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Sample records for real-time quantitative analysis

  1. Quantitative analysis of real-time radiographic systems

    International Nuclear Information System (INIS)

    Barker, M.D.; Condon, P.E.; Barry, R.C.; Betz, R.A.; Klynn, L.M.

    1988-01-01

    A method was developed which yields quantitative information on the spatial resolution, contrast sensitivity, image noise, and focal spot size from real time radiographic images. The method uses simple image quality indicators and computer programs which make it possible to readily obtain quantitative performance measurements of single or multiple radiographic systems. It was used for x-ray and optical images to determine which component of the system was not operating up to standard. Focal spot size was monitored by imaging a bar pattern. This paper constitutes the second progress report on the development of the camera and radiation image quality indicators

  2. Mass spectrometry for real-time quantitative breath analysis

    Czech Academy of Sciences Publication Activity Database

    Smith, D.; Španěl, Patrik; Herbig, J.; Beauchamp, J.

    2014-01-01

    Roč. 8, č. 2 (2014), 027101 ISSN 1752-7155 Institutional support: RVO:61388955 Keywords : breath analysis * proton transfer reaction mass spectrometry * selected ion flow tube mass spectrometry Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 4.631, year: 2014

  3. Quantitative (real-time) PCR

    International Nuclear Information System (INIS)

    Denman, S.E.; McSweeney, C.S.

    2005-01-01

    Many nucleic acid-based probe and PCR assays have been developed for the detection tracking of specific microbes within the rumen ecosystem. Conventional PCR assays detect PCR products at the end stage of each PCR reaction, where exponential amplification is no longer being achieved. This approach can result in different end product (amplicon) quantities being generated. In contrast, using quantitative, or real-time PCR, quantification of the amplicon is performed not at the end of the reaction, but rather during exponential amplification, where theoretically each cycle will result in a doubling of product being created. For real-time PCR, the cycle at which fluorescence is deemed to be detectable above the background during the exponential phase is termed the cycle threshold (Ct). The Ct values obtained are then used for quantitation, which will be discussed later

  4. Exploring Valid Reference Genes for Quantitative Real-Time PCR Analysis in Sesamia inferens (Lepidoptera: Noctuidae)

    OpenAIRE

    Sun, Meng; Lu, Ming-Xing; Tang, Xiao-Tian; Du, Yu-Zhou

    2015-01-01

    The pink stem borer, Sesamia inferens, which is endemic in China and other parts of Asia, is a major pest of rice and causes significant yield loss in this host plant. Very few studies have addressed gene expression in S. inferens. Quantitative real-time PCR (qRT-PCR) is currently the most accurate and sensitive method for gene expression analysis. In qRT-PCR, data are normalized using reference genes, which help control for internal differences and reduce error between samples. In this study...

  5. Real Time Text Analysis

    Science.gov (United States)

    Senthilkumar, K.; Ruchika Mehra Vijayan, E.

    2017-11-01

    This paper aims to illustrate real time analysis of large scale data. For practical implementation we are performing sentiment analysis on live Twitter feeds for each individual tweet. To analyze sentiments we will train our data model on sentiWordNet, a polarity assigned wordNet sample by Princeton University. Our main objective will be to efficiency analyze large scale data on the fly using distributed computation. Apache Spark and Apache Hadoop eco system is used as distributed computation platform with Java as development language

  6. Exploring Valid Reference Genes for Quantitative Real-time PCR Analysis in Plutella xylostella (Lepidoptera: Plutellidae)

    Science.gov (United States)

    Fu, Wei; Xie, Wen; Zhang, Zhuo; Wang, Shaoli; Wu, Qingjun; Liu, Yong; Zhou, Xiaomao; Zhou, Xuguo; Zhang, Youjun

    2013-01-01

    Abstract: Quantitative real-time PCR (qRT-PCR), a primary tool in gene expression analysis, requires an appropriate normalization strategy to control for variation among samples. The best option is to compare the mRNA level of a target gene with that of reference gene(s) whose expression level is stable across various experimental conditions. In this study, expression profiles of eight candidate reference genes from the diamondback moth, Plutella xylostella, were evaluated under diverse experimental conditions. RefFinder, a web-based analysis tool, integrates four major computational programs including geNorm, Normfinder, BestKeeper, and the comparative ΔCt method to comprehensively rank the tested candidate genes. Elongation factor 1 (EF1) was the most suited reference gene for the biotic factors (development stage, tissue, and strain). In contrast, although appropriate reference gene(s) do exist for several abiotic factors (temperature, photoperiod, insecticide, and mechanical injury), we were not able to identify a single universal reference gene. Nevertheless, a suite of candidate reference genes were specifically recommended for selected experimental conditions. Our finding is the first step toward establishing a standardized qRT-PCR analysis of this agriculturally important insect pest. PMID:23983612

  7. Real time spectrum analysis

    International Nuclear Information System (INIS)

    Blunden, A.; O'Prey, D.G.; Tait, W.H.

    1983-01-01

    A method is described for the separation of a composite pulse-height spectrum into its unresolved component parts, which belong to a set of measured library spectra. The method allows real-time estimation giving running estimates during acquisition of the spectrum, minimises computation space, especially for a number of parallel calculations, estimates in advance the rms errors, and produces a significance measure for the hypothesis that the composite contains only the library spectra. Least squares curve-fitting, and other methods, can be compared, with the formalism developed, allowing analytical comparison of the effect of detector energy resolution and detection efficiency. A rational basis for the choice between the various methods of spectrum analysis follows from the theory, minimising rms estimation errors. The method described is applicable for very low numbers of counts and poor resolution. (orig.)

  8. Evaluation of Housekeeping Genes for Quantitative Real-Time PCR Analysis of Bradysia odoriphaga (Diptera: Sciaridae

    Directory of Open Access Journals (Sweden)

    Caihua Shi

    2016-07-01

    Full Text Available The soil insect Bradysia odoriphaga (Diptera: Sciaridae causes substantial damage to Chinese chive. Suitable reference genes in B. odoriphaga (Bradysia odoriphaga have yet to be identified for normalizing target gene expression among samples by quantitative real-time PCR (qRT-PCR. This study was focused on identifying the expression stability of 12 candidate housekeeping genes in B. odoriphaga under various experiment conditions. The final stability ranking of 12 housekeeping genes was obtained with RefFinder, and the most suitable number of reference genes was analyzed by GeNorm. The results revealed that the most appropriate sets of internal controls were RPS15, RPL18, and RPS18 across developmental phases; RPS15, RPL28, and GAPDH across temperatures; RPS15 and RPL18 across pesticide treatments; RSP5, RPS18, and SDHA across photoperiods; ACTb, RPS18, and RPS15 across diets; RPS13 and RPL28 across populations; and RPS15, ACTb, and RPS18 across all samples. The use of the most suitable reference genes versus an arbitrarily selected reference gene resulted in significant differences in the analysis of a target gene expression. HSP23 in B. odoriphaga was found to be up-regulated under low temperatures. These results will contribute to the standardization of qRT-PCR and will also be valuable for further research on gene function in B. odoriphaga.

  9. Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data

    NARCIS (Netherlands)

    Ramakers, Christian; Ruijter, Jan M.; Deprez, Ronald H. Lekanne; Moorman, Antoon F. M.

    2003-01-01

    Quantification of mRNAs using real-time polymerase chain reaction (PCR) by monitoring the product formation with the fluorescent dye SYBR Green I is being extensively used in neurosciences, developmental biology, and medical diagnostics. Most PCR data analysis procedures assume that the PCR

  10. Exploring valid reference genes for quantitative real-time PCR analysis in Sesamia inferens (Lepidoptera: Noctuidae.

    Directory of Open Access Journals (Sweden)

    Meng Sun

    Full Text Available The pink stem borer, Sesamia inferens, which is endemic in China and other parts of Asia, is a major pest of rice and causes significant yield loss in this host plant. Very few studies have addressed gene expression in S. inferens. Quantitative real-time PCR (qRT-PCR is currently the most accurate and sensitive method for gene expression analysis. In qRT-PCR, data are normalized using reference genes, which help control for internal differences and reduce error between samples. In this study, seven candidate reference genes, 18S ribosomal RNA (18S rRNA, elongation factor 1 (EF1, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, ribosomal protein S13 (RPS13, ribosomal protein S20 (RPS20, tubulin (TUB, and β-actin (ACTB were evaluated for their suitability in normalizing gene expression under different experimental conditions. The results indicated that three genes (RPS13, RPS20, and EF1 were optimal for normalizing gene expression in different insect tissues (head, epidermis, fat body, foregut, midgut, hindgut, Malpighian tubules, haemocytes, and salivary glands. 18S rRNA, EF1, and GAPDH were best for normalizing expression with respect to developmental stages and sex (egg masses; first, second, third, fourth, fifth, and sixth instar larvae; male and female pupae; and one-day-old male and female adults. 18S rRNA, RPS20, and TUB were optimal for fifth instars exposed to different temperatures (-8, -6, -4, -2, 0, and 27°C. To validate this recommendation, the expression profile of a target gene heat shock protein 83 gene (hsp83 was investigated, and results showed the selection was necessary and effective. In conclusion, this study describes reference gene sets that can be used to accurately measure gene expression in S. inferens.

  11. Exploring valid reference genes for quantitative real-time PCR analysis in Sesamia inferens (Lepidoptera: Noctuidae).

    Science.gov (United States)

    Sun, Meng; Lu, Ming-Xing; Tang, Xiao-Tian; Du, Yu-Zhou

    2015-01-01

    The pink stem borer, Sesamia inferens, which is endemic in China and other parts of Asia, is a major pest of rice and causes significant yield loss in this host plant. Very few studies have addressed gene expression in S. inferens. Quantitative real-time PCR (qRT-PCR) is currently the most accurate and sensitive method for gene expression analysis. In qRT-PCR, data are normalized using reference genes, which help control for internal differences and reduce error between samples. In this study, seven candidate reference genes, 18S ribosomal RNA (18S rRNA), elongation factor 1 (EF1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), ribosomal protein S13 (RPS13), ribosomal protein S20 (RPS20), tubulin (TUB), and β-actin (ACTB) were evaluated for their suitability in normalizing gene expression under different experimental conditions. The results indicated that three genes (RPS13, RPS20, and EF1) were optimal for normalizing gene expression in different insect tissues (head, epidermis, fat body, foregut, midgut, hindgut, Malpighian tubules, haemocytes, and salivary glands). 18S rRNA, EF1, and GAPDH were best for normalizing expression with respect to developmental stages and sex (egg masses; first, second, third, fourth, fifth, and sixth instar larvae; male and female pupae; and one-day-old male and female adults). 18S rRNA, RPS20, and TUB were optimal for fifth instars exposed to different temperatures (-8, -6, -4, -2, 0, and 27°C). To validate this recommendation, the expression profile of a target gene heat shock protein 83 gene (hsp83) was investigated, and results showed the selection was necessary and effective. In conclusion, this study describes reference gene sets that can be used to accurately measure gene expression in S. inferens.

  12. On-Orbit Quantitative Real-Time Gene Expression Analysis Using the Wetlab-2 System

    Science.gov (United States)

    Parra, Macarena; Jung, Jimmy; Almeida, Eduardo; Boone, Travis; Tran, Luan; Schonfeld, Julie

    2015-01-01

    NASA Ames Research Center's WetLab-2 Project enables on-orbit quantitative Reverse Transcriptase PCR (qRT-PCR) analysis without the need for sample return. The WetLab-2 system is capable of processing sample types ranging from microbial cultures to animal tissues dissected on-orbit. The project developed a RNA preparation module that can lyse cells and extract RNA of sufficient quality and quantity for use as templates in qRT-PCR reactions. Our protocol has the advantage of using non-toxic chemicals and does not require alcohols or other organics. The resulting RNA is dispensed into reaction tubes that contain all lyophilized reagents needed to perform qRT-PCR reactions. System operations require simple and limited crew actions including syringe pushes, valve turns and pipette dispenses. The project selected the Cepheid SmartCycler (TradeMark), a Commercial-Off-The-Shelf (COTS) qRT-PCR unit, because of its advantages including rugged modular design, low power consumption, rapid thermal ramp times and four-color multiplex detection. Single tube multiplex assays can be used to normalize for RNA concentration and integrity, and to study multiple genes of interest in each module. The WetLab-2 system can downlink data from the ISS to the ground after a completed run and uplink new thermal cycling programs. The ability to conduct qRT-PCR and generate results on-orbit is an important step towards utilizing the ISS as a National Laboratory facility. Specifically, the ability to get on-orbit data will provide investigators with the opportunity to adjust experimental parameters in real time without the need for sample return and re-flight. On orbit gene expression analysis can also eliminate the confounding effects on gene expression of reentry stresses and shock acting on live cells and organisms or the concern of RNA degradation of fixed samples and provide on-orbit gene expression benchmarking prior to sample return. Finally, the system can also be used for analysis of

  13. Quantitative analysis of the dystrophin gene by real-time PCR

    Directory of Open Access Journals (Sweden)

    Maksimovic Nela

    2012-01-01

    Full Text Available Duchenne and Becker muscular dystrophy (DMD/BMD are severe X-linked neuromuscular disorders caused by mutations in the dystrophin gene. Our aim was to optimize a quantitative real-time PCR method based on SYBR® Green I chemistry for routine diagnostics of DMD/BMD deletion carriers. Twenty female relatives of DMD/BMD patients with previously detected partial gene deletions were studied. The relative quantity of the target exons was calculated by a comparative threshold cycle method (ΔΔCt. The carrier status of all subjects was successfully determined. The gene dosage ratio for non-carriers was 1.07±0.20, and for carriers 0.56±0.11. This assay proved to be simple, rapid, reliable and cost-effective.

  14. Verus: A Tool for Quantitative Analysis of Finite-State Real-Time Systems.

    Science.gov (United States)

    1996-08-12

    Symbolic model checking is a technique for verifying finite-state concurrent systems that has been extended to handle real - time systems . Models with...up to 10(exp 30) states can often be verified in minutes. In this paper, we present a new tool to analyze real - time systems , based on this technique...We have designed a language, called Verus, for the description of real - time systems . Such a description is compiled into a state-transition graph and

  15. Expression analysis of fusarium wilt resistance gene in melon by real-time quantitative pcr

    International Nuclear Information System (INIS)

    Wang, X.; Xu, B.; Zhao, L.; Gao, P.; Luan, F.

    2014-01-01

    Melon Actin gene was used as a reference gene, to explore the gene expression profiles of the Fom-2 gene in roots, stems, and leaves of melon MR-1 under induction by Fusarium oxysporum f. sp. melonis. Monitoring using real-time quantitative PCR showed similar accumulation patterns of Fom-2 in roots, stems, and leaves over the observation period of 1 to 11 days; the expression level in stems was the highest. The expression of the Fom-2 gene was strengthened by the prolongation of induction time. In stems, the expression of Fom-2 was 5.737 times higher than in the control at three days; in roots, expression of Fom-2 was 5.617 times higher than in the control at five days. Similarly, the expression of Fom-2 in leaves obviously increased. It was 4.441 times higher than in the control at 5 days. The expression of Fom-2 was non-tissue specific, up-regulated under induction by Fusarium, and related to early resistance to Fusarium wilt. (author)

  16. An integrated real-time diagnostic concept using expert systems, qualitative reasoning and quantitative analysis

    International Nuclear Information System (INIS)

    Edwards, R.M.; Lee, K.Y.; Kumara, S.; Levine, S.H.

    1989-01-01

    An approach for an integrated real-time diagnostic system is being developed for inclusion as an integral part of a power plant automatic control system. In order to participate in control decisions and automatic closed loop operation, the diagnostic system must operate in real-time. Thus far, an expert system with real-time capabilities has been developed and installed on a subsystem at the Experimental Breeder Reactor (EBR-II) in Idaho, USA. Real-time simulation testing of advanced power plant concepts at the Pennsylvania State University has been developed and was used to support the expert system development and installation at EBR-II. Recently, the US National Science Foundation (NSF) and the US Department of Energy (DOE) have funded a Penn State research program to further enhance application of real-time diagnostic systems by pursuing implementation in a distributed power plant computer system including microprocessor based controllers. This paper summarizes past, current, planned, and possible future approaches to power plant diagnostic systems research at Penn State. 34 refs., 9 figs

  17. Visualisation and quantitative analysis of the rodent malaria liver stage by real time imaging.

    NARCIS (Netherlands)

    Ploemen, I.H.J.; Prudencio, M.; Douradinha, B.G.; Ramesar, J.; Fonager, J.; Gemert, G.J.A. van; Luty, A.J.F.; Hermsen, C.C.; Sauerwein, R.W.; Baptista, F.G.; Mota, M.M.; Waters, A.P.; Que, I.; Lowik, C.W.G.M.; Khan, S.M.; Janse, C.J.; Franke-Fayard, B.

    2009-01-01

    The quantitative analysis of Plasmodium development in the liver in laboratory animals in cultured cells is hampered by low parasite infection rates and the complicated methods required to monitor intracellular development. As a consequence, this important phase of the parasite's life cycle has been

  18. Cloning and evaluation of reference genes for quantitative real-time PCR analysis in Amorphophallus

    Directory of Open Access Journals (Sweden)

    Kai Wang

    2017-04-01

    Full Text Available Quantitative real-time reverse transcription PCR (RT-qPCR has been widely used in the detection and quantification of gene expression levels because of its high accuracy, sensitivity, and reproducibility as well as its large dynamic range. However, the reliability and accuracy of RT-qPCR depends on accurate transcript normalization using stably expressed reference genes. Amorphophallus is a perennial plant with a high content of konjac glucomannan (KGM in its corm. This crop has been used as a food source and as a traditional medicine for thousands of years. Without adequate knowledge of gene expression profiles, there has been no report of validated reference genes in Amorphophallus. In this study, nine genes that are usually used as reference genes in other crops were selected as candidate reference genes. These putative sequences of these genes Amorphophallus were cloned by the use of degenerate primers. The expression stability of each gene was assessed in different tissues and under two abiotic stresses (heat and waterlogging in A. albus and A. konjac. Three distinct algorithms were used to evaluate the expression stability of the candidate reference genes. The results demonstrated that EF1-a, EIF4A, H3 and UBQ were the best reference genes under heat stress in Amorphophallus. Furthermore, EF1-a, EIF4A, TUB, and RP were the best reference genes in waterlogged conditions. By comparing different tissues from all samples, we determined that EF1-α, EIF4A, and CYP were stable in these sets. In addition, the suitability of these reference genes was confirmed by validating the expression of a gene encoding the small heat shock protein SHSP, which is related to heat stress in Amorphophallus. In sum, EF1-α and EIF4A were the two best reference genes for normalizing mRNA levels in different tissues and under various stress treatments, and we suggest using one of these genes in combination with 1 or 2 reference genes associated with different

  19. A novel universal real-time PCR system using the attached universal duplex probes for quantitative analysis of nucleic acids

    Directory of Open Access Journals (Sweden)

    Wilson Zoe A

    2008-06-01

    Full Text Available Abstract Background Real-time PCR techniques are being widely used for nucleic acids analysis, but one limitation of current frequently employed real-time PCR is the high cost of the labeled probe for each target molecule. Results We describe a real-time PCR technique employing attached universal duplex probes (AUDP, which has the advantage of generating fluorescence by probe hydrolysis and strand displacement over current real-time PCR methods. AUDP involves one set of universal duplex probes in which the 5' end of the fluorescent probe (FP and a complementary quenching probe (QP lie in close proximity so that fluorescence can be quenched. The PCR primer pair with attached universal template (UT and the FP are identical to the UT sequence. We have shown that the AUDP technique can be used for detecting multiple target DNA sequences in both simplex and duplex real-time PCR assays for gene expression analysis, genotype identification, and genetically modified organism (GMO quantification with comparable sensitivity, reproducibility, and repeatability with other real-time PCR methods. Conclusion The results from GMO quantification, gene expression analysis, genotype identification, and GMO quantification using AUDP real-time PCR assays indicate that the AUDP real-time PCR technique has been successfully applied in nucleic acids analysis, and the developed AUDP real-time PCR technique will offer an alternative way for nucleic acid analysis with high efficiency, reliability, and flexibility at low cost.

  20. Interlaboratory validation of quantitative duplex real-time PCR method for screening analysis of genetically modified maize.

    Science.gov (United States)

    Takabatake, Reona; Koiwa, Tomohiro; Kasahara, Masaki; Takashima, Kaori; Futo, Satoshi; Minegishi, Yasutaka; Akiyama, Hiroshi; Teshima, Reiko; Oguchi, Taichi; Mano, Junichi; Furui, Satoshi; Kitta, Kazumi

    2011-01-01

    To reduce the cost and time required to routinely perform the genetically modified organism (GMO) test, we developed a duplex quantitative real-time PCR method for a screening analysis simultaneously targeting an event-specific segment for GA21 and Cauliflower Mosaic Virus 35S promoter (P35S) segment [Oguchi et al., J. Food Hyg. Soc. Japan, 50, 117-125 (2009)]. To confirm the validity of the method, an interlaboratory collaborative study was conducted. In the collaborative study, conversion factors (Cfs), which are required to calculate the GMO amount (%), were first determined for two real-time PCR instruments, the ABI PRISM 7900HT and the ABI PRISM 7500. A blind test was then conducted. The limit of quantitation for both GA21 and P35S was estimated to be 0.5% or less. The trueness and precision were evaluated as the bias and reproducibility of the relative standard deviation (RSD(R)). The determined bias and RSD(R) were each less than 25%. We believe the developed method would be useful for the practical screening analysis of GM maize.

  1. Real time analysis under EDS

    International Nuclear Information System (INIS)

    Schneberk, D.

    1985-07-01

    This paper describes the analysis component of the Enrichment Diagnostic System (EDS) developed for the Atomic Vapor Laser Isotope Separation Program (AVLIS) at Lawrence Livermore National Laboratory (LLNL). Four different types of analysis are performed on data acquired through EDS: (1) absorption spectroscopy on laser-generated spectral lines, (2) mass spectrometer analysis, (3) general purpose waveform analysis, and (4) separation performance calculations. The information produced from this data includes: measures of particle density and velocity, partial pressures of residual gases, and overall measures of isotope enrichment. The analysis component supports a variety of real-time modeling tasks, a means for broadcasting data to other nodes, and a great degree of flexibility for tailoring computations to the exact needs of the process. A particular data base structure and program flow is common to all types of analysis. Key elements of the analysis component are: (1) a fast access data base which can configure all types of analysis, (2) a selected set of analysis routines, (3) a general purpose data manipulation and graphics package for the results of real time analysis. Each of these components are described with an emphasis upon how each contributes to overall system capability. 3 figs

  2. Validation of Reference Genes for Quantitative Expression Analysis by Real-Time RT-PCR in Four Lepidopteran Insects

    OpenAIRE

    Teng, Xiaolu; Zhang, Zan; He, Guiling; Yang, Liwen; Li, Fei

    2012-01-01

    Quantitative real-time polymerase chain reaction (qPCR) is an efficient and widely used technique to monitor gene expression. Housekeeping genes (HKGs) are often empirically selected as the reference genes for data normalization. However, the suitability of HKGs used as the reference genes has been seldom validated. Here, six HKGs were chosen (actin A3, actin A1, GAPDH, G3PDH, E2F, rp49) in four lepidopteran insects Bombyx mori L. (Lepidoptera: Bombycidae), Plutella xylostella L. (Plutellidae...

  3. Identification of reference genes for expression analysis by real-time quantitative PCR in drought-stressed soybean

    Directory of Open Access Journals (Sweden)

    Renata Stolf-Moreira

    2011-01-01

    Full Text Available The objective of this work was to validate, by quantitative PCR in real time (RT-qPCR, genes to be used as reference in studies of gene expression in soybean in drought-stressed trials. Four genes commonly used in soybean were evaluated: Gmβ-actin, GmGAPDH, GmLectin and GmRNAr18S. Total RNA was extracted from six samples: three from roots in a hydroponic system with different drought intensities (0, 25, 50, 75 and 100 minutes of water stress, and three from leaves of plants grown in sand with different soil moistures (15, 5 and 2.5% gravimetric humidity. The raw cycle threshold (Ct data were analyzed, and the efficiency of each primer was calculated for an overall analysis of the Ct range among the different samples. The GeNorm application was used to evaluate the best reference gene, according to its stability. The GmGAPDH was the least stable gene, with the highest mean values of expression stability (M, and the most stable genes, with the lowest M values, were the Gmβ-actin and GmRNAr18S, when both root and leaves samples were tested. These genes can be used in RT-qPCR as reference gene for expression analysis.

  4. Quantitative analysis of diet structure by real-time PCR, reveals different feeding patterns by two dominant grasshopper species

    Science.gov (United States)

    Huang, Xunbing; Wu, Huihui; McNeill, Mark Richard; Qin, Xinghu; Ma, Jingchuan; Tu, Xiongbing; Cao, Guangchun; Wang, Guangjun; Nong, Xiangqun; Zhang, Zehua

    2016-01-01

    Studies on grasshopper diets have historically employed a range of methodologies, each with certain advantages and disadvantages. For example, some methodologies are qualitative instead of quantitative. Others require long experimental periods or examine population-level effects, only. In this study, we used real-time PCR to examine diets of individual grasshoppers. The method has the advantage of being both fast and quantitative. Using two grasshopper species, Oedaleus asiaticus and Dasyhippus barbipes, we designed ITS primer sequences for their three main host plants, Stipa krylovii, Leymus chinensis and Cleistogenes squarrosa and used real-time PCR method to test diet structure both qualitatively and quantitatively. The lowest detection efficiency of the three grass species was ~80% with a strong correlation between actual and PCR-measured food intake. We found that Oedaleus asiaticus maintained an unchanged diet structure across grasslands with different grass communities. By comparison, Dasyhippus barbipes changed its diet structure. These results revealed why O. asiaticus distribution is mainly confined to Stipa-dominated grassland, and D. barbipes is more widely distributed across Inner Mongolia. Overall, real-time PCR was shown to be a useful tool for investigating grasshopper diets, which in turn offers some insight into grasshopper distributions and improved pest management. PMID:27562455

  5. WetLab-2: Tools for Conducting On-Orbit Quantitative Real-Time Gene Expression Analysis on ISS

    Science.gov (United States)

    Parra, Macarena; Almeida, Eduardo; Boone, Travis; Jung, Jimmy; Schonfeld, Julie

    2014-01-01

    The objective of NASA Ames Research Centers WetLab-2 Project is to place on the ISS a research platform capable of conducting gene expression analysis via quantitative real-time PCR (qRT-PCR) of biological specimens sampled or cultured on orbit. The project has selected a Commercial-Off-The-Shelf (COTS) qRT-PCR system, the Cepheid SmartCycler and will fly it in its COTS configuration. The SmartCycler has a number of advantages including modular design (16 independent PCR modules), low power consumption, rapid ramp times and the ability to detect up to four separate fluorescent channels at one time enabling multiplex assays that can be used for normalization and to study multiple genes of interest in each module. The team is currently working with Cepheid to enable the downlink of data from the ISS to the ground and provide uplink capabilities for programming, commanding, monitoring, and instrument maintenance. The project has adapted commercial technology to design a module that can lyse cells and extract RNA of sufficient quality and quantity for use in qRT-PCR reactions while using a housekeeping gene to normalize RNA concentration and integrity. The WetLab-2 system is capable of processing multiple sample types ranging from microbial cultures to animal tissues dissected on-orbit. The ability to conduct qRT-PCR on-orbit eliminates the confounding effects on gene expression of reentry stresses and shock acting on live cells and organisms or the concern of RNA degradation of fixed samples. The system can be used to validate terrestrial analyses of samples returned from ISS by providing on-orbit gene expression benchmarking prior to sample return. The ability to get on orbit data will provide investigators with the opportunity to adjust experiment parameters for subsequent trials based on the real-time data analysis without need for sample return and re-flight. Researchers will also be able to sample multigenerational changes in organisms. Finally, the system can be

  6. Validation of reference genes for quantitative expression analysis by real-time rt-PCR in four lepidopteran insects.

    Science.gov (United States)

    Teng, Xiaolu; Zhang, Zan; He, Guiling; Yang, Liwen; Li, Fei

    2012-01-01

    Quantitative real-time polymerase chain reaction (qPCR) is an efficient and widely used technique to monitor gene expression. Housekeeping genes (HKGs) are often empirically selected as the reference genes for data normalization. However, the suitability of HKGs used as the reference genes has been seldom validated. Here, six HKGs were chosen (actin A3, actin A1, GAPDH, G3PDH, E2F, rp49) in four lepidopteran insects Bombyx mori L. (Lepidoptera: Bombycidae), Plutella xylostella L. (Plutellidae), Chilo suppressalis Walker (Crambidae), and Spodoptera exigua Hübner (Noctuidae) to study their expression stability. The algorithms of geNorm, NormFinder, stability index, and ΔCt analysis were used to evaluate these HKGs. Across different developmental stages, actin A1 was the most stable in P. xylostella and C. suppressalis, but it was the least stable in B. mori and S. exigua. Rp49 and GAPDH were the most stable in B. mori and S. exigua, respectively. In different tissues, GAPDH, E2F, and Rp49 were the most stable in B. mori, S. exigua, and C. suppressalis, respectively. The relative abundances of Siwi genes estimated by 2(-ΔΔCt) method were tested with different HKGs as the reference gene, proving the importance of internal controls in qPCR data analysis. The results not only presented a list of suitable reference genes in four lepidopteran insects, but also proved that the expression stabilities of HKGs were different among evolutionarily close species. There was no single universal reference gene that could be used in all situations. It is indispensable to validate the expression of HKGs before using them as the internal control in qPCR.

  7. Selection and validation of appropriate reference genes for quantitative real-time PCR analysis in Salvia hispanica.

    Directory of Open Access Journals (Sweden)

    Rahul Gopalam

    Full Text Available Quantitative real-time polymerase chain reaction (qRT-PCR has become the most popular choice for gene expression studies. For accurate expression analysis, it is pertinent to select a stable reference gene to normalize the data. It is now known that the expression of internal reference genes varies considerably during developmental stages and under different experimental conditions. For Salvia hispanica, an economically important oilseed crop, there are no reports of stable reference genes till date. In this study, we chose 13 candidate reference genes viz. Actin11 (ACT, Elongation factor 1-alpha (EF1-α, Eukaryotic translation initiation factor 3E (ETIF3E, alpha tubulin (α-TUB, beta tubulin (β-TUB, Glyceraldehyde 3-phosphate dehydrogenase (GAPDH, Cyclophilin (CYP, Clathrin adaptor complex (CAC, Serine/threonine-protein phosphatase 2A (PP2A, FtsH protease (FtsH, 18S ribosomal RNA (18S rRNA, S-adenosyl methionine decarboxylase (SAMDC and Rubisco activase (RCA and the expression levels of these genes were assessed in a diverse set of tissue samples representing vegetative stages, reproductive stages and various abiotic stress treatments. Two of the widely used softwares, geNorm and Normfinder were used to evaluate the expression stabilities of these 13 candidate reference genes under different conditions. Results showed that GAPDH and CYP expression remain stable throughout in the different abiotic stress treatments, CAC and PP2A expression were relatively stable under reproductive stages and α-TUB, PP2A and ETIF3E were found to be stably expressed in vegetative stages. Further, the expression levels of Diacylglycerol acyltransferase (DGAT1, a key enzyme in triacylglycerol synthesis was analyzed to confirm the validity of reference genes identified in the study. This is the first systematic study of selection of reference genes in S. hispanica, and will benefit future expression studies in this crop.

  8. Detection and quantification of Renibacterium salmoninarum DNA in salmonid tissues by real-time quantitative polymerase chain reaction analysis

    Science.gov (United States)

    Chase, D.M.; Elliott, D.G.; Pascho, R.J.

    2006-01-01

    Renibacterium salmoninarum is an important salmonid pathogen that is difficult to culture. We developed and assessed a real-time, quantitative, polymerase chain reaction (qPCR) assay for the detection and enumeration of R. salmoninarum. The qPCR is based on TaqMan technology and amplifies a 69-base pair (bp) region of the gene encoding the major soluble antigen (MSA) of R. salmoninarum. The qPCR assay consistently detected as few as 5 R. salmoninarum cells per reaction in kidney tissue. The specificity of the qPCR was confirmed by testing the DNA extracts from a panel of microorganisms that were either common fish pathogens or reported to cause false-positive reactions in the enzyme-linked immunosorbent assay (ELISA). Kidney samples from 38 juvenile Chinook salmon (Oncorhynchus tshawytscha) in a naturally infected population were examined by real-time qPCR, a nested PCR, and ELISA, and prevalences of R. salmoninarum detected were 71, 66, and 71%, respectively. The qPCR should be a valuable tool for evaluating the R. salmoninarum infection status of salmonids.

  9. Screening suitable reference genes for normalization in reverse transcription quantitative real-time PCR analysis in melon.

    Directory of Open Access Journals (Sweden)

    Qiusheng Kong

    Full Text Available Melon (Cucumis melo. L is not only an economically important cucurbitaceous crop but also an attractive model for studying many biological characteristics. Screening appropriate reference genes is essential to reverse transcription quantitative real-time PCR (RT-qPCR, which is key to many studies involving gene expression analysis. In this study, 14 candidate reference genes were selected, and the variations in their expression in roots and leaves of plants subjected to biotic stress, abiotic stress, and plant growth regulator treatment were assessed by RT-qPCR. The stability of the expression of the selected genes was determined and ranked using geNorm and NormFinder. geNorm identified the two most stable genes for each set of conditions: CmADP and CmUBIep across all samples, CmUBIep and CmRPL in roots, CmRAN and CmACT in leaves, CmADP and CmRPL under abiotic stress conditions, CmTUA and CmACT under biotic stress conditions, and CmRAN and CmACT under plant growth regulator treatments. NormFinder determined CmRPL to be the best reference gene in roots and under biotic stress conditions and CmADP under the other experimental conditions. CmUBC2 and CmPP2A were not found to be suitable under many experimental conditions. The catalase family genes CmCAT1, CmCAT2, and CmCAT3 were identified in melon genome and used as target genes to validate the reliability of identified reference genes. The catalase family genes showed the most upregulation 3 days after inoculation with Fusarium wilt in roots, after which they were downregulated. Their levels of expression were significantly overestimated when the unsuitable reference gene was used for normalization. These results not only provide guidelines for the selection of reference genes for gene expression analyses in melons but may also provide valuable information for studying the functions of catalase family genes in stress responses.

  10. Recombinant plasmid-based quantitative Real-Time PCR analysis of Salmonella enterica serotypes and its application to milk samples.

    Science.gov (United States)

    Gokduman, Kurtulus; Avsaroglu, M Dilek; Cakiris, Aris; Ustek, Duran; Gurakan, G Candan

    2016-03-01

    The aim of the current study was to develop, a new, rapid, sensitive and quantitative Salmonella detection method using a Real-Time PCR technique based on an inexpensive, easy to produce, convenient and standardized recombinant plasmid positive control. To achieve this, two recombinant plasmids were constructed as reference molecules by cloning the two most commonly used Salmonella-specific target gene regions, invA and ttrRSBC. The more rapid detection enabled by the developed method (21 h) compared to the traditional culture method (90 h) allows the quantitative evaluation of Salmonella (quantification limits of 10(1)CFU/ml and 10(0)CFU/ml for the invA target and the ttrRSBC target, respectively), as illustrated using milk samples. Three advantages illustrated by the current study demonstrate the potential of the newly developed method to be used in routine analyses in the medical, veterinary, food and water/environmental sectors: I--The method provides fast analyses including the simultaneous detection and determination of correct pathogen counts; II--The method is applicable to challenging samples, such as milk; III--The method's positive controls (recombinant plasmids) are reproducible in large quantities without the need to construct new calibration curves. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Reference gene selection for real-time quantitative PCR analysis of the mouse uterus in the peri-implantation period.

    Directory of Open Access Journals (Sweden)

    Pengfei Lin

    Full Text Available The study of uterine gene expression patterns is valuable for understanding the biological and molecular mechanisms that occur during embryo implantation. Real-time quantitative RT-PCR (qRT-PCR is an extremely sensitive technique that allows for the precise quantification of mRNA abundance; however, selecting stable reference genes suitable for the normalization of qRT-PCR data is required to avoid the misinterpretation of experimental results and erroneous analyses. This study employs several mouse models, including an early pregnancy, a pseudopregnancy, a delayed implantation and activation, an artificial decidualization and a hormonal treatment model; ten candidate reference genes (PPIA, RPLP0, HPRT1, GAPDH, ACTB, TBP, B2M, 18S, UBC and TUBA that are found in uterine tissues were assessed for their suitability as internal controls for relative qRT-PCR quantification. GeNorm(PLUS, NormFinder, and BestKeeper were used to evaluate these candidate reference genes, and all of these methods identified RPLP0 and GAPDH as the most stable candidates and B2M and 18S as the least stable candidates. However, when the different models were analyzed separately, the reference genes exhibited some variation in their expression levels.

  12. Quantitative Real-Time Fluoroscopy Analysis on Measurement of the Hepatic Arterial Flow During Transcatheter Arterial Chemoembolization of Hepatocellular Carcinoma: Comparison with Quantitative Digital Subtraction Angiography Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Yi-Yang; Lee, Rheun-Chuan, E-mail: rclee@vghtpe.gov.tw; Guo, Wan-Yuo, E-mail: wyguo@vghtpe.gov.tw; Chu, Wei-Fa [Taipei Veterans General Hospital, Department of Radiology (China); Wu, Frank Chun-Hsien [Siemens Healthcare Ltd. (China); Gehrisch, Sonja [Siemens Healthcare GmbH (Germany)

    2016-11-15

    PurposeTo quantify the arterial flow change during transcatheter arterial chemoembolization (TACE) for hepatocellular carcinoma (HCC) using digital subtraction angiography, quantitative color-coding analysis (d-QCA), and real-time subtraction fluoroscopy QCA (f-QCA).Materials and MethodsThis prospective study enrolled 20 consecutive patients with HCC who had undergone TACE via a subsegmental approach between February 2014 and April 2015. The TACE endpoint was a sluggish antegrade tumor-feeding arterial flow. d-QCA and f-QCA were used for determining the relative maximal density time (rT{sub max}) of the selected arteries. The rT{sub max} of the selected arteries was analyzed in d-QCA and f-QCA before and after TACE, and its correlation in both analyses was evaluated.ResultsThe pre- and post-TACE rT{sub max} of the embolized segmental artery in d-QCA and f-QCA were 1.59 ± 0.81 and 2.97 ± 1.80 s (P < 0.001) and 1.44 ± 0.52 and 2.28 ± 1.02 s (P < 0.01), respectively. The rT{sub max} of the proximal hepatic artery did not significantly change during TACE in d-QCA and f-QCA. The Spearman correlation coefficients of the pre- and post-TACE rT{sub max} of the embolized segmental artery between d-QCA and f-QCA were 0.46 (P < 0.05) and 0.80 (P < 0.001). Radiation doses in one series of d-QCA and f-QCA were 140.7 ± 51.5 milligray (mGy) and 2.5 ± 0.7 mGy, respectively.Conclusionsf-QCA can quantify arterial flow changes with a higher temporal resolution and lower radiation dose. Flow quantification of the embolized segmental artery using f-QCA and d-QCA is highly correlated.

  13. Quantitative Real-Time Fluoroscopy Analysis on Measurement of the Hepatic Arterial Flow During Transcatheter Arterial Chemoembolization of Hepatocellular Carcinoma: Comparison with Quantitative Digital Subtraction Angiography Analysis

    International Nuclear Information System (INIS)

    Lin, Yi-Yang; Lee, Rheun-Chuan; Guo, Wan-Yuo; Chu, Wei-Fa; Wu, Frank Chun-Hsien; Gehrisch, Sonja

    2016-01-01

    PurposeTo quantify the arterial flow change during transcatheter arterial chemoembolization (TACE) for hepatocellular carcinoma (HCC) using digital subtraction angiography, quantitative color-coding analysis (d-QCA), and real-time subtraction fluoroscopy QCA (f-QCA).Materials and MethodsThis prospective study enrolled 20 consecutive patients with HCC who had undergone TACE via a subsegmental approach between February 2014 and April 2015. The TACE endpoint was a sluggish antegrade tumor-feeding arterial flow. d-QCA and f-QCA were used for determining the relative maximal density time (rT_m_a_x) of the selected arteries. The rT_m_a_x of the selected arteries was analyzed in d-QCA and f-QCA before and after TACE, and its correlation in both analyses was evaluated.ResultsThe pre- and post-TACE rT_m_a_x of the embolized segmental artery in d-QCA and f-QCA were 1.59 ± 0.81 and 2.97 ± 1.80 s (P < 0.001) and 1.44 ± 0.52 and 2.28 ± 1.02 s (P < 0.01), respectively. The rT_m_a_x of the proximal hepatic artery did not significantly change during TACE in d-QCA and f-QCA. The Spearman correlation coefficients of the pre- and post-TACE rT_m_a_x of the embolized segmental artery between d-QCA and f-QCA were 0.46 (P < 0.05) and 0.80 (P < 0.001). Radiation doses in one series of d-QCA and f-QCA were 140.7 ± 51.5 milligray (mGy) and 2.5 ± 0.7 mGy, respectively.Conclusionsf-QCA can quantify arterial flow changes with a higher temporal resolution and lower radiation dose. Flow quantification of the embolized segmental artery using f-QCA and d-QCA is highly correlated.

  14. Quantitative analysis of the improvement in high zoom maritime tracking due to real-time image enhancement

    CSIR Research Space (South Africa)

    Bachoo, AK

    2011-04-01

    Full Text Available This work aims to evaluate the improvement in the performance of tracking small maritime targets due to real-time enhancement of the video streams from high zoom cameras on pan-tilt pedestal. Due to atmospheric conditions these images can frequently...

  15. Identification of internal control genes for quantitative expression analysis by real-time PCR in bovine peripheral lymphocytes.

    Science.gov (United States)

    Spalenza, Veronica; Girolami, Flavia; Bevilacqua, Claudia; Riondato, Fulvio; Rasero, Roberto; Nebbia, Carlo; Sacchi, Paola; Martin, Patrice

    2011-09-01

    Gene expression studies in blood cells, particularly lymphocytes, are useful for monitoring potential exposure to toxicants or environmental pollutants in humans and livestock species. Quantitative PCR is the method of choice for obtaining accurate quantification of mRNA transcripts although variations in the amount of starting material, enzymatic efficiency, and the presence of inhibitors can lead to evaluation errors. As a result, normalization of data is of crucial importance. The most common approach is the use of endogenous reference genes as an internal control, whose expression should ideally not vary among individuals and under different experimental conditions. The accurate selection of reference genes is therefore an important step in interpreting quantitative PCR studies. Since no systematic investigation in bovine lymphocytes has been performed, the aim of the present study was to assess the expression stability of seven candidate reference genes in circulating lymphocytes collected from 15 dairy cows. Following the characterization by flow cytometric analysis of the cell populations obtained from blood through a density gradient procedure, three popular softwares were used to evaluate the gene expression data. The results showed that two genes are sufficient for normalization of quantitative PCR studies in cattle lymphocytes and that YWAHZ, S24 and PPIA are the most stable genes. Copyright © 2010 Elsevier Ltd. All rights reserved.

  16. Progesterone receptor isoform analysis by quantitative real-time polymerase chain reaction in formalin-fixed, paraffin-embedded canine mammary dysplasias and tumors

    DEFF Research Database (Denmark)

    Guil-Luna, S.; Stenvang, Jan; Brünner, Nils

    2014-01-01

    and its isoforms in formalin-fixed, paraffin-embedded tissue samples from canine mammary lesions (4 dysplasias, 10 benign tumors, and 46 carcinomas) using 1-step SYBR Green quantitative real-time polymerase chain reaction (RT-qPCR). Progesterone receptor was expressed in 75% of dysplasias, all benign...... in the expression of isoform A versus B. Analysis of progesterone receptor mRNA isoforms by RT-qPCR was successful in routinely formalin-fixed, paraffin-embedded tissue samples and enabled the distribution of isoforms A and B to be identified for the first time in dysplasias, benign tumors, and malignant tumors...

  17. Diagnosis of aerobic vaginitis by quantitative real-time PCR

    OpenAIRE

    Rumyantseva, T. A.; Bellen, G.; Savochkina, Y. A.; Guschin, A. E.; Donders, G.G.G.

    2016-01-01

    Abstract: Purpose To evaluate a real-time PCR-based technique to quantify bacteria associated with aerobic vaginitis (AV) as a potential test. Methods Vaginal samples from 100 women were tested by wet-mount microscopy, gram stain and quantitative real-time PCR targeting Enterobacteriacea, Staphylococcus spp., Streptococcus spp., Enterococcus spp., Escherichia coli, Streptococcus agalactiae, S. aureus; Lactobacillus spp. AV diagnosis obtained by wet-mount microscopy was used as reference. Resu...

  18. Identification of appropriate reference genes for human mesenchymal stem cell analysis by quantitative real-time PCR.

    Science.gov (United States)

    Li, Xiuying; Yang, Qiwei; Bai, Jinping; Xuan, Yali; Wang, Yimin

    2015-01-01

    Normalization to a reference gene is the method of choice for quantitative reverse transcription-PCR (RT-qPCR) analysis. The stability of reference genes is critical for accurate experimental results and conclusions. We have evaluated the expression stability of eight commonly used reference genes found in four different human mesenchymal stem cells (MSC). Using geNorm, NormFinder and BestKeeper algorithms, we show that beta-2-microglobulin and peptidyl-prolylisomerase A were the optimal reference genes for normalizing RT-qPCR data obtained from MSC, whereas the TATA box binding protein was not suitable due to its extensive variability in expression. Our findings emphasize the significance of validating reference genes for qPCR analyses. We offer a short list of reference genes to use for normalization and recommend some commercially-available software programs as a rapid approach to validate reference genes. We also demonstrate that the two reference genes, β-actin and glyceraldehyde-3-phosphate dehydrogenase, are frequently used are not always successful in many cases.

  19. Failure analysis of real-time systems

    International Nuclear Information System (INIS)

    Jalashgar, A.; Stoelen, K.

    1998-01-01

    This paper highlights essential aspects of real-time software systems that are strongly related to the failures and their course of propagation. The significant influence of means-oriented and goal-oriented system views in the description, understanding and analysing of those aspects is elaborated. The importance of performing failure analysis prior to reliability analysis of real-time systems is equally addressed. Problems of software reliability growth models taking the properties of such systems into account are discussed. Finally, the paper presents a preliminary study of a goal-oriented approach to model the static and dynamic characteristics of real-time systems, so that the corresponding analysis can be based on a more descriptive and informative picture of failures, their effects and the possibility of their occurrence. (author)

  20. A two-stage method of quantitative flood risk analysis for reservoir real-time operation using ensemble-based hydrologic forecasts

    Science.gov (United States)

    Liu, P.

    2013-12-01

    Quantitative analysis of the risk for reservoir real-time operation is a hard task owing to the difficulty of accurate description of inflow uncertainties. The ensemble-based hydrologic forecasts directly depict the inflows not only the marginal distributions but also their persistence via scenarios. This motivates us to analyze the reservoir real-time operating risk with ensemble-based hydrologic forecasts as inputs. A method is developed by using the forecast horizon point to divide the future time into two stages, the forecast lead-time and the unpredicted time. The risk within the forecast lead-time is computed based on counting the failure number of forecast scenarios, and the risk in the unpredicted time is estimated using reservoir routing with the design floods and the reservoir water levels of forecast horizon point. As a result, a two-stage risk analysis method is set up to quantify the entire flood risks by defining the ratio of the number of scenarios that excessive the critical value to the total number of scenarios. The China's Three Gorges Reservoir (TGR) is selected as a case study, where the parameter and precipitation uncertainties are implemented to produce ensemble-based hydrologic forecasts. The Bayesian inference, Markov Chain Monte Carlo, is used to account for the parameter uncertainty. Two reservoir operation schemes, the real operated and scenario optimization, are evaluated for the flood risks and hydropower profits analysis. With the 2010 flood, it is found that the improvement of the hydrologic forecast accuracy is unnecessary to decrease the reservoir real-time operation risk, and most risks are from the forecast lead-time. It is therefore valuable to decrease the avarice of ensemble-based hydrologic forecasts with less bias for a reservoir operational purpose.

  1. Real-time systems design and analysis

    CERN Document Server

    Laplante, Phillip A

    2004-01-01

    "Real-Time Systems Design and Analysis, Third Edition is essential for students and practicing software engineers who want improved designs, faster computation, and ultimate cost savings. Chapters discuss hardware considerations and software requirements, software systems design, the software production process, performance estimation and optimization, and engineering considerations."--Jacket.

  2. Human fecal source identification with real-time quantitative PCR

    Science.gov (United States)

    Waterborne diseases represent a significant public health risk worldwide, and can originate from contact with water contaminated with human fecal material. We describe a real-time quantitative PCR (qPCR) method that targets a Bacteroides dori human-associated genetic marker for...

  3. Identification and evaluation of reliable reference genes for quantitative real-time PCR analysis in tea plant (Camellia sinensis (L.) O. Kuntze)

    Science.gov (United States)

    Quantitative real-time polymerase chain reaction (qRT-PCR) is a commonly used technique for measuring gene expression levels due to its simplicity, specificity, and sensitivity. Reliable reference selection for the accurate quantification of gene expression under various experimental conditions is a...

  4. Statistical aspects of quantitative real-time PCR experiment design

    Czech Academy of Sciences Publication Activity Database

    Kitchen, R.R.; Kubista, Mikael; Tichopád, Aleš

    2010-01-01

    Roč. 50, č. 4 (2010), s. 231-236 ISSN 1046-2023 R&D Projects: GA AV ČR IAA500520809 Institutional research plan: CEZ:AV0Z50520701 Keywords : Real-time PCR * Experiment design * Nested analysis of variance Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.527, year: 2010

  5. Quantitative detection and typing of hepatitis D virus in human serum by real-time polymerase chain reaction and melting curve analysis.

    Science.gov (United States)

    Hofmann, Joerg; Frenzel, Katrin; Minh, Bui Q; von Haeseler, Arndt; Edelmann, Anke; Ross, Stefan R; Berg, Thomas; Krüger, Detlev H; Meisel, Helga

    2010-06-01

    Hepatitis D virus (HDV) infection is an important etiologic agent of fulminant hepatitis and may aggravate the clinical course of chronic hepatitis B infection resulting in cirrhosis and liver failure. This report describes the establishment of a real-time reverse transcriptase polymerase chain reaction method that allows the quantitative detection of HDV-1 and HDV-3 with a sensitivity in a linear range of 2 x 10(3) to 10(8) copies/mL. Additionally, the new assay provides the opportunity to distinguish HDV-1 from HDV-3 by a subsequent melting curve analysis, an important option because these HDV types are highly associated with severe clinical outcome. The results of the melting curve analysis of 42 HDV sequences obtained in this study and the phylogenetic analysis based on 139 full-length sequences from GenBank were consistent and showed that all sequences described here cluster within the HDV-1 clade. Therefore, this assay is useful for monitoring of antiviral treatment and molecular epidemiologic studies of HDV distribution. Copyright 2010 Elsevier Inc. All rights reserved.

  6. Microgravity validation of a novel system for RNA isolation and multiplex quantitative real time PCR analysis of gene expression on the International Space Station.

    Directory of Open Access Journals (Sweden)

    Macarena Parra

    Full Text Available The International Space Station (ISS National Laboratory is dedicated to studying the effects of space on life and physical systems, and to developing new science and technologies for space exploration. A key aspect of achieving these goals is to operate the ISS National Lab more like an Earth-based laboratory, conducting complex end-to-end experimentation, not limited to simple microgravity exposure. Towards that end NASA developed a novel suite of molecular biology laboratory tools, reagents, and methods, named WetLab-2, uniquely designed to operate in microgravity, and to process biological samples for real-time gene expression analysis on-orbit. This includes a novel fluidic RNA Sample Preparation Module and fluid transfer devices, all-in-one lyophilized PCR assays, centrifuge, and a real-time PCR thermal cycler. Here we describe the results from the WetLab-2 validation experiments conducted in microgravity during ISS increment 47/SPX-8. Specifically, quantitative PCR was performed on a concentration series of DNA calibration standards, and Reverse Transcriptase-quantitative PCR was conducted on RNA extracted and purified on-orbit from frozen Escherichia coli and mouse liver tissue. Cycle threshold (Ct values and PCR efficiencies obtained on-orbit from DNA standards were similar to Earth (1 g controls. Also, on-orbit multiplex analysis of gene expression from bacterial cells and mammalian tissue RNA samples was successfully conducted in about 3 h, with data transmitted within 2 h of experiment completion. Thermal cycling in microgravity resulted in the trapping of gas bubbles inside septa cap assay tubes, causing small but measurable increases in Ct curve noise and variability. Bubble formation was successfully suppressed in a rapid follow-up on-orbit experiment using standard caps to pressurize PCR tubes and reduce gas release during heating cycles. The WetLab-2 facility now provides a novel operational on-orbit research capability for

  7. Evaluation of Reference Genes for Real-Time Quantitative PCR Analysis in Larvae of Spodoptera litura Exposed to Azadirachtin Stress Conditions

    OpenAIRE

    Benshui Shu; Jingjing Zhang; Gaofeng Cui; Ranran Sun; Veeran Sethuraman; Xin Yi; Guohua Zhong

    2018-01-01

    Azadirachtin is an efficient and broad-spectrum botanical insecticide against more than 150 kinds of agricultural pests with the effects of mortality, antifeedant and growth regulation. Real-time quantitative polymerase chain reaction (RT-qPCR) could be one of the powerful tools to analyze the gene expression level and investigate the mechanism of azadirachtin at transcriptional level, however, the ideal reference genes are needed to normalize the expression profiling of target genes. In this...

  8. Evaluation of Reference Genes for Real-Time Quantitative PCR Analysis in Larvae of Spodoptera litura Exposed to Azadirachtin Stress Conditions

    Directory of Open Access Journals (Sweden)

    Benshui Shu

    2018-04-01

    Full Text Available Azadirachtin is an efficient and broad-spectrum botanical insecticide against more than 150 kinds of agricultural pests with the effects of mortality, antifeedant and growth regulation. Real-time quantitative polymerase chain reaction (RT-qPCR could be one of the powerful tools to analyze the gene expression level and investigate the mechanism of azadirachtin at transcriptional level, however, the ideal reference genes are needed to normalize the expression profiling of target genes. In this present study, the fragments of eight candidate reference genes were cloned and identified from the pest Spodoptera litura. In addition, the expression stability of these genes in different samples from larvae of control and azadirachtin treatments were evaluated by the computational methods of NormFinder, BestKeeper, Delta CT, geNorm, and RefFinder. According to our results, two of the reference genes should be the optimal number for RT-qPCR analysis. Furthermore, the best reference genes for different samples were showed as followed: EF-1α and EF2 for cuticle, β-Tubulin and RPL7A for fat body, EF2 and Actin for midgut, EF2 and RPL13A for larva and RPL13A and RPL7A for all the samples. Our results established a reliable normalization for RT-qPCR experiments in S. litura and ensure the data more accurate for the mechanism analysis of azadirachtin.

  9. Evaluation of Reference Genes for Real-Time Quantitative PCR Analysis in Larvae of Spodoptera litura Exposed to Azadirachtin Stress Conditions.

    Science.gov (United States)

    Shu, Benshui; Zhang, Jingjing; Cui, Gaofeng; Sun, Ranran; Sethuraman, Veeran; Yi, Xin; Zhong, Guohua

    2018-01-01

    Azadirachtin is an efficient and broad-spectrum botanical insecticide against more than 150 kinds of agricultural pests with the effects of mortality, antifeedant and growth regulation. Real-time quantitative polymerase chain reaction (RT-qPCR) could be one of the powerful tools to analyze the gene expression level and investigate the mechanism of azadirachtin at transcriptional level, however, the ideal reference genes are needed to normalize the expression profiling of target genes. In this present study, the fragments of eight candidate reference genes were cloned and identified from the pest Spodoptera litura . In addition, the expression stability of these genes in different samples from larvae of control and azadirachtin treatments were evaluated by the computational methods of NormFinder, BestKeeper, Delta CT, geNorm, and RefFinder. According to our results, two of the reference genes should be the optimal number for RT-qPCR analysis. Furthermore, the best reference genes for different samples were showed as followed: EF-1α and EF2 for cuticle, β-Tubulin and RPL7A for fat body, EF2 and Actin for midgut, EF2 and RPL13A for larva and RPL13A and RPL7A for all the samples. Our results established a reliable normalization for RT-qPCR experiments in S. litura and ensure the data more accurate for the mechanism analysis of azadirachtin.

  10. MATtrack: A MATLAB-Based Quantitative Image Analysis Platform for Investigating Real-Time Photo-Converted Fluorescent Signals in Live Cells.

    Science.gov (United States)

    Courtney, Jane; Woods, Elena; Scholz, Dimitri; Hall, William W; Gautier, Virginie W

    2015-01-01

    We introduce here MATtrack, an open source MATLAB-based computational platform developed to process multi-Tiff files produced by a photo-conversion time lapse protocol for live cell fluorescent microscopy. MATtrack automatically performs a series of steps required for image processing, including extraction and import of numerical values from Multi-Tiff files, red/green image classification using gating parameters, noise filtering, background extraction, contrast stretching and temporal smoothing. MATtrack also integrates a series of algorithms for quantitative image analysis enabling the construction of mean and standard deviation images, clustering and classification of subcellular regions and injection point approximation. In addition, MATtrack features a simple user interface, which enables monitoring of Fluorescent Signal Intensity in multiple Regions of Interest, over time. The latter encapsulates a region growing method to automatically delineate the contours of Regions of Interest selected by the user, and performs background and regional Average Fluorescence Tracking, and automatic plotting. Finally, MATtrack computes convenient visualization and exploration tools including a migration map, which provides an overview of the protein intracellular trajectories and accumulation areas. In conclusion, MATtrack is an open source MATLAB-based software package tailored to facilitate the analysis and visualization of large data files derived from real-time live cell fluorescent microscopy using photoconvertible proteins. It is flexible, user friendly, compatible with Windows, Mac, and Linux, and a wide range of data acquisition software. MATtrack is freely available for download at eleceng.dit.ie/courtney/MATtrack.zip.

  11. Statistical aspects of quantitative real-time PCR experiment design.

    Science.gov (United States)

    Kitchen, Robert R; Kubista, Mikael; Tichopad, Ales

    2010-04-01

    Experiments using quantitative real-time PCR to test hypotheses are limited by technical and biological variability; we seek to minimise sources of confounding variability through optimum use of biological and technical replicates. The quality of an experiment design is commonly assessed by calculating its prospective power. Such calculations rely on knowledge of the expected variances of the measurements of each group of samples and the magnitude of the treatment effect; the estimation of which is often uninformed and unreliable. Here we introduce a method that exploits a small pilot study to estimate the biological and technical variances in order to improve the design of a subsequent large experiment. We measure the variance contributions at several 'levels' of the experiment design and provide a means of using this information to predict both the total variance and the prospective power of the assay. A validation of the method is provided through a variance analysis of representative genes in several bovine tissue-types. We also discuss the effect of normalisation to a reference gene in terms of the measured variance components of the gene of interest. Finally, we describe a software implementation of these methods, powerNest, that gives the user the opportunity to input data from a pilot study and interactively modify the design of the assay. The software automatically calculates expected variances, statistical power, and optimal design of the larger experiment. powerNest enables the researcher to minimise the total confounding variance and maximise prospective power for a specified maximum cost for the large study. Copyright 2010 Elsevier Inc. All rights reserved.

  12. Prospective Qualitative and Quantitative Analysis of Real-Time Peer Review Quality Assurance Rounds Incorporating Direct Physical Examination for Head and Neck Cancer Radiation Therapy.

    Science.gov (United States)

    Cardenas, Carlos E; Mohamed, Abdallah S R; Tao, Randa; Wong, Andrew J R; Awan, Mussadiq J; Kuruvila, Shirly; Aristophanous, Michalis; Gunn, G Brandon; Phan, Jack; Beadle, Beth M; Frank, Steven J; Garden, Adam S; Morrison, William H; Fuller, Clifton D; Rosenthal, David I

    2017-07-01

    Our department has a long-established comprehensive quality assurance (QA) planning clinic for patients undergoing radiation therapy (RT) for head and neck cancer. Our aim is to assess the impact of a real-time peer review QA process on the quantitative and qualitative radiation therapy plan changes in the era of intensity modulated RT (IMRT). Prospective data for 85 patients undergoing head and neck IMRT who presented at a biweekly QA clinic after simulation and contouring were collected. A standard data collection form was used to document alterations made during this process. The original pre-QA clinical target volumes (CTVs) approved by the treating-attending physicians were saved before QA and compared with post-QA consensus CTVs. Qualitative assessment was done according to predefined criteria. Dice similarity coefficients (DSC) and other volume overlap metrics were calculated for each CTV level and were used for quantitative comparison. Changes are categorized as major, minor, and trivial according to the degree of overlap. Patterns of failure were analyzed and correlated to plan changes. All 85 patients were examined by at least 1 head and neck subspecialist radiation oncologist who was not the treating-attending physician; 80 (94%) were examined by ≥3 faculty members. New clinical findings on physical examination were found in 12 patients (14%) leading to major plan changes. Quantitative DSC analysis revealed significantly better agreement in CTV1 (0.94 ± 0.10) contours than in CTV2 (0.82 ± 0.25) and CTV3 (0.86 ± 0.2) contours (P=.0002 and P=.03, respectively; matched-pair Wilcoxon test). The experience of the treating-attending radiation oncologist significantly affected DSC values when all CTV levels were considered (P=.012; matched-pair Wilcoxon text). After a median follow-up time of 38 months, only 10 patients (12%) had local recurrence, regional recurrence, or both, mostly in central high-dose areas. Comprehensive peer review planning

  13. A plastome primer set for comprehensive quantitative real time RT-PCR analysis of Zea mays: a starter primer set for other Poaceae species

    Directory of Open Access Journals (Sweden)

    Dunn Sade N

    2008-06-01

    Full Text Available Abstract Background Quantitative Real Time RT-PCR (q2(RTPCR is a maturing technique which gives researchers the ability to quantify and compare very small amounts of nucleic acids. Primer design and optimization is an essential yet time consuming aspect of using q2(RTPCR. In this paper we describe the design and empirical optimization of primers to amplify and quantify plastid RNAs from Zea mays that are robust enough to use with other closely related species. Results Primers were designed and successfully optimized for 57 of the 104 reported genes in the maize plastome plus two nuclear genes. All 59 primer pairs produced single amplicons after end-point reverse transcriptase polymerase chain reactions (RT-PCR as visualized on agarose gels and subsequently verified by q2(RTPCR. Primer pairs were divided into several categories based on the optimization requirements or the uniqueness of the target gene. An in silico test suggested the majority of the primer sets should work with other members of the Poaceae family. An in vitro test of the primer set on two unsequenced species (Panicum virgatum and Miscanthus sinensis supported this assumption by successfully producing single amplicons for each primer pair. Conclusion Due to the highly conserved chloroplast genome in plant families it is possible to utilize primer pairs designed against one genomic sequence to detect the presence and abundance of plastid genes or transcripts from genomes that have yet to be sequenced. Analysis of steady state transcription of vital system genes is a necessary requirement to comprehensively elucidate gene expression in any organism. The primer pairs reported in this paper were designed for q2(RTPCR of maize chloroplast genes but should be useful for other members of the Poaceae family. Both in silico and in vitro data are presented to support this assumption.

  14. Adhesion of mutans streptococci to self-ligating ceramic brackets: in vivo quantitative analysis with real-time polymerase chain reaction.

    Science.gov (United States)

    Jung, Woo-Sun; Yang, Il-Hyung; Lim, Won Hee; Baek, Seung-Hak; Kim, Tae-Woo; Ahn, Sug-Joon

    2015-12-01

    To analyze in vivo mutans streptococci (MS) adhesion to self-ligating ceramic brackets [Clarity-SL (CSL) and Clippy-C (CC)] and the relationships between bacterial adhesion and oral hygiene indices. Four central incisor brackets from the maxilla and mandible were collected from 40 patients (20 patients per each bracket type) at debonding immediately after plaque and gingival indices were measured. Adhesions of Streptococcus mutans, S. sobrinus, and total bacteria were quantitatively determined using real-time polymerase chain reaction after genomic DNA was extracted. Factorial analysis of variance was used to analyze bacterial adhesion to the brackets with respect to the bracket type and jaw position. Correlation coefficients were calculated to determine the relationships of bacterial adhesion to oral hygiene indices. Adhesion of total bacteria and S. mutans to CSL was higher than that to CC (P brackets was higher than that to the maxillary ones (P brackets were higher than that in the mandibular ones (P brackets and jaw positions. Interestingly, no significant relationships were found between bacterial adhesions and oral hygiene indices. Complex bracket configurations may significantly influence bacterial adhesion to orthodontic brackets. Further in vivo study using bracket raw materials will help to define the relationships between bacteria adhesion and enamel demineralization. Because oral hygiene indices were not significantly correlated with adhesions of MS to self-ligating ceramic brackets, careful examinations around the brackets should be needed to prevent enamel demineralization, regardless of oral hygiene status. © The Author 2015. Published by Oxford University Press on behalf of the European Orthodontic Society. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  15. Selection and validation of reference genes for gene expression analysis in switchgrass (Panicum virgatum using quantitative real-time RT-PCR.

    Directory of Open Access Journals (Sweden)

    Jacinta Gimeno

    Full Text Available Switchgrass (Panicum virgatum has received a lot of attention as a forage and bioenergy crop during the past few years. Gene expression studies are in progress to improve new traits and develop new cultivars. Quantitative real time PCR (qRT-PCR has emerged as an important technique to study gene expression analysis. For accurate and reliable results, normalization of data with reference genes is essential. In this work, we evaluate the stability of expression of genes to use as reference for qRT-PCR in the grass P. virgatum. Eleven candidate reference genes, including eEF-1α, UBQ6, ACT12, TUB6, eIF-4a, GAPDH, SAMDC, TUA6, CYP5, U2AF, and FTSH4, were validated for qRT-PCR normalization in different plant tissues and under different stress conditions. The expression stability of these genes was verified by the use of two distinct algorithms, geNorm and NormFinder. Differences were observed after comparison of the ranking of the candidate reference genes identified by both programs but eEF-1α, eIF-4a, CYP5 and U2AF are ranked as the most stable genes in the samples sets under study. Both programs discard the use of SAMDC and TUA6 for normalization. Validation of the reference genes proposed by geNorm and NormFinder were performed by normalization of transcript abundance of a group of target genes in different samples. Results show similar expression patterns when the best reference genes selected by both programs were used but differences were detected in the transcript abundance of the target genes. Based on the above research, we recommend the use of different statistical algorithms to identify the best reference genes for expression data normalization. The best genes selected in this study will help to improve the quality of gene expression data in a wide variety of samples in switchgrass.

  16. Diagnosis of aerobic vaginitis by quantitative real-time PCR.

    Science.gov (United States)

    Rumyantseva, T A; Bellen, G; Savochkina, Y A; Guschin, A E; Donders, G G G

    2016-07-01

    To evaluate a real-time PCR-based technique to quantify bacteria associated with aerobic vaginitis (AV) as a potential test. Vaginal samples from 100 women were tested by wet-mount microscopy, gram stain and quantitative real-time PCR targeting Enterobacteriacea, Staphylococcus spp., Streptococcus spp., Enterococcus spp., Escherichia coli, Streptococcus agalactiae, S. aureus; Lactobacillus spp. AV diagnosis obtained by wet-mount microscopy was used as reference. Some level of AV was diagnosed in 23 (23.7 %) cases. Various concentrations of Enterobacteriacea, Staphylococcus spp., Streptococcus spp. were detected an all patients. Enterococcus spp. were detected in 76 (78.3 %) cases. Summarized concentrations of aerobes were tenfold higher in AV-positive compared to AV-negative cases [7.30lg vs 6.06lg (p = 0.02)]. Concentrations of aerobes in severe, moderate and light AV cases did not vary significantly (p = 0.14). Concentration of lactobacilli was 1000-fold lower in AV-positive cases compared to normal cases (5.3lg vs 8.3lg, p AV-positive cases [19/22 (86.4 %) samples]. The relation of high loads of aerobes to the low numbers of Lactobacilli are a reliable marker for the presence of AV and could substitute microscopy as a test. PCR may be a good standardized substitution for AV diagnosis in settings where well-trained microscopists are lacking.

  17. Quantification of Human Fecal Bifidobacterium Species by Use of Quantitative Real-Time PCR Analysis Targeting the groEL Gene

    Science.gov (United States)

    Junick, Jana

    2012-01-01

    Quantitative real-time PCR assays targeting the groEL gene for the specific enumeration of 12 human fecal Bifidobacterium species were developed. The housekeeping gene groEL (HSP60 in eukaryotes) was used as a discriminative marker for the differentiation of Bifidobacterium adolescentis, B. angulatum, B. animalis, B. bifidum, B. breve, B. catenulatum, B. dentium, B. gallicum, B. longum, B. pseudocatenulatum, B. pseudolongum, and B. thermophilum. The bifidobacterial chromosome contains a single copy of the groEL gene, allowing the determination of the cell number by quantification of the groEL copy number. Real-time PCR assays were validated by comparing fecal samples spiked with known numbers of a given Bifidobacterium species. Independent of the Bifidobacterium species tested, the proportion of groEL copies recovered from fecal samples spiked with 5 to 9 log10 cells/g feces was approximately 50%. The quantification limit was 5 to 6 log10 groEL copies/g feces. The interassay variability was less than 10%, and variability between different DNA extractions was less than 23%. The method developed was applied to fecal samples from healthy adults and full-term breast-fed infants. Bifidobacterial diversity in both adults and infants was low, with mostly ≤3 Bifidobacterium species and B. longum frequently detected. The predominant species in infant and adult fecal samples were B. breve and B. adolescentis, respectively. It was possible to distinguish B. catenulatum and B. pseudocatenulatum. We conclude that the groEL gene is a suitable molecular marker for the specific and accurate quantification of human fecal Bifidobacterium species by real-time PCR. PMID:22307308

  18. Multiplex real-time quantitative PCR, microscopy and rapid diagnostic immuno-chromatographic tests for the detection of Plasmodium spp: performance, limit of detection analysis and quality assurance

    Directory of Open Access Journals (Sweden)

    Ralevski Filip

    2009-12-01

    Full Text Available Abstract Background Accurate laboratory diagnosis of malaria species in returning travelers is paramount in the treatment of this potentially fatal infectious disease. Materials and methods A total of 466 blood specimens from returning travelers to Africa, Asia, and South/Central America with suspected malaria infection were collected between 2007 and 2009 at the reference public health laboratory. These specimens were assessed by reference microscopy, multipex real-time quantitative polymerase chain reaction (QPCR, and two rapid diagnostic immuno-chromatographic tests (ICT in a blinded manner. Key clinical laboratory parameters such as limit of detection (LOD analysis on clinical specimens by parasite stage, inter-reader variability of ICTs, staffing implications, quality assurance and cost analysis were evaluated. Results QPCR is the most analytically sensitive method (sensitivity 99.41%, followed by CARESTART (sensitivity 88.24%, and BINAXNOW (sensitivity 86.47% for the diagnosis of malaria in returning travelers when compared to reference microscopy. However, microscopy was unable to specifically identify Plasmodia spp. in 18 out of 170 positive samples by QPCR. Moreover, the 17 samples that were negative by microscopy and positive by QPCR were also positive by ICTs. Quality assurance was achieved for QPCR by exchanging a blinded proficiency panel with another reference laboratory. The Kappa value of inter-reader variability among three readers for BINAXNOW and CARESTART was calculated to be 0.872 and 0.898 respectively. Serial dilution studies demonstrated that the QPCR cycle threshold correlates linearly with parasitemia (R2 = 0.9746 in a clinically relevant dynamic range and retains a LOD of 11 rDNA copies/μl for P. falciparum, which was several log lower than reference microscopy and ICTs. LOD for QPCR is affected not only by parasitemia but the parasite stage distribution of each clinical specimen. QPCR was approximately 6-fold more

  19. Real-time quantitative reverse transcription-PCR analysis of expression stability of Actinobacillus pleuropneumoniae housekeeping genes during in vitro growth under iron-depleted conditions

    DEFF Research Database (Denmark)

    Nielsen, K. K.; Boye, Mette

    2005-01-01

    up-regulation under iron-restricted conditions compared to bacteria grown in medium with sufficient iron. The observed expression patterns of the genes of interest were consistent with previous observations. This study therefore lends further support to the use of real-time quantitative RT...... controls, as such controls have not been defined yet for this bacterium. Bacterial gene expression was studied during in vitro exponential and early stationary growth in medium with and without sufficient iron, respectively. First, the stability of expression of five genes, the glyA, tpiA, pykA, rec......F, and rhoAP genes involved in basic housekeeping, was evaluated on the basis of the mean pairwise variation. All the housekeeping genes included were stably expressed under the conditions investigated and consequently were included in the normalization procedure. Next, the geometric mean of the internal...

  20. High-level expression of podoplanin in benign and malignant soft tissue tumors: immunohistochemical and quantitative real-time RT-PCR analysis.

    Science.gov (United States)

    Xu, Yongjun; Ogose, Akira; Kawashima, Hiroyuki; Hotta, Tetsuo; Ariizumi, Takashi; Li, Guidong; Umezu, Hajime; Endo, Naoto

    2011-03-01

    Podoplanin is a 38 kDa mucin-type transmembrane glycoprotein that was first identified in rat glomerular epithelial cells (podocytes). It is expressed in normal lymphatic endothelium, but is absent from vascular endothelial cells. D2-40 is a commercially available mouse monoclonal antibody which binds to an epitope on human podoplanin. D2-40 immunoreactivity is therefore highly sensitive and specific for lymphatic endothelium. Recent investigations have shown widespread applications of immunohistochemical staining with D2-40 in evaluating podoplanin expression as an immunohistochemical marker for diagnosis and prognosis in various tumors. To determine whether the podoplanin (D2-40) antibody may be useful for the diagnosis of soft tissue tumors, 125 cases, including 4 kinds of benign tumors, 15 kinds of malignant tumors and 3 kinds of tumor-like lesions were immunostained using the D2-40 antibody. Total RNA was extracted from frozen tumor tissue obtained from 41 corresponding soft tissue tumor patients and 12 kinds of soft tissue tumor cell lines. Quantitative real-time PCR reactions were performed. Immunohistochemical and quantitative real-time RT-PCR analyses demonstrated the expression of the podoplanin protein and mRNA in the majority of benign and malignant soft tissue tumors and tumor-like lesions examined, with the exception of alveolar soft part sarcoma, embryonal and alveolar rhabdomyosarcoma, extraskeletal Ewing's sarcoma/peripheral primitive neuro-ectodermal tumor and lipoma, which were completely negative for podoplanin. Since it is widely and highly expressed in nearly all kinds of soft tissue tumors, especially in spindle cell sarcoma, myxoid type soft tissue tumors and soft tissue tumors of the nervous system, podoplanin is considered to have little value in the differential diagnosis of soft tissue tumors.

  1. Real-time Forensic Disaster Analysis

    Science.gov (United States)

    Wenzel, F.; Daniell, J.; Khazai, B.; Mühr, B.; Kunz-Plapp, T.; Markus, M.; Vervaeck, A.

    2012-04-01

    The Center for Disaster Management and Risk Reduction Technology (CEDIM, www.cedim.de) - an interdisciplinary research center founded by the German Research Centre for Geoscience (GFZ) and Karlsruhe Institute of Technology (KIT) - has embarked on a new style of disaster research known as Forensic Disaster Analysis. The notion has been coined by the Integrated Research on Disaster Risk initiative (IRDR, www.irdrinternational.org) launched by ICSU in 2010. It has been defined as an approach to studying natural disasters that aims at uncovering the root causes of disasters through in-depth investigations that go beyond the reconnaissance reports and case studies typically conducted after disasters. In adopting this comprehensive understanding of disasters CEDIM adds a real-time component to the assessment and evaluation process. By comprehensive we mean that most if not all relevant aspects of disasters are considered and jointly analysed. This includes the impact (human, economy, and infrastructure), comparisons with recent historic events, social vulnerability, reconstruction and long-term impacts on livelihood issues. The forensic disaster analysis research mode is thus best characterized as "event-based research" through systematic investigation of critical issues arising after a disaster across various inter-related areas. The forensic approach requires (a) availability of global data bases regarding previous earthquake losses, socio-economic parameters, building stock information, etc.; (b) leveraging platforms such as the EERI clearing house, relief-web, and the many sources of local and international sources where information is organized; and (c) rapid access to critical information (e.g., crowd sourcing techniques) to improve our understanding of the complex dynamics of disasters. The main scientific questions being addressed are: What are critical factors that control loss of life, of infrastructure, and for economy? What are the critical interactions

  2. Real-time quantitative PCR of microdissected paraffin-embedded breast carcinoma

    DEFF Research Database (Denmark)

    Gjerdrum, Lise Mette; Sorensen, Boe Sandahl; Kjeldsen, Eigil

    2004-01-01

    We studied the feasibility of using real-time quantitative PCR to determine HER-2 DNA amplification and mRNA expression in microdissected formalin-fixed, paraffin-embedded breast tumors and compared this with standard immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) methods...... tumors as being amplified. Interestingly, all these scored 2+ with the HercepTest, but were negative using FISH. We believe that real-time quantitative PCR analysis of HER-2 DNA amplification following microdissection represents a useful supplementary or perhaps even an alternative technique...

  3. The numbers game: quantitative analysis of Neorickettsia sp. propagation through complex life cycle of its digenean host using real-time qPCR.

    Science.gov (United States)

    Greiman, Stephen E; Tkach, Vasyl V

    2016-07-01

    Bacteria of the genus Neorickettsia are obligate intracellular endosymbionts of parasitic flukes (Digenea) and are passed through the entire complex life cycle of the parasite by vertical transmission. Several species of Neorickettsia are known to cause diseases in domestic animals, wildlife, and humans. Quantitative data on the transmission of the bacteria through the digenean life cycle is almost completely lacking. This study quantified for the first time the abundance of Neorickettsia within multiple stages of the life cycle of the digenean Plagiorchis elegans. Snails Lymnaea stagnalis collected from a pond in North Dakota were screened for the presence of digenean cercariae, which were subsequently tested for the presence of Neorickettsia. Three L. stagnalis were found shedding P. elegans cercariae infected with Neorickettsia. These snails were used to initiate three separate laboratory life cycles and obtain all life cycle stages for bacterial quantification. A quantitative real-time PCR assay targeting the GroEL gene was developed to enumerate Neorickettsia sp. within different stages of the digenean life cycle. The number of bacteria significantly increased throughout all stages, from eggs to adults. The two largest increases in number of bacteria occurred during the period from eggs to cercariae and from 6-day metacercariae to 48-h juvenile worms. These two periods seem to be the most important for Neorickettsia propagation through the complex digenean life cycle and maturation in the definitive host.

  4. Comparison of three-way and four-way calibration for the real-time quantitative analysis of drug hydrolysis in complex dynamic samples by excitation-emission matrix fluorescence.

    Science.gov (United States)

    Yin, Xiao-Li; Gu, Hui-Wen; Liu, Xiao-Lu; Zhang, Shan-Hui; Wu, Hai-Long

    2018-03-05

    Multiway calibration in combination with spectroscopic technique is an attractive tool for online or real-time monitoring of target analyte(s) in complex samples. However, how to choose a suitable multiway calibration method for the resolution of spectroscopic-kinetic data is a troubling problem in practical application. In this work, for the first time, three-way and four-way fluorescence-kinetic data arrays were generated during the real-time monitoring of the hydrolysis of irinotecan (CPT-11) in human plasma by excitation-emission matrix fluorescence. Alternating normalization-weighted error (ANWE) and alternating penalty trilinear decomposition (APTLD) were used as three-way calibration for the decomposition of the three-way kinetic data array, whereas alternating weighted residual constraint quadrilinear decomposition (AWRCQLD) and alternating penalty quadrilinear decomposition (APQLD) were applied as four-way calibration to the four-way kinetic data array. The quantitative results of the two kinds of calibration models were fully compared from the perspective of predicted real-time concentrations, spiked recoveries of initial concentration, and analytical figures of merit. The comparison study demonstrated that both three-way and four-way calibration models could achieve real-time quantitative analysis of the hydrolysis of CPT-11 in human plasma under certain conditions. However, it was also found that both of them possess some critical advantages and shortcomings during the process of dynamic analysis. The conclusions obtained in this paper can provide some helpful guidance for the reasonable selection of multiway calibration models to achieve the real-time quantitative analysis of target analyte(s) in complex dynamic systems. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Comparison of three-way and four-way calibration for the real-time quantitative analysis of drug hydrolysis in complex dynamic samples by excitation-emission matrix fluorescence

    Science.gov (United States)

    Yin, Xiao-Li; Gu, Hui-Wen; Liu, Xiao-Lu; Zhang, Shan-Hui; Wu, Hai-Long

    2018-03-01

    Multiway calibration in combination with spectroscopic technique is an attractive tool for online or real-time monitoring of target analyte(s) in complex samples. However, how to choose a suitable multiway calibration method for the resolution of spectroscopic-kinetic data is a troubling problem in practical application. In this work, for the first time, three-way and four-way fluorescence-kinetic data arrays were generated during the real-time monitoring of the hydrolysis of irinotecan (CPT-11) in human plasma by excitation-emission matrix fluorescence. Alternating normalization-weighted error (ANWE) and alternating penalty trilinear decomposition (APTLD) were used as three-way calibration for the decomposition of the three-way kinetic data array, whereas alternating weighted residual constraint quadrilinear decomposition (AWRCQLD) and alternating penalty quadrilinear decomposition (APQLD) were applied as four-way calibration to the four-way kinetic data array. The quantitative results of the two kinds of calibration models were fully compared from the perspective of predicted real-time concentrations, spiked recoveries of initial concentration, and analytical figures of merit. The comparison study demonstrated that both three-way and four-way calibration models could achieve real-time quantitative analysis of the hydrolysis of CPT-11 in human plasma under certain conditions. However, it was also found that both of them possess some critical advantages and shortcomings during the process of dynamic analysis. The conclusions obtained in this paper can provide some helpful guidance for the reasonable selection of multiway calibration models to achieve the real-time quantitative analysis of target analyte(s) in complex dynamic systems.

  6. A quantitative method to track protein translocation between intracellular compartments in real-time in live cells using weighted local variance image analysis.

    Directory of Open Access Journals (Sweden)

    Guillaume Calmettes

    Full Text Available The genetic expression of cloned fluorescent proteins coupled to time-lapse fluorescence microscopy has opened the door to the direct visualization of a wide range of molecular interactions in living cells. In particular, the dynamic translocation of proteins can now be explored in real time at the single-cell level. Here we propose a reliable, easy-to-implement, quantitative image processing method to assess protein translocation in living cells based on the computation of spatial variance maps of time-lapse images. The method is first illustrated and validated on simulated images of a fluorescently-labeled protein translocating from mitochondria to cytoplasm, and then applied to experimental data obtained with fluorescently-labeled hexokinase 2 in different cell types imaged by regular or confocal microscopy. The method was found to be robust with respect to cell morphology changes and mitochondrial dynamics (fusion, fission, movement during the time-lapse imaging. Its ease of implementation should facilitate its application to a broad spectrum of time-lapse imaging studies.

  7. Real-time quantitative PCR of Staphylococcus aureus and application in restaurant meals.

    Science.gov (United States)

    Berrada, H; Soriano, J M; Mañes, J; Picó, Y

    2006-01-01

    Staphylococcus aureus is considered the second most common pathogen to cause outbreaks of food poisoning, exceeded only by Campylobacter. Consumption of foods containing this microorganism is often identified as the cause of illness. In this study, a rapid, reliable, and sensitive real-time quantitative PCR was developed and compared with conventional culture methods. Real-time quantitative PCR was carried out by purifying DNA extracts of S. aureus with a Staphylococcus sample preparation kit and quantifying it in the LightCycler system with hybridization probes. The assay was linear from a range of 10 to 10(6) S. aureus cells (r2 > 0.997). The PCR reaction presented an efficiency of >85%. Accuracy of the PCR-based assay, expressed as percent bias, was around 13%, and the precision, expressed as a percentage of the coefficient of variation, was 7 to 10%. Intraday and interday variability were studied at 10(2) CFU/g and was 12 and 14%, respectively. The proposed method was applied to the analysis of 77 samples of restaurant meals in Valencia (Spain). In 11.6% of samples S. aureus was detected by real-time quantitative PCR, as well as by the conventional microbiological method. An excellent correspondence between real-time quantitative PCR and microbiological numbers (CFU/g) was observed with deviations of < 28%.

  8. Human parvovirus B19, varicella zoster virus, and human herpesvirus-6 in mesenchymal stem cells of patients with osteoarthritis: analysis with quantitative real-time polymerase chain reaction.

    Science.gov (United States)

    Rollín, R; Alvarez-Lafuente, R; Marco, F; Jover, J A; Hernández-García, C; Rodríguez-Navas, C; López-Durán, L; Fernández-Gutiérrez, B

    2007-04-01

    To investigate whether there is a possible viral transmission using mesenchymal stem cells (MSCs) in autologous or allogeneic transplantation in the context of osteoarthritis (OA) patients. The presence of parvovirus B19 (B19), varicella zoster virus (VZV), and human herpesvirus-6 (HHV-6) was studied in MSCs from bone marrow of patients with OA and healthy controls. MSCs were prepared from bone marrow aspirates obtained from 18 patients undergoing joint replacement as a result of OA and from 10 healthy controls. DNA was extracted from primary MSCs' culture established from these cells and quantitative real-time polymerase chain reaction was performed to analyse the prevalence and viral load of B19, VZV and HHV-6. The prevalence of total viral DNA among patients with OA was 16.7% (3/18), with a mean viral load of 29.7 copies/microg of DNA. One out of 18 was positive for B19 (viral load, 61.2 copies/microg of DNA), two for VZV (mean viral load, 14.4 copies/microg of DNA), and none for HHV-6. The prevalence of total viral DNA in the control group was 20% (2/10), with a mean viral load of 13.4 copies/microg of DNA. Both positive results were of B19 parvoviruses. There were no statistically significant differences among patients and controls. This first approach to the viral prevalence in MSCs of bone marrow in OA patients and healthy controls seems to show a very low risk of viral transmission or reactivation in a possible MSCs' transplantation.

  9. Identification of normalization factors for quantitative real-time RT-PCR analysis of gene expression in Pacific abalone Haliotis discus hannai

    Science.gov (United States)

    Qiu, Reng; Sun, Boguang; Fang, Shasha; Sun, Li; Liu, Xiao

    2013-03-01

    Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) is widely used in studies of gene expression. In most of these studies, housekeeping genes are used as internal references without validation. To identify appropriate reference genes for qRT-PCR in Pacific abalone Haliotis discus hannai, we examined the transcription stability of six housekeeping genes in abalone tissues in the presence and absence of bacterial infection. For this purpose, abalone were infected with the bacterial pathogen Vibrio anguillarum for 12 h and 48 h. The mRNA levels of the housekeeping genes in five tissues (digestive glands, foot muscle, gill, hemocyte, and mantle) were determined by qRT-PCR. The PCR data was subsequently analyzed with the geNorm and NormFinder algorithms. The results show that in the absence of bacterial infection, elongation factor-1-alpha and beta-actin were the most stably expressed genes in all tissues, and thus are suitable as cross-tissue type normalization factors. However, we did not identify any universal reference genes post infection because the most stable genes varied between tissue types. Furthermore, for most tissues, the optimal reference genes identified by both algorithms at 12 h and 48 h post-infection differed. These results indicate that bacterial infection induced significant changes in the expression of abalone housekeeping genes in a manner that is dependent on tissue type and duration of infection. As a result, different normalization factors must be used for different tissues at different infection points.

  10. Identification and validation of quantitative real-time reverse transcription PCR reference genes for gene expression analysis in teak (Tectona grandis L.f.).

    Science.gov (United States)

    Galeano, Esteban; Vasconcelos, Tarcísio Sales; Ramiro, Daniel Alves; De Martin, Valentina de Fátima; Carrer, Helaine

    2014-07-22

    Teak (Tectona grandis L.f.) is currently the preferred choice of the timber trade for fabrication of woody products due to its extraordinary qualities and is widely grown around the world. Gene expression studies are essential to explore wood formation of vascular plants, and quantitative real-time reverse transcription PCR (qRT-PCR) is a sensitive technique employed for quantifying gene expression levels. One or more appropriate reference genes are crucial to accurately compare mRNA transcripts through different tissues/organs and experimental conditions. Despite being the focus of some genetic studies, a lack of molecular information has hindered genetic exploration of teak. To date, qRT-PCR reference genes have not been identified and validated for teak. Identification and cloning of nine commonly used qRT-PCR reference genes from teak, including ribosomal protein 60s (rp60s), clathrin adaptor complexes medium subunit family (Cac), actin (Act), histone 3 (His3), sand family (Sand), β-Tubulin (Β-Tub), ubiquitin (Ubq), elongation factor 1-α (Ef-1α), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Expression profiles of these genes were evaluated by qRT-PCR in six tissue and organ samples (leaf, flower, seedling, root, stem and branch secondary xylem) of teak. Appropriate gene cloning and sequencing, primer specificity and amplification efficiency was verified for each gene. Their stability as reference genes was validated by NormFinder, BestKeeper, geNorm and Delta Ct programs. Results obtained from all programs showed that TgUbq and TgEf-1α are the most stable genes to use as qRT-PCR reference genes and TgAct is the most unstable gene in teak. The relative expression of the teak cinnamyl alcohol dehydrogenase (TgCAD) gene in lignified tissues at different ages was assessed by qRT-PCR, using TgUbq and TgEf-1α as internal controls. These analyses exposed a consistent expression pattern with both reference genes. This study proposes a first broad

  11. Quantitative analysis of aortic regurgitation: real-time 3-dimensional and 2-dimensional color Doppler echocardiographic method--a clinical and a chronic animal study

    Science.gov (United States)

    Shiota, Takahiro; Jones, Michael; Tsujino, Hiroyuki; Qin, Jian Xin; Zetts, Arthur D.; Greenberg, Neil L.; Cardon, Lisa A.; Panza, Julio A.; Thomas, James D.

    2002-01-01

    BACKGROUND: For evaluating patients with aortic regurgitation (AR), regurgitant volumes, left ventricular (LV) stroke volumes (SV), and absolute LV volumes are valuable indices. AIM: The aim of this study was to validate the combination of real-time 3-dimensional echocardiography (3DE) and semiautomated digital color Doppler cardiac flow measurement (ACM) for quantifying absolute LV volumes, LVSV, and AR volumes using an animal model of chronic AR and to investigate its clinical applicability. METHODS: In 8 sheep, a total of 26 hemodynamic states were obtained pharmacologically 20 weeks after the aortic valve noncoronary (n = 4) or right coronary (n = 4) leaflet was incised to produce AR. Reference standard LVSV and AR volume were determined using the electromagnetic flow method (EM). Simultaneous epicardial real-time 3DE studies were performed to obtain LV end-diastolic volumes (LVEDV), end-systolic volumes (LVESV), and LVSV by subtracting LVESV from LVEDV. Simultaneous ACM was performed to obtain LVSV and transmitral flows; AR volume was calculated by subtracting transmitral flow volume from LVSV. In a total of 19 patients with AR, real-time 3DE and ACM were used to obtain LVSVs and these were compared with each other. RESULTS: A strong relationship was found between LVSV derived from EM and those from the real-time 3DE (r = 0.93, P <.001, mean difference (3D - EM) = -1.0 +/- 9.8 mL). A good relationship between LVSV and AR volumes derived from EM and those by ACM was found (r = 0.88, P <.001). A good relationship between LVSV derived from real-time 3DE and that from ACM was observed (r = 0.73, P <.01, mean difference = 2.5 +/- 7.9 mL). In patients, a good relationship between LVSV obtained by real-time 3DE and ACM was found (r = 0.90, P <.001, mean difference = 0.6 +/- 9.8 mL). CONCLUSION: The combination of ACM and real-time 3DE for quantifying LV volumes, LVSV, and AR volumes was validated by the chronic animal study and was shown to be clinically applicable.

  12. Real time quantitative amplification detection on a microarray: towards high multiplex quantitative PCR.

    NARCIS (Netherlands)

    Pierik, A.; Moamfa, M; van Zelst, M.; Clout, D.; Stapert, H.; Dijksman, Johan Frederik; Broer, D.; Wimberger-Friedl, R.

    2012-01-01

    Quantitative real-time polymerase chain reaction (qrtPCR) is widely used as a research and diagnostic tool. Notwithstanding its many powerful features, the method is limited in the degree of multiplexing to about 6 due to spectral overlap of the available fluorophores. A new method is presented that

  13. Real time quantitative amplification detection on a microarray : towards high multiplex quantitative PCR

    NARCIS (Netherlands)

    Pierik, Anke; Boamfa, M.; Zelst, van M.; Clout, D.; Stapert, H.R.; Dijksman, J.F.; Broer, D.J.; Wimberger-Friedl, R.

    2012-01-01

    Quantitative real-time polymerase chain reaction (qrtPCR) is widely used as a research and diagnostic tool. Notwithstanding its many powerful features, the method is limited in the degree of multiplexing to about 6 due to spectral overlap of the available fluorophores. A new method is presented that

  14. Enabling Real-Time Analysis at LHCb

    CERN Multimedia

    Govorkova, Katya

    2017-01-01

    A new streaming strategy of the LHCb experiment includes the possibility to perform the physics analysis with candidates reconstructed in the trigger, thus bypassing the offline reconstruction. In the Turbo stream the trigger writes out a compact summary of physics objects containing all information necessary for analyses. This allows an increased output rate and thus higher average efficiencies. The Turbo stream was introduced in 2015 and has allowed for and expanded physics program in Run 2 of the LHC.

  15. A Quantitative Approach to the Formal Verification of Real-Time Systems.

    Science.gov (United States)

    1996-09-01

    Computer Science A Quantitative Approach to the Formal Verification of Real - Time Systems Sergio Vale Aguiar Campos September 1996 CMU-CS-96-199...ptisiic raieaiSI v Diambimos Lboiamtad _^ A Quantitative Approach to the Formal Verification of Real - Time Systems Sergio Vale Aguiar Campos...implied, of NSF, the Semiconduc- tor Research Corporation, ARPA or the U.S. government. Keywords: real - time systems , formal verification, symbolic

  16. Analysis and Synthesis of Distributed Real-Time Embedded Systems

    DEFF Research Database (Denmark)

    Pop, Paul; Eles, Petru; Peng, Zebo

    like automotive electronics, real-time multimedia, avionics, medical equipment, and factory systems. The proposed analysis and synthesis techniques derive optimized implementations that fulfill the imposed design constraints. An important part of the implementation process is the synthesis...

  17. Embryonation of Ostertagia ostertagi eggs affects the outcome of real-time quantitative PCR

    DEFF Research Database (Denmark)

    Drag, Markus; Höglund, Johan; Nejsum, Peter

    prior to detection and quantification by real-time quantitative polymerase chain reaction (qPCR). Fresh O. ostertagi eggs were isolated from cattle faeces and stored at 4°C or 25°C under aerobic or anaerobic conditions. Embryonation was monitored by microscopy and the ITS2 copies were determined by q...... the outcome of qPCR analysis for the quantitative determination of O. ostertagi eggs in cattle faeces. Cold storage at 4°C for up to 3 days or anaerobicvacuum packing at 25°C for up to 336 h will entail no undesirable effects on ITS2 copies....

  18. Embryonation of Ostertagia ostertagi eggs affects the outcome of real-time quantitative PCR

    DEFF Research Database (Denmark)

    Drag, Markus; Höglund, Johan; Nejsum, Peter

    prior to detection and quantification by real-time quantitative polymerase chain reaction (qPCR) . Fresh O. ostertagi eggs were isolated from cattle faeces and stored at 4°C or 25°C under aerobic or anaerobic conditions. Embryonation was monitored by microscopy and the ITS2 copies were determined by q...... the outcome of qPCR analysis for the quantitative determination of O. ostertagi eggs in cattle faeces. Cold storage at 4°C for up to 3 days or anaerobic vacuum packing at 25°C for up to 336 h will entail no undesirable effects on ITS2 copies....

  19. Development of real-time PCR for detection and quantitation of Streptococcus parauberis.

    Science.gov (United States)

    Nguyen, T L; Lim, Y J; Kim, D-H; Austin, B

    2016-01-01

    Streptococcus parauberis is an increasing threat to aquaculture of olive flounder, Paralichthys olivaceus Temminck & Schlegel, in South Korea. We developed a real-time polymerase chain reaction (PCR) method using the TaqMan probe assay to detect and quantify S. parauberis by targeting the gyrB gene sequences, which are effective for molecular analysis of the genus Streptococcus. Our real-time PCR assay is capable of detecting 10 fg of genomic DNA per reaction. The intra- and interassay coefficient of variation (CV) values ranged from 0.42-1.95%, demonstrating that the assay has good reproducibility. There was not any cross-reactivity to Streptococcus iniae or to other streptococcal/lactococcal fish pathogens, such as S. agalactiae and Lactococcus garvieae, indicating that the assay is highly specific to S. parauberis. The results of the real-time PCR assay corresponded well to those of conventional culture assays for S. parauberis from inoculated tissue homogenates (r = 0.957; P < 0.05). Hence, this sensitive and specific real-time PCR is a valuable tool for diagnostic quantitation of S. parauberis in clinical samples. © 2014 John Wiley & Sons Ltd.

  20. Primary quantitative analysis of the mtDNA4977bp deletion induced by lonizing radiation in human peripheral blood u-sing real-time PCR

    International Nuclear Information System (INIS)

    Duan Zhikai; Liu Jiangong; Guo Wanlong; Zhang Shuxian

    2011-01-01

    Objective: To observe the influence of mtDNA4977bp deletion induced by different dose of γ ray in human peripheral blood in order to explore the feasibility of mtDNA4977bp deletion as biodosimeter. Methods: Human peripheral blood samples were collected from three healthy donors and irradiated by γ ray, MtDNA4977bp deletion was detected by real-time PCR. Results: It indicated that that from the range of 0 ∼ 8 Gy, the relationship between mtDNA4977bp deletion and irradiation dose represents certain curvilinear correlation (Y=1.2693+1.0660X+0.0198X 2 ). Conclusion: We find that γ ray has influence on the mtDNA4977bp deletion, so it may be an important biodosmeter in future. (authors)

  1. Reference gene selection for quantitative real-time PCR analysis in virus infected cells: SARS corona virus, Yellow fever virus, Human Herpesvirus-6, Camelpox virus and Cytomegalovirus infections

    Directory of Open Access Journals (Sweden)

    Müller Marcel A

    2005-02-01

    Full Text Available Abstract Ten potential reference genes were compared for their use in experiments investigating cellular mRNA expression of virus infected cells. Human cell lines were infected with Cytomegalovirus, Human Herpesvirus-6, Camelpox virus, SARS coronavirus or Yellow fever virus. The expression levels of these genes and the viral replication were determined by real-time PCR. Genes were ranked by the BestKeeper tool, the GeNorm tool and by criteria we reported previously. Ranking lists of the genes tested were tool dependent. However, over all, β-actin is an unsuitable as reference gene, whereas TATA-Box binding protein and peptidyl-prolyl-isomerase A are stable reference genes for expression studies in virus infected cells.

  2. Quantitative real-time PCR analysis of Anopheles dirus TEP1 and NOS during Plasmodium berghei infection, using three reference genes

    Directory of Open Access Journals (Sweden)

    Jonathan W.K. Liew

    2017-07-01

    Full Text Available Quantitative reverse transcription PCR (qRT-PCR has been an integral part of characterizing the immunity of Anopheles mosquitoes towards Plasmodium invasion. Two anti-Plasmodium factors of Anopheles, thioester-containing protein 1 (TEP1 and nitric oxide synthase (NOS, play a role in the refractoriness of Anopheles towards Plasmodium infection and are generally expressed during infection. However, these are less studied in Anopheles dirus, a dominant malaria vector in Southeast Asia. Furthermore, most studies used a single reference gene for normalization during gene expression analysis without proper validation. This may lead to erroneous quantification of expression levels. Therefore, the present study characterized and investigated the expression profiles of TEP1 and NOS of Anopheles dirus during P. berghei infection. Prior to that, the elongation factor 1-alpha (EF1, actin 1 (Act and ribosomal protein S7 (S7 genes were validated for their suitability as a set of reference genes. TEP1 and NOS expressions in An. dirus were found to be significantly induced after P. berghei infection.

  3. Optimization and analysis of a quantitative real-time PCR-based technique to determine microRNA expression in formalin-fixed paraffin-embedded samples

    Directory of Open Access Journals (Sweden)

    Reis Patricia P

    2010-06-01

    Full Text Available Abstract Background MicroRNAs (miRs are non-coding RNA molecules involved in post-transcriptional regulation, with diverse functions in tissue development, differentiation, cell proliferation and apoptosis. miRs may be less prone to degradation during formalin fixation, facilitating miR expression studies in formalin-fixed paraffin-embedded (FFPE tissue. Results Our study demonstrates that the TaqMan Human MicroRNA Array v1.0 (Early Access platform is suitable for miR expression analysis in FFPE tissue with a high reproducibility (correlation coefficients of 0.95 between duplicates, p 35, we show that reproducibility between technical replicates, equivalent dilutions, and FFPE vs. frozen samples is best in the high abundance stratum. We also demonstrate that the miR expression profiles of FFPE samples are comparable to those of fresh-frozen samples, with a correlation of up to 0.87 (p Conclusion Our study thus demonstrates the utility, reproducibility, and optimization steps needed in miR expression studies using FFPE samples on a high-throughput quantitative PCR-based miR platform, opening up a realm of research possibilities for retrospective studies.

  4. Validation of reference genes for gene expression analysis in olive (Olea europaea) mesocarp tissue by quantitative real-time RT-PCR

    Science.gov (United States)

    2014-01-01

    Background Gene expression analysis using quantitative reverse transcription PCR (qRT-PCR) is a robust method wherein the expression levels of target genes are normalised using internal control genes, known as reference genes, to derive changes in gene expression levels. Although reference genes have recently been suggested for olive tissues, combined/independent analysis on different cultivars has not yet been tested. Therefore, an assessment of reference genes was required to validate the recent findings and select stably expressed genes across different olive cultivars. Results A total of eight candidate reference genes [glyceraldehyde 3-phosphate dehydrogenase (GAPDH), serine/threonine-protein phosphatase catalytic subunit (PP2A), elongation factor 1 alpha (EF1-alpha), polyubiquitin (OUB2), aquaporin tonoplast intrinsic protein (TIP2), tubulin alpha (TUBA), 60S ribosomal protein L18-3 (60S RBP L18-3) and polypyrimidine tract-binding protein homolog 3 (PTB)] were chosen based on their stability in olive tissues as well as in other plants. Expression stability was examined by qRT-PCR across 12 biological samples, representing mesocarp tissues at various developmental stages in three different olive cultivars, Barnea, Frantoio and Picual, independently and together during the 2009 season with two software programs, GeNorm and BestKeeper. Both software packages identified GAPDH, EF1-alpha and PP2A as the three most stable reference genes across the three cultivars and in the cultivar, Barnea. GAPDH, EF1-alpha and 60S RBP L18-3 were found to be most stable reference genes in the cultivar Frantoio while 60S RBP L18-3, OUB2 and PP2A were found to be most stable reference genes in the cultivar Picual. Conclusions The analyses of expression stability of reference genes using qRT-PCR revealed that GAPDH, EF1-alpha, PP2A, 60S RBP L18-3 and OUB2 are suitable reference genes for expression analysis in developing Olea europaea mesocarp tissues, displaying the highest level

  5. GNSS global real-time augmentation positioning: Real-time precise satellite clock estimation, prototype system construction and performance analysis

    Science.gov (United States)

    Chen, Liang; Zhao, Qile; Hu, Zhigang; Jiang, Xinyuan; Geng, Changjiang; Ge, Maorong; Shi, Chuang

    2018-01-01

    Lots of ambiguities in un-differenced (UD) model lead to lower calculation efficiency, which isn't appropriate for the high-frequency real-time GNSS clock estimation, like 1 Hz. Mixed differenced model fusing UD pseudo-range and epoch-differenced (ED) phase observations has been introduced into real-time clock estimation. In this contribution, we extend the mixed differenced model for realizing multi-GNSS real-time clock high-frequency updating and a rigorous comparison and analysis on same conditions are performed to achieve the best real-time clock estimation performance taking the efficiency, accuracy, consistency and reliability into consideration. Based on the multi-GNSS real-time data streams provided by multi-GNSS Experiment (MGEX) and Wuhan University, GPS + BeiDou + Galileo global real-time augmentation positioning prototype system is designed and constructed, including real-time precise orbit determination, real-time precise clock estimation, real-time Precise Point Positioning (RT-PPP) and real-time Standard Point Positioning (RT-SPP). The statistical analysis of the 6 h-predicted real-time orbits shows that the root mean square (RMS) in radial direction is about 1-5 cm for GPS, Beidou MEO and Galileo satellites and about 10 cm for Beidou GEO and IGSO satellites. Using the mixed differenced estimation model, the prototype system can realize high-efficient real-time satellite absolute clock estimation with no constant clock-bias and can be used for high-frequency augmentation message updating (such as 1 Hz). The real-time augmentation message signal-in-space ranging error (SISRE), a comprehensive accuracy of orbit and clock and effecting the users' actual positioning performance, is introduced to evaluate and analyze the performance of GPS + BeiDou + Galileo global real-time augmentation positioning system. The statistical analysis of real-time augmentation message SISRE is about 4-7 cm for GPS, whlile 10 cm for Beidou IGSO/MEO, Galileo and about 30 cm

  6. Quantitative Real-Time PCR Fecal Source Identification in the ...

    Science.gov (United States)

    Rivers in the Tillamook Basin play a vital role in supporting a thriving dairy and cheese-making industry, as well as providing a safe water resource for local human and wildlife populations. Historical concentrations of fecal bacteria in these waters are at times too high to allow for safe use leading to economic loss, endangerment of local wildlife, and poor conditions for recreational use. In this study, we employ host-associated qPCR methods for human (HF183/BacR287 and HumM2), ruminant (Rum2Bac), cattle (CowM2 and CowM3), canine (DG3 and DG37), and avian (GFD) fecal pollution combined with high-resolution geographic information system (GIS) land use data and general indicator bacteria measurements to elucidatewater quality spatial and temporal trends. Water samples (n=584) were collected over a 1-year period at 29 sites along the Trask, Kilchis, and Tillamook rivers and tributaries (Tillamook Basin, OR). A total of 16.6% of samples (n=97) yielded E. coli levels considered impaired based on Oregon Department of Environmental Quality bacteria criteria (406 MPN/100mL). Hostassociated genetic indicators were detected at frequencies of 39.2% (HF183/BacR287), 16.3% (HumM2), 74.6% (Rum2Bac), 13.0% (CowM2), 26.7% (CowM3), 19.8% (DG3), 3.2% (DG37), and 53.4% (GFD) across all water samples (n=584). Seasonal trends in avian, cattle, and human fecal pollution sources were evident over the study area. On a sample site basis, quantitative fecal source identification and

  7. Real Time Engineering Analysis Based on a Generative Component Implementation

    DEFF Research Database (Denmark)

    Kirkegaard, Poul Henning; Klitgaard, Jens

    2007-01-01

    The present paper outlines the idea of a conceptual design tool with real time engineering analysis which can be used in the early conceptual design phase. The tool is based on a parametric approach using Generative Components with embedded structural analysis. Each of these components uses the g...

  8. Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-time quantitative PCR data

    NARCIS (Netherlands)

    van der Velden, V. H. J.; Cazzaniga, G.; Schrauder, A.; Hancock, J.; Bader, P.; Panzer-Grumayer, E. R.; Flohr, T.; Sutton, R.; Cave, H.; Madsen, H. O.; Cayuela, J. M.; Trka, J.; Eckert, C.; Foroni, L.; Zur Stadt, U.; Beldjord, K.; Raff, T.; van der Schoot, C. E.; van Dongen, J. J. M.

    2007-01-01

    Most modern treatment protocols for acute lymphoblastic leukaemia (ALL) include the analysis of minimal residual disease (MRD). To ensure comparable MRD results between different MRD-polymerase chain reaction (PCR) laboratories, standardization and quality control are essential. The European Study

  9. Real-time elastography with a novel quantitative technology for assessment of liver fibrosis in chronic hepatitis B

    International Nuclear Information System (INIS)

    Wang Juan; Guo Long; Shi Xiuying; Pan Wenqian; Bai Yunfei; Ai Hong

    2012-01-01

    Background: The accurate evaluation of liver fibrosis stage is important in determining the treatment strategy. The limitations of percutaneous liver biopsy as the gold standard are obvious for invasion. Real-time elastography with conventional ultrasound probes and a new quantitative technology for diffuse histological lesion is a novel approach for staging of liver fibrosis. Purpose: This study aimed to evaluate the value of real-time tissue elastography with a new quantitative technology for the assessment of liver fibrosis stage. Materials and methods: Real-time elastography was performed in 55 patients with liver fibrosis and chronic hepatitis B and in 20 healthy volunteers. Eleven parameters for every patient in colorcode image obtained from the real-time elastography were analyzed with principal components analysis. We analyzed the correlation between elasticity index and liver fibrosis stage and the accuracy of real-time elastography for liver fibrosis staging. Additionally, aspartate transaminase-to-platelet ratio index was also included in the analysis. Results: The Spearman's correlation coefficient between the elasticity index and the histologic fibrosis stage was 0.81, which is highly significant (p 0.05), respectively. Conclusions: Real-time elastography with a new quantitative technology for diffuse histological lesion is a new and promising sonography-based noninvasive method for the assessment of liver fibrosis in patients with chronic hepatitis B.

  10. Modelling and analysis of real-time and hybrid systems

    Energy Technology Data Exchange (ETDEWEB)

    Olivero, A

    1994-09-29

    This work deals with the modelling and analysis of real-time and hybrid systems. We first present the timed-graphs as model for the real-time systems and we recall the basic notions of the analysis of real-time systems. We describe the temporal properties on the timed-graphs using TCTL formulas. We consider two methods for property verification: in one hand we study the symbolic model-checking (based on backward analysis) and in the other hand we propose a verification method derived of the construction of the simulation graph (based on forward analysis). Both methods have been implemented within the KRONOS verification tool. Their application for the automatic verification on several real-time systems confirms the practical interest of our approach. In a second part we study the hybrid systems, systems combining discrete components with continuous ones. As in the general case the analysis of this king of systems is not decidable, we identify two sub-classes of hybrid systems and we give a construction based method for the generation of a timed-graph from an element into the sub-classes. We prove that in one case the timed-graph obtained is bi-similar with the considered system and that there exists a simulation in the other case. These relationships allow the application of the described technics on the hybrid systems into the defined sub-classes. (authors). 60 refs., 43 figs., 8 tabs., 2 annexes.

  11. Resource-Parameterized Timing Analysis of Real-Time Systems

    DEFF Research Database (Denmark)

    Kim, Jin Hyun; Legay, Axel; Larsen, Kim Guldstrand

    2015-01-01

    on a specic platform. For the same reason, a configuration of platforms cannot be independent from applications in most cases. This paper proposes a new analysis framework of real-time systems where an application and a platform can be analyzed in a fully independent way such that not only the application...

  12. Real-time regression analysis with deep convolutional neural networks

    OpenAIRE

    Huerta, E. A.; George, Daniel; Zhao, Zhizhen; Allen, Gabrielle

    2018-01-01

    We discuss the development of novel deep learning algorithms to enable real-time regression analysis for time series data. We showcase the application of this new method with a timely case study, and then discuss the applicability of this approach to tackle similar challenges across science domains.

  13. Identification and Evaluation of Reliable Reference Genes for Quantitative Real-Time PCR Analysis in Tea Plant (Camellia sinensis (L.) O. Kuntze)

    Science.gov (United States)

    Hao, Xinyuan; Horvath, David P.; Chao, Wun S.; Yang, Yajun; Wang, Xinchao; Xiao, Bin

    2014-01-01

    Reliable reference selection for the accurate quantification of gene expression under various experimental conditions is a crucial step in qRT-PCR normalization. To date, only a few housekeeping genes have been identified and used as reference genes in tea plant. The validity of those reference genes are not clear since their expression stabilities have not been rigorously examined. To identify more appropriate reference genes for qRT-PCR studies on tea plant, we examined the expression stability of 11 candidate reference genes from three different sources: the orthologs of Arabidopsis traditional reference genes and stably expressed genes identified from whole-genome GeneChip studies, together with three housekeeping gene commonly used in tea plant research. We evaluated the transcript levels of these genes in 94 experimental samples. The expression stabilities of these 11 genes were ranked using four different computation programs including geNorm, Normfinder, BestKeeper, and the comparative ∆CT method. Results showed that the three commonly used housekeeping genes of CsTUBULIN1, CsACINT1 and Cs18S rRNA1 together with CsUBQ1 were the most unstable genes in all sample ranking order. However, CsPTB1, CsEF1, CsSAND1, CsCLATHRIN1 and CsUBC1 were the top five appropriate reference genes for qRT-PCR analysis in complex experimental conditions. PMID:25474086

  14. Selection of reference genes for expression analysis using quantitative real-time PCR in the pea aphid, Acyrthosiphon pisum (Harris (Hemiptera, Aphidiae.

    Directory of Open Access Journals (Sweden)

    Chunxiao Yang

    Full Text Available To facilitate gene expression study and obtain accurate qRT-PCR analysis, normalization relative to stable expressed housekeeping genes is required. In this study, expression profiles of 11 candidate reference genes, including actin (Actin, elongation factor 1 α (EF1A, TATA-box-binding protein (TATA, ribosomal protein L12 (RPL12, β-tubulin (Tubulin, NADH dehydrogenase (NADH, vacuolar-type H+-ATPase (v-ATPase, succinate dehydrogenase B (SDHB, 28S ribosomal RNA (28S, 16S ribosomal RNA (16S, and 18S ribosomal RNA (18S from the pea aphid Acyrthosiphon pisum, under different developmental stages and temperature conditions, were investigated. A total of four analytical tools, geNorm, Normfinder, BestKeeper, and the ΔCt method, were used to evaluate the suitability of these genes as endogenous controls. According to RefFinder, a web-based software tool which integrates all four above-mentioned algorithms to compare and rank the reference genes, SDHB, 16S, and NADH were the three most stable house-keeping genes under different developmental stages and temperatures. This work is intended to establish a standardized qRT-PCR protocol in pea aphid and serves as a starting point for the genomics and functional genomics research in this emerging insect model.

  15. Quantitative analysis of waterfowl parvoviruses in geese and Muscovy ducks by real-time polymerase chain reaction: correlation between age, clinical symptoms and DNA copy number of waterfowl parvoviruses

    Directory of Open Access Journals (Sweden)

    Woźniakowski Grzegorz

    2012-03-01

    Full Text Available Abstract Background Waterfowl parvoviruses cause serious loss in geese and ducks production. Goose parvovirus (GPV is infectious for geese and ducks while Muscovy duck parvovirus (MDPV infects Muscovy ducks only. So far, for these viruses' sensitive detection polymerase chain reaction (PCR and loop-mediated isothermal amplification (LAMP were applied. However, there was no molecular biology method for both waterfowl parvoviruses detection and quantification which could unify the laboratory procedures. The level of GPV and MDPV replication and distribution plays a significant role in the parvoviral infection progress and is strictly correlated to clinical symptoms. Meanwhile, experiments conducted previously on GPV distribution in geese, performed as animal trial, did not involve epidemiological data from the disease field cases. The study on the correlation between age, clinical symptoms and viral DNA copy number may be benefitable in understanding the GPV and MDPV infection. Such data may also aid in determination of the stage and severity of the infection with parvoviruses. Therefore the aim of this study was to develop quantitative real-time PCR for parallel detection of GPV and MDPV in geese and Muscovy ducks and to determine the correlation between the age of the infected birds, clinical symptoms and DNA copy number for the estimation of the disease stage or severity. Results In order to develop quantitative real-time PCR the viral material was collected from 13 farms of geese and 3 farms of Muscovy ducks. The designed primers and Taqman probe for real-time PCR were complementary to GPV and MDPV inverted terminal repeats region. The pITR plasmid was constructed, purified and used to prepare dilutions for standard curve preparation and DNA quantification. The applied method detected both GPV and MDPV in all the examined samples extracted from the heart and liver of the infected birds. The conducted correlation tests have shown relationship

  16. Ultrafast Screening and Quantitation of Pesticides in Food and Environmental Matrices by Solid-Phase Microextraction-Transmission Mode (SPME-TM) and Direct Analysis in Real Time (DART).

    Science.gov (United States)

    Gómez-Ríos, Germán Augusto; Gionfriddo, Emanuela; Poole, Justen; Pawliszyn, Janusz

    2017-07-05

    The direct interface of microextraction technologies to mass spectrometry (MS) has unquestionably revolutionized the speed and efficacy at which complex matrices are analyzed. Solid Phase Micro Extraction-Transmission Mode (SPME-TM) is a technology conceived as an effective synergy between sample preparation and ambient ionization. Succinctly, the device consists of a mesh coated with polymeric particles that extracts analytes of interest present in a given sample matrix. This coated mesh acts as a transmission-mode substrate for Direct Analysis in Real Time (DART), allowing for rapid and efficient thermal desorption/ionization of analytes previously concentrated on the coating, and dramatically lowering the limits of detection attained by sole DART analysis. In this study, we present SPME-TM as a novel tool for the ultrafast enrichment of pesticides present in food and environmental matrices and their quantitative determination by MS via DART ionization. Limits of quantitation in the subnanogram per milliliter range can be attained, while total analysis time does not exceed 2 min per sample. In addition to target information obtained via tandem MS, retrospective studies of the same sample via high-resolution mass spectrometry (HRMS) were accomplished by thermally desorbing a different segment of the microextraction device.

  17. Analysis and optimisation of heterogeneous real-time embedded systems

    DEFF Research Database (Denmark)

    Pop, Paul; Eles, Petru; Peng, Zebo

    2005-01-01

    . The success of such new design methods depends on the availability of analysis and optimisation techniques. Analysis and optimisation techniques for heterogeneous real-time embedded systems are presented in the paper. The authors address in more detail a particular class of such systems called multi...... of application messages to frames. Optimisation heuristics for frame packing aimed at producing a schedulable system are presented. Extensive experiments and a real-life example show the efficiency of the frame-packing approach....

  18. Analysis and optimisation of heterogeneous real-time embedded systems

    DEFF Research Database (Denmark)

    Pop, Paul; Eles, Petru; Peng, Zebo

    2006-01-01

    . The success of such new design methods depends on the availability of analysis and optimisation techniques. Analysis and optimisation techniques for heterogeneous real-time embedded systems are presented in the paper. The authors address in more detail a particular class of such systems called multi...... of application messages to frames. Optimisation heuristics for frame packing aimed at producing a schedulable system are presented. Extensive experiments and a real-life example show the efficiency of the frame-packing approach....

  19. Analysis and Optimization of Heterogeneous Real-Time Embedded Systems

    DEFF Research Database (Denmark)

    Pop, Paul; Eles, Petru; Peng, Zebo

    2005-01-01

    . The success of such new design methods depends on the availability of analysis and optimization techniques. In this paper, we present analysis and optimization techniques for heterogeneous real-time embedded systems. We address in more detail a particular class of such systems called multi-clusters, composed...... to frames. Optimization heuristics for frame packing aiming at producing a schedulable system are presented. Extensive experiments and a real-life example show the efficiency of the frame-packing approach....

  20. Evaluation of Rice Resistance to Southern Rice Black-Streaked Dwarf Virus and Rice Ragged Stunt Virus through Combined Field Tests, Quantitative Real-Time PCR, and Proteome Analysis.

    Science.gov (United States)

    Wang, Zhenchao; Yu, Lu; Jin, Linhong; Wang, Wenli; Zhao, Qi; Ran, Longlu; Li, Xiangyang; Chen, Zhuo; Guo, Rong; Wei, Yongtian; Yang, Zhongcheng; Liu, Enlong; Hu, Deyu; Song, Baoan

    2017-02-22

    Diseases caused by southern rice black-streaked dwarf virus (SRBSDV) and rice ragged stunt virus (RRSV) considerably decrease grain yield. Therefore, determining rice cultivars with high resistance to SRBSDV and RRSV is necessary. In this study, rice cultivars with high resistance to SRBSDV and RRSV were evaluated through field trials in Shidian and Mangshi county, Yunnan province, China. SYBR Green I-based quantitative real-time polymerase chain reaction (qRT-PCR) analysis was used to quantitatively detect virus gene expression levels in different rice varieties. The following parameters were applied to evaluate rice resistance: acre yield (A.Y.), incidence of infected plants (I.I.P.), virus load (V.L.), disease index (D.I.), and insect quantity (I.Q.) per 100 clusters. Zhongzheyou1 (Z1) and Liangyou2186 (L2186) were considered the most suitable varieties with integrated higher A.Y., lower I.I.P., V.L., D.I. and I.Q. In order to investigate the mechanism of rice resistance, comparative label-free shotgun liquid chromatography tandem-mass spectrometry (LC-MS/MS) proteomic approaches were applied to comprehensively describe the proteomics of rice varieties' SRBSDV tolerance. Systemic acquired resistance (SAR)-related proteins in Z1 and L2186 may result in the superior resistance of these varieties compared with Fengyouxiangzhan (FYXZ).

  1. Low-level processing for real-time image analysis

    Science.gov (United States)

    Eskenazi, R.; Wilf, J. M.

    1979-01-01

    A system that detects object outlines in television images in real time is described. A high-speed pipeline processor transforms the raw image into an edge map and a microprocessor, which is integrated into the system, clusters the edges, and represents them as chain codes. Image statistics, useful for higher level tasks such as pattern recognition, are computed by the microprocessor. Peak intensity and peak gradient values are extracted within a programmable window and are used for iris and focus control. The algorithms implemented in hardware and the pipeline processor architecture are described. The strategy for partitioning functions in the pipeline was chosen to make the implementation modular. The microprocessor interface allows flexible and adaptive control of the feature extraction process. The software algorithms for clustering edge segments, creating chain codes, and computing image statistics are also discussed. A strategy for real time image analysis that uses this system is given.

  2. Real-time analysis of water movement in plant sample

    International Nuclear Information System (INIS)

    Yokota, Harumi; Furukawa, Jun; Tanoi, Keitaro

    2000-01-01

    To know the effect of drought stress on two cultivars of cowpea, drought tolerant (DT) and drought sensitive (DS), and to estimate vanadium treatment on plant activity, we performed real time 18 F labeled water uptake measurement by PETIS. Fluoride-18 was produced by bombarding a cubic ice target with 50 MeV protons using TIARA AVF cyclotron. Then 18 F labeled water was applied to investigate water movement in a cowpea plant. Real time water uptake manner could be monitored by PETIS. After the analysis by PETIS, we also measured the distribution of 18 F in a whole plant by BAS. When a cowpea plant was treated with drought stress, there was a difference in water uptake manner between DT and DS cultivar. When a cowpea plant was treated with V for 20 hours before the water uptake experiment, the total amount of 18 F labeled water absorption was found to be drastically decreased. (author)

  3. Real-time analysis of water movement in plant sample

    Energy Technology Data Exchange (ETDEWEB)

    Yokota, Harumi; Furukawa, Jun; Tanoi, Keitaro [Graduate School, Tokyo Univ. (Japan)

    2000-07-01

    To know the effect of drought stress on two cultivars of cowpea, drought tolerant (DT) and drought sensitive (DS), and to estimate vanadium treatment on plant activity, we performed real time{sup 18}F labeled water uptake measurement by PETIS. Fluoride-18 was produced by bombarding a cubic ice target with 50 MeV protons using TIARA AVF cyclotron. Then {sup 18}F labeled water was applied to investigate water movement in a cowpea plant. Real time water uptake manner could be monitored by PETIS. After the analysis by PETIS, we also measured the distribution of {sup 18}F in a whole plant by BAS. When a cowpea plant was treated with drought stress, there was a difference in water uptake manner between DT and DS cultivar. When a cowpea plant was treated with V for 20 hours before the water uptake experiment, the total amount of {sup 18}F labeled water absorption was found to be drastically decreased. (author)

  4. Real-time quantitative PCR of microdissected paraffin-embedded breast carcinoma

    DEFF Research Database (Denmark)

    Gjerdrum, Lise Mette; Sorensen, Boe Sandahl; Kjeldsen, Eigil

    2004-01-01

    We studied the feasibility of using real-time quantitative PCR to determine HER-2 DNA amplification and mRNA expression in microdissected formalin-fixed, paraffin-embedded breast tumors and compared this with standard immunohistochemistry (IHC) and fluorescent in situ hybridization (FISH) methods...

  5. Quantitative real-time RT-PCR and chromogenic in situ hybridization

    DEFF Research Database (Denmark)

    Rosa, Fabíola E; Silveira, Sara M; Silveira, Cássia G T

    2009-01-01

    . METHODS: To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR) and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. RESULTS: The concordance...

  6. Periodontal pathogens: a quantitative comparison of anaerobic culture and real-time PCR

    NARCIS (Netherlands)

    Boutaga, Khalil; van Winkelhoff, Arie Jan; Vandenbroucke-Grauls, Christina M. J. E.; Savelkoul, Paul H. M.

    2005-01-01

    Periodontitis is a multi-factorial chronic inflammatory and destructive disease of the tooth-supporting tissues. Quantitative anaerobic culture techniques have been used for microbial diagnosis of the different forms of the disease. The aim of this study was to compare real-time PCR with

  7. Protein Analysis Using Real-Time PCR Instrumentation: Incorporation in an Integrated, Inquiry-Based Project

    Science.gov (United States)

    Southard, Jonathan N.

    2014-01-01

    Instrumentation for real-time PCR is used primarily for amplification and quantitation of nucleic acids. The capability to measure fluorescence while controlling temperature in multiple samples can also be applied to the analysis of proteins. Conformational stability and changes in stability due to ligand binding are easily assessed. Protein…

  8. Quantitative density measurements from a real-time neutron radiography system

    International Nuclear Information System (INIS)

    McRae, D.D.; Jenkins, R.W. Jr.; Brenizer, J.S.; Tobin, K.W.; Hosticka, B.; Sulcoski, M.F.

    1986-01-01

    An advanced video system has been assembled from commercially available equipment to support the real-time neutron radiography facility established jointly by the University of Virginia Department of Nuclear Engineering and Engineering Physics, and the Philip Morris Research Center. A schematic diagram of the equipment used for real-time neutron radiography is presented. To obtain quantitative density measurements with this system, several modifications of both hardware and image processing software were required. After implementation of these changes, the system was capable of determining material densities by measuring the degree of neutron attenuation

  9. ClockWork: a Real-Time Feasibility Analysis Tool

    NARCIS (Netherlands)

    Jansen, P.G.; Hanssen, F.T.Y.; Mullender, Sape J.

    ClockWork shows that we can improve the flexibility and efficiency of real-time kernels. We do this by proposing methods for scheduling based on so-called Real-Time Transactions. ClockWork uses Real-Time Transactions which allow scheduling decisions to be taken by the system. A programmer does not

  10. Phase-shifting Real-time Holographic Microscopy applied in micro-structures surface analysis

    International Nuclear Information System (INIS)

    Brito, I V; Gesualdi, M R R; Muramatsu, M; Ricardo, J

    2011-01-01

    The microscopic real-time analysis of micro structured materials is of great importance in various domains of science and technology. For other hand, the holographic interferometry comprises a group of powerful optical methods for non-destructive testing in surface analysis. The holographic microscopy uses the holographic interferometric techniques to obtain quantitative intensity and phase information of the optical waves by microscopic systems. With the development of CCD cameras, computers (hardware and software), and new materials for holographic recording, these techniques can be used to replace the classical form of registration and became promising tools in surface analysis. In this work, we developed a prototype of Photorefractive and Digital Holographic Microscope for real-time analysis of micro-structured systems based on the phase-shifting real-time holographic interferometry techniques. Using this apparatus, we are made analysis of shapes and surfaces to obtain the phase maps and the 3D profiles of some samples.

  11. Single-Use Poly(etheretherketone) Solid-Phase Microextraction-Transmission Mode Devices for Rapid Screening and Quantitation of Drugs of Abuse in Oral Fluid and Urine via Direct Analysis in Real-Time Tandem Mass Spectrometry.

    Science.gov (United States)

    Vasiljevic, Tijana; Gómez-Ríos, Germán Augusto; Pawliszyn, Janusz

    2018-01-02

    The analysis of oral fluid (OF) and urine samples to detect drug consumption has garnered considerable attention as alternative biomatrices. Efficient implementation of microextraction and ambient ionization technologies for rapid detection of target compounds in such biomatrices creates a need for biocompatible devices which can be implemented for in vivo sampling and easily interfaced with mass spectrometry (MS) analyzers. This study introduces a novel solid-phase microextraction-transmission mode (SPME-TM) device made of poly(etheretherketone) (PEEK) mesh that can rapidly detect prohibited substances in biofluids via direct analysis in real-time tandem MS (DART-MS/MS). PEEK mesh was selected due to its biocompatibility, excellent resistance to various organic solvents, and its ability to withstand relatively high temperatures (≤350 °C). The meshes were coated with hydrophilic-lipophilic-balance particle-poly(acrylonitrile) (HLB-PAN) slurry. The robustness of the coated meshes was tested by performing rapid vortex agitation (≥3200 rpm) in LC/MS-grade solvents and by exposing them to the DART source jet stream at typical operational temperatures (∼250-350 °C). PEEK SPME-TM devices proved to be robust and were therefore used to perform ex vivo analysis of drugs of abuse spiked in urine and OF samples. Excellent results were obtained for all analytes under study; furthermore, the tests yielded satisfactory limits of quantitation (median, ∼0.5 ng mL -1 ), linearity (≥0.99), and accuracy (80-120%) over the evaluated range (0.5-200 ng mL -1 ). This research highlights plastic SPME-TM's potential usefulness as a method for rapidly screening for prohibited substances in on-site/in vivo scenarios, such as roadside or workplace drug testing, antidoping controls, and pain management programs.

  12. A quantitative method to evaluate mesenchymal stem cell lipofection using real-time PCR.

    Science.gov (United States)

    Ribeiro, S C; Mendes, R; Madeira, C; Monteiro, G A; da Silva, C L; Cabral, J M S

    2010-01-01

    Genetic modification of human mesenchymal stem cells (MSC) is a powerful tool to improve the therapeutic utility of these cells and to increase the knowledge on their regulation mechanisms. In this context, strong efforts have been made recently to develop efficient nonviral gene delivery systems. Although several studies addressed this question most of them use the end product of a reporter gene instead of the DNA uptake quantification to test the transfection efficiency. In this study, we established a method based on quantitative real-time PCR (RT-PCR) to determine the intracellular plasmid DNA copy number in human MSC after lipofection. The procedure requires neither specific cell lysis nor DNA purification. The influence of cell number on the RT-PCR sensitivity was evaluated. The method showed good reproducibility, high sensitivity, and a wide linear range of 75-2.5 x 10⁶ plasmid DNA copies per cell. RT-PCR results were then compared with the percentage of transfected cells assessed by flow cytometry analysis, which showed that flow cytometry-based results are not always proportional to plasmid cellular uptake determined by RT-PCR. This work contributed for the establishment of a rapid quantitative assay to determine intracellular plasmid DNA in stem cells, which will be extremely beneficial for the optimization of gene delivery strategies. © 2010 American Institute of Chemical Engineers

  13. Implementation and Analysis of Real-Time Streaming Protocols.

    Science.gov (United States)

    Santos-González, Iván; Rivero-García, Alexandra; Molina-Gil, Jezabel; Caballero-Gil, Pino

    2017-04-12

    Communication media have become the primary way of interaction thanks to the discovery and innovation of many new technologies. One of the most widely used communication systems today is video streaming, which is constantly evolving. Such communications are a good alternative to face-to-face meetings, and are therefore very useful for coping with many problems caused by distance. However, they suffer from different issues such as bandwidth limitation, network congestion, energy efficiency, cost, reliability and connectivity. Hence, the quality of service and the quality of experience are considered the two most important issues for this type of communication. This work presents a complete comparative study of two of the most used protocols of video streaming, Real Time Streaming Protocol (RTSP) and the Web Real-Time Communication (WebRTC). In addition, this paper proposes two new mobile applications that implement those protocols in Android whose objective is to know how they are influenced by the aspects that most affect the streaming quality of service, which are the connection establishment time and the stream reception time. The new video streaming applications are also compared with the most popular video streaming applications for Android, and the experimental results of the analysis show that the developed WebRTC implementation improves the performance of the most popular video streaming applications with respect to the stream packet delay.

  14. Portable real time analysis system for regional cerebral blood flow

    International Nuclear Information System (INIS)

    Tiernan, T.; Entine, G.; Stump, D.A.; Prough, D.S.

    1988-01-01

    A very portable, regional cerebral blood flow (rCBF) analysis instrument system suitable for use in the operating theater during surgery is under development. Cadmium telluride (CdTe) solid state radiation detectors, an 8086 based data acquisition and communications module and a DEC Microvax computer are used so that the instrument is very compact, yet has the computational power to provide real time data analysis in the clinical environment. The instrument is currently being used at Bowman Gray School of Medicine to study rCBF during cardiopulmonary bypass surgery (CPB). Preliminary studies indicate that monitoring rCBF during this surgical procedure may provide insights into the mechanism that causes a significant fraction of these patients to suffer post operative neuropsychological deficit

  15. Real-time analysis of nitrogen translocation in plants

    International Nuclear Information System (INIS)

    Hayashi, Hiroaki

    2000-01-01

    Nitrogen absorbed by roots is transported to the leaves through xylem vessels and then retranslocated to the new leaves, such as root and storage organs through sieve tubes. It is very important to know how this nitrogen movement occurs in the plants and what mechanisms are involved in controlling this movement in order to increase the efficiency of fertilizer. In this experiments, 13 N and 15 N was used to detect the nitrogen circulation in plants, in combination with the technique for positron detection in real time and for collection of sap in sieve tubes and analysis of 15 N in it. By using 13 N, nitrogen movement from root to shoot was analyzed within 10 min after 13 N was applied to the roots. On the other hand, nitrogen retranslocation through sieve tubes was detected by the analysis of 15 N in the phloem sap over 6 hrs. All data suggest the dynamic translocation of nitrogen in rice plants. (author)

  16. Oxidative stress provokes distinct transcriptional responses in the stress-tolerant atr7 and stress-sensitive loh2 Arabidopsis thaliana mutants as revealed by multi-parallel quantitative real-time PCR analysis of ROS marker and antioxidant genes.

    Science.gov (United States)

    Mehterov, Nikolay; Balazadeh, Salma; Hille, Jacques; Toneva, Valentina; Mueller-Roeber, Bernd; Gechev, Tsanko

    2012-10-01

    The Arabidopsis thaliana atr7 mutant is tolerant to oxidative stress induced by paraquat (PQ) or the catalase inhibitor aminotriazole (AT), while its original background loh2 and wild-type plants are sensitive. Both, AT and PQ, which stimulate the intracellular formation of H₂O₂ or superoxide anions, respectively, trigger cell death in loh2 but do not lead to visible damage in atr7. To study gene expression during oxidative stress and ROS-induced programmed cell death, two platforms for multi-parallel quantitative real-time PCR (qRT-PCR) analysis of 217 antioxidant and 180 ROS marker genes were employed. The qRT-PCR analyses revealed AT- and PQ-induced expression of many ROS-responsive genes mainly in loh2, confirming that an oxidative burst plays a role in the activation of the cell death in this mutant. Some of the genes were specifically regulated by either AT or PQ, serving as markers for particular types of ROS. Genes significantly induced by both AT and PQ in loh2 included transcription factors (ANAC042/JUB1, ANAC102, DREB19, HSFA2, RRTF1, ZAT10, ZAT12, ethylene-responsive factors), signaling compounds, ferritins, alternative oxidases, and antioxidant enzymes. Many of these genes were upregulated in atr7 compared to loh2 under non-stress conditions at the first time point, indicating that higher basal levels of ROS and higher antioxidant capacity in atr7 are responsible for the enhanced tolerance to oxidative stress and suggesting a possible tolerance against multiple stresses of this mutant. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  17. Legionellosis and Lung Abscesses: Contribution of Legionella Quantitative Real-Time PCR to an Adapted Followup

    Directory of Open Access Journals (Sweden)

    G. Descours

    2013-01-01

    Full Text Available We report a case of severe Legionnaires' disease (LD complicated by a lung abscess in an immunocompetent patient who required ECMO therapy and thoracic surgery. The results of repeated Legionella quantitative real-time PCR performed on both sera and respiratory samples correlated with the LD severity and the poor clinical outcome. Moreover, the PCR allowed for the detection of Legionella DNA in the lung abscess specimen, which was negative when cultured for Legionella. This case report provides a logical basis for further investigations to examine whether the Legionella quantitative PCR could improve the assessment of LD severity and constitute a prognostic marker.

  18. Identification of circulating miRNA biomarkers based on global quantitative real-time PCR profiling

    Directory of Open Access Journals (Sweden)

    Kang Kang

    2012-02-01

    Full Text Available Abstract MicroRNAs (miRNAs are small noncoding RNAs (18-25 nucleotides that regulate gene expression at the post-transcriptional level. Recent studies have demonstrated the presence of miRNAs in the blood circulation. Deregulation of miRNAs in serum or plasma has been associated with many diseases including cancers and cardiovascular diseases, suggesting the possible use of miRNAs as diagnostic biomarkers. However, the detection of the small amount of miRNAs found in serum or plasma requires a method with high sensitivity and accuracy. Therefore, the current study describes polymerase chain reaction (PCR-based methods for measuring circulating miRNAs. Briefly, the procedure involves four major steps: (1 sample collection and preparation; (2 global miRNAs profiling using quantitative real-time PCR (qRT-PCR; (3 data normalization and analysis; and (4 selection and validation of miRNA biomarkers. In conclusion, qRT-PCR is a promising method for profiling of circulating miRNAs as biomarkers.

  19. Selecting and validating reference genes for quantitative real-time PCR in Plutella xylostella (L.).

    Science.gov (United States)

    You, Yanchun; Xie, Miao; Vasseur, Liette; You, Minsheng

    2018-05-01

    Gene expression analysis provides important clues regarding gene functions, and quantitative real-time PCR (qRT-PCR) is a widely used method in gene expression studies. Reference genes are essential for normalizing and accurately assessing gene expression. In the present study, 16 candidate reference genes (ACTB, CyPA, EF1-α, GAPDH, HSP90, NDPk, RPL13a, RPL18, RPL19, RPL32, RPL4, RPL8, RPS13, RPS4, α-TUB, and β-TUB) from Plutella xylostella were selected to evaluate gene expression stability across different experimental conditions using five statistical algorithms (geNorm, NormFinder, Delta Ct, BestKeeper, and RefFinder). The results suggest that different reference genes or combinations of reference genes are suitable for normalization in gene expression studies of P. xylostella according to the different developmental stages, strains, tissues, and insecticide treatments. Based on the given experimental sets, the most stable reference genes were RPS4 across different developmental stages, RPL8 across different strains and tissues, and EF1-α across different insecticide treatments. A comprehensive and systematic assessment of potential reference genes for gene expression normalization is essential for post-genomic functional research in P. xylostella, a notorious pest with worldwide distribution and a high capacity to adapt and develop resistance to insecticides.

  20. Quantitative real-time monitoring of dryer effluent using fiber optic near-infrared spectroscopy.

    Science.gov (United States)

    Harris, S C; Walker, D S

    2000-09-01

    This paper describes a method for real-time quantitation of the solvents evaporating from a dryer. The vapor stream in the vacuum line of a dryer was monitored in real time using a fiber optic-coupled acousto-optic tunable filter near-infrared (AOTF-NIR) spectrometer. A balance was placed in the dryer, and mass readings were recorded for every scan of the AOTF-NIR. A partial least-squares (PLS) calibration was subsequently built based on change in mass over change in time for solvents typically used in a chemical manufacturing plant. Controlling software for the AOTF-NIR was developed. The software collects spectra, builds the PLS calibration model, and continuously fits subsequently collected spectra to the calibration, allowing the operator to follow the mass loss of solvent from the dryer. The results indicate that solvent loss can be monitored and quantitated in real time using NIR for the optimization of drying times. These time-based mass loss values have also been used to calculate "dynamic" vapor density values for the solvents. The values calculated are in agreement with values determined from the ideal gas law and could prove valuable as tools to measure temperature or pressure indirectly.

  1. Development of a real time polymerase chain reaction for quantitation of Schistosoma mansoni DNA

    Directory of Open Access Journals (Sweden)

    Ana Lisa do Vale Gomes

    2006-10-01

    Full Text Available This report describes the development of a SYBR Green I based real time polymerase chain reaction (PCR protocol for detection on the ABI Prism 7000 instrument. Primers targeting the gene encoding the SSU rRNA were designed to amplify with high specificity DNA from Schistosoma mansoni, in a real time quantitative PCR system. The limit of detection of parasite DNA for the system was 10 fg of purified genomic DNA, that means less than the equivalent to one parasite cell (genome ~580 fg DNA. The efficiency was 0.99 and the correlation coefficient (R² was 0.97. When different copy numbers of the target amplicon were used as standards, the assay could detect at least 10 copies of the specific target. The primers used were designed to amplify a 106 bp DNA fragment (Tm 83ºC. The assay was highly specific for S. mansoni, and did not recognize DNA from closely related non-schistosome trematodes. The real time PCR allowed for accurate quantification of S. mansoni DNA and no time-consuming post-PCR detection of amplification products by gel electrophoresis was required. The assay is potentially able to quantify S. mansoni DNA (and indirectly parasite burden in a number of samples, such as snail tissue, serum and feces from patients, and cercaria infested water. Thus, these PCR protocols have potential to be used as tools for monitoring of schistosome transmission and quantitative diagnosis of human infection.

  2. Real-time quantitative phase reconstruction in off-axis digital holography using multiplexing.

    Science.gov (United States)

    Girshovitz, Pinhas; Shaked, Natan T

    2014-04-15

    We present a new approach for obtaining significant speedup in the digital processing of extracting unwrapped phase profiles from off-axis digital holograms. The new technique digitally multiplexes two orthogonal off-axis holograms, where the digital reconstruction, including spatial filtering and two-dimensional phase unwrapping on a decreased number of pixels, can be performed on both holograms together, without redundant operations. Using this technique, we were able to reconstruct, for the first time to our knowledge, unwrapped phase profiles from off-axis holograms with 1 megapixel in more than 30 frames per second using a standard single-core personal computer on a MATLAB platform, without using graphic-processing-unit programming or parallel computing. This new technique is important for real-time quantitative visualization and measurements of highly dynamic samples and is applicable for a wide range of applications, including rapid biological cell imaging and real-time nondestructive testing. After comparing the speedups obtained by the new technique for holograms of various sizes, we present experimental results of real-time quantitative phase visualization of cells flowing rapidly through a microchannel.

  3. Petroleomics by Direct Analysis in Real Time-Mass Spectrometry.

    Science.gov (United States)

    Romão, Wanderson; Tose, Lilian V; Vaz, Boniek G; Sama, Sara G; Lobinski, Ryszard; Giusti, Pierre; Carrier, Hervé; Bouyssiere, Brice

    2016-01-01

    The analysis of crude oil and its fractions by applying ambient ionization techniques remains underexplored in mass spectrometry (MS). Direct analysis in real time (DART) in the positive-ion mode was coupled to a linear quadrupole ion trap Orbitrap mass spectrometer (LTQ Orbitrap) to analyze crude oil, paraffin samples, and porphyrin standard compounds. The ionization parameters of DART-MS were optimized for crude oil analysis. DART-MS rendered the optimum conditions of the operation using paper as the substrate, T = 400°C, helium as the carrier gas, and a sample concentration ≥6 mg mL(-1). In the crude oils analysis, the DART(+)-Orbitrap mass spectra detected the typical N, NO, and O-containing compounds. In the paraffin samples, oxidized hydrocarbon species (Ox classes, where x = 1-4) with double-bond equivalent of 1-4 were detected, and their structures and connectivity were confirmed by collision-induced dissociation (CID) experiments. DART(+)-MS has identified the porphyrin standard compounds as [M + H](+) ions of m/z 615.2502 and 680.1763, where M = C44H30N4 and C44H28N4OV, respectively, based on the formula assignment and by phenyl losses observed on CID experiments.

  4. A method for accurate detection of genomic microdeletions using real-time quantitative PCR

    Directory of Open Access Journals (Sweden)

    Bassett Anne S

    2005-12-01

    Full Text Available Abstract Background Quantitative Polymerase Chain Reaction (qPCR is a well-established method for quantifying levels of gene expression, but has not been routinely applied to the detection of constitutional copy number alterations of human genomic DNA. Microdeletions or microduplications of the human genome are associated with a variety of genetic disorders. Although, clinical laboratories routinely use fluorescence in situ hybridization (FISH to identify such cryptic genomic alterations, there remains a significant number of individuals in which constitutional genomic imbalance is suspected, based on clinical parameters, but cannot be readily detected using current cytogenetic techniques. Results In this study, a novel application for real-time qPCR is presented that can be used to reproducibly detect chromosomal microdeletions and microduplications. This approach was applied to DNA from a series of patient samples and controls to validate genomic copy number alteration at cytoband 22q11. The study group comprised 12 patients with clinical symptoms of chromosome 22q11 deletion syndrome (22q11DS, 1 patient trisomic for 22q11 and 4 normal controls. 6 of the patients (group 1 had known hemizygous deletions, as detected by standard diagnostic FISH, whilst the remaining 6 patients (group 2 were classified as 22q11DS negative using the clinical FISH assay. Screening of the patients and controls with a set of 10 real time qPCR primers, spanning the 22q11.2-deleted region and flanking sequence, confirmed the FISH assay results for all patients with 100% concordance. Moreover, this qPCR enabled a refinement of the region of deletion at 22q11. Analysis of DNA from chromosome 22 trisomic sample demonstrated genomic duplication within 22q11. Conclusion In this paper we present a qPCR approach for the detection of chromosomal microdeletions and microduplications. The strategic use of in silico modelling for qPCR primer design to avoid regions of repetitive

  5. Detection of medically important Candida species by absolute quantitation real-time polymerase chain reaction.

    Science.gov (United States)

    Than, Leslie Thian Lung; Chong, Pei Pei; Ng, Kee Peng; Seow, Heng Fong

    2015-01-01

    The number of invasive candidiasis cases has risen especially with an increase in the number of immunosuppressed and immunocom promised patients. The early detection of Candida species which is specific and sensitive is important in determining the correct administration of antifungal drugs to patients. This study aims to develop a method for the detection, identification and quantitation of medically important Candida species through quantitative polymerase chain reaction (qPCR). The isocitrate lyase (ICL) gene which is not found in mammals was chosen as the target gene of real-time PCR. Absolute quantitation of the gene copy number was achieved by constructing the plasmid containing the ICL gene which is used to generate standard curve. Twenty fungal species, two bacterial species and human DNA were tested to check the specificity of the detection method. All eight Candida species were successfully detected, identified and quantitated based on the ICL gene. A seven-log range of the gene copy number and a minimum detection limit of 10(3) copies were achieved. A one-tube absolute quantification real-time PCR that differentiates medically important Candida species via individual unique melting temperature was achieved. Analytical sensitivity and specificity were not compromised.

  6. Real time physics analysis with the ATLAS tau trigger system

    International Nuclear Information System (INIS)

    Casado Lechuga, M. P.

    2009-01-01

    The scope of the ATLAS tau trigger system at the LHC is most ambitious. It aims at reconstructing in real time, a matter of seconds, a detailed picture of the high energy proton proton collisions at the LHC. Such system is mandatory in order to select efficiently data needed for discovery of new physics in a proton proton collision environment where the rates of jets observed in the detector are high and the tau identification is difficult. New physics scenarios targeted specifically by the the ATLAS tau trigger system are Standard Model or Supersymmetric Higgs production, and production of new exotic resonances. This contribution will detail how the analysis techniques developed offline for efficient data analysis have been implemented in the algorithms which run online at the trigger. In particular, the focus will be on how to satisfy the requirements imposed by the physics goals while addressing the limitations from the overall event rate and latency allowed. The prospects for early running during the first LHC collisions and trigger evolution from first collisions to stable running will be also summarized, following change of trigger goals from commissioning of detector to measurement of Standard Model physics and discoveries. (author)

  7. Cluster Computing For Real Time Seismic Array Analysis.

    Science.gov (United States)

    Martini, M.; Giudicepietro, F.

    A seismic array is an instrument composed by a dense distribution of seismic sen- sors that allow to measure the directional properties of the wavefield (slowness or wavenumber vector) radiated by a seismic source. Over the last years arrays have been widely used in different fields of seismological researches. In particular they are applied in the investigation of seismic sources on volcanoes where they can be suc- cessfully used for studying the volcanic microtremor and long period events which are critical for getting information on the volcanic systems evolution. For this reason arrays could be usefully employed for the volcanoes monitoring, however the huge amount of data produced by this type of instruments and the processing techniques which are quite time consuming limited their potentiality for this application. In order to favor a direct application of arrays techniques to continuous volcano monitoring we designed and built a small PC cluster able to near real time computing the kinematics properties of the wavefield (slowness or wavenumber vector) produced by local seis- mic source. The cluster is composed of 8 Intel Pentium-III bi-processors PC working at 550 MHz, and has 4 Gigabytes of RAM memory. It runs under Linux operating system. The developed analysis software package is based on the Multiple SIgnal Classification (MUSIC) algorithm and is written in Fortran. The message-passing part is based upon the LAM programming environment package, an open-source imple- mentation of the Message Passing Interface (MPI). The developed software system includes modules devote to receiving date by internet and graphical applications for the continuous displaying of the processing results. The system has been tested with a data set collected during a seismic experiment conducted on Etna in 1999 when two dense seismic arrays have been deployed on the northeast and the southeast flanks of this volcano. A real time continuous acquisition system has been simulated by

  8. Noncontact optical motion sensing for real-time analysis

    Science.gov (United States)

    Fetzer, Bradley R.; Imai, Hiromichi

    1990-08-01

    The adaptation of an image dissector tube (IDT) within the OPTFOLLOW system provides high resolution displacement measurement of a light discontinuity. Due to the high speed response of the IDT and the advanced servo loop circuitry, the system is capable of real time analysis of the object under test. The image of the discontinuity may be contoured by direct or reflected light and ranges spectrally within the field of visible light. The image is monitored to 500 kHz through a lens configuration which transposes the optical image upon the photocathode of the IDT. The photoelectric effect accelerates the resultant electrons through a photomultiplier and an enhanced current is emitted from the anode. A servo loop controls the electron beam, continually centering it within the IDT using magnetic focusing of deflection coils. The output analog voltage from the servo amplifier is thereby proportional to the displacement of the target. The system is controlled by a microprocessor with a 32kbyte memory and provides a digital display as well as instructional readout on a color monitor allowing for offset image tracking and automatic system calibration.

  9. Resistive MHD Stability Analysis in Near Real-time

    Science.gov (United States)

    Glasser, Alexander; Kolemen, Egemen

    2017-10-01

    We discuss the feasibility of a near real-time calculation of the tokamak Δ' matrix, which summarizes MHD stability to resistive modes, such as tearing and interchange modes. As the operational phase of ITER approaches, solutions for active feedback tokamak stability control are needed. It has been previously demonstrated that an ideal MHD stability analysis is achievable on a sub- O (1 s) timescale, as is required to control phenomena comparable with the MHD-evolution timescale of ITER. In the present work, we broaden this result to incorporate the effects of resistive MHD modes. Such modes satisfy ideal MHD equations in regions outside narrow resistive layers that form at singular surfaces. We demonstrate that the use of asymptotic expansions at the singular surfaces, as well as the application of state transition matrices, enable a fast, parallelized solution to the singular outer layer boundary value problem, and thereby rapidly compute Δ'. Sponsored by US DOE under DE-SC0015878 and DE-FC02-04ER54698.

  10. Rapid quantitative detection of Lactobacillus sakei in meat and fermented sausages by real-time PCR.

    Science.gov (United States)

    Martín, Belén; Jofré, Anna; Garriga, Margarita; Pla, Maria; Aymerich, Teresa

    2006-09-01

    A quick and simple method for quantitative detection of Lactobacillus sakei in fermented sausages was successfully developed. It is based on Chelex-100-based DNA purification and real-time PCR enumeration using a TaqMan fluorescence probe. Primers and probes were designed in the L. sakei 16S-23S rRNA intergenic transcribed spacer region, and the assay was evaluated using L. sakei genomic DNA and an artificially inoculated sausage model. The detection limit of this technique was approximately 3 cells per reaction mixture using both purified DNA and the inoculated sausage model. The quantification limit was established at 30 cells per reaction mixture in both models. The assay was then applied to enumerate L. sakei in real samples, and the results were compared to the MRS agar count method followed by confirmation of the percentage of L. sakei colonies. The results obtained by real-time PCR were not statistically significantly different than those obtained by plate count on MRS agar (P > 0.05), showing a satisfactory agreement between both methods. Therefore, the real-time PCR assay developed can be considered a promising rapid alternative method for the quantification of L. sakei and evaluation of the implantation of starter strains of L. sakei in fermented sausages.

  11. Exploring valid reference genes for quantitative real - time rt - pce studies of hydrogenperoxide signaling in arabidopsis

    International Nuclear Information System (INIS)

    Zhou, H.; Han, B.; Xie, Y.; Zhang, J.; Shen, W.

    2015-01-01

    Hydrogen peroxide (H/sub 2/O/sub 2/ ) acts as a signaling molecule modulating the expression of various genes in plants. However, the reference gene(s) used for gene expression analysis of H/sub 2/O/sub 2/ signaling is still arbitrary. A reliable result obtained by quantitative real-time RT-PCR (RT-qPCR) highly depends on accurate transcript normalization using stably expressed reference genes, whereas the inaccurate normalization could easily lead to the false conclusions. In this report, by using geNorm and NormFinder algorithms, 12 candidate reference genes were evaluated and compared in root and shoot tissues of Arabidopsis upon different doses of H/sub 2/O/sub 2/. The results revealed that, in our experimental conditions, three novel reference genes (TIP41-like, UKN, and UBC21) were identified and validated as suitable reference genes for RT-qPCR normalization in both root and shoot tissues under oxidative stress. This conclusion was further confirmed by publicly available microarray data of methyl viologen and drought stress. In comparison with a single reference gene (EF-1a), the expression pattern of ZAT12 modulated by H/sub 2/O/sub 2/, when using TIP41-like, UKN, and UBC21 as multiple reference gene(s), was similar with the previous reports by using northern blotting. Thus, we proposed that these three reference genes might be good candidates for other researchers to include in their reference gene validation in gene expression studies under H/sub 2/O/sub 2/ related oxidative stress. (author)

  12. Comparison of Abbott and Da-an real-time PCR for quantitating serum HBV DNA.

    Science.gov (United States)

    Qiu, Ning; Li, Rui; Yu, Jian-Guo; Yang, Wen; Zhang, Wei; An, Yong; Li, Tong; Liu, Xue-En; Zhuang, Hui

    2014-09-07

    To compare the performance of the Da-an real-time hepatitis B virus (HBV) DNA assay and Abbott RealTime HBV assay. HBV DNA standards as well as a total of 180 clinical serum samples from patients with chronic hepatitis B were measured using the Abbott and Da-an real-time polymerase chain reaction (PCR) assays. Correlation and Bland-Altman plot analysis was used to compare the performance of the Abbott and Da-an assays. The HBV DNA levels were logarithmically transformed for analysis. All statistical analyses were performed using SPSS for Windows version 18.0. The correlation between the two assays was analyzed by Pearson's correlation and linear regression. The Bland-Altman plots were used for the analysis of agreement between the two assays. A P value of Da-an assay were significantly correlated with the expected values of HBV DNA standards (r = 0.999, for Abbott; r = 0.987, for Da-an, P Da-an assay. Moreover, HBV DNA levels measured by the Abbott assay were significantly higher than those of the Da-an assay (6.23 ± 1.76 log IU/mL vs 5.46 ± 1.55 log IU/mL, P Da-an assay presented lower sensitivity and a narrower linear range as compared to the Abbott assay, suggesting the need to be improved.

  13. Modelling and analysis of real-time coordination patterns

    NARCIS (Netherlands)

    Kemper, Stephanie

    2011-01-01

    Present-day embedded software systems need to support an increasing number of features and formalisms; the two most important ones being handling of real-time, and the possibility to develop the system in a modular, component-based way. To ensure that the behaviour of the final system is correct

  14. ANALYSIS OF REAL-TIME VEHICLE HYDROCARBON EMISSIONS DATA

    Science.gov (United States)

    The report gives results of analyses using real-time dynamometer test emissions data from 13 passenger cars to examine variations in emissions during different speeds or modes of travel. The resulting data provided a way to separately identify idle, cruise, acceleration, and dece...

  15. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos

    Directory of Open Access Journals (Sweden)

    Van Zeveren Alex

    2005-12-01

    Full Text Available Abstract Background Real-time quantitative PCR is a sensitive and very efficient technique to examine gene transcription patterns in preimplantation embryos, in order to gain information about embryo development and to optimize assisted reproductive technologies. Critical to the succesful application of real-time PCR is careful assay design, reaction optimization and validation to maximize sensitivity and accuracy. In most of the studies published GAPD, ACTB or 18S rRNA have been used as a single reference gene without prior verification of their expression stability. Normalization of the data using unstable controls can result in erroneous conclusions, especially when only one reference gene is used. Results In this study the transcription levels of 8 commonly used reference genes (ACTB, GAPD, Histone H2A, TBP, HPRT1, SDHA, YWHAZ and 18S rRNA were determined at different preimplantation stages (2-cell, 8-cell, blastocyst and hatched blastocyst in order to select the most stable genes to normalize quantitative data within different preimplantation embryo stages. Conclusion Using the geNorm application YWHAZ, GAPD and SDHA were found to be the most stable genes across the examined embryonic stages, while the commonly used ACTB was shown to be highly regulated. We recommend the use of the geometric mean of those 3 reference genes as an accurate normalization factor, which allows small expression differences to be reliably measured.

  16. Molecular quantification of lactic acid bacteria in fermented milk products using real-time quantitative PCR.

    Science.gov (United States)

    Furet, Jean-Pierre; Quénée, Pascal; Tailliez, Patrick

    2004-12-15

    Real-time quantitative PCR assays were developed for the absolute quantification of lactic acid bacteria (LAB) (Streptococcus thermophilus, Lactobacillus delbrueckii, L. casei, L. paracasei, L. rhamnosus, L. acidophilus and L. johnsonii) in fermented milk products. The results of molecular quantification and classic bacterial enumeration did not differ significantly with respect to S. thermophilus and the species of the L. casei group which were detected in the six commercial fermented products tested, thus showing that DNA extraction was efficient and that genomic DNA solutions were free of PCR inhibitors. For L. delbrueckii, the results of bacterial enumeration were generally lower by a factor 10 to 100 than those of PCR quantification, suggesting a loss of viability during storage of the dairy products at 1-8 degrees C for most of the strains in this species. Real-time quantitative assays enabled identification of the species of lactic acid bacterial strains initially present in commercial fermented milk products and their accurate quantification with a detection threshold of 10(3) cells per ml of product.

  17. Quantification bias caused by plasmid DNA conformation in quantitative real-time PCR assay.

    Science.gov (United States)

    Lin, Chih-Hui; Chen, Yu-Chieh; Pan, Tzu-Ming

    2011-01-01

    Quantitative real-time PCR (qPCR) is the gold standard for the quantification of specific nucleic acid sequences. However, a serious concern has been revealed in a recent report: supercoiled plasmid standards cause significant over-estimation in qPCR quantification. In this study, we investigated the effect of plasmid DNA conformation on the quantification of DNA and the efficiency of qPCR. Our results suggest that plasmid DNA conformation has significant impact on the accuracy of absolute quantification by qPCR. DNA standard curves shifted significantly among plasmid standards with different DNA conformations. Moreover, the choice of DNA measurement method and plasmid DNA conformation may also contribute to the measurement error of DNA standard curves. Due to the multiple effects of plasmid DNA conformation on the accuracy of qPCR, efforts should be made to assure the highest consistency of plasmid standards for qPCR. Thus, we suggest that the conformation, preparation, quantification, purification, handling, and storage of standard plasmid DNA should be described and defined in the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) to assure the reproducibility and accuracy of qPCR absolute quantification.

  18. Complementary techniques: validation of gene expression data by quantitative real time PCR.

    Science.gov (United States)

    Provenzano, Maurizio; Mocellin, Simone

    2007-01-01

    Microarray technology can be considered the most powerful tool for screening gene expression profiles of biological samples. After data mining, results need to be validated with highly reliable biotechniques allowing for precise quantitation of transcriptional abundance of identified genes. Quantitative real time PCR (qrt-PCR) technology has recently reached a level of sensitivity, accuracy and practical ease that support its use as a routine bioinstrumentation for gene level measurement. Currently, qrt-PCR is considered by most experts the most appropriate method to confirm or confute microarray-generated data. The knowledge of the biochemical principles underlying qrt-PCR as well as some related technical issues must be beard in mind when using this biotechnology.

  19. Real time quantitative phase microscopy based on single-shot transport of intensity equation (ssTIE) method

    Science.gov (United States)

    Yu, Wei; Tian, Xiaolin; He, Xiaoliang; Song, Xiaojun; Xue, Liang; Liu, Cheng; Wang, Shouyu

    2016-08-01

    Microscopy based on transport of intensity equation provides quantitative phase distributions which opens another perspective for cellular observations. However, it requires multi-focal image capturing while mechanical and electrical scanning limits its real time capacity in sample detections. Here, in order to break through this restriction, real time quantitative phase microscopy based on single-shot transport of the intensity equation method is proposed. A programmed phase mask is designed to realize simultaneous multi-focal image recording without any scanning; thus, phase distributions can be quantitatively retrieved in real time. It is believed the proposed method can be potentially applied in various biological and medical applications, especially for live cell imaging.

  20. Development of quantitative real-time PCR for detection and enumeration of Enterobacteriaceae.

    Science.gov (United States)

    Takahashi, Hajime; Saito, Rumi; Miya, Satoko; Tanaka, Yuichiro; Miyamura, Natsumi; Kuda, Takashi; Kimura, Bon

    2017-04-04

    The family Enterobacteriaceae, members of which are widely distributed in the environment, includes many important human pathogens. In this study, a rapid real-time PCR method targeting rplP, coding for L16 protein, a component of the ribosome large subunit, was developed for enumerating Enterobacteriaceae strains, and its efficiency was evaluated using naturally contaminated food products. The rplP-targeted real-time PCR amplified Enterobacteriaceae species with Ct values of 14.0-22.8, whereas the Ct values for non-Enterobacteriaceae species were >30, indicating the specificity of this method for the Enterobacteriaceae. Using a calibration curve of Ct=-3.025 (log CFU/g)+37.35, which was calculated from individual plots of the cell numbers in different concentrations of 5 Enterobacteriaceae species, the rplP-targeted real-time PCR was applied to 51 food samples. A Enterobacteriaceae species in foods rapidly and accurately, and therefore, it can be used for the microbiological risk analysis of foods. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Quantitative Real-time PCR detection of putrescine-producing Gram-negative bacteria

    Directory of Open Access Journals (Sweden)

    Kristýna Maršálková

    2017-01-01

    Full Text Available Biogenic amines are indispensable components of living cells; nevertheless these compounds could be toxic for human health in higher concentrations. Putrescine is supposed to be the major biogenic amine associated with microbial food spoilage. Development of reliable, fast and culture-independent molecular methods to detect bacteria producing biogenic amines deserves the attention, especially of the food industry in purpose to protect health. The objective of this study was to verify the newly designed primer sets for detection of two inducible genes adiA and speF together in Salmonella enterica and Escherichia coli genome by Real-time PCR. These forenamed genes encode enzymes in the metabolic pathway which leads to production of putrescine in Gram-negative bacteria. Moreover, relative expression of these genes was studied in E. coli CCM 3954 strain using Real-time PCR. In this study, sets of new primers for the detection two inducible genes (speF and adiA in Salmonella enterica and E. coli by Real-time PCR were designed and tested. Amplification efficiency of a Real-time PCR was calculated from the slope of the standard curves (adiA, speF, gapA. An efficiency in a range from 95 to 105 % for all tested reactions was achieved. The gene expression (R of adiA and speF genes in E. coli was varied depending on culture conditions. The highest gene expression of adiA and speF was observed at 6, 24 and 36 h (RadiA ~ 3, 5, 9; RspeF ~11, 10, 9; respectively after initiation of growth of this bacteria in nutrient broth medium enchired with amino acids. The results show that these primers could be used for relative quantification analysis of E. coli.

  2. Real-time impedance analysis of silica nanowire toxicity on epithelial breast cancer cells.

    Science.gov (United States)

    Alexander, Frank A; Huey, Eric G; Price, Dorielle T; Bhansali, Shekhar

    2012-12-21

    Silica nanowires have great potential for usage in the development of highly sensitive in vivo biosensors used for biomarker monitoring. However, careful analysis of nanowire toxicity is required prior to placing these sensors within the human body. This paper describes a real-time and quantitative analysis of nanowire cytotoxicity using impedance spectroscopy; improving upon studies that have utilized traditional endpoint assays. Silica nanowires were grown using the vapor liquid solid (VLS) method, mixed with Dulbecco's Modified Eagle Medium (DMEM) and exposed to Hs578T epithelial breast cancer cells at concentrations of 0 μg ml(-1), 1 μg ml(-1), 50 μg ml(-1) and 100 μg ml(-1). Real-time cellular responses to silica nanowires confirm that while not cytotoxic, silica nanowires at high concentrations (≥50 μg ml(-1)) are toxic to cells, and also suggest that cell death is due to mechanical disturbances of high numbers of nanowires.

  3. Real-time quantitative PCR for retrovirus-like particle quantification in CHO cell culture.

    Science.gov (United States)

    de Wit, C; Fautz, C; Xu, Y

    2000-09-01

    Chinese hamster ovary (CHO) cells have been widely used to manufacture recombinant proteins intended for human therapeutic uses. Retrovirus-like particles, which are apparently defective and non-infectious, have been detected in all CHO cells by electron microscopy (EM). To assure viral safety of CHO cell-derived biologicals, quantification of retrovirus-like particles in production cell culture and demonstration of sufficient elimination of such retrovirus-like particles by the down-stream purification process are required for product market registration worldwide. EM, with a detection limit of 1x10(6) particles/ml, is the standard retrovirus-like particle quantification method. The whole process, which requires a large amount of sample (3-6 litres), is labour intensive, time consuming, expensive, and subject to significant assay variability. In this paper, a novel real-time quantitative PCR assay (TaqMan assay) has been developed for the quantification of retrovirus-like particles. Each retrovirus particle contains two copies of the viral genomic particle RNA (pRNA) molecule. Therefore, quantification of retrovirus particles can be achieved by quantifying the pRNA copy number, i.e. every two copies of retroviral pRNA is equivalent to one retrovirus-like particle. The TaqMan assay takes advantage of the 5'-->3' exonuclease activity of Taq DNA polymerase and utilizes the PRISM 7700 Sequence Detection System of PE Applied Biosystems (Foster City, CA, U.S.A.) for automated pRNA quantification through a dual-labelled fluorogenic probe. The TaqMan quantification technique is highly comparable to the EM analysis. In addition, it offers significant advantages over the EM analysis, such as a higher sensitivity of less than 600 particles/ml, greater accuracy and reliability, higher sample throughput, more flexibility and lower cost. Therefore, the TaqMan assay should be used as a substitute for EM analysis for retrovirus-like particle quantification in CHO cell

  4. Analysis of real-time reservoir monitoring : reservoirs, strategies, & modeling.

    Energy Technology Data Exchange (ETDEWEB)

    Mani, Seethambal S.; van Bloemen Waanders, Bart Gustaaf; Cooper, Scott Patrick; Jakaboski, Blake Elaine; Normann, Randy Allen; Jennings, Jim (University of Texas at Austin, Austin, TX); Gilbert, Bob (University of Texas at Austin, Austin, TX); Lake, Larry W. (University of Texas at Austin, Austin, TX); Weiss, Chester Joseph; Lorenz, John Clay; Elbring, Gregory Jay; Wheeler, Mary Fanett (University of Texas at Austin, Austin, TX); Thomas, Sunil G. (University of Texas at Austin, Austin, TX); Rightley, Michael J.; Rodriguez, Adolfo (University of Texas at Austin, Austin, TX); Klie, Hector (University of Texas at Austin, Austin, TX); Banchs, Rafael (University of Texas at Austin, Austin, TX); Nunez, Emilio J. (University of Texas at Austin, Austin, TX); Jablonowski, Chris (University of Texas at Austin, Austin, TX)

    2006-11-01

    The project objective was to detail better ways to assess and exploit intelligent oil and gas field information through improved modeling, sensor technology, and process control to increase ultimate recovery of domestic hydrocarbons. To meet this objective we investigated the use of permanent downhole sensors systems (Smart Wells) whose data is fed real-time into computational reservoir models that are integrated with optimized production control systems. The project utilized a three-pronged approach (1) a value of information analysis to address the economic advantages, (2) reservoir simulation modeling and control optimization to prove the capability, and (3) evaluation of new generation sensor packaging to survive the borehole environment for long periods of time. The Value of Information (VOI) decision tree method was developed and used to assess the economic advantage of using the proposed technology; the VOI demonstrated the increased subsurface resolution through additional sensor data. Our findings show that the VOI studies are a practical means of ascertaining the value associated with a technology, in this case application of sensors to production. The procedure acknowledges the uncertainty in predictions but nevertheless assigns monetary value to the predictions. The best aspect of the procedure is that it builds consensus within interdisciplinary teams The reservoir simulation and modeling aspect of the project was developed to show the capability of exploiting sensor information both for reservoir characterization and to optimize control of the production system. Our findings indicate history matching is improved as more information is added to the objective function, clearly indicating that sensor information can help in reducing the uncertainty associated with reservoir characterization. Additional findings and approaches used are described in detail within the report. The next generation sensors aspect of the project evaluated sensors and packaging

  5. Quantitative detection of Campylobacter jejuni on fresh chicken carcasses by real-time PCR.

    Science.gov (United States)

    Rönner, Anna-Clara; Lindmark, Hans

    2007-06-01

    Campylobacter jejuni infection is a significant cause of foodborne gastroenteritis worldwide. Consumption and handling of poultry products is believed to be the primary risk factor for campylobacteriosis. Risk assessments require quantitative data, and C. jejuni is enumerated usually by direct plating, which sometimes allows growth of non-Campylobacter bacteria. The objective of the present study was to develop a quantitative real-time PCR method (q-PCR) for enumerating C. jejuni in chicken rinse without a culturing step. The procedure to obtain the template for the PCR assay involved (i) filtration of 10 ml of chicken rinse, (ii) centrifugation of the sample, and (iii) total DNA extraction from the pellet obtained using a commercial DNA extraction kit. The detection limit of the method was comparable to that for plating 100 microl of chicken rinse on modified charcoal cefoperazone deoxycholate agar, and the detection limit could be further improved 10-fold by concentrating the DNA eluate by ethanol precipitation. A close correlation for spiked chicken rinse was obtained for the results of the quantitative real-time PCR method and direct plating (r = 0.99). The coefficient of correlation for the methods was 0.87 when samples from chicken carcasses on the slaughter line were analyzed, whereas a lower correlation (r = 0.76) was obtained when samples from retail carcasses were analyzed. Greater variation in the proportion of dead and/or viable but not culturable Campylobacter types in the retail samples may explain the decreased correlation between the methods. Overall, the new method is simple and fast and the results obtained are closely correlated with those for direct plating for samples containing a low proportion of dead Campylobacter cells.

  6. Portable Dew Point Mass Spectrometry System for Real-Time Gas and Moisture Analysis

    Science.gov (United States)

    Arkin, C.; Gillespie, Stacey; Ratzel, Christopher

    2010-01-01

    A portable instrument incorporates both mass spectrometry and dew point measurement to provide real-time, quantitative gas measurements of helium, nitrogen, oxygen, argon, and carbon dioxide, along with real-time, quantitative moisture analysis. The Portable Dew Point Mass Spectrometry (PDP-MS) system comprises a single quadrupole mass spectrometer and a high vacuum system consisting of a turbopump and a diaphragm-backing pump. A capacitive membrane dew point sensor was placed upstream of the MS, but still within the pressure-flow control pneumatic region. Pressure-flow control was achieved with an upstream precision metering valve, a capacitance diaphragm gauge, and a downstream mass flow controller. User configurable LabVIEW software was developed to provide real-time concentration data for the MS, dew point monitor, and sample delivery system pressure control, pressure and flow monitoring, and recording. The system has been designed to include in situ, NIST-traceable calibration. Certain sample tubing retains sufficient water that even if the sample is dry, the sample tube will desorb water to an amount resulting in moisture concentration errors up to 500 ppm for as long as 10 minutes. It was determined that Bev-A-Line IV was the best sample line to use. As a result of this issue, it is prudent to add a high-level humidity sensor to PDP-MS so such events can be prevented in the future.

  7. Investigations on abundance and activity of microbial sponge symbionts using quantitative real - time PCR

    DEFF Research Database (Denmark)

    Kumala, Lars; Hentschel, Ute; Bayer, Kristina

    Marine sponges are hosts to dense and diverse microbial consortia that are likely to play a key role in the metabolic processes of the host sponge due to their enormous abundance. Common symbioses between nitrogen transforming microorganisms and sponges indicate complex nitrogen cycling within...... the host. Of particular interest is determining the community structure and function of microbial symbionts in order to gain deeper insight into host-symbiont interactions. We investigated the abundance and activity of microbial symbionts in two Mediterranean sponge species using quantitative real-time PCR....... An absolute quantification of functional genes and transcripts in archaeal and bacterial symbionts was conducted to determine their involvement in nitrification and denitrification, comparing the low microbial abundance (LMA) sponge Dysidea avara with the high microbial abundance (HMA) representative Aplysina...

  8. Comparison of high and low virulence serotypes of Actinobacillus pleuropneumoniae by quantitative real-time PCR

    DEFF Research Database (Denmark)

    Schou, Kirstine Klitgaard; Angen, Øystein; Boye, Mette

    PCR data. Preliminary results showed that in both serotype 2 and serotype 6, the toxin producing gene apxIV was the most highly expressed of the investigated genes. The major difference observed between the two serotypes was that apfA, involved in type IV vili production, was significantly upregulated...... of high virulence while serotype 6 strains are normally found to be less pathogenic. To gain an understanding of the differential virulence of serotype 2 and 6, the expression of a panel of Ap genes during infection of porcine epithelial lung cells (SJPL) were examined by quantitative real-time PCR (q...... to be important for early establishment of the bacteria in the host were examined by qPCR. The genes examined were apfA, coding for a subunit of Type IV pili, kdsB coding for a gene involved in lippopolysacceride biosynthesis, and pgaB which is involved in biofilm formation, all three believed to be important...

  9. Fast real-time polymerase chain reaction for quantitative detection of Lactobacillus delbrueckii bacteriophages in milk.

    Science.gov (United States)

    Martín, Maria Cruz; del Rio, Beatriz; Martínez, Noelia; Magadán, Alfonso H; Alvarez, Miguel A

    2008-12-01

    One of the main microbiological problems of the dairy industry is the susceptibility of starter bacteria to virus infections. Lactobacillus delbrueckii, a component of thermophilic starter cultures used in the manufacture of several fermented dairy products, including yogurt, is also sensitive to bacteriophage attacks. To avoid the problems associated with these viruses, quick and sensitive detection methods are necessary. In the present study, a fast real-time quantitative polymerase chain reaction assay for the direct detection and quantification of L. delbrueckii phages in milk was developed. A set of primers and a TaqMan MGB probe was designed, based on the lysin gene sequence of different L. delbrueckii phages. The results show the proposed method to be a rapid (total processing time 30 min), specific and highly sensitive technique for detecting L. delbrueckii phages in milk.

  10. Real-time and quantitative isotropic spatial resolution susceptibility imaging for magnetic nanoparticles

    Science.gov (United States)

    Pi, Shiqiang; Liu, Wenzhong; Jiang, Tao

    2018-03-01

    The magnetic transparency of biological tissue allows the magnetic nanoparticle (MNP) to be a promising functional sensor and contrast agent. The complex susceptibility of MNPs, strongly influenced by particle concentration, excitation magnetic field and their surrounding microenvironment, provides significant implications for biomedical applications. Therefore, magnetic susceptibility imaging of high spatial resolution will give more detailed information during the process of MNP-aided diagnosis and therapy. In this study, we present a novel spatial magnetic susceptibility extraction method for MNPs under a gradient magnetic field, a low-frequency drive magnetic field, and a weak strength high-frequency magnetic field. Based on this novel method, a magnetic particle susceptibility imaging (MPSI) of millimeter-level spatial resolution (<3 mm) was achieved using our homemade imaging system. Corroborated by the experimental results, the MPSI shows real-time (1 s per frame acquisition) and quantitative abilities, and isotropic high resolution.

  11. Mathematical analysis of the real time array PCR (RTA PCR) process

    NARCIS (Netherlands)

    Dijksman, Johan Frederik; Pierik, A.

    2012-01-01

    Real time array PCR (RTA PCR) is a recently developed biochemical technique that measures amplification curves (like with quantitative real time Polymerase Chain Reaction (qRT PCR)) of a multitude of different templates in a sample. It combines two different methods in order to profit from the

  12. Real-time PCR assays for the quantitation of rDNA from apricot and other plant species in marzipan.

    Science.gov (United States)

    Haase, Ilka; Brüning, Philipp; Matissek, Reinhard; Fischer, Markus

    2013-04-10

    Marzipan or marzipan raw paste is a typical German sweet which is consumed directly or is used as an ingredient in the bakery industry/confectionery (e.g., in stollen) and as filling for chocolate candies. Almonds (blanched and pealed) and sugar are the only ingredients for marzipan production according to German food guidelines. Especially for the confectionery industry, the use of persipan, which contains apricot or peach kernels instead of almonds, is preferred due to its stronger aroma. In most of the companies, both raw pastes are produced, in most cases on the same production line, running the risk of an unintended cross contamination. Additionally, due to high almond market values, dilutions of marzipan with cheaper seeds may occur. Especially in the case of apricot and almond, the close relationship of both species is a challenge for the analysis. DNA based methods for the qualitative detection of apricot, peach, pea, bean, lupine, soy, cashew, pistachio, and chickpea in marzipan have recently been published. In this study, different quantitation strategies on the basis of real-time PCR have been evaluated and a relative quantitation method with a reference amplification product was shown to give the best results. As the real-time PCR is based on the high copy rDNA-cluster, even contaminations <1% can be reliably quantitated.

  13. Tendency for interlaboratory precision in the GMO analysis method based on real-time PCR.

    Science.gov (United States)

    Kodama, Takashi; Kurosawa, Yasunori; Kitta, Kazumi; Naito, Shigehiro

    2010-01-01

    The Horwitz curve estimates interlaboratory precision as a function only of concentration, and is frequently used as a method performance criterion in food analysis with chemical methods. The quantitative biochemical methods based on real-time PCR require an analogous criterion to progressively promote method validation. We analyzed the tendency of precision using a simplex real-time PCR technique in 53 collaborative studies of seven genetically modified (GM) crops. Reproducibility standard deviation (SR) and repeatability standard deviation (Sr) of the genetically modified organism (GMO) amount (%) was more or less independent of GM crops (i.e., maize, soybean, cotton, oilseed rape, potato, sugar beet, and rice) and evaluation procedure steps. Some studies evaluated whole steps consisting of DNA extraction and PCR quantitation, whereas others focused only on the PCR quantitation step by using DNA extraction solutions. Therefore, SR and Sr for GMO amount (%) are functions only of concentration similar to the Horwitz curve. We proposed S(R) = 0.1971C 0.8685 and S(r) = 0.1478C 0.8424, where C is the GMO amount (%). We also proposed a method performance index in GMO quantitative methods that is analogous to the Horwitz Ratio.

  14. On-line real time gamma analysis of primary coolant

    International Nuclear Information System (INIS)

    Kalechstein, W.; Kupca, S.; Lipsett, J.J.

    1985-10-01

    The evolution of failed fuel monitoring at CANDU power stations is briefly summarized and the design of the latest system for failed fuel detection at a multi-unit power station is described. At each reactor, the system employs a germanium spectrometer combined with a novel spectrum analyzer that simultaneously accumulates the gamma-ray spectrum of the coolant and provides the control room with the concentration of radioisotope activity in the coolant for the gaseous fission products Xe-133, Xe-135, Kr-88 and I-131 in real time and with statistical precision independent of count rate. A gross gamma monitor is included to provide independent information on the level of radioactivity in the coolant and extend the measurement range at very high count rates. A central computer system archives spectra received from all four spectrum analyzers and provides both the activity concentrations and the release rates of specified isotopes. Compared with previous systems the current design offers improvements in that the activity concentrations are updated much more frequently, improved tools are provided for long term surveillance of the heat transport system and the monitor is more reliable and less costly

  15. Quantitative Real Time PCR approach to study gene expression profile during prenatal growth of skeletal muscle in pig of Duroc and Pietrain breeds

    Directory of Open Access Journals (Sweden)

    M. Cagnazzo

    2010-01-01

    Full Text Available The quantitative real time-PCR (QRT-PCR is a very sensitive method used to quantify mRNA level in gene expression analysis. Combining amplification, detection and quantification in a single step, allows a more accurate measurement compared to the traditional PCR end point analysis (Pfaffl, 2001; Bustin, 2002.

  16. REAL TIME ANALYSIS OF WIRELESS CONTROLLER AREA NETWORK

    Directory of Open Access Journals (Sweden)

    Gerardine Immaculate Mary

    2014-09-01

    Full Text Available It is widely known that Control Area Networks (CAN are used in real-time, distributed and parallel processing which cover manufacture plants, humanoid robots, networking fields, etc., In applications where wireless conditions are encountered it is convenient to continue the exchange of CAN frames within the Wireless CAN (WCAN. The WCAN considered in this research is based on wireless token ring protocol (WTRP; a MAC protocol for wireless networks to reduce the number of retransmissions due to collision and the wired counterpart CAN attribute on message based communication. WCAN uses token frame method to provide channel access to the nodes in the system. This method allow all the nodes to share common broadcast channel by taken turns in transmitting upon receiving the token frame which is circulating within the network for specified amount of time. This method provides high throughput in bounded latency environment, consistent and predictable delays and good packet delivery ratio. The most important factor to consider when evaluating a control network is the end-to-end time delay between sensors, controllers, and actuators. The correct operation of a control system depends on the timeliness of the data coming over the network, and thus, a control network should be able to guarantee message delivery within a bounded transmission time. The proposed WCAN is modeled and simulated using QualNet, and its average end to end delay and packet delivery ratio (PDR are calculated. The parameters boundaries of WCAN are evaluated to guarantee a maximum throughput and a minimum latency time, in the case of wireless communications, precisely WCAN.

  17. Selection of Suitable Reference Genes for Quantitative Real-time PCR in Sapium sebiferum

    Directory of Open Access Journals (Sweden)

    Xue Chen

    2017-05-01

    Full Text Available Chinese tallow (Sapium sebiferum L. is a promising landscape and bioenergy plant. Measuring gene expression by quantitative real-time polymerase chain reaction (qRT-PCR can provide valuable information on gene function. Stably expressed reference genes for normalization are a prerequisite for ensuring the accuracy of the target gene expression level among different samples. However, the reference genes in Chinese tallow have not been systematically validated. In this study, 12 candidate reference genes (18S, GAPDH, UBQ, RPS15, SAND, TIP41, 60S, ACT7, PDF2, APT, TBP, and TUB were investigated with qRT-PCR in 18 samples, including those from different tissues, from plants treated with sucrose and cold stresses. The data were calculated with four common algorithms, geNorm, BestKeeper, NormFinder, and the delta cycle threshold (ΔCt. TIP41 and GAPDH were the most stable for the tissue-specific experiment, GAPDH and 60S for cold treatment, and GAPDH and UBQ for sucrose stresses, while the least stable genes were 60S, TIP41, and 18S respectively. The comprehensive results showed APT, GAPDH, and UBQ to be the top-ranked stable genes across all the samples. The stability of 60S was the lowest during all experiments. These selected reference genes were further validated by comparing the expression profiles of the chalcone synthase gene in Chinese tallow in different samples. The results will help to improve the accuracy of gene expression studies in Chinese tallow.

  18. Detection of Ophiocordyceps sinensis in soil by quantitative real-time PCR.

    Science.gov (United States)

    Peng, Qingyun; Zhong, Xin; Lei, Wei; Zhang, Guren; Liu, Xin

    2013-03-01

    Ophiocordyceps sinensis, one of the best known entomopathogenic fungi in traditional Chinese medicine, parasitizes larvae of the moth genus Thitarodes, which lives in soil tunnels. However, little is known about the spatial distribution of O. sinensis in the soil. We established a protocol for DNA extraction, purification, and quantification of O. sinensis in soil with quantitative real-time PCR targeting the internal transcribed spacer region. The method was assessed using 34 soil samples from Tibet. No inhibitory effects in purified soil DNA extracts were detected. The standard curve method for absolute DNA quantification generated crossing point values that were strongly and linearly correlated to the log10 of the initial amount of O. sinensis genomic DNA (r(2) = 0.999) over 7 orders of magnitude (4 × 10(1) to 4 × 10(7) fg). The amplification efficiency and y-intercept value of the standard curve were 1.953 and 37.70, respectively. The amount of O. sinensis genomic DNA decreased with increasing soil depth and horizontal distance from a sclerotium (P protocol is rapid, specific, sensitive, and provides a powerful tool for quantification of O. sinensis from soil.

  19. Quantitative Real-Time PCR using the Thermo Scientific Solaris qPCR Assay

    Science.gov (United States)

    Ogrean, Christy; Jackson, Ben; Covino, James

    2010-01-01

    The Solaris qPCR Gene Expression Assay is a novel type of primer/probe set, designed to simplify the qPCR process while maintaining the sensitivity and accuracy of the assay. These primer/probe sets are pre-designed to >98% of the human and mouse genomes and feature significant improvements from previously available technologies. These improvements were made possible by virtue of a novel design algorithm, developed by Thermo Scientific bioinformatics experts. Several convenient features have been incorporated into the Solaris qPCR Assay to streamline the process of performing quantitative real-time PCR. First, the protocol is similar to commonly employed alternatives, so the methods used during qPCR are likely to be familiar. Second, the master mix is blue, which makes setting the qPCR reactions easier to track. Third, the thermal cycling conditions are the same for all assays (genes), making it possible to run many samples at a time and reducing the potential for error. Finally, the probe and primer sequence information are provided, simplifying the publication process. Here, we demonstrate how to obtain the appropriate Solaris reagents using the GENEius product search feature found on the ordering web site (www.thermo.com/solaris) and how to use the Solaris reagents for performing qPCR using the standard curve method. PMID:20567213

  20. Species identification of Cannabis sativa using real-time quantitative PCR (qPCR).

    Science.gov (United States)

    Johnson, Christopher E; Premasuthan, Amritha; Satkoski Trask, Jessica; Kanthaswamy, Sree

    2013-03-01

    Most narcotics-related cases in the United States involve Cannabis sativa. Material is typically identified based on the cystolithic hairs on the leaves and with chemical tests to identify of the presence of cannabinoids. Suspect seeds are germinated into a viable plant so that morphological and chemical tests can be conducted. Seed germination, however, causes undue analytical delays. DNA analyses that involve the chloroplast and nuclear genomes have been developed for identification of C. sativa materials, but they require several nanograms of template DNA. Using the trnL 3' exon-trnF intragenic spacer regions within the C. sativa chloroplast, we have developed a real-time quantitative PCR assay that is capable of identifying picogram amounts of chloroplast DNA for species determination of suspected C. sativa material. This assay provides forensic science laboratories with a quick and reliable method to identify an unknown sample as C. sativa. © 2013 American Academy of Forensic Sciences.

  1. Identification of Reference Genes for Normalizing Quantitative Real-Time PCR in Urechis unicinctus

    Science.gov (United States)

    Bai, Yajiao; Zhou, Di; Wei, Maokai; Xie, Yueyang; Gao, Beibei; Qin, Zhenkui; Zhang, Zhifeng

    2018-06-01

    The reverse transcription quantitative real-time PCR (RT-qPCR) has become one of the most important techniques of studying gene expression. A set of valid reference genes are essential for the accurate normalization of data. In this study, five candidate genes were analyzed with geNorm, NormFinder, BestKeeper and ΔCt methods to identify the genes stably expressed in echiuran Urechis unicinctus, an important commercial marine benthic worm, under abiotic (sulfide stress) and normal (adult tissues, embryos and larvae at different development stages) conditions. The comprehensive results indicated that the expression of TBP was the most stable at sulfide stress and in developmental process, while the expression of EF- 1- α was the most stable at sulfide stress and in various tissues. TBP and EF- 1- α were recommended as a suitable reference gene combination to accurately normalize the expression of target genes at sulfide stress; and EF- 1- α, TBP and TUB were considered as a potential reference gene combination for normalizing the expression of target genes in different tissues. No suitable gene combination was obtained among these five candidate genes for normalizing the expression of target genes for developmental process of U. unicinctus. Our results provided a valuable support for quantifying gene expression using RT-qPCR in U. unicinctus.

  2. Identification of four squid species by quantitative real-time polymerase chain reaction.

    Science.gov (United States)

    Ye, Jian; Feng, Junli; Liu, Shasha; Zhang, Yanping; Jiang, Xiaona; Dai, Zhiyuan

    2016-02-01

    Squids are distributed worldwide, including many species of commercial importance, and they are often made into varieties of flavor foods. The rapid identification methods for squid species especially their processed products, however, have not been well developed. In this study, quantitative real-time PCR (qPCR) systems based on specific primers and TaqMan probes have been established for rapid and accurate identification of four common squid species (Ommastrephes bartramii, Dosidicus gigas, Illex argentinus, Todarodes pacificus) in Chinese domestic market. After analyzing mitochondrial genes reported in GenBank, the mitochondrial cytochrome b (Cytb) gene was selected for O. bartramii detection, cytochrome c oxidase subunit I (COI) gene for D. gigas and T. Pacificus detection, ATPase subunit 6 (ATPase 6) gene for I. Argentinus detection, and 12S ribosomal RNA (12S rDNA) gene for designing Ommastrephidae-specific primers and probe. As a result, all the TaqMan systems are of good performance, and efficiency of each reaction was calculated by making standard curves. This method could detect target species either in single or mixed squid specimen, and it was applied to identify 12 squid processed products successfully. Thus, it would play an important role in fulfilling labeling regulations and squid fishery control. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. A real-time, quantitative PCR protocol for assessing the relative parasitemia of Leucocytozoon in waterfowl

    Science.gov (United States)

    Smith, Matthew M.; Schmutz, Joel A.; Apelgren, Chloe; Ramey, Andy M.

    2015-01-01

    Microscopic examination of blood smears can be effective at diagnosing and quantifying hematozoa infections. However, this method requires highly trained observers, is time consuming, and may be inaccurate for detection of infections at low levels of parasitemia. To develop a molecular methodology for identifying and quantifying Leucocytozoon parasite infection in wild waterfowl (Anseriformes), we designed a real-time, quantitative PCR protocol to amplify Leucocytozoon mitochondrial DNA using TaqMan fluorogenic probes and validated our methodology using blood samples collected from waterfowl in interior Alaska during late summer and autumn (n = 105). By comparing our qPCR results to those derived from a widely used nested PCR protocol, we determined that our assay showed high levels of sensitivity (91%) and specificity (100%) in detecting Leucocytozoon DNA from host blood samples. Additionally, results of a linear regression revealed significant correlation between the raw measure of parasitemia produced by our qPCR assay (Ct values) and numbers of parasites observed on blood smears (R2 = 0.694, P = 0.003), indicating that our assay can reliably determine the relative parasitemia levels among samples. This methodology provides a powerful new tool for studies assessing effects of haemosporidian infection in wild avian species.

  4. GETPrime: a gene- or transcript-specific primer database for quantitative real-time PCR.

    Science.gov (United States)

    Gubelmann, Carine; Gattiker, Alexandre; Massouras, Andreas; Hens, Korneel; David, Fabrice; Decouttere, Frederik; Rougemont, Jacques; Deplancke, Bart

    2011-01-01

    The vast majority of genes in humans and other organisms undergo alternative splicing, yet the biological function of splice variants is still very poorly understood in large part because of the lack of simple tools that can map the expression profiles and patterns of these variants with high sensitivity. High-throughput quantitative real-time polymerase chain reaction (qPCR) is an ideal technique to accurately quantify nucleic acid sequences including splice variants. However, currently available primer design programs do not distinguish between splice variants and also differ substantially in overall quality, functionality or throughput mode. Here, we present GETPrime, a primer database supported by a novel platform that uniquely combines and automates several features critical for optimal qPCR primer design. These include the consideration of all gene splice variants to enable either gene-specific (covering the majority of splice variants) or transcript-specific (covering one splice variant) expression profiling, primer specificity validation, automated best primer pair selection according to strict criteria and graphical visualization of the latter primer pairs within their genomic context. GETPrime primers have been extensively validated experimentally, demonstrating high transcript specificity in complex samples. Thus, the free-access, user-friendly GETPrime database allows fast primer retrieval and visualization for genes or groups of genes of most common model organisms, and is available at http://updepla1srv1.epfl.ch/getprime/. Database URL: http://deplanckelab.epfl.ch.

  5. [Quantitative fluorogenic real-time PCR assay for respiratory syncytial virus detection].

    Science.gov (United States)

    Zhang, Qi-wei; You, Shang-you; Sun, Ji-min; Wu, Qi; Yu, Chun-hua; Zhang, Chu-yu

    2005-07-01

    To Establish a rapid and objective quantitative fluorogenic real-time PCR assay for early detection of human respiratory syncytial virus (hRSV). Two pairs of primers and one TaqMan Fluorogenic probe that are specific for the recognition of the most conservative N gene of hRSV for virus detection with LighCycler PCR in 93 nasopharyngeal secretion specimens collected from infants and young children. The assay was compared with virus isolation, routine PCR, nested PCR, and enzyme-linked immunosorbent assay (ELISA). This TaqMan assay had a sensitivity of 1 x 10(2) cDNA copies/microl with a dynamic range between 1 x 10(2) and 1 x 10(7) cDNA copies/microl, which was the same as that of nested PCR, but 10 times more sensitive than routine PCR. The specificity of the assay was evaluated by comparing hRSV with polivirus type 1, coxsackie virus type 2, influenza A, influenza B and adenovirus type 7. A PCR product of the expected size (195 bp) was produced and fluorescence signal detected for hRSV, but not for any of the other viruses. The results in LightCycler and Rotor-Gene instrument were consistent. Forty-four specimens (43.9%) were hRSV-positive with this assay and 4 (4/93,4.3%) were hRSV-positive with ELISA, showing rather low correlation between the two methods. No visible relation was found between the concentration of hRSV RNA and severity of the disease. This assay is rapid, sensitive, specific and quantitative, and has the potential of wide application for early diagnosis of hRSV infection and evaluation of the therapeutic effect.

  6. Selection of internal control genes for quantitative real-time RT-PCR studies during tomato development process

    Directory of Open Access Journals (Sweden)

    Borges-Pérez Andrés

    2008-12-01

    Full Text Available Abstract Background The elucidation of gene expression patterns leads to a better understanding of biological processes. Real-time quantitative RT-PCR has become the standard method for in-depth studies of gene expression. A biologically meaningful reporting of target mRNA quantities requires accurate and reliable normalization in order to identify real gene-specific variation. The purpose of normalization is to control several variables such as different amounts and quality of starting material, variable enzymatic efficiencies of retrotranscription from RNA to cDNA, or differences between tissues or cells in overall transcriptional activity. The validity of a housekeeping gene as endogenous control relies on the stability of its expression level across the sample panel being analysed. In the present report we describe the first systematic evaluation of potential internal controls during tomato development process to identify which are the most reliable for transcript quantification by real-time RT-PCR. Results In this study, we assess the expression stability of 7 traditional and 4 novel housekeeping genes in a set of 27 samples representing different tissues and organs of tomato plants at different developmental stages. First, we designed, tested and optimized amplification primers for real-time RT-PCR. Then, expression data from each candidate gene were evaluated with three complementary approaches based on different statistical procedures. Our analysis suggests that SGN-U314153 (CAC, SGN-U321250 (TIP41, SGN-U346908 ("Expressed" and SGN-U316474 (SAND genes provide superior transcript normalization in tomato development studies. We recommend different combinations of these exceptionally stable housekeeping genes for suited normalization of different developmental series, including the complete tomato development process. Conclusion This work constitutes the first effort for the selection of optimal endogenous controls for quantitative real-time

  7. Detection of Haemophilus influenzae in respiratory secretions from pneumonia patients by quantitative real-time polymerase chain reaction.

    Science.gov (United States)

    Abdeldaim, Guma M K; Strålin, Kristoffer; Kirsebom, Leif A; Olcén, Per; Blomberg, Jonas; Herrmann, Björn

    2009-08-01

    A quantitative real-time polymerase chain reaction (PCR) based on the omp P6 gene was developed to detect Haemophilus influenzae. Its specificity was determined by analysis of 29 strains of 11 different Haemophilus spp. and was compared with PCR assays having other target genes: rnpB, 16S rRNA, and bexA. The method was evaluated on nasopharyngeal aspirates from 166 adult patients with community-acquired pneumonia. When 10(4) DNA copies/mL was used as cutoff limit for the method, P6 PCR had a sensitivity of 97.5% and a specificity of 96.0% compared with the culture. Of 20 culture-negative but P6 PCR-positive cases, 18 were confirmed by fucK PCR as H. influenzae. Five (5.9%) of 84 nasopharyngeal aspirates from adult controls tested PCR positive. We conclude that the P6 real-time PCR is both sensitive and specific for identification of H. influenzae in respiratory secretions. Quantification facilitates discrimination between disease-causing H. influenzae strains and commensal colonization.

  8. Ultrasensitive quantitation of human papillomavirus type 16 E6 oncogene sequences by nested real time PCR

    Directory of Open Access Journals (Sweden)

    López-Revilla Rubén

    2010-05-01

    Full Text Available Abstract Background We have developed an ultrasensitive method based on conventional PCR preamplification followed by nested amplification through real time PCR (qPCR in the presence of the DNA intercalating agent EvaGreen. Results Amplification mixtures calibrated with a known number of pHV101 copies carrying a 645 base pair (bp-long insert of the human papillomavirus type 16 (HPV16 E6 oncogene were used to generate the E6-1 amplicon of 645 bp by conventional PCR and then the E6-2 amplicon of 237 bp by nested qPCR. Direct and nested qPCR mixtures for E6-2 amplification corresponding to 2.5 × 102-2.5 × 106 initial pHV101 copies had threshold cycle (Ct values in the ranges of 18.7-29.0 and 10.0-25.0, respectively. The Ct of qPCR mixtures prepared with 1/50 volumes of preamplified mixtures containing 50 ng of DNA of the SiHa cell line (derived from an invasive cervical cancer with one HPV16 genome per cell was 19.9. Thermal fluorescence extinction profiles of E6-2 amplicons generated from pHV101 and SiHa DNA were identical, with a peak at 85.5°C. Conclusions Our method based on conventional preamplification for 15 cycles increased 10,750 times the sensitivity of nested qPCR for the quantitation of the E6 viral oncogene and confirmed that the SiHa cell line contains one E6-HPV16 copy per cell.

  9. The quantification of spermatozoa by real-time quantitative PCR, spectrophotometry, and spermatophore cap size.

    Science.gov (United States)

    Doyle, Jacqueline M; McCormick, Cory R; DeWoody, J Andrew

    2011-01-01

    Many animals, such as crustaceans, insects, and salamanders, package their sperm into spermatophores, and the number of spermatozoa contained in a spermatophore is relevant to studies of sexual selection and sperm competition. We used two molecular methods, real-time quantitative polymerase chain reaction (RT-qPCR) and spectrophotometry, to estimate sperm numbers from spermatophores. First, we designed gene-specific primers that produced a single amplicon in four species of ambystomatid salamanders. A standard curve generated from cloned amplicons revealed a strong positive relationship between template DNA quantity and cycle threshold, suggesting that RT-qPCR could be used to quantify sperm in a given sample. We then extracted DNA from multiple Ambystoma maculatum spermatophores, performed RT-qPCR on each sample, and estimated template copy numbers (i.e. sperm number) using the standard curve. Second, we used spectrophotometry to determine the number of sperm per spermatophore by measuring DNA concentration relative to the genome size. We documented a significant positive relationship between the estimates of sperm number based on RT-qPCR and those based on spectrophotometry. When these molecular estimates were compared to spermatophore cap size, which in principle could predict the number of sperm contained in the spermatophore, we also found a significant positive relationship between sperm number and spermatophore cap size. This linear model allows estimates of sperm number strictly from cap size, an approach which could greatly simplify the estimation of sperm number in future studies. These methods may help explain variation in fertilization success where sperm competition is mediated by sperm quantity. © 2010 Blackwell Publishing Ltd.

  10. Quantitative assessment of hematopoietic chimerism by quantitative real-time polymerase chain reaction of sequence polymorphism systems after hematopoietic stem cell transplantation.

    Science.gov (United States)

    Qin, Xiao-ying; Li, Guo-xuan; Qin, Ya-zhen; Wang, Yu; Wang, Feng-rong; Liu, Dai-hong; Xu, Lan-ping; Chen, Huan; Han, Wei; Wang, Jing-zhi; Zhang, Xiao-hui; Li, Jin-lan; Li, Ling-di; Liu, Kai-yan; Huang, Xiao-jun

    2011-08-01

    Analysis of changes in recipient and donor hematopoietic cell origin is extremely useful to monitor the effect of hematopoietic stem cell transplantation (HSCT) and sequential adoptive immunotherapy by donor lymphocyte infusions. We developed a sensitive, reliable and rapid real-time PCR method based on sequence polymorphism systems to quantitatively assess the hematopoietic chimerism after HSCT. A panel of 29 selected sequence polymorphism (SP) markers was screened by real-time PCR in 101 HSCT patients with leukemia and other hematological diseases. The chimerism kinetics of bone marrow samples of 8 HSCT patients in remission and relapse situations were followed longitudinally. Recipient genotype discrimination was possible in 97.0% (98 of 101) with a mean number of 2.5 (1-7) informative markers per recipient/donor pair. Using serial dilutions of plasmids containing specific SP markers, the linear correlation (r) of 0.99, the slope between -3.2 and -3.7 and the sensitivity of 0.1% were proved reproducible. By this method, it was possible to very accurately detect autologous signals in the range from 0.1% to 30%. The accuracy of the method in the very important range of autologous signals below 5% was extraordinarily high (standard deviation real-time PCR method over short tandem repeat PCR chimerism assays is the absence of PCR competition and plateau biases, with demonstrated greater sensitivity and linearity. Finally, we prospectively analyzed bone marrow samples of 8 patients who received allografts and presented the chimerism kinetics of remission and relapse situations that illustrated the sensitivity level and the promising clinical application of this method. This SP-based real-time PCR assay provides a rapid, sensitive, and accurate quantitative assessment of mixed chimerism that can be useful in predicting graft rejection and early relapse.

  11. Intra-laboratory validation of chronic bee paralysis virus quantitation using an accredited standardised real-time quantitative RT-PCR method.

    Science.gov (United States)

    Blanchard, Philippe; Regnault, Julie; Schurr, Frank; Dubois, Eric; Ribière, Magali

    2012-03-01

    Chronic bee paralysis virus (CBPV) is responsible for chronic bee paralysis, an infectious and contagious disease in adult honey bees (Apis mellifera L.). A real-time RT-PCR assay to quantitate the CBPV load is now available. To propose this assay as a reference method, it was characterised further in an intra-laboratory study during which the reliability and the repeatability of results and the performance of the assay were confirmed. The qPCR assay alone and the whole quantitation method (from sample RNA extraction to analysis) were both assessed following the ISO/IEC 17025 standard and the recent XP U47-600 standard issued by the French Standards Institute. The performance of the qPCR assay and of the overall CBPV quantitation method were validated over a 6 log range from 10(2) to 10(8) with a detection limit of 50 and 100 CBPV RNA copies, respectively, and the protocol of the real-time RT-qPCR assay for CBPV quantitation was approved by the French Accreditation Committee. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Image analysis of multiple moving wood pieces in real time

    Science.gov (United States)

    Wang, Weixing

    2006-02-01

    This paper presents algorithms for image processing and image analysis of wood piece materials. The algorithms were designed for auto-detection of wood piece materials on a moving conveyor belt or a truck. When wood objects on moving, the hard task is to trace the contours of the objects in n optimal way. To make the algorithms work efficiently in the plant, a flexible online system was designed and developed, which mainly consists of image acquisition, image processing, object delineation and analysis. A number of newly-developed algorithms can delineate wood objects with high accuracy and high speed, and in the wood piece analysis part, each wood piece can be characterized by a number of visual parameters which can also be used for constructing experimental models directly in the system.

  13. Analysis and Optimization of Distributed Real-Time Embedded Systems

    DEFF Research Database (Denmark)

    Pop, Paul; Eles, Petru; Peng, Zebo

    2006-01-01

    and scheduling policies. In this context, the task of designing such systems is becoming increasingly difficult. The success of new adequate design methods depends on the availability of efficient analysis as well as optimization techniques. In this paper, we present both analysis and optimization approaches...... characteristic to this class of systems: mapping of functionality, the optimization of the access to the communication channel, and the assignment of scheduling policies to processes. Optimization heuristics aiming at producing a schedulable system, with a given amount of resources, are presented....

  14. Real Time Analysis: Does Navy Have a Plan?

    Science.gov (United States)

    2015-06-12

    our militaries face. Peter Tran, senior director of the worldwide advanced cyber defense practice at RSA , commented that “Historically, the...should look like for analysis on the data Navy needs to analyze when he stated: Computers, communications links, encryption devices, and other IT allow

  15. Interactive Construction Digital Tools With Real Time Analysis

    DEFF Research Database (Denmark)

    Klitgaard, Jens; Kirkegaard, Poul Henning

    2007-01-01

    . The aim of this research is to look into integrated digital design and analysis tools in order to find out if it is suited for use by architects and designers or only by specialists and technicians - and if not, then to look at what can be done to make them more available to architects and designers...... an architect-engineer or hybrid practitioner works simultaneously with both aesthetic and technical design requirements. In this paper the problem of a vague or not existing link between digital design tools, used by architects and designers, and the analysis tools developed by and for engineers is considered......The recent developments in computational design tools have evolved into a sometimes purely digital process which opens up for new perspectives and problems in the sketching process. One of the interesting possibilities lay within the hybrid practitioner- or architect-engineer approach, where...

  16. A human fecal contamination index for ranking impaired recreational watersusing the HF183 quantitative real-time PCR method

    Science.gov (United States)

    Human fecal pollution of surface water remains a public health concern worldwide. As a result, there is a growing interest in the application of human-associated fecal source identification quantitative real-time PCR (qPCR) technologies for recreational water quality risk managem...

  17. Data Acceptance Criteria for Standardized Human-Associated Fecal Source Identification Quantitative Real-Time PCR Methods

    Science.gov (United States)

    There is a growing interest in the application of human-associated fecal sourceidentification quantitative real-time PCR (qPCR) technologies for water quality management. The transition from a research tool to a standardized protocol requires a high degree of confidence in data q...

  18. Quantitation of RHD by real-time polymerase chain reaction for determination of RHD zygosity and RHD mosaicism/chimerism

    DEFF Research Database (Denmark)

    Krog, Grethe Risum; Clausen, Frederik Banch; Dziegiel, Morten Hanefeld

    2007-01-01

    Determination of RHD zygosity of the spouse is crucial in preconception counseling of families with history of hemolytic disease of the fetus and newborn caused by anti-D. RHD zygosity can be determined by quantitative real-time polymerase chain reaction (PCR) basically by determining RHD dosage,......, and this feature is relevant in investigating RHD mosaicism and chimerism....

  19. Development of a non invasion real-time PCR assay for the quantitation of chicken parvovirus in fecal swabs

    Science.gov (United States)

    The present study describes the development of a real time Taqman polymerase chain reaction (PCR) assay using a fluorescent labeled probe for the detection and quantitation of chicken parvovirus (ChPV) in feces. The primers and probes were designed based on the nucleotide sequence of the non struct...

  20. EVALUATION OF A RAPID, QUANTITATIVE REAL-TIME PCR METHOD FOR ENUMERATION OF PATHOGENIC CANDIDA CELLS IN WATER

    Science.gov (United States)

    Quantitative Real-Time PCR (QRT-PCR) technology, incorporating fluorigenic 5' nuclease (TaqMan?) chemistry, was developed for the specific detection and quantification of six pathogenic species of Candida (C. albicans, C. tropicalis, C. krusei, C. parapsilosis, C. glabrata and C....

  1. Decay Of Bacterial Pathogens, Fecal Indicators, And Real-Time Quantitative PCR Genetic Markers In Manure-Amended Soils

    Science.gov (United States)

    This study examined persistence and decay of bacterial pathogens, fecal indicator bacteria (FIB), and emerging real-time quantitative PCR (qPCR) genetic markers for rapid detection of fecal pollution in manure-amended agricultural soils. Known concentrations of transformed green...

  2. Decay Of Bacterial Pathogen, Fecal Indicators, And Real-Time Quantitative PCR Genetic Markers In Manure Amended Soils

    Science.gov (United States)

    This study examined persistence and decay of bacterial pathogens, fecal indicator bacteria, and emerging real-time quantitative PCR (qPCR) genetic markers for rapid detection of fecal pollution in manre-amended agricultural soils. Known concentrations of transformed green fluore...

  3. Surrogate analysis and index developer (SAID) tool and real-time data dissemination utilities

    Science.gov (United States)

    Domanski, Marian M.; Straub, Timothy D.; Wood, Molly S.; Landers, Mark N.; Wall, Gary R.; Brady, Steven J.

    2015-01-01

    The use of acoustic and other parameters as surrogates for suspended-sediment concentrations (SSC) in rivers has been successful in multiple applications across the Nation. Critical to advancing the operational use of surrogates are tools to process and evaluate the data along with the subsequent development of regression models from which real-time sediment concentrations can be made available to the public. Recent developments in both areas are having an immediate impact on surrogate research, and on surrogate monitoring sites currently in operation. The Surrogate Analysis and Index Developer (SAID) standalone tool, under development by the U.S. Geological Survey (USGS), assists in the creation of regression models that relate response and explanatory variables by providing visual and quantitative diagnostics to the user. SAID also processes acoustic parameters to be used as explanatory variables for suspended-sediment concentrations. The sediment acoustic method utilizes acoustic parameters from fixed-mount stationary equipment. The background theory and method used by the tool have been described in recent publications, and the tool also serves to support sediment-acoustic-index methods being drafted by the multi-agency Sediment Acoustic Leadership Team (SALT), and other surrogate guidelines like USGS Techniques and Methods 3-C4 for turbidity and SSC. The regression models in SAID can be used in utilities that have been developed to work with the USGS National Water Information System (NWIS) and for the USGS National Real-Time Water Quality (NRTWQ) Web site. The real-time dissemination of predicted SSC and prediction intervals for each time step has substantial potential to improve understanding of sediment-related water-quality and associated engineering and ecological management decisions.

  4. The economic analysis of power market architectures: application to real-time market design

    International Nuclear Information System (INIS)

    Saguan, M.

    2007-04-01

    This work contributes to the economic analysis of power market architectures. A modular framework is used to separate problems of market design in different modules. The work's goal is to study real-time market design. A two-stage market equilibrium model is used to analyse the two main real-time designs: the 'market' and the 'mechanism' (with penalty). Numerical simulations show that design applied in real-time is not neutral vis-a-vis of energy markets sequence and the competition dynamic. Designs using penalty (mechanisms) cause distortions, inefficiencies and can create barriers to entry. The size of distortions is given by the temporal position of the gate that closure the forward markets. This model has also allowed us to show the key role of real-time integration between zones and the importance of good harmonization between real-time designs of each zone. (author)

  5. A triplex quantitative real-time PCR assay for differential detection of human adenovirus serotypes 2, 3 and 7.

    Science.gov (United States)

    Qiu, Fang-Zhou; Shen, Xin-Xin; Zhao, Meng-Chuan; Zhao, Li; Duan, Su-Xia; Chen, Chen; Qi, Ju-Ju; Li, Gui-Xia; Wang, Le; Feng, Zhi-Shan; Ma, Xue-Jun

    2018-05-02

    Human adenovirus (HAdV) serotypes 2, 3 and 7 are more prevalent than other serotypes and have been associated with severe pneumonia in pediatric children. Molecular typing of HAdV is not routinely performed in clinical diagnostic laboratories as it is time-consuming and labor-intensive. In the present study, we developed a triplex quantitative real-time PCR assay (tq-PCR) in a single closed tube for differential detection and quantitative analysis of HAdV serotypes 2, 3 and 7. The sensitivity, specificity, reproducibility and clinical performance of tq-PCR were evaluated. The analytical sensitivity of the tq-PCR was 100 copies/reaction for each of HAdV serotypes 2, 3 and 7, and no cross-reaction with other common respiratory viruses or HAdV serotypes 1,4,5,6,31,55 and 57 was observed. The coefficients of variation (CV) of intra-assay and inter-assay were between 0.6% to 3.6%. Of 138 previously-defined HAdV-positive nasopharyngeal aspirates samples tested, the detection agreement between tq-PCR and nested PCR was 96.38% (133/138). The proposed tq-PCR assay is a sensitive, specific and reproducible method and has the potential for clinical use in the rapid and differential detection and quantitation of HAdV serotypes 2, 3 and 7.

  6. Quantitative real-time monitoring of multi-elements in airborne particulates by direct introduction into an inductively coupled plasma mass spectrometer

    International Nuclear Information System (INIS)

    Suzuki, Yoshinari; Sato, Hikaru; Hiyoshi, Katsuhiro; Furuta, Naoki

    2012-01-01

    A new calibration system for real-time determination of trace elements in airborne particulates was developed. Airborne particulates were directly introduced into an inductively coupled plasma mass spectrometer, and the concentrations of 15 trace elements were determined by means of an external calibration method. External standard solutions were nebulized by an ultrasonic nebulizer (USN) coupled with a desolvation system, and the resulting aerosol was introduced into the plasma. The efficiency of sample introduction via the USN was calculated by two methods: (1) the introduction of a Cr standard solution via the USN was compared with introduction of a Cr(CO) 6 standard gas via a standard gas generator and (2) the aerosol generated by the USN was trapped on filters and then analyzed. The Cr introduction efficiencies obtained by the two methods were the same, and the introduction efficiencies of the other elements were equal to the introduction efficiency of Cr. Our results indicated that our calibration method for introduction efficiency worked well for the 15 elements (Ti, V, Cr, Mn, Co, Ni, Cu, Zn, As, Mo, Sn, Sb, Ba, Tl and Pb). The real-time data and the filter-collection data agreed well for elements with low-melting oxides (V, Co, As, Mo, Sb, Tl, and Pb). In contrast, the real-time data were smaller than the filter-collection data for elements with high-melting oxides (Ti, Cr, Mn, Ni, Cu, Zn, Sn, and Ba). This result implies that the oxides of these 8 elements were not completely fused, vaporized, atomized, and ionized in the initial radiation zone of the inductively coupled plasma. However, quantitative real-time monitoring can be realized after correction for the element recoveries which can be calculated from the ratio of real-time data/filter-collection data. - Highlights: ► APs were directly introduced into ICP-MS and real-time analysis was performed. ► The real-time data were calibrated by a multi-element standard solution from USN. ► During real-time

  7. Estimating marginal properties of quantitative real-time PCR data using nonlinear mixed models

    DEFF Research Database (Denmark)

    Gerhard, Daniel; Bremer, Melanie; Ritz, Christian

    2014-01-01

    A unified modeling framework based on a set of nonlinear mixed models is proposed for flexible modeling of gene expression in real-time PCR experiments. Focus is on estimating the marginal or population-based derived parameters: cycle thresholds and ΔΔc(t), but retaining the conditional mixed mod...

  8. Development of a real-time quantitative assay for detection of Epstein-Barr virus

    NARCIS (Netherlands)

    Niesters, H. G.; van Esser, J.; Fries, E.; Wolthers, K. C.; Cornelissen, J.; Osterhaus, A. D.

    2000-01-01

    With the use of real-time PCR, we developed and evaluated a rapid, sensitive, specific, and reproducible method for the detection of Epstein-Barr virus (EBV) DNA in plasma samples. This method allowed us to screen plasma and serum samples over a range between 100 and 10(7) copies of DNA per ml using

  9. Development of a real-time quantitative assay for detection of Epstein-Barr virus

    NARCIS (Netherlands)

    H.G.M. Niesters (Bert); E. Fries; K.C. Wolthers (Katja); A.D.M.E. Osterhaus (Albert); J.J. Cornelissen (Jan); J.W.J. van Esser (Joost)

    2000-01-01

    textabstractWith the use of real-time PCR, we developed and evaluated a rapid, sensitive, specific, and reproducible method for the detection of Epstein-Barr virus (EBV) DNA in plasma samples. This method allowed us to screen plasma and serum samples over a range between 100 and

  10. Comparison of Hybrid Capture 2 Assay with Real-time-PCR for Detection and Quantitation of Hepatitis B Virus DNA.

    Science.gov (United States)

    Majid, Farjana; Jahan, Munira; Lutful Moben, Ahmed; Tabassum, Shahina

    2014-01-01

    Both real-time-polymerase chain reaction (PCR) and hybrid capture 2 (HC2) assay can detect and quantify hepatitis B virus (HBV) DNA. However, real-time-PCR can detect a wide range of HBV DNA, while HC2 assay could not detect lower levels of viremia. The present study was designed to detect and quantify HBV DNA by real-time-PCR and HC2 assay and compare the quantitative data of these two assays. A cross-sectional study was conducted in between July 2010 and June 2011. A total of 66 serologically diagnosed chronic hepatitis B (CHB) patients were selected for the study. Real-time-PCR and HC2 assay was done to detect HBV DNA. Data were analyzed by statistical Package for the social sciences (SPSS). Among 66 serologically diagnosed chronic hepatitis B patients 40 (60.61%) patients had detectable and 26 (39.39%) had undetectable HBV DNA by HC2 assay. Concordant results were obtained for 40 (60.61%) out of these 66 patients by real-time-PCR and HC2 assay with mean viral load of 7.06 ± 1.13 log 10 copies/ml and 6.95 ± 1.08 log 10 copies/ml, respectively. In the remaining 26 patients, HBV DNA was detectable by real-time-PCR in 20 patients (mean HBV DNA level was 3.67 ± 0.72 log 10 copies/ml. However, HBV DNA could not be detectable in six cases by the both assays. The study showed strong correlation (r = 0.915) between real-time-PCR and HC2 assay for the detection and quantification of HBV DNA. HC2 assay may be used as an alternative to real-time-PCR for CHB patients. How to cite this article: Majid F, Jahan M, Moben AL, Tabassum S. Comparison of Hybrid Capture 2 Assay with Real-time-PCR for Detection and Quantitation of Hepatitis B Virus DNA. Euroasian J Hepato-Gastroenterol 2014;4(1):31-35.

  11. Sequential processing of quantitative phase images for the study of cell behaviour in real-time digital holographic microscopy.

    Science.gov (United States)

    Zikmund, T; Kvasnica, L; Týč, M; Křížová, A; Colláková, J; Chmelík, R

    2014-11-01

    Transmitted light holographic microscopy is particularly used for quantitative phase imaging of transparent microscopic objects such as living cells. The study of the cell is based on extraction of the dynamic data on cell behaviour from the time-lapse sequence of the phase images. However, the phase images are affected by the phase aberrations that make the analysis particularly difficult. This is because the phase deformation is prone to change during long-term experiments. Here, we present a novel algorithm for sequential processing of living cells phase images in a time-lapse sequence. The algorithm compensates for the deformation of a phase image using weighted least-squares surface fitting. Moreover, it identifies and segments the individual cells in the phase image. All these procedures are performed automatically and applied immediately after obtaining every single phase image. This property of the algorithm is important for real-time cell quantitative phase imaging and instantaneous control of the course of the experiment by playback of the recorded sequence up to actual time. Such operator's intervention is a forerunner of process automation derived from image analysis. The efficiency of the propounded algorithm is demonstrated on images of rat fibrosarcoma cells using an off-axis holographic microscope. © 2014 The Authors Journal of Microscopy © 2014 Royal Microscopical Society.

  12. Using quantitative real-time PCR to detect chimeras in transgenic tobacco and apricot and to monitor their dissociation

    Directory of Open Access Journals (Sweden)

    Burgos Lorenzo

    2010-07-01

    Full Text Available Abstract Background The routine generation of transgenic plants involves analysis of transgene integration into the host genome by means of Southern blotting. However, this technique cannot distinguish between uniformly transformed tissues and the presence of a mixture of transgenic and non-transgenic cells in the same tissue. On the other hand, the use of reporter genes often fails to accurately detect chimerical tissues because their expression can be affected by several factors, including gene silencing and plant development. So, new approaches based on the quantification of the amount of the transgene are needed urgently. Results We show here that chimeras are a very frequent phenomenon observed after regenerating transgenic plants. Spatial and temporal analyses of transformed tobacco and apricot plants with a quantitative, real-time PCR amplification of the neomycin phosphotransferase (nptII transgene as well as of an internal control (β-actin, used to normalise the amount of target DNA at each reaction, allowed detection of chimeras at unexpected rates. The amount of the nptII transgene differed greatly along with the sub-cultivation period of these plants and was dependent on the localisation of the analysed leaves; being higher in roots and basal leaves, while in the apical leaves it remained at lower levels. These data demonstrate that, unlike the use of the gus marker gene, real-time PCR is a powerful tool for detection of chimeras. Although some authors have proposed a consistent, positive Southern analysis as an alternative methodology for monitoring the dissociation of chimeras, our data show that it does not provide enough proof of uniform transformation. In this work, however, real-time PCR was applied successfully to monitor the dissociation of chimeras in tobacco plants and apricot callus. Conclusions We have developed a rapid and reliable method to detect and estimate the level of chimeras in transgenic tobacco and apricot

  13. Real time computer control of a nonlinear Multivariable System via Linearization and Stability Analysis

    International Nuclear Information System (INIS)

    Raza, K.S.M.

    2004-01-01

    This paper demonstrates that if a complicated nonlinear, non-square, state-coupled multi variable system is smartly linearized and subjected to a thorough stability analysis then we can achieve our design objectives via a controller which will be quite simple (in term of resource usage and execution time) and very efficient (in terms of robustness). Further the aim is to implement this controller via computer in a real time environment. Therefore first a nonlinear mathematical model of the system is achieved. An intelligent work is done to decouple the multivariable system. Linearization and stability analysis techniques are employed for the development of a linearized and mathematically sound control law. Nonlinearities like the saturation in actuators are also been catered. The controller is then discretized using Runge-Kutta integration. Finally the discretized control law is programmed in a computer in a real time environment. The programme is done in RT -Linux using GNU C for the real time realization of the control scheme. The real time processes, like sampling and controlled actuation, and the non real time processes, like graphical user interface and display, are programmed as different tasks. The issue of inter process communication, between real time and non real time task is addressed quite carefully. The results of this research pursuit are presented graphically. (author)

  14. Video digitizer (real time-frame grabber) with region of interest suitable for quantitative data analysis used on the infrared and H alpha cameras installed on the DIII-D experiment

    International Nuclear Information System (INIS)

    Ferguson, S.W.; Kevan, D.K.; Hill, D.N.; Allen, S.L.

    1987-01-01

    This paper describes a CAMAC based video digitizer with region of interest (ROI) capability that was designed for use with the infrared and H alpha cameras installed by Lawrence Livermore Laboratory on the DIII-D experiment at G.A. Technologies in San Diego, California. The video digitizer uses a custom built CAMAC video synchronizer module to clock data into a CAMAC transient recorder on a line-by-line basis starting at the beginning of a field. The number of fields that are recorded is limited only by the available transient recorder memory. In order to conserve memory, the CAMAC video synchronizer module provides for the alternative selection of a specific region of interest in each successive field to be recorded. Memory conservation can be optimized by specifying lines in the field, start time, stop time, and the number of data samples per line. This video frame grabber has proved versatile for capturing video in such diverse applications as recording video fields from a video tape recorder played in slow motion or recording video fields in real time during a DIII-D shot. In other cases, one or more lines of video are recorded per frame to give a cross sectional slice of the plasma. Since all the data in the digitizer memory is synchronized to video fields and lines, the data can be read directly into the control computer in the proper matrix format to facilitate rapid processing, display, and permanent storage

  15. Effectiveness of quantitative real time PCR in long-term follow-up of chronic myeloid leukemia patients

    International Nuclear Information System (INIS)

    Savasoglu, K.; Berber, B.

    2015-01-01

    To determine the use of the Quantitative Real Time PCR (RQ-PCR) assay follow-up with Chronic Myeloid Leukemia (CML) patients. Study Design: Cross-sectional observational. Place and Duration of Study: Izmir Ataturk Education and Research Hospital, Izmir, Turkey, from 2009 to 2013. Methodology: Cytogenetic, FISH, RQ-PCR test results from 177 CML patients materials selected between 2009 - 2013 years was set up for comparison analysis. Statistical analysis was performed to compare between FISH, karyotype and RQ-PCR results of the patients. Karyotyping and FISH specificity and sensitivity rates determined by ROC analysis compared with RQ-PCR results. Chi-square test was used to compare test failure rates. Results:Sensitivity and specificity values were determined for karyotyping 17.6 - 98% (p=0.118, p > 0.05) and for FISH 22.5 - 96% (p=0.064, p > 0.05) respectively. FISH sensitivity was slightly higher than karyotyping but there was calculated a strong correlation between them (p < 0.001). RQ-PCR test failure rate did not correlate with other two tests (p > 0.05); however, karyotyping and FISH test failure rate was statistically significant (p < 0.001). Conclusion: Besides, the situation needed for karyotype analysis, RQ-PCR assay can be used alone in the follow-up of CML disease. (author)

  16. Effectiveness of Quantitative Real Time PCR in Long-Term Follow-up of Chronic Myeloid Leukemia Patients.

    Science.gov (United States)

    Savasoglu, Kaan; Payzin, Kadriye Bahriye; Ozdemirkiran, Fusun; Berber, Belgin

    2015-08-01

    To determine the use of the Quantitative Real Time PCR (RQ-PCR) assay follow-up with Chronic Myeloid Leukemia (CML) patients. Cross-sectional observational. Izmir Ataturk Education and Research Hospital, Izmir, Turkey, from 2009 to 2013. Cytogenetic, FISH, RQ-PCR test results from 177 CMLpatients' materials selected between 2009 - 2013 years was set up for comparison analysis. Statistical analysis was performed to compare between FISH, karyotype and RQ-PCR results of the patients. Karyotyping and FISH specificity and sensitivity rates determined by ROC analysis compared with RQ-PCR results. Chi-square test was used to compare test failure rates. Sensitivity and specificity values were determined for karyotyping 17.6 - 98% (p=0.118, p > 0.05) and for FISH 22.5 - 96% (p=0.064, p > 0.05) respectively. FISH sensitivity was slightly higher than karyotyping but there was calculated a strong correlation between them (p 0.05); however, karyotyping and FISH test failure rate was statistically significant (p < 0.001). Besides, the situation needed for karyotype analysis, RQ-PCR assay can be used alone in the follow-up of CMLdisease.

  17. Cost Analysis for Real-time Java Scoped-memory Areas

    Directory of Open Access Journals (Sweden)

    Delvin Defoe

    2007-08-01

    Full Text Available Java has recently joined C and C++ as a development platform for real-time and embedded applications. Java's garbage collection, while generally a useful feature, can be problematic for these applications: garbage collection occurs at unpredictable times and its latency is typically unbounded. This can compromise necessary real-time guarantees. To overcome these limitations, the Real-Time for Java Expert Group (RTJEG proposed the Real-Time Specification for Java (RTSJ, which introduced new memory models and new threads to utilize those models. One such memory model uses scoped-memory areas, which work best in the context of a NoHeapRealtimeThread (NHRT. Although much work has been done with scoped-memory areas and NHRTs, there is no system-independent analysis of their costs. In this article we present an asymptotic analysis for RTSJ scoped-memory areas and NHRTs.

  18. Development and evaluation of a real-time one step Reverse-Transcriptase PCR for quantitation of Chandipura Virus

    Directory of Open Access Journals (Sweden)

    Tandale Babasaheb V

    2008-12-01

    Full Text Available Abstract Background Chandipura virus (CHPV, a member of family Rhabdoviridae was attributed to an explosive outbreak of acute encephalitis in children in Andhra Pradesh, India in 2003 and a small outbreak among tribal children from Gujarat, Western India in 2004. The case-fatality rate ranged from 55–75%. Considering the rapid progression of the disease and high mortality, a highly sensitive method for quantifying CHPV RNA by real-time one step reverse transcriptase PCR (real-time one step RT-PCR using TaqMan technology was developed for rapid diagnosis. Methods Primers and probe for P gene were designed and used to standardize real-time one step RT-PCR assay for CHPV RNA quantitation. Standard RNA was prepared by PCR amplification, TA cloning and run off transcription. The optimized real-time one step RT-PCR assay was compared with the diagnostic nested RT-PCR and different virus isolation systems [in vivo (mice in ovo (eggs, in vitro (Vero E6, PS, RD and Sand fly cell line] for the detection of CHPV. Sensitivity and specificity of real-time one step RT-PCR assay was evaluated with diagnostic nested RT-PCR, which is considered as a gold standard. Results Real-time one step RT-PCR was optimized using in vitro transcribed (IVT RNA. Standard curve showed linear relationship for wide range of 102-1010 (r2 = 0.99 with maximum Coefficient of variation (CV = 5.91% for IVT RNA. The newly developed real-time RT-PCR was at par with nested RT-PCR in sensitivity and superior to cell lines and other living systems (embryonated eggs and infant mice used for the isolation of the virus. Detection limit of real-time one step RT-PCR and nested RT-PCR was found to be 1.2 × 100 PFU/ml. RD cells, sand fly cells, infant mice, and embryonated eggs showed almost equal sensitivity (1.2 × 102 PFU/ml. Vero and PS cell-lines (1.2 × 103 PFU/ml were least sensitive to CHPV infection. Specificity of the assay was found to be 100% when RNA from other viruses or healthy

  19. QPCR: Application for real-time PCR data management and analysis

    Directory of Open Access Journals (Sweden)

    Eichhorn Heiko

    2009-08-01

    Full Text Available Abstract Background Since its introduction quantitative real-time polymerase chain reaction (qPCR has become the standard method for quantification of gene expression. Its high sensitivity, large dynamic range, and accuracy led to the development of numerous applications with an increasing number of samples to be analyzed. Data analysis consists of a number of steps, which have to be carried out in several different applications. Currently, no single tool is available which incorporates storage, management, and multiple methods covering the complete analysis pipeline. Results QPCR is a versatile web-based Java application that allows to store, manage, and analyze data from relative quantification qPCR experiments. It comprises a parser to import generated data from qPCR instruments and includes a variety of analysis methods to calculate cycle-threshold and amplification efficiency values. The analysis pipeline includes technical and biological replicate handling, incorporation of sample or gene specific efficiency, normalization using single or multiple reference genes, inter-run calibration, and fold change calculation. Moreover, the application supports assessment of error propagation throughout all analysis steps and allows conducting statistical tests on biological replicates. Results can be visualized in customizable charts and exported for further investigation. Conclusion We have developed a web-based system designed to enhance and facilitate the analysis of qPCR experiments. It covers the complete analysis workflow combining parsing, analysis, and generation of charts into one single application. The system is freely available at http://genome.tugraz.at/QPCR

  20. MTF analysis of the near-real time neutron radiography facility at MURR

    International Nuclear Information System (INIS)

    Lindsay, J.T.; Alger, D.M.; Bull, S.R.; Miller, W.H.

    1983-01-01

    Several neutron radiography systems designed to view transient processes on a real-time basis have been developed. With the advent of these different real-time systems comes the necessity to develop a means to quantitatively evaluate and compare these systems. A suitable method for measuring the resolution capabilities of the image-forming system is the determination of the modulation transfer function (MTF). The MTF is a measure of an imaging system's ability to reproduce the spatial frequencies present in an image. The system in use at the University of Missouri Research Reactor is described. (Auth.)

  1. Development of Transient-Reactor Analysis Code (TRAC) for real-time applications

    International Nuclear Information System (INIS)

    Niederauer, G.F.; Giguere, P.T.; Lime, J.F.; Knight, T.D.; Ashy, O.; Fakory, R.

    1997-01-01

    This is the final report of a six-month, Laboratory-Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). Nuclear-plant training simulators employ simplified one-dimensional thermal-hydraulics codes because of the demands to run in real time and with limited computing power. The objective of this project was to investigate the feasibility of using the advanced Transient-Reactor Analysis Code (TRAC) in a simulator to increase the fidelity of a simulator. Many issues need to be addressed to take such a complex code from a batch engineering environment to a real-time environment. Working with simulator vendor, GSE, the authors investigated the technical issues relating to integrating TRAC into a real-time environment. They also modified a nuclear power plant model for simulator purposes and investigated its performance in real time

  2. Application of droplet digital PCR for quantitative detection of Spiroplasma citri in comparison with real time PCR.

    Directory of Open Access Journals (Sweden)

    Yogita Maheshwari

    Full Text Available Droplet digital polymerase chain reaction (ddPCR is a method for performing digital PCR that is based on water-oil emulsion droplet technology. It is a unique approach to measure the absolute copy number of nucleic acid targets without the need of external standards. This study evaluated the applicability of ddPCR as a quantitative detection tool for the Spiroplasma citri, causal agent of citrus stubborn disease (CSD in citrus. Two sets of primers, SP1, based on the spiral in housekeeping gene, and a multicopy prophage gene, SpV1 ORF1, were used to evaluate ddPCR in comparison with real time (quantitative PCR (qPCR for S. citri detection in citrus tissues. Standard curve analyses on tenfold dilution series showed that both ddPCR and qPCR exhibited good linearity and efficiency. However, ddPCR had a tenfold greater sensitivity than qPCR and accurately quantified up to one copy of spiralin gene. Receiver operating characteristic analysis indicated that the ddPCR methodology was more robust for diagnosis of CSD and the area under the curve was significantly broader compared to qPCR. Field samples were used to validate ddPCR efficacy and demonstrated that it was equal or better than qPCR to detect S. citri infection in fruit columella due to a higher pathogen titer. The ddPCR assay detected both the S. citri spiralin and the SpV1 ORF1 targets quantitatively with high precision and accuracy compared to qPCR assay. The ddPCR was highly reproducible and repeatable for both the targets and showed higher resilience to PCR inhibitors in citrus tissue extract for the quantification of S. citri compare to qPCR.

  3. Effect of ionizing radiation on the quantitative detection of Salmonella using real-time PCR

    Energy Technology Data Exchange (ETDEWEB)

    Lim, Sangyong; Jung, Jinwoo [Radiation Research Center for Biotechnology, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of); Kim, Minjeong; Ryu, Sangryeol [Department of Food and Animal Biotechnology, School of Agricultural Biotechnology, Center for Agricultural Biomaterials, Seoul National University, Seoul 151-921 (Korea, Republic of); Kim, Dongho [Radiation Research Center for Biotechnology, Korea Atomic Energy Research Institute, Jeongeup 580-185 (Korea, Republic of)], E-mail: fungikim@kaeri.re.kr

    2008-09-15

    Food irradiation is an economically viable technology for inactivating foodborne pathogens, but irradiation can mask pathogens in unhygienically prepared food. The aim of this study was to investigate the effect of irradiation treatment on the detection of Salmonella using real-time PCR. Three commercially available kits were tested, of which the InstaGene Matrix procedure was most effective in preparing template DNA from Salmonella exposed to radiation in broth culture. The minimum level of detection by real-time PCR combined with InstaGene Matrix was 3 log units of Salmonella per milliliter. However, when pure cultures of Salmonella were irradiated at 3 and 5 kGy, the cycle threshold (C{sub T}) increased 1-1.5-fold compared to irradiation at 0 and 1 kGy. This indicated that irradiation treatment may result in an underestimation of bacterial counts due to radiation-induced DNA lesions. We also compared C{sub T} values in inoculated chicken homogenates before and after irradiation, which in this model caused a 1.3-3.3-fold underestimation of bacterial counts with respect to irradiation dose.

  4. Design and Validation of Real-Time PCR: Quantitative Diagnosis of Common Leishmania Species in Iran.

    Science.gov (United States)

    Fekri Soofi Abadi, Maryam; Dabiri, Shahriar; Fotouhi Ardakani, Reza; Fani Malaki, Lina; Amirpoor Rostami, Sahar; Ziasistani, Mahsa; Dabiri, Donya

    2016-07-01

    Design and validation of Real-time PCR on the protected gene region ITS2 to quantify the parasite load in common leishmania (L) species. Probe and primer were designed from the ITS2 region between the rRNA genes with minimum gene variation in three common leishmania species followed by a Real-time PCR using the Taq man probe method in the form of absolute quantification. A series of different concentrations of leishmania were analyzed. After the purified PCR product was successfully placed in a PTG19-T plasmid vector, specialized ITS2 region was cloned in this plasmid. In the last phase, the cloned gene was transferred to the Ecoli.Top10F bacteria. The standard plasmid was provided in 10(7) to 10(1) copies/rxn concentrations. The specification and clinical sensitivity of the data was analyzed using inter and intra scales. The probe and primer were designed using three species, including L. infantum, L. major, and L.tropica. Seven concentrations of purified parasite in culture media showed that the selected region for quantifying the parasite is suitable. Clinical and analytical specificity and sensitivity were both 100%, respectively. The Taq man method for the ITS2 region in leishmania is one the most sensitive diagnostic test for identifying the parasite load and is suggested as a tool for fast identification and quantification of species.

  5. Determination of drugs and drug-like compounds in different samples with direct analysis in real time mass spectrometry.

    Science.gov (United States)

    Chernetsova, Elena S; Morlock, Gertrud E

    2011-01-01

    Direct analysis in real time (DART), a relatively new ionization source for mass spectrometry, ionizes small-molecule components from different kinds of samples without any sample preparation and chromatographic separation. The current paper reviews the published data available on the determination of drugs and drug-like compounds in different matrices with DART-MS, including identification and quantitation issues. Parameters that affect ionization efficiency and mass spectra composition are also discussed. Copyright © 2011 Wiley Periodicals, Inc.

  6. Identification and validation of reference genes for quantitative real-time PCR in Drosophila suzukii (Diptera: Drosophilidae.

    Directory of Open Access Journals (Sweden)

    Yifan Zhai

    Full Text Available To accurately evaluate gene expression levels and obtain more accurate quantitative real-time RT-PCR (qRT-PCR data, normalization relative to reliable reference gene(s is required. Drosophila suzukii, is an invasive fruit pest native to East Asia, and recently invaded Europe and North America, the stability of its reference genes have not been previously investigated. In this study, ten candidate reference genes (RPL18, RPS3, AK, EF-1β, TBP, NADH, HSP22, GAPDH, Actin, α-Tubulin, were evaluated for their suitability as normalization genes under different biotic (developmental stage, tissue and population, and abiotic (photoperiod, temperature conditions. The three statistical approaches (geNorm, NormFinder and BestKeeper and one web-based comprehensive tool (RefFinder were used to normalize analysis of the ten candidate reference genes identified α-Tubulin, TBP and AK as the most stable candidates, while HSP22 and Actin showed the lowest expression stability. We used three most stable genes (α-Tubulin, TBP and AK and one unstably expressed gene to analyze the expression of P-glycoprotein in abamectin-resistant and sensitive strains, and the results were similar to reference genes α-Tubulin, TBP and AK, which show good stability, while the result of HSP22 has a certain bias. The three validated reference genes can be widely used for quantification of target gene expression with qRT-PCR technology in D.suzukii.

  7. Standardisation and evaluation of a quantitative multiplex real-time PCR assay for the rapid identification of Streptococcus pneumoniae

    Directory of Open Access Journals (Sweden)

    Feroze Ahmed Ganaie

    2015-01-01

    Full Text Available Rapid diagnosis of Streptococcus pneumoniae can play a significant role in decreasing morbidity and mortality of infection. The accurate diagnosis of pneumococcal disease is hampered by the difficulties in growing the isolates from clinical specimens and also by misidentification. Molecular methods have gained popularity as they offer improvement in the detection of causative pathogens with speed and ease. The present study aims at validating and standardising the use of 4 oligonucleotide primer-probe sets (pneumolysin [ply], autolysin [lytA], pneumococcal surface adhesion A [psaA] and Spn9802 [DNA fragment] in a single-reaction mixture for the detection and discrimination of S. pneumoniae. Here, we validate a quantitative multiplex real-time PCR (qmPCR assay with a panel consisting of 43 S. pneumoniae and 29 non-pneumococcal isolates, 20 culture positive, 26 culture negative and 30 spiked serum samples. A standard curve was obtained using S. pneumoniae ATCC 49619 strain and glyceraldehyde 3-phosphate dehydrogenase (GAPDH gene was used as an endogenous internal control. The experiment showed high sensitivity with lower limit of detection equivalent to 4 genome copies/µl. The efficiency of the reaction was 100% for ply, lytA, Spn9802 and 97% for psaA. The test showed sensitivity and specificity of 100% with culture isolates and serum specimens. This study demonstrates that qmPCR analysis of sera using 4 oligonucleotide primers appears to be an appropriate method for the genotypic identification of S. pneumoniae infection.

  8. Real-time visualization and analysis of airflow field by use of digital holography

    Science.gov (United States)

    Di, Jianglei; Wu, Bingjing; Chen, Xin; Liu, Junjiang; Wang, Jun; Zhao, Jianlin

    2013-04-01

    The measurement and analysis of airflow field is very important in fluid dynamics. For airflow, smoke particles can be added to visually observe the turbulence phenomena by particle tracking technology, but the effect of smoke particles to follow the high speed airflow will reduce the measurement accuracy. In recent years, with the advantage of non-contact, nondestructive, fast and full-field measurement, digital holography has been widely applied in many fields, such as deformation and vibration analysis, particle characterization, refractive index measurement, and so on. In this paper, we present a method to measure the airflow field by use of digital holography. A small wind tunnel model made of acrylic glass is built to control the velocity and direction of airflow. Different shapes of samples such as aircraft wing and cylinder are placed in the wind tunnel model to produce different forms of flow field. With a Mach-Zehnder interferometer setup, a series of digital holograms carrying the information of airflow filed distributions in different states are recorded by CCD camera and corresponding holographic images are numerically reconstructed from the holograms by computer. Then we can conveniently obtain the velocity or pressure information of the airflow deduced from the quantitative phase information of holographic images and visually display the airflow filed and its evolution in the form of a movie. The theory and experiment results show that digital holography is a robust and feasible approach for real-time visualization and analysis of airflow field.

  9. Detection and identification of Rift Valley fever virus in mosquito vectors by quantitative real-time PCR.

    Science.gov (United States)

    Mwaengo, D; Lorenzo, G; Iglesias, J; Warigia, M; Sang, R; Bishop, R P; Brun, A

    2012-10-01

    Diagnostic methods allowing for rapid identification of pathogens are crucial for controlling and preventing dissemination after disease outbreaks as well as for use in surveillance programs. For arboviruses, detection of the presence of virus in their arthropod hosts is important for monitoring of viral activity and quantitative information is useful for modeling of transmission dynamics. In this study, molecular detection of Rift Valley fever virus (RVFV) in mosquito samples from the 2006 to 2007 East African outbreaks was performed using quantitative real-time PCR assay (qRT-PCR). Specific RVFV sequence-based primer/fluorogenic (TaqMan) probe sets were derived from the L and S RNA segments of the virus. Both primer-probe L and S segment-based combinations detected genomic RVFV sequences, with generally comparable levels of sensitivity. Viral loads from three mosquito species, Aedes mcintoshi, Aedes ochraceus and Mansonia uniformis were estimated and significant differences of between 5- and 1000-fold were detected between Ae. mcintoshi and M. uniformis using both the L and S primer-probe-based assays. The genetic relationships of the viral sequences in mosquito samples were established by partial M segment sequencing and assigned to the two previously described viral lineages defined by analysis of livestock isolates obtained during the 2006-2007 outbreak, confirming that similar viruses were present in both the vector and mammalian host. The data confirms the utility of qRT-PCR for identification and initial quantification of virus in mosquito samples during RVFV outbreaks. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. The Real-time Frequency Spectrum Analysis of Neutron Pulse Signal Series

    International Nuclear Information System (INIS)

    Tang Yuelin; Ren Yong; Wei Biao; Feng Peng; Mi Deling; Pan Yingjun; Li Jiansheng; Ye Cenming

    2009-01-01

    The frequency spectrum analysis of neutron pulse signal is a very important method in nuclear stochastic signal processing Focused on the special '0' and '1' of neutron pulse signal series, this paper proposes new rotation-table and realizes a real-time frequency spectrum algorithm under 1G Hz sample rate based on PC with add, address and SSE. The numerical experimental results show that under the count rate of 3X10 6 s -1 , this algorithm is superior to FFTW in time-consumption and can meet the real-time requirement of frequency spectrum analysis. (authors)

  11. Flipping the analytical coin : closing the information flow loop in high speed (real time) analysis

    NARCIS (Netherlands)

    K.E. Shahroudi

    1997-01-01

    textabstractAnalysis modules tend to be set up as one way flow of information, i.e a clear distinction between cause and effect or input and output. However, as the speed of analysis approaches real time (or faster than movie rate), it becomes increasingly difficult for an external user to

  12. Validation of an intermediate heat exchanger model for real time analysis

    International Nuclear Information System (INIS)

    Tzanos, C.P.

    1986-11-01

    A new method was presented for LMFBR intermediate heat exchanger (IHX) analysis in real time for purposes of continuous on-line data validation, plant state verification and fault identification. For the validation of this methodology the EBR-II IHX transient during Test 8A was analyzed. This paper presents the results of this analysis

  13. Quantitative real-time PCR as a sensitive protein-protein interaction quantification method and a partial solution for non-accessible autoactivator and false-negative molecule analysis in the yeast two-hybrid system.

    Science.gov (United States)

    Maier, Richard H; Maier, Christina J; Hintner, Helmut; Bauer, Johann W; Onder, Kamil

    2012-12-01

    Many functional proteomic experiments make use of high-throughput technologies such as mass spectrometry combined with two-dimensional polyacrylamide gel electrophoresis and the yeast two-hybrid (Y2H) system. Currently there are even automated versions of the Y2H system available that can be used for proteome-wide research. The Y2H system has the capacity to deliver a profusion of Y2H positive colonies from a single library screen. However, subsequent analysis of these numerous primary candidates with complementary methods can be overwhelming. Therefore, a method to select the most promising candidates with strong interaction properties might be useful to reduce the number of candidates requiring further analysis. The method described here offers a new way of quantifying and rating the performance of positive Y2H candidates. The novelty lies in the detection and measurement of mRNA expression instead of proteins or conventional Y2H genetic reporters. This method correlates well with the direct genetic reporter readouts usually used in the Y2H system, and has greater sensitivity for detecting and quantifying protein-protein interactions (PPIs) than the conventional Y2H system, as demonstrated by detection of the Y2H false-negative PPI of RXR/PPARG. Approximately 20% of all proteins are not suitable for the Y2H system, the so-called autoactivators. A further advantage of this method is the possibility to evaluate molecules that usually cannot be analyzed in the Y2H system, exemplified by a VDR-LXXLL motif peptide interaction. Copyright © 2012 Elsevier Inc. All rights reserved.

  14. Noninvasive Strategy Based on Real-Time in Vivo Cataluminescence Monitoring for Clinical Breath Analysis.

    Science.gov (United States)

    Zhang, Runkun; Huang, Wanting; Li, Gongke; Hu, Yufei

    2017-03-21

    The development of noninvasive methods for real-time in vivo analysis is of great significant, which provides powerful tools for medical research and clinical diagnosis. In the present work, we described a new strategy based on cataluminescence (CTL) for real-time in vivo clinical breath analysis. To illustrate such strategy, a homemade real-time CTL monitoring system characterized by coupling an online sampling device with a CTL sensor for sevoflurane (SVF) was designed, and a real-time in vivo method for the monitoring of SVF in exhaled breath was proposed. The accuracy of the method was evaluated by analyzing the real exhaled breath samples, and the results were compared with those obtained by GC/MS. The measured data obtained by the two methods were in good agreement. Subsequently, the method was applied to real-time monitoring of SVF in exhaled breath from rat models of the control group to investigate elimination pharmacokinetics. In order to further probe the potential of the method for clinical application, the elimination pharmacokinetics of SVF from rat models of control group, liver fibrosis group alcohol liver group, and nonalcoholic fatty liver group were monitored by the method. The raw data of pharmacokinetics of different groups were normalized and subsequently subjected to linear discriminant analysis (LDA). These data were transformed to canonical scores which were visualized as well-clustered with the classification accuracy of 100%, and the overall accuracy of leave-one-out cross-validation procedure is 88%, thereby indicating the utility of the potential of the method for liver disease diagnosis. Our strategy undoubtedly opens up a new door for real-time clinical analysis in a pain-free and noninvasive way and also guides a promising development direction for CTL.

  15. Real-Time Shop-Floor Production Performance Analysis Method for the Internet of Manufacturing Things

    Directory of Open Access Journals (Sweden)

    Yingfeng Zhang

    2014-04-01

    Full Text Available Typical challenges that manufacturing enterprises are facing now are compounded by lack of timely, accurate, and consistent information of manufacturing resources. As a result, it is difficult to analyze the real-time production performance for the shop-floor. In this paper, the definition and overall architecture of the internet of manufacturing things is presented to provide a new paradigm by extending the techniques of internet of things (IoT to manufacturing field. Under this architecture, the real-time primitive events which occurred at different manufacturing things such as operators, machines, pallets, key materials, and so forth can be easily sensed. Based on these distributed primitive events, a critical event model is established to automatically analyze the real-time production performance. Here, the up-level production performance analysis is regarded as a series of critical events, and the real-time value of each critical event can be easily calculated according to the logical and sequence relationships among these multilevel events. Finally, a case study is used to illustrate how to apply the designed methods to analyze the real-time production performance.

  16. Simultaneous quantitative assessment of circulating cell-free mitochondrial and nuclear DNA by multiplex real-time PCR

    Directory of Open Access Journals (Sweden)

    Peng Xia

    2009-01-01

    Full Text Available Quantification of circulating nucleic acids in plasma and serum could be used as a non-invasive diagnostic tool for monitoring a wide variety of diseases and conditions. We describe here a rapid, simple and accurate multiplex real-time PCR method for direct synchronized analysis of circulating cell-free (ccf mitochondrial (mtDNA and nuclear (nDNA DNA in plasma and serum samples. The method is based on one-step multiplex real-time PCR using a FAM-labeled MGB probe and primers to amplify the mtDNA sequence of the ATP 8 gene, and a VIC-labeled MGB probe and primers to amplify the nDNA sequence of the glycerinaldehyde-3-phosphate-dehydrogenase (GAPDH gene, in plasma and serum samples simultaneously. The efficiencies of the multiplex assays were measured in serial dilutions. Based on the simulation of the PCR reaction kinetics, the relative quantities of ccf mtDNA were calculated using a very simple equation. Using our optimised real-time PCR conditions, close to 100% efficiency was obtained from the two assays. The two assays performed in the dilution series showed very good and reproducible correlation to each other. This optimised multiplex real-time PCR protocol can be widely used for synchronized quantification of mtDNA and nDNA in different samples, with a very high rate of efficiency.

  17. Development and validation of a real-time quantitative PCR assay to detect Xanthomonas axonopodis pv. allii from onion seed.

    Science.gov (United States)

    Robène, Isabelle; Perret, Marion; Jouen, Emmanuel; Escalon, Aline; Maillot, Marie-Véronique; Chabirand, Aude; Moreau, Aurélie; Laurent, Annie; Chiroleu, Frédéric; Pruvost, Olivier

    2015-07-01

    Bacterial blight of onion is an emerging disease threatening world onion production. The causal agent Xanthomonas axonopodis pv. allii is seed transmitted and a reliable and sensitive tool is needed to monitor seed exchanges. A triplex quantitative real-time PCR assay was developed targeting two X. axonopodis pv. allii-specific markers and an internal control chosen in 5.8S rRNA gene from Alliaceae. Amplification of at least one marker indicates the presence of the bacterium in seed extracts. This real-time PCR assay detected all the 79 X. axonopodis pv. allii strains tested and excluded 85.2% of the 135 non-target strains and particularly all 39 saprophytic and pathogenic bacteria associated with onion. Cross-reactions were mainly obtained for strains assigned to nine phylogenetically related X. axonopodis pathovars. The cycle cut-off was estimated statistically at 36.3 considering a risk of false positive of 1%. The limit of detection obtained in at least 95% of the time (LOD 95%) was 5×10(3) CFU/g (colony forming unit/g). The sensitivity threshold was found to be 1 infected seed in 32,790 seeds. This real-time PCR assay should be useful for preventing the long-distance spread of X. axonopodis pv. allii via contaminated seed lots and determining the epidemiology of the bacterium. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Development of real-time multitask OSS based on cognitive task analysis

    International Nuclear Information System (INIS)

    Wang He; Cheng Shouyu

    2010-01-01

    A Real-time Multi-task Operator Support System (RMOSS) has been developed to support the operator's decision making process in the control room of NPP. VxWorks, one embedded real-time operation system, is used for RMOSS software development. According to the SRK modeling analysis result of the operator' decision making process, RMOSS is divided into five system subtasks, including Data Collection and Validation Task (DCVT), Operation Monitor Task (OMT), Fault Diagnostic Task (FDT), Operation Guideline Task (OGT) and Human Machine Interface Task (HMIT). The task test of RMOSS has been done in a real-time full scope simulator. The results showed that each task of RMOSS is capable of accomplishing their functions. (authors)

  19. Analysis and Synthesis of Communication-Intensive Heterogeneous Real-Time Systems

    DEFF Research Database (Denmark)

    Pop, Paul

    2003-01-01

    Embedded computer systems are now everywhere: from alarm clocks to PDAs, from mobile phones to cars, almost all the devices we use are controlled by embedded computer systems. An important class of embedded computer systems is that of real-time systems, which have to fulfill strict timing...... requirements. As realtime systems become more complex, they are often implemented using distributed heterogeneous architectures. The main objective of this thesis is to develop analysis and synthesis methods for communication-intensive heterogeneous hard real-time systems. The systems are heterogeneous...... is the synthesis of the communication infrastructure, which has a significant impact on the overall system performance and cost. To reduce the time-to-market of products, the design of real-time systems seldom starts from scratch. Typically, designers start from an already existing system, running certain...

  20. MAC-Level Communication Time Modeling and Analysis for Real-Time WSNs

    Directory of Open Access Journals (Sweden)

    STANGACIU, V.

    2016-02-01

    Full Text Available Low-level communication protocols and their timing behavior are essential to developing wireless sensor networks (WSNs able to provide the support and operating guarantees required by many current real-time applications. Nevertheless, this aspect still remains an issue in the state-of-the-art. In this paper we provide a detailed analysis of a recently proposed MAC-level communication timing model and demonstrate its usability in designing real-time protocols. The results of a large set of measurements are also presented and discussed here, in direct relation to the main time parameters of the analyzed model.

  1. Chemisorption of iodine-125 to gold nanoparticles allows for real-time quantitation and potential use in nanomedicine

    Energy Technology Data Exchange (ETDEWEB)

    Walsh, Adrian A, E-mail: a.walsh@nanobiosols.com [Liverpool Science Park, Nano Biosols Ltd (United Kingdom)

    2017-04-15

    Gold nanoparticles have been available for many years as a research tool in the life sciences due to their electron density and optical properties. New applications are continually being developed, particularly in nanomedicine. One drawback is the need for an easy, real-time quantitation method for gold nanoparticles so that the effects observed in in vitro cell toxicity assays and cell uptake studies can be interpreted quantitatively in terms of nanoparticle loading. One potential method of quantifying gold nanoparticles in real time is by chemisorption of iodine-125, a gamma emitter, to the nanoparticles. This paper revisits the labelling of gold nanoparticles with iodine-125, first described 30 years ago and never fully exploited since. We explore the chemical properties and usefulness in quantifying bio-functionalised gold nanoparticle binding in a quick and simple manner. The gold particles were labelled specifically and quantitatively simply by mixing the two items. The nature of the labelling is chemisorption and is robust, remaining bound over several weeks in a variety of cell culture media. Chemisorption was confirmed as potassium iodide can remove the label whereas sodium chloride and many other buffers had no effect. Particles precoated in polymers or proteins can be labelled just as efficiently allowing for post-labelling experiments in situ rather than using radioactive gold atoms in the production process. We also demonstrate that interparticle exchange of I-125 between different size particles does not appear to take place confirming the affinity of the binding.

  2. Chemisorption of iodine-125 to gold nanoparticles allows for real-time quantitation and potential use in nanomedicine

    Science.gov (United States)

    Walsh, Adrian A.

    2017-04-01

    Gold nanoparticles have been available for many years as a research tool in the life sciences due to their electron density and optical properties. New applications are continually being developed, particularly in nanomedicine. One drawback is the need for an easy, real-time quantitation method for gold nanoparticles so that the effects observed in in vitro cell toxicity assays and cell uptake studies can be interpreted quantitatively in terms of nanoparticle loading. One potential method of quantifying gold nanoparticles in real time is by chemisorption of iodine-125, a gamma emitter, to the nanoparticles. This paper revisits the labelling of gold nanoparticles with iodine-125, first described 30 years ago and never fully exploited since. We explore the chemical properties and usefulness in quantifying bio-functionalised gold nanoparticle binding in a quick and simple manner. The gold particles were labelled specifically and quantitatively simply by mixing the two items. The nature of the labelling is chemisorption and is robust, remaining bound over several weeks in a variety of cell culture media. Chemisorption was confirmed as potassium iodide can remove the label whereas sodium chloride and many other buffers had no effect. Particles precoated in polymers or proteins can be labelled just as efficiently allowing for post-labelling experiments in situ rather than using radioactive gold atoms in the production process. We also demonstrate that interparticle exchange of I-125 between different size particles does not appear to take place confirming the affinity of the binding.

  3. Chemisorption of iodine-125 to gold nanoparticles allows for real-time quantitation and potential use in nanomedicine

    International Nuclear Information System (INIS)

    Walsh, Adrian A

    2017-01-01

    Gold nanoparticles have been available for many years as a research tool in the life sciences due to their electron density and optical properties. New applications are continually being developed, particularly in nanomedicine. One drawback is the need for an easy, real-time quantitation method for gold nanoparticles so that the effects observed in in vitro cell toxicity assays and cell uptake studies can be interpreted quantitatively in terms of nanoparticle loading. One potential method of quantifying gold nanoparticles in real time is by chemisorption of iodine-125, a gamma emitter, to the nanoparticles. This paper revisits the labelling of gold nanoparticles with iodine-125, first described 30 years ago and never fully exploited since. We explore the chemical properties and usefulness in quantifying bio-functionalised gold nanoparticle binding in a quick and simple manner. The gold particles were labelled specifically and quantitatively simply by mixing the two items. The nature of the labelling is chemisorption and is robust, remaining bound over several weeks in a variety of cell culture media. Chemisorption was confirmed as potassium iodide can remove the label whereas sodium chloride and many other buffers had no effect. Particles precoated in polymers or proteins can be labelled just as efficiently allowing for post-labelling experiments in situ rather than using radioactive gold atoms in the production process. We also demonstrate that interparticle exchange of I-125 between different size particles does not appear to take place confirming the affinity of the binding.

  4. Real-time PCR analysis of carbon catabolite repression of cellobiose gene transcription in Trametes versicolor

    Energy Technology Data Exchange (ETDEWEB)

    Stapleton, P. C.; O' Mahoney, J.; Dobson, A. D. W. [National University of Ireland, Microbiology Department, Cork (Ireland)

    2004-02-01

    Previous reports indicate that in white rot fungi such as Trametes versicolor, the production of cellobiose dehydrogenase (CDH), an extracellular haemo-flavo-enzyme, is subject to carbon catabolite repression by both glucose and maltose, and that the repression is mediated at the transcriptional level. This paper describes the results of an investigation of CDH gene transcription in cellulolytic cultures of T. versicolor, in the presence of other additional carbon sources such as glucose, arabinose, and xylose. Using real time polymerase chain reaction (RT-PCR) assay methods in the presence of these other additional carbon sources, the levels of repression observed are quantitatively determined in an effort to obtain more accurate measurements of carbon catabolite repression of CDH production in this ligninolytic fungus. Ninety-six hours after addition, results of the analysis showed reduction in CDH transcript levels of 19-fold for galactose, 92-fold for arabinose and 114-fold for xylose. The greatest repressive effect was exhibited by glucose. In this case the reduction in CDH transcript levels was 3400-fold. CDH plays an important role in lignin degradation, and there is also substantial interest in the biotechnological applications of CDH, most particularly in the pulp and paper industry. 24 refs., 4 figs.

  5. Stacker’s Crane Position Fixing Based on Real Time Image Processing and Analysis

    Directory of Open Access Journals (Sweden)

    Kmeid Saad

    2015-06-01

    Full Text Available This study illustrates the usage of stacker cranes and image processing in automated warehouse systems. The aim is to use real time image processing and analysis for a stacker’s crane position fixing in order to use it as a pick-up and delivery system (P/D, to be controlled by a programmable logic controller unit (PLC.

  6. Improved SIRAP analysis for synchronization in hierarchical scheduled real-time systems

    NARCIS (Netherlands)

    Behnam, M.; Bril, R.J.; Nolte, T.

    2009-01-01

    We present our ongoing work on synchronization in hierarchical scheduled real-time systems, where tasks are scheduled using fixed-priority pre-emptive scheduling. In this paper, we show that the original local schedulability analysis of the synchronization protocol SIRAP [4] is very pessimistic when

  7. Hybrid Real-time Zero-day Malware Analysis and Reporting System

    OpenAIRE

    Ratinder Kaur; Maninder Singh

    2016-01-01

    To understand completely the malicious intents of a zero-day malware there is really no automated way. There is no single best approach for malware analysis so it demands to combine existing static, dynamic and manual malware analysis techniques in a single unit. In this paper a hybrid real-time analysis and reporting system is presented. The proposed system integrates various malware analysis tools and utilities in a component-based architecture. The system automatica...

  8. Evaluation and validation of candidate endogenous control genes for real-time quantitative PCR studies of breast cancer

    Directory of Open Access Journals (Sweden)

    Miller Nicola

    2007-11-01

    Full Text Available Abstract Background Real-time quantitative PCR (RQ-PCR forms the basis of many breast cancer biomarker studies and novel prognostic assays, paving the way towards personalised cancer treatments. Normalisation of relative RQ-PCR data is required to control for non-biological variation introduced during sample preparation. Endogenous control (EC genes, used in this context, should ideally be expressed constitutively and uniformly across treatments in all test samples. Despite widespread recognition that the accuracy of the normalised data is largely dependent on the reliability of the EC, there are no reports of the systematic validation of genes commonly used for this purpose in the analysis of gene expression by RQ-PCR in primary breast cancer tissues. The aim of this study was to identify the most suitable endogenous control genes for RQ-PCR analysis of primary breast tissue from a panel of eleven candidates in current use. Oestrogen receptor alpha (ESR1 was used a target gene to compare the effect of choice of EC on the estimate of gene quantity. Results The expression and validity of candidate ECs (GAPDH, TFRC, ABL, PPIA, HPRT1, RPLP0, B2M, GUSB, MRPL19, PUM1 and PSMC4 was determined in 6 benign and 21 malignant primary breast cancer tissues. Gene expression data was analysed using two different statistical models. MRPL19 and PPIA were identified as the most stable and reliable EC genes, while GUSB, RPLP0 and ABL were least stable. There was a highly significant difference in variance between ECs. ESR1 expression was appreciably higher in malignant compared to benign tissues and there was a significant effect of EC on the magnitude of the error associated with the relative quantity of ESR1. Conclusion We have validated two endogenous control genes, MRPL19 and PPIA, for RQ-PCR analysis of gene expression in primary breast tissue. Of the genes in current use in this field, the above combination offers increased accuracy and resolution in the

  9. Real-Time Analysis and Forecasting of Multisite River Flow Using a Distributed Hydrological Model

    Directory of Open Access Journals (Sweden)

    Mingdong Sun

    2014-01-01

    Full Text Available A spatial distributed hydrological forecasting system was developed to promote the analysis of river flow dynamic state in a large basin. The research presented the real-time analysis and forecasting of multisite river flow in the Nakdong River Basin using a distributed hydrological model with radar rainfall forecast data. A real-time calibration algorithm of hydrological distributed model was proposed to investigate the particular relationship between the water storage and basin discharge. Demonstrate the approach of simulating multisite river flow using a distributed hydrological model couple with real-time calibration and forecasting of multisite river flow with radar rainfall forecasts data. The hydrographs and results exhibit that calibrated flow simulations are very approximate to the flow observation at all sites and the accuracy of forecasting flow is gradually decreased with lead times extending from 1 hr to 3 hrs. The flow forecasts are lower than the flow observation which is likely caused by the low estimation of radar rainfall forecasts. The research has well demonstrated that the distributed hydrological model is readily applicable for multisite real-time river flow analysis and forecasting in a large basin.

  10. Comparison between quantitative nucleic acid sequence-based amplification, real-time reverse transcriptase PCR, and real-time PCR for quantification of Leishmania parasites

    NARCIS (Netherlands)

    van der Meide, Wendy; Guerra, Jorge; Schoone, Gerard; Farenhorst, Marit; Coelho, Leila; Faber, William; Peekel, Inge; Schallig, Henk

    2008-01-01

    DNA or RNA amplification methods for detection of Leishmania parasites have advantages regarding sensitivity and potential quantitative characteristics in comparison with conventional diagnostic methods but are often still not routinely applied. However, the use and application of molecular assays

  11. TaqMan MGB probe fluorescence real-time quantitative PCR for rapid detection of Chinese Sacbrood virus.

    Directory of Open Access Journals (Sweden)

    Ma Mingxiao

    Full Text Available Sacbrood virus (SBV is a picorna-like virus that affects honey bees (Apis mellifera and results in the death of the larvae. Several procedures are available to detect Chinese SBV (CSBV in clinical samples, but not to estimate the level of CSBV infection. The aim of this study was develop an assay for rapid detection and quantification of this virus. Primers and probes were designed that were specific for CSBV structural protein genes. A TaqMan minor groove binder (MGB probe-based, fluorescence real-time quantitative PCR was established. The specificity, sensitivity and stability of the assay were assessed; specificity was high and there were no cross-reactivity with healthy larvae or other bee viruses. The assay was applied to detect CSBV in 37 clinical samples and its efficiency was compared with clinical diagnosis, electron microscopy observation, and conventional RT-PCR. The TaqMan MGB-based probe fluorescence real-time quantitative PCR for CSBV was more sensitive than other methods tested. This assay was a reliable, fast, and sensitive method that was used successfully to detect CSBV in clinical samples. The technology can provide a useful tool for rapid detection of CSBV. This study has established a useful protocol for CSBV testing, epidemiological investigation, and development of animal models.

  12. Methodology for object-oriented real-time systems analysis and design: Software engineering

    Science.gov (United States)

    Schoeffler, James D.

    1991-01-01

    Successful application of software engineering methodologies requires an integrated analysis and design life-cycle in which the various phases flow smoothly 'seamlessly' from analysis through design to implementation. Furthermore, different analysis methodologies often lead to different structuring of the system so that the transition from analysis to design may be awkward depending on the design methodology to be used. This is especially important when object-oriented programming is to be used for implementation when the original specification and perhaps high-level design is non-object oriented. Two approaches to real-time systems analysis which can lead to an object-oriented design are contrasted: (1) modeling the system using structured analysis with real-time extensions which emphasizes data and control flows followed by the abstraction of objects where the operations or methods of the objects correspond to processes in the data flow diagrams and then design in terms of these objects; and (2) modeling the system from the beginning as a set of naturally occurring concurrent entities (objects) each having its own time-behavior defined by a set of states and state-transition rules and seamlessly transforming the analysis models into high-level design models. A new concept of a 'real-time systems-analysis object' is introduced and becomes the basic building block of a series of seamlessly-connected models which progress from the object-oriented real-time systems analysis and design system analysis logical models through the physical architectural models and the high-level design stages. The methodology is appropriate to the overall specification including hardware and software modules. In software modules, the systems analysis objects are transformed into software objects.

  13. Real-time quantitative Schlieren imaging by fast Fourier demodulation of a checkered backdrop

    Science.gov (United States)

    Wildeman, Sander

    2018-06-01

    A quantitative synthetic Schlieren imaging (SSI) method based on fast Fourier demodulation is presented. Instead of a random dot pattern (as usually employed in SSI), a 2D periodic pattern (such as a checkerboard) is used as a backdrop to the refractive object of interest. The range of validity and accuracy of this "Fast Checkerboard Demodulation" (FCD) method are assessed using both synthetic data and experimental recordings of patterns optically distorted by small waves on a water surface. It is found that the FCD method is at least as accurate as sophisticated, multi-stage, digital image correlation (DIC) or optical flow (OF) techniques used with random dot patterns, and it is significantly faster. Efficient, fully vectorized, implementations of both the FCD and DIC/OF schemes developed for this study are made available as open source Matlab scripts.

  14. Design base transient analysis using the real-time nuclear reactor simulator model

    International Nuclear Information System (INIS)

    Tien, K.K.; Yakura, S.J.; Morin, J.P.; Gregory, M.V.

    1987-01-01

    A real-time simulation model has been developed to describe the dynamic response of all major systems in a nuclear process reactor. The model consists of a detailed representation of all hydraulic components in the external coolant circulating loops consisting of piping, valves, pumps and heat exchangers. The reactor core is described by a three-dimensional neutron kinetics model with detailed representation of assembly coolant and moderator thermal hydraulics. The models have been developed to support a real-time training simulator, therefore, they reproduce system parameters characteristic of steady state normal operation with high precision. The system responses for postulated severe transients such as large pipe breaks, loss of pumping power, piping leaks, malfunctions in control rod insertion, and emergency injection of neutron absorber are calculated to be in good agreement with reference safety analyses. Restrictions were imposed by the requirement that the resulting code be able to run in real-time with sufficient spare time to allow interfacing with secondary systems and simulator hardware. Due to hardware set-up and real plant instrumentation, simplifications due to symmetry were not allowed. The resulting code represents a coarse-node engineering model in which the level of detail has been tailored to the available computing power of a present generation super-minicomputer. Results for several significant transients, as calculated by the real-time model, are compared both to actual plant data and to results generated by fine-mesh analysis codes

  15. Design base transient analysis using the real-time nuclear reactor simulator model

    International Nuclear Information System (INIS)

    Tien, K.K.; Yakura, S.J.; Morin, J.P.; Gregory, M.V.

    1987-01-01

    A real-time simulation model has been developed to describe the dynamic response of all major systems in a nuclear process reactor. The model consists of a detailed representation of all hydraulic components in the external coolant circulating loops consisting of piping, valves, pumps and heat exchangers. The reactor core is described by a three-dimensional neutron kinetics model with detailed representation of assembly coolant and mode-rator thermal hydraulics. The models have been developed to support a real-time training simulator, therefore, they reproduce system parameters characteristic of steady state normal operation with high precision. The system responses for postulated severe transients such as large pipe breaks, loss of pumping power, piping leaks, malfunctions in control rod insertion, and emergency injection of neutron absorber are calculated to be in good agreement with reference safety analyses. Restrictions were imposed by the requirement that the resulting code be able to run in real-time with sufficient spare time to allow interfacing with secondary systems and simulator hardware. Due to hardware set-up and real plant instrumentation, simplifications due to symmetry were not allowed. The resulting code represents a coarse-node engineering model in which the level of detail has been tailored to the available computing power of a present generation super-minicomputer. Results for several significant transients, as calculated by the real-time model, are compared both to actual plant data and to results generated by fine-mesh analysis codes

  16. An Analysis of a Hard Real-Time Execution Environment Extension for FreeRTOS

    Directory of Open Access Journals (Sweden)

    STANGACIU, C.

    2015-08-01

    Full Text Available FreeRTOS is a popular real-time operating system, which has been under a significant attention in the last years due to its main advantages: it is open source, portable, well documented and implemented on more than 30 architectures. FreeRTOS execution environment is dynamic, preemptive and priority based, but it is not suitable for hard real-time tasks, because it provides task execution determinism only to a certain degree and cannot guarantee the absence of task execution jitter. As a solution to this problem, we propose a hard real time execution extension to FreeRTOS in order to support a particular model of HRT tasks, called ModXs, which are executed with no jitter. This article presents a detailed analysis, in terms of scheduling, task execution and memory usage of this hard real time execution environment extension. The article is concluding with the advantages this extension brings to the system compared to the small memory and timing overhead introduced.

  17. Comparison of lung cancer cell lines representing four histopathological subtypes with gene expression profiling using quantitative real-time PCR

    Directory of Open Access Journals (Sweden)

    Kawaguchi Makoto

    2010-01-01

    Full Text Available Abstract Background Lung cancers are the most common type of human malignancy and are intractable. Lung cancers are generally classified into four histopathological subtypes: adenocarcinoma (AD, squamous cell carcinoma (SQ, large cell carcinoma (LC, and small cell carcinoma (SC. Molecular biological characterization of these subtypes has been performed mainly using DNA microarrays. In this study, we compared the gene expression profiles of these four subtypes using twelve human lung cancer cell lines and the more reliable quantitative real-time PCR (qPCR. Results We selected 100 genes from public DNA microarray data and examined them by DNA microarray analysis in eight test cell lines (A549, ABC-1, EBC-1, LK-2, LU65, LU99, STC 1, RERF-LC-MA and a normal control lung cell line (MRC-9. From this, we extracted 19 candidate genes. We quantified the expression of the 19 genes and a housekeeping gene, GAPDH, with qPCR, using the same eight cell lines plus four additional validation lung cancer cell lines (RERF-LC-MS, LC-1/sq, 86-2, and MS-1-L. Finally, we characterized the four subtypes of lung cancer cell lines using principal component analysis (PCA of gene expression profiling for 12 of the 19 genes (AMY2A, CDH1, FOXG1, IGSF3, ISL1, MALL, PLAU, RAB25, S100P, SLCO4A1, STMN1, and TGM2. The combined PCA and gene pathway analyses suggested that these genes were related to cell adhesion, growth, and invasion. S100P in AD cells and CDH1 in AD and SQ cells were identified as candidate markers of these lung cancer subtypes based on their upregulation and the results of PCA analysis. Immunohistochemistry for S100P and RAB25 was closely correlated to gene expression. Conclusions These results show that the four subtypes, represented by 12 lung cancer cell lines, were well characterized using qPCR and PCA for the 12 genes examined. Certain genes, in particular S100P and CDH1, may be especially important for distinguishing the different subtypes. Our results

  18. Real-time GPS seismology using a single receiver: method comparison, error analysis and precision validation

    Science.gov (United States)

    Li, Xingxing

    2014-05-01

    Earthquake monitoring and early warning system for hazard assessment and mitigation has traditional been based on seismic instruments. However, for large seismic events, it is difficult for traditional seismic instruments to produce accurate and reliable displacements because of the saturation of broadband seismometers and problematic integration of strong-motion data. Compared with the traditional seismic instruments, GPS can measure arbitrarily large dynamic displacements without saturation, making them particularly valuable in case of large earthquakes and tsunamis. GPS relative positioning approach is usually adopted to estimate seismic displacements since centimeter-level accuracy can be achieved in real-time by processing double-differenced carrier-phase observables. However, relative positioning method requires a local reference station, which might itself be displaced during a large seismic event, resulting in misleading GPS analysis results. Meanwhile, the relative/network approach is time-consuming, particularly difficult for the simultaneous and real-time analysis of GPS data from hundreds or thousands of ground stations. In recent years, several single-receiver approaches for real-time GPS seismology, which can overcome the reference station problem of the relative positioning approach, have been successfully developed and applied to GPS seismology. One available method is real-time precise point positioning (PPP) relied on precise satellite orbit and clock products. However, real-time PPP needs a long (re)convergence period, of about thirty minutes, to resolve integer phase ambiguities and achieve centimeter-level accuracy. In comparison with PPP, Colosimo et al. (2011) proposed a variometric approach to determine the change of position between two adjacent epochs, and then displacements are obtained by a single integration of the delta positions. This approach does not suffer from convergence process, but the single integration from delta positions to

  19. Evaluation of methods for oligonucleotide array data via quantitative real-time PCR.

    Science.gov (United States)

    Qin, Li-Xuan; Beyer, Richard P; Hudson, Francesca N; Linford, Nancy J; Morris, Daryl E; Kerr, Kathleen F

    2006-01-17

    There are currently many different methods for processing and summarizing probe-level data from Affymetrix oligonucleotide arrays. It is of great interest to validate these methods and identify those that are most effective. There is no single best way to do this validation, and a variety of approaches is needed. Moreover, gene expression data are collected to answer a variety of scientific questions, and the same method may not be best for all questions. Only a handful of validation studies have been done so far, most of which rely on spike-in datasets and focus on the question of detecting differential expression. Here we seek methods that excel at estimating relative expression. We evaluate methods by identifying those that give the strongest linear association between expression measurements by array and the "gold-standard" assay. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) is generally considered the "gold-standard" assay for measuring gene expression by biologists and is often used to confirm findings from microarray data. Here we use qRT-PCR measurements to validate methods for the components of processing oligo array data: background adjustment, normalization, mismatch adjustment, and probeset summary. An advantage of our approach over spike-in studies is that methods are validated on a real dataset that was collected to address a scientific question. We initially identify three of six popular methods that consistently produced the best agreement between oligo array and RT-PCR data for medium- and high-intensity genes. The three methods are generally known as MAS5, gcRMA, and the dChip mismatch mode. For medium- and high-intensity genes, we identified use of data from mismatch probes (as in MAS5 and dChip mismatch) and a sequence-based method of background adjustment (as in gcRMA) as the most important factors in methods' performances. However, we found poor reliability for methods using mismatch probes for low-intensity genes

  20. Evaluation of methods for oligonucleotide array data via quantitative real-time PCR

    Directory of Open Access Journals (Sweden)

    Morris Daryl E

    2006-01-01

    Full Text Available Abstract Background There are currently many different methods for processing and summarizing probe-level data from Affymetrix oligonucleotide arrays. It is of great interest to validate these methods and identify those that are most effective. There is no single best way to do this validation, and a variety of approaches is needed. Moreover, gene expression data are collected to answer a variety of scientific questions, and the same method may not be best for all questions. Only a handful of validation studies have been done so far, most of which rely on spike-in datasets and focus on the question of detecting differential expression. Here we seek methods that excel at estimating relative expression. We evaluate methods by identifying those that give the strongest linear association between expression measurements by array and the "gold-standard" assay. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR is generally considered the "gold-standard" assay for measuring gene expression by biologists and is often used to confirm findings from microarray data. Here we use qRT-PCR measurements to validate methods for the components of processing oligo array data: background adjustment, normalization, mismatch adjustment, and probeset summary. An advantage of our approach over spike-in studies is that methods are validated on a real dataset that was collected to address a scientific question. Results We initially identify three of six popular methods that consistently produced the best agreement between oligo array and RT-PCR data for medium- and high-intensity genes. The three methods are generally known as MAS5, gcRMA, and the dChip mismatch mode. For medium- and high-intensity genes, we identified use of data from mismatch probes (as in MAS5 and dChip mismatch and a sequence-based method of background adjustment (as in gcRMA as the most important factors in methods' performances. However, we found poor reliability for methods

  1. Assessment of Real-Time Compaction Quality Test Indexes for Rockfill Material Based on Roller Vibratory Acceleration Analysis

    Directory of Open Access Journals (Sweden)

    Tianbo Hua

    2018-01-01

    Full Text Available Compaction quality is directly related to the structure and seepage stability of a rockfill dam. To timely and accurately test the compaction quality of the rockfill material, four real-time test indexes were chosen to characterize the soil compaction degree based on the analysis of roller vibratory acceleration, including acceleration peak value (ap, acceleration root mean square value (arms, crest factor value (CF, and compaction meter value (CMV. To determine which of these indexes is the most appropriate, a two-part field compaction experiment was conducted using a vibratory roller in different filling zones of the dam body. Data on rolling parameters, real-time test indexes, and compaction quality indexes were collected to perform statistical regression analyses. Combined with the spectrum analysis of the acceleration signal, it was found that the CF index best characterizes the compaction degree of the rockfill material among the four indexes. Furthermore, the quantitative relations between the real-time index and compaction quality index were established to determine the control criterion of CF, which can instruct the site work of compaction quality control in the rockfill rolling process.

  2. Evaluation of reference genes for real-time quantitative PCR studies in Candida glabrata following azole treatment

    Directory of Open Access Journals (Sweden)

    Li Qingdi

    2012-06-01

    Full Text Available Abstract Background The selection of stable and suitable reference genes for real-time quantitative PCR (RT-qPCR is a crucial prerequisite for reliable gene expression analysis under different experimental conditions. The present study aimed to identify reference genes as internal controls for gene expression studies by RT-qPCR in azole-stimulated Candida glabrata. Results The expression stability of 16 reference genes under fluconazole stress was evaluated using fold change and standard deviation computations with the hkgFinder tool. Our data revealed that the mRNA expression levels of three ribosomal RNAs (RDN5.8, RDN18, and RDN25 remained stable in response to fluconazole, while PGK1, UBC7, and UBC13 mRNAs showed only approximately 2.9-, 3.0-, and 2.5-fold induction by azole, respectively. By contrast, mRNA levels of the other 10 reference genes (ACT1, EF1α, GAPDH, PPIA, RPL2A, RPL10, RPL13A, SDHA, TUB1, and UBC4 were dramatically increased in C. glabrata following antifungal treatment, exhibiting changes ranging from 4.5- to 32.7-fold. We also assessed the expression stability of these reference genes using the 2-ΔΔCT method and three other software packages. The stability rankings of the reference genes by geNorm and the 2-ΔΔCT method were identical to those by hkgFinder, whereas the stability rankings by BestKeeper and NormFinder were notably different. We then validated the suitability of six candidate reference genes (ACT1, PGK1, RDN5.8, RDN18, UBC7, and UBC13 as internal controls for ten target genes in this system using the comparative CT method. Our validation experiments passed for all six reference genes analyzed except RDN18, where the amplification efficiency of RDN18 was different from that of the ten target genes. Finally, we demonstrated that the relative quantification of target gene expression varied according to the endogenous control used, highlighting the importance of the choice of internal controls in such

  3. The PWI [plutonium waste incinerator] expert system: Real time, PC-based process analysis

    International Nuclear Information System (INIS)

    Brown, K.G.; Smith, F.G.

    1987-01-01

    A real time, microcomputer-based expert system is being developed for a prototype plutonium waste incinerator (PWI) process at Du Pont's Savannah River Laboratory. The expert system will diagnose instrumentation problems, assist operator training, serve as a repository for engineering knowledge about the process, and provide continuous operation and performance information. A set of necessary operational criteria was developed from process and engineering constraints; it was used to define hardware and software needs. The most important criterion is operating speed because the analysis operates in real time. TURBO PROLOG by Borland International was selected. The analysis system is divided into three sections: the user-system interface, the inference engine and rule base, and the files representing the blackboard information center

  4. Real-time spectral analysis of HRV signals: an interactive and user-friendly PC system.

    Science.gov (United States)

    Basano, L; Canepa, F; Ottonello, P

    1998-01-01

    We present a real-time system, built around a PC and a low-cost data acquisition board, for the spectral analysis of the heart rate variability signal. The Windows-like operating environment on which it is based makes the computer program very user-friendly even for non-specialized personnel. The Power Spectral Density is computed through the use of a hybrid method, in which a classical FFT analysis follows an autoregressive finite-extension of data; the stationarity of the sequence is continuously checked. The use of this algorithm gives a high degree of robustness of the spectral estimation. Moreover, always in real time, the FFT of every data block is computed and displayed in order to corroborate the results as well as to allow the user to interactively choose a proper AR model order.

  5. Detecting spatial patterns of rivermouth processes using a geostatistical framework for near-real-time analysis

    Science.gov (United States)

    Xu, Wenzhao; Collingsworth, Paris D.; Bailey, Barbara; Carlson Mazur, Martha L.; Schaeffer, Jeff; Minsker, Barbara

    2017-01-01

    This paper proposes a geospatial analysis framework and software to interpret water-quality sampling data from towed undulating vehicles in near-real time. The framework includes data quality assurance and quality control processes, automated kriging interpolation along undulating paths, and local hotspot and cluster analyses. These methods are implemented in an interactive Web application developed using the Shiny package in the R programming environment to support near-real time analysis along with 2- and 3-D visualizations. The approach is demonstrated using historical sampling data from an undulating vehicle deployed at three rivermouth sites in Lake Michigan during 2011. The normalized root-mean-square error (NRMSE) of the interpolation averages approximately 10% in 3-fold cross validation. The results show that the framework can be used to track river plume dynamics and provide insights on mixing, which could be related to wind and seiche events.

  6. Real-time measurement and control at JET signal processing and physics analysis for diagnostics

    International Nuclear Information System (INIS)

    Felton, R.; Joffrin, E.; Murari, A.

    2005-01-01

    To meet the requirements of the scientific programme, the EFDA JET real-time measurement and control project has developed an integrated set of real-time plasma measurements, experiment control and communication facilities. Traditional experiments collected instrument data during the plasma pulse and calculated physics data after the pulse. The challenge for continuous tokamak operation is to calculate the physics data in real-time, keeping up with the evolution of the plasma. In JET, many plasma diagnostics have been augmented with extra data acquisition and signal-processing systems so that they can both capture instrument data for conventional post-pulse analysis and calculate calibrated, validated physics results in real-time. During the pulse, the systems send sampled data sets into a network, which distributes the data to several destinations. The receiving systems may do further analysis, integrating data from several measurements, or may control the plasma scenario by heating or fuelling. The simplest real-time diagnostic systems apply scale factors to the signals, as with the electron cyclotron emission (ECE) diagnostic's 96 tuned radiometer channels, giving the electron temperature profile. In various spectroscopy diagnostics, spectral features are least-squares-fitted to measure spectra from several lines of sight, within 50 ms. Ion temperatures and rotation speed can be calculated from the line widths and shifts. For diagnostics using modulation techniques, the systems implement digital-signal processing phase trackers, lock-in amplifiers and filters, e.g., the far infrared (FIR) interferometer samples 15 channels at 400 kHz for 30 s, i.e., six million samples per second. Diagnostics have specific lines of sight, spatial channels, and various sampling rates. The heating/fuelling systems have relatively coarse spatial localisation. Analysis systems have been developed to integrate the basic physics data into smaller sets of controllable parameters on a

  7. Progress on RTSS simulation-based analysis for real-time systems development at two laboratories

    International Nuclear Information System (INIS)

    Shu, Y.; Jia, M.; Fei, Y.; Zhang, Y.; Liu, G.; Yang, S.; Chen, Y.

    1996-01-01

    A new object-oriented Real Time System Simulator (RTSS) with the capability for simulation graphics and animation, has been developed and used for modeling the distributed data acquisition and processing systems at JET and ASIPP. Simulation allows estimates of response time, throughput and resource utilization for a variety of configurations to be investigated. Performance measurements, simulation and analysis are used together to calibrate and validate each other

  8. Analysis of the LTE Access Reservation Protocol for Real-Time Traffic

    DEFF Research Database (Denmark)

    Thomsen, Henning; Kiilerich Pratas, Nuno; Stefanovic, Cedomir

    2013-01-01

    LTE is increasingly seen as a system for serving real-time Machine-to-Machine (M2M) communication needs. The asynchronous M2M user access in LTE is obtained through a two-phase access reservation protocol (contention and data phase). Existing analysis related to these protocols is based...... of the two-phase LTE reservation protocol and asses its performance, when assumptions (1) and (2) do not hold....

  9. An Analysis of Input/Output Paradigms for Real-Time Systems

    Science.gov (United States)

    1990-07-01

    timing and concurrency aspects of real - time systems . This paper illustrates how to build a mathematical model of the schedulability of a real-time...various design alternatives. The primary characteristic that distinguishes real-time system from non- real - time systems is the importance of time. The

  10. Stability Analysis and Variational Integrator for Real-Time Formation Based on Potential Field

    Directory of Open Access Journals (Sweden)

    Shengqing Yang

    2014-01-01

    Full Text Available This paper investigates a framework of real-time formation of autonomous vehicles by using potential field and variational integrator. Real-time formation requires vehicles to have coordinated motion and efficient computation. Interactions described by potential field can meet the former requirement which results in a nonlinear system. Stability analysis of such nonlinear system is difficult. Our methodology of stability analysis is discussed in error dynamic system. Transformation of coordinates from inertial frame to body frame can help the stability analysis focus on the structure instead of particular coordinates. Then, the Jacobian of reduced system can be calculated. It can be proved that the formation is stable at the equilibrium point of error dynamic system with the effect of damping force. For consideration of calculation, variational integrator is introduced. It is equivalent to solving algebraic equations. Forced Euler-Lagrange equation in discrete expression is used to construct a forced variational integrator for vehicles in potential field and obstacle environment. By applying forced variational integrator on computation of vehicles' motion, real-time formation of vehicles in obstacle environment can be implemented. Algorithm based on forced variational integrator is designed for a leader-follower formation.

  11. Comparison of real-time quantitative PCR and culture for the diagnosis of emerging Rickettsioses.

    Directory of Open Access Journals (Sweden)

    Emmanouil Angelakis

    Full Text Available BACKGROUND: Isolation of Rickettsia species from skin biopsies may be replaced by PCR. We evaluated culture sensitivity compared to PCR based on sampling delay and previous antibiotic treatment. METHODOLOGY/PRINCIPAL FINDINGS: Skin biopsies and ticks from patients with suspected Rickettsia infection were screened for Rickettsia spp. using qPCR, and positive results were amplified and sequenced for the gltA and ompA genes. Immunofluorescence for spotted fever group rickettsial antigens was done for 79 patients. All skin biopsies and only ticks that tested positive using qPCR were cultured in human embryonic lung (HEL fibroblasts using the centrifugation-shell vial technique. Patients and ticks were classified as definitely having rickettsioses if there was direct evidence of infection with a Rickettsia sp. using culture or molecular assays or in patients if serology was positive. Data on previous antibiotic treatments were obtained for patients with rickettsiosis. Rickettsia spp. infection was diagnosed in 47 out of 145 patients (32%, 41 by PCR and 12 by culture, whereas 3 isolates were obtained from PCR negative biopsies. For 3 of the patients serology was positive although PCR and culture were negative. Rickettsia africae was the most common detected species (n = 25, [17.2%] and isolated bacterium (n = 5, [3.4%]. The probability of isolating Rickettsia spp. was 12 times higher in untreated patients and 5.4 times higher in patients from our hometown. Rickettsia spp. was amplified in 24 out of 95 ticks (25% and we isolated 7 R. slovaca and 1 R. raoultii from Dermacentor marginatus. CONCLUSIONS/SIGNIFICANCE: We found a positive correlation between the bacteria copies and the isolation success in skin biopsies and ticks. Culture remains critical for strain analysis but is less sensitive than serology and PCR for the diagnosis of a Rickettsia infection.

  12. Real-Time PCR-Based Quantitation Method for the Genetically Modified Soybean Line GTS 40-3-2.

    Science.gov (United States)

    Kitta, Kazumi; Takabatake, Reona; Mano, Junichi

    2016-01-01

    This chapter describes a real-time PCR-based method for quantitation of the relative amount of genetically modified (GM) soybean line GTS 40-3-2 [Roundup Ready(®) soybean (RRS)] contained in a batch. The method targets a taxon-specific soybean gene (lectin gene, Le1) and the specific DNA construct junction region between the Petunia hybrida chloroplast transit peptide sequence and the Agrobacterium 5-enolpyruvylshikimate-3-phosphate synthase gene (epsps) sequence present in GTS 40-3-2. The method employs plasmid pMulSL2 as a reference material in order to quantify the relative amount of GTS 40-3-2 in soybean samples using a conversion factor (Cf) equal to the ratio of the RRS-specific DNA to the taxon-specific DNA in representative genuine GTS 40-3-2 seeds.

  13. Species-specific detection and quantification of common barnacle larvae from the Japanese coast using quantitative real-time PCR.

    Science.gov (United States)

    Endo, Noriyuki; Sato, Kana; Matsumura, Kiyotaka; Yoshimura, Erina; Odaka, Yukiko; Nogata, Yasuyuki

    2010-11-01

    Species-specific detection and quantification methods for barnacle larvae using quantitative real-time polymerase chain reaction (qPCR) were developed. Species-specific primers for qPCR were designed for 13 barnacle species in the mitochondrial 12S ribosomal RNA gene region. Primer specificity was examined by PCR using template DNA extracted from each of the 13 barnacle species, other unidentified barnacle species, and field collected zooplankton samples. The resulting PCR products comprised single bands following agarose gel electrophoresis when the templates corresponded to primers. The amplifications were highly species-specific even for the field plankton samples. The field plankton samples were subjected to qPCR assay. The calculated DNA contents for each barnacle species were closely correlated with the number of larvae measured by microscopic examination. The method could be applied to quantify barnacle larvae in natural plankton samples.

  14. Evaluation of reference genes for quantitative real-time PCR in oil palm elite planting materials propagated by tissue culture.

    Directory of Open Access Journals (Sweden)

    Pek-Lan Chan

    Full Text Available BACKGROUND: The somatic embryogenesis tissue culture process has been utilized to propagate high yielding oil palm. Due to the low callogenesis and embryogenesis rates, molecular studies were initiated to identify genes regulating the process, and their expression levels are usually quantified using reverse transcription quantitative real-time PCR (RT-qPCR. With the recent release of oil palm genome sequences, it is crucial to establish a proper strategy for gene analysis using RT-qPCR. Selection of the most suitable reference genes should be performed for accurate quantification of gene expression levels. RESULTS: In this study, eight candidate reference genes selected from cDNA microarray study and literature review were evaluated comprehensively across 26 tissue culture samples using RT-qPCR. These samples were collected from two tissue culture lines and media treatments, which consisted of leaf explants cultures, callus and embryoids from consecutive developmental stages. Three statistical algorithms (geNorm, NormFinder and BestKeeper confirmed that the expression stability of novel reference genes (pOP-EA01332, PD00380 and PD00569 outperformed classical housekeeping genes (GAPDH, NAD5, TUBULIN, UBIQUITIN and ACTIN. PD00380 and PD00569 were identified as the most stably expressed genes in total samples, MA2 and MA8 tissue culture lines. Their applicability to validate the expression profiles of a putative ethylene-responsive transcription factor 3-like gene demonstrated the importance of using the geometric mean of two genes for normalization. CONCLUSIONS: Systematic selection of the most stably expressed reference genes for RT-qPCR was established in oil palm tissue culture samples. PD00380 and PD00569 were selected for accurate and reliable normalization of gene expression data from RT-qPCR. These data will be valuable to the research associated with the tissue culture process. Also, the method described here will facilitate the selection

  15. Evaluation of Reference Genes for Quantitative Real-Time PCR in Oil Palm Elite Planting Materials Propagated by Tissue Culture

    Science.gov (United States)

    Chan, Pek-Lan; Rose, Ray J.; Abdul Murad, Abdul Munir; Zainal, Zamri; Leslie Low, Eng-Ti; Ooi, Leslie Cheng-Li; Ooi, Siew-Eng; Yahya, Suzaini; Singh, Rajinder

    2014-01-01

    Background The somatic embryogenesis tissue culture process has been utilized to propagate high yielding oil palm. Due to the low callogenesis and embryogenesis rates, molecular studies were initiated to identify genes regulating the process, and their expression levels are usually quantified using reverse transcription quantitative real-time PCR (RT-qPCR). With the recent release of oil palm genome sequences, it is crucial to establish a proper strategy for gene analysis using RT-qPCR. Selection of the most suitable reference genes should be performed for accurate quantification of gene expression levels. Results In this study, eight candidate reference genes selected from cDNA microarray study and literature review were evaluated comprehensively across 26 tissue culture samples using RT-qPCR. These samples were collected from two tissue culture lines and media treatments, which consisted of leaf explants cultures, callus and embryoids from consecutive developmental stages. Three statistical algorithms (geNorm, NormFinder and BestKeeper) confirmed that the expression stability of novel reference genes (pOP-EA01332, PD00380 and PD00569) outperformed classical housekeeping genes (GAPDH, NAD5, TUBULIN, UBIQUITIN and ACTIN). PD00380 and PD00569 were identified as the most stably expressed genes in total samples, MA2 and MA8 tissue culture lines. Their applicability to validate the expression profiles of a putative ethylene-responsive transcription factor 3-like gene demonstrated the importance of using the geometric mean of two genes for normalization. Conclusions Systematic selection of the most stably expressed reference genes for RT-qPCR was established in oil palm tissue culture samples. PD00380 and PD00569 were selected for accurate and reliable normalization of gene expression data from RT-qPCR. These data will be valuable to the research associated with the tissue culture process. Also, the method described here will facilitate the selection of appropriate

  16. Comparative Evaluation of Four Real-Time PCR Methods for the Quantitative Detection of Epstein-Barr Virus from Whole Blood Specimens.

    Science.gov (United States)

    Buelow, Daelynn; Sun, Yilun; Tang, Li; Gu, Zhengming; Pounds, Stanley; Hayden, Randall

    2016-07-01

    Monitoring of Epstein-Barr virus (EBV) load in immunocompromised patients has become integral to their care. An increasing number of reagents are available for quantitative detection of EBV; however, there are little published comparative data. Four real-time PCR systems (one using laboratory-developed reagents and three using analyte-specific reagents) were compared with one another for detection of EBV from whole blood. Whole blood specimens seeded with EBV were used to determine quantitative linearity, analytical measurement range, lower limit of detection, and CV for each assay. Retrospective testing of 198 clinical samples was performed in parallel with all methods; results were compared to determine relative quantitative and qualitative performance. All assays showed similar performance. No significant difference was found in limit of detection (3.12-3.49 log10 copies/mL; P = 0.37). A strong qualitative correlation was seen with all assays that used clinical samples (positive detection rates of 89.5%-95.8%). Quantitative correlation of clinical samples across assays was also seen in pairwise regression analysis, with R(2) ranging from 0.83 to 0.95. Normalizing clinical sample results to IU/mL did not alter the quantitative correlation between assays. Quantitative EBV detection by real-time PCR can be performed over a wide linear dynamic range, using three different commercially available reagents and laboratory-developed methods. EBV was detected with comparable sensitivity and quantitative correlation for all assays. Copyright © 2016 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  17. Practical Testing and Performance Analysis of Phasor Measurement Unit Using Real Time Digital Simulator (RTDS)

    DEFF Research Database (Denmark)

    Liu, Leo; Rather, Zakir Hussain; Stearn, Nathen

    2012-01-01

    Wide Area Measurement Systems (WAMS) and Wide Area Monitoring, Protection and Control Systems (WAMPACS) have evolved rapidly over the last two decades [1]. This fast emerging technology enables real time synchronized monitoring of power systems. Presently, WAMS are mainly used for real time...... visualisation and post event analysis of power systems. It is expected however, that through integration with traditional Supervisory Control and Data Acquisition (SCADA) systems, closed loop control applications will be possible. Phasor Measurement Units (PMUs) are fundamental components of WAMS. Large WAMS...... proposed to realize highly precise phasoreasurements. Further a comparative study based on features of PMUs from different major manufacturers is presented. The selection of optimal parameters, such as phasor format and filter length is also discussed for various applications....

  18. Analysis of fault tolerance and reliability in distributed real-time system architectures

    International Nuclear Information System (INIS)

    Philippi, Stephan

    2003-01-01

    Safety critical real-time systems are becoming ubiquitous in many areas of our everyday life. Failures of such systems potentially have catastrophic consequences on different scales, in the worst case even the loss of human life. Therefore, safety critical systems have to meet maximum fault tolerance and reliability requirements. As the design of such systems is far from being trivial, this article focuses on concepts to specifically support the early architectural design. In detail, a simulation based approach for the analysis of fault tolerance and reliability in distributed real-time system architectures is presented. With this approach, safety related features can be evaluated in the early development stages and thus prevent costly redesigns in later ones

  19. A real-time data-acquisition and analysis system with distributed UNIX workstations

    International Nuclear Information System (INIS)

    Yamashita, H.; Miyamoto, K.; Maruyama, K.; Hirosawa, H.; Nakayoshi, K.; Emura, T.; Sumi, Y.

    1996-01-01

    A compact data-acquisition system using three RISC/UNIX TM workstations (SUN TM /SPARCstation TM ) with real-time capabilities of monitoring and analysis has been developed for the study of photonuclear reactions with the large-acceptance spectrometer TAGX. One workstation acquires data from memory modules in the front-end electronics (CAMAC and TKO) with a maximum speed of 300 Kbytes/s, where data size times instantaneous rate is 1 Kbyte x 300 Hz. Another workstation, which has real-time capability for run monitoring, gets the data with a buffer manager called NOVA. The third workstation analyzes the data and reconstructs the event. In addition to a general hardware and software description, priority settings and run control by shell scripts are described. This system has recently been used successfully in a two month long experiment. (orig.)

  20. Use of quantitative real-time PCR for direct detection of serratia marcescens in marine and other aquatic environments.

    Science.gov (United States)

    Joyner, Jessica; Wanless, David; Sinigalliano, Christopher D; Lipp, Erin K

    2014-03-01

    Serratia marcescens is the etiological agent of acroporid serratiosis, a distinct form of white pox disease in the threatened coral Acropora palmata. The pathogen is commonly found in untreated human waste in the Florida Keys, which may contaminate both nearshore and offshore waters. Currently there is no direct method for detection of this bacterium in the aquatic or reef environment, and culture-based techniques may underestimate its abundance in marine waters. A quantitative real-time PCR assay was developed to detect S. marcescens directly from environmental samples, including marine water, coral mucus, sponge tissue, and wastewater. The assay targeted the luxS gene and was able to distinguish S. marcescens from other Serratia species with a reliable quantitative limit of detection of 10 cell equivalents (CE) per reaction. The method could routinely discern the presence of S. marcescens for as few as 3 CE per reaction, but it could not be reliably quantified at this level. The assay detected environmental S. marcescens in complex sewage influent samples at up to 761 CE ml(-1) and in septic system-impacted residential canals in the Florida Keys at up to 4.1 CE ml(-1). This detection assay provided rapid quantitative abilities and good sensitivity and specificity, which should offer an important tool for monitoring this ubiquitous pathogen that can potentially impact both human health and coral health.

  1. Comparison between culture and a multiplex quantitative real-time polymerase chain reaction assay detecting Ureaplasma urealyticum and U. parvum.

    Science.gov (United States)

    Frølund, Maria; Björnelius, Eva; Lidbrink, Peter; Ahrens, Peter; Jensen, Jørgen Skov

    2014-01-01

    A novel multiplex quantitative real-time polymerase chain reaction (qPCR) for simultaneous detection of U. urealyticum and U. parvum was developed and compared with quantitative culture in Shepard's 10 C medium for ureaplasmas in urethral swabs from 129 men and 66 women, and cervical swabs from 61 women. Using culture as the gold standard, the sensitivity of the qPCR was 96% and 95% for female urethral and cervical swabs, respectively. In male urethral swabs the sensitivity was 89%. The corresponding specificities were 100%, 87% and 99%. The qPCR showed a linear increasing DNA copy number with increasing colour-changing units. Although slightly less sensitive than culture, this multiplex qPCR assay detecting U. urealyticum and U. parvum constitutes a simple and fast alternative to the traditional methods for identification of ureaplasmas and allows simultaneous species differentiation and quantitation in clinical samples. Furthermore, specimens overgrown by other bacteria using the culture method can be evaluated in the qPCR.

  2. Comparison between culture and a multiplex quantitative real-time polymerase chain reaction assay detecting Ureaplasma urealyticum and U. parvum.

    Directory of Open Access Journals (Sweden)

    Maria Frølund

    Full Text Available A novel multiplex quantitative real-time polymerase chain reaction (qPCR for simultaneous detection of U. urealyticum and U. parvum was developed and compared with quantitative culture in Shepard's 10 C medium for ureaplasmas in urethral swabs from 129 men and 66 women, and cervical swabs from 61 women. Using culture as the gold standard, the sensitivity of the qPCR was 96% and 95% for female urethral and cervical swabs, respectively. In male urethral swabs the sensitivity was 89%. The corresponding specificities were 100%, 87% and 99%. The qPCR showed a linear increasing DNA copy number with increasing colour-changing units. Although slightly less sensitive than culture, this multiplex qPCR assay detecting U. urealyticum and U. parvum constitutes a simple and fast alternative to the traditional methods for identification of ureaplasmas and allows simultaneous species differentiation and quantitation in clinical samples. Furthermore, specimens overgrown by other bacteria using the culture method can be evaluated in the qPCR.

  3. Assessment of Multiple GNSS Real-Time SSR Products from Different Analysis Centers

    Directory of Open Access Journals (Sweden)

    Zhiyu Wang

    2018-03-01

    Full Text Available The real-time State Space Representation (SSR product of the GNSS (Global Navigation Satellite System orbit and clock is one of the most essential corrections for real-time precise point positioning (PPP. In this work, the performance of current SSR products from eight analysis centers were assessed by comparing it with the final product and the accuracy of real-time PPP. Numerical results showed that (1 the accuracies of the GPS SSR product were better than 8 cm for the satellite orbit and 0.3 ns for the satellite clock; (2 the accuracies of the GLONASS (GLObalnaya NAvigatsionnaya Sputnikovaya Sistema SSR product were better than 10 cm for orbit RMS (Root Mean Square and 0.6 ns for clock STD (Standard Deviation; and (3 the accuracies of the BDS (BeiDou Navigation Satellite System and Galileo SSR products from CLK93 were about 14.54 and 4.42 cm for the orbit RMS and 0.32 and 0.18 ns for the clock STD, respectively. The simulated kinematic PPP results obtained using the SSR products from CLK93 and CLK51 performed better than those using other SSR products; and the accuracy of PPP based on all products was better than 6 and 10 cm in the horizontal and vertical directions, respectively. The real-time kinematic PPP experiment carried out in Beijing, Tianjin, and Shijiazhuang, China indicated that the SSR product CLK93 from Centre National d’Etudes Spatiales (CNES had a better performance than CAS01. Moreover, the PPP with GPS + BDS dual systems had a higher accuracy than those with only a GPS single system.

  4. Combining real-time monitoring and knowledge-based analysis in MARVEL

    Science.gov (United States)

    Schwuttke, Ursula M.; Quan, A. G.; Angelino, R.; Veregge, J. R.

    1993-01-01

    Real-time artificial intelligence is gaining increasing attention for applications in which conventional software methods are unable to meet technology needs. One such application area is the monitoring and analysis of complex systems. MARVEL, a distributed monitoring and analysis tool with multiple expert systems, was developed and successfully applied to the automation of interplanetary spacecraft operations at NASA's Jet Propulsion Laboratory. MARVEL implementation and verification approaches, the MARVEL architecture, and the specific benefits that were realized by using MARVEL in operations are described.

  5. Analysis of writing inks on paper using direct analysis in real time mass spectrometry.

    Science.gov (United States)

    Jones, Roger W; McClelland, John F

    2013-09-10

    Ink analysis is central to questioned document examination. We applied direct analysis in real time mass spectrometry (DART MS) to ballpoint, gel, and fluid writing ink analysis. DART MS acquires the mass spectrum of an ink while it is still on a document without altering the appearance of the document. Spectra were acquired from ink on a variety of papers, and the spectrum of the blank paper could be subtracted out to produce a cleanly isolated ink spectrum in most cases. Only certain heavy or heavily processed papers interfered. The time since an ink is written on paper has a large effect on its spectrum. DART spectra change radically during the first few months after an ink is written as the more volatile components evaporate, but the spectra stabilize after that. A library-search study involving 166 well-aged inks assessed the ability to identify inks from their DART spectra. The aggregate success rate was 92%. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  6. Quantitative Determination of Plasmodium vivax Gametocytes by Real-Time Quantitative Nucleic Acid Sequence-Based Amplification in Clinical Samples

    NARCIS (Netherlands)

    Beurskens, Martijn; Mens, Pètra; Schallig, Henk; Syafruddin, Din; Asih, Puji Budi Setia; Hermsen, Rob; Sauerwein, Robert

    2009-01-01

    Microscopic detection of Plasmodium vivax gametocytes, the sexual life stage of this malaria parasite, is insensitive because P vivax parasitaemia is low. To detect and quantity gametocytes a more sensitive, quantitative realtime Pvs25-QT-NASBA based oil Pvs25 mRNA was developed and tested in two

  7. Applying real-time quantitative PCR to diagnosis of freemartin in Holstein cattle by quantifying SRY gene: a comparison experiment

    Directory of Open Access Journals (Sweden)

    Qinghua Qiu

    2018-04-01

    Full Text Available Background Freemartinism generally occurs in female offspring of dizygotic twins in a mixed-sex pregnancy. Most bovine heterosexual twin females are freemartins. However, about 10% of bovine heterosexual twin females are fertile. Farmers mostly cull bovine fertile heterosexual twin females due to the lack of a practical diagnostic approach. Culling of such animals results in economic and genetic-material losses both for dairy and beef industry. Methods In this study, a comparative test, including qualitative detection of SRY gene by polymerase chain reaction (PCR, quantitative detection of relative content of SRY by real-time quantitative polymerase chain reaction (qPCR, and quantitative detection of H-Y antigen, was performed to establish the most accurate diagnosis for freemartin. Twelve Holstein heterosexual twin females were used in this study, while three normal Holstein bulls and three normal Holstein cows were used as a positive and negative control, respectively. Results Polymerase chain reaction results revealed that SRY gene were absent in three heterosexual twin females and only two of them were verified as fertile in later age. The qPCR results showed that relative content of SRY was more than 14.2% in freemartins and below 0.41% in fertile heterosexual twin females. The H-Y antigen test showed no significant numerical difference between freemartin and fertile heterosexual twin female. Discussion Our results show that relative content of SRY quantified by qPCR is a better detection method for diagnosis of freemartin in Holstein cattle as compare to qualitative detection of SRY gene by PCR or quantitative detection of H-Y antigen. To the authors’ knowledge, this is the first time we applied qPCR to diagnosing freemartin by quantifying SRY gene and got relative SRY content of each freemartin and fertile heterosexual twin female. We concluded that low-level of SRY would not influence fertility of bovine heterosexual twin female.

  8. Micromagnetic Cancer Cell Immobilization and Release for Real-Time Single Cell Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Jaiswal, Devina; Rad, Armin Tahmasbi [Department of Biomedical Engineering, University of Connecticut, Storrs, CT, 06269 (United States); Nieh, Mu-Ping [Department of Biomedical Engineering, University of Connecticut, Storrs, CT, 06269 (United States); Department of Chemical and Biomolecular Engineering, University of Connecticut, Storrs, CT 06269 (United States); Polymer Program, Institute of Materials Science, University of Connecticut, Storrs, CT 06269 (United States); Claffey, Kevin P. [Department of Cell Biology, University of Connecticut Health Center, Farmington, CT 06030 (United States); Hoshino, Kazunori, E-mail: hoshino@engr.uconn.edu [Department of Biomedical Engineering, University of Connecticut, Storrs, CT, 06269 (United States)

    2017-04-01

    Understanding the interaction of live cells with macromolecules is crucial for designing efficient therapies. Considering the functional heterogeneity found in cancer cells, real-time single cell analysis is necessary to characterize responses. In this study, we have designed and fabricated a microfluidic channel with patterned micromagnets which can temporarily immobilize the cells during analysis and release them after measurements. The microchannel is composed of plain coverslip top and bottom panels to facilitate easy microscopic observation and undisturbed application of analytes to the cells. Cells labeled with functionalized magnetic beads were immobilized in the device with an efficiency of 90.8±3.6%. Since the micromagnets are made of soft magnetic material (Ni), they released cells when external magnetic field was turned off from the channel. This allows the reuse of the channel for a new sample. As a model drug analysis, the immobilized breast cancer cells (MCF7) were exposed to fluorescent lipid nanoparticles and association and dissociation were measured through fluorescence analysis. Two concentrations of nanoparticles, 0.06 µg/ml and 0.08 µg/ml were tested and time lapse images were recorded and analyzed. The microfluidic device was able to provide a microenvironment for sample analysis, making it an efficient platform for real-time analysis.

  9. A two-step real-time PCR assay for quantitation and genotyping of human parvovirus 4.

    Science.gov (United States)

    Väisänen, E; Lahtinen, A; Eis-Hübinger, A M; Lappalainen, M; Hedman, K; Söderlund-Venermo, M

    2014-01-01

    Human parvovirus 4 (PARV4) of the family Parvoviridae was discovered in a plasma sample of a patient with an undiagnosed acute infection in 2005. Currently, three PARV4 genotypes have been identified, however, with an unknown clinical significance. Interestingly, these genotypes seem to differ in epidemiology. In Northern Europe, USA and Asia, genotypes 1 and 2 have been found to occur mainly in persons with a history of injecting drug use or other parenteral exposure. In contrast, genotype 3 appears to be endemic in sub-Saharan Africa, where it infects children and adults without such risk behaviour. In this study, a novel straightforward and cost-efficient molecular assay for both quantitation and genotyping of PARV4 DNA was developed. The two-step method first applies a single-probe pan-PARV4 qPCR for screening and quantitation of this relatively rare virus, and subsequently, only the positive samples undergo a real-time PCR-based multi-probe genotyping. The new qPCR-GT method is highly sensitive and specific regardless of the genotype, and thus being suitable for studying the clinical impact and occurrence of the different PARV4 genotypes. Copyright © 2013 Elsevier B.V. All rights reserved.

  10. Evaluation of changes in periodontal bacteria in healthy dogs over 6 months using quantitative real-time PCR.

    Science.gov (United States)

    Maruyama, N; Mori, A; Shono, S; Oda, H; Sako, T

    2018-03-01

    Porphyromonas gulae, Tannerella forsythia and Campylobacter rectus are considered dominant periodontal pathogens in dogs. Recently, quantitative real-time PCR (qRT-PCR) methods have been used for absolute quantitative determination of oral bacterial counts. The purpose of the present study was to establish a standardized qRT-PCR procedure to quantify bacterial counts of the three target periodontal bacteria (P. gulae, T. forsythia and C. rectus). Copy numbers of the three target periodontal bacteria were evaluated in 26 healthy dogs. Then, changes in bacterial counts of the three target periodontal bacteria were evaluated for 24 weeks in 7 healthy dogs after periodontal scaling. Analytical evaluation of each self-designed primer indicated acceptable analytical imprecision. All 26 healthy dogs were found to be positive for P. gulae, T. forsythia and C. rectus. Median total bacterial counts (copies/ng) of each target genes were 385.612 for P. gulae, 25.109 for T. forsythia and 5.771 for C. rectus. Significant differences were observed between the copy numbers of the three target periodontal bacteria. Periodontal scaling reduced median copy numbers of the three target periodontal bacteria in 7 healthy dogs. However, after periodontal scaling, copy numbers of all three periodontal bacteria significantly increased over time (pperiodontal bacteria in dogs. Furthermore, the present study has revealed that qRT-PCR method can be considered as a new objective evaluation system for canine periodontal disease. Copyright© by the Polish Academy of Sciences.

  11. A multiplex calibrated real-time PCR assay for quantitation of DNA of EBV-1 and 2.

    Science.gov (United States)

    Gatto, Francesca; Cassina, Giulia; Broccolo, Francesco; Morreale, Giuseppe; Lanino, Edoardo; Di Marco, Eddi; Vardas, Efthiya; Bernasconi, Daniela; Buttò, Stefano; Principi, Nicola; Esposito, Susanna; Scarlatti, Gabriella; Lusso, Paolo; Malnati, Mauro S

    2011-12-01

    Accurate and highly sensitive tests for the diagnosis of active Epstein-Barr virus (EBV) infection are essential for the clinical management of individuals infected with EBV. A calibrated quantitative real-time PCR assay for the measurement of EBV DNA of both EBV-1 and 2 subtypes was developed, combining the detection of the EBV DNA and a synthetic DNA calibrator in a multiplex PCR format. The assay displays a wide dynamic range and a high degree of accuracy even in the presence of 1μg of human genomic DNA. This assay measures with the same efficiency EBV DNA from strains prevalent in different geographic areas. The clinical sensitivity and specificity of the system were evaluated by testing 181 peripheral blood mononuclear cell (PBMCs) and plasma specimens obtained from 21 patients subjected to bone marrow transplantation, 70 HIV-seropositive subjects and 23 healthy controls. Patients affected by EBV-associated post-transplant lymphoprolipherative disorders had the highest frequency of EBV detection and the highest viral load. Persons infected with HIV had higher levels of EBV DNA load in PBMCs and a higher frequency of EBV plasma viremia compared to healthy controls. In conclusion, this new assay provides a reliable high-throughput method for the quantitation of EBV DNA in clinical samples. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Quantitative real-time PCR approaches for microbial community studies in wastewater treatment systems: applications and considerations.

    Science.gov (United States)

    Kim, Jaai; Lim, Juntaek; Lee, Changsoo

    2013-12-01

    Quantitative real-time PCR (qPCR) has been widely used in recent environmental microbial ecology studies as a tool for detecting and quantifying microorganisms of interest, which aids in better understandings of the complexity of wastewater microbial communities. Although qPCR can be used to provide more specific and accurate quantification than other molecular techniques, it does have limitations that must be considered when applying it in practice. This article reviews the principle of qPCR quantification and its applications to microbial ecology studies in various wastewater treatment environments. Here we also address several limitations of qPCR-based approaches that can affect the validity of quantification data: template nucleic acid quality, nucleic acid extraction efficiency, specificity of group-specific primers and probes, amplification of nonviable DNA, gene copy number variation, and limited number of sequences in the database. Even with such limitations, qPCR is reportedly among the best methods for quantitatively investigating environmental microbial communities. The application of qPCR is and will continue to be increasingly common in studies of wastewater treatment systems. To obtain reliable analyses, however, the limitations that have often been overlooked must be carefully considered when interpreting the results. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. [Comparison of two quantitative methods of endobronchial ultrasound real-time elastography for evaluating intrathoracic lymph nodes].

    Science.gov (United States)

    Mao, X W; Yang, J Y; Zheng, X X; Wang, L; Zhu, L; Li, Y; Xiong, H K; Sun, J Y

    2017-06-12

    Objective: To compare the clinical value of two quantitative methods in analyzing endobronchial ultrasound real-time elastography (EBUS-RTE) images for evaluating intrathoracic lymph nodes. Methods: From January 2014 to April 2014, EBUS-RTE examination was performed in patients who received EBUS-TBNA examination in Shanghai Chest Hospital. Each intrathoracic lymph node had a selected EBUS-RTE image. Stiff area ratio and mean hue value of region of interest (ROI) in each image were calculated respectively. The final diagnosis of lymph node was based on the pathologic/microbiologic results of EBUS-TBNA, pathologic/microbiologic results of other examinations and clinical following-up. The sensitivity, specificity, positive predictive value, negative predictive value and accuracy were evaluated for distinguishing malignant and benign lesions. Results: Fifty-six patients and 68 lymph nodes were enrolled in this study, of which 35 lymph nodes were malignant and 33 lymph nodes were benign. The stiff area ratio and mean hue value of benign and malignant lesions were 0.32±0.29, 0.62±0.20 and 109.99±28.13, 141.62±17.52, respectively, and statistical differences were found in both of those two methods ( t =-5.14, P methods can be used for analyzing EBUS-RTE images quantitatively, having the value of differentiating benign and malignant intrathoracic lymph nodes, and the stiff area ratio is better than the mean hue value between the two methods.

  14. Development and application of an oligonucleotide microarray and real-time quantitative PCR for detection of wastewater bacterial pathogens

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Dae-Young [National Water Research Institute, Environment Canada, 867 Lakeshore Road, Burlington, Ontario, L7R 4A6 (Canada)], E-mail: daeyoung.lee@ec.gc.ca; Lauder, Heather; Cruwys, Heather; Falletta, Patricia [National Water Research Institute, Environment Canada, 867 Lakeshore Road, Burlington, Ontario, L7R 4A6 (Canada); Beaudette, Lee A. [Environmental Science and Technology Centre, Environment Canada, 335 River Road South, Ottawa, Ontario, K1A 0H3 (Canada)], E-mail: lee.beaudette@ec.gc.ca

    2008-07-15

    Conventional microbial water quality test methods are well known for their technical limitations, such as lack of direct pathogen detection capacity and low throughput capability. The microarray assay has recently emerged as a promising alternative for environmental pathogen monitoring. In this study, bacterial pathogens were detected in municipal wastewater using a microarray equipped with short oligonucleotide probes targeting 16S rRNA sequences. To date, 62 probes have been designed against 38 species, 4 genera, and 1 family of pathogens. The detection sensitivity of the microarray for a waterborne pathogen Aeromonas hydrophila was determined to be approximately 1.0% of the total DNA, or approximately 10{sup 3}A. hydrophila cells per sample. The efficacy of the DNA microarray was verified in a parallel study where pathogen genes and E. coli cells were enumerated using real-time quantitative PCR (qPCR) and standard membrane filter techniques, respectively. The microarray and qPCR successfully detected multiple wastewater pathogen species at different stages of the disinfection process (i.e. secondary effluents vs. disinfected final effluents) and at two treatment plants employing different disinfection methods (i.e. chlorination vs. UV irradiation). This result demonstrates the effectiveness of the DNA microarray as a semi-quantitative, high throughput pathogen monitoring tool for municipal wastewater.

  15. A quantitative real time polymerase chain reaction approach for estimating processed animal proteins in feed: preliminary data

    Directory of Open Access Journals (Sweden)

    Maria Cesarina Abete

    2013-04-01

    Full Text Available Lifting of the ban on the use of processed animal proteins (PAPs from non-ruminants in non-ruminant feed is in the wind, avoiding intraspecies recycling. Discrimination of species will be performed through polymerase chain reaction (PCR, which is at a moment a merely qualitative method. Nevertheless, quantification of PAPs in feed is needed. The aim of this study was to approach the quantitative determination of PAPs in feed through Real Time (RT-PCR technique; three different protocols picked up from the literature were tested. Three different kind of matrices were examined: pure animal meals (bovine, chicken and pork; one feed sample certified by the European reference laboratory on animal proteins (EURL AP in feed spiked with 0.1% bovine meal; and genomic DNAs from bovine, chicken and pork muscles. The limit of detection (LOD of the three protocols was set up. All the results obtained from the three protocols considered failed in the quantification process, most likely due to the uncertain copy numbers of the analytical targets chosen. This preliminary study will allow us to address further investigations, with the purpose of developing a RT-PCR quantitative method.

  16. RFLP Analysis and Allelic Discrimination with Real-Time PCR Using the Human Lactase Persistence Trait: A Pair of Molecular Genetic Investigations

    Science.gov (United States)

    Weinlander, Kenneth M.; Hall, David J.; De Stasio, Elizabeth A.

    2010-01-01

    We describe here two open-ended laboratory investigations for an undergraduate laboratory course that uses students' DNA as templates for quantitative real-time PCR and for traditional PCR followed by RFLP analysis. Students are captivated by the immediacy of the application and the relevance of the genotypes and traits, lactase persistence or…

  17. Real time video processing software for the analysis of endoscopic guided-biopsies

    International Nuclear Information System (INIS)

    Ordoñez, C; Bouchet, A; Pastore, J; Blotta, E

    2011-01-01

    The severity in Barrett esophagus disease is, undoubtedly, the possibility of its malignization. To make an early diagnosis in order to avoid possible complications, it is absolutely necessary collect biopsies to make a histological analysis. This should be done under endoscopic control to avoid mucus areas that may co-exist within the columnar epithelial, which could lead to a false diagnosis. This paper presents a video processing software in real-time in order to delineate and enhance areas of interest to facilitate the work of the expert.

  18. Method for Hot Real-Time Analysis of Pyrolysis Vapors at Pilot Scale

    Energy Technology Data Exchange (ETDEWEB)

    Pomeroy, Marc D [National Renewable Energy Laboratory (NREL), Golden, CO (United States)

    2017-09-29

    Pyrolysis oils contain more than 400 compounds, up to 60% of which do not re-volatilize for subsequent chemical analysis. Vapor chemical composition is also complicated as additional condensation reactions occur during quenching and collection of the product. Due to the complexity of the pyrolysis oil, and a desire to catalytically upgrade the vapor composition before condensation, online real-time analytical techniques such as Molecular Beam Mass Spectrometry (MBMS) are of great use. However, in order to properly sample hot pyrolysis vapors at the pilot scale, many challenges must be overcome.

  19. Optically selected GRB afterglows, a real time analysis system at the CFHT

    International Nuclear Information System (INIS)

    Malacrino, F.; Atteia, J.-L.; Klotz, A.; Boer, M.; Kavelaars, J.J.; Cuillandre, J.-C.

    2005-01-01

    We attempt to detect optical GRB afterglows on images taken by the Canada France Hawaii Telescope for the Very Wide survey, component of the Legacy Survey. To do so, a Real Time Analysis System called Optically Selected GRB Afterglows has been installed on a dedicated computer in Hawaii. This pipeline automatically and quickly analyzes Mega cam images and extracts from them a list of variable objects which is displayed on a web page far validation by a member of the collaboration. The Very Wide survey covers 1200 square degrees down to i 1 = 23.5. This paper briefly explain the RTAS process

  20. Real-time data analysis at the LHC: present and future

    CERN Document Server

    Gligorov, V.V.

    2015-01-01

    The Large Hadron Collider (LHC), which collides protons at an energy of 14 TeV, produces hundreds of exabytes of data per year, making it one of the largest sources of data in the world today. At present it is not possible to even transfer most of this data from the four main particle detectors at the LHC to "offline" data facilities, much less to permanently store it for future processing. For this reason the LHC detectors are equipped with real-time analysis systems, called triggers, which process this volume of data and select the most interesting proton-proton collisions. The LHC experiment triggers reduce the data produced by the LHC by between 1/1000 and 1/100000, to tens of petabytes per year, allowing its economical storage and further analysis. The bulk of the data-reduction is performed by custom electronics which ignores most of the data in its decision making, and is therefore unable to exploit the most powerful known data analysis strategies. I cover the present status of real-time data analysis ...

  1. Real-Time Earthquake Intensity Estimation Using Streaming Data Analysis of Social and Physical Sensors

    Science.gov (United States)

    Kropivnitskaya, Yelena; Tiampo, Kristy F.; Qin, Jinhui; Bauer, Michael A.

    2017-06-01

    Earthquake intensity is one of the key components of the decision-making process for disaster response and emergency services. Accurate and rapid intensity calculations can help to reduce total loss and the number of casualties after an earthquake. Modern intensity assessment procedures handle a variety of information sources, which can be divided into two main categories. The first type of data is that derived from physical sensors, such as seismographs and accelerometers, while the second type consists of data obtained from social sensors, such as witness observations of the consequences of the earthquake itself. Estimation approaches using additional data sources or that combine sources from both data types tend to increase intensity uncertainty due to human factors and inadequate procedures for temporal and spatial estimation, resulting in precision errors in both time and space. Here we present a processing approach for the real-time analysis of streams of data from both source types. The physical sensor data is acquired from the U.S. Geological Survey (USGS) seismic network in California and the social sensor data is based on Twitter user observations. First, empirical relationships between tweet rate and observed Modified Mercalli Intensity (MMI) are developed using data from the M6.0 South Napa, CAF earthquake that occurred on August 24, 2014. Second, the streams of both data types are analyzed together in simulated real-time to produce one intensity map. The second implementation is based on IBM InfoSphere Streams, a cloud platform for real-time analytics of big data. To handle large processing workloads for data from various sources, it is deployed and run on a cloud-based cluster of virtual machines. We compare the quality and evolution of intensity maps from different data sources over 10-min time intervals immediately following the earthquake. Results from the joint analysis shows that it provides more complete coverage, with better accuracy and higher

  2. Real time analysis with the upgraded LHCb trigger in Run III

    Science.gov (United States)

    Szumlak, Tomasz

    2017-10-01

    The current LHCb trigger system consists of a hardware level, which reduces the LHC bunch-crossing rate of 40 MHz to 1.1 MHz, a rate at which the entire detector is read out. A second level, implemented in a farm of around 20k parallel processing CPUs, the event rate is reduced to around 12.5 kHz. The LHCb experiment plans a major upgrade of the detector and DAQ system in the LHC long shutdown II (2018-2019). In this upgrade, a purely software based trigger system is being developed and it will have to process the full 30 MHz of bunch crossings with inelastic collisions. LHCb will also receive a factor of 5 increase in the instantaneous luminosity, which further contributes to the challenge of reconstructing and selecting events in real time with the CPU farm. We discuss the plans and progress towards achieving efficient reconstruction and selection with a 30 MHz throughput. Another challenge is to exploit the increased signal rate that results from removing the 1.1 MHz readout bottleneck, combined with the higher instantaneous luminosity. Many charm hadron signals can be recorded at up to 50 times higher rate. LHCb is implementing a new paradigm in the form of real time data analysis, in which abundant signals are recorded in a reduced event format that can be fed directly to the physics analyses. These data do not need any further offline event reconstruction, which allows a larger fraction of the grid computing resources to be devoted to Monte Carlo productions. We discuss how this real-time analysis model is absolutely critical to the LHCb upgrade, and how it will evolve during Run-II.

  3. Near Real-time Scientific Data Analysis and Visualization with the ArcGIS Platform

    Science.gov (United States)

    Shrestha, S. R.; Viswambharan, V.; Doshi, A.

    2017-12-01

    Scientific multidimensional data are generated from a variety of sources and platforms. These datasets are mostly produced by earth observation and/or modeling systems. Agencies like NASA, NOAA, USGS, and ESA produce large volumes of near real-time observation, forecast, and historical data that drives fundamental research and its applications in larger aspects of humanity from basic decision making to disaster response. A common big data challenge for organizations working with multidimensional scientific data and imagery collections is the time and resources required to manage and process such large volumes and varieties of data. The challenge of adopting data driven real-time visualization and analysis, as well as the need to share these large datasets, workflows, and information products to wider and more diverse communities, brings an opportunity to use the ArcGIS platform to handle such demand. In recent years, a significant effort has put in expanding the capabilities of ArcGIS to support multidimensional scientific data across the platform. New capabilities in ArcGIS to support scientific data management, processing, and analysis as well as creating information products from large volumes of data using the image server technology are becoming widely used in earth science and across other domains. We will discuss and share the challenges associated with big data by the geospatial science community and how we have addressed these challenges in the ArcGIS platform. We will share few use cases, such as NOAA High Resolution Refresh Radar (HRRR) data, that demonstrate how we access large collections of near real-time data (that are stored on-premise or on the cloud), disseminate them dynamically, process and analyze them on-the-fly, and serve them to a variety of geospatial applications. We will also share how on-the-fly processing using raster functions capabilities, can be extended to create persisted data and information products using raster analytics

  4. Real-time Position Based Population Data Analysis and Visualization Using Heatmap for Hazard Emergency Response

    Science.gov (United States)

    Ding, R.; He, T.

    2017-12-01

    With the increased popularity in mobile applications and services, there has been a growing demand for more advanced mobile technologies that utilize real-time Location Based Services (LBS) data to support natural hazard response efforts. Compared to traditional sources like the census bureau that often can only provide historical and static data, an LBS service can provide more current data to drive a real-time natural hazard response system to more accurately process and assess issues such as population density in areas impacted by a hazard. However, manually preparing or preprocessing the data to suit the needs of the particular application would be time-consuming. This research aims to implement a population heatmap visual analytics system based on real-time data for natural disaster emergency management. System comprised of a three-layered architecture, including data collection, data processing, and visual analysis layers. Real-time, location-based data meeting certain polymerization conditions are collected from multiple sources across the Internet, then processed and stored in a cloud-based data store. Parallel computing is utilized to provide fast and accurate access to the pre-processed population data based on criteria such as the disaster event and to generate a location-based population heatmap as well as other types of visual digital outputs using auxiliary analysis tools. At present, a prototype system, which geographically covers the entire region of China and combines population heat map based on data from the Earthquake Catalogs database has been developed. It Preliminary results indicate that the generation of dynamic population density heatmaps based on the prototype system has effectively supported rapid earthquake emergency rescue and evacuation efforts as well as helping responders and decision makers to evaluate and assess earthquake damage. Correlation analyses that were conducted revealed that the aggregation and movement of people

  5. Real-time label-free quantitative fluorescence microscopy-based detection of ATP using a tunable fluorescent nano-aptasensor platform.

    Science.gov (United States)

    Shrivastava, Sajal; Sohn, Il-Yung; Son, Young-Min; Lee, Won-Il; Lee, Nae-Eung

    2015-12-14

    Although real-time label-free fluorescent aptasensors based on nanomaterials are increasingly recognized as a useful strategy for the detection of target biomolecules with high fidelity, the lack of an imaging-based quantitative measurement platform limits their implementation with biological samples. Here we introduce an ensemble strategy for a real-time label-free fluorescent graphene (Gr) aptasensor platform. This platform employs aptamer length-dependent tunability, thus enabling the reagentless quantitative detection of biomolecules through computational processing coupled with real-time fluorescence imaging data. We demonstrate that this strategy effectively delivers dose-dependent quantitative readouts of adenosine triphosphate (ATP) concentration on chemical vapor deposited (CVD) Gr and reduced graphene oxide (rGO) surfaces, thereby providing cytotoxicity assessment. Compared with conventional fluorescence spectrometry methods, our highly efficient, universally applicable, and rational approach will facilitate broader implementation of imaging-based biosensing platforms for the quantitative evaluation of a range of target molecules.

  6. Real-time breath analysis with active capillary plasma ionization-ambient mass spectrometry.

    Science.gov (United States)

    Bregy, Lukas; Sinues, Pablo Martinez-Lozano; Nudnova, Maryia M; Zenobi, Renato

    2014-06-01

    On-line analysis of exhaled human breath is a growing area in analytical science, for applications such as fast and non-invasive medical diagnosis and monitoring. In this work, we present a novel approach based on ambient ionization of compounds in breath and subsequent real-time mass spectrometric analysis. We introduce a plasma ionization source for this purpose, which has no need for additional gases, is very small, and is easily interfaced with virtually any commercial atmospheric pressure ionization mass spectrometer (API-MS) without major modifications. If an API-MS instrument exists in a laboratory, the cost to implement this technology is only around [Formula: see text]500, far less than the investment for a specialized mass spectrometric system designed for volatile organic compounds (VOCs) analysis. In this proof-of-principle study we were able to measure mass spectra of exhaled human breath and found these to be comparable to spectra obtained with other electrospray-based methods. We detected over 100 VOCs, including relevant metabolites like fatty acids, with molecular weights extending up to 340 Da. In addition, we were able to monitor the time-dependent evolution of the peaks and show the enhancement of the metabolism after a meal. We conclude that this approach may complement current methods to analyze breath or other types of vapors, offering an affordable option to upgrade any pre-existing API-MS to a real-time breath analyzer.

  7. Impact Analysis of Land Use on Traffic Congestion Using Real-Time Traffic and POI

    Directory of Open Access Journals (Sweden)

    Tianqi Zhang

    2017-01-01

    Full Text Available This paper proposed a new method to describe, compare, and classify the traffic congestion points in Beijing, China, by using the online map data and further revealed the relationship between traffic congestion and land use. The data of the point of interest (POI and the real-time traffic was extracted from an electronic map of the area in the fourth ring road of Beijing. The POIs were quantified based on the architectural area of the land use; the congestion points were identified based on real-time traffic. Then, the cluster analysis using the attributes of congestion time was conducted to identify the main traffic congestion areas. The result of a linear regression analysis between the congestion time and the land use showed that the influence of the high proportion of commercial land use on the traffic congestion was significant. Also, we considered five types of land use through performing a linear regression analysis between the congestion time and the ratio of four types of land use. The results showed that the reasonable ratio of land use types could efficiently reduce congestion time. This study makes contributions to the policy-making of urban land use.

  8. Real-time label-free quantitative fluorescence microscopy-based detection of ATP using a tunable fluorescent nano-aptasensor platform

    Science.gov (United States)

    Shrivastava, Sajal; Sohn, Il-Yung; Son, Young-Min; Lee, Won-Il; Lee, Nae-Eung

    2015-11-01

    Although real-time label-free fluorescent aptasensors based on nanomaterials are increasingly recognized as a useful strategy for the detection of target biomolecules with high fidelity, the lack of an imaging-based quantitative measurement platform limits their implementation with biological samples. Here we introduce an ensemble strategy for a real-time label-free fluorescent graphene (Gr) aptasensor platform. This platform employs aptamer length-dependent tunability, thus enabling the reagentless quantitative detection of biomolecules through computational processing coupled with real-time fluorescence imaging data. We demonstrate that this strategy effectively delivers dose-dependent quantitative readouts of adenosine triphosphate (ATP) concentration on chemical vapor deposited (CVD) Gr and reduced graphene oxide (rGO) surfaces, thereby providing cytotoxicity assessment. Compared with conventional fluorescence spectrometry methods, our highly efficient, universally applicable, and rational approach will facilitate broader implementation of imaging-based biosensing platforms for the quantitative evaluation of a range of target molecules.Although real-time label-free fluorescent aptasensors based on nanomaterials are increasingly recognized as a useful strategy for the detection of target biomolecules with high fidelity, the lack of an imaging-based quantitative measurement platform limits their implementation with biological samples. Here we introduce an ensemble strategy for a real-time label-free fluorescent graphene (Gr) aptasensor platform. This platform employs aptamer length-dependent tunability, thus enabling the reagentless quantitative detection of biomolecules through computational processing coupled with real-time fluorescence imaging data. We demonstrate that this strategy effectively delivers dose-dependent quantitative readouts of adenosine triphosphate (ATP) concentration on chemical vapor deposited (CVD) Gr and reduced graphene oxide (r

  9. Twitter data analysis: temporal and term frequency analysis with real-time event

    Science.gov (United States)

    Yadav, Garima; Joshi, Mansi; Sasikala, R.

    2017-11-01

    From the past few years, World Wide Web (www) has become a prominent and huge source for user generated content and opinionative data. Among various social media, Twitter gained popularity as it offers a fast and effective way of sharing users’ perspective towards various critical and other issues in different domain. As the data is hugely generated on cloud, it has opened doors for the researchers in the field of data science and analysis. There are various domains such as ‘Political’ domain, ‘Entertainment’ domain and ‘Business’ domain. Also there are various APIs that Twitter provides for developers 1) Search API, focus on the old tweets 2) Rest API, focuses on user details and allow to collect the user profile, friends and followers 3) Streaming API, which collects details like tweets, hashtags, geo locations. In our work we are accessing Streaming API in order to fetch real-time tweets for the dynamic happening event. For this we are focusing on ‘Entertainment’ domain especially ‘Sports’ as IPL-T20 is currently the trending on-going event. We are collecting these numerous amounts of tweets and storing them in MongoDB database where the tweets are stored in JSON document format. On this document we are performing time-series analysis and term frequency analysis using different techniques such as filtering, information extraction for text-mining that fulfils our objective of finding interesting moments for temporal data in the event and finding the ranking among the players or the teams based on popularity which helps people in understanding key influencers on the social media platform.

  10. Direct analysis in real time mass spectrometry for analysis of sexual assault evidence.

    Science.gov (United States)

    Musah, Rabi A; Cody, Robert B; Dane, A John; Vuong, Angela L; Shepard, Jason R E

    2012-05-15

    Sexual assault crimes are vastly underreported and suffer from alarmingly low prosecution and conviction rates. The key scientific method to aid in prosecution of such cases is forensic DNA analysis, where biological evidence such as semen collected using a rape test kit is used to determine a suspect's DNA profile. However, the growing awareness by criminals of the importance of DNA in the prosecution of sexual assaults has resulted in increased condom use by assailants as a means to avoid leaving behind their DNA. Thus, other types of trace evidence are important to help corroborate victims' accounts, exonerate the innocent, link suspects to the crime, or confirm penetration. Direct Analysis in Real Time Mass Spectrometry (DART-MS) was employed for the comprehensive characterization of non-DNA trace evidence associated with sexual assault. The ambient ionization method associated with DART-MS is extremely rapid and samples are processed instantaneously, without the need for extraction, sample preparation, or other means that might compromise forensic evidence for future analyses. In a single assay, we demonstrated the ability to identify lubricant formulations associated with sexual assault, such as the spermicide nonoxynol-9, compounds used in condom manufacture, and numerous other trace components as probative evidence. In addition, the method can also serve to identify compounds within trace biological residues, such as fatty acids commonly identified in latent fingerprints. Characterization of lubricant residues as probative evidence serves to establish a connection between the victim and the perpetrator, and the availability of these details may lead to higher rates of prosecution and conviction, as well as more severe penalties. The methodology described here opens the way for the adoption of a comprehensive, rapid, and sensitive analysis for use in crime labs, while providing knowledge that can inform and guide criminal justice policy and practice

  11. Comparison of commercial DNA preparation kits for the detection of Brucellae in tissue using quantitative real-time PCR

    Directory of Open Access Journals (Sweden)

    Straube Eberhard

    2010-04-01

    Full Text Available Abstract Background The detection of Brucellae in tissue specimens using PCR assays is difficult because the amount of bacteria is usually low. Therefore, optimised DNA extraction methods are critical. The aim of this study was to assess the performance of commercial kits for the extraction of Brucella DNA. Methods Five kits were evaluated using clinical specimens: QIAamp™ DNA Mini Kit (QIAGEN, peqGold™ Tissue DNA Mini Kit (PeqLab, UltraClean™ Tissue and Cells DNA Isolation Kit (MoBio, DNA Isolation Kit for Cells and Tissues (Roche, and NucleoSpin™ Tissue (Macherey-Nagel. DNA yield was determined using a quantitative real-time PCR assay targeting IS711 that included an internal amplification control. Results Kits of QIAGEN and Roche provided the highest amount of DNA, Macherey-Nagel and Peqlab products were intermediate whereas MoBio yielded the lowest amount of DNA. Differences were significant (p Conclusions We observed differences in DNA yield as high as two orders of magnitude for some samples between the best and the worst DNA extraction kits and inhibition was observed occasionally. This indicates that DNA purification may be more relevant than expected when the amount of DNA in tissue is very low.

  12. Quantitative real-time PCR technique for the identification of E. coli residual DNA in streptokinase recombinant product.

    Science.gov (United States)

    Fazelahi, Mansoureh; Kia, Vahid; Kaghazian, Hooman; Paryan, Mahdi

    2017-11-26

    Recombinant streptokinase is a biopharmaceutical which is usually produced in E. coli. Residual DNA as a contamination and risk factor may remain in the product. It is necessary to control the production procedure to exclude any possible contamination. The aim of the present study was to develop a highly specific and sensitive quantitative real-time PCR-based method to determine the amount of E. coli DNA in recombinant streptokinase. A specific primers and a probe was designed to detect all strains of E. coli. To determine the specificity, in addition to using NCBI BLASTn, 28 samples including human, bacterial, and viral genomes were used. The results confirmed that the assay detects no genomic DNA but E. coli's and the specificity was determined to be 100%. To determine the sensitivity and limit of detection of the assay, a 10-fold serial dilution (10 1 to 10 7 copies/µL) was tested in triplicate. The sensitivity of the test was determined to be 101 copies/µL or 35 fg/µL. Inter-assay and intra-assay were determined to be 0.86 and 1.69%, respectively. Based on the results, this assay can be used as an accurate method to evaluate the contamination of recombinant streptokinase in E. coli.

  13. Development of real-time quantitative polymerase chain reaction assays to track treatment response in retinoid resistant acute promyelocytic leukemia

    Directory of Open Access Journals (Sweden)

    Jelena V Jovanovic

    2011-10-01

    Full Text Available Molecular detection of minimal residual disease (MRD has become established to assess remission status and guide therapy in patients with PML-RARA+ acute promyelocytic leukemia (APL. However, there are few data on tracking disease response in patients with rarer retinoid resistant subtypes of APL, characterized by PLZF-RARA and STAT5b-RARA. Despite their relative rarity (<1% of APL we identified 6 cases (PLZF-RARA, n=5; STAT5b-RARA, n=1, established the respective breakpoint junction regions and designed real-time quantitative polymerase chain reaction (RQ-PCR assays to detect leukemic transcripts. The relative level of fusion gene expression in diagnostic samples was comparable to that observed in t(15;17-associated APL, affording assay sensitivities of ~1 in 104-105. Serial samples were available from 2 PLZF-RARA APL patients. One showed persistent PCR positivity, predicting subsequent relapse, and remains in CR2, ~11 years post-autograft. The other, achieved molecular remission (CRm with combination chemotherapy, remaining in CR1 at 6 years. The STAT5b-RARA patient failed to achieve CRm following frontline combination chemotherapy and ultimately proceeded to allogeneic transplant on the basis of a steadily rising fusion transcript level. These data highlight the potential of RQ-PCR detection of MRD to facilitate development of more individualized approaches to the management of rarer molecularly-defined subsets of acute leukemia.

  14. Accurate, Fast and Cost-Effective Diagnostic Test for Monosomy 1p36 Using Real-Time Quantitative PCR

    Directory of Open Access Journals (Sweden)

    Pricila da Silva Cunha

    2014-01-01

    Full Text Available Monosomy 1p36 is considered the most common subtelomeric deletion syndrome in humans and it accounts for 0.5–0.7% of all the cases of idiopathic intellectual disability. The molecular diagnosis is often made by microarray-based comparative genomic hybridization (aCGH, which has the drawback of being a high-cost technique. However, patients with classic monosomy 1p36 share some typical clinical characteristics that, together with its common prevalence, justify the development of a less expensive, targeted diagnostic method. In this study, we developed a simple, rapid, and inexpensive real-time quantitative PCR (qPCR assay for targeted diagnosis of monosomy 1p36, easily accessible for low-budget laboratories in developing countries. For this, we have chosen two target genes which are deleted in the majority of patients with monosomy 1p36: PRKCZ and SKI. In total, 39 patients previously diagnosed with monosomy 1p36 by aCGH, fluorescent in situ hybridization (FISH, and/or multiplex ligation-dependent probe amplification (MLPA all tested positive on our qPCR assay. By simultaneously using these two genes we have been able to detect 1p36 deletions with 100% sensitivity and 100% specificity. We conclude that qPCR of PRKCZ and SKI is a fast and accurate diagnostic test for monosomy 1p36, costing less than 10 US dollars in reagent costs.

  15. Detection of exogenous gene doping of IGF-I by a real-time quantitative PCR assay.

    Science.gov (United States)

    Zhang, Jin-Ju; Xu, Jing-Feng; Shen, Yong-Wei; Ma, Shi-Jiao; Zhang, Ting-Ting; Meng, Qing-Lin; Lan, Wen-Jun; Zhang, Chun; Liu, Xiao-Mei

    2017-07-01

    Gene doping can be easily concealed since its product is similar to endogenous protein, making its effective detection very challenging. In this study, we selected insulin-like growth factor I (IGF-I) exogenous gene for gene doping detection. First, the synthetic IGF-I gene was subcloned to recombinant adeno-associated virus (rAAV) plasmid to produce recombinant rAAV2/IGF-I-GFP vectors. Second, in an animal model, rAAV2/IGF-I-GFP vectors were injected into the thigh muscle tissue of mice, and then muscle and blood specimens were sampled at different time points for total DNA isolation. Finally, real-time quantitative PCR was employed to detect the exogenous gene doping of IGF-I. In view of the characteristics of endogenous IGF-I gene sequences, a TaqMan probe was designed at the junction of exons 2 and 3 of IGF-I gene to distinguish it from the exogenous IGF-I gene. In addition, an internal reference control plasmid and its probe were used in PCR to rule out false-positive results through comparison of their threshold cycle (Ct) values. Thus, an accurate exogenous IGF-I gene detection approach was developed in this study. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

  16. Identification of Reference Genes for Quantitative Real Time PCR Assays in Aortic Tissue of Syrian Hamsters with Bicuspid Aortic Valve.

    Science.gov (United States)

    Rueda-Martínez, Carmen; Fernández, M Carmen; Soto-Navarrete, María Teresa; Jiménez-Navarro, Manuel; Durán, Ana Carmen; Fernández, Borja

    2016-01-01

    Bicuspid aortic valve (BAV) is the most frequent congenital cardiac malformation in humans, and appears frequently associated with dilatation of the ascending aorta. This association is likely the result of a common aetiology. Currently, a Syrian hamster strain with a relatively high (∼40%) incidence of BAV constitutes the only spontaneous animal model of BAV disease. The characterization of molecular alterations in the aorta of hamsters with BAV may serve to identify pathophysiological mechanisms and molecular markers of disease in humans. In this report, we evaluate the expression of ten candidate reference genes in aortic tissue of hamsters in order to identify housekeeping genes for normalization using quantitative real time PCR (RT-qPCR) assays. A total of 51 adult (180-240 days old) and 56 old (300-440 days old) animals were used. They belonged to a control strain of hamsters with normal, tricuspid aortic valve (TAV; n = 30), or to the affected strain of hamsters with TAV (n = 45) or BAV (n = 32). The expression stability of the candidate reference genes was determined by RT-qPCR using three statistical algorithms, GeNorm, NormFinder and Bestkeeper. The expression analyses showed that the most stable reference genes for the three algorithms employed were Cdkn1β, G3pdh and Polr2a. We propose the use of Cdkn1β, or both Cdkn1β and G3pdh as reference genes for mRNA expression analyses in Syrian hamster aorta.

  17. Identification of Reference Genes for Quantitative Real Time PCR Assays in Aortic Tissue of Syrian Hamsters with Bicuspid Aortic Valve.

    Directory of Open Access Journals (Sweden)

    Carmen Rueda-Martínez

    Full Text Available Bicuspid aortic valve (BAV is the most frequent congenital cardiac malformation in humans, and appears frequently associated with dilatation of the ascending aorta. This association is likely the result of a common aetiology. Currently, a Syrian hamster strain with a relatively high (∼40% incidence of BAV constitutes the only spontaneous animal model of BAV disease. The characterization of molecular alterations in the aorta of hamsters with BAV may serve to identify pathophysiological mechanisms and molecular markers of disease in humans. In this report, we evaluate the expression of ten candidate reference genes in aortic tissue of hamsters in order to identify housekeeping genes for normalization using quantitative real time PCR (RT-qPCR assays. A total of 51 adult (180-240 days old and 56 old (300-440 days old animals were used. They belonged to a control strain of hamsters with normal, tricuspid aortic valve (TAV; n = 30, or to the affected strain of hamsters with TAV (n = 45 or BAV (n = 32. The expression stability of the candidate reference genes was determined by RT-qPCR using three statistical algorithms, GeNorm, NormFinder and Bestkeeper. The expression analyses showed that the most stable reference genes for the three algorithms employed were Cdkn1β, G3pdh and Polr2a. We propose the use of Cdkn1β, or both Cdkn1β and G3pdh as reference genes for mRNA expression analyses in Syrian hamster aorta.

  18. Selection of reliable reference genes for quantitative real-time PCR in human T cells and neutrophils

    Directory of Open Access Journals (Sweden)

    Ledderose Carola

    2011-10-01

    Full Text Available Abstract Background The choice of reliable reference genes is a prerequisite for valid results when analyzing gene expression with real-time quantitative PCR (qPCR. This method is frequently applied to study gene expression patterns in immune cells, yet a thorough validation of potential reference genes is still lacking for most leukocyte subtypes and most models of their in vitro stimulation. In the current study, we evaluated the expression stability of common reference genes in two widely used cell culture models-anti-CD3/CD28 activated T cells and lipopolysaccharide stimulated neutrophils-as well as in unselected untreated leukocytes. Results The mRNA expression of 17 (T cells, 7 (neutrophils or 8 (unselected leukocytes potential reference genes was quantified by reverse transcription qPCR, and a ranking of the preselected candidate genes according to their expression stability was calculated using the programs NormFinder, geNorm and BestKeeper. IPO8, RPL13A, TBP and SDHA were identified as suitable reference genes in T cells. TBP, ACTB and SDHA were stably expressed in neutrophils. TBP and SDHA were also the most stable genes in untreated total blood leukocytes. The critical impact of reference gene selection on the estimated target gene expression is demonstrated for IL-2 and FIH expression in T cells. Conclusions The study provides a shortlist of suitable reference genes for normalization of gene expression data in unstimulated and stimulated T cells, unstimulated and stimulated neutrophils and in unselected leukocytes.

  19. Development of Real-Time Quantitative Polymerase Chain Reaction Assays to Track Treatment Response in Retinoid Resistant Acute Promyelocytic Leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Jovanovic, Jelena V. [Cancer Genetics Laboratory, Department of Medical and Molecular Genetics, King’s College London School of Medicine, London (United Kingdom); Rennie, Kristian [GSTS Pathology, Guy’s Hospital, London (United Kingdom); Culligan, Dominic [Department of Haematology, Aberdeen Royal Infirmary, Aberdeen (United Kingdom); Peniket, Andrew [Department of Haematology, John Radcliffe Hospital, Oxford (United Kingdom); Lennard, Anne [Department of Haematology, Royal Victoria Infirmary, Newcastle (United Kingdom); Harrison, Justin [Department of Haematology, Hemel Hempstead Hospital, Hemel Hempstead (United Kingdom); Vyas, Paresh [Medical Research Council Molecular Haematology Unit, Weatherall Institute of Molecular Medicine, Oxford (United Kingdom); Grimwade, David, E-mail: david.grimwade@genetics.kcl.ac.uk [Cancer Genetics Laboratory, Department of Medical and Molecular Genetics, King’s College London School of Medicine, London (United Kingdom)

    2011-10-25

    Molecular detection of minimal residual disease (MRD) has become established to assess remission status and guide therapy in patients with ProMyelocytic Leukemia–RARA+ acute promyelocytic leukemia (APL). However, there are few data on tracking disease response in patients with rarer retinoid resistant subtypes of APL, characterized by PLZF–RARA and STAT5b–RARA. Despite their rarity (<1% of APL) we identified 6 cases (PLZF–RARA, n = 5; STAT5b–RARA, n = 1), established the respective breakpoint junction regions and designed reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) assays to detect leukemic transcripts. The relative level of fusion gene expression in diagnostic samples was comparable to that observed in t(15;17) – associated APL, affording assay sensitivities of ∼1 in 10{sup 4}−10{sup 5}. Serial samples were available from two PLZF–RARA APL patients. One showed persistent polymerase chain reaction positivity, predicting subsequent relapse, and remains in CR2, ∼11 years post-autograft. The other, achieved molecular remission (CRm) with combination chemotherapy, remaining in CR1 at 6 years. The STAT5b–RARA patient failed to achieve CRm following frontline combination chemotherapy and ultimately proceeded to allogeneic transplant on the basis of a steadily rising fusion transcript level. These data highlight the potential of RT-qPCR detection of MRD to facilitate development of more individualized approaches to the management of rarer molecularly defined subsets of acute leukemia.

  20. Development of Real-Time Quantitative Polymerase Chain Reaction Assays to Track Treatment Response in Retinoid Resistant Acute Promyelocytic Leukemia

    International Nuclear Information System (INIS)

    Jovanovic, Jelena V.; Rennie, Kristian; Culligan, Dominic; Peniket, Andrew; Lennard, Anne; Harrison, Justin; Vyas, Paresh; Grimwade, David

    2011-01-01

    Molecular detection of minimal residual disease (MRD) has become established to assess remission status and guide therapy in patients with ProMyelocytic Leukemia–RARA+ acute promyelocytic leukemia (APL). However, there are few data on tracking disease response in patients with rarer retinoid resistant subtypes of APL, characterized by PLZF–RARA and STAT5b–RARA. Despite their rarity (<1% of APL) we identified 6 cases (PLZF–RARA, n = 5; STAT5b–RARA, n = 1), established the respective breakpoint junction regions and designed reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) assays to detect leukemic transcripts. The relative level of fusion gene expression in diagnostic samples was comparable to that observed in t(15;17) – associated APL, affording assay sensitivities of ∼1 in 10 4 −10 5 . Serial samples were available from two PLZF–RARA APL patients. One showed persistent polymerase chain reaction positivity, predicting subsequent relapse, and remains in CR2, ∼11 years post-autograft. The other, achieved molecular remission (CRm) with combination chemotherapy, remaining in CR1 at 6 years. The STAT5b–RARA patient failed to achieve CRm following frontline combination chemotherapy and ultimately proceeded to allogeneic transplant on the basis of a steadily rising fusion transcript level. These data highlight the potential of RT-qPCR detection of MRD to facilitate development of more individualized approaches to the management of rarer molecularly defined subsets of acute leukemia.

  1. Development of a multiplex real-time PCR assay for phylogenetic analysis of Uropathogenic Escherichia coli.

    Science.gov (United States)

    Hasanpour, Mojtaba; Najafi, Akram

    2017-06-01

    Uropathogenic Escherichia coli (UPEC) is among major pathogens causing 80-90% of all episodes of urinary tract infections (UTIs). Recently, E. coli strains are divided into eight main phylogenetic groups including A, B1, B2, C, D, E, F, and clade I. This study was aimed to develop a rapid, sensitive, and specific multiplex real time PCR method capable of detecting phylogenetic groups of E. coli strains. This study was carried out on E. coli strains (isolated from the patient with UTI) in which the presence of all seven target genes had been confirmed in our previous phylogenetic study. An EvaGreen-based singleplex and multiplex real-time PCR with melting curve analysis was designed for simultaneous detection and differentiation of these genes. The primers were selected mainly based on the production of amplicons with melting temperatures (T m ) ranging from 82°C to 93°C and temperature difference of more than 1.5°C between each peak.The multiplex real-time PCR assays that have been developed in the present study were successful in detecting the eight main phylogenetic groups. Seven distinct melting peaks were discriminated, with Tm value of 93±0.8 for arpA, 89.2±0.1for chuA, 86.5±0.1 for yjaA, 82.3±0.2 for TspE4C2, 87.8±0.1for trpAgpC, 85.4±0.6 for arpAgpE genes, and 91±0.5 for the internal control. To our knowledge, this study is the first melting curve-based real-time PCR assay developed for simultaneous and discrete detection of these seven target genes. Our findings showed that this assay has the potential to be a rapid, reliable and cost-effective alternative for routine phylotyping of E. coli strains. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Development of Geometrical Quality Control Real-time Analysis Program using an Electronic Portal Imaging

    International Nuclear Information System (INIS)

    Lee, Sang Rok; Jung, Kyung Yong; Jang, Min Sun; Lee, Byung Gu; Kwon, Young Ho

    2012-01-01

    To develop a geometrical quality control real-time analysis program using an electronic portal imaging to replace film evaluation method. A geometrical quality control item was established with the Eclipse treatment planning system (Version 8.1, Varian, USA) after the Electronic Portal Imaging Device (EPID) took care of the problems occurring from the fixed substructure of the linear accelerator (CL-iX, Varian, USA). Electronic portal image (single exposure before plan) was created at the treatment room's 4DTC (Version 10.2, Varian, USA) and a beam was irradiated in accordance with each item. The gaining the entire electronic portal imaging at the Off-line review and was evaluated by a self-developed geometrical quality control real-time analysis program. As for evaluation methods, the intra-fraction error was analyzed by executing 5 times in a row under identical conditions and procedures on the same day, and in order to confirm the infer-fraction error, it was executed for 10 days under identical conditions of all procedures and was compared with the film evaluation method using an Iso-align quality control device. Measurement and analysis time was measured by sorting the time into from the device setup to data achievement and the time amount after the time until the completion of analysis and the convenience of the users and execution processes were compared. The intra-fraction error values for each average 0.1, 0.2, 0.3, 0.2 mm at light-radiation field coincidence, collimator rotation axis, couch rotation axis and gantry rotation axis. By checking the infer-fraction error through 10 days of continuous quality control, the error values obtained were average 1.7, 1.4, 0.7, 1.1 mm for each item. Also, the measurement times were average 36 minutes, 15 minutes for the film evaluation method and electronic portal imaging system, and the analysis times were average 30 minutes, 22 minutes. When conducting a geometrical quality control using an electronic portal imaging

  3. Real time analysis of electromagnetic radiation in a very wide frequency range

    International Nuclear Information System (INIS)

    Peralta, J.A.; Reyes L, P.; Yepez, E.

    2001-01-01

    In this work, we present an electronic apparatus that facilitates the monitoring and analysis of electromagnetic radiation in a very wide frequency range. The device is a combination of real and virtual instruments, taking advantage of new hardware and software; the measurable range of frequencies depends on the speed of an analog/digital converter, reaching tens of Megahertz. The device has been successfully used to monitor the environmental electromagnetic radiation at very low frequency, a very useful parameter in the research of electromagnetic precursors of earthquakes. The apparatus is a new configuration and has advantages with respect to those previously used: when the attached computer is fast, Fourier analysis can be done in real time, can display simultaneously several bands, the digitized data allow a variety of methods of analysis, and the apparatus is very cheap. (Author)

  4. Profiling of Piper betle Linn. cultivars by direct analysis in real time mass spectrometric technique.

    Science.gov (United States)

    Bajpai, Vikas; Sharma, Deepty; Kumar, Brijesh; Madhusudanan, K P

    2010-12-01

    Piper betle Linn. is a traditional plant associated with the Asian and southeast Asian cultures. Its use is also recorded in folk medicines in these regions. Several of its medicinal properties have recently been proven. Phytochemical analysis showed the presence of mainly terpenes and phenols in betel leaves. These constituents vary in the different cultivars of Piper betle. In this paper we have attempted to profile eight locally available betel cultivars using the recently developed mass spectral ionization technique of direct analysis in real time (DART). Principal component analysis has also been employed to analyze the DART MS data of these betel cultivars. The results show that the cultivars of Piper betle could be differentiated using DART MS data. Copyright © 2010 John Wiley & Sons, Ltd.

  5. Validation of the direct analysis in real time source for use in forensic drug screening.

    Science.gov (United States)

    Steiner, Robert R; Larson, Robyn L

    2009-05-01

    The Direct Analysis in Real Time (DART) ion source is a relatively new mass spectrometry technique that is seeing widespread use in chemical analyses world-wide. DART studies include such diverse topics as analysis of flavors and fragrances, melamine in contaminated dog food, differentiation of writing inks, characterization of solid counterfeit drugs, and as a detector for planar chromatography. Validation of this new technique for the rapid screening of forensic evidence for drugs of abuse, utilizing the DART source coupled to an accurate mass time-of-flight mass spectrometer, was conducted. The study consisted of the determination of the lower limit of detection for the method, determination of selectivity and a comparison of this technique to established analytical protocols. Examples of DART spectra are included. The results of this study have allowed the Virginia Department of Forensic Science to incorporate this new technique into their analysis scheme for the screening of solid dosage forms of drugs of abuse.

  6. Real time analysis of electromagnetic radiation in a very wide frequency range

    Energy Technology Data Exchange (ETDEWEB)

    Peralta, J.A.; Reyes L, P.; Yepez, E. [Escuela Superior de Fisica y Matematicas, Instituto Politecnico Nacional, Edificio 9, U.P. Adolfo Lopez Mateos, Zacatenco, 07738 Mexico D.F. (Mexico)

    2001-07-01

    In this work, we present an electronic apparatus that facilitates the monitoring and analysis of electromagnetic radiation in a very wide frequency range. The device is a combination of real and virtual instruments, taking advantage of new hardware and software; the measurable range of frequencies depends on the speed of an analog/digital converter, reaching tens of Megahertz. The device has been successfully used to monitor the environmental electromagnetic radiation at very low frequency, a very useful parameter in the research of electromagnetic precursors of earthquakes. The apparatus is a new configuration and has advantages with respect to those previously used: when the attached computer is fast, Fourier analysis can be done in real time, can display simultaneously several bands, the digitized data allow a variety of methods of analysis, and the apparatus is very cheap. (Author)

  7. A CAMAC based real-time noise analysis system for nuclear reactors

    International Nuclear Information System (INIS)

    Ciftcioglu, O.

    1987-01-01

    A CAMAC based real-time noise analysis system was designed for the TRIGA MARK II nuclear reactor at the Institute for Nuclear Energy, Istanbul. The input analog signals obtained from the radiation detectors are introduced to the system through CAMAC interface. The signals coverted into digital form are processed by a PDP-11 computer. The fast data processing based on auto/cross power spectral density computations is carried out by means of assembly written FFT algorithms in real-time and the spectra obtained are displayed on a CAMAC driven display system as an additional monitoring device. The system has the advantage of being software programmable and controlled by a CAMAC system so that it is operated under porgram control for reactor surveillance, anomaly detection and diagnosis. The system can also be used for the identification of nonstationary operational characteristics of the reactor in long term by comparing the noise power spectra with the corresponding reference noise patterns prepared in advance. (orig.)

  8. Real time data analysis with the ATLAS Trigger at the LHC in Run-2

    CERN Document Server

    Beauchemin, Pierre-Hugues; The ATLAS collaboration

    2018-01-01

    The trigger selection capabilities of the ATLAS detector have been significantly enhanced for the LHC Run- 2 in order to cope with the higher event rates and with the large number of simultaneous interactions (pile-up) per protonproton bunch crossing. A new hardware system, designed to analyse real time event-topologies at Level-1 came to full use in 2017. A hardware-based track reconstruction system, expected to be used real-time in 2018, is designed to provide track information to the high-level software trigger at its full input rate. The high-level trigger selections are largely relying on offline-like reconstruction techniques, and in some cases multivariate analysis methods. Despite the sudden change in LHC operations during the second half of 2017, which caused an increase in pile-up and therefore also in CPU usage of the trigger algorithms, the set of triggers (so called trigger menu) running online has undergone only minor modifications thanks to the robustness and redundancy of the trigger system, a...

  9. Real time data analysis with the ATLAS trigger at the LHC in Run-2

    CERN Document Server

    Beauchemin, Pierre-Hugues; The ATLAS collaboration

    2018-01-01

    The trigger selection capabilities of the ATLAS detector have been significantly enhanced for the LHC Run-2 in order to cope with the higher event rates and with the large number of simultaneous interactions (pile-up) per proton-proton bunch crossing. A new hardware system, designed to analyse real time event-topologies at Level-1 came to full use in 2017. A hardware-based track reconstruction system, expected to be used real-time in 2018, is designed to provide track information to the high-level software trigger at its full input rate. The high-level trigger selections are largely relying on offline-like reconstruction techniques, and in some cases multi-variate analysis methods. Despite the sudden change in LHC operations during the second half of 2017, which caused an increase in pile-up and therefore also in CPU usage of the trigger algorithms, the set of triggers (so called trigger menu) running online has undergone only minor modifications thanks to the robustness and redundancy of the trigger system, ...

  10. Distributed Resource Energy Analysis and Management System (DREAMS) Development for Real-time Grid Operations

    Energy Technology Data Exchange (ETDEWEB)

    Nakafuji, Dora [Hawaiian Electric Company, Honululu, HI (United States); Gouveia, Lauren [Hawaiian Electric Company, Honululu, HI (United States)

    2016-10-24

    This project supports development of the next generation, integrated energy management infrastructure (EMS) able to incorporate advance visualization of behind-the-meter distributed resource information and probabilistic renewable energy generation forecasts to inform real-time operational decisions. The project involves end-users and active feedback from an Utility Advisory Team (UAT) to help inform how information can be used to enhance operational functions (e.g. unit commitment, load forecasting, Automatic Generation Control (AGC) reserve monitoring, ramp alerts) within two major EMS platforms. Objectives include: Engaging utility operations personnel to develop user input on displays, set expectations, test and review; Developing ease of use and timeliness metrics for measuring enhancements; Developing prototype integrated capabilities within two operational EMS environments; Demonstrating an integrated decision analysis platform with real-time wind and solar forecasting information and timely distributed resource information; Seamlessly integrating new 4-dimensional information into operations without increasing workload and complexities; Developing sufficient analytics to inform and confidently transform and adopt new operating practices and procedures; Disseminating project lessons learned through industry sponsored workshops and conferences;Building on collaborative utility-vendor partnership and industry capabilities

  11. Analysis of actuator delay and its effect on uncertainty quantification for real-time hybrid simulation

    Science.gov (United States)

    Chen, Cheng; Xu, Weijie; Guo, Tong; Chen, Kai

    2017-10-01

    Uncertainties in structure properties can result in different responses in hybrid simulations. Quantification of the effect of these uncertainties would enable researchers to estimate the variances of structural responses observed from experiments. This poses challenges for real-time hybrid simulation (RTHS) due to the existence of actuator delay. Polynomial chaos expansion (PCE) projects the model outputs on a basis of orthogonal stochastic polynomials to account for influences of model uncertainties. In this paper, PCE is utilized to evaluate effect of actuator delay on the maximum displacement from real-time hybrid simulation of a single degree of freedom (SDOF) structure when accounting for uncertainties in structural properties. The PCE is first applied for RTHS without delay to determine the order of PCE, the number of sample points as well as the method for coefficients calculation. The PCE is then applied to RTHS with actuator delay. The mean, variance and Sobol indices are compared and discussed to evaluate the effects of actuator delay on uncertainty quantification for RTHS. Results show that the mean and the variance of the maximum displacement increase linearly and exponentially with respect to actuator delay, respectively. Sensitivity analysis through Sobol indices also indicates the influence of the single random variable decreases while the coupling effect increases with the increase of actuator delay.

  12. Collaborative real-time motion video analysis by human observer and image exploitation algorithms

    Science.gov (United States)

    Hild, Jutta; Krüger, Wolfgang; Brüstle, Stefan; Trantelle, Patrick; Unmüßig, Gabriel; Heinze, Norbert; Peinsipp-Byma, Elisabeth; Beyerer, Jürgen

    2015-05-01

    Motion video analysis is a challenging task, especially in real-time applications. In most safety and security critical applications, a human observer is an obligatory part of the overall analysis system. Over the last years, substantial progress has been made in the development of automated image exploitation algorithms. Hence, we investigate how the benefits of automated video analysis can be integrated suitably into the current video exploitation systems. In this paper, a system design is introduced which strives to combine both the qualities of the human observer's perception and the automated algorithms, thus aiming to improve the overall performance of a real-time video analysis system. The system design builds on prior work where we showed the benefits for the human observer by means of a user interface which utilizes the human visual focus of attention revealed by the eye gaze direction for interaction with the image exploitation system; eye tracker-based interaction allows much faster, more convenient, and equally precise moving target acquisition in video images than traditional computer mouse selection. The system design also builds on prior work we did on automated target detection, segmentation, and tracking algorithms. Beside the system design, a first pilot study is presented, where we investigated how the participants (all non-experts in video analysis) performed in initializing an object tracking subsystem by selecting a target for tracking. Preliminary results show that the gaze + key press technique is an effective, efficient, and easy to use interaction technique when performing selection operations on moving targets in videos in order to initialize an object tracking function.

  13. Novel Quantitative Real-Time LCR for the Sensitive Detection of SNP Frequencies in Pooled DNA: Method Development, Evaluation and Application

    Science.gov (United States)

    Psifidi, Androniki; Dovas, Chrysostomos; Banos, Georgios

    2011-01-01

    Background Single nucleotide polymorphisms (SNP) have proven to be powerful genetic markers for genetic applications in medicine, life science and agriculture. A variety of methods exist for SNP detection but few can quantify SNP frequencies when the mutated DNA molecules correspond to a small fraction of the wild-type DNA. Furthermore, there is no generally accepted gold standard for SNP quantification, and, in general, currently applied methods give inconsistent results in selected cohorts. In the present study we sought to develop a novel method for accurate detection and quantification of SNP in DNA pooled samples. Methods The development and evaluation of a novel Ligase Chain Reaction (LCR) protocol that uses a DNA-specific fluorescent dye to allow quantitative real-time analysis is described. Different reaction components and thermocycling parameters affecting the efficiency and specificity of LCR were examined. Several protocols, including gap-LCR modifications, were evaluated using plasmid standard and genomic DNA pools. A protocol of choice was identified and applied for the quantification of a polymorphism at codon 136 of the ovine PRNP gene that is associated with susceptibility to a transmissible spongiform encephalopathy in sheep. Conclusions The real-time LCR protocol developed in the present study showed high sensitivity, accuracy, reproducibility and a wide dynamic range of SNP quantification in different DNA pools. The limits of detection and quantification of SNP frequencies were 0.085% and 0.35%, respectively. Significance The proposed real-time LCR protocol is applicable when sensitive detection and accurate quantification of low copy number mutations in DNA pools is needed. Examples include oncogenes and tumour suppressor genes, infectious diseases, pathogenic bacteria, fungal species, viral mutants, drug resistance resulting from point mutations, and genetically modified organisms in food. PMID:21283808

  14. A Real-Time PCR with Melting Curve Analysis for Molecular Typing of Vibrio parahaemolyticus.

    Science.gov (United States)

    He, Peiyan; Wang, Henghui; Luo, Jianyong; Yan, Yong; Chen, Zhongwen

    2018-05-23

    Foodborne disease caused by Vibrio parahaemolyticus is a serious public health problem in many countries. Molecular typing has a great scientific significance and application value for epidemiological research of V. parahaemolyticus. In this study, a real-time PCR with melting curve analysis was established for molecular typing of V. parahaemolyticus. Eighteen large variably presented gene clusters (LVPCs) of V. parahaemolyticus which have different distributions in the genome of different strains were selected as targets. Primer pairs of 18 LVPCs were distributed into three tubes. To validate this newly developed assay, we tested 53 Vibrio parahaemolyticus strains, which were classified in 13 different types. Furthermore, cluster analysis using NTSYS PC 2.02 software could divide 53 V. parahaemolyticus strains into six clusters at a relative similarity coefficient of 0.85. This method is fast, simple, and conveniently for molecular typing of V. parahaemolyticus.

  15. Timing Analysis of Mixed-Criticality Hard Real-Time Applications Implemented on Distributed Partitioned Architectures

    DEFF Research Database (Denmark)

    Marinescu, Sorin Ovidiu; Tamas-Selicean, Domitian; Acretoaie, Vlad

    In this paper we are interested in the timing analysis of mixed-criticality embedded real-time applications mapped on distributed heterogeneous architectures. Mixedcriticality tasks can be integrated onto the same architecture only if there is enough spatial and temporal separation among them. We...... in partitions using fixedpriority preemptive scheduling. We have extended the stateof- the-art algorithms for schedulability analysis to take into account the partitions. The proposed algorithm has been evaluated using several synthetic and real-life benchmarks....... consider that the separation is provided by partitioning, such that applications run in separate partitions, and each partition is allocated several time slots on a processor. Each partition can have its own scheduling policy. We are interested to determine the worst-case response times of tasks scheduled...

  16. Direct analysis in real time mass spectrometry, a process analytical technology tool for real-time process monitoring in botanical drug manufacturing.

    Science.gov (United States)

    Wang, Lu; Zeng, Shanshan; Chen, Teng; Qu, Haibin

    2014-03-01

    A promising process analytical technology (PAT) tool has been introduced for batch processes monitoring. Direct analysis in real time mass spectrometry (DART-MS), a means of rapid fingerprint analysis, was applied to a percolation process with multi-constituent substances for an anti-cancer botanical preparation. Fifteen batches were carried out, including ten normal operations and five abnormal batches with artificial variations. The obtained multivariate data were analyzed by a multi-way partial least squares (MPLS) model. Control trajectories were derived from eight normal batches, and the qualification was tested by R(2) and Q(2). Accuracy and diagnosis capability of the batch model were then validated by the remaining batches. Assisted with high performance liquid chromatography (HPLC) determination, process faults were explained by corresponding variable contributions. Furthermore, a batch level model was developed to compare and assess the model performance. The present study has demonstrated that DART-MS is very promising in process monitoring in botanical manufacturing. Compared with general PAT tools, DART-MS offers a particular account on effective compositions and can be potentially used to improve batch quality and process consistency of samples in complex matrices. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Ex vivo screening for immunodominant viral epitopes by quantitative real time polymerase chain reaction (qRT-PCR

    Directory of Open Access Journals (Sweden)

    Nagorsen Dirk

    2003-12-01

    Full Text Available Abstract The identification and characterization of viral epitopes across the Human Leukocyte Antigen (HLA polymorphism is critical for the development of actives-specific or adoptive immunotherapy of virally-mediated diseases. This work investigates whether cytokine mRNA transcripts could be used to identify epitope-specific HLA-restricted memory T lymphocytes reactivity directly in fresh peripheral blood mononuclear cells (PBMCs from viral-seropositive individuals in response to ex vivo antigen recall. PBMCs from HLA-A*0201 healthy donors, seropositive for Cytomegalovirus (CMV and Influenza (Flu, were exposed for different periods and at different cell concentrations to the HLA-A*0201-restricted viral FluM158–66 and CMVpp65495–503 peptides. Quantitative real time PCR (qRT-PCR was employed to evaluate memory T lymphocyte immune reactivation by measuring the production of mRNA encoding four cytokines: Interferon-γ (IFN-γ, Interleukin-2 (IL-2, Interleukin-4 (IL-4, and Interleukin-10 (IL-10. We could characterize cytokine expression kinetics that illustrated how cytokine mRNA levels could be used as ex vivo indicators of T cell reactivity. Particularly, IFN-γ mRNA transcripts could be consistently detected within 3 to 12 hours of short-term stimulation in levels sufficient to screen for HLA-restricted viral immune responses in seropositive subjects. This strategy will enhance the efficiency of the identification of viral epitopes independently of the individual HLA phenotype and could be used to follow the intensity of immune responses during disease progression or in response to in vivo antigen-specific immunization.

  18. Ex vivo screening for immunodominant viral epitopes by quantitative real time polymerase chain reaction (qRT-PCR)

    Science.gov (United States)

    Provenzano, Maurizio; Mocellin, Simone; Bonginelli, Paola; Nagorsen, Dirk; Kwon, Seog-Woon; Stroncek, David

    2003-01-01

    The identification and characterization of viral epitopes across the Human Leukocyte Antigen (HLA) polymorphism is critical for the development of actives-specific or adoptive immunotherapy of virally-mediated diseases. This work investigates whether cytokine mRNA transcripts could be used to identify epitope-specific HLA-restricted memory T lymphocytes reactivity directly in fresh peripheral blood mononuclear cells (PBMCs) from viral-seropositive individuals in response to ex vivo antigen recall. PBMCs from HLA-A*0201 healthy donors, seropositive for Cytomegalovirus (CMV) and Influenza (Flu), were exposed for different periods and at different cell concentrations to the HLA-A*0201-restricted viral FluM158–66 and CMVpp65495–503 peptides. Quantitative real time PCR (qRT-PCR) was employed to evaluate memory T lymphocyte immune reactivation by measuring the production of mRNA encoding four cytokines: Interferon-γ (IFN-γ), Interleukin-2 (IL-2), Interleukin-4 (IL-4), and Interleukin-10 (IL-10). We could characterize cytokine expression kinetics that illustrated how cytokine mRNA levels could be used as ex vivo indicators of T cell reactivity. Particularly, IFN-γ mRNA transcripts could be consistently detected within 3 to 12 hours of short-term stimulation in levels sufficient to screen for HLA-restricted viral immune responses in seropositive subjects. This strategy will enhance the efficiency of the identification of viral epitopes independently of the individual HLA phenotype and could be used to follow the intensity of immune responses during disease progression or in response to in vivo antigen-specific immunization. PMID:14675481

  19. Evaluation of candidate reference genes for gene expression normalization in Brassica juncea using real time quantitative RT-PCR.

    Directory of Open Access Journals (Sweden)

    Ruby Chandna

    Full Text Available The real time quantitative reverse transcription PCR (qRT-PCR is becoming increasingly important to gain insight into function of genes. Given the increased sensitivity, ease and reproducibility of qRT-PCR, the requirement of suitable reference genes for normalization has become important and stringent. It is now known that the expression of internal control genes in living organism vary considerably during developmental stages and under different experimental conditions. For economically important Brassica crops, only a couple of reference genes are reported till date. In this study, expression stability of 12 candidate reference genes including ACT2, ELFA, GAPDH, TUA, UBQ9 (traditional housekeeping genes, ACP, CAC, SNF, TIPS-41, TMD, TSB and ZNF (new candidate reference genes, in a diverse set of 49 tissue samples representing different developmental stages, stress and hormone treated conditions and cultivars of Brassica juncea has been validated. For the normalization of vegetative stages the ELFA, ACT2, CAC and TIPS-41 combination would be appropriate whereas TIPS-41 along with CAC would be suitable for normalization of reproductive stages. A combination of GAPDH, TUA, TIPS-41 and CAC were identified as the most suitable reference genes for total developmental stages. In various stress and hormone treated samples, UBQ9 and TIPS-41 had the most stable expression. Across five cultivars of B. juncea, the expression of CAC and TIPS-41 did not vary significantly and were identified as the most stably expressed reference genes. This study provides comprehensive information that the new reference genes selected herein performed better than the traditional housekeeping genes. The selection of most suitable reference genes depends on the experimental conditions, and is tissue and cultivar-specific. Further, to attain accuracy in the results more than one reference genes are necessary for normalization.

  20. SNPs and real-time quantitative PCR method for constitutional allelic copy number determination, the VPREB1 marker case

    Directory of Open Access Journals (Sweden)

    Costa Elena

    2011-05-01

    Full Text Available Abstract Background 22q11.2 microdeletion is responsible for the DiGeorge Syndrome, characterized by heart defects, psychiatric disorders, endocrine and immune alterations and a 1 in 4000 live birth prevalence. Real-time quantitative PCR (qPCR approaches for allelic copy number determination have recently been investigated in 22q11.2 microdeletions detection. The qPCR method was performed for 22q11.2 microdeletions detection as a first-level screening approach in a genetically unknown series of patients with congenital heart defects. A technical issue related to the VPREB1 qPCR marker was pointed out. Methods A set of 100 unrelated Italian patients with congenital heart defects were tested for 22q11.2 microdeletions by a qPCR method using six different markers. Fluorescence In Situ Hybridization technique (FISH was used for confirmation. Results qPCR identified six patients harbouring the 22q11.2 microdeletion, confirmed by FISH. The VPREB1 gene marker presented with a pattern consistent with hemideletion in one 3 Mb deleted patient, suggestive for a long distal deletion, and in additional five non-deleted patients. The long distal 22q11.2 deletion was not confirmed by Comparative Genomic Hybridization. Indeed, the VPREB1 gene marker generated false positive results in association with the rs1320 G/A SNP, a polymorphism localized within the VPREB1 marker reverse primer sequence. Patients heterozygous for rs1320 SNP, showed a qPCR profile consistent with the presence of a hemideletion. Conclusions Though the qPCR technique showed advantages as a screening approach in terms of cost and time, the VPREB1 marker case revealed that single nucleotide polymorphisms can interfere with qPCR data generating erroneous allelic copy number interpretations.

  1. Application of propidium monoazide quantitative real-time PCR to quantify the viability of Lactobacillus delbrueckii ssp. bulgaricus.

    Science.gov (United States)

    Shao, Yuyu; Wang, Zhaoxia; Bao, Qiuhua; Zhang, Heping

    2016-12-01

    In this study, a combination of propidium monoazide (PMA) and quantitative real-time PCR (qPCR) was used to develop a method to determine the viability of cells of Lactobacillus delbrueckii ssp. bulgaricus ND02 (L. bulgaricus) that may have entered into a viable but nonculturable state. This can happen due to its susceptibility to cold shock during lyophilization and storage. Propidium monoazide concentration, PMA incubation time, and light exposure time were optimized to fully exploit the PMA-qPCR approach to accurately assess the total number of living L. bulgaricus ND02. Although PMA has little influence on living cells, when concentrations of PMA were higher than 30μg/mL the number of PCR-positive living bacteria decreased from 10 6 to 10 5 cfu/mL in comparison with qPCR enumeration. Mixtures of living and dead cells were used as method verification samples for enumeration by PMA-qPCR, demonstrating that this method was feasible and effective for distinguishing living cells of L. bulgaricus when mixed with a known number of dead cells. We suggest that several conditions need to be studied further before PMA-qPCR methods can be accurately used to distinguish living from dead cells for enumeration under more realistic sampling situations. However, this research provides a rapid way to enumerate living cells of L. bulgaricus and could be used to optimize selection of cryoprotectants in the lyophilization process and develop technologies for high cell density cultivation and optimal freeze-drying processes. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  2. Quantitative Real-time Polymerase Chain Reaction for Enteropathogenic Escherichia coli: A Tool for Investigation of Asymptomatic Versus Symptomatic Infections

    Science.gov (United States)

    Barletta, Francesca; Mercado, Erik; Ruiz, Joaquim; Ecker, Lucie; Lopez, Giovanni; Mispireta, Monica; Gil, Ana I.; Lanata, Claudio F.; Cleary, Thomas G.

    2011-01-01

    Background. Enteropathogenic Escherichia coli (EPEC) strains are pediatric pathogens commonly isolated from both healthy and sick children with diarrhea in areas of endemicity. The aim of this study was to compare the bacterial load of EPEC isolated from stool samples from children with and without diarrhea to determine whether bacterial load might be a useful tool for further study of this phenomenon. Methods. EPEC was detected by polymerase chain reaction (PCR) of colonies isolated on MacConkey plates from 53 diarrheal and 90 healthy children aged <2 years. DNA was isolated from stool samples by cetyltrimethylammonium bromide extraction. To standardize quantification by quantitative real-time PCR (qRT-PCR), the correlation between fluorescence threshold cycle and copy number of the intimin gene of EPEC E2348/69 was determined. Results. The detection limit of qRT-PCR was 5 bacteria/mg stool. The geometric mean load in diarrhea was 299 bacteria/mg (95% confidence interval [CI], 77–1164 bacteria/mg), compared with 29 bacteria/mg (95% CI, 10–87 bacteria/mg) in control subjects (P = .016). Bacterial load was significantly higher in children with diarrhea than in control subjects among children <12 months of age (178 vs 5 bacteria/mg; P = .006) and among children with EPEC as the sole pathogen (463 vs 24 bacteria/mg; P = .006). Conclusions. EPEC load measured by qRT-PCR is higher in diarrheal than in healthy children. qRT-PCR may be useful to study the relationship between disease and colonization in settings of endemicity. PMID:22028433

  3. Evaluation and analysis of real-time precise orbits and clocks products from different IGS analysis centers

    Science.gov (United States)

    Zhang, Liang; Yang, Hongzhou; Gao, Yang; Yao, Yibin; Xu, Chaoqian

    2018-06-01

    To meet the increasing demands from the real-time Precise Point Positioning (PPP) users, the real-time satellite orbit and clock products are generated by different International GNSS Service (IGS) real-time analysis centers and can be publicly received through the Internet. Based on different data sources and processing strategies, the real-time products from different analysis centers therefore differ in availability and accuracy. The main objective of this paper is to evaluate availability and accuracy of different real-time products and their effects on real-time PPP. A total of nine commonly used Real-Time Service (RTS) products, namely IGS01, IGS03, CLK01, CLK15, CLK22, CLK52, CLK70, CLK81 and CLK90, will be evaluated in this paper. Because not all RTS products support multi-GNSS, only GPS products are analyzed in this paper. Firstly, the availability of all RTS products is analyzed in two levels. The first level is the epoch availability, indicating whether there is outage for that epoch. The second level is the satellite availability, which defines the available satellite number for each epoch. Then the accuracy of different RTS products is investigated on nominal accuracy and the accuracy degradation over time. Results show that Root-Mean-Square Error (RMSE) of satellite orbit ranges from 3.8 cm to 7.5 cm for different RTS products. While the mean Standard Deviations of Errors (STDE) of satellite clocks range from 1.9 cm to 5.6 cm. The modified Signal In Space Range Error (SISRE) for all products are from 1.3 cm to 5.5 cm for different RTS products. The accuracy degradation of the orbit has the linear trend for all RTS products and the satellite clock degradation depends on the satellite clock types. The Rb clocks on board of GPS IIF satellites have the smallest degradation rate of less than 3 cm over 10 min while the Cs clocks on board of GPS IIF have the largest degradation rate of more than 10 cm over 10 min. Finally, the real-time kinematic PPP is

  4. Relationship Between Ebola Virus Real-Time Quantitative Polymerase Chain Reaction-Based Threshold Cycle Value and Virus Isolation From Human Plasma.

    Science.gov (United States)

    Spengler, Jessica R; McElroy, Anita K; Harmon, Jessica R; Ströher, Ute; Nichol, Stuart T; Spiropoulou, Christina F

    2015-10-01

    We performed a longitudinal analysis of plasma samples obtained from 4 patients with Ebola virus (EBOV) disease (EVD) to determine the relationship between the real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR)-based threshold cycle (Ct) value and the presence of infectious EBOV. EBOV was not isolated from plasma samples with a Ct value of >35.5 or >12 days after onset of symptoms. EBOV was not isolated from plasma samples in which anti-EBOV nucleoprotein immunoglobulin G was detected. These data demonstrate the utility of interpreting qRT-PCR results in the context of the course of EBOV infection and associated serological responses for patient-management decisions. Published by Oxford University Press on behalf of the Infectious Diseases Society of America 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  5. CyNC - towards a General Tool for Performance Analysis of Complex Distributed Real Time Systems

    DEFF Research Database (Denmark)

    Schiøler, Henrik; Jessen, Jan Jakob; Nielsen, Jens F. Dalsgaard

    2005-01-01

    The paper addresses the current state and the ongoing activities of a tool for performance analysis of complex real time systems. The tool named CyNC is based on network calculus allowing for the computation of backlogs and delays in a system from specified lower and upper bounds of external...... workflow and computational resources. The current version of the tool implements an extension to previous work in that it allows for general workflow and resource bounds and provides optimal solutions even to systems with cyclic dependencies. Despite the virtues of the current tool, improvements...... and extensions still remain, which are in focus of ongoing activities. Improvements include accounting for phase information to improve bounds, whereas the tool awaits extension to include flow control models, which both depend on the possibility of accounting for propagation delay. Since the current version...

  6. Quasi-real-time photon pulse duration measurement by analysis of FEL radiation spectra

    Energy Technology Data Exchange (ETDEWEB)

    Engel, Robin, E-mail: robin.engel@uni-oldenburg.de [Deutsches Elektronen-Synchrotron, Notkestrasse 85, D-22603 Hamburg (Germany); Institut für Physik, Carl von Ossietzky Universität Oldenburg, D-26111 Oldenburg (Germany); Institut für Laser und Optik, Hochschule Emden/Leer, University of Applied Sciences, Constantiaplatz 4, D-26723 Emden (Germany); Düsterer, Stefan; Brenner, Günter [Deutsches Elektronen-Synchrotron, Notkestrasse 85, D-22603 Hamburg (Germany); Teubner, Ulrich [Deutsches Elektronen-Synchrotron, Notkestrasse 85, D-22603 Hamburg (Germany); Institut für Physik, Carl von Ossietzky Universität Oldenburg, D-26111 Oldenburg (Germany); Institut für Laser und Optik, Hochschule Emden/Leer, University of Applied Sciences, Constantiaplatz 4, D-26723 Emden (Germany)

    2016-01-01

    Considering the second-order spectral correlation function of SASE-FEL radiation allows a real-time observation of the photon pulse duration during spectra acquisition. For photon diagnostics at free-electron lasers (FELs), the determination of the photon pulse duration is an important challenge and a complex task. This is especially true for SASE FELs with strongly fluctuating pulse parameters. However, most techniques require an extensive experimental setup, data acquisition and evaluation time, limiting the usability in all-day operation. In contrast, the presented work uses an existing approach based on the analysis of statistical properties of measured SASE FEL spectra and implements it as a software tool, integrated in FLASH’s data acquisition system. This allows the calculation of the average pulse durations from a set of measured spectral distributions with only seconds of delay, whenever high-resolution spectra are recorded.

  7. Off-line real-time FTIR analysis of a process step in imipenem production

    Science.gov (United States)

    Boaz, Jhansi R.; Thomas, Scott M.; Meyerhoffer, Steven M.; Staskiewicz, Steven J.; Lynch, Joseph E.; Egan, Richard S.; Ellison, Dean K.

    1992-08-01

    We have developed an FT-IR method, using a Spectra-Tech Monit-IR 400 systems, to monitor off-line the completion of a reaction in real-time. The reaction is moisture-sensitive and analysis by more conventional methods (normal-phase HPLC) is difficult to reproduce. The FT-IR method is based on the shift of a diazo band when a conjugated beta-diketone is transformed into a silyl enol ether during the reaction. The reaction mixture is examined directly by IR and does not require sample workup. Data acquisition time is less than one minute. The method has been validated for specificity, precision and accuracy. The results obtained by the FT-IR method for known mixtures and in-process samples compare favorably with those from a normal-phase HPLC method.

  8. Tradeoff analysis for Dependable Real-Time Embedded Systems during the Early Design Phases

    DEFF Research Database (Denmark)

    Gan, Junhe

    Embedded systems are becoming increasingly complex and have tight competing constraints in terms of performance, cost, energy consumption, dependability, flexibility, security, etc. The objective of this thesis is to propose design methods and tools for supporting the tradeoff analysis of competing...... to processing elements, as well as the processor voltage and frequency levels for executing each task, such that transient faults are tolerated, the real-time constraints of the application are satisfied, and the energy consumed is minimized. In this thesis, we target the early design phases, when decisions...... have a high impact on the subsequent implementation choices. However, due to a lack of information, the early design phases are characterized by uncertainties, e.g., in the worst-case execution times (WCETs), in the functionality requirements, or in the hardware component costs. In this context, we...

  9. AEP Ohio gridSMART Demonstration Project Real-Time Pricing Demonstration Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Widergren, Steven E.; Subbarao, Krishnappa; Fuller, Jason C.; Chassin, David P.; Somani, Abhishek; Marinovici, Maria C.; Hammerstrom, Janelle L.

    2014-02-01

    This report contributes initial findings from an analysis of significant aspects of the gridSMART® Real-Time Pricing (RTP) – Double Auction demonstration project. Over the course of four years, Pacific Northwest National Laboratory (PNNL) worked with American Electric Power (AEP), Ohio and Battelle Memorial Institute to design, build, and operate an innovative system to engage residential consumers and their end-use resources in a participatory approach to electric system operations, an incentive-based approach that has the promise of providing greater efficiency under normal operating conditions and greater flexibility to react under situations of system stress. The material contained in this report supplements the findings documented by AEP Ohio in the main body of the gridSMART report. It delves into three main areas: impacts on system operations, impacts on households, and observations about the sensitivity of load to price changes.

  10. Adaptive pattern recognition in real-time video-based soccer analysis

    DEFF Research Database (Denmark)

    Schlipsing, Marc; Salmen, Jan; Tschentscher, Marc

    2017-01-01

    are taken into account. Our contribution is twofold: (1) the deliberate use of machine learning and pattern recognition techniques allows us to achieve high classification accuracy in varying environments. We systematically evaluate combinations of image features and learning machines in the given online......Computer-aided sports analysis is demanded by coaches and the media. Image processing and machine learning techniques that allow for "live" recognition and tracking of players exist. But these methods are far from collecting and analyzing event data fully autonomously. To generate accurate results......, human interaction is required at different stages including system setup, calibration, supervision of classifier training, and resolution of tracking conflicts. Furthermore, the real-time constraints are challenging: in contrast to other object recognition and tracking applications, we cannot treat data...

  11. Real Time Analysis of Bioanalytes in Healthcare, Food, Zoology and Botany.

    Science.gov (United States)

    Wang, Tianqi; Ramnarayanan, Ashwin; Cheng, Huanyu

    2017-12-21

    The growing demand for real time analysis of bioanalytes has spurred development in the field of wearable technology to offer non-invasive data collection at a low cost. The manufacturing processes for creating these sensing systems vary significantly by the material used, the type of sensors needed and the subject of study as well. The methods predominantly involve stretchable electronic sensors to monitor targets and transmit data mainly through flexible wires or short-range wireless communication devices. Capable of conformal contact, the application of wearable technology goes beyond the healthcare to fields of food, zoology and botany. With a brief review of wearable technology and its applications to various fields, we believe this mini review would be of interest to the reader in broad fields of materials, sensor development and areas where wearable sensors can provide data that are not available elsewhere.

  12. Study on Real-Time Simulation Analysis and Inverse Analysis System for Temperature and Stress of Concrete Dam

    Directory of Open Access Journals (Sweden)

    Lei Zhang

    2015-01-01

    Full Text Available In the concrete dam construction, it is very necessary to strengthen the real-time monitoring and scientific management of concrete temperature control. This paper constructs the analysis and inverse analysis system of temperature stress simulation, which is based on various useful data collected in real time in the process of concrete construction. The system can produce automatically data file of temperature and stress calculation and then achieve the remote real-time simulation calculation of temperature stress by using high performance computing techniques, so the inverse analysis can be carried out based on a basis of monitoring data in the database; it fulfills the automatic feedback calculation according to the error requirement and generates the corresponding curve and chart after the automatic processing and analysis of corresponding results. The system realizes the automation and intellectualization of complex data analysis and preparation work in simulation process and complex data adjustment in the inverse analysis process, which can facilitate the real-time tracking simulation and feedback analysis of concrete temperature stress in construction process and enable you to discover problems timely, take measures timely, and adjust construction scheme and can well instruct you how to ensure project quality.

  13. Statistical quality analysis of schedulers under soft-real-time constraints

    NARCIS (Netherlands)

    Baarsma, H.E.; Hurink, Johann L.; Jansen, P.G.

    2007-01-01

    This paper describes an algorithm to determine the performance of real-time systems with tasks using stochastic processing times. Such an algorithm can be used for guaranteeing Quality of Service of periodic tasks with soft real-time constraints. We use a discrete distribution model of processing

  14. WCET Analysis of ARM Processors using Real-Time Model Checking

    DEFF Research Database (Denmark)

    Toft, Martin; Olesen, Mads Christian; Dalsgaard, Andreas

    2009-01-01

    This paper presents a flexible method that utilises real-time model checking to determine safe and sharp WCETs for processes running on hardware platforms featuring pipelining and caching.......This paper presents a flexible method that utilises real-time model checking to determine safe and sharp WCETs for processes running on hardware platforms featuring pipelining and caching....

  15. A novel approach for evaluating the performance of real time quantitative loop-mediated isothermal amplification-based methods.

    Science.gov (United States)

    Nixon, Gavin J; Svenstrup, Helle F; Donald, Carol E; Carder, Caroline; Stephenson, Judith M; Morris-Jones, Stephen; Huggett, Jim F; Foy, Carole A

    2014-12-01

    Molecular diagnostic measurements are currently underpinned by the polymerase chain reaction (PCR). There are also a number of alternative nucleic acid amplification technologies, which unlike PCR, work at a single temperature. These 'isothermal' methods, reportedly offer potential advantages over PCR such as simplicity, speed and resistance to inhibitors and could also be used for quantitative molecular analysis. However there are currently limited mechanisms to evaluate their quantitative performance, which would assist assay development and study comparisons. This study uses a sexually transmitted infection diagnostic model in combination with an adapted metric termed isothermal doubling time (IDT), akin to PCR efficiency, to compare quantitative PCR and quantitative loop-mediated isothermal amplification (qLAMP) assays, and to quantify the impact of matrix interference. The performance metric described here facilitates the comparison of qLAMP assays that could assist assay development and validation activities.

  16. A novel approach for evaluating the performance of real time quantitative loop-mediated isothermal amplification-based methods

    Directory of Open Access Journals (Sweden)

    Gavin J. Nixon

    2014-12-01

    Full Text Available Molecular diagnostic measurements are currently underpinned by the polymerase chain reaction (PCR. There are also a number of alternative nucleic acid amplification technologies, which unlike PCR, work at a single temperature. These ‘isothermal’ methods, reportedly offer potential advantages over PCR such as simplicity, speed and resistance to inhibitors and could also be used for quantitative molecular analysis. However there are currently limited mechanisms to evaluate their quantitative performance, which would assist assay development and study comparisons. This study uses a sexually transmitted infection diagnostic model in combination with an adapted metric termed isothermal doubling time (IDT, akin to PCR efficiency, to compare quantitative PCR and quantitative loop-mediated isothermal amplification (qLAMP assays, and to quantify the impact of matrix interference. The performance metric described here facilitates the comparison of qLAMP assays that could assist assay development and validation activities.

  17. Validation of reference genes for quantitative real-time PCR in Périgord black truffle (Tuber melanosporum) developmental stages.

    Science.gov (United States)

    Zarivi, Osvaldo; Cesare, Patrizia; Ragnelli, Anna Maria; Aimola, Pierpaolo; Leonardi, Marco; Bonfigli, Antonella; Colafarina, Sabrina; Poma, Anna Maria; Miranda, Michele; Pacioni, Giovanni

    2015-08-01

    The symbiotic fungus Tuber melanosporum Vittad. (Périgord black truffle) belongs to the Ascomycota and forms mutualistic symbiosis with tree and shrub roots. This truffle has a high value in a global market and is cultivated in many countries of both hemispheres. The publication of the T. melanosporum genome has given researchers unique opportunities to learn more about the biology of the fungus. Real-time quantitative PCR (qRT-PCR) is a definitive technique for quantitating differences in transcriptional gene expression levels between samples. To facilitate gene expression studies and obtain more accurate qRT-PCR data, normalization relative to stable housekeeping genes is required. These housekeeping genes must show stable expression under given experimental conditions for the qRT-PCR results to be accurate. Unfortunately, there are no studies on the stability of housekeeping genes used in T. melanosporum development. In this study, we present a morphological and microscopical classification of the developmental stages of T. melanosporum fruit body, and investigate the expression levels of 12 candidate reference genes (18S rRNA; 5.8S rRNA; Elongation factor 1-alpha; Elongation factor 1-beta; α-tubulin; 60S ribosomal protein L29; β-tubulin; 40S ribosomal protein S1; 40S ribosomal protein S3; Glucose-6-phosphate dehydrogenase; β-actin; Ubiquitin-conjugating enzyme). To evaluate the suitability of these genes as endogenous controls, five software-based approaches and one web-based comprehensive tool (RefFinder) were used to analyze and rank the tested genes. We demonstrate here that the 18S rRNA gene shows the most stable expression during T. melanosporum development and that a set of three genes, 18S rRNA, Elongation factor 1-alpha and 40S ribosomal protein S3, is the most suitable to normalize qRT-PCR data from all the analyzed developmental stages; conversely, 18S rRNA, Glucose-6-phosphate dehydrogenase and Elongation factor 1-alpha are the most suitable

  18. Trends and advances in food analysis by real-time polymerase chain reaction.

    Science.gov (United States)

    Salihah, Nur Thaqifah; Hossain, Mohammad Mosharraf; Lubis, Hamadah; Ahmed, Minhaz Uddin

    2016-05-01

    Analyses to ensure food safety and quality are more relevant now because of rapid changes in the quantity, diversity and mobility of food. Food-contamination must be determined to maintain health and up-hold laws, as well as for ethical and cultural concerns. Real-time polymerase chain reaction (RT-PCR), a rapid and inexpensive quantitative method to detect the presence of targeted DNA-segments in samples, helps in determining both accidental and intentional adulterations of foods by biological contaminants. This review presents recent developments in theory, techniques, and applications of RT-PCR in food analyses, RT-PCR addresses the limitations of traditional food analyses in terms of sensitivity, range of analytes, multiplexing ability, cost, time, and point-of-care applications. A range of targets, including species of plants or animals which are used as food ingredients, food-borne bacteria or viruses, genetically modified organisms, and allergens, even in highly processed foods can be identified by RT-PCR, even at very low concentrations. Microfluidic RT-PCR eliminates the separate sample-processing step to create opportunities for point-of-care analyses. We also cover the challenges related to using RT-PCR for food analyses, such as the need to further improve sample handling.

  19. Comparison of real-time and quantitative polymerase chain reaction assays in detection of cytomegalovirus DNA in clinical specimens

    International Nuclear Information System (INIS)

    Gokahmetoglu, S.; Deniz, E.

    2007-01-01

    To compare the real-time (RT) and qualitative (Q) polymerase chain reaction (PCR) assays for detection of Cytomegalovirus (CMV) DNA. The study took place in the Department of Microbiology, Erciyes University, Kayseri and in Iontek Laboratory, Istanbul, Turkey, from August to December 2006. One hundred and seven clinical specimens from 67 patients were included in the study. Cytomegalovirus DNA was investigated using RT-PCR kit (Fluorion Iontek, Turkey) and Q-PCR kit (Fluorion Iontek, Turkey). Deoxyribonucleic acid sequencing was applied to the samples that yielded discrepant results in both assays. Mac Nema's Chi Square test was used for statistical analysis. Of the specimens, 27 were found positive with both assays: 9 with only RT-PCR, and 11 with only Q-PCR assay. Both assays were found negative in 60 of the specimens. There was a good agreement between the 2 assays in 87(81.3%) of the specimens. There was no statistical significant difference between the assays (p>0.05). Two of the 11 samples that RT-PCR negative Q-PCR positive, and 3 of 9 samples that RT-PCR positive Q-PCR negative were found to be CMV DNA positive by DNA sequencing. A good level of concordance between RT-PCR and Q-PCR assays for CMV DNA detection has been found. (author)

  20. Rapid analysis of Δ-9-tetrahydrocannabinol in hair using direct analysis in real time ambient ionization orbitrap mass spectrometry.

    Science.gov (United States)

    Duvivier, Wilco F; van Beek, Teris A; Pennings, Ed J M; Nielen, Michel W F

    2014-04-15

    Forensic hair analysis methods are laborious, time-consuming and provide only a rough retrospective estimate of the time of drug intake. Recently, hair imaging methods using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) were reported, but these methods require the application of MALDI matrix and are performed under vacuum. Direct analysis of entire locks of hair without any sample pretreatment and with improved spatial resolution would thus address a need. Hair samples were attached to stainless steel mesh screens and scanned in the X-direction using direct analysis in real time (DART) ambient ionization orbitrap MS. The DART gas temperature and the accuracy of the probed hair zone were optimized using Δ-9-tetrahydrocannabinol (THC) as a model compound. Since external contamination is a major issue in forensic hair analysis, sub-samples were measured before and after dichloromethane decontamination. The relative intensity of the THC signal in spiked blank hair versus that of quinine as the internal standard showed good reproducibility (26% RSD) and linearity of the method (R(2)  = 0.991). With the DART hair scan THC could be detected in hair samples from different chronic cannabis users. The presence of THC was confirmed by quantitative liquid chromatography/tandem mass spectrometry. Zones with different THC content could be clearly distinguished, indicating that the method might be used for retrospective timeline assessments. Detection of THC in decontaminated drug user hair showed that the DART hair scan not only probes THC on the surface of hair, but penetrates deeply enough to measure incorporated THC. A new approach in forensic hair analysis has been developed by probing complete locks of hair using DART-MS. Longitudinal scanning enables detection of incorporated compounds and can be used as pre-screening for THC without sample preparation. The method could also be adjusted for the analysis of other drugs of abuse. Copyright

  1. Tier-1 and Tier-2 real-time analysis experience in CMS Data Challenge 2004

    CERN Document Server

    De Filippis, N; Pierro, A; Silvestris, L; Fanfani, A; Grandi, C; Hernández, J M; Bonacorsi, D; Corvo, M; Fanzago, F

    2005-01-01

    During the CMS Data Challenge 2004 a real-time analysis was attempted at INFN and PIC Tier-1 and Tier-2s in order to test the ability of the instrumented methods to quickly process the data. Several agents and automatic procedures were implemented to perform the analysis at the Tier-1/2 synchronously with the data transfer from Tier-0 at CERN. The system was implemented in the LCG-2 Grid environment and allowed on-the-fly job preparation and subsequent submission to the Resource Broker as new data came along. Running job accessed data from the Storage Elements via remote file protocol, whenever possible, or copying them locally with replica manager commands. Details of the procedures adopted to run the analysis jobs and the expected results are described. An evaluation of the ability of the system to maintain an analysis rate at Tier-1 and Tier-2 comparable with the data transfer rate is also presented. The results on the analysis timeline, the statistics of submitted jobs, the overall efficiency of the GRID ...

  2. Rapid quality assessment of Radix Aconiti Preparata using direct analysis in real time mass spectrometry.

    Science.gov (United States)

    Zhu, Hongbin; Wang, Chunyan; Qi, Yao; Song, Fengrui; Liu, Zhiqiang; Liu, Shuying

    2012-11-08

    This study presents a novel and rapid method to identify chemical markers for the quality control of Radix Aconiti Preparata, a world widely used traditional herbal medicine. In the method, the samples with a fast extraction procedure were analyzed using direct analysis in real time mass spectrometry (DART MS) combined with multivariate data analysis. At present, the quality assessment approach of Radix Aconiti Preparata was based on the two processing methods recorded in Chinese Pharmacopoeia for the purpose of reducing the toxicity of Radix Aconiti and ensuring its clinical therapeutic efficacy. In order to ensure the safety and effectivity in clinical use, the processing degree of Radix Aconiti should be well controlled and assessed. In the paper, hierarchical cluster analysis and principal component analysis were performed to evaluate the DART MS data of Radix Aconiti Preparata samples in different processing times. The results showed that the well processed Radix Aconiti Preparata, unqualified processed and the raw Radix Aconiti could be clustered reasonably corresponding to their constituents. The loading plot shows that the main chemical markers having the most influence on the discrimination amongst the qualified and unqualified samples were mainly some monoester diterpenoid aconitines and diester diterpenoid aconitines, i.e. benzoylmesaconine, hypaconitine, mesaconitine, neoline, benzoylhypaconine, benzoylaconine, fuziline, aconitine and 10-OH-mesaconitine. The established DART MS approach in combination with multivariate data analysis provides a very flexible and reliable method for quality assessment of toxic herbal medicine. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Reachability analysis of real-time systems using time Petri nets.

    Science.gov (United States)

    Wang, J; Deng, Y; Xu, G

    2000-01-01

    Time Petri nets (TPNs) are a popular Petri net model for specification and verification of real-time systems. A fundamental and most widely applied method for analyzing Petri nets is reachability analysis. The existing technique for reachability analysis of TPNs, however, is not suitable for timing property verification because one cannot derive end-to-end delay in task execution, an important issue for time-critical systems, from the reachability tree constructed using the technique. In this paper, we present a new reachability based analysis technique for TPNs for timing property analysis and verification that effectively addresses the problem. Our technique is based on a concept called clock-stamped state class (CS-class). With the reachability tree generated based on CS-classes, we can directly compute the end-to-end time delay in task execution. Moreover, a CS-class can be uniquely mapped to a traditional state class based on which the conventional reachability tree is constructed. Therefore, our CS-class-based analysis technique is more general than the existing technique. We show how to apply this technique to timing property verification of the TPN model of a command and control (C2) system.

  4. Real-time gene expression analysis in carp (Cyprinus carpio) skin: inflammatory responses to injury mimicking infection with ectoparasites

    NARCIS (Netherlands)

    Gonzalez, S.F.; Huising, M.O.; Stakauska, R.; Forlenza, M.; Verburg-van Kemenade, B.M.L.; Buchmann, K.; Nielsen, M.E.; Wiegertjes, G.F.

    2007-01-01

    We studied a predictive model of gene expression induced by mechanical injury of fish skin, to resolve the confounding effects on the immune system induced by injury and skin parasite-specific molecules. We applied real time quantitative PCR (RQ-PCR) to measure the expression of the pro-inflammatory

  5. Real-time gene expression analysis in carp (Cyprinus carpio L.) skin: Inflammatory responses to injury mimicking infection with ectoparasites

    NARCIS (Netherlands)

    Gonzalez, S.F.; Huising, M.O.; Stakauskas, R.; Forlenza, M.; Verburg-van Kemenade, B.M.L.; Buchmann, K.; Nielsen, M.E.; Wiegertjes, G.F.

    2007-01-01

    We studied a predictive model of gene expression induced by mechanical injury of fish skin, to resolve the confounding effects on the immune system induced by injury and skin parasite-specific molecules. We applied real time quantitative PCR (RQ-PCR) to measure the expression of the pro-inflammatory

  6. DEVELOPMENT OF REAL-TIME MULTIPLEX PCR FOR THE QUANTITATIVE DETERMINATION OF TREC'S AND KREC'S IN WHOLE BLOOD AND IN DRIED BLOOD SPOTS

    Directory of Open Access Journals (Sweden)

    M. A. Gordukova

    2015-01-01

    Full Text Available Primary immunodeficiencies (PID such as severe combined immunodeficiency (SCID and X-linked agammaglobulinemia are characterized by the lack of functional Tand B-cells, respectively. Without early diagnosis and prompt treatment children with PID suffer from severe infectious diseases, leading to their death or disability. Our purpose was developing of simple, inexpensive, high throughput technique based on the quantitative determination of TREC and KREC molecules by real-time PCR, and its validation in a group of children with a verified diagnosis of SCID and X-linked agammaglobulinemia.In this study, we developed and validated multiplex real-time PCR for the TREC’s and KREC’s quantitative analysis. We have shown that linear range of Ct changes depending on the concentrations of targets with a correlation coefficient R2 not worse than 0.98 was observed at concentrations from 109 to 5 × 104 copies per ml. The lowest amount of targets reliably detected in a reaction volume was 10 TREC’s copies, 5 KREC ‘s copies and 5 copies of internal control (IL17RA. We determined the age-depended reference values of TRECs and KRECs in whole blood in 29 boys and 27 girls with normal immunological parameters. The normal cut-offs for TRECs and KRECs were defined in dry blood spots depending on the method of extraction.The proposed method showed 100% diagnostic sensitivity and specificity in the studied group. The method can be proposed as a screening tool for the diagnosis of SCID and X-linked agammaglobulinemia both in whole blood and in the dry blood spots. The further investigation is required with larger number of samples. 

  7. Strategy for Extracting DNA from Clay Soil and Detecting a Specific Target Sequence via Selective Enrichment and Real-Time (Quantitative) PCR Amplification ▿

    Science.gov (United States)

    Yankson, Kweku K.; Steck, Todd R.

    2009-01-01

    We present a simple strategy for isolating and accurately enumerating target DNA from high-clay-content soils: desorption with buffers, an optional magnetic capture hybridization step, and quantitation via real-time PCR. With the developed technique, μg quantities of DNA were extracted from mg samples of pure kaolinite and a field clay soil. PMID:19633108

  8. MIQE précis: Practical implementation of minimum standard guidelines for fluorescence-based quantitative real-time PCR experiments

    NARCIS (Netherlands)

    Bustin, S.A.; Beaulieu, J.F.; Huggett, J.; Jaggi, R.; Kibenge, F.S.; Olsvik, P.A.; Penning, L.C.; Toegel, S.

    2010-01-01

    MIQE précis: Practical implementation of minimum standard guidelines for fluorescence-based quantitative real-time PCR experiments Stephen A Bustin1 , Jean-François Beaulieu2 , Jim Huggett3 , Rolf Jaggi4 , Frederick SB Kibenge5 , Pål A Olsvik6 , Louis C Penning7 and Stefan Toegel8 1 Centre for

  9. A Human Fecal Contamination Score for Ranking Recreational Sites using the HF183/BacR287 Quantitative Real-Time PCR Method

    Science.gov (United States)

    Human fecal pollution of recreational waters remains a public health concern worldwide. As a result, there is a growing interest in the application of human-associated fecal source identification quantitative real-time PCR (qPCR) technologies for water quality research and manag...

  10. QUANTITATION OF INTRACELLULAR NAD(P)H IN LIVING CELLS CAN MONITOR AN IMBALANCE OF DNA SINGLE STRAND BREAK REPAIR IN REAL TIME

    Science.gov (United States)

    Quantitation of intracellular NAD(P)H in living cells can monitor an imbalance of DNA single strand break repair in real time.ABSTRACTDNA single strand breaks (SSBs) are one of the most frequent DNA lesions in genomic DNA generated either by oxidative stress or du...

  11. Direct analysis in real time-Mass spectrometry (DART-MS) in forensic and security applications.

    Science.gov (United States)

    Pavlovich, Matthew J; Musselman, Brian; Hall, Adam B

    2018-03-01

    Over the last decade, direct analysis in real time (DART) has emerged as a viable method for fast, easy, and reliable "ambient ionization" for forensic analysis. The ability of DART to generate ions from chemicals that might be present at the scene of a criminal activity, whether they are in the gas, liquid, or solid phase, with limited sample preparation has made the technology a useful analytical tool in numerous forensic applications. This review paper summarizes many of those applications, ranging from the analysis of trace evidence to security applications, with a focus on providing the forensic scientist with a resource for developing their own applications. The most common uses for DART in forensics are in studying seized drugs, drugs of abuse and their metabolites, bulk and detonated explosives, toxic chemicals, chemical warfare agents, inks and dyes, and commercial plant and animal products that have been adulterated for economic gain. This review is meant to complement recent reviews that have described the fundamentals of the ionization mechanism and the general use of DART. We describe a wide range of forensic applications beyond the field of analyzing drugs of abuse, which dominates the literature, including common experimental and data analysis methods. © 2016 Wiley Periodicals, Inc. Mass Spec Rev 37:171-187, 2018. © 2016 Wiley Periodicals, Inc.

  12. The Maximum Entropy Method for Optical Spectrum Analysis of Real-Time TDDFT

    International Nuclear Information System (INIS)

    Toogoshi, M; Kano, S S; Zempo, Y

    2015-01-01

    The maximum entropy method (MEM) is one of the key techniques for spectral analysis. The major feature is that spectra in the low frequency part can be described by the short time-series data. Thus, we applied MEM to analyse the spectrum from the time dependent dipole moment obtained from the time-dependent density functional theory (TDDFT) calculation in real time. It is intensively studied for computing optical properties. In the MEM analysis, however, the maximum lag of the autocorrelation is restricted by the total number of time-series data. We proposed that, as an improved MEM analysis, we use the concatenated data set made from the several-times repeated raw data. We have applied this technique to the spectral analysis of the TDDFT dipole moment of ethylene and oligo-fluorene with n = 8. As a result, the higher resolution can be obtained, which is closer to that of FT with practically time-evoluted data as the same total number of time steps. The efficiency and the characteristic feature of this technique are presented in this paper. (paper)

  13. O-5S quantitative real-time PCR: a new diagnostic tool for laboratory confirmation of human onchocerciasis.

    Science.gov (United States)

    Mekonnen, Solomon A; Beissner, Marcus; Saar, Malkin; Ali, Solomon; Zeynudin, Ahmed; Tesfaye, Kassahun; Adbaru, Mulatu G; Battke, Florian; Poppert, Sven; Hoelscher, Michael; Löscher, Thomas; Bretzel, Gisela; Herbinger, Karl-Heinz

    2017-10-02

    Onchocerciasis is a parasitic disease caused by the filarial nematode Onchocerca volvulus. In endemic areas, the diagnosis is commonly confirmed by microscopic examination of skin snip samples, though this technique is considered to have low sensitivity. The available melting-curve based quantitative real-time PCR (qPCR) using degenerated primers targeting the O-150 repeat of O. volvulus was considered insufficient for confirming the individual diagnosis, especially in elimination studies. This study aimed to improve detection of O. volvulus DNA in clinical samples through the development of a highly sensitive qPCR assay. A novel hydrolysis probe based qPCR assay was designed targeting the specific sequence of the O. volvulus O-5S rRNA gene. A total of 200 clinically suspected onchocerciasis cases were included from Goma district in South-west Ethiopia, from October 2012 through May 2013. Skin snip samples were collected and subjected to microscopy, O-150 qPCR, and the novel O-5S qPCR. Among the 200 individuals, 133 patients tested positive (positivity rate of 66.5%) and 67 negative by O-5S qPCR, 74 tested positive by microscopy (37.0%) and 78 tested positive by O-150 qPCR (39.0%). Among the 133 O-5S qPCR positive individuals, microscopy and O-150 qPCR detected 55.6 and 59.4% patients, respectively, implying a higher sensitivity of O-5S qPCR than microscopy and O-150 qPCR. None of the 67 individuals who tested negative by O-5S qPCR tested positive by microscopy or O-150 qPCR, implying 100% specificity of the newly designed O-5S qPCR assay. The novel O-5S qPCR assay is more sensitive than both microscopic examination and the existing O-150 qPCR for the detection of O. volvulus from skin snip samples. The newly designed assay is an important step towards appropriate individual diagnosis and control of onchocerciasis.

  14. A real-time digital control, data acquisition and analysis system for the DIII-D multipulse Thomson scattering diagnostic

    International Nuclear Information System (INIS)

    Greenfield, C.M.; Campbell, G.L.; Carlstrom, T.N.; DeBoo, J.C.; Hsieh, C.-L.; Snider, R.T.; Trost, P.K.

    1990-10-01

    A VME-based real-time computer systems for laser control, data acquisition and analysis for the DIII-D multipulse Thomson scattering diagnostic is described. The laser control task requires precise timing of up to 8 Nd:YAG lasers, each with an average firing rate of 20 Hz. A cpu module in real-time multiprocessing computer system will operate the lasers with evenly staggered laser pulses or in a ''burst mode'', where all available (fully charged) lasers can be fired at 50--100 μsec intervals upon receipt of an external event trigger signal. One of more cpu modules, along with a LeCroy FERA (Fast Encoding and Readout ADC) system, will perform real-time data acquisition and analysis. Partial electron temperature and density profiles will be available for plasma feedback control within 1 msec following each laser pulse. The VME-based computer system consists of 2 or more target processor modules (25 MHz Motorola 68030) running the VMEexec real-time operating system connected to a Unix based host system (also a 68030). All real-time software is fully interrupt driven to maximize system efficiency. Operator interaction and (non real-time) data analysis takes place on a MicroVAX 3400 connected via DECnet. 17 refs., 1 fig

  15. Real-time digital control, data acquisition, and analysis system for the DIII-D multipulse Thomson scattering diagnostic

    International Nuclear Information System (INIS)

    Greenfield, C.M.; Campbell, G.L.; Carlstrom, T.N.; DeBoo, J.C.; Hsieh, C.; Snider, R.T.; Trost, P.K.

    1990-01-01

    A VME-based real-time computer system for laser control, data acquisition, and analysis for the DIII-D multipulse Thomson scattering diagnostic is described. The laser control task requires precise timing of up to eight Nd:YAG lasers, each with an average firing rate of 20 Hz. A cpu module in a real-time multiprocessing computer system will operate the lasers with evenly staggered laser pulses or in a ''burst mode,'' where all available (fully charged) lasers can be fired at 50--100 μs intervals upon receipt of an external event trigger signal. One or more cpu modules, along with a LeCroy FERA (fast encoding and readout ADC) system, will perform real-time data acquisition and analysis. Partial electron temperature and density profiles will be available for plasma feedback control within 1 ms following each laser pulse. The VME-based computer system consists of two or more target processor modules (25 MHz Motorola 68030) running the VMEexec real-time operating system connected to a Unix-based host system (also a 68030). All real-time software is fully interrupt driven to maximize system efficiency. Operator interaction and (non-real-time) data analysis takes place on a MicroVAX 3400 connected via DECnet

  16. REAL-TIME ANALYSIS AND SELECTION BIASES IN THE SUPERNOVA LEGACY SURVEY

    International Nuclear Information System (INIS)

    Perrett, K.; Conley, A.; Carlberg, R.; Balam, D.; Hook, I. M.; Sullivan, M.; Pritchet, C.; Astier, P.; Balland, C.; Guy, J.; Hardin, D.; Pain, R.; Regnault, N.; Basa, S.; Fouchez, D.; Howell, D. A.

    2010-01-01

    The Supernova Legacy Survey (SNLS) has produced a high-quality, homogeneous sample of Type Ia supernovae (SNe Ia) out to redshifts greater than z = 1. In its first four years of full operation (to 2007 June), the SNLS discovered more than 3000 transient candidates, 373 of which have been spectroscopically confirmed as SNe Ia. Use of these SNe Ia in precision cosmology critically depends on an analysis of the observational biases incurred in the SNLS survey due to the incomplete sampling of the underlying SN Ia population. This paper describes our real-time supernova detection and analysis procedures, and uses detailed Monte Carlo simulations to examine the effects of Malmquist bias and spectroscopic sampling. Such sampling effects are found to become apparent at z ∼ 0.6, with a significant shift in the average magnitude of the spectroscopically confirmed SN Ia sample toward brighter values for z ∼> 0.75. We describe our approach to correct for these selection biases in our three-year SNLS cosmological analysis (SNLS3) and present a breakdown of the systematic uncertainties involved.

  17. Direct Analysis in Real Time Mass Spectrometry for Characterization of Large Saccharides.

    Science.gov (United States)

    Ma, Huiying; Jiang, Qing; Dai, Diya; Li, Hongli; Bi, Wentao; Da Yong Chen, David

    2018-03-06

    Polysaccharide characterization posts the most difficult challenge to available analytical technologies compared to other types of biomolecules. Plant polysaccharides are reported to have numerous medicinal values, but their effect can be different based on the types of plants, and even regions of productions and conditions of cultivation. However, the molecular basis of the differences of these polysaccharides is largely unknown. In this study, direct analysis in real time mass spectrometry (DART-MS) was used to generate polysaccharide fingerprints. Large saccharides can break down into characteristic small fragments in the DART source via pyrolysis, and the products are then detected by high resolution MS. Temperature was shown to be a crucial parameter for the decomposition of large polysaccharide. The general behavior of carbohydrates in DART-MS was also studied through the investigation of a number of mono- and oligosaccharide standards. The chemical formula and putative ionic forms of the fragments were proposed based on accurate mass with less than 10 ppm mass errors. Multivariate data analysis shows the clear differentiation of different plant species. Intensities of marker ions compared among samples also showed obvious differences. The combination of DART-MS analysis and mechanochemical extraction method used in this work demonstrates a simple, fast, and high throughput analytical protocol for the efficient evaluation of molecular features in plant polysaccharides.

  18. A Real-Time Analysis Method for Pulse Rate Variability Based on Improved Basic Scale Entropy

    Directory of Open Access Journals (Sweden)

    Yongxin Chou

    2017-01-01

    Full Text Available Base scale entropy analysis (BSEA is a nonlinear method to analyze heart rate variability (HRV signal. However, the time consumption of BSEA is too long, and it is unknown whether the BSEA is suitable for analyzing pulse rate variability (PRV signal. Therefore, we proposed a method named sliding window iterative base scale entropy analysis (SWIBSEA by combining BSEA and sliding window iterative theory. The blood pressure signals of healthy young and old subjects are chosen from the authoritative international database MIT/PhysioNet/Fantasia to generate PRV signals as the experimental data. Then, the BSEA and the SWIBSEA are used to analyze the experimental data; the results show that the SWIBSEA reduces the time consumption and the buffer cache space while it gets the same entropy as BSEA. Meanwhile, the changes of base scale entropy (BSE for healthy young and old subjects are the same as that of HRV signal. Therefore, the SWIBSEA can be used for deriving some information from long-term and short-term PRV signals in real time, which has the potential for dynamic PRV signal analysis in some portable and wearable medical devices.

  19. Real time spectroscopic ellipsometry for analysis and control of thin film polycrystalline semiconductor deposition in photovoltaics

    Energy Technology Data Exchange (ETDEWEB)

    Koirala, Prakash; Attygalle, Dinesh; Aryal, Puruswottam; Pradhan, Puja; Chen, Jie [Center for Photovoltaics Innovation and Commercialization and Department of Physics and Astronomy, University of Toledo, Toledo, OH 43606 (United States); Marsillac, Sylvain [Department of Electrical and Computer Engineering, Old Dominion University, Norfolk, VA 23529 (United States); Ferlauto, Andre S.; Podraza, Nikolas J.; Collins, Robert W. [Center for Photovoltaics Innovation and Commercialization and Department of Physics and Astronomy, University of Toledo, Toledo, OH 43606 (United States)

    2014-11-28

    Real time spectroscopic ellipsometry (RTSE) from the near-infrared to ultraviolet has been applied for analysis of the deposition of polycrystalline thin films that form the basis of two key photovoltaic heterojunction configurations, superstrate SnO{sub 2}/CdS/CdTe and substrate Mo/Cu(In{sub 1−x}Ga{sub x})Se{sub 2}/CdS. The focus of this work is to develop capabilities for monitoring and controlling the key steps in the fabrication of these device structures. Analysis of RTSE data collected during sputter deposition of CdS on a rough SnO{sub 2} transparent top contact provides the time evolution of the CdS effective thickness, or film volume per unit substrate area. This thickness includes interface, bulk, and surface roughness layer components and affects the CdS/CdTe heterojunction performance and the quantum efficiency of the solar cell in the blue region of the solar spectrum. Similarly, analysis of RTSE data collected during co-evaporation of Cu(In{sub 1−x}Ga{sub x})Se{sub 2} (CIGS; x ∼ 0.3) on a rough Mo back contact provides the evolution of a second phase of Cu{sub 2−x}Se within the CIGS layer. During the last stage of CIGS deposition, the In, Ga, and Se co-evaporants convert this Cu{sub 2−x}Se phase to CIGS, and RTSE identifies the endpoint, specifically the time at which complete conversion occurs and single-phase, large-grain CIGS is obtained in this key stage. - Highlights: • Real time spectroscopic ellipsometry (RTSE) study of CdS and CuIn{sub 1−x}Ga{sub x}Se{sub 2} (CIGS) films. • RTSE during CdS deposition provides the evolution of the CdS effective thickness. • RTSE for CIGS film enables to measure and control the composition and thickness. • The work leads to the development of optical models for processing steps.

  20. DAG expression: high-throughput gene expression analysis of real-time PCR data using standard curves for relative quantification.

    Directory of Open Access Journals (Sweden)

    María Ballester

    Full Text Available BACKGROUND: Real-time quantitative PCR (qPCR is still the gold-standard technique for gene-expression quantification. Recent technological advances of this method allow for the high-throughput gene-expression analysis, without the limitations of sample space and reagent used. However, non-commercial and user-friendly software for the management and analysis of these data is not available. RESULTS: The recently developed commercial microarrays allow for the drawing of standard curves of multiple assays using the same n-fold diluted samples. Data Analysis Gene (DAG Expression software has been developed to perform high-throughput gene-expression data analysis using standard curves for relative quantification and one or multiple reference genes for sample normalization. We discuss the application of DAG Expression in the analysis of data from an experiment performed with Fluidigm technology, in which 48 genes and 115 samples were measured. Furthermore, the quality of our analysis was tested and compared with other available methods. CONCLUSIONS: DAG Expression is a freely available software that permits the automated analysis and visualization of high-throughput qPCR. A detailed manual and a demo-experiment are provided within the DAG Expression software at http://www.dagexpression.com/dage.zip.

  1. A kinetic-based sigmoidal model for the polymerase chain reaction and its application to high-capacity absolute quantitative real-time PCR

    Directory of Open Access Journals (Sweden)

    Stewart Don

    2008-05-01

    Full Text Available Abstract Background Based upon defining a common reference point, current real-time quantitative PCR technologies compare relative differences in amplification profile position. As such, absolute quantification requires construction of target-specific standard curves that are highly resource intensive and prone to introducing quantitative errors. Sigmoidal modeling using nonlinear regression has previously demonstrated that absolute quantification can be accomplished without standard curves; however, quantitative errors caused by distortions within the plateau phase have impeded effective implementation of this alternative approach. Results Recognition that amplification rate is linearly correlated to amplicon quantity led to the derivation of two sigmoid functions that allow target quantification via linear regression analysis. In addition to circumventing quantitative errors produced by plateau distortions, this approach allows the amplification efficiency within individual amplification reactions to be determined. Absolute quantification is accomplished by first converting individual fluorescence readings into target quantity expressed in fluorescence units, followed by conversion into the number of target molecules via optical calibration. Founded upon expressing reaction fluorescence in relation to amplicon DNA mass, a seminal element of this study was to implement optical calibration using lambda gDNA as a universal quantitative standard. Not only does this eliminate the need to prepare target-specific quantitative standards, it relegates establishment of quantitative scale to a single, highly defined entity. The quantitative competency of this approach was assessed by exploiting "limiting dilution assay" for absolute quantification, which provided an independent gold standard from which to verify quantitative accuracy. This yielded substantive corroborating evidence that absolute accuracies of ± 25% can be routinely achieved. Comparison

  2. Real-Time Analysis of Beats in Music for Entertainment Robots

    OpenAIRE

    Yue-Der Lin; Ting-Tsao Wu; Yu-Ren Chen; Yen-Ting Lin; Wen-Hsiu Chen; Shih-Fan Wang; Jinghom Chakhap

    2012-01-01

    The dancing actions for entertainment robots are usually designed in advance and saved in a database according to the beats and rhythm of the given music. This research is devoted to developing a real-time algorithm that can detect the primary information of the music needed for the actions of entertainment robots. The computation of the proposed algorithm is very efficient and can satisfy the requirement of real-time processing by a digital signal controller. The digitized music signal is fi...

  3. A novel method to compensate for different amplification efficiencies between patient DNA samples in quantitative real-time PCR

    NARCIS (Netherlands)

    J.P.P. Meijerink (Jules); C. Mandigers; L. van de Locht; E. Tonnissen; F. Goodsaid; J. Raemaekers (John)

    2001-01-01

    textabstractQuantification of residual disease by real-time polymerase chain reaction (PCR) will become a pivotal tool in the development of patient-directed therapy. In recent years, various protocols to quantify minimal residual disease in leukemia or lymphoma patients have been

  4. A novel method to compensate for different amplification efficiencies between patient DNA samples in quantitative real-time PCR.

    NARCIS (Netherlands)

    Meijerink, J.P.P.; Mandigers, C.M.P.W.; Locht, A.T.F. van de; Tonnissen, E.L.R.T.M.; Goodsaid, F.; Raemaekers, J.M.M.

    2001-01-01

    Quantification of residual disease by real-time polymerase chain reaction (PCR) will become a pivotal tool in the development of patient-directed therapy. In recent years, various protocols to quantify minimal residual disease in leukemia or lymphoma patients have been developed. These assays assume

  5. Partial validation of a TaqMan real-time quantitative PCR for the detection of ranaviruses

    DEFF Research Database (Denmark)

    Stilwell, Natalie K.; Whittington, Richard J.; Hick, Paul M.

    2018-01-01

    Ranaviruses are globally emerging pathogens negatively impacting wild and cultured fish, amphibians, and reptiles. Although conventional and diagnostic real-time PCR (qPCR) assays have been developed to detect ranaviruses, these assays often have not been tested against the known diversity of ran...

  6. User satisfaction with a real-time automated feedback system for general practitioners: a quantitative and qualitative study

    NARCIS (Netherlands)

    Bindels, Rianne; Hasman, Arie; Derickx, Mieke; van Wersch, Jan W. J.; Winkens, Ron A. G.

    2003-01-01

    OBJECTIVE: The GRIF automated feedback system produces real-time comments on the appropriateness of diagnostic tests ordered by general practitioners (GPs) based on recommendations from accepted national and regional practice guidelines. We investigated the experiences of GPs with this system and,

  7. Two quantitative real-time PCR assays for the detection of penaeid shrimp and blue crab, crustacean shellfish allergens.

    Science.gov (United States)

    Eischeid, Anne C; Kim, Bang-hyun; Kasko, Sasha M

    2013-06-19

    Food allergen detection methods must be able to specifically detect minute quantities of an allergenic food in a complex food matrix. One technique that can be used is real-time PCR. For the work described here, real-time PCR assays were developed to detect penaeid shrimp and blue crab, crustacean shellfish allergens. The method was tested using shrimp meat and crab meat spiked into several types of foods, including canned soups, deli foods, meat, seafood, and prepared seafood products. Foods were spiked with either shrimp or crab at levels ranging from 0.1 to 10⁶ parts per million (ppm) and analyzed either raw or cooked by a variety of methods. Real-time PCR data were used to generate linear standard curves, and assays were evaluated with respect to linear range and reaction efficiency. Results indicate that both assays performed well in a variety of food types. High reaction efficiencies were achieved across a linear range of 6-8 orders of magnitude. Limits of detection were generally between 0.1 and 1 ppm. Cooking methods used to simulate thermal processing of foods had little effect on assay performance. This work demonstrates that real-time PCR can be a valuable tool in the detection of crustacean shellfish.

  8. Gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of Japanese encephalitis virus

    International Nuclear Information System (INIS)

    Huang, S-H; Tsai, M-H; Lin, C-W; Yang, T-C; Chuang, P-H; Tsai, I-S; Lu, H-C; Wan Lei; Lin, Y-J; Lai, C-H

    2008-01-01

    Virus isolation and antibody detection are routinely used for diagnosis of Japanese encephalitis virus (JEV) infection, but the low level of transient viremia in some JE patients makes JEV isolation from clinical and surveillance samples very difficult. We describe the use of gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of JEV from its RNA genome. We tested the effect of gold nanoparticles on four different PCR systems, including conventional PCR, reverse-transcription PCR (RT-PCR), and SYBR green real-time PCR and RT-PCR assays for diagnosis in the acute phase of JEV infection. Gold nanoparticles increased the amplification yield of the PCR product and shortened the PCR time compared to the conventional reaction. In addition, nanogold-based real-time RT-PCR showed a linear relationship between Ct and template amount using ten-fold dilutions of JEV. The nanogold-based RT-PCR and real-time quantitative RT-PCR assays were able to detect low levels (1-10 000 copies) of the JEV RNA genomes extracted from culture medium or whole blood, providing early diagnostic tools for the detection of low-level viremia in the acute-phase infection. The assays described here were simple, sensitive, and rapid approaches for detection and quantitation of JEV in tissue cultured samples as well as clinical samples

  9. Fast analysis of glycosides based on HKUST-1-coated monolith solid-phase microextraction and direct analysis in real-time mass spectrometry.

    Science.gov (United States)

    Li, Xianjiang; Wang, Xin; Ma, Wen; Ai, Wanpeng; Bai, Yu; Ding, Li; Liu, Huwei

    2017-04-01

    Glycosides are a kind of highly important natural aromatic precursors in tobacco leaves. In this study, a novel HKUST-1-coated monolith dip-it sampler was designed for the fast and sensitive analysis of trace glycosides using direct analysis in real-time mass spectrometry. This device was prepared in two steps: in situ polymerization of monolith in a glass capillary of dip-it and layer-by-layer growth of HKUST-1 on the surface of monolith. Sufficient extraction was realized by immersing the tip to solution and in situ desorption was carried out by plasma direct analysis in real time. Compared with traditional solid-phase microextraction protocols, sample desorption was not needed anymore, and only extraction conditions were needed to be optimized in this method, including the gas temperature of direct analysis in real time, extraction time, and CH 3 COONH 4 additive concentration. This method enabled the simultaneous detection of six kinds of glycosides with the limits of detection of 0.02-0.05 μg/mL and the linear ranges covering two orders of magnitude with the limits of quantitation of 0.05-0.1 μg/mL. Moreover, the developed method was applied for the glycosides analysis of three tobacco samples, which only took about 2 s for every sample. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Real-time analysis, visualization, and steering of microtomography experiments at photon sources

    International Nuclear Information System (INIS)

    Laszeski, G. von; Insley, J.A.; Foster, I.; Bresnahan, J.; Kesselman, C.; Su, M.; Thiebaux, M.; Rivers, M.L.; Wang, S.; Tieman, B.; McNulty, I.

    2000-01-01

    A new generation of specialized scientific instruments called synchrotron light sources allow the imaging of materials at very fine scales. However, in contrast to a traditional microscope, interactive use has not previously been possible because of the large amounts of data generated and the considerable computation required translating this data into a useful image. The authors describe a new software architecture that uses high-speed networks and supercomputers to enable quasi-real-time and hence interactive analysis of synchrotron light source data. This architecture uses technologies provided by the Globus computational grid toolkit to allow dynamic creation of a reconstruction pipeline that transfers data from a synchrotron source beamline to a preprocessing station, next to a parallel reconstruction system, and then to multiple visualization stations. Collaborative analysis tools allow multiple users to control data visualization. As a result, local and remote scientists can see and discuss preliminary results just minutes after data collection starts. The implications for more efficient use of this scarce resource and for more effective science appear tremendous

  11. Detection of Low Molecular Weight Adulterants in Beverages by Direct Analysis in Real Time Mass Spectrometry.

    Science.gov (United States)

    Sisco, Edward; Dake, Jeffrey

    2016-04-14

    Direct Analysis in Real Time Mass Spectrometry (DART-MS) has been used to detect the presence of non-narcotic adulterants in beverages. The non-narcotic adulterants that were examined in this work incorporated a number low molecular weight alcohols, acetone, ammonium hydroxide, and sodium hypochlorite. Analysis of the adulterants was completed by pipetting 1 µL deposits onto glass microcapillaries along with an appropriate dopant species followed by introduction into the DART gas stream. It was found that detection of these compounds in the complex matrices of common beverages (soda, energy drinks, etc.) was simplified through the use of a dopant species to allow for adduct formation with the desired compound(s) of interest. Other parameters that were investigated included DART gas stream temperature, in source collision induced dissociation, ion polarity, and DART needle voltage. Sensitivities of the technique were found to range from 0.001 % volume fraction to 0.1 % volume fraction, comparable to traditional analyses completed using headspace gas chromatography mass spectrometry (HS-GC/MS). Once a method was established using aqueous solutions, , fifteen beverages were spiked with each of the nine adulterants, to simulate real world detection, and in nearly all cases the adulterant could be detected either in pure form, or complexed with the added dopant species. This technique provides a rapid way to directly analyze beverages believed to be contaminated with non-narcotic adulterants at sensitivities similar to or exceeding those of traditional confirmatory analyses.

  12. Two-dimensional statistical linear discriminant analysis for real-time robust vehicle-type recognition

    Science.gov (United States)

    Zafar, I.; Edirisinghe, E. A.; Acar, S.; Bez, H. E.

    2007-02-01

    Automatic vehicle Make and Model Recognition (MMR) systems provide useful performance enhancements to vehicle recognitions systems that are solely based on Automatic License Plate Recognition (ALPR) systems. Several car MMR systems have been proposed in literature. However these approaches are based on feature detection algorithms that can perform sub-optimally under adverse lighting and/or occlusion conditions. In this paper we propose a real time, appearance based, car MMR approach using Two Dimensional Linear Discriminant Analysis that is capable of addressing this limitation. We provide experimental results to analyse the proposed algorithm's robustness under varying illumination and occlusions conditions. We have shown that the best performance with the proposed 2D-LDA based car MMR approach is obtained when the eigenvectors of lower significance are ignored. For the given database of 200 car images of 25 different make-model classifications, a best accuracy of 91% was obtained with the 2D-LDA approach. We use a direct Principle Component Analysis (PCA) based approach as a benchmark to compare and contrast the performance of the proposed 2D-LDA approach to car MMR. We conclude that in general the 2D-LDA based algorithm supersedes the performance of the PCA based approach.

  13. Real-Time EEG Signal Enhancement Using Canonical Correlation Analysis and Gaussian Mixture Clustering

    Directory of Open Access Journals (Sweden)

    Chin-Teng Lin

    2018-01-01

    Full Text Available Electroencephalogram (EEG signals are usually contaminated with various artifacts, such as signal associated with muscle activity, eye movement, and body motion, which have a noncerebral origin. The amplitude of such artifacts is larger than that of the electrical activity of the brain, so they mask the cortical signals of interest, resulting in biased analysis and interpretation. Several blind source separation methods have been developed to remove artifacts from the EEG recordings. However, the iterative process for measuring separation within multichannel recordings is computationally intractable. Moreover, manually excluding the artifact components requires a time-consuming offline process. This work proposes a real-time artifact removal algorithm that is based on canonical correlation analysis (CCA, feature extraction, and the Gaussian mixture model (GMM to improve the quality of EEG signals. The CCA was used to decompose EEG signals into components followed by feature extraction to extract representative features and GMM to cluster these features into groups to recognize and remove artifacts. The feasibility of the proposed algorithm was demonstrated by effectively removing artifacts caused by blinks, head/body movement, and chewing from EEG recordings while preserving the temporal and spectral characteristics of the signals that are important to cognitive research.

  14. Automated methods for real-time analysis of spent-fuel measurement data

    International Nuclear Information System (INIS)

    Bosler, G.E.; Rinard, P.M.; Klosterbuer, S.F.; Painter, J.

    1988-01-01

    Software has been developed for ''real-time'' analysis of neutron and gamma data from GRAND-1/fork measurements on spent-fuel assemblies. Three modules compose the software package. The modules are linked through a database system of files. The first module is part of a general data-base processing code. This module prepares input data files with inventory and correction-factor information for the second module. The second module, called OLAF, operates on a computer attached to the GRAND-1 electronics unit. In this second module, neutron and gamma data from spent-fuel assemblies are analyzed to verify consistency in the facility operator declarations for exposure (burnup) and cooling time. From the analysis, potential discrepancies in the measurement data are questioned while equipment is still installed at the facility and is available for additional measurements. During the measurements, data are written to an output file, called a results file, which can be processed by the third module of the software package. In the third module, printed reports summarizing the data and results are prepared, and neutron and gamma data are written to files that are process by the Deming curve-fitting code

  15. Real-time myoelectric control of a multi-fingered hand prosthesis using principal components analysis

    Directory of Open Access Journals (Sweden)

    Matrone Giulia C

    2012-06-01

    Full Text Available Abstract Background In spite of the advances made in the design of dexterous anthropomorphic hand prostheses, these sophisticated devices still lack adequate control interfaces which could allow amputees to operate them in an intuitive and close-to-natural way. In this study, an anthropomorphic five-fingered robotic hand, actuated by six motors, was used as a prosthetic hand emulator to assess the feasibility of a control approach based on Principal Components Analysis (PCA, specifically conceived to address this problem. Since it was demonstrated elsewhere that the first two principal components (PCs can describe the whole hand configuration space sufficiently well, the controller here employed reverted the PCA algorithm and allowed to drive a multi-DoF hand by combining a two-differential channels EMG input with these two PCs. Hence, the novelty of this approach stood in the PCA application for solving the challenging problem of best mapping the EMG inputs into the degrees of freedom (DoFs of the prosthesis. Methods A clinically viable two DoFs myoelectric controller, exploiting two differential channels, was developed and twelve able-bodied participants, divided in two groups, volunteered to control the hand in simple grasp trials, using forearm myoelectric signals. Task completion rates and times were measured. The first objective (assessed through one group of subjects was to understand the effectiveness of the approach; i.e., whether it is possible to drive the hand in real-time, with reasonable performance, in different grasps, also taking advantage of the direct visual feedback of the moving hand. The second objective (assessed through a different group was to investigate the intuitiveness, and therefore to assess statistical differences in the performance throughout three consecutive days. Results Subjects performed several grasp, transport and release trials with differently shaped objects, by operating the hand with the myoelectric

  16. Rapid quality assessment of Radix Aconiti Preparata using direct analysis in real time mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Zhu Hongbin; Wang Chunyan; Qi Yao [Changchun Center of Mass Spectrometry and Chemical Biology Laboratory, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022 (China); University of Chinese Academy of Sciences, Beijing 100039 (China); Song Fengrui, E-mail: songfr@ciac.jl.cn [Changchun Center of Mass Spectrometry and Chemical Biology Laboratory, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022 (China); Liu Zhiqiang; Liu Shuying [Changchun Center of Mass Spectrometry and Chemical Biology Laboratory, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun 130022 (China)

    2012-11-08

    Highlights: Black-Right-Pointing-Pointer DART MS combined with PCA and HCA was used to rapidly identify markers of Radix Aconiti. Black-Right-Pointing-Pointer The DART MS behavior of six aconitine-type alkaloids was investigated. Black-Right-Pointing-Pointer Chemical markers were recognized between the qualified and unqualified samples. Black-Right-Pointing-Pointer DART MS was shown to be an effective tool for quality control of Radix Aconiti Preparata. - Abstract: This study presents a novel and rapid method to identify chemical markers for the quality control of Radix Aconiti Preparata, a world widely used traditional herbal medicine. In the method, the samples with a fast extraction procedure were analyzed using direct analysis in real time mass spectrometry (DART MS) combined with multivariate data analysis. At present, the quality assessment approach of Radix Aconiti Preparata was based on the two processing methods recorded in Chinese Pharmacopoeia for the purpose of reducing the toxicity of Radix Aconiti and ensuring its clinical therapeutic efficacy. In order to ensure the safety and effectivity in clinical use, the processing degree of Radix Aconiti should be well controlled and assessed. In the paper, hierarchical cluster analysis and principal component analysis were performed to evaluate the DART MS data of Radix Aconiti Preparata samples in different processing times. The results showed that the well processed Radix Aconiti Preparata, unqualified processed and the raw Radix Aconiti could be clustered reasonably corresponding to their constituents. The loading plot shows that the main chemical markers having the most influence on the discrimination amongst the qualified and unqualified samples were mainly some monoester diterpenoid aconitines and diester diterpenoid aconitines, i.e. benzoylmesaconine, hypaconitine, mesaconitine, neoline, benzoylhypaconine, benzoylaconine, fuziline, aconitine and 10-OH-mesaconitine. The established DART MS approach in

  17. Rapid quality assessment of Radix Aconiti Preparata using direct analysis in real time mass spectrometry

    International Nuclear Information System (INIS)

    Zhu Hongbin; Wang Chunyan; Qi Yao; Song Fengrui; Liu Zhiqiang; Liu Shuying

    2012-01-01

    Highlights: ► DART MS combined with PCA and HCA was used to rapidly identify markers of Radix Aconiti. ► The DART MS behavior of six aconitine-type alkaloids was investigated. ► Chemical markers were recognized between the qualified and unqualified samples. ► DART MS was shown to be an effective tool for quality control of Radix Aconiti Preparata. - Abstract: This study presents a novel and rapid method to identify chemical markers for the quality control of Radix Aconiti Preparata, a world widely used traditional herbal medicine. In the method, the samples with a fast extraction procedure were analyzed using direct analysis in real time mass spectrometry (DART MS) combined with multivariate data analysis. At present, the quality assessment approach of Radix Aconiti Preparata was based on the two processing methods recorded in Chinese Pharmacopoeia for the purpose of reducing the toxicity of Radix Aconiti and ensuring its clinical therapeutic efficacy. In order to ensure the safety and effectivity in clinical use, the processing degree of Radix Aconiti should be well controlled and assessed. In the paper, hierarchical cluster analysis and principal component analysis were performed to evaluate the DART MS data of Radix Aconiti Preparata samples in different processing times. The results showed that the well processed Radix Aconiti Preparata, unqualified processed and the raw Radix Aconiti could be clustered reasonably corresponding to their constituents. The loading plot shows that the main chemical markers having the most influence on the discrimination amongst the qualified and unqualified samples were mainly some monoester diterpenoid aconitines and diester diterpenoid aconitines, i.e. benzoylmesaconine, hypaconitine, mesaconitine, neoline, benzoylhypaconine, benzoylaconine, fuziline, aconitine and 10-OH-mesaconitine. The established DART MS approach in combination with multivariate data analysis provides a very flexible and reliable method for quality

  18. The U-tube sampling methodology and real-time analysis of geofluids

    International Nuclear Information System (INIS)

    Freifeld, Barry; Perkins, Ernie; Underschultz, James; Boreham, Chris

    2009-01-01

    The U-tube geochemical sampling methodology, an extension of the porous cup technique proposed by Wood (1973), provides minimally contaminated aliquots of multiphase fluids from deep reservoirs and allows for accurate determination of dissolved gas composition. The initial deployment of the U-tube during the Frio Brine Pilot CO 2 storage experiment, Liberty County, Texas, obtained representative samples of brine and supercritical CO 2 from a depth of 1.5 km. A quadrupole mass spectrometer provided real-time analysis of dissolved gas composition. Since the initial demonstration, the U-tube has been deployed for (1) sampling of fluids down gradient of the proposed Yucca Mountain High-Level Waste Repository, Armagosa Valley, Nevada (2) acquiring fluid samples beneath permafrost in Nunuvut Territory, Canada, and (3) at a CO 2 storage demonstration project within a depleted gas reservoir, Otway Basin, Victoria, Australia. The addition of in-line high-pressure pH and EC sensors allows for continuous monitoring of fluid during sample collection. Difficulties have arisen during U-tube sampling, such as blockage of sample lines from naturally occurring waxes or from freezing conditions; however, workarounds such as solvent flushing or heating have been used to address these problems. The U-tube methodology has proven to be robust, and with careful consideration of the constraints and limitations, can provide high quality geochemical samples.

  19. Expression analysis of fiber related genes in cotton (gossypium hirsutum l.) through real time pcr

    International Nuclear Information System (INIS)

    Iqbal, N.; Khatoon, A.; Asif, M.; Bashir, A.

    2016-01-01

    Cotton fibers are unicellular seed trichomes and the largest known plant cells. Fiber morphogenesis in cotton is a complex process involving a large number of genes expressed throughout fiber development process. The expression profiling of five gene families in various cotton tissues was carried out through real time PCR. Expression analysis revealed that transcripts of expansin, tubulin and E6 were elevated from 5 to 20 days post anthesis (DPA) fibers. Three Lipid transfer proteins (LTPs) including LTP1, LTP3, LTP7 exhibited highest expression in 10 - 20 DPA fibers. Transcripts of LTP3 were detected in fibers and non fiber tissues that of LTP7 were almost negligible in non fiber tissues. Sucrose phosphate synthase gene showed highest expression in 10 DPA fibers while sucrose synthse (susy) expressed at higher rate in 5-20 DPA fibers as well as roots. The results reveal that most of fiber related genes showed high expression in 5-20 DPA fibers. Comprehensive expression study may help to determine tissue and stage specificity of genes under study. The study may also help to explore complex process of fiber development and understand the role of these genes in fiber development process. Highly expressed genes in fibers may be transformed in cotton for improvement of fiber quality traits. Genes that were expressed specifically in fibers or other tissues could be used for isolation of upstream regulatory sequences. (author)

  20. Compositional schedulability analysis of real-time actor-based systems.

    Science.gov (United States)

    Jaghoori, Mohammad Mahdi; de Boer, Frank; Longuet, Delphine; Chothia, Tom; Sirjani, Marjan

    2017-01-01

    We present an extension of the actor model with real-time, including deadlines associated with messages, and explicit application-level scheduling policies, e.g.,"earliest deadline first" which can be associated with individual actors. Schedulability analysis in this setting amounts to checking whether, given a scheduling policy for each actor, every task is processed within its designated deadline. To check schedulability, we introduce a compositional automata-theoretic approach, based on maximal use of model checking combined with testing. Behavioral interfaces define what an actor expects from the environment, and the deadlines for messages given these assumptions. We use model checking to verify that actors match their behavioral interfaces. We extend timed automata refinement with the notion of deadlines and use it to define compatibility of actor environments with the behavioral interfaces. Model checking of compatibility is computationally hard, so we propose a special testing process. We show that the analyses are decidable and automate the process using the Uppaal model checker.

  1. Real-time systems

    OpenAIRE

    Badr, Salah M.; Bruztman, Donald P.; Nelson, Michael L.; Byrnes, Ronald Benton

    1992-01-01

    This paper presents an introduction to the basic issues involved in real-time systems. Both real-time operating sys and real-time programming languages are explored. Concurrent programming and process synchronization and communication are also discussed. The real-time requirements of the Naval Postgraduate School Autonomous Under Vehicle (AUV) are then examined. Autonomous underwater vehicle (AUV), hard real-time system, real-time operating system, real-time programming language, real-time sy...

  2. Performance of real-time elastography for the staging of hepatic fibrosis: a meta-analysis.

    Directory of Open Access Journals (Sweden)

    Huisuo Hong

    Full Text Available BACKGROUND: With the rapid development of real-time elastography (RTE, a variety of measuring methods have been developed for the assessment of hepatic fibrosis. We evaluated the overall performance of four methods based on RTE by performing meta-analysis of published literature. METHODS: Online journal databases and a manual search from April 2000 to April 2014 were used. Studies from different databases that meet inclusion criteria were enrolled. The statistical analysis was performed using a random-effects model and fixed-effects model for the overall effectiveness of RTE. The area under the receiver operating characteristic curve (AUROC was calculated for various means. Fagan plot analysis was used to estimate the clinical utility of RTE, and the heterogeneity of the studies was explored with meta-regression analysis. RESULTS: Thirteen studies from published articles were enrolled and analyzed. The combined AUROC of the liver fibrosis index (LFI for the evaluation of significant fibrosis (F≥2, advanced fibrosis (F≥3, and cirrhosis (F = 4 were 0.79, 0.94, and 0.85, respectively. The AUROC of the elasticity index (EI ranged from 0.75 to 0.92 for F≥2 and 0.66 to 0.85 for F = 4. The overall AUROC of the elastic ratio of the liver for the intrahepatic venous vessels were 0.94, 0.93, and 0.96, respectively. The AUROC of the elastic ratio of the liver for the intercostal muscle in diagnosing advanced fibrosis and cirrhosis were 0.96 and 0.92, respectively. There was significant heterogeneity in the diagnostic odds ratio (DOR for F≥2 of LFI mainly due to etiology (p<0.01. CONCLUSION: The elastic ratio of the liver for the intrahepatic vein has excellent precision in differentiating each stage of hepatic fibrosis and is recommend to be applied to the clinic.

  3. [Construction and analysis of a monitoring system with remote real-time multiple physiological parameters based on cloud computing].

    Science.gov (United States)

    Zhu, Lingyun; Li, Lianjie; Meng, Chunyan

    2014-12-01

    There have been problems in the existing multiple physiological parameter real-time monitoring system, such as insufficient server capacity for physiological data storage and analysis so that data consistency can not be guaranteed, poor performance in real-time, and other issues caused by the growing scale of data. We therefore pro posed a new solution which was with multiple physiological parameters and could calculate clustered background data storage and processing based on cloud computing. Through our studies, a batch processing for longitudinal analysis of patients' historical data was introduced. The process included the resource virtualization of IaaS layer for cloud platform, the construction of real-time computing platform of PaaS layer, the reception and analysis of data stream of SaaS layer, and the bottleneck problem of multi-parameter data transmission, etc. The results were to achieve in real-time physiological information transmission, storage and analysis of a large amount of data. The simulation test results showed that the remote multiple physiological parameter monitoring system based on cloud platform had obvious advantages in processing time and load balancing over the traditional server model. This architecture solved the problems including long turnaround time, poor performance of real-time analysis, lack of extensibility and other issues, which exist in the traditional remote medical services. Technical support was provided in order to facilitate a "wearable wireless sensor plus mobile wireless transmission plus cloud computing service" mode moving towards home health monitoring for multiple physiological parameter wireless monitoring.

  4. Development of real-time plasma analysis and control algorithms for the TCV tokamak using SIMULINK

    International Nuclear Information System (INIS)

    Felici, F.; Le, H.B.; Paley, J.I.; Duval, B.P.; Coda, S.; Moret, J.-M.; Bortolon, A.; Federspiel, L.; Goodman, T.P.; Hommen, G.; Karpushov, A.; Piras, F.; Pitzschke, A.; Romero, J.; Sevillano, G.; Sauter, O.; Vijvers, W.

    2014-01-01

    Highlights: • A new digital control system for the TCV tokamak has been commissioned. • The system is entirely programmable by SIMULINK, allowing rapid algorithm development. • Different control system nodes can run different algorithms at varying sampling times. • The previous control system functions have been emulated and improved. • New capabilities include MHD control, profile control, equilibrium reconstruction. - Abstract: One of the key features of the new digital plasma control system installed on the TCV tokamak is the possibility to rapidly design, test and deploy real-time algorithms. With this flexibility the new control system has been used for a large number of new experiments which exploit TCV's powerful actuators consisting of 16 individually controllable poloidal field coils and 7 real-time steerable electron cyclotron (EC) launchers. The system has been used for various applications, ranging from event-based real-time MHD control to real-time current diffusion simulations. These advances have propelled real-time control to one of the cornerstones of the TCV experimental program. Use of the SIMULINK graphical programming language to directly program the control system has greatly facilitated algorithm development and allowed a multitude of different algorithms to be deployed in a short time. This paper will give an overview of the developed algorithms and their application in physics experiments

  5. Development of Quantitative Competitive PCR and Absolute Based Real-Time PCR Assays for Quantification of The Butyrate Producing Bacterium: Butyrivibrio fibrisolvens

    Directory of Open Access Journals (Sweden)

    Mojtaba Tahmoorespur

    2016-04-01

    Full Text Available Introduction Butyrivibrio fibrisolvens strains are presently recognized as the major butyrate-producing bacteria found in the rumen and digestive track of many animals and also in the human gut. In this study we reported the development of two DNA based techniques, quantitative competitive (QC PCR and absolute based Real-Time PCR, for enumerating Butyrivibrio fibrisolvens strains. Despite the recent introduction of real-time PCR method for the rapid quantification of the target DNA sequences, use of quantitative competitive PCR (QC-PCR technique continues to play an important role in nucleic acid quantification since it is more cost effective. The procedure relies on the co-amplification of the sequence of interest with a serially diluted synthetic DNA fragment of the known concentration (competitor, using the single set primers. A real-time polymerase chain reaction is a laboratory technique of molecular biology based on the polymerase chain reaction (PCR. It monitors the amplification of a targeted DNA molecule during the PCR. Materials and Methods At first reported species-specific primers targeting the 16S rDNA region of the bacterium Butyrivibrio fibrisolvens were used for amplifying a 213 bp fragment. A DNA competitor differing by 50 bp in length from the 213 bp fragment was constructed and cloned into pTZ57R/T vector. The competitor was quantified by NanoDrop spectrophotometer and serially diluted and co-amplified by PCR with total extracted DNA from rumen fluid samples. PCR products were quantified by photographing agarose gels and analyzed with Image J software and the amount of amplified target DNA was log plotted against the amount of amplified competitor. Coefficient of determination (R2 was used as a criterion of methodology precision. For developing the Real-time PCR technique, the 213 bp fragment was amplified and cloned into pTZ57R/T was used to draw a standard curve. Results and Discussion The specific primers of Butyrivibrio

  6. Development Status of the WetLab-2 Project: New Tools for On-orbit Real-time Quantitative Gene Expression.

    Science.gov (United States)

    Jung, Jimmy; Parra, Macarena P.; Almeida, Eduardo; Boone, Travis; Chinn, Tori; Ricco, Antonio; Souza, Kenneth; Hyde, Liz; Rukhsana, Yousuf; Richey, C. Scott

    2013-01-01

    The primary objective of NASA Ames Research Centers WetLab-2 Project is to place on the ISS a research platform to facilitate gene expression analysis via quantitative real-time PCR (qRT-PCR) of biological specimens grown or cultured on orbit. The WetLab-2 equipment will be capable of processing multiple sample types ranging from microbial cultures to animal tissues dissected on-orbit. In addition to the logistical benefits of in-situ sample processing and analysis, conducting qRT-PCR on-orbit eliminates the confounding effects on gene expression of reentry stresses and shock acting on live cells and organisms. The system can also validate terrestrial analyses of samples returned from ISS by providing quantitative on-orbit gene expression benchmarking prior to sample return. The ability to get on orbit data will provide investigators with the opportunity to adjust experimental parameters for subsequent trials based on the real-time data analysis without need for sample return and re-flight. Finally, WetLab-2 can be used for analysis of air, surface, water, and clinical samples to monitor environmental contaminants and crew health. The verification flight of the instrument is scheduled to launch on SpaceX-5 in Aug. 2014.Progress to date: The WetLab-2 project completed a thorough study of commercially available qRT-PCR systems and performed a downselect based on both scientific and engineering requirements. The selected instrument, the Cepheid SmartCycler, has advantages including modular design (16 independent PCR modules), low power consumption, and rapid ramp times. The SmartCycler has multiplex capabilities, assaying up to four genes of interest in each of the 16 modules. The WetLab-2 team is currently working with Cepheid to modify the unit for housing within an EXPRESS rack locker on the ISS. This will enable the downlink of data to the ground and provide uplink capabilities for programming, commanding, monitoring, and instrument maintenance. The project is

  7. PRIMAS: a real-time 3D motion-analysis system

    Science.gov (United States)

    Sabel, Jan C.; van Veenendaal, Hans L. J.; Furnee, E. Hans

    1994-03-01

    The paper describes a CCD TV-camera-based system for real-time multicamera 2D detection of retro-reflective targets and software for accurate and fast 3D reconstruction. Applications of this system can be found in the fields of sports, biomechanics, rehabilitation research, and various other areas of science and industry. The new feature of real-time 3D opens an even broader perspective of application areas; animations in virtual reality are an interesting example. After presenting an overview of the hardware and the camera calibration method, the paper focuses on the real-time algorithms used for matching of the images and subsequent 3D reconstruction of marker positions. When using a calibrated setup of two cameras, it is now possible to track at least ten markers at 100 Hz. Limitations in the performance are determined by the visibility of the markers, which could be improved by adding a third camera.

  8. St. Patrick's Day 2015 geomagnetic storm analysis based on Real Time Ionosphere Monitoring

    Science.gov (United States)

    García-Rigo, Alberto

    2017-04-01

    Ionosphere Monitoring (RTIM) is a new Working Group within the International Association of Geodesy (IAG) Sub-Commission 4.3 "Atmosphere Remote Sensing". The complementary expertise of the participating research groups allows to analyse the ionospheric behaviour from a broad perspective, taking benefit of comparing multiple independent real time and near real time ionospheric approaches. In this context, a detailed analysis will be presented for the days in March, 2015 surrounding St. Patrick's Day 2015 geomagnetic storm, based on the existing ionospheric models (global or regional) within the group, which are mainly based on Global Navigation Satellite Systems (GNSS) and ionosonde data. For this purpose, a variety of ionospheric parameters will be considered, including Total Electron Content (TEC), F2 layer critical frequency (foF2), F2 layer peak (hmF2), bottomside half-thickness (B0) and ionospheric disturbance W-index. Also, ionospheric high-frequency perturbations such as Travelling Ionospheric Disturbances (TIDs), scintillations and the impact of solar flares facing the Earth will be presented to derive a clear picture of the ionospheric dynamics. Among other sources of information to take part in the comparisons, there will be (1) scintillation results -from MONITOR ESA/ESTEC-funded project- derived by means of S4 index and Sigma Phi (IEEA), specially significant in the African sector and European high latitudes, (2) dynamics of the global maps of W-index with 1h resolution derived from JPL Global Ionospheric Maps (GIMs; IZMIRAN), (3) deviations from expected quiet-time behavior analysed in terms of foF2, hmF2, B0 and B1 based on IRTAM and GIRO network of digisondes (Lowell), showing F2 layer peculiar changes due to the storm, (4) statistics based on the median of the VTEC for the 15 previous days considering VTEC european regional maps (ROB), (5) time series of VTEC data that are derived by running the NRT ionosphere model of DGFI-TUM in offline mode, which show

  9. Feasibility of iFlow color-coding technique in quantitative real-time measurement of hemodynamic changes after transarterial chemoembolization for hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    WANG Yuzhe

    2018-01-01

    Full Text Available Objective To investigate the value of iFlow color-coding technique in quantitative real-time analysis of hemodynamic changes after transarterial chemoembolization (TACE for hepatocellular carcinoma (HCC. Methods A total of 31 patients who were diagnosed with HCC in Shanghai Fifth People′s Hospital from December 2015 to January 2017 were enrolled. No patient underwent surgical operation or ablation. All patients underwent TACE with the same contrast agent, high-pressure injector parameters, and place of angiographic catheter. The iFlow technique was used to generate two-dimensional color-coded images and time-density curve (TDC before and after surgery and measure the opening of the angiographic catheter and the time to peak (TTP of the starting and ending points of the major tumor feeding arteries, as well as the ratio of the areas under the curve (AUC of TDC of tumor tissue and the opening of the angiographic catheter. The paired t-test was used for comparison of continuous data between groups. Results TTP of the major tumor feeding arteries was 4.64±0.49 s before TACE and 5.97±0.84 s after TACE (t=11.57, P<0.01, and there was a significant difference in AUC between the tumor tissue and the opening of the angiographic catheter (0.53±0.15 vs 0.16±0.12, t=25.85, P<0.01. There was no significant difference in TTP between the opening of the angiographic catheter and the major tumor feeding arteries before and after TACE (P>0.05. Before TACE, the TDC of tumor feeding arteries had a shape of “rapid increase-rapid reduction” with relatively high slope and peak value, while after TACE, the TDC had a shape of “increase-flat-reduction” with reductions in slope and peak value. Conclusion The iFlow technique can perform real-time measurement of TTP and TDC of the region of interest and helps with quantitative evaluation of hemodynamic changes in HCC. Therefore, it can provide objective quantitative indices for evaluating the degree of

  10. Real-time face and gesture analysis for human-robot interaction

    Science.gov (United States)

    Wallhoff, Frank; Rehrl, Tobias; Mayer, Christoph; Radig, Bernd

    2010-05-01

    Human communication relies on a large number of different communication mechanisms like spoken language, facial expressions, or gestures. Facial expressions and gestures are one of the main nonverbal communication mechanisms and pass large amounts of information between human dialog partners. Therefore, to allow for intuitive human-machine interaction, a real-time capable processing and recognition of facial expressions, hand and head gestures are of great importance. We present a system that is tackling these challenges. The input features for the dynamic head gestures and facial expressions are obtained from a sophisticated three-dimensional model, which is fitted to the user in a real-time capable manner. Applying this model different kinds of information are extracted from the image data and afterwards handed over to a real-time capable data-transferring framework, the so-called Real-Time DataBase (RTDB). In addition to the head and facial-related features, also low-level image features regarding the human hand - optical flow, Hu-moments are stored into the RTDB for the evaluation process of hand gestures. In general, the input of a single camera is sufficient for the parallel evaluation of the different gestures and facial expressions. The real-time capable recognition of the dynamic hand and head gestures are performed via different Hidden Markov Models, which have proven to be a quick and real-time capable classification method. On the other hand, for the facial expressions classical decision trees or more sophisticated support vector machines are used for the classification process. These obtained results of the classification processes are again handed over to the RTDB, where other processes (like a Dialog Management Unit) can easily access them without any blocking effects. In addition, an adjustable amount of history can be stored by the RTDB buffer unit.

  11. Automated multivariate analysis of multi-sensor data submitted online: Real-time environmental monitoring.

    Science.gov (United States)

    Eide, Ingvar; Westad, Frank

    2018-01-01

    A pilot study demonstrating real-time environmental monitoring with automated multivariate analysis of multi-sensor data submitted online has been performed at the cabled LoVe Ocean Observatory located at 258 m depth 20 km off the coast of Lofoten-Vesterålen, Norway. The major purpose was efficient monitoring of many variables simultaneously and early detection of changes and time-trends in the overall response pattern before changes were evident in individual variables. The pilot study was performed with 12 sensors from May 16 to August 31, 2015. The sensors provided data for chlorophyll, turbidity, conductivity, temperature (three sensors), salinity (calculated from temperature and conductivity), biomass at three different depth intervals (5-50, 50-120, 120-250 m), and current speed measured in two directions (east and north) using two sensors covering different depths with overlap. A total of 88 variables were monitored, 78 from the two current speed sensors. The time-resolution varied, thus the data had to be aligned to a common time resolution. After alignment, the data were interpreted using principal component analysis (PCA). Initially, a calibration model was established using data from May 16 to July 31. The data on current speed from two sensors were subject to two separate PCA models and the score vectors from these two models were combined with the other 10 variables in a multi-block PCA model. The observations from August were projected on the calibration model consecutively one at a time and the result was visualized in a score plot. Automated PCA of multi-sensor data submitted online is illustrated with an attached time-lapse video covering the relative short time period used in the pilot study. Methods for statistical validation, and warning and alarm limits are described. Redundant sensors enable sensor diagnostics and quality assurance. In a future perspective, the concept may be used in integrated environmental monitoring.

  12. Automated multivariate analysis of multi-sensor data submitted online: Real-time environmental monitoring.

    Directory of Open Access Journals (Sweden)

    Ingvar Eide

    Full Text Available A pilot study demonstrating real-time environmental monitoring with automated multivariate analysis of multi-sensor data submitted online has been performed at the cabled LoVe Ocean Observatory located at 258 m depth 20 km off the coast of Lofoten-Vesterålen, Norway. The major purpose was efficient monitoring of many variables simultaneously and early detection of changes and time-trends in the overall response pattern before changes were evident in individual variables. The pilot study was performed with 12 sensors from May 16 to August 31, 2015. The sensors provided data for chlorophyll, turbidity, conductivity, temperature (three sensors, salinity (calculated from temperature and conductivity, biomass at three different depth intervals (5-50, 50-120, 120-250 m, and current speed measured in two directions (east and north using two sensors covering different depths with overlap. A total of 88 variables were monitored, 78 from the two current speed sensors. The time-resolution varied, thus the data had to be aligned to a common time resolution. After alignment, the data were interpreted using principal component analysis (PCA. Initially, a calibration model was established using data from May 16 to July 31. The data on current speed from two sensors were subject to two separate PCA models and the score vectors from these two models were combined with the other 10 variables in a multi-block PCA model. The observations from August were projected on the calibration model consecutively one at a time and the result was visualized in a score plot. Automated PCA of multi-sensor data submitted online is illustrated with an attached time-lapse video covering the relative short time period used in the pilot study. Methods for statistical validation, and warning and alarm limits are described. Redundant sensors enable sensor diagnostics and quality assurance. In a future perspective, the concept may be used in integrated environmental monitoring.

  13. GENEASE: Real time bioinformatics tool for multi-omics and disease ontology exploration, analysis and visualization.

    Science.gov (United States)

    Ghandikota, Sudhir; Hershey, Gurjit K Khurana; Mersha, Tesfaye B

    2018-03-24

    Advances in high-throughput sequencing technologies have made it possible to generate multiple omics data at an unprecedented rate and scale. The accumulation of these omics data far outpaces the rate at which biologists can mine and generate new hypothesis to test experimentally. There is an urgent need to develop a myriad of powerful tools to efficiently and effectively search and filter these resources to address specific post-GWAS functional genomics questions. However, to date, these resources are scattered across several databases and often lack a unified portal for data annotation and analytics. In addition, existing tools to analyze and visualize these databases are highly fragmented, resulting researchers to access multiple applications and manual interventions for each gene or variant in an ad hoc fashion until all the questions are answered. In this study, we present GENEASE, a web-based one-stop bioinformatics tool designed to not only query and explore multi-omics and phenotype databases (e.g., GTEx, ClinVar, dbGaP, GWAS Catalog, ENCODE, Roadmap Epigenomics, KEGG, Reactome, Gene and Phenotype Ontology) in a single web interface but also to perform seamless post genome-wide association downstream functional and overlap analysis for non-coding regulatory variants. GENEASE accesses over 50 different databases in public domain including model organism-specific databases to facilitate gene/variant and disease exploration, enrichment and overlap analysis in real time. It is a user-friendly tool with point-and-click interface containing links for support information including user manual and examples. GENEASE can be accessed freely at http://research.cchmc.org/mershalab/genease_new/login.html. Tesfaye.Mersha@cchmc.org, Sudhir.Ghandikota@cchmc.org. Supplementary data are available at Bioinformatics online.

  14. Improving practices in nanomedicine through near real-time pharmacokinetic analysis

    Science.gov (United States)

    Magafia, Isidro B.

    More than a decade into the development of gold nanoparticles, with multiple clinical trials underway, ongoing pre-clinical research continues towards better understanding in vivo interactions. The goal is treatment optimization through improved best practices. In an effort to collect information for healthcare providers enabling informed decisions in a relevant time frame, instrumentation for real-time plasma concentration (multi-wavelength photoplethysmography) and protocols for rapid elemental analysis (energy dispersive X-Ray fluorescence) of biopsied tumor tissue have been developed in a murine model. An initial analysis, designed to demonstrate the robust nature and utility of the techniques, revealed that area under the bioavailability curve (AUC) alone does not currently inform tumor accumulation with a high degree of accuracy (R2=0.56), marginally better than injected dose (R2=0.46). This finding suggests that the control of additional experimental and physiological variables (chosen through modeling efforts) may yield more predictable tumor accumulation. Subject core temperature, blood pressure, and tumor perfusion are evaluated relative to particle uptake in a murine tumor model. New research efforts are also focused on adjuvant therapies that are employed to modify circulation parameters, including the AUC, of nanorods and gold nanoshells. Preliminary studies demonstrated a greater than 300% increase in average AUC using a reticuloendothelial blockade agent versus control groups. Given a better understanding of the relative importance of the physiological factors that influence rates of tumor accumulation, a set of experimental best practices is presented. This dissertation outlines the experimental protocols conducted, and discusses the real-world needs discovered and how these needs became specifications of developed protocols.

  15. Application of quantitative real-time PCR compared to filtration methods for the enumeration of Escherichia coli in surface waters within Vietnam.

    Science.gov (United States)

    Vital, Pierangeli G; Van Ha, Nguyen Thi; Tuyet, Le Thi Hong; Widmer, Kenneth W

    2017-02-01

    Surface water samples in Vietnam were collected from the Saigon River, rural and suburban canals, and urban runoff canals in Ho Chi Minh City, Vietnam, and were processed to enumerate Escherichia coli. Quantification was done through membrane filtration and quantitative real-time polymerase chain reaction (PCR). Mean log colony-forming unit (CFU)/100 ml E. coli counts in the dry season for river/suburban canals and urban canals were log 2.8 and 3.7, respectively, using a membrane filtration method, while using Taqman quantitative real-time PCR they were log 2.4 and 2.8 for river/suburban canals and urban canals, respectively. For the wet season, data determined by the membrane filtration method in river/suburban canals and urban canals samples had mean counts of log 3.7 and 4.1, respectively. While mean log CFU/100 ml counts in the wet season using quantitative PCR were log 3 and 2, respectively. Additionally, the urban canal samples were significantly lower than those determined by conventional culture methods for the wet season. These results show that while quantitative real-time PCR can be used to determine levels of fecal indicator bacteria in surface waters, there are some limitations to its application and it may be impacted by sources of runoff based on surveyed samples.

  16. Real-time systems design and analysis tools for the practitioner

    CERN Document Server

    Laplante, Phillip A

    2012-01-01

    An important resource, this book offers an introduction and overview of real-time systems: systems where timeliness is a crucial part of the correctness of the system. It contains a pragmatic overview of key topics (computer architecture and organization, operating systems, software engineering, programming languages, and compiler theory) from the perspective of the real-time systems designer and is organized into chapters that are essentially self-contained. In addition, each chapter contains both basic and more challenging exercises that will help the reader to confront actual problems.

  17. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods.

    Science.gov (United States)

    Edagawa, Akiko; Kimura, Akio; Kawabuchi-Kurata, Takako; Adachi, Shinichi; Furuhata, Katsunori; Miyamoto, Hiroshi

    2015-10-19

    We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR), and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%). Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%). In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8%) compared with real-time qPCR alone (46/68, 67.6%). Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1%) compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%). Legionella was not detected in the remaining six samples (6/68, 8.8%), irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  18. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods

    Directory of Open Access Journals (Sweden)

    Akiko Edagawa

    2015-10-01

    Full Text Available We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR, and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%. Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%. In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8% compared with real-time qPCR alone (46/68, 67.6%. Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1% compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%. Legionella was not detected in the remaining six samples (6/68, 8.8%, irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  19. Application of clone library analysis and real-time PCR for comparison of microbial communities in a low-grade copper sulfide ore bioheap leachate.

    Science.gov (United States)

    Bowei, Chen; Xingyu, Liu; Wenyan, Liu; Jiankang, Wen

    2009-11-01

    The microbial communities of leachate from a bioleaching heap located in China were analyzed using the 16S rRNA gene clone library and real-time quantitative PCR. Both methods showed that Leptospirillum spp. were the dominant bacteria, and Ferroplasma acidiphilum were the only archaea detected in the leachate. Clone library results indicated that nine operational taxonomic units (OTUs) were obtained, which fell into four divisions, the Nitrospirae (74%), the gamma-Proteobacteria (14%), the Actinobacteria (6%) and the Euryarchaeota (6%). The results obtained by real-time PCR in some ways were the same as clone library analysis. Furthermore, Sulfobacillus spp., detected only by real-time PCR, suggests that real-time PCR was a reliable technology to study the microbial communities in bioleaching environments. It is a useful tool to assist clone library analysis, to further understand microbial consortia and to have comprehensive and exact microbiological information about bioleaching environments. Finally, the interactions among the microorganisms detected in the leachate were summarized according to the characteristics of these species.

  20. Chemometric brand differentiation of commercial spices using direct analysis in real time mass spectrometry.

    Science.gov (United States)

    Pavlovich, Matthew J; Dunn, Emily E; Hall, Adam B

    2016-05-15

    Commercial spices represent an emerging class of fuels for improvised explosives. Being able to classify such spices not only by type but also by brand would represent an important step in developing methods to analytically investigate these explosive compositions. Therefore, a combined ambient mass spectrometric/chemometric approach was developed to quickly and accurately classify commercial spices by brand. Direct analysis in real time mass spectrometry (DART-MS) was used to generate mass spectra for samples of black pepper, cayenne pepper, and turmeric, along with four different brands of cinnamon, all dissolved in methanol. Unsupervised learning techniques showed that the cinnamon samples clustered according to brand. Then, we used supervised machine learning algorithms to build chemometric models with a known training set and classified the brands of an unknown testing set of cinnamon samples. Ten independent runs of five-fold cross-validation showed that the training set error for the best-performing models (i.e., the linear discriminant and neural network models) was lower than 2%. The false-positive percentages for these models were 3% or lower, and the false-negative percentages were lower than 10%. In particular, the linear discriminant model perfectly classified the testing set with 0% error. Repeated iterations of training and testing gave similar results, demonstrating the reproducibility of these models. Chemometric models were able to classify the DART mass spectra of commercial cinnamon samples according to brand, with high specificity and low classification error. This method could easily be generalized to other classes of spices, and it could be applied to authenticating questioned commercial samples of spices or to examining evidence from improvised explosives. Copyright © 2016 John Wiley & Sons, Ltd.

  1. The role of real-time in biomedical science: a meta-analysis on computational complexity, delay and speedup.

    Science.gov (United States)

    Faust, Oliver; Yu, Wenwei; Rajendra Acharya, U

    2015-03-01

    The concept of real-time is very important, as it deals with the realizability of computer based health care systems. In this paper we review biomedical real-time systems with a meta-analysis on computational complexity (CC), delay (Δ) and speedup (Sp). During the review we found that, in the majority of papers, the term real-time is part of the thesis indicating that a proposed system or algorithm is practical. However, these papers were not considered for detailed scrutiny. Our detailed analysis focused on papers which support their claim of achieving real-time, with a discussion on CC or Sp. These papers were analyzed in terms of processing system used, application area (AA), CC, Δ, Sp, implementation/algorithm (I/A) and competition. The results show that the ideas of parallel processing and algorithm delay were only recently introduced and journal papers focus more on Algorithm (A) development than on implementation (I). Most authors compete on big O notation (O) and processing time (PT). Based on these results, we adopt the position that the concept of real-time will continue to play an important role in biomedical systems design. We predict that parallel processing considerations, such as Sp and algorithm scaling, will become more important. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Specification and Analysis of Soft Real-Time Systems: Quantity and Quality

    NARCIS (Netherlands)

    d' Argenio, P.R.; Katoen, Joost P.; Brinksma, Hendrik

    This paper presents a process algebra for specifying soft real-time constraints in a compositional way. For these soft constraints we take a stochastic point of view and allow arbitrary probability distributions to express delays of activities. The semantics of this process algebra is given in terms

  3. Analysis and optimization techniques for real-time streaming image processing software on general purpose systems

    NARCIS (Netherlands)

    Westmijze, Mark

    2018-01-01

    Commercial Off The Shelf (COTS) Chip Multi-Processor (CMP) systems are for cost reasons often used in industry for soft real-time stream processing. COTS CMP systems typically have a low timing predictability, which makes it difficult to develop software applications for these systems with tight

  4. Temporal analysis and scheduling of hard real-time radios running on a multi-processor

    NARCIS (Netherlands)

    Moreira, O.

    2012-01-01

    On a multi-radio baseband system, multiple independent transceivers must share the resources of a multi-processor, while meeting each its own hard real-time requirements. Not all possible combinations of transceivers are known at compile time, so a solution must be found that either allows for

  5. Developmental stage of strongyle eggs affects the outcome variations of real-time PCR analysis

    DEFF Research Database (Denmark)

    Andersen, Ulla Vestergaard; Haakansson, I. T.; Roust, Tina

    2013-01-01

    extent developmental stages can affect the variation of diagnostic test results. This study investigated the influence of developmental stages of strongyle eggs on the variation real-time polymerase chain reaction (PCR) results. Mixed species strongyle eggs were obtained from the faeces of a naturally...

  6. Reliable 5-min real-time MR technique for left-ventricular-wall motion analysis

    International Nuclear Information System (INIS)

    Katoh, Marcus; Spuentrup, Elmar; Guenther, Rolf W.; Buecker, Arno; Kuehl, Harald P.; Lipke, Claudia S.A.

    2007-01-01

    The aim of this study was to investigate the value of a real-time magnetic resonance imaging (MRI) approach for the assessment of left-ventricular-wall motion in patients with insufficient transthoracic echocardiography in terms of accuracy and temporal expenditure. Twenty-five consecutive patients were examined on a 1.5-Tesla whole-body MR system (ACS-NT, Philips Medical Systems, Best, NL) using a real-time and ECG-gated (the current gold standard) steady-state free-precession (SSFP) sequence. Wall motion was analyzed by three observers by consensus interpretation. In addition, the preparation, scanning, and overall examination times were measured. The assessment of the wall motion demonstrated a close agreement between the two modalities resulting in a mean κ coefficient of 0.8. At the same time, each stage of the examination was significantly shortened using the real-time MR approach. Real-time imaging allows for accurate assessment of left-ventricular-wall motion with the added benefit of decreased examination time. Therefore, it may serve as a cost-efficient alternative in patients with insufficient echocardiography. (orig.)

  7. WCET analysis in shared resources real-time systems with TDMA buses

    NARCIS (Netherlands)

    Rihani, H.; Moy, M.; Maiza, C.; Altmeyer, S.

    2015-01-01

    Predictability is an important aspect in real-time and safety-critical systems, where non-functional properties -- such as the timing behavior -- have high impact on the system correctness. As many safety-critical systems have a growing performance demand, simple, but outdated architectures are not

  8. Performance analysis of a real-time database with optimistic concurrency control

    NARCIS (Netherlands)

    Sassen, S.A.E.; Wal, van der J.

    1997-01-01

    For a real-time shared-memory database with Optimistic Concurrency Control (OCC), an approximation for the transaction response-time distribution and thus for the deadline miss probability is obtained. Transactions arrive at the database according to a Poisson process. There is a limited number of

  9. Design-time performance analysis of component-based real-time systems

    NARCIS (Netherlands)

    Bondarev, E.

    2009-01-01

    In current real-time systems, performance metrics are one of the most challenging properties to specify, predict and measure. Performance properties depend on various factors, like environmental context, load profile, middleware, operating system, hardware platform and sharing of internal resources.

  10. Feasibility analysis of real-time physical modeling using WaveCore processor technology on FPGA

    NARCIS (Netherlands)

    Verstraelen, Martinus Johannes Wilhelmina; Pfeifle, Florian; Bader, Rolf

    2015-01-01

    WaveCore is a scalable many-core processor technology. This technology is specifically developed and optimized for real-time acoustical modeling applications. The programmable WaveCore soft-core processor is silicon-technology independent and hence can be targeted to ASIC or FPGA technologies. The

  11. INTEGRITY ANALYSIS OF REAL-TIME PPP TECHNIQUE WITH IGS-RTS SERVICE FOR MARITIME NAVIGATION

    Directory of Open Access Journals (Sweden)

    M. El-Diasty

    2017-10-01

    Full Text Available Open sea and inland waterways are the most widely used mode for transporting goods worldwide. It is the International Maritime Organization (IMO that defines the requirements for position fixing equipment for a worldwide radio-navigation system, in terms of accuracy, integrity, continuity, availability and coverage for the various phases of navigation. Satellite positioning systems can contribute to meet these requirements, as well as optimize marine transportation. Marine navigation usually consists of three major phases identified as Ocean/Coastal/Port approach/Inland waterway, in port navigation and automatic docking with alert limit ranges from 25 m to 0.25 m. GPS positioning is widely used for many applications and is currently recognized by IMO for a future maritime navigation. With the advancement in autonomous GPS positioning techniques such as Precise Point Positioning (PPP and with the advent of new real-time GNSS correction services such as IGS-Real-Time-Service (RTS, it is necessary to investigate the integrity of the PPP-based positioning technique along with IGS-RTS service in terms of availability and reliability for safe navigation in maritime application. This paper monitors the integrity of an autonomous real-time PPP-based GPS positioning system using the IGS real-time service (RTS for maritime applications that require minimum availability of integrity of 99.8 % to fulfil the IMO integrity standards. To examine the integrity of the real-time IGS-RTS PPP-based technique for maritime applications, kinematic data from a dual frequency GPS receiver is collected onboard a vessel and investigated with the real-time IGS-RTS PPP-based GPS positioning technique. It is shown that the availability of integrity of the real-time IGS-RTS PPP-based GPS solution is 100 % for all navigation phases and therefore fulfil the IMO integrity standards (99.8 % availability immediately (after 1 second, after 2 minutes and after 42 minutes

  12. Integrity Analysis of Real-Time Ppp Technique with Igs-Rts Service for Maritime Navigation

    Science.gov (United States)

    El-Diasty, M.

    2017-10-01

    Open sea and inland waterways are the most widely used mode for transporting goods worldwide. It is the International Maritime Organization (IMO) that defines the requirements for position fixing equipment for a worldwide radio-navigation system, in terms of accuracy, integrity, continuity, availability and coverage for the various phases of navigation. Satellite positioning systems can contribute to meet these requirements, as well as optimize marine transportation. Marine navigation usually consists of three major phases identified as Ocean/Coastal/Port approach/Inland waterway, in port navigation and automatic docking with alert limit ranges from 25 m to 0.25 m. GPS positioning is widely used for many applications and is currently recognized by IMO for a future maritime navigation. With the advancement in autonomous GPS positioning techniques such as Precise Point Positioning (PPP) and with the advent of new real-time GNSS correction services such as IGS-Real-Time-Service (RTS), it is necessary to investigate the integrity of the PPP-based positioning technique along with IGS-RTS service in terms of availability and reliability for safe navigation in maritime application. This paper monitors the integrity of an autonomous real-time PPP-based GPS positioning system using the IGS real-time service (RTS) for maritime applications that require minimum availability of integrity of 99.8 % to fulfil the IMO integrity standards. To examine the integrity of the real-time IGS-RTS PPP-based technique for maritime applications, kinematic data from a dual frequency GPS receiver is collected onboard a vessel and investigated with the real-time IGS-RTS PPP-based GPS positioning technique. It is shown that the availability of integrity of the real-time IGS-RTS PPP-based GPS solution is 100 % for all navigation phases and therefore fulfil the IMO integrity standards (99.8 % availability) immediately (after 1 second), after 2 minutes and after 42 minutes of convergence

  13. Response of a diuron-degrading community to diuron exposure assessed by real-time quantitative PCR monitoring of phenylurea hydrolase A and B encoding genes

    OpenAIRE

    Pesce , S.; Beguet , J.; Rouard , N.; Devers Lamrani , M.; Martin Laurent , F.

    2013-01-01

    A real-time quantitative PCR method was developed to detect and quantify phenlylurea hydrolase genes' (puhA and puhB) sequences from environmental DNA samples to assess diuron-degrading genetic potential in some soil and sediment microbial communities. In the soil communities, mineralization rates (determined with [ring-14C]-labeled diuron) were linked to diuron-degrading genetic potentials estimated from puhB number copies, which increased following repeated diuron treatments. In the sedimen...

  14. Real-Time QoS Routing Protocols in Wireless Multimedia Sensor Networks: Study and Analysis.

    Science.gov (United States)

    Alanazi, Adwan; Elleithy, Khaled

    2015-09-02

    Many routing protocols have been proposed for wireless sensor networks. These routing protocols are almost always based on energy efficiency. However, recent advances in complementary metal-oxide semiconductor (CMOS) cameras and small microphones have led to the development of Wireless Multimedia Sensor Networks (WMSN) as a class of wireless sensor networks which pose additional challenges. The transmission of imaging and video data needs routing protocols with both energy efficiency and Quality of Service (QoS) characteristics in order to guarantee the efficient use of the sensor nodes and effective access to the collected data. Also, with integration of real time applications in Wireless Senor Networks (WSNs), the use of QoS routing protocols is not only becoming a significant topic, but is also gaining the attention of researchers. In designing an efficient QoS routing protocol, the reliability and guarantee of end-to-end delay are critical events while conserving energy. Thus, considerable research has been focused on designing energy efficient and robust QoS routing protocols. In this paper, we present a state of the art research work based on real-time QoS routing protocols for WMSNs that have already been proposed. This paper categorizes the real-time QoS routing protocols into probabilistic and deterministic protocols. In addition, both categories are classified into soft and hard real time protocols by highlighting the QoS issues including the limitations and features of each protocol. Furthermore, we have compared the performance of mobility-aware query based real-time QoS routing protocols from each category using Network Simulator-2 (NS2). This paper also focuses on the design challenges and future research directions as well as highlights the characteristics of each QoS routing protocol.

  15. Relative quantification of mRNA: comparison of methods currently used for real-time PCR data analysis

    Directory of Open Access Journals (Sweden)

    Koppel Juraj

    2007-12-01

    Full Text Available Abstract Background Fluorescent data obtained from real-time PCR must be processed by some method of data analysis to obtain the relative quantity of target mRNA. The method chosen for data analysis can strongly influence results of the quantification. Results To compare the performance of six techniques which are currently used for analysing fluorescent data in real-time PCR relative quantification, we quantified four cytokine transcripts (IL-1β, IL-6 TNF-α, and GM-CSF in an in vivo model of colonic inflammation. Accuracy of the methods was tested by quantification on samples with known relative amounts of target mRNAs. Reproducibility of the methods was estimated by the determination of the intra-assay and inter-assay variability. Cytokine expression normalized to the expression of three reference genes (ACTB, HPRT, SDHA was then determined using the six methods for data analysis. The best results were obtained with the relative standard curve method, comparative Ct method and with DART-PCR, LinRegPCR and Liu & Saint exponential methods when average amplification efficiency was used. The use of individual amplification efficiencies in DART-PCR, LinRegPCR and Liu & Saint exponential methods significantly impaired the results. The sigmoid curve-fitting (SCF method produced medium performance; the results indicate that the use of appropriate type of fluorescence data and in some instances manual selection of the number of amplification cycles included in the analysis is necessary when the SCF method is applied. We also compared amplification efficiencies (E and found that although the E values determined by different methods of analysis were not identical, all the methods were capable to identify two genes whose E values significantly differed from other genes. Conclusion Our results show that all the tested methods can provide quantitative values reflecting the amounts of measured mRNA in samples, but they differ in their accuracy and

  16. Exogenous reference gene normalization for real-time reverse transcription-polymerase chain reaction analysis under dynamic endogenous transcription.

    Science.gov (United States)

    Johnston, Stephen; Gallaher, Zachary; Czaja, Krzysztof

    2012-05-15

    Quantitative real-time reverse transcription-polymerase chain reaction (qPCR) is widely used to investigate transcriptional changes following experimental manipulations to the nervous system. Despite the widespread utilization of qPCR, the interpretation of results is marred by the lack of a suitable reference gene due to the dynamic nature of endogenous transcription. To address this inherent deficiency, we investigated the use of an exogenous spike-in mRNA, luciferase, as an internal reference gene for the 2(-∆∆Ct) normalization method. To induce dynamic transcription, we systemically administered capsaicin, a neurotoxin selective for C-type sensory neurons expressing the TRPV-1 receptor, to adult male Sprague-Dawley rats. We later isolated nodose ganglia for qPCR analysis with the reference being either exogenous luciferase mRNA or the commonly used endogenous reference β-III tubulin. The exogenous luciferase mRNA reference clearly demonstrated the dynamic expression of the endogenous reference. Furthermore, variability of the endogenous reference would lead to misinterpretation of other genes of interest. In conclusion, traditional reference genes are often unstable under physiologically normal situations, and certainly unstable following the damage to the nervous system. The use of exogenous spike-in reference provides a consistent and easily implemented alternative for the analysis of qPCR data.

  17. Identification of the major capsid protein of erythrocytic necrosis virus (ENV) and development of quantitative real-time PCR assays for quantification of ENV DNA

    Science.gov (United States)

    Purcell, Maureen K.; Pearman-Gillman, Schuyler; Thompson, Rachel L.; Gregg, Jacob L.; Hart, Lucas M.; Winton, James R.; Emmenegger, Eveline J.; Hershberger, Paul K.

    2016-01-01

    Viral erythrocytic necrosis (VEN) is a disease of marine and anadromous fish that is caused by the erythrocytic necrosis virus (ENV), which was recently identified as a novel member of family Iridoviridae by next-generation sequencing. Phylogenetic analysis of the ENV DNA polymerase grouped ENV with other erythrocytic iridoviruses from snakes and lizards. In the present study, we identified the gene encoding the ENV major capsid protein (MCP) and developed a quantitative real-time PCR (qPCR) assay targeting this gene. Phylogenetic analysis of the MCP gene sequence supported the conclusion that ENV does not group with any of the currently described iridovirus genera. Because there is no information regarding genetic variation of the MCP gene across the reported host and geographic range for ENV, we also developed a second qPCR assay for a more conserved ATPase-like gene region. The MCP and ATPase qPCR assays demonstrated good analytical and diagnostic sensitivity and specificity based on samples from laboratory challenges of Pacific herring Clupea pallasii. The qPCR assays had similar diagnostic sensitivity and specificity as light microscopy of stained blood smears for the presence of intraerythrocytic inclusion bodies. However, the qPCR assays may detect viral DNA early in infection prior to the formation of inclusion bodies. Both qPCR assays appear suitable for viral surveillance or as a confirmatory test for ENV in Pacific herring from the Salish Sea.

  18. Comparative evaluation of a laboratory developed real-time PCR assay and the RealStar® HHV-6 PCR Kit for quantitative detection of human herpesvirus 6.

    Science.gov (United States)

    Yip, Cyril C Y; Sridhar, Siddharth; Cheng, Andrew K W; Fung, Ami M Y; Cheng, Vincent C C; Chan, Kwok-Hung; Yuen, Kwok-Yung

    2017-08-01

    HHV-6 reactivation in immunocompromised patients is common and may be associated with serious morbidity and mortality; therefore, early detection and initiation of therapy might be of benefit. Real-time PCR assays allow for early identification of HHV-6 reactivation to assist in providing a timely response. Thus, we compared the performance of an in-house developed HHV-6 quantitative PCR assay with a commercially available kit, the RealStar ® HHV-6 PCR Kit. The analytical sensitivity, analytical specificity, linearity, precision and accuracy of the in-house developed HHV-6 qPCR assay were evaluated. The diagnostic performance of the in-house HHV-6 qPCR assay was compared with the RealStar ® HHV-6 PCR Kit, using 72 clinical specimens and 17 proficiency testing samples. Linear regression analysis of the quantitative results showed a dynamic range from 2 to 10 log 10 copies/ml and a coefficient of determination (R 2 ) of 0.999 for the in-house assay. A dilution series demonstrated a limit of detection and a limit of quantification of 1.7 log 10 and 2 log 10 copies/ml, respectively. The precision of the assay was highly reproducible among runs with coefficients of variance (CV) ranging from 0.27% to 4.37%. A comparison of 27 matched samples showed an excellent correlation between the quantitative viral loads measured by the in-house HHV-6 qPCR assay and the RealStar ® HHV-6 PCR Kit (R 2 =0.926; PPCR Kit. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Soil Baiting, Rapid PCR Assay and Quantitative Real Time PCR to Diagnose Late Blight of Potato in Quarantine Programs

    Directory of Open Access Journals (Sweden)

    Touseef Hussain

    2018-05-01

    Full Text Available Phytophthora infestans (mont de Bary is a pathogen of great concern across the globe, and accurate detection is an important component in responding to the outbreaks of potential disease. Although the molecular diagnostic protocol used in regulatory programs has been evaluated but till date methods implying direct comparison has rarely used. In this study, a known area soil samples from potato fields where light blight appear every year (both A1 and A2 mating type was assayed by soil bait method, PCR assay detection and quantification of the inoculums. Suspected disease symptoms appeared on bait tubers were further confirmed by rapid PCR, inoculums were quantified through Real Time PCR, which confirms presence of P. infestans. These diagnostic methods can be highly correlated with one another. Potato tuber baiting increased the sensitivity of the assay compared with direct extraction of DNA from tuber and soil samples. Our study determines diagnostic sensitivity and specificity of the assays to determine the performance of each method. Overall, molecular techniques based on different types of PCR amplification and Real-time PCR can lead to high throughput, faster and more accurate detection method which can be used in quarantine programmes in potato industry and diagnostic laboratory.

  20. Real-Time Measurement of Electronic Cigarette Aerosol Size Distribution and Metals Content Analysis.

    Science.gov (United States)

    Mikheev, Vladimir B; Brinkman, Marielle C; Granville, Courtney A; Gordon, Sydney M; Clark, Pamela I

    2016-09-01

    Electronic cigarette (e-cigarette) use is increasing worldwide and is highest among both daily and nondaily smokers. E-cigarettes are perceived as a healthier alternative to combustible tobacco products, but their health risk factors have not yet been established, and one of them is lack of data on aerosol size generated by e-cigarettes. We applied a real-time, high-resolution aerosol differential mobility spectrometer to monitor the evolution of aerosol size and concentration during puff development. Particles generated by e-cigarettes were immediately delivered for analysis with minimal dilution and therefore with minimal sample distortion, which is critically important given the highly dynamic aerosol/vapor mixture inherent to e-cigarette emissions. E-cigarette aerosols normally exhibit a bimodal particle size distribution: nanoparticles (11-25nm count median diameter) and submicron particles (96-175nm count median diameter). Each mode has comparable number concentrations (10(7)-10(8) particles/cm(3)). "Dry puff" tests conducted with no e-cigarette liquid (e-liquid) present in the e-cigarette tank demonstrated that under these conditions only nanoparticles were generated. Analysis of the bulk aerosol collected on the filter showed that e-cigarette emissions contained a variety of metals. E-cigarette aerosol size distribution is different from that of combustible tobacco smoke. E-cigarettes generate high concentrations of nanoparticles and their chemical content requires further investigation. Despite the small mass of nanoparticles, their toxicological impact could be significant. Toxic chemicals that are attached to the small nanoparticles may have greater adverse health effects than when attached to larger submicron particles. The e-cigarette aerosol size distribution is different from that of combustible tobacco smoke and typically exhibits a bimodal behavior with comparable number concentrations of nanoparticles and submicron particles. While vaping the e

  1. Modeling and analysis of real-time and embedded systems with UML and MARTE developing cyber-physical systems

    CERN Document Server

    Selic, Bran

    2013-01-01

    Modeling and Analysis of Real-Time and Embedded Systems with UML and MARTE explains how to apply the complex MARTE standard in practical situations. This approachable reference provides a handy user guide, illustrating with numerous examples how you can use MARTE to design and develop real-time and embedded systems and software. Expert co-authors Bran Selic and Sébastien Gérard lead the team that drafted and maintain the standard and give you the tools you need apply MARTE to overcome the limitations of cyber-physical systems. The functional sophistication required of modern cyber-physical

  2. Quantitation of hepatitis B virus DNA in plasma using a sensitive cost-effective "in-house" real-time PCR assay

    Directory of Open Access Journals (Sweden)

    Daniel Hubert Darius

    2009-01-01

    Full Text Available Background: Sensitive nucleic acid testing for the detection and accurate quantitation of hepatitis B virus (HBV is necessary to reduce transmission through blood and blood products and for monitoring patients on antiviral therapy. The aim of this study is to standardize an "in-house" real-time HBV polymerase chain reaction (PCR for accurate quantitation and screening of HBV. Materials and Methods: The "in-house" real-time assay was compared with a commercial assay using 30 chronically infected individuals and 70 blood donors who are negative for hepatitis B surface antigen, hepatitis C virus (HCV antibody and human immunodeficiency virus (HIV antibody. Further, 30 HBV-genotyped samples were tested to evaluate the "in-house" assay′s capacity to detect genotypes prevalent among individuals attending this tertiary care hospital. Results: The lower limit of detection of this "in-house" HBV real-time PCR was assessed against the WHO international standard and found to be 50 IU/mL. The interassay and intra-assay coefficient of variation (CV of this "in-house" assay ranged from 1.4% to 9.4% and 0.0% to 2.3%, respectively. Virus loads as estimated with this "in-house" HBV real-time assay correlated well with the commercial artus HBV RG PCR assay ( r = 0.95, P < 0.0001. Conclusion: This assay can be used for the detection and accurate quantitation of HBV viral loads in plasma samples. This assay can be employed for the screening of blood donations and can potentially be adapted to a multiplex format for simultaneous detection of HBV, HIV and HCV to reduce the cost of testing in blood banks.

  3. Analysis of Real Time Technical Data Obtained While Shotcreting: An Approach Towards Automation

    OpenAIRE

    Rodríguez, Ángel; Río, Olga

    2010-01-01

    Automation of shotcreting process is a key factor in both improving the working conditions and increasing productivity; as well as in increasing the quality of shotcrete. The confidence in the quality of the automation process itself and shotcrete linings can be improved by real time monitoring of pumping as well as other shotcreting machine related parameters. Prediction of how the difIerent technical parameters of application are governing the whole process is being a subject of increasing ...

  4. A Schedulability Analysis Framework for Real-time Infrastructure Systems Managing Heterogeneous Resources

    OpenAIRE

    Kim, Jinhyun; Kang, Sungwon

    2012-01-01

    REACTION 2012. 1st International workshop on Real-time and distributed computing in emerging applications. December 4th, 2012, San Juan, Puerto Rico. Electricity generating systems, such as smart grid systems, and water management systems are infrastructure systems that manage resources critical to human life. In the systems, resources are produced and managed to supply them to various consumers, such as building, car, factory, and household, according to their needs and pri...

  5. Efficient Feasibility Analysis for Real-Time Systems with EDF Scheduling

    OpenAIRE

    Albers , Karsten; Slomka , Frank

    2005-01-01

    Submitted on behalf of EDAA (http://www.edaa.com/); International audience; This paper presents new fast exact feasibility tests for uniprocessor real-time systems using preemptive EDF scheduling. Task sets which are accepted by previously described sufficient tests will be evaluated in nearly the same time as with the old tests by the new algorithms. Many task sets are not accepted by the earlier tests despite them beeing feasible. These task sets will be evaluated by the new algorithms a lo...

  6. RCNF: Real-time Collaborative Network Forensic Scheme for Evidence Analysis

    OpenAIRE

    Moustafa, Nour; Slay, Jill

    2017-01-01

    Network forensic techniques help in tracking different types of cyber attack by monitoring and inspecting network traffic. However, with the high speed and large sizes of current networks, and the sophisticated philosophy of attackers, in particular mimicking normal behaviour and/or erasing traces to avoid detection, investigating such crimes demands intelligent network forensic techniques. This paper suggests a real-time collaborative network Forensic scheme (RCNF) that can monitor and inves...

  7. A Platform for Real-time Acquisition and Analysis of Physiological Data in Hospital Emergency Departments

    Science.gov (United States)

    2014-08-01

    with the Department of Emergency Medicine, Massachusetts General Hospital, Boston, MA 02114 USA (corresponding author; phone: 617 -726-2241; e-mail...programming interface ( API ). Algorithms are used to determine the reliability of waveform (e.g., electrocardiogram) and vital-sign data (e.g., heart rate...and comparing of real-time decision- support algorithms in mobile environments," Conf Proc IEEE Eng Med Biol Soc, vol. 2009 , pp. 3417-20, 2009 . [3

  8. Quantitative Microbiological Study of Human Carious Dentine by Culture and Real-Time PCR: Association of Anaerobes with Histopathological Changes in Chronic Pulpitis

    Science.gov (United States)

    Martin, F. Elizabeth; Nadkarni, Mangala A.; Jacques, Nicholas A.; Hunter, Neil

    2002-01-01

    The bacteria found in carious dentine were correlated with the tissue response of the dental pulps of 65 teeth extracted from patients with advanced caries and pulpitis. Standardized homogenates of carious dentine were plated onto selective and nonselective media under anaerobic and microaerophilic conditions. In addition, real-time PCR was used to quantify the recovery of anaerobic bacteria. Primers and fluorogenic probes were designed to detect the total anaerobic microbial load, the genera Prevotella and Fusobacterium, and the species Prevotella melaninogenica, Porphyromonas endodontalis, Porphyromonas gingivalis, and Micromonas (formerly Peptostreptococcus) micros. The pulpal pathology was categorized according to the cellular response and degenerative changes. Analysis of cultured bacteria showed a predominance of gram-positive microorganisms, particularly lactobacilli. Gram-negative bacteria were also present in significant numbers with Prevotella spp., the most numerous anaerobic group cultured. Real-time PCR analysis indicated a greater microbial load than that determined by colony counting. The total number of anaerobes detected was 41-fold greater by real-time PCR than by colony counting, while the numbers of Prevotella and Fusobacterium spp. detected were 82- and 2.4-fold greater by real-time PCR than by colony counting, respectively. Real-time PCR also identified M. micros, P. endodontalis, and P. gingivalis in 71, 60, and 52% of carious samples, respectively. Correlation matrices of the real-time PCR data revealed significant positive associations between M. micros and P. endodontalis detection and inflammatory degeneration of pulpal tissues. These anaerobes have been strongly implicated in endodontic infections that occur as sequelae to carious pulpitis. Accordingly, the data suggest that the presence of high levels of these bacteria in carious lesions may be indicative of irreversible pulpal pathology. PMID:11980945

  9. MTF analysis of the MURR real-time neutron radiography facility

    International Nuclear Information System (INIS)

    Lindsay, J.T.

    1982-01-01

    In neutron radiography, as in other forms of NDE, it is sometimes desirable to observe dynamic events. This need has generated increased interest in real-time neutron radiography systems. As in other forms of radiography, a standard method for measuring the image forming capability of real-time systems is necessary in order to compare the various methods and systems used. A technique which has been used extensively in general photography and has been applied in the characterization of several screen-film combinations used in conventional neutron radiography is to determine the imaging system's modulation transfer function (MTF). This gives a graphical representation of the system's spatial resolution capabilities and was therefore chosen as the method for evaluation of the real-time neutron radiography facility at the University of Missouri Research Reactor (MURR). The method used was to image a knife-edge, differentiate the edge gradient to obtain the line spread function (LSF) to obtain the system MTF. A Gd foil was used for the knife-edge on several neutron converter screens and was imaged by a low-light level ISIT camera. The video signal was then digitized and presented to a PDP-11/05 microcomputer for the numerical calculations

  10. Near-real-time Forensic Disaster Analysis: experiences from hurricane Sandy

    Science.gov (United States)

    Kunz, Michael; Mühr, Bernhard; Schröter, Kai; Kunz-Plapp, Tina; Daniell, James; Khazai, Bijan; Wenzel, Friedemann; Vannieuwenhuyse, Marjorie; Comes, Tina; Münzberg, Thomas; Elmer, Florian; Fohringer, Joachim; Lucas, Christian; Trieselmann, Werner; Zschau, Jochen

    2013-04-01

    Hurricane Sandy was the last tropical cyclone of the 2012 Northern Atlantic Hurricane season that made landfall. It moved on an unusual track from the Caribbean to the East Coast of the United States from 24 to 30 October as a Category 1 and 2 Hurricane according to the Saffir-Simpson Scale. Along its path, the severe storm event caused widespread damage including almost 200 fatalities. In the early hours of 30 October, Sandy made landfall near Atlantic City, N.J. Sandy was an extraordinary event due to its multihazard nature and several cascading effects in the aftermath. From the hydro-meteorological perspective, most unusual was the very large spatial extent of up to 1,700 km. High wind speeds were associated with record breaking storm surges at the U.S. Mid- Atlantic and New England Coast during high (astronomical) tide, leading to widespread flooding. Though Sandy was not the most severe storm event in terms of wind speed and precipitation, the impact in the U.S. was enormous with total damage estimates of up to 90 billion US (own estimate from Dec. 2012). Although much better data emerge weeks after such an event, the Forensic Disaster Analysis (FDA) Task Force of the Center for Disaster Management and Risk Reduction Technology (CEDIM) made an effort to obtain a comprehensive and holistic overview of the causes, hazardous effects and consequences associated with Sandy immediately after landfall at the U.S. coast on 30 October 2012. This was done in an interdisciplinary way by collecting and compiling scattered and distributed information from available databases and sources via the Internet, by applying own methodologies and models for near-real time analyses developed in recent years, and by expert knowledge. This contribution gives an overview about the CEDIM-FDA analyses' results. It describes the situation that led to the extraordinary event, highlights the interaction of the tropical cyclone with other hydro-meteorological events, and examines the

  11. Urethra-Sparing, Intraoperative, Real-Time Planned, Permanent-Seed Prostate Brachytherapy: Toxicity Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Zilli, Thomas [Department of Radiation Oncology, Centre hospitalier de l' Universite de Montreal-Hopital Notre-Dame, Montreal, QC (Canada); Taussky, Daniel, E-mail: daniel.taussky.chum@ssss.gouv.qc.ca [Department of Radiation Oncology, Centre hospitalier de l' Universite de Montreal-Hopital Notre-Dame, Montreal, QC (Canada); Donath, David; Le, Hoa Phong; Larouche, Renee-Xaviere; Beliveau-Nadeau, Dominique; Hervieux, Yannick; Delouya, Guila [Department of Radiation Oncology, Centre hospitalier de l' Universite de Montreal-Hopital Notre-Dame, Montreal, QC (Canada)

    2011-11-15

    Purpose: To report the toxicity outcome in patients with localized prostate cancer undergoing {sup 125}I permanent-seed brachytherapy (BT) according to a urethra-sparing, intraoperative (IO), real-time planned conformal technique. Methods and Materials: Data were analyzed on 250 patients treated consecutively for low- or intermediate-risk prostate cancer between 2005 and 2009. The planned goal was urethral V{sub 150} = 0. Acute and late genitourinary (GU), gastrointestinal (GI), and erectile toxicities were scored with the International Prostate Symptom Score (IPSS) questionnaire and Common Terminology Criteria for Adverse Events (version 3.0). Median follow-up time for patients with at least 2 years of follow-up (n = 130) was 34.4 months (range, 24-56.9 months). Results: Mean IO urethra V{sub 150} was 0.018% {+-} 0.08%. Mean prostate D{sub 90} and V{sub 100} on day-30 computed tomography scan were 158.0 {+-} 27.0 Gy and 92.1% {+-} 7.2%, respectively. Mean IPSS peak was 9.5 {+-} 6.3 1 month after BT (mean difference from baseline IPSS, 5.3). No acute GI toxicity was observed in 86.8% of patients. The 3-year probability of Grade {>=}2 late GU toxicity-free survival was 77.4% {+-} 4.0%, with Grade 3 late GU toxicity encountered in only 3 patients. Three-year Grade 1 late GI toxicity-free survival was 86.1% {+-} 3.2%. No patient presented Grade {>=}2 late GI toxicity. Of patients with normal sexual status at baseline, 20.7% manifested Grade {>=}2 erectile dysfunction after BT. On multivariate analysis, elevated baseline IPSS (p = 0.016) and high-activity sources (median 0.61 mCi) (p = 0.033) predicted increased Grade {>=}2 late GU toxicity. Conclusions: Urethra-sparing IO BT results in low acute and late GU toxicity compared with the literature. High seed activity and elevated IPSS at baseline increased long-term GU toxicity.

  12. Selection of suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using quantitative real-time polymerase chain reaction.

    Science.gov (United States)

    Zornhagen, K W; Kristensen, A T; Hansen, A E; Oxboel, J; Kjaer, A

    2015-12-01

    Quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) is a sensitive technique for quantifying gene expression. Stably expressed reference genes are necessary for normalization of RT-qPCR data. Only a few articles have been published on reference genes in canine tumours. The objective of this study was to demonstrate how to identify suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using RT-qPCR. Primer pairs for 17 potential reference genes were designed and tested in archival tumour biopsies from six dogs. The geNorm algorithm was used to analyse the most suitable reference genes. Eight potential reference genes were excluded from this final analysis because of their dissociation curves. β-Glucuronidase (GUSB) and proteasome subunit, beta type, 6 (PSMB6) were most stably expressed with an M value of 0.154 and a CV of 0.053 describing their average stability. We suggest that choice of reference genes should be based on specific testing in every new experimental set-up. © 2014 John Wiley & Sons Ltd.

  13. A human fecal contamination score for ranking recreational sites using the HF183/BacR287 quantitative real-time PCR method.

    Science.gov (United States)

    Cao, Yiping; Sivaganesan, Mano; Kelty, Catherine A; Wang, Dan; Boehm, Alexandria B; Griffith, John F; Weisberg, Stephen B; Shanks, Orin C

    2018-01-01

    Human fecal pollution of recreational waters remains a public health concern worldwide. As a result, there is a growing interest in the application of human-associated fecal source identification quantitative real-time PCR (qPCR) technologies for water quality research and management. However, there are currently no standardized approaches for field implementation and interpretation of qPCR data. In this study, a standardized HF183/BacR287 qPCR method was combined with a water sampling strategy and a novel Bayesian weighted average approach to establish a human fecal contamination score (HFS) that can be used to prioritize sampling sites for remediation based on measured human waste levels. The HFS was then used to investigate 975 study design scenarios utilizing different combinations of sites with varying sampling intensities (daily to once per week) and number of qPCR replicates per sample (2-14 replicates). Findings demonstrate that site prioritization with HFS is feasible and that both sampling intensity and number of qPCR replicates influence reliability of HFS estimates. The novel data analysis strategy presented here provides a prescribed approach for the implementation and interpretation of human-associated HF183/BacR287 qPCR data with the goal of site prioritization based on human fecal pollution levels. In addition, information is provided for future users to customize study designs for optimal HFS performance. Published by Elsevier Ltd.

  14. Identification and validation of biomarkers of IgV(H) mutation status in chronic lymphocytic leukemia using microfluidics quantitative real-time polymerase chain reaction technology.

    Science.gov (United States)

    Abruzzo, Lynne V; Barron, Lynn L; Anderson, Keith; Newman, Rachel J; Wierda, William G; O'brien, Susan; Ferrajoli, Alessandra; Luthra, Madan; Talwalkar, Sameer; Luthra, Rajyalakshmi; Jones, Dan; Keating, Michael J; Coombes, Kevin R

    2007-09-01

    To develop a model incorporating relevant prognostic biomarkers for untreated chronic lymphocytic leukemia patients, we re-analyzed the raw data from four published gene expression profiling studies. We selected 88 candidate biomarkers linked to immunoglobulin heavy-chain variable region gene (IgV(H)) mutation status and produced a reliable and reproducible microfluidics quantitative real-time polymerase chain reaction array. We applied this array to a training set of 29 purified samples from previously untreated patients. In an unsupervised analysis, the samples clustered into two groups. Using a cutoff point of 2% homology to the germline IgV(H) sequence, one group contained all 14 IgV(H)-unmutated samples; the other contained all 15 mutated samples. We confirmed the differential expression of 37 of the candidate biomarkers using two-sample t-tests. Next, we constructed 16 different models to predict IgV(H) mutation status and evaluated their performance on an independent test set of 20 new samples. Nine models correctly classified 11 of 11 IgV(H)-mutated cases and eight of nine IgV(H)-unmutated cases, with some models using three to seven genes. Thus, we can classify cases with 95% accuracy based on the expression of as few as three genes.

  15. Gene expression profile and immunological evaluation of unique hypothetical unknown proteins of Mycobacterium leprae by using quantitative real-time PCR.

    Science.gov (United States)

    Kim, Hee Jin; Prithiviraj, Kalyani; Groathouse, Nathan; Brennan, Patrick J; Spencer, John S

    2013-02-01

    The cell-mediated immunity (CMI)-based in vitro gamma interferon release assay (IGRA) of Mycobacterium leprae-specific antigens has potential as a promising diagnostic means to detect those individuals in the early stages of M. leprae infection. Diagnosis of leprosy is a major obstacle toward ultimate disease control and has been compromised in the past by the lack of specific markers. Comparative bioinformatic analysis among mycobacterial genomes identified potential M. leprae-specific proteins called "hypothetical unknowns." Due to massive gene decay and the prevalence of pseudogenes, it is unclear whether any of these proteins are expressed or are immunologically relevant. In this study, we performed cDNA-based quantitative real-time PCR to investigate the expression status of 131 putative open reading frames (ORFs) encoding hypothetical unknowns. Twenty-six of the M. leprae-specific antigen candidates showed significant levels of gene expression compared to that of ESAT-6 (ML0049), which is an important T cell antigen of low abundance in M. leprae. Fifteen of 26 selected antigen candidates were expressed and purified in Escherichia coli. The seroreactivity to these proteins of pooled sera from lepromatous leprosy patients and cavitary tuberculosis patients revealed that 9 of 15 recombinant hypothetical unknowns elicited M. leprae-specific immune responses. These nine proteins may be good diagnostic reagents to improve both the sensitivity and specificity of detection of individuals with asymptomatic leprosy.

  16. Quantitation of O6-methylguanine-DNA methyltransferase gene messenger RNA in gliomas by means of real-time RT-PCR and clinical response to nitrosoureas.

    Science.gov (United States)

    Tanaka, Satoshi; Oka, Hidehiro; Fujii, Kiyotaka; Watanabe, Kaoru; Nagao, Kumi; Kakimoto, Atsushi

    2005-09-01

    1. O6-methylguanine-DNA methyltransferase (MGMT) mRNA was measured in 50 malignant gliomas that had received 1-(4-amino-2-methyl-5-pyrimidynyl) methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU) after the resection of the tumor by real-time reverse transcription-polymerase chain reaction (RT-PCR) using TaqMan probe. 2. The mean absolute value of MGMTmRNA normalized to the level of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) for 50 tumors was 1.29 x 10(4)+/- 1.28 x 10(4) copy/microg RNA (mean +/- SD). The amount of MGMTmRNA less than 6 x 10(3) copy/microg RNA was the most significant factor in predicting the initial effect of treatment with ACNU by multi-variant regression analysis (p = 0.0157). 3. These results suggest that quantitation of MGMTmRNA is the excellent method for predicting for the effect of ACNU in glioma therapy.

  17. Assessing reference genes for accurate transcript normalization using quantitative real-time PCR in pearl millet [Pennisetum glaucum (L. R. Br].

    Directory of Open Access Journals (Sweden)

    Prasenjit Saha

    Full Text Available Pearl millet [Pennisetum glaucum (L. R.Br.], a close relative of Panicoideae food crops and bioenergy grasses, offers an ideal system to perform functional genomics studies related to C4 photosynthesis and abiotic stress tolerance. Quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR provides a sensitive platform to conduct such gene expression analyses. However, the lack of suitable internal control reference genes for accurate transcript normalization during qRT-PCR analysis in pearl millet is the major limitation. Here, we conducted a comprehensive assessment of 18 reference genes on 234 samples which included an array of different developmental tissues, hormone treatments and abiotic stress conditions from three genotypes to determine appropriate reference genes for accurate normalization of qRT-PCR data. Analyses of Ct values using Stability Index, BestKeeper, ΔCt, Normfinder, geNorm and RefFinder programs ranked PP2A, TIP41, UBC2, UBQ5 and ACT as the most reliable reference genes for accurate transcript normalization under different experimental conditions. Furthermore, we validated the specificity of these genes for precise quantification of relative gene expression and provided evidence that a combination of the best reference genes are required to obtain optimal expression patterns for both endogeneous genes as well as transgenes in pearl millet.

  18. Quantitative Tetraplex Real-Time Polymerase Chain Reaction Assay with TaqMan Probes Discriminates Cattle, Buffalo, and Porcine Materials in Food Chain.

    Science.gov (United States)

    Hossain, M A Motalib; Ali, Md Eaqub; Sultana, Sharmin; Asing; Bonny, Sharmin Quazi; Kader, Md Abdul; Rahman, M Aminur

    2017-05-17

    Cattle, buffalo, and porcine materials are widely adulterated, and their quantification might safeguard health, religious, economic, and social sanctity. Recently, conventional polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism (RFLP) assays have been documented but they are just suitable for identification, cannot quantify adulterations. We described here a quantitative tetraplex real-time PCR assay with TaqMan Probes to quantify contributions from cattle, buffalo, and porcine materials simultaneously. Amplicon-sizes were very short (106-, 90-, and 146-bp for cattle, buffalo, and porcine) because longer targets could be broken down, bringing serious ambiguity in molecular diagnostics. False negative detection was eliminated through an endogenous control (141-bp site of eukaryotic 18S rRNA). Analysis of 27 frankfurters and 27 meatballs reflected 84-115% target recovery at 0.1-10% adulterations. Finally, a test of 36 commercial products revealed 71% beef frankfurters, 100% meatballs, and 85% burgers contained buffalo adulteration, but no porcine was found in beef products.

  19. On the relationship between fiscal plans in the European Union: An empirical analysis based on real-time data

    NARCIS (Netherlands)

    Giuliodori, M.; Beetsma, R.M.W.J.

    2007-01-01

    We investigate the interdependence of fiscal policies, and in particular deficits, in the European Union using an empirical analysis based on real-time fiscal data. There are many potential reasons why fiscal policies could be interdependent, such as direct externalities due to cross-border public

  20. On the relationship between fiscal plans in the European Union: an empirical analysis based on real-time data

    NARCIS (Netherlands)

    Giuliodori, M.; Beetsma, R.

    2008-01-01

    We investigate the interdependence of fiscal policies, and in particular deficits, in the European Union using an empirical analysis based on real-time fiscal data. There are many potential reasons why fiscal policies could be interdependent, such as direct externalities due to cross-border public

  1. Combining real-time PCR and next-generation DNA sequencing to provide quantitative comparisons of fungal aerosol populations

    Science.gov (United States)

    Dannemiller, Karen C.; Lang-Yona, Naama; Yamamoto, Naomichi; Rudich, Yinon; Peccia, Jordan

    2014-02-01

    We examined fungal communities associated with the PM10 mass of Rehovot, Israel outdoor air samples collected in the spring and fall seasons. Fungal communities were described by 454 pyrosequencing of the internal transcribed spacer (ITS) region of the fungal ribosomal RNA encoding gene. To allow for a more quantitative comparison of fungal exposure in humans, the relative abundance values of specific taxa were transformed to absolute concentrations through multiplying these values by the sample's total fungal spore concentration (derived from universal fungal qPCR). Next, the sequencing-based absolute concentrations for Alternaria alternata, Cladosporium cladosporioides, Epicoccum nigrum, and Penicillium/Aspergillus spp. were compared to taxon-specific qPCR concentrations for A. alternata, C. cladosporioides, E. nigrum, and Penicillium/Aspergillus spp. derived from the same spring and fall aerosol samples. Results of these comparisons showed that the absolute concentration values generated from pyrosequencing were strongly associated with the concentration values derived from taxon-specific qPCR (for all four species, p 0.70). The correlation coefficients were greater for species present in higher concentrations. Our microbial aerosol population analyses demonstrated that fungal diversity (number of fungal operational taxonomic units) was higher in the spring compared to the fall (p = 0.02), and principal coordinate analysis showed distinct seasonal differences in taxa distribution (ANOSIM p = 0.004). Among genera containing allergenic and/or pathogenic species, the absolute concentrations of Alternaria, Aspergillus, Fusarium, and Cladosporium were greater in the fall, while Cryptococcus, Penicillium, and Ulocladium concentrations were greater in the spring. The transformation of pyrosequencing fungal population relative abundance data to absolute concentrations can improve next-generation DNA sequencing-based quantitative aerosol exposure assessment.

  2. [Real time 3D echocardiography

    Science.gov (United States)

    Bauer, F.; Shiota, T.; Thomas, J. D.

    2001-01-01

    Three-dimensional representation of the heart is an old concern. Usually, 3D reconstruction of the cardiac mass is made by successive acquisition of 2D sections, the spatial localisation and orientation of which require complex guiding systems. More recently, the concept of volumetric acquisition has been introduced. A matricial emitter-receiver probe complex with parallel data processing provides instantaneous of a pyramidal 64 degrees x 64 degrees volume. The image is restituted in real time and is composed of 3 planes (planes B and C) which can be displaced in all spatial directions at any time during acquisition. The flexibility of this system of acquisition allows volume and mass measurement with greater accuracy and reproducibility, limiting inter-observer variability. Free navigation of the planes of investigation allows reconstruction for qualitative and quantitative analysis of valvular heart disease and other pathologies. Although real time 3D echocardiography is ready for clinical usage, some improvements are still necessary to improve its conviviality. Then real time 3D echocardiography could be the essential tool for understanding, diagnosis and management of patients.

  3. Real Time Processing

    CERN Multimedia

    CERN. Geneva; ANDERSON, Dustin James; DOGLIONI, Caterina

    2015-01-01

    The LHC provides experiments with an unprecedented amount of data. Experimental collaborations need to meet storage and computing requirements for the analysis of this data: this is often a limiting factor in the physics program that would be achievable if the whole dataset could be analysed. In this talk, I will describe the strategies adopted by the LHCb, CMS and ATLAS collaborations to overcome these limitations and make the most of LHC data: data parking, data scouting, and real-time analysis.

  4. An Integrated Hot-Stage Microscope-Direct Analysis in Real Time-Mass Spectrometry System for Studying the Thermal Behavior of Materials.

    Science.gov (United States)

    Ashton, Gage P; Harding, Lindsay P; Parkes, Gareth M B

    2017-12-19

    This paper describes a new analytical instrument that combines a precisely temperature-controlled hot-stage with digital microscopy and Direct Analysis in Real Time-mass spectrometry (DART-MS) detection. The novelty of the instrument lies in its ability to monitor processes as a function of temperature through the simultaneous recording of images, quantitative color changes, and mass spectra. The capability of the instrument was demonstrated through successful application to four very varied systems including profiling an organic reaction, decomposition of silicone polymers, and the desorption of rhodamine B from an alumina surface. The multidimensional, real-time analytical data provided by this instrument allow for a much greater insight into thermal processes than could be achieved previously.

  5. Simulation and real-time analysis of pulse shapes from segmented HPGe-detectors

    Energy Technology Data Exchange (ETDEWEB)

    Schlarb, Michael Christian

    2009-11-17

    The capabilities of future HPGe arrays consisting of highly segmented detectors, like AGATA will depend heavily on the performance of {gamma}-ray tracking. The most crucial component in the whole concept is the pulse shape analysis (PSA). The working principle of PSA is to compare the experimental signal shape with signals available from a basis set with known interaction locations. The efficiency of the tracking algorithm hinges on the ability of the PSA to reconstruct the interaction locations accurately, especially for multiple {gamma}-interactions. Given the size of the arrays the PSA algorithm must be run in a real-time environment. A prerequisite to a successful PSA is an accurate knowledge of the detectors response. Making a full coincidence scan of a single AGATA detector, however takes between two and three months, which is too long to produce an experimental signal basis for all detector elements. A straight forward possibility is to use a precise simulation of the detector and to provide a basis of simulated signals. For this purpose the Java Agata Signal Simulation (JASS) was developed in the course of this thesis. The geometry of the detector is given with numerical precision and models describing the anisotropic mobilities of the charge carriers in germanium were taken from the literature. The pulse shapes of the transient and net-charge signals are calculated using weighting potentials on a finite grid. Special care was taken that the interpolation routine not only reproduces the weighting potentials precisely in the highly varying areas of the segment boundaries but also that its performance is independent of the location within the detector. Finally data from a coincidence scan and a pencil beam experiment were used to verify JASS. The experimental signals are reproduced accurately by the simulation. Pulse Shape Analysis (PSA) reconstructs the positions of the individual interactions and the corresponding energy deposits within the detector. This

  6. Simulation and real-time analysis of pulse shapes from segmented HPGe-detectors

    International Nuclear Information System (INIS)

    Schlarb, Michael Christian

    2009-01-01

    The capabilities of future HPGe arrays consisting of highly segmented detectors, like AGATA will depend heavily on the performance of γ-ray tracking. The most crucial component in the whole concept is the pulse shape analysis (PSA). The working principle of PSA is to compare the experimental signal shape with signals available from a basis set with known interaction locations. The efficiency of the tracking algorithm hinges on the ability of the PSA to reconstruct the interaction locations accurately, especially for multiple γ-interactions. Given the size of the arrays the PSA algorithm must be run in a real-time environment. A prerequisite to a successful PSA is an accurate knowledge of the detectors response. Making a full coincidence scan of a single AGATA detector, however takes between two and three months, which is too long to produce an experimental signal basis for all detector elements. A straight forward possibility is to use a precise simulation of the detector and to provide a basis of simulated signals. For this purpose the Java Agata Signal Simulation (JASS) was developed in the course of this thesis. The geometry of the detector is given with numerical precision and models describing the anisotropic mobilities of the charge carriers in germanium were taken from the literature. The pulse shapes of the transient and net-charge signals are calculated using weighting potentials on a finite grid. Special care was taken that the interpolation routine not only reproduces the weighting potentials precisely in the highly varying areas of the segment boundaries but also that its performance is independent of the location within the detector. Finally data from a coincidence scan and a pencil beam experiment were used to verify JASS. The experimental signals are reproduced accurately by the simulation. Pulse Shape Analysis (PSA) reconstructs the positions of the individual interactions and the corresponding energy deposits within the detector. This is

  7. Impact Analysis of Land Use on Traffic Congestion Using Real-Time Traffic and POI

    OpenAIRE

    Zhang, Tianqi; Sun, Lishan; Yao, Liya; Rong, Jian

    2017-01-01

    This paper proposed a new method to describe, compare, and classify the traffic congestion points in Beijing, China, by using the online map data and further revealed the relationship between traffic congestion and land use. The data of the point of interest (POI) and the real-time traffic was extracted from an electronic map of the area in the fourth ring road of Beijing. The POIs were quantified based on the architectural area of the land use; the congestion points were identified based on ...

  8. Real-time analysis and display of reactor system mass inventory

    International Nuclear Information System (INIS)

    Dao, L.T.; Meachum, T.R.

    1982-01-01

    A mass inventory system (MIS) to evaluate, in real-time, the coolant distribution within the primary coolant system of the Loss-of-Fluid Test (LOFT) reactor has been developed. The computer-based system calculates and displays the coolant levels by two methods: using level measurements and performing a mass balance. The MIS is designed to provide up-to-date, intelligible information on the coolant distribution during any LOFT experiment. During LOFT experiments in which the primary coolant pumps are on, the method also provides void fraction information and the anticipated liquid level in the reactor vessel should the pumps be turned off

  9. The use of real-time polymerase chain reaction with high resolution melting (real-time PCR-HRM) analysis for the detection and discrimination of nematodes Bursaphelenchus xylophilus and Bursaphelenchus mucronatus.

    Science.gov (United States)

    Filipiak, Anna; Hasiów-Jaroszewska, Beata

    2016-04-01

    The real-time PCR-HRM analysis was developed for the detection and discrimination of the quarantine nematode Bursaphelenchus xylophilus and Bursaphelenchus mucronatus. A set of primers was designed to target the ITS region of rDNA. The results have demonstrated that this analysis is a valuable tool for differentiation of these both species. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Isothermal microcalorimetry - A quantitative method to monitor Trypanosoma congolense growth and growth inhibition by trypanocidal drugs in real time.

    Science.gov (United States)

    Gysin, M; Braissant, O; Gillingwater, K; Brun, R; Mäser, P; Wenzler, T

    2018-03-16

    Trypanosoma congolense is a protozoan parasite that is transmitted by tsetse flies, causing African Animal Trypanosomiasis, also known as Nagana, in sub-Saharan Africa. Nagana is a fatal disease of livestock that causes severe economic losses. Two drugs are available, diminazene and isometamidium, yet successful treatment is jeopardized by drug resistant T. congolense. Isothermal microcalorimetry is a highly sensitive tool that can be used to study growth of the extracellular T. congolense parasites or to study parasite growth inhibition after the addition of antitrypanosomal drugs. Time of drug action and time to kill can be quantified in a simple way by real time heat flow measurements. We established a robust protocol for the microcalorimetric studies of T. congolense and developed mathematical computations in R to calculate different parameters related to growth and the kinetics of drug action. We demonstrate the feasibility and benefit of the method exemplary with the two standard drugs, diminazene aceturate and isometamidium chloride. The method and the mathematical approach can be translated to study other pathogenic or non-pathogenic cells if they are metabolically active and grow under axenic conditions. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

  11. Quantitative evaluation of alternatively spliced mRNA isoforms by label-free real-time plasmonic sensing.

    Science.gov (United States)

    Huertas, César S; Carrascosa, L G; Bonnal, S; Valcárcel, J; Lechuga, L M

    2016-04-15

    Alternative splicing of mRNA precursors enables cells to generate different protein outputs from the same gene depending on their developmental or homeostatic status. Its deregulation is strongly linked to disease onset and progression. Current methodologies for monitoring alternative splicing demand elaborate procedures and often present difficulties in discerning between closely related isoforms, e.g. due to cross-hybridization during their detection. Herein, we report a general methodology using a Surface Plasmon Resonance (SPR) biosensor for label-free monitoring of alternative splicing events in real-time, without any cDNA synthesis or PCR amplification requirements. We applied this methodology to RNA isolated from HeLa cells for the quantification of alternatively spliced isoforms of the Fas gene, involved in cancer progression through regulation of programmed cell death. We demonstrate that our methodology is isoform-specific, with virtually no cross-hybridization, achieving limits of detection (LODs) in the picoMolar (pM) range. Similar results were obtained for the detection of the BCL-X gene mRNA isoforms. The results were independently validated by RT-qPCR, with excellent concordance in the determination of isoform ratios. The simplicity and robustness of this biosensor technology can greatly facilitate the exploration of alternative splicing biomarkers in disease diagnosis and therapy. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Selection of suitable housekeeping genes for real-time quantitative PCR in CD4(+ lymphocytes from asthmatics with or without depression.

    Directory of Open Access Journals (Sweden)

    Ting Wang

    Full Text Available OBJECTIVE: No optimal housekeeping genes (HKGs have been identified for CD4(+ T cells from non-depressive asthmatic and depressive asthmatic adults for normalizing quantitative real-time PCR (qPCR assays. The aim of present study was to select appropriate HKGs for gene expression analysis in purified CD4(+ T cells from these asthmatics. METHODS: Three groups of subjects (Non-depressive asthmatic, NDA, n = 10, Depressive asthmatic, DA, n = 11, and Healthy control, HC, n = 10 respectively were studied. qPCR for 9 potential HKGs, namely RNA, 28S ribosomal 1 (RN28S1, ribosomal protein, large, P0 (RPLP0, actin, beta (ACTB, cyclophilin A (PPIA, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, phosphoglycerate kinase 1 (PGK1, beta-2-microglobulin (B2M, glucuronidase, beta (GUSB and ribosomal protein L13a (RPL13A, was performed. Then the data were analyzed with three different applications namely BestKeeper, geNorm, and NormFinder. RESULTS: The analysis of gene expression data identified B2M and RPLP0 as the most stable reference genes and showed that the level of PPIA was significantly different among subjects of three groups when the two best HKGs identified were applied. Post-hoc analysis by Student-Newman-Keuls correction shows that depressive asthmatics and non-depressive asthmatics exhibited lower expression level of PPIA than healthy controls (p<0.05. CONCLUSIONS: B2M and RPLP0 were identified as the most optimal HKGs in gene expression studies involving human blood CD4(+ T cells derived from normal, depressive asthmatics and non-depressive asthmatics. The suitability of using the PPIA gene as the HKG for such studies was questioned due to its low expression in asthmatics.

  13. Performance analysis and kernel size study of the Lynx real-time operating system

    Science.gov (United States)

    Liu, Yuan-Kwei; Gibson, James S.; Fernquist, Alan R.

    1993-01-01

    This paper analyzes the Lynx real-time operating system (LynxOS), which has been selected as the operating system for the Space Station Freedom Data Management System (DMS). The features of LynxOS are compared to other Unix-based operating system (OS). The tools for measuring the performance of LynxOS, which include a high-speed digital timer/counter board, a device driver program, and an application program, are analyzed. The timings for interrupt response, process creation and deletion, threads, semaphores, shared memory, and signals are measured. The memory size of the DMS Embedded Data Processor (EDP) is limited. Besides, virtual memory is not suitable for real-time applications because page swap timing may not be deterministic. Therefore, the DMS software, including LynxOS, has to fit in the main memory of an EDP. To reduce the LynxOS kernel size, the following steps are taken: analyzing the factors that influence the kernel size; identifying the modules of LynxOS that may not be needed in an EDP; adjusting the system parameters of LynxOS; reconfiguring the device drivers used in the LynxOS; and analyzing the symbol table. The reductions in kernel disk size, kernel memory size and total kernel size reduction from each step mentioned above are listed and analyzed.

  14. Analysis of relationships between hourly electricity price and load in deregulated real-time power markets

    International Nuclear Information System (INIS)

    Lo, K.L.; Wu, Y.K.

    2004-01-01

    Risk management in the electric power industry involves measuring the risk for all instruments owned by a company. The value of many of these instruments depends directly on electricity prices. In theory, the wholesale price in a real-time market should reflect the short-run marginal cost. However, most markets are not perfectly competitive, therefore by understanding the degree of correlation between price and physical drivers, electric traders and consumers can manage their risk more effectively and efficiently. Market data from two power-pool architectures, both pre-2003 ISO-NE and Australia's NEM, have been studied. The dynamic character of electricity price is mean-reverting, and consists of intra-day and weekly variations, seasonal fluctuations, and instant jumps. Parts of them are affected by load demands. Hourly signals on both price and load are divided into deterministic and random components with a discrete Fourier transform algorithm. Next, the real-time price-load relationship for periodic and random signals is examined. In addition, time-varying volatility models are constructed on random price and random load with the GARCH model, and the correlation between them analysed. Volatility plays a critical role on evaluating option pricing and risk management. (author)

  15. Real-time analysis of total, elemental, and total speciated mercury

    International Nuclear Information System (INIS)

    Schlager, R.J.; Wilson, K.G.; Sappey, A.D.

    1995-01-01

    ADA Technologies, Inc., is developing a continuous emissions monitoring system that measures the concentrations of mercury in flue gas. Mercury is emitted as an air pollutant from a number of industrial processes. The largest contributors of these emissions are coal and oil combustion, municipal waste combustion, medical waste combustion, and the thermal treatment of hazardous materials. It is difficult, time consuming, and expensive to measure mercury emissions using current testing methods. Part of the difficulty lies in the fact that mercury is emitted from sources in several different forms, such as elemental mercury and mercuric chloride. The ADA analyzer measures these emissions in real time, thus providing a number of advantages over existing test methods: (1) it will provide a real-time measure of emission rates, (2) it will assure facility operators, regulators, and the public that emissions control systems are working at peak efficiency, and (3) it will provide information as to the nature of the emitted mercury (elemental mercury or speciated compounds). This update presents an overview of the CEM and describes features of key components of the monitoring system--the mercury detector, a mercury species converter, and the analyzer calibration system

  16. Comparative Analysis of Neural Network Training Methods in Real-time Radiotherapy

    Directory of Open Access Journals (Sweden)

    Nouri S.

    2017-03-01

    Full Text Available Background: The motions of body and tumor in some regions such as chest during radiotherapy treatments are one of the major concerns protecting normal tissues against high doses. By using real-time radiotherapy technique, it is possible to increase the accuracy of delivered dose to the tumor region by means of tracing markers on the body of patients. Objective: This study evaluates the accuracy of some artificial intelligence methods including neural network and those of combination with genetic algorithm as well as particle swarm optimization (PSO estimating tumor positions in real-time radiotherapy. Method: One hundred recorded signals of three external markers were used as input data. The signals from 3 markers thorough 10 breathing cycles of a patient treated via a cyber-knife for a lung tumor were used as data input. Then, neural network method and its combination with genetic or PSO algorithms were applied determining the tumor locations using MATLAB© software program. Results: The accuracies were obtained 0.8%, 12% and 14% in neural network, genetic and particle swarm optimization algorithms, respectively. Conclusion: The internal target volume (ITV should be determined based on the applied neural network algorithm on training steps.

  17. Energy-aware embedded classifier design for real-time emotion analysis.

    Science.gov (United States)

    Padmanabhan, Manoj; Murali, Srinivasan; Rincon, Francisco; Atienza, David

    2015-01-01

    Detection and classification of human emotions from multiple bio-signals has a wide variety of applications. Though electronic devices are available in the market today that acquire multiple body signals, the classification of human emotions in real-time, adapted to the tight energy budgets of wearable embedded systems is a big challenge. In this paper we present an embedded classifier for real-time emotion classification. We propose a system that operates at different energy budgeted modes, depending on the available energy, where each mode is constrained by an operating energy bound. The classifier has an offline training phase where feature selection is performed for each operating mode, with an energy-budget aware algorithm that we propose. Across the different operating modes, the classification accuracy ranges from 95% - 75% and 89% - 70% for arousal and valence respectively. The accuracy is traded off for less power consumption, which results in an increased battery life of up to 7.7 times (from 146.1 to 1126.9 hours).

  18. [Exoskeleton robot system based on real-time gait analysis for walking assist].

    Science.gov (United States)

    Xie, Zheng; Wang, Mingjiang; Huang, Wulong; Yong, Shanshan; Wang, Xin'an

    2017-04-01

    This paper presents a wearable exoskeleton robot system to realize walking assist function, which oriented toward the patients or the elderly with the mild impairment of leg movement function, due to illness or natural aging. It reduces the loads of hip, knee, ankle and leg muscles during walking by way of weight support. In consideration of the characteristics of the psychological demands and the disease, unlike the weight loss system in the fixed or followed rehabilitation robot, the structure of the proposed exoskeleton robot is artistic, lightweight and portable. The exoskeleton system analyzes the user's gait real-timely by the plantar pressure sensors to divide gait phases, and present different control strategies for each gait phase. The pressure sensors in the seat of the exoskeleton system provide real-time monitoring of the support efforts. And the drive control uses proportion-integral-derivative (PID) control technology for torque control. The total weight of the robot system is about 12.5 kg. The average of the auxiliary support is about 10 kg during standing, and it is about 3 kg during walking. The system showed, in the experiments, a certain effect of weight support, and reduction of the pressure on the lower limbs to walk and stand.

  19. 3D Hand Gesture Analysis through a Real-Time Gesture Search Engine

    Directory of Open Access Journals (Sweden)

    Shahrouz Yousefi

    2015-06-01

    Full Text Available 3D gesture recognition and tracking are highly desired features of interaction design in future mobile and smart environments. Specifically, in virtual/augmented reality applications, intuitive interaction with the physical space seems unavoidable and 3D gestural interaction might be the most effective alternative for the current input facilities such as touchscreens. In this paper, we introduce a novel solution for real-time 3D gesture-based interaction by finding the best match from an extremely large gesture database. This database includes images of various articulated hand gestures with the annotated 3D position/orientation parameters of the hand joints. Our unique matching algorithm is based on the hierarchical scoring of the low-level edge-orientation features between the query frames and database and retrieving the best match. Once the best match is found from the database in each moment, the pre-recorded 3D motion parameters can instantly be used for natural interaction. The proposed bare-hand interaction technology performs in real time with high accuracy using an ordinary camera.

  20. Real-time analysis of total, elemental, and total speciated mercury

    Energy Technology Data Exchange (ETDEWEB)

    Schlager, R.J.; Wilson, K.G.; Sappey, A.D. [ADA Technologies, Inc., Englewood, CO (United States)

    1995-11-01

    ADA Technologies, Inc., is developing a continuous emissions monitoring system that measures the concentrations of mercury in flue gas. Mercury is emitted as an air pollutant from a number of industrial processes. The largest contributors of these emissions are coal and oil combustion, municipal waste combustion, medical waste combustion, and the thermal treatment of hazardous materials. It is difficult, time consuming, and expensive to measure mercury emissions using current testing methods. Part of the difficulty lies in the fact that mercury is emitted from sources in several different forms, such as elemental mercury and mercuric chloride. The ADA analyzer measures these emissions in real time, thus providing a number of advantages over existing test methods: (1) it will provide a real-time measure of emission rates, (2) it will assure facility operators, regulators, and the public that emissions control systems are working at peak efficiency, and (3) it will provide information as to the nature of the emitted mercury (elemental mercury or speciated compounds). This update presents an overview of the CEM and describes features of key components of the monitoring system--the mercury detector, a mercury species converter, and the analyzer calibration system.

  1. Power system analysis of Hanlim superconducting HVDC system using real time digital simulator

    International Nuclear Information System (INIS)

    Won, Y.J.; Kim, J.G.; Kim, A.R.; Kim, G.H.; Park, M.; Yu, I.K.; Sim, K.D.; Cho, J.; Lee, S.; Jeong, K.W.; Watanabe, K.

    2011-01-01

    KEPCO has planned to construct a test site for renewable energy in Jeju power system. One kilometer length of total 8 km was designed as superconducting DC cable. We have developed a simulation model of the 8 km HVDC system using real time digital simulator. The simulation result shows that the HVDC line was not affected by wind power variation. Jeju island is located approximately 100 km south from the mainland of Korea, and had a peak load of about 553 MW in 2008. The demand increases 7.2% a year over the last 5 years. Since the wind profiles of Jeju island are more favorable than mainland of Korea, many companies have shown interest in the wind power business at the Jeju island. Moreover KEPCO has a plan for renewable energy test too whose power will be delivered by HVDC system. One kilometer length of total 8 km was designed as superconducting DC cable. Rest 7 km will be the conventional overhead line. In this paper, the authors have developed a simulation model of the power network around 8 km HVDC system using real time digital simulator (RTDS).

  2. Exploring inter-frame correlation analysis and wavelet-domain modeling for real-time caption detection in streaming video

    Science.gov (United States)

    Li, Jia; Tian, Yonghong; Gao, Wen

    2008-01-01

    In recent years, the amount of streaming video has grown rapidly on the Web. Often, retrieving these streaming videos offers the challenge of indexing and analyzing the media in real time because the streams must be treated as effectively infinite in length, thus precluding offline processing. Generally speaking, captions are important semantic clues for video indexing and retrieval. However, existing caption detection methods often have difficulties to make real-time detection for streaming video, and few of them concern on the differentiation of captions from scene texts and scrolling texts. In general, these texts have different roles in streaming video retrieval. To overcome these difficulties, this paper proposes a novel approach which explores the inter-frame correlation analysis and wavelet-domain modeling for real-time caption detection in streaming video. In our approach, the inter-frame correlation information is used to distinguish caption texts from scene texts and scrolling texts. Moreover, wavelet-domain Generalized Gaussian Models (GGMs) are utilized to automatically remove non-text regions from each frame and only keep caption regions for further processing. Experiment results show that our approach is able to offer real-time caption detection with high recall and low false alarm rate, and also can effectively discern caption texts from the other texts even in low resolutions.

  3. Direct Analysis in Real Time Mass Spectrometry of Potential By-Products from Homemade Nitrate Ester Explosive Synthesis

    OpenAIRE

    Sisco, Edward; Forbes, Thomas P.

    2015-01-01

    This work demonstrates the coupling of direct analysis in real time (DART) ionization with time-of-flight mass spectrometry (MS) in an off-axis configuration for the trace detection and analysis of potential partially nitrated and dimerized by-products of homemade nitrate ester explosive synthesis. Five compounds relating to the synthesis of nitroglycerin (NG) and pentaerythritol tetranitrate (PETN) were examined. Deprotonated ions and adducts with molecular oxygen, nitrite, and nitrate were ...

  4. Analysis of the phenomena associated with structural damage using real time vibration analysis

    International Nuclear Information System (INIS)

    Garcia Peyrano, O; Cismondi, L; Damiani, H; Torres, E

    2004-01-01

    It is of interest to have analytical methodologies available for the dynamic behavior of large mechanical structures like those in thermal cycle systems of nuclear power plants or in transport systems during the experimental stage prior to their construction, as happens in aeronautics, where prototypes are tested in experimental banks on a scale of 1 to 1. The same does not occur with systems for the generation of electrical energy such as a nuclear power plant or in ships, competition automobiles, railway systems, etc. Not because of the technical impossibility but because of the high costs involved. This work aims to implement a technology based on the analysis of the vibrations to obtain a profile of the modal dynamic response and its influence on the critical components of the mechanisms with the particularity of detecting the preventive location of the component that may suffer a potential damage. The Vibrations Analysis Laboratory has resolved different cases in the Embalse Nuclear Plant, in the Atucha Nuclear Plant, in the Heavy Water Industrial Plant, in the automobile industry and in other industrial areas (CW)

  5. The use of laser microdissection in the identification of suitable reference genes for normalization of quantitative real-time PCR in human FFPE epithelial ovarian tissue samples.

    Directory of Open Access Journals (Sweden)

    Jing Cai

    Full Text Available Quantitative real-time PCR (qPCR is a powerful and reproducible method of gene expression analysis in which expression levels are quantified by normalization against reference genes. Therefore, to investigate the potential biomarkers and therapeutic targets for epithelial ovarian cancer by qPCR, it is critical to identify stable reference genes. In this study, twelve housekeeping genes (ACTB, GAPDH, 18S rRNA, GUSB, PPIA, PBGD, PUM1, TBP, HRPT1, RPLP0, RPL13A, and B2M were analyzed in 50 ovarian samples from normal, benign, borderline, and malignant tissues. For reliable results, laser microdissection (LMD, an effective technique used to prepare homogeneous starting material, was utilized to precisely excise target tissues or cells. One-way analysis of variance (ANOVA and nonparametric (Kruskal-Wallis tests were used to compare the expression differences. NormFinder and geNorm software were employed to further validate the suitability and stability of the candidate genes. Results showed that epithelial cells occupied a small percentage of the normal ovary indeed. The expression of ACTB, PPIA, RPL13A, RPLP0, and TBP were stable independent of the disease progression. In addition, NormFinder and geNorm identified the most stable combination (ACTB, PPIA, RPLP0, and TBP and the relatively unstable reference gene GAPDH from the twelve commonly used housekeeping genes. Our results highlight the use of homogeneous ovarian tissues and multiple-reference normalization strategy, e.g. the combination of ACTB, PPIA, RPLP0, and TBP, for qPCR in epithelial ovarian tissues, whereas GAPDH, the most commonly used reference gene, is not recommended, especially as a single reference gene.

  6. Selection of Suitable Internal Control Genes for Accurate Normalization of Real-Time Quantitative PCR Data of Buffalo (Bubalus bubalis) Blastocysts Produced by SCNT and IVF.

    Science.gov (United States)

    Sood, Tanushri Jerath; Lagah, Swati Viviyan; Sharma, Ankita; Singla, Suresh Kumar; Mukesh, Manishi; Chauhan, Manmohan Singh; Manik, Radheysham; Palta, Prabhat

    2017-10-01

    We evaluated the suitability of 10 candidate internal control genes (ICGs), belonging to different functional classes, namely ACTB, EEF1A1, GAPDH, HPRT1, HMBS, RPS15, RPS18, RPS23, SDHA, and UBC for normalizing the real-time quantitative polymerase chain reaction (qPCR) data of blastocyst-stage buffalo embryos produced by hand-made cloning and in vitro fertilization (IVF). Total RNA was isolated from three pools, each of cloned and IVF blastocysts (n = 50/pool) for cDNA synthesis. Two different statistical algorithms geNorm and NormFinder were used for evaluating the stability of these genes. Based on gene stability measure (M value) and pairwise variation (V value), calculated by geNorm analysis, the most stable ICGs were RPS15, HPRT1, and ACTB for cloned blastocysts, HMBS, UBC, and HPRT1 for IVF blastocysts and RPS15, GAPDH, and HPRT1 for both the embryo types analyzed together. RPS18 was the least stable gene for both cloned and IVF blastocysts. Following NormFinder analysis, the order of stability was RPS15 = HPRT1>GAPDH for cloned blastocysts, HMBS = UBC>RPS23 for IVF blastocysts, and HPRT1>GAPDH>RPS15 for cloned and IVF blastocysts together. These results suggest that despite overlapping of the three most stable ICGs between cloned and IVF blastocysts, the panel of ICGs selected for normalization of qPCR data of cloned and IVF blastocyst-stage embryos should be different.

  7. The use of laser microdissection in the identification of suitable reference genes for normalization of quantitative real-time PCR in human FFPE epithelial ovarian tissue samples.

    Science.gov (United States)

    Cai, Jing; Li, Tao; Huang, Bangxing; Cheng, Henghui; Ding, Hui; Dong, Weihong; Xiao, Man; Liu, Ling; Wang, Zehua

    2014-01-01

    Quantitative real-time PCR (qPCR) is a powerful and reproducible method of gene expression analysis in which expression levels are quantified by normalization against reference genes. Therefore, to investigate the potential biomarkers and therapeutic targets for epithelial ovarian cancer by qPCR, it is critical to identify stable reference genes. In this study, twelve housekeeping genes (ACTB, GAPDH, 18S rRNA, GUSB, PPIA, PBGD, PUM1, TBP, HRPT1, RPLP0, RPL13A, and B2M) were analyzed in 50 ovarian samples from normal, benign, borderline, and malignant tissues. For reliable results, laser microdissection (LMD), an effective technique used to prepare homogeneous starting material, was utilized to precisely excise target tissues or cells. One-way analysis of variance (ANOVA) and nonparametric (Kruskal-Wallis) tests were used to compare the expression differences. NormFinder and geNorm software were employed to further validate the suitability and stability of the candidate genes. Results showed that epithelial cells occupied a small percentage of the normal ovary indeed. The expression of ACTB, PPIA, RPL13A, RPLP0, and TBP were stable independent of the disease progression. In addition, NormFinder and geNorm identified the most stable combination (ACTB, PPIA, RPLP0, and TBP) and the relatively unstable reference gene GAPDH from the twelve commonly used housekeeping genes. Our results highlight the use of homogeneous ovarian tissues and multiple-reference normalization strategy, e.g. the combination of ACTB, PPIA, RPLP0, and TBP, for qPCR in epithelial ovarian tissues, whereas GAPDH, the most commonly used reference gene, is not recommended, especially as a single reference gene.

  8. Hyperspectral Imaging Using Intracellular Spies: Quantitative Real-Time Measurement of Intracellular Parameters In Vivo during Interaction of the Pathogenic Fungus Aspergillus fumigatus with Human Monocytes.

    Directory of Open Access Journals (Sweden)

    Sara Mohebbi

    Full Text Available Hyperspectral imaging (HSI is a technique based on the combination of classical spectroscopy and conventional digital image processing. It is also well suited for the biological assays and quantitative real-time analysis since it provides spectral and spatial data of samples. The method grants detailed information about a sample by recording the entire spectrum in each pixel of the whole image. We applied HSI to quantify the constituent pH variation in a single infected apoptotic monocyte as a model system. Previously, we showed that the human-pathogenic fungus Aspergillus fumigatus conidia interfere with the acidification of phagolysosomes. Here, we extended this finding to monocytes and gained a more detailed analysis of this process. Our data indicate that melanised A. fumigatus conidia have the ability to interfere with apoptosis in human monocytes as they enable the apoptotic cell to recover from mitochondrial acidification and to continue with the cell cycle. We also showed that this ability of A. fumigatus is dependent on the presence of melanin, since a non-pigmented mutant did not stop the progression of apoptosis and consequently, the cell did not recover from the acidic pH. By conducting the current research based on the HSI, we could measure the intracellular pH in an apoptotic infected human monocyte and show the pattern of pH variation during 35 h of measurements. As a conclusion, we showed the importance of melanin for determining the fate of intracellular pH in a single apoptotic cell.

  9. Real-time cellular exometabolome analysis with a microfluidic-mass spectrometry platform.

    Directory of Open Access Journals (Sweden)

    Christina C Marasco

    Full Text Available To address the challenges of tracking the multitude of signaling molecules and metabolites that is the basis of biological complexity, we describe a strategy to expand the analytical techniques for dynamic systems biology. Using microfluidics, online desalting, and mass spectrometry technologies, we constructed and validated a platform well suited for sampling the cellular microenvironment with high temporal resolution. Our platform achieves success in: automated cellular stimulation and microenvironment control; reduced non-specific adsorption to polydimethylsiloxane due to surface passivation; real-time online sample collection; near real-time sample preparation for salt removal; and real-time online mass spectrometry. When compared against the benchmark of "in-culture" experiments combined with ultraperformance liquid chromatography-electrospray ionization-ion mobility-mass spectrometry (UPLC-ESI-IM-MS, our platform alleviates the volume challenge issues caused by dilution of autocrine and paracrine signaling and dramatically reduces sample preparation and data collection time, while reducing undesirable external influence from various manual methods of manipulating cells and media (e.g., cell centrifugation. To validate this system biologically, we focused on cellular responses of Jurkat T cells to microenvironmental stimuli. Application of these stimuli, in conjunction with the cell's metabolic processes, results in changes in consumption of nutrients and secretion of biomolecules (collectively, the exometabolome, which enable communication with other cells or tissues and elimination of waste. Naïve and experienced T-cell metabolism of cocaine is used as an exemplary system to confirm the platform's capability, highlight its potential for metabolite discovery applications, and explore immunological memory of T-cell drug exposure. Our platform proved capable of detecting metabolomic variations between naïve and experienced Jurkat T cells

  10. Analysis of carbohydrates in Fusarium verticillioides using size-exclusion HPLC – DRI and direct analysis in real time ionization – time-of-flight – mass spectrometry (DART-MS)

    Science.gov (United States)

    Direct analysis in real time ionization – time-of-flight – mass spectrometry (DART-MS) and size-exclusion HPLC – DRI are used, respectively, to qualitatively and quantitatively determine the carbohydrates extracted from the corn rot fungus Fusarium verticillioides. In situ permethylation in the DART...

  11. Selection of suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using quantitative real-time polymerase chain reaction

    DEFF Research Database (Denmark)

    Zornhagen, K. W.; Kristensen, A. T.; Hansen, Anders Elias

    2015-01-01

    Quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) is a sensitive technique for quantifying gene expression. Stably expressed reference genes are necessary for normalization of RT-qPCR data. Only a few articles have been published on reference genes in canine tumours....... The objective of this study was to demonstrate how to identify suitable reference genes for normalization of genes of interest in canine soft tissue sarcomas using RT-qPCR. Primer pairs for 17 potential reference genes were designed and tested in archival tumour biopsies from six dogs. The geNorm algorithm...

  12. The developmental stage of strongyle eggs affects the outcome of real-time PCR analysis

    DEFF Research Database (Denmark)

    Roust, Tina; Haakansson, Ida T.; Rhod, Maria

    Several molecular diagnostic tests are based upon measuring and quantifying DNA obtained from parasite eggs. It is well-known that such eggs undergo development during storage, but it remains unknown to which extent the stage of development can affect the diagnostic test result. This project...... investigated whether the developmental stage of strongyle eggs affects real-time polymerase chain reaction (PCR) results. Mixed species strongyle eggs were obtained from the faeces of a naturally infected horse. Eggs were isolated and placed in microtiter plates with demineralised water. A total of 25 wells...... containing 100 eggs each were set up and kept refrigerated for up to five days. Once daily, five wells were microscopied on an inverted microscope, the developmental stages of the eggs were noted, and the eggs harvested for DNA extraction. The protocol was repeated three times. Genomic DNA was extracted...

  13. Analysis of several digital network technologies for hard real-time communications in nuclear plant

    International Nuclear Information System (INIS)

    Song, Ki Sang; No, Hee Chun

    1999-01-01

    Applying digital network technology for advanced nuclear plant requires deterministic communication for tight safety requirements, timely and reliable data delivery for operation critical and mission-critical characteristics of nuclear plant. Communication protocols, such as IEEE 802/4 Tiken Bus, IEEE 802/5 Token Ring, FDDI, and ARCnet, which have deterministic communication capability are partially applied to several nuclear power plants. Although digital communication technologies have many advantages, it is necessary to consider the noise immunity form electromagnetic interference (EMI), electrical interference, impulse noise, and heat noise before selecting specific digital network technology for nuclear plant. In this paper, we consider the token frame loss and data frame loss rate due to the link error event, frame size, and link data rate in different protocols, and evaluate the possibility of failure to meet the hard real-time requirement in nuclear plant. (author). 11 refs., 3 figs., 4 tabs

  14. Power system analysis of Hanlim superconducting HVDC system using real time digital simulator

    Science.gov (United States)

    Won, Y. J.; Kim, J. G.; Kim, A. R.; Kim, G. H.; Park, M.; Yu, I. K.; Sim, K. D.; Cho, J.; Lee, S.; Jeong, K. W.; Watanabe, K.

    2011-11-01

    Jeju island is located approximately 100 km south from the mainland of Korea, and had a peak load of about 553 MW in 2008. The demand increases 7.2% a year over the last 5 years. Since the wind profiles of Jeju island are more favorable than mainland of Korea, many companies have shown interest in the wind power business at the Jeju island. Moreover KEPCO has a plan for renewable energy test too whose power will be delivered by HVDC system. One kilometer length of total 8 km was designed as superconducting DC cable. Rest 7 km will be the conventional overhead line. In this paper, the authors have developed a simulation model of the power network around 8 km HVDC system using real time digital simulator (RTDS).

  15. Measurement and analysis of workload effects on fault latency in real-time systems

    Science.gov (United States)

    Woodbury, Michael H.; Shin, Kang G.

    1990-01-01

    The authors demonstrate the need to address fault latency in highly reliable real-time control computer systems. It is noted that the effectiveness of all known recovery mechanisms is greatly reduced in the presence of multiple latent faults. The presence of multiple latent faults increases the possibility of multiple errors, which could result in coverage failure. The authors present experimental evidence indicating that the duration of fault latency is dependent on workload. A synthetic workload generator is used to vary the workload, and a hardware fault injector is applied to inject transient faults of varying durations. This method makes it possible to derive the distribution of fault latency duration. Experimental results obtained from the fault-tolerant multiprocessor at the NASA Airlab are presented and discussed.

  16. Assessing HER2 amplification by IHC, FISH, and real-time polymerase chain reaction analysis (real-time PCR) following LCM in formalin-fixed paraffin embedded tissue from 40 women with ovarian cancer

    DEFF Research Database (Denmark)

    Hillig, Thore; Thode, Jørgen; Breinholt, Ellen Marie

    2012-01-01

    . Only few ovarian cancer patients were HER2 overexpressed measured by IHC or FISH and thus could be eligible for antibody-based therapy with trastuzumab (Herceptin). Interestingly, we find an increased number of HER2 positive patients by real-time PCR analysis on microdissected cancer cells, suggesting...

  17. Analysis of hairy root culture of Rauvolfia serpentina using direct analysis in real time mass spectrometric technique.

    Science.gov (United States)

    Madhusudanan, K P; Banerjee, Suchitra; Khanuja, Suman P S; Chattopadhyay, Sunil K

    2008-06-01

    The applicability of a new mass spectrometric technique, DART (direct analysis in real time) has been studied in the analysis of the hairy root culture of Rauvolfia serpentina. The intact hairy roots were analyzed by holding them in the gap between the DART source and the mass spectrometer for measurements. Two nitrogen-containing compounds, vomilenine and reserpine, were characterized from the analysis of the hairy roots almost instantaneously. The confirmation of the structures of the identified compounds was made through their accurate molecular formula determinations. This is the first report of the application of DART technique for the characterization of compounds that are expressed in the hairy root cultures of Rauvolfia serpentina. Moreover, this also constitutes the first report of expression of reserpine in the hairy root culture of Rauvolfia serpentina. Copyright (c) 2008 John Wiley & Sons, Ltd.

  18. Development of a Rapid Real-Time PCR Assay for Quantitation of Pneumocystis carinii f. sp. Carinii

    DEFF Research Database (Denmark)

    Larsen, Hans Henrik; Kovacs, Joseph A; Stock, Frida

    2002-01-01

    A method for reliable quantification of Pneumocystis carinii in research models of P. carinii pneumonia (PCP) that is more convenient and reproducible than microscopic enumeration of organisms would greatly facilitate investigations of this organism. We developed a rapid quantitative touchdown (QTD......) PCR assay for detecting P. carinii f. sp. carinii, the subspecies of P. carinii commonly used in research models of PCP. The assay was based on the single-copy dihydrofolate reductase gene and was able to detect ... 6 log values for standards containing > or =5 copies/tube. Application of the assay to a series of 10-fold dilutions of P. carinii organisms isolated from rat lung demonstrated that it was reproducibly quantitative over 5 log values (r = 0.99). The assay was applied to a recently reported in vitro...

  19. Development of a Rapid Real-Time PCR Assay for Quantitation of Pneumocystis carinii f. sp. Carinii

    DEFF Research Database (Denmark)

    Larsen, Hans Henrik; Kovacs, Joseph A; Stock, Frida

    2002-01-01

    6 log values for standards containing > or =5 copies/tube. Application of the assay to a series of 10-fold dilutions of P. carinii organisms isolated from rat lung demonstrated that it was reproducibly quantitative over 5 log values (r = 0.99). The assay was applied to a recently reported in vitro...... axenic cultivation system for P. carinii and confirmed our microscopy findings that no organism multiplication had occurred during culture. For all cultures analyzed, QTD PCR assays showed a decrease in P. carinii DNA that exceeded the expected decrease due to dilution of the inoculum upon transfer......A method for reliable quantification of Pneumocystis carinii in research models of P. carinii pneumonia (PCP) that is more convenient and reproducible than microscopic enumeration of organisms would greatly facilitate investigations of this organism. We developed a rapid quantitative touchdown (QTD...

  20. Development of a Rapid Real-Time PCR Assay for Quantitation of Pneumocystis carinii f. sp. Carinii

    DEFF Research Database (Denmark)

    Larsen, Hans Henrik; Kovacs, Joseph A; Stock, Frida

    2002-01-01

    ) PCR assay for detecting P. carinii f. sp. carinii, the subspecies of P. carinii commonly used in research models of PCP. The assay was based on the single-copy dihydrofolate reductase gene and was able to detect r = 0.99) over...... 6 log values for standards containing > or =5 copies/tube. Application of the assay to a series of 10-fold dilutions of P. carinii organisms isolated from rat lung demonstrated that it was reproducibly quantitative over 5 log values (r = 0.99). The assay was applied to a recently reported in vitro....... In conclusion, a rapid, sensitive, and reproducible quantitative PCR assay for P. carinii f. sp. carinii has been developed and is applicable to in vivo as well as in vitro systems. The assay should prove useful for conducting studies in which quantification of organism burden or growth assessment is critical...

  1. Quantification of viable bacterial starter cultures of Virgibacillus sp. and Tetragenococcus halophilus in fish sauce fermentation by real-time quantitative PCR.

    Science.gov (United States)

    Udomsil, Natteewan; Chen, Shu; Rodtong, Sureelak; Yongsawatdigul, Jirawat

    2016-08-01

    Real-time quantitative polymerase chain reaction (qPCR) methods were developed for the quantification of Virgibacillus sp. SK37 and Tetragenococcus halophilus MS33, which were added as starter cultures in fish sauce fermentation. The PCR assays were coupled with propidium monoazide (PMA) treatment of samples to selectively quantify viable cells and integrated with exogenous recombinant Escherichia coli cells to control variabilities in analysis procedures. The qPCR methods showed species-specificity for both Virgibacillus halodenitrificans and T. halophilus as evaluated using 6 reference strains and 28 strains of bacteria isolated from fish sauce fermentation. The qPCR efficiencies were 101.1% for V. halodenitrificans and 90.2% for T. halophilus. The quantification limits of the assays were 10(3) CFU/mL and 10(2) CFU/mL in fish sauce samples with linear correlations over 4 Logs for V. halodenitrificans and T. halophilus, respectively. The matrix effect was not observed when evaluated using fish sauce samples fermented for 1-6 months. The developed PMA-qPCR methods were successfully applied to monitor changes of Virgibacillus sp. SK37 and T. halophilus MS33 in a mackerel fish sauce fermentation model where culture-dependent techniques failed to quantify the starter cultures. The results demonstrated the usability of the methods as practical tools for monitoring the starter cultures in fish sauce fermentation. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Selection of Reliable Reference Genes for Gene Expression Studies in the Biofuel Plant Jatropha curcas Using Real-Time Quantitative PCR

    Directory of Open Access Journals (Sweden)

    Lu Zhang

    2013-12-01

    Full Text Available Jatropha curcas is a promising renewable feedstock for biodiesel and bio-jet fuel production. To study gene expression in Jatropha in different tissues throughout development and under stress conditions, we examined a total of 11 typical candidate reference genes using real-time quantitative polymerase chain reaction (RT-qPCR analysis, which is widely used for validating transcript levels in gene expression studies. The expression stability of these candidate reference genes was assessed across a total of 20 samples, including various tissues at vegetative and reproductive stages and under desiccation and cold stress treatments. The results obtained using software qBasePLUS showed that the top-ranked reference genes differed across the sample subsets. The combination of actin, GAPDH, and EF1α would be appropriate as a reference panel for normalizing gene expression data across samples at different developmental stages; the combination of actin, GAPDH, and TUB5 should be used as a reference panel for normalizing gene expression data across samples under various abiotic stress treatments. With regard to different developmental stages, we recommend the use of actin and TUB8 for normalization at the vegetative stage and GAPDH and EF1α for normalization at the reproductive stage. For abiotic stress treatments, we recommend the use of TUB5 and TUB8 for normalization under desiccation stress and GAPDH and actin for normalization under cold stress. These results are valuable for future research on gene expression during development or under abiotic stress in Jatropha. To our knowledge, this is the first report on the stability of reference genes in Jatropha.

  3. Identification of human intestinal parasites affecting an asymptomatic peri-urban Argentinian population using multi-parallel quantitative real-time polymerase chain reaction.

    Science.gov (United States)

    Cimino, Rubén O; Jeun, Rebecca; Juarez, Marisa; Cajal, Pamela S; Vargas, Paola; Echazú, Adriana; Bryan, Patricia E; Nasser, Julio; Krolewiecki, Alejandro; Mejia, Rojelio

    2015-07-17

    In resource-limited countries, stool microscopy is the diagnostic test of choice for intestinal parasites (soil-transmitted helminths and/or intestinal protozoa). However, sensitivity and specificity is low. Improved diagnosis of intestinal parasites is especially important for accurate measurements of prevalence and intensity of infections in endemic areas. The study was carried out in Orán, Argentina. A total of 99 stool samples from a local surveillance campaign were analyzed by concentration microscopy and McMaster egg counting technique compared to the analysis by multi-parallel quantitative real-time polymerase chain reaction (qPCR). This study compared the performance of qPCR assay and stool microscopy for 8 common intestinal parasites that infect humans including the helminths Ascaris lumbricoides, Ancylostoma duodenale, Necator americanus, Strongyloides stercoralis, Trichuris trichiura, and the protozoa Giardia lamblia, Cryptosporidium parvum/hominis, and Entamoeba histolytica, and investigated the prevalence of polyparasitism in an endemic area. qPCR showed higher detection rates for all parasites as compared to stool microscopy except T. trichiura. Species-specific primers and probes were able to distinguish between A. duodenale (19.1%) and N. americanus (36.4%) infections. There were 48.6% of subjects co-infected with both hookworms, and a significant increase in hookworm DNA for A. duodenale versus N. americanus (119.6 fg/μL: 0.63 fg/μL, P parasites in an endemic area that has improved diagnostic accuracy compared to stool microscopy. This first time use of multi-parallel qPCR in Argentina has demonstrated the high prevalence of intestinal parasites in a peri-urban area. These results will contribute to more accurate epidemiological survey, refined treatment strategies on a public scale, and better health outcomes in endemic settings.

  4. Selection of Reliable Reference Genes for Gene Expression Studies of a Promising Oilseed Crop, Plukenetia volubilis, by Real-Time Quantitative PCR

    Directory of Open Access Journals (Sweden)

    Longjian Niu

    2015-06-01

    Full Text Available Real-time quantitative PCR (RT-qPCR is a reliable and widely used method for gene expression analysis. The accuracy of the determination of a target gene expression level by RT-qPCR demands the use of appropriate reference genes to normalize the mRNA levels among different samples. However, suitable reference genes for RT-qPCR have not been identified in Sacha inchi (Plukenetia volubilis, a promising oilseed crop known for its polyunsaturated fatty acid (PUFA-rich seeds. In this study, using RT-qPCR, twelve candidate reference genes were examined in seedlings and adult plants, during flower and seed development and for the entire growth cycle of Sacha inchi. Four statistical algorithms (delta cycle threshold (ΔCt, BestKeeper, geNorm, and NormFinder were used to assess the expression stabilities of the candidate genes. The results showed that ubiquitin-conjugating enzyme (UCE, actin (ACT and phospholipase A22 (PLA were the most stable genes in Sacha inchi seedlings. For roots, stems, leaves, flowers, and seeds from adult plants, 30S ribosomal protein S13 (RPS13, cyclophilin (CYC and elongation factor-1alpha (EF1α were recommended as reference genes for RT-qPCR. During the development of reproductive organs, PLA, ACT and UCE were the optimal reference genes for flower development, whereas UCE, RPS13 and RNA polymerase II subunit (RPII were optimal for seed development. Considering the entire growth cycle of Sacha inchi, UCE, ACT and EF1α were sufficient for the purpose of normalization. Our results provide useful guidelines for the selection of reliable reference genes for the normalization of RT-qPCR data for seedlings and adult plants, for reproductive organs, and for the entire growth cycle of Sacha inchi.

  5. Selection of Reliable Reference Genes for Gene Expression Studies of a Promising Oilseed Crop, Plukenetia volubilis, by Real-Time Quantitative PCR

    Science.gov (United States)

    Niu, Longjian; Tao, Yan-Bin; Chen, Mao-Sheng; Fu, Qiantang; Li, Chaoqiong; Dong, Yuling; Wang, Xiulan; He, Huiying; Xu, Zeng-Fu

    2015-01-01

    Real-time quantitative PCR (RT-qPCR) is a reliable and widely used method for gene expression analysis. The accuracy of the determination of a target gene expression level by RT-qPCR demands the use of appropriate reference genes to normalize the mRNA levels among different samples. However, suitable reference genes for RT-qPCR have not been identified in Sacha inchi (Plukenetia volubilis), a promising oilseed crop known for its polyunsaturated fatty acid (PUFA)-rich seeds. In this study, using RT-qPCR, twelve candidate reference genes were examined in seedlings and adult plants, during flower and seed development and for the entire growth cycle of Sacha inchi. Four statistical algorithms (delta cycle threshold (ΔCt), BestKeeper, geNorm, and NormFinder) were used to assess the expression stabilities of the candidate genes. The results showed that ubiquitin-conjugating enzyme (UCE), actin (ACT) and phospholipase A22 (PLA) were the most stable genes in Sacha inchi seedlings. For roots, stems, leaves, flowers, and seeds from adult plants, 30S ribosomal protein S13 (RPS13), cyclophilin (CYC) and elongation factor-1alpha (EF1α) were recommended as reference genes for RT-qPCR. During the development of reproductive organs, PLA, ACT and UCE were the optimal reference genes for flower development, whereas UCE, RPS13 and RNA polymerase II subunit (RPII) were optimal for seed development. Considering the entire growth cycle of Sacha inchi, UCE, ACT and EF1α were sufficient for the purpose of normalization. Our results provide useful guidelines for the selection of reliable reference genes for the normalization of RT-qPCR data for seedlings and adult plants, for reproductive organs, and for the entire growth cycle of Sacha inchi. PMID:26047338

  6. Rapid Detection and Differentiation of Clonorchis sinensis and Opisthorchis viverrini Using Real-Time PCR and High Resolution Melting Analysis

    OpenAIRE

    Cai, Xian-Quan; Yu, Hai-Qiong; Li, Rong; Yue, Qiao-Yun; Liu, Guo-Hua; Bai, Jian-Shan; Deng, Yan; Qiu, De-Yi; Zhu, Xing-Quan

    2014-01-01

    Clonorchis sinensis and Opisthorchis viverrini are both important fish-borne pathogens, causing serious public health problem in Asia. The present study developed an assay integrating real-time PCR and high resolution melting (HRM) analysis for the specific detection and rapid identification of C. sinensis and O. viverrini. Primers targeting COX1 gene were highly specific for these liver flukes, as evidenced by the negative amplification of closely related trematodes. Assays using genomic DNA...

  7. [Usefulness of a real-time quantitative polymerase-chain reaction (PCR) assay for the diagnosis of congenital and postnatal cytomegalovirus infection].

    Science.gov (United States)

    Reina, J; Weber, I; Riera, E; Busquets, M; Morales, C

    2014-05-01

    Cytomegalovirus (CMV) is the main virus causing congenital and postnatal infections in the pediatric population. The aim of this study is to evaluate the usefulness of a quantitative real-time PCR in the diagnosis of these infections using urine as a single sample. We studied all the urine samples of newborns (< 7 days) with suspected congenital infection, and urine of patients with suspected postnatal infection (urine negative at birth). Urines were simultaneously studied by cell culture, qualitative PCR (PCRc), and quantitative real-time PCR (PCRq). We analyzed 332 urine samples (270 to rule out congenital infection and 62 postnatal infections). Of the first, 22 were positive in the PCRq, 19 in the PCRc, and 17 in the culture. PCRq had a sensitivity of 100%, on comparing the culture with the rest of the techniques. Using the PCRq as a reference method, culture had a sensitivity of 77.2%, and PCRc 86.3%. In cases of postnatal infection, PCRq detected 16 positive urines, the PCRq 12, and the cell culture 10. The urines showed viral loads ranging from 2,178 to 116,641 copies/ml. The genomic amplification technique PCRq in real time was more sensitive than the other techniques evaluated. This technique should be considered as a reference (gold standard), leaving the cell culture as a second diagnostic level. The low cost and the automation of PCRq would enable the screening for CMV infection in large neonatal and postnatal populations. Copyright © 2013 Asociación Española de Pediatría. Published by Elsevier Espana. All rights reserved.

  8. Real-time dynamic analysis for complete loop of direct steam generation solar trough collector

    International Nuclear Information System (INIS)

    Guo, Su; Liu, Deyou; Chu, Yinghao; Chen, Xingying; Shen, Bingbing; Xu, Chang; Zhou, Ling; Wang, Pei

    2016-01-01

    Highlights: • A nonlinear distribution parameter dynamic model has been developed. • Real-time local heat transfer coefficient and friction coefficient are adopted. • The dynamic behavior of the solar trough collector loop are simulated. • High-frequency chattering of outlet fluid flow are analyzed and modeled. • Irradiance disturbance at subcooled water region generates larger influence. - Abstract: Direct steam generation is a potential approach to further reduce the levelized electricity cost of solar trough. Dynamic modeling of the collector loop is essential for operation and control of direct steam generation solar trough. However, the dynamic behavior of fluid based on direct steam generation is complex because of the two-phase flow in the pipeline. In this work, a nonlinear distribution parameter model has been developed to model the dynamic behaviors of direct steam generation parabolic trough collector loops under either full or partial solar irradiance disturbance. Compared with available dynamic model, the proposed model possesses two advantages: (1) real-time local values of heat transfer coefficient and friction resistance coefficient, and (2) considering of the complete loop of collectors, including subcooled water region, two-phase flow region and superheated steam region. The proposed model has shown superior performance, particularly in case of sensitivity study of fluid parameters when the pipe is partially shaded. The proposed model has been validated using experimental data from Solar Thermal Energy Laboratory of University of New South Wales, with an outlet fluid temperature relative error of only 1.91%. The validation results show that: (1) The proposed model successfully outperforms two reference models in predicting the behavior of direct steam generation solar trough. (2) The model theoretically predicts that, during solar irradiance disturbance, the discontinuities of fluid physical property parameters and the moving back and

  9. Detection and semi-quantification of Strongylus vulgaris DNA in equine faeces by real-time quantitative PCR.

    Science.gov (United States)

    Nielsen, Martin K; Peterson, David S; Monrad, Jesper; Thamsborg, Stig M; Olsen, Susanne N; Kaplan, Ray M

    2008-03-01

    Strongylus vulgaris is an important strongyle nematode with high pathogenic potential infecting horses world-wide. Several decades of intensive anthelmintic use has virtually eliminated clinical disease caused by S. vulgaris, but has also caused high levels of anthelmintic resistance in equine small strongyle (cyathostomin) nematodes. Recommendations aimed at limiting the development of anthelmintic resistance by reducing treatment intensity raises a simultaneous demand for reliable and accurate diagnostic tools for detecting important parasitic pathogens. Presently, the only means available to differentiate among strongyle species in a faecal sample is by identifying individual L3 larvae following a two week coproculture procedure. The aim of the present study is to overcome this diagnostic