100 micrograms/ml effectively sterilized up to 10(11) amplicons, the compound was inhibitory to PCR. IP-10 at a concentration of 25 micrograms/ml had slight inhibitory effect on PCR and did not completely sterilized all of the amplicons. Therefore, in subsequent experiments AmpliWax was substituted for mineral oil, and PCR was performed on 10(9) to 10(3) amplicons as described above. Following the amplification, the PCR tubes were cooled to solidify the AmpliWax and inoculated with various concentrations of IP-10. With this technique, PCR products produced from as many as 10(9) target amplicons were effectively sterilized with 200 micrograms of IP-10 per ml. Similarly, the addition of IP-10 (50 micrograms/ml) before and after PCR was evaluated for the detection of B. burgdorferi in 62 ticks from a region of Southern Connecticut where the organism is highly endemic. PCR performed in the presence of 50 micrograms of IP-10 per ml detected B. burgdorferi-specific DNA in 17 of 62 ticks (27%) following gel electrophoresis and in 34 of 62 ticks (55%) following Southern blot hybridization of the PCR products. In contrast, post-PCR addition of IP-10 detected borrelia-specific DNA in 31 of 62 ticks (50%) following gel electrophoresis and in

  2. Unlabeled Oligonucleotides as Internal Temperature Controls for Genotyping by Amplicon Melting

    OpenAIRE

    Seipp, Michael T.; Durtschi, Jacob D; Liew, Michael A.; Williams, Jamie; Damjanovich, Kristy; Pont-Kingdon, Genevieve; Lyon, Elaine; Voelkerding, Karl V.; Wittwer, Carl T.

    2007-01-01

    Amplicon melting is a closed-tube method for genotyping that does not require probes, real-time analysis, or allele-specific polymerase chain reaction. However, correct differentiation of homozygous mutant and wild-type samples by melting temperature (Tm) requires high-resolution melting and closely controlled reaction conditions. When three different DNA extraction methods were used to isolate DNA from whole blood, amplicon Tm differences of 0.03 to 0.39°C attributable to the extractions wer...

  3. Overexpressed Genes/ESTs and Characterization of Distinct Amplicons on 17823 in Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ayse E. Erson

    2001-01-01

    Full Text Available 17823 is a frequent site of gene amplification in breast cancer. Several lines of evidence suggest the presence of multiple amplicons on 17823. To characterize distinct amplicons on 17823 and localize putative oncogenes, we screened genes and expressed sequence tags (ESTs in existing physical and radiation hybrid maps for amplification and overexpression in breast cancer cell lines by semiquantitative duplex PCR, semiquantitative duplex RT-PCR, Southern blot, Northern blot analyses. We identified two distinct amplicons on 17823, one including TBX2 and another proximal region including RPS6KB1 (PS6K and MUL. In addition to these previously reported overexpressed genes, we also identified amplification and overexpression of additional uncharacterized genes and ESTs, some of which suggest potential oncogenic activity. In conclusion, we have further defined two distinct regions of gene amplification and overexpression on 17823 with identification of new potential oncogene candidates. Based on the amplification and overexpression patterns of known and as of yet unrecognized genes on 17823, it is likely that some of these genes mapping to the discrete amplicons function as oncogenes and contribute to tumor progression in breast cancer cells.

  4. Bacterial metabarcoding by 16S rRNA gene ion torrent amplicon sequencing.

    Science.gov (United States)

    Fantini, Elio; Gianese, Giulio; Giuliano, Giovanni; Fiore, Alessia

    2015-01-01

    Ion Torrent is a next generation sequencing technology based on the detection of hydrogen ions produced during DNA chain elongation; this technology allows analyzing and characterizing genomes, genes, and species. Here, we describe an Ion Torrent procedure applied to the metagenomic analysis of 16S rRNA gene amplicons to study the bacterial diversity in food and environmental samples. PMID:25343859

  5. Evaluation of Hybridization Capture Versus Amplicon-Based Methods for Whole-Exome Sequencing.

    Science.gov (United States)

    Samorodnitsky, Eric; Jewell, Benjamin M; Hagopian, Raffi; Miya, Jharna; Wing, Michele R; Lyon, Ezra; Damodaran, Senthilkumar; Bhatt, Darshna; Reeser, Julie W; Datta, Jharna; Roychowdhury, Sameek

    2015-09-01

    Next-generation sequencing has aided characterization of genomic variation. While whole-genome sequencing may capture all possible mutations, whole-exome sequencing remains cost-effective and captures most phenotype-altering mutations. Initial strategies for exome enrichment utilized a hybridization-based capture approach. Recently, amplicon-based methods were designed to simplify preparation and utilize smaller DNA inputs. We evaluated two hybridization capture-based and two amplicon-based whole-exome sequencing approaches, utilizing both Illumina and Ion Torrent sequencers, comparing on-target alignment, uniformity, and variant calling. While the amplicon methods had higher on-target rates, the hybridization capture-based approaches demonstrated better uniformity. All methods identified many of the same single-nucleotide variants, but each amplicon-based method missed variants detected by the other three methods and reported additional variants discordant with all three other technologies. Many of these potential false positives or negatives appear to result from limited coverage, low variant frequency, vicinity to read starts/ends, or the need for platform-specific variant calling algorithms. All methods demonstrated effective copy-number variant calling when evaluated against a single-nucleotide polymorphism array. This study illustrates some differences between whole-exome sequencing approaches, highlights the need for selecting appropriate variant calling based on capture method, and will aid laboratories in selecting their preferred approach. PMID:26110913

  6. High-throughput amplicon sequencing reveals distinct communities within a corroding concrete sewer system.

    Science.gov (United States)

    Cayford, Barry I; Dennis, Paul G; Keller, Jurg; Tyson, Gene W; Bond, Philip L

    2012-10-01

    Microbially induced concrete corrosion (MICC) is an important problem in sewers. Here, small-subunit (SSU) rRNA gene amplicon pyrosequencing was used to characterize MICC communities. Microbial community composition differed between wall- and ceiling-associated MICC layers. Acidithiobacillus spp. were present at low abundances, and the communities were dominated by other sulfur-oxidizing-associated lineages.

  7. Bacterial metabarcoding by 16S rRNA gene ion torrent amplicon sequencing.

    Science.gov (United States)

    Fantini, Elio; Gianese, Giulio; Giuliano, Giovanni; Fiore, Alessia

    2015-01-01

    Ion Torrent is a next generation sequencing technology based on the detection of hydrogen ions produced during DNA chain elongation; this technology allows analyzing and characterizing genomes, genes, and species. Here, we describe an Ion Torrent procedure applied to the metagenomic analysis of 16S rRNA gene amplicons to study the bacterial diversity in food and environmental samples.

  8. Improved sensitivity of circulating tumor DNA measurement using short PCR amplicons

    DEFF Research Database (Denmark)

    Andersen, Rikke Fredslund; Spindler, Karen-Lise Garm; Brandslund, Ivan;

    2015-01-01

    , however, presents a number of challenges that require attention. The amount of DNA is low and highly fragmented and analyses need to be optimized accordingly. KRAS ARMS-qPCR assays with amplicon lengths of 120 and 85 base pairs, respectively, were compared using positive control material (PCR fragments...

  9. Genomic and expression array profiling of chromosome 20q amplicon in human colon cancer cells

    Directory of Open Access Journals (Sweden)

    Carter Jennifer

    2005-01-01

    Full Text Available Background: Gain of the q arm of chromosome 20 in human colorectal cancer has been associated with poorer survival time and has been reported to increase in frequency from adenomas to metastasis. The increasing frequency of chromosome 20q amplification during colorectal cancer progression and the presence of this amplification in carcinomas of other tissue origin has lead us to hypothesize that 20q11-13 harbors one or more genes which, when over expressed promote tumor invasion and metastasis. Aims: Generate genomic and expression profiles of the 20q amplicon in human cancer cell lines in order to identify genes with increased copy number and expression. Materials and Methods: Utilizing genomic sequencing clones and amplification mapping data from our lab and other previous studies, BAC/ PAC tiling paths spanning the 20q amplicon and genomic microarrays were generated. Array-CGH on the custom array with human cancer cell line DNAs was performed to generate genomic profiles of the amplicon. Expression array analysis with RNA from these cell lines using commercial oligo microarrays generated expression profiles of the amplicon. The data were then combined in order to identify genes with increased copy number and expression. Results: Over expressed genes in regions of increased copy number were identified and a list of potential novel genetic tumor markers was assembled based on biological functions of these genes Conclusions: Performing high-resolution genomic microarray profiling in conjunction with expression analysis is an effective approach to identify potential tumor markers.

  10. Comparison of normalization methods for construction of large, multiplex amplicon pools for next-generation sequencing.

    Science.gov (United States)

    Harris, J Kirk; Sahl, Jason W; Castoe, Todd A; Wagner, Brandie D; Pollock, David D; Spear, John R

    2010-06-01

    Constructing mixtures of tagged or bar-coded DNAs for sequencing is an important requirement for the efficient use of next-generation sequencers in applications where limited sequence data are required per sample. There are many applications in which next-generation sequencing can be used effectively to sequence large mixed samples; an example is the characterization of microbial communities where management costs associated with construction of multiplex pools are also scalable. One critical step in multiplex pool construction is the normalization process, whereby equimolar amounts of each amplicon are mixed. Here we compare three approaches (spectroscopy, size-restricted spectroscopy, and quantitative binding) for normalization of large, multiplex amplicon pools for performance and efficiency. We found that the quantitative binding approach was superior and represents an efficient scalable process for construction of very large, multiplex pools with hundreds and perhaps thousands of individual amplicons included. We demonstrate the increased sequence diversity identified with higher throughput. Massively parallel sequencing can dramatically accelerate microbial ecology studies by allowing appropriate replication of sequence acquisition to account for temporal and spatial variations. Further, population studies to examine genetic variation, which require even lower levels of sequencing, should be possible where thousands of individual bar-coded amplicons are examined in parallel. PMID:20418443

  11. Using expected sequence features to improve basecalling accuracy of amplicon pyrosequencing data

    DEFF Research Database (Denmark)

    Rask, Thomas Salhøj; Petersen, Bent; Chen, Donald S.;

    2016-01-01

    insertions and deletions, are on the other hand likely to disrupt open reading frames. Such an inverse relationship between errors and expectation based on prior knowledge can be used advantageously to guide the process known as basecalling, i.e. the inference of nucleotide sequence from raw sequencing data......Amplicon pyrosequencing targets a known genetic region and thus inherently produces reads highly anticipated to have certain features, such as conserved nucleotide sequence, and in the case of protein coding DNA, an open reading frame. Pyrosequencing errors, consisting mainly of nucleotide....... This probabilistic approach enables integration of basecalling into a larger model where other parameters can be incorporated, such as the likelihood for observing a full-length open reading frame at the targeted region. We apply the method to 454 amplicon pyrosequencing data obtained from a malaria...

  12. Nitrogenase gene amplicons from global marine surface waters are dominated by genes of non-cyanobacteria

    DEFF Research Database (Denmark)

    Farnelid, Hanna; Andersson, Anders F.; Bertilsson, Stefan;

    2011-01-01

    analysis of 79,090 nitrogenase (nifH) PCR amplicons encoding 7,468 unique proteins from surface samples (ten DNA samples and two RNA samples) collected at ten marine locations world-wide provides the first in-depth survey of a functional bacterial gene and yield insights into the composition and diversity...... of the nifH gene pool in marine waters. Great divergence in nifH composition was observed between sites. Cyanobacteria-like genes were most frequent among amplicons from the warmest waters, but overall the data set was dominated by nifH sequences most closely related to non-cyanobacteria. Clusters related...... to Alpha-, Beta-, Gamma-, and Delta-Proteobacteria were most common and showed distinct geographic distributions. Sequences related to anaerobic bacteria (nifH Cluster III) were generally rare, but preponderant in cold waters, especially in the Arctic. Although the two transcript samples were dominated...

  13. One-way sequencing of multiple amplicons from tandem repetitive mitochondrial DNA control region.

    Science.gov (United States)

    Xu, Jiawu; Fonseca, Dina M

    2011-10-01

    Repetitive DNA sequences not only exist abundantly in eukaryotic nuclear genomes, but also occur as tandem repeats in many animal mitochondrial DNA (mtDNA) control regions. Due to concerted evolution, these repetitive sequences are highly similar or even identical within a genome. When long repetitive regions are the targets of amplification for the purpose of sequencing, multiple amplicons may result if one primer has to be located inside the repeats. Here, we show that, without separating these amplicons by gel purification or cloning, directly sequencing the mitochondrial repeats with the primer outside repetitive region is feasible and efficient. We exemplify it by sequencing the mtDNA control region of the mosquito Aedes albopictus, which harbors typical large tandem DNA repeats. This one-way sequencing strategy is optimal for population surveys.

  14. Surface density dependence of PCR amplicon hybridization on PNA/DNA probe layers

    DEFF Research Database (Denmark)

    Yao, Danfeng; Kim, Junyoung; Yu, Fang;

    2005-01-01

    Surface plasmon field-enhanced fluorescence spectroscopy was employed to extensively investigate the hybridization behaviors of polymerase chain reaction (PCR) amplicons on a peptide nucleic acid (PNA) or DNA probe layer that was previously attached on a streptavidin-modified gold surface via...... biotin/streptavidin interaction. Despite the neutral backbone of PNA, the hybridization reactions were strongly influenced by the variation of ionic strength. The association rates exhibited a monotonic decrease with ionic strength increase and the maximum hybridization signal was achieved...... at an intermediate sodium concentration (approximately 100 mM). These effects were mainly ascribed to the electrostatic cross talk among the hybridized DNA molecules and the secondary structure of PCR amplicons. For the negatively charged DNA probes, the hybridization reaction was subjected additionally to the DNA/DNA...

  15. Fast, accurate and easy-to-pipeline methods for amplicon sequence processing

    Science.gov (United States)

    Antonielli, Livio; Sessitsch, Angela

    2016-04-01

    Next generation sequencing (NGS) technologies established since years as an essential resource in microbiology. While on the one hand metagenomic studies can benefit from the continuously increasing throughput of the Illumina (Solexa) technology, on the other hand the spreading of third generation sequencing technologies (PacBio, Oxford Nanopore) are getting whole genome sequencing beyond the assembly of fragmented draft genomes, making it now possible to finish bacterial genomes even without short read correction. Besides (meta)genomic analysis next-gen amplicon sequencing is still fundamental for microbial studies. Amplicon sequencing of the 16S rRNA gene and ITS (Internal Transcribed Spacer) remains a well-established widespread method for a multitude of different purposes concerning the identification and comparison of archaeal/bacterial (16S rRNA gene) and fungal (ITS) communities occurring in diverse environments. Numerous different pipelines have been developed in order to process NGS-derived amplicon sequences, among which Mothur, QIIME and USEARCH are the most well-known and cited ones. The entire process from initial raw sequence data through read error correction, paired-end read assembly, primer stripping, quality filtering, clustering, OTU taxonomic classification and BIOM table rarefaction as well as alternative "normalization" methods will be addressed. An effective and accurate strategy will be presented using the state-of-the-art bioinformatic tools and the example of a straightforward one-script pipeline for 16S rRNA gene or ITS MiSeq amplicon sequencing will be provided. Finally, instructions on how to automatically retrieve nucleotide sequences from NCBI and therefore apply the pipeline to targets other than 16S rRNA gene (Greengenes, SILVA) and ITS (UNITE) will be discussed.

  16. Massively parallel sequencing of 17 commonly used forensic autosomal STRs and amelogenin with small amplicons.

    Science.gov (United States)

    Kim, Eun Hye; Lee, Hwan Young; Yang, In Seok; Jung, Sang-Eun; Yang, Woo Ick; Shin, Kyoung-Jin

    2016-05-01

    The next-generation sequencing (NGS) method has been utilized to analyze short tandem repeat (STR) markers, which are routinely used for human identification purposes in the forensic field. Some researchers have demonstrated the successful application of the NGS system to STR typing, suggesting that NGS technology may be an alternative or additional method to overcome limitations of capillary electrophoresis (CE)-based STR profiling. However, there has been no available multiplex PCR system that is optimized for NGS analysis of forensic STR markers. Thus, we constructed a multiplex PCR system for the NGS analysis of 18 markers (13CODIS STRs, D2S1338, D19S433, Penta D, Penta E and amelogenin) by designing amplicons in the size range of 77-210 base pairs. Then, PCR products were generated from two single-sources, mixed samples and artificially degraded DNA samples using a multiplex PCR system, and were prepared for sequencing on the MiSeq system through construction of a subsequent barcoded library. By performing NGS and analyzing the data, we confirmed that the resultant STR genotypes were consistent with those of CE-based typing. Moreover, sequence variations were detected in targeted STR regions. Through the use of small-sized amplicons, the developed multiplex PCR system enables researchers to obtain successful STR profiles even from artificially degraded DNA as well as STR loci which are analyzed with large-sized amplicons in the CE-based commercial kits. In addition, successful profiles can be obtained from mixtures up to a 1:19 ratio. Consequently, the developed multiplex PCR system, which produces small size amplicons, can be successfully applied to STR NGS analysis of forensic casework samples such as mixtures and degraded DNA samples. PMID:26799314

  17. From Benchtop to Desktop: Important Considerations when Designing Amplicon Sequencing Workflows

    OpenAIRE

    Murray, Dáithí C; Megan L Coghlan; Michael Bunce

    2015-01-01

    Amplicon sequencing has been the method of choice in many high-throughput DNA sequencing (HTS) applications. To date there has been a heavy focus on the means by which to analyse the burgeoning amount of data afforded by HTS. In contrast, there has been a distinct lack of attention paid to considerations surrounding the importance of sample preparation and the fidelity of library generation. No amount of high-end bioinformatics can compensate for poorly prepared samples and it is therefore im...

  18. Potential of pmoA Amplicon Pyrosequencing for Methanotroph Diversity Studies ▿†

    OpenAIRE

    Lüke, Claudia; Frenzel, Peter

    2011-01-01

    We analyzed the potential of pmoA amplicon pyrosequencing compared to that of Sanger sequencing with paddy soils as a model environment. We defined operational taxonomic unit (OTU) cutoff values of 7% and 18%, reflecting methanotrophic species and major phylogenetic pmoA lineages, respectively. Major lineages were already well covered by clone libraries; nevertheless, pyrosequencing provided a higher level of diversity at the species level.

  19. Characterization of FGFR1 Locus in sqNSCLC Reveals a Broad and Heterogeneous Amplicon.

    Science.gov (United States)

    Rooney, Claire; Geh, Catherine; Williams, Victoria; Heuckmann, Johannes M; Menon, Roopika; Schneider, Petra; Al-Kadhimi, Katherine; Dymond, Michael; Smith, Neil R; Baker, Dawn; French, Tim; Smith, Paul D; Harrington, Elizabeth A; Barrett, J Carl; Kilgour, Elaine

    2016-01-01

    FGFR1 amplification occurs in ~20% of sqNSCLC and trials with FGFR inhibitors have selected FGFR1 amplified patients by FISH. Lung cancer cell lines were profiled for sensitivity to AZD4547, a potent, selective inhibitor of FGFRs 1-3. Sensitivity to FGFR inhibition was associated with but not wholly predicted by increased FGFR1 gene copy number. Additional biomarker assays evaluating expression of FGFRs and correlation between amplification and expression in clinical tissues are therefore warranted. We validated nanoString for mRNA expression analysis of 194 genes, including FGFRs, from clinical tumour tissue. In a panel of sqNSCLC tumours 14.4% (13/90) were FGFR1 amplified by FISH. Although mean FGFR1 expression was significantly higher in amplified samples, there was significant overlap in the range of expression levels between the amplified and non-amplified cohorts with several non-amplified samples expressing FGFR1 to levels equivalent to amplified samples. Statistical analysis revealed increased expression of FGFR1 neighboring genes on the 8p12 amplicon (BAG4, LSM1 and WHSC1L1) in FGFR1 amplified tumours, suggesting a broad rather than focal amplicon and raises the potential for codependencies. High resolution aCGH analysis of pre-clinical and clinical samples supported the presence of a broad and heterogeneous amplicon around the FGFR1 locus. In conclusion, the range of FGFR1 expression levels in both FGFR1 amplified and non-amplified NSCLC tissues, together with the breadth and intra-patient heterogeneity of the 8p amplicon highlights the need for gene expression analysis of clinical samples to inform the understanding of determinants of response to FGFR inhibitors. In this respect the nanoString platform provides an attractive option for RNA analysis of FFPE clinical samples. PMID:26905262

  20. Characterization of FGFR1 Locus in sqNSCLC Reveals a Broad and Heterogeneous Amplicon.

    Directory of Open Access Journals (Sweden)

    Claire Rooney

    Full Text Available FGFR1 amplification occurs in ~20% of sqNSCLC and trials with FGFR inhibitors have selected FGFR1 amplified patients by FISH. Lung cancer cell lines were profiled for sensitivity to AZD4547, a potent, selective inhibitor of FGFRs 1-3. Sensitivity to FGFR inhibition was associated with but not wholly predicted by increased FGFR1 gene copy number. Additional biomarker assays evaluating expression of FGFRs and correlation between amplification and expression in clinical tissues are therefore warranted. We validated nanoString for mRNA expression analysis of 194 genes, including FGFRs, from clinical tumour tissue. In a panel of sqNSCLC tumours 14.4% (13/90 were FGFR1 amplified by FISH. Although mean FGFR1 expression was significantly higher in amplified samples, there was significant overlap in the range of expression levels between the amplified and non-amplified cohorts with several non-amplified samples expressing FGFR1 to levels equivalent to amplified samples. Statistical analysis revealed increased expression of FGFR1 neighboring genes on the 8p12 amplicon (BAG4, LSM1 and WHSC1L1 in FGFR1 amplified tumours, suggesting a broad rather than focal amplicon and raises the potential for codependencies. High resolution aCGH analysis of pre-clinical and clinical samples supported the presence of a broad and heterogeneous amplicon around the FGFR1 locus. In conclusion, the range of FGFR1 expression levels in both FGFR1 amplified and non-amplified NSCLC tissues, together with the breadth and intra-patient heterogeneity of the 8p amplicon highlights the need for gene expression analysis of clinical samples to inform the understanding of determinants of response to FGFR inhibitors. In this respect the nanoString platform provides an attractive option for RNA analysis of FFPE clinical samples.

  1. Single genome amplification and direct amplicon sequencing of Plasmodium spp. DNA from ape fecal specimens

    OpenAIRE

    Liu, Weimin; Li, Yingying; Peeters, Martine; Rayner, Julian; Sharp, Paul; Shaw, George; Hahn, Beatrice

    2010-01-01

    Conventional PCR followed by molecular cloning and sequencing of amplified products is commonly used to test clinical specimens for target sequences of interest, such as viral, bacterial or parasite nucleic acids. However, this approach has serious limitations when used to analyze mixtures of genetically divergent templates1–9. This is because Taq polymerase is prone to switch templates during the amplification process, thereby generating recombinants that do not exist in vivo4. When amplicon...

  2. From benchtop to desktop: important considerations when designing amplicon sequencing workflows.

    Directory of Open Access Journals (Sweden)

    Dáithí C Murray

    Full Text Available Amplicon sequencing has been the method of choice in many high-throughput DNA sequencing (HTS applications. To date there has been a heavy focus on the means by which to analyse the burgeoning amount of data afforded by HTS. In contrast, there has been a distinct lack of attention paid to considerations surrounding the importance of sample preparation and the fidelity of library generation. No amount of high-end bioinformatics can compensate for poorly prepared samples and it is therefore imperative that careful attention is given to sample preparation and library generation within workflows, especially those involving multiple PCR steps. This paper redresses this imbalance by focusing on aspects pertaining to the benchtop within typical amplicon workflows: sample screening, the target region, and library generation. Empirical data is provided to illustrate the scope of the problem. Lastly, the impact of various data analysis parameters is also investigated in the context of how the data was initially generated. It is hoped this paper may serve to highlight the importance of pre-analysis workflows in achieving meaningful, future-proof data that can be analysed appropriately. As amplicon sequencing gains traction in a variety of diagnostic applications from forensics to environmental DNA (eDNA it is paramount workflows and analytics are both fit for purpose.

  3. Electrochemical DNA sensor for anthrax toxin activator gene atxA-detection of PCR amplicons.

    Science.gov (United States)

    Das, Ritu; Goel, Ajay K; Sharma, Mukesh K; Upadhyay, Sanjay

    2015-12-15

    We report the DNA probe functionalized electrochemical genosensor for the detection of Bacillus anthracis, specific towards the regulatory gene atxA. The DNA sensor is fabricated on electrochemically deposited gold nanoparticle on self assembled layer of (3-Mercaptopropyl) trimethoxysilane (MPTS) on GC electrode. DNA hybridization is monitored by differential pulse voltammogram (DPV). The modified GC electrode is characterized by atomic force microscopy (AFM), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS) method. We also quantified the DNA probe density on electrode surface by the chronocoulometric method. The detection is specific and selective for atxA gene by DNA probe on the electrode surface. No report is available for the detection of B. anthracis by using atxA an anthrax toxin activator gene. In the light of real and complex sample, we have studied the PCR amplicons of 303, 361 and 568 base pairs by using symmetric and asymmetric PCR approaches. The DNA probe of atxA gene efficiently hybridizes with different base pairs of PCR amplicons. The detection limit is found to be 1.0 pM (S/N ratio=3). The results indicate that the DNA sensor is able to detect synthetic target as well as PCR amplicons of different base pairs. PMID:26257186

  4. Impact of Mutation Type and Amplicon Characteristics on Genetic Diversity Measures Generated Using a High-Resolution Melting Diversity Assay

    OpenAIRE

    Cousins, Matthew M.; Donnell, Deborah; Eshleman, Susan H.

    2013-01-01

    We adapted high-resolution melting (HRM) technology to measure genetic diversity without sequencing. Diversity is measured as a single numeric HRM score. Herein, we determined the impact of mutation types and amplicon characteristics on HRM diversity scores. Plasmids were generated with single-base changes, insertions, and deletions. Different primer sets were used to vary the position of mutations within amplicons. Plasmids and plasmid mixtures were analyzed to determine the impact of mutati...

  5. Clinical impact of targeted amplicon sequencing for meningioma as a practical clinical-sequencing system.

    Science.gov (United States)

    Yuzawa, Sayaka; Nishihara, Hiroshi; Yamaguchi, Shigeru; Mohri, Hiromi; Wang, Lei; Kimura, Taichi; Tsuda, Masumi; Tanino, Mishie; Kobayashi, Hiroyuki; Terasaka, Shunsuke; Houkin, Kiyohiro; Sato, Norihiro; Tanaka, Shinya

    2016-07-01

    Recent genetic analyses using next-generation sequencers have revealed numerous genetic alterations in various tumors including meningioma, which is the most common primary brain tumor. However, their use as routine laboratory examinations in clinical applications for tumor genotyping is not cost effective. To establish a clinical sequencing system for meningioma and investigate the clinical significance of genotype, we retrospectively performed targeted amplicon sequencing on 103 meningiomas and evaluated the association with clinicopathological features. We designed amplicon-sequencing panels targeting eight genes including NF2 (neurofibromin 2), TRAF7, KLF4, AKT1, and SMO. Libraries prepared with genomic DNA extracted from PAXgene-fixed paraffin-embedded tissues of 103 meningioma specimens were sequenced using the Illumina MiSeq. NF2 loss in some cases was also confirmed by interphase-fluorescent in situ hybridization. We identified NF2 loss and/or at least one mutation in NF2, TRAF7, KLF4, AKT1, and SMO in 81 out of 103 cases (79%) by targeted amplicon sequencing. On the basis of genetic status, we categorized meningiomas into three genotype groups: NF2 type, TRAKLS type harboring mutation in TRAF7, AKT1, KLF4, and/or SMO, and 'not otherwise classified' type. Genotype significantly correlated with tumor volume, tumor location, and magnetic resonance imaging findings such as adjacent bone change and heterogeneous gadolinium enhancement, as well as histopathological subtypes. In addition, multivariate analysis revealed that genotype was independently associated with risk of recurrence. In conclusion, we established a rapid clinical sequencing system that enables final confirmation of meningioma genotype within 7 days turnaround time. Our method will bring multiple benefits to neuropathologists and neurosurgeons for accurate diagnosis and appropriate postoperative management. PMID:27102344

  6. Shedding Light on the Microbial Community of the Macropod Foregut Using 454-Amplicon Pyrosequencing

    OpenAIRE

    Gulino, Lisa-Maree; Ouwerkerk, Diane; Alicia Y H Kang; Maguire, Anita J.; Kienzle, Marco; Klieve, Athol V.

    2013-01-01

    Twenty macropods from five locations in Queensland, Australia, grazing on a variety of native pastures were surveyed and the bacterial community of the foregut was examined using 454-amplicon pyrosequencing. Specifically, the V3/V4 region of 16S rRNA gene was examined. A total of 5040 OTUs were identified in the data set (post filtering). Thirty-two OTUs were identified as ‘shared’ OTUS (i.e. present in all samples) belonging to either Firmicutes or Bacteroidetes (Clostridiales/Bacteroidales)...

  7. Analysis and Visualization Tool for Targeted Amplicon Bisulfite Sequencing on Ion Torrent Sequencers.

    Science.gov (United States)

    Pabinger, Stephan; Ernst, Karina; Pulverer, Walter; Kallmeyer, Rainer; Valdes, Ana M; Metrustry, Sarah; Katic, Denis; Nuzzo, Angelo; Kriegner, Albert; Vierlinger, Klemens; Weinhaeusel, Andreas

    2016-01-01

    Targeted sequencing of PCR amplicons generated from bisulfite deaminated DNA is a flexible, cost-effective way to study methylation of a sample at single CpG resolution and perform subsequent multi-target, multi-sample comparisons. Currently, no platform specific protocol, support, or analysis solution is provided to perform targeted bisulfite sequencing on a Personal Genome Machine (PGM). Here, we present a novel tool, called TABSAT, for analyzing targeted bisulfite sequencing data generated on Ion Torrent sequencers. The workflow starts with raw sequencing data, performs quality assessment, and uses a tailored version of Bismark to map the reads to a reference genome. The pipeline visualizes results as lollipop plots and is able to deduce specific methylation-patterns present in a sample. The obtained profiles are then summarized and compared between samples. In order to assess the performance of the targeted bisulfite sequencing workflow, 48 samples were used to generate 53 different Bisulfite-Sequencing PCR amplicons from each sample, resulting in 2,544 amplicon targets. We obtained a mean coverage of 282X using 1,196,822 aligned reads. Next, we compared the sequencing results of these targets to the methylation level of the corresponding sites on an Illumina 450k methylation chip. The calculated average Pearson correlation coefficient of 0.91 confirms the sequencing results with one of the industry-leading CpG methylation platforms and shows that targeted amplicon bisulfite sequencing provides an accurate and cost-efficient method for DNA methylation studies, e.g., to provide platform-independent confirmation of Illumina Infinium 450k methylation data. TABSAT offers a novel way to analyze data generated by Ion Torrent instruments and can also be used with data from the Illumina MiSeq platform. It can be easily accessed via the Platomics platform, which offers a web-based graphical user interface along with sample and parameter storage. TABSAT is freely

  8. Amplicon-based metagenomics identified candidate organisms in soils that caused yield decline in strawberry

    OpenAIRE

    Xiangming Xu; Thomas Passey; Feng Wei; Robert Saville; Harrison, Richard J.

    2015-01-01

    A phenomenon of yield decline due to weak plant growth in strawberry was recently observed in non-chemo-fumigated soils, which was not associated with the soil fungal pathogen Verticillium dahliae, the main target of fumigation. Amplicon-based metagenomics was used to profile soil microbiota in order to identify microbial organisms that may have caused the yield decline. A total of 36 soil samples were obtained in 2013 and 2014 from four sites for metagenomic studies; two of the four sites ha...

  9. Simultaneous digital quantification and fluorescence-based size characterization of massively parallel sequencing libraries.

    Science.gov (United States)

    Laurie, Matthew T; Bertout, Jessica A; Taylor, Sean D; Burton, Joshua N; Shendure, Jay A; Bielas, Jason H

    2013-08-01

    Due to the high cost of failed runs and suboptimal data yields, quantification and determination of fragment size range are crucial steps in the library preparation process for massively parallel sequencing (or next-generation sequencing). Current library quality control methods commonly involve quantification using real-time quantitative PCR and size determination using gel or capillary electrophoresis. These methods are laborious and subject to a number of significant limitations that can make library calibration unreliable. Herein, we propose and test an alternative method for quality control of sequencing libraries using droplet digital PCR (ddPCR). By exploiting a correlation we have discovered between droplet fluorescence and amplicon size, we achieve the joint quantification and size determination of target DNA with a single ddPCR assay. We demonstrate the accuracy and precision of applying this method to the preparation of sequencing libraries.

  10. Analysis of the microbiome: Advantages of whole genome shotgun versus 16S amplicon sequencing.

    Science.gov (United States)

    Ranjan, Ravi; Rani, Asha; Metwally, Ahmed; McGee, Halvor S; Perkins, David L

    2016-01-22

    The human microbiome has emerged as a major player in regulating human health and disease. Translational studies of the microbiome have the potential to indicate clinical applications such as fecal transplants and probiotics. However, one major issue is accurate identification of microbes constituting the microbiota. Studies of the microbiome have frequently utilized sequencing of the conserved 16S ribosomal RNA (rRNA) gene. We present a comparative study of an alternative approach using whole genome shotgun sequencing (WGS). In the present study, we analyzed the human fecal microbiome compiling a total of 194.1 × 10(6) reads from a single sample using multiple sequencing methods and platforms. Specifically, after establishing the reproducibility of our methods with extensive multiplexing, we compared: 1) The 16S rRNA amplicon versus the WGS method, 2) the Illumina HiSeq versus MiSeq platforms, 3) the analysis of reads versus de novo assembled contigs, and 4) the effect of shorter versus longer reads. Our study demonstrates that whole genome shotgun sequencing has multiple advantages compared with the 16S amplicon method including enhanced detection of bacterial species, increased detection of diversity and increased prediction of genes. In addition, increased length, either due to longer reads or the assembly of contigs, improved the accuracy of species detection.

  11. Multiplex amplicon sequencing for microbe identification in community-based culture collections.

    Science.gov (United States)

    Armanhi, Jaderson Silveira Leite; de Souza, Rafael Soares Correa; de Araújo, Laura Migliorini; Okura, Vagner Katsumi; Mieczkowski, Piotr; Imperial, Juan; Arruda, Paulo

    2016-01-01

    Microbiome analysis using metagenomic sequencing has revealed a vast microbial diversity associated with plants. Identifying the molecular functions associated with microbiome-plant interaction is a significant challenge concerning the development of microbiome-derived technologies applied to agriculture. An alternative to accelerate the discovery of the microbiome benefits to plants is to construct microbial culture collections concomitant with accessing microbial community structure and abundance. However, traditional methods of isolation, cultivation, and identification of microbes are time-consuming and expensive. Here we describe a method for identification of microbes in culture collections constructed by picking colonies from primary platings that may contain single or multiple microorganisms, which we named community-based culture collections (CBC). A multiplexing 16S rRNA gene amplicon sequencing based on two-step PCR amplifications with tagged primers for plates, rows, and columns allowed the identification of the microbial composition regardless if the well contains single or multiple microorganisms. The multiplexing system enables pooling amplicons into a single tube. The sequencing performed on the PacBio platform led to recovery near-full-length 16S rRNA gene sequences allowing accurate identification of microorganism composition in each plate well. Cross-referencing with plant microbiome structure and abundance allowed the estimation of diversity and abundance representation of microorganism in the CBC. PMID:27404280

  12. AMPLISAS: a web server for multilocus genotyping using next-generation amplicon sequencing data.

    Science.gov (United States)

    Sebastian, Alvaro; Herdegen, Magdalena; Migalska, Magdalena; Radwan, Jacek

    2016-03-01

    Next-generation sequencing (NGS) technologies are revolutionizing the fields of biology and medicine as powerful tools for amplicon sequencing (AS). Using combinations of primers and barcodes, it is possible to sequence targeted genomic regions with deep coverage for hundreds, even thousands, of individuals in a single experiment. This is extremely valuable for the genotyping of gene families in which locus-specific primers are often difficult to design, such as the major histocompatibility complex (MHC). The utility of AS is, however, limited by the high intrinsic sequencing error rates of NGS technologies and other sources of error such as polymerase amplification or chimera formation. Correcting these errors requires extensive bioinformatic post-processing of NGS data. Amplicon Sequence Assignment (AMPLISAS) is a tool that performs analysis of AS results in a simple and efficient way, while offering customization options for advanced users. AMPLISAS is designed as a three-step pipeline consisting of (i) read demultiplexing, (ii) unique sequence clustering and (iii) erroneous sequence filtering. Allele sequences and frequencies are retrieved in excel spreadsheet format, making them easy to interpret. AMPLISAS performance has been successfully benchmarked against previously published genotyped MHC data sets obtained with various NGS technologies.

  13. Amplicon-based metagenomics identified candidate organisms in soils that caused yield decline in strawberry.

    Science.gov (United States)

    Xu, Xiangming; Passey, Thomas; Wei, Feng; Saville, Robert; Harrison, Richard J

    2015-01-01

    A phenomenon of yield decline due to weak plant growth in strawberry was recently observed in non-chemo-fumigated soils, which was not associated with the soil fungal pathogen Verticillium dahliae, the main target of fumigation. Amplicon-based metagenomics was used to profile soil microbiota in order to identify microbial organisms that may have caused the yield decline. A total of 36 soil samples were obtained in 2013 and 2014 from four sites for metagenomic studies; two of the four sites had a yield-decline problem, the other two did not. More than 2000 fungal or bacterial operational taxonomy units (OTUs) were found in these samples. Relative abundance of individual OTUs was statistically compared for differences between samples from sites with or without yield decline. A total of 721 individual comparisons were statistically significant - involving 366 unique bacterial and 44 unique fungal OTUs. Based on further selection criteria, we focused on 34 bacterial and 17 fungal OTUs and found that yield decline resulted probably from one or more of the following four factors: (1) low abundance of Bacillus and Pseudomonas populations, which are well known for their ability of supressing pathogen development and/or promoting plant growth; (2) lack of the nematophagous fungus (Paecilomyces species); (3) a high level of two non-specific fungal root rot pathogens; and (4) wet soil conditions. This study demonstrated the usefulness of an amplicon-based metagenomics approach to profile soil microbiota and to detect differential abundance in microbes. PMID:26504572

  14. AMPLISAS: a web server for multilocus genotyping using next-generation amplicon sequencing data.

    Science.gov (United States)

    Sebastian, Alvaro; Herdegen, Magdalena; Migalska, Magdalena; Radwan, Jacek

    2016-03-01

    Next-generation sequencing (NGS) technologies are revolutionizing the fields of biology and medicine as powerful tools for amplicon sequencing (AS). Using combinations of primers and barcodes, it is possible to sequence targeted genomic regions with deep coverage for hundreds, even thousands, of individuals in a single experiment. This is extremely valuable for the genotyping of gene families in which locus-specific primers are often difficult to design, such as the major histocompatibility complex (MHC). The utility of AS is, however, limited by the high intrinsic sequencing error rates of NGS technologies and other sources of error such as polymerase amplification or chimera formation. Correcting these errors requires extensive bioinformatic post-processing of NGS data. Amplicon Sequence Assignment (AMPLISAS) is a tool that performs analysis of AS results in a simple and efficient way, while offering customization options for advanced users. AMPLISAS is designed as a three-step pipeline consisting of (i) read demultiplexing, (ii) unique sequence clustering and (iii) erroneous sequence filtering. Allele sequences and frequencies are retrieved in excel spreadsheet format, making them easy to interpret. AMPLISAS performance has been successfully benchmarked against previously published genotyped MHC data sets obtained with various NGS technologies. PMID:26257385

  15. ICO amplicon NGS data analysis: a Web tool for variant detection in common high-risk hereditary cancer genes analyzed by amplicon GS Junior next-generation sequencing.

    Science.gov (United States)

    Lopez-Doriga, Adriana; Feliubadaló, Lídia; Menéndez, Mireia; Lopez-Doriga, Sergio; Morón-Duran, Francisco D; del Valle, Jesús; Tornero, Eva; Montes, Eva; Cuesta, Raquel; Campos, Olga; Gómez, Carolina; Pineda, Marta; González, Sara; Moreno, Victor; Capellá, Gabriel; Lázaro, Conxi

    2014-03-01

    Next-generation sequencing (NGS) has revolutionized genomic research and is set to have a major impact on genetic diagnostics thanks to the advent of benchtop sequencers and flexible kits for targeted libraries. Among the main hurdles in NGS are the difficulty of performing bioinformatic analysis of the huge volume of data generated and the high number of false positive calls that could be obtained, depending on the NGS technology and the analysis pipeline. Here, we present the development of a free and user-friendly Web data analysis tool that detects and filters sequence variants, provides coverage information, and allows the user to customize some basic parameters. The tool has been developed to provide accurate genetic analysis of targeted sequencing of common high-risk hereditary cancer genes using amplicon libraries run in a GS Junior System. The Web resource is linked to our own mutation database, to assist in the clinical classification of identified variants. We believe that this tool will greatly facilitate the use of the NGS approach in routine laboratories.

  16. Parallel tagged amplicon sequencing of relatively long PCR products using the Illumina HiSeq platform and transcriptome assembly.

    Science.gov (United States)

    Feng, Yan-Jie; Liu, Qing-Feng; Chen, Meng-Yun; Liang, Dan; Zhang, Peng

    2016-01-01

    In phylogenetics and population genetics, a large number of loci are often needed to accurately resolve species relationships. Normally, loci are enriched by PCR and sequenced by Sanger sequencing, which is expensive when the number of amplicons is large. Next-generation sequencing (NGS) techniques are increasingly used for parallel amplicon sequencing, which reduces sequencing costs tremendously, but has not reduced preparation costs very much. Moreover, for most current NGS methods, amplicons need to be purified and quantified before sequencing and their lengths are also restricted (normally HiSeq paired-end 90-bp data. Overall, we validate a rapid, cost-effective and scalable approach to sequence a large number of targeted loci from a large number of samples that is particularly suitable for both phylogenetics and population genetics studies that require a modest scale of data.

  17. Amplicon restriction patterns associated with nitrogenase activity of root nodules for selection of superior Myrica seedlings

    Indian Academy of Sciences (India)

    Mhathung Yanthan; Arvind K Misran

    2013-11-01

    Trees of Myrica sp. grow abundantly in the forests of Meghalaya, India. These trees are actinorhizal and harbour nitrogen-fixing Frankia in their root nodules and contribute positively towards the enhancement of nitrogen status of forest areas. They can be used in rejuvenation of mine spoils and nitrogen-depleted fallow lands generated due to slash and burn agriculture practiced in the area. We have studied the association of amplicon restriction patterns (ARPs) of Myrica ribosomal RNA gene and internal transcribed spacer (ITS) region and nitrogenase activity of its root nodules. We found that ARPs thus obtained could be used as markers for early screening of seedlings that could support strains of Frankia that fix atmospheric nitrogen more efficiently.

  18. 16S rRNA amplicon sequencing dataset for conventionalized and conventionally raised zebrafish larvae.

    Science.gov (United States)

    Davis, Daniel J; Bryda, Elizabeth C; Gillespie, Catherine H; Ericsson, Aaron C

    2016-09-01

    Data presented here contains metagenomic analysis regarding the sequential conventionalization of germ-free zebrafish embryos. Zebrafish embryos that underwent a germ-free sterilization process immediately after fertilization were promptly exposed to and raised to larval stage in conventional fish water. At 6 days postfertilization (dpf), these "conventionalized" larvae were compared to zebrafish larvae that were raised in conventional fish water never undergoing the initial sterilization process. Bacterial 16S rRNA amplicon sequencing was performed on DNA isolated from homogenates of the larvae revealing distinct microbiota variations between the two groups. The dataset described here is also related to the research article entitled "Microbial modulation of behavior and stress responses in zebrafish larvae" (Davis et al., 2016) [1]. PMID:27508247

  19. Rare amplicons implicate frequent deregulation of cell fate specification pathways in oral squamous cell carcinoma.

    Science.gov (United States)

    Snijders, Antoine M; Schmidt, Brian L; Fridlyand, Jane; Dekker, Nusi; Pinkel, Daniel; Jordan, Richard C K; Albertson, Donna G

    2005-06-16

    Genomes of solid tumors are characterized by gains and losses of regions, which may contribute to tumorigenesis by altering gene expression. Often the aberrations are extensive, encompassing whole chromosome arms, which makes identification of candidate genes in these regions difficult. Here, we focused on narrow regions of gene amplification to facilitate identification of genetic pathways important in oral squamous cell carcinoma (SCC) development. We used array comparative genomic hybridization (array CGH) to define minimum common amplified regions and then used expression analysis to identify candidate driver genes in amplicons that spanned LAMA3, MMP7), as well as members of the hedgehog (GLI2) and notch (JAG1, RBPSUH, FJX1) pathways to be amplified and overexpressed. Deregulation of these and other members of the hedgehog and notch pathways (HHIP, SMO, DLL1, NOTCH4) implicates deregulation of developmental and differentiation pathways, cell fate misspecification, in oral SCC development. PMID:15824737

  20. Amplicon restriction patterns associated with nitrogenase activity of root nodules for selection of superior Myrica seedlings.

    Science.gov (United States)

    Yanthan, Mhathung; Misra, Arvind K

    2013-11-01

    Trees of Myrica sp. grow abundantly in the forests of Meghalaya, India. These trees are actinorhizal and harbour nitrogen-fixing Frankia in their root nodules and contribute positively towards the enhancement of nitrogen status of forest areas. They can be used in rejuvenation of mine spoils and nitrogen-depleted fallow lands generated due to slash and burn agriculture practiced in the area. We have studied the association of amplicon restriction patterns (ARPs) of Myrica ribosomal RNA gene and internal transcribed spacer (ITS) region and nitrogenase activity of its root nodules. We found that ARPs thus obtained could be used as markers for early screening of seedlings that could support strains of Frankia that fix atmospheric nitrogen more efficiently. PMID:24287658

  1. Similar diversity of Alphaproteobacteria and nitrogenase gene amplicons on two related Sphagnum mosses

    Directory of Open Access Journals (Sweden)

    Anastasia eBragina

    2012-01-01

    Full Text Available Sphagnum mosses represent a main component in ombrotrophic wetlands. They harbor a specific and diverse microbial community with essential functions for the host. To understand extend and degree of host specificity, Sphagnum fallax and S. angustifolium, two phylogenetically closely related species, which show distinct habitat preference with respect to the nutrient level, were analyzed by a multifaceted approach. Microbial fingerprints obtained by PCR-SSCP (single-strand conformation polymorphism using universal, group-specific and functional primers were highly similar. Similarity was confirmed for colonization patterns obtained by fluorescence in situ hybridization (FISH coupled with confocal laser scanning microscopy (CLSM: Alphaproteobacteria were the main colonizers inside the hyaline cells of Sphagnum leaves. A deeper survey of Alphaproteobacteria by 16S rRNA gene amplicon sequencing reveals a high diversity with Acidocella, Acidisphaera, Rhodopila and Phenylobacterium as major genera for both mosses. Pathogen defense and nitrogen fixation are important functions of Sphagnum-associated bacteria, which are fulfilled by microbial communities of both Sphagna in a similar way. NifH libraries of Sphagnum-associated microbial communities were characterized by high diversity and abundance of Alphaproteobacteria but contained also diverse amplicons of other taxa, e.g. Cyanobacteria, Geobacter and Spirochaeta. Statistically significant differences between the microbial communities of both Sphagnum species could not be discovered in any of the experimental approach. Our results show that the same close relationship, which exists between the physical, morphological and chemical characteristics of Sphagnum mosses and the ecology and function of bog ecosystems, also connects moss plantlets with their associated bacterial communities.

  2. Long amplicon (LA)-qPCR for the discrimination of infectious and noninfectious phix174 bacteriophages after UV inactivation.

    Science.gov (United States)

    Ho, Johannes; Seidel, Michael; Niessner, Reinhard; Eggers, Jutta; Tiehm, Andreas

    2016-10-15

    Waterborne viruses are increasingly being considered in risk assessment schemes. In general, virus detection by culture methods is time consuming. In contrast, detection by quantitative polymerase chain reaction (qPCR) is more rapid and therefore, more suitable for monitoring. At present, qPCR lacks the essential ability for discriminating between infectious and non-infectious viruses, thus limiting its applicability for monitoring disinfection processes. In this study, a method was developed to quantify UV inactivation by long amplicon (LA)-qPCR. Bacteriophage phiX174 was used as a surrogate for human pathogenic viruses. A qPCR protocol was developed with new sets of primers, resulting in amplicon lengths of 108, 250, 456, 568, 955, 1063, 1544, and 1764 nucleotides. The log reduction of gene copies increased with increasing amplicon length. Additional treatment with the intercalating dye, PMA, had no effect, indicating that the bacteriophage capsids were not damaged by low pressure UV irradiation. A qPCR of nearly the complete genome (approx. 5000 nucleotides) showed similar results to the plaque assay. The log reduction in qPCR correlates with [specific amplicon length x UV dose]. The normalized DNA effect constant can be applied to calculate phiX174 inactivation based on qPCR detection.

  3. Next-generation sequencing of multiple individuals per barcoded library by deconvolution of sequenced amplicons using endonuclease fragment analysis

    DEFF Research Database (Denmark)

    Andersen, Jeppe D; Pereira, Vania; Pietroni, Carlotta;

    2014-01-01

    digestion of PCR amplicons prior to library preparation, creating a specific fragment pattern for each individual that can be resolved after sequencing. By using both barcodes and restriction fragment patterns, we demonstrate the ability to sequence the human melanocortin 1 receptor (MC1R) genes from 72...

  4. Introduction to uncertainty quantification

    CERN Document Server

    Sullivan, T J

    2015-01-01

    Uncertainty quantification is a topic of increasing practical importance at the intersection of applied mathematics, statistics, computation, and numerous application areas in science and engineering. This text provides a framework in which the main objectives of the field of uncertainty quantification are defined, and an overview of the range of mathematical methods by which they can be achieved. Complete with exercises throughout, the book will equip readers with both theoretical understanding and practical experience of the key mathematical and algorithmic tools underlying the treatment of uncertainty in modern applied mathematics. Students and readers alike are encouraged to apply the mathematical methods discussed in this book to their own favourite problems to understand their strengths and weaknesses, also making the text suitable as a self-study. This text is designed as an introduction to uncertainty quantification for senior undergraduate and graduate students with a mathematical or statistical back...

  5. Digital fragment analysis of short tandem repeats by high-throughput amplicon sequencing.

    Science.gov (United States)

    Darby, Brian J; Erickson, Shay F; Hervey, Samuel D; Ellis-Felege, Susan N

    2016-07-01

    High-throughput sequencing has been proposed as a method to genotype microsatellites and overcome the four main technical drawbacks of capillary electrophoresis: amplification artifacts, imprecise sizing, length homoplasy, and limited multiplex capability. The objective of this project was to test a high-throughput amplicon sequencing approach to fragment analysis of short tandem repeats and characterize its advantages and disadvantages against traditional capillary electrophoresis. We amplified and sequenced 12 muskrat microsatellite loci from 180 muskrat specimens and analyzed the sequencing data for precision of allele calling, propensity for amplification or sequencing artifacts, and for evidence of length homoplasy. Of the 294 total alleles, we detected by sequencing, only 164 alleles would have been detected by capillary electrophoresis as the remaining 130 alleles (44%) would have been hidden by length homoplasy. The ability to detect a greater number of unique alleles resulted in the ability to resolve greater population genetic structure. The primary advantages of fragment analysis by sequencing are the ability to precisely size fragments, resolve length homoplasy, multiplex many individuals and many loci into a single high-throughput run, and compare data across projects and across laboratories (present and future) with minimal technical calibration. A significant disadvantage of fragment analysis by sequencing is that the method is only practical and cost-effective when performed on batches of several hundred samples with multiple loci. Future work is needed to optimize throughput while minimizing costs and to update existing microsatellite allele calling and analysis programs to accommodate sequence-aware microsatellite data. PMID:27386092

  6. Extracellular DNA amplicon sequencing reveals high levels of benthic eukaryotic diversity in the central Red Sea.

    Science.gov (United States)

    Pearman, John K; Irigoien, Xabier; Carvalho, Susana

    2016-04-01

    The present study aims to characterize the benthic eukaryotic biodiversity patterns at a coarse taxonomic level in three areas of the central Red Sea (a lagoon, an offshore area in Thuwal and a shallow coastal area near Jeddah) based on extracellular DNA. High-throughput amplicon sequencing targeting the V9 region of the 18S rRNA gene was undertaken for 32 sediment samples. High levels of alpha-diversity were detected with 16,089 operational taxonomic units (OTUs) being identified. The majority of the OTUs were assigned to Metazoa (29.2%), Alveolata (22.4%) and Stramenopiles (17.8%). Stramenopiles (Diatomea) and Alveolata (Ciliophora) were frequent in a lagoon and in shallower coastal stations, whereas metazoans (Arthropoda: Maxillopoda) were dominant in deeper offshore stations. Only 24.6% of total OTUs were shared among all areas. Beta-diversity was generally lower between the lagoon and Jeddah (nearshore) than between either of those and the offshore area, suggesting a nearshore-offshore biodiversity gradient. The current approach allowed for a broad-range of benthic eukaryotic biodiversity to be analysed with significantly less labour than would be required by other traditional taxonomic approaches. Our findings suggest that next generation sequencing techniques have the potential to provide a fast and standardised screening of benthic biodiversity at large spatial and temporal scales.

  7. The development of reduced size STR amplicons as tools for analysis of degraded DNA.

    Science.gov (United States)

    Butler, John M; Shen, Yin; McCord, Bruce R

    2003-09-01

    New multiplex PCR sets of commonly used short tandem repeat (STR) markers have been developed to produce PCR products that are reduced in size when compared to standard commercial STR kits. The reduction in size of these amplicons can facilitate the examination and analysis of degraded DNA evidence by improving amplification efficiency. This "miniSTR" approach will permit current forensic practitioners to use STR markers and instrumentation already present in their laboratories and to generate genotyping data that is directly comparable to reference samples and searchable through the FBI's Combined DNA Index System (CODIS) databases. This paper discusses the development of these new primer sets and presents some initial results in the analysis of degraded and aged DNA samples. A method for removal of problematic fluorescent dye artifacts is also described. Comparison studies in over 100 samples have verified that these miniSTR primers can provide fully concordant results to commercial STR kits and can provide improved signal from degraded DNA specimens. These miniplex sets should prove valuable in the analysis of samples where allele dropout and reduced sensitivity of larger STR alleles occurs.

  8. Accuracy of the high-throughput amplicon sequencing to identify species within the genus Aspergillus.

    Science.gov (United States)

    Lee, Seungeun; Yamamoto, Naomichi

    2015-12-01

    This study characterized the accuracy of high-throughput amplicon sequencing to identify species within the genus Aspergillus. To this end, we sequenced the internal transcribed spacer 1 (ITS1), β-tubulin (BenA), and calmodulin (CaM) gene encoding sequences as DNA markers from eight reference Aspergillus strains with known identities using 300-bp sequencing on the Illumina MiSeq platform, and compared them with the BLASTn outputs. The identifications with the sequences longer than 250 bp were accurate at the section rank, with some ambiguities observed at the species rank due to mostly cross detection of sibling species. Additionally, in silico analysis was performed to predict the identification accuracy for all species in the genus Aspergillus, where 107, 210, and 187 species were predicted to be identifiable down to the species rank based on ITS1, BenA, and CaM, respectively. Finally, air filter samples were analysed to quantify the relative abundances of Aspergillus species in outdoor air. The results were reproducible across biological duplicates both at the species and section ranks, but not strongly correlated between ITS1 and BenA, suggesting the Aspergillus detection can be taxonomically biased depending on the selection of the DNA markers and/or primers.

  9. Functional Overexpression of Vomeronasal Receptors Using a Herpes Simplex Virus Type 1 (HSV-1)-Derived Amplicon.

    Science.gov (United States)

    Stein, Benjamin; Alonso, María Teresa; Zufall, Frank; Leinders-Zufall, Trese; Chamero, Pablo

    2016-01-01

    In mice, social behaviors such as mating and aggression are mediated by pheromones and related chemosignals. The vomeronasal organ (VNO) detects olfactory information from other individuals by sensory neurons tuned to respond to specific chemical cues. Receptors expressed by vomeronasal neurons are implicated in selective detection of these cues. Nearly 400 receptor genes have been identified in the mouse VNO, but the tuning properties of individual receptors remain poorly understood, in part due to the lack of a robust heterologous expression system. Here we develop a herpes virus-based amplicon delivery system to overexpress three types of vomeronasal receptor genes and to characterize cell responses to their proposed ligands. Through Ca2+ imaging in native VNO cells we show that virus-induced overexpression of V1rj2, V2r1b or Fpr3 caused a pronounced increase of responsivity to sulfated steroids, MHC-binding peptide or the synthetic hexapeptide W-peptide, respectively. Other related ligands were not recognized by infected individual neurons, indicating a high degree of selectivity by the overexpressed receptor. Removal of G-protein signaling eliminates Ca2+ responses, indicating that the endogenous second messenger system is essential for observing receptor activation. Our results provide a novel expression system for vomeronasal receptors that should be useful for understanding the molecular logic of VNO ligand detection. Functional expression of vomeronasal receptors and their deorphanization provides an essential requirement for deciphering the neural mechanisms controlling behavior. PMID:27195771

  10. Functional Overexpression of Vomeronasal Receptors Using a Herpes Simplex Virus Type 1 (HSV-1-Derived Amplicon.

    Directory of Open Access Journals (Sweden)

    Benjamin Stein

    Full Text Available In mice, social behaviors such as mating and aggression are mediated by pheromones and related chemosignals. The vomeronasal organ (VNO detects olfactory information from other individuals by sensory neurons tuned to respond to specific chemical cues. Receptors expressed by vomeronasal neurons are implicated in selective detection of these cues. Nearly 400 receptor genes have been identified in the mouse VNO, but the tuning properties of individual receptors remain poorly understood, in part due to the lack of a robust heterologous expression system. Here we develop a herpes virus-based amplicon delivery system to overexpress three types of vomeronasal receptor genes and to characterize cell responses to their proposed ligands. Through Ca2+ imaging in native VNO cells we show that virus-induced overexpression of V1rj2, V2r1b or Fpr3 caused a pronounced increase of responsivity to sulfated steroids, MHC-binding peptide or the synthetic hexapeptide W-peptide, respectively. Other related ligands were not recognized by infected individual neurons, indicating a high degree of selectivity by the overexpressed receptor. Removal of G-protein signaling eliminates Ca2+ responses, indicating that the endogenous second messenger system is essential for observing receptor activation. Our results provide a novel expression system for vomeronasal receptors that should be useful for understanding the molecular logic of VNO ligand detection. Functional expression of vomeronasal receptors and their deorphanization provides an essential requirement for deciphering the neural mechanisms controlling behavior.

  11. Extracellular DNA amplicon sequencing reveals high levels of benthic eukaryotic diversity in the central Red Sea

    KAUST Repository

    Pearman, John K.

    2015-11-01

    The present study aims to characterize the benthic eukaryotic biodiversity patterns at a coarse taxonomic level in three areas of the central Red Sea (a lagoon, an offshore area in Thuwal and a shallow coastal area near Jeddah) based on extracellular DNA. High-throughput amplicon sequencing targeting the V9 region of the 18S rRNA gene was undertaken for 32 sediment samples. High levels of alpha-diversity were detected with 16,089 operational taxonomic units (OTUs) being identified. The majority of the OTUs were assigned to Metazoa (29.2%), Alveolata (22.4%) and Stramenopiles (17.8%). Stramenopiles (Diatomea) and Alveolata (Ciliophora) were frequent in a lagoon and in shallower coastal stations, whereas metazoans (Arthropoda: Maxillopoda) were dominant in deeper offshore stations. Only 24.6% of total OTUs were shared among all areas. Beta-diversity was generally lower between the lagoon and Jeddah (nearshore) than between either of those and the offshore area, suggesting a nearshore–offshore biodiversity gradient. The current approach allowed for a broad-range of benthic eukaryotic biodiversity to be analysed with significantly less labour than would be required by other traditional taxonomic approaches. Our findings suggest that next generation sequencing techniques have the potential to provide a fast and standardised screening of benthic biodiversity at large spatial and temporal scales.

  12. Shedding light on the microbial community of the macropod foregut using 454-amplicon pyrosequencing.

    Directory of Open Access Journals (Sweden)

    Lisa-Maree Gulino

    Full Text Available Twenty macropods from five locations in Queensland, Australia, grazing on a variety of native pastures were surveyed and the bacterial community of the foregut was examined using 454-amplicon pyrosequencing. Specifically, the V3/V4 region of 16S rRNA gene was examined. A total of 5040 OTUs were identified in the data set (post filtering. Thirty-two OTUs were identified as 'shared' OTUS (i.e. present in all samples belonging to either Firmicutes or Bacteroidetes (Clostridiales/Bacteroidales. These phyla predominated the general microbial community in all macropods. Genera represented within the shared OTUs included: unclassified Ruminococcaceae, unclassified Lachnospiraceae, unclassified Clostridiales, Peptococcus sp. Coprococcus spp., Streptococcus spp., Blautia sp., Ruminoccocus sp., Eubacterium sp., Dorea sp., Oscillospira sp. and Butyrivibrio sp. The composition of the bacterial community of the foregut samples of each the host species (Macropus rufus, Macropus giganteus and Macropus robustus was significantly different allowing differentiation between the host species based on alpha and beta diversity measures. Specifically, eleven dominant OTUs that separated the three host species were identified and classified as: unclassified Ruminococcaceae, unclassified Bacteroidales, Prevotella spp. and a Syntrophococcus sucromutans. Putative reductive acetogens and fibrolytic bacteria were also identified in samples. Future work will investigate the presence and role of fibrolytics and acetogens in these ecosystems. Ideally, the isolation and characterization of these organisms will be used for enhanced feed efficiency in cattle, methane mitigation and potentially for other industries such as the biofuel industry.

  13. Flow cytometry community fingerprinting and amplicon sequencing for the assessment of landfill leachate cellulolytic bioaugmentation.

    Science.gov (United States)

    Kinet, R; Dzaomuho, P; Baert, J; Taminiau, B; Daube, G; Nezer, C; Brostaux, Y; Nguyen, F; Dumont, G; Thonart, P; Delvigne, F

    2016-08-01

    Flow cytometry (FCM) is a high throughput single cell technology that is actually becoming widely used for studying phenotypic and genotypic diversity among microbial communities. This technology is considered in this work for the assessment of a bioaugmentation treatment in order to enhance cellulolytic potential of landfill leachate. The experimental results reveal the relevant increase of leachate cellulolytic potential due to bioaugmentation. Cytometric monitoring of microbial dynamics along these assays is then realized. The flow FP package is used to establish microbial samples fingerprint from initial 2D cytometry histograms. This procedure allows highlighting microbial communities' variation along the assays. Cytometric and 16S rRNA gene sequencing fingerprinting methods are then compared. The two approaches give same evidence about microbial dynamics throughout digestion assay. There are however a lack of significant correlation between cytometric and amplicon sequencing fingerprint at genus or species level. Same phenotypical profiles of microbiota during assays matched to several 16S rRNA gene sequencing ones. Flow cytometry fingerprinting can thus be considered as a promising routine on-site method suitable for the detection of stability/variation/disturbance of complex microbial communities involved in bioprocesses. PMID:27160955

  14. Amplicon pyrosequencing reveals the soil microbial diversity associated with invasive Japanese barberry (Berberis thunbergii DC.).

    Science.gov (United States)

    Coats, V C; Pelletreau, K N; Rumpho, M E

    2014-03-01

    The soil microbial community acts as a reservoir of microbes that directly influences the structure and composition of the aboveground plant community, promotes plant growth, increases stress tolerance and mediates local patterns of nutrient cycling. Direct interactions between plants and rhizosphere-dwelling microorganisms occur at, or near, the surface of the root. Upon introduction and establishment, invasive plants modify the soil microbial communities and soil biochemistry affecting bioremediation efforts and future plant communities. Here, we used tag-encoded FLX amplicon 454 pyrosequencing (TEFAP) to characterize the bacterial and fungal community diversity in the rhizosphere of Berberis thunbergii DC. (Japanese barberry) from invasive stands in coastal Maine to investigate effects of soil type, soil chemistry and surrounding plant cover on the soil microbial community structure. Acidobacteria, Actinobacteria, Proteobacteria and Verrucomicrobia were the dominant bacterial phyla, whereas fungal communities were comprised mostly of Ascomycota and Basidiomycota phyla members, including Agaricomycetes and Sordariomycetes. Bulk soil chemistry had more effect on the bacterial community structure than the fungal community. An effect of geographic location was apparent in the rhizosphere microbial communities, yet it was less significant than the effect of surrounding plant cover. These data demonstrate a high degree of spatial variation in the rhizosphere microbial communities of Japanese barberry with apparent effects of soil chemistry, location and canopy cover on the microbial community structure. PMID:24118303

  15. Implementing amplicon-based next generation sequencing in the diagnosis of small cell lung carcinoma metastases.

    Science.gov (United States)

    Meder, Lydia; König, Katharina; Fassunke, Jana; Ozretić, Luka; Wolf, Jürgen; Merkelbach-Bruse, Sabine; Heukamp, Lukas C; Buettner, Reinhard

    2015-12-01

    Small cell lung carcinoma (SCLC) is the most aggressive entity of lung cancer. Rapid cancer progression and early formation of systemic metastases drive the deadly outcome of SCLC. Recent advances in identifying oncogenes by cancer whole genome sequencing improved the understanding of SCLC carcinogenesis. However, tumor material is often limited in the clinic. Thus, it is a compulsive issue to improve SCLC diagnostics by combining established immunohistochemistry and next generation sequencing. We implemented amplicon-based next generation deep sequencing in our routine diagnostics pipeline to analyze RB1, TP53, EP300 and CREBBP, frequently mutated in SCLC. Thereby, our pipeline combined routine SCLC histology and identification of somatic mutations. We comprehensively analyzed fifty randomly collected SCLC metastases isolated from trachea and lymph nodes in comparison to specimens derived from primary SCLC. SCLC lymph node metastases showed enhanced proliferation and frequently a collapsed keratin cytoskeleton compared to SCLC metastases isolated from trachea. We identified characteristic synchronous mutations in RB1 and TP53 and non-synchronous CREBBP and EP300 mutations. Our data showed the benefit of implementing deep sequencing into routine diagnostics. We here identify oncogenic drivers and simultaneously gain further insights into SCLC tumor biology.

  16. Comparative analyses of amplicon migration behavior in differing denaturing gradient gel electrophoresis (DGGE) systems

    Science.gov (United States)

    Thornhill, D. J.; Kemp, D. W.; Sampayo, E. M.; Schmidt, G. W.

    2010-03-01

    Denaturing gradient gel electrophoresis (DGGE) is commonly utilized to identify and quantify microbial diversity, but the conditions required for different electrophoretic systems to yield equivalent results and optimal resolution have not been assessed. Herein, the influence of different DGGE system configuration parameters on microbial diversity estimates was tested using Symbiodinium, a group of marine eukaryotic microbes that are important constituents of coral reef ecosystems. To accomplish this, bacterial clone libraries were constructed and sequenced from cultured isolates of Symbiodinium for the ribosomal DNA internal transcribed spacer 2 (ITS2) region. From these, 15 clones were subjected to PCR with a GC clamped primer set for DGGE analyses. Migration behaviors of the resulting amplicons were analyzed using a range of conditions, including variation in the composition of the denaturing gradient, electrophoresis time, and applied voltage. All tests were conducted in parallel on two commercial DGGE systems, a C.B.S. Scientific DGGE-2001, and the Bio-Rad DCode system. In this context, identical nucleotide fragments exhibited differing migration behaviors depending on the model of apparatus utilized, with fragments denaturing at a lower gradient concentration and applied voltage on the Bio-Rad DCode system than on the C.B.S. Scientific DGGE-2001 system. Although equivalent PCR-DGGE profiles could be achieved with both brands of DGGE system, the composition of the denaturing gradient and application of electrophoresis time × voltage must be appropriately optimized to achieve congruent results across platforms.

  17. Groundtruthing next-gen sequencing for microbial ecology-biases and errors in community structure estimates from PCR amplicon pyrosequencing.

    Directory of Open Access Journals (Sweden)

    Charles K Lee

    Full Text Available Analysis of microbial communities by high-throughput pyrosequencing of SSU rRNA gene PCR amplicons has transformed microbial ecology research and led to the observation that many communities contain a diverse assortment of rare taxa-a phenomenon termed the Rare Biosphere. Multiple studies have investigated the effect of pyrosequencing read quality on operational taxonomic unit (OTU richness for contrived communities, yet there is limited information on the fidelity of community structure estimates obtained through this approach. Given that PCR biases are widely recognized, and further unknown biases may arise from the sequencing process itself, a priori assumptions about the neutrality of the data generation process are at best unvalidated. Furthermore, post-sequencing quality control algorithms have not been explicitly evaluated for the accuracy of recovered representative sequences and its impact on downstream analyses, reducing useful discussion on pyrosequencing reads to their diversity and abundances. Here we report on community structures and sequences recovered for in vitro-simulated communities consisting of twenty 16S rRNA gene clones tiered at known proportions. PCR amplicon libraries of the V3-V4 and V6 hypervariable regions from the in vitro-simulated communities were sequenced using the Roche 454 GS FLX Titanium platform. Commonly used quality control protocols resulted in the formation of OTUs with >1% abundance composed entirely of erroneous sequences, while over-aggressive clustering approaches obfuscated real, expected OTUs. The pyrosequencing process itself did not appear to impose significant biases on overall community structure estimates, although the detection limit for rare taxa may be affected by PCR amplicon size and quality control approach employed. Meanwhile, PCR biases associated with the initial amplicon generation may impose greater distortions in the observed community structure.

  18. Assessment of DNA degradation induced by thermal and UV radiation processing: implications for quantification of genetically modified organisms.

    Science.gov (United States)

    Ballari, Rajashekhar V; Martin, Asha

    2013-12-01

    DNA quality is an important parameter for the detection and quantification of genetically modified organisms (GMO's) using the polymerase chain reaction (PCR). Food processing leads to degradation of DNA, which may impair GMO detection and quantification. This study evaluated the effect of various processing treatments such as heating, baking, microwaving, autoclaving and ultraviolet (UV) irradiation on the relative transgenic content of MON 810 maize using pRSETMON-02, a dual target plasmid as a model system. Amongst all the processing treatments examined, autoclaving and UV irradiation resulted in the least recovery of the transgenic (CaMV 35S promoter) and taxon-specific (zein) target DNA sequences. Although a profound impact on DNA degradation was seen during the processing, DNA could still be reliably quantified by Real-time PCR. The measured mean DNA copy number ratios of the processed samples were in agreement with the expected values. Our study confirms the premise that the final analytical value assigned to a particular sample is independent of the degree of DNA degradation since the transgenic and the taxon-specific target sequences possessing approximately similar lengths degrade in parallel. The results of our study demonstrate that food processing does not alter the relative quantification of the transgenic content provided the quantitative assays target shorter amplicons and the difference in the amplicon size between the transgenic and taxon-specific genes is minimal.

  19. Disease quantification in dermatology

    DEFF Research Database (Denmark)

    Greve, Tanja Maria; Kamp, Søren; Jemec, Gregor B E

    2013-01-01

    Accurate documentation of disease severity is a prerequisite for clinical research and the practice of evidence-based medicine. The quantification of skin diseases such as psoriasis currently relies heavily on clinical scores. Although these clinical scoring methods are well established and very ...

  20. Analysis of the mouse gut microbiome using full-length 16S rRNA amplicon sequencing.

    Science.gov (United States)

    Shin, Jongoh; Lee, Sooin; Go, Min-Jeong; Lee, Sang Yup; Kim, Sun Chang; Lee, Chul-Ho; Cho, Byung-Kwan

    2016-01-01

    Demands for faster and more accurate methods to analyze microbial communities from natural and clinical samples have been increasing in the medical and healthcare industry. Recent advances in next-generation sequencing technologies have facilitated the elucidation of the microbial community composition with higher accuracy and greater throughput than was previously achievable; however, the short sequencing reads often limit the microbial composition analysis at the species level due to the high similarity of 16S rRNA amplicon sequences. To overcome this limitation, we used the nanopore sequencing platform to sequence full-length 16S rRNA amplicon libraries prepared from the mouse gut microbiota. A comparison of the nanopore and short-read sequencing data showed that there were no significant differences in major taxonomic units (89%) except one phylotype and three taxonomic units. Moreover, both sequencing data were highly similar at all taxonomic resolutions except the species level. At the species level, nanopore sequencing allowed identification of more species than short-read sequencing, facilitating the accurate classification of the bacterial community composition. Therefore, this method of full-length 16S rRNA amplicon sequencing will be useful for rapid, accurate and efficient detection of microbial diversity in various biological and clinical samples. PMID:27411898

  1. Voltammetric detection of sequence-selective DNA hybridization related to Toxoplasma gondii in PCR amplicons.

    Science.gov (United States)

    Gokce, Gultekin; Erdem, Arzum; Ceylan, Cagdas; Akgöz, Muslum

    2016-03-01

    This work describes the single-use electrochemical DNA biosensor technology developed for voltammetric detection of sequence selective DNA hybridization related to important human and veterinary pathogen; Toxoplasma gondii. In the principle of electrochemical label-free detection assay, the duplex of DNA hybrid formation was detected by measuring guanine oxidation signal occured in the presence of DNA hybridization. The biosensor design consisted of the immobilization of an inosine-modified (guanine-free) probe onto the surface of pencil graphite electrode (PGE), and the detection of the duplex formation in connection with the differential pulse voltammetry(DPV) by measuring the guanine signal. Toxoplasma gondii capture probe was firstly immobilized onto the surface of the activated PGE by wet adsorption. The extent of hybridization at PGE surface between the probe and the target was then determined by measuring the guanine signal observed at +1.0V. The electrochemical monitoring of optimum DNA hybridization has been performed in the target concentration of 40µg/mL in 50min of hybridization time. The specificity of the electrochemical biosensor was then tested using non-complementary, or mismatch short DNA sequences. Under the optimum conditions, the guanine oxidation signal indicating full hybridization was measured in various target concentration from 0.5 to 25µg/mL and a detection limit was found to be 1.78µg/mL. This single-use biosensor platform was successfully applied for the voltammetric detection of DNA hybridization related to Toxoplasma gondii in PCR amplicons. PMID:26717837

  2. Amplicon-based semiconductor sequencing of human exomes: performance evaluation and optimization strategies.

    Science.gov (United States)

    Damiati, E; Borsani, G; Giacopuzzi, Edoardo

    2016-05-01

    The Ion Proton platform allows to perform whole exome sequencing (WES) at low cost, providing rapid turnaround time and great flexibility. Products for WES on Ion Proton system include the AmpliSeq Exome kit and the recently introduced HiQ sequencing chemistry. Here, we used gold standard variants from GIAB consortium to assess the performances in variants identification, characterize the erroneous calls and develop a filtering strategy to reduce false positives. The AmpliSeq Exome kit captures a large fraction of bases (>94 %) in human CDS, ClinVar genes and ACMG genes, but with 2,041 (7 %), 449 (13 %) and 11 (19 %) genes not fully represented, respectively. Overall, 515 protein coding genes contain hard-to-sequence regions, including 90 genes from ClinVar. Performance in variants detection was maximum at mean coverage >120×, while at 90× and 70× we measured a loss of variants of 3.2 and 4.5 %, respectively. WES using HiQ chemistry showed ~71/97.5 % sensitivity, ~37/2 % FDR and ~0.66/0.98 F1 score for indels and SNPs, respectively. The proposed low, medium or high-stringency filters reduced the amount of false positives by 10.2, 21.2 and 40.4 % for indels and 21.2, 41.9 and 68.2 % for SNP, respectively. Amplicon-based WES on Ion Proton platform using HiQ chemistry emerged as a competitive approach, with improved accuracy in variants identification. False-positive variants remain an issue for the Ion Torrent technology, but our filtering strategy can be applied to reduce erroneous variants.

  3. Amplicon-based semiconductor sequencing of human exomes: performance evaluation and optimization strategies.

    Science.gov (United States)

    Damiati, E; Borsani, G; Giacopuzzi, Edoardo

    2016-05-01

    The Ion Proton platform allows to perform whole exome sequencing (WES) at low cost, providing rapid turnaround time and great flexibility. Products for WES on Ion Proton system include the AmpliSeq Exome kit and the recently introduced HiQ sequencing chemistry. Here, we used gold standard variants from GIAB consortium to assess the performances in variants identification, characterize the erroneous calls and develop a filtering strategy to reduce false positives. The AmpliSeq Exome kit captures a large fraction of bases (>94 %) in human CDS, ClinVar genes and ACMG genes, but with 2,041 (7 %), 449 (13 %) and 11 (19 %) genes not fully represented, respectively. Overall, 515 protein coding genes contain hard-to-sequence regions, including 90 genes from ClinVar. Performance in variants detection was maximum at mean coverage >120×, while at 90× and 70× we measured a loss of variants of 3.2 and 4.5 %, respectively. WES using HiQ chemistry showed ~71/97.5 % sensitivity, ~37/2 % FDR and ~0.66/0.98 F1 score for indels and SNPs, respectively. The proposed low, medium or high-stringency filters reduced the amount of false positives by 10.2, 21.2 and 40.4 % for indels and 21.2, 41.9 and 68.2 % for SNP, respectively. Amplicon-based WES on Ion Proton platform using HiQ chemistry emerged as a competitive approach, with improved accuracy in variants identification. False-positive variants remain an issue for the Ion Torrent technology, but our filtering strategy can be applied to reduce erroneous variants. PMID:27003585

  4. Analysis of a Drosophila amplicon in follicle cells highlights the diversity of metazoan replication origins.

    Science.gov (United States)

    Kim, Jane C; Orr-Weaver, Terry L

    2011-10-01

    To investigate the properties of metazoan replication origins, recent studies in cell culture have adopted the strategy of identifying origins using genome-wide approaches and assessing correlations with such features as transcription and histone modifications. Drosophila amplicon in follicle cells (DAFCs), genomic regions that undergo repeated rounds of DNA replication to increase DNA copy number, serve as powerful in vivo model replicons. Because there are six DAFCs, compared with thousands of origins activated in the typical S phase, close molecular characterization of all DAFCs is possible. To determine the extent to which the six DAFCs are different or similar, we investigated the developmental and replication properties of the newly identified DAFC-34B. DAFC-34B contains two genes expressed in follicle cells, although the timing and spatial patterns of expression suggest that amplification is not a strategy to promote high expression at this locus. Like the previously characterized DAFC-62D, DAFC-34B displays origin activation at two separate stages of development. However, unlike DAFC-62D, amplification at the later stage is not transcription-dependent. We mapped the DAFC-34B amplification origin to 1 kb by nascent strand analysis and delineated cis requirements for origin activation, finding that a 6-kb region, but not the 1-kb origin alone, is sufficient for amplification. We analyzed the developmental localization of the origin recognition complex (ORC) and the minichromosome maintenance (MCM)2-7 complex, the replicative helicase. Intriguingly, the final round of origin activation at DAFC-34B occurs in the absence of detectable ORC, although MCMs are present, suggesting a new amplification initiation mechanism. PMID:21933960

  5. Competitive amplification of differentially melting amplicons (CADMA improves KRAS hotspot mutation testing in colorectal cancer

    Directory of Open Access Journals (Sweden)

    Kristensen Lasse

    2012-11-01

    Full Text Available Abstract Background Cancer is an extremely heterogeneous group of diseases traditionally categorized according to tissue of origin. However, even among patients with the same cancer subtype the cellular alterations at the molecular level are often very different. Several new therapies targeting specific molecular changes found in individual patients have initiated the era of personalized therapy and significantly improved patient care. In metastatic colorectal cancer (mCRC a selected group of patients with wild-type KRAS respond to antibodies against the epidermal growth factor receptor (EGFR. Testing for KRAS mutations is now required prior to anti-EGFR treatment, however, less sensitive methods based on conventional PCR regularly fail to detect KRAS mutations in clinical samples. Methods We have developed sensitive and specific assays for detection of the seven most common KRAS mutations based on a novel methodology named Competitive Amplification of Differentially Melting Amplicons (CADMA. The clinical applicability of these assays was assessed by analyzing 100 colorectal cancer samples, for which KRAS mutation status has been evaluated by the commercially available TheraScreen® KRAS mutation kit. Results The CADMA assays were sensitive to at least 0.5% mutant alleles in a wild-type background when using 50 nanograms of DNA in the reactions. Consensus between CADMA and the TheraScreen kit was observed in 96% of the colorectal cancer samples. In cases where disagreement was observed the CADMA result could be confirmed by a previously published assay based on TaqMan probes and by fast COLD-PCR followed by Sanger sequencing. Conclusions The high analytical sensitivity and specificity of CADMA may increase diagnostic sensitivity and specificity of KRAS mutation testing in mCRC patients.

  6. De novo origin of VCY2 from autosome to Y-transposed amplicon.

    Directory of Open Access Journals (Sweden)

    Peng-Rong Cao

    Full Text Available The formation of new genes is a primary driving force of evolution in all organisms. The de novo evolution of new genes from non-protein-coding genomic regions is emerging as an important additional mechanism for novel gene creation. Y chromosomes underlie sex determination in mammals and contain genes that are required for male-specific functions. In this study, a search was undertaken for Y chromosome de novo genes derived from non-protein-coding sequences. The Y chromosome orphan gene variable charge, Y-linked (VCY2, is an autosome-derived gene that has sequence similarity to large autosomal fragments but lacks an autosomal protein-coding homolog. VCY2 locates in the amplicon containing long DNA fragments that were transposed from autosomes to the Y chromosome before the ape-monkey split. We confirmed that VCY2 cannot be encoded by autosomes due to the presence of multiple disablers that disrupt the open reading frame, such as the absence of start or stop codons and the presence of premature stop codons. Similar observations have been made for homologs in the autosomes of the chimpanzee, gorilla, rhesus macaque, baboon and out-group marmoset, which suggests that there was a non-protein-coding ancestral VCY2 that was common to apes and monkeys that predated the transposition event. Furthermore, while protein-coding orthologs are absent, a putative non-protein-coding VCY2 with conserved disablers was identified in the rhesus macaque Y chromosome male-specific region. This finding implies that VCY2 might have not acquired its protein-coding ability before the ape-monkey split. VCY2 encodes a testis-specific expressed protein and is involved in the pathologic process of male infertility, and the acquisition of this gene might improve male fertility. This is the first evidence that de novo genes can be generated from transposed autosomal non-protein-coding segments, and this evidence provides novel insights into the evolutionary history of the Y

  7. De novo origin of VCY2 from autosome to Y-transposed amplicon.

    Science.gov (United States)

    Cao, Peng-Rong; Wang, Lei; Jiang, Yu-Chao; Yi, Yin-Sha; Qu, Fang; Liu, Tao-Cheng; Lv, Yuan

    2015-01-01

    The formation of new genes is a primary driving force of evolution in all organisms. The de novo evolution of new genes from non-protein-coding genomic regions is emerging as an important additional mechanism for novel gene creation. Y chromosomes underlie sex determination in mammals and contain genes that are required for male-specific functions. In this study, a search was undertaken for Y chromosome de novo genes derived from non-protein-coding sequences. The Y chromosome orphan gene variable charge, Y-linked (VCY)2, is an autosome-derived gene that has sequence similarity to large autosomal fragments but lacks an autosomal protein-coding homolog. VCY2 locates in the amplicon containing long DNA fragments that were transposed from autosomes to the Y chromosome before the ape-monkey split. We confirmed that VCY2 cannot be encoded by autosomes due to the presence of multiple disablers that disrupt the open reading frame, such as the absence of start or stop codons and the presence of premature stop codons. Similar observations have been made for homologs in the autosomes of the chimpanzee, gorilla, rhesus macaque, baboon and out-group marmoset, which suggests that there was a non-protein-coding ancestral VCY2 that was common to apes and monkeys that predated the transposition event. Furthermore, while protein-coding orthologs are absent, a putative non-protein-coding VCY2 with conserved disablers was identified in the rhesus macaque Y chromosome male-specific region. This finding implies that VCY2 might have not acquired its protein-coding ability before the ape-monkey split. VCY2 encodes a testis-specific expressed protein and is involved in the pathologic process of male infertility, and the acquisition of this gene might improve male fertility. This is the first evidence that de novo genes can be generated from transposed autosomal non-protein-coding segments, and this evidence provides novel insights into the evolutionary history of the Y chromosome. PMID

  8. Genotyping-in-Thousands by sequencing (GT-seq): A cost effective SNP genotyping method based on custom amplicon sequencing.

    Science.gov (United States)

    Campbell, Nathan R; Harmon, Stephanie A; Narum, Shawn R

    2015-07-01

    Genotyping-in-Thousands by sequencing (GT-seq) is a method that uses next-generation sequencing of multiplexed PCR products to generate genotypes from relatively small panels (50-500) of targeted single-nucleotide polymorphisms (SNPs) for thousands of individuals in a single Illumina HiSeq lane. This method uses only unlabelled oligos and PCR master mix in two thermal cycling steps for amplification of targeted SNP loci. During this process, sequencing adapters and dual barcode sequence tags are incorporated into the amplicons enabling thousands of individuals to be pooled into a single sequencing library. Post sequencing, reads from individual samples are split into individual files using their unique combination of barcode sequences. Genotyping is performed with a simple perl script which counts amplicon-specific sequences for each allele, and allele ratios are used to determine the genotypes. We demonstrate this technique by genotyping 2068 individual steelhead trout (Oncorhynchus mykiss) samples with a set of 192 SNP markers in a single library sequenced in a single Illumina HiSeq lane. Genotype data were 99.9% concordant to previously collected TaqMan(™) genotypes at the same 192 loci, but call rates were slightly lower with GT-seq (96.4%) relative to Taqman (99.0%). Of the 192 SNPs, 187 were genotyped in ≥90% of the individual samples and only 3 SNPs were genotyped in <70% of samples. This study demonstrates amplicon sequencing with GT-seq greatly reduces the cost of genotyping hundreds of targeted SNPs relative to existing methods by utilizing a simple library preparation method and massive efficiency of scale.

  9. The cytological manifestation of gene amplification in multidrug-resistant mouse leukemia P388 sublines is correlated with amplicon content

    Energy Technology Data Exchange (ETDEWEB)

    Il`inskaya, G.V.; Kopnin, B.P. [Institute of Carcinogenesis, Moscow (Russian Federation); Demidova, N.S. [Institute of Chemical Physics, Moscow (Russian Federation)

    1995-10-01

    Previously, we showed that development of multidrug resistance (MDR) in mouse P388 leukemia cells is often associated with the appearance of newly-formed chromosomelike structures that contain amplified copies of the mdr1 gene. In the present study, we compared amplicon content in P388 sublines showing different types of these structures. A strong correlation between the formation of specific acentric markers consisting of two identical arms and the absence of the sorcin gene coamplification was found. In all the sublines containing other types of chromosomelike structures, the sorcin gene is coamplified. 9 refs., 2 figs., 1 tab.

  10. A Portable Automatic Endpoint Detection System for Amplicons of Loop Mediated Isothermal Amplification on Microfluidic Compact Disk Platform

    Directory of Open Access Journals (Sweden)

    Shah Mukim Uddin

    2015-03-01

    Full Text Available In recent years, many improvements have been made in foodborne pathogen detection methods to reduce the impact of food contamination. Several rapid methods have been developed with biosensor devices to improve the way of performing pathogen detection. This paper presents an automated endpoint detection system for amplicons generated by loop mediated isothermal amplification (LAMP on a microfluidic compact disk platform. The developed detection system utilizes a monochromatic ultraviolet (UV emitter for excitation of fluorescent labeled LAMP amplicons and a color sensor to detect the emitted florescence from target. Then it processes the sensor output and displays the detection results on liquid crystal display (LCD. The sensitivity test has been performed with detection limit up to 2.5 × 10−3 ng/µL with different DNA concentrations of Salmonella bacteria. This system allows a rapid and automatic endpoint detection which could lead to the development of a point-of-care diagnosis device for foodborne pathogens detection in a resource-limited environment.

  11. RiboFR-Seq: a novel approach to linking 16S rRNA amplicon profiles to metagenomes.

    Science.gov (United States)

    Zhang, Yanming; Ji, Peifeng; Wang, Jinfeng; Zhao, Fangqing

    2016-06-01

    16S rRNA amplicon analysis and shotgun metagenome sequencing are two main culture-independent strategies to explore the genetic landscape of various microbial communities. Recently, numerous studies have employed these two approaches together, but downstream data analyses were performed separately, which always generated incongruent or conflict signals on both taxonomic and functional classifications. Here we propose a novel approach, RiboFR-Seq (Ribosomal RNA gene flanking region sequencing), for capturing both ribosomal RNA variable regions and their flanking protein-coding genes simultaneously. Through extensive testing on clonal bacterial strain, salivary microbiome and bacterial epibionts of marine kelp, we demonstrated that RiboFR-Seq could detect the vast majority of bacteria not only in well-studied microbiomes but also in novel communities with limited reference genomes. Combined with classical amplicon sequencing and shotgun metagenome sequencing, RiboFR-Seq can link the annotations of 16S rRNA and metagenomic contigs to make a consensus classification. By recognizing almost all 16S rRNA copies, the RiboFR-seq approach can effectively reduce the taxonomic abundance bias resulted from 16S rRNA copy number variation. We believe that RiboFR-Seq, which provides an integrated view of 16S rRNA profiles and metagenomes, will help us better understand diverse microbial communities. PMID:26984526

  12. CLOTU: An online pipeline for processing and clustering of 454 amplicon reads into OTUs followed by taxonomic annotation

    Directory of Open Access Journals (Sweden)

    Shalchian-Tabrizi Kamran

    2011-05-01

    Full Text Available Abstract Background The implementation of high throughput sequencing for exploring biodiversity poses high demands on bioinformatics applications for automated data processing. Here we introduce CLOTU, an online and open access pipeline for processing 454 amplicon reads. CLOTU has been constructed to be highly user-friendly and flexible, since different types of analyses are needed for different datasets. Results In CLOTU, the user can filter out low quality sequences, trim tags, primers, adaptors, perform clustering of sequence reads, and run BLAST against NCBInr or a customized database in a high performance computing environment. The resulting data may be browsed in a user-friendly manner and easily forwarded to downstream analyses. Although CLOTU is specifically designed for analyzing 454 amplicon reads, other types of DNA sequence data can also be processed. A fungal ITS sequence dataset generated by 454 sequencing of environmental samples is used to demonstrate the utility of CLOTU. Conclusions CLOTU is a flexible and easy to use bioinformatics pipeline that includes different options for filtering, trimming, clustering and taxonomic annotation of high throughput sequence reads. Some of these options are not included in comparable pipelines. CLOTU is implemented in a Linux computer cluster and is freely accessible to academic users through the Bioportal web-based bioinformatics service (http://www.bioportal.uio.no.

  13. Improved Efficiency and Reliability of NGS Amplicon Sequencing Data Analysis for Genetic Diagnostic Procedures Using AGSA Software

    Directory of Open Access Journals (Sweden)

    Axel Poulet

    2016-01-01

    Full Text Available Screening for BRCA mutations in women with familial risk of breast or ovarian cancer is an ideal situation for high-throughput sequencing, providing large amounts of low cost data. However, 454, Roche, and Ion Torrent, Thermo Fisher, technologies produce homopolymer-associated indel errors, complicating their use in routine diagnostics. We developed software, named AGSA, which helps to detect false positive mutations in homopolymeric sequences. Seventy-two familial breast cancer cases were analysed in parallel by amplicon 454 pyrosequencing and Sanger dideoxy sequencing for genetic variations of the BRCA genes. All 565 variants detected by dideoxy sequencing were also detected by pyrosequencing. Furthermore, pyrosequencing detected 42 variants that were missed with Sanger technique. Six amplicons contained homopolymer tracts in the coding sequence that were systematically misread by the software supplied by Roche. Read data plotted as histograms by AGSA software aided the analysis considerably and allowed validation of the majority of homopolymers. As an optimisation, additional 250 patients were analysed using microfluidic amplification of regions of interest (Access Array Fluidigm of the BRCA genes, followed by 454 sequencing and AGSA analysis. AGSA complements a complete line of high-throughput diagnostic sequence analysis, reducing time and costs while increasing reliability, notably for homopolymer tracts.

  14. A Method for Amplicon Deep Sequencing of Drug Resistance Genes in Plasmodium falciparum Clinical Isolates from India.

    Science.gov (United States)

    Rao, Pavitra N; Uplekar, Swapna; Kayal, Sriti; Mallick, Prashant K; Bandyopadhyay, Nabamita; Kale, Sonal; Singh, Om P; Mohanty, Akshaya; Mohanty, Sanjib; Wassmer, Samuel C; Carlton, Jane M

    2016-06-01

    A major challenge to global malaria control and elimination is early detection and containment of emerging drug resistance. Next-generation sequencing (NGS) methods provide the resolution, scalability, and sensitivity required for high-throughput surveillance of molecular markers of drug resistance. We have developed an amplicon sequencing method on the Ion Torrent PGM platform for targeted resequencing of a panel of six Plasmodium falciparum genes implicated in resistance to first-line antimalarial therapy, including artemisinin combination therapy, chloroquine, and sulfadoxine-pyrimethamine. The protocol was optimized using 12 geographically diverse P. falciparum reference strains and successfully applied to multiplexed sequencing of 16 clinical isolates from India. The sequencing results from the reference strains showed 100% concordance with previously reported drug resistance-associated mutations. Single-nucleotide polymorphisms (SNPs) in clinical isolates revealed a number of known resistance-associated mutations and other nonsynonymous mutations that have not been implicated in drug resistance. SNP positions containing multiple allelic variants were used to identify three clinical samples containing mixed genotypes indicative of multiclonal infections. The amplicon sequencing protocol has been designed for the benchtop Ion Torrent PGM platform and can be operated with minimal bioinformatics infrastructure, making it ideal for use in countries that are endemic for the disease to facilitate routine large-scale surveillance of the emergence of drug resistance and to ensure continued success of the malaria treatment policy. PMID:27008882

  15. Improved Efficiency and Reliability of NGS Amplicon Sequencing Data Analysis for Genetic Diagnostic Procedures Using AGSA Software

    Science.gov (United States)

    Poulet, Axel; Privat, Maud; Viala, Sandrine; Decousus, Stephanie; Perin, Axel; Lafarge, Laurence; Ollier, Marie; El Saghir, Nagi S.

    2016-01-01

    Screening for BRCA mutations in women with familial risk of breast or ovarian cancer is an ideal situation for high-throughput sequencing, providing large amounts of low cost data. However, 454, Roche, and Ion Torrent, Thermo Fisher, technologies produce homopolymer-associated indel errors, complicating their use in routine diagnostics. We developed software, named AGSA, which helps to detect false positive mutations in homopolymeric sequences. Seventy-two familial breast cancer cases were analysed in parallel by amplicon 454 pyrosequencing and Sanger dideoxy sequencing for genetic variations of the BRCA genes. All 565 variants detected by dideoxy sequencing were also detected by pyrosequencing. Furthermore, pyrosequencing detected 42 variants that were missed with Sanger technique. Six amplicons contained homopolymer tracts in the coding sequence that were systematically misread by the software supplied by Roche. Read data plotted as histograms by AGSA software aided the analysis considerably and allowed validation of the majority of homopolymers. As an optimisation, additional 250 patients were analysed using microfluidic amplification of regions of interest (Access Array Fluidigm) of the BRCA genes, followed by 454 sequencing and AGSA analysis. AGSA complements a complete line of high-throughput diagnostic sequence analysis, reducing time and costs while increasing reliability, notably for homopolymer tracts.

  16. A Method for Amplicon Deep Sequencing of Drug Resistance Genes in Plasmodium falciparum Clinical Isolates from India.

    Science.gov (United States)

    Rao, Pavitra N; Uplekar, Swapna; Kayal, Sriti; Mallick, Prashant K; Bandyopadhyay, Nabamita; Kale, Sonal; Singh, Om P; Mohanty, Akshaya; Mohanty, Sanjib; Wassmer, Samuel C; Carlton, Jane M

    2016-06-01

    A major challenge to global malaria control and elimination is early detection and containment of emerging drug resistance. Next-generation sequencing (NGS) methods provide the resolution, scalability, and sensitivity required for high-throughput surveillance of molecular markers of drug resistance. We have developed an amplicon sequencing method on the Ion Torrent PGM platform for targeted resequencing of a panel of six Plasmodium falciparum genes implicated in resistance to first-line antimalarial therapy, including artemisinin combination therapy, chloroquine, and sulfadoxine-pyrimethamine. The protocol was optimized using 12 geographically diverse P. falciparum reference strains and successfully applied to multiplexed sequencing of 16 clinical isolates from India. The sequencing results from the reference strains showed 100% concordance with previously reported drug resistance-associated mutations. Single-nucleotide polymorphisms (SNPs) in clinical isolates revealed a number of known resistance-associated mutations and other nonsynonymous mutations that have not been implicated in drug resistance. SNP positions containing multiple allelic variants were used to identify three clinical samples containing mixed genotypes indicative of multiclonal infections. The amplicon sequencing protocol has been designed for the benchtop Ion Torrent PGM platform and can be operated with minimal bioinformatics infrastructure, making it ideal for use in countries that are endemic for the disease to facilitate routine large-scale surveillance of the emergence of drug resistance and to ensure continued success of the malaria treatment policy.

  17. The HER2 amplicon includes several genes required for the growth and survival of HER2 positive breast cancer cells — A data description

    Directory of Open Access Journals (Sweden)

    Vesa Hongisto

    2014-12-01

    Full Text Available A large number of breast cancers are characterized by amplification and overexpression of the chromosome segment surrounding the HER2 (ERBB2 oncogene. As the HER2 amplicon at 17q12 contains multiple genes, we have systematically explored the role of the HER2 co-amplified genes in breast cancer cell growth and their relation to trastuzumab resistance. We integrated array comparative genomic hybridization (aCGH data of the HER2 amplicon from 71 HER2 positive breast tumors and 10 cell lines with systematic functional RNA interference analysis of 23 core amplicon genes with several phenotypic endpoints in a panel of trastuzumab responding and non-responding HER2 positive breast cancer cells. In this Data in Brief we give a detailed description of the experimental procedures and the data analysis methods used in the study (1.

  18. Optimisation of 16S rDNA amplicon sequencing protocols for microbial community profiling of anaerobic digesters

    DEFF Research Database (Denmark)

    Kirkegaard, Rasmus Hansen; McIlroy, Simon Jon; Larsen, Poul;

    of the community composition. As such sample specific optimisation and standardisation of DNA extraction, as well PCR primer selection, are essential to minimising the potential for such biases. The aim of this study was to develop a protocol for optimized community profiling of anaerobic digesters. The FastDNA......RNA gene amplicon sequencing is rapid, cheap, high throughput, and has high taxonomic resolution. However, biases are introduced in multiple steps of this approach, including non-representative DNA extraction and uneven taxonomic coverage of selected PCR primers, potentially giving a skewed view...... SPIN kit was selected and the mechanical lysis parameters optimised for extraction of genomic DNA from mesophilic and thermophilic anaerobic digester samples. Different primer sets were compared for targeting the archaea and bacteria, both together and individually. Shotgun sequencing of a TruSeq PCR...

  19. Obtaining representative community profiles of anaerobic digesters through optimisation of 16S rRNA amplicon sequencing protocols

    DEFF Research Database (Denmark)

    Kirkegaard, Rasmus Hansen; McIlroy, Simon Jon; Karst, Søren Michael;

    of the community composition . As such sample specific optimisation and standardisation of DNA extraction, as well PCR primer selection, are essential to minimising the potential for such biases. The aim of this study was to develop a protocol for optimized community profiling of anaerobic digesters. The FastDNA......RNA gene amplicon sequencing is rapid, cheap, high throughput, and has high taxonomic resolution. However, biases are introduced in multiple steps of this approach, including non-representative DNA extraction and uneven taxonomic coverage of selected PCR primers, potentially giving a skewed view...... SPIN kit was selected and the mechanical lysis parameters optimised for extraction of genomic DNA from mesophilic and thermophilic anaerobic digester samples. Different primer sets were compared for targeting the archaea and bacteria, both together and individually . Shotgun sequencing...

  20. Network analysis of the microorganism in 25 Danish wastewater treatment plants over 7 years using high-throughput amplicon sequencing

    DEFF Research Database (Denmark)

    Albertsen, Mads; Larsen, Poul; Saunders, Aaron Marc;

    Wastewater treatment is the world’s largest biotechnological processes and a perfect model system for microbial ecology as the habitat is well defined and replicated all over the world. Extensive investigations on Danish wastewater treatment plants using fluorescent in situ hybridization have...... identified 38 probe-defined core genera, which are shared among all investigated Danish plants. A large body of knowledge exists on many of the core genera, however few attempts have been made to integrate the knowledge on a system-level understanding of the process. In this work we aimed to integrate...... the existing knowledge on core genera with high-throughput amplicon sequencing, plant design and process data in order to identify interactions and community shaping factors. We investigated 25 Danish full-scale wastewater treatment plants with nutrient removal over a period of 7 years with two to four samples...

  1. Shotgun metagenomes and multiple primer pair-barcode combinations of amplicons reveal biases in metabarcoding analyses of fungi

    Directory of Open Access Journals (Sweden)

    Leho Tedersoo

    2015-05-01

    Full Text Available Rapid development of high-throughput (HTS molecular identification methods has revolutionized our knowledge about taxonomic diversity and ecology of fungi. However, PCR-based methods exhibit multiple technical shortcomings that may bias our understanding of the fungal kingdom. This study was initiated to quantify potential biases in fungal community ecology by comparing the relative performance of amplicon-free shotgun metagenomics and amplicons of nine primer pairs over seven nuclear ribosomal DNA (rDNA regions often used in metabarcoding analyses. The internal transcribed spacer (ITS barcodes ITS1 and ITS2 provided greater taxonomic and functional resolution and richness of operational taxonomic units (OTUs at the 97% similarity threshold compared to barcodes located within the ribosomal small subunit (SSU and large subunit (LSU genes. All barcode-primer pair combinations provided consistent results in ranking taxonomic richness and recovering the importance of floristic variables in driving fungal community composition in soils of Papua New Guinea. The choice of forward primer explained up to 2.0% of the variation in OTU-level analysis of the ITS1 and ITS2 barcode data sets. Across the whole data set, barcode-primer pair combination explained 37.6–38.1% of the variation, which surpassed any environmental signal. Overall, the metagenomics data set recovered a similar taxonomic overview, but resulted in much lower fungal rDNA sequencing depth, inability to infer OTUs, and high uncertainty in identification. We recommend the use of ITS2 or the whole ITS region for metabarcoding and we advocate careful choice of primer pairs in consideration of the relative proportion of fungal DNA and expected dominant groups.

  2. Short-read assembly of full-length 16S amplicons reveals bacterial diversity in subsurface sediments.

    Directory of Open Access Journals (Sweden)

    Christopher S Miller

    Full Text Available In microbial ecology, a fundamental question relates to how community diversity and composition change in response to perturbation. Most studies have had limited ability to deeply sample community structure (e.g. Sanger-sequenced 16S rRNA libraries, or have had limited taxonomic resolution (e.g. studies based on 16S rRNA hypervariable region sequencing. Here, we combine the higher taxonomic resolution of near-full-length 16S rRNA gene amplicons with the economics and sensitivity of short-read sequencing to assay the abundance and identity of organisms that represent as little as 0.01% of sediment bacterial communities. We used a new version of EMIRGE optimized for large data size to reconstruct near-full-length 16S rRNA genes from amplicons sheared and sequenced with Illumina technology. The approach allowed us to differentiate the community composition among samples acquired before perturbation, after acetate amendment shifted the predominant metabolism to iron reduction, and once sulfate reduction began. Results were highly reproducible across technical replicates, and identified specific taxa that responded to the perturbation. All samples contain very high alpha diversity and abundant organisms from phyla without cultivated representatives. Surprisingly, at the time points measured, there was no strong loss of evenness, despite the selective pressure of acetate amendment and change in the terminal electron accepting process. However, community membership was altered significantly. The method allows for sensitive, accurate profiling of the "long tail" of low abundance organisms that exist in many microbial communities, and can resolve population dynamics in response to environmental change.

  3. Analysis of 16S rRNA amplicon sequencing options on the Roche/454 next-generation titanium sequencing platform.

    Directory of Open Access Journals (Sweden)

    Hideyuki Tamaki

    Full Text Available BACKGROUND: 16S rRNA gene pyrosequencing approach has revolutionized studies in microbial ecology. While primer selection and short read length can affect the resulting microbial community profile, little is known about the influence of pyrosequencing methods on the sequencing throughput and the outcome of microbial community analyses. The aim of this study is to compare differences in output, ease, and cost among three different amplicon pyrosequencing methods for the Roche/454 Titanium platform METHODOLOGY/PRINCIPAL FINDINGS: The following three pyrosequencing methods for 16S rRNA genes were selected in this study: Method-1 (standard method is the recommended method for bi-directional sequencing using the LIB-A kit; Method-2 is a new option designed in this study for unidirectional sequencing with the LIB-A kit; and Method-3 uses the LIB-L kit for unidirectional sequencing. In our comparison among these three methods using 10 different environmental samples, Method-2 and Method-3 produced 1.5-1.6 times more useable reads than the standard method (Method-1, after quality-based trimming, and did not compromise the outcome of microbial community analyses. Specifically, Method-3 is the most cost-effective unidirectional amplicon sequencing method as it provided the most reads and required the least effort in consumables management. CONCLUSIONS: Our findings clearly demonstrated that alternative pyrosequencing methods for 16S rRNA genes could drastically affect sequencing output (e.g. number of reads before and after trimming but have little effect on the outcomes of microbial community analysis. This finding is important for both researchers and sequencing facilities utilizing 16S rRNA gene pyrosequencing for microbial ecological studies.

  4. Verb aspect, alternations and quantification

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    Svetla Koeva

    2015-11-01

    Full Text Available Verb aspect, alternations and quantification In this paper we are briefly discuss the nature of Bulgarian verb aspect and argue that the verb aspect pairs are different lexical units with different (although related meaning, different argument structure (reflecting categories, explicitness and referential status of arguments and different sets of semantic and syntactic alternations. The verb prefixes resulting in perfective verbs derivation in some cases can be interpreted as lexical quantifiers as well. Thus the Bulgarian verb aspect is related (in different way both with the potential for the generation of alternations and with the prefixal lexical quantification. It is shown that the scope of the lexical quantification by means of verbal prefixes is the quantified verb phrase and the scope remains constant in all derived alternations. The paper concerns the basic issues of these complex problems, while the detailed description of the conditions satisfying particular alternation or particular lexical quantification are subject of a more detailed study.

  5. Quantification of human responses

    Science.gov (United States)

    Steinlage, R. C.; Gantner, T. E.; Lim, P. Y. W.

    1992-01-01

    Human perception is a complex phenomenon which is difficult to quantify with instruments. For this reason, large panels of people are often used to elicit and aggregate subjective judgments. Print quality, taste, smell, sound quality of a stereo system, softness, and grading Olympic divers and skaters are some examples of situations where subjective measurements or judgments are paramount. We usually express what is in our mind through language as a medium but languages are limited in available choices of vocabularies, and as a result, our verbalizations are only approximate expressions of what we really have in mind. For lack of better methods to quantify subjective judgments, it is customary to set up a numerical scale such as 1, 2, 3, 4, 5 or 1, 2, 3, ..., 9, 10 for characterizing human responses and subjective judgments with no valid justification except that these scales are easy to understand and convenient to use. But these numerical scales are arbitrary simplifications of the complex human mind; the human mind is not restricted to such simple numerical variations. In fact, human responses and subjective judgments are psychophysical phenomena that are fuzzy entities and therefore difficult to handle by conventional mathematics and probability theory. The fuzzy mathematical approach provides a more realistic insight into understanding and quantifying human responses. This paper presents a method for quantifying human responses and subjective judgments without assuming a pattern of linear or numerical variation for human responses. In particular, quantification and evaluation of linguistic judgments was investigated.

  6. Nitrogen quantification with SNMS

    Science.gov (United States)

    Goschnick, J.; Natzeck, C.; Sommer, M.

    1999-04-01

    Plasma-based secondary neutral mass spectrometry (plasma SNMS) is a powerful analytical method for determining the elemental concentrations of almost any kind of material at low cost by using a cheap quadrupole mass filter. However, a quadrupole-based mass spectrometer is limited to nominal mass resolution. Atomic signals are sometimes superimposed by molecular signals (2 or 3 atomic clusters such as CH +, CH 2+ or metal oxide clusters) and/or intensities of double-charged species. Especially in the case of nitrogen several interferences can impede the quantification. This article reports on methods to recognize and deconvolute superpositions of N + with CH 2+, Li 2+, and Si 2+ at mass 14 D (Debye) occurring during analysis of organic and inorganic substances. The recognition is based on the signal pattern of N +, Li +, CH +, and Si +. The latter serve as indicators for a probable interference of molecular or double-charged species with N on mass 14 D. The subsequent deconvolution use different shapes of atomic and cluster kinetic energy distributions (kEDs) to determine the quantities of the intensity components by a linear fit of N + and non-atomic kEDs obtained from several organic and inorganic standards into the measured kED. The atomic intensity fraction yields a much better nitrogen concentration than the total intensity of mass 14 D after correction.

  7. Investigation of Microbial Diversity in Geothermal Hot Springs in Unkeshwar, India, Based on 16S rRNA Amplicon Metagenome Sequencing.

    Science.gov (United States)

    Mehetre, Gajanan T; Paranjpe, Aditi; Dastager, Syed G; Dharne, Mahesh S

    2016-02-25

    Microbial diversity in geothermal waters of the Unkeshwar hot springs in Maharashtra, India, was studied using 16S rRNA amplicon metagenomic sequencing. Taxonomic analysis revealed the presence of Bacteroidetes, Proteobacteria, Cyanobacteria, Actinobacteria, Archeae, and OD1 phyla. Metabolic function prediction analysis indicated a battery of biological information systems indicating rich and novel microbial diversity, with potential biotechnological applications in this niche.

  8. A one-step real-time multiplex PCR for screening Y-chromosomal microdeletions without downstream amplicon size analysis.

    Directory of Open Access Journals (Sweden)

    Viviana Kozina

    Full Text Available BACKGROUND: Y-chromosomal microdeletions (YCMD are one of the major genetic causes for non-obstructive azoospermia. Genetic testing for YCMD by multiplex polymerase chain reaction (PCR is an established method for quick and robust screening of deletions in the AZF regions of the Y-chromosome. Multiplex PCRs have the advantage of including a control gene in every reaction and significantly reducing the number of reactions needed to screen the relevant genomic markers. PRINCIPAL FINDINGS: The widely established "EAA/EMQN best practice guidelines for molecular diagnosis of Y-chromosomal microdeletions (2004" were used as a basis for designing a real-time multiplex PCR system, in which the YCMD can simply be identified by their melting points. For this reason, some AZF primers were substituted by primers for regions in their genomic proximity, and the ZFX/ZFY control primer was exchanged by the AMELX/AMELY control primer. Furthermore, we substituted the classical SybrGreen I dye by the novel and high-performing DNA-binding dye EvaGreen™ and put substantial effort in titrating the primer combinations in respect to optimal melting peak separation and peak size. SIGNIFICANCE: With these changes, we were able to develop a platform-independent and robust real-time based multiplex PCR, which makes the need for amplicon identification by electrophoretic sizing expendable. By using an open-source system for real-time PCR analysis, we further demonstrate the applicability of automated melting point and YCMD detection.

  9. Amplicon-Based Pyrosequencing Reveals High Diversity of Protistan Parasites in Ships' Ballast Water: Implications for Biogeography and Infectious Diseases.

    Science.gov (United States)

    Pagenkopp Lohan, K M; Fleischer, R C; Carney, K J; Holzer, K K; Ruiz, G M

    2016-04-01

    Ships' ballast water (BW) commonly moves macroorganisms and microorganisms across the world's oceans and along coasts; however, the majority of these microbial transfers have gone undetected. We applied high-throughput sequencing methods to identify microbial eukaryotes, specifically emphasizing the protistan parasites, in ships' BW collected from vessels calling to the Chesapeake Bay (Virginia and Maryland, USA) from European and Eastern Canadian ports. We utilized tagged-amplicon 454 pyrosequencing with two general primer sets, amplifying either the V4 or V9 domain of the small subunit (SSU) of the ribosomal RNA (rRNA) gene complex, from total DNA extracted from water samples collected from the ballast tanks of bulk cargo vessels. We detected a diverse group of protistan taxa, with some known to contain important parasites in marine systems, including Apicomplexa (unidentified apicomplexans, unidentified gregarines, Cryptosporidium spp.), Dinophyta (Blastodinium spp., Euduboscquella sp., unidentified syndinids, Karlodinium spp., Syndinium spp.), Perkinsea (Parvilucifera sp.), Opisthokonta (Ichthyosporea sp., Pseudoperkinsidae, unidentified ichthyosporeans), and Stramenopiles (Labyrinthulomycetes). Further characterization of groups with parasitic taxa, consisting of phylogenetic analyses for four taxa (Cryptosporidium spp., Parvilucifera spp., Labyrinthulomycetes, and Ichthyosporea), revealed that sequences were obtained from both known and novel lineages. This study demonstrates that high-throughput sequencing is a viable and sensitive method for detecting parasitic protists when present and transported in the ballast water of ships. These data also underscore the potential importance of human-aided dispersal in the biogeography of these microbes and emerging diseases in the world's oceans. PMID:26476551

  10. Simultaneous amplicon sequencing to explore co-occurrence patterns of bacterial, archaeal and eukaryotic microorganisms in rumen microbial communities.

    Science.gov (United States)

    Kittelmann, Sandra; Seedorf, Henning; Walters, William A; Clemente, Jose C; Knight, Rob; Gordon, Jeffrey I; Janssen, Peter H

    2013-01-01

    Ruminants rely on a complex rumen microbial community to convert dietary plant material to energy-yielding products. Here we developed a method to simultaneously analyze the community's bacterial and archaeal 16S rRNA genes, ciliate 18S rRNA genes and anaerobic fungal internal transcribed spacer 1 genes using 12 DNA samples derived from 11 different rumen samples from three host species (Ovis aries, Bos taurus, Cervus elephas) and multiplex 454 Titanium pyrosequencing. We show that the mixing ratio of the group-specific DNA templates before emulsion PCR is crucial to compensate for differences in amplicon length. This method, in contrast to using a non-specific universal primer pair, avoids sequencing non-targeted DNA, such as plant- or endophyte-derived rRNA genes, and allows increased or decreased levels of community structure resolution for each microbial group as needed. Communities analyzed with different primers always grouped by sample origin rather than by the primers used. However, primer choice had a greater impact on apparent archaeal community structure than on bacterial community structure, and biases for certain methanogen groups were detected. Co-occurrence analysis of microbial taxa from all three domains of life suggested strong within- and between-domain correlations between different groups of microorganisms within the rumen. The approach used to simultaneously characterize bacterial, archaeal and eukaryotic components of a microbiota should be applicable to other communities occupying diverse habitats.

  11. A molecular-beacon-based asymmetric PCR assay for easy visualization of amplicons in the diagnosis of trichomoniasis.

    Science.gov (United States)

    Sonkar, Subash C; Sachdev, Divya; Mishra, Prashant K; Kumar, Anita; Mittal, Pratima; Saluja, Daman

    2016-12-15

    The currently available nucleic acid amplification tests (NAATs) for trichomoniasis are accurate, quick and confirmative with superior sensitivity than traditional culture-based microbiology assays. However, these assays are associated with problems of carry over contamination, false positive results, requirement of technical expertise for performance and detection of end product. Hence, a diagnostic assay with easy visualization of the amplified product will be profitable. An in-house, rapid, sensitive, specific molecular-beacon-based PCR assay, using primers against pfoB gene of Trichomonas vaginalis, was developed and evaluated using dry ectocervical swabs (n=392) from symptomatic females with vaginal discharge. Total DNA was isolated and used as template for the PCR assays. The performance and reproducibility of PCR assay was evaluated by composite reference standard (CRS). For easy visualization of the amplified product, molecular-beacon was designed and amplicons were visualized directly using fluorescent handheld dark reader or by Micro-Plate Reader. Molecular-beacons are single-stranded hairpin shaped nucleic acid probes composed of a stem, with fluorophore/quencher pair and a loop region complementary to the desired DNA. The beacon-based PCR assay designed in the present study is highly specific as confirmed by competition experiments and extremely sensitive with detection limit of 20fg of genomic DNA (3-4 pathogens). The minimum infrastructure requirement and ease to perform the assay makes this method highly useful for resource poor countries for better disease management.

  12. Factors that affect large subunit ribosomal DNA amplicon sequencing studies of fungal communities: classification method, primer choice, and error.

    Directory of Open Access Journals (Sweden)

    Teresita M Porter

    Full Text Available Nuclear large subunit ribosomal DNA is widely used in fungal phylogenetics and to an increasing extent also amplicon-based environmental sequencing. The relatively short reads produced by next-generation sequencing, however, makes primer choice and sequence error important variables for obtaining accurate taxonomic classifications. In this simulation study we tested the performance of three classification methods: 1 a similarity-based method (BLAST + Metagenomic Analyzer, MEGAN; 2 a composition-based method (Ribosomal Database Project naïve bayesian classifier, NBC; and, 3 a phylogeny-based method (Statistical Assignment Package, SAP. We also tested the effects of sequence length, primer choice, and sequence error on classification accuracy and perceived community composition. Using a leave-one-out cross validation approach, results for classifications to the genus rank were as follows: BLAST + MEGAN had the lowest error rate and was particularly robust to sequence error; SAP accuracy was highest when long LSU query sequences were classified; and, NBC runs significantly faster than the other tested methods. All methods performed poorly with the shortest 50-100 bp sequences. Increasing simulated sequence error reduced classification accuracy. Community shifts were detected due to sequence error and primer selection even though there was no change in the underlying community composition. Short read datasets from individual primers, as well as pooled datasets, appear to only approximate the true community composition. We hope this work informs investigators of some of the factors that affect the quality and interpretation of their environmental gene surveys.

  13. A molecular-beacon-based asymmetric PCR assay for easy visualization of amplicons in the diagnosis of trichomoniasis.

    Science.gov (United States)

    Sonkar, Subash C; Sachdev, Divya; Mishra, Prashant K; Kumar, Anita; Mittal, Pratima; Saluja, Daman

    2016-12-15

    The currently available nucleic acid amplification tests (NAATs) for trichomoniasis are accurate, quick and confirmative with superior sensitivity than traditional culture-based microbiology assays. However, these assays are associated with problems of carry over contamination, false positive results, requirement of technical expertise for performance and detection of end product. Hence, a diagnostic assay with easy visualization of the amplified product will be profitable. An in-house, rapid, sensitive, specific molecular-beacon-based PCR assay, using primers against pfoB gene of Trichomonas vaginalis, was developed and evaluated using dry ectocervical swabs (n=392) from symptomatic females with vaginal discharge. Total DNA was isolated and used as template for the PCR assays. The performance and reproducibility of PCR assay was evaluated by composite reference standard (CRS). For easy visualization of the amplified product, molecular-beacon was designed and amplicons were visualized directly using fluorescent handheld dark reader or by Micro-Plate Reader. Molecular-beacons are single-stranded hairpin shaped nucleic acid probes composed of a stem, with fluorophore/quencher pair and a loop region complementary to the desired DNA. The beacon-based PCR assay designed in the present study is highly specific as confirmed by competition experiments and extremely sensitive with detection limit of 20fg of genomic DNA (3-4 pathogens). The minimum infrastructure requirement and ease to perform the assay makes this method highly useful for resource poor countries for better disease management. PMID:27318568

  14. Metagenomic Analysis of Slovak Bryndza Cheese Using Next-Generation 16S rDNA Amplicon Sequencing

    Directory of Open Access Journals (Sweden)

    Planý Matej

    2016-06-01

    Full Text Available Knowledge about diversity and taxonomic structure of the microbial population present in traditional fermented foods plays a key role in starter culture selection, safety improvement and quality enhancement of the end product. Aim of this study was to investigate microbial consortia composition in Slovak bryndza cheese. For this purpose, we used culture-independent approach based on 16S rDNA amplicon sequencing using next generation sequencing platform. Results obtained by the analysis of three commercial (produced on industrial scale in winter season and one traditional (artisanal, most valued, produced in May Slovak bryndza cheese sample were compared. A diverse prokaryotic microflora composed mostly of the genera Lactococcus, Streptococcus, Lactobacillus, and Enterococcus was identified. Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris were the dominant taxons in all tested samples. Second most abundant species, detected in all bryndza cheeses, were Lactococcus fujiensis and Lactococcus taiwanensis, independently by two different approaches, using different reference 16S rRNA genes databases (Greengenes and NCBI respectively. They have been detected in bryndza cheese samples in substantial amount for the first time. The narrowest microbial diversity was observed in a sample made with a starter culture from pasteurised milk. Metagenomic analysis by high-throughput sequencing using 16S rRNA genes seems to be a powerful tool for studying the structure of the microbial population in cheeses.

  15. Combined subtractive cDNA cloning and array CGH: an efficient approach for identification of overexpressed genes in DNA amplicons

    Directory of Open Access Journals (Sweden)

    De Paepe Anne

    2004-02-01

    Full Text Available Abstract Background Activation of proto-oncogenes by DNA amplification is an important mechanism in the development and maintenance of cancer cells. Until recently, identification of the targeted genes relied on labour intensive and time consuming positional cloning methods. In this study, we outline a straightforward and efficient strategy for fast and comprehensive cloning of amplified and overexpressed genes. Results As a proof of principle, we analyzed neuroblastoma cell line IMR-32, with at least two amplification sites along the short arm of chromosome 2. In a first step, overexpressed cDNA clones were isolated using a PCR based subtractive cloning method. Subsequent deposition of these clones on a custom microarray and hybridization with IMR-32 DNA, resulted in the identification of clones that were overexpressed due to gene amplification. Using this approach, amplification of all previously reported amplified genes in this cell line was detected. Furthermore, four additional clones were found to be amplified, including the TEM8 gene on 2p13.3, two anonymous transcripts, and a fusion transcript, resulting from 2p13.3 and 2p24.3 fused sequences. Conclusions The combinatorial strategy of subtractive cDNA cloning and array CGH analysis allows comprehensive amplicon dissection, which opens perspectives for improved identification of hitherto unknown targeted oncogenes in cancer cells.

  16. Fungi Sailing the Arctic Ocean: Speciose Communities in North Atlantic Driftwood as Revealed by High-Throughput Amplicon Sequencing.

    Science.gov (United States)

    Rämä, Teppo; Davey, Marie L; Nordén, Jenni; Halvorsen, Rune; Blaalid, Rakel; Mathiassen, Geir H; Alsos, Inger G; Kauserud, Håvard

    2016-08-01

    High amounts of driftwood sail across the oceans and provide habitat for organisms tolerating the rough and saline environment. Fungi have adapted to the extremely cold and saline conditions which driftwood faces in the high north. For the first time, we applied high-throughput sequencing to fungi residing in driftwood to reveal their taxonomic richness, community composition, and ecology in the North Atlantic. Using pyrosequencing of ITS2 amplicons obtained from 49 marine logs, we found 807 fungal operational taxonomic units (OTUs) based on clustering at 97 % sequence similarity cut-off level. The phylum Ascomycota comprised 74 % of the OTUs and 20 % belonged to Basidiomycota. The richness of basidiomycetes decreased with prolonged submersion in the sea, supporting the general view of ascomycetes being more extremotolerant. However, more than one fourth of the fungal OTUs remained unassigned to any fungal class, emphasising the need for better DNA reference data from the marine habitat. Different fungal communities were detected in coniferous and deciduous logs. Our results highlight that driftwood hosts a considerably higher fungal diversity than currently known. The driftwood fungal community is not a terrestrial relic but a speciose assemblage of fungi adapted to the stressful marine environment and different kinds of wooden substrates found in it. PMID:27147245

  17. MAMA Software Features: Quantification Verification Documentation-1

    Energy Technology Data Exchange (ETDEWEB)

    Ruggiero, Christy E. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Porter, Reid B. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2014-05-21

    This document reviews the verification of the basic shape quantification attributes in the MAMA software against hand calculations in order to show that the calculations are implemented mathematically correctly and give the expected quantification results.

  18. Strategy for the maximization of clinically relevant information from hepatitis C virus, RT-PCR quantification.

    LENUS (Irish Health Repository)

    Levis, J

    2012-02-03

    BACKGROUND: The increasing clinical application of viral load assays for monitoring viral infections has been an incentive for the development of standardized tests for the hepatitis C virus. OBJECTIVE: To develop a simple model for the prediction of baseline viral load in individuals infected with the hepatitis C virus. METHODOLOGY: Viral load quantification of each patient\\'s first sample was assessed by RT-PCR-ELISA using the Roche MONITOR assay in triplicate. Genotype of the infecting virus was identified by reverse line probe hybridization, using amplicons resulting from the qualitative HCV Roche AMPLICOR assay. RESULTS: Retrospective evaluation of first quantitative values suggested that 82.4% (n=168\\/204) of individuals had a viral load between 4.3 and 6.7 log(10) viral copies per ml. A few patients (3.4%; n=7\\/204) have a serum viremia less than the lower limit of the linear range of the RT-PCR assay. Subsequent, prospective evaluation of hepatitis C viral load of all new patients using a model based on the dynamic range of viral load in the retrospective group correctly predicted the dynamic range in 75.9% (n=33\\/54). CONCLUSION: The dynamic range of hepatitis C viremia extends beyond the linear range of the Roche MONITOR assay. Accurate determination of serum viremia is substantially improved by dilution of specimens prior to quantification.

  19. FasL and FADD delivery by a glioma-specific and cell cycle-dependent HSV-1 amplicon virus enhanced apoptosis in primary human brain tumors

    Directory of Open Access Journals (Sweden)

    Lam Paula Y

    2010-10-01

    Full Text Available Abstract Background Glioblastoma multiforme is the most malignant cancer of the brain and is notoriously difficult to treat due to the highly proliferative and infiltrative nature of the cells. Herein, we explored the combination treatment of pre-established human glioma xenograft using multiple therapeutic genes whereby the gene expression is regulated by both cell-type and cell cycle-dependent transcriptional regulatory mechanism conferred by recombinant HSV-1 amplicon vectors. Results We demonstrated for the first time that Ki67-positive proliferating primary human glioma cells cultured from biopsy samples were effectively induced into cell death by the dual-specific function of the pG8-FasL amplicon vectors. These vectors were relatively stable and exhibited minimal cytotoxicity in vivo. Intracranial implantation of pre-transduced glioma cells resulted in better survival outcome when compared with viral vectors inoculated one week post-implantation of tumor cells, indicating that therapeutic efficacy is dependent on the viral spread and mode of viral vectors administration. We further showed that pG8-FasL amplicon vectors are functional in the presence of commonly used treatment regimens for human brain cancer. In fact, the combined therapies of pG8-FasL and pG8-FADD in the presence of temozolomide significantly improved the survival of mice bearing intracranial high-grade gliomas. Conclusion Taken together, our results showed that the glioma-specific and cell cycle-dependent HSV-1 amplicon vector is potentially useful as an adjuvant therapy to complement the current gene therapy strategy for gliomas.

  20. Development of a control region-based mtDNA SNaPshot™ selection tool, integrated into a mini amplicon sequencing method.

    Science.gov (United States)

    Weiler, Natalie E C; de Vries, Gerda; Sijen, Titia

    2016-03-01

    Mitochondrial DNA (mtDNA) analysis is regularly applied to forensic DNA samples with limited amounts of nuclear DNA (nDNA), such as hair shafts and bones. Generally, this mtDNA analysis involves examination of the hypervariable control region by Sanger sequencing of amplified products. When samples are severely degraded, small-sized amplicons can be applied and an earlier described mini-mtDNA method by Eichmann et al. [1] that accommodates ten mini amplicons in two multiplexes is found to be a very robust approach. However, in cases with large numbers of samples, like when searching for hairs with an mtDNA profile deviant from that of the victim, the method is time (and cost) consuming. Previously, Chemale et al. [2] described a SNaPshot™-based screening tool for a Brazilian population that uses standard-size amplicons for HVS-I and HVS-II. Here, we describe a similar tool adapted to the full control region and compatible with mini-mtDNA amplicons. Eighteen single nucleotide polymorphisms (SNPs) were selected based on their relative frequencies in a European population. They showed a high discriminatory power in a Dutch population (97.2%). The 18 SNPs are assessed in two SNaPshot™ multiplexes that pair to the two mini-mtDNA amplification multiplexes. Degenerate bases are included to limit allele dropout due to SNPs at primer binding site positions. Three SNPs provide haplogroup information. Reliability testing showed no differences with Sanger sequencing results. Since mini-mtSNaPshot screening uses only a small portion of the same PCR products used for Sanger sequencing, no additional DNA extract is consumed, which is forensically advantageous. PMID:26976467

  1. Development of a control region-based mtDNA SNaPshot™ selection tool, integrated into a mini amplicon sequencing method.

    Science.gov (United States)

    Weiler, Natalie E C; de Vries, Gerda; Sijen, Titia

    2016-03-01

    Mitochondrial DNA (mtDNA) analysis is regularly applied to forensic DNA samples with limited amounts of nuclear DNA (nDNA), such as hair shafts and bones. Generally, this mtDNA analysis involves examination of the hypervariable control region by Sanger sequencing of amplified products. When samples are severely degraded, small-sized amplicons can be applied and an earlier described mini-mtDNA method by Eichmann et al. [1] that accommodates ten mini amplicons in two multiplexes is found to be a very robust approach. However, in cases with large numbers of samples, like when searching for hairs with an mtDNA profile deviant from that of the victim, the method is time (and cost) consuming. Previously, Chemale et al. [2] described a SNaPshot™-based screening tool for a Brazilian population that uses standard-size amplicons for HVS-I and HVS-II. Here, we describe a similar tool adapted to the full control region and compatible with mini-mtDNA amplicons. Eighteen single nucleotide polymorphisms (SNPs) were selected based on their relative frequencies in a European population. They showed a high discriminatory power in a Dutch population (97.2%). The 18 SNPs are assessed in two SNaPshot™ multiplexes that pair to the two mini-mtDNA amplification multiplexes. Degenerate bases are included to limit allele dropout due to SNPs at primer binding site positions. Three SNPs provide haplogroup information. Reliability testing showed no differences with Sanger sequencing results. Since mini-mtSNaPshot screening uses only a small portion of the same PCR products used for Sanger sequencing, no additional DNA extract is consumed, which is forensically advantageous.

  2. Very high resolution single pass HLA genotyping using amplicon sequencing on the 454 next generation DNA sequencers: Comparison with Sanger sequencing.

    Science.gov (United States)

    Yamamoto, F; Höglund, B; Fernandez-Vina, M; Tyan, D; Rastrou, M; Williams, T; Moonsamy, P; Goodridge, D; Anderson, M; Erlich, H A; Holcomb, C L

    2015-12-01

    Compared to Sanger sequencing, next-generation sequencing offers advantages for high resolution HLA genotyping including increased throughput, lower cost, and reduced genotype ambiguity. Here we describe an enhancement of the Roche 454 GS GType HLA genotyping assay to provide very high resolution (VHR) typing, by the addition of 8 primer pairs to the original 14, to genotype 11 HLA loci. These additional amplicons help resolve common and well-documented alleles and exclude commonly found null alleles in genotype ambiguity strings. Simplification of workflow to reduce the initial preparation effort using early pooling of amplicons or the Fluidigm Access Array™ is also described. Performance of the VHR assay was evaluated on 28 well characterized cell lines using Conexio Assign MPS software which uses genomic, rather than cDNA, reference sequence. Concordance was 98.4%; 1.6% had no genotype assignment. Of concordant calls, 53% were unambiguous. To further assess the assay, 59 clinical samples were genotyped and results compared to unambiguous allele assignments obtained by prior sequence-based typing supplemented with SSO and/or SSP. Concordance was 98.7% with 58.2% as unambiguous calls; 1.3% could not be assigned. Our results show that the amplicon-based VHR assay is robust and can replace current Sanger methodology. Together with software enhancements, it has the potential to provide even higher resolution HLA typing. PMID:26037172

  3. Nuclear species-diagnostic SNP markers mined from 454 amplicon sequencing reveal admixture genomic structure of modern citrus varieties.

    Directory of Open Access Journals (Sweden)

    Franck Curk

    Full Text Available Most cultivated Citrus species originated from interspecific hybridisation between four ancestral taxa (C. reticulata, C. maxima, C. medica, and C. micrantha with limited further interspecific recombination due to vegetative propagation. This evolution resulted in admixture genomes with frequent interspecific heterozygosity. Moreover, a major part of the phenotypic diversity of edible citrus results from the initial differentiation between these taxa. Deciphering the phylogenomic structure of citrus germplasm is therefore essential for an efficient utilization of citrus biodiversity in breeding schemes. The objective of this work was to develop a set of species-diagnostic single nucleotide polymorphism (SNP markers for the four Citrus ancestral taxa covering the nine chromosomes, and to use these markers to infer the phylogenomic structure of secondary species and modern cultivars. Species-diagnostic SNPs were mined from 454 amplicon sequencing of 57 gene fragments from 26 genotypes of the four basic taxa. Of the 1,053 SNPs mined from 28,507 kb sequence, 273 were found to be highly diagnostic for a single basic taxon. Species-diagnostic SNP markers (105 were used to analyse the admixture structure of varieties and rootstocks. This revealed C. maxima introgressions in most of the old and in all recent selections of mandarins, and suggested that C. reticulata × C. maxima reticulation and introgression processes were important in edible mandarin domestication. The large range of phylogenomic constitutions between C. reticulata and C. maxima revealed in mandarins, tangelos, tangors, sweet oranges, sour oranges, grapefruits, and orangelos is favourable for genetic association studies based on phylogenomic structures of the germplasm. Inferred admixture structures were in agreement with previous hypotheses regarding the origin of several secondary species and also revealed the probable origin of several acid citrus varieties. The developed species

  4. Nuclear species-diagnostic SNP markers mined from 454 amplicon sequencing reveal admixture genomic structure of modern citrus varieties.

    Science.gov (United States)

    Curk, Franck; Ancillo, Gema; Ollitrault, Frédérique; Perrier, Xavier; Jacquemoud-Collet, Jean-Pierre; Garcia-Lor, Andres; Navarro, Luis; Ollitrault, Patrick

    2015-01-01

    Most cultivated Citrus species originated from interspecific hybridisation between four ancestral taxa (C. reticulata, C. maxima, C. medica, and C. micrantha) with limited further interspecific recombination due to vegetative propagation. This evolution resulted in admixture genomes with frequent interspecific heterozygosity. Moreover, a major part of the phenotypic diversity of edible citrus results from the initial differentiation between these taxa. Deciphering the phylogenomic structure of citrus germplasm is therefore essential for an efficient utilization of citrus biodiversity in breeding schemes. The objective of this work was to develop a set of species-diagnostic single nucleotide polymorphism (SNP) markers for the four Citrus ancestral taxa covering the nine chromosomes, and to use these markers to infer the phylogenomic structure of secondary species and modern cultivars. Species-diagnostic SNPs were mined from 454 amplicon sequencing of 57 gene fragments from 26 genotypes of the four basic taxa. Of the 1,053 SNPs mined from 28,507 kb sequence, 273 were found to be highly diagnostic for a single basic taxon. Species-diagnostic SNP markers (105) were used to analyse the admixture structure of varieties and rootstocks. This revealed C. maxima introgressions in most of the old and in all recent selections of mandarins, and suggested that C. reticulata × C. maxima reticulation and introgression processes were important in edible mandarin domestication. The large range of phylogenomic constitutions between C. reticulata and C. maxima revealed in mandarins, tangelos, tangors, sweet oranges, sour oranges, grapefruits, and orangelos is favourable for genetic association studies based on phylogenomic structures of the germplasm. Inferred admixture structures were in agreement with previous hypotheses regarding the origin of several secondary species and also revealed the probable origin of several acid citrus varieties. The developed species-diagnostic SNP

  5. Uncultured bacterial diversity in a seawater recirculating aquaculture system revealed by 16S rRNA gene amplicon sequencing.

    Science.gov (United States)

    Lee, Da-Eun; Lee, Jinhwan; Kim, Young-Mog; Myeong, Jeong-In; Kim, Kyoung-Ho

    2016-04-01

    Bacterial diversity in a seawater recirculating aquaculture system (RAS) was investigated using 16S rRNA amplicon sequencing to understand the roles of bacterial communities in the system. The RAS was operated at nine different combinations of temperature (15°C, 20°C, and 25°C) and salinity (20‰, 25‰, and 32.5‰). Samples were collected from five or six RAS tanks (biofilters) for each condition. Fifty samples were analyzed. Proteobacteria and Bacteroidetes were most common (sum of both phyla: 67.2% to 99.4%) and were inversely proportional to each other. Bacteria that were present at an average of ≥ 1% included Actinobacteria (2.9%) Planctomycetes (2.0%), Nitrospirae (1.5%), and Acidobacteria (1.0%); they were preferentially present in packed bed biofilters, mesh biofilters, and maturation biofilters. The three biofilters showed higher diversity than other RAS tanks (aerated biofilters, floating bed biofilters, and fish tanks) from phylum to operational taxonomic unit (OTU) level. Samples were clustered into several groups based on the bacterial communities. Major taxonomic groups related to family Rhodobacteraceae and Flavobacteriaceae were distributed widely in the samples. Several taxonomic groups like [Saprospiraceae], Cytophagaceae, Octadecabacter, and Marivita showed a cluster-oriented distribution. Phaeobacter and Sediminicola-related reads were detected frequently and abundantly at low temperature. Nitrifying bacteria were detected frequently and abundantly in the three biofilters. Phylogenetic analysis of the nitrifying bacteria showed several similar OTUs were observed widely through the biofilters. The diverse bacterial communities and the minor taxonomic groups, except for Proteobacteria and Bacteroidetes, seemed to play important roles and seemed necessary for nitrifying activity in the RAS, especially in packed bed biofilters, mesh biofilters, and maturation biofilters.

  6. Classification of pmoA amplicon pyrosequences using BLAST and the lowest common ancestor method in MEGAN

    Directory of Open Access Journals (Sweden)

    Marc Gregory Dumont

    2014-02-01

    Full Text Available The classification of high-throughput sequencing data of protein-encoding genes is not as well established as for 16S rRNA. The objective of this work was to develop a simple and accurate method of classifying large datasets of pmoA sequences, a common marker for methanotrophic bacteria. A taxonomic system for pmoA was developed based on a phylogenetic analysis of available sequences. The taxonomy incorporates the known diversity of pmoA present in public databases, including both sequences from cultivated and uncultivated organisms. Representative sequences from closely related genes, such as those encoding the bacterial ammonia monooxygenase, were also included in the pmoA taxonomy. In total, 53 low-level taxa (genus-level are included. Using previously published datasets of high-throughput pmoA amplicon sequence data, we tested two approaches for classifying pmoA: a naïve Bayesian classifier and BLAST. Classification of pmoA sequences based on BLAST analyses was performed using the lowest common ancestor (LCA algorithm in MEGAN, a software program commonly used for the analysis of metagenomic data. Both the naïve Bayesian and BLAST methods were able to classify pmoA sequences and provided similar classifications; however, the naïve Bayesian classifier was prone to misclassifying contaminant sequences present in the datasets. Another advantage of the BLAST/LCA method was that it provided a user-interpretable output and enabled novelty detection at various levels, from highly divergent pmoA sequences to genus-level novelty.  

  7. Nuclear species-diagnostic SNP markers mined from 454 amplicon sequencing reveal admixture genomic structure of modern citrus varieties.

    Science.gov (United States)

    Curk, Franck; Ancillo, Gema; Ollitrault, Frédérique; Perrier, Xavier; Jacquemoud-Collet, Jean-Pierre; Garcia-Lor, Andres; Navarro, Luis; Ollitrault, Patrick

    2015-01-01

    Most cultivated Citrus species originated from interspecific hybridisation between four ancestral taxa (C. reticulata, C. maxima, C. medica, and C. micrantha) with limited further interspecific recombination due to vegetative propagation. This evolution resulted in admixture genomes with frequent interspecific heterozygosity. Moreover, a major part of the phenotypic diversity of edible citrus results from the initial differentiation between these taxa. Deciphering the phylogenomic structure of citrus germplasm is therefore essential for an efficient utilization of citrus biodiversity in breeding schemes. The objective of this work was to develop a set of species-diagnostic single nucleotide polymorphism (SNP) markers for the four Citrus ancestral taxa covering the nine chromosomes, and to use these markers to infer the phylogenomic structure of secondary species and modern cultivars. Species-diagnostic SNPs were mined from 454 amplicon sequencing of 57 gene fragments from 26 genotypes of the four basic taxa. Of the 1,053 SNPs mined from 28,507 kb sequence, 273 were found to be highly diagnostic for a single basic taxon. Species-diagnostic SNP markers (105) were used to analyse the admixture structure of varieties and rootstocks. This revealed C. maxima introgressions in most of the old and in all recent selections of mandarins, and suggested that C. reticulata × C. maxima reticulation and introgression processes were important in edible mandarin domestication. The large range of phylogenomic constitutions between C. reticulata and C. maxima revealed in mandarins, tangelos, tangors, sweet oranges, sour oranges, grapefruits, and orangelos is favourable for genetic association studies based on phylogenomic structures of the germplasm. Inferred admixture structures were in agreement with previous hypotheses regarding the origin of several secondary species and also revealed the probable origin of several acid citrus varieties. The developed species-diagnostic SNP

  8. Responses of soil microeukaryotic communities to short-term fumigation-incubation revealed by MiSeq amplicon sequencing

    Directory of Open Access Journals (Sweden)

    Lin eChen

    2015-10-01

    Full Text Available In soil microbiology, there is a ‘paradox’ of soil organic carbon (SOC mineralization, which is that even though chloroform fumigation destroys majority of the soil microbial biomass, SOC mineralization continues at the same rate as in the non-fumigated soil during the incubation period. Soil microeukaryotes as important SOC decomposers, however, their community-level responses to chloroform fumigation are not well understood. Using the 18S rRNA gene amplicon sequencing, we analyzed the composition, diversity and C-metabolic functions of a grassland soil and an arable soil microeukaryotic community in response to fumigation followed by a 30-day incubation. The grassland and arable soil microeukaryotic communities were dominated by the fungal Ascomycota (80.5–93.1% of the fungal sequences, followed by the protistan Cercozoa and Apicomplexa. In the arable soil fungal community, the predominance of the class Sordariomycetes was replaced by the class Eurotiomycetes after fumigation at days 7 and 30 of the incubation. Fumigation changed the microeukaryotic α-diversity in the grassland soil at days 0 and 7, and β-diversity in the arable soil at days 7 and 30. Network analysis indicated that after fumigation fungi were important groups closely related to other taxa. Most phylotypes (especially Sordariomycetes, Dothideomycetes, Coccidia and uncultured Chytridiomycota were inhibited, and only a few were positively stimulated by fumigation. Despite the inhibited Sordariomycetes, the fumigated communities mainly consisted of Eurotiomycetes and Sordariomycetes (21.9% and 36.5% relative frequency, respectively, which are able to produce hydrolytic enzymes associated with SOC mineralization. Our study suggests that fumigation not only decreases biomass size, but modulates the composition and diversity of the soil microeukaryotic communities, which are capable of driving SOC mineralization by release of hydrolytic enzymes during short-term fumigation-incubation.

  9. Uncultured bacterial diversity in a seawater recirculating aquaculture system revealed by 16S rRNA gene amplicon sequencing.

    Science.gov (United States)

    Lee, Da-Eun; Lee, Jinhwan; Kim, Young-Mog; Myeong, Jeong-In; Kim, Kyoung-Ho

    2016-04-01

    Bacterial diversity in a seawater recirculating aquaculture system (RAS) was investigated using 16S rRNA amplicon sequencing to understand the roles of bacterial communities in the system. The RAS was operated at nine different combinations of temperature (15°C, 20°C, and 25°C) and salinity (20‰, 25‰, and 32.5‰). Samples were collected from five or six RAS tanks (biofilters) for each condition. Fifty samples were analyzed. Proteobacteria and Bacteroidetes were most common (sum of both phyla: 67.2% to 99.4%) and were inversely proportional to each other. Bacteria that were present at an average of ≥ 1% included Actinobacteria (2.9%) Planctomycetes (2.0%), Nitrospirae (1.5%), and Acidobacteria (1.0%); they were preferentially present in packed bed biofilters, mesh biofilters, and maturation biofilters. The three biofilters showed higher diversity than other RAS tanks (aerated biofilters, floating bed biofilters, and fish tanks) from phylum to operational taxonomic unit (OTU) level. Samples were clustered into several groups based on the bacterial communities. Major taxonomic groups related to family Rhodobacteraceae and Flavobacteriaceae were distributed widely in the samples. Several taxonomic groups like [Saprospiraceae], Cytophagaceae, Octadecabacter, and Marivita showed a cluster-oriented distribution. Phaeobacter and Sediminicola-related reads were detected frequently and abundantly at low temperature. Nitrifying bacteria were detected frequently and abundantly in the three biofilters. Phylogenetic analysis of the nitrifying bacteria showed several similar OTUs were observed widely through the biofilters. The diverse bacterial communities and the minor taxonomic groups, except for Proteobacteria and Bacteroidetes, seemed to play important roles and seemed necessary for nitrifying activity in the RAS, especially in packed bed biofilters, mesh biofilters, and maturation biofilters. PMID:27033205

  10. Haplotyping and copy number estimation of the highly polymorphic human beta-defensin locus on 8p23 by 454 amplicon sequencing

    Directory of Open Access Journals (Sweden)

    Rosenstiel Philip

    2010-04-01

    Full Text Available Abstract Background The beta-defensin gene cluster (DEFB at chromosome 8p23.1 is one of the most copy number (CN variable regions of the human genome. Whereas individual DEFB CNs have been suggested as independent genetic risk factors for several diseases (e.g. psoriasis and Crohn's disease, the role of multisite sequence variations (MSV is less well understood and to date has only been reported for prostate cancer. Simultaneous assessment of MSVs and CNs can be achieved by PCR, cloning and Sanger sequencing, however, these methods are labour and cost intensive as well as prone to methodological bias introduced by bacterial cloning. Here, we demonstrate that amplicon sequencing of pooled individual PCR products by the 454 technology allows in-depth determination of MSV haplotypes and estimation of DEFB CNs in parallel. Results Six PCR products spread over ~87 kb of DEFB and harbouring 24 known MSVs were amplified from 11 DNA samples, pooled and sequenced on a Roche 454 GS FLX sequencer. From ~142,000 reads, ~120,000 haplotype calls (HC were inferred that identified 22 haplotypes ranging from 2 to 7 per amplicon. In addition to the 24 known MSVs, two additional sequence variations were detected. Minimal CNs were estimated from the ratio of HCs and compared to absolute CNs determined by alternative methods. Concordance in CNs was found for 7 samples, the CNs differed by one in 2 samples and the estimated minimal CN was half of the absolute in one sample. For 7 samples and 2 amplicons, the 454 haplotyping results were compared to those by cloning/Sanger sequencing. Intrinsic problems related to chimera formation during PCR and differences between haplotyping by 454 and cloning/Sanger sequencing are discussed. Conclusion Deep amplicon sequencing using the 454 technology yield thousands of HCs per amplicon for an affordable price and may represent an effective method for parallel haplotyping and CN estimation in small to medium-sized cohorts. The

  11. Risk Quantification and Evaluation Modelling

    OpenAIRE

    Manmohan Singh; M.D. Jaybhaye; S. K. Basu

    2014-01-01

    In this paper authors have discussed risk quantification methods and evaluation of risks and decision parameter to be used for deciding on ranking of the critical items, for prioritization of condition monitoring based risk and reliability centered maintenance (CBRRCM). As time passes any equipment or any product degrades into lower effectiveness and the rate of failure or malfunctioning increases, thereby lowering the reliability. Thus with the passage of time or a number of active tests or ...

  12. qPCR-based mitochondrial DNA quantification: Influence of template DNA fragmentation on accuracy

    Energy Technology Data Exchange (ETDEWEB)

    Jackson, Christopher B., E-mail: Christopher.jackson@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland); Gallati, Sabina, E-mail: sabina.gallati@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland); Schaller, Andre, E-mail: andre.schaller@insel.ch [Division of Human Genetics, Departements of Pediatrics and Clinical Research, Inselspital, University of Berne, Freiburgstrasse, CH-3010 Berne (Switzerland)

    2012-07-06

    Highlights: Black-Right-Pointing-Pointer Serial qPCR accurately determines fragmentation state of any given DNA sample. Black-Right-Pointing-Pointer Serial qPCR demonstrates different preservation of the nuclear and mitochondrial genome. Black-Right-Pointing-Pointer Serial qPCR provides a diagnostic tool to validate the integrity of bioptic material. Black-Right-Pointing-Pointer Serial qPCR excludes degradation-induced erroneous quantification. -- Abstract: Real-time PCR (qPCR) is the method of choice for quantification of mitochondrial DNA (mtDNA) by relative comparison of a nuclear to a mitochondrial locus. Quantitative abnormal mtDNA content is indicative of mitochondrial disorders and mostly confines in a tissue-specific manner. Thus handling of degradation-prone bioptic material is inevitable. We established a serial qPCR assay based on increasing amplicon size to measure degradation status of any DNA sample. Using this approach we can exclude erroneous mtDNA quantification due to degraded samples (e.g. long post-exicision time, autolytic processus, freeze-thaw cycles) and ensure abnormal DNA content measurements (e.g. depletion) in non-degraded patient material. By preparation of degraded DNA under controlled conditions using sonification and DNaseI digestion we show that erroneous quantification is due to the different preservation qualities of the nuclear and the mitochondrial genome. This disparate degradation of the two genomes results in over- or underestimation of mtDNA copy number in degraded samples. Moreover, as analysis of defined archival tissue would allow to precise the molecular pathomechanism of mitochondrial disorders presenting with abnormal mtDNA content, we compared fresh frozen (FF) with formalin-fixed paraffin-embedded (FFPE) skeletal muscle tissue of the same sample. By extrapolation of measured decay constants for nuclear DNA ({lambda}{sub nDNA}) and mtDNA ({lambda}{sub mtDNA}) we present an approach to possibly correct measurements in

  13. Polymicrobial nature of chronic diabetic foot ulcer biofilm infections determined using bacterial tag encoded FLX amplicon pyrosequencing (bTEFAP.

    Directory of Open Access Journals (Sweden)

    Scot E Dowd

    Full Text Available BACKGROUND: Diabetic extremity ulcers are associated with chronic infections. Such ulcer infections are too often followed by amputation because there is little or no understanding of the ecology of such infections or how to control or eliminate this type of chronic infection. A primary impediment to the healing of chronic wounds is biofilm phenotype infections. Diabetic foot ulcers are the most common, disabling, and costly complications of diabetes. Here we seek to derive a better understanding of the polymicrobial nature of chronic diabetic extremity ulcer infections. METHODS AND FINDINGS: Using a new bacterial tag encoded FLX amplicon pyrosequencing (bTEFAP approach we have evaluated the bacterial diversity of 40 chronic diabetic foot ulcers from different patients. The most prevalent bacterial genus associated with diabetic chronic wounds was Corynebacterium spp. Findings also show that obligate anaerobes including Bacteroides, Peptoniphilus, Fingoldia, Anaerococcus, and Peptostreptococcus spp. are ubiquitous in diabetic ulcers, comprising a significant portion of the wound biofilm communities. Other major components of the bacterial communities included commonly cultured genera such as Streptococcus, Serratia, Staphylococcus and Enterococcus spp. CONCLUSIONS: In this article, we highlight the patterns of population diversity observed in the samples and introduce preliminary evidence to support the concept of functional equivalent pathogroups (FEP. Here we introduce FEP as consortia of genotypically distinct bacteria that symbiotically produce a pathogenic community. According to this hypothesis, individual members of these communities when they occur alone may not cause disease but when they coaggregate or consort together into a FEP the synergistic effect provides the functional equivalence of well-known pathogens, such as Staphylococcus aureus, giving the biofilm community the factors necessary to maintain chronic biofilm infections

  14. Gastrointestinal Bacterial and Methanogenic Archaea Diversity Dynamics Associated with Condensed Tannin-Containing Pine Bark Diet in Goats Using 16S rDNA Amplicon Pyrosequencing

    OpenAIRE

    Min, Byeng R.; Sandra Solaiman; Raymon Shange; Jong-Su Eun

    2014-01-01

    Eighteen Kiko-cross meat goats (n=6) were used to collect gastrointestinal (GI) bacteria and methanogenic archaea for diversity measures when fed condensed tannin-containing pine bark (PB). Three dietary treatments were tested: control diet (0% PB and 30% wheat straw (WS); 0.17% condensed tannins (CT) dry matter (DM)); 15% PB and 15% WS (1.6% CT DM), and 30% PB and 0% WS (3.2% CT DM). A 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing technique was used to characterize and elucida...

  15. Quantification of wastewater sludge dewatering.

    Science.gov (United States)

    Skinner, Samuel J; Studer, Lindsay J; Dixon, David R; Hillis, Peter; Rees, Catherine A; Wall, Rachael C; Cavalida, Raul G; Usher, Shane P; Stickland, Anthony D; Scales, Peter J

    2015-10-01

    Quantification and comparison of the dewatering characteristics of fifteen sewage sludges from a range of digestion scenarios are described. The method proposed uses laboratory dewatering measurements and integrity analysis of the extracted material properties. These properties were used as inputs into a model of filtration, the output of which provides the dewatering comparison. This method is shown to be necessary for quantification and comparison of dewaterability as the permeability and compressibility of the sludges varies by up to ten orders of magnitude in the range of solids concentration of interest to industry. This causes a high sensitivity of the dewaterability comparison to the starting concentration of laboratory tests, thus simple dewaterability comparison based on parameters such as the specific resistance to filtration is difficult. The new approach is demonstrated to be robust relative to traditional methods such as specific resistance to filtration analysis and has an in-built integrity check. Comparison of the quantified dewaterability of the fifteen sludges to the relative volatile solids content showed a very strong correlation in the volatile solids range from 40 to 80%. The data indicate that the volatile solids parameter is a strong indicator of the dewatering behaviour of sewage sludges. PMID:26003332

  16. Detection and Quantification of Neurotransmitters in Dialysates

    OpenAIRE

    Zapata, Agustin; Chefer, Vladimir I.; Shippenberg, Toni S.; Denoroy, Luc

    2009-01-01

    Sensitive analytical methods are needed for the separation and quantification of neurotransmitters obtained in microdialysate studies. This unit describes methods that permit quantification of nanomolar concentrations of monoamines and their metabolites (high-pressure liquid chromatography electrochemical detection), acetylcholine (HPLC-coupled to an enzyme reactor), and amino acids (HPLC-fluorescence detection; capillary electrophoresis with laser-induced fluorescence detection).

  17. Conservative fragments in bacterial 16S rRNA genes and primer design for 16S ribosomal DNA amplicons in metagenomic studies

    KAUST Repository

    Wang, Yong

    2009-10-09

    Bacterial 16S ribosomal DNA (rDNA) amplicons have been widely used in the classification of uncultured bacteria inhabiting environmental niches. Primers targeting conservative regions of the rDNAs are used to generate amplicons of variant regions that are informative in taxonomic assignment. One problem is that the percentage coverage and application scope of the primers used in previous studies are largely unknown. In this study, conservative fragments of available rDNA sequences were first mined and then used to search for candidate primers within the fragments by measuring the coverage rate defined as the percentage of bacterial sequences containing the target. Thirty predicted primers with a high coverage rate (>90%) were identified, which were basically located in the same conservative regions as known primers in previous reports, whereas 30% of the known primers were associated with a coverage rate of <90%. The application scope of the primers was also examined by calculating the percentages of failed detections in bacterial phyla. Primers A519-539, E969- 983, E1063-1081, U515 and E517, are highly recommended because of their high coverage in almost all phyla. As expected, the three predominant phyla, Firmicutes, Gemmatimonadetes and Proteobacteria, are best covered by the predicted primers. The primers recommended in this report shall facilitate a comprehensive and reliable survey of bacterial diversity in metagenomic studies. © 2009 Wang, Qian.

  18. Development of Quantification Method for Bioluminescence Imaging

    International Nuclear Information System (INIS)

    Optical molecular luminescence imaging is widely used for detection and imaging of bio-photons emitted by luminescent luciferase activation. The measured photons in this method provide the degree of molecular alteration or cell numbers with the advantage of high signal-to-noise ratio. To extract useful information from the measured results, the analysis based on a proper quantification method is necessary. In this research, we propose a quantification method presenting linear response of measured light signal to measurement time. We detected the luminescence signal by using lab-made optical imaging equipment of animal light imaging system (ALIS) and different two kinds of light sources. One is three bacterial light-emitting sources containing different number of bacteria. The other is three different non-bacterial light sources emitting very weak light. By using the concept of the candela and the flux, we could derive simplified linear quantification formula. After experimentally measuring light intensity, the data was processed with the proposed quantification function. We could obtain linear response of photon counts to measurement time by applying the pre-determined quantification function. The ratio of the re-calculated photon counts and measurement time present a constant value although different light source was applied. The quantification function for linear response could be applicable to the standard quantification process. The proposed method could be used for the exact quantitative analysis in various light imaging equipment with presenting linear response behavior of constant light emitting sources to measurement time

  19. Advancing agricultural greenhouse gas quantification*

    Science.gov (United States)

    Olander, Lydia; Wollenberg, Eva; Tubiello, Francesco; Herold, Martin

    2013-03-01

    1. Introduction Better information on greenhouse gas (GHG) emissions and mitigation potential in the agricultural sector is necessary to manage these emissions and identify responses that are consistent with the food security and economic development priorities of countries. Critical activity data (what crops or livestock are managed in what way) are poor or lacking for many agricultural systems, especially in developing countries. In addition, the currently available methods for quantifying emissions and mitigation are often too expensive or complex or not sufficiently user friendly for widespread use. The purpose of this focus issue is to capture the state of the art in quantifying greenhouse gases from agricultural systems, with the goal of better understanding our current capabilities and near-term potential for improvement, with particular attention to quantification issues relevant to smallholders in developing countries. This work is timely in light of international discussions and negotiations around how agriculture should be included in efforts to reduce and adapt to climate change impacts, and considering that significant climate financing to developing countries in post-2012 agreements may be linked to their increased ability to identify and report GHG emissions (Murphy et al 2010, CCAFS 2011, FAO 2011). 2. Agriculture and climate change mitigation The main agricultural GHGs—methane and nitrous oxide—account for 10%-12% of anthropogenic emissions globally (Smith et al 2008), or around 50% and 60% of total anthropogenic methane and nitrous oxide emissions, respectively, in 2005. Net carbon dioxide fluxes between agricultural land and the atmosphere linked to food production are relatively small, although significant carbon emissions are associated with degradation of organic soils for plantations in tropical regions (Smith et al 2007, FAO 2012). Population growth and shifts in dietary patterns toward more meat and dairy consumption will lead to

  20. A new amplicon based approach of whole mitogenome sequencing for phylogenetic and phylogeographic analysis: An example of East African white-eyes (Aves, Zosteropidae).

    Science.gov (United States)

    Meimberg, Harald; Schachtler, Christina; Curto, Manuel; Husemann, Martin; Habel, Jan Christian

    2016-09-01

    Classical Sanger sequencing is still frequently used to generate sequence data for phylogenetic and phylogeographic inference. In this contribution we present a novel approach to genotype whole mitogenomic haplotypes using Illumina MiSeq reads from indexed amplicons. Our new approach reduces preparation time by multiplexing loci within a single or few PCR reactions and by plate format library construction. The use of paired-end reads allows covering amplicons of about 0.5kb and thus no nebulisation and assembly are necessary. We tested the power and effectiveness of this technique by analysing the mitogenomic diversity of East African white-eye bird species (Zosteropidae), a taxonomically highly diverse and complex species flock found in various ecosystems spread across major parts of Africa. We compare the newly generated mitogenomic data set with published data of three mitochondrial genes for a similar set of populations and taxa. The comparison demonstrates that our new procedure represents a cost effective use of NGS for medium throughput phylogenetic analyses. Using this method, we were able to increase the amount of phylogenetic information significantly, while reducing the costs and effort in the laboratory. The mitogenomic data show a higher resolution than previous studies providing higher support and new insights in the relationships of Zosterops species. Our data suggest to split Z. poliogaster into four distinct species, three of which had previously been proposed: Z. silvanus, Z. mbulensis, Z. kikyuensis and Z. kulalensis. Our approach allows the genotyping of whole mitogenomes for a large number of individuals and thus allows more reliable reconstruction of phylogenetic and phylogeographic relationships - also for non-model organisms. PMID:27233440

  1. Uncertainty quantification theory, implementation, and applications

    CERN Document Server

    Smith, Ralph C

    2014-01-01

    The field of uncertainty quantification is evolving rapidly because of increasing emphasis on models that require quantified uncertainties for large-scale applications, novel algorithm development, and new computational architectures that facilitate implementation of these algorithms. Uncertainty Quantification: Theory, Implementation, and Applications provides readers with the basic concepts, theory, and algorithms necessary to quantify input and response uncertainties for simulation models arising in a broad range of disciplines. The book begins with a detailed discussion of applications where uncertainty quantification is critical for both scientific understanding and policy. It then covers concepts from probability and statistics, parameter selection techniques, frequentist and Bayesian model calibration, propagation of uncertainties, quantification of model discrepancy, surrogate model construction, and local and global sensitivity analysis. The author maintains a complementary web page where readers ca...

  2. Uncertainty Quantification in Aerodynamics Simulations Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The objective of the proposed work (Phases I and II) is to develop uncertainty quantification methodologies and software suitable for use in CFD simulations of...

  3. MAMA Software Features: Visual Examples of Quantification

    Energy Technology Data Exchange (ETDEWEB)

    Ruggiero, Christy E. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Porter, Reid B. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2014-05-20

    This document shows examples of the results from quantifying objects of certain sizes and types in the software. It is intended to give users a better feel for some of the quantification calculations, and, more importantly, to help users understand the challenges with using a small set of ‘shape’ quantification calculations for objects that can vary widely in shapes and features. We will add more examples to this in the coming year.

  4. Risk Quantification and Evaluation Modelling

    Directory of Open Access Journals (Sweden)

    Manmohan Singh

    2014-07-01

    Full Text Available In this paper authors have discussed risk quantification methods and evaluation of risks and decision parameter to be used for deciding on ranking of the critical items, for prioritization of condition monitoring based risk and reliability centered maintenance (CBRRCM. As time passes any equipment or any product degrades into lower effectiveness and the rate of failure or malfunctioning increases, thereby lowering the reliability. Thus with the passage of time or a number of active tests or periods of work, the reliability of the product or the system, may fall down to a low value known as a threshold value, below which the reliability should not be allowed to dip. Hence, it is necessary to fix up the normal basis for determining the appropriate points in the product life cycle where predictive preventive maintenance may be applied in the programme so that the reliability (the probability of successful functioning can be enhanced, preferably to its original value, by reducing the failure rate and increasing the mean time between failure. It is very important for defence application where reliability is a prime work. An attempt is made to develop mathematical model for risk assessment and ranking them. Based on likeliness coefficient β1 and risk coefficient β2 ranking of the sub-systems can be modelled and used for CBRRCM.Defence Science Journal, Vol. 64, No. 4, July 2014, pp. 378-384, DOI:http://dx.doi.org/10.14429/dsj.64.6366 

  5. Lung involvement quantification in chest radiographs

    International Nuclear Information System (INIS)

    Tuberculosis (TB) caused by Mycobacterium tuberculosis, is an infectious disease which remains a global health problem. The chest radiography is the commonly method employed to assess the TB's evolution. The methods for quantification of abnormalities of chest are usually performed on CT scans (CT). This quantification is important to assess the TB evolution and treatment and comparing different treatments. However, precise quantification is not feasible for the amount of CT scans required. The purpose of this work is to develop a methodology for quantification of lung damage caused by TB through chest radiographs. It was developed an algorithm for computational processing of exams in Matlab, which creates a lungs' 3D representation, with compromised dilated regions inside. The quantification of lung lesions was also made for the same patients through CT scans. The measurements from the two methods were compared and resulting in strong correlation. Applying statistical Bland and Altman, all samples were within the limits of agreement, with a confidence interval of 95%. The results showed an average variation of around 13% between the two quantification methods. The results suggest the effectiveness and applicability of the method developed, providing better risk-benefit to the patient and cost-benefit ratio for the institution. (author)

  6. Uncertainty Quantification for Safeguards Measurements

    International Nuclear Information System (INIS)

    Part of the scientific method requires all calculated and measured results to be accompanied by a description that meets user needs and provides an adequate statement of the confidence one can have in the results. The scientific art of generating quantitative uncertainty statements is closely related to the mathematical disciplines of applied statistics, sensitivity analysis, optimization, and inversion, but in the field of non-destructive assay, also often draws heavily on expert judgment based on experience. We call this process uncertainty quantification, (UQ). Philosophical approaches to UQ along with the formal tools available for UQ have advanced considerably over recent years and these advances, we feel, may be useful to include in the analysis of data gathered from safeguards instruments. This paper sets out what we hope to achieve during a three year US DOE NNSA research project recently launched to address the potential of advanced UQ to improve safeguards conclusions. By way of illustration we discuss measurement of uranium enrichment by the enrichment meter principle (also known as the infinite thickness technique), that relies on gamma counts near the 186 keV peak directly from 235U. This method has strong foundations in fundamental physics and so we have a basis for the choice of response model — although in some implementations, peak area extraction may result in a bias when applied over a wide dynamic range. It also allows us to describe a common but usually neglected aspect of applying a calibration curve, namely the error structure in the predictors. We illustrate this using a combination of measured data and simulation. (author)

  7. Separation and quantification of microalgal carbohydrates.

    Science.gov (United States)

    Templeton, David W; Quinn, Matthew; Van Wychen, Stefanie; Hyman, Deborah; Laurens, Lieve M L

    2012-12-28

    Structural carbohydrates can constitute a large fraction of the dry weight of algal biomass and thus accurate identification and quantification is important for summative mass closure. Two limitations to the accurate characterization of microalgal carbohydrates are the lack of a robust analytical procedure to hydrolyze polymeric carbohydrates to their respective monomers and the subsequent identification and quantification of those monosaccharides. We address the second limitation, chromatographic separation of monosaccharides, here by identifying optimum conditions for the resolution of a synthetic mixture of 13 microalgae-specific monosaccharides, comprised of 8 neutral, 2 amino sugars, 2 uronic acids and 1 alditol (myo-inositol as an internal standard). The synthetic 13-carbohydrate mix showed incomplete resolution across 11 traditional high performance liquid chromatography (HPLC) methods, but showed improved resolution and accurate quantification using anion exchange chromatography (HPAEC) as well as alditol acetate derivatization followed by gas chromatography (for the neutral- and amino-sugars only). We demonstrate the application of monosaccharide quantification using optimized chromatography conditions after sulfuric acid analytical hydrolysis for three model algae strains and compare the quantification and complexity of monosaccharides in analytical hydrolysates relative to a typical terrestrial feedstock, sugarcane bagasse. PMID:23177152

  8. Comparison of five DNA quantification methods

    DEFF Research Database (Denmark)

    Nielsen, Karsten; Mogensen, Helle Smidt; Hedman, Johannes;

    2008-01-01

    Six commercial preparations of human genomic DNA were quantified using five quantification methods: UV spectrometry, SYBR-Green dye staining, slot blot hybridization with the probe D17Z1, Quantifiler Human DNA Quantification kit and RB1 rt-PCR. All methods measured higher DNA concentrations than...... expected based on the information by the manufacturers. UV spectrometry, SYBR-Green dye staining, slot blot and RB1 rt-PCR gave 39, 27, 11 and 12%, respectively, higher concentrations than expected based on the manufacturers' information. The DNA preparations were quantified using the Quantifiler Human DNA...... Quantification kit in two experiments. The measured DNA concentrations with Quantifiler were 125 and 160% higher than expected based on the manufacturers' information. When the Quantifiler human DNA standard (Raji cell line) was replaced by the commercial human DNA preparation G147A (Promega) to generate the DNA...

  9. Advancing agricultural greenhouse gas quantification*

    Science.gov (United States)

    Olander, Lydia; Wollenberg, Eva; Tubiello, Francesco; Herold, Martin

    2013-03-01

    1. Introduction Better information on greenhouse gas (GHG) emissions and mitigation potential in the agricultural sector is necessary to manage these emissions and identify responses that are consistent with the food security and economic development priorities of countries. Critical activity data (what crops or livestock are managed in what way) are poor or lacking for many agricultural systems, especially in developing countries. In addition, the currently available methods for quantifying emissions and mitigation are often too expensive or complex or not sufficiently user friendly for widespread use. The purpose of this focus issue is to capture the state of the art in quantifying greenhouse gases from agricultural systems, with the goal of better understanding our current capabilities and near-term potential for improvement, with particular attention to quantification issues relevant to smallholders in developing countries. This work is timely in light of international discussions and negotiations around how agriculture should be included in efforts to reduce and adapt to climate change impacts, and considering that significant climate financing to developing countries in post-2012 agreements may be linked to their increased ability to identify and report GHG emissions (Murphy et al 2010, CCAFS 2011, FAO 2011). 2. Agriculture and climate change mitigation The main agricultural GHGs—methane and nitrous oxide—account for 10%-12% of anthropogenic emissions globally (Smith et al 2008), or around 50% and 60% of total anthropogenic methane and nitrous oxide emissions, respectively, in 2005. Net carbon dioxide fluxes between agricultural land and the atmosphere linked to food production are relatively small, although significant carbon emissions are associated with degradation of organic soils for plantations in tropical regions (Smith et al 2007, FAO 2012). Population growth and shifts in dietary patterns toward more meat and dairy consumption will lead to

  10. The utility of diversity profiling using Illumina 18S rRNA gene amplicon deep sequencing to detect and discriminate Toxoplasma gondii among the cyst-forming coccidia.

    Science.gov (United States)

    Cooper, Madalyn K; Phalen, David N; Donahoe, Shannon L; Rose, Karrie; Šlapeta, Jan

    2016-01-30

    Next-generation sequencing (NGS) has the capacity to screen a single DNA sample and detect pathogen DNA from thousands of host DNA sequence reads, making it a versatile and informative tool for investigation of pathogens in diseased animals. The technique is effective and labor saving in the initial identification of pathogens, and will complement conventional diagnostic tests to associate the candidate pathogen with a disease process. In this report, we investigated the utility of the diversity profiling NGS approach using Illumina small subunit ribosomal RNA (18S rRNA) gene amplicon deep sequencing to detect Toxoplasma gondii in previously confirmed cases of toxoplasmosis. We then tested the diagnostic approach with species-specific PCR genotyping, histopathology and immunohistochemistry of toxoplasmosis in a Risso's dolphin (Grampus griseus) to systematically characterise the disease and associate causality. We show that the Euk7A/Euk570R primer set targeting the V1-V3 hypervariable region of the 18S rRNA gene can be used as a species-specific assay for cyst-forming coccidia and discriminate T. gondii. Overall, the approach is cost-effective and improves diagnostic decision support by narrowing the differential diagnosis list with more certainty than was previously possible. Furthermore, it supplements the limitations of cryptic protozoan morphology and surpasses the need for species-specific PCR primer combinations.

  11. Gene-specific amplicons from metagenomes as an alternative to directed evolution for enzyme screening: a case study using phenylacetaldehyde reductases.

    Science.gov (United States)

    Itoh, Nobuya; Kazama, Miki; Takeuchi, Nami; Isotani, Kentaro; Kurokawa, Junji

    2016-06-01

    Screening gene-specific amplicons from metagenomes (S-GAM) is a highly promising technique for the isolation of genes encoding enzymes for biochemical and industrial applications. From metagenomes, we isolated phenylacetaldehyde reductase (par) genes, which code for an enzyme that catalyzes the production of various Prelog's chiral alcohols. Nearly full-length par genes were amplified by PCR from metagenomic DNA, the products of which were fused with engineered par sequences at both terminal regions of the expression vector to ensure proper expression and then used to construct Escherichia coli plasmid libraries. Sequence- and activity-based screening of these libraries identified different homologous par genes, Hpar-001 to -036, which shared more than 97% amino acid sequence identity with PAR. Comparative characterization of these active homologs revealed a wide variety of enzymatic properties including activity, substrate specificity, and thermal stability. Moreover, amino acid substitutions in these genes coincided with those of Sar268 and Har1 genes, which were independently engineered by error-prone PCR to exhibit increased activity in the presence of concentrated 2-propanol. The comparative data from both approaches suggest that sequence information from homologs isolated from metagenomes is quite useful for enzyme engineering. Furthermore, by examining the GAM-based sequence dataset derived from soil metagenomes, we easily found amino acid substitutions that increase the thermal stability of PAR/PAR homologs. Thus, GAM-based approaches can provide not only useful homologous enzymes but also an alternative to directed evolution methodologies. PMID:27419059

  12. Cross-comparison of exome analysis, next-generation sequencing of amplicons, and the iPLEX® ADME PGx Panel for pharmacogenomic profiling

    Directory of Open Access Journals (Sweden)

    Eng Wee eChua

    2016-01-01

    Full Text Available Whole-exome sequencing (WES has been widely used for analysis of human genetic diseases, but its value for the pharmacogenomic profiling of individuals is not well studied. Initially, we performed an in-depth evaluation of the accuracy of WES variant calling in the pharmacogenes CYP2D6 and CYP2C19 by comparison with MiSeq® amplicon sequencing data (n = 36. This analysis revealed that the concordance rate between WES and MiSeq® was high, achieving 99.60% for variants that were called without exceeding the truth-sensitivity threshold (99%, defined during variant quality score recalibration. Beyond this threshold, the proportion of discordant calls increased markedly. Subsequently, we expanded our findings beyond CYP2D6 and CYP2C19 to include more genes genotyped by the iPLEX® ADME PGx Panel in the subset of twelve samples. WES performed well, agreeing with the genotyping panel in approximately 99% of the selected pass-filter variant calls. Overall, our results have demonstrated WES to be a promising approach for pharmacogenomic profiling, with an estimated error rate of lower than 1%. Quality filters, particularly variant quality score recalibration, are important for reducing the number of false variants. Future studies may benefit from examining the role of WES in the clinical setting for guiding drug therapy.

  13. The utility of diversity profiling using Illumina 18S rRNA gene amplicon deep sequencing to detect and discriminate Toxoplasma gondii among the cyst-forming coccidia.

    Science.gov (United States)

    Cooper, Madalyn K; Phalen, David N; Donahoe, Shannon L; Rose, Karrie; Šlapeta, Jan

    2016-01-30

    Next-generation sequencing (NGS) has the capacity to screen a single DNA sample and detect pathogen DNA from thousands of host DNA sequence reads, making it a versatile and informative tool for investigation of pathogens in diseased animals. The technique is effective and labor saving in the initial identification of pathogens, and will complement conventional diagnostic tests to associate the candidate pathogen with a disease process. In this report, we investigated the utility of the diversity profiling NGS approach using Illumina small subunit ribosomal RNA (18S rRNA) gene amplicon deep sequencing to detect Toxoplasma gondii in previously confirmed cases of toxoplasmosis. We then tested the diagnostic approach with species-specific PCR genotyping, histopathology and immunohistochemistry of toxoplasmosis in a Risso's dolphin (Grampus griseus) to systematically characterise the disease and associate causality. We show that the Euk7A/Euk570R primer set targeting the V1-V3 hypervariable region of the 18S rRNA gene can be used as a species-specific assay for cyst-forming coccidia and discriminate T. gondii. Overall, the approach is cost-effective and improves diagnostic decision support by narrowing the differential diagnosis list with more certainty than was previously possible. Furthermore, it supplements the limitations of cryptic protozoan morphology and surpasses the need for species-specific PCR primer combinations. PMID:26801593

  14. Enhanced detection of polymorphic DNA by multiple arbitrary amplicon profiling of endonuclease-digested DNA: identification of markers tightly linked to the supernodulation locus in soybean.

    Science.gov (United States)

    Caetano-Anollés, G; Bassam, B J; Gresshoff, P M

    1993-10-01

    Multiple endonuclease digestion of template DNA or amplification products can increase significantly the detection of polymorphic DNA in fingerprints generated by multiple arbitrary amplicon profiling (MAAP). This coupling of endonuclease cleavage and amplification of arbitrary stretches of DNA, directed by short oligonucleotide primers, readily allowed distinction of closely related fungal and bacterial isolates and plant cultivars. MAAP analysis of cleaved template DNA enabled the identification of molecular markers linked to a developmental locus of soybean (Glycine max L. Merrill). Ethyl methane sulfonate (EMS)-induced supernodulating, near-isogenic lines altered in the nts locus, which controls nodule formation, could be distinguished from each other and from the parent cultivar by amplification of template pre-digested with 2-3 restriction enzymes. A total of 42 DNA polymorphisms were detected using only 19 octamer primers. In the absence of digestion, 25 primers failed to differentiate these soybean genotypes. Several polymorphic products co-segregated tightly with the nts locus in F2 families from crosses between the allelic mutants nts382 and nts1007 and the ancestral G. soja Sieb. & Succ. PI468.397. Our results suggest that EMS is capable of inducing extensive DNA alterations, probably around discrete mutational hot-spots. EMS-induced DNA polymorphisms may constitute sequence-tagged markers diagnostic of specific genomic regions.

  15. Gastrointestinal Bacterial and Methanogenic Archaea Diversity Dynamics Associated with Condensed Tannin-Containing Pine Bark Diet in Goats Using 16S rDNA Amplicon Pyrosequencing

    Directory of Open Access Journals (Sweden)

    Byeng R. Min

    2014-01-01

    Full Text Available Eighteen Kiko-cross meat goats (n=6 were used to collect gastrointestinal (GI bacteria and methanogenic archaea for diversity measures when fed condensed tannin-containing pine bark (PB. Three dietary treatments were tested: control diet (0% PB and 30% wheat straw (WS; 0.17% condensed tannins (CT dry matter (DM; 15% PB and 15% WS (1.6% CT DM, and 30% PB and 0% WS (3.2% CT DM. A 16S rDNA bacterial tag-encoded FLX amplicon pyrosequencing technique was used to characterize and elucidate changes in GI bacteria and methanogenic archaea diversity among the diets. Proteobacteria was the most dominant phylum in goats with mean relative abundance values ranging from 39.7 (30% PB to 46.5% (control and 47.1% (15% PB. Other phyla individually accounted for fewer than 25% of the relative abundance observed. Predominant methanogens were Methanobrevibacter (75, 72, and 49%, Methanosphaera (3.3, 2.3, and 3.4%, and Methanobacteriaceae (1.2, 0.6, and 0.7% population in control, 15, and 30% PB, respectively. Among methanogens, Methanobrevibacter was linearly decreased (P=0.05 with increasing PB supplementation. These results indicate that feeding PB selectively altered bacteria and methanogenic archaeal populations in the GI tract of goats.

  16. Simultaneous DNA-RNA Extraction from Coastal Sediments and Quantification of 16S rRNA Genes and Transcripts by Real-time PCR.

    Science.gov (United States)

    Tatti, Enrico; McKew, Boyd A; Whitby, Corrine; Smith, Cindy J

    2016-06-11

    Real Time Polymerase Chain Reaction also known as quantitative PCR (q-PCR) is a widely used tool in microbial ecology to quantify gene abundances of taxonomic and functional groups in environmental samples. Used in combination with a reverse transcriptase reaction (RT-q-PCR), it can also be employed to quantify gene transcripts. q-PCR makes use of highly sensitive fluorescent detection chemistries that allow quantification of PCR amplicons during the exponential phase of the reaction. Therefore, the biases associated with 'end-point' PCR detected in the plateau phase of the PCR reaction are avoided. A protocol to quantify bacterial 16S rRNA genes and transcripts from coastal sediments via real-time PCR is provided. First, a method for the co-extraction of DNA and RNA from coastal sediments, including the additional steps required for the preparation of DNA-free RNA, is outlined. Second, a step-by-step guide for the quantification of 16S rRNA genes and transcripts from the extracted nucleic acids via q-PCR and RT-q-PCR is outlined. This includes details for the construction of DNA and RNA standard curves. Key considerations for the use of RT-q-PCR assays in microbial ecology are included.

  17. Efficient uncertainty quantification in unsteady aeroelastic simulations

    NARCIS (Netherlands)

    Witteveen, J.A.S.; Bijl, H.

    2009-01-01

    An efficient uncertainty quantification method for unsteady problems is presented in order to achieve a constant accuracy in time for a constant number of samples. The approach is applied to the aeroelastic problems of a transonic airfoil flutter system and the AGARD 445.6 wing benchmark with uncert

  18. Quantification of topological concepts using ideals

    Directory of Open Access Journals (Sweden)

    Robert Lowen

    2001-01-01

    Full Text Available We introduce certain ideals of real-valued functions as a natural generalization of filters. We show that these ideals establish a canonical framework for the quantification of topological concepts, such as closedness, adherence, and compactness, in the setting of approach spaces.

  19. Perfusion Quantification Using Gaussian Process Deconvolution

    DEFF Research Database (Denmark)

    Andersen, Irene Klærke; Have, Anna Szynkowiak; Rasmussen, Carl Edward;

    2002-01-01

    The quantification of perfusion using dynamic susceptibility contrast MRI (DSC-MRI) requires deconvolution to obtain the residual impulse response function (IRF). In this work, a method using the Gaussian process for deconvolution (GPD) is proposed. The fact that the IRF is smooth is incorporated...

  20. Quantification of Cannabinoid Content in Cannabis

    Science.gov (United States)

    Tian, Y.; Zhang, F.; Jia, K.; Wen, M.; Yuan, Ch.

    2015-09-01

    Cannabis is an economically important plant that is used in many fields, in addition to being the most commonly consumed illicit drug worldwide. Monitoring the spatial distribution of cannabis cultivation and judging whether it is drug- or fiber-type cannabis is critical for governments and international communities to understand the scale of the illegal drug trade. The aim of this study was to investigate whether the cannabinoids content in cannabis could be spectrally quantified using a spectrometer and to identify the optimal wavebands for quantifying the cannabinoid content. Spectral reflectance data of dried cannabis leaf samples and the cannabis canopy were measured in the laboratory and in the field, respectively. Correlation analysis and the stepwise multivariate regression method were used to select the optimal wavebands for cannabinoid content quantification based on the laboratory-measured spectral data. The results indicated that the delta-9-tetrahydrocannabinol (THC) content in cannabis leaves could be quantified using laboratory-measured spectral reflectance data and that the 695 nm band is the optimal band for THC content quantification. This study provides prerequisite information for designing spectral equipment to enable immediate quantification of THC content in cannabis and to discriminate drug- from fiber-type cannabis based on THC content quantification in the field.

  1. Recurrence quantification analysis in Liu's attractor

    International Nuclear Information System (INIS)

    Recurrence Quantification Analysis is used to detect transitions chaos to periodical states or chaos to chaos in a new dynamical system proposed by Liu et al. This system contains a control parameter in the second equation and was originally introduced to investigate the forming mechanism of the compound structure of the chaotic attractor which exists when the control parameter is zero

  2. HPC Analytics Support. Requirements for Uncertainty Quantification Benchmarks

    Energy Technology Data Exchange (ETDEWEB)

    Paulson, Patrick R. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Purohit, Sumit [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Rodriguez, Luke R. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2015-05-01

    This report outlines techniques for extending benchmark generation products so they support uncertainty quantification by benchmarked systems. We describe how uncertainty quantification requirements can be presented to candidate analytical tools supporting SPARQL. We describe benchmark data sets for evaluating uncertainty quantification, as well as an approach for using our benchmark generator to produce data sets for generating benchmark data sets.

  3. Development of an assay for rapid detection and quantification of Verticillium dahliae in soil.

    Science.gov (United States)

    Bilodeau, Guillaume J; Koike, Steven T; Uribe, Pedro; Martin, Frank N

    2012-03-01

    ABSTRACT Verticillium dahliae is responsible for Verticillium wilt on a wide range of hosts, including strawberry, on which low soil inoculum densities can cause significant crop loss. Determination of inoculum density is currently done by soil plating but this can take 6 to 8 weeks to complete and delay the grower's ability to make planting decisions. To provide a faster means for estimating pathogen populations in the soil, a multiplexed TaqMan real-time polymerase chain reaction (PCR) assay based on the ribosomal DNA (rDNA) intergenic spacer (IGS) was developed for V. dahliae. The assay was specific for V. dahliae and included an internal control for evaluation of inhibition due to the presence of PCR inhibitors in DNA extracted from soil samples. An excellent correlation was observed in regression analysis (R(2) = 0.96) between real-time PCR results and inoculum densities determined by soil plating in a range of field soils with pathogen densities as low as 1 to 2 microsclerotia/g of soil. Variation in copy number of the rDNA was also evaluated among isolates by SYBR Green real-time PCR amplification of the V. dahliae-specific amplicon compared with amplification of several single-copy genes and was estimated to range from ≈24 to 73 copies per haploid genome, which translated into possible differences in results among isolates of ≈1.8 cycle thresholds. Analysis of the variation in results of V. dahliae quantification among extractions of the same soil sample indicated that assaying four replicate DNA extractions for each field sample would provide accurate results. A TaqMan assay also was developed to help identify colonies of V. tricorpus on soil plates.

  4. Development of an assay for rapid detection and quantification of Verticillium dahliae in soil.

    Science.gov (United States)

    Bilodeau, Guillaume J; Koike, Steven T; Uribe, Pedro; Martin, Frank N

    2012-03-01

    ABSTRACT Verticillium dahliae is responsible for Verticillium wilt on a wide range of hosts, including strawberry, on which low soil inoculum densities can cause significant crop loss. Determination of inoculum density is currently done by soil plating but this can take 6 to 8 weeks to complete and delay the grower's ability to make planting decisions. To provide a faster means for estimating pathogen populations in the soil, a multiplexed TaqMan real-time polymerase chain reaction (PCR) assay based on the ribosomal DNA (rDNA) intergenic spacer (IGS) was developed for V. dahliae. The assay was specific for V. dahliae and included an internal control for evaluation of inhibition due to the presence of PCR inhibitors in DNA extracted from soil samples. An excellent correlation was observed in regression analysis (R(2) = 0.96) between real-time PCR results and inoculum densities determined by soil plating in a range of field soils with pathogen densities as low as 1 to 2 microsclerotia/g of soil. Variation in copy number of the rDNA was also evaluated among isolates by SYBR Green real-time PCR amplification of the V. dahliae-specific amplicon compared with amplification of several single-copy genes and was estimated to range from ≈24 to 73 copies per haploid genome, which translated into possible differences in results among isolates of ≈1.8 cycle thresholds. Analysis of the variation in results of V. dahliae quantification among extractions of the same soil sample indicated that assaying four replicate DNA extractions for each field sample would provide accurate results. A TaqMan assay also was developed to help identify colonies of V. tricorpus on soil plates. PMID:22066673

  5. Real-time PCR assay for the simultaneous quantification of nitrifying and denitrifying bacteria in activated sludge

    Energy Technology Data Exchange (ETDEWEB)

    Geets, J.; Cooman, M. de; Wittebolle, L.; Verstraete, W.; Boon, N. [Ghent Univ. (BE). Lab. of Microbial Ecology and Technology (LabMET); Heylen, K.; Vanparys, B.; Vos, P. de [Ghent Univ. (Belgium). Dept. of Biochemistry, Physiology and Microbiology

    2007-05-15

    In order to improve wastewater treatment processes, a need exists for tools that rapidly give detailed insight into the community structure of activated sludge, supplementary to chemical and physical data. In this study, the advantages of microarrays and quantitative polymerase chin reaction (PCR) methods were combined into a real-time PCR assay that allows the simultaneous quantification of phylogenetic and functional genes involved in nitrification and denitrification processes. Simultaneous quantification was possible along a 5-log dynamic range and with high linear correlation (R{sup 2}>0.98). The specificity of the assay was confirmed by cloning and sequencing analyses of PCR amplicons obtained from activated sludge. The real-time assay was validated on mixed liquid samples of different treatment plants, which varied in nitrogen removal rate. The abundance of ammonia oxidizers was in the order of magnitude of 10{sup 6} down to 10{sup 4} ml{sup -1}, whereas nitrite oxidizers were less abundant (10{sup 3}-10{sup 1} order of magnitude). The results were in correspondence with the nitrite oxidation rate in the sludge types. As for the nirS, nirK, and nosZ gene copy numbers, their abundance was generally in the order of magnitude of 10{sup 8}-10{sup 5}. When sludge samples were subjected to lab-scale perturbations, a decrease in nitrification rate was reflected within 18 h in the copy numbers of nitrifier genes (decrease with 1 to 5 log units), whereas denitrification genes remained rather unaffected. These results demonstrate that the method is a fast and accurate tool for the analysis of the (de)nitrifying community structure and size in both natural and engineered environmental samples. (orig.)

  6. TaqMan probe real-time polymerase chain reaction assay for the quantification of canine DNA in chicken nugget.

    Science.gov (United States)

    Rahman, Md Mahfujur; Hamid, Sharifah Bee Abd; Basirun, Wan Jefrey; Bhassu, Subha; Rashid, Nur Raifana Abdul; Mustafa, Shuhaimi; Mohd Desa, Mohd Nasir; Ali, Md Eaqub

    2016-01-01

    This paper describes a short-amplicon-based TaqMan probe quantitative real-time PCR (qPCR) assay for the quantitative detection of canine meat in chicken nuggets, which are very popular across the world, including Malaysia. The assay targeted a 100-bp fragment of canine cytb gene using a canine-specific primer and TaqMan probe. Specificity against 10 different animals and plants species demonstrated threshold cycles (Ct) of 16.13 ± 0.12 to 16.25 ± 0.23 for canine DNA and negative results for the others in a 40-cycle reaction. The assay was tested for the quantification of up to 0.01% canine meat in deliberately spiked chicken nuggets with 99.7% PCR efficiency and 0.995 correlation coefficient. The analysis of the actual and qPCR predicted values showed a high recovery rate (from 87% ± 28% to 112% ± 19%) with a linear regression close to unity (R(2) = 0.999). Finally, samples of three halal-branded commercial chicken nuggets collected from different Malaysian outlets were screened for canine meat, but no contamination was demonstrated. PMID:26458055

  7. Detection and quantification of root-knot nematode (Meloidogyne javanica), lesion nematode (Pratylenchus zeae) and dagger nematode (Xiphinema elongatum) parasites of sugarcane using real-time PCR.

    Science.gov (United States)

    Berry, Shaun D; Fargette, Mireille; Spaull, Vaughan W; Morand, Serge; Cadet, Patrice

    2008-06-01

    A number of different plant parasitic nematode species are found associated with sugarcane in South Africa. Of these, the root-knot nematode (Meloidogyne javanica), the lesion nematode (Pratylenchus zeae) and the dagger nematode (Xiphinema elongatum) are potentially the most damaging pests. Identification and enumeration of the number of these nematodes are necessary for providing advice to farmers as well as studying the effects of various treatments in field and glasshouse trials. We report on the development, use, and extent of specificity of three sets of primers, for M. javanica, P. zeae and X. elongatum, and on tests to detect and quantify the number of these nematodes in soil samples using SYBR Green I dye and real-time PCR technology. Amplicons from the three target species (obtained with their respective primer sets) are discernible in size by gel electrophoresis (380bp for M. javanica, 250bp for P. zeae and 500bp for X. elongatum). Also, these amplicons have characteristic melting temperatures of 83.8 degrees C (M. javanica), 86.6 degrees C (P. zeae) and 86.1 degrees C (X. elongatum). Investigations into multiplex reactions found competition between species with M. javanica competing with P. zeae and X. elongatum. Subsequent single tube (simplex) assays, enabled the construction of calibration curves for each of the three species. These were then used for quantification of the numbers of each of these species in nematode samples extracted from the field, with a high (R2=0.83) and significant positive correlation between real-time PCR and counts performed with microscopy.

  8. Rapid Drug Susceptibility Testing of Drug-Resistant Mycobacterium tuberculosis Isolates Directly from Clinical Samples by Use of Amplicon Sequencing: a Proof-of-Concept Study

    Science.gov (United States)

    Anderson, Julia; Lemmer, Darrin; Lehmkuhl, Erik; Georghiou, Sophia B.; Heaton, Hannah; Wiggins, Kristin; Gillece, John D.; Schupp, James M.; Catanzaro, Donald G.; Crudu, Valeriu; Cohen, Ted; Rodwell, Timothy C.; Engelthaler, David M.

    2016-01-01

    Increasingly complex drug-resistant tuberculosis (DR-TB) is a major global health concern and one of the primary reasons why TB is now the leading infectious cause of death worldwide. Rapid characterization of a DR-TB patient's complete drug resistance profile would facilitate individualized treatment in place of empirical treatment, improve treatment outcomes, prevent amplification of resistance, and reduce the transmission of DR-TB. The use of targeted next-generation sequencing (NGS) to obtain drug resistance profiles directly from patient sputum samples has the potential to enable comprehensive evidence-based treatment plans to be implemented quickly, rather than in weeks to months, which is currently needed for phenotypic drug susceptibility testing (DST) results. In this pilot study, we evaluated the performance of amplicon sequencing of Mycobacterium tuberculosis DNA from patient sputum samples using a tabletop NGS technology and automated data analysis to provide a rapid DST solution (the Next Gen-RDST assay). One hundred sixty-six out of 176 (94.3%) sputum samples from the Republic of Moldova yielded complete Next Gen-RDST assay profiles for 7 drugs of interest. We found a high level of concordance of our Next Gen-RDST assay results with phenotypic DST (97.0%) and pyrosequencing (97.8%) results from the same clinical samples. Our Next Gen-RDST assay was also able to estimate the proportion of resistant-to-wild-type alleles down to mixtures of ≤1%, which demonstrates the ability to detect very low levels of resistant variants not detected by pyrosequencing and possibly below the threshold for phenotypic growth methods. The assay as described here could be used as a clinical or surveillance tool. PMID:27225403

  9. Rapid Drug Susceptibility Testing of Drug-Resistant Mycobacterium tuberculosis Isolates Directly from Clinical Samples by Use of Amplicon Sequencing: a Proof-of-Concept Study.

    Science.gov (United States)

    Colman, Rebecca E; Anderson, Julia; Lemmer, Darrin; Lehmkuhl, Erik; Georghiou, Sophia B; Heaton, Hannah; Wiggins, Kristin; Gillece, John D; Schupp, James M; Catanzaro, Donald G; Crudu, Valeriu; Cohen, Ted; Rodwell, Timothy C; Engelthaler, David M

    2016-08-01

    Increasingly complex drug-resistant tuberculosis (DR-TB) is a major global health concern and one of the primary reasons why TB is now the leading infectious cause of death worldwide. Rapid characterization of a DR-TB patient's complete drug resistance profile would facilitate individualized treatment in place of empirical treatment, improve treatment outcomes, prevent amplification of resistance, and reduce the transmission of DR-TB. The use of targeted next-generation sequencing (NGS) to obtain drug resistance profiles directly from patient sputum samples has the potential to enable comprehensive evidence-based treatment plans to be implemented quickly, rather than in weeks to months, which is currently needed for phenotypic drug susceptibility testing (DST) results. In this pilot study, we evaluated the performance of amplicon sequencing of Mycobacterium tuberculosis DNA from patient sputum samples using a tabletop NGS technology and automated data analysis to provide a rapid DST solution (the Next Gen-RDST assay). One hundred sixty-six out of 176 (94.3%) sputum samples from the Republic of Moldova yielded complete Next Gen-RDST assay profiles for 7 drugs of interest. We found a high level of concordance of our Next Gen-RDST assay results with phenotypic DST (97.0%) and pyrosequencing (97.8%) results from the same clinical samples. Our Next Gen-RDST assay was also able to estimate the proportion of resistant-to-wild-type alleles down to mixtures of ≤1%, which demonstrates the ability to detect very low levels of resistant variants not detected by pyrosequencing and possibly below the threshold for phenotypic growth methods. The assay as described here could be used as a clinical or surveillance tool. PMID:27225403

  10. Uncertainty Quantification with Applications to Engineering Problems

    DEFF Research Database (Denmark)

    Bigoni, Daniele

    in measurements, predictions and manufacturing, and we can say that any dynamical system used in engineering is subject to some of these uncertainties. The first part of this work presents an overview of the mathematical framework used in Uncertainty Quantification (UQ) analysis and introduces the spectral tensor......The systematic quantification of the uncertainties affecting dynamical systems and the characterization of the uncertainty of their outcomes is critical for engineering design and analysis, where risks must be reduced as much as possible. Uncertainties stem naturally from our limitations......-train (STT) decomposition, a novel high-order method for the effective propagation of uncertainties which aims at providing an exponential convergence rate while tackling the curse of dimensionality. The curse of dimensionality is a problem that afflicts many methods based on meta-models, for which...

  11. SPECT quantification of regional radionuclide distributions

    International Nuclear Information System (INIS)

    SPECT quantification of regional radionuclide activities within the human body is affected by several physical and instrumental factors including attenuation of photons within the patient, Compton scattered events, the system's finite spatial resolution and object size, finite number of detected events, partial volume effects, the radiopharmaceutical biokinetics, and patient and/or organ motion. Furthermore, other instrumentation factors such as calibration of the center-of-rotation, sampling, and detector nonuniformities will affect the SPECT measurement process. These factors are described, together with examples of compensation methods that are currently available for improving SPECT quantification. SPECT offers the potential to improve in vivo estimates of absorbed dose, provided the acquisition, reconstruction, and compensation procedures are adequately implemented and utilized. 53 references, 2 figures

  12. Automated Quantification of Pneumothorax in CT

    OpenAIRE

    Synho Do; Kristen Salvaggio; Supriya Gupta; Mannudeep Kalra; Ali, Nabeel U.; Homer Pien

    2012-01-01

    An automated, computer-aided diagnosis (CAD) algorithm for the quantification of pneumothoraces from Multidetector Computed Tomography (MDCT) images has been developed. Algorithm performance was evaluated through comparison to manual segmentation by expert radiologists. A combination of two-dimensional and three-dimensional processing techniques was incorporated to reduce required processing time by two-thirds (as compared to similar techniques). Volumetric measurements on relative pneumothor...

  13. A Tableaux Calculus for Ambiguous Quantification

    OpenAIRE

    Monz, Christof; de Rijke, Maarten

    2000-01-01

    Coping with ambiguity has recently received a lot of attention in natural language processing. Most work focuses on the semantic representation of ambiguous expressions. In this paper we complement this work in two ways. First, we provide an entailment relation for a language with ambiguous expressions. Second, we give a sound and complete tableaux calculus for reasoning with statements involving ambiguous quantification. The calculus interleaves partial disambiguation steps with steps in a t...

  14. Whitepaper on Uncertainty Quantification for MPACT

    Energy Technology Data Exchange (ETDEWEB)

    Williams, Mark L. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

    2015-12-17

    The MPACT code provides the ability to perform high-fidelity deterministic calculations to obtain a wide variety of detailed results for very complex reactor core models. However MPACT currently does not have the capability to propagate the effects of input data uncertainties to provide uncertainties in the calculated results. This white paper discusses a potential method for MPACT uncertainty quantification (UQ) based on stochastic sampling.

  15. Uncertainty quantification of effective nuclear interactions

    CERN Document Server

    Perez, R Navarro; Arriola, E Ruiz

    2016-01-01

    We give a brief review on the development of phenomenological NN interactions and the corresponding quantification of statistical uncertainties. We look into the uncertainty of effective interactions broadly used in mean field calculations through the Skyrme parameters and effective field theory counter-terms by estimating both statistical and systematic uncertainties stemming from the NN interaction. We also comment on the role played by different fitting strategies on the light of recent developments.

  16. Automated quantification of synapses by fluorescence microscopy.

    Science.gov (United States)

    Schätzle, Philipp; Wuttke, René; Ziegler, Urs; Sonderegger, Peter

    2012-02-15

    The quantification of synapses in neuronal cultures is essential in studies of the molecular mechanisms underlying synaptogenesis and synaptic plasticity. Conventional counting of synapses based on morphological or immunocytochemical criteria is extremely work-intensive. We developed a fully automated method which quantifies synaptic elements and complete synapses based on immunocytochemistry. Pre- and postsynaptic elements are detected by their corresponding fluorescence signals and their proximity to dendrites. Synapses are defined as the combination of a pre- and postsynaptic element within a given distance. The analysis is performed in three dimensions and all parameters required for quantification can be easily adjusted by a graphical user interface. The integrated batch processing enables the analysis of large datasets without any further user interaction and is therefore efficient and timesaving. The potential of this method was demonstrated by an extensive quantification of synapses in neuronal cultures from DIV 7 to DIV 21. The method can be applied to all datasets containing a pre- and postsynaptic labeling plus a dendritic or cell surface marker.

  17. Automated Template Quantification for DNA Sequencing Facilities

    Science.gov (United States)

    Ivanetich, Kathryn M.; Yan, Wilson; Wunderlich, Kathleen M.; Weston, Jennifer; Walkup, Ward G.; Simeon, Christian

    2005-01-01

    The quantification of plasmid DNA by the PicoGreen dye binding assay has been automated, and the effect of quantification of user-submitted templates on DNA sequence quality in a core laboratory has been assessed. The protocol pipets, mixes and reads standards, blanks and up to 88 unknowns, generates a standard curve, and calculates template concentrations. For pUC19 replicates at five concentrations, coefficients of variance were 0.1, and percent errors were from 1% to 7% (n = 198). Standard curves with pUC19 DNA were nonlinear over the 1 to 1733 ng/μL concentration range required to assay the majority (98.7%) of user-submitted templates. Over 35,000 templates have been quantified using the protocol. For 1350 user-submitted plasmids, 87% deviated by ≥ 20% from the requested concentration (500 ng/μL). Based on data from 418 sequencing reactions, quantification of user-submitted templates was shown to significantly improve DNA sequence quality. The protocol is applicable to all types of double-stranded DNA, is unaffected by primer (1 pmol/μL), and is user modifiable. The protocol takes 30 min, saves 1 h of technical time, and costs approximately $0.20 per unknown. PMID:16461949

  18. Fluorescence-linked Antigen Quantification (FLAQ) Assay for Fast Quantification of HIV-1 p24Gag

    Science.gov (United States)

    Gesner, Marianne; Maiti, Mekhala; Grant, Robert; Cavrois, Marielle

    2016-01-01

    The fluorescence-linked antigen quantification (FLAQ) assay allows a fast quantification of HIV-1 p24Gag antigen. Viral supernatant are lysed and incubated with polystyrene microspheres coated with polyclonal antibodies against HIV-1 p24Gag and detector antibodies conjugated to fluorochromes (Figure 1). After washes, the fluorescence of microspheres is measured by flow cytometry and reflects the abundance of the antigen in the lysate. The speed, simplicity, and wide dynamic range of the FLAQ assay are optimum for many applications performed in HIV-1 research laboratories.

  19. Illumina MiSeq Phylogenetic Amplicon Sequencing Shows a Large Reduction of an Uncharacterised Succinivibrionaceae and an Increase of the Methanobrevibacter gottschalkii Clade in Feed Restricted Cattle.

    Directory of Open Access Journals (Sweden)

    Matthew Sean McCabe

    Full Text Available Periodic feed restriction is used in cattle production to reduce feed costs. When normal feed levels are resumed, cattle catch up to a normal weight by an acceleration of normal growth rate, known as compensatory growth, which is not yet fully understood. Illumina Miseq Phylogenetic marker amplicon sequencing of DNA extracted from rumen contents of 55 bulls showed that restriction of feed (70% concentrate, 30% grass silage for 125 days, to levels that caused a 60% reduction of growth rate, resulted in a large increase of relative abundance of Methanobrevibacter gottschalkii clade (designated as OTU-M7, and a large reduction of an uncharacterised Succinivibrionaceae species (designated as OTU-S3004. There was a strong negative Spearman correlation (ρ = -0.72, P = <1x10(-20 between relative abundances of OTU-3004 and OTU-M7 in the liquid rumen fraction. There was also a significant increase in acetate:propionate ratio (A:P in feed restricted animals that showed a negative Spearman correlation (ρ = -0.69, P = <1x10(-20 with the relative abundance of OTU-S3004 in the rumen liquid fraction but not the solid fraction, and a strong positive Spearman correlation with OTU-M7 in the rumen liquid (ρ = 0.74, P = <1x10(-20 and solid (ρ = 0.69, P = <1x10(-20 fractions. Reduced A:P ratios in the rumen are associated with increased feed efficiency and reduced production of methane which has a global warming potential (GWP 100 years of 28. Succinivibrionaceae growth in the rumen was previously suggested to reduce methane emissions as some members of this family utilise hydrogen, which is also utilised by methanogens for methanogenesis, to generate succinate which is converted to propionate. Relative abundance of OTU-S3004 showed a positive Spearman correlation with propionate (ρ = 0.41, P = <0.01 but not acetate in the liquid rumen fraction.

  20. Aerodynamic Modeling with Heterogeneous Data Assimilation and Uncertainty Quantification Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Clear Science Corp. proposes to develop an aerodynamic modeling tool that assimilates data from different sources and facilitates uncertainty quantification. The...

  1. Efficient Quantification of Uncertainties in Complex Computer Code Results Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Propagation of parameter uncertainties through large computer models can be very resource intensive. Frameworks and tools for uncertainty quantification are...

  2. Efficient Quantification of Uncertainties in Complex Computer Code Results Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This proposal addresses methods for efficient quantification of margins and uncertainties (QMU) for models that couple multiple, large-scale commercial or...

  3. Adjoint-Based Uncertainty Quantification with MCNP

    Energy Technology Data Exchange (ETDEWEB)

    Seifried, Jeffrey E. [Univ. of California, Berkeley, CA (United States)

    2011-09-01

    This work serves to quantify the instantaneous uncertainties in neutron transport simulations born from nuclear data and statistical counting uncertainties. Perturbation and adjoint theories are used to derive implicit sensitivity expressions. These expressions are transformed into forms that are convenient for construction with MCNP6, creating the ability to perform adjoint-based uncertainty quantification with MCNP6. These new tools are exercised on the depleted-uranium hybrid LIFE blanket, quantifying its sensitivities and uncertainties to important figures of merit. Overall, these uncertainty estimates are small (< 2%). Having quantified the sensitivities and uncertainties, physical understanding of the system is gained and some confidence in the simulation is acquired.

  4. Tutorial examples for uncertainty quantification methods.

    Energy Technology Data Exchange (ETDEWEB)

    De Bord, Sarah [Univ. of California, Davis, CA (United States)

    2015-08-01

    This report details the work accomplished during my 2015 SULI summer internship at Sandia National Laboratories in Livermore, CA. During this internship, I worked on multiple tasks with the common goal of making uncertainty quantification (UQ) methods more accessible to the general scientific community. As part of my work, I created a comprehensive numerical integration example to incorporate into the user manual of a UQ software package. Further, I developed examples involving heat transfer through a window to incorporate into tutorial lectures that serve as an introduction to UQ methods.

  5. Uncertainty quantification and stochastic modeling with Matlab

    CERN Document Server

    Souza de Cursi, Eduardo

    2015-01-01

    Uncertainty Quantification (UQ) is a relatively new research area which describes the methods and approaches used to supply quantitative descriptions of the effects of uncertainty, variability and errors in simulation problems and models. It is rapidly becoming a field of increasing importance, with many real-world applications within statistics, mathematics, probability and engineering, but also within the natural sciences. Literature on the topic has up until now been largely based on polynomial chaos, which raises difficulties when considering different types of approximation and does no

  6. Quantification of thermal damage in skin tissue

    Institute of Scientific and Technical Information of China (English)

    Xu Feng; Wen Ting; Lu Tianjian; Seffen Keith

    2008-01-01

    Skin thermal damage or skin burns are the most commonly encountered type of trauma in civilian and military communities. Besides, advances in laser, microwave and similar technologies have led to recent developments of thermal treatments for disease and damage involving skin tissue, where the objective is to induce thermal damage precisely within targeted tissue structures but without affecting the surrounding, healthy tissue. Further, extended pain sensation induced by thermal damage has also brought great problem for burn patients. Thus, it is of great importance to quantify the thermal damage in skin tissue. In this paper, the available models and experimental methods for quantification of thermal damage in skin tissue are discussed.

  7. Quantification of prebiotics in commercial infant formulas.

    Science.gov (United States)

    Sabater, Carlos; Prodanov, Marin; Olano, Agustín; Corzo, Nieves; Montilla, Antonia

    2016-03-01

    Since breastfeeding is not always possible, infant formulas (IFs) are supplemented with prebiotic oligosaccharides, such as galactooligosaccharides (GOS) and/or fructooligosaccharides (FOS) to exert similar effects to those of the breast milk. Nowadays, a great number of infant formulas enriched with prebiotics are disposal in the market, however there are scarce data about their composition. In this study, the combined use of two chromatographic methods (GC-FID and HPLC-RID) for the quantification of carbohydrates present in commercial infant formulas have been used. According to the results obtained by GC-FID for products containing prebiotics, the content of FOS, GOS and GOS/FOS was in the ranges of 1.6-5.0, 1.7-3.2, and 0.08-0.25/2.3-3.8g/100g of product, respectively. HPLC-RID analysis allowed quantification of maltodextrins with degree of polymerization (DP) up to 19. The methodology proposed here may be used for routine quality control of infant formula and other food ingredients containing prebiotics.

  8. Quantification of prebiotics in commercial infant formulas.

    Science.gov (United States)

    Sabater, Carlos; Prodanov, Marin; Olano, Agustín; Corzo, Nieves; Montilla, Antonia

    2016-03-01

    Since breastfeeding is not always possible, infant formulas (IFs) are supplemented with prebiotic oligosaccharides, such as galactooligosaccharides (GOS) and/or fructooligosaccharides (FOS) to exert similar effects to those of the breast milk. Nowadays, a great number of infant formulas enriched with prebiotics are disposal in the market, however there are scarce data about their composition. In this study, the combined use of two chromatographic methods (GC-FID and HPLC-RID) for the quantification of carbohydrates present in commercial infant formulas have been used. According to the results obtained by GC-FID for products containing prebiotics, the content of FOS, GOS and GOS/FOS was in the ranges of 1.6-5.0, 1.7-3.2, and 0.08-0.25/2.3-3.8g/100g of product, respectively. HPLC-RID analysis allowed quantification of maltodextrins with degree of polymerization (DP) up to 19. The methodology proposed here may be used for routine quality control of infant formula and other food ingredients containing prebiotics. PMID:26471520

  9. Quantification of abdominal aortic deformation after EVAR

    Science.gov (United States)

    Demirci, Stefanie; Manstad-Hulaas, Frode; Navab, Nassir

    2009-02-01

    Quantification of abdominal aortic deformation is an important requirement for the evaluation of endovascular stenting procedures and the further refinement of stent graft design. During endovascular aortic repair (EVAR) treatment, the aortic shape is subject to severe deformation that is imposed by medical instruments such as guide wires, catheters, and, the stent graft. This deformation can affect the flow characteristics and morphology of the aorta which have been shown to be elicitors for stent graft failures and be reason for reappearance of aneurysms. We present a method for quantifying the deformation of an aneurysmatic aorta imposed by an inserted stent graft device. The outline of the procedure includes initial rigid alignment of the two abdominal scans, segmentation of abdominal vessel trees, and automatic reduction of their centerline structures to one specified region of interest around the aorta. This is accomplished by preprocessing and remodeling of the pre- and postoperative aortic shapes before performing a non-rigid registration. We further narrow the resulting displacement fields to only include local non-rigid deformation and therefore, eliminate all remaining global rigid transformations. Finally, deformations for specified locations can be calculated from the resulting displacement fields. In order to evaluate our method, experiments for the extraction of aortic deformation fields are conducted on 15 patient datasets from endovascular aortic repair (EVAR) treatment. A visual assessment of the registration results and evaluation of the usage of deformation quantification were performed by two vascular surgeons and one interventional radiologist who are all experts in EVAR procedures.

  10. 大豆小片段法HRM基因分型体系优化%Optimization of small amplicon of HRM genotyping system in soybean

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    SNP genotyping by High Resolution Melting Curve( HRM)is a simple and effective,higher sensi-tivity and stronger specificity method. Optimization and establishment of soybean HRM-based SNP genotyping sys-tem is the precondition and basis for soybean SNP research in future. In this study,small amplicon after adding the internal calibration was used to optimize and establish SNP genotyping system. The results showed that it was better to make PCR product length 50 -80bp,the melting temperature at 72 -82℃ and Tm value between 55 -62℃when designing a SNP primer. A total of 10μL PCR reaction system should contain 25ng DNA template,0. 6pmol SNP primers and 1μL LC Green dye. After PCR reaction,3 pmol high and low temperature internal calibration should be added to each wells,and then denatured for HRM analysis. Moreover,the optimized SNP genotyping system was used to genotyping a recombinant inbred lines( RILs)population,and the results demonstrated that the RILs could be completely divided into 2 genotypes of their parents by the optimized system,with improved accuracy and efficiency. The optimized SNP genotyping system established in this study will provide a useful foundation for developing SNP markers,constructing high density genetic maps,QTL identification in the future.%应用高分辨率熔解曲线( high resolution melting curve,HRM)技术进行基因分型简单有效,灵敏度高、特异性强。优化并建立基于HRM的大豆SNP基因分型体系,是准确进行SNP研究的前提和基础。本研究利用添加内标后的小片段法进行SNP基因分型体系优化研究,结果表明在设计SNP引物时,最好使其PCR产物长度为50~80bp,熔解温度在72~82℃,Tm值介于55~62℃之间;10μL PCR反应体系中应包含25ng DNA模板,0.6pmol SNP引物,1μL LC Green染料;PCR反应后,应在各点样孔分别加入3pmol的高、低温内标,然后进行变性处理和HRM分析。同时利用建立的基因分型体

  11. Comparative studies of modern methods for caries detection and quantification

    OpenAIRE

    Shi, Xie-Qi

    2001-01-01

    In clinical dentistry proper treatment of caries lesions is highly dependent on diagnostic accuracy. Aim The present dissertation aimed at the evaluation and comparisons between several modem methods for caries detection and quantification. Methods The employed methods for detection and quantification of caries lesions may be divided into two categories, namely laser fluorescence based methods and radiographic methods. Laser fluorescence The performance of the...

  12. 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...

  13. Molecular quantification of genes encoding for green-fluorescent proteins

    DEFF Research Database (Denmark)

    Felske, A; Vandieken, V; Pauling, B V;

    2003-01-01

    A quantitative PCR approach is presented to analyze the amount of recombinant green fluorescent protein (gfp) genes in environmental DNA samples. The quantification assay is a combination of specific PCR amplification and temperature gradient gel electrophoresis (TGGE). Gene quantification...... PCR strategy is a highly specific and sensitive way to monitor recombinant DNA in environments like the efflux of a biotechnological plant....

  14. Results of an explorative research into value quantification methods

    NARCIS (Netherlands)

    Drechsler, M.; Ridder, H. de

    2006-01-01

    This paper presents the results of an explorative research into value quantification methods called the Quick Scan. An overview of value quantification methods was needed in order to be able to establish the economically most advantageous tender, which in turn is needed for improvements in the Dutch

  15. An improved competitive inhibition enzymatic immunoassay method for tetrodotoxin quantification

    OpenAIRE

    Stokes Amber N; Williams Becky L; French Susannah S

    2012-01-01

    Abstract Quantifying tetrodotoxin (TTX) has been a challenge in both ecological and medical research due to the cost, time and training required of most quantification techniques. Here we present a modified Competitive Inhibition Enzymatic Immunoassay for the quantification of TTX, and to aid researchers in the optimization of this technique for widespread use with a high degree of accuracy and repeatability.

  16. An improved competitive inhibition enzymatic immunoassay method for tetrodotoxin quantification

    Directory of Open Access Journals (Sweden)

    Stokes Amber N

    2012-03-01

    Full Text Available Abstract Quantifying tetrodotoxin (TTX has been a challenge in both ecological and medical research due to the cost, time and training required of most quantification techniques. Here we present a modified Competitive Inhibition Enzymatic Immunoassay for the quantification of TTX, and to aid researchers in the optimization of this technique for widespread use with a high degree of accuracy and repeatability.

  17. An improved competitive inhibition enzymatic immunoassay method for tetrodotoxin quantification.

    Science.gov (United States)

    Stokes, Amber N; Williams, Becky L; French, Susannah S

    2012-01-01

    Quantifying tetrodotoxin (TTX) has been a challenge in both ecological and medical research due to the cost, time and training required of most quantification techniques. Here we present a modified Competitive Inhibition Enzymatic Immunoassay for the quantification of TTX, and to aid researchers in the optimization of this technique for widespread use with a high degree of accuracy and repeatability.

  18. Influence of amplicon size on the polymerase chain reaction of Parvovirus B19 genome in formalin-fixed specimens Influência do tamanho do amplicon na reação em cadeia da polimerase na detecção do genoma do PB19 em amostras fixadas em formalina

    Directory of Open Access Journals (Sweden)

    Paulo Roberto Veiga Quemelo

    2009-04-01

    Full Text Available The polymerase chain reaction (PCR has provided diagnosis of archival material, but some fixation methods such as formalin damage DNA and, subsequently, affect PCR analysis, particularly paraffin-embedded tissues. PCR is known due to its high specificity and sensitivity, although some difficulties arise when formalinfixed and paraffin-embedded tissue is used. Not only does this occur due to protein cross-linking, which increases with longer fixation time, but it also happens due to the direct damage that formalin causes in the DNA itself. PCR was used to analyze placenta and fetal organs from 34 samples with suspected Parvovirus B19 infection. It was not possible to amplify Parvovirus B19 DNA using nested-PCR, probably due to the size of the amplicon generated with the first set of primers. We approached this problem by using only the second set of primers. Two out of 34 tissue samples (5,9% were positive by PCR. However, PCR performed on corresponding fetal organs was negative in one of the two. We also observed a negative relation between the thickness of the tissue fragment and the positivity of the samples. In conclusion, although PCR is highly specific and sensitive in fresh or ideally fixed material, a careful standardization of PCR assays is necessary when using formalin fixed paraffin-embedded tissues by applying primers that require smaller DNA fragments for amplification.A reação em cadeia da polimerase (PCR tem fornecido diagnóstico de material de arquivo, mas alguns métodos de fixação, tais como formalina, provocam danos ao DNA e subsequentemente afetam sua análise, particularmente tecidos embebidos em parafina. A PCR é conhecida pela sua alta especificidade e sensibilidade, embora algumas dificuldades ocorram quando o material utilizado foi fixado em formalina e embebido em parafina. Isso não se deve somente pela formação de cross-linkings com proteínas, a qual aumenta com o maior tempo de fixação, mas também pelo

  19. Survey and Evaluate Uncertainty Quantification Methodologies

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Guang; Engel, David W.; Eslinger, Paul W.

    2012-02-01

    The Carbon Capture Simulation Initiative (CCSI) is a partnership among national laboratories, industry and academic institutions that will develop and deploy state-of-the-art computational modeling and simulation tools to accelerate the commercialization of carbon capture technologies from discovery to development, demonstration, and ultimately the widespread deployment to hundreds of power plants. The CCSI Toolset will provide end users in industry with a comprehensive, integrated suite of scientifically validated models with uncertainty quantification, optimization, risk analysis and decision making capabilities. The CCSI Toolset will incorporate commercial and open-source software currently in use by industry and will also develop new software tools as necessary to fill technology gaps identified during execution of the project. The CCSI Toolset will (1) enable promising concepts to be more quickly identified through rapid computational screening of devices and processes; (2) reduce the time to design and troubleshoot new devices and processes; (3) quantify the technical risk in taking technology from laboratory-scale to commercial-scale; and (4) stabilize deployment costs more quickly by replacing some of the physical operational tests with virtual power plant simulations. The goal of CCSI is to deliver a toolset that can simulate the scale-up of a broad set of new carbon capture technologies from laboratory scale to full commercial scale. To provide a framework around which the toolset can be developed and demonstrated, we will focus on three Industrial Challenge Problems (ICPs) related to carbon capture technologies relevant to U.S. pulverized coal (PC) power plants. Post combustion capture by solid sorbents is the technology focus of the initial ICP (referred to as ICP A). The goal of the uncertainty quantification (UQ) task (Task 6) is to provide a set of capabilities to the user community for the quantification of uncertainties associated with the carbon

  20. A newly developed real-time PCR assay for detection and quantification of Fusarium oxysporum and its use in compatible and incompatible interactions with grafted melon genotypes.

    Science.gov (United States)

    Haegi, Anita; Catalano, Valentina; Luongo, Laura; Vitale, Salvatore; Scotton, Michele; Ficcadenti, Nadia; Belisario, Alessandra

    2013-08-01

    A reliable and species-specific real-time quantitative polymerase chain reaction (qPCR) assay was developed for detection of the complex soilborne anamorphic fungus Fusarium oxysporum. The new primer pair, designed on the translation elongation factor 1-α gene with an amplicon of 142 bp, was highly specific to F. oxysporum without cross reactions with other Fusarium spp. The protocol was applied to grafted melon plants for the detection and quantification of F. oxysporum f. sp. melonis, a devastating pathogen of this cucurbit. Grafting technologies are widely used in melon to confer resistance against new virulent races of F. oxysporum f. sp. melonis, while maintaining the properties of valuable commercial varieties. However, the effects on the vascular pathogen colonization have not been fully investigated. Analyses were performed on 'Charentais-T' (susceptible) and 'Nad-1' (resistant) melon cultivars, both used either as rootstock and scion, and inoculated with F. oxysporum f. sp. melonis race 1 and race 1,2. Pathogen development was compared using qPCR and isolations from stem tissues. Early asymptomatic melon infections were detected with a quantification limit of 1 pg of fungal DNA. The qPCR protocol clearly showed that fungal development was highly affected by host-pathogen interaction (compatible or incompatible) and time (days postinoculation). The principal significant effect (P ≤ 0.01) on fungal development was due to the melon genotype used as rootstock, and this effect had a significant interaction with time and F. oxysporum f. sp. melonis race. In particular, the amount of race 1,2 DNA was significantly higher compared with that estimated for race 1 in the incompatible interaction at 18 days postinoculation. The two fungal races were always present in both the rootstock and scion of grafted plants in either the compatible or incompatible interaction.

  1. Quantification of adipose tissue insulin sensitivity

    DEFF Research Database (Denmark)

    Søndergaard, Esben; Jensen, Michael D

    2016-01-01

    In metabolically healthy humans, adipose tissue is exquisitely sensitive to insulin. Similar to muscle and liver, adipose tissue lipolysis is insulin resistant in adults with central obesity and type 2 diabetes. Perhaps uniquely, however, insulin resistance in adipose tissue may directly contribute...... to development of insulin resistance in muscle and liver because of the increased delivery of free fatty acids to those tissues. It has been hypothesized that insulin adipose tissue resistance may precede other metabolic defects in obesity and type 2 diabetes. Therefore, precise and reproducible...... quantification of adipose tissue insulin sensitivity, in vivo, in humans, is an important measure. Unfortunately, no consensus exists on how to determine adipose tissue insulin sensitivity. We review the methods available to quantitate adipose tissue insulin sensitivity and will discuss their strengths and...

  2. Quantification of Osteon Morphology Using Geometric Histomorphometrics.

    Science.gov (United States)

    Dillon, Scott; Cunningham, Craig; Felts, Paul

    2016-03-01

    Many histological methods in forensic anthropology utilize combinations of traditional histomorphometric parameters which may not accurately describe the morphology of microstructural features. Here, we report the novel application of a geometric morphometric method suitable when considering structures without anatomically homologous landmarks for the quantification of complete secondary osteon size and morphology. The method is tested for its suitability in the measurement of intact secondary osteons using osteons digitized from transverse femoral diaphyseal sections prepared from two human individuals. The results of methodological testing demonstrate the efficacy of the technique when applied to intact secondary osteons. In providing accurate characterization of micromorphology within the robust mathematical framework of geometric morphometrics, this method may surpass traditional histomorphometric variables currently employed in forensic research and practice. A preliminary study of the intersectional histomorphometric variation within the femoral diaphysis is made using this geometric histomorphometric method to demonstrate its potential. PMID:26478136

  3. Recurrence quantification analysis of global stock markets

    Science.gov (United States)

    Bastos, João A.; Caiado, Jorge

    2011-04-01

    This study investigates the presence of deterministic dependencies in international stock markets using recurrence plots and recurrence quantification analysis (RQA). The results are based on a large set of free float-adjusted market capitalization stock indices, covering a period of 15 years. The statistical tests suggest that the dynamics of stock prices in emerging markets is characterized by higher values of RQA measures when compared to their developed counterparts. The behavior of stock markets during critical financial events, such as the burst of the technology bubble, the Asian currency crisis, and the recent subprime mortgage crisis, is analyzed by performing RQA in sliding windows. It is shown that during these events stock markets exhibit a distinctive behavior that is characterized by temporary decreases in the fraction of recurrence points contained in diagonal and vertical structures.

  4. On Uncertainty Quantification in Particle Accelerators Modelling

    CERN Document Server

    Adelmann, Andreas

    2015-01-01

    Using a cyclotron based model problem, we demonstrate for the first time the applicability and usefulness of a uncertainty quantification (UQ) approach in order to construct surrogate models for quantities such as emittance, energy spread but also the halo parameter, and construct a global sensitivity analysis together with error propagation and $L_{2}$ error analysis. The model problem is selected in a way that it represents a template for general high intensity particle accelerator modelling tasks. The presented physics problem has to be seen as hypothetical, with the aim to demonstrate the usefulness and applicability of the presented UQ approach and not solving a particulate problem. The proposed UQ approach is based on sparse polynomial chaos expansions and relies on a small number of high fidelity particle accelerator simulations. Within this UQ framework, the identification of most important uncertainty sources is achieved by performing a global sensitivity analysis via computing the so-called Sobols' ...

  5. Quantification Methods of Management Skills in Shipping

    Directory of Open Access Journals (Sweden)

    Riana Iren RADU

    2012-04-01

    Full Text Available Romania can not overcome the financial crisis without business growth, without finding opportunities for economic development and without attracting investment into the country. Successful managers find ways to overcome situations of uncertainty. The purpose of this paper is to determine the managerial skills developed by the Romanian fluvial shipping company NAVROM (hereinafter CNFR NAVROM SA, compared with ten other major competitors in the same domain, using financial information of these companies during the years 2005-2010. For carrying out the work it will be used quantification methods of managerial skills to CNFR NAVROM SA Galati, Romania, as example mentioning the analysis of financial performance management based on profitability ratios, net profit margin, suppliers management, turnover.

  6. Quantification of adipose tissue insulin sensitivity.

    Science.gov (United States)

    Søndergaard, Esben; Jensen, Michael D

    2016-06-01

    In metabolically healthy humans, adipose tissue is exquisitely sensitive to insulin. Similar to muscle and liver, adipose tissue lipolysis is insulin resistant in adults with central obesity and type 2 diabetes. Perhaps uniquely, however, insulin resistance in adipose tissue may directly contribute to development of insulin resistance in muscle and liver because of the increased delivery of free fatty acids to those tissues. It has been hypothesized that insulin adipose tissue resistance may precede other metabolic defects in obesity and type 2 diabetes. Therefore, precise and reproducible quantification of adipose tissue insulin sensitivity, in vivo, in humans, is an important measure. Unfortunately, no consensus exists on how to determine adipose tissue insulin sensitivity. We review the methods available to quantitate adipose tissue insulin sensitivity and will discuss their strengths and weaknesses.

  7. [Quantification of motor activity in biomedicine].

    Science.gov (United States)

    Giannazzo, E

    1993-01-01

    A computer-assisted analysis of motor activity was carried out using ultrasound waves, that are not invasive and free from any kind of interference, because of their specific characteristics. We worked out the Doppler's effect which determines a frequency variation on the reflected wave from any body in motion. That variation is linked to the velocity of the moving body and the superimposition of the emitted wave with those reflected, results in beats, which have a frequency proportional to the motor activity velocity. Our research group planned and carried out an electronic quantification apparatus that can be interfaced with a personal computer system by means of an Analog to Digital acquisition card. The performed test on the apparatus confirmed that the theory that the number of antinodes detected was proportional to the space covered by the moving body. The equipment was also tested on several types of animals.

  8. Uncertainty quantification in DIC with Kriging regression

    Science.gov (United States)

    Wang, Dezhi; DiazDelaO, F. A.; Wang, Weizhuo; Lin, Xiaoshan; Patterson, Eann A.; Mottershead, John E.

    2016-03-01

    A Kriging regression model is developed as a post-processing technique for the treatment of measurement uncertainty in classical subset-based Digital Image Correlation (DIC). Regression is achieved by regularising the sample-point correlation matrix using a local, subset-based, assessment of the measurement error with assumed statistical normality and based on the Sum of Squared Differences (SSD) criterion. This leads to a Kriging-regression model in the form of a Gaussian process representing uncertainty on the Kriging estimate of the measured displacement field. The method is demonstrated using numerical and experimental examples. Kriging estimates of displacement fields are shown to be in excellent agreement with 'true' values for the numerical cases and in the experimental example uncertainty quantification is carried out using the Gaussian random process that forms part of the Kriging model. The root mean square error (RMSE) on the estimated displacements is produced and standard deviations on local strain estimates are determined.

  9. Recurrence quantification analysis theory and best practices

    CERN Document Server

    Jr, Jr; Marwan, Norbert

    2015-01-01

    The analysis of recurrences in dynamical systems by using recurrence plots and their quantification is still an emerging field.  Over the past decades recurrence plots have proven to be valuable data visualization and analysis tools in the theoretical study of complex, time-varying dynamical systems as well as in various applications in biology, neuroscience, kinesiology, psychology, physiology, engineering, physics, geosciences, linguistics, finance, economics, and other disciplines.   This multi-authored book intends to comprehensively introduce and showcase recent advances as well as established best practices concerning both theoretical and practical aspects of recurrence plot based analysis.  Edited and authored by leading researcher in the field, the various chapters address an interdisciplinary readership, ranging from theoretical physicists to application-oriented scientists in all data-providing disciplines.

  10. Theoretical model and quantification of reflectance photometer

    Institute of Scientific and Technical Information of China (English)

    Lihua Huang; Youbao Zhang; Chengke Xie; Jianfeng Qu; Huijie Huang; Xiangzhao Wang

    2009-01-01

    @@ The surface morphology of lateral flow (LF) strip is examined by scanning electron microscope (SEM) and the diffuse reflection of porous strip with or without nanogold particles is investigated.Based on the scattering and absorption of nanogold particles, a reflectance photometer is developed for quantification of LF strip with nanogold particles as reporter.The integration of reflection optical density is to indicate the signals of test line and control line.As an example, serial dilutions of microalbunminuria (MAU) solution are used to calibrate the performance of the reflectance photometer.The dose response curve is fitted with a four-parameter logistic mathematical model for the determination of an unknown MAU concentration.The response curve spans a dynamic range of 5 to 200 μg/ml.The developed reflectance photometer can realize simple and quantitative detection of analyte on nanogold-labeled LF strip.

  11. Quantification of Condylar Resorption in TMJ Osteoarthritis

    Science.gov (United States)

    Cevidanes, LHS; Hajati, A-K; Paniagua, B; Lim, PF; Walker, DG; Palconet, G; Nackley, AG; Styner, M; Ludlow, JB; Zhu, H; Phillips, C

    2010-01-01

    OBJECTIVE This study was performed to determine the condylar morphological variation of osteoarthritic (OA) and asymptomatic temporomandibular joints (TMJ) and to determine its correlation with pain intensity and duration. STUDY DESIGN Three dimensional surface models of mandibular condyles were constructed from Cone-Beam CT images of 29 female patients with TMJ OA (Research Diagnostic Criteria for Temporomandibular Disorders Group III) and 36 female asymptomatic subjects. Shape Correspondence was used to localize and quantify the condylar morphology. Statistical analysis was performed with MANCOVA analysis using Hotelling T2 metric based on covariance matrices, and Pearson correlation. RESULTS OA condylar morphology was statistically significantly different from the asymptomatic condyles (p<0.05). 3D morphological variation of the OA condyles was significantly correlated with pain intensity and duration. CONCLUSION 3D quantification of condylar morphology revealed profound differences between OA and asymptomatic condyles and the extent of the resorptive changes paralleled pain severity and duration. PMID:20382043

  12. Quantification of uncertainties for application in detonation simulation

    Science.gov (United States)

    Zheng, Miao; Ma, Zhibo

    2016-06-01

    Numerical simulation has become an important means in designing detonation systems, and the quantification of its uncertainty is also necessary to reliability certification. As to quantifying the uncertainty, it is the most important to analyze how the uncertainties occur and develop, and how the simulations develop from benchmark models to new models. Based on the practical needs of engineering and the technology of verification & validation, a framework of QU(quantification of uncertainty) is brought forward in the case that simulation is used on detonation system for scientific prediction. An example is offered to describe the general idea of quantification of simulation uncertainties.

  13. Toolbox Approaches Using Molecular Markers and 16S rRNA Gene Amplicon Data Sets for Identification of Fecal Pollution in Surface Water.

    Science.gov (United States)

    Ahmed, W; Staley, C; Sadowsky, M J; Gyawali, P; Sidhu, J P S; Palmer, A; Beale, D J; Toze, S

    2015-10-01

    In this study, host-associated molecular markers and bacterial 16S rRNA gene community analysis using high-throughput sequencing were used to identify the sources of fecal pollution in environmental waters in Brisbane, Australia. A total of 92 fecal and composite wastewater samples were collected from different host groups (cat, cattle, dog, horse, human, and kangaroo), and 18 water samples were collected from six sites (BR1 to BR6) along the Brisbane River in Queensland, Australia. Bacterial communities in the fecal, wastewater, and river water samples were sequenced. Water samples were also tested for the presence of bird-associated (GFD), cattle-associated (CowM3), horse-associated, and human-associated (HF183) molecular markers, to provide multiple lines of evidence regarding the possible presence of fecal pollution associated with specific hosts. Among the 18 water samples tested, 83%, 33%, 17%, and 17% were real-time PCR positive for the GFD, HF183, CowM3, and horse markers, respectively. Among the potential sources of fecal pollution in water samples from the river, DNA sequencing tended to show relatively small contributions from wastewater treatment plants (up to 13% of sequence reads). Contributions from other animal sources were rarely detected and were very small (pollution in an urban river. This study is a proof of concept, and based on the results, we recommend using bacterial community analysis (where possible) along with PCR detection or quantification of host-associated molecular markers to provide information on the sources of fecal pollution in waterways. PMID:26231650

  14. Extended Forward Sensitivity Analysis for Uncertainty Quantification

    Energy Technology Data Exchange (ETDEWEB)

    Haihua Zhao; Vincent A. Mousseau

    2008-09-01

    This report presents the forward sensitivity analysis method as a means for quantification of uncertainty in system analysis. The traditional approach to uncertainty quantification is based on a “black box” approach. The simulation tool is treated as an unknown signal generator, a distribution of inputs according to assumed probability density functions is sent in and the distribution of the outputs is measured and correlated back to the original input distribution. This approach requires large number of simulation runs and therefore has high computational cost. Contrary to the “black box” method, a more efficient sensitivity approach can take advantage of intimate knowledge of the simulation code. In this approach equations for the propagation of uncertainty are constructed and the sensitivity is solved for as variables in the same simulation. This “glass box” method can generate similar sensitivity information as the above “black box” approach with couples of runs to cover a large uncertainty region. Because only small numbers of runs are required, those runs can be done with a high accuracy in space and time ensuring that the uncertainty of the physical model is being measured and not simply the numerical error caused by the coarse discretization. In the forward sensitivity method, the model is differentiated with respect to each parameter to yield an additional system of the same size as the original one, the result of which is the solution sensitivity. The sensitivity of any output variable can then be directly obtained from these sensitivities by applying the chain rule of differentiation. We extend the forward sensitivity method to include time and spatial steps as special parameters so that the numerical errors can be quantified against other physical parameters. This extension makes the forward sensitivity method a much more powerful tool to help uncertainty analysis. By knowing the relative sensitivity of time and space steps with other

  15. Synthesis and Review: Advancing agricultural greenhouse gas quantification

    International Nuclear Information System (INIS)

    Reducing emissions of agricultural greenhouse gases (GHGs), such as methane and nitrous oxide, and sequestering carbon in the soil or in living biomass can help reduce the impact of agriculture on climate change while improving productivity and reducing resource use. There is an increasing demand for improved, low cost quantification of GHGs in agriculture, whether for national reporting to the United Nations Framework Convention on Climate Change (UNFCCC), underpinning and stimulating improved practices, establishing crediting mechanisms, or supporting green products. This ERL focus issue highlights GHG quantification to call attention to our existing knowledge and opportunities for further progress. In this article we synthesize the findings of 21 papers on the current state of global capability for agricultural GHG quantification and visions for its improvement. We conclude that strategic investment in quantification can lead to significant global improvement in agricultural GHG estimation in the near term. (paper)

  16. Quantification of Uncertainties in Integrated Spacecraft System Models Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed effort is to investigate a novel uncertainty quantification (UQ) approach based on non-intrusive polynomial chaos (NIPC) for computationally efficient...

  17. Uncertainty Quantification for Production Navier-Stokes Solvers Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The uncertainty quantification methods developed under this program are designed for use with current state-of-the-art flow solvers developed by and in use at NASA....

  18. A Micropillar Compression Methodology for Ductile Damage Quantification

    NARCIS (Netherlands)

    Tasan, C.C.; Hoefnagels, J.P.M.; Geers, M.G.D.

    2012-01-01

    Microstructural damage evolution is reported to influence significantly the failures of new high-strength alloys. Its accurate quantification is, therefore, critical for (1) microstructure optimization and (2) continuum damage models to predict failures of these materials. As existing methodologies

  19. Quantification of Uncertainties in Integrated Spacecraft System Models Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The objective for the Phase II effort will be to develop a comprehensive, efficient, and flexible uncertainty quantification (UQ) framework implemented within a...

  20. Construction of recombinant pseudorabies viruses by using PRV BACs deficient in IE180 or pac sequences: Application of vBAC90D recombinant virus to production of PRV amplicons.

    Science.gov (United States)

    Lerma, L; Muñoz, A L; Wagner, S; Dinu, M; Martín, B; Tabarés, E

    2016-02-01

    We describe a simple and efficient method to obtain recombinant pseudorabies virus (PRV) in mammalian cells by using the PRV BACs, PBAC80 deficient in pac sequences and PBAC90 deficient in the IE180 gene. These essential viral sequences were used as targets to obtain viable recombinant viruses. PBAC80 was constructed, confirmed to encode a copy of the IE180 gene regulated by the inducible Ptet promoter, and used to obtain recombinant attenuated PRV viruses that express the EGFP protein (PRV-BT80GF virus). PBAC90 was used to obtain the vBAC90D virus, deficient in IE180 and free of replication-competent revertants, and which can be used as a helper in the production of PRV amplicons. PMID:26756577

  1. Droplet digital PCR, the new tool in HIV reservoir quantification?

    OpenAIRE

    De Spiegelaere, Ward; Kiselinova, Maja; Malatinkova, Eva; Pasternak, Alexander; Berkhout, Ben; Vandekerckhove, Linos

    2013-01-01

    Background: Digital PCR is a relatively old concept for absolute quantification of DNA using PCR, but recent technological developments allowed its wide use. The current state of art technique for performing digital PCR is based on microdroplet technology. Direct absolute quantification relieves the necessity of standard curves and increases assay accuracy. In addition, the end point PCR set-up allows higher assay flexibility and decreases quantitative bias due to variations in PCR efficiency...

  2. Volumetric motion quantification by 3D tissue phase mapped CMR

    OpenAIRE

    Lutz Anja; Paul Jan; Bornstedt Axel; Nienhaus G; Etyngier Patrick; Bernhardt Peter; Rottbauer Wolfgang; Rasche Volker

    2012-01-01

    Abstract Background The objective of this study was the quantification of myocardial motion from 3D tissue phase mapped (TPM) CMR. Recent work on myocardial motion quantification by TPM has been focussed on multi-slice 2D acquisitions thus excluding motion information from large regions of the left ventricle. Volumetric motion assessment appears an important next step towards the understanding of the volumetric myocardial motion and hence may further improve diagnosis and treatments in patien...

  3. Quantification of bromophenols in Islay whiskies.

    Science.gov (United States)

    Bendig, Paul; Lehnert, Katja; Vetter, Walter

    2014-04-01

    Two single malt whiskies from the Scottish island Islay, i.e., Laphroiag and Lagavulin, are characterized by an iodine-like flavor associated with marine environments. In this study we investigated if this flavor impression could be due to bromophenols which are character impact compounds of marine fish and shrimps. In this study we developed a method suited for the determination of dibromo- and tribromophenols in whisky. Aliquots were O-acetylated, and quantification was carried out with gas chromatography with electron-capture negative ion mass spectrometry (GC/ECNI-MS). Both Islay whiskies contained more than 400 ng/L bromophenols with 2,6-dibromophenol being the most relevant homologue (>300 ng/L, respectively). These concentrations are at least 1 order of magnitude higher than the taste threshold of 2,6-dibromophenol in water. A third Islay whisky, Bowmore, contained ∼100 ng/L bromophenols while seventeen other whiskies from other regions in Scotland as well as from the USA, Ireland, and Germany contained at least 1 order of magnitude less than the two whiskies with the marine taste. Accordingly, bromophenols may contribute to the marine flavor and taste of Laphroaig and Lagavulin.

  4. Quantification of variability in trichome patterns

    Directory of Open Access Journals (Sweden)

    Bettina eGreese

    2014-11-01

    Full Text Available While pattern formation is studied in various areas of biology, little is known about the intrinsic noise leading to variations between individual realizations of the pattern. One prominent example for de novo pattern formation in plants is the patterning of trichomes on Arabidopsis leaves, which involves genetic regulation and cell-to-cell communication. These processes are potentially variable due to , e.g., the abundance of cell components or environmental conditions. To elevate the understanding of the regulatory processes underlying the pattern formation it is crucial to quantitatively analyze the variability in naturally occurring patterns. Here, we review recent approaches towards characterization of noise on trichome initiation. We present methods for the quantification of spatial patterns, which are the basis for data-driven mathematical modeling and enable the analysis of noise from different sources. Besides the insight gained on trichome formation, the examination of observed trichome patterns also shows that highly regulated biological processes can be substantially affected by variability.

  5. Legionella spp. isolation and quantification from greywater.

    Science.gov (United States)

    Rodríguez-Martínez, Sara; Blanky, Marina; Friedler, Eran; Halpern, Malka

    2015-01-01

    Legionella, an opportunistic human pathogen whose natural environment is water, is transmitted to humans through inhalation of contaminated aerosols. Legionella has been isolated from a high diversity of water types. Due its importance as a pathogen, two ISO protocols have been developed for its monitoring. However, these two protocols are not suitable for analyzing Legionella in greywater (GW). GW is domestic wastewater excluding the inputs from toilets and kitchen. It can serve as an alternative water source, mainly for toilet flushing and garden irrigation; both producing aerosols that can cause a risk for Legionella infection. Hence, before reuse, GW has to be treated and its quality needs to be monitored. The difficulty of Legionella isolation from GW strives in the very high load of contaminant bacteria. Here we describe a modification of the ISO protocol 11731:1998 that enables the isolation and quantification of Legionella from GW samples. The following modifications were made:•To enable isolation of Legionella from greywater, a pre-filtration step that removes coarse matter is recommended.•Legionella can be isolated after a combined acid-thermic treatment that eliminates the high load of contaminant bacteria in the sample. PMID:26740925

  6. Low cost high performance uncertainty quantification

    KAUST Repository

    Bekas, C.

    2009-01-01

    Uncertainty quantification in risk analysis has become a key application. In this context, computing the diagonal of inverse covariance matrices is of paramount importance. Standard techniques, that employ matrix factorizations, incur a cubic cost which quickly becomes intractable with the current explosion of data sizes. In this work we reduce this complexity to quadratic with the synergy of two algorithms that gracefully complement each other and lead to a radically different approach. First, we turned to stochastic estimation of the diagonal. This allowed us to cast the problem as a linear system with a relatively small number of multiple right hand sides. Second, for this linear system we developed a novel, mixed precision, iterative refinement scheme, which uses iterative solvers instead of matrix factorizations. We demonstrate that the new framework not only achieves the much needed quadratic cost but in addition offers excellent opportunities for scaling at massively parallel environments. We based our implementation on BLAS 3 kernels that ensure very high processor performance. We achieved a peak performance of 730 TFlops on 72 BG/P racks, with a sustained performance 73% of theoretical peak. We stress that the techniques presented in this work are quite general and applicable to several other important applications. Copyright © 2009 ACM.

  7. Tissue quantification for development of pediatric phantom

    International Nuclear Information System (INIS)

    The optimization of the risk- benefit ratio is a major concern in the pediatric radiology, due to the greater vulnerability of children to the late somatic effects and genetic effects of exposure to radiation compared to adults. In Brazil, it is estimated that the causes of death from head trauma are 18 % for the age group between 1-5 years and the radiograph is the primary diagnostic test for the detection of skull fracture . Knowing that the image quality is essential to ensure the identification of structures anatomical and minimizing errors diagnostic interpretation, this paper proposed the development and construction of homogeneous phantoms skull, for the age group 1-5 years. The construction of the phantoms homogeneous was performed using the classification and quantification of tissue present in the skull of pediatric patients. In this procedure computational algorithms were used, using Matlab, to quantify distinct biological tissues present in the anatomical regions studied , using pictures retrospective CT scans. Preliminary data obtained from measurements show that between the ages of 1-5 years, assuming an average anteroposterior diameter of the pediatric skull region of the 145.73 ± 2.97 mm, can be represented by 92.34 mm ± 5.22 of lucite and 1.75 ± 0:21 mm of aluminum plates of a provision of PEP (Pacient equivalent phantom). After its construction, the phantoms will be used for image and dose optimization in pediatric protocols process to examinations of computerized radiography

  8. Uncertainty Quantification for Cargo Hold Fires

    CERN Document Server

    DeGennaro, Anthony M; Martinelli, Luigi; Rowley, Clarence W

    2015-01-01

    The purpose of this study is twofold -- first, to introduce the application of high-order discontinuous Galerkin methods to buoyancy-driven cargo hold fire simulations, second, to explore statistical variation in the fluid dynamics of a cargo hold fire given parameterized uncertainty in the fire source location and temperature. Cargo hold fires represent a class of problems that require highly-accurate computational methods to simulate faithfully. Hence, we use an in-house discontinuous Galerkin code to treat these flows. Cargo hold fires also exhibit a large amount of uncertainty with respect to the boundary conditions. Thus, the second aim of this paper is to quantify the resulting uncertainty in the flow, using tools from the uncertainty quantification community to ensure that our efforts require a minimal number of simulations. We expect that the results of this study will provide statistical insight into the effects of fire location and temperature on cargo fires, and also assist in the optimization of f...

  9. Quantification of perceived macro-uniformity

    Science.gov (United States)

    Lee, Ki-Youn; Bang, Yousun; Choh, Heui-Keun

    2011-01-01

    Macro-uniformity refers to the subjective impression of overall uniformity in the print sample. By the efforts of INCITS W1.1 team, macro-uniformity is categorized into five types of attributes: banding, streaks, mottle, gradients, and moiré patterns, and the ruler samples are generated with perceptual scales. W1.1 macro-uniformity ruler is useful for judging the levels of print defect, but it is not an easy task to reproduce the samples having the same perceptual scales at different times in different places. An objective quantification method is more helpful and convenient for developers to analyze print quality and design printing system components. In this paper, we propose a method for measuring perceived macro-uniformity for a given print using a flat-bed scanner. First, banding, 2D noise, and gradients are separately measured, and they are converted to the perceptual scales based on subjective results of each attribute. The correlation coefficients between the measured values of the attributes and the perceptual scales are 0.92, 0.97, and 0.86, respectively. Another subjective test is performed to find the relationship between the overall macro-uniformity and the three attributes. The weighting factors are obtained by the experimental result, and the final macro-uniformity grade is determined by the weighted sums of each attribute.

  10. Legionella spp. isolation and quantification from greywater

    Science.gov (United States)

    Rodríguez-Martínez, Sara; Blanky, Marina; Friedler, Eran; Halpern, Malka

    2015-01-01

    Legionella, an opportunistic human pathogen whose natural environment is water, is transmitted to humans through inhalation of contaminated aerosols. Legionella has been isolated from a high diversity of water types. Due its importance as a pathogen, two ISO protocols have been developed for its monitoring. However, these two protocols are not suitable for analyzing Legionella in greywater (GW). GW is domestic wastewater excluding the inputs from toilets and kitchen. It can serve as an alternative water source, mainly for toilet flushing and garden irrigation; both producing aerosols that can cause a risk for Legionella infection. Hence, before reuse, GW has to be treated and its quality needs to be monitored. The difficulty of Legionella isolation from GW strives in the very high load of contaminant bacteria. Here we describe a modification of the ISO protocol 11731:1998 that enables the isolation and quantification of Legionella from GW samples. The following modifications were made:•To enable isolation of Legionella from greywater, a pre-filtration step that removes coarse matter is recommended.•Legionella can be isolated after a combined acid-thermic treatment that eliminates the high load of contaminant bacteria in the sample. PMID:26740925

  11. Quantification of food intake in Drosophila.

    Directory of Open Access Journals (Sweden)

    Richard Wong

    Full Text Available Measurement of food intake in the fruit fly Drosophila melanogaster is often necessary for studies of behaviour, nutrition and drug administration. There is no reliable and agreed method for measuring food intake of flies in undisturbed, steady state, and normal culture conditions. We report such a method, based on measurement of feeding frequency by proboscis-extension, validated by short-term measurements of food dye intake. We used the method to demonstrate that (a female flies feed more frequently than males, (b flies feed more often when housed in larger groups and (c fly feeding varies at different times of the day. We also show that alterations in food intake are not induced by dietary restriction or by a null mutation of the fly insulin receptor substrate chico. In contrast, mutation of takeout increases food intake by increasing feeding frequency while mutation of ovo(D increases food intake by increasing the volume of food consumed per proboscis-extension. This approach provides a practical and reliable method for quantification of food intake in Drosophila under normal, undisturbed culture conditions.

  12. Shape regression for vertebra fracture quantification

    Science.gov (United States)

    Lund, Michael Tillge; de Bruijne, Marleen; Tanko, Laszlo B.; Nielsen, Mads

    2005-04-01

    Accurate and reliable identification and quantification of vertebral fractures constitute a challenge both in clinical trials and in diagnosis of osteoporosis. Various efforts have been made to develop reliable, objective, and reproducible methods for assessing vertebral fractures, but at present there is no consensus concerning a universally accepted diagnostic definition of vertebral fractures. In this project we want to investigate whether or not it is possible to accurately reconstruct the shape of a normal vertebra, using a neighbouring vertebra as prior information. The reconstructed shape can then be used to develop a novel vertebra fracture measure, by comparing the segmented vertebra shape with its reconstructed normal shape. The vertebrae in lateral x-rays of the lumbar spine were manually annotated by a medical expert. With this dataset we built a shape model, with equidistant point distribution between the four corner points. Based on the shape model, a multiple linear regression model of a normal vertebra shape was developed for each dataset using leave-one-out cross-validation. The reconstructed shape was calculated for each dataset using these regression models. The average prediction error for the annotated shape was on average 3%.

  13. Standardless quantification methods in electron probe microanalysis

    Energy Technology Data Exchange (ETDEWEB)

    Trincavelli, Jorge, E-mail: trincavelli@famaf.unc.edu.ar [Facultad de Matemática, Astronomía y Física, Universidad Nacional de Córdoba, Ciudad Universitaria, 5000 Córdoba (Argentina); Instituto de Física Enrique Gaviola, Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina, Medina Allende s/n, Ciudad Universitaria, 5000 Córdoba (Argentina); Limandri, Silvina, E-mail: s.limandri@conicet.gov.ar [Facultad de Matemática, Astronomía y Física, Universidad Nacional de Córdoba, Ciudad Universitaria, 5000 Córdoba (Argentina); Instituto de Física Enrique Gaviola, Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina, Medina Allende s/n, Ciudad Universitaria, 5000 Córdoba (Argentina); Bonetto, Rita, E-mail: bonetto@quimica.unlp.edu.ar [Centro de Investigación y Desarrollo en Ciencias Aplicadas Dr. Jorge Ronco, Consejo Nacional de Investigaciones Científicas y Técnicas de la República Argentina, Facultad de Ciencias Exactas, de la Universidad Nacional de La Plata, Calle 47 N° 257, 1900 La Plata (Argentina)

    2014-11-01

    The elemental composition of a solid sample can be determined by electron probe microanalysis with or without the use of standards. The standardless algorithms are quite faster than the methods that require standards; they are useful when a suitable set of standards is not available or for rough samples, and also they help to solve the problem of current variation, for example, in equipments with cold field emission gun. Due to significant advances in the accuracy achieved during the last years, product of the successive efforts made to improve the description of generation, absorption and detection of X-rays, the standardless methods have increasingly become an interesting option for the user. Nevertheless, up to now, algorithms that use standards are still more precise than standardless methods. It is important to remark, that care must be taken with results provided by standardless methods that normalize the calculated concentration values to 100%, unless an estimate of the errors is reported. In this work, a comprehensive discussion of the key features of the main standardless quantification methods, as well as the level of accuracy achieved by them is presented. - Highlights: • Standardless methods are a good alternative when no suitable standards are available. • Their accuracy reaches 10% for 95% of the analyses when traces are excluded. • Some of them are suitable for the analysis of rough samples.

  14. Quantification of the vocal folds’ dynamic displacements

    Science.gov (United States)

    del Socorro Hernández-Montes, María; Muñoz, Silvino; De La Torre, Manuel; Flores, Mauricio; Pérez, Carlos; Mendoza-Santoyo, Fernando

    2016-05-01

    Fast dynamic data acquisition techniques are required to investigate the motional behavior of the vocal folds (VFs) when they are subjected to a steady air-flow through the trachea. High-speed digital holographic interferometry (DHI) is a non-invasive full-field-of-view technique that has proved its usefulness to study rapid and non-repetitive object movements. Hence it is an ideal technique used here to measure VF displacements and vibration patterns at 2000 fps. Analyses from a set of 200 displacement images showed that VFs’ vibration cycles are established along their width (y) and length (x). Furthermore, the maximum deformation for the right and left VFs’ area may be quantified from these images, which in itself represents an important result in the characterization of this structure. At a controlled air pressure, VF displacements fall within the range ~100-1740 nm, with a calculated precision and accuracy that yields a variation coefficient of 1.91%. High-speed acquisition of full-field images of VFs and their displacement quantification are on their own significant data in the study of their functional and physiological behavior since voice quality and production depend on how they vibrate, i.e. their displacement amplitude and frequency. Additionally, the use of high speed DHI avoids prolonged examinations and represents a significant scientific and technological alternative contribution in advancing the knowledge and working mechanisms of these tissues.

  15. Quantification of biological aging in young adults

    Science.gov (United States)

    Belsky, Daniel W.; Caspi, Avshalom; Houts, Renate; Cohen, Harvey J.; Corcoran, David L.; Danese, Andrea; Harrington, HonaLee; Israel, Salomon; Levine, Morgan E.; Schaefer, Jonathan D.; Sugden, Karen; Williams, Ben; Yashin, Anatoli I.; Poulton, Richie; Moffitt, Terrie E.

    2015-01-01

    Antiaging therapies show promise in model organism research. Translation to humans is needed to address the challenges of an aging global population. Interventions to slow human aging will need to be applied to still-young individuals. However, most human aging research examines older adults, many with chronic disease. As a result, little is known about aging in young humans. We studied aging in 954 young humans, the Dunedin Study birth cohort, tracking multiple biomarkers across three time points spanning their third and fourth decades of life. We developed and validated two methods by which aging can be measured in young adults, one cross-sectional and one longitudinal. Our longitudinal measure allows quantification of the pace of coordinated physiological deterioration across multiple organ systems (e.g., pulmonary, periodontal, cardiovascular, renal, hepatic, and immune function). We applied these methods to assess biological aging in young humans who had not yet developed age-related diseases. Young individuals of the same chronological age varied in their “biological aging” (declining integrity of multiple organ systems). Already, before midlife, individuals who were aging more rapidly were less physically able, showed cognitive decline and brain aging, self-reported worse health, and looked older. Measured biological aging in young adults can be used to identify causes of aging and evaluate rejuvenation therapies. PMID:26150497

  16. Cross recurrence quantification for cover song identification

    Energy Technology Data Exchange (ETDEWEB)

    Serra, Joan; Serra, Xavier; Andrzejak, Ralph G [Department of Information and Communication Technologies, Universitat Pompeu Fabra, Roc Boronat 138, 08018 Barcelona (Spain)], E-mail: joan.serraj@upf.edu

    2009-09-15

    There is growing evidence that nonlinear time series analysis techniques can be used to successfully characterize, classify, or process signals derived from real-world dynamics even though these are not necessarily deterministic and stationary. In the present study, we proceed in this direction by addressing an important problem our modern society is facing, the automatic classification of digital information. In particular, we address the automatic identification of cover songs, i.e. alternative renditions of a previously recorded musical piece. For this purpose, we here propose a recurrence quantification analysis measure that allows the tracking of potentially curved and disrupted traces in cross recurrence plots (CRPs). We apply this measure to CRPs constructed from the state space representation of musical descriptor time series extracted from the raw audio signal. We show that our method identifies cover songs with a higher accuracy as compared to previously published techniques. Beyond the particular application proposed here, we discuss how our approach can be useful for the characterization of a variety of signals from different scientific disciplines. We study coupled Roessler dynamics with stochastically modulated mean frequencies as one concrete example to illustrate this point.

  17. Quantification of moving target cyber defenses

    Science.gov (United States)

    Farris, Katheryn A.; Cybenko, George

    2015-05-01

    Current network and information systems are static, making it simple for attackers to maintain an advantage. Adaptive defenses, such as Moving Target Defenses (MTD) have been developed as potential "game-changers" in an effort to increase the attacker's workload. With many new methods being developed, it is difficult to accurately quantify and compare their overall costs and effectiveness. This paper compares the tradeoffs between current approaches to the quantification of MTDs. We present results from an expert opinion survey on quantifying the overall effectiveness, upfront and operating costs of a select set of MTD techniques. We find that gathering informed scientific opinions can be advantageous for evaluating such new technologies as it offers a more comprehensive assessment. We end by presenting a coarse ordering of a set of MTD techniques from most to least dominant. We found that seven out of 23 methods rank as the more dominant techniques. Five of which are techniques of either address space layout randomization or instruction set randomization. The remaining two techniques are applicable to software and computer platforms. Among the techniques that performed the worst are those primarily aimed at network randomization.

  18. Targeted Amplicon Sequencing for Single-Nucleotide-Polymorphism Genotyping of Attaching and Effacing Escherichia coli O26:H11 Cattle Strains via a High-Throughput Library Preparation Technique.

    Science.gov (United States)

    Ison, Sarah A; Delannoy, Sabine; Bugarel, Marie; Nagaraja, Tiruvoor G; Renter, David G; den Bakker, Henk C; Nightingale, Kendra K; Fach, Patrick; Loneragan, Guy H

    2015-11-13

    Enterohemorrhagic Escherichia coli (EHEC) O26:H11, a serotype within Shiga toxin-producing E. coli (STEC) that causes severe human disease, has been considered to have evolved from attaching and effacing E. coli (AEEC) O26:H11 through the acquisition of a Shiga toxin-encoding gene. Targeted amplicon sequencing using next-generation sequencing technology of 48 phylogenetically informative single-nucleotide polymorphisms (SNPs) and three SNPs differentiating Shiga toxin-positive (stx-positive) strains from Shiga toxin-negative (stx-negative) strains were used to infer the phylogenetic relationships of 178 E. coli O26:H11 strains (6 stx-positive strains and 172 stx-negative AEEC strains) from cattle feces to 7 publically available genomes of human clinical strains. The AEEC cattle strains displayed synonymous SNP genotypes with stx2-positive sequence type 29 (ST29) human O26:H11 strains, while stx1 ST21 human and cattle strains clustered separately, demonstrating the close phylogenetic relatedness of these Shiga toxin-negative AEEC cattle strains and human clinical strains. With the exception of seven stx-negative strains, five of which contained espK, three stx-related SNPs differentiated the STEC strains from non-STEC strains, supporting the hypothesis that these AEEC cattle strains could serve as a potential reservoir for new or existing pathogenic human strains. Our results support the idea that targeted amplicon sequencing for SNP genotyping expedites strain identification and genetic characterization of E. coli O26:H11, which is important for food safety and public health.

  19. Molecular identification of CTX-M and blaOXY/K1 β-lactamase genes in Enterobacteriaceae by sequencing of universal M13-sequence tagged PCR-amplicons

    Directory of Open Access Journals (Sweden)

    Tärnberg Maria

    2009-01-01

    Full Text Available Abstract Background Plasmid encoded blaCTX-M enzymes represent an important sub-group of class A β-lactamases causing the ESBL phenotype which is increasingly found in Enterobacteriaceae including Klebsiella spp. Molecular typing of clinical ESBL-isolates has become more and more important for prevention of the dissemination of ESBL-producers among nosocomial environment. Methods Multiple displacement amplified DNA derived from 20 K. pneumoniae and 34 K. oxytoca clinical isolates with an ESBL-phenotype was used in a universal CTX-M PCR amplification assay. Identification and differentiation of blaCTX-M and blaOXY/K1 sequences was obtained by DNA sequencing of M13-sequence-tagged CTX-M PCR-amplicons using a M13-specific sequencing primer. Results Nine out of 20 K. pneumoniae clinical isolates had a blaCTX-M genotype. Interestingly, we found that the universal degenerated primers also amplified the chromosomally located K1-gene in all 34 K. oxytoca clinical isolates. Molecular identification and differentiation between blaCTX-M and blaOXY/K1-genes could only been achieved by sequencing of the PCR-amplicons. In silico analysis revealed that the universal degenerated CTX-M primer-pair used here might also amplify the chromosomally located blaOXY and K1-genes in Klebsiella spp. and K1-like genes in other Enterobacteriaceae. Conclusion The PCR-based molecular typing method described here enables a rapid and reliable molecular identification of blaCTX-M, and blaOXY/K1-genes. The principles used in this study could also be applied to any situation in which antimicrobial resistance genes would need to be sequenced.

  20. Volumetric motion quantification by 3D tissue phase mapped CMR

    Directory of Open Access Journals (Sweden)

    Lutz Anja

    2012-10-01

    Full Text Available Abstract Background The objective of this study was the quantification of myocardial motion from 3D tissue phase mapped (TPM CMR. Recent work on myocardial motion quantification by TPM has been focussed on multi-slice 2D acquisitions thus excluding motion information from large regions of the left ventricle. Volumetric motion assessment appears an important next step towards the understanding of the volumetric myocardial motion and hence may further improve diagnosis and treatments in patients with myocardial motion abnormalities. Methods Volumetric motion quantification of the complete left ventricle was performed in 12 healthy volunteers and two patients applying a black-blood 3D TPM sequence. The resulting motion field was analysed regarding motion pattern differences between apical and basal locations as well as for asynchronous motion pattern between different myocardial segments in one or more slices. Motion quantification included velocity, torsion, rotation angle and strain derived parameters. Results All investigated motion quantification parameters could be calculated from the 3D-TPM data. Parameters quantifying hypokinetic or asynchronous motion demonstrated differences between motion impaired and healthy myocardium. Conclusions 3D-TPM enables the gapless volumetric quantification of motion abnormalities of the left ventricle, which can be applied in future application as additional information to provide a more detailed analysis of the left ventricular function.

  1. Quantification of isotopic turnover in agricultural systems

    Science.gov (United States)

    Braun, A.; Auerswald, K.; Schnyder, H.

    2012-04-01

    The isotopic turnover, which is a proxy for the metabolic rate, is gaining scientific importance. It is quantified for an increasing range of organisms, from microorganisms over plants to animals including agricultural livestock. Additionally, the isotopic turnover is analyzed on different scales, from organs to organisms to ecosystems and even to the biosphere. In particular, the quantification of the isotopic turnover of specific tissues within the same organism, e.g. organs like liver and muscle and products like milk and faeces, has brought new insights to improve understanding of nutrient cycles and fluxes, respectively. Thus, the knowledge of isotopic turnover is important in many areas, including physiology, e.g. milk synthesis, ecology, e.g. soil retention time of water, and medical science, e.g. cancer diagnosis. So far, the isotopic turnover is quantified by applying time, cost and expertise intensive tracer experiments. Usually, this comprises two isotopic equilibration periods. A first equilibration period with a constant isotopic input signal is followed by a second equilibration period with a distinct constant isotopic input signal. This yields a smooth signal change from the first to the second signal in the object under consideration. This approach reveals at least three major problems. (i) The input signals must be controlled isotopically, which is almost impossible in many realistic cases like free ranging animals. (ii) Both equilibration periods may be very long, especially when the turnover rate of the object under consideration is very slow, which aggravates the first problem. (iii) The detection of small or slow pools is improved by large isotopic signal changes, but large isotopic changes also involve a considerable change in the input material; e.g. animal studies are usually carried out as diet-switch experiments, where the diet is switched between C3 and C4 plants, since C3 and C4 plants differ strongly in their isotopic signal. The

  2. Quantification of nanowire uptake by live cells

    KAUST Repository

    Margineanu, Michael B.

    2015-05-01

    Nanostructures fabricated by different methods have become increasingly important for various applications at the cellular level. In order to understand how these nanostructures “behave” and for studying their internalization kinetics, several attempts have been made at tagging and investigating their interaction with living cells. In this study, magnetic iron nanowires with an iron oxide layer are coated with (3-Aminopropyl)triethoxysilane (APTES), and subsequently labeled with a fluorogenic pH-dependent dye pHrodo™ Red, covalently bound to the aminosilane surface. Time-lapse live imaging of human colon carcinoma HCT 116 cells interacting with the labeled iron nanowires is performed for 24 hours. As the pHrodo™ Red conjugated nanowires are non-fluorescent outside the cells but fluoresce brightly inside, internalized nanowires are distinguished from non-internalized ones and their behavior inside the cells can be tracked for the respective time length. A machine learning-based computational framework dedicated to automatic analysis of live cell imaging data, Cell Cognition, is adapted and used to classify cells with internalized and non-internalized nanowires and subsequently determine the uptake percentage by cells at different time points. An uptake of 85 % by HCT 116 cells is observed after 24 hours incubation at NW-to-cell ratios of 200. While the approach of using pHrodo™ Red for internalization studies is not novel in the literature, this study reports for the first time the utilization of a machine-learning based time-resolved automatic analysis pipeline for quantification of nanowire uptake by cells. This pipeline has also been used for comparison studies with nickel nanowires coated with APTES and labeled with pHrodo™ Red, and another cell line derived from the cervix carcinoma, HeLa. It has thus the potential to be used for studying the interaction of different types of nanostructures with potentially any live cell types.

  3. Extended quantification of the generalized recurrence plot

    Science.gov (United States)

    Riedl, Maik; Marwan, Norbert; Kurths, Jürgen

    2016-04-01

    The generalized recurrence plot is a modern tool for quantification of complex spatial patterns. Its application spans the analysis of trabecular bone structures, Turing structures, turbulent spatial plankton patterns, and fractals. But, it is also successfully applied to the description of spatio-temporal dynamics and the detection of regime shifts, such as in the complex Ginzburg-Landau- equation. The recurrence plot based determinism is a central measure in this framework quantifying the level of regularities in temporal and spatial structures. We extend this measure for the generalized recurrence plot considering additional operations of symmetry than the simple translation. It is tested not only on two-dimensional regular patterns and noise but also on complex spatial patterns reconstructing the parameter space of the complex Ginzburg-Landau-equation. The extended version of the determinism resulted in values which are consistent to the original recurrence plot approach. Furthermore, the proposed method allows a split of the determinism into parts which based on laminar and non-laminar regions of the two-dimensional pattern of the complex Ginzburg-Landau-equation. A comparison of these parts with a standard method of image classification, the co-occurrence matrix approach, shows differences especially in the description of patterns associated with turbulence. In that case, it seems that the extended version of the determinism allows a distinction of phase turbulence and defect turbulence by means of their spatial patterns. This ability of the proposed method promise new insights in other systems with turbulent dynamics coming from climatology, biology, ecology, and social sciences, for example.

  4. Quantification of asphaltene precipitation by scaling equation

    Science.gov (United States)

    Janier, Josefina Barnachea; Jalil, Mohamad Afzal B. Abd.; Samin, Mohamad Izhar B. Mohd; Karim, Samsul Ariffin B. A.

    2015-02-01

    Asphaltene precipitation from crude oil is one of the issues for the oil industry. The deposition of asphaltene occurs during production, transportation and separating process. The injection of carbon dioxide (CO2) during enhance oil recovery (EOR) is believed to contribute much to the precipitation of asphaltene. Precipitation can be affected by the changes in temperature and pressure on the crude oil however, reduction in pressure contribute much to the instability of asphaltene as compared to temperature. This paper discussed the quantification of precipitated asphaltene in crude oil at different high pressures and at constant temperature. The derived scaling equation was based on the reservoir condition with variation in the amount of carbon dioxide (CO2) mixed with Dulang a light crude oil sample used in the experiment towards the stability of asphaltene. A FluidEval PVT cell with Solid Detection System (SDS) was the instrument used to gain experimental knowledge on the behavior of fluid at reservoir conditions. Two conditions were followed in the conduct of the experiment. Firstly, a 45cc light crude oil was mixed with 18cc (40%) of CO2 and secondly, the same amount of crude oil sample was mixed with 27cc (60%) of CO2. Results showed that for a 45cc crude oil sample combined with 18cc (40%) of CO2 gas indicated a saturation pressure of 1498.37psi and asphaltene onset point was 1620psi. Then for the same amount of crude oil combined with 27cc (60%) of CO2, the saturation pressure was 2046.502psi and asphaltene onset point was 2230psi. The derivation of the scaling equation considered reservoir temperature, pressure, bubble point pressure, mole percent of the precipitant the injected gas CO2, and the gas molecular weight. The scaled equation resulted to a third order polynomial that can be used to quantify the amount of asphaltene in crude oil.

  5. Rapid digital quantification of microfracture populations

    Science.gov (United States)

    Gomez, Leonel A.; Laubach, Stephen E.

    2006-03-01

    Populations of microfractures are a structural fabric in many rocks deformed at upper crustal conditions. In some cases these fractures are visible in transmitted-light microscopy as fluid-inclusion planes or cement filled microfractures, but because SEM-based cathodoluminescence (CL) reveals more fractures and delineates their shapes, sizes, and crosscutting relations, it is a more effective structural tool. Yet at magnifications of 150-300×, at which many microfractures are visible, SEM-CL detectors image only small sample areas (0.5-0.1 mm 2) relative to fracture population patterns. The substantial effort required to image and measure centimeter-size areas at high-magnification has impeded quantitative study of microfractures. We present a method for efficient collection of mosaics of high-resolution CL imagery, a preparation method that allows samples to be any size while retaining continuous imagery of rock (no gaps), and software that facilitates fracture mapping and data reduction. Although the method introduced here was developed for CL imagery, it can be used with any other kind of images, including mosaics from petrographic microscopes. Compared with manual measurements, the new method increases several fold the number of microfractures imaged without a proportional increase in level of effort, increases the accuracy and repeatability of fracture measurements, and speeds quantification and display of fracture population attributes. We illustrate the method on microfracture arrays in dolostone from NE Mexico and sandstone from NW Scotland. We show that key aspects of microfracture population attributes are only fully manifest at scales larger than a single thin section.

  6. Methodological considerations in quantification of oncological FDG PET studies

    Energy Technology Data Exchange (ETDEWEB)

    Vriens, Dennis; Visser, Eric P.; Geus-Oei, Lioe-Fee de; Oyen, Wim J.G. [Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine, Nijmegen (Netherlands)

    2010-07-15

    This review aims to provide insight into the factors that influence quantification of glucose metabolism by FDG PET images in oncology as well as their influence on repeated measures studies (i.e. treatment response assessment), offering improved understanding both for clinical practice and research. Structural PubMed searches have been performed for the many factors affecting quantification of glucose metabolism by FDG PET. Review articles and references lists have been used to supplement the search findings. Biological factors such as fasting blood glucose level, FDG uptake period, FDG distribution and clearance, patient motion (breathing) and patient discomfort (stress) all influence quantification. Acquisition parameters should be adjusted to maximize the signal to noise ratio without exposing the patient to a higher than strictly necessary radiation dose. This is especially challenging in pharmacokinetic analysis, where the temporal resolution is of significant importance. The literature is reviewed on the influence of attenuation correction on parameters for glucose metabolism, the effect of motion, metal artefacts and contrast agents on quantification of CT attenuation-corrected images. Reconstruction settings (analytical versus iterative reconstruction, post-reconstruction filtering and image matrix size) all potentially influence quantification due to artefacts, noise levels and lesion size dependency. Many region of interest definitions are available, but increased complexity does not necessarily result in improved performance. Different methods for the quantification of the tissue of interest can introduce systematic and random inaccuracy. This review provides an up-to-date overview of the many factors that influence quantification of glucose metabolism by FDG PET. (orig.) 3.

  7. Automated lobar quantification of emphysema in patients with severe COPD

    International Nuclear Information System (INIS)

    Automated lobar quantification of emphysema has not yet been evaluated. Unenhanced 64-slice MDCT was performed in 47 patients evaluated before bronchoscopic lung-volume reduction. CT images reconstructed with a standard (B20) and high-frequency (B50) kernel were analyzed using a dedicated prototype software (MevisPULMO) allowing lobar quantification of emphysema extent. Lobar quantification was obtained following (a) a fully automatic delineation of the lobar limits by the software and (b) a semiautomatic delineation with manual correction of the lobar limits when necessary and was compared with the visual scoring of emphysema severity per lobe. No statistically significant difference existed between automated and semiautomated lobar quantification (p>0.05 in the five lobes), with differences ranging from 0.4 to 3.9%. The agreement between the two methods (intraclass correlation coefficient, ICC) was excellent for left upper lobe (ICC=0.94), left lower lobe (ICC=0.98), and right lower lobe (ICC=0.80). The agreement was good for right upper lobe (ICC=0.68) and moderate for middle lobe (IC=0.53). The Bland and Altman plots confirmed these results. A good agreement was observed between the software and visually assessed lobar predominance of emphysema (kappa 0.78; 95% CI 0.64-0.92). Automated and semiautomated lobar quantifications of emphysema are concordant and show good agreement with visual scoring. (orig.)

  8. GMO quantification: valuable experience and insights for the future.

    Science.gov (United States)

    Milavec, Mojca; Dobnik, David; Yang, Litao; Zhang, Dabing; Gruden, Kristina; Zel, Jana

    2014-10-01

    Cultivation and marketing of genetically modified organisms (GMOs) have been unevenly adopted worldwide. To facilitate international trade and to provide information to consumers, labelling requirements have been set up in many countries. Quantitative real-time polymerase chain reaction (qPCR) is currently the method of choice for detection, identification and quantification of GMOs. This has been critically assessed and the requirements for the method performance have been set. Nevertheless, there are challenges that should still be highlighted, such as measuring the quantity and quality of DNA, and determining the qPCR efficiency, possible sequence mismatches, characteristics of taxon-specific genes and appropriate units of measurement, as these remain potential sources of measurement uncertainty. To overcome these problems and to cope with the continuous increase in the number and variety of GMOs, new approaches are needed. Statistical strategies of quantification have already been proposed and expanded with the development of digital PCR. The first attempts have been made to use new generation sequencing also for quantitative purposes, although accurate quantification of the contents of GMOs using this technology is still a challenge for the future, and especially for mixed samples. New approaches are needed also for the quantification of stacks, and for potential quantification of organisms produced by new plant breeding techniques.

  9. GMO quantification: valuable experience and insights for the future.

    Science.gov (United States)

    Milavec, Mojca; Dobnik, David; Yang, Litao; Zhang, Dabing; Gruden, Kristina; Zel, Jana

    2014-10-01

    Cultivation and marketing of genetically modified organisms (GMOs) have been unevenly adopted worldwide. To facilitate international trade and to provide information to consumers, labelling requirements have been set up in many countries. Quantitative real-time polymerase chain reaction (qPCR) is currently the method of choice for detection, identification and quantification of GMOs. This has been critically assessed and the requirements for the method performance have been set. Nevertheless, there are challenges that should still be highlighted, such as measuring the quantity and quality of DNA, and determining the qPCR efficiency, possible sequence mismatches, characteristics of taxon-specific genes and appropriate units of measurement, as these remain potential sources of measurement uncertainty. To overcome these problems and to cope with the continuous increase in the number and variety of GMOs, new approaches are needed. Statistical strategies of quantification have already been proposed and expanded with the development of digital PCR. The first attempts have been made to use new generation sequencing also for quantitative purposes, although accurate quantification of the contents of GMOs using this technology is still a challenge for the future, and especially for mixed samples. New approaches are needed also for the quantification of stacks, and for potential quantification of organisms produced by new plant breeding techniques. PMID:25182968

  10. Uncertainty Quantification in Climate Modeling and Projection

    Energy Technology Data Exchange (ETDEWEB)

    Qian, Yun; Jackson, Charles S.; Giorgi, Filippo; Booth, Ben; Duan, Qingyun; Forest , Chris; Higdon, Dave; Hou, Zhangshuan; Huerta, Gabriel

    2016-05-01

    The projection of future climate is one of the most complex problems undertaken by the scientific community. Although scientists have been striving to better understand the physical basis of the climate system and to improve climate models, the overall uncertainty in projections of future climate has not been significantly reduced (e.g., from the IPCC AR4 to AR5). With the rapid increase of complexity in Earth system models, reducing uncertainties in climate projections becomes extremely challenging. Since uncertainties always exist in climate models, interpreting the strengths and limitations of future climate projections is key to evaluating risks, and climate change information for use in Vulnerability, Impact, and Adaptation (VIA) studies should be provided with both well-characterized and well-quantified uncertainty. The workshop aimed at providing participants, many of them from developing countries, information on strategies to quantify the uncertainty in climate model projections and assess the reliability of climate change information for decision-making. The program included a mixture of lectures on fundamental concepts in Bayesian inference and sampling, applications, and hands-on computer laboratory exercises employing software packages for Bayesian inference, Markov Chain Monte Carlo methods, and global sensitivity analyses. The lectures covered a range of scientific issues underlying the evaluation of uncertainties in climate projections, such as the effects of uncertain initial and boundary conditions, uncertain physics, and limitations of observational records. Progress in quantitatively estimating uncertainties in hydrologic, land surface, and atmospheric models at both regional and global scales was also reviewed. The application of Uncertainty Quantification (UQ) concepts to coupled climate system models is still in its infancy. The Coupled Model Intercomparison Project (CMIP) multi-model ensemble currently represents the primary data for

  11. Quantification of carbon nanomaterials in vivo.

    Science.gov (United States)

    Wang, Haifang; Yang, Sheng-Tao; Cao, Aoneng; Liu, Yuanfang

    2013-03-19

    this Account, we review the in vivo quantification methods of carbon NMs, focusing on isotopic labeling and tracing methods, and summarize the related labeling, purification, bio-sampling, and detection of carbon NMs. We also address the advantages, applicable situations, and limits of various labeling and tracing methods and propose guidelines for choosing suitable labeling methods. A collective analysis of the ADME information on various carbon NMs in vivo would provide general principles for understanding the fate of carbon NMs and the effects of chemical functionalization and aggregation of carbon NMs on their ADME/T in vivo and their implications in nanotoxicology and biosafety evaluations.

  12. Extended Forward Sensitivity Analysis for Uncertainty Quantification

    Energy Technology Data Exchange (ETDEWEB)

    Haihua Zhao; Vincent A. Mousseau

    2011-09-01

    Verification and validation (V&V) are playing more important roles to quantify uncertainties and realize high fidelity simulations in engineering system analyses, such as transients happened in a complex nuclear reactor system. Traditional V&V in the reactor system analysis focused more on the validation part or did not differentiate verification and validation. The traditional approach to uncertainty quantification is based on a 'black box' approach. The simulation tool is treated as an unknown signal generator, a distribution of inputs according to assumed probability density functions is sent in and the distribution of the outputs is measured and correlated back to the original input distribution. The 'black box' method mixes numerical errors with all other uncertainties. It is also not efficient to perform sensitivity analysis. Contrary to the 'black box' method, a more efficient sensitivity approach can take advantage of intimate knowledge of the simulation code. In these types of approaches equations for the propagation of uncertainty are constructed and the sensitivities are directly solved for as variables in the simulation. This paper presents the forward sensitivity analysis as a method to help uncertainty qualification. By including time step and potentially spatial step as special sensitivity parameters, the forward sensitivity method is extended as one method to quantify numerical errors. Note that by integrating local truncation errors over the whole system through the forward sensitivity analysis process, the generated time step and spatial step sensitivity information reflect global numerical errors. The discretization errors can be systematically compared against uncertainties due to other physical parameters. This extension makes the forward sensitivity method a much more powerful tool to help uncertainty qualification. By knowing the relative sensitivity of time and space steps with other interested physical

  13. Methane bubbling: from speculation to quantification

    Science.gov (United States)

    Grinham, A. R.; Dunbabin, M.; Yuan, Z.

    2013-12-01

    magnitude from 500 to 100 000 mg m-2 d-1 depending on time of day and water depth. Average storage bubble flux rates between reservoirs varied by two orders of magnitude from 1 200 to 15 000 mg m-2 d-1, with the primary driver likely to be catchment forest cover. The relative contribution of bubbling to total fluxes varied from 10% to more than 90% depending on the reservoir and time of sampling. This method was consistently shown to greatly improve the spatial mapping and quantification of methane bubbling rates from reservoir surfaces and reduces the uncertainty associated with the determining the relative contribution of bubbling to total flux.

  14. Superlattice band structure: New and simple energy quantification condition

    International Nuclear Information System (INIS)

    Assuming an approximated effective mass and using Bastard's boundary conditions, a simple method is used to calculate the subband structure for periodic semiconducting heterostructures. Our method consists to derive and solve the energy quantification condition (EQC), this is a simple real equation, composed of trigonometric and hyperbolic functions, and does not need any programming effort or sophistic machine to solve it. For less than ten wells heterostructures, we have derived and simplified the energy quantification conditions. The subband is build point by point; each point presents an energy level. Our simple energy quantification condition is used to calculate the subband structure of the GaAs/Ga0.5Al0.5As heterostructures, and build its subband point by point for 4 and 20 wells. Our finding shows a good agreement with previously published results

  15. Iron overload in the liver diagnostic and quantification

    Energy Technology Data Exchange (ETDEWEB)

    Alustiza, Jose M. [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain)]. E-mail: jmalustiza@osatek.es; Castiella, Agustin [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain); Juan, Maria D. de [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain); Emparanza, Jose I. [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain); Artetxe, Jose [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain); Uranga, Maite [Osatek SA, P Dr. Beguiristain 109, 20014, San Sebastian, Guipuzcoa (Spain)

    2007-03-15

    Hereditary Hemochromatosis is the most frequent modality of iron overload. Since 1996 genetic tests have facilitated significantly the non-invasive diagnosis of the disease. There are however many cases of negative genetic tests that require confirmation by hepatic iron quantification which is traditionally performed by hepatic biopsy. There are many studies that have demonstrated the possibility of performing hepatic iron quantification with Magnetic Resonance. However, a consensus has not been reached yet regarding the technique or the possibility to reproduce the same method of calculus in different machines. This article reviews the state of the art of the question and delineates possible future lines to standardise this non-invasive method of hepatic iron quantification.

  16. Stereological quantification of mast cells in human synovium

    DEFF Research Database (Denmark)

    Damsgaard, T E; Sørensen, Flemming Brandt; Herlin, T;

    1999-01-01

    synovium, using stereological techniques. Different methods of staining and quantification have previously been used for mast cell quantification in human synovium. Stereological techniques provide precise and unbiased information on the number of cell profiles in two-dimensional tissue sections of, in......Mast cells participate in both the acute allergic reaction as well as in chronic inflammatory diseases. Earlier studies have revealed divergent results regarding the quantification of mast cells in the human synovium. The aim of the present study was therefore to quantify these cells in the human...... this case, human synovium. In 10 patients suffering from osteoarthritis a median of 3.6 mast cells/mm2 synovial membrane was found. The total number of cells (synoviocytes, fibroblasts, lymphocytes, leukocytes) present was 395.9 cells/mm2 (median). The mast cells constituted 0.8% of all the cell...

  17. Superlattice band structure: New and simple energy quantification condition

    Energy Technology Data Exchange (ETDEWEB)

    Maiz, F., E-mail: fethimaiz@gmail.com [University of Cartage, Nabeul Engineering Preparatory Institute, Merazka, 8000 Nabeul (Tunisia); King Khalid University, Faculty of Science, Physics Department, P.O. Box 9004, Abha 61413 (Saudi Arabia)

    2014-10-01

    Assuming an approximated effective mass and using Bastard's boundary conditions, a simple method is used to calculate the subband structure for periodic semiconducting heterostructures. Our method consists to derive and solve the energy quantification condition (EQC), this is a simple real equation, composed of trigonometric and hyperbolic functions, and does not need any programming effort or sophistic machine to solve it. For less than ten wells heterostructures, we have derived and simplified the energy quantification conditions. The subband is build point by point; each point presents an energy level. Our simple energy quantification condition is used to calculate the subband structure of the GaAs/Ga{sub 0.5}Al{sub 0.5}As heterostructures, and build its subband point by point for 4 and 20 wells. Our finding shows a good agreement with previously published results.

  18. Large-Scale Inverse Problems and Quantification of Uncertainty

    CERN Document Server

    Biegler, Lorenz; Ghattas, Omar

    2010-01-01

    Large-scale inverse problems and associated uncertainty quantification has become an important area of research, central to a wide range of science and engineering applications. Written by leading experts in the field, Large-scale Inverse Problems and Quantification of Uncertainty focuses on the computational methods used to analyze and simulate inverse problems. The text provides PhD students, researchers, advanced undergraduate students, and engineering practitioners with the perspectives of researchers in areas of inverse problems and data assimilation, ranging from statistics and large-sca

  19. Brief review of uncertainty quantification for particle image velocimetry

    Science.gov (United States)

    Farias, M. H.; Teixeira, R. S.; Koiller, J.; Santos, A. M.

    2016-07-01

    Metrological studies for particle image velocimetry (PIV) are recent in literature. An attempt to evaluate the uncertainty quantifications (UQ) of the PIV velocity field are in evidence. Therefore, a short review on main sources of uncertainty in PIV and available methodologies for its quantification are presented. In addition, the potential of some mathematical techniques, coming from the area of geometric mechanics and control, that could interest the fluids UQ community are highlighted as good possibilities. “We must measure what is measurable and make measurable what cannot be measured” (Galileo)

  20. A quick colorimetric method for total lipid quantification in microalgae.

    Science.gov (United States)

    Byreddy, Avinesh R; Gupta, Adarsha; Barrow, Colin J; Puri, Munish

    2016-06-01

    Discovering microalgae with high lipid productivity are among the key milestones for achieving sustainable biodiesel production. Current methods of lipid quantification are time intensive and costly. A rapid colorimetric method based on sulfo-phospho-vanillin (SPV) reaction was developed for the quantification of microbial lipids to facilitate screening for lipid producing microalgae. This method was successfully tested on marine thraustochytrid strains and vegetable oils. The colorimetric method results correlated well with gravimetric method estimates. The new method was less time consuming than gravimetric analysis and is quantitative for lipid determination, even in the presence of carbohydrates, proteins and glycerol. PMID:27050419

  1. Remote sensing for quantification of agronomic properties

    Science.gov (United States)

    Sullivan, Dana Grace

    Remote sensing (RS) may be used to rapidly assess surface features and facilitate natural resource management, precision agriculture and soil survey. Information obtained in such a way would streamline data collection and improve diagnostic capabilities. Current RS technology has had limited testing, particularly within the Southeast. Our study was designed to evaluate RS as a rapid assessment tool in three different natural resource applications: nitrogen (N) management in a corn crop (Zea mays L.), assessment of in situ crop residue cover, and quantification of near-surface soil properties. In 2000, study sites were established in four physiographic provinces of Alabama: Tennessee Valley, Ridge and Valley, Appalachian Plateau, and Coastal Plain. Spectral measurements were acquired via spectroradiometer (350--1050 nm), airborne ATLAS multispectral scanner (400--12,500 nm), and IKONOS satellite (450--900 nm). Corn plots were established from fresh-tilled ground in a completely randomized design at the Appalachian Plateau and Coastal Plain study sites in 2000. Plots received four N rates (0, 56, 112, and 168 kg N ha-1 ), and were maintained for three consecutive growing seasons. Spectroradiometer data were acquired biweekly from V6-R2 and ATLAS and IKONOS were acquired per availability. Results showed vegetation indices derived from hand-held spectroradiometer measurements as early as V6-V8 were linearly related to yield and tissue N. ATLAS imagery showed promise at the AP site during the V6 stage (r2 = 0.66), but no significant relationships between plant N and IKONOS imagery were observed. Residue plots (15m x 15m) were established at the Appalachian Plateau and Coastal Plain in 2000 and 200. Residue treatments consisted of hand applied wheat straw cover (0, 10 20, 50, or 80%) arranged in a completely randomized design. Spectroradiometer data were acquired monthly and ATLAS and IKONOS were acquired per availability. Residue cover estimates were best with ATLAS

  2. A monomial chaos approach for efficient uncertainty quantification on nonlinear problems

    NARCIS (Netherlands)

    Witteveen, J.A.S.; Bijl, H.

    2008-01-01

    A monomial chaos approach is presented for efficient uncertainty quantification in nonlinear computational problems. Propagating uncertainty through nonlinear equations can be computationally intensive for existing uncertainty quantification methods. It usually results in a set of nonlinear equation

  3. A Monomial Chaos Approach for Efficient Uncertainty Quantification in Computational Fluid Dynamics

    NARCIS (Netherlands)

    Witteveen, J.A.S.; Bijl, H.

    2006-01-01

    A monomial chaos approach is proposed for efficient uncertainty quantification in nonlinear computational problems. Propagating uncertainty through nonlinear equations can still be computationally intensive for existing uncertainty quantification methods. It usually results in a set of nonlinear equ

  4. Rapid Quantification of Myocardial Fibrosis: A New Macro-Based Automated Analysis

    Directory of Open Access Journals (Sweden)

    Awal M. Hadi

    2010-01-01

    Full Text Available Background: Fibrosis is associated with various cardiac pathologies and dysfunction. Current quantification methods are time-consuming and laborious. We describe a semi-automated quantification technique for myocardial fibrosis and validated this using traditional methods.

  5. Rapid Quantification and Validation of Lipid Concentrations within Liposomes.

    Science.gov (United States)

    Roces, Carla B; Kastner, Elisabeth; Stone, Peter; Lowry, Deborah; Perrie, Yvonne

    2016-01-01

    Quantification of the lipid content in liposomal adjuvants for subunit vaccine formulation is of extreme importance, since this concentration impacts both efficacy and stability. In this paper, we outline a high performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD) method that allows for the rapid and simultaneous quantification of lipid concentrations within liposomal systems prepared by three liposomal manufacturing techniques (lipid film hydration, high shear mixing, and microfluidics). The ELSD system was used to quantify four lipids: 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), cholesterol, dimethyldioctadecylammonium (DDA) bromide, and ᴅ-(+)-trehalose 6,6'-dibehenate (TDB). The developed method offers rapidity, high sensitivity, direct linearity, and a good consistency on the responses (R² > 0.993 for the four lipids tested). The corresponding limit of detection (LOD) and limit of quantification (LOQ) were 0.11 and 0.36 mg/mL (DMPC), 0.02 and 0.80 mg/mL (cholesterol), 0.06 and 0.20 mg/mL (DDA), and 0.05 and 0.16 mg/mL (TDB), respectively. HPLC-ELSD was shown to be a rapid and effective method for the quantification of lipids within liposome formulations without the need for lipid extraction processes. PMID:27649231

  6. Striga hermonthica seed bank dynamics: process quantification and modelling

    NARCIS (Netherlands)

    Mourik, van T.A.

    2007-01-01

    Key words:    Weed control, integrated management, parasitic weed, population, sorghum, millet.   This thesis presents a study on the quantification of seed bank dynamics of the parasitic weed Striga hermonthica. The main objectives were to quantify transition rates between different stages of the l

  7. Methane emission quantification from landfills using a double tracer approach

    DEFF Research Database (Denmark)

    Scheutz, Charlotte; Samuelsson, J.; Fredenslund, Anders Michael;

    2007-01-01

    A tracer method was successfully used for quantification of the whole methane (CH4) emission from Fakse landfill. By using two different tracers the emission from different sections of the landfill could be quantified. Furthermore, is was possible to determine the emissions from local on site...

  8. Automatic quantification of subarachnoid hemorrhage on noncontrast CT

    NARCIS (Netherlands)

    Boers, A.M.; Zijlstra, I.A.; Gathier, C.S.; Berg, van den R.; Slump, C.H.; Marquering, H.A.; Majoie, C.B.

    2014-01-01

    Quantification of blood after SAH on initial NCCT is an important radiologic measure to predict patient outcome and guide treatment decisions. In current scales, hemorrhage volume and density are not accounted for. The purpose of this study was to develop and validate a fully automatic method for SA

  9. Temperature dependence of postmortem MR quantification for soft tissue discrimination

    Energy Technology Data Exchange (ETDEWEB)

    Zech, Wolf-Dieter; Schwendener, Nicole; Jackowski, Christian [University of Bern, From the Institute of Forensic Medicine, Bern (Switzerland); Persson, Anders; Warntjes, Marcel J. [University of Linkoeping, The Center for Medical Image Science and Visualization (CMIV), Linkoeping (Sweden)

    2015-08-15

    To investigate and correct the temperature dependence of postmortem MR quantification used for soft tissue characterization and differentiation in thoraco-abdominal organs. Thirty-five postmortem short axis cardiac 3-T MR examinations were quantified using a quantification sequence. Liver, spleen, left ventricular myocardium, pectoralis muscle and subcutaneous fat were analysed in cardiac short axis images to obtain mean T1, T2 and PD tissue values. The core body temperature was measured using a rectally inserted thermometer. The tissue-specific quantitative values were related to the body core temperature. Equations to correct for temperature differences were generated. In a 3D plot comprising the combined data of T1, T2 and PD, different organs/tissues could be well differentiated from each other. The quantitative values were influenced by the temperature. T1 in particular exhibited strong temperature dependence. The correction of quantitative values to a temperature of 37 C resulted in better tissue discrimination. Postmortem MR quantification is feasible for soft tissue discrimination and characterization of thoraco-abdominal organs. This provides a base for computer-aided diagnosis and detection of tissue lesions. The temperature dependence of the T1 values challenges postmortem MR quantification. Equations to correct for the temperature dependence are provided. (orig.)

  10. Quantification of the auditory startle reflex in children

    NARCIS (Netherlands)

    Bakker, Mirte J.; Boer, Frits; van der Meer, Johan N.; Koelman, Johannes H. T. M.; Boeree, Thijs; Bour, Lo; Tijssen, Marina A. J.

    2009-01-01

    Objective: To find an adequate tool to assess the auditory startle reflex (ASR) in children. Methods: We investigated the effect of stimulus repetition, gender and age on several quantifications of the ASR. ASR's were elicited by eight consecutive auditory stimuli in 27 healthy children. Electromyog

  11. 15 CFR 990.52 - Injury assessment-quantification.

    Science.gov (United States)

    2010-01-01

    ... 15 Commerce and Foreign Trade 3 2010-01-01 2010-01-01 false Injury assessment-quantification. 990... REGULATIONS NATURAL RESOURCE DAMAGE ASSESSMENTS Restoration Planning Phase § 990.52 Injury assessment... extent of injury to a natural resource, with subsequent translation of that adverse change to a...

  12. Real-Time PCR for Universal Phytoplasma Detection and Quantification

    DEFF Research Database (Denmark)

    Christensen, Nynne Meyn; Nyskjold, Henriette; Nicolaisen, Mogens

    2013-01-01

    Currently, the most efficient detection and precise quantification of phytoplasmas is by real-time PCR. Compared to nested PCR, this method is less sensitive to contamination and is less work intensive. Therefore, a universal real-time PCR method will be valuable in screening programs and in other...

  13. Improved perfusion quantification in FAIR imaging by offset correction

    DEFF Research Database (Denmark)

    Sidaros, Karam; Andersen, Irene K.; Gesmar, Henrik;

    2001-01-01

    Perfusion quantification using pulsed arterial spin labeling has been shown to be sensitive to the RF pulse slice profiles. Therefore, in Flow-sensitive Alternating-Inversion Recovery (FAIR) imaging the slice selective (ss) inversion slab is usually three to four times thicker than the imaging...

  14. Hepatocellular carcinoma: perfusion quantification with dynamic contrast-enhanced MRI

    NARCIS (Netherlands)

    Taouli, B.; Johnson, R.S.; Hajdu, C.H.; Oei, M.T.H.; Merad, M.; Yee, H.; Rusinek, H.

    2013-01-01

    The objective of our study was to report our initial experience with dynamic contrast-enhanced MRI (DCE-MRI) for perfusion quantification of hepatocellular carcinoma (HCC) and surrounding liver.DCE-MRI of the liver was prospectively performed on 31 patients with HCC (male-female ratio, 26:5; mean ag

  15. Macroscopic inspection of ape feces: what's in a quantification method?

    Science.gov (United States)

    Phillips, Caroline A; McGrew, William C

    2014-06-01

    Macroscopic inspection of feces has been used to investigate primate diet. The limitations of this method to identify food-items to species level have long been recognized, but ascertaining aspects of diet (e.g., folivory) are achievable by quantifying food-items in feces. Quantification methods applied include rating food-items using a scale of abundance, estimating their percentage volume, and weighing food-items. However, verification as to whether or not composition data differ, depending on which quantification method is used during macroscopic inspection, has not been done. We analyzed feces collected from ten adult chimpanzees (Pan troglodytes schweinfurthii) of the Kanyawara community in Kibale National Park, Uganda. We compare dietary composition totals obtained from using different quantification methods and ascertain if sieve mesh size influences totals calculated. Finally, this study validates findings from direct observation of feeding by the same individuals from whom the fecal samples had been collected. Contrasting diet composition totals obtained by using different quantification methods and sieve mesh sizes can influence folivory and frugivory estimates. However, our findings were based on the assumption that fibrous matter contained pith and leaf fragments only, which remains to be verified. We advocate macroscopic inspection of feces can be a valuable tool to provide a generalized overview of dietary composition for primate populations. As most populations remain unhabituated, scrutinizing and validating indirect measures are important if they are to be applied to further understand inter- and intra-species dietary variation. PMID:24482001

  16. The Role of Uncertainty Quantification for Reactor Physics

    Energy Technology Data Exchange (ETDEWEB)

    Salvatores, Massimo [Idaho National Lab. (INL), Idaho Falls, ID (United States); Palmiotti, Giuseppe [Idaho National Lab. (INL), Idaho Falls, ID (United States); Aliberti, G. [Argonne National Lab. (ANL), Argonne, IL (United States)

    2015-01-01

    The quantification of uncertainties is a crucial step in design. The comparison of a-priori uncertainties with the target accuracies, allows to define needs and priorities for uncertainty reduction. In view of their impact, the uncertainty analysis requires a reliability assessment of the uncertainty data used. The choice of the appropriate approach and the consistency of different approaches are discussed.

  17. Optimal purification and sensitive quantification of DNA from fecal samples

    DEFF Research Database (Denmark)

    Jensen, Annette Nygaard; Hoorfar, Jeffrey

    2002-01-01

    , the detection range of X-DNA of a spectrophotometric and a fluorometric (PicoGreen) method was compared. The PicoGreen showed a quantification limit of 1 ng/mL, consistent triplicate measurements, and finally a linear relationship between the concentrations of DNA standards and the fluorescence readings (R-2...

  18. Improved perfusion quantification in FAIR imaging by offset correction

    DEFF Research Database (Denmark)

    Sidaros, K; Andersen, I K; Gesmar, H;

    2001-01-01

    Perfusion quantification using pulsed arterial spin labeling has been shown to be sensitive to the RF pulse slice profiles. Therefore, in Flow-sensitive Alternating-Inversion Recovery (FAIR) imaging the slice selective (ss) inversion slab is usually three to four times thicker than the imaging sl...

  19. Quantification of Photoperiodic Effects on Growth of Phleum pratense

    OpenAIRE

    WU, ZUOLI; SKJELVÅG A.o.; BAADSHAUG, O. H.

    2004-01-01

    • Background and Aims Accurate quantifications of plant responses to photoperiod are useful for physiological studies, in growth modelling and in other studies of environmental effects. The objective of the current work was a mathematical description of photoperiodic influence on plant morphological traits, using functions with few and common parameters related to key plant characteristics and typical response patterns.

  20. Unilateral condylar hyperplasia: a 3-dimensional quantification of asymmetry

    NARCIS (Netherlands)

    T.J. Verhoeven; J.W. Nolte; T.J.J. Maal; S.J. Bergé; A.G. Becking

    2013-01-01

    Purpose Objective quantifications of facial asymmetry in patients with Unilateral Condylar Hyperplasia (UCH) have not yet been described in literature. The aim of this study was to objectively quantify soft-tissue asymmetry in patients with UCH and to compare the findings with a control group using

  1. Unilateral condylar hyperplasia: a 3-dimensional quantification of asymmetry

    NARCIS (Netherlands)

    Verhoeven, T.J.; Nolte, J.W.; Maal, T.J.J.; Berge, S.J.; Becking, A.G.

    2013-01-01

    PURPOSE: Objective quantifications of facial asymmetry in patients with Unilateral Condylar Hyperplasia (UCH) have not yet been described in literature. The aim of this study was to objectively quantify soft-tissue asymmetry in patients with UCH and to compare the findings with a control group using

  2. Identification and Quantification Soil Redoximorphic Features by Digital Image Processing

    Science.gov (United States)

    Soil redoximorphic features (SRFs) have provided scientists and land managers with insight into relative soil moisture for approximately 60 years. The overall objective of this study was to develop a new method of SRF identification and quantification from soil cores using a digital camera and imag...

  3. Current Issues in the Quantification of Federal Reserved Water Rights

    Science.gov (United States)

    Brookshire, David S.; Watts, Gary L.; Merrill, James L.

    1985-11-01

    This paper examines the quantification of federal reserved water rights from legal, institutional, and economic perspectives. Special attention is directed toward Indian reserved water rights and the concept of practicably irrigable acreage. We conclude by examining current trends and exploring alternative approaches to the dilemma of quantifying Indian reserved water rights.

  4. Damage Localization and Quantification of Earthquake Excited RC-Frames

    DEFF Research Database (Denmark)

    Skjærbæk, P.S.; Nielsen, Søren R.K.; Kirkegaard, Poul Henning;

    In the paper a recently proposed method for damage localization and quantification of RC-structures from response measurements is tested on experimental data. The method investigated requires at least one response measurement along the structure and the ground surface acceleration. Further, the t...

  5. StudyonMathematicalModeofQuantification Performance-to-Price Ratio

    Institute of Scientific and Technical Information of China (English)

    裴兰华; 史德战; 王冠

    2007-01-01

    Nowadays, consumers often compare the same kind of commodities and decide what to pick out when they purchase merchandise including the service. The paper discusses the mathematical mode of quantification performance-to-price ratio according to which product can be made in order to increase the competitiveness in the market.

  6. Quantification and Negation in Event Semantics

    Directory of Open Access Journals (Sweden)

    Lucas Champollion

    2010-12-01

    Full Text Available Recently, it has been claimed that event semantics does not go well together with quantification, especially if one rejects syntactic, LF-based approaches to quantifier scope. This paper shows that such fears are unfounded, by presenting a simple, variable-free framework which combines a Neo-Davidsonian event semantics with a type-shifting based account of quantifier scope. The main innovation is that the event variable is bound inside the verbal denotation, rather than at sentence level by existential closure. Quantifiers can then be interpreted in situ. The resulting framework combines the strengths of event semantics and type-shifting accounts of quantifiers and thus does not force the semanticist to posit either a default underlying word order or a syntactic LF-style level. It is therefore well suited for applications to languages where word order is free and quantifier scope is determined by surface order. As an additional benefit, the system leads to a straightforward account of negation, which has also been claimed to be problematic for event-based frameworks.ReferencesBarker, Chris. 2002. ‘Continuations and the nature of quantification’. Natural Language Semantics 10: 211–242.http://dx.doi.org/10.1023/A:1022183511876Barker, Chris & Shan, Chung-chieh. 2008. ‘Donkey anaphora is in-scope binding’. Semantics and Pragmatics 1: 1–46.Beaver, David & Condoravdi, Cleo. 2007. ‘On the logic of verbal modification’. In Maria Aloni, Paul Dekker & Floris Roelofsen (eds. ‘Proceedings of the Sixteenth Amsterdam Colloquium’, 3–9. Amsterdam, Netherlands: University of Amsterdam.Beghelli, Filippo & Stowell, Tim. 1997. ‘Distributivity and negation: The syntax of each and every’. In Anna Szabolcsi (ed. ‘Ways of scope taking’, 71–107. Dordrecht, Netherlands: Kluwer.Brasoveanu, Adrian. 2010. ‘Modified Numerals as Post-Suppositions’. In Maria Aloni, Harald Bastiaanse, Tikitu de Jager & Katrin Schulz (eds. ‘Logic, Language

  7. The Maqāṣid approach and rethinking political rights in modern society

    Directory of Open Access Journals (Sweden)

    Louay Safi

    2010-12-01

    Full Text Available This paper examines political rights in Islam by focusing on freedom of religion and the extent to which the state is empowered to enforce faith and religious law on society. It starts by comparing the notion of law in both Western and Islamic traditions, and then analyzes the difference between the ethical and legal within Sharī‘ah. The paper illustrates how Islamic law grew historically by working to limit the power of the state, and points out the need to maintain the distinction between the state and civil society for the proper implementation of Sharī‘ah. The paper also contends that those who call on the state to enforce all rules of Sharī‘ah on society rely on a faulty theory of right and concludes that Islamic law fully recognizes the right of individuals to adopt and practice their faith freely. Freedom of religion, it stresses, is an intrinsic aspect of Islamic law and all efforts to limit this freedom is bound to violate its purpose and dictates.

  8. The Dark Side of the Mushroom Spring Microbial Mat: Life in the Shadow of Chlorophototrophs. I. Microbial Diversity Based on 16S rRNA Gene Amplicons and Metagenomic Sequencing.

    Science.gov (United States)

    Thiel, Vera; Wood, Jason M; Olsen, Millie T; Tank, Marcus; Klatt, Christian G; Ward, David M; Bryant, Donald A

    2016-01-01

    Microbial-mat communities in the effluent channels of Octopus and Mushroom Springs within the Lower Geyser Basin at Yellowstone National Park have been studied for nearly 50 years. The emphasis has mostly focused on the chlorophototrophic bacterial organisms of the phyla Cyanobacteria and Chloroflexi. In contrast, the diversity and metabolic functions of the heterotrophic community in the microoxic/anoxic region of the mat are not well understood. In this study we analyzed the orange-colored undermat of the microbial community of Mushroom Spring using metagenomic and rRNA-amplicon (iTag) analyses. Our analyses disclosed a highly diverse community exhibiting a high degree of unevenness, strongly dominated by a single taxon, the filamentous anoxygenic phototroph, Roseiflexus spp. The second most abundant organisms belonged to the Thermotogae, which have been hypothesized to be a major source of H2 from fermentation that could enable photomixotrophic metabolism by Chloroflexus and Roseiflexus spp. Other abundant organisms include two members of the Armatimonadetes (OP10); Thermocrinis sp.; and phototrophic and heterotrophic members of the Chloroflexi. Further, an Atribacteria (OP9/JS1) member; a sulfate-reducing Thermodesulfovibrio sp.; a Planctomycetes member; a member of the EM3 group tentatively affiliated with the Thermotogae, as well as a putative member of the Arminicenantes (OP8) represented ≥1% of the reads. Archaea were not abundant in the iTag analysis, and no metagenomic bin representing an archaeon was identified. A high microdiversity of 16S rRNA gene sequences was identified for the dominant taxon, Roseiflexus spp. Previous studies demonstrated that highly similar Synechococcus variants in the upper layer of the mats represent ecological species populations with specific ecological adaptations. This study suggests that similar putative ecotypes specifically adapted to different niches occur within the undermat community, particularly for Roseiflexus

  9. The Dark Side of the Mushroom Spring Microbial Mat: Life in the Shadow of Chlorophototrophs. I. Microbial Diversity Based on 16S rRNA Gene Amplicons and Metagenomic Sequencing

    Science.gov (United States)

    Thiel, Vera; Wood, Jason M.; Olsen, Millie T.; Tank, Marcus; Klatt, Christian G.; Ward, David M.; Bryant, Donald A.

    2016-01-01

    Microbial-mat communities in the effluent channels of Octopus and Mushroom Springs within the Lower Geyser Basin at Yellowstone National Park have been studied for nearly 50 years. The emphasis has mostly focused on the chlorophototrophic bacterial organisms of the phyla Cyanobacteria and Chloroflexi. In contrast, the diversity and metabolic functions of the heterotrophic community in the microoxic/anoxic region of the mat are not well understood. In this study we analyzed the orange-colored undermat of the microbial community of Mushroom Spring using metagenomic and rRNA-amplicon (iTag) analyses. Our analyses disclosed a highly diverse community exhibiting a high degree of unevenness, strongly dominated by a single taxon, the filamentous anoxygenic phototroph, Roseiflexus spp. The second most abundant organisms belonged to the Thermotogae, which have been hypothesized to be a major source of H2 from fermentation that could enable photomixotrophic metabolism by Chloroflexus and Roseiflexus spp. Other abundant organisms include two members of the Armatimonadetes (OP10); Thermocrinis sp.; and phototrophic and heterotrophic members of the Chloroflexi. Further, an Atribacteria (OP9/JS1) member; a sulfate-reducing Thermodesulfovibrio sp.; a Planctomycetes member; a member of the EM3 group tentatively affiliated with the Thermotogae, as well as a putative member of the Arminicenantes (OP8) represented ≥1% of the reads. Archaea were not abundant in the iTag analysis, and no metagenomic bin representing an archaeon was identified. A high microdiversity of 16S rRNA gene sequences was identified for the dominant taxon, Roseiflexus spp. Previous studies demonstrated that highly similar Synechococcus variants in the upper layer of the mats represent ecological species populations with specific ecological adaptations. This study suggests that similar putative ecotypes specifically adapted to different niches occur within the undermat community, particularly for Roseiflexus

  10. Quantification of Si in silicone oils by ICP-OES

    DEFF Research Database (Denmark)

    Wang, Qian; Yang, Zhenyu

    2014-01-01

    A new simple and low cost method of quantification of trace amounts of silicon (Si) in silicone oils has been developed by combining the silicone emulsion and inductively coupled plasma optical emission spectroscopy (ICP-OES) analysis. Silicone oils that contained phenyl groups in the viscosity...... range from 20 to 1000 mPa.s formed stable oil in water emulsions in the presence of Tween 80 surfactant and methylisobutylketone (MIBK). The Si in the emulsions was further quantified by ICP-OES. The calibration was performed using spiked inorganic silicon standard in the emulsions and the method...... (LOD) in the emulsion as 0.5 ppm Si. Compared to the Si determination by the direct organic solvent ICP-OES, this method is much more convenient, where a regular ICP-OES instrument can be directly used for the quantification of Si in the silicone oils obtained via extraction by organic solvents from...

  11. Collaborative framework for PIV uncertainty quantification: comparative assessment of methods

    International Nuclear Information System (INIS)

    A posteriori uncertainty quantification of particle image velocimetry (PIV) data is essential to obtain accurate estimates of the uncertainty associated with a given experiment. This is particularly relevant when measurements are used to validate computational models or in design and decision processes. In spite of the importance of the subject, the first PIV uncertainty quantification (PIV-UQ) methods have been developed only in the last three years. The present work is a comparative assessment of four approaches recently proposed in the literature: the uncertainty surface method (Timmins et al 2012), the particle disparity approach (Sciacchitano et al 2013), the peak ratio criterion (Charonko and Vlachos 2013) and the correlation statistics method (Wieneke 2015). The analysis is based upon experiments conducted for this specific purpose, where several measurement techniques are employed simultaneously. The performances of the above approaches are surveyed across different measurement conditions and flow regimes. (paper)

  12. Reliability of resistivity quantification for shallow subsurface water processes

    CERN Document Server

    Rings, Joerg; 10.1016/j.jappgeo.2009.03.008

    2009-01-01

    The reliability of surface-based electrical resistivity tomography (ERT) for quantifying resistivities for shallow subsurface water processes is analysed. A method comprising numerical simulations of water movement in soil and forward-inverse modeling of ERT surveys for two synthetic data sets is presented. Resistivity contrast, e.g. by changing water content, is shown to have large influence on the resistivity quantification. An ensemble and clustering approach is introduced in which ensembles of 50 different inversion models for one data set are created by randomly varying the parameters for a regularisation based inversion routine. The ensemble members are sorted into five clusters of similar models and the mean model for each cluster is computed. Distinguishing persisting features in the mean models from singular artifacts in individual tomograms can improve the interpretation of inversion results. Especially in the presence of large resistivity contrasts in high sensitivity areas, the quantification of r...

  13. CT quantification of pleuropulmonary lesions in severe thoracic trauma

    International Nuclear Information System (INIS)

    Purpose: Computed quantification of the extent of pleuropulmonary trauma by CT and comparison with conventional chest X-ray - Impact on therapy and correlation with mechanical ventilation support and clinical outcome. Method: In a prospective trial, 50 patients with clinically suspicious blunt chest trauma were evaluated using CT and conventional chest X-ray. The computed quantification of ventilated lung provided by CT volumetry was correlated with the consecutive artificial respiration parameters and the clinical outcome. Results: We found a high correlation between CT volumetry and artificial ventilation concerning maximal pressures and inspiratory oxygen concentration (FiO2, Goris-Score) (r=0.89, Pearson). The graduation of thoracic trauma correlated highly with the duration of mechanical ventilation (r=0.98, Pearson). Especially with regard to atelectases and lung contusions CT is superior compared to conventional chest X-ray; only 32% and 43%, respectively, were identified by conventional chest X-ray. (orig./AJ)

  14. Quantification of aortic regurgitation by magnetic resonance velocity mapping

    DEFF Research Database (Denmark)

    Søndergaard, Lise; Lindvig, K; Hildebrandt, P;

    1993-01-01

    The use of magnetic resonance (MR) velocity mapping in the quantification of aortic valvular blood flow was examined in 10 patients with angiographically verified aortic regurgitation. MR velocity mapping succeeded in identifying and quantifying the regurgitation in all patients, and the regurgit......The use of magnetic resonance (MR) velocity mapping in the quantification of aortic valvular blood flow was examined in 10 patients with angiographically verified aortic regurgitation. MR velocity mapping succeeded in identifying and quantifying the regurgitation in all patients......, and the regurgitant volume determined with MR velocity mapping agreed well with the grade obtained by aortic root angiography (p velocity mapping...... and calculated from MR imaging of the left ventricular end-diastolic and end-systolic volumes in eight patients (Y = 0.89 x X + 11, r = 0.97, p velocity mapping and simultaneous 125I...

  15. A highly sensitive method for quantification of iohexol

    DEFF Research Database (Denmark)

    Schulz, A.; Boeringer, F.; Swifka, J.;

    2014-01-01

    lohexol (1-N,3-N-bis(2,3-dihydroxypropyl)-5-IN-(2,3-dihydroxypropyl) acetamide-2,4,6-triiodobenzene1,3-dicarboxamide) is used for accurate determination of the glomerular filtration rate (GFR) in chronic kidney disease (CKD) patients. However, high iohexol amounts might lead to adverse effects in...... in organisms. In order to minimize the iohexol dosage required for the GFR determination in humans, the development of a sensitive quantification method is essential. Therefore, the objective of our preclinical study was to establish and validate a simple and robust liquid...... ng (r2 = 0.998). The lowest limit of quantification (LLOQ) was 50 pg. The intraand inter-day accuracies were between 91.2% and 98.7%. The intra- and inter-day precisions were between 2.7% and 9.2%. The recovery rate of iohexol was determined in the range of 100.8 10.9%. The gene expression test...

  16. Uncertainty Quantification for Large-Scale Ice Sheet Modeling

    Energy Technology Data Exchange (ETDEWEB)

    Ghattas, Omar [Univ. of Texas, Austin, TX (United States)

    2016-02-05

    This report summarizes our work to develop advanced forward and inverse solvers and uncertainty quantification capabilities for a nonlinear 3D full Stokes continental-scale ice sheet flow model. The components include: (1) forward solver: a new state-of-the-art parallel adaptive scalable high-order-accurate mass-conservative Newton-based 3D nonlinear full Stokes ice sheet flow simulator; (2) inverse solver: a new adjoint-based inexact Newton method for solution of deterministic inverse problems governed by the above 3D nonlinear full Stokes ice flow model; and (3) uncertainty quantification: a novel Hessian-based Bayesian method for quantifying uncertainties in the inverse ice sheet flow solution and propagating them forward into predictions of quantities of interest such as ice mass flux to the ocean.

  17. Quantification of landscape multifunctionality based on farm functionality indices

    DEFF Research Database (Denmark)

    Andersen, Peter Stubkjær; Vejre, Henrik; Dalgaard, Tommy;

    2011-01-01

    landscapes differ in the capacity to provide such goods and services (Willemen et al. 2008). The quantification of different functions in comparable units is challenging. Willemen et al. (2010) presented a top-down method in which interactions of functions are quantified based on national survey data. We...... and according coldspots are identified. The distribution of functions is compared with the landscapes biophysical factors to identify structural-functional correlations. ( 281 words) Keywords: multifunctionality, quantification, landscape, functional distribution References O’Farrell PJ, Andersen PML, 2010......The term multifunctionality is used to indicate that agricultural landscapes simultanously provide goods and services (functions) e.g., food security, livelihood opportunities, life support system maintenance, and space for cultural and recreational activities (O'Farrell & Anderson 2010). Different...

  18. Statistical approach for uncertainty quantification of experimental modal model parameters

    DEFF Research Database (Denmark)

    Luczak, M.; Peeters, B.; Kahsin, M.;

    2014-01-01

    estimates obtained from vibration experiments. Modal testing results are influenced by numerous factors introducing uncertainty to the measurement results. Different experimental techniques applied to the same test item or testing numerous nominally identical specimens yields different test results....... This paper aims at a systematic approach for uncertainty quantification of the parameters of the modal models estimated from experimentally obtained data. Statistical analysis of modal parameters is implemented to derive an assessment of the entire modal model uncertainty measure. Investigated structures...

  19. RNA quantification using gold nanoprobes - application to cancer diagnostics

    Directory of Open Access Journals (Sweden)

    Baptista Pedro V

    2010-02-01

    Full Text Available Abstract Molecular nanodiagnostics applied to cancer may provide rapid and sensitive detection of cancer related molecular alterations, which would enable early detection even when those alterations occur only in a small percentage of cells. The use of gold nanoparticles derivatized with thiol modified oligonucleotides (Au-nanoprobes for the detection of specific nucleic acid targets has been gaining momentum as an alternative to more traditional methodologies. Here, we present an Au-nanoparticles based approach for the molecular recognition and quantification of the BCR-ABL fusion transcript (mRNA, which is responsible for chronic myeloid leukemia (CML, and to the best of our knowledge it is the first time quantification of a specific mRNA directly in cancer cells is reported. This inexpensive and very easy to perform Au-nanoprobe based method allows quantification of unamplified total human RNA and specific detection of the oncogene transcript. The sensitivity settled by the Au-nanoprobes allows differential gene expression from 10 ng/μl of total RNA and takes less than 30 min to complete after total RNA extraction, minimizing RNA degradation. Also, at later stages, accumulation of malignant mutations may lead to resistance to chemotherapy and consequently poor outcome. Such a method, allowing for fast and direct detection and quantification of the chimeric BCR-ABL mRNA, could speed up diagnostics and, if appropriate, revision of therapy. This assay may constitute a promising tool in early diagnosis of CML and could easily be extended to further target genes with proven involvement in cancer development.

  20. Frequency feature based quantification of defect depth and thickness

    Science.gov (United States)

    Tian, Shulin; Chen, Kai; Bai, Libing; Cheng, Yuhua; Tian, Lulu; Zhang, Hong

    2014-06-01

    This study develops a frequency feature based pulsed eddy current method. A frequency feature, termed frequency to zero, is proposed for subsurface defects and metal loss quantification in metallic specimens. A curve fitting method is also employed to generate extra frequency components and improve the accuracy of the proposed method. Experimental validation is carried out. Conclusions and further work are derived on the basis of the studies.

  1. Methodological Issues in the Quantification of Respiratory Sinus Arrhythmia

    OpenAIRE

    Denver, John W.; Reed, Shawn F.; Porges, Stephen W.

    2006-01-01

    Although respiratory sinus arrhythmia (RSA) is a commonly quantified physiological variable, the methods for quantification are not consistent. This manuscript questions the assumption that respiration frequency needs to be manipulated or monitored to generate an accurate measure of RSA amplitude. A review of recent papers is presented that contrast RSA amplitude with measures that use respiratory parameters to adjust RSA amplitude. In addition, data from two studies are presented to evaluate...

  2. A Novel Immunological Assay for Hepcidin Quantification in Human Serum

    OpenAIRE

    Koliaraki, V.; Marinou, M.; Vassilakopoulos, T P; Vavourakis, E.; Tsochatzis, E.; Pangalis, G. A.; Papatheodoridis, G; Stamoulakatou, A.; Swinkels, D. W.; Papanikolaou, G; A. Mamalaki

    2009-01-01

    BACKGROUND: Hepcidin is a 25-aminoacid cysteine-rich iron regulating peptide. Increased hepcidin concentrations lead to iron sequestration in macrophages, contributing to the pathogenesis of anaemia of chronic disease whereas decreased hepcidin is observed in iron deficiency and primary iron overload diseases such as hereditary hemochromatosis. Hepcidin quantification in human blood or urine may provide further insights for the pathogenesis of disorders of iron homeostasis and might prove a v...

  3. Quantification of clay minerals by combined EWA/XRD method

    Institute of Scientific and Technical Information of China (English)

    XU; Jianhong; (徐建红); XU; Jianhong; (徐建红); T.; R.; Astin; PAN; Mao; (潘懋)

    2001-01-01

    Illite has been considered the main constraint on permeability in the Morecambe Gas Field, East Irish Sea, UK. Previous research has emphasized the morphology rather than the amount of clay minerals. By applying a new method of clay mineral quantification, EWA/XRD, and applying statistical analysis methods, we are able to establish a quantitative model of illite distribution in the field. The result also leads to a better understanding of permeability distribution in reservoir sandstones.

  4. QUANTIFICATIONS OF THE DETRIMENTAL HEALTHEFFECTS OF IONISING RADIATION

    OpenAIRE

    Walsh, Linda

    2013-01-01

    This thesis, presented to The University of Manchester in 2012 by Dr. Linda Walsh, is in fulfilment of the requirements for the degree of Doctor of Science (DSc) and is entitled “Quantifications of the detrimental health effects of ionising radiation.” A body of work and ensuing publications covering 2000–2012 are presented, predominantly concerning studies of various cohorts of people exposed to ionising radiation. The major areas cover epidemiological and statistical studies on the Life spa...

  5. Color correction for automatic fibrosis quantification in liver biopsy specimens

    Directory of Open Access Journals (Sweden)

    Yuri Murakami

    2013-01-01

    Full Text Available Context: For a precise and objective quantification of liver fibrosis, quantitative evaluations through image analysis have been utilized. However, manual operations are required in most cases for extracting fiber areas because of color variation included in digital pathology images. Aims: The purpose of this research is to propose a color correction method for whole slide images (WSIs of Elastica van Gieson (EVG stained liver biopsy tissue specimens and to realize automated operation of image analysis for fibrosis quantification. Materials and Methods: Our experimental dataset consisted of 38 WSIs of liver biopsy specimens collected from 38 chronic viral hepatitis patients from multiple medical facilities, stained with EVG and scanned at ×20 using a Nano Zoomer 2.0 HT (Hamamatsu Photonics K.K., Hamamatsu, Japan. Color correction was performed by modifying the color distribution of a target WSI so as to fit to the reference, where the color distribution was modeled by a set of two triangle pyramids. Using color corrected WSIs; fibrosis quantification was performed based on tissue classification analysis. Statistical Analysis Used: Spearman′s rank correlation coefficients were calculated between liver stiffness measured by transient elastography and median area ratio of collagen fibers calculated based on tissue classification results. Results: Statistical analysis results showed a significant correlation r = 0.61-0.68 even when tissue classifiers were trained by using a subset of WSIs, while the correlation coefficients were reduced to r = 0.40-0.50 without color correction. Conclusions: Fibrosis quantification accompanied with the proposed color correction method could provide an objective evaluation tool for liver fibrosis, which complements semi-quantitative histologic evaluation systems.

  6. Standardless quantification by parameter optimization in electron probe microanalysis

    Energy Technology Data Exchange (ETDEWEB)

    Limandri, Silvina P. [Instituto de Fisica Enrique Gaviola (IFEG), CONICET (Argentina); Facultad de Matematica, Astronomia y Fisica, Universidad Nacional de Cordoba, Medina Allende s/n, (5016) Cordoba (Argentina); Bonetto, Rita D. [Centro de Investigacion y Desarrollo en Ciencias Aplicadas Dr. Jorge Ronco (CINDECA), CONICET, 47 Street 257, (1900) La Plata (Argentina); Facultad de Ciencias Exactas, Universidad Nacional de La Plata, 1 and 47 Streets (1900) La Plata (Argentina); Josa, Victor Galvan; Carreras, Alejo C. [Instituto de Fisica Enrique Gaviola (IFEG), CONICET (Argentina); Facultad de Matematica, Astronomia y Fisica, Universidad Nacional de Cordoba, Medina Allende s/n, (5016) Cordoba (Argentina); Trincavelli, Jorge C., E-mail: trincavelli@famaf.unc.edu.ar [Instituto de Fisica Enrique Gaviola (IFEG), CONICET (Argentina); Facultad de Matematica, Astronomia y Fisica, Universidad Nacional de Cordoba, Medina Allende s/n, (5016) Cordoba (Argentina)

    2012-11-15

    A method for standardless quantification by parameter optimization in electron probe microanalysis is presented. The method consists in minimizing the quadratic differences between an experimental spectrum and an analytical function proposed to describe it, by optimizing the parameters involved in the analytical prediction. This algorithm, implemented in the software POEMA (Parameter Optimization in Electron Probe Microanalysis), allows the determination of the elemental concentrations, along with their uncertainties. The method was tested in a set of 159 elemental constituents corresponding to 36 spectra of standards (mostly minerals) that include trace elements. The results were compared with those obtained with the commercial software GENESIS Spectrum Registered-Sign for standardless quantification. The quantifications performed with the method proposed here are better in the 74% of the cases studied. In addition, the performance of the method proposed is compared with the first principles standardless analysis procedure DTSA for a different data set, which excludes trace elements. The relative deviations with respect to the nominal concentrations are lower than 0.04, 0.08 and 0.35 for the 66% of the cases for POEMA, GENESIS and DTSA, respectively. - Highlights: Black-Right-Pointing-Pointer A method for standardless quantification in EPMA is presented. Black-Right-Pointing-Pointer It gives better results than the commercial software GENESIS Spectrum. Black-Right-Pointing-Pointer It gives better results than the software DTSA. Black-Right-Pointing-Pointer It allows the determination of the conductive coating thickness. Black-Right-Pointing-Pointer It gives an estimation for the concentration uncertainties.

  7. Quantification of myocardial perfusion by cardiovascular magnetic resonance

    OpenAIRE

    Jerosch-Herold Michael

    2010-01-01

    Abstract The potential of contrast-enhanced cardiovascular magnetic resonance (CMR) for a quantitative assessment of myocardial perfusion has been explored for more than a decade now, with encouraging results from comparisons with accepted "gold standards", such as microspheres used in the physiology laboratory. This has generated an increasing interest in the requirements and methodological approaches for the non-invasive quantification of myocardial blood flow by CMR. This review provides a...

  8. Universal quantification for deterministic chaos in dynamical systems

    OpenAIRE

    Selvam, A. Mary

    2000-01-01

    A cell dynamical system model for deterministic chaos enables precise quantification of the round-off error growth,i.e., deterministic chaos in digital computer realizations of mathematical models of continuum dynamical systems. The model predicts the following: (a) The phase space trajectory (strange attractor) when resolved as a function of the computer accuracy has intrinsic logarithmic spiral curvature with the quasiperiodic Penrose tiling pattern for the internal structure. (b) The unive...

  9. The necessity of operational risk management and quantification

    OpenAIRE

    Barbu Teodora Cristina; Olteanu (Puiu) Ana Cornelia; Radu Alina Nicoleta

    2008-01-01

    Beginning with the fact that performant strategies of the financial institutions have programmes and management procedures for the banking risks, which have as main objective to minimize the probability of risk generation and the bank’s potential exposure, this paper wants to present the operational risk management and quantification methods. Also it presents the modality of minimum capital requirement for the operational risk. Therefore, the first part presents the conceptual approach of the...

  10. A "Toy" Model for Operational Risk Quantification using Credibility Theory

    OpenAIRE

    Hans B\\"uhlmann; Shevchenko, Pavel V.; Mario V. W\\"uthrich

    2009-01-01

    To meet the Basel II regulatory requirements for the Advanced Measurement Approaches in operational risk, the bank's internal model should make use of the internal data, relevant external data, scenario analysis and factors reflecting the business environment and internal control systems. One of the unresolved challenges in operational risk is combining of these data sources appropriately. In this paper we focus on quantification of the low frequency high impact losses exceeding some high thr...

  11. Single-Molecule Fluorescence Quantification with a Photobleached Internal Standard

    OpenAIRE

    Gadd, Jennifer C.; Fujimoto, Bryant S.; Sandra M Bajjalieh; Chiu, Daniel T.

    2012-01-01

    In cellular and molecular biology, fluorophores are employed to aid in tracking and quantifying molecules involved in cellular function. We previously developed a sensitive single-molecule quantification technique to count the number of proteins and the variation of the protein number over the population of individual sub-cellular organelles. However, environmental effects on the fluorescent intensity of fluorophores can make it difficult to accurately quantify proteins using these sensitive ...

  12. Lessons in uncertainty quantification for turbulent dynamical systems

    OpenAIRE

    Majda, A. J.; M. Branicki

    2012-01-01

    The modus operandi of modern applied mathematics in developing very recent mathematical strategies for uncertainty quantification in partially observed high-dimensional turbulent dynamical systems is emphasized here. The approach involves the synergy of rigorous mathematical guidelines with a suite of physically relevant and progressively more complex test models which are mathematically tractable while possessing such important features as the two-way coupling between the resolved dynamics a...

  13. Initial water quantification results using neutron computed tomography

    Energy Technology Data Exchange (ETDEWEB)

    Heller, A.K. [Department of Mechanical and Nuclear Engineering, Pennsylvania State University (United States)], E-mail: axh174@psu.edu; Shi, L.; Brenizer, J.S.; Mench, M.M. [Department of Mechanical and Nuclear Engineering, Pennsylvania State University (United States)

    2009-06-21

    Neutron computed tomography is an important imaging tool in the field of non-destructive testing and in fundamental research for many engineering applications. Contrary to X-rays, neutrons can be attenuated by some light materials, such as hydrogen, but can penetrate many heavy materials. Thus, neutron computed tomography is useful in obtaining important three-dimensional information about a sample's interior structure and material properties that other traditional methods cannot provide. The neutron computed tomography system at Pennsylvania State University's Radiation Science and Engineering Center is being utilized to develop a water quantification technique for investigation of water distribution in fuel cells under normal conditions. A hollow aluminum cylinder test sample filled with a known volume of water was constructed for purposes of testing the quantification technique. Transmission images of the test sample at different angles were easily acquired through the synthesis of a dedicated image acquisition computer driving a rotary table controller and an in-house developed synchronization software package. After data acquisition, Octopus (version 8.2) and VGStudio Max (version 1.2) were used to perform cross-sectional and three-dimensional reconstructions of the sample, respectively. The initial reconstructions and water quantification results are presented.

  14. Assessment methods for angiogenesis and current approaches for its quantification

    Directory of Open Access Journals (Sweden)

    Waleed Hassan AlMalki

    2014-01-01

    Full Text Available Angiogenesis is a physiological process which describes the development of new blood vessels from the existing vessels. It is a common and the most important process in the formation and development of blood vessels, so it is supportive in the healing of wounds and granulation of tissues. The different assays for the evaluation of angiogenesis have been described with distinct advantages and some limitations. In order to develop angiogenic and antiangiogenic techniques, continuous efforts have been resulted to give animal models for more quantitative analysis of angiogenesis. Most of the studies on angiogenic inducers and inhibitors rely on various models, both in vitro, in vivo and in ova, as indicators of efficacy. The angiogenesis assays are very much helpful to test efficacy of both pro- and anti- angiogenic agents. The development of non-invasive procedures for quantification of angiogenesis will facilitate this process significantly. The main objective of this review article is to focus on the novel and existing methods of angiogenesis and their quantification techniques. These findings will be helpful to establish the most convenient methods for the detection, quantification of angiogenesis and to develop a novel, well tolerated and cost effective anti-angiogenic treatment in the near future.

  15. Leveraging transcript quantification for fast computation of alternative splicing profiles.

    Science.gov (United States)

    Alamancos, Gael P; Pagès, Amadís; Trincado, Juan L; Bellora, Nicolás; Eyras, Eduardo

    2015-09-01

    Alternative splicing plays an essential role in many cellular processes and bears major relevance in the understanding of multiple diseases, including cancer. High-throughput RNA sequencing allows genome-wide analyses of splicing across multiple conditions. However, the increasing number of available data sets represents a major challenge in terms of computation time and storage requirements. We describe SUPPA, a computational tool to calculate relative inclusion values of alternative splicing events, exploiting fast transcript quantification. SUPPA accuracy is comparable and sometimes superior to standard methods using simulated as well as real RNA-sequencing data compared with experimentally validated events. We assess the variability in terms of the choice of annotation and provide evidence that using complete transcripts rather than more transcripts per gene provides better estimates. Moreover, SUPPA coupled with de novo transcript reconstruction methods does not achieve accuracies as high as using quantification of known transcripts, but remains comparable to existing methods. Finally, we show that SUPPA is more than 1000 times faster than standard methods. Coupled with fast transcript quantification, SUPPA provides inclusion values at a much higher speed than existing methods without compromising accuracy, thereby facilitating the systematic splicing analysis of large data sets with limited computational resources. The software is implemented in Python 2.7 and is available under the MIT license at https://bitbucket.org/regulatorygenomicsupf/suppa. PMID:26179515

  16. In vivo behavior of NTBI revealed by automated quantification system.

    Science.gov (United States)

    Ito, Satoshi; Ikuta, Katsuya; Kato, Daisuke; Lynda, Addo; Shibusa, Kotoe; Niizeki, Noriyasu; Toki, Yasumichi; Hatayama, Mayumi; Yamamoto, Masayo; Shindo, Motohiro; Iizuka, Naomi; Kohgo, Yutaka; Fujiya, Mikihiro

    2016-08-01

    Non-Tf-bound iron (NTBI), which appears in serum in iron overload, is thought to contribute to organ damage; the monitoring of serum NTBI levels may therefore be clinically useful in iron-overloaded patients. However, NTBI quantification methods remain complex, limiting their use in clinical practice. To overcome the technical difficulties often encountered, we recently developed a novel automated NTBI quantification system capable of measuring large numbers of samples. In the present study, we investigated the in vivo behavior of NTBI in human and animal serum using this newly established automated system. Average NTBI in healthy volunteers was 0.44 ± 0.076 μM (median 0.45 μM, range 0.28-0.66 μM), with no significant difference between sexes. Additionally, serum NTBI rapidly increased after iron loading, followed by a sudden disappearance. NTBI levels also decreased in inflammation. The results indicate that NTBI is a unique marker of iron metabolism, unlike other markers of iron metabolism, such as serum ferritin. Our new automated NTBI quantification method may help to reveal the clinical significance of NTBI and contribute to our understanding of iron overload. PMID:27086349

  17. A fast and robust hepatocyte quantification algorithm including vein processing

    Directory of Open Access Journals (Sweden)

    Homeyer André

    2010-03-01

    Full Text Available Abstract Background Quantification of different types of cells is often needed for analysis of histological images. In our project, we compute the relative number of proliferating hepatocytes for the evaluation of the regeneration process after partial hepatectomy in normal rat livers. Results Our presented automatic approach for hepatocyte (HC quantification is suitable for the analysis of an entire digitized histological section given in form of a series of images. It is the main part of an automatic hepatocyte quantification tool that allows for the computation of the ratio between the number of proliferating HC-nuclei and the total number of all HC-nuclei for a series of images in one processing run. The processing pipeline allows us to obtain desired and valuable results for a wide range of images with different properties without additional parameter adjustment. Comparing the obtained segmentation results with a manually retrieved segmentation mask which is considered to be the ground truth, we achieve results with sensitivity above 90% and false positive fraction below 15%. Conclusions The proposed automatic procedure gives results with high sensitivity and low false positive fraction and can be applied to process entire stained sections.

  18. Gas plume quantification in downlooking hyperspectral longwave infrared images

    Science.gov (United States)

    Turcotte, Caroline S.; Davenport, Michael R.

    2010-10-01

    Algorithms have been developed to support quantitative analysis of a gas plume using down-looking airborne hyperspectral long-wave infrared (LWIR) imagery. The resulting gas quantification "GQ" tool estimates the quantity of one or more gases at each pixel, and estimates uncertainty based on factors such as atmospheric transmittance, background clutter, and plume temperature contrast. GQ uses gas-insensitive segmentation algorithms to classify the background very precisely so that it can infer gas quantities from the differences between plume-bearing pixels and similar non-plume pixels. It also includes MODTRAN-based algorithms to iteratively assess various profiles of air temperature, water vapour, and ozone, and select the one that implies smooth emissivity curves for the (unknown) materials on the ground. GQ then uses a generalized least-squares (GLS) algorithm to simultaneously estimate the most likely mixture of background (terrain) material and foreground plume gases. Cross-linking of plume temperature to the estimated gas quantity is very non-linear, so the GLS solution was iteratively assessed over a range of plume temperatures to find the best fit to the observed spectrum. Quantification errors due to local variations in the camera-topixel distance were suppressed using a subspace projection operator. Lacking detailed depth-maps for real plumes, the GQ algorithm was tested on synthetic scenes generated by the Digital Imaging and Remote Sensing Image Generation (DIRSIG) software. Initial results showed pixel-by-pixel gas quantification errors of less than 15% for a Freon 134a plume.

  19. Initial water quantification results using neutron computed tomography

    International Nuclear Information System (INIS)

    Neutron computed tomography is an important imaging tool in the field of non-destructive testing and in fundamental research for many engineering applications. Contrary to X-rays, neutrons can be attenuated by some light materials, such as hydrogen, but can penetrate many heavy materials. Thus, neutron computed tomography is useful in obtaining important three-dimensional information about a sample's interior structure and material properties that other traditional methods cannot provide. The neutron computed tomography system at Pennsylvania State University's Radiation Science and Engineering Center is being utilized to develop a water quantification technique for investigation of water distribution in fuel cells under normal conditions. A hollow aluminum cylinder test sample filled with a known volume of water was constructed for purposes of testing the quantification technique. Transmission images of the test sample at different angles were easily acquired through the synthesis of a dedicated image acquisition computer driving a rotary table controller and an in-house developed synchronization software package. After data acquisition, Octopus (version 8.2) and VGStudio Max (version 1.2) were used to perform cross-sectional and three-dimensional reconstructions of the sample, respectively. The initial reconstructions and water quantification results are presented.

  20. Gas chromatographic validated method for quantification of ayurvedic polyherbal formulation

    Directory of Open Access Journals (Sweden)

    Navdeep Saini

    2015-01-01

    Full Text Available A new gas chromatographic-flame ionization detector (GC-FID method was developed for quantification of ayurvedic polyherbal formulation. The GC-FID method was found highly accurate, sensitive, simple and precise. This method was validated as per international conference on harmonization (ICH guidelines. Experimental work was performed by nonpolar capillary column (Zb-5, 5%-Phenyl-95%-dimethylpolysiloxane. Film thickness of capillary column (Zb-5 was (0.25 μm and length 30 m × 0.25 mm i.d. The temperature of the oven, injector and detector were 200, 210 and 280°C respectively. Data processing system was applied to obtain data. The standards and test samples were prepared in absolute ethanol. The principle constituents t-Anethol, d-Limonene, cuminaldehyde and thymol were found in ayurvedic polyherbal formulation. The ICH validation parameters for the proposed procedure, recovery (limit 98.85-100.76%, precision (<1.00%, limits of detection, limits of quantification and linearity (r2 = 0.995 ± 0.002 were observed under acceptance limit. Validation results were statistically calculated. The result shows that method is selective and reproducible for quantification of ayurvedic polyherbal formulation. The presented GC method can be applied for the routine analysis of principle constituents as well as ayurvedic polyherbal formulation.

  1. Quantification of rice bran oil in oil blends

    Energy Technology Data Exchange (ETDEWEB)

    Mishra, R.; Sharma, H. K.; Sengar, G.

    2012-11-01

    Blends consisting of physically refined rice bran oil (PRBO): sunflower oil (SnF) and PRBO: safflower oil (SAF) in different proportions were analyzed for various physicochemical parameters. The quantification of pure rice bran oil in the blended oils was carried out using different methods including gas chromatographic, HPLC, ultrasonic velocity and methods based on physico-chemical parameters. The physicochemical parameters such as ultrasonic velocity, relative association and acoustic impedance at 2 MHz, iodine value, palmitic acid content and oryzanol content reflected significant changes with increased proportions of PRBO in the blended oils. These parameters were selected as dependent parameters and % PRBO proportion was selected as independent parameters. The study revealed that regression equations based on the oryzanol content, palmitic acid composition, ultrasonic velocity, relative association, acoustic impedance, and iodine value can be used for the quantification of rice bran oil in blended oils. The rice bran oil can easily be quantified in the blended oils based on the oryzanol content by HPLC even at a 1% level. The palmitic acid content in blended oils can also be used as an indicator to quantify rice bran oil at or above the 20% level in blended oils whereas the method based on ultrasonic velocity, acoustic impedance and relative association showed initial promise in the quantification of rice bran oil. (Author) 23 refs.

  2. Automated Erythema Quantification in Radiation Therapy - a Java Based Tool

    Directory of Open Access Journals (Sweden)

    Paul Martin PUTORA

    2010-03-01

    Full Text Available Introduction: In radiotherapy, erythema is a common side-effect, especially during radiotherapy treatment regimes that last several weeks. The measurement of erythema might be of clinical relevance, especially when standardized interpretation is possible. Aim: The aim of this article is to present a tool that can be implemented for automatized and time efficient quantification of erythema from digital images taken during radiotherapy treatment. Method: Instead of relying on commercially available graphic editors and performing manual operations on the images within these programs we developed a java based tool that can automatically evaluate the “redness” of images. These erythema values receive a score number, are connected with the date and time the pictures were taken and are exported into a comma separated values (CSV file. Results: The Erythema values of images could be quickly evaluated with the developed tool. With spreadsheet software the exported file could be easily manipulated to produce graphical representations of erythema rise. Conclusion: Erythema quantification from digital images can be easily performed by custom developed java tools. An automated quantification provides a method of detecting an increase in erythema that may not be visible to the naked eye.

  3. Automatic quantification of neo-vasculature from micro-CT

    Science.gov (United States)

    Mallya, Yogish; Narayanan, A. K.; Zagorchev, Lyubomir

    2009-02-01

    Angiogenesis is the process of formation of new blood vessels as outgrowths of pre-existing ones. It occurs naturally during development, tissue repair, and abnormally in pathologic diseases such as cancer. It is associated with proliferation of blood vessels/tubular sprouts that penetrate deep into tissues to supply nutrients and remove waste products. The process starts with migration of endothelial cells. As the cells move towards the target area they form small tubular sprouts recruited from the parent vessel. The sprouts grow in length due to migration, proliferation, and recruitment of new endothelial cells and the process continues until the target area becomes fully vascular. Accurate quantification of sprout formation is very important for evaluation of treatments for ischemia as well as angiogenesis inhibitors and plays a key role in the battle against cancer. This paper presents a technique for automatic quantification of newly formed blood vessels from Micro-CT volumes of tumor samples. A semiautomatic technique based on interpolation of Bezier curves was used to segment out the cancerous growths. Small vessels as determined by their diameter within the segmented tumors were enhanced and quantified with a multi-scale 3-D line detection filter. The same technique can be easily extended for quantification of tubular structures in other 3-D medical imaging modalities. Experimental results are presented and discussed.

  4. Automatic quantification in lung scintigraphy: functional atlas; Quantification automatique en scintigraphie pulmonaire: elaboration d'atlas fonctionnels

    Energy Technology Data Exchange (ETDEWEB)

    Ledee, R.; Therain, F. [Orleans Univ., Lab. d' Electronique - Signaux - Images (LESI), 45 (France); Debrun, D. [Centre Hospitalier Regional de la Source (CHRO), Medecine Nucleaire, 45 - Orleans (France)

    2003-06-01

    In spite of the development of new techniques, the ventilation perfusion lung scintigraphy keeps a good place for the diagnosis of pulmonary embolism (PE). In the context of an improvement of reliability and reproducibility of the diagnosis, this study proposes to realize an automatic quantification of the distribution of the radioactive tracers by pulmonary segments. Measurements are made following a procedure of non-rigid matching of morphological 2-D charts of the lungs on the scintigraphic images. The adaptation of these charts to the patients' morphology is carried out by exploiting iso-contour information of the images and using Fourier descriptors to determine the parameters of the transformation. The study was performed on a population of 30 patients with a probability of nil of the pulmonary embolism. After a study of the robustness of the quantification, 2-D segmental functional reference charts (according to the conditions of acquisition) were proposed. In the perfusion case and four views, the following lobar distribution, in relative value, is measured: Right Inferior Lobar = 23,39%, Medial Lobar = 10,41 %, Right Superior Lobar = 20,37%, Left Inferior Lobar 20,6% and Left Superior Lobar = 25.6% with culmen = 18,8% and lingula = 6,8%; values comparable with those of the publication. The process of quantification is adaptable to the ventilation lung scans. The segmental quantifications of a patient carried out under the same conditions of acquisition as the functional reference charts, could be compared with the reference data and provide indicators for the diagnosis but also for patient follow-up and preoperative evaluation of lung cancers. (authors)

  5. Systematic development of a group quantification method using evaporative light scattering detector for relative quantification of ginsenosides in ginseng products.

    Science.gov (United States)

    Lee, Gwang Jin; Shin, Byong-Kyu; Yu, Yun-Hyun; Ahn, Jongsung; Kwon, Sung Won; Park, Jeong Hill

    2016-09-01

    The determination for the contents of multi-components in ginseng products has come to the fore by demands of in-depth information, but the associated industries confront the high cost of securing pure standards for the continuous quality evaluation of the products. This study aimed to develop a prospective high-performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD) method for relative quantification of ginsenosides in ginseng products without a considerable change from the conventional gradient analysis. We investigated the effects of mobile phase composition and elution bandwidth, which are potential variables affecting the ELSD response in the gradient analysis. Similar ELSD response curves of nine major ginsenosides were obtained under the identical flow injection conditions, and the response increased as the percentage of organic solvent increased. The nine ginsenosides were divided into three groups to confirm the effect of elution bandwidth. The ELSD response significantly decreased in case of the late eluted ginsenoside in the individual groups under the isocratic conditions. With the consideration of the two important effects, stepwise changes of the gradient condition were carried out to reach a group quantification method. The inconsistent responses of the nine ginsenosides were reconstituted to three normalized responses by the stepwise changes of the gradient condition, and this result actualized relative quantification in the individual groups. The availability was confirmed by comparing the ginsenoside contents in a base material of ginseng products determined by the direct and group quantification method. The largest difference in the determination results from the two methods was 8.26%, and the difference of total contents was only 0.91%. PMID:27262109

  6. Amplicon DNA melting analysis for the simultaneous detection of Brucella spp and Mycobacterium tuberculosis complex. Potential use in rapid differential diagnosis between extrapulmonary tuberculosis and focal complications of brucellosis.

    Directory of Open Access Journals (Sweden)

    Rocio Sanjuan-Jimenez

    Full Text Available Some sites of extrapulmonary tuberculosis and focal complications of brucellosis are very difficult to differentiate clinically, radiologically, and even histopathologically. Conventional microbiological methods for the diagnosis of extrapulmonary tuberculosis and complicated brucellosis not only lack adequate sensitivity, they are also time consuming, which could lead to an unfavourable prognosis. The aim of this work was to develop a multiplex real-time PCR assay based on SYBR Green I to simultaneously detect Brucella spp and Mycobacterium tuberculosis complex and evaluate the efficacy of the technique with different candidate genes. The IS711, bcsp31 and omp2a genes were used for the identification of Brucella spp and the IS6110, senX3-regX3 and cfp31 genes were targeted for the detection of the M. tuberculosis complex. As a result of the different combinations of primers, nine different reactions were evaluated. A test was defined as positive only when the gene combinations were capable of co-amplifying both pathogens in a single reaction tube and showed distinguishable melting temperatures for each microorganism. According to the melting analysis, only three combinations of amplicons (senX3-regX3+bcsp31, senX3-regX3+IS711 and IS6110+IS711 were visible. Detection limits of senX3-regX3+bcsp31 and senX3-regX3+IS711 were of 2 and 3 genome equivalents for M. tuberculosis complex and Brucella while for IS6110+IS711 they were of 200 and 300 genome equivalents, respectively. The three assays correctly identified all the samples, showing negative results for the control patients. The presence of multicopy elements and GC content were the components most influencing the efficiency of the test; this should be taken into account when designing a multiplex-based SYBR Green I assay. In conclusion, multiplex real time PCR assays based on the targets senX3-regX3+bcsp31 and senX3-regX3+IS711 using SYBR Green I are highly sensitive and reproducible. This may

  7. A method for simultaneous quantification of phospholipid species by routine 31P NMR

    DEFF Research Database (Denmark)

    Brinkmann-Trettenes, Ulla; Stein, Paul C.; Klösgen, Beate Maria;

    2012-01-01

    We report a 31P NMR assay for quantification of aqueous phospholipid samples. Using a capillary with trimethylphosphate as internal standard, the limit of quantification is 1.30mM. Comparison of the 31P NMR quantification method in aqueous buffer and in organic solvent revealed that the two metho...... fast results of a limited number of samples are requested. © 2012 Elsevier B.V.....

  8. The sociology of quantification - perspectives on an emerging field in the social sciences

    OpenAIRE

    Diaz-Bone, Rainer; Didier, Emmanuel

    2016-01-01

    The introductory article to this HSR Special Issue presents the emerging field of sociology of quantification, which can be regarded as a transdisciplinary approach to the analysis of processes of quantification. Processes of categorization and classification are included because they can result in processes of generating figures and numbers also. The contribution sketches the science-historical development of this field. It is argued that processes of quantification are related in many ways ...

  9. Learning-guided automatic three dimensional synapse quantification for drosophila neurons

    OpenAIRE

    Sanders, Jonathan; Singh, Anil; Sterne, Gabriella; Ye, Bing; Zhou, Jie

    2015-01-01

    Background The subcellular distribution of synapses is fundamentally important for the assembly, function, and plasticity of the nervous system. Automated and effective quantification tools are a prerequisite to large-scale studies of the molecular mechanisms of subcellular synapse distribution. Common practices for synapse quantification in neuroscience labs remain largely manual or semi-manual. This is mainly due to computational challenges in automatic quantification of synapses, including...

  10. QUANTIFICATION OF GENETICALLY MODIFIED MAIZE MON 810 IN PROCESSED FOODS

    Directory of Open Access Journals (Sweden)

    Peter Siekel

    2012-12-01

    Full Text Available 800x600 Normal 0 21 false false false SK X-NONE X-NONE MicrosoftInternetExplorer4 Maize MON 810 (Zea mays L. represents the majority of genetically modified food crops. It is the only transgenic cultivar grown in the EU (European Union countries and food products with its content higher than 0.9 % must be labelled. This study was aimed at impact of food processing (temperature, pH and pressure on DNA degradation and quantification of the genetically modified maize MON 810. The transgenic DNA was quantified by the real-time polymerase chain reaction method. Processing as is high temperature (121 °C, elevated pressure (0.1 MPa and low pH 2.25 fragmented DNA. A consequence of two order difference in the species specific gene content compared to the transgenic DNA content in plant materials used has led to false negative results in the quantification of transgenic DNA. The maize containing 4.2 % of the transgene after processing appeared to be as low as 3.0 % (100 °C and 1.9 % (121 °C, 0.1 MPa. The 2.1 % amount of transgene dropped at 100 °C to 1.0 % and at 121 °C, 0.1 MPa to 0.6 %. Under such make up the DNA degradation of transgenic content showed up 2 or 3 time higher decrease a consequence of unequal gene presence. Such genes disparity is expressed as considerable decrease of transgenic content while the decrease of species specific gene content remains unnoticed. Based on our findings we conclude that high degree of processing might have led to false negative results of the transgenic constituent quantification. Determination of GMO content in processed foods may leads to incorrect statement and labelling in these cases could misleads consumers.doi:10.5219/212

  11. Quantification of Carnosine-Aldehyde Adducts in Human Urine.

    Science.gov (United States)

    da Silva Bispo, Vanderson; Di Mascio, Paolo; Medeiros, Marisa

    2014-10-01

    Lipid peroxidation generates several reactive carbonyl species, including 4-hydroxy-2-nonenal (HNE), acrolein (ACR), 4-hydroxy-2-hexenal (HHE) and malondialdehyde. One major pathwayof aldehydes detoxification is through conjugation with glutathione catalyzed by glutathione-S-transferases or, alternatively, by conjugation with endogenous histidine containing dipeptides, such as carnosine (CAR). In this study, on-line reverse-phase high-performance liquid chromatography (HPLC) separation with tandem mass spectrometry detection was utilized for the accurate quantification of CAR- ACR, CAR-HHE and CAR-HNE adducts in human urinary samples from non-smokers young adults. Standard adducts were prepared and isolated by HPLC. The results showed the presence of a new product from the reaction of CAR with ACR. This new adduct was completely characterized by HPLC/MS-MSn, 1H RMN, COSY and HSQC. The new HPLC/MS/MS methodology employing stable isotope-labeled internal standards (CAR-HHEd5 and CAR-HNEd11) was developed for adducts quantification. This methodology permits quantification of 10pmol CAR-HHE and 1pmol of CAR-ACR and CAR-HNE. Accurate determinations in human urine sample were performed and showed 4.65±1.71 to CAR-ACR, 5.13±1.76 to CAR-HHE and 5.99±3.19nmol/mg creatinine to CAR-HNE. Our results indicate that carnosine pathways can be an important detoxification route of a, ß -unsaturated aldehydes. Moreover, carnosine adducts may be useful as redox stress indicator. PMID:26461323

  12. DNA imaging and quantification using chemi-luminescent probes

    International Nuclear Information System (INIS)

    During this interdisciplinary study we have developed an ultra sensitive and reliable imaging system of DNA labelled by chemiluminescence. Based on a liquid nitrogen cooled CCD, the system achieves sensitivities down to 10 fg/mm2 labelled DNA over a surface area of 25 x 25 cm2 with a sub-millimeter resolution. Commercially available chemi-luminescent - and enhancer molecules are compared and their reaction conditions optimized for best signal-to-noise ratios. Double labelling was performed to verify quantification with radioactive probes. (authors)

  13. MM98.57 Quantification of Combined Strain Paths

    DEFF Research Database (Denmark)

    Nielsen, Morten Sturgård; Wanheim, Tarras

    1998-01-01

    Hitherto the quantification of material properties of a material submitted to a deformation has been taken as a function of one or more internal variables. When making measurements of yield properties as a function of a deformation a convenient way of describing the deformation in an experiment is...... surface at a given point in a new strain history. A simple example of this concept is to take the length of the strain curve as describing scalar relation: E.g. to use the equivalent strain as parameter for describing the yield stress. This paper focuses on the strain curve concept and the possibilities...

  14. HPTLC densitometric quantification of stigmasterol and lupeol from Ficus religiosa

    OpenAIRE

    Deepti Rathee; Sushila Rathee; Permender Rathee; Aakash Deep; Sheetal Anandjiwala; Dharmender Rathee

    2015-01-01

    This study presents the first report of TLC densitometric method, which has been developed and validated for simultaneous quantification of the two marker compounds (stigmasterol and lupeol) from methanolic extract using the solvent system of toluene:methanol (9:1, v/v). The method employed TLC aluminum plates precoated with silica gel 60 F254 as the stationary phase. Densitometric analysis of stigmasterol and lupeol was carried out in the reflectance mode at 525 nm. The system was found to g...

  15. Trace cancer biomarker quantification using polystyrene-functionalized gold nanorods

    Science.gov (United States)

    Wu, Jian; Li, Wei; Hajisalem, Ghazal; Lukach, Ariella; Kumacheva, Eugenia; Hof, Fraser; Gordon, Reuven

    2014-01-01

    We demonstrate the application of polystyrene-functionalized gold nanorods (AuNRs) as a platform for surface enhanced Raman scattering (SERS) quantification of the exogenous cancer biomarker Acetyl Amantadine (AcAm). We utilize the hydrophobicity of the polystyrene attached to the AuNR surface to capture the hydrophobic AcAm from solution, followed by drying and detection using SERS. We achieve a detection limit of 16 ng/mL using this platform. This result shows clinical potential for low-cost early cancer detection. PMID:25574423

  16. Method for quantification of aerobic anoxygenic phototrophic bacteria

    Institute of Scientific and Technical Information of China (English)

    ZHANG Yao; JIAO Nianzhi

    2004-01-01

    Accurate quantification of aerobic anoxygenic phototrophic bacteria (AAPB) is of crucial importance for estimation of the role of AAPB in the carbon cycling in marine ecosystems. The normally used method "epifiuorescence microscope-infrared photography (EFM-IRP)"is, however, subject to positive errors introduced by mistaking cyanobacteria as AAPB due to the visibility of cyanobacteria under infrared photographic conditions for AAPB. This error could be up to 30% in the coast of the East China Sea. Such bias should be avoided by either subtracting cyanobacteira from the total infrared counts or using a fiowcytometer equipped with specific detectors for discrimination between cyanobacteria and AAPB.

  17. Protocol for Quantification of Defects in Natural Fibres for Composites

    DEFF Research Database (Denmark)

    Mortensen, Ulrich Andreas; Madsen, Bo

    2014-01-01

    Natural bast-type plant fibres are attracting increasing interest for being used for structural composite applications where high quality fibres with good mechanical properties are required. A protocol for the quantification of defects in natural fibres is presented. The protocol is based...... of defect size by width, and it is shown that both definitions can be used to give unbiased findings for the comparison between fibre types. Finally, considerations are given with respect to true measures of defect content, number of determinations, and number of significant figures used for the descriptive...

  18. Multi data reservior history matching and uncertainty quantification framework

    KAUST Repository

    Katterbauer, Klemens

    2015-11-26

    A multi-data reservoir history matching and uncertainty quantification framework is provided. The framework can utilize multiple data sets such as production, seismic, electromagnetic, gravimetric and surface deformation data for improving the history matching process. The framework can consist of a geological model that is interfaced with a reservoir simulator. The reservoir simulator can interface with seismic, electromagnetic, gravimetric and surface deformation modules to predict the corresponding observations. The observations can then be incorporated into a recursive filter that subsequently updates the model state and parameters distributions, providing a general framework to quantify and eventually reduce with the data, uncertainty in the estimated reservoir state and parameters.

  19. Stereometric Measurement System For Quantification Of Object Forms

    Science.gov (United States)

    Fischer, P.; Mesqui, F.; Kaeser, F.

    1986-07-01

    A three-dimensional camera unit has been developed at the Dental School of the University of Zurich for jaw motion recording. Since the system is designed to detect and reconstruct the spatial position of a light source, it can easily be used to digitize an object surface being scanned by a laser beam. This is performed by an additional device which has been developed in order to enlarge the application field of the camera unit as well as to prove the feasibility and find out the limits of such a simple and low-cost quantification system.

  20. Quantification in histopathology-Can magnetic particles help?

    International Nuclear Information System (INIS)

    Every year, more than 270,000 people are diagnosed with cancer in the UK alone; this means that one in three people worldwide contract cancer within their lifetime. Histopathology is the principle method for confirming cancer and directing treatment. In this paper, a novel application of magnetic particles is proposed to help address the problem of subjectivity in histopathology. Preliminary results indicate that magnetic nanoparticles cannot only be used to assist diagnosis through improving quantification but also potentially increase throughput, hence offering a way of dramatically reducing costs within the routine histopathology laboratory

  1. Preliminary Results on Uncertainty Quantification for Pattern Analytics

    Energy Technology Data Exchange (ETDEWEB)

    Stracuzzi, David John [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Brost, Randolph [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Chen, Maximillian Gene [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Malinas, Rebecca [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Peterson, Matthew Gregor [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Phillips, Cynthia A. [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Robinson, David G. [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States); Woodbridge, Diane [Sandia National Laboratories (SNL-NM), Albuquerque, NM (United States)

    2015-09-01

    This report summarizes preliminary research into uncertainty quantification for pattern ana- lytics within the context of the Pattern Analytics to Support High-Performance Exploitation and Reasoning (PANTHER) project. The primary focus of PANTHER was to make large quantities of remote sensing data searchable by analysts. The work described in this re- port adds nuance to both the initial data preparation steps and the search process. Search queries are transformed from does the specified pattern exist in the data? to how certain is the system that the returned results match the query? We show example results for both data processing and search, and discuss a number of possible improvements for each.

  2. Progressive damage state evolution and quantification in composites

    Science.gov (United States)

    Patra, Subir; Banerjee, Sourav

    2016-04-01

    Precursor damage state quantification can be helpful for safety and operation of aircraft and defense equipment's. Damage develops in the composite material in the form of matrix cracking, fiber breakages and deboning, etc. However, detection and quantification of the damage modes at their very early stage is not possible unless modifications of the existing indispensable techniques are conceived, particularly for the quantification of multiscale damages at their early stage. Here, we present a novel nonlocal mechanics based damage detection technique for precursor damage state quantification. Micro-continuum physics is used by modifying the Christoffel equation. American society of testing and materials (ASTM) standard woven carbon fiber (CFRP) specimens were tested under Tension-Tension fatigue loading at the interval of 25,000 cycles until 500,000 cycles. Scanning Acoustic Microcopy (SAM) and Optical Microscopy (OM) were used to examine the damage development at the same interval. Surface Acoustic Wave (SAW) velocity profile on a representative volume element (RVE) of the specimen were calculated at the regular interval of 50,000 cycles. Nonlocal parameters were calculated form the micromorphic wave dispersion curve at a particular frequency of 50 MHz. We used a previously formulated parameter called "Damage entropy" which is a measure of the damage growth in the material calculated with the loading cycle. Damage entropy (DE) was calculated at every pixel on the RVE and the mean of DE was plotted at the loading interval of 25,000 cycle. Growth of DE with fatigue loading cycles was observed. Optical Imaging also performed at the interval of 25,000 cycles to investigate the development of damage inside the materials. We also calculated the mean value of the Surface Acoustic Wave (SAW) velocity and plotted with fatigue cycle which is correlated further with Damage Entropy (DE). Statistical analysis of the Surface Acoustic Wave profile (SAW) obtained at different

  3. A recipe for EFT uncertainty quantification in nuclear physics

    International Nuclear Information System (INIS)

    The application of effective field theory (EFT) methods to nuclear systems provides the opportunity to rigorously estimate the uncertainties originating in the nuclear Hamiltonian. Yet this is just one source of uncertainty in the observables predicted by calculations based on nuclear EFTs. We discuss the goals of uncertainty quantification in such calculations and outline a recipe to obtain statistically meaningful error bars for their predictions. We argue that the different sources of theory error can be accounted for within a Bayesian framework, as we illustrate using a toy model. (paper)

  4. Predicting human age with bloodstains by sjTREC quantification.

    Directory of Open Access Journals (Sweden)

    Xue-ling Ou

    Full Text Available The age-related decline of signal joint T-cell receptor rearrangement excision circles (sjTRECs in human peripheral blood has been demonstrated in our previous study and other reports. Until now, only a few studies on sjTREC detection in bloodstain samples were reported, which were based on a small sample of subjects of a limited age range, although bloodstains are much more frequently encountered in forensic practice. In this present study, we adopted the sensitive Taqman real-time quantitative polymerase chain reaction (qPCR method to perform sjTREC quantification in bloodstains from individuals ranging from 0-86 years old (n = 264. The results revealed that sjTREC contents in human bloodstains were declined in an age-dependent manner (r = -0.8712. The formula of age estimation was Age = -7.1815Y-42.458 ± 9.42 (Y dCt(TBP-sjTREC; 9.42 standard error. Furthermore, we tested for the influence of short- or long- storage time by analyzing fresh and stored bloodstains from the same individuals. Remarkably, no statistically significant difference in sjTREC contents was found between the fresh and old DNA samples over a 4-week of storage time. However, significant loss (0.16-1.93 dCt in sjTREC contents was detected after 1.5 years of storage in 31 samples. Moreover, preliminary sjTREC quantification from up to 20-year-old bloodstains showed that though the sjTREC contents were detectable in all samples and highly correlated with donor age, a time-dependent decrease in the correlation coefficient r was found, suggesting the predicting accuracy of this described assay would be deteriorated in aged samples. Our findings show that sjTREC quantification might be also suitable for age prediction in bloodstains, and future researches into the time-dependent or other potential impacts on sjTREC quantification might allow further improvement of the predicting accuracy.

  5. Recurrence plots and recurrence quantification analysis of human motion data

    Science.gov (United States)

    Josiński, Henryk; Michalczuk, Agnieszka; Świtoński, Adam; Szczesna, Agnieszka; Wojciechowski, Konrad

    2016-06-01

    The authors present exemplary application of recurrence plots, cross recurrence plots and recurrence quantification analysis for the purpose of exploration of experimental time series describing selected aspects of human motion. Time series were extracted from treadmill gait sequences which were recorded in the Human Motion Laboratory (HML) of the Polish-Japanese Academy of Information Technology in Bytom, Poland by means of the Vicon system. Analysis was focused on the time series representing movements of hip, knee, ankle and wrist joints in the sagittal plane.

  6. Quantification of Lung Metastases from In Vivo Mouse Models

    DEFF Research Database (Denmark)

    Chang, Joan; Erler, Janine T

    2016-01-01

    Cancer research has made significant progress in terms of understanding and targeting primary tumors; however, the challenge remains for the successful treatment of metastatic cancers. This highlights the importance to use in vivo models to study the metastatic process, as well as for preclinical...... testing of compounds that could inhibit metastasis. As a result, proper quantification of metastases from in vivo models is of the utmost significance. Here, we provide a detailed protocol for collecting and handling lung tissues from mice, and guidance for subsequent analysis of metastases, as well...

  7. An EPGPT-based approach for uncertainty quantification

    Energy Technology Data Exchange (ETDEWEB)

    Wang, C.; Abdel-Khalik, H. S. [Dept. of Nuclear Engineering, North Caroline State Univ., Raleigh, NC 27695 (United States)

    2012-07-01

    Generalized Perturbation Theory (GPT) has been widely used by many scientific disciplines to perform sensitivity analysis and uncertainty quantification. This manuscript employs recent developments in GPT theory, collectively referred to as Exact-to-Precision Generalized Perturbation Theory (EPGPT), to enable uncertainty quantification for computationally challenging models, e.g. nonlinear models associated with many input parameters and many output responses and with general non-Gaussian parameters distributions. The core difference between EPGPT and existing GPT is in the way the problem is formulated. GPT formulates an adjoint problem that is dependent on the response of interest. It tries to capture via the adjoint solution the relationship between the response of interest and the constraints on the state variations. EPGPT recasts the problem in terms of a smaller set of what is referred to as the 'active' responses which are solely dependent on the physics model and the boundary and initial conditions rather than on the responses of interest. The objective of this work is to apply an EPGPT methodology to propagate cross-sections variations in typical reactor design calculations. The goal is to illustrate its use and the associated impact for situations where the typical Gaussian assumption for parameters uncertainties is not valid and when nonlinear behavior must be considered. To allow this demonstration, exaggerated variations will be employed to stimulate nonlinear behavior in simple prototypical neutronics models. (authors)

  8. HPTLC densitometric quantification of stigmasterol and lupeol from Ficus religiosa

    Directory of Open Access Journals (Sweden)

    Deepti Rathee

    2015-05-01

    Full Text Available This study presents the first report of TLC densitometric method, which has been developed and validated for simultaneous quantification of the two marker compounds (stigmasterol and lupeol from methanolic extract using the solvent system of toluene:methanol (9:1, v/v. The method employed TLC aluminum plates precoated with silica gel 60 F254 as the stationary phase. Densitometric analysis of stigmasterol and lupeol was carried out in the reflectance mode at 525 nm. The system was found to give compact spots for stigmasterol and lupeol (Rf value of 0.37 and 0.60, respectively. The method was validated using ICH guidelines in terms of precision, repeatability and accuracy. Linearity range for stigmasterol and lupeol was 80–480 ng/spot and 150–900 ng/spot and the contents were found to be 0.06 ± 0.005% w/w and 0.12 ± 0.02% w/w, respectively. The limit of detection (LOD value for stigmasterol and lupeol were found to be 20 and 50 ng, and limit of quantification (LOQ value were 60 and 100 ng, respectively. This simple, precise and accurate method gave good resolution from other constituents present in the extract. The method has been successfully applied in the analysis and routine quality control of herbal material and formulations containing Ficus religiosa.

  9. Guided Wave Delamination Detection and Quantification With Wavefield Data Analysis

    Science.gov (United States)

    Tian, Zhenhua; Campbell Leckey, Cara A.; Seebo, Jeffrey P.; Yu, Lingyu

    2014-01-01

    Unexpected damage can occur in aerospace composites due to impact events or material stress during off-nominal loading events. In particular, laminated composites are susceptible to delamination damage due to weak transverse tensile and inter-laminar shear strengths. Developments of reliable and quantitative techniques to detect delamination damage in laminated composites are imperative for safe and functional optimally-designed next-generation composite structures. In this paper, we investigate guided wave interactions with delamination damage and develop quantification algorithms by using wavefield data analysis. The trapped guided waves in the delamination region are observed from the wavefield data and further quantitatively interpreted by using different wavenumber analysis methods. The frequency-wavenumber representation of the wavefield shows that new wavenumbers are present and correlate to trapped waves in the damage region. These new wavenumbers are used to detect and quantify the delamination damage through the wavenumber analysis, which can show how the wavenumber changes as a function of wave propagation distance. The location and spatial duration of the new wavenumbers can be identified, providing a useful means not only for detecting the presence of delamination damage but also allowing for estimation of the delamination size. Our method has been applied to detect and quantify real delamination damage with complex geometry (grown using a quasi-static indentation technique). The detection and quantification results show the location, size, and shape of the delamination damage.

  10. An uncertainty inventory demonstration - a primary step in uncertainty quantification

    Energy Technology Data Exchange (ETDEWEB)

    Langenbrunner, James R. [Los Alamos National Laboratory; Booker, Jane M [Los Alamos National Laboratory; Hemez, Francois M [Los Alamos National Laboratory; Salazar, Issac F [Los Alamos National Laboratory; Ross, Timothy J [UNM

    2009-01-01

    Tools, methods, and theories for assessing and quantifying uncertainties vary by application. Uncertainty quantification tasks have unique desiderata and circumstances. To realistically assess uncertainty requires the engineer/scientist to specify mathematical models, the physical phenomena of interest, and the theory or framework for assessments. For example, Probabilistic Risk Assessment (PRA) specifically identifies uncertainties using probability theory, and therefore, PRA's lack formal procedures for quantifying uncertainties that are not probabilistic. The Phenomena Identification and Ranking Technique (PIRT) proceeds by ranking phenomena using scoring criteria that results in linguistic descriptors, such as importance ranked with words, 'High/Medium/Low.' The use of words allows PIRT to be flexible, but the analysis may then be difficult to combine with other uncertainty theories. We propose that a necessary step for the development of a procedure or protocol for uncertainty quantification (UQ) is the application of an Uncertainty Inventory. An Uncertainty Inventory should be considered and performed in the earliest stages of UQ.

  11. Amperometric quantification based on serial dilution microfluidic systems.

    Science.gov (United States)

    Stephan, Khaled; Pittet, Patrick; Sigaud, Monique; Renaud, Louis; Vittori, Olivier; Morin, Pierre; Ouaini, Naim; Ferrigno, Rosaria

    2009-03-01

    This paper describes a microfluidic device fabricated in poly(dimethylsiloxane) that was employed to perform amperometric quantifications using on-chip calibration curves and on-chip standard addition methods. This device integrated a network of Au electrodes within a microfluidic structure designed for automatic preparation of a series of solutions containing an electroactive molecule at a concentration linearly decreasing. This device was first characterized by fluorescence microscopy and then evaluated with a model electroactive molecule such as Fe(CN(6))(4-). Operating a quantification in this microfluidic parallel approach rather than in batch mode allows a reduced analysis time to be achieved. Moreover, the microfluidic approach is compatible with the on-chip calibration of sensors simultaneously to the analysis, therefore preventing problems due to sensor response deviation with time. When using the on-chip calibration and on-chip standard addition method, we reached concentration estimation better than 5%. We also demonstrated that compared to the calibration curve approach, the standard addition mode is less complex to operate. Indeed, in this case, it is not necessary to take into account flow rate discrepancies as in the calibration approach. PMID:19238282

  12. Ultrasound strain imaging for quantification of tissue function: cardiovascular applications

    Science.gov (United States)

    de Korte, Chris L.; Lopata, Richard G. P.; Hansen, Hendrik H. G.

    2013-03-01

    With ultrasound imaging, the motion and deformation of tissue can be measured. Tissue can be deformed by applying a force on it and the resulting deformation is a function of its mechanical properties. Quantification of this resulting tissue deformation to assess the mechanical properties of tissue is called elastography. If the tissue under interrogation is actively deforming, the deformation is directly related to its function and quantification of this deformation is normally referred as `strain imaging'. Elastography can be used for atherosclerotic plaques characterization, while the contractility of the heart or skeletal muscles can be assessed with strain imaging. We developed radio frequency (RF) based ultrasound methods to assess the deformation at higher resolution and with higher accuracy than commercial methods using conventional image data (Tissue Doppler Imaging and 2D speckle tracking methods). However, the improvement in accuracy is mainly achieved when measuring strain along the ultrasound beam direction, so 1D. We further extended this method to multiple directions and further improved precision by using compounding of data acquired at multiple beam steered angles. In arteries, the presence of vulnerable plaques may lead to acute events like stroke and myocardial infarction. Consequently, timely detection of these plaques is of great diagnostic value. Non-invasive ultrasound strain compounding is currently being evaluated as a diagnostic tool to identify the vulnerability of plaques. In the heart, we determined the strain locally and at high resolution resulting in a local assessment in contrary to conventional global functional parameters like cardiac output or shortening fraction.

  13. Volumetric loss quantification using ultrasonic inductively coupled transducers

    Science.gov (United States)

    Gong, Peng; Hay, Thomas R.; Greve, David W.; Oppenheim, Irving J.

    2015-03-01

    The pulse-echo method is widely used for plate and pipe thickness measurement. However, the pulse echo method does not work well for detecting localized volumetric loss in thick-wall tubes, as created by erosion damage, when the morphology of volumetric loss is irregular and can reflect ultrasonic pulses away from the transducer, making it difficult to detect an echo. In this paper, we propose a novel method using an inductively coupled transducer to generate longitudinal waves propagating in a thick-wall aluminum tube for the volumetric loss quantification. In the experiment, longitudinal waves exhibit diffraction effects during the propagation which can be explained by the Huygens-Fresnel principle. The diffractive waves are also shown to be significantly delayed by the machined volumetric loss on the inside surface of the thick-wall aluminum tube. It is also shown that the inductively coupled transducers can generate and receive similar ultrasonic waves to those from wired transducers, and the inductively coupled transducers perform as well as the wired transducers in the volumetric loss quantification when other conditions are the same.

  14. Generation, Quantification, and Tracing of Metabolically Labeled Fluorescent Exosomes.

    Science.gov (United States)

    Coscia, Carolina; Parolini, Isabella; Sanchez, Massimo; Biffoni, Mauro; Boussadia, Zaira; Zanetti, Cristiana; Fiani, Maria Luisa; Sargiacomo, Massimo

    2016-01-01

    Over the last 10 years, the constant progression in exosome (Exo)-related studies highlighted the importance of these cell-derived nano-sized vesicles in cell biology and pathophysiology. Functional studies on Exo uptake and intracellular trafficking require accurate quantification to assess sufficient and/or necessary Exo particles quantum able to elicit measurable effects on target cells. We used commercially available BODIPY(®) fatty acid analogues to label a primary melanoma cell line (Me501) that highly and spontaneously secrete nanovesicles. Upon addition to cell culture, BODIPY fatty acids are rapidly incorporated into major phospholipid classes ultimately producing fluorescent Exo as direct result of biogenesis. Our metabolic labeling protocol produced bright fluorescent Exo that can be examined and quantified with conventional non-customized flow cytometry (FC) instruments by exploiting their fluorescent emission rather than light-scattering detection. Furthermore, our methodology permits the measurement of single Exo-associated fluorescence transfer to cells making quantitative the correlation between Exo uptake and activation of cellular processes. Thus the protocol presented here appears as an appropriate tool to who wants to investigate mechanisms of Exo functions in that it allows for direct and rapid characterization and quantification of fluorescent Exo number, intensity, size, and eventually evaluation of their kinetic of uptake/secretion in target cells. PMID:27317184

  15. Damage Detection and Quantification Using Transmissibility Coherence Analysis

    Directory of Open Access Journals (Sweden)

    Yun-Lai Zhou

    2015-01-01

    Full Text Available A new transmissibility-based damage detection and quantification approach is proposed. Based on the operational modal analysis, the transmissibility is extracted from system responses and transmissibility coherence is defined and analyzed. Afterwards, a sensitive-damage indicator is defined in order to detect and identify the severity of damage and compared with an indicator developed by other authors. The proposed approach is validated on data from a physics-based numerical model as well as experimental data from a three-story aluminum frame structure. For both numerical simulation and experiment the results of the new indicator reveal a better performance than coherence measure proposed in Rizos et al., 2008, Rizos et al., 2002, Fassois and Sakellariou, 2007, especially when nonlinearity occurs, which might be further used in real engineering. The main contribution of this study is the construction of the relation between transmissibility coherence and frequency response function coherence and the construction of an effective indicator based on the transmissibility modal assurance criteria for damage (especially for minor nonlinearity detection as well as quantification.

  16. Unilateral condylar hyperplasia: a 3-dimensional quantification of asymmetry.

    Directory of Open Access Journals (Sweden)

    Tim J Verhoeven

    Full Text Available PURPOSE: Objective quantifications of facial asymmetry in patients with Unilateral Condylar Hyperplasia (UCH have not yet been described in literature. The aim of this study was to objectively quantify soft-tissue asymmetry in patients with UCH and to compare the findings with a control group using a new method. MATERIAL AND METHODS: Thirty 3D photographs of patients diagnosed with UCH were compared with 30 3D photographs of healthy controls. As UCH presents particularly in the mandible, a new method was used to isolate the lower part of the face to evaluate asymmetry of this part separately. The new method was validated by two observers using 3D photographs of five patients and five controls. RESULTS: A significant difference (0.79 mm between patients and controls whole face asymmetry was found. Intra- and inter-observer differences of 0.011 mm (-0.034-0.011 and 0.017 mm (-0.007-0.042 respectively were found. These differences are irrelevant in clinical practice. CONCLUSION: After objective quantification, a significant difference was identified in soft-tissue asymmetry between patients with UCH and controls. The method used to isolate mandibular asymmetry was found to be valid and a suitable tool to evaluate facial asymmetry.

  17. A Spanish model for quantification and management of construction waste

    International Nuclear Information System (INIS)

    Currently, construction and demolition waste (C and D waste) is a worldwide issue that concerns not only governments but also the building actors involved in construction activity. In Spain, a new national decree has been regulating the production and management of C and D waste since February 2008. The present work describes the waste management model that has inspired this decree: the Alcores model implemented with good results in Los Alcores Community (Seville, Spain). A detailed model is also provided to estimate the volume of waste that is expected to be generated on the building site. The quantification of C and D waste volume, from the project stage, is essential for the building actors to properly plan and control its disposal. This quantification model has been developed by studying 100 dwelling projects, especially their bill of quantities, and defining three coefficients to estimate the demolished volume (CT), the wreckage volume (CR) and the packaging volume (CE). Finally, two case studies are included to illustrate the usefulness of the model to estimate C and D waste volume in both new construction and demolition projects.

  18. Mixture quantification using PLS in plastic scintillation measurements

    Energy Technology Data Exchange (ETDEWEB)

    Bagan, H.; Tarancon, A.; Rauret, G. [Departament de Quimica Analitica, Universitat de Barcelona, Diagonal 647, E-08028 Barcelona (Spain); Garcia, J.F., E-mail: jfgarcia@ub.ed [Departament de Quimica Analitica, Universitat de Barcelona, Diagonal 647, E-08028 Barcelona (Spain)

    2011-06-15

    This article reports the capability of plastic scintillation (PS) combined with multivariate calibration (Partial least squares; PLS) to detect and quantify alpha and beta emitters in mixtures. While several attempts have been made with this purpose in mind using liquid scintillation (LS), no attempt was done using PS that has the great advantage of not producing mixed waste after the measurements are performed. Following this objective, ternary mixtures of alpha and beta emitters ({sup 241}Am, {sup 137}Cs and {sup 90}Sr/{sup 90}Y) have been quantified. Procedure optimisation has evaluated the use of the net spectra or the sample spectra, the inclusion of different spectra obtained at different values of the Pulse Shape Analysis parameter and the application of the PLS1 or PLS2 algorithms. The conclusions show that the use of PS+PLS2 applied to the sample spectra, without the use of any pulse shape discrimination, allows quantification of the activities with relative errors less than 10% in most of the cases. This procedure not only allows quantification of mixtures but also reduces measurement time (no blanks are required) and the application of this procedure does not require detectors that include the pulse shape analysis parameter.

  19. A critical view on microplastic quantification in aquatic organisms.

    Science.gov (United States)

    Vandermeersch, Griet; Van Cauwenberghe, Lisbeth; Janssen, Colin R; Marques, Antonio; Granby, Kit; Fait, Gabriella; Kotterman, Michiel J J; Diogène, Jorge; Bekaert, Karen; Robbens, Johan; Devriese, Lisa

    2015-11-01

    Microplastics, plastic particles and fragments smaller than 5mm, are ubiquitous in the marine environment. Ingestion and accumulation of microplastics have previously been demonstrated for diverse marine species ranging from zooplankton to bivalves and fish, implying the potential for microplastics to accumulate in the marine food web. In this way, microplastics can potentially impact food safety and human health. Although a few methods to quantify microplastics in biota have been described, no comparison and/or intercalibration of these techniques have been performed. Here we conducted a literature review on all available extraction and quantification methods. Two of these methods, involving wet acid destruction, were used to evaluate the presence of microplastics in field-collected mussels (Mytilus galloprovincialis) from three different "hotspot" locations in Europe (Po estuary, Italy; Tagus estuary, Portugal; Ebro estuary, Spain). An average of 0.18±0.14 total microplastics g(-1) w.w. for the Acid mix Method and 0.12±0.04 total microplastics g(-1) w.w. for the Nitric acid Method was established. Additionally, in a pilot study an average load of 0.13±0.14 total microplastics g(-1) w.w. was recorded in commercial mussels (Mytilus edulis and M. galloprovincialis) from five European countries (France, Italy, Denmark, Spain and The Netherlands). A detailed analysis and comparison of methods indicated the need for further research to develop a standardised operating protocol for microplastic quantification and monitoring. PMID:26249746

  20. An on-bacterium flow cytometric immunoassay for protein quantification.

    Science.gov (United States)

    Lan, Wen-Jun; Lan, Wei; Wang, Hai-Yan; Yan, Lei; Wang, Zhe-Li

    2013-09-01

    The polystyrene bead-based flow cytometric immunoassay has been widely reported. However, the preparation of functional polystyrene bead is still inconvenient. This study describes a simple and easy on-bacterium flow cytometric immunoassay for protein quantification, in which Staphylococcus aureus (SAC) is used as an antibody-antigen carrier to replace the polystyrene bead. The SAC beads were prepared by carboxyfluorescein diacetate succinimidyl ester (CFSE) labeling, paraformaldehyde fixation and antibody binding. Carcinoembryonic antigen (CEA) and cytokeratin-19 fragment (CYFRA 21-1) proteins were used as models in the test system. Using prepared SAC beads, biotinylated proteins, and streptavidin-phycoerythrin (SA-PE), the on-bacterium flow cytometric immunoassay was validated by quantifying CEA and CYFRA 21-1 in sample. Obtained data demonstrated a concordant result between the logarithm of the protein concentration and the logarithm of the PE mean fluorescence intensity (MFI). The limit of detection (LOD) in this immunoassay was at least 0.25 ng/ml. Precision and accuracy assessments appeared that either the relative standard deviation (R.S.D.) or the relative error (R.E.) was CYFRA 21-1. In conclusion, the on-bacterium flow cytometric immunoassay may be of use in the quantification of serum protein. PMID:23739299

  1. Simple and inexpensive quantification of ammonia in whole blood.

    Science.gov (United States)

    Ayyub, Omar B; Behrens, Adam M; Heligman, Brian T; Natoli, Mary E; Ayoub, Joseph J; Cunningham, Gary; Summar, Marshall; Kofinas, Peter

    2015-01-01

    Quantification of ammonia in whole blood has applications in the diagnosis and management of many hepatic diseases, including cirrhosis and rare urea cycle disorders, amounting to more than 5 million patients in the United States. Current techniques for ammonia measurement suffer from limited range, poor resolution, false positives or large, complex sensor set-ups. Here we demonstrate a technique utilizing inexpensive reagents and simple methods for quantifying ammonia in 100 μL of whole blood. The sensor comprises a modified form of the indophenol reaction, which resists sources of destructive interference in blood, in conjunction with a cation-exchange membrane. The presented sensing scheme is selective against other amine containing molecules such as amino acids and has a shelf life of at least 50 days. Additionally, the resulting system has high sensitivity and allows for the accurate reliable quantification of ammonia in whole human blood samples at a minimum range of 25 to 500 μM, which is clinically for rare hyperammonemic disorders and liver disease. Furthermore, concentrations of 50 and 100 μM ammonia could be reliably discerned with p = 0.0001. PMID:25936660

  2. An EPGPT-based approach for uncertainty quantification

    International Nuclear Information System (INIS)

    Generalized Perturbation Theory (GPT) has been widely used by many scientific disciplines to perform sensitivity analysis and uncertainty quantification. This manuscript employs recent developments in GPT theory, collectively referred to as Exact-to-Precision Generalized Perturbation Theory (EPGPT), to enable uncertainty quantification for computationally challenging models, e.g. nonlinear models associated with many input parameters and many output responses and with general non-Gaussian parameters distributions. The core difference between EPGPT and existing GPT is in the way the problem is formulated. GPT formulates an adjoint problem that is dependent on the response of interest. It tries to capture via the adjoint solution the relationship between the response of interest and the constraints on the state variations. EPGPT recasts the problem in terms of a smaller set of what is referred to as the 'active' responses which are solely dependent on the physics model and the boundary and initial conditions rather than on the responses of interest. The objective of this work is to apply an EPGPT methodology to propagate cross-sections variations in typical reactor design calculations. The goal is to illustrate its use and the associated impact for situations where the typical Gaussian assumption for parameters uncertainties is not valid and when nonlinear behavior must be considered. To allow this demonstration, exaggerated variations will be employed to stimulate nonlinear behavior in simple prototypical neutronics models. (authors)

  3. Comparison of techniques for quantification of next-generation sequencing libraries

    DEFF Research Database (Denmark)

    Hussing, Christian; Kampmann, Marie-Louise; Mogensen, Helle Smidt;

    2015-01-01

    To ensure efficient sequencing, the DNA of next-generation sequencing (NGS) libraries must be quantified correctly. Therefore, an accurate, sensitive and stable method for DNA quantification is crucial. In this study, seven different methods for DNA quantification were compared to each other...

  4. New Approach for Segmentation and Quantification of Two-Dimensional Gel Electrophoresis Images

    DEFF Research Database (Denmark)

    Anjo, Antonio dos; Laurell Blom Møller, Anders; Ersbøll, Bjarne Kjær;

    2011-01-01

    of background due to poor segmentation. Other software applications use a fixed window for this task, resulting in omission of part of the protein spot, or including background in the quantification. The approach presented here for the segmentation and quantification of 2-DE aims to minimize these problems...

  5. PCR amplification of repetitive sequences as a possible approach in relative species quantification

    DEFF Research Database (Denmark)

    Ballin, Nicolai Zederkopff; Vogensen, Finn Kvist; Karlsson, Anders H

    2012-01-01

    Abstract Both relative and absolute quantifications are possible in species quantification when single copy genomic DNA is used. However, amplification of single copy genomic DNA does not allow a limit of detection as low as one obtained from amplification of repetitive sequences. Amplification...... of repetitive sequences is therefore frequently used in absolute quantification but problems occur in relative quantification as the number of repetitive sequences is unknown. A promising approach was developed where data from amplification of repetitive sequences were used in relative quantification of species...... in binary mixtures. PCR LUX primers were designed that amplify repetitive and single copy sequences to establish the species dependent number (constants) (SDC) of amplified repetitive sequences per genome. The SDCs and data from amplification of repetitive sequences were tested for their applicability...

  6. Critical points of DNA quantification by real-time PCR – effects of DNA extraction method and sample matrix on quantification of genetically modified organisms

    Directory of Open Access Journals (Sweden)

    Žel Jana

    2006-08-01

    Full Text Available Abstract Background Real-time PCR is the technique of choice for nucleic acid quantification. In the field of detection of genetically modified organisms (GMOs quantification of biotech products may be required to fulfil legislative requirements. However, successful quantification depends crucially on the quality of the sample DNA analyzed. Methods for GMO detection are generally validated on certified reference materials that are in the form of powdered grain material, while detection in routine laboratories must be performed on a wide variety of sample matrixes. Due to food processing, the DNA in sample matrixes can be present in low amounts and also degraded. In addition, molecules of plant origin or from other sources that affect PCR amplification of samples will influence the reliability of the quantification. Further, the wide variety of sample matrixes presents a challenge for detection laboratories. The extraction method must ensure high yield and quality of the DNA obtained and must be carefully selected, since even components of DNA extraction solutions can influence PCR reactions. GMO quantification is based on a standard curve, therefore similarity of PCR efficiency for the sample and standard reference material is a prerequisite for exact quantification. Little information on the performance of real-time PCR on samples of different matrixes is available. Results Five commonly used DNA extraction techniques were compared and their suitability for quantitative analysis was assessed. The effect of sample matrix on nucleic acid quantification was assessed by comparing 4 maize and 4 soybean matrixes. In addition 205 maize and soybean samples from routine analysis were analyzed for PCR efficiency to assess variability of PCR performance within each sample matrix. Together with the amount of DNA needed for reliable quantification, PCR efficiency is the crucial parameter determining the reliability of quantitative results, therefore it was

  7. Quantification of tidal parameters from Solar system data

    CERN Document Server

    Lainey, Valéry

    2016-01-01

    Tidal dissipation is the main driver of orbital evolution of natural satellites and a key point to understand the exoplanetary system configurations. Despite its importance, its quantification from observations still remains difficult for most objects of our own Solar system. In this work, we overview the method that has been used to determine, directly from observations, the tidal parameters, with emphasis on the Love number k2 and the tidal quality factor Q. Up-to-date values of these tidal parameters are summarized. Last, an assessment on the possible determination of the tidal ratio k2/Q of Uranus and Neptune is done. This may be particularly relevant for coming astrometric campaigns and future space missions focused on these systems.

  8. Dielectrophoretic immobilization of proteins: Quantification by atomic force microscopy.

    Science.gov (United States)

    Laux, Eva-Maria; Knigge, Xenia; Bier, Frank F; Wenger, Christian; Hölzel, Ralph

    2015-09-01

    The combination of alternating electric fields with nanometer-sized electrodes allows the permanent immobilization of proteins by dielectrophoretic force. Here, atomic force microscopy is introduced as a quantification method, and results are compared with fluorescence microscopy. Experimental parameters, for example the applied voltage and duration of field application, are varied systematically, and the influence on the amount of immobilized proteins is investigated. A linear correlation to the duration of field application was found by atomic force microscopy, and both microscopical methods yield a square dependence of the amount of immobilized proteins on the applied voltage. While fluorescence microscopy allows real-time imaging, atomic force microscopy reveals immobilized proteins obscured in fluorescence images due to low S/N. Furthermore, the higher spatial resolution of the atomic force microscope enables the visualization of the protein distribution on single nanoelectrodes. The electric field distribution is calculated and compared to experimental results with very good agreement to atomic force microscopy measurements. PMID:26010162

  9. Automated image analysis for quantification of filamentous bacteria

    DEFF Research Database (Denmark)

    Fredborg, Marlene; Rosenvinge, Flemming Schønning; Spillum, Erik;

    2015-01-01

    Background Antibiotics of the β-lactam group are able to alter the shape of the bacterial cell wall, e.g. filamentation or a spheroplast formation. Early determination of antimicrobial susceptibility may be complicated by filamentation of bacteria as this can be falsely interpreted as growth...... in systems relying on colorimetry or turbidometry (such as Vitek-2, Phoenix, MicroScan WalkAway). The objective was to examine an automated image analysis algorithm for quantification of filamentous bacteria using the 3D digital microscopy imaging system, oCelloScope. Results Three E. coli strains displaying...... different resistant profiles and differences in filamentation kinetics were used to study a novel image analysis algorithm to quantify length of bacteria and bacterial filamentation. A total of 12 β-lactam antibiotics or β-lactam–β-lactamase inhibitor combinations were analyzed for their ability to induce...

  10. An approximation approach for uncertainty quantification using evidence theory

    Energy Technology Data Exchange (ETDEWEB)

    Bae, Ha-Rok; Grandhi, Ramana V.; Canfield, Robert A

    2004-12-01

    Over the last two decades, uncertainty quantification (UQ) in engineering systems has been performed by the popular framework of probability theory. However, many scientific and engineering communities realize that there are limitations in using only one framework for quantifying the uncertainty experienced in engineering applications. Recently evidence theory, also called Dempster-Shafer theory, was proposed to handle limited and imprecise data situations as an alternative to the classical probability theory. Adaptation of this theory for large-scale engineering structures is a challenge due to implicit nature of simulations and excessive computational costs. In this work, an approximation approach is developed to improve the practical utility of evidence theory in UQ analysis. The techniques are demonstrated on composite material structures and airframe wing aeroelastic design problem.

  11. Reliability and discriminatory power of methods for dental plaque quantification

    Directory of Open Access Journals (Sweden)

    Daniela Prócida Raggio

    2010-04-01

    Full Text Available OBJECTIVE: This in situ study evaluated the discriminatory power and reliability of methods of dental plaque quantification and the relationship between visual indices (VI and fluorescence camera (FC to detect plaque. MATERIAL AND METHODS: Six volunteers used palatal appliances with six bovine enamel blocks presenting different stages of plaque accumulation. The presence of plaque with and without disclosing was assessed using VI. Images were obtained with FC and digital camera in both conditions. The area covered by plaque was assessed. Examinations were done by two independent examiners. Data were analyzed by Kruskal-Wallis and Kappa tests to compare different conditions of samples and to assess the inter-examiner reproducibility. RESULTS: Some methods presented adequate reproducibility. The Turesky index and the assessment of area covered by disclosed plaque in the FC images presented the highest discriminatory powers. CONCLUSION: The Turesky index and images with FC with disclosing present good reliability and discriminatory power in quantifying dental plaque.

  12. Capacitive immunosensor for C-reactive protein quantification

    KAUST Repository

    Sapsanis, Christos

    2015-08-02

    We report an agglutination-based immunosensor for the quantification of C-reactive protein (CRP). The developed immunoassay sensor requires approximately 15 minutes of assay time per sample and provides a sensitivity of 0.5 mg/L. We have measured the capacitance of interdigitated electrodes (IDEs) and quantified the concentration of added analyte. The proposed method is a label free detection method and hence provides rapid measurement preferable in diagnostics. We have so far been able to quantify the concentration to as low as 0.5 mg/L and as high as 10 mg/L. By quantifying CRP in serum, we can assess whether patients are prone to cardiac diseases and monitor the risk associated with such diseases. The sensor is a simple low cost structure and it can be a promising device for rapid and sensitive detection of disease markers at the point-of-care stage.

  13. Mitral and aortic valvular flow: quantification with MR phase mapping

    DEFF Research Database (Denmark)

    Søndergaard, Lise; Thomsen, C; Ståhlberg, F;

    1992-01-01

    of the flow-encoding gradient to 2.2 msec. Tested on a flow phantom simulating a severe valvular stenosis, the sequence was found to be robust for higher-order motion within the clinical velocity range. In eight healthy volunteers, mitral and aortic volume flow rates and peak velocities were quantified...... mapping is also valid for pathologic conditions. Phase mapping may be used as a noninvasive clinical tool for flow quantification in heart valve disease.......When magnetic resonance phase mapping is used to quantitate valvular blood flow, the presence of higher-order-motion terms may cause a loss of phase information. To overcome this problem, a sequence with reduced encoding for higher-order motion was used, achieved by decreasing the duration...

  14. Quantification of Human Movement for Assessment in Automated Exercise Coaching

    CERN Document Server

    Hagler, Stuart; Bajczy, Ruzena; Pavel, Misha

    2016-01-01

    Quantification of human movement is a challenge in many areas, ranging from physical therapy to robotics. We quantify of human movement for the purpose of providing automated exercise coaching in the home. We developed a model-based assessment and inference process that combines biomechanical constraints with movement assessment based on the Microsoft Kinect camera. To illustrate the approach, we quantify the performance of a simple squatting exercise using two model-based metrics that are related to strength and endurance, and provide an estimate of the strength and energy-expenditure of each exercise session. We look at data for 5 subjects, and show that for some subjects the metrics indicate a trend consistent with improved exercise performance.

  15. Automated quantification of one-dimensional nanostructure alignment on surfaces

    CERN Document Server

    Dong, Jianjin; Abukhdeir, Nasser Mohieddin

    2016-01-01

    A method for automated quantification of the alignment of one-dimensional nanostructures from microscopy imaging is presented. Nanostructure alignment metrics are formulated and shown to able to rigorously quantify the orientational order of nanostructures within a two-dimensional domain (surface). A complementary image processing method is also presented which enables robust processing of microscopy images where overlapping nanostructures might be present. Scanning electron microscopy (SEM) images of nanowire-covered surfaces are analyzed using the presented methods and it is shown that past single parameter alignment metrics are insufficient for highly aligned domains. Through the use of multiple parameter alignment metrics, automated quantitative analysis of SEM images is shown to be possible and the alignment characteristics of different samples are able to be rigorously compared using a similarity metric. The results of this work provide researchers in nanoscience and nanotechnology with a rigorous metho...

  16. Experimental investigations for uncertainty quantification in brake squeal analysis

    Science.gov (United States)

    Renault, A.; Massa, F.; Lallemand, B.; Tison, T.

    2016-04-01

    The aim of this paper is to improve the correlation between the experimental and the numerical prediction of unstable frequencies for automotive brake systems considering uncertainty. First, an experimental quantification of uncertainty and a discussion analysing the contributions of uncertainty to a numerical squeal simulation are proposed. Frequency and transient simulations are performed considering nominal values of model parameters, determined experimentally. The obtained results are compared with those derived from experimental tests to highlight the limitation of deterministic simulations. The effects of the different kinds of uncertainty detected in working conditions of brake system, the pad boundary condition, the brake system material properties and the pad surface topography are discussed by defining different unstable mode classes. Finally, a correlation between experimental and numerical results considering uncertainty is successfully proposed for an industrial brake system. Results from the different comparisons reveal also a major influence of the pad topography and consequently the contact distribution.

  17. Uncertainty quantification for CO2 sequestration and enhanced oil recovery

    CERN Document Server

    Dai, Zhenxue; Fessenden-Rahn, Julianna; Middleton, Richard; Pan, Feng; Jia, Wei; Lee, Si-Yong; McPherson, Brian; Ampomah, William; Grigg, Reid

    2014-01-01

    This study develops a statistical method to perform uncertainty quantification for understanding CO2 storage potential within an enhanced oil recovery (EOR) environment at the Farnsworth Unit of the Anadarko Basin in northern Texas. A set of geostatistical-based Monte Carlo simulations of CO2-oil-water flow and reactive transport in the Morrow formation are conducted for global sensitivity and statistical analysis of the major uncertainty metrics: net CO2 injection, cumulative oil production, cumulative gas (CH4) production, and net water injection. A global sensitivity and response surface analysis indicates that reservoir permeability, porosity, and thickness are the major intrinsic reservoir parameters that control net CO2 injection/storage and oil/gas recovery rates. The well spacing and the initial water saturation also have large impact on the oil/gas recovery rates. Further, this study has revealed key insights into the potential behavior and the operational parameters of CO2 sequestration at CO2-EOR s...

  18. Tannins quantification in barks of Mimosa tenuiflora and Acacia mearnsii

    Directory of Open Access Journals (Sweden)

    Leandro Calegari

    2016-03-01

    Full Text Available Due to its chemical complexity, there are several methodologies for vegetable tannins quantification. Thus, this work aims at quantifying both tannin and non-tannin substances present in the barks of Mimosa tenuiflora and Acacia mearnsii by two different methods. From bark particles of both species, analytical solutions were produced by using a steam-jacketed extractor. The solution was analyzed by Stiasny and hide-powder (no chromed methods. For both species, tannin levels were superior when analyzed by hide-powder method, reaching 47.8% and 24.1% for A. mearnsii and M. tenuiflora, respectively. By Stiasny method, the tannins levels considered were 39.0% for A. mearnsii, and 15.5% for M. tenuiflora. Despite the best results presented by A. mearnsii, the bark of M. tenuiflora also showed great potential due to its considerable amount of tannin and the availability of the species at Caatinga biome.

  19. Quantification of hydroxyacetone and glycolaldehyde using chemical ionization mass spectrometry

    Directory of Open Access Journals (Sweden)

    K. M. Spencer

    2011-08-01

    Full Text Available Chemical ionization mass spectrometry (CIMS enables online, fast, in situ detection and quantification of hydroxyacetone and glycolaldehyde. Two different CIMS approaches are demonstrated employing the strengths of single quadrupole mass spectrometry and triple quadrupole (tandem mass spectrometry. Both methods are capable of the measurement of hydroxyacetone, an analyte with minimal isobaric interferences. Tandem mass spectrometry provides direct separation of the isobaric compounds glycolaldehyde and acetic acid using distinct, collision-induced dissociation daughter ions. Measurement of hydroxyacetone and glycolaldehyde by these methods was demonstrated during the ARCTAS-CARB 2008 campaign and the BEARPEX 2009 campaign. Enhancement ratios of these compounds in ambient biomass burning plumes are reported for the ARCTAS-CARB campaign. BEARPEX observations are compared to simple photochemical box model predictions of biogenic volatile organic compound oxidation at the site.

  20. Enhanced techniques for asymmetry quantification in brain imagery

    Science.gov (United States)

    Liu, Xin; Imielinska, Celina; Rosiene, Joel; Connolly, E. S.; D'Ambrosio, Anthony L.

    2006-03-01

    We present an automated generic methodology for symmetry identification and asymmetry quantification, novel method of identifying and delineation of brain pathology by analyzing the opposing sides of the brain utilizing of inherent left-right symmetry in the brain. After symmetry axis has been detected, we apply non-parametric statistical tests operating on the pairs of samples to identify initial seeds points which is defined defined as the pixels where the most statistically significant difference appears. Local region growing is performed on the difference map, from where the seeds are aggregating until it captures all 8-way connected high signals from the difference map. We illustrate the capability of our method with examples ranging from tumors in patient MR data to animal stroke data. The validation results on Rat stroke data have shown that this approach has promise to achieve high precision and full automation in segmenting lesions in reflectional symmetrical objects.

  1. Quantification of projection angle in fragment generator warhead

    Institute of Scientific and Technical Information of China (English)

    K.D.DHOTE; K.P.S.MURTHY; K.M.RAJAN; M.M.SUCHEENDRAN

    2014-01-01

    Tactical Ballistic Missile (TBM) class target neutralization by the fragment spray of a Fragment Generator Warhead (FGW) calls for quantification of fragment projection angle scatter to finalize the end game engagement logic. For conventional axi-symmetric warhead, dispersion is assumed to be normal with a standard deviation of 30. However, such information is not available in case of FGW. Hence, a set of experiments are conducted to determine the dispersion of fragments. The experiments are conducted with a specific configuration of FGW in an identical arena to quantify the scatter and then verified its applicability to other configurations having a range of L/D and C/M ratios, and contoured fragmenting discs. From the experimental study, it is concluded that the scatter in projection angle follows normal distribution with a standard deviation of 0.75? at Chi-square significance level of 0.01(c20.99).

  2. Uncertainty quantification in computational fluid dynamics and aircraft engines

    CERN Document Server

    Montomoli, Francesco; D'Ammaro, Antonio; Massini, Michela; Salvadori, Simone

    2015-01-01

    This book introduces novel design techniques developed to increase the safety of aircraft engines. The authors demonstrate how the application of uncertainty methods can overcome problems in the accurate prediction of engine lift, caused by manufacturing error. This in turn ameliorates the difficulty of achieving required safety margins imposed by limits in current design and manufacturing methods. This text shows that even state-of-the-art computational fluid dynamics (CFD) are not able to predict the same performance measured in experiments; CFD methods assume idealised geometries but ideal geometries do not exist, cannot be manufactured and their performance differs from real-world ones. By applying geometrical variations of a few microns, the agreement with experiments improves dramatically, but unfortunately the manufacturing errors in engines or in experiments are unknown. In order to overcome this limitation, uncertainty quantification considers the probability density functions of manufacturing errors...

  3. Uncertainty quantification of an inflatable/rigidizable torus

    Science.gov (United States)

    Lew, Jiann-Shiun; Horta, Lucas G.; Reaves, Mercedes C.

    2006-06-01

    There is an increasing interest in lightweight inflatable structures for space missions. The dynamic testing and model updating of these types of structures present many challenges in terms of model uncertainty and structural nonlinearity. This paper presents an experimental study of uncertainty quantification of a 3m-diameter inflatable torus. Model uncertainty can be thought of as coming from two different sources, uncertainty due to changes in controlled conditions, such as temperature and input force level, and uncertainty associated with others random factors, such as measurement noise, etc. To precisely investigate and quantify model uncertainty from different sources, experiments, using sine-sweep excitation in the specified narrow frequency bands, are conducted to collect frequency response function (FRF) under various test conditions. To model the variation of the identified parameters, a singular value decomposition technique is applied to extract the principal components of the parameter change.

  4. Quantification of osteolytic bone lesions in a preclinical rat trial

    Science.gov (United States)

    Fränzle, Andrea; Bretschi, Maren; Bäuerle, Tobias; Giske, Kristina; Hillengass, Jens; Bendl, Rolf

    2013-10-01

    In breast cancer, most of the patients who died, have developed bone metastasis as disease progression. Bone metastases in case of breast cancer are mainly bone destructive (osteolytic). To understand pathogenesis and to analyse response to different treatments, animal models, in our case rats, are examined. For assessment of treatment response to bone remodelling therapies exact segmentations of osteolytic lesions are needed. Manual segmentations are not only time-consuming but lack in reproducibility. Computerized segmentation tools are essential. In this paper we present an approach for the computerized quantification of osteolytic lesion volumes using a comparison to a healthy reference model. The presented qualitative and quantitative evaluation of the reconstructed bone volumes show, that the automatically segmented lesion volumes complete missing bone in a reasonable way.

  5. Temporal and spatial quantification of farm and landscape functions

    DEFF Research Database (Denmark)

    Andersen, Peter Stubkjær

    This PhD thesis presents a study on the spatial distribution of agricultural functions at farm and landscape levels. The study focuses on conceptualization of multifunctionality. The concrete conceptual steps include: identification of indicators of four farm and landscape functions – production......, residence, habitat, and recreation; development of a method for quantifying farm functionality and assessing multifunctionality; and definition of a farm typology based on multifunctionality strategies. Empirical data from farm interviews were used in the study to test the developed methods. The results...... related to quantification shows large variation of functional scores among farms of different sizes and different occupational statuses of the farmer. Furthermore, in the study area medium-large farms are the most multifunctional. Farm functionality has changed from the 1990s to 2008. Multifunctionality...

  6. A surrogate accelerated multicanonical Monte Carlo method for uncertainty quantification

    Science.gov (United States)

    Wu, Keyi; Li, Jinglai

    2016-09-01

    In this work we consider a class of uncertainty quantification problems where the system performance or reliability is characterized by a scalar parameter y. The performance parameter y is random due to the presence of various sources of uncertainty in the system, and our goal is to estimate the probability density function (PDF) of y. We propose to use the multicanonical Monte Carlo (MMC) method, a special type of adaptive importance sampling algorithms, to compute the PDF of interest. Moreover, we develop an adaptive algorithm to construct local Gaussian process surrogates to further accelerate the MMC iterations. With numerical examples we demonstrate that the proposed method can achieve several orders of magnitudes of speedup over the standard Monte Carlo methods.

  7. Quantification of tidal parameters from Solar System data

    Science.gov (United States)

    Lainey, Valéry

    2016-05-01

    Tidal dissipation is the main driver of orbital evolution of natural satellites and a key point to understand the exoplanetary system configurations. Despite its importance, its quantification from observations still remains difficult for most objects of our own Solar System. In this work, we overview the method that has been used to determine, directly from observations, the tidal parameters, with emphasis on the Love number k_2 and the tidal quality factor Q. Up-to-date values of these tidal parameters are summarized. Last, an assessment on the possible determination of the tidal ratio k_2/Q of Uranus and Neptune is done. This may be particularly relevant for coming astrometric campaigns and future space missions focused on these systems.

  8. Quantification of Diffuse Hydrothermal Flows Using Multibeam Sonar

    Science.gov (United States)

    Ivakin, A. N.; Jackson, D. R.; Bemis, K. G.; Xu, G.

    2014-12-01

    The Cabled Observatory Vent Imaging Sonar (COVIS) deployed at the Main Endeavour node of the NEPTUNE Canada observatory has provided acoustic time series extending over 2 years. This includes 3D images of plume scattering strength and Doppler velocity measurements as well as 2D images showing regions of diffuse flow. The diffuse-flow images display the level of decorrelation between sonar echos with transmissions separated by 0.2 s. The present work aims to provide further information on the strength of diffuse flows. Two approaches are used: Measurement of the dependence of decorrelation on lag and measurement of phase shift of sonar echos, with lags in 3-hour increments up to several days. The phase shifts and decorrelation are linked to variations of temperature above the seabed, which allows quantification of those variations, their magnitudes, spatial and temporal scales, and energy spectra. These techniques are illustrated using COVIS data obtained near the Grotto vent complex.

  9. Nanoscale elemental quantification in heterostructured SiGe nanowires.

    Science.gov (United States)

    Hourani, W; Periwal, P; Bassani, F; Baron, T; Patriarche, G; Martinez, E

    2015-05-14

    The nanoscale chemical characterization of axial heterostructured Si1-xGex nanowires (NWs) has been performed using scanning Auger microscopy (SAM) through local spectroscopy, line-scan and depth profile measurements. Local Auger profiles are realized with sufficient lateral resolution to resolve individual nanowires. Axial and radial composition heterogeneities are highlighted. Our results confirm the phenomenon of Ge radial growth forming a Ge shell around the nanowire. Moreover, quantification is performed after verifying the absence of preferential sputtering of Si or Ge on a bulk SiGe sample. Hence, reliable results are obtained for heterostructured NW diameters higher than 100 nm. However, for smaller sizes, we have noticed that the sensitivity factors evaluated from bulk samples cannot be used because of edge effects occurring for highly topographical features and a modified contribution of backscattered electrons.

  10. Quantification of rutile in anatase by X-ray diffraction

    International Nuclear Information System (INIS)

    Nowadays the discovering of new and better materials required in all areas of the industry has been lead to the human being to introduce him to this small and great world. The crystalline materials, have properties markedly directional. When it is necessary to realize a quantitative analysis to these materials the task is not easy. The main objective of this work is the research of a real problem, its solution and perfecting of a technique involving the theoretical and experimental principles which allow the quantification of crystalline phases. The chapter 1 treats about the study of crystalline state during the last century, by means of the X-ray diffraction technique. The chapter 2 studies the nature and production of X-rays, the chapter 3 expounds the principles of the diffraction technique which to carry out when it is satisfied the Bragg law studying the powder diffraction method and its applications. In the chapter 4 it is explained how the intensities of the beams diffracted are determined by the atoms positions inside of the elemental cell of the crystal. The properties of the crystalline samples of anatase and rutile are described in the chapter 5. The results of this last analysis are the information which will be processed by means of the auxiliary software: Diffrac AT, Axum and Peakfit as well as the TAFOR and CUANTI software describing this part with more detail in the chapters 6 and 7 where it is mentioned step by step the function of each software until to reach the quantification of crystalline phases, objective of this work. Finally, in the chapter 8 there are a results analysis and conclusions. The contribution of this work is for those learned institutions of limited resources which can tackle in this way the characterization of materials. (Author)

  11. A novel immunological assay for hepcidin quantification in human serum.

    Directory of Open Access Journals (Sweden)

    Vasiliki Koliaraki

    Full Text Available BACKGROUND: Hepcidin is a 25-aminoacid cysteine-rich iron regulating peptide. Increased hepcidin concentrations lead to iron sequestration in macrophages, contributing to the pathogenesis of anaemia of chronic disease whereas decreased hepcidin is observed in iron deficiency and primary iron overload diseases such as hereditary hemochromatosis. Hepcidin quantification in human blood or urine may provide further insights for the pathogenesis of disorders of iron homeostasis and might prove a valuable tool for clinicians for the differential diagnosis of anaemia. This study describes a specific and non-operator demanding immunoassay for hepcidin quantification in human sera. METHODS AND FINDINGS: An ELISA assay was developed for measuring hepcidin serum concentration using a recombinant hepcidin25-His peptide and a polyclonal antibody against this peptide, which was able to identify native hepcidin. The ELISA assay had a detection range of 10-1500 microg/L and a detection limit of 5.4 microg/L. The intra- and interassay coefficients of variance ranged from 8-15% and 5-16%, respectively. Mean linearity and recovery were 101% and 107%, respectively. Mean hepcidin levels were significantly lower in 7 patients with juvenile hemochromatosis (12.8 microg/L and 10 patients with iron deficiency anemia (15.7 microg/L and higher in 7 patients with Hodgkin lymphoma (116.7 microg/L compared to 32 age-matched healthy controls (42.7 microg/L. CONCLUSIONS: We describe a new simple ELISA assay for measuring hepcidin in human serum with sufficient accuracy and reproducibility.

  12. Bayesian Proteoform Modeling Improves Protein Quantification of Global Proteomic Measurements

    Energy Technology Data Exchange (ETDEWEB)

    Webb-Robertson, Bobbie-Jo M.; Matzke, Melissa M.; Datta, Susmita; Payne, Samuel H.; Kang, Jiyun; Bramer, Lisa M.; Nicora, Carrie D.; Shukla, Anil K.; Metz, Thomas O.; Rodland, Karin D.; Smith, Richard D.; Tardiff, Mark F.; McDermott, Jason E.; Pounds, Joel G.; Waters, Katrina M.

    2014-12-01

    As the capability of mass spectrometry-based proteomics has matured, tens of thousands of peptides can be measured simultaneously, which has the benefit of offering a systems view of protein expression. However, a major challenge is that with an increase in throughput, protein quantification estimation from the native measured peptides has become a computational task. A limitation to existing computationally-driven protein quantification methods is that most ignore protein variation, such as alternate splicing of the RNA transcript and post-translational modifications or other possible proteoforms, which will affect a significant fraction of the proteome. The consequence of this assumption is that statistical inference at the protein level, and consequently downstream analyses, such as network and pathway modeling, have only limited power for biomarker discovery. Here, we describe a Bayesian model (BP-Quant) that uses statistically derived peptides signatures to identify peptides that are outside the dominant pattern, or the existence of multiple over-expressed patterns to improve relative protein abundance estimates. It is a research-driven approach that utilizes the objectives of the experiment, defined in the context of a standard statistical hypothesis, to identify a set of peptides exhibiting similar statistical behavior relating to a protein. This approach infers that changes in relative protein abundance can be used as a surrogate for changes in function, without necessarily taking into account the effect of differential post-translational modifications, processing, or splicing in altering protein function. We verify the approach using a dilution study from mouse plasma samples and demonstrate that BP-Quant achieves similar accuracy as the current state-of-the-art methods at proteoform identification with significantly better specificity. BP-Quant is available as a MatLab ® and R packages at https://github.com/PNNL-Comp-Mass-Spec/BP-Quant.

  13. Rapid quantification method for Legionella pneumophila in surface water.

    Science.gov (United States)

    Wunderlich, Anika; Torggler, Carmen; Elsässer, Dennis; Lück, Christian; Niessner, Reinhard; Seidel, Michael

    2016-03-01

    World-wide legionellosis outbreaks caused by evaporative cooling systems have shown that there is a need for rapid screening methods for Legionella pneumophila in water. Antibody-based methods for the quantification of L. pneumophila are rapid, non-laborious, and relatively cheap but not sensitive enough for establishment as a screening method for surface and drinking water. Therefore, preconcentration methods have to be applied in advance to reach the needed sensitivity. In a basic test, monolithic adsorption filtration (MAF) was used as primary preconcentration method that adsorbs L. pneumophila with high efficiency. Ten-liter water samples were concentrated in 10 min and further reduced to 1 mL by centrifugal ultrafiltration (CeUF). The quantification of L. pneumophila strains belonging to the monoclonal subtype Bellingham was performed via flow-based chemiluminescence sandwich microarray immunoassays (CL-SMIA) in 36 min. The whole analysis process takes 90 min. A polyclonal antibody (pAb) against L. pneumophila serogroup 1-12 and a monoclonal antibody (mAb) against L. pneumophila SG 1 strain Bellingham were immobilized on a microarray chip. Without preconcentration, the detection limit was 4.0 × 10(3) and 2.8 × 10(3) CFU/mL determined by pAb and mAb 10/6, respectively. For samples processed by MAF-CeUF prior to SMIA detection, the limit of detection (LOD) could be decreased to 8.7 CFU/mL and 0.39 CFU/mL, respectively. A recovery of 99.8 ± 15.9% was achieved for concentrations between 1-1000 CFU/mL. The established combined analytical method is sensitive for rapid screening of surface and drinking water to allow fast hygiene control of L. pneumophila. PMID:26873217

  14. Quantification of rat kisspeptin using a novel radioimmunoassay.

    Directory of Open Access Journals (Sweden)

    James S Kinsey-Jones

    Full Text Available Kisspeptin is a hypothalamic peptide hormone that plays a pivotal role in pubertal onset and reproductive function. Previous studies have examined hypothalamic kisspeptin mRNA expression, either through in situ hybridisation or real-time RT-PCR, as a means quantifying kisspeptin gene expression. However, mRNA expression levels are not always reflected in levels of the translated protein. Kisspeptin-immunoreactivity (IR has been extensively examined using immunohistochemistry, enabling detection and localisation of kisspeptin perikaya in the arcuate nucleus (ARC and anteroventral periventricular nucleus (AVPV. However, quantification of kisspeptin-IR remains challenging. We developed a specific rodent radioimmunoassay assay (RIA capable of detecting and quantifying kisspeptin-IR in rodent tissues. The RIA uses kisspeptin-10 as a standard and radioactive tracer, combined with a commercially available antibody raised to the kisspeptin-10 fragment. Adult female wistar rat brain samples were sectioned at 300 µm and the ARC and AVPV punch micro-dissected. Brain punches were homogenised in extraction buffer and assayed with rodent kisspeptin-RIA. In accord with the pattern of kisspeptin mRNA expression, kisspeptin-IR was detected in both the ARC (47.1±6.2 fmol/punch, mean±SEM n = 15 and AVPV (7.6±1.3 fmol/punch, mean±SEM n = 15. Kisspeptin-IR was also detectable in rat placenta (1.26±0.15 fmol/mg. Reverse phase high pressure liquid chromatography analysis showed that hypothalamic kisspeptin-IR had the same elution profile as a synthetic rodent kisspeptin standard. A specific rodent kisspeptin-RIA will allow accurate quantification of kisspeptin peptide levels within specific tissues in rodent experimental models.

  15. Quantification and Localization of Mast Cells in Periapical Lesions

    Science.gov (United States)

    Mahita, VN; Manjunatha, BS; Shah, R; Astekar, M; Purohit, S; Kovvuru, S

    2015-01-01

    Background: Periapical lesions occur in response to chronic irritation in periapical tissue, generally resulting from an infected root canal. Specific etiological agents of induction, participating cell population and growth factors associated with maintenance and resolution of periapical lesions are incompletely understood. Among the cells found in periapical lesions, mast cells have been implicated in the inflammatory mechanism. Aim: Quantifications and the possible role played by mast cells in the periapical granuloma and radicular cyst. Hence, this study is to emphasize the presence (localization) and quantification of mast cells in periapical granuloma and radicular cyst. Materials and Methods: A total of 30 cases and out of which 15 of periapical granuloma and 15 radicular cyst, each along with the case details from the previously diagnosed cases in the department of oral pathology were selected for the study. The gender distribution showed male 8 (53.3%) and females 7 (46.7%) in periapical granuloma cases and male 10 (66.7%) and females 5 (33.3%) in radicular cyst cases. The statistical analysis used was unpaired t-test. Results: Mean mast cell count in periapical granuloma subepithelial and deeper connective tissue, was 12.40 (0.99%) and 7.13 (0.83%), respectively. The mean mast cell counts in subepithelial and deeper connective tissue of radicular cyst were 17.64 (1.59%) and 12.06 (1.33%) respectively, which was statistically significant. No statistical significant difference was noted among males and females. Conclusion: Mast cells were more in number in radicular cyst. Based on the concept that mast cells play a critical role in the induction of inflammation, it is logical to use therapeutic agents to alter mast cell function and secretion, to thwart inflammation at its earliest phases. These findings may suggest the possible role of mast cells in the pathogenesis of periapical lesions. PMID:25861530

  16. Quantification of Carbohydrates in Grape Tissues Using Capillary Zone Electrophoresis

    Science.gov (United States)

    Zhao, Lu; Chanon, Ann M.; Chattopadhyay, Nabanita; Dami, Imed E.; Blakeslee, Joshua J.

    2016-01-01

    Soluble sugars play an important role in freezing tolerance in both herbaceous and woody plants, functioning in both the reduction of freezing-induced dehydration and the cryoprotection of cellular constituents. The quantification of soluble sugars in plant tissues is, therefore, essential in understanding freezing tolerance. While a number of analytical techniques and methods have been used to quantify sugars, most of these are expensive and time-consuming due to complex sample preparation procedures which require the derivatization of the carbohydrates being analyzed. Analysis of soluble sugars using capillary zone electrophoresis (CZE) under alkaline conditions with direct UV detection has previously been used to quantify simple sugars in fruit juices. However, it was unclear whether CZE-based methods could be successfully used to quantify the broader range of sugars present in complex plant extracts. Here, we present the development of an optimized CZE method capable of separating and quantifying mono-, di-, and tri-saccharides isolated from plant tissues. This optimized CZE method employs a column electrolyte buffer containing 130 mM NaOH, pH 13.0, creating a current of 185 μA when a separation voltage of 10 kV is employed. The optimized CZE method provides limits-of-detection (an average of 1.5 ng/μL) for individual carbohydrates comparable or superior to those obtained using gas chromatography–mass spectrometry, and allows resolution of non-structural sugars and cell wall components (structural sugars). The optimized CZE method was successfully used to quantify sugars from grape leaves and buds, and is a robust tool for the quantification of plant sugars found in vegetative and woody tissues. The increased analytical efficiency of this CZE method makes it ideal for use in high-throughput metabolomics studies designed to quantify plant sugars. PMID:27379118

  17. BioMaS: a modular pipeline for Bioinformatic analysis of Metagenomic AmpliconS

    OpenAIRE

    Fosso, Bruno; Santamaria, Monica; Marzano, Marinella; Alonso-Alemany, Daniel; Valiente Feruglio, Gabriel Alejandro; Donvito, Giacinto; Monaco, Alfonso; Notarangelo, Pasquale; Pesole, Graziano

    2015-01-01

    Background Substantial advances in microbiology, molecular evolution and biodiversity have been carried out in recent years thanks to Metagenomics, which allows to unveil the composition and functions of mixed microbial communities in any environmental niche. If the investigation is aimed only at the microbiome taxonomic structure, a target-based metagenomic approach, here also referred as Meta-barcoding, is generally applied. This approach commonly involves the selective amplification of a s...

  18. [Remarks on application of acupuncture instruments in acupuncture quantification and normalization studies].

    Science.gov (United States)

    Liu, Jian; Fan, Xiao-Nong; Wang, Shu; Shi, Xue-Min

    2009-01-01

    Acupuncture manipulation quantification is an important link of acupuncture normalization study. Because traditional acupuncture manipulation are complicated with difficulty of quantification, acupuncture instruments provide a new way for acupuncture manipulation quantification and acupuncture normalization. It is necessary to increase the understanding of the importance of developing acupuncture instruments, strengthen the study of related theory and development of acupuncture measurement instruments and acupuncture imitation instruments, enlarge serviceable range, verify efficacy, develop the theory of acupuncture manipulation, richen the study methods of acupuncture normalization, so as to promote internationalization of acupuncture.

  19. Quantification of rice bran oil in oil blends

    Directory of Open Access Journals (Sweden)

    Mishra, R.

    2012-03-01

    Full Text Available Blends consisting of physically refined rice bran oil (PRBO: sunflower oil (SnF and PRBO: safflower oil (SAF in different proportions were analyzed for various physicochemical parameters. The quantification of pure rice bran oil in the blended oils was carried out using different methods including gas chromatographic, HPLC, ultrasonic velocity and methods based on physico-chemical parameters. The physicochemical parameters such as ultrasonic velocity, relative association and acoustic impedance at 2 MHz, iodine value, palmitic acid content and oryzanol content reflected significant changes with increased proportions of PRBO in the blended oils. These parameters were selected as dependent parameters and % PRBO proportion was selected as independent parameters. The study revealed that regression equations based on the oryzanol content, palmitic acid composition, ultrasonic velocity, relative association, acoustic impedance, and iodine value can be used for the quantification of rice bran oil in blended oils. The rice bran oil can easily be quantified in the blended oils based on the oryzanol content by HPLC even at a 1% level. The palmitic acid content in blended oils can also be used as an indicator to quantify rice bran oil at or above the 20% level in blended oils whereas the method based on ultrasonic velocity, acoustic impedance and relative association showed initial promise in the quantification of rice bran oil.

    Se analizaron diversos parámetros físico-químicos para la evaluación de mezclas de aceites en diferentes proporciones que incluyen: aceite de salvado de arroz físícamente refinado (PRBO: aceite de girasol (SNF y las mezclas PRBO: aceite de cártamo (SAF en diferentes proporciones. La cuantificación de la presencia del aceite de salvado de arroz en las mezclas se llevó a cabo por diferentes métodos, como cromatografía de gases (GC, cromatografía líquida (HPLC, ultrasonidos y métodos basados en otros parámetros f

  20. Quantification of glucosylceramide in plasma of Gaucher disease patients

    Directory of Open Access Journals (Sweden)

    Maria Viviane Gomes Muller

    2010-12-01

    Full Text Available Gaucher disease is a sphingolipidosis that leads to an accumulation of glucosylceramide. The objective of this study was to develop a methodology, based on the extraction, purification and quantification of glucosylceramide from blood plasma, for use in clinical research laboratories. Comparison of the glucosylceramide content in plasma from Gaucher disease patients, submitted to enzyme replacement therapy or otherwise, against that from normal individuals was also carried out. The glucosylceramide, separated from other glycosphingolipids by high performance thin layer chromatography (HPTLC was chemically developed (CuSO4 / H3PO4 and the respective band confirmed by immunostaining (human anti-glucosylceramide antibody / peroxidase-conjugated secondary antibody. Chromatogram quantification by densitometry demonstrated that the glucosylceramide content in Gaucher disease patients was seventeen times higher than that in normal individuals, and seven times higher than that in patients on enzyme replacement therapy. The results obtained indicate that the methodology established can be used in complementary diagnosis and for treatment monitoring of Gaucher disease patients.A doença de Gaucher é uma esfingolipidose caracterizada pelo acúmulo de glicosilceramida. O objetivo deste estudo foi desenvolver metodologia baseada na extração, purificação e quantificação da glicosilceramida plasmática a qual possa ser usada em laboratórios de pesquisa clínica. Após o desenvolvimento desta metodologia, foi proposto, também, comparar o conteúdo de glicosilceramida presente no plasma de pacientes com doença de Gaucher, submetidos ou não a tratamento, com aquele de indivíduos normais. A glicosilceramida, separada de outros glicoesfingolipídios por cromatografia de camada delgada de alto desempenho (HPTLC, foi revelada quimicamente (CuSO4/H3PO4 e a respectiva banda foi confirmada por imunorrevelação (anticorpo anti-glicosilceramida humana

  1. Quantification of clarithromycin polymorphs in presence of tablet excipients.

    Directory of Open Access Journals (Sweden)

    Swathi Kuncham

    2014-03-01

    Full Text Available Excipients can cause a considerable challenge when developing a solid form of an active pharmaceutical ingredient (API. The aim of this present study was to analyze the polymorphs of clarithromycin (CAM mixed with excipients using powder X-ray diffraction (PXRD. Polymorphic Form I (CAM-1, Form II (CAM-2 and an amorphous phase of CAM were characterized using thermal and crystallographic methods. CAM-1 and CAM-2 were monotropically related, with CAM-2 being the stable form. PXRD instrument related parameters were optimized for the characterization of CAM polymorphic forms using a variety of excipients. Calibration curves for CAM-1 and CAM-2 mixed with excipients were also prepared. Analytical methods based on the differences in the diffraction patterns of CAM-1, CAM-2 and the excipients were developed. Sodium methyl paraben, sodium propyl paraben, microcrystalline cellulose and magnesium stearate were crystalline showing characteristic diffraction patterns. Starch, croscarmellose sodium, talc and sodium starch glycolate were semicrystalline in nature, while colloidal silicon dioxide was amorphous. A diffraction peak at 8.7° 2θ provided a quantification of CAM-2 when mixed with excipients. The analytical method was evaluated and validated for accuracy, precision, inter- and intra-day variation, variability due to sample repacking and instrument reproducibility. The method for quantification of CAM-2 in the range of 80 to 100% w/w was linear with R2 = 0.998. Relative standard deviation (RSD, due to sample repacking, was 2.77% indicating good homogeneity of mixing of the samples. RSD due to assay errors was 1.66%. PXRD analysis of the commercial tablet showed the CAM-2 as a major polymorph being 98% of the overall content of the API. CAM-1 was found to be present as an impurity at trace levels shown by peaks at 2θ values of 5.2° and 6.7°. This method provides a method for characterization of the polymorphic forms of CAM in the presence of

  2. Development of hydrate risk quantification in oil and gas production

    Science.gov (United States)

    Chaudhari, Piyush N.

    order to reduce the parametric study that may require a long duration of time using The Colorado School of Mines Hydrate Kinetic Model (CSMHyK). The evolution of the hydrate plugging risk along flowline-riser systems is modeled for steady state and transient operations considering the effect of several critical parameters such as oil-hydrate slip, duration of shut-in, and water droplet size on a subsea tieback system. This research presents a novel platform for quantification of the hydrate plugging risk, which in-turn will play an important role in improving and optimizing current hydrate management strategies. The predictive strength of the hydrate risk quantification and hydrate prediction models will have a significant impact on flow assurance engineering and design with respect to building safe and efficient hydrate management techniques for future deep-water developments.

  3. Coral Pigments: Quantification Using HPLC and Detection by Remote Sensing

    Science.gov (United States)

    Cottone, Mary C.

    1995-01-01

    Widespread coral bleaching (loss of pigments of symbiotic dinoflagellates), and the corresponding decline in coral reef health worldwide, mandates the monitoring of coral pigmentation. Samples of the corals Porites compressa and P. lobata were collected from a healthy reef at Puako, Hawaii, and chlorophyll (chl) a, peridinin, and Beta-carotene (Beta-car) were quantified using reverse-phase high performance liquid chromatography (HPLC). Detailed procedures are presented for the extraction of the coral pigments in 90% acetone, and the separation, identification, and quantification of the major zooxanthellar pigments using spectrophotometry and a modification of the HPLC system described by Mantoura and Llewellyn (1983). Beta-apo-8-carotenal was found to be inadequate as in internal standard, due to coelution with chl b and/or chl a allomer in the sample extracts. Improvements are suggested, which may result in better resolution of the major pigments and greater accuracy in quantification. Average concentrations of peridinin, chl a, and Beta-car in corals on the reef were 5.01, 8.59, and 0.29, micro-grams/cm(exp 2), respectively. Average concentrations of peridinin and Beta-car did not differ significantly between the two coral species sampled; however, the mean chl a concentration in P. compressa specimens (7.81 ,micro-grams/cm(exp 2) was significantly lower than that in P. lobata specimens (9.96 11g/cm2). Chl a concentrations determined spectrophotometrically were significantly higher than those generated through HPLC, suggesting that spectrophotometry overestimates chl a concentrations. The average ratio of chl a-to-peridinin concentrations was 1.90, with a large (53%) coefficient of variation and a significant difference between the two species sampled. Additional data are needed before conclusions can be drawn regarding average pigment concentrations in healthy corals and the consistency of the chl a/peridinin ratio. The HPLC pigment concentration values

  4. Collaborative framework for PIV uncertainty quantification: the experimental database

    International Nuclear Information System (INIS)

    The uncertainty quantification of particle image velocimetry (PIV) measurements has recently become a topic of great interest as shown by the recent appearance of several different methods within the past few years. These approaches have different working principles, merits and limitations, which have been speculated upon in subsequent studies. This paper reports a unique experiment that has been performed specifically to test the efficacy of PIV uncertainty methods. The case of a rectangular jet, as previously studied by Timmins et al (2012) and Wilson and Smith (2013b), is used. The novel aspect of the experiment is simultaneous velocity measurements using two different time-resolved PIV systems and a hot-wire anemometry (HWA) system. The first PIV system, called the PIV measurement system (‘PIV-MS’), is intended for nominal measurements of which the uncertainty is to be evaluated. It is based on a single camera and features a dynamic velocity range (DVR) representative of typical PIV experiments. The second PIV system, called the ‘PIV-HDR’ (high dynamic range) system, features a significantly higher DVR obtained with a higher digital imaging resolution. The hot-wire is placed in close proximity to the PIV measurement domain. The three measurement systems were carefully set to simultaneously measure the flow velocity at the same time and location. The comparison between the PIV-HDR system and the HWA provides an estimate of the measurement precision of the reference velocity for evaluation of the instantaneous error in the measurement system. The discrepancy between the PIV-MS and the reference data provides the measurement error, which is later used to assess the different uncertainty quantification methods proposed in the literature. A detailed comparison of the uncertainty estimation methods based on the present datasets is presented in a second paper from Sciacchitano et al (2015). Furthermore, this database offers the potential to be used for

  5. Dispersoid quantification and size distribution in hot and cold processed AA3103

    International Nuclear Information System (INIS)

    A quantification procedure coupling Field Emission Gun Scanning Electron Microscope (FEGSEM) and image analysis software has been developed to obtain size distributions of secondary phase dispersoids. The procedure is described in detail for aluminium alloys, and each involved parameter is evaluated separately. The first part of the procedure is the acquisition of the micrograph; both microscope and software parameters are important in this stage. The second part is the most critical step, where the micrograph is transformed to a binary image. Here, the choice of the threshold value is of great importance because it corresponds to the maximum loss of information from the original image. The quantification procedure is then presented with considerations and examples of statistical meaning. The quantification method is applied to a series of AA3103 samples taken from different industrial processing steps. The results obtained are consistent with the expected trends and prove the quantification procedure to be valid

  6. The Effect of AOP on Software Engineering, with Particular Attention to OIF and Event Quantification

    Science.gov (United States)

    Havelund, Klaus; Filman, Robert; Korsmeyer, David (Technical Monitor)

    2003-01-01

    We consider the impact of Aspect-Oriented Programming on Software Engineering, and, in particular, analyze two AOP systems, one of which does component wrapping and the other, quantification over events, for their software engineering effects.

  7. Quantification of synovial and erosive changes in rheumatoid arthritis with ultrasound-Revisited

    Energy Technology Data Exchange (ETDEWEB)

    Schueller-Weidekamm, Claudia [Department of Radiology, Medical University of Vienna, Vienna General Hospital, Waehringer Guertel 18-20, A-1090 Vienna (Austria)], E-mail: claudia.schueller-weidekamm@meduniwien.ac.at

    2009-08-15

    Synovitis is a predictive factor of irreversible changes in the joints, tendons, and ligaments in patients with rheumatoid arthritis (RA). Therefore, the early demonstration of reversible, pre-erosive inflammatory features to diagnose RA, the monitoring of disease activity, and the response to therapy are of great importance. Technical developments in ultrasound now allow the quantification of synovitis and erosions, and enable the assessment and follow-up of disease activity. However, both the subjective and objective quantification techniques are associated with different problems. This review article highlights the advantages and disadvantages of sonographic quantification, and revisits the somewhat controversial positions apparent in the current literature. Familiarity with the imaging findings and the scoring systems used to characterize erosive changes are prerequisites for considerably improving the detection and monitoring of synovitis and erosions. The role of ultrasound in the diagnostic approach to RA, particularly in the quantification of synovial and erosive changes, will be explored and the current literature will be reviewed.

  8. The Effect of Human Genome Annotation Complexity on RNA-Seq Gene Expression Quantification

    Science.gov (United States)

    Wu, Po-Yen; Phan, John H.; Wang, May D.

    2016-01-01

    Next-generation sequencing (NGS) has brought human genomic research to an unprecedented era. RNA-Seq is a branch of NGS that can be used to quantify gene expression and depends on accurate annotation of the human genome (i.e., the definition of genes and all of their variants or isoforms). Multiple annotations of the human genome exist with varying complexity. However, it is not clear how the choice of genome annotation influences RNA-Seq gene expression quantification. We assess the effect of different genome annotations in terms of (1) mapping quality, (2) quantification variation, (3) quantification accuracy (i.e., by comparing to qRT-PCR data), and (4) the concordance of detecting differentially expressed genes. External validation with qRT-PCR suggests that more complex genome annotations result in higher quantification variation.

  9. Nanoparticle-based detection and quantification of DNA with single nucleotide polymorphism (SNP) discrimination selectivity

    OpenAIRE

    Qin, Wei Jie; Yung, Lin Yue Lanry

    2007-01-01

    Sequence-specific DNA detection is important in various biomedical applications such as gene expression profiling, disease diagnosis and treatment, drug discovery and forensic analysis. Here we report a gold nanoparticle-based method that allows DNA detection and quantification and is capable of single nucleotide polymorphism (SNP) discrimination. The precise quantification of single-stranded DNA is due to the formation of defined nanoparticle-DNA conjugate groupings in the presence of target...

  10. Enhanced Information Output From Shotgun Proteomics Data by Protein Quantification and Peptide Quality Control (PQPQ)

    OpenAIRE

    Forshed, J.; Johansson, H. J.; Pernemalm, M; Branca, R. M. M.; Sandberg, A.; Lehtio, J.

    2011-01-01

    We present a tool to improve quantitative accuracy and precision in mass spectrometry based on shotgun proteomics: protein quantification by peptide quality control, PQPQ. The method is based on the assumption that the quantitative pattern of peptides derived from one protein will correlate over several samples. Dissonant patterns arise either from outlier peptides or because of the presence of different protein species. By correlation analysis, protein quantification by peptide quality contr...

  11. Quantification of Abscisic Acid, Cytokinin, and Auxin Content in Salt-Stressed Plant Tissues

    OpenAIRE

    Dobrev, P.; Vaňková, R. (Radomíra)

    2012-01-01

    Plant hormones cytokinins, auxin (indole-3-acetic acid), and abscisic acid are central to regulation of plant growth and defence to abiotic stresses such as salinity. Quantification of the hormone levels and determination of their ratios can reveal different plant strategies to cope with the stress, e.g., suppression of growth or mobilization of plant metabolism. This chapter describes a procedure enabling such quantification. Due to the high variability of these hormones in plant tissues, it...

  12. MR Spectroscopy: Real-Time Quantification of in-vivo MR Spectroscopic data

    OpenAIRE

    Massé, Kunal

    2009-01-01

    In the last two decades, magnetic resonance spectroscopy (MRS) has had an increasing success in biomedical research. This technique has the faculty of discerning several metabolites in human tissue non-invasively and thus offers a multitude of medical applications. In clinical routine, quantification plays a key role in the evaluation of the different chemical elements. The quantification of metabolites characterizing specific pathologies helps physicians establish the patient's diagnosis. E...

  13. Evaluation of chromium concentration in cattle feces using different acid digestion and spectrophotometric quantification techniques

    OpenAIRE

    N.K.P. Souza; E. Detmann; D. S. Pina; Valadares Filho, S. C.; C.B. Sampaio; A.C. Queiroz; C.M. Veloso

    2013-01-01

    The objective of this work was to evaluate combinations between acid digestion techniques and spectrophotometric quantification to measure chromium concentration in cattle feces. Digestion techniques were evaluated based on the use of nitric and perchloric acids, sulfuric and perchloric acids, and phosphoric acid. The chromium quantification in the solutions was performed by colorimetry and by atomic absorption spectrophotometry (AAS). When AAS was used, the addition of calcium chloride to th...

  14. Stochastic modeling for magnetic resonance quantification of myocardial blood flow

    Science.gov (United States)

    Seethamraju, Ravi T.; Muehling, Olaf; Panse, Prasad M.; Wilke, Norbert M.; Jerosch-Herold, Michael

    2000-10-01

    Quantification of myocardial blood flow is useful for determining the functional severity of coronary artery lesions. With advances in MR imaging it has become possible to assess myocardial perfusion and blood flow in a non-invasive manner by rapid serial imaging following injection of contrast agent. To date most approaches reported in the literature relied mostly on deriving relative indices of myocardial perfusion directly from the measured signal intensity curves. The central volume principle on the other hand states that it is possible to derive absolute myocardial blood flow from the tissue impulse response. Because of the sensitivity involved in deconvolution due to noise in measured data, conventional methods are sub-optimal, hence, we propose to use stochastic time series modeling techniques like ARMA to obtain a robust impulse response estimate. It is shown that these methods when applied for the optical estimation of the transfer function give accurate estimates of myocardial blood flow. The most significant advantage of this approach, compared with compartmental tracer kinetic models, is the use of a minimum set of prior assumptions on data. The bottleneck in assessing myocardial blood flow, does not lie in the MRI acquisition, but rather in the effort or time for post processing. It is anticipated that the very limited requirements for user input and interaction will be of significant advantage for the clinical application of these methods. The proposed methods are validated by comparison with mean blood flow measurements obtained from radio-isotope labeled microspheres.

  15. On Uncertainty Quantification of Lithium-ion Batteries

    CERN Document Server

    Hadigol, Mohammad; Doostan, Alireza

    2015-01-01

    In this work, a stochastic, physics-based model for Lithium-ion batteries (LIBs) is presented in order to study the effects of model uncertainties on the cell capacity, voltage, and concentrations. To this end, the proposed uncertainty quantification (UQ) approach, based on sparse polynomial chaos expansions, relies on a small number of battery simulations. Within this UQ framework, the identification of most important uncertainty sources is achieved by performing a global sensitivity analysis via computing the so-called Sobol' indices. Such information aids in designing more efficient and targeted quality control procedures, which consequently may result in reducing the LIB production cost. An LiC$_6$/LiCoO$_2$ cell with 19 uncertain parameters discharged at 0.25C, 1C and 4C rates is considered to study the performance and accuracy of the proposed UQ approach. The results suggest that, for the considered cell, the battery discharge rate is a key factor affecting not only the performance variability of the ce...

  16. Numerical Continuation Methods for Intrusive Uncertainty Quantification Studies

    Energy Technology Data Exchange (ETDEWEB)

    Safta, Cosmin [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Najm, Habib N. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Phipps, Eric Todd [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2014-09-01

    Rigorous modeling of engineering systems relies on efficient propagation of uncertainty from input parameters to model outputs. In recent years, there has been substantial development of probabilistic polynomial chaos (PC) Uncertainty Quantification (UQ) methods, enabling studies in expensive computational models. One approach, termed ”intrusive”, involving reformulation of the governing equations, has been found to have superior computational performance compared to non-intrusive sampling-based methods in relevant large-scale problems, particularly in the context of emerging architectures. However, the utility of intrusive methods has been severely limited due to detrimental numerical instabilities associated with strong nonlinear physics. Previous methods for stabilizing these constructions tend to add unacceptably high computational costs, particularly in problems with many uncertain parameters. In order to address these challenges, we propose to adapt and improve numerical continuation methods for the robust time integration of intrusive PC system dynamics. We propose adaptive methods, starting with a small uncertainty for which the model has stable behavior and gradually moving to larger uncertainty where the instabilities are rampant, in a manner that provides a suitable solution.

  17. A surrogate-based uncertainty quantification with quantifiable errors

    Energy Technology Data Exchange (ETDEWEB)

    Bang, Y.; Abdel-Khalik, H. S. [North Carolina State Univ., Raleigh, NC 27695 (United States)

    2012-07-01

    Surrogate models are often employed to reduce the computational cost required to complete uncertainty quantification, where one is interested in propagating input parameters uncertainties throughout a complex engineering model to estimate responses uncertainties. An improved surrogate construction approach is introduced here which places a premium on reducing the associated computational cost. Unlike existing methods where the surrogate is constructed first, then employed to propagate uncertainties, the new approach combines both sensitivity and uncertainty information to render further reduction in the computational cost. Mathematically, the reduction is described by a range finding algorithm that identifies a subspace in the parameters space, whereby parameters uncertainties orthogonal to the subspace contribute negligible amount to the propagated uncertainties. Moreover, the error resulting from the reduction can be upper-bounded. The new approach is demonstrated using a realistic nuclear assembly model and compared to existing methods in terms of computational cost and accuracy of uncertainties. Although we believe the algorithm is general, it will be applied here for linear-based surrogates and Gaussian parameters uncertainties. The generalization to nonlinear models will be detailed in a separate article. (authors)

  18. Quantification of acidic compounds in complex biomass-derived streams

    Energy Technology Data Exchange (ETDEWEB)

    Karp, Eric M.; Nimlos, Claire T.; Deutch, Steve; Salvachúa, Davinia; Cywar, Robin M.; Beckham, Gregg T.

    2016-01-01

    Biomass-derived streams that contain acidic compounds from the degradation of lignin and polysaccharides (e.g. black liquor, pyrolysis oil, pyrolytic lignin, etc.) are chemically complex solutions prone to instability and degradation during analysis, making quantification of compounds within them challenging. Here we present a robust analytical method to quantify acidic compounds in complex biomass-derived mixtures using ion exchange, sample reconstitution in pyridine and derivatization with BSTFA. The procedure is based on an earlier method originally reported for kraft black liquors and, in this work, is applied to identify and quantify a large slate of acidic compounds in corn stover derived alkaline pretreatment liquor (APL) as a function of pretreatment severity. Analysis of the samples is conducted with GCxGC-TOFMS to achieve good resolution of the components within the complex mixture. The results reveal the dominant low molecular weight components and their concentrations as a function of pretreatment severity. Application of this method is also demonstrated in the context of lignin conversion technologies by applying it to track the microbial conversion of an APL substrate. Here too excellent results are achieved, and the appearance and disappearance of compounds is observed in agreement with the known metabolic pathways of two bacteria, indicating the sample integrity was maintained throughout analysis. Finally, it is shown that this method applies more generally to lignin-rich materials by demonstrating its usefulness in analysis of pyrolysis oil and pyrolytic lignin.

  19. Experimental quantification of contact freezing in an electrodynamic balance

    Directory of Open Access Journals (Sweden)

    N. Hoffmann

    2013-04-01

    Full Text Available Heterogeneous nucleation of ice in a supercooled water droplet induced by an external contact with a dry aerosol particle has long been known to be more effective than freezing induced by the same nucleus immersed in the droplet. However, the experimental quantification of contact freezing is challenging. Here we report an experimental method allowing to determine the temperature dependent ice nucleation probability of size selected aerosol particles. The method uses supercooled charged water droplets suspended in a laminar flow of air containing aerosol particles as contact freezing nuclei. The rate of droplet–particle collisions is calculated numerically with account for Coulomb attraction, drag force and induced dipole interaction between charged droplet and aerosol particles. The calculation is verified by direct counting of aerosol particles collected by a levitated droplet. By repeating the experiment on individual droplets for a sufficient number of times, we are able to reproduce the statistical freezing behavior of a large ensemble of supercooled droplets and measure the average rate of freezing events. The freezing rate is equal to the product of the droplet–particle collision rate and the probability of freezing on a single contact, the latter being a function of temperature, size and composition of the contact ice nuclei. Based on these observations, we show that for the types of particles investigated so far, contact freezing is the dominating freezing mechanism on the time scale of our experiment.

  20. Experimental quantification of contact freezing in an electrodynamic balance

    Directory of Open Access Journals (Sweden)

    N. Hoffmann

    2013-09-01

    Full Text Available Heterogeneous nucleation of ice in a supercooled water droplet induced by external contact with a dry aerosol particle has long been known to be more effective than freezing induced by the same nucleus immersed in the droplet. However, the experimental quantification of contact freezing is challenging. Here we report an experimental method to determine the temperature-dependent ice nucleation probability of size-selected aerosol particles. The method is based on the suspension of supercooled charged water droplets in a laminar flow of air containing aerosol particles as contact freezing nuclei. The rate of droplet–particle collisions is calculated numerically with account for Coulomb attraction, drag force and induced dipole interaction between charged droplet and aerosol particles. The calculation is verified by direct counting of aerosol particles collected by a levitated droplet. By repeating the experiment on individual droplets for a sufficient number of times, we are able to reproduce the statistical freezing behavior of a large ensemble of supercooled droplets and measure the average rate of freezing events. The freezing rate is equal to the product of the droplet–particle collision rate and the probability of freezing on a single contact, the latter being a function of temperature, size and composition of the contact ice nuclei. Based on these observations, we show that for the types of particles investigated so far, contact freezing is the dominating freezing mechanism on the timescale of our experiment.

  1. Quantification of sugars in breakfast cereals using capillary electrophoresis.

    Science.gov (United States)

    Toutounji, Michelle R; Van Leeuwen, Matthew P; Oliver, James D; Shrestha, Ashok K; Castignolles, Patrice; Gaborieau, Marianne

    2015-05-18

    About 80% of the Australian population consumes breakfast cereal (BC) at least five days a week. With high prevalence rates of obesity and other diet-related diseases, improved methods for monitoring sugar levels in breakfast cereals would be useful in nutrition research. The heterogeneity of the complex matrix of BCs can make carbohydrate analysis challenging or necessitate tedious sample preparation leading to potential sugar loss or starch degradation into sugars. A recently established, simple and robust free solution capillary electrophoresis (CE) method was used in a new application to 13 BCs (in Australia) and compared with several established methods for quantification of carbohydrates. Carbohydrates identified in BCs by CE included sucrose, maltose, glucose and fructose. The CE method is simple requiring no sample preparation or derivatization and carbohydrates are detected by direct UV detection. CE was shown to be a more robust and accurate method for measuring carbohydrates than Fehling method, DNS (3,5-dinitrosalicylic acid) assay and HPLC (high performance liquid chromatography).

  2. The Method of Manufactured Universes for validating uncertainty quantification methods

    KAUST Repository

    Stripling, H.F.

    2011-09-01

    The Method of Manufactured Universes is presented as a validation framework for uncertainty quantification (UQ) methodologies and as a tool for exploring the effects of statistical and modeling assumptions embedded in these methods. The framework calls for a manufactured reality from which experimental data are created (possibly with experimental error), an imperfect model (with uncertain inputs) from which simulation results are created (possibly with numerical error), the application of a system for quantifying uncertainties in model predictions, and an assessment of how accurately those uncertainties are quantified. The application presented in this paper manufactures a particle-transport universe, models it using diffusion theory with uncertain material parameters, and applies both Gaussian process and Bayesian MARS algorithms to make quantitative predictions about new experiments within the manufactured reality. The results of this preliminary study indicate that, even in a simple problem, the improper application of a specific UQ method or unrealized effects of a modeling assumption may produce inaccurate predictions. We conclude that the validation framework presented in this paper is a powerful and flexible tool for the investigation and understanding of UQ methodologies. © 2011 Elsevier Ltd. All rights reserved.

  3. Uncertainty quantification for large-scale ocean circulation predictions.

    Energy Technology Data Exchange (ETDEWEB)

    Safta, Cosmin; Debusschere, Bert J.; Najm, Habib N.; Sargsyan, Khachik

    2010-09-01

    Uncertainty quantificatio in climate models is challenged by the sparsity of the available climate data due to the high computational cost of the model runs. Another feature that prevents classical uncertainty analyses from being easily applicable is the bifurcative behavior in the climate data with respect to certain parameters. A typical example is the Meridional Overturning Circulation in the Atlantic Ocean. The maximum overturning stream function exhibits discontinuity across a curve in the space of two uncertain parameters, namely climate sensitivity and CO{sub 2} forcing. We develop a methodology that performs uncertainty quantificatio in the presence of limited data that have discontinuous character. Our approach is two-fold. First we detect the discontinuity location with a Bayesian inference, thus obtaining a probabilistic representation of the discontinuity curve location in presence of arbitrarily distributed input parameter values. Furthermore, we developed a spectral approach that relies on Polynomial Chaos (PC) expansions on each sides of the discontinuity curve leading to an averaged-PC representation of the forward model that allows efficient uncertainty quantification and propagation. The methodology is tested on synthetic examples of discontinuous data with adjustable sharpness and structure.

  4. Relative Quantification of Several Plasma Proteins during Liver Transplantation Surgery

    Science.gov (United States)

    Parviainen, Ville; Joenväärä, Sakari; Tukiainen, Eija; Ilmakunnas, Minna; Isoniemi, Helena; Renkonen, Risto

    2011-01-01

    Plasma proteome is widely used in studying changes occurring in human body during disease or other disturbances. Immunological methods are commonly used in such studies. In recent years, mass spectrometry has gained popularity in high-throughput analysis of plasma proteins. In this study, we tested whether mass spectrometry and iTRAQ-based protein quantification might be used in proteomic analysis of human plasma during liver transplantation surgery to characterize changes in protein abundances occurring during early graft reperfusion. We sampled blood from systemic circulation as well as blood entering and exiting the liver. After immunodepletion of six high-abundant plasma proteins, trypsin digestion, iTRAQ labeling, and cation-exchange fractionation, the peptides were analyzed by reverse phase nano-LC-MS/MS. In total, 72 proteins were identified of which 31 could be quantified in all patient specimens collected. Of these 31 proteins, ten, mostly medium-to-high abundance plasma proteins with a concentration range of 50–2000 mg/L, displayed relative abundance change of more than 10%. The changes in protein abundance observed in this study allow further research on the role of several proteins in ischemia-reperfusion injury during liver transplantation and possibly in other surgery. PMID:22187521

  5. Micelle Mediated Trace Level Sulfide Quantification through Cloud Point Extraction

    Directory of Open Access Journals (Sweden)

    Samrat Devaramani

    2012-01-01

    Full Text Available A simple cloud point extraction protocol has been proposed for the quantification of sulfide at trace level. The method is based on the reduction of iron (III to iron (II by the sulfide and the subsequent complexation of metal ion with nitroso-R salt in alkaline medium. The resulting green-colored complex was extracted through cloud point formation using cationic surfactant, that is, cetylpyridinium chloride, and the obtained surfactant phase was homogenized by ethanol before its absorbance measurement at 710 nm. The reaction variables like metal ion, ligand, surfactant concentration, and medium pH on the cloud point extraction of the metal-ligand complex have been optimized. The interference effect of the common anions and cations was studied. The proposed method has been successfully applied to quantify the trace level sulfide in the leachate samples of the landfill and water samples from bore wells and ponds. The validity of the proposed method has been studied by spiking the samples with known quantities of sulfide as well as comparing with the results obtained by the standard method.

  6. Uncertainty Quantification applied to flow simulations in thoracic aortic aneurysms

    Science.gov (United States)

    Boccadifuoco, Alessandro; Mariotti, Alessandro; Celi, Simona; Martini, Nicola; Salvetti, Maria Vittoria

    2015-11-01

    The thoracic aortic aneurysm is a progressive dilatation of the thoracic aorta causing a weakness in the aortic wall, which may eventually cause life-threatening events. Clinical decisions on treatment strategies are currently based on empiric criteria, like the aortic diameter value or its growth rate. Numerical simulations can give the quantification of important indexes which are impossible to be obtained through in-vivo measurements and can provide supplementary information. Hemodynamic simulations are carried out by using the open-source tool SimVascular and considering patient-specific geometries. One of the main issues in these simulations is the choice of suitable boundary conditions, modeling the organs and vessels not included in the computational domain. The current practice is to use outflow conditions based on resistance and capacitance, whose values are tuned to obtain a physiological behavior of the patient pressure. However it is not known a priori how this choice affects the results of the simulation. The impact of the uncertainties in these outflow parameters is investigated here by using the generalized Polynomial Chaos approach. This analysis also permits to calibrate the outflow-boundary parameters when patient-specific in-vivo data are available.

  7. Crystal Violet and XTT Assays on Staphylococcus aureus Biofilm Quantification.

    Science.gov (United States)

    Xu, Zhenbo; Liang, Yanrui; Lin, Shiqi; Chen, Dingqiang; Li, Bing; Li, Lin; Deng, Yang

    2016-10-01

    Staphylococcus aureus (S. Aureus) is a common food-borne pathogenic microorganism. Biofilm formation remains the major obstruction for bacterial elimination. The study aims at providing a basis for determining S. aureus biofilm formation. 257 clinical samples of S. aureus isolates were identified by routine analysis and multiplex PCR detection and found to contain 227 MRSA, 16 MSSA, 11 MRCNS, and 3 MSCNS strains. Two assays for quantification of S. aureus biofilm formation, the crystal violet (CV) assay and the XTT (tetrazolium salt reduction) assay, were optimized, evaluated, and further compared. In CV assay, most isolates formed weak biofilm 74.3 %), while the rest formed moderate biofilm (23.3 %) or strong biofilm (2.3 %). However, most isolates in XTT assay showed weak metabolic activity (77.0 %), while the rest showed moderate metabolic activity (17.9 %) or high metabolic activity (5.1 %). In this study, we found a distinct strain-to-strain dissimilarity in terms of both biomass formation and metabolic activity, and it was concluded from this study that two assays were mutual complementation rather than being comparison. PMID:27324342

  8. Quantification of mouse pulmonary cancer models by microcomputed tomography imaging

    International Nuclear Information System (INIS)

    The advances in preclinical cancer models, including orthotopic implantation models or genetically engineered mouse models of cancer, enable pursuing the molecular mechanism of cancer disease that might mimic genetic and biological processes in humans. Lung cancer is the major cause of cancer deaths; therefore, the treatment and prevention of lung cancer are expected to be improved by a better understanding of the complex mechanism of disease. In this study, we have examined the quantification of two distinct mouse lung cancer models by utilizing imaging modalities for monitoring tumor progression and drug efficacy evaluation. The utility of microcomputed tomography (micro-CT) for real-time/non-invasive monitoring of lung cancer progression has been confirmed by combining bioluminescent imaging and histopathological analyses. Further, we have developed a more clinically relevant lung cancer model by utilizing K-rasLSL-G12D/p53LSL-R270H mutant mice. Using micro-CT imaging, we monitored the development and progression of solitary lung tumor in K-rasLSL-G12D/p53LSL-R270H mutant mouse, and further demonstrated tumor growth inhibition by anticancer drug treatment. These results clearly indicate that imaging-guided evaluation of more clinically relevant tumor models would improve the process of new drug discovery and increase the probability of success in subsequent clinical studies. (author)

  9. Automated angiogenesis quantification through advanced image processing techniques.

    Science.gov (United States)

    Doukas, Charlampos N; Maglogiannis, Ilias; Chatziioannou, Aristotle; Papapetropoulos, Andreas

    2006-01-01

    Angiogenesis, the formation of blood vessels in tumors, is an interactive process between tumor, endothelial and stromal cells in order to create a network for oxygen and nutrients supply, necessary for tumor growth. According to this, angiogenic activity is considered a suitable method for both tumor growth or inhibition detection. The angiogenic potential is usually estimated by counting the number of blood vessels in particular sections. One of the most popular assay tissues to study the angiogenesis phenomenon is the developing chick embryo and its chorioallantoic membrane (CAM), which is a highly vascular structure lining the inner surface of the egg shell. The aim of this study was to develop and validate an automated image analysis method that would give an unbiased quantification of the micro-vessel density and growth in angiogenic CAM images. The presented method has been validated by comparing automated results to manual counts over a series of digital chick embryo photos. The results indicate the high accuracy of the tool, which has been thus extensively used for tumor growth detection at different stages of embryonic development. PMID:17946107

  10. Comprehensive quantification of ceramide species in human stratum corneum.

    Science.gov (United States)

    Masukawa, Yoshinori; Narita, Hirofumi; Sato, Hirayuki; Naoe, Ayano; Kondo, Naoki; Sugai, Yoshiya; Oba, Tsuyoshi; Homma, Rika; Ishikawa, Junko; Takagi, Yutaka; Kitahara, Takashi

    2009-08-01

    One of the key challenges in lipidomics is to quantify lipidomes of interest, as it is practically impossible to collect all authentic materials covering the targeted lipidomes. For diverse ceramides (CER) in human stratum corneum (SC) that play important physicochemical roles in the skin, we developed a novel method for quantification of the overall CER species by improving our previously reported profiling technique using normal-phase liquid chromatography-electrospray ionization-mass spectrometry (NPLC-ESI-MS). The use of simultaneous selected ion monitoring measurement of as many as 182 kinds of molecular-related ions enables the highly sensitive detection of the overall CER species, as they can be analyzed in only one SC-stripped tape as small as 5 mm x 10 mm. To comprehensively quantify CERs, including those not available as authentic species, we designed a procedure to estimate their levels using relative responses of representative authentic species covering the species targeted, considering the systematic error based on intra-/inter-day analyses. The CER levels obtained by this method were comparable to those determined by conventional thin-layer chromatography (TLC), which guarantees the validity of this method. This method opens lipidomics approaches for CERs in the SC. PMID:19349641

  11. Accurate 3D quantification of the bronchial parameters in MDCT

    Science.gov (United States)

    Saragaglia, A.; Fetita, C.; Preteux, F.; Brillet, P. Y.; Grenier, P. A.

    2005-08-01

    The assessment of bronchial reactivity and wall remodeling in asthma plays a crucial role in better understanding such a disease and evaluating therapeutic responses. Today, multi-detector computed tomography (MDCT) makes it possible to perform an accurate estimation of bronchial parameters (lumen and wall areas) by allowing a quantitative analysis in a cross-section plane orthogonal to the bronchus axis. This paper provides the tools for such an analysis by developing a 3D investigation method which relies on 3D reconstruction of bronchial lumen and central axis computation. Cross-section images at bronchial locations interactively selected along the central axis are generated at appropriate spatial resolution. An automated approach is then developed for accurately segmenting the inner and outer bronchi contours on the cross-section images. It combines mathematical morphology operators, such as "connection cost", and energy-controlled propagation in order to overcome the difficulties raised by vessel adjacencies and wall irregularities. The segmentation accuracy was validated with respect to a 3D mathematically-modeled phantom of a pair bronchus-vessel which mimics the characteristics of real data in terms of gray-level distribution, caliber and orientation. When applying the developed quantification approach to such a model with calibers ranging from 3 to 10 mm diameter, the lumen area relative errors varied from 3.7% to 0.15%, while the bronchus area was estimated with a relative error less than 5.1%.

  12. Detection and quantification of MS lesions using fuzzy topological principles

    Science.gov (United States)

    Udupa, Jayaram K.; Wei, Luogang; Samarasekera, Supun; Miki, Yukio; van Buchem, M. A.; Grossman, Robert I.

    1996-04-01

    Quantification of the severity of the multiple sclerosis (MS) disease through estimation of lesion volume via MR imaging is vital for understanding and monitoring the disease and its treatment. This paper presents a novel methodology and a system that can be routinely used for segmenting and estimating the volume of MS lesions via dual-echo spin-echo MR imagery. An operator indicates a few points in the images by pointing to the white matter, the gray matter, and the CSF. Each of these objects is then detected as a fuzzy connected set. The holes in the union of these objects correspond to potential lesion sites which are utilized to detect each potential lesion as a fuzzy connected object. These 3D objects are presented to the operator who indicates acceptance/rejection through the click of a mouse button. The volume of accepted lesions is then computed and output. Based on several evaluation studies and over 300 3D data sets that were processed, we conclude that the methodology is highly reliable and consistent, with a coefficient of variation (due to subjective operator actions) of less than 1.0% for volume.

  13. Uncertainty Quantification for Airfoil Icing using Polynomial Chaos Expansions

    CERN Document Server

    DeGennaro, Anthony M; Martinelli, Luigi

    2014-01-01

    The formation and accretion of ice on the leading edge of a wing can be detrimental to airplane performance. Complicating this reality is the fact that even a small amount of uncertainty in the shape of the accreted ice may result in a large amount of uncertainty in aerodynamic performance metrics (e.g., stall angle of attack). The main focus of this work concerns using the techniques of Polynomial Chaos Expansions (PCE) to quantify icing uncertainty much more quickly than traditional methods (e.g., Monte Carlo). First, we present a brief survey of the literature concerning the physics of wing icing, with the intention of giving a certain amount of intuition for the physical process. Next, we give a brief overview of the background theory of PCE. Finally, we compare the results of Monte Carlo simulations to PCE-based uncertainty quantification for several different airfoil icing scenarios. The results are in good agreement and confirm that PCE methods are much more efficient for the canonical airfoil icing un...

  14. Automated quantification of one-dimensional nanostructure alignment on surfaces

    Science.gov (United States)

    Dong, Jianjin; Goldthorpe, Irene A.; Mohieddin Abukhdeir, Nasser

    2016-06-01

    A method for automated quantification of the alignment of one-dimensional (1D) nanostructures from microscopy imaging is presented. Nanostructure alignment metrics are formulated and shown to be able to rigorously quantify the orientational order of nanostructures within a two-dimensional domain (surface). A complementary image processing method is also presented which enables robust processing of microscopy images where overlapping nanostructures might be present. Scanning electron microscopy (SEM) images of nanowire-covered surfaces are analyzed using the presented methods and it is shown that past single parameter alignment metrics are insufficient for highly aligned domains. Through the use of multiple parameter alignment metrics, automated quantitative analysis of SEM images is shown to be possible and the alignment characteristics of different samples are able to be quantitatively compared using a similarity metric. The results of this work provide researchers in nanoscience and nanotechnology with a rigorous method for the determination of structure/property relationships, where alignment of 1D nanostructures is significant.

  15. Quantification and prediction of rare events in nonlinear waves

    Science.gov (United States)

    Sapsis, Themistoklis; Cousins, Will; Mohamad, Mustafa

    2014-11-01

    The scope of this work is the quantification and prediction of rare events characterized by extreme intensity, in nonlinear dispersive models that simulate water waves. In particular we are interested for the understanding and the short-term prediction of rogue waves in the ocean and to this end, we consider 1-dimensional nonlinear models of the NLS type. To understand the energy transfers that occur during the development of an extreme event we perform a spatially localized analysis of the energy distribution along different wavenumbers by means of the Gabor transform. A stochastic analysis of the Gabor coefficients reveals i) the low-dimensionality of the intermittent structures, ii) the interplay between non-Gaussian statistical properties and nonlinear energy transfers between modes, as well as iii) the critical scales (or Gabor coefficients) where a critical energy can trigger the formation of an extreme event. The unstable character of these critical localized modes is analysed directly through the system equation and it is shown that it is defined as the result of the system nonlinearity and the wave dissipation (that mimics wave breaking). These unstable modes are randomly triggered through the dispersive ``heat bath'' of random waves that propagate in the nonlinear medium. Using these properties we formulate low-dimensional functionals of these Gabor coefficients that allow for the prediction of extreme event well before the strongly nonlinear interactions begin to occur. The prediction window is further enhanced by the combination of the developed scheme with traditional filtering schemes.

  16. Uranium quantification in semen by inductively coupled plasma mass spectrometry.

    Science.gov (United States)

    Todorov, Todor I; Ejnik, John W; Guandalini, Gustavo; Xu, Hanna; Hoover, Dennis; Anderson, Larry; Squibb, Katherine; McDiarmid, Melissa A; Centeno, Jose A

    2013-01-01

    In this study we report uranium analysis for human semen samples. Uranium quantification was performed by inductively coupled plasma mass spectrometry. No additives, such as chymotrypsin or bovine serum albumin, were used for semen liquefaction, as they showed significant uranium content. For method validation we spiked 2g aliquots of pooled control semen at three different levels of uranium: low at 5 pg/g, medium at 50 pg/g, and high at 1000 pg/g. The detection limit was determined to be 0.8 pg/g uranium in human semen. The data reproduced within 1.4-7% RSD and spike recoveries were 97-100%. The uranium level of the unspiked, pooled control semen was 2.9 pg/g of semen (n=10). In addition six semen samples from a cohort of Veterans exposed to depleted uranium (DU) in the 1991 Gulf War were analyzed with no knowledge of their exposure history. Uranium levels in the Veterans' semen samples ranged from undetectable (<0.8 pg/g) to 3350 pg/g. This wide concentration range for uranium in semen is consistent with known differences in current DU body burdens in these individuals, some of whom have retained embedded DU fragments.

  17. Accurate quantification of cells recovered by bronchoalveolar lavage.

    Science.gov (United States)

    Saltini, C; Hance, A J; Ferrans, V J; Basset, F; Bitterman, P B; Crystal, R G

    1984-10-01

    Quantification of the differential cell count and total number of cells recovered from the lower respiratory tract by bronchoalveolar lavage is a valuable technique for evaluating the alveolitis of patients with inflammatory disorders of the lower respiratory tract. The most commonly used technique for the evaluation of cells recovered by lavage has been to concentrate cells by centrifugation and then to determine total cell number using a hemocytometer and differential cell count from a Wright-Glemsa-stained cytocentrifuge preparation. However, we have noted that the percentage of small cells present in the original cell suspension recovered by lavage is greater than the percentage of lymphocytes identified on cytocentrifuge preparations. Therefore, we developed procedures for determining differential cell counts on lavage cells collected on Millipore filters and stained with hematoxylin-eosin (filter preparations) and compared the results of differential cell counts performed on filter preparations with those obtained using cytocentrifuge preparations. When cells recovered by lavage were collected on filter preparations, accurate differential cell counts were obtained, as confirmed by performing differential cell counts on cell mixtures of known composition, and by comparing differential cell counts obtained using filter preparations stained with hematoxylin-eosin with those obtained using filter preparations stained with a peroxidase cytochemical stain. The morphology of cells displayed on filter preparations was excellent, and interobserver variability in quantitating cell types recovered by lavage was less than 3%.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:6385789

  18. Pinpower uncertainty quantification of LWR-PROTEUS Phase III experiments

    International Nuclear Information System (INIS)

    CASMO-5MX is a set of Perl-based tools built around the lattice code CASMO-5 and developed at the Paul Scherrer Institut, used to perform sensitivity analysis (SA) and uncertainty quantification (UQ). The paper describes the recent developments to the CASMO-5MX methodology to perform pinpower UQ with the Stochastic Sampling methodology while perturbing nuclear data and technological parameters. The CASMO-5MX SS pinpower UQ is applied to the re-analysis of the LWR-PROTEUS Phase III campaign which featured SVEA-96 Optima2 fresh BWR fuel assemblies in an attempt to explain some discrepancy between the fission rate measurements (E) and their associated predictions with CASMO-5M (C). The pinpower UQ results have shown that the pinpower uncertainty due to nuclear data and technological parameters is in the range of 0.4% to 0.8%. This additional source of uncertainty, added to the existing experimental uncertainty, do not fully explain the remaining few C/E discrepancies in the pinpower map. The magnitude of nuclear data and technological parameters uncertainty is either underestimated or an additional source of uncertainty has not been taken into account. (author)

  19. Quantification of Covariance in Tropical Cyclone Activity across Teleconnected Basins

    Science.gov (United States)

    Tolwinski-Ward, S. E.; Wang, D.

    2015-12-01

    Rigorous statistical quantification of natural hazard covariance across regions has important implications for risk management, and is also of fundamental scientific interest. We present a multivariate Bayesian Poisson regression model for inferring the covariance in tropical cyclone (TC) counts across multiple ocean basins and across Saffir-Simpson intensity categories. Such covariability results from the influence of large-scale modes of climate variability on local environments that can alternately suppress or enhance TC genesis and intensification, and our model also simultaneously quantifies the covariance of TC counts with various climatic modes in order to deduce the source of inter-basin TC covariability. The model explicitly treats the time-dependent uncertainty in observed maximum sustained wind data, and hence the nominal intensity category of each TC. Differences in annual TC counts as measured by different agencies are also formally addressed. The probabilistic output of the model can be probed for probabilistic answers to such questions as: - Does the relationship between different categories of TCs differ statistically by basin? - Which climatic predictors have significant relationships with TC activity in each basin? - Are the relationships between counts in different basins conditionally independent given the climatic predictors, or are there other factors at play affecting inter-basin covariability? - How can a portfolio of insured property be optimized across space to minimize risk? Although we present results of our model applied to TCs, the framework is generalizable to covariance estimation between multivariate counts of natural hazards across regions and/or across peril types.

  20. In vivo cell tracking and quantification method in adult zebrafish

    Science.gov (United States)

    Zhang, Li; Alt, Clemens; Li, Pulin; White, Richard M.; Zon, Leonard I.; Wei, Xunbin; Lin, Charles P.

    2012-03-01

    Zebrafish have become a powerful vertebrate model organism for drug discovery, cancer and stem cell research. A recently developed transparent adult zebrafish using double pigmentation mutant, called casper, provide unparalleled imaging power in in vivo longitudinal analysis of biological processes at an anatomic resolution not readily achievable in murine or other systems. In this paper we introduce an optical method for simultaneous visualization and cell quantification, which combines the laser scanning confocal microscopy (LSCM) and the in vivo flow cytometry (IVFC). The system is designed specifically for non-invasive tracking of both stationary and circulating cells in adult zebrafish casper, under physiological conditions in the same fish over time. The confocal imaging part in this system serves the dual purposes of imaging fish tissue microstructure and a 3D navigation tool to locate a suitable vessel for circulating cell counting. The multi-color, multi-channel instrument allows the detection of multiple cell populations or different tissues or organs simultaneously. We demonstrate initial testing of this novel instrument by imaging vasculature and tracking circulating cells in CD41: GFP/Gata1: DsRed transgenic casper fish whose thrombocytes/erythrocytes express the green and red fluorescent proteins. Circulating fluorescent cell incidents were recorded and counted repeatedly over time and in different types of vessels. Great application opportunities in cancer and stem cell researches are discussed.

  1. Experimental model for civilian ballistic brain injury biomechanics quantification.

    Science.gov (United States)

    Zhang, Jiangyue; Yoganandan, Narayan; Pintar, Frank A; Guan, Yabo; Gennarelli, Thomas A

    2007-01-01

    Biomechanical quantification of projectile penetration using experimental head models can enhance the understanding of civilian ballistic brain injury and advance treatment. Two of the most commonly used handgun projectiles (25-cal, 275 m/s and 9 mm, 395 m/s) were discharged to spherical head models with gelatin and Sylgard simulants. Four ballistic pressure transducers recorded temporal pressure distributions at 308kHz, and temporal cavity dynamics were captured at 20,000 frames/second (fps) using high-speed digital video images. Pressures ranged from 644.6 to -92.8 kPa. Entry pressures in gelatin models were higher than exit pressures, whereas in Sylgard models entry pressures were lower or equivalent to exit pressures. Gelatin responded with brittle-type failure, while Sylgard demonstrated a ductile pattern through formation of micro-bubbles along projectile path. Temporary cavities in Sylgard models were 1.5-2x larger than gelatin models. Pressures in Sylgard models were more sensitive to projectile velocity and diameter increase, indicating Sylgard was more rate sensitive than gelatin. Based on failure patterns and brain tissue rate-sensitive characteristics, Sylgard was found to be an appropriate simulant. Compared with spherical projectile data, full-metal jacket (FMJ) projectiles produced different temporary cavity and pressures, demonstrating shape effects. Models using Sylgard gel and FMJ projectiles are appropriate to enhance understanding and mechanisms of ballistic brain injury.

  2. Quantification of brodifacoum in plasma and liver tissue by HPLC.

    Science.gov (United States)

    O'Bryan, S M; Constable, D J

    1991-01-01

    A simple high-performance liquid chromatographic method has been developed for detection and quantification of brodifacoum in plasma and liver tissue. After adding difenacoum as the internal standard, brodifacoum and difenacoum are extracted from 2 mL of plasma with two sequential 10-mL volumes of acetonitrile-ethyl ether (9:1) and from 2 g of liver tissue by grinding the tissue with 10 mL acetonitrile. The extracts are evaporated to dryness under nitrogen, 2 mL of acetonitrile is added to reconstitute the residues, and the resulting solution is analyzed using reversed-phase chromatography and fluorescence detection. The limits of detection for plasma and tissue are 2 micrograms/L and 5 ng/g, respectively. Using internal standardization, the mean intra-assay recovery from plasma is 92% and the mean inter-assay recoveries is 109%. The mean intra-assay and inter-assay recoveries from tissue are 96%. No interferences were observed with any of the following related compounds: brodifacoum, bromadiolone, coumarin, difenacoum, diphacinone, warfarin, and vitamin K1. PMID:1943058

  3. Relative quantification of several plasma proteins during liver transplantation surgery.

    Science.gov (United States)

    Parviainen, Ville; Joenväärä, Sakari; Tukiainen, Eija; Ilmakunnas, Minna; Isoniemi, Helena; Renkonen, Risto

    2011-01-01

    Plasma proteome is widely used in studying changes occurring in human body during disease or other disturbances. Immunological methods are commonly used in such studies. In recent years, mass spectrometry has gained popularity in high-throughput analysis of plasma proteins. In this study, we tested whether mass spectrometry and iTRAQ-based protein quantification might be used in proteomic analysis of human plasma during liver transplantation surgery to characterize changes in protein abundances occurring during early graft reperfusion. We sampled blood from systemic circulation as well as blood entering and exiting the liver. After immunodepletion of six high-abundant plasma proteins, trypsin digestion, iTRAQ labeling, and cation-exchange fractionation, the peptides were analyzed by reverse phase nano-LC-MS/MS. In total, 72 proteins were identified of which 31 could be quantified in all patient specimens collected. Of these 31 proteins, ten, mostly medium-to-high abundance plasma proteins with a concentration range of 50-2000 mg/L, displayed relative abundance change of more than 10%. The changes in protein abundance observed in this study allow further research on the role of several proteins in ischemia-reperfusion injury during liver transplantation and possibly in other surgery.

  4. Relative Quantification of Several Plasma Proteins during Liver Transplantation Surgery

    Directory of Open Access Journals (Sweden)

    Ville Parviainen

    2011-01-01

    Full Text Available Plasma proteome is widely used in studying changes occurring in human body during disease or other disturbances. Immunological methods are commonly used in such studies. In recent years, mass spectrometry has gained popularity in high-throughput analysis of plasma proteins. In this study, we tested whether mass spectrometry and iTRAQ-based protein quantification might be used in proteomic analysis of human plasma during liver transplantation surgery to characterize changes in protein abundances occurring during early graft reperfusion. We sampled blood from systemic circulation as well as blood entering and exiting the liver. After immunodepletion of six high-abundant plasma proteins, trypsin digestion, iTRAQ labeling, and cation-exchange fractionation, the peptides were analyzed by reverse phase nano-LC-MS/MS. In total, 72 proteins were identified of which 31 could be quantified in all patient specimens collected. Of these 31 proteins, ten, mostly medium-to-high abundance plasma proteins with a concentration range of 50–2000 mg/L, displayed relative abundance change of more than 10%. The changes in protein abundance observed in this study allow further research on the role of several proteins in ischemia-reperfusion injury during liver transplantation and possibly in other surgery.

  5. Atomic Resolution Imaging and Quantification of Chemical Functionality of Surfaces

    Energy Technology Data Exchange (ETDEWEB)

    Schwarz, Udo [Yale University

    2014-12-10

    The work carried out from 2006-2014 under DoE support was targeted at developing new approaches to the atomic-scale characterization of surfaces that include species-selective imaging and an ability to quantify chemical surface interactions with site-specific accuracy. The newly established methods were subsequently applied to gain insight into the local chemical interactions that govern the catalytic properties of model catalysts of interest to DoE. The foundation of our work was the development of three-dimensional atomic force microscopy (3D-AFM), a new measurement mode that allows the mapping of the complete surface force and energy fields with picometer resolution in space (x, y, and z) and piconewton/millielectron volts in force/energy. From this experimental platform, we further expanded by adding the simultaneous recording of tunneling current (3D-AFM/STM) using chemically well-defined tips. Through comparison with simulations, we were able to achieve precise quantification and assignment of local chemical interactions to exact positions within the lattice. During the course of the project, the novel techniques were applied to surface-oxidized copper, titanium dioxide, and silicon oxide. On these materials, defect-induced changes to the chemical surface reactivity and electronic charge density were characterized with site-specific accuracy.

  6. VESGEN Software for Mapping and Quantification of Vascular Regulators

    Science.gov (United States)

    Parsons-Wingerter, Patricia A.; Vickerman, Mary B.; Keith, Patricia A.

    2012-01-01

    VESsel GENeration (VESGEN) Analysis is an automated software that maps and quantifies effects of vascular regulators on vascular morphology by analyzing important vessel parameters. Quantification parameters include vessel diameter, length, branch points, density, and fractal dimension. For vascular trees, measurements are reported as dependent functions of vessel branching generation. VESGEN maps and quantifies vascular morphological events according to fractal-based vascular branching generation. It also relies on careful imaging of branching and networked vascular form. It was developed as a plug-in for ImageJ (National Institutes of Health, USA). VESGEN uses image-processing concepts of 8-neighbor pixel connectivity, skeleton, and distance map to analyze 2D, black-and-white (binary) images of vascular trees, networks, and tree-network composites. VESGEN maps typically 5 to 12 (or more) generations of vascular branching, starting from a single parent vessel. These generations are tracked and measured for critical vascular parameters that include vessel diameter, length, density and number, and tortuosity per branching generation. The effects of vascular therapeutics and regulators on vascular morphology and branching tested in human clinical or laboratory animal experimental studies are quantified by comparing vascular parameters with control groups. VESGEN provides a user interface to both guide and allow control over the users vascular analysis process. An option is provided to select a morphological tissue type of vascular trees, network or tree-network composites, which determines the general collections of algorithms, intermediate images, and output images and measurements that will be produced.

  7. Quantification of motility of carabid beetles in farmland.

    Science.gov (United States)

    Allema, A B; van der Werf, W; Groot, J C J; Hemerik, L; Gort, G; Rossing, W A H; van Lenteren, J C

    2015-04-01

    Quantification of the movement of insects at field and landscape levels helps us to understand their ecology and ecological functions. We conducted a meta-analysis on movement of carabid beetles (Coleoptera: Carabidae), to identify key factors affecting movement and population redistribution. We characterize the rate of redistribution using motility μ (L2 T-1), which is a measure for diffusion of a population in space and time that is consistent with ecological diffusion theory and which can be used for upscaling short-term data to longer time frames. Formulas are provided to calculate motility from literature data on movement distances. A field experiment was conducted to measure the redistribution of mass-released carabid, Pterostichus melanarius in a crop field, and derive motility by fitting a Fokker-Planck diffusion model using inverse modelling. Bias in estimates of motility from literature data is elucidated using the data from the field experiment as a case study. The meta-analysis showed that motility is 5.6 times as high in farmland as in woody habitat. Species associated with forested habitats had greater motility than species associated with open field habitats, both in arable land and woody habitat. The meta-analysis did not identify consistent differences in motility at the species level, or between clusters of larger and smaller beetles. The results presented here provide a basis for calculating time-varying distribution patterns of carabids in farmland and woody habitat. The formulas for calculating motility can be used for other taxa. PMID:25673121

  8. An improved method for the quantification of SOA bound peroxides

    Science.gov (United States)

    Mutzel, Anke; Rodigast, Maria; Iinuma, Yoshiteru; Böge, Olaf; Herrmann, Hartmut

    2013-03-01

    An improvement is made to a method for the quantification of SOA-bound peroxides. The procedure is based on an iodometric-spectrophotometric method that has been commonly used for the determination of peroxides in a wide range of biological and environmental samples. The improved method was applied to determine the peroxide content of laboratory-generated SOA from α-pinene ozonolysis. Besides main improvements for the detection conditions, the use of more environmentally sound solvents is considered instead of carcinogenic solvents. In addition to the improved method for peroxide determination, the present study provides evidence for artefact formation caused by ultrasonic agitation for the extraction of organic compounds in SOA filter samples. The concentration of SOA-bound peroxides in the extracts from ultrasonic agitation were up to three times higher than those from a laboratory orbital shaker under the same extraction conditions, indicating peroxide formation caused by acoustic cavitation during extraction. In contrast, pinic acid, terebic acid and terpenylic acid showed significantly lower concentrations in the sample extract prepared using ultrasonic agitation, indicating that these compounds react with OH radicals that are formed from acoustic cavitation. Great care should be taken when extracting SOA samples and the use of ultrasound should be avoided.

  9. Electrochemical Quantification of Single Nucleotide Polymorphisms Using Nanoparticle Probes

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Guodong; Lin, Yuehe

    2007-08-29

    We report a new approach for electrochemical quantification of single-nucleotide polymorphisms (SNPs) using nanoparticle probes. The principle is based on DNA polymerase I (klenow fragment)-induced coupling of the nucleotide-modified nanoparticle probe to the mutant sites of duplex DNA under the Watson-Crick base pairing rule. After liquid hybridization events occurred among biotinylated DNA probes, mutant DNA, and complementary DNA, the resulting duplex DNA helixes were captured to the surface of magnetic beads through a biotin-avidin affinity reaction and magnetic separation. A cadmium phosphate-loaded apoferritin nanoparticle probe, which is modified with nucleotides and is complementary to the mutant site, is coupled to the mutant sites of the formed duplex DNA in the presence of DNA polymerase. Subsequent electrochemical stripping analysis of the cadmium component of coupled nanoparticle probes provides a means to quantify the concentration of mutant DNA. The method is sensitive enough to detect 21.5 attomol mutant DNA, which will enable the quantitative analysis of nucleic acid without polymerase chain reaction pre-amplification. The approach was challenged with constructed samples containing mutant and complementary DNA. The results indicated that it was possible to accurately determine SNPs with frequencies as low 0.01. The proposed approach has a great potential for realizing an accurate, sensitive, rapid, and low-cost method of SNP detection.

  10. Quantification of Safety-Critical Software Test Uncertainty

    International Nuclear Information System (INIS)

    The method, conservatively assumes that the failure probability of a software for the untested inputs is 1, and the failure probability turns in 0 for successful testing of all test cases. However, in reality the chance of failure exists due to the test uncertainty. Some studies have been carried out to identify the test attributes that affect the test quality. Cao discussed the testing effort, testing coverage, and testing environment. Management of the test uncertainties was discussed in. In this study, the test uncertainty has been considered to estimate the software failure probability because the software testing process is considered to be inherently uncertain. A reliability estimation of software is very important for a probabilistic safety analysis of a digital safety critical system of NPPs. This study focused on the estimation of the probability of a software failure that considers the uncertainty in software testing. In our study, BBN has been employed as an example model for software test uncertainty quantification. Although it can be argued that the direct expert elicitation of test uncertainty is much simpler than BBN estimation, however the BBN approach provides more insights and a basis for uncertainty estimation

  11. Quantification of Safety-Critical Software Test Uncertainty

    Energy Technology Data Exchange (ETDEWEB)

    Khalaquzzaman, M.; Cho, Jaehyun; Lee, Seung Jun; Jung, Wondea [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2015-05-15

    The method, conservatively assumes that the failure probability of a software for the untested inputs is 1, and the failure probability turns in 0 for successful testing of all test cases. However, in reality the chance of failure exists due to the test uncertainty. Some studies have been carried out to identify the test attributes that affect the test quality. Cao discussed the testing effort, testing coverage, and testing environment. Management of the test uncertainties was discussed in. In this study, the test uncertainty has been considered to estimate the software failure probability because the software testing process is considered to be inherently uncertain. A reliability estimation of software is very important for a probabilistic safety analysis of a digital safety critical system of NPPs. This study focused on the estimation of the probability of a software failure that considers the uncertainty in software testing. In our study, BBN has been employed as an example model for software test uncertainty quantification. Although it can be argued that the direct expert elicitation of test uncertainty is much simpler than BBN estimation, however the BBN approach provides more insights and a basis for uncertainty estimation.

  12. Lung involvement quantification in chest radiographs; Quantificacao de comprometimento pulmonar em radiografias de torax

    Energy Technology Data Exchange (ETDEWEB)

    Giacomini, Guilherme; Alvarez, Matheus; Oliveira, Marcela de; Miranda, Jose Ricardo A. [Universidade Estadual Paulista Julio de Mesquita Filho (UNESP), Botucatu, SP (Brazil). Instituto de Biociencias. Departamento de Fisica e Biofisica; Pina, Diana R.; Pereira, Paulo C.M.; Ribeiro, Sergio M., E-mail: giacomini@ibb.unesp.br [Universidade Estadual Paulista Julio de Mesquita Filho (UNESP), Botucatu, SP (Brazil). Faculdade de Medicina. Departamento de Doencas Tropicais e Diagnostico por Imagem

    2014-12-15

    Tuberculosis (TB) caused by Mycobacterium tuberculosis, is an infectious disease which remains a global health problem. The chest radiography is the commonly method employed to assess the TB's evolution. The methods for quantification of abnormalities of chest are usually performed on CT scans (CT). This quantification is important to assess the TB evolution and treatment and comparing different treatments. However, precise quantification is not feasible for the amount of CT scans required. The purpose of this work is to develop a methodology for quantification of lung damage caused by TB through chest radiographs. It was developed an algorithm for computational processing of exams in Matlab, which creates a lungs' 3D representation, with compromised dilated regions inside. The quantification of lung lesions was also made for the same patients through CT scans. The measurements from the two methods were compared and resulting in strong correlation. Applying statistical Bland and Altman, all samples were within the limits of agreement, with a confidence interval of 95%. The results showed an average variation of around 13% between the two quantification methods. The results suggest the effectiveness and applicability of the method developed, providing better risk-benefit to the patient and cost-benefit ratio for the institution. (author)

  13. freeQuant: A Mass Spectrometry Label-Free Quantification Software Tool for Complex Proteome Analysis.

    Science.gov (United States)

    Deng, Ning; Li, Zhenye; Pan, Chao; Duan, Huilong

    2015-01-01

    Study of complex proteome brings forward higher request for the quantification method using mass spectrometry technology. In this paper, we present a mass spectrometry label-free quantification tool for complex proteomes, called freeQuant, which integrated quantification with functional analysis effectively. freeQuant consists of two well-integrated modules: label-free quantification and functional analysis with biomedical knowledge. freeQuant supports label-free quantitative analysis which makes full use of tandem mass spectrometry (MS/MS) spectral count, protein sequence length, shared peptides, and ion intensity. It adopts spectral count for quantitative analysis and builds a new method for shared peptides to accurately evaluate abundance of isoforms. For proteins with low abundance, MS/MS total ion count coupled with spectral count is included to ensure accurate protein quantification. Furthermore, freeQuant supports the large-scale functional annotations for complex proteomes. Mitochondrial proteomes from the mouse heart, the mouse liver, and the human heart were used to evaluate the usability and performance of freeQuant. The evaluation showed that the quantitative algorithms implemented in freeQuant can improve accuracy of quantification with better dynamic range.

  14. Method for Indirect Quantification of CH4 Production via H2O Production Using Hydrogenotrophic Methanogens.

    Science.gov (United States)

    Taubner, Ruth-Sophie; Rittmann, Simon K-M R

    2016-01-01

    Hydrogenotrophic methanogens are an intriguing group of microorganisms from the domain Archaea. Methanogens exhibit extraordinary ecological, biochemical, and physiological characteristics and possess a huge biotechnological potential. Yet, the only possibility to assess the methane (CH4) production potential of hydrogenotrophic methanogens is to apply gas chromatographic quantification of CH4. In order to be able to effectively screen pure cultures of hydrogenotrophic methanogens regarding their CH4 production potential we developed a novel method for indirect quantification of the volumetric CH4 production rate by measuring the volumetric water production rate. This method was established in serum bottles for cultivation of methanogens in closed batch cultivation mode. Water production was estimated by determining the difference in mass increase in a quasi-isobaric setting. This novel CH4 quantification method is an accurate and precise analytical technique, which can be used to rapidly screen pure cultures of methanogens regarding their volumetric CH4 evolution rate. It is a cost effective alternative determining CH4 production of methanogens over CH4 quantification by using gas chromatography, especially if applied as a high throughput quantification method. Eventually, the method can be universally applied for quantification of CH4 production from psychrophilic, thermophilic and hyperthermophilic hydrogenotrophic methanogens. PMID:27199898

  15. Hepatic Iron Quantification on 3 Tesla (3 T) Magnetic Resonance (MR): Technical Challenges and Solutions

    International Nuclear Information System (INIS)

    MR has become a reliable and noninvasive method of hepatic iron quantification. Currently, most of the hepatic iron quantification is performed on 1.5 T MR, and the biopsy measurements have been paired with R2 and R2* values for 1.5 T MR. As the use of 3 T MR scanners is steadily increasing in clinical practice, it has become important to evaluate the practicality of calculating iron burden at 3 T MR. Hepatic iron quantification on 3 T MR requires a better understanding of the process and more stringent technical considerations. The purpose of this work is to focus on the technical challenges in establishing a relationship between T2* values at 1.5 T MR and 3 T MR for hepatic iron concentration (HIC) and to develop an appropriately optimized MR protocol for the evaluation of T2* values in the liver at 3 T magnetic field strength. We studied 22 sickle cell patients using multiecho fast gradient-echo sequence (MFGRE) 3 T MR and compared the results with serum ferritin and liver biopsy results. Our study showed that the quantification of hepatic iron on 3 T MRI in sickle cell disease patients correlates well with clinical blood test results and biopsy results. 3 T MR liver iron quantification based on MFGRE can be used for hepatic iron quantification in transfused patients

  16. freeQuant: A Mass Spectrometry Label-Free Quantification Software Tool for Complex Proteome Analysis

    Directory of Open Access Journals (Sweden)

    Ning Deng

    2015-01-01

    Full Text Available Study of complex proteome brings forward higher request for the quantification method using mass spectrometry technology. In this paper, we present a mass spectrometry label-free quantification tool for complex proteomes, called freeQuant, which integrated quantification with functional analysis effectively. freeQuant consists of two well-integrated modules: label-free quantification and functional analysis with biomedical knowledge. freeQuant supports label-free quantitative analysis which makes full use of tandem mass spectrometry (MS/MS spectral count, protein sequence length, shared peptides, and ion intensity. It adopts spectral count for quantitative analysis and builds a new method for shared peptides to accurately evaluate abundance of isoforms. For proteins with low abundance, MS/MS total ion count coupled with spectral count is included to ensure accurate protein quantification. Furthermore, freeQuant supports the large-scale functional annotations for complex proteomes. Mitochondrial proteomes from the mouse heart, the mouse liver, and the human heart were used to evaluate the usability and performance of freeQuant. The evaluation showed that the quantitative algorithms implemented in freeQuant can improve accuracy of quantification with better dynamic range.

  17. Bacterial adhesion force quantification by fluidic force microscopy

    Science.gov (United States)

    Potthoff, Eva; Ossola, Dario; Zambelli, Tomaso; Vorholt, Julia A.

    2015-02-01

    Quantification of detachment forces between bacteria and substrates facilitates the understanding of the bacterial adhesion process that affects cell physiology and survival. Here, we present a method that allows for serial, single bacterial cell force spectroscopy by combining the force control of atomic force microscopy with microfluidics. Reversible bacterial cell immobilization under physiological conditions on the pyramidal tip of a microchanneled cantilever is achieved by underpressure. Using the fluidic force microscopy technology (FluidFM), we achieve immobilization forces greater than those of state-of-the-art cell-cantilever binding as demonstrated by the detachment of Escherichia coli from polydopamine with recorded forces between 4 and 8 nN for many cells. The contact time and setpoint dependence of the adhesion forces of E. coli and Streptococcus pyogenes, as well as the sequential detachment of bacteria out of a chain, are shown, revealing distinct force patterns in the detachment curves. This study demonstrates the potential of the FluidFM technology for quantitative bacterial adhesion measurements of cell-substrate and cell-cell interactions that are relevant in biofilms and infection biology.Quantification of detachment forces between bacteria and substrates facilitates the understanding of the bacterial adhesion process that affects cell physiology and survival. Here, we present a method that allows for serial, single bacterial cell force spectroscopy by combining the force control of atomic force microscopy with microfluidics. Reversible bacterial cell immobilization under physiological conditions on the pyramidal tip of a microchanneled cantilever is achieved by underpressure. Using the fluidic force microscopy technology (FluidFM), we achieve immobilization forces greater than those of state-of-the-art cell-cantilever binding as demonstrated by the detachment of Escherichia coli from polydopamine with recorded forces between 4 and 8 nN for many

  18. Christiansen Revisited: Rethinking Quantification of Uniformity in Rainfall Simulator Studies

    Science.gov (United States)

    Green, Daniel; Pattison, Ian

    2016-04-01

    Rainfall simulators, whether based within a laboratory or field setting are used extensively within a number of fields of research, including plot-scale runoff, infiltration and erosion studies, irrigation and crop management and scaled investigations into urban flooding. Rainfall simulators offer a number of benefits, including the ability to create regulated and repeatable rainfall characteristics (e.g. intensity, duration, drop size distribution and kinetic energy) without relying on unpredictable natural precipitation regimes. Ensuring and quantifying spatially uniform simulated rainfall across the entirety of the plot area is of particular importance to researchers undertaking rainfall simulation. As a result, numerous studies have focused on the quantification and improvement of uniformity values. Several statistical methods for the assessment of rainfall simulator uniformity have been developed. However, the Christiansen Uniformity Coefficient (CUC) suggested by Christiansen (1942) is most frequently used. Despite this, there is no set methodology and researchers can adapt or alter factors such as the quantity, as well as the spacing, distance and location of the measuring beakers used to derive CUC values. Because CUC values are highly sensitive to the resolution of the data, i.e. the number of observations taken, many densely distributed measuring containers subjected to the same experimental conditions may generate a significantly lower CUC value than fewer, more sparsely distributed measuring containers. Thus, the simulated rainfall under a higher resolution sampling method could appear less uniform than when using a coarser resolution sampling method, despite being derived from the same initial rainfall conditions. Expressing entire plot uniformity as a single, simplified percentage value disregards valuable qualitative information about plot uniformity, such as the small-scale spatial distribution of rainfall over the plot surface and whether these

  19. Streptavidin conjugation and quantification-a method evaluation for nanoparticles.

    Science.gov (United States)

    Quevedo, Pablo Darío; Behnke, Thomas; Resch-Genger, Ute

    2016-06-01

    Aiming at the development of validated protocols for protein conjugation of nanomaterials and the determination of protein labeling densities, we systematically assessed the conjugation of the model protein streptavidin (SAv) to 100-, 500-, and 1000-nm-sized polystyrene and silica nanoparticles and dye-encoded polymer particles with two established conjugation chemistries, based upon achievable coupling efficiencies and labeling densities. Bioconjugation reactions compared included EDC/sulfo NHS ester chemistry for direct binding of the SAv to carboxyl groups at the particle surface and maleimide-thiol chemistry in conjunction with heterobifunctional PEG linkers and aminated nanoparticles (NPs). Quantification of the total and functional amounts of SAv on these nanomaterials and unreacted SAv in solution was performed with the BCA assay and the biotin-FITC (BF) titration, relying on different signal generation principles, which are thus prone to different interferences. Our results revealed a clear influence of the conjugation chemistry on the amount of NP crosslinking, yet under optimized reaction conditions, EDC/sulfo NHS ester chemistry and the attachment via heterobifunctional PEG linkers led to comparably efficient SAv coupling and good labeling densities. Particle size can obviously affect protein labeling densities and particularly protein functionality, especially for larger particles. For unstained nanoparticles, direct bioconjugation seems to be the most efficient strategy, whereas for dye-encoded nanoparticles, PEG linkers are to be favored for the prevention of dye-protein interactions which can affect protein functionality specifically in the case of direct SAv binding. Moreover, an influence of particle size on achievable protein labeling densities and protein functionality could be demonstrated. PMID:27038055

  20. Semi-automated quantification of living cells with internalized nanostructures

    KAUST Repository

    Margineanu, Michael Bogdan

    2016-01-15

    Background Nanostructures fabricated by different methods have become increasingly important for various applications in biology and medicine, such as agents for medical imaging or cancer therapy. In order to understand their interaction with living cells and their internalization kinetics, several attempts have been made in tagging them. Although methods have been developed to measure the number of nanostructures internalized by the cells, there are only few approaches aimed to measure the number of cells that internalize the nanostructures, and they are usually limited to fixed-cell studies. Flow cytometry can be used for live-cell assays on large populations of cells, however it is a single time point measurement, and does not include any information about cell morphology. To date many of the observations made on internalization events are limited to few time points and cells. Results In this study, we present a method for quantifying cells with internalized magnetic nanowires (NWs). A machine learning-based computational framework, CellCognition, is adapted and used to classify cells with internalized and no internalized NWs, labeled with the fluorogenic pH-dependent dye pHrodo™ Red, and subsequently to determine the percentage of cells with internalized NWs at different time points. In a “proof-of-concept”, we performed a study on human colon carcinoma HCT 116 cells and human epithelial cervical cancer HeLa cells interacting with iron (Fe) and nickel (Ni) NWs. Conclusions This study reports a novel method for the quantification of cells that internalize a specific type of nanostructures. This approach is suitable for high-throughput and real-time data analysis and has the potential to be used to study the interaction of different types of nanostructures in live-cell assays.

  1. Local quantification of numerically-induced mixing and dissipation

    Science.gov (United States)

    Klingbeil, Knut; Mohammadi-Aragh, Mahdi; Gräwe, Ulf; Burchard, Hans

    2016-04-01

    The discretisation of the advection terms in transport equations introduces truncation errors in numerical models. These errors are usually associated with spurious diffusion, i.e. numerically-induced mixing of the advected quantities or dissipation of kinetic energy associated with the advection of momentum. Especially the numerically-induced diapycnal mixing part is very problematic for realistic model simulations. Since any diapycnal mixing of temperature and salinity increases the reference potential energy (RPE), numerically-induced mixing is often quantified in terms of RPE. However, this global bulk measure does not provide any information about the local amount of numerically-induced mixing of a single advected quantity. In this talk we will present a recently developed analysis method that quantifies the numerically-induced mixing of a single advected quantity locally (Klingbeil et al., 2014***). The method is based on the local tracer variance decay in terms of variance fluxes associated with the corresponding advective tracer fluxes. Because of its physically sound definition, this analysis method provides a reliable diagnostic tool, e.g., to assess the performance of advection schemes and to identify hotspots of numerically-induced mixing. At these identified positions the model could be adapted in terms of resolution or the applied numerical schemes. In this context we will demonstrate how numerically-induced mixing of temperature and salinity can be substantially reduced by vertical meshes adapting towards stratification. *** Klingbeil, K., M. Mohammadi-Aragh, U. Gräwe, H. Burchard (2014) . Quantification of spurious dissipation and mixing -- Discrete Variance Decay in a Finite-Volume framework. Ocean Modelling. doi:10.1016/j.ocemod.2014.06.001.

  2. Quantification of the adrenal cortex hormones with radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Badillo A, V.; Carrera D, A. A.; Ibarra M, C. M., E-mail: vbadillocren@hotmail.co [Universidad Autonoma de Zacatecas, Unidad Academica de Estudios Nucleares, Calle Cipres No. 10, Fracc. La Penuela, 98068 Zacatecas (Mexico)

    2010-10-15

    The pathologies of the adrenal cortex -adrenal insufficiency and Cushing syndrome- have their origin on the deficit or hypersecretion of some of the hormones that are secreted by the adrenal cortex, which is divided in three zones anatomically defined: the external zone, also called the zona glomerulosa, which is the main production site of aldosterone and mineralocorticoids; the internal zone, or zona reticularis, that produces androgens; and the external zone, or zone 1 orticotrop, which is responsible for producing glucocorticoids. In this work, a quantitative analysis of those hormones and their pathologic trigger was made; the quantification was made in the laboratory by means of highly sensitive and specific techniques, in this case, the radioimmunoassay, in which a radioisotope I-125 is used. This technique is based on the biochemical bond-type reaction, because it requires of a substance called the linker, which bonds to another called ligand. This reaction is also known as antigen-antibody (Ag-Ab), where the results of the reaction will depend on the quantity of antigen in the sample and on its affinity for the antibody. In this work, a 56 patients (of which 13 were men and 43 women) study was made. The cortisol, the ACTH, the androsterone and the DHEA values were very elevated in the majority of the cases corresponding to women, predominating cortisol; while in men, a notorious elevation of the 17 {alpha}-OH-PRG and of the DHEA-SO{sub 4} was observed. Based on that, we can conclude that 51 of them did not have mayor complications, because they just went to the laboratory once, while the remaining 5 had a medical monitoring, and they visited the laboratory more than one occasion, tell about a difficulty on their improvement. According to the results, an approximate relation of 8:2 women:men, respectively, becomes clear to the hormonal pathologies of the adrenal cortex. (Author)

  3. Quantification of ocular inflammation with technetium-99m glucoheptonate

    International Nuclear Information System (INIS)

    Histological and morphometric evaluation of ocular inflammation is difficult, particularly when there is extensive ocular involvement with abscess formation and necrosis. A quantitative imaging procedure applicable to humans would be important clinically. To establish such a procedure, turpentine-induced ocular inflammation was obtained by subconjunctival injection in the right eye of 55 rabbits. The left eye was used as control and injected with a volume of saline equal to the volume of turpentine in the right eye. Volumes of turpentine or saline were 0.02, 0.04, 0.06, 0.2 and 0.6 ml, and the rabbits were divided into groups 1-5, according to these volumes. Imaging was performed 48 h after turpentine injection and 6 h after intravenous injection of 10 mCi of technetium 99m glucoheptonate (99mTc-GH). An inflammatory reaction index (IRI), defined as the ratio of counts of the right eye divided by counts of the left eye, was used. IRIs were proportional to the degree of inflammation and allowed the distinction of 3 subgroups: One represented by group 4, one by group 5 and one by groups 1, 2 and 3. This method of quantification of ocular inflammatory processes using 99mTc-GH is original, rapid, non-invasive, reproducible and safe, although unable to differentiate inflammatory processes caused by doses of turpentine which are very small and close to each other. It is conceivable that its application to humans will bring new insight into the ocular inflammatory process and response to therapy. (orig.)

  4. Automated renal histopathology: digital extraction and quantification of renal pathology

    Science.gov (United States)

    Sarder, Pinaki; Ginley, Brandon; Tomaszewski, John E.

    2016-03-01

    The branch of pathology concerned with excess blood serum proteins being excreted in the urine pays particular attention to the glomerulus, a small intertwined bunch of capillaries located at the beginning of the nephron. Normal glomeruli allow moderate amount of blood proteins to be filtered; proteinuric glomeruli allow large amount of blood proteins to be filtered. Diagnosis of proteinuric diseases requires time intensive manual examination of the structural compartments of the glomerulus from renal biopsies. Pathological examination includes cellularity of individual compartments, Bowman's and luminal space segmentation, cellular morphology, glomerular volume, capillary morphology, and more. Long examination times may lead to increased diagnosis time and/or lead to reduced precision of the diagnostic process. Automatic quantification holds strong potential to reduce renal diagnostic time. We have developed a computational pipeline capable of automatically segmenting relevant features from renal biopsies. Our method first segments glomerular compartments from renal biopsies by isolating regions with high nuclear density. Gabor texture segmentation is used to accurately define glomerular boundaries. Bowman's and luminal spaces are segmented using morphological operators. Nuclei structures are segmented using color deconvolution, morphological processing, and bottleneck detection. Average computation time of feature extraction for a typical biopsy, comprising of ~12 glomeruli, is ˜69 s using an Intel(R) Core(TM) i7-4790 CPU, and is ~65X faster than manual processing. Using images from rat renal tissue samples, automatic glomerular structural feature estimation was reproducibly demonstrated for 15 biopsy images, which contained 148 individual glomeruli images. The proposed method holds immense potential to enhance information available while making clinical diagnoses.

  5. Uncertainty Quantification of Calculated Temperatures for the AGR-1 Experiment

    Energy Technology Data Exchange (ETDEWEB)

    Binh T. Pham; Jeffrey J. Einerson; Grant L. Hawkes

    2013-03-01

    This report documents an effort to quantify the uncertainty of the calculated temperature data for the first Advanced Gas Reactor (AGR-1) fuel irradiation experiment conducted in the INL’s Advanced Test Reactor (ATR) in support of the Next Generation Nuclear Plant (NGNP) R&D program. Recognizing uncertainties inherent in physics and thermal simulations of the AGR-1 test, the results of the numerical simulations can be used in combination with the statistical analysis methods to improve qualification of measured data. Additionally, the temperature simulation data for AGR tests can be used for validation of the fuel transport and fuel performance simulation models. The crucial roles of the calculated fuel temperatures in ensuring achievement of the AGR experimental program objectives require accurate determination of the model temperature uncertainties. The report is organized into three chapters. Chapter 1 introduces the AGR Fuel Development and Qualification program and provides overviews of AGR-1 measured data, AGR-1 test configuration and test procedure, and thermal simulation. Chapters 2 describes the uncertainty quantification procedure for temperature simulation data of the AGR-1 experiment, namely, (i) identify and quantify uncertainty sources; (ii) perform sensitivity analysis for several thermal test conditions; (iii) use uncertainty propagation to quantify overall response temperature uncertainty. A set of issues associated with modeling uncertainties resulting from the expert assessments are identified. This also includes the experimental design to estimate the main effects and interactions of the important thermal model parameters. Chapter 3 presents the overall uncertainty results for the six AGR-1 capsules. This includes uncertainties for the daily volume-average and peak fuel temperatures, daily average temperatures at TC locations, and time-average volume-average and time-average peak fuel temperatures.

  6. Uncertainty Quantification of Calculated Temperatures for the AGR-1 Experiment

    Energy Technology Data Exchange (ETDEWEB)

    Binh T. Pham; Jeffrey J. Einerson; Grant L. Hawkes

    2012-04-01

    This report documents an effort to quantify the uncertainty of the calculated temperature data for the first Advanced Gas Reactor (AGR-1) fuel irradiation experiment conducted in the INL's Advanced Test Reactor (ATR) in support of the Next Generation Nuclear Plant (NGNP) R&D program. Recognizing uncertainties inherent in physics and thermal simulations of the AGR-1 test, the results of the numerical simulations can be used in combination with the statistical analysis methods to improve qualification of measured data. Additionally, the temperature simulation data for AGR tests can be used for validation of the fuel transport and fuel performance simulation models. The crucial roles of the calculated fuel temperatures in ensuring achievement of the AGR experimental program objectives require accurate determination of the model temperature uncertainties. The report is organized into three chapters. Chapter 1 introduces the AGR Fuel Development and Qualification program and provides overviews of AGR-1 measured data, AGR-1 test configuration and test procedure, and thermal simulation. Chapters 2 describes the uncertainty quantification procedure for temperature simulation data of the AGR-1 experiment, namely, (i) identify and quantify uncertainty sources; (ii) perform sensitivity analysis for several thermal test conditions; (iii) use uncertainty propagation to quantify overall response temperature uncertainty. A set of issues associated with modeling uncertainties resulting from the expert assessments are identified. This also includes the experimental design to estimate the main effects and interactions of the important thermal model parameters. Chapter 3 presents the overall uncertainty results for the six AGR-1 capsules. This includes uncertainties for the daily volume-average and peak fuel temperatures, daily average temperatures at TC locations, and time-average volume-average and time-average peak fuel temperatures.

  7. Direct quantification of negatively charged functional groups on membrane surfaces

    KAUST Repository

    Tiraferri, Alberto

    2012-02-01

    Surface charge plays an important role in membrane-based separations of particulates, macromolecules, and dissolved ionic species. In this study, we present two experimental methods to determine the concentration of negatively charged functional groups at the surface of dense polymeric membranes. Both techniques consist of associating the membrane surface moieties with chemical probes, followed by quantification of the bound probes. Uranyl acetate and toluidine blue O dye, which interact with the membrane functional groups via complexation and electrostatic interaction, respectively, were used as probes. The amount of associated probes was quantified using liquid scintillation counting for uranium atoms and visible light spectroscopy for the toluidine blue dye. The techniques were validated using self-assembled monolayers of alkanethiols with known amounts of charged moieties. The surface density of negatively charged functional groups of hand-cast thin-film composite polyamide membranes, as well as commercial cellulose triacetate and polyamide membranes, was quantified under various conditions. Using both techniques, we measured a negatively charged functional group density of 20-30nm -2 for the hand-cast thin-film composite membranes. The ionization behavior of the membrane functional groups, determined from measurements with toluidine blue at varying pH, was consistent with published data for thin-film composite polyamide membranes. Similarly, the measured charge densities on commercial membranes were in general agreement with previous investigations. The relative simplicity of the two methods makes them a useful tool for quantifying the surface charge concentration of a variety of surfaces, including separation membranes. © 2011 Elsevier B.V.

  8. Robust quantification of polymerase chain reactions using global fitting.

    Directory of Open Access Journals (Sweden)

    Ana C Carr

    Full Text Available BACKGROUND: Quantitative polymerase chain reactions (qPCR are used to monitor relative changes in very small amounts of DNA. One drawback to qPCR is reproducibility: measuring the same sample multiple times can yield data that is so noisy that important differences can be dismissed. Numerous analytical methods have been employed that can extract the relative template abundance between samples. However, each method is sensitive to baseline assignment and to the unique shape profiles of individual reactions, which gives rise to increased variance stemming from the analytical procedure itself. PRINCIPAL FINDINGS: We developed a simple mathematical model that accurately describes the entire PCR reaction profile using only two reaction variables that depict the maximum capacity of the reaction and feedback inhibition. This model allows quantification that is more accurate than existing methods and takes advantage of the brighter fluorescence signals from later cycles. Because the model describes the entire reaction, the influences of baseline adjustment errors, reaction efficiencies, template abundance, and signal loss per cycle could be formalized. We determined that the common cycle-threshold method of data analysis introduces unnecessary variance because of inappropriate baseline adjustments, a dynamic reaction efficiency, and also a reliance on data with a low signal-to-noise ratio. SIGNIFICANCE: Using our model, fits to raw data can be used to determine template abundance with high precision, even when the data contains baseline and signal loss defects. This improvement reduces the time and cost associated with qPCR and should be applicable in a variety of academic, clinical, and biotechnological settings.

  9. Quantification of Structural Isomers via Mode-Selective Irmpd

    Science.gov (United States)

    Polfer, Nicolas C.

    2016-06-01

    Mixtures of structural isomers can pose a challenge for vibrational ion spectroscopy. In cases where particular structures display diagnostic vibrations, these structures can be selectively "burned away". In ion traps, the ion population can be subjected to multiple laser shots, in order to fully deplete a particular structure, in effect allowing a quantification of this structure. Protonated para-amino benzoic acid (PABA) serves as an illustrative example. PABA is known to preferentially exist in the N-protonated (N-prot) form in solution, but in the gas phase it is energetically favorable in the O-protonated (O-prot) form. As shown in Figure 1, the N-prot structure can be kinetically trapped in the gas phase when sprayed from non-protic solvent, whereas the O-prot structure is obtained when sprayed from protic solvents, analogous to results by others [1,2]. y parking the light source on the diagnostic 3440 wn mode, the percentage of the O-prot structure can be determined, and by default the remainder is assumed to adopt the N-prot structure. It will be shown that the relative percentages of O-prot vs N-prot are highly dependent on the solvent mixture, going from close to 0% O-prot in non-protic solvents, to 99% in protic solvents. Surprisingly, water behaves much more like a non-protic solvent than methanol. It is observed that the capillary temperature, which aids droplet desolvation by black-body radiation in the ESI source, is critical to promote the appearance of O-prot structures. These results are consistent with the picture that a protic bridge mechanism is at play to facilitate proton transfer, and thus allow conversion from N-prot to O-prot, but that this mechanism is subject to appreciable kinetic barriers on the timescale of solvent evaporation. 1. J. Phys. Chem. A 2011, 115, 7625. 2. Anal. Chem. 2012, 84, 7857.

  10. Quantification of glacial till chemical composition by reflectance spectroscopy

    International Nuclear Information System (INIS)

    Chemometric modelling of soil element concentrations from diffuse visible and near-infrared (VSWIR, 350–2500 nm) reflectance spectroscopic measurements holds potential for soil element analyses. Research has demonstrated it particularly for organic agricultural soils, yet little is known about the VSWIR response of glacial tills. Soils with low organic matter content developed on unstratified glacial materials were studied at two geologically similar sites on the mafic metavolcanic rocks of the Lapland Greenstone belt in northern Finland. The till samples (n = 217) were composed primarily of quartz, plagioclase and amphibole having less than 3% of clinochlore, talc and illite. VSWIR spectra of mineral powder (2 = 0.80–0.89) of several soil chemical elements such as Al (validation RMSE 1802 mg kg−1), Ba (5.85 mg kg−1), Co (0.86 mg kg−1), Cr (6.94 mg kg−1), Cu (2.54 mg kg−1), Fe (2088 mg kg−1), Mg (449.6 mg kg−1), Mn (0.82 mg kg−1), Ni (3.24 mg kg−1), V (4.88 mg kg−1), and Zn (0.80 mg kg−1). The electronic and vibrational molecular processes causing absorption might be responsible for accurate predictions of major elements such as Al, Fe and Mg. However, the concentrations of other major and trace elements could be predicted by the PLSR because they were cross-correlated to spectrally active soil elements or extraneous soil properties. Therefore, the applicability of the results is highly sample set specific. Further, the results show that in local scale studies at geologically fairly homogenous areas the limited spread of the data may restrict the use of the spectroscopic–chemometric approach. This paper demonstrates the capability of laboratory VSWIR spectroscopy for determining element concentrations of glacial tills. Further work should focus on overcoming the issues of sampling scale and understanding the causality for cross-correlation in quantification of the elements.

  11. Automated quantification reveals hyperglycemia inhibits endothelial angiogenic function.

    Directory of Open Access Journals (Sweden)

    Anthony R Prisco

    Full Text Available Diabetes Mellitus (DM has reached epidemic levels globally. A contributing factor to the development of DM is high blood glucose (hyperglycemia. One complication associated with DM is a decreased angiogenesis. The Matrigel tube formation assay (TFA is the most widely utilized in vitro assay designed to assess angiogenic factors and conditions. In spite of the widespread use of Matrigel TFAs, quantification is labor-intensive and subjective, often limiting experiential design and interpretation of results. This study describes the development and validation of an open source software tool for high throughput, morphometric analysis of TFA images and the validation of an in vitro hyperglycemic model of DM.Endothelial cells mimic angiogenesis when placed onto a Matrigel coated surface by forming tube-like structures. The goal of this study was to develop an open-source software algorithm requiring minimal user input (Pipeline v1.3 to automatically quantify tubular metrics from TFA images. Using Pipeline, the ability of endothelial cells to form tubes was assessed after culture in normal or high glucose for 1 or 2 weeks. A significant decrease in the total tube length and number of branch points was found when comparing groups treated with high glucose for 2 weeks versus normal glucose or 1 week of high glucose.Using Pipeline, it was determined that hyperglycemia inhibits formation of endothelial tubes in vitro. Analysis using Pipeline was more accurate and significantly faster than manual analysis. The Pipeline algorithm was shown to have additional applications, such as detection of retinal vasculature.

  12. Quantification of radionuclide uptake levels for primary bone tumors

    Directory of Open Access Journals (Sweden)

    Hasford Francis

    2015-04-01

    Full Text Available The purpose of the study is to quantify the level of uptake of administered radionuclide in primary bone tumors for patients undergoing bone scintigraphy. Retrospective study on 48 patient's scintigrams to quantify the uptake levels of administered radiopharmaceuticals was performed in a nuclear medicine unit in Ghana. Patients were administered with activity ranging between 0.555 and 1.110 MBq (15–30 mCi, and scanned on Siemens e.cam SPECT system. Analyses on scintigrams were performed with Image J software by drawing regions of interest (ROIs over identified hot spots (pathologic sites. Nine skeletal parts namely cranium, neck, shoulder, sacrum, sternum, vertebra, femur, ribcage, and knee were considered in the study, which involved 96 identified primary tumors. Radionuclide uptakes were quantified in terms of the estimated counts of activity per patient for identified tumor sites. Average normalized counts of activity (nGMC per patient ranged from 5.2759 ± 0.6590 cts/mm2/MBq in the case of cranium tumors to 72.7569 ± 17.8786 cts/mm2/MBq in the case of ribcage tumors. The differences in uptake levels could be attributed to different mechanisms of Tc-99m MDP uptake in different types of bones, which is directly related to blood flow and degree of osteoblastic activity. The overall normalized count of activity for the 96 identified tumors was estimated to be 23.0350 ± 19.5424 cts/mm2/MBq. The study revealed highest uptake of activity in ribcage and least uptake in cranium. Quantification of radionuclide uptakes in tumors is important and recommended in assessing patient's response to therapy, doses to critical organs and in diagnosing tumors.

  13. Quantification of leukocyte migration: improvement of a method.

    Science.gov (United States)

    Sunder-Plassmann, G; Hofbauer, R; Sengoelge, G; Hörl, W H

    1996-01-01

    Eighteen different permeable membrane supports with and without confluent endothelial cell monolayers were incubated with normal donor derived neutrophils in the upper chambers of a 24 multiwell double chamber system. In order to study transmembrane or transendothelial leukocyte migration leukocytes were stimulated by chemoattractants, or endothelial cells were activated by IL-1. After coincubation the membrane supports building the upper chambers were discarded. Using this technique, leukocytes that had migrated into the lower chamber were exposed to the fluorescent dye calcein AM without additional washing or transfer steps. Absolute cell counts were determined computer assisted using dilution series of calcein AM labeled leukocytes as standards. Serial dilutions of neutrophils exposed to calcein AM showed reproducible linear fluorescence intensity, and relative fluorescence intensity correlated significant with cell counts (r2 = 0.974, p polyethylene terephtalate with a pore size of 3 mm at a pore density of 0.8 x 10(6)/cm2. Stimulation of leukocytes or endothelium by FMLP or IL-1 revealed an increase of transendothelial migration to 7.2 +/- 1.8 x 10(5) PMN and 5.1 +/- 0.7 x 10(5) PMN respectively if compared with medium (0.6 +/- 0.2 x 10(5) PMN). IL-1 induced migration of neutrophils was inhibited by anti IL-1 autoantibodies derived from chronic renal failure patients (IL-1: 100% of PMN migrated, anti IL-1 antibody: 39% of PMN migrated, control antibody: 84% of PMN migrated). In summary, a simple fluorimetric assay was established for the quantification of transmembrane and transendothelial leukocyte migration. PMID:8675234

  14. Combination radioimmunotherapy approaches and quantification of immuno-PET

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jin Su [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2016-06-15

    Monoclonal antibodies (mAbs), which play a prominent role in cancer therapy, can interact with specific antigens on cancer cells, thereby enhancing the patient' immune response via various mechanisms, or mAbs can act against cell growth factors and, thereby, arrest the proliferation of tumor cells. Radionuclide-labeled mAbs, which are used in radioimmunotherapy (RIT), are effective for cancer treatment because tumor associated-mAbs linked to cytotoxic radionuclides can selectively bind to tumor antigens and release targeted cytotoxic radiation. Immunological positron emission tomography (immuno-PET), which is the combination of PET with mAb, is an attractive option for improving tumor detection and mAb quantification. However, RIT remains a challenge because of the limited delivery of mAb into tumors. The transport and uptake of mAb into tumors is slow and heterogeneous. The tumor microenvironment contributed to the limited delivery of the mAb. During the delivery process of mAb to tumor, mechanical drug resistance such as collagen distribution or physiological drug resistance such as high intestinal pressure or absence of lymphatic vessel would be the limited factor of mAb delivery to the tumor at a potentially lethal mAb concentration. When α-emitter-labeled mAbs were used, deeper penetration of α-emitter-labeled mAb inside tumors was more important because of the short range of the α emitter. Therefore, combination therapy strategies aimed at improving mAb tumor penetration and accumulation would be beneficial for maximizing their therapeutic efficacy against solid tumors.

  15. Towards the quantification of rockfall risk assessment for urban areas

    Science.gov (United States)

    Mavrouli, Olga; Corominas, Jordi

    2010-05-01

    In many mountainous inhabited areas rockfalls are a major threat for structures and population. The quantification of the risk gives an estimate of the potential consequences that allows the analysis of different scenarios, minimizing the subjectivity and the uncertainties that derive from judgmental and qualitative approaches. The four main phases of the rockfall phenomenon have to be determined including: a. the calculation of the frequency of the rock block volumes falling down the slope, b. the calculation of the probability of the rock blocks reaching a reference section with a certain level of kinetic energy; c. the calculation of the spatio-temporal probability of the exposed elements; and d. the calculation of the probability that an exposed element will suffer a certain degree of damage. Here, a step-by-step methodology for the quantification of risk is presented. The methodology focuses on steps (b) to (d). An example of an urban area that is situated at the toe of a talus cone below of a rocky slope is considered. Three different rock diameters are considered with their respective frequencies (step a). For the calculation of the spatial probability of a given rock size reaching a location, a probabilistic 3D trajectory analysis is performed using the software ROTOMAP. The inputs are the topographic relief, the rockfall source and velocity and the soil parameters (restitution coefficient and friction coefficients). The latter are evaluated by back analysis using historical events. The probability of a given rock magnitude reaching a critical section of the talus cone with a certain level of kinetic energy is evaluated. For the step (c), the spatio-temporal probability of the element at risk is calculated taking into account both the trajectographic analysis of the rock blocks and the location of the elements at risk on the talus cone. For the step (d), the probability of a certain degree of structural damage in the buildings is calculated. To this purpose

  16. A statistical modeling approach to computer-aided quantification of dental biofilm.

    Science.gov (United States)

    Mansoor, Awais; Patsekin, Valery; Scherl, Dale; Robinson, J Paul; Rajwa, Bartlomiej

    2015-01-01

    Biofilm is a formation of microbial material on tooth substrata. Several methods to quantify dental biofilm coverage have recently been reported in the literature, but at best they provide a semiautomated approach to quantification with significant input from a human grader that comes with the grader's bias of what is foreground, background, biofilm, and tooth. Additionally,human assessment indices limit the resolution of the quantification scale; most commercial scales use five levels of quantification for biofilm coverage (0%, 25%, 50%, 75%, and 100%). On the other hand, current state-of-the-art techniques in automatic plaque quantification fail to make their way into practical applications owing to their inability to incorporate human input to handle misclassifications. This paper proposes a new interactive method for biofilm quantification in Quantitative light-induced fluorescence(QLF) images of canine teeth that is independent of the perceptual bias of the grader. The method partitions a QLF image into segments of uniform texture and intensity called superpixels; every superpixel is statistically modeled as a realization of a single 2-D Gaussian Markov random field (GMRF) whose parameters are estimated; the superpixel is then assigned to one of three classes (background, biofilm, tooth substratum) based on the training set of data. The quantification results show a high degree of consistency and precision. At the same time, the proposed method gives pathologists full control to postprocess the automatic quantification by flipping misclassified superpixels to a different state (background,tooth, biofilm) with a single click, providing greater usability than simply marking the boundaries of biofilm and tooth as done by current state-of-the-art methods.

  17. Elemental labelling combined with liquid chromatography inductively coupled plasma mass spectrometry for quantification of biomolecules: A review

    OpenAIRE

    Kretschy, Daniela; Koellensperger, Gunda; Hann, Stephan

    2012-01-01

    This article reviews novel quantification concepts where elemental labelling is combined with flow injection inductively coupled plasma mass spectrometry (FI-ICP-MS) or liquid chromatography inductively coupled plasma mass spectrometry (LC–ICP-MS), and employed for quantification of biomolecules such as proteins, peptides and related molecules in challenging sample matrices. In the first sections an overview on general aspects of biomolecule quantification, as well as of labelling will be pre...

  18. Pore REconstruction and Segmentation (PORES) method for improved porosity quantification of nanoporous materials

    Energy Technology Data Exchange (ETDEWEB)

    Van Eyndhoven, G., E-mail: geert.vaneyndhoven@uantwerpen.be [iMinds-Vision Lab, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk (Belgium); Kurttepeli, M. [EMAT, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp (Belgium); Van Oers, C.J.; Cool, P. [Laboratory of Adsorption and Catalysis, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk (Belgium); Bals, S. [EMAT, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp (Belgium); Batenburg, K.J. [iMinds-Vision Lab, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk (Belgium); Centrum Wiskunde and Informatica, Science Park 123, NL-1090 GB Amsterdam (Netherlands); Mathematical Institute, Universiteit Leiden, Niels Bohrweg 1, NL-2333 CA Leiden (Netherlands); Sijbers, J. [iMinds-Vision Lab, University of Antwerp, Universiteitsplein 1, B-2610 Wilrijk (Belgium)

    2015-01-15

    Electron tomography is currently a versatile tool to investigate the connection between the structure and properties of nanomaterials. However, a quantitative interpretation of electron tomography results is still far from straightforward. Especially accurate quantification of pore-space is hampered by artifacts introduced in all steps of the processing chain, i.e., acquisition, reconstruction, segmentation and quantification. Furthermore, most common approaches require subjective manual user input. In this paper, the PORES algorithm “POre REconstruction and Segmentation” is introduced; it is a tailor-made, integral approach, for the reconstruction, segmentation, and quantification of porous nanomaterials. The PORES processing chain starts by calculating a reconstruction with a nanoporous-specific reconstruction algorithm: the Simultaneous Update of Pore Pixels by iterative REconstruction and Simple Segmentation algorithm (SUPPRESS). It classifies the interior region to the pores during reconstruction, while reconstructing the remaining region by reducing the error with respect to the acquired electron microscopy data. The SUPPRESS reconstruction can be directly plugged into the remaining processing chain of the PORES algorithm, resulting in accurate individual pore quantification and full sample pore statistics. The proposed approach was extensively validated on both simulated and experimental data, indicating its ability to generate accurate statistics of nanoporous materials. - Highlights: • An electron tomography reconstruction/segmentation method for nanoporous materials. • The method exploits the porous nature of the scanned material. • Validated extensively on both simulation and real data experiments. • Results in increased image resolution and improved porosity quantification.

  19. RNAontheBENCH: computational and empirical resources for benchmarking RNAseq quantification and differential expression methods

    KAUST Repository

    Germain, Pierre-Luc

    2016-06-20

    RNA sequencing (RNAseq) has become the method of choice for transcriptome analysis, yet no consensus exists as to the most appropriate pipeline for its analysis, with current benchmarks suffering important limitations. Here, we address these challenges through a rich benchmarking resource harnessing (i) two RNAseq datasets including ERCC ExFold spike-ins; (ii) Nanostring measurements of a panel of 150 genes on the same samples; (iii) a set of internal, genetically-determined controls; (iv) a reanalysis of the SEQC dataset; and (v) a focus on relative quantification (i.e. across-samples). We use this resource to compare different approaches to each step of RNAseq analysis, from alignment to differential expression testing. We show that methods providing the best absolute quantification do not necessarily provide good relative quantification across samples, that count-based methods are superior for gene-level relative quantification, and that the new generation of pseudo-alignment-based software performs as well as established methods, at a fraction of the computing time. We also assess the impact of library type and size on quantification and differential expression analysis. Finally, we have created a R package and a web platform to enable the simple and streamlined application of this resource to the benchmarking of future methods.

  20. Sulfur-based absolute quantification of proteins using isotope dilution inductively coupled plasma mass spectrometry

    Science.gov (United States)

    Lee, Hyun-Seok; Heun Kim, Sook; Jeong, Ji-Seon; Lee, Yong-Moon; Yim, Yong-Hyeon

    2015-10-01

    An element-based reductive approach provides an effective means of realizing International System of Units (SI) traceability for high-purity biological standards. Here, we develop an absolute protein quantification method using double isotope dilution (ID) inductively coupled plasma mass spectrometry (ICP-MS) combined with microwave-assisted acid digestion for the first time. We validated the method and applied it to certify the candidate protein certified reference material (CRM) of human growth hormone (hGH). The concentration of hGH was determined by analysing the total amount of sulfur in hGH. Next, the size-exclusion chromatography method was used with ICP-MS to characterize and quantify sulfur-containing impurities. By subtracting the contribution of sulfur-containing impurities from the total sulfur content in the hGH CRM, we obtained a SI-traceable certification value. The quantification result obtained with the present method based on sulfur analysis was in excellent agreement with the result determined via a well-established protein quantification method based on amino acid analysis using conventional acid hydrolysis combined with an ID liquid chromatography-tandem mass spectrometry. The element-based protein quantification method developed here can be generally used for SI-traceable absolute quantification of proteins, especially pure-protein standards.