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Sample records for quantitative detection approach

  1. Quantitative Surface Chirality Detection with Sum Frequency Generation Vibrational Spectroscopy: Twin Polarization Angle Approach

    SciTech Connect

    Wei, Feng; Xu, Yanyan; Guo, Yuan; Liu, Shi-lin; Wang, Hongfei

    2009-12-27

    Here we report a novel twin polarization angle (TPA) approach in the quantitative chirality detection with the surface sum-frequency generation vibrational spectroscopy (SFG-VS). Generally, the achiral contribution dominates the surface SFG-VS signal, and the pure chiral signal is usually two or three orders of magnitude smaller. Therefore, it has been difficult to make quantitative detection and analysis of the chiral contributions to the surface SFG- VS signal. In the TPA method, by varying together the polarization angles of the incoming visible light and the sum frequency signal at fixed s or p polarization of the incoming infrared beam, the polarization dependent SFG signal can give not only direct signature of the chiral contribution in the total SFG-VS signal, but also the accurate measurement of the chiral and achiral components in the surface SFG signal. The general description of the TPA method is presented and the experiment test of the TPA approach is also presented for the SFG-VS from the S- and R-limonene chiral liquid surfaces. The most accurate degree of chiral excess values thus obtained for the 2878 cm⁻¹ spectral peak of the S- and R-limonene liquid surfaces are (23.7±0.4)% and ({25.4±1.3)%, respectively.

  2. [Quantitative Approach to Melamine Detection in Egg White with Surface-Enhanced Raman Spectroscopy].

    PubMed

    Wang, Qiao-hua; Liu, Ya-li; Ma, Mei-hu; Wang, Hong

    2015-04-01

    Due to the harmfulness of melamine to human, the quantitative detection of melamine in egg is very necessary. In the present study, the surface enhanced Raman spectra technology combined with chemometric analysis method was used to conduct melamine quantitative detection in egg white. Firstly, the melamine egg sample could be got by the method of artificial feeding hens usingdifferent feeding formulation. Then the surface enhanced Raman spectra of egg white was determined using portable Raman spectroscopy (Opto Trace RamTracer-200) and Raman enhancement reagents, and the melamine content within the white eggs was measured with gas chromatography mass spectrometry technology. The software of Raman Analyzer was used for baseline correction of Raman spectra. The correlation coefficient method was used to choose 320 spectral variables from the surface enhanced Raman spectroscopy as input variables to establish partial least squares quantitative calibration model . And the peaks-decomposition method was used to establish peaks-decomposition quantitative calibration model. Both models selected 90 and 44 samples respectively as calibration sets and validation sets during model establishment, and both models achieved good prediction effect. The determination coefficient between predicted values of partial least squares quantitative calibration model and measured values of gas chromatography mass spectrometry was 0.856, and root mean square error of prediction was 1.547. The determination coefficient was 0.947 and RMSEP was 0.893 for the peaks-decomposition quantitative calibration model. This study demonstrated that the method can effectively quantitatively detect melamine in eggs. Testing a sample only takes 15 minutes, which can provide a new way for the melamine egg detection. PMID:26197575

  3. Approach to quantitative detection of CD146 with the label-free protein biosensor based on imaging ellipsometry

    NASA Astrophysics Data System (ADS)

    Niu, Yu; Liu, Li; Yan, Xiyun; Jin, Gang

    2011-03-01

    CD146 glycoprotein belonging to cell adhesion molecules is considered to be a novel target on endothelial cell involved in tumor angiogenesis. The biosensor based on imaging ellipsometry (BIE) which is performed in null and off-null mode is used for CD146 detection as a trial by the following steps. Firstly, anti-CD146 antibody as ligand is immobilized on Protein G modified silicon substrate. Then, CD146 test is carried out and its calibration curve is established for the requirement of quantitative detection. Finally, 18 serum samples are detected quantitatively and their results are validated by ELISA's. The sensitivity for CD146 detection achieves the order of ng/ml and the relationship between BIE signal y (grayscale value) and CD146 concentration x (ng/ml) is y=3.3ln(x) +91.3. Compared with ELISA's, the majority of results are in agreement, and the results of two approaches have significant statistic relevance.

  4. Novel Approach to Quantitative Detection of Specific rRNA in a Microbial Community, Using Catalytic DNA

    PubMed Central

    Suenaga, Hikaru; Liu, Rui; Shiramasa, Yuko; Kanagawa, Takahiro

    2005-01-01

    We developed a novel method for the quantitative detection of the 16S rRNA of a specific bacterial species in the microbial community by using deoxyribozyme (DNAzyme), which possesses the catalytic function to cleave RNA in a sequence-specific manner. A mixture of heterogeneous 16S rRNA containing the target 16S rRNA was incubated with a species-specific DNAzyme. The cleaved target 16S rRNA was separated from the intact 16S rRNA by electrophoresis, and then their amounts were compared for the quantitative detection of target 16S rRNA. This method was used to determine the abundance of the 16S rRNA of a filamentous bacterium, Sphaerotilus natans, in activated sludge, which is a microbial mixture used in wastewater treatment systems. The result indicated that this DNAzyme-based approach would be applicable to actual microbial communities. PMID:16085888

  5. Computational vaccinology: quantitative approaches.

    PubMed

    Flower, Darren R; McSparron, Helen; Blythe, Martin J; Zygouri, Christianna; Taylor, Debra; Guan, Pingping; Wan, Shouzhan; Coveney, Peter V; Walshe, Valerie; Borrow, Persephone; Doytchinova, Irini A

    2003-01-01

    The immune system is hierarchical and has many levels, exhibiting much emergent behaviour. However, at its heart are molecular recognition events that are indistinguishable from other types of biomacromolecular interaction. These can be addressed well by quantitative experimental and theoretical biophysical techniques, and particularly by methods from drug design. We review here our approach to computational immunovaccinology. In particular, we describe the JenPep database and two new techniques for T cell epitope prediction. One is based on quantitative structure-activity relationships (a 3D-QSAR method based on CoMSIA and another 2D method based on the Free-Wilson approach) and the other on atomistic molecular dynamic simulations using high performance computing. JenPep (http://www.jenner.ar.uk/ JenPep) is a relational database system supporting quantitative data on peptide binding to major histocompatibility complexes, TAP transporters, TCR-pMHC complexes, and an annotated list of B cell and T cell epitopes. Our 2D-QSAR method factors the contribution to peptide binding from individual amino acids as well as 1-2 and 1-3 residue interactions. In the 3D-QSAR approach, the influence of five physicochemical properties (volume, electrostatic potential, hydrophobicity, hydrogen-bond donor and acceptor abilities) on peptide affinity were considered. Both methods are exemplified through their application to the well-studied problem of peptide binding to the human class I MHC molecule HLA-A*0201. PMID:14712934

  6. Quantitative detection of protein arrays.

    PubMed

    Levit-Binnun, Nava; Lindner, Ariel B; Zik, Ory; Eshhar, Zelig; Moses, Elisha

    2003-03-15

    We introduce a quantitative method that utilizes scanning electron microscopy for the analysis of protein chips (SEMPC). SEMPC is based upon counting target-coated gold particles interacting specifically with ligands or proteins arrayed on a derivative microscope glass slide by utilizing backscattering electron detection. As model systems, we quantified the interactions of biotin and streptavidin and of an antibody with its cognate hapten. Our method gives quantitative molecule-counting capabilities with an excellent signal-to-noise ratio and demonstrates a broad dynamic range while retaining easy sample preparation and realistic automation capability. Increased sensitivity and dynamic range are achieved in comparison to currently used array detection methods such as fluorescence, with no signal bleaching, affording high reproducibility and compatibility with miniaturization. Thus, our approach facilitates the determination of the absolute number of molecules bound to the chip rather than their relative amounts, as well as the use of smaller samples.

  7. Effect of genetic cross on the detection of quantitative trait loci and a novel approach to mapping QTLs.

    PubMed

    Hitzemann, R; Demarest, K; Koyner, J; Cipp, L; Patel, N; Rasmussen, E; McCaughran, J

    2000-12-01

    A genome-wide scan was conducted in two F(2) intercrosses, C57BL/6J (B6)xDBA/2J (D2) and BALB/cJ (C)xLP/J (LP), for three different phenotypes: basal locomotor activity, ethanol-induced locomotor activity, and haloperidol-induced catalepsy. For basal activity, significant quantitative trait loci (QTLs, LOD> or =4.3) were detected on chromosomes 9 and 19 for the CxLP intercross and chromosome 1 for the B6xD2 intercross. Significant QTLs for ethanol-induced activation were detected on chromosome 6 for the CxLP intercross, and on chromosomes 1 and 2 for the B6xD2 intercross. For haloperidol-induced catalepsy, significant QTLs were detected on chromosome 14 (two different QTLs) in the CxLP intercross, and chromosomes 1 and 9 in the B6xD2 intercross. These data illustrate the importance of the genetic cross for QTL detection. Finally, the data reported here, and elsewhere, are also used to demonstrate a novel approach to QTL detection and localization.

  8. Quantitative assessment of hyperspectral imaging in detection of plasmonic nanoparticles: a modified contrast-detail analysis approach

    NASA Astrophysics Data System (ADS)

    Wang, Jianting; Chen, Yu; Pfefer, T. Joshua

    2016-03-01

    Hyperspectral reflectance imaging (HRI) is an emerging imaging modality being applied for clinical indications such as tissue oximetry, and cancer detection based on endogenous biological constituents including plasmonic nanoparticles. However, there is currently a lack of standardized test methods for objective, quantitative evaluation of HRI system performance. Contrast-detail analysis (CDA) is a phantom-based test method commonly used to evaluate medical imaging devices (e.g., mammography systems) in terms of their lower detection limit. We investigated a modified CDA (mCDA) method to quantify the detectability of gold nanoparticles by HRI systems. Silicone-based turbid phantoms containing micro-fluidic channels were developed for the mCDA tests. Polydimethylsiloxane (PDMS) phantom materials were doped with chromophores and scatterers to achieve biologically relevant optical properties (OPs). Molds were used to produce cylindrical channels of diameters 0.3 to 1.65 mm and depths of 0.2 mm inside the phantoms. Channels were filled with a mixture of hemoglobin and concentrations of gold nanorods (GNR) and measured with our HRI system. The contrast of GNRs was solved with a spectral unmixing algorithm from the reflectance spectra. The lowest detectable concentration was determined as a function of inclusion size and depth and plotted as modified contrast detail curve (mCDC). mCDCs were used to compare the detectabilities of the HRI system with different data processing algorithms. It is demonstrated that our mCDA test method involving turbid microchannel phantoms can help to elucidate the combined performance of imaging devices and plasmonic nanoparticle contrast agents. This approach may be useful for performing clinical trial standardization and device re-calibration, thus ensuring quality control and clinical performance.

  9. Time-series tropical forest change detection: a visual and quantitative approach

    NASA Astrophysics Data System (ADS)

    Sader, Steven A.; Sever, Thomas; Smoot, James C.

    1996-11-01

    Forest change detection over a decadal time frame was conducted for the Maya Biosphere Reserve in northern Guatemala. A simple and logical method of visualizing and quantifying forest change is presented. Analysis of time- series Landsat-Thematic Mapper imagery provided estimates of forest change at three time periods; prior to 1990, 1990 to 1993 and 1993 to 1995. Four dates of Landsat imagery were pre-processed, co-registered to a UTM projection and the normalized difference vegetation index was computed for each date. An unsupervised classification was performed and cluster classes were grouped into time-series change/no change categories. A color coded image was generated which resembled the RBG-NDVI color composite of the 1990, 1993, and 1995 imagery. Land cover information and Geographic Information System (GIS) editing techniques were applied to resolve some confusions between forest change and change in non-forest types. Results indicated that forest clearing rates in the reserve were less than 0.5 percent per year in the early to mid 1990s but the buffer zone clearing rates, at over two percent, were much higher.

  10. Quantitative multiplex detection of pathogen biomarkers

    DOEpatents

    Mukundan, Harshini; Xie, Hongzhi; Swanson, Basil I; Martinez, Jennifer; Grace, Wynne K

    2014-10-14

    The present invention addresses the simultaneous detection and quantitative measurement of multiple biomolecules, e.g., pathogen biomarkers through either a sandwich assay approach or a lipid insertion approach. The invention can further employ a multichannel, structure with multi-sensor elements per channel.

  11. Quantitative multiplex detection of pathogen biomarkers

    DOEpatents

    Mukundan, Harshini; Xie, Hongzhi; Swanson, Basil I.; Martinez, Jennifer; Grace, Wynne K.

    2016-02-09

    The present invention addresses the simultaneous detection and quantitative measurement of multiple biomolecules, e.g., pathogen biomarkers through either a sandwich assay approach or a lipid insertion approach. The invention can further employ a multichannel, structure with multi-sensor elements per channel.

  12. Rapid and direct quantitative detection of viable bifidobacteria in probiotic yogurt by combination of ethidium monoazide and real-time PCR using a molecular beacon approach.

    PubMed

    Meng, X C; Pang, R; Wang, C; Wang, L Q

    2010-11-01

    The potential of ethidium monoazide (EMA) real-time PCR method based on molecular beacon probe for rapid detection of viable bifidobacteria present in probiotic yogurt was evaluated in this work. A real-time PCR with molecular beacon assay was developed to determine genus Bifidobacterium quantitatively in order to increase the sensitivity and specificity of assay. EMA was used to treat probiotic yogurt prior to DNA extraction and real-time PCR detection to allow detection of only viable bacteria. The primer set of Bif-F/Bif-R which is genus-specific for Bifid. was designed. The specificity of the probes ensures that no signal is generated by non-target amplicons. Linear regression analysis demonstrated a good correlation (R² = 0·9948) between the EMA real-time PCR results and the plate counting, and real-time quantitative PCR results correlated adequately with enumeration of bifidobacteria by culture for commercial probiotic yogurt. This culture-independent approach is promising for the direct and rapid detection of viable bifidobacteria in commercial probiotic yogurt, and the detection can be carried out within 4 h. The detection limit for this method is about 10⁴ cell/ml. In conclusion, the direct quantitative EMA real-time PCR assay based on molecular beacon described in this research is a rapid and quantitative method.

  13. A microfluidic immunoassay platform for the detection of free prostate specific antigen: a systematic and quantitative approach.

    PubMed

    Madaboosi, Narayanan; Soares, Ruben R G; Chu, Virginia; Conde, João Pedro

    2015-07-01

    As a leading cause of cancer-related deaths in men globally, prostate cancer (PCa) demands immense attention for theranostic purposes. There is an increasing need for the development of rapid, sensitive, economical, miniaturized and multiplexable assays. Towards this goal, we present a systematic approach for the optimisation of a microfluidic sandwich immunoassay, which can be applied as a generic biosensor platform for PCa detection. Prostate specific antigen (PSA) was used as the model biomarker, and its free form was captured using commercially available antibodies and detected using chemiluminescence, both in spiked buffer and matrix solutions. Along with the optimisation of surface chemistry and microfluidic parameters, we report a bio-affinity amplification strategy based on biotin-streptavidin chemistry to bring the limits of detection for free-PSA from 21.4 ng mL(-1) down to 2.7 ng mL(-1), within the clinically relevant range. An estimate of the surface coverage and simulations of the interactions taking place in the microfluidic biosensor during the assay are also presented. This novel platform using a simple passive adsorption-based bio-affinity strategy, when coupled with multiplexing and integrated detection, can serve as a promising point-of-care diagnostic tool for PCa. PMID:25988197

  14. Quantitative approaches to computational vaccinology.

    PubMed

    Doytchinova, Irini A; Flower, Darren R

    2002-06-01

    This article reviews the newly released JenPep database and two new powerful techniques for T-cell epitope prediction: (i) the additive method; and (ii) a 3D-Quantitative Structure Activity Relationships (3D-QSAR) method, based on Comparative Molecular Similarity Indices Analysis (CoMSIA). The JenPep database is a family of relational databases supporting the growing need of immunoinformaticians for quantitative data on peptide binding to major histocompatibility complexes and to the Transporters associated with Antigen Processing (TAP). It also contains an annotated list of T-cell epitopes. The database is available free via the Internet (http://www.jenner.ac.uk/JenPep). The additive prediction method is based on the assumption that the binding affinity of a peptide depends on the contributions from each amino acid as well as on the interactions between the adjacent and every second side-chain. In the 3D-QSAR approach, the influence of five physicochemical properties (steric bulk, electrostatic potential, local hydrophobicity, hydrogen-bond donor and hydrogen-bond acceptor abilities) on the affinity of peptides binding to MHC molecules were considered. Both methods were exemplified through their application to the well-studied problem of peptides binding to the human class I MHC molecule HLA-A*0201. PMID:12067414

  15. Cancer detection by quantitative fluorescence image analysis.

    PubMed

    Parry, W L; Hemstreet, G P

    1988-02-01

    Quantitative fluorescence image analysis is a rapidly evolving biophysical cytochemical technology with the potential for multiple clinical and basic research applications. We report the application of this technique for bladder cancer detection and discuss its potential usefulness as an adjunct to methods used currently by urologists for the diagnosis and management of bladder cancer. Quantitative fluorescence image analysis is a cytological method that incorporates 2 diagnostic techniques, quantitation of nuclear deoxyribonucleic acid and morphometric analysis, in a single semiautomated system to facilitate the identification of rare events, that is individual cancer cells. When compared to routine cytopathology for detection of bladder cancer in symptomatic patients, quantitative fluorescence image analysis demonstrated greater sensitivity (76 versus 33 per cent) for the detection of low grade transitional cell carcinoma. The specificity of quantitative fluorescence image analysis in a small control group was 94 per cent and with the manual method for quantitation of absolute nuclear fluorescence intensity in the screening of high risk asymptomatic subjects the specificity was 96.7 per cent. The more familiar flow cytometry is another fluorescence technique for measurement of nuclear deoxyribonucleic acid. However, rather than identifying individual cancer cells, flow cytometry identifies cellular pattern distributions, that is the ratio of normal to abnormal cells. Numerous studies by others have shown that flow cytometry is a sensitive method to monitor patients with diagnosed urological disease. Based upon results in separate quantitative fluorescence image analysis and flow cytometry studies, it appears that these 2 fluorescence techniques may be complementary tools for urological screening, diagnosis and management, and that they also may be useful separately or in combination to elucidate the oncogenic process, determine the biological potential of tumors

  16. Reproducibility and quantitation of amplicon sequencing-based detection.

    PubMed

    Zhou, Jizhong; Wu, Liyou; Deng, Ye; Zhi, Xiaoyang; Jiang, Yi-Huei; Tu, Qichao; Xie, Jianping; Van Nostrand, Joy D; He, Zhili; Yang, Yunfeng

    2011-08-01

    To determine the reproducibility and quantitation of the amplicon sequencing-based detection approach for analyzing microbial community structure, a total of 24 microbial communities from a long-term global change experimental site were examined. Genomic DNA obtained from each community was used to amplify 16S rRNA genes with two or three barcode tags as technical replicates in the presence of a small quantity (0.1% wt/wt) of genomic DNA from Shewanella oneidensis MR-1 as the control. The technical reproducibility of the amplicon sequencing-based detection approach is quite low, with an average operational taxonomic unit (OTU) overlap of 17.2%±2.3% between two technical replicates, and 8.2%±2.3% among three technical replicates, which is most likely due to problems associated with random sampling processes. Such variations in technical replicates could have substantial effects on estimating β-diversity but less on α-diversity. A high variation was also observed in the control across different samples (for example, 66.7-fold for the forward primer), suggesting that the amplicon sequencing-based detection approach could not be quantitative. In addition, various strategies were examined to improve the comparability of amplicon sequencing data, such as increasing biological replicates, and removing singleton sequences and less-representative OTUs across biological replicates. Finally, as expected, various statistical analyses with preprocessed experimental data revealed clear differences in the composition and structure of microbial communities between warming and non-warming, or between clipping and non-clipping. Taken together, these results suggest that amplicon sequencing-based detection is useful in analyzing microbial community structure even though it is not reproducible and quantitative. However, great caution should be taken in experimental design and data interpretation when the amplicon sequencing-based detection approach is used for quantitative

  17. A novel quantitation approach for maximizing detectable targets for offensive/volatile odorants with diverse functional groups by thermal desorption-gas chromatography-mass spectrometry

    PubMed Central

    Kim, Yong-Hyun; Kim, Ki-Hyun

    2016-01-01

    A multitude of analytical systems are needed to analyze diverse odorants with various functionalities. In this study, an experimental method was developed to assess the maximum covering range of odorants using a single experimental setup consisting of a thermal desorber-gas chromatography-mass spectrometry system. To this end, a total of 20 offensive odorants (aldehyde, ketone, ester, alcohol, aromatic, sulfide, amine, and carboxyl) were selected and tested by a single system. The analytical results of standards and environmental samples were evaluated in a number of respects. In the analysis of the standards, all targets were quantified via Carbopack (C + B + X) tube sampling while operating the thermal desorber at −25 °C. The method detection limits of 18 targets (exception of 2 out of the 20 targets: acetaldehyde and methanethiol) were excellent (mean 0.04 ± 0.03 ppb) in terms of their odor threshold values (74.7 ± 140 ~ 624 ± 1,729 ppb). The analysis of organic fertilizer plant samples at a pig farm (slurry treatment facility, compost facility, and ambient air) confirmed the presence of 18 odorants from 0.03 ppb (dimethyldisulfide, ambient sample) to 522 ppb (methyl ethyl ketone, slurry treatment facility). As such, our method allowed simultaneous quantitation of most key odorants with sufficient reliability and sensitivity. PMID:27404037

  18. A novel quantitation approach for maximizing detectable targets for offensive/volatile odorants with diverse functional groups by thermal desorption-gas chromatography-mass spectrometry

    NASA Astrophysics Data System (ADS)

    Kim, Yong-Hyun; Kim, Ki-Hyun

    2016-07-01

    A multitude of analytical systems are needed to analyze diverse odorants with various functionalities. In this study, an experimental method was developed to assess the maximum covering range of odorants using a single experimental setup consisting of a thermal desorber-gas chromatography-mass spectrometry system. To this end, a total of 20 offensive odorants (aldehyde, ketone, ester, alcohol, aromatic, sulfide, amine, and carboxyl) were selected and tested by a single system. The analytical results of standards and environmental samples were evaluated in a number of respects. In the analysis of the standards, all targets were quantified via Carbopack (C + B + X) tube sampling while operating the thermal desorber at ‑25 °C. The method detection limits of 18 targets (exception of 2 out of the 20 targets: acetaldehyde and methanethiol) were excellent (mean 0.04 ± 0.03 ppb) in terms of their odor threshold values (74.7 ± 140 ~ 624 ± 1,729 ppb). The analysis of organic fertilizer plant samples at a pig farm (slurry treatment facility, compost facility, and ambient air) confirmed the presence of 18 odorants from 0.03 ppb (dimethyldisulfide, ambient sample) to 522 ppb (methyl ethyl ketone, slurry treatment facility). As such, our method allowed simultaneous quantitation of most key odorants with sufficient reliability and sensitivity.

  19. A novel quantitation approach for maximizing detectable targets for offensive/volatile odorants with diverse functional groups by thermal desorption-gas chromatography-mass spectrometry.

    PubMed

    Kim, Yong-Hyun; Kim, Ki-Hyun

    2016-01-01

    A multitude of analytical systems are needed to analyze diverse odorants with various functionalities. In this study, an experimental method was developed to assess the maximum covering range of odorants using a single experimental setup consisting of a thermal desorber-gas chromatography-mass spectrometry system. To this end, a total of 20 offensive odorants (aldehyde, ketone, ester, alcohol, aromatic, sulfide, amine, and carboxyl) were selected and tested by a single system. The analytical results of standards and environmental samples were evaluated in a number of respects. In the analysis of the standards, all targets were quantified via Carbopack (C + B + X) tube sampling while operating the thermal desorber at -25 °C. The method detection limits of 18 targets (exception of 2 out of the 20 targets: acetaldehyde and methanethiol) were excellent (mean 0.04 ± 0.03 ppb) in terms of their odor threshold values (74.7 ± 140 ~ 624 ± 1,729 ppb). The analysis of organic fertilizer plant samples at a pig farm (slurry treatment facility, compost facility, and ambient air) confirmed the presence of 18 odorants from 0.03 ppb (dimethyldisulfide, ambient sample) to 522 ppb (methyl ethyl ketone, slurry treatment facility). As such, our method allowed simultaneous quantitation of most key odorants with sufficient reliability and sensitivity. PMID:27404037

  20. Toward a quantitative approach to migrants integration

    NASA Astrophysics Data System (ADS)

    Barra, A.; Contucci, P.

    2010-03-01

    Migration phenomena and all the related issues, like integration of different social groups, are intrinsically complex problems since they strongly depend on several competitive mechanisms as economic factors, cultural differences and many others. By identifying a few essential assumptions, and using the statistical mechanics of complex systems, we propose a novel quantitative approach that provides a minimal theory for those phenomena. We show that the competitive interactions in decision making between a population of N host citizens and P immigrants, a bi-partite spin-glass, give rise to a social consciousness inside the host community in the sense of the associative memory of neural networks. The theory leads to a natural quantitative definition of migrant's "integration" inside the community. From the technical point of view this minimal picture assumes, as control parameters, only general notions like the strength of the random interactions, the ratio between the sizes of the two parties and the cultural influence. Few steps forward, toward more refined models, which include a digression on the kind of the felt experiences and some structure on the random interaction topology (as dilution to avoid the plain mean-field approach) and correlations of experiences felt between the two parties (biasing the distribution of the coupling) are discussed at the end, where we show the robustness of our approach.

  1. [Quantitative detection of Vibrio vulnificus in seafood].

    PubMed

    Hara-Kud, Yukiko; Miwa, Norinaga; Yamasaki, Syougo; Yatsuyanagi, Jun; Iwade, Yoshito; Takahash, Hajime; Miyasaka, Jiro

    2005-12-01

    To quantify the number of Vibrio vulnificus in shellfish, we compared the most probable number (MPN) combined with a culture (MPN-culture) or polymerase-chain reaction (PCR) assay (MPN-PCR) to a quantitative PCR assay. Enrichment in alkaline peptone water by MPN was conducted at 25 and 35 degrees C. Enrichment at 35 degrees C was superior or similar to enrichment at 25 degrees C in over 65% of samples by MPNculture and in more than 75% of samples by MPN-PCR assay. V. vulnificus was more easily isolated on chromogenic agar medium during culture, MPN-PCR assay was superior or similar to MPNculture in over 90% of samples by enrichment at 25 degrees C and to over 88% of samples by enrichment at 35 degrees C. The number of V. vulnificus by quantitative PCR assay was similar to that of MPN-PCR assay in 6 of 8 samples but not from MPNculture. V. vulnificus contamination was frequently detected in samples from Kyushu Island.

  2. A novel approach to the quantitative detection of anabolic steroids in bovine muscle tissue by means of a hybrid quadrupole time-of-flight-mass spectrometry instrument.

    PubMed

    Bussche, Julie Vanden; Decloedt, Anneleen; Van Meulebroek, Lieven; De Clercq, Nathalie; Lock, Stephen; Stahl-Zeng, Jianru; Vanhaecke, Lynn

    2014-09-19

    In recent years, the analysis of veterinary drugs and growth-promoting agents has shifted from target-oriented procedures, mainly based on liquid chromatography coupled to triple-quadrupole mass spectrometry (LC-QqQ-MS), towards accurate mass full scan MS (such as Time-of-Flight (ToF) and Fourier Transform (FT)-MS). In this study, the performance of a hybrid analysis instrument (i.e. UHPLC-QuadrupoleTime-of-Flight-MS (QqToF-MS)), able to exploit both full scan HR and MS/MS capabilities within a single analytical platform, was evaluated for confirmatory analysis of anabolic steroids (gestagens, estrogens including stilbenes and androgens) in meat. The validation data was compared to previously obtained results (CD 2002/657/EC) for QqQ-MS and single stage Orbitrap-MS. Additionally, a fractional factorial design was used to shorten and optimize the sample extraction. Validation according to CD 2002/657/EC demonstrated that steroid analysis using QqToF has a higher competing value towards QqQ-MS in terms of selectivity/specificity, compared to single stage Orbitrap-MS. While providing excellent linearity, based on lack-of-fit calculations (F-test, α=0.05 for all steroids except 17β-ethinylestradiol: α=0.01), the sensitivity of QqToF-MS proved for 61.8% and 85.3% of the compounds more sensitive compared to QqQ-MS and Orbitrap-MS, respectively. Indeed, the CCα values, obtained upon ToF-MS/MS detection, ranged from 0.02 to 1.74μgkg(-1) for the 34 anabolic steroids, while for QqQ-MS and Orbitrap-MS values ranged from 0.04 to 0.88μgkg(-1) and from 0.07 to 2.50μgkg(-1), respectively. Using QqToF-MS and QqQ-MS, adequate precision was obtained as relative standard deviations for repeatability and within-laboratory reproducibility, were below 20%. In case of Orbitrap-MS, some compounds (i.e. some estrogens) displayed poor precision, which was possibly caused by some lack of sensitivity at lower concentrations and the absence of MRM-like experiments. Overall, it can be

  3. The detection and quantitation of protein oligomerization.

    PubMed

    Gell, David A; Grant, Richard P; Mackay, Joel P

    2012-01-01

    There are many different techniques available to biologists and biochemists that can be used to detect and characterize the self-association of proteins. Each technique has strengths and weaknesses and it is often useful to combine several approaches to maximize the former and minimize the latter. Here we review a range of methodologies that identify protein self-association and/or allow the stoichiometry and affinity of the interaction to be determined, placing an emphasis on what type of information can be obtained and outlining the advantages and disadvantages involved. In general, in vitro biophysical techniques, such as size exclusion chromatography, analytical ultracentrifugation, scattering techniques, NMR spectroscopy, isothermal titration calorimetry, fluorescence anisotropy and mass spectrometry, provide information on stoichiometry and/or binding affinities. Other approaches such as cross-linking, fluorescence methods (e.g., fluorescence correlation spectroscopy, FCS; Förster resonance energy transfer, FRET; fluorescence recovery after photobleaching, FRAP; and proximity imaging, PRIM) and complementation approaches (e.g., yeast two hybrid assays and bimolecular fluorescence complementation, BiFC) can be used to detect protein self-association in a cellular context.

  4. A Quantitative Approach to Assessing System Evolvability

    NASA Technical Reports Server (NTRS)

    Christian, John A., III

    2004-01-01

    When selecting a system from multiple candidates, the customer seeks the one that best meets his or her needs. Recently the desire for evolvable systems has become more important and engineers are striving to develop systems that accommodate this need. In response to this search for evolvability, we present a historical perspective on evolvability, propose a refined definition of evolvability, and develop a quantitative method for measuring this property. We address this quantitative methodology from both a theoretical and practical perspective. This quantitative model is then applied to the problem of evolving a lunar mission to a Mars mission as a case study.

  5. Nested-quantitative PCR approach with improved sensitivity for the detection of low titer levels of Candidatus Liberibacter asiaticus in the Asian citrus psyllid, Diaphorina citri Kuwayama.

    PubMed

    Coy, M R; Hoffmann, M; Kingdom Gibbard, H N; Kuhns, E H; Pelz-Stelinski, K S; Stelinski, L L

    2014-07-01

    Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay using the CLas 16S rDNA gene target. Although qPCR is highly sensitive, low bacterial titers or suboptimal qPCR conditions can result in false-negatives. Using a nested qPCR assay, we determined the false-negative rate of the 16S presence/absence qPCR assay was greater than 50%. Studies to determine the performance parameters of the qPCR assays for CLas 16S and Wingless (Wg), the D. citri endogenous gene, using plasmid and psyllid DNA, revealed suboptimal and variable performance of the 16S assay in psyllid samples. Average efficiencies and sensitivity limits of the plasmid assays were 99.0% and 2.7 copies of template for Wg, respectively, and 98.5% and 2.2-22.1 copies for 16S, respectively. Variability in efficiency was significantly greater in psyllid samples for both gene targets compared to the corresponding plasmid assays, and efficiencies as low as 76% were obtained for 16S. A secondary structure analysis revealed the formation of two stem-loop structures that block the forward and probe binding sites in the 16S template, which could hinder amplification. In summary, our results suggest that suboptimal qPCR efficiency is not uncommon for the 16S presence/absence qPCR assay, which combined with lowCLas titers in some samples, could contribute significantly to the under-reporting of CLas infection in psyllid and plant samples.

  6. Nested-quantitative PCR approach with improved sensitivity for the detection of low titer levels of Candidatus Liberibacter asiaticus in the Asian citrus psyllid, Diaphorina citri Kuwayama.

    PubMed

    Coy, M R; Hoffmann, M; Kingdom Gibbard, H N; Kuhns, E H; Pelz-Stelinski, K S; Stelinski, L L

    2014-07-01

    Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay using the CLas 16S rDNA gene target. Although qPCR is highly sensitive, low bacterial titers or suboptimal qPCR conditions can result in false-negatives. Using a nested qPCR assay, we determined the false-negative rate of the 16S presence/absence qPCR assay was greater than 50%. Studies to determine the performance parameters of the qPCR assays for CLas 16S and Wingless (Wg), the D. citri endogenous gene, using plasmid and psyllid DNA, revealed suboptimal and variable performance of the 16S assay in psyllid samples. Average efficiencies and sensitivity limits of the plasmid assays were 99.0% and 2.7 copies of template for Wg, respectively, and 98.5% and 2.2-22.1 copies for 16S, respectively. Variability in efficiency was significantly greater in psyllid samples for both gene targets compared to the corresponding plasmid assays, and efficiencies as low as 76% were obtained for 16S. A secondary structure analysis revealed the formation of two stem-loop structures that block the forward and probe binding sites in the 16S template, which could hinder amplification. In summary, our results suggest that suboptimal qPCR efficiency is not uncommon for the 16S presence/absence qPCR assay, which combined with lowCLas titers in some samples, could contribute significantly to the under-reporting of CLas infection in psyllid and plant samples. PMID:24769405

  7. Towards automated early cancer detection: Non-invasive, fluorescence-based approaches for quantitative assessment of cells and tissue to identify pre-cancers

    NASA Astrophysics Data System (ADS)

    Levitt, Jonathan Michael

    Cancer is the second leading cause of death globally, second only to heart disease. As in many diseases, patient survival is directly related to how early lesions are detected. Using conventional screening methods, the early changes associated with cancer, which occur on the microscopic scale, can easily go overlooked. Due to the inherent drawbacks of conventional techniques we present non-invasive, optically based methods to acquire high resolution images from live samples and assess cellular function associated with the onset of disease. Specifically, we acquired fluorescence images from NADH and FAD to quantify morphology and metabolic activity. We first conducted studies to monitor monolayers of keratinocytes in response to apoptosis which has been shown to be disrupted during cancer progression. We found that as keratinocytes undergo apoptosis there are populations of mitochondria that exhibit a higher metabolic activity that become progressively confined to a gradually smaller perinuclear region. To further assess the changes associated with early cancer growth we developed automated methods to rapidly quantify fluorescence images and extract morphological and metabolic information from life tissue. In this study, we simultaneously quantified mitochondrial organization, metabolic activity, nuclear size distribution, and the localization of the structural protein keratin, to differentiate between normal and pre-cancerous engineered tissues. We found the degree mitochondrial organization, as determined from the fractal derived Hurst parameter, was well correlated to level of cellular differentiation. We also found that the metabolic activity in the pre-cancerous cells was greater and more consistent throughout tissue depths in comparison to normal tissue. Keratin localization, also quantified from the fluorescence images, we found it to be confined to the uppermost layers of normal tissue while it was more evenly distributed in the precancerous tissues. To

  8. Anomaly Detection Using Behavioral Approaches

    NASA Astrophysics Data System (ADS)

    Benferhat, Salem; Tabia, Karim

    Behavioral approaches, which represent normal/abnormal activities, have been widely used during last years in intrusion detection and computer security. Nevertheless, most works showed that they are ineffective for detecting novel attacks involving new behaviors. In this paper, we first study this recurring problem due on one hand to inadequate handling of anomalous and unusual audit events and on other hand to insufficient decision rules which do not meet behavioral approach objectives. We then propose to enhance the standard decision rules in order to fit behavioral approach requirements and better detect novel attacks. Experimental studies carried out on real and simulated http traffic show that these enhanced decision rules improve detecting most novel attacks without triggering higher false alarm rates.

  9. Quantitative evaluation of phase processing approaches in susceptibility weighted imaging

    NASA Astrophysics Data System (ADS)

    Li, Ningzhi; Wang, Wen-Tung; Sati, Pascal; Pham, Dzung L.; Butman, John A.

    2012-03-01

    Susceptibility weighted imaging (SWI) takes advantage of the local variation in susceptibility between different tissues to enable highly detailed visualization of the cerebral venous system and sensitive detection of intracranial hemorrhages. Thus, it has been increasingly used in magnetic resonance imaging studies of traumatic brain injury as well as other intracranial pathologies. In SWI, magnitude information is combined with phase information to enhance the susceptibility induced image contrast. Because of global susceptibility variations across the image, the rate of phase accumulation varies widely across the image resulting in phase wrapping artifacts that interfere with the local assessment of phase variation. Homodyne filtering is a common approach to eliminate this global phase variation. However, filter size requires careful selection in order to preserve image contrast and avoid errors resulting from residual phase wraps. An alternative approach is to apply phase unwrapping prior to high pass filtering. A suitable phase unwrapping algorithm guarantees no residual phase wraps but additional computational steps are required. In this work, we quantitatively evaluate these two phase processing approaches on both simulated and real data using different filters and cutoff frequencies. Our analysis leads to an improved understanding of the relationship between phase wraps, susceptibility effects, and acquisition parameters. Although homodyne filtering approaches are faster and more straightforward, phase unwrapping approaches perform more accurately in a wider variety of acquisition scenarios.

  10. An overview of quantitative approaches in Gestalt perception.

    PubMed

    Jäkel, Frank; Singh, Manish; Wichmann, Felix A; Herzog, Michael H

    2016-09-01

    Gestalt psychology is often criticized as lacking quantitative measurements and precise mathematical models. While this is true of the early Gestalt school, today there are many quantitative approaches in Gestalt perception and the special issue of Vision Research "Quantitative Approaches in Gestalt Perception" showcases the current state-of-the-art. In this article we give an overview of these current approaches. For example, ideal observer models are one of the standard quantitative tools in vision research and there is a clear trend to try and apply this tool to Gestalt perception and thereby integrate Gestalt perception into mainstream vision research. More generally, Bayesian models, long popular in other areas of vision research, are increasingly being employed to model perceptual grouping as well. Thus, although experimental and theoretical approaches to Gestalt perception remain quite diverse, we are hopeful that these quantitative trends will pave the way for a unified theory.

  11. Quantitative spectroscopic imaging for noninvasive early cancer detection

    PubMed Central

    Yu, Chung-Chieh; Lau, Condon; O’Donoghue, Geoff; Mirkovic, Jelena; McGee, Sasha; Galindo, Luis; Elackattu, Alphi; Stier, Elizabeth; Grillone, Gregory; Badizadegan, Kamran; Dasari, Ramachandra R.; Feld, Michael S.

    2008-01-01

    We report a fully quantitative spectroscopy imaging instrument for wide area detection of early cancer (dysplasia). This instrument provides quantitative maps of tissue biochemistry and morphology, making it a potentially powerful surveillance tool for objective early cancer detection. We describe the design, construction, calibration, and first clinical application of this new system. We demonstrate its accuracy using physical tissue models. We validate its diagnostic ability on a resected colon adenoma, and demonstrate feasibility of in vivo imaging in the oral cavity. PMID:18825262

  12. Whole genome approaches to quantitative genetics.

    PubMed

    Visscher, Peter M

    2009-06-01

    Apart from parent-offspring pairs and clones, relative pairs vary in the proportion of the genome that they share identical by descent. In the past, quantitative geneticists have used the expected value of sharing genes by descent to estimate genetic parameters and predict breeding values. With the possibility to genotype individuals for many markers across the genome it is now possible to empirically estimate the actual relationship between relatives. We review some of the theory underlying the variation in genetic identity, show applications to estimating genetic variance for height in humans and discuss other applications.

  13. A quantitative approach to painting styles

    NASA Astrophysics Data System (ADS)

    Vieira, Vilson; Fabbri, Renato; Sbrissa, David; da Fontoura Costa, Luciano; Travieso, Gonzalo

    2015-01-01

    This research extends a method previously applied to music and philosophy (Vilson Vieira et al., 2012), representing the evolution of art as a time-series where relations like dialectics are measured quantitatively. For that, a corpus of paintings of 12 well-known artists from baroque and modern art is analyzed. A set of 99 features is extracted and the features which most contributed to the classification of painters are selected. The projection space obtained provides the basis to the analysis of measurements. These quantitative measures underlie revealing observations about the evolution of painting styles, specially when compared with other humanity fields already analyzed: while music evolved along a master-apprentice tradition (high dialectics) and philosophy by opposition, painting presents another pattern: constant increasing skewness, low opposition between members of the same movement and opposition peaks in the transition between movements. Differences between baroque and modern movements are also observed in the projected "painting space": while baroque paintings are presented as an overlapped cluster, the modern paintings present minor overlapping and are disposed more widely in the projection than the baroque counterparts. This finding suggests that baroque painters shared aesthetics while modern painters tend to "break rules" and develop their own style.

  14. Developing a Research Program Using Qualitative and Quantitative Approaches.

    ERIC Educational Resources Information Center

    Beck, Cheryl Tatano

    1997-01-01

    A research program on postpartum depression is used to illustrate the use of both qualitative and quantitative approaches. The direction of a research program is thus not limited by the type of methods in which a researcher has expertise. (SK)

  15. Pain relief by touch: a quantitative approach.

    PubMed

    Mancini, Flavia; Nash, Thomas; Iannetti, Gian Domenico; Haggard, Patrick

    2014-03-01

    Pain relief by touch has been studied for decades in pain neuroscience. Human perceptual studies revealed analgesic effects of segmental tactile stimulation, as compared to extrasegmental touch. However, the spatial organisation of touch-pain interactions within a single human dermatome has not been investigated yet. In 2 experiments we tested whether, how, and where within a dermatome touch modulates the perception of laser-evoked pain. We measured pain perception using intensity ratings, qualitative descriptors, and signal detection measures of sensitivity and response bias. Touch concurrent with laser pulses produced a significant analgesia, and reduced the sensitivity in detecting the energy of laser stimulation, implying a functional loss of information within the ascending Aδ pathway. Touch also produced a bias to judge laser stimuli as less painful. This bias decreased linearly when the distance between the laser and tactile stimuli increased. Thus, our study provides evidence for a spatial organisation of intrasegmental touch-pain interactions.

  16. Quantitative Multi-color Detection Strategies for Bioorthogonally Labeled GPCRs.

    PubMed

    Park, Minyoung; Tian, He; Naganathan, Saranga; Sakmar, Thomas P; Huber, Thomas

    2015-01-01

    We describe multiple bioorthogonal approaches to label G protein-coupled receptors (GPCRs) heterologously expressed in mammalian cells. The use of genetically encoded unnatural amino acids as bioorthogonal tags results in receptors that are expressed at lower levels than even their low abundance wild-type counterparts. Therefore, reproducible and sensitive quantification of the labeled GPCRs is extremely important and conventional methods are simply not sufficiently accurate and precise. Silver stains lack reproducibility, spectroscopic methods using fluorescent ligands are limited to quantifying only functional receptor molecules, and immunoassays using epitope tags derived from rhodopsin are particularly variable for low-abundance GPCRs. To avoid these shortcomings, we employ near infrared (NIR) imaging-based methods that enable simultaneous multi-color detection of two different antigens, thus facilitating the ratiometric analysis of bioorthogonally modified GPCRs. We anticipate that these multi-color detection strategies will provide new tools for quantitatively assessing stoichiometrically labeled GPCRs for studies of signalosomes and for structure-function relationships at a single molecule level.

  17. Quantitative Proteomic Approaches for Studying Phosphotyrosine Signaling

    SciTech Connect

    Ding, Shi-Jian; Qian, Weijun; Smith, Richard D.

    2007-02-01

    Protein tyrosine phosphorylation is a fundamental mechanism for controlling many aspects of cellular processes, as well as aspects of human health and diseases. Compared to phosphoserine (pSer) and phosphothreonine (pThr), phosphotyrosine (pTyr) signaling is more tightly regulated, but often more challenging to characterize due to significantly lower level of tyrosine phosphorylation (a relative abundance of 1800:200:1 was estimated for pSer/pThr/pTyr in vertebrate cells[1]). In this review, we outline the recent advances in analytical methodologies for enrichment, identification, and accurate quantitation of tyrosine phosphorylated proteins and peptides using antibody-based technologies, capillary liquid chromatography (LC) coupled with mass spectrometry (MS), and various stable isotope labeling strategies, as well as non-MS-based methods such as protein or peptide array methods. These proteomic technological advances provide powerful tools for potentially understanding signal transduction at the system level and provide a basis for discovering novel drug targets for human diseases. [1] Hunter, T. (1998) The Croonian Lecture 1997. The phosphorylation of proteins on tyrosine: its role in cell growth and disease. Philos. Trans. R. Soc. Lond. B Biol. Sci. 353, 583–605

  18. [Quantitative Detection of Chinese Cabbage Clubroot Based on FTIR Spectroscopy].

    PubMed

    Wang, Wei-ping; Chai, A-li; Shi, Yan-xia; Xie, Xue-wen; Li, Bao-ju

    2015-05-01

    Clubroot, caused by Plasmodiophora brassicae, is considered the most devastating soilborne disease in Brassica crops. It has emerged as a serious disease threatening the cruciferous crop production industry in China. Nowadays, the detection techniques for P. brassicae are laborious, time-consuming and low sensitivity. Rapid and effective detection methods are needed. The objective of this study is to develop a Fourier transform infrared spectrometer (FTIR) technique for detection of P. brassicae effectively and accurately. FTIR and Real-time PCR techniques were applied in quantitative detection of P. brassicae. Chinese cabbages were inoculated with P. brassicae. By analyzing the FTIR spectra of P. brassicae, infected clubroots and healthy roots, three specific bands 1 105, 1 145 and 1 228 cm-1 were selected. According to the correlation between the peak areas at these sensitive bands and Real-time PCR Ct value, quantitative evaluation model of P. brassicae was established based on FTIR y=34. 17 +12. 24x - 9. 81x2 - 6. 05x3, r=0. 98 (p<0. 05). To validate accuracy of the model, 10 clubroot samples were selected randomly from field, and detected by FTIR spectrum model, the results showed that the average error is 1. 60%. This demonstrated that the FTIR technology is an available one for the quantitative detection of P. brassicae in clubroot, and it provides a new method for quantitative and quickly detection of Chinese cabbage clubroot.

  19. New approaches in GMO detection.

    PubMed

    Querci, Maddalena; Van den Bulcke, Marc; Zel, Jana; Van den Eede, Guy; Broll, Hermann

    2010-03-01

    The steady rate of development and diffusion of genetically modified plants and their increasing diversification of characteristics, genes and genetic control elements poses a challenge in analysis of genetically modified organisms (GMOs). It is expected that in the near future the picture will be even more complex. Traditional approaches, mostly based on the sequential detection of one target at a time, or on a limited multiplexing, allowing only a few targets to be analysed at once, no longer meet the testing requirements. Along with new analytical technologies, new approaches for the detection of GMOs authorized for commercial purposes in various countries have been developed that rely on (1) a smart and accurate strategy for target selection, (2) the use of high-throughput systems or platforms for the detection of multiple targets and (3) algorithms that allow the conversion of analytical results into an indication of the presence of individual GMOs potentially present in an unknown sample. This paper reviews the latest progress made in GMO analysis, taking examples from the most recently developed strategies and tools, and addresses some of the critical aspects related to these approaches. PMID:19876618

  20. New approaches in GMO detection.

    PubMed

    Querci, Maddalena; Van den Bulcke, Marc; Zel, Jana; Van den Eede, Guy; Broll, Hermann

    2010-03-01

    The steady rate of development and diffusion of genetically modified plants and their increasing diversification of characteristics, genes and genetic control elements poses a challenge in analysis of genetically modified organisms (GMOs). It is expected that in the near future the picture will be even more complex. Traditional approaches, mostly based on the sequential detection of one target at a time, or on a limited multiplexing, allowing only a few targets to be analysed at once, no longer meet the testing requirements. Along with new analytical technologies, new approaches for the detection of GMOs authorized for commercial purposes in various countries have been developed that rely on (1) a smart and accurate strategy for target selection, (2) the use of high-throughput systems or platforms for the detection of multiple targets and (3) algorithms that allow the conversion of analytical results into an indication of the presence of individual GMOs potentially present in an unknown sample. This paper reviews the latest progress made in GMO analysis, taking examples from the most recently developed strategies and tools, and addresses some of the critical aspects related to these approaches.

  1. Current Approaches Toward Quantitative Mapping of the Interactome

    PubMed Central

    Buntru, Alexander; Trepte, Philipp; Klockmeier, Konrad; Schnoegl, Sigrid; Wanker, Erich E.

    2016-01-01

    Protein–protein interactions (PPIs) play a key role in many, if not all, cellular processes. Disease is often caused by perturbation of PPIs, as recently indicated by studies of missense mutations. To understand the associations of proteins and to unravel the global picture of PPIs in the cell, different experimental detection techniques for PPIs have been established. Genetic and biochemical methods such as the yeast two-hybrid system or affinity purification-based approaches are well suited to high-throughput, proteome-wide screening and are mainly used to obtain qualitative results. However, they have been criticized for not reflecting the cellular situation or the dynamic nature of PPIs. In this review, we provide an overview of various genetic methods that go beyond qualitative detection and allow quantitative measuring of PPIs in mammalian cells, such as dual luminescence-based co-immunoprecipitation, Förster resonance energy transfer or luminescence-based mammalian interactome mapping with bait control. We discuss the strengths and weaknesses of different techniques and their potential applications in biomedical research. PMID:27200083

  2. Quantitative vs. Qualitative Approaches to Quality Special Education Program Evaluation.

    ERIC Educational Resources Information Center

    Council of Administrators of Special Education, Inc.

    One in a series of issue papers commissioned by the Council of Administrators of Special Education (CASE), this document presents a comparison of contemporary evaluation approaches for special education programs. The first section describes the two approaches to be compared: (1) traditional scientific inquiry which emphasizes quantitative methods;…

  3. Quantitation and detection of vanadium in biologic and pollution materials

    NASA Technical Reports Server (NTRS)

    Gordon, W. A.

    1974-01-01

    A review is presented of special considerations and methodology for determining vanadium in biological and air pollution materials. In addition to descriptions of specific analysis procedures, general sections are included on quantitation of analysis procedures, sample preparation, blanks, and methods of detection of vanadium. Most of the information presented is applicable to the determination of other trace elements in addition to vanadium.

  4. Frequency-domain approaches to quantitative brain SPECT

    NASA Astrophysics Data System (ADS)

    Cheng, Jui-Hsi

    1997-12-01

    Quantitative SPECT (Single Photon Emission Computed Tomography) has been limited mainly by (1) inadequate numbers of detected photons which are contaminated by Poisson noise in projections, (2) photon attenuation in the body, (3) inclusion of scattered photons in the projections, and (4) depth-dependent blurring due to the finite size of collimator holes. Various methods to compensate for the above effects via either spatial- domain approaches or frequency-domain approaches have been proposed. However, most of the proposed methods focus only on individual effects. A reconstruction method which can compensate for all of the effects simultaneously is necessary. We have developed an algorithm to compensate for all of the effects simultaneously using frequency-domain approaches. For noise suppression, a method to convert the signal-dependent Poisson noise into signal- independent Gaussian white noise was first applied. Then the Wiener filter with a designed butterfly window was used. For scatter-photon removal, a subtraction technique based on multiple energy-window acquisitions was employed. For collimation deblurring, a direct inverse filter was designed via stationary phase condition (depth-frequency relationship). For attenuation compensation, an exact analytical solution was derived by Fourier analysis. Our algorithm was executed on a HP/730 workstation. An improvement was shown in computing time, noise suppression, recognition of phantom features, and quantification of concentrations of regions of interest (ROI). In addition to simulation, we performed a series of experiments to verify our models and test our algorithm. These include (1) investigation of the characteristics of Poisson noise of SPECT, (2) comparison of the performance of dual-energy window and triple-energy window methods for scatter correction, (3) measurement of a depth- dependent PSF, (4) reconstruction of the attenuation map using scatter-window data, and (5) implementation of a simultaneous

  5. Problems in Achieving a Quantitative Approach to Technologic Proliferation Resistance

    SciTech Connect

    Wiborg, James C.; Omberg, Ronald P.; Zentner, Michael D.

    2001-07-06

    In spite of setbacks, substantial success has been achieved by the various nonproliferation efforts over the past 50 years. Because the pace of technology evolution remains high and the cost of entry to nuclear weapons technology is decreasing, improved approaches are critical if similar success is to be achieved over the next 20 years. Recent analyses have been published that provide a semi-quantitative assessment of proliferation risk, which can serve as the foundation for a meaningful quantitative approach to assessing proliferation risk. These methods represent an important step, but represent only one step in the work that must be achieved in the next few years. This paper presents perspectives on evaluating the merits of institutional arrangements and the role of design versus institutional features in proliferation prevention. It concludes by proposing methodology and quantitative approaches to be considered for evaluating proliferation-resistant measures in innovative reactor and fuel cycle technologies.

  6. Accounting for Imperfect Detection in Ecology: A Quantitative Review

    PubMed Central

    Kellner, Kenneth F.; Swihart, Robert K.

    2014-01-01

    Detection in studies of species abundance and distribution is often imperfect. Assuming perfect detection introduces bias into estimation that can weaken inference upon which understanding and policy are based. Despite availability of numerous methods designed to address this assumption, many refereed papers in ecology fail to account for non-detection error. We conducted a quantitative literature review of 537 ecological articles to measure the degree to which studies of different taxa, at various scales, and over time have accounted for imperfect detection. Overall, just 23% of articles accounted for imperfect detection. The probability that an article incorporated imperfect detection increased with time and varied among taxa studied; studies of vertebrates were more likely to incorporate imperfect detection. Among articles that reported detection probability, 70% contained per-survey estimates of detection that were less than 0.5. For articles in which constancy of detection was tested, 86% reported significant variation. We hope that our findings prompt more ecologists to consider carefully the detection process when designing studies and analyzing results, especially for sub-disciplines where incorporation of imperfect detection in study design and analysis so far has been lacking. PMID:25356904

  7. Quantitative proteomics: assessing the spectrum of in-gel protein detection methods

    PubMed Central

    Gauci, Victoria J.; Wright, Elise P.

    2010-01-01

    Proteomics research relies heavily on visualization methods for detection of proteins separated by polyacrylamide gel electrophoresis. Commonly used staining approaches involve colorimetric dyes such as Coomassie Brilliant Blue, fluorescent dyes including Sypro Ruby, newly developed reactive fluorophores, as well as a plethora of others. The most desired characteristic in selecting one stain over another is sensitivity, but this is far from the only important parameter. This review evaluates protein detection methods in terms of their quantitative attributes, including limit of detection (i.e., sensitivity), linear dynamic range, inter-protein variability, capacity for spot detection after 2D gel electrophoresis, and compatibility with subsequent mass spectrometric analyses. Unfortunately, many of these quantitative criteria are not routinely or consistently addressed by most of the studies published to date. We would urge more rigorous routine characterization of stains and detection methodologies as a critical approach to systematically improving these critically important tools for quantitative proteomics. In addition, substantial improvements in detection technology, particularly over the last decade or so, emphasize the need to consider renewed characterization of existing stains; the quantitative stains we need, or at least the chemistries required for their future development, may well already exist. PMID:21686332

  8. An approach for quantitative image quality analysis for CT

    NASA Astrophysics Data System (ADS)

    Rahimi, Amir; Cochran, Joe; Mooney, Doug; Regensburger, Joe

    2016-03-01

    An objective and standardized approach to assess image quality of Compute Tomography (CT) systems is required in a wide variety of imaging processes to identify CT systems appropriate for a given application. We present an overview of the framework we have developed to help standardize and to objectively assess CT image quality for different models of CT scanners used for security applications. Within this framework, we have developed methods to quantitatively measure metrics that should correlate with feature identification, detection accuracy and precision, and image registration capabilities of CT machines and to identify strengths and weaknesses in different CT imaging technologies in transportation security. To that end we have designed, developed and constructed phantoms that allow for systematic and repeatable measurements of roughly 88 image quality metrics, representing modulation transfer function, noise equivalent quanta, noise power spectra, slice sensitivity profiles, streak artifacts, CT number uniformity, CT number consistency, object length accuracy, CT number path length consistency, and object registration. Furthermore, we have developed a sophisticated MATLAB based image analysis tool kit to analyze CT generated images of phantoms and report these metrics in a format that is standardized across the considered models of CT scanners, allowing for comparative image quality analysis within a CT model or between different CT models. In addition, we have developed a modified sparse principal component analysis (SPCA) method to generate a modified set of PCA components as compared to the standard principal component analysis (PCA) with sparse loadings in conjunction with Hotelling T2 statistical analysis method to compare, qualify, and detect faults in the tested systems.

  9. Infusing Quantitative Approaches throughout the Biological Sciences Curriculum

    ERIC Educational Resources Information Center

    Thompson, Katerina V.; Cooke, Todd J.; Fagan, William F.; Gulick, Denny; Levy, Doron; Nelson, Kären C.; Redish, Edward F.; Smith, Robert F.; Presson, Joelle

    2013-01-01

    A major curriculum redesign effort at the University of Maryland is infusing all levels of our undergraduate biological sciences curriculum with increased emphasis on interdisciplinary connections and quantitative approaches. The curriculum development efforts have largely been guided by recommendations in the National Research Council's…

  10. Rapid and quantitative detection of hepatitis B virus

    PubMed Central

    Liu, Yue-Ping; Yao, Chun-Yan

    2015-01-01

    Despite availability of a universal vaccine, hepatitis B virus (HBV) infection has a huge impact on public health worldwide. Accurate and timely diagnosis of HBV infection is needed. Rapid developments have been made in the diagnostic and monitoring methods for HBV infection, including serological and molecular assays. In clinical practice, qualitative hepatitis B surface antigen (HBsAg) testing has long served as a diagnostic marker for individuals infected with HBV. More recently, HBsAg level has been used to predict treatment outcome when determined early during treatment or at baseline. However, identification of HBV DNA positive cases that do not have detectable HBsAg has encouraged the application of molecular tests. Hence, combination of quantitative detection of HBV DNA and HBsAg can be used to discriminate patients during the course of HBV infection and to monitor therapy. This article reviews the most commonly used quantitative methods for HBsAg and HBV DNA. PMID:26576084

  11. Fast and Accurate Detection of Multiple Quantitative Trait Loci

    PubMed Central

    Nettelblad, Carl; Holmgren, Sverker

    2013-01-01

    Abstract We present a new computational scheme that enables efficient and reliable quantitative trait loci (QTL) scans for experimental populations. Using a standard brute-force exhaustive search effectively prohibits accurate QTL scans involving more than two loci to be performed in practice, at least if permutation testing is used to determine significance. Some more elaborate global optimization approaches, for example, DIRECT have been adopted earlier to QTL search problems. Dramatic speedups have been reported for high-dimensional scans. However, since a heuristic termination criterion must be used in these types of algorithms, the accuracy of the optimization process cannot be guaranteed. Indeed, earlier results show that a small bias in the significance thresholds is sometimes introduced. Our new optimization scheme, PruneDIRECT, is based on an analysis leading to a computable (Lipschitz) bound on the slope of a transformed objective function. The bound is derived for both infinite- and finite-size populations. Introducing a Lipschitz bound in DIRECT leads to an algorithm related to classical Lipschitz optimization. Regions in the search space can be permanently excluded (pruned) during the optimization process. Heuristic termination criteria can thus be avoided. Hence, PruneDIRECT has a well-defined error bound and can in practice be guaranteed to be equivalent to a corresponding exhaustive search. We present simulation results that show that for simultaneous mapping of three QTLS using permutation testing, PruneDIRECT is typically more than 50 times faster than exhaustive search. The speedup is higher for stronger QTL. This could be used to quickly detect strong candidate eQTL networks. PMID:23919387

  12. A microfabrication-based approach to quantitative isothermal titration calorimetry.

    PubMed

    Wang, Bin; Jia, Yuan; Lin, Qiao

    2016-04-15

    Isothermal titration calorimetry (ITC) directly measures heat evolved in a chemical reaction to determine equilibrium binding properties of biomolecular systems. Conventional ITC instruments are expensive, use complicated design and construction, and require long analysis times. Microfabricated calorimetric devices are promising, although they have yet to allow accurate, quantitative ITC measurements of biochemical reactions. This paper presents a microfabrication-based approach to integrated, quantitative ITC characterization of biomolecular interactions. The approach integrates microfabricated differential calorimetric sensors with microfluidic titration. Biomolecules and reagents are introduced at each of a series of molar ratios, mixed, and allowed to react. The reaction thermal power is differentially measured, and used to determine the thermodynamic profile of the biomolecular interactions. Implemented in a microdevice featuring thermally isolated, well-defined reaction volumes with minimized fluid evaporation as well as highly sensitive thermoelectric sensing, the approach enables accurate and quantitative ITC measurements of protein-ligand interactions under different isothermal conditions. Using the approach, we demonstrate ITC characterization of the binding of 18-Crown-6 with barium chloride, and the binding of ribonuclease A with cytidine 2'-monophosphate within reaction volumes of approximately 0.7 µL and at concentrations down to 2mM. For each binding system, the ITC measurements were completed with considerably reduced analysis times and material consumption, and yielded a complete thermodynamic profile of the molecular interaction in agreement with published data. This demonstrates the potential usefulness of our approach for biomolecular characterization in biomedical applications. PMID:26655185

  13. A microfabrication-based approach to quantitative isothermal titration calorimetry.

    PubMed

    Wang, Bin; Jia, Yuan; Lin, Qiao

    2016-04-15

    Isothermal titration calorimetry (ITC) directly measures heat evolved in a chemical reaction to determine equilibrium binding properties of biomolecular systems. Conventional ITC instruments are expensive, use complicated design and construction, and require long analysis times. Microfabricated calorimetric devices are promising, although they have yet to allow accurate, quantitative ITC measurements of biochemical reactions. This paper presents a microfabrication-based approach to integrated, quantitative ITC characterization of biomolecular interactions. The approach integrates microfabricated differential calorimetric sensors with microfluidic titration. Biomolecules and reagents are introduced at each of a series of molar ratios, mixed, and allowed to react. The reaction thermal power is differentially measured, and used to determine the thermodynamic profile of the biomolecular interactions. Implemented in a microdevice featuring thermally isolated, well-defined reaction volumes with minimized fluid evaporation as well as highly sensitive thermoelectric sensing, the approach enables accurate and quantitative ITC measurements of protein-ligand interactions under different isothermal conditions. Using the approach, we demonstrate ITC characterization of the binding of 18-Crown-6 with barium chloride, and the binding of ribonuclease A with cytidine 2'-monophosphate within reaction volumes of approximately 0.7 µL and at concentrations down to 2mM. For each binding system, the ITC measurements were completed with considerably reduced analysis times and material consumption, and yielded a complete thermodynamic profile of the molecular interaction in agreement with published data. This demonstrates the potential usefulness of our approach for biomolecular characterization in biomedical applications.

  14. Biotechnology Approaches to Life Detection

    NASA Technical Reports Server (NTRS)

    Steele, Andrew; McKay, David; Schweitzer, Mary

    2001-01-01

    The direct detection of organic biomarkers for living or fossil microbes on Mars by an in situ instrument is a worthy goal for future lander missions. Several new and innovative biotechnology approaches are being explored. Firstly we have proposed an instrument based on immunological reactions to specific antibodies to cause activation of fluorescent stains. Antibodies are raised or acquired to a variety of general and specific substances that might be in Mars soil. These antibodies are then combined with various fluorescent stains and applied to micron sized numbered spots on a small (2-3 cm) test plate where they become firmly attached after freeze drying. Using technology that has been developed for gene mining in DNA technology up to 10,000 tests per square inch can now be applied to a test plate. On Mars or the planet/moon of interest, a sample of soil from a trench or drill core is extracted with water and/or an organic solvent and ultrasonication and then applied to the test plate. Any substance, which has an antibody on the test plate, will react with its antibody and activate its fluorescent stain. At the moment a small UV light source will illuminate the test plate, which is observed with a small CCD camera, although other detection systems will be applied. The numbered spots that fluoresce indicate the presence of the tested-for substance, and the intensity indicates relative amounts. Furthermore with up to a thousand test plates available false positives and several variations of antibody can also be screened for. The entire instrument can be quite small and light, on the order of 10 cm in each dimension. A possible choice for light source may be small UV lasers at several wavelengths. Some of the wells or spots can contain simply standard fluorescent stains used to detect live cells, dead cells, DNA, etc. The stains in these spots may be directly activated, with no antibodies being necessary. The proposed system will look for three classes of

  15. Quantitative rotor damage detection based on piezoelectric impedance

    NASA Astrophysics Data System (ADS)

    Qin, Yi; Tao, Yi; Mao, Yongfang; Tang, Baoping

    2015-12-01

    To realize the quantitative damage detection of a rotor, firstly an impedance analytic model is built. Then the change of bending stiffness is introduced as the damage index. Given the circular boundary condition of a rotor, annular elements are used as the analyzed objects and spectral element method is used. The electro-mechanical (E/M) coupled impedance expression of an undamaged rotor is derived with the application of a low-cost impedance test circuit. A Taylor expansion method is used to obtain the approximate E/M coupled impedance expression for the damaged rotor. After obtaining the difference between the undamaged and damaged rotor impedance, a rotor damage detection algorithm is proposed. In this paper, a preset damage configuration is used for the numerical simulation and experiment validation. The detection results have shown that the quantitative damage detection algorithm based on spectral element method and piezoelectric impedance proposed in this paper can identify the location and the severity of the damaged rotor accurately.

  16. Quantitative Proteomic Approaches for Analysis of Protein S-Nitrosylation.

    PubMed

    Qu, Zhe; Greenlief, C Michael; Gu, Zezong

    2016-01-01

    S-Nitrosylation is a redox-based post-translational modification of a protein in response to nitric oxide (NO) signaling, and it participates in a variety of processes in diverse biological systems. The significance of this type of protein modification in health and diseases is increasingly recognized. In the central nervous system, aberrant S-nitrosylation, due to excessive NO production, is known to cause protein misfolding, mitochondrial dysfunction, transcriptional dysregulation, and neuronal death. This leads to an altered physiological state and consequently contributes to pathogenesis of neurodegenerative disorders. To date, much effort has been made to understand the mechanisms underlying protein S-nitrosylation, and several approaches have been developed to unveil S-nitrosylated proteins from different organisms. Interest in determining the dynamic changes of protein S-nitrosylation under different physiological and pathophysiological conditions has underscored the need for the development of quantitative proteomic approaches. Currently, both gel-based and gel-free mass spectrometry-based quantitative methods are widely used, and they each have advantages and disadvantages but may also be used together to produce complementary data. This review evaluates current available quantitative proteomic techniques for the analysis of protein S-nitrosylation and highlights recent advances, with emphasis on applications in neurodegenerative diseases. An important goal is to provide a comprehensive guide of feasible quantitative proteomic methodologies for examining protein S-nitrosylation in research to yield insights into disease mechanisms, diagnostic biomarkers, and drug discovery.

  17. Quantitative Genetic Interaction Mapping Using the E-MAP Approach

    PubMed Central

    Collins, Sean R.; Roguev, Assen; Krogan, Nevan J.

    2010-01-01

    Genetic interactions represent the degree to which the presence of one mutation modulates the phenotype of a second mutation. In recent years, approaches for measuring genetic interactions systematically and quantitatively have proven to be effective tools for unbiased characterization of gene function and have provided valuable data for analyses of evolution. Here, we present protocols for systematic measurement of genetic interactions with respect to organismal growth rate for two yeast species. PMID:20946812

  18. Quantitative detection of settled dust over green canopy

    NASA Astrophysics Data System (ADS)

    Brook, Anna

    2016-04-01

    NMF (SS-NMF), 6) Alternating Least-Square (ALS), and 2) Lin's Projected Gradient (LPG). The performance is evaluated on real hyperspectral imagery data via detailed experimental assessment. The study showed that in certain compression tasks content-adapted sparse representation is provided by state-of-the-art solutions. The NMF algorithm estimates endmembers that are used to remove spurious information. If computationally feasible, it should include interaction terms to make the model more flexible. The optimal NMF algorithms, such as ALS and LPG, are assumed to be the simplest methods that achieve the minimum error on the test set. In summary, this work shows that sediment dust can be assessed using airborne HSI data, making it a potentially powerful tool for environmental studies. References Keshava, N., Mustard, J. (2002). Spectral unmixing. IEEE Signal Process. Mag., 19(1), 44-57. Chudnovsky, A., & Ben-Dor, E. (2009). Reflectance spectroscopy as a tool for settled dust monitoring in office environment. International Journal of Environment and Waste Management, 4(1), 32-49. Brook, A. (2014). Quantitative Detection of Settled dust over Green Canopy using Sparse Unmixing of Airborne Hyperspectral Data. IEEE-Whispers 6th Workshop on Hyperspectral Image and Signal Processing: Evolution in Remote Sensing, 2014, Switzerland, 4-8. Keshava, N., Mustard, J. (2002). Spectral unmixing. IEEE Signal Process. Mag., 19(1), 44-57. Bioucas-Dias et al. (2012). Hyperspectral unmixing overview: Geometrical, statistical, and sparse regression-based approaches, IEEE Journal of Selected Topics in Applied Earth Observations and Remote Sensing, 5(2), 354 -379.

  19. Quantitative optical imaging for the detection of early cancer

    NASA Astrophysics Data System (ADS)

    Wu, Tao

    The objectives of this thesis are to provide insight of fundamental mechanisms of acetowhitening effect, upon which the colposcopic diagnosis of human cervical cancer is based and to develop novel quantitative optical imaging technologies supplementing colposcopy to improve its performance in detecting early cancer. Firstly, the temporal characteristics of acetowhitening process are studied on monolayer cell cultures. It is found that the dynamic acetowhitening processes in normal and cancerous cells are significantly different. Secondly, the changes in light scattering induced by acetic acid in intact cells and isolated cellular fractions are investigated by using confocal microscopy and light scattering spectroscopy. The results provide evidence that the small-sized components in the cytoplasm are the major contributors to the acetowhitening effect. Thirdly, a unified Mie and fractal model is proposed to interpret light scattering by biological cells. It is found that light scattering in forward directions is dominated by Mie scattering by bare cells and nuclei, whereas light scattering at large angles is determined by fractal scattering by subcellular structures. Fourthly, an optical imaging system based on active stereo vision and motion tracking is built to measure the 3-D surface topology of cervix and track the motion of patient. The information of motion tracking is used to register the time-sequenced images of cervix recorded during colposcopic examination. The imaging system is evaluated by tracking the movements of cervix models. The results demonstrate that the imaging technique holds the promise to enable the quantitative mapping of the acetowhitening kinetics over cervical surface for more accurate diagnosis of cervical cancer. At last, a calibrated autofluorescence imaging system is instrumented for detecting neoplasia in vivo. It is found that the calibrated autofluorescence signals from neoplasia are generally lower than signals from normal

  20. A quantitative approach to evolution of music and philosophy

    NASA Astrophysics Data System (ADS)

    Vieira, Vilson; Fabbri, Renato; Travieso, Gonzalo; Oliveira, Osvaldo N., Jr.; da Fontoura Costa, Luciano

    2012-08-01

    The development of new statistical and computational methods is increasingly making it possible to bridge the gap between hard sciences and humanities. In this study, we propose an approach based on a quantitative evaluation of attributes of objects in fields of humanities, from which concepts such as dialectics and opposition are formally defined mathematically. As case studies, we analyzed the temporal evolution of classical music and philosophy by obtaining data for 8 features characterizing the corresponding fields for 7 well-known composers and philosophers, which were treated with multivariate statistics and pattern recognition methods. A bootstrap method was applied to avoid statistical bias caused by the small sample data set, with which hundreds of artificial composers and philosophers were generated, influenced by the 7 names originally chosen. Upon defining indices for opposition, skewness and counter-dialectics, we confirmed the intuitive analysis of historians in that classical music evolved according to a master-apprentice tradition, while in philosophy changes were driven by opposition. Though these case studies were meant only to show the possibility of treating phenomena in humanities quantitatively, including a quantitative measure of concepts such as dialectics and opposition, the results are encouraging for further application of the approach presented here to many other areas, since it is entirely generic.

  1. Comparison of Droplet Digital PCR and Quantitative PCR Assays for Quantitative Detection of Xanthomonas citri Subsp. citri

    PubMed Central

    Yin, Youping; Wang, Zhongkang

    2016-01-01

    Droplet digital polymerase chain reaction (ddPCR) is a novel molecular biology technique providing absolute quantification of target nucleic acids without the need for an external calibrator. Despite its emerging applications in medical diagnosis, there are few reports of its use for the detection of plant pathogens. This work was designed to assess the diagnosis potential of the ddPCR for absolute quantitative detection of Xanthomonas citri subsp. citri, a quarantine plant pathogenic bacterium that causes citrus bacterial canker in susceptible Citrus species. We transferred an established quantitative PCR (qPCR) assay for citrus bacterial canker diagnosis directly to the ddPCR format and compared the performance of the two methods. The qPCR assay has a broader dynamic range compared to the ddPCR assay and the ddPCR assay has a significantly higher degree of sensitivity compared to the qPCR assay. The influence of PCR inhibitors can be reduced considerably in the ddPCR assay because the collection of end-point fluorescent signals and the counting of binomial events (positive or negative droplets) are associated with a Poisson algorithm. The ddPCR assay also shows lower coefficient of variation compared to the qPCR assay especially in low target concentration. The linear association of the measurements by ddPCR and qPCR assays is strong (Pearson correlation = 0.8633; P<0.001). Receiver operating characteristic analysis indicates the ddPCR methodology is a more robust approach for diagnosis of citrus bacterial canker. In summary, the results demonstrated that the ddPCR assay has the potential for the quantitative detection of X. citri subsp. citri with high precision and accuracy as compared with the results from qPCR assay. Further studies are required to evaluate and validate the value of ddPCR technology in the diagnosis of plant disease and quarantine applications. PMID:27427975

  2. Comparison of Droplet Digital PCR and Quantitative PCR Assays for Quantitative Detection of Xanthomonas citri Subsp. citri.

    PubMed

    Zhao, Yun; Xia, Qingyan; Yin, Youping; Wang, Zhongkang

    2016-01-01

    Droplet digital polymerase chain reaction (ddPCR) is a novel molecular biology technique providing absolute quantification of target nucleic acids without the need for an external calibrator. Despite its emerging applications in medical diagnosis, there are few reports of its use for the detection of plant pathogens. This work was designed to assess the diagnosis potential of the ddPCR for absolute quantitative detection of Xanthomonas citri subsp. citri, a quarantine plant pathogenic bacterium that causes citrus bacterial canker in susceptible Citrus species. We transferred an established quantitative PCR (qPCR) assay for citrus bacterial canker diagnosis directly to the ddPCR format and compared the performance of the two methods. The qPCR assay has a broader dynamic range compared to the ddPCR assay and the ddPCR assay has a significantly higher degree of sensitivity compared to the qPCR assay. The influence of PCR inhibitors can be reduced considerably in the ddPCR assay because the collection of end-point fluorescent signals and the counting of binomial events (positive or negative droplets) are associated with a Poisson algorithm. The ddPCR assay also shows lower coefficient of variation compared to the qPCR assay especially in low target concentration. The linear association of the measurements by ddPCR and qPCR assays is strong (Pearson correlation = 0.8633; P<0.001). Receiver operating characteristic analysis indicates the ddPCR methodology is a more robust approach for diagnosis of citrus bacterial canker. In summary, the results demonstrated that the ddPCR assay has the potential for the quantitative detection of X. citri subsp. citri with high precision and accuracy as compared with the results from qPCR assay. Further studies are required to evaluate and validate the value of ddPCR technology in the diagnosis of plant disease and quarantine applications. PMID:27427975

  3. Hyperspectral imaging and quantitative analysis for prostate cancer detection

    PubMed Central

    Akbari, Hamed; Halig, Luma V.; Schuster, David M.; Osunkoya, Adeboye; Master, Viraj; Nieh, Peter T.; Chen, Georgia Z.

    2012-01-01

    Abstract. Hyperspectral imaging (HSI) is an emerging modality for various medical applications. Its spectroscopic data might be able to be used to noninvasively detect cancer. Quantitative analysis is often necessary in order to differentiate healthy from diseased tissue. We propose the use of an advanced image processing and classification method in order to analyze hyperspectral image data for prostate cancer detection. The spectral signatures were extracted and evaluated in both cancerous and normal tissue. Least squares support vector machines were developed and evaluated for classifying hyperspectral data in order to enhance the detection of cancer tissue. This method was used to detect prostate cancer in tumor-bearing mice and on pathology slides. Spatially resolved images were created to highlight the differences of the reflectance properties of cancer versus those of normal tissue. Preliminary results with 11 mice showed that the sensitivity and specificity of the hyperspectral image classification method are 92.8% to 2.0% and 96.9% to 1.3%, respectively. Therefore, this imaging method may be able to help physicians to dissect malignant regions with a safe margin and to evaluate the tumor bed after resection. This pilot study may lead to advances in the optical diagnosis of prostate cancer using HSI technology. PMID:22894488

  4. Mass spectrometric-based approaches in quantitative proteomics.

    PubMed

    Ong, Shao-En; Foster, Leonard J; Mann, Matthias

    2003-02-01

    Classically, experiments aimed at studying changes in protein expression have always followed a small set of proteins. This focused approach was necessary since tools to efficiently analyze large numbers of proteins were simply not available. Large-scale quantitative proteomics promises to produce reams of data that previously would have taken decades to measure with classical methods. Mass spectrometry is already a well-established protein identification tool and recent methodological developments indicate that it can also be successfully applied to extract quantitative data of protein abundance. From the first reports 4 years ago, numerous schemes to take advantage of stable isotope nuclei incorporation in proteins and peptides have been developed. Here we review the benefits and pitfalls of some of the most commonly used protocols, focusing on a procedure now being used extensively in our laboratory, stable isotope labeling with amino acids in cell culture (SILAC). The basic theory, application, and data analysis of a SILAC experiment are discussed. The emerging nature of these techniques and the rapid pace of technological development make forecasting the directions of the field difficult but we speculate that SILAC will soon be a key tool of quantitative proteomics.

  5. Detecting contingencies: an infomax approach.

    PubMed

    Butko, Nicholas J; Movellan, Javier R

    2010-01-01

    The ability to detect social contingencies plays an important role in the social and emotional development of infants. Analyzing this problem from a computational perspective may provide important clues for understanding social development, as well as for the synthesis of social behavior in robots. In this paper, we show that the turn-taking behaviors observed in infants during contingency detection situations are tuned to optimally gather information as to whether a person is responsive to them. We show that simple reinforcement learning mechanisms can explain how infants acquire these efficient contingency detection schemas. The key is to use the reduction of uncertainty (information gain) as a reward signal. The result is an interesting form of learning in which the learner rewards itself for conducting actions that help reduce its own sense of uncertainty. This paper illustrates the possibilities of an emerging area of computer science and engineering that focuses on the computational understanding of human behavior and on its synthesis in robots. We believe that the theory of stochastic optimal control will play a key role providing a formal mathematical foundation for this newly emerging discipline.

  6. Distance-based microfluidic quantitative detection methods for point-of-care testing.

    PubMed

    Tian, Tian; Li, Jiuxing; Song, Yanling; Zhou, Leiji; Zhu, Zhi; Yang, Chaoyong James

    2016-04-01

    Equipment-free devices with quantitative readout are of great significance to point-of-care testing (POCT), which provides real-time readout to users and is especially important in low-resource settings. Among various equipment-free approaches, distance-based visual quantitative detection methods rely on reading the visual signal length for corresponding target concentrations, thus eliminating the need for sophisticated instruments. The distance-based methods are low-cost, user-friendly and can be integrated into portable analytical devices. Moreover, such methods enable quantitative detection of various targets by the naked eye. In this review, we first introduce the concept and history of distance-based visual quantitative detection methods. Then, we summarize the main methods for translation of molecular signals to distance-based readout and discuss different microfluidic platforms (glass, PDMS, paper and thread) in terms of applications in biomedical diagnostics, food safety monitoring, and environmental analysis. Finally, the potential and future perspectives are discussed. PMID:26928571

  7. Distance-based microfluidic quantitative detection methods for point-of-care testing.

    PubMed

    Tian, Tian; Li, Jiuxing; Song, Yanling; Zhou, Leiji; Zhu, Zhi; Yang, Chaoyong James

    2016-04-01

    Equipment-free devices with quantitative readout are of great significance to point-of-care testing (POCT), which provides real-time readout to users and is especially important in low-resource settings. Among various equipment-free approaches, distance-based visual quantitative detection methods rely on reading the visual signal length for corresponding target concentrations, thus eliminating the need for sophisticated instruments. The distance-based methods are low-cost, user-friendly and can be integrated into portable analytical devices. Moreover, such methods enable quantitative detection of various targets by the naked eye. In this review, we first introduce the concept and history of distance-based visual quantitative detection methods. Then, we summarize the main methods for translation of molecular signals to distance-based readout and discuss different microfluidic platforms (glass, PDMS, paper and thread) in terms of applications in biomedical diagnostics, food safety monitoring, and environmental analysis. Finally, the potential and future perspectives are discussed.

  8. Fast detection of deletion breakpoints using quantitative PCR

    PubMed Central

    Abildinova, Gulshara; Abdrakhmanova, Zhanara; Tuchinsky, Helena; Nesher, Elimelech; Pinhasov, Albert; Raskin, Leon

    2016-01-01

    Abstract The routine detection of large and medium copy number variants (CNVs) is well established. Hemizygotic deletions or duplications in the large Duchenne muscular dystrophy DMD gene responsible for Duchenne and Becker muscular dystrophies are routinely identified using multiple ligation probe amplification and array-based comparative genomic hybridization. These methods only map deleted or duplicated exons, without providing the exact location of breakpoints. Commonly used methods for the detection of CNV breakpoints include long-range PCR and primer walking, their success being limited by the deletion size, GC content and presence of DNA repeats. Here, we present a strategy for detecting the breakpoints of medium and large CNVs regardless of their size. The hemizygous deletion of exons 45-50 in the DMD gene and the large autosomal heterozygous PARK2 deletion were used to demonstrate the workflow that relies on real-time quantitative PCR to narrow down the deletion region and Sanger sequencing for breakpoint confirmation. The strategy is fast, reliable and cost-efficient, making it amenable to widespread use in genetic laboratories. PMID:27560363

  9. Fast detection of deletion breakpoints using quantitative PCR.

    PubMed

    Abildinova, Gulshara; Abdrakhmanova, Zhanara; Tuchinsky, Helena; Nesher, Elimelech; Pinhasov, Albert; Raskin, Leon

    2016-01-01

    The routine detection of large and medium copy number variants (CNVs) is well established. Hemizygotic deletions or duplications in the large Duchenne muscular dystrophy DMD gene responsible for Duchenne and Becker muscular dystrophies are routinely identified using multiple ligation probe amplification and array-based comparative genomic hybridization. These methods only map deleted or duplicated exons, without providing the exact location of breakpoints. Commonly used methods for the detection of CNV breakpoints include long-range PCR and primer walking, their success being limited by the deletion size, GC content and presence of DNA repeats. Here, we present a strategy for detecting the breakpoints of medium and large CNVs regardless of their size. The hemizygous deletion of exons 45-50 in the DMD gene and the large autosomal heterozygous PARK2 deletion were used to demonstrate the workflow that relies on real-time quantitative PCR to narrow down the deletion region and Sanger sequencing for breakpoint confirmation. The strategy is fast, reliable and cost-efficient, making it amenable to widespread use in genetic laboratories. PMID:27560363

  10. A Nonparametric Approach for Mapping Quantitative Trait Loci

    PubMed Central

    Kruglyak, L.; Lander, E. S.

    1995-01-01

    Genetic mapping of quantitative trait loci (QTLs) is performed typically by using a parametric approach, based on the assumption that the phenotype follows a normal distribution. Many traits of interest, however, are not normally distributed. In this paper, we present a nonparametric approach to QTL mapping applicable to any phenotypic distribution. The method is based on a statistic Z(w), which generalizes the nonparametric Wilcoxon rank-sum test to the situation of whole-genome search by interval mapping. We determine the appropriate significance level for the statistic Z(w), by showing that its asymptotic null distribution follows an Ornstein-Uhlenbeck process. These results provide a robust, distribution-free method for mapping QTLs. PMID:7768449

  11. Salient object detection approach in UAV video

    NASA Astrophysics Data System (ADS)

    Zhang, Yueqiang; Su, Ang; Zhu, Xianwei; Zhang, Xiaohu; Shang, Yang

    2013-10-01

    The automatic detection of visually salient information from abundant video imagery is crucial, as it plays an important role in surveillance and reconnaissance tasks for Unmanned Aerial Vehicle (UAV). A real-time approach for the detection of salient objects on road, e.g. stationary and moving vehicle or people, is proposed, which is based on region segmentation and saliency detection within related domains. Generally, the traditional method specifically depends upon additional scene information and auxiliary thermal or IR sensing for secondary confirmation. However, this proposed approach can detect the interesting objects directly from video imagery captured by optical camera fixed on the small level UAV platform. To validate this proposed salient object detection approach, the 25 Hz video data from our low speed small UAV are tested. The results have demonstrated the proposed approach performs excellently in isolated rural environments.

  12. A novel logic-based approach for quantitative toxicology prediction.

    PubMed

    Amini, Ata; Muggleton, Stephen H; Lodhi, Huma; Sternberg, Michael J E

    2007-01-01

    There is a pressing need for accurate in silico methods to predict the toxicity of molecules that are being introduced into the environment or are being developed into new pharmaceuticals. Predictive toxicology is in the realm of structure activity relationships (SAR), and many approaches have been used to derive such SAR. Previous work has shown that inductive logic programming (ILP) is a powerful approach that circumvents several major difficulties, such as molecular superposition, faced by some other SAR methods. The ILP approach reasons with chemical substructures within a relational framework and yields chemically understandable rules. Here, we report a general new approach, support vector inductive logic programming (SVILP), which extends the essentially qualitative ILP-based SAR to quantitative modeling. First, ILP is used to learn rules, the predictions of which are then used within a novel kernel to derive a support-vector generalization model. For a highly heterogeneous dataset of 576 molecules with known fathead minnow fish toxicity, the cross-validated correlation coefficients (R2CV) from a chemical descriptor method (CHEM) and SVILP are 0.52 and 0.66, respectively. The ILP, CHEM, and SVILP approaches correctly predict 55, 58, and 73%, respectively, of toxic molecules. In a set of 165 unseen molecules, the R2 values from the commercial software TOPKAT and SVILP are 0.26 and 0.57, respectively. In all calculations, SVILP showed significant improvements in comparison with the other methods. The SVILP approach has a major advantage in that it uses ILP automatically and consistently to derive rules, mostly novel, describing fragments that are toxicity alerts. The SVILP is a general machine-learning approach and has the potential of tackling many problems relevant to chemoinformatics including in silico drug design.

  13. Quantum dots assisted laser desorption/ionization mass spectrometric detection of carbohydrates: qualitative and quantitative analysis.

    PubMed

    Bibi, Aisha; Ju, Huangxian

    2016-04-01

    A quantum dots (QDs) assisted laser desorption/ionization mass spectrometric (QDA-LDI-MS) strategy was proposed for qualitative and quantitative analysis of a series of carbohydrates. The adsorption of carbohydrates on the modified surface of different QDs as the matrices depended mainly on the formation of hydrogen bonding, which led to higher MS intensity than those with conventional organic matrix. The effects of QDs concentration and sample preparation method were explored for improving the selective ionization process and the detection sensitivity. The proposed approach offered a new dimension to the application of QDs as matrices for MALDI-MS research of carbohydrates. It could be used for quantitative measurement of glucose concentration in human serum with good performance. The QDs served as a matrix showed the advantages of low background, higher sensitivity, convenient sample preparation and excellent stability under vacuum. The QDs assisted LDI-MS approach has promising application to the analysis of carbohydrates in complex biological samples.

  14. Glucose Oxidase-Catalyzed Growth of Gold Nanoparticles Enables Quantitative Detection of Attomolar Cancer Biomarkers

    PubMed Central

    2015-01-01

    Ultrasensitive and quantitative detection of cancer biomarkers is an unmet challenge because of their ultralow concentrations in clinical samples. Although gold nanoparticle (AuNP)-based immunoassays offer high sensitivity, they were unable to quantitatively detect targets of interest most likely due to their very narrow linear ranges. This article describes a quantitative colorimetric immunoassay based on glucose oxidase (GOx)-catalyzed growth of 5 nm AuNPs that can detect cancer biomarkers from attomolar to picomolar levels. In addition, the limit of detection (LOD) of prostate-specific antigen (PSA) of this approach (93 aM) exceeds that of commercial enzyme-linked immunosorbent assay (ELISA) (6.3 pM) by more than 4 orders of magnitude. The emergence of red or purple color based on enzyme-catalyzed growth of 5 nm AuNPs in the presence of target antigen is particularly suitable for point-of-care (POC) diagnostics in both resource-rich and resource-limited settings. PMID:24896231

  15. The use of selection experiments for detecting quantitative trait loci.

    PubMed

    Ollivier, L; Messer, L A; Rothschild, M F; Legault, C

    1997-06-01

    Gene frequency changes following selection may reveal the existence of gene effects on the trait selected. Loci for the selected quantitative trait (SQTL) may thus be detected. Additionally, one can estimate the average effect (alpha) of a marker allele associated with an SQTL from the allele frequency change (delta q) due to selection of given intensity (i). In a sample of unrelated individuals, it is optimal to select the upper and lower 27% for generating delta q in order to estimate alpha. For a given number of individuals genotyped, this estimator is 0.25i2 times more efficient than the classical estimator of alpha, based on the regression of the trait on the genotype at the marker locus. The method is extended to selection criteria using information from relatives, showing that combined selection considerably increases the efficiency of estimation for traits of low heritability. The method has been applied to the detection of SQTL in a selection experiment in which the trait selected was pig litter size averaged over the first four parities, with i = 3. Results for four genes are provided, one of which yielded a highly significant effect. The conditions required for valid application of the method are discussed, including selection experiments over several generations. Additional advantages of the method can be anticipated from determining gene frequencies on pooled samples of blood or DNA.

  16. Quantitative PCR detection for abalone shriveling syndrome-associated virus.

    PubMed

    Jiang, Jing-Zhe; Zhu, Zhen-Ni; Zhang, Han; Liang, Ya-Yu; Guo, Zhi-Xun; Liu, Guang-Feng; Su, You-Lu; Wang, Jiang-Yong

    2012-09-01

    Haliotis diversicolor (small abalone) is an important seafood found along the southern coast of China. Since 1999, the yields of cultured abalone in China have been severely affected by an epidemic of continuous outbreaks of a fatal disease. A novel double-stranded DNA virus, abalone shriveling syndrome-associated virus (AbSV), was proven to be one of the main causative agent. Although the pathogenicity and genome of AbSV has been ascertained, the epidemiology of AbSV remains to be investigated. In this study, four pairs of AbSV-specific primers were designed on the basis of the AbSV genome, and were tested for their specificities and sensitivities in quantitative real-time PCRs (qPCRs) after optimization of the annealing temperature. The 3F3/3B3 primer pair was finally chosen with a good specificity and high efficiency of amplification, with a detection limit of about 10 copies of recombinant plasmid containing AbSV genes in a 20-μL reaction mixture. In the detection of AbSV in abalone samples along the southern coast of China, most of the diseased samples had more than 80 virus copies in 1ng host genome DNA. AbSV was also demonstrated in mature hybrid (LY) and juvenile (JH) abalones from assays of healthy animals collected in recent years.

  17. A gold nanoparticle-based semi-quantitative and quantitative ultrasensitive paper sensor for the detection of twenty mycotoxins.

    PubMed

    Kong, Dezhao; Liu, Liqiang; Song, Shanshan; Suryoprabowo, Steven; Li, Aike; Kuang, Hua; Wang, Libing; Xu, Chuanlai

    2016-03-01

    A semi-quantitative and quantitative multi-immunochromatographic (ICA) strip detection assay was developed for the simultaneous detection of twenty types of mycotoxins from five classes, including zearalenones (ZEAs), deoxynivalenols (DONs), T-2 toxins (T-2s), aflatoxins (AFs), and fumonisins (FBs), in cereal food samples. Sensitive and specific monoclonal antibodies were selected for this assay. The semi-quantitative results were obtained within 20 min by the naked eye, with visual limits of detection for ZEAs, DONs, T-2s, AFs and FBs of 0.1-0.5, 2.5-250, 0.5-1, 0.25-1 and 2.5-10 μg kg(-1), and cut-off values of 0.25-1, 5-500, 1-10, 0.5-2.5 and 5-25 μg kg(-1), respectively. The quantitative results were obtained using a hand-held strip scan reader, with the calculated limits of detection for ZEAs, DONs, T-2s, AFs and FBs of 0.04-0.17, 0.06-49, 0.15-0.22, 0.056-0.49 and 0.53-1.05 μg kg(-1), respectively. The analytical results of spiked samples were in accordance with the accurate content in the simultaneous detection analysis. This newly developed ICA strip assay is suitable for the on-site detection and rapid initial screening of mycotoxins in cereal samples, facilitating both semi-quantitative and quantitative determination.

  18. Quantitative corrosion monitoring and detection using ultrasonic Lamb waves

    NASA Astrophysics Data System (ADS)

    Gordon, Grant A.; Braunling, Russ

    2005-05-01

    Corrosion is a major problem for airframe operators. For the aircraft industry in general, the direct costs of corrosion are estimated at $2.2 billion. As part of their strategy to control corrosion, airframe operators constantly seek to improve their ability to anticipate, manage and identify corrosion activity. Motivated by the need for an on-line real-time corrosion-monitoring tool for industry and aircraft a prototype system and analysis approach is presented. The tool employs ultrasonic Lamb waves along with a dispersion compensated synthetic aperture focusing technique (SAFT) to detect emerging pitting damage. In order to develop an automated detection approach the noise sources of the SAFT processed defect maps were examined and modeled. The random noise was found to be neither stationary nor normally distributed. Locally varying Weibull distribution parameters are used to characterize the image noise. An algorithm is developed to quantify the uncertainty in the corrosion detection and to allow assignment of a constant false alarm probability to any region of the monitored area.

  19. Quantitative proteomic approach for cellulose degradation by Neurospora crassa.

    PubMed

    Phillips, Christopher M; Iavarone, Anthony T; Marletta, Michael A

    2011-09-01

    Conversion of plant biomass to soluble sugars is the primary bottleneck associated with production of economically viable cellulosic fuels and chemicals. To better understand the biochemical route that filamentous fungi use to degrade plant biomass, we have taken a quantitative proteomics approach to characterizing the secretome of Neurospora crassa during growth on microcrystalline cellulose. Thirteen proteins were quantified in the N. crassa secretome using a combination of Absolute Quantification (AQUA) and Absolute SILAC to verify protein concentrations. Four of these enzymes including 2 cellobiohydrolases (CBH-1 and GH6-2), an endoglucanase (GH5-1), and a β-glucosidase (GH3-4) were then chosen to reconstitute a defined cellulase mixture in vitro. These enzymes were assayed alone and in mixtures and the activity of the reconstituted set was then compared to the crude mixture of N. crassa secretome proteins. Results show that while these 4 proteins represent 63-65% of the total secretome by weight, they account for just 43% of the total activity on microcrystalline cellulose after 24 h of hydrolysis. This result and quantitative proteomic data on other less abundant proteins secreted by Neurospora suggest that proteins other than canonical fungal cellulases may play an important role in cellulose degradation by fungi. PMID:21744778

  20. New approach to quantitative angiographic assessment after stent implantation.

    PubMed

    Reimers, B; Di Mario, C; Di Francesco, L; Moussa, I; Blengino, S; Martini, G; Reiber, J H; Colombo, A

    1997-04-01

    The new generation quantitative angiographic systems apply the interpolated technique to calculate the reference diameter at the site of the stenosis by integrating measurements of the segments proximal and distal to the stenosis. After stent implantation these measurements can be misleading as the treated segment, which is frequently larger than the adjacent not stented segments, is included in the measurements. The consequence is an overestimation of the reference diameter and the residual diameter stenosis. The present study was performed to compare this conventional technique of measurement with a new method which excludes the stented segment for the calculation of the reference diameter. Fifty-two lesions treated with poorly radiopaque stents (56% Palmaz-Schatz, 28% NIR, 10% Gianturco-Roubin, 6% Wallstent) expanded at high pressure (> = or 16 atm) were analyzed according to the conventional and stent excluded method. After stent implantation the reference diameter was 3.39 +/- 0.48 mm with conventional measurements and 3.02 +/- 0.45 mm with the stent excluded method (P < 0.05). The corresponding % diameter stenosis was 13 +/- 9 for the conventional technique and 1 +/- 13 for the stent excluded analysis (P < 0.05). The new approach to quantitative coronary analysis after stenting provides higher accuracy in reference diameter calculations and allows a more appropriate matching of stented segments with adjacent normal segments.

  1. A New, Principled Approach to Anomaly Detection

    SciTech Connect

    Ferragut, Erik M; Laska, Jason A; Bridges, Robert A

    2012-01-01

    Intrusion detection is often described as having two main approaches: signature-based and anomaly-based. We argue that only unsupervised methods are suitable for detecting anomalies. However, there has been a tendency in the literature to conflate the notion of an anomaly with the notion of a malicious event. As a result, the methods used to discover anomalies have typically been ad hoc, making it nearly impossible to systematically compare between models or regulate the number of alerts. We propose a new, principled approach to anomaly detection that addresses the main shortcomings of ad hoc approaches. We provide both theoretical and cyber-specific examples to demonstrate the benefits of our more principled approach.

  2. Detection of Saccharopolyspora rectivirgula by quantitative real-time PCR.

    PubMed

    Schäfer, Jenny; Kämpfer, Peter; Jäckel, Udo

    2011-07-01

    The thermophilic actinomycete species Saccharopolyspora rectivirgula has been associated with the exogen allergic alveolitis (EAA). EAA is caused by the inhalation of high amounts of airborne spores that can be found for example in environments of agricultural production, compost facilities, mushroom cultivation rooms, or rooms with technical air moistening. Because of the medical relevance of S. rectivirgula, a reliable detection system is needed. Therefore, a quantitative real-time polymerase chain reaction (qPCR) primer system was designed, targeting the 16S rRNA gene of the type strain S. rectivirgula DSM 43747(T) and six other S. rectivirgula reference strains. Our investigation showed that S. rectivirgula presumably own four operons of the 16S rRNA gene, which has to be considered for estimation of cell equivalents. Furthermore, the DNA recovery efficiency from these strains was tested in combination with bioaerosol or material sample as well as the influence of non-target DNA to the recovery rate. Results showed a recovery DNA efficiency of 7-55%. The recovery rate of DNA in a mixture with non-target DNA resulted in ∼87%. In summary, a high amplification efficiency using real-time PCR was found, for which estimated concentrations revealed cell numbers of 2.7 × 10(5) cells m(-3) in bioaerosol and 2.8 × 10(6) cells g(-1) fw(-1) in material samples from a duck house. The specificity of the new developed quantification system was shown by generation of two clone libraries from bioarosol samples, from a duck house, and from a composting plant. Totally, the results clearly show the specificity and practicability of the established qPCR assay for detection of S. rectivirgula.

  3. Human lymphocyte polymorphisms detected by quantitative two-dimensional electrophoresis

    SciTech Connect

    Goldman, D.; Merril, C.R.

    1983-09-01

    A survey of 186 soluble lymphocyte proteins for genetic polymorphism was carried out utilizing two-dimensional electrophoresis of /sup 14/C-labeled phytohemagglutinin (PHA)-stimulated human lymphocyte proteins. Nineteen of these proteins exhibited positional variation consistent with independent genetic polymorphism in a primary sample of 28 individuals. Each of these polymorphisms was characterized by quantitative gene-dosage dependence insofar as the heterozygous phenotype expressed approximately 50% of each allelic gene product as was seen in homozygotes. Patterns observed were also identical in monozygotic twins, replicate samples, and replicate gels. The three expected phenotypes (two homozygotes and a heterozygote) were observed in each of 10 of these polymorphisms while the remaining nine had one of the homozygous classes absent. The presence of the three phenotypes, the demonstration of gene-dosage dependence, and our own and previous pedigree analysis of certain of these polymorphisms supports the genetic basis of these variants. Based on this data, the frequency of polymorphic loci for man is: P . 19/186 . .102, and the average heterozygosity is .024. This estimate is approximately 1/3 to 1/2 the rate of polymorphism previously estimated for man in other studies using one-dimensional electrophoresis of isozyme loci. The newly described polymorphisms and others which should be detectable in larger protein surveys with two-dimensional electrophoresis hold promise as genetic markers of the human genome for use in gene mapping and pedigree analyses.

  4. High-throughput and quantitative approaches for measuring circadian rhythms in cyanobacteria using bioluminescence.

    PubMed

    Shultzaberger, Ryan K; Paddock, Mark L; Katsuki, Takeo; Greenspan, Ralph J; Golden, Susan S

    2015-01-01

    The temporal measurement of a bioluminescent reporter has proven to be one of the most powerful tools for characterizing circadian rhythms in the cyanobacterium Synechococcus elongatus. Primarily, two approaches have been used to automate this process: (1) detection of cell culture bioluminescence in 96-well plates by a photomultiplier tube-based plate-cycling luminometer (TopCount Microplate Scintillation and Luminescence Counter, Perkin Elmer) and (2) detection of individual colony bioluminescence by iteratively rotating a Petri dish under a cooled CCD camera using a computer-controlled turntable. Each approach has distinct advantages. The TopCount provides a more quantitative measurement of bioluminescence, enabling the direct comparison of clock output levels among strains. The computer-controlled turntable approach has a shorter set-up time and greater throughput, making it a more powerful phenotypic screening tool. While the latter approach is extremely useful, only a few labs have been able to build such an apparatus because of technical hurdles involved in coordinating and controlling both the camera and the turntable, and in processing the resulting images. This protocol provides instructions on how to construct, use, and process data from a computer-controlled turntable to measure the temporal changes in bioluminescence of individual cyanobacterial colonies. Furthermore, we describe how to prepare samples for use with the TopCount to minimize experimental noise and generate meaningful quantitative measurements of clock output levels for advanced analysis. PMID:25662451

  5. High-throughput and quantitative approaches for measuring circadian rhythms in cyanobacteria using bioluminescence.

    PubMed

    Shultzaberger, Ryan K; Paddock, Mark L; Katsuki, Takeo; Greenspan, Ralph J; Golden, Susan S

    2015-01-01

    The temporal measurement of a bioluminescent reporter has proven to be one of the most powerful tools for characterizing circadian rhythms in the cyanobacterium Synechococcus elongatus. Primarily, two approaches have been used to automate this process: (1) detection of cell culture bioluminescence in 96-well plates by a photomultiplier tube-based plate-cycling luminometer (TopCount Microplate Scintillation and Luminescence Counter, Perkin Elmer) and (2) detection of individual colony bioluminescence by iteratively rotating a Petri dish under a cooled CCD camera using a computer-controlled turntable. Each approach has distinct advantages. The TopCount provides a more quantitative measurement of bioluminescence, enabling the direct comparison of clock output levels among strains. The computer-controlled turntable approach has a shorter set-up time and greater throughput, making it a more powerful phenotypic screening tool. While the latter approach is extremely useful, only a few labs have been able to build such an apparatus because of technical hurdles involved in coordinating and controlling both the camera and the turntable, and in processing the resulting images. This protocol provides instructions on how to construct, use, and process data from a computer-controlled turntable to measure the temporal changes in bioluminescence of individual cyanobacterial colonies. Furthermore, we describe how to prepare samples for use with the TopCount to minimize experimental noise and generate meaningful quantitative measurements of clock output levels for advanced analysis.

  6. High throughput and quantitative approaches for measuring circadian rhythms in cyanobacteria using bioluminescence

    PubMed Central

    Shultzaberger, Ryan K.; Paddock, Mark L.; Katsuki, Takeo; Greenspan, Ralph J.; Golden, Susan S.

    2016-01-01

    The temporal measurement of a bioluminescent reporter has proven to be one of the most powerful tools for characterizing circadian rhythms in the cyanobacterium Synechococcus elongatus. Primarily, two approaches have been used to automate this process: (1) detection of cell culture bioluminescence in 96-well plates by a photomultiplier tube-based plate-cycling luminometer (TopCount Microplate Scintillation and Luminescence Counter, Perkin Elmer) and (2) detection of individual colony bioluminescence by iteratively rotating a Petri dish under a cooled CCD camera using a computer-controlled turntable. Each approach has distinct advantages. The TopCount provides a more quantitative measurement of bioluminescence, enabling the direct comparison of clock output levels among strains. The computer-controlled turntable approach has a shorter set-up time and greater throughput, making it a more powerful phenotypic screening tool. While the latter approach is extremely useful, only a few labs have been able to build such an apparatus because of technical hurdles involved in coordinating and controlling both the camera and the turntable, and in processing the resulting images. This protocol provides instructions on how to construct, use, and process data from a computer-controlled turntable to measure the temporal changes in bioluminescence of individual cyanobacterial colonies. Furthermore, we describe how to prepare samples for use with the TopCount to minimize experimental noise, and generate meaningful quantitative measurements of clock output levels for advanced analysis. PMID:25662451

  7. Quantitative Laughter Detection, Measurement, and Classification-A Critical Survey.

    PubMed

    Cosentino, Sarah; Sessa, Salvatore; Takanishi, Atsuo

    2016-01-01

    The study of human nonverbal social behaviors has taken a more quantitative and computational approach in recent years due to the development of smart interfaces and virtual agents or robots able to interact socially. One of the most interesting nonverbal social behaviors, producing a characteristic vocal signal, is laughing. Laughter is produced in several different situations: in response to external physical, cognitive, or emotional stimuli; to negotiate social interactions; and also, pathologically, as a consequence of neural damage. For this reason, laughter has attracted researchers from many disciplines. A consequence of this multidisciplinarity is the absence of a holistic vision of this complex behavior: the methods of analysis and classification of laughter, as well as the terminology used, are heterogeneous; the findings sometimes contradictory and poorly documented. This survey aims at collecting and presenting objective measurement methods and results from a variety of different studies in different fields, to contribute to build a unified model and taxonomy of laughter. This could be successfully used for advances in several fields, from artificial intelligence and human-robot interaction to medicine and psychiatry. PMID:26887012

  8. Radioactive Contraband Detection: A Bayesian Approach

    SciTech Connect

    Candy, J; Breitfeller, E; Guidry, B; Manatt, D; Sale, K; Chambers, D; Axelrod, M; Meyer, A

    2009-03-16

    Radionuclide emissions from nuclear contraband challenge both detection and measurement technologies to capture and record each event. The development of a sequential Bayesian processor incorporating both the physics of gamma-ray emissions and the measurement of photon energies offers a physics-based approach to attack this challenging problem. It is shown that a 'physics-based' structure can be used to develop an effective detection technique, but also motivates the implementation of this approach using or particle filters to enhance and extract the required information.

  9. A gold nanoparticle-based semi-quantitative and quantitative ultrasensitive paper sensor for the detection of twenty mycotoxins

    NASA Astrophysics Data System (ADS)

    Kong, Dezhao; Liu, Liqiang; Song, Shanshan; Suryoprabowo, Steven; Li, Aike; Kuang, Hua; Wang, Libing; Xu, Chuanlai

    2016-02-01

    A semi-quantitative and quantitative multi-immunochromatographic (ICA) strip detection assay was developed for the simultaneous detection of twenty types of mycotoxins from five classes, including zearalenones (ZEAs), deoxynivalenols (DONs), T-2 toxins (T-2s), aflatoxins (AFs), and fumonisins (FBs), in cereal food samples. Sensitive and specific monoclonal antibodies were selected for this assay. The semi-quantitative results were obtained within 20 min by the naked eye, with visual limits of detection for ZEAs, DONs, T-2s, AFs and FBs of 0.1-0.5, 2.5-250, 0.5-1, 0.25-1 and 2.5-10 μg kg-1, and cut-off values of 0.25-1, 5-500, 1-10, 0.5-2.5 and 5-25 μg kg-1, respectively. The quantitative results were obtained using a hand-held strip scan reader, with the calculated limits of detection for ZEAs, DONs, T-2s, AFs and FBs of 0.04-0.17, 0.06-49, 0.15-0.22, 0.056-0.49 and 0.53-1.05 μg kg-1, respectively. The analytical results of spiked samples were in accordance with the accurate content in the simultaneous detection analysis. This newly developed ICA strip assay is suitable for the on-site detection and rapid initial screening of mycotoxins in cereal samples, facilitating both semi-quantitative and quantitative determination.A semi-quantitative and quantitative multi-immunochromatographic (ICA) strip detection assay was developed for the simultaneous detection of twenty types of mycotoxins from five classes, including zearalenones (ZEAs), deoxynivalenols (DONs), T-2 toxins (T-2s), aflatoxins (AFs), and fumonisins (FBs), in cereal food samples. Sensitive and specific monoclonal antibodies were selected for this assay. The semi-quantitative results were obtained within 20 min by the naked eye, with visual limits of detection for ZEAs, DONs, T-2s, AFs and FBs of 0.1-0.5, 2.5-250, 0.5-1, 0.25-1 and 2.5-10 μg kg-1, and cut-off values of 0.25-1, 5-500, 1-10, 0.5-2.5 and 5-25 μg kg-1, respectively. The quantitative results were obtained using a hand-held strip scan

  10. A Novel Quantitative Approach to Concept Analysis: The Internomological Network

    PubMed Central

    Cook, Paul F.; Larsen, Kai R.; Sakraida, Teresa J.; Pedro, Leli

    2012-01-01

    Background When a construct such as patients’ transition to self-management of chronic illness is studied by researchers across multiple disciplines, the meaning of key terms can become confused. This results from inherent problems in language where a term can have multiple meanings (polysemy) and different words can mean the same thing (synonymy). Objectives To test a novel quantitative method for clarifying the meaning of constructs by examining the similarity of published contexts in which they are used. Method Published terms related to the concept transition to self-management of chronic illness were analyzed using the internomological network (INN), a type of latent semantic analysis to calculate the mathematical relationships between constructs based on the contexts in which researchers use each term. This novel approach was tested by comparing results to those from concept analysis, a best-practice qualitative approach to clarifying meanings of terms. By comparing results of the two methods, the best synonyms of transition to self-management, as well as key antecedent, attribute, and consequence terms, were identified. Results Results from INN analysis were consistent with those from concept analysis. The potential synonyms self-management, transition, and adaptation had the greatest utility. Adaptation was the clearest overall synonym, but had lower cross-disciplinary use. The terms coping and readiness had more circumscribed meanings. The INN analysis confirmed key features of transition to self-management, and suggested related concepts not found by the previous review. Discussion The INN analysis is a promising novel methodology that allows researchers to quantify the semantic relationships between constructs. The method works across disciplinary boundaries, and may help to integrate the diverse literature on self-management of chronic illness. PMID:22592387

  11. Quantitative proteomic approaches in biomarker discovery of inflammatory bowel disease.

    PubMed

    Han, Na-Young; Kim, Eun Hee; Choi, Joon; Lee, Hookeun; Hahm, Ki-Baik

    2012-10-01

    Proteomics offers considerable opportunities for either enhancing our biological understanding or discovering biomarkers, blood and biopsied specimen-based proteomic approaches, provide reproducible and quantitative tools that can complement clinical assessments and aid clinicians in the diagnosis and treatment of inflammatory bowel disease (IBD). Sometimes a differential diagnosis of Crohn's disease (CD) and ulcerative colitis (UC) and the prediction of treatment response can be deduced by finding meaningful biomarkers, for which the central platform for proteomics is tandem mass spectrometry (MS/MS). A range of workflows are available for protein (or peptide) separation prior to MS/MS as well as bioinformatics analysis to achieve protein identification, for which two-dimensional electrophoresis (2-DE) and subsequent mass spectrometry (MS), liquid chromatography-MS, difference gel electrophoresis following 2-DE, isobaric tags for relative and absolute quantification (iTRAQ), stable isotope labeling by amino acids and label-free quantification are under development. In this article, the current status and perspective of these advanced proteomic technologies are introduced, with examples of recent biomarkers focused on the diagnosis, treatment response, prognosis of IBD, and even colitis-associated carcinogenesis in both animal models and human patients. PMID:22988922

  12. Active contour approach for accurate quantitative airway analysis

    NASA Astrophysics Data System (ADS)

    Odry, Benjamin L.; Kiraly, Atilla P.; Slabaugh, Greg G.; Novak, Carol L.; Naidich, David P.; Lerallut, Jean-Francois

    2008-03-01

    Chronic airway disease causes structural changes in the lungs including peribronchial thickening and airway dilatation. Multi-detector computed tomography (CT) yields detailed near-isotropic images of the lungs, and thus the potential to obtain quantitative measurements of lumen diameter and airway wall thickness. Such measurements would allow standardized assessment, and physicians to diagnose and locate airway abnormalities, adapt treatment, and monitor progress over time. However, due to the sheer number of airways per patient, systematic analysis is infeasible in routine clinical practice without automation. We have developed an automated and real-time method based on active contours to estimate both airway lumen and wall dimensions; the method does not require manual contour initialization but only a starting point on the targeted airway. While the lumen contour segmentation is purely region-based, the estimation of the outer diameter considers the inner wall segmentation as well as local intensity variation, in order anticipate the presence of nearby arteries and exclude them. These properties make the method more robust than the Full-Width Half Maximum (FWHM) approach. Results are demonstrated on a phantom dataset with known dimensions and on a human dataset where the automated measurements are compared against two human operators. The average error on the phantom measurements was 0.10mm and 0.14mm for inner and outer diameters, showing sub-voxel accuracy. Similarly, the mean variation from the average manual measurement was 0.14mm and 0.18mm for inner and outer diameters respectively.

  13. Quantitative detection of settled dust over green canopy

    NASA Astrophysics Data System (ADS)

    Brook, Anna

    2016-04-01

    The main task of environmental and geoscience applications are efficient and accurate quantitative classification of earth surfaces and spatial phenomena. In the past decade, there has been a significant interest in employing hyperspectral unmixing in order to retrieve accurate quantitative information latent in hyperspectral imagery data. Recently, the ground-truth and laboratory measured spectral signatures promoted by advanced algorithms are proposed as a new path toward solving the unmixing problem of hyperspectral imagery in semi-supervised fashion. This paper suggests that the sensitivity of sparse unmixing techniques provides an ideal approach to extract and identify dust settled over/upon green vegetation canopy using hyperspectral airborne data. Atmospheric dust transports a variety of chemicals, some of which pose a risk to the ecosystem and human health (Kaskaoutis, et al., 2008). Many studies deal with the impact of dust on particulate matter (PM) and atmospheric pollution. Considering the potential impact of industrial pollutants, one of the most important considerations is the fact that suspended PM can have both a physical and a chemical impact on plants, soils, and water bodies. Not only can the particles covering surfaces cause physical distortion, but particles of diverse origin and different chemistries can also serve as chemical stressors and cause irreversible damage. Sediment dust load in an indoor environment can be spectrally assessed using reflectance spectroscopy (Chudnovsky and Ben-Dor, 2009). Small amounts of particulate pollution that may carry a signature of a forthcoming environmental hazard are of key interest when considering the effects of pollution. According to the most basic distribution dynamics, dust consists of suspended particulate matter in a fine state of subdivision that are raised and carried by wind. In this context, it is increasingly important to first, understand the distribution dynamics of pollutants, and

  14. Quantitative Analysis of Mutant Subclones in Chronic Myeloid Leukemia: Comparison of Different Methodological Approaches.

    PubMed

    Preuner, Sandra; Barna, Agnes; Frommlet, Florian; Czurda, Stefan; Konstantin, Byrgazov; Alikian, Mary; Machova Polakova, Katerina; Sacha, Tomasz; Richter, Johan; Lion, Thomas; Gabriel, Christian

    2016-01-01

    Identification and quantitative monitoring of mutant BCR-ABL1 subclones displaying resistance to tyrosine kinase inhibitors (TKIs) have become important tasks in patients with Ph-positive leukemias. Different technologies have been established for patient screening. Various next-generation sequencing (NGS) platforms facilitating sensitive detection and quantitative monitoring of mutations in the ABL1-kinase domain (KD) have been introduced recently, and are expected to become the preferred technology in the future. However, broad clinical implementation of NGS methods has been hampered by the limited accessibility at different centers and the current costs of analysis which may not be regarded as readily affordable for routine diagnostic monitoring. It is therefore of interest to determine whether NGS platforms can be adequately substituted by other methodological approaches. We have tested three different techniques including pyrosequencing, LD (ligation-dependent)-PCR and NGS in a series of peripheral blood specimens from chronic myeloid leukemia (CML) patients carrying single or multiple mutations in the BCR-ABL1 KD. The proliferation kinetics of mutant subclones in serial specimens obtained during the course of TKI-treatment revealed similar profiles via all technical approaches, but individual specimens showed statistically significant differences between NGS and the other methods tested. The observations indicate that different approaches to detection and quantification of mutant subclones may be applicable for the monitoring of clonal kinetics, but careful calibration of each method is required for accurate size assessment of mutant subclones at individual time points. PMID:27136541

  15. Quantitative Analysis of Mutant Subclones in Chronic Myeloid Leukemia: Comparison of Different Methodological Approaches

    PubMed Central

    Preuner, Sandra; Barna, Agnes; Frommlet, Florian; Czurda, Stefan; Konstantin, Byrgazov; Alikian, Mary; Machova Polakova, Katerina; Sacha, Tomasz; Richter, Johan; Lion, Thomas; Gabriel, Christian

    2016-01-01

    Identification and quantitative monitoring of mutant BCR-ABL1 subclones displaying resistance to tyrosine kinase inhibitors (TKIs) have become important tasks in patients with Ph-positive leukemias. Different technologies have been established for patient screening. Various next-generation sequencing (NGS) platforms facilitating sensitive detection and quantitative monitoring of mutations in the ABL1-kinase domain (KD) have been introduced recently, and are expected to become the preferred technology in the future. However, broad clinical implementation of NGS methods has been hampered by the limited accessibility at different centers and the current costs of analysis which may not be regarded as readily affordable for routine diagnostic monitoring. It is therefore of interest to determine whether NGS platforms can be adequately substituted by other methodological approaches. We have tested three different techniques including pyrosequencing, LD (ligation-dependent)-PCR and NGS in a series of peripheral blood specimens from chronic myeloid leukemia (CML) patients carrying single or multiple mutations in the BCR-ABL1 KD. The proliferation kinetics of mutant subclones in serial specimens obtained during the course of TKI-treatment revealed similar profiles via all technical approaches, but individual specimens showed statistically significant differences between NGS and the other methods tested. The observations indicate that different approaches to detection and quantification of mutant subclones may be applicable for the monitoring of clonal kinetics, but careful calibration of each method is required for accurate size assessment of mutant subclones at individual time points. PMID:27136541

  16. Quantitative analysis of in situ hybridization methods for the detection of actin gene expression.

    PubMed Central

    Lawrence, J B; Singer, R H

    1985-01-01

    We have implemented an efficient, quantitative approach for the optimization of in situ hybridization using double-stranded recombinant DNA probes. The model system studied was actin mRNA expression in chicken embryonic muscle cultures. Actin and control (pBR322) probes were nick-translated with p32 labeled nucleotides, hybridized to cells grown on coverslips, and quantitated in a scintillation counter. Cellular RNA retention was monitored via the incorporation of H3-Uridine into RNA prior to cell fixation. Over a thousand samples were analyzed, and among the technical variables examined were the fixation protocol, proteolytic cell pretreatment, the time course of hybridization, saturation kinetics, hybridization efficiency, and effect of probe size on hybridization and network formation. Results have allowed us to develop a reproducible in situ hybridization methodology which is simpler and less destructive to cellular RNA and morphology than other protocols. Moreover, this technique is highly sensitive and efficient in detection of cellular RNAs. Lastly, the rapid quantitative approach used for this analysis is valuable in itself as a potential alternative to filter or solution hybridizations. Images PMID:3889842

  17. Hyperspectral target detection using sequential approach

    NASA Astrophysics Data System (ADS)

    Haskett, Hanna T.; Sood, Arun K.; Habib, Mohammad K.

    1999-08-01

    This paper describes an automatic target detection algorithm based on the sequential multi-stage approach. Each stage of the algorithm uses more spectral bands than the previous stage. To ensure high probability of detection and low false alarm rate, Chebyshev's inequality test is applied. The sequential approach enables a significant reduction in computational time of a hyperspectral detection system. The Forest Radiance I database collected with the HYDICE hyperspectral sensor at the U.S. Army Proving Ground in Aberdeen, Maryland is utilized. Scenarios include targets in the open, with footprints of 1 m and different times of day. The total area coverage and the number of targets used in this evaluation are approximately 6 km2 and 126, respectively.

  18. Nonlinear sensors: an approach to the residence time detection strategy.

    PubMed

    Dari, A; Bosi, L; Gammaitoni, L

    2010-01-01

    The monitoring of the residence time difference in bistable sensors has been recently proposed as a valid scheme for improving the detection capabilities of sensors as diverse as fluxgate magnetometers, ferroelectric sensors and mechanical sensors. In this paper we propose an approach to the residence time based detection strategy based on the measurement of the slope m of the sensor output integral. We demonstrate that such a method, far from degrading the detection performances can provide an easier way to realize fast and reliable sensors without the computationally demanding task related with the computation of the residence time difference. We introduce the receiver operating characteristic curve as a quantitative estimator for the comparison of the two methods and show that the detector performances increase with increasing the periodic bias amplitude A up to a maximum value. This condition has potentially relevant consequences in the future detectors design. PMID:20365331

  19. High sensitivity detection of active botulinum neurotoxin by glyco-quantitative polymerase chain-reaction.

    PubMed

    Kwon, Seok Joon; Jeong, Eun Ji; Yoo, Yung Choon; Cai, Chao; Yang, Gi-Hyeok; Lee, Jae Chul; Dordick, Jonathan S; Linhardt, Robert J; Lee, Kyung Bok

    2014-03-01

    The sensitive detection of highly toxic botulinum neurotoxin (BoNT) from Clostridium botulinum is of critical importance because it causes human illnesses if foodborne or introduced in wounds and as an iatrogenic substance. Moreover, it has been recently considered a possible biological warfare agent. Over the past decade, significant progress has been made in BoNT detection technologies, including mouse lethality assays, enzyme-linked immunosorbent assays, and endopeptidase assays and by mass spectrometry. Critical assay requirements, including rapid assay, active toxin detection, sensitive and accurate detection, still remain challenging. Here, we present a novel method to detect active BoNTs using a Glyco-quantitative polymerase chain-reaction (qPCR) approach. Sialyllactose, which interacts with the binding-domain of BoNTs, is incorporated into a sialyllactose-DNA conjugate as a binding-probe for active BoNT and recovered through BoNT-immunoprecipitation. Glyco-qPCR analysis of the bound sialyllactose-DNA is then used to detect low attomolar concentrations of BoNT and attomolar to femtomolar concentrations of BoNT in honey, the most common foodborne source of infant botulism.

  20. Anomalous human behavior detection: an adaptive approach

    NASA Astrophysics Data System (ADS)

    van Leeuwen, Coen; Halma, Arvid; Schutte, Klamer

    2013-05-01

    Detection of anomalies (outliers or abnormal instances) is an important element in a range of applications such as fault, fraud, suspicious behavior detection and knowledge discovery. In this article we propose a new method for anomaly detection and performed tested its ability to detect anomalous behavior in videos from DARPA's Mind's Eye program, containing a variety of human activities. In this semi-unsupervised task a set of normal instances is provided for training, after which unknown abnormal behavior has to be detected in a test set. The features extracted from the video data have high dimensionality, are sparse and inhomogeneously distributed in the feature space making it a challenging task. Given these characteristics a distance-based method is preferred, but choosing a threshold to classify instances as (ab)normal is non-trivial. Our novel aproach, the Adaptive Outlier Distance (AOD) is able to detect outliers in these conditions based on local distance ratios. The underlying assumption is that the local maximum distance between labeled examples is a good indicator of the variation in that neighborhood, and therefore a local threshold will result in more robust outlier detection. We compare our method to existing state-of-art methods such as the Local Outlier Factor (LOF) and the Local Distance-based Outlier Factor (LDOF). The results of the experiments show that our novel approach improves the quality of the anomaly detection.

  1. Fluorescent microscopy approaches of quantitative soil microbial analysis

    NASA Astrophysics Data System (ADS)

    Ivanov, Konstantin; Polyanskaya, Lubov

    2015-04-01

    Classical fluorescent microscopy method was used during the last decades in various microbiological studies of terrestrial ecosystems. The method provides representative results and simple application which is allow to use it both as routine part of amplitudinous research and in small-scaled laboratories. Furthermore, depending on research targets a lot of modifications of fluorescent microscopy method were established. Combination and comparison of several approaches is an opportunity of quantitative estimation of microbial community in soil. The first analytical part of the study was dedicated to soil bacterial density estimation by fluorescent microscopy in dynamic of several 30-days experiments. The purpose of research was estimation of changes in soil bacterial community on the different soil horizons under aerobic and anaerobic conditions with adding nutrients in two experimental sets: cellulose and chitin. Was modified the nalidixic acid method for inhibition of DNA division of gram-negative bacteria, and the method provides the quantification of this bacterial group by fluorescent microscopy. Established approach allowed to estimate 3-4 times more cells of gram-negative bacteria in soil. The functions of actinomyces in soil polymer destruction are traditionally considered as dominant in comparison to gram-negative bacterial group. However, quantification of gram-negative bacteria in chernozem and peatland provides underestimation of classical notion for this bacterial group. Chitin introduction had no positive effect to gram-negative bacterial population density changes in chernozem but concurrently this nutrient provided the fast growing dynamics at the first 3 days of experiment both under aerobic and anaerobic conditions. This is confirming chitinolytic activity of gram-negative bacteria in soil organic matter decomposition. At the next part of research modified method for soil gram-negative bacteria quantification was compared to fluorescent in situ

  2. Single-Plex Quantitative Assays for the Detection and Quantification of Most Pneumococcal Serotypes

    PubMed Central

    Chochua, Sopio; Satzke, Catherine; Dunne, Eileen M.; Mulholland, Kim; Klugman, Keith P.

    2015-01-01

    Streptococcus pneumoniae globally kills more children than any other infectious disease every year. A prerequisite for pneumococcal disease and transmission is colonization of the nasopharynx. While the introduction of pneumococcal conjugate vaccines has reduced the burden of pneumococcal disease, understanding the impact of vaccination on nasopharyngeal colonization has been hampered by the lack of sensitive quantitative methods for the detection of >90 known S. pneumoniae serotypes. In this work, we developed 27 new quantitative (q)PCR reactions and optimized 26 for a total of 53 qPCR reactions targeting pneumococcal serotypes or serogroups, including all vaccine types. Reactions proved to be target-specific with a limit of detection of 2 genome equivalents per reaction. Given the number of probes required for these assays and their unknown shelf-life, the stability of cryopreserved reagents was evaluated. Our studies demonstrate that two-year cryopreserved probes had similar limit of detection as freshly-diluted probes. Moreover, efficiency and limit of detection of 1-month cryopreserved, ready-to-use, qPCR reaction mixtures were similar to those of freshly prepared mixtures. Using these reactions, our proof-of-concept studies utilizing nasopharyngeal samples (N=30) collected from young children detected samples containing ≥2 serotypes/serogroups. Samples colonized by multiple serotypes/serogroups always had a serotype that contributes at least 50% of the pneumococcal load. In addition, a molecular approach called S6-q(PCR)2 was developed and proven to individually detect and quantify epidemiologically-important serogroup 6 strains including 6A, 6B, 6C and 6D. This technology will be useful for epidemiological studies, diagnostic platforms and to study the pneumobiome. PMID:25798884

  3. Single-plex quantitative assays for the detection and quantification of most pneumococcal serotypes.

    PubMed

    Sakai, Fuminori; Chochua, Sopio; Satzke, Catherine; Dunne, Eileen M; Mulholland, Kim; Klugman, Keith P; Vidal, Jorge E

    2015-01-01

    Streptococcus pneumoniae globally kills more children than any other infectious disease every year. A prerequisite for pneumococcal disease and transmission is colonization of the nasopharynx. While the introduction of pneumococcal conjugate vaccines has reduced the burden of pneumococcal disease, understanding the impact of vaccination on nasopharyngeal colonization has been hampered by the lack of sensitive quantitative methods for the detection of >90 known S. pneumoniae serotypes. In this work, we developed 27 new quantitative (q)PCR reactions and optimized 26 for a total of 53 qPCR reactions targeting pneumococcal serotypes or serogroups, including all vaccine types. Reactions proved to be target-specific with a limit of detection of 2 genome equivalents per reaction. Given the number of probes required for these assays and their unknown shelf-life, the stability of cryopreserved reagents was evaluated. Our studies demonstrate that two-year cryopreserved probes had similar limit of detection as freshly-diluted probes. Moreover, efficiency and limit of detection of 1-month cryopreserved, ready-to-use, qPCR reaction mixtures were similar to those of freshly prepared mixtures. Using these reactions, our proof-of-concept studies utilizing nasopharyngeal samples (N=30) collected from young children detected samples containing ≥2 serotypes/serogroups. Samples colonized by multiple serotypes/serogroups always had a serotype that contributes at least 50% of the pneumococcal load. In addition, a molecular approach called S6-q(PCR)2 was developed and proven to individually detect and quantify epidemiologically-important serogroup 6 strains including 6A, 6B, 6C and 6D. This technology will be useful for epidemiological studies, diagnostic platforms and to study the pneumobiome.

  4. Quantitative asymmetric-detection time-stretch optical microscopy (Q-ATOM) for ultrafast quantitative phase imaging flow cytometry (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Lau, Andy K. S.; Tang, Anson H. L.; Chung, Bob M. F.; Tsang, Kwok Yeung; Chan, Antony C. S.; Wei, Xiaoming; Wong, Kenneth K.; Lam, Edmund Y.; Cheah, Kathryn S. E.; Shum, Anderson H. C.; Tsia, Kevin K.

    2016-03-01

    Based on the interferometric or holographic approaches, recent QPM techniques provide quantitative-phase information, e.g cell volume, dry mass and optical scattering properties for label-free cellular physical phenotyping. These approaches generally rely on iterative phase-retrieval algorithms to obtain quantitative-phase information, which are computationally intensive. Moreover, current QPM techniques can only offer limited image acquisition rate by using CMOS/CCD image sensors, these two limitations hinder QPM for high-throughput quantitative image-based single-cell analysis in real-time. To this end, we demonstrate an interferometry-free quantitative phase microscopy developed on a new generation of time-stretch microscopy, asymmetric-detection time-stretch optical microscopy (ATOM), which is coined quantitative ATOM (Q-ATOM) - featuring an unprecedented cell measurement throughput together with the assorted intrinsic optical phenotypes (e.g. angular light scattering profile) and the derived physical properties of the cells (e.g. cell size, dry mass density etc.). Based on a similar concept to Schlieren imaging, Q-ATOM retrieves quantitative-phase information through multiple off-axis light-beam detection at a line-scan rate of <10 MHz - a speed unachievable by any existing QPM techniques. Phase retrieval in Q-ATOM relies on a non-iterative method, significantly reducing the computational complexity of the technique. It is a particularly important feature which facilitates real-time continuous label-free single-cell analysis in Q-ATOM. With the use of a non-interferometric configuration, we demonstrate ultrafast Q-ATOM of mouse chondrocytes and hypertrophic chondrocytes in ultrafast microfluidic flow with sub-cellular resolution at an imaging throughput equivalent to ~100,000 cells/sec without image blur. This technique shows a great potential for ultrahigh throughput label-free image-based single-cell biophysical phentotyping.

  5. Simultaneous imaging of locus coeruleus and substantia nigra with a quantitative neuromelanin MRI approach.

    PubMed

    Chen, Xiangchuan; Huddleston, Daniel E; Langley, Jason; Ahn, Sinyeob; Barnum, Christopher J; Factor, Stewart A; Levey, Allan I; Hu, Xiaoping

    2014-12-01

    Quantitative MRI of neuromelanin (NM) containing structures (referred to as NM-MRI) in the brainstem, namely the locus coeruleus (LC) and substantia nigra (SN), may assist with the early detection of Parkinson's disease (PD) and Alzheimer's disease (AD) as well as differential diagnosis in the early disease stages. In this study, two gradient echo (GRE) sequences with magnetization transfer contrast (MTC) preparation pulses were developed to simultaneously image the LC and SN. This has been a challenge with NM-MRI techniques used in previous studies due to the relatively high specific absorption rate (SAR) induced by these techniques. In addition, a semi-automated quantitative analysis scheme was applied to estimate volumes and contrast-to-noise ratios (CNR) of the LC and SN based on segmentation of both structures. Compared to a T1-weighted turbo spin echo (TSE) sequence typically used for simultaneous imaging of the LC and SN, the two GRE-MTC sequences exhibited improved performance in terms of higher sensitivity (in CNR) in imaging the SN and lower SAR during the scans. A multiple-measurement protocol was adopted as well so that motion degraded measurements could be removed and artifacts associated with motion could be corrected. The present approach has demonstrated advantages in image acquisition (lower SAR and higher sensitivity), image pre-processing (with motion correction) and quantitative image analysis (segmentation-based estimation of volume and CNR) when compared with existing NM-MRI approaches. This approach has potential for detection and monitoring of neurodegeneration in LC and SN in disease states including AD and PD.

  6. A morphogenetic quantitative approach to species-typical behaviour.

    PubMed

    Irsigler, F J

    1991-01-01

    The numerical ratio of the allocortical, late-myelinized infra-temporal sector (Flechsig) to the neocortical convexity of the temporal lobe in the human brain is proposed as a quantitative invariant of the human species, called "Neocorticalisation Index NI". Its morphogenetic nature derives from (1) the dual, that is the reptilian-palaeomammalian origin of the primate cortex (Kappers 1909, Vogt 1910. Kuhlenbeck 1924, Filimonoff 1947, Northcutt 1967, Stephan 1975). The two layers differentiate at independent rates forming a transitional belt, called "periallocortex" (limbic) and "mesocortex" (insular). They approach the most the general cyto- and myeloarchitectonic scheme of Brodmann (1909) and C. and O. Vogt (1919), and in the human brain are the latest to reach maturity (Flechsig 1920, Kahle 1969, Sanides 1975); (2) measurements in fossilized human brain endocasts (Taung, Rhodesian man) reported earlier (Irsigler 1984a and b); (3) clinico-pathological evidence gained from cases of the Pick-type of presenile dementia confirming the anthropogenetic character of the frontotemporal "basal forebrain" (Spatz and his school since 1937, Lüers 1947, Jakob 1979). The latter is considered to be in the mammals the major counterpart of the hormonally and reciprocally controlled reptilian forebrain (Schepers 1948, Sanides 1970, MacLean 1978). The results of NI measurements in 128 human brains are tabulated and interpreted in the light of species-typical behaviour. It turns out that the left hemisphere is the one possessing the greater share of allocorticity showing precocious development in the non-European (Negro) infant. There are gender-related differences of the NI parameters. The Indo-Europeans (Caucasians) undergo, in their ontogeny, a kind of "secondary" allocorticalisation in both hemispheres which, hypothetically, may be correlated with the "substantializing" (reifying) property inherent in their languages.

  7. Automatic detection and quantitative analysis of cells in the mouse primary motor cortex

    NASA Astrophysics Data System (ADS)

    Meng, Yunlong; He, Yong; Wu, Jingpeng; Chen, Shangbin; Li, Anan; Gong, Hui

    2014-09-01

    Neuronal cells play very important role on metabolism regulation and mechanism control, so cell number is a fundamental determinant of brain function. Combined suitable cell-labeling approaches with recently proposed three-dimensional optical imaging techniques, whole mouse brain coronal sections can be acquired with 1-μm voxel resolution. We have developed a completely automatic pipeline to perform cell centroids detection, and provided three-dimensional quantitative information of cells in the primary motor cortex of C57BL/6 mouse. It involves four principal steps: i) preprocessing; ii) image binarization; iii) cell centroids extraction and contour segmentation; iv) laminar density estimation. Investigations on the presented method reveal promising detection accuracy in terms of recall and precision, with average recall rate 92.1% and average precision rate 86.2%. We also analyze laminar density distribution of cells from pial surface to corpus callosum from the output vectorizations of detected cell centroids in mouse primary motor cortex, and find significant cellular density distribution variations in different layers. This automatic cell centroids detection approach will be beneficial for fast cell-counting and accurate density estimation, as time-consuming and error-prone manual identification is avoided.

  8. Using multiple PCR and CE with chemiluminescence detection for simultaneous qualitative and quantitative analysis of genetically modified organism.

    PubMed

    Guo, Longhua; Qiu, Bin; Chi, Yuwu; Chen, Guonan

    2008-09-01

    In this paper, an ultrasensitive CE-CL detection system coupled with a novel double-on-column coaxial flow detection interface was developed for the detection of PCR products. A reliable procedure based on this system had been demonstrated for qualitative and quantitative analysis of genetically modified organism-the detection of Roundup Ready Soy (RRS) samples was presented as an example. The promoter, terminator, function and two reference genes of RRS were amplified with multiplex PCR simultaneously. After that, the multiplex PCR products were labeled with acridinium ester at the 5'-terminal through an amino modification and then analyzed by the proposed CE-CL system. Reproducibility of analysis times and peak heights for the CE-CL analysis were determined to be better than 0.91 and 3.07% (RSD, n=15), respectively, for three consecutive days. It was shown that this method could accurately and qualitatively detect RRS standards and the simulative samples. The evaluation in terms of quantitative analysis of RRS provided by this new method was confirmed by comparing our assay results with those of the standard real-time quantitative PCR (RT-QPCR) using SYBR Green I dyes. The results showed a good coherence between the two methods. This approach demonstrated the possibility for accurate qualitative and quantitative detection of GM plants in a single run.

  9. Column precipitation chromatography: an approach to quantitative analysis of eigencolloids.

    PubMed

    Breynaert, E; Maes, A

    2005-08-01

    A new column precipitation chromatography (CPC) technique, capable of quantitatively measuring technetium eigencolloids in aqueous solutions, is presented. The CPC technique is based on the destabilization and precipitation of eigencolloids by polycations in a confined matrix. Tc(IV) colloids can be quantitatively determined from their precipitation onto the CPC column (separation step) and their subsequent elution upon oxidation to pertechnetate by peroxide (elution step). A clean-bed particle removal model was used to explain the experimental results. PMID:16053321

  10. Quantitative multiplex detection of biomarkers on a waveguide-based biosensor using quantum dots

    SciTech Connect

    Xie, Hongzhi; Mukundan, Harshini; Martinez, Jennifer S; Swanson, Basil I; Anderson, Aaron S; Grace, Kevin

    2009-01-01

    imaging of live cells using QD-bioconjugates [Jaiswal 2003]. Gao [Gao 2004] and So [So 2006] have used QDs as probes for in-vivo cancer targeting and imaging. Medintz et al. reported self-assembled QD-based biosensors for detection of analytes based on energy transfer [Medintz 2003]. Others have developed an approach for multiplex optical encoding of biomolecules using QDs [Han 2001]. Immunoassays have also benefited from the advantages of QDs. Recently, dihydrolipoic acid (DHLA) capped-QDs have been attached to antibodies and used as fluorescence reporters in plate-based multiplex immunoassays [Goodman 2004]. However, DHLA-QDs are associated with low quantum efficiency and are unstable at neutral pH. These problems limit the application of this technology to the sensitive detection of biomolecules, especially in complex biological samples. Thus, the development of a rapid, sensitive, quantitative, and specific multiplex platform for the detection of biomarkers in difficult samples remains an elusive target. The goal stated above has applications in many fields including medical diagnostics, biological research, and threat reduction. The current decade alone has seen the development of a need to rapidly and accurately detect potential biological warfare agents. For example, current methods for the detection of anthrax are grossly inadequate for a variety of reasons including long incubation time (5 days from time of exposure to onset of symptoms) and non-specific ('flu-like') symptoms. When five employees of the United State Senate were exposed to B. anthracis in the mail (2001), only one patient had a confirmed diagnosis before death. Since then, sandwich immunoassays using both colorimetric and fluorescence detectors have been developed for key components of the anthrax lethal toxin, namely protective antigen (PA), lethal factor (LF), and the edema factor [Mourez 2001]. While these platforms were successful in assays against anthrax toxins, the sensitivity was poor

  11. A quantitative strategy to detect changes in accessibility of protein regions to chemical modification on heterodimerization.

    PubMed

    Dreger, Mathias; Leung, Bo Wah; Brownlee, George G; Deng, Tao

    2009-07-01

    We describe a method for studying quantitative changes in accessibility of surface lysine residues of the PB1 subunit of the influenza RNA polymerase as a result of association with the PA subunit to form a PB1-PA heterodimer. Our method combines two established methods: (i) the chemical modification of surface lysine residues of native proteins by N-hydroxysuccinimidobiotin (NHS-biotin) and (ii) the stable isotope labeling of amino acids in cell culture (SILAC) followed by tryptic digestion and mass spectrometry. By linking the chemical modification with the SILAC methodology for the first time, we obtain quantitative data on chemical modification allowing subtle changes in accessibility to be described. Five regions in the PB1 monomer showed altered reactivity to NHS-biotin when compared with the [PB1-PA] heterodimer. Mutational analysis of residues in two such regions-at K265 and K481 of PB1, which were about three- and twofold, respectively, less accessible to biotinylation in the PB1-PA heterodimer compared with the PB1 monomer, demonstrated that both K265 and K481 were crucial for polymerase function. This novel assay of quantitative profiling of biotinylation patterns (Q-POP assay) highlights likely conformational changes at important functional sites, as observed here for PB1, and may provide information on protein-protein interaction interfaces. The Q-POP assay should be a generally applicable approach and may detect novel functional sites suitable for targeting by drugs. PMID:19517532

  12. A quantitative strategy to detect changes in accessibility of protein regions to chemical modification on heterodimerization

    PubMed Central

    Dreger, Mathias; Leung, Bo Wah; Brownlee, George G; Deng, Tao

    2009-01-01

    We describe a method for studying quantitative changes in accessibility of surface lysine residues of the PB1 subunit of the influenza RNA polymerase as a result of association with the PA subunit to form a PB1-PA heterodimer. Our method combines two established methods: (i) the chemical modification of surface lysine residues of native proteins by N-hydroxysuccinimidobiotin (NHS-biotin) and (ii) the stable isotope labeling of amino acids in cell culture (SILAC) followed by tryptic digestion and mass spectrometry. By linking the chemical modification with the SILAC methodology for the first time, we obtain quantitative data on chemical modification allowing subtle changes in accessibility to be described. Five regions in the PB1 monomer showed altered reactivity to NHS-biotin when compared with the [PB1-PA] heterodimer. Mutational analysis of residues in two such regions—at K265 and K481 of PB1, which were about three- and twofold, respectively, less accessible to biotinylation in the PB1-PA heterodimer compared with the PB1 monomer, demonstrated that both K265 and K481 were crucial for polymerase function. This novel assay of quantitative profiling of biotinylation patterns (Q-POP assay) highlights likely conformational changes at important functional sites, as observed here for PB1, and may provide information on protein–protein interaction interfaces. The Q-POP assay should be a generally applicable approach and may detect novel functional sites suitable for targeting by drugs. PMID:19517532

  13. Quantitation of dissolved gas content in emulsions and in blood using mass spectrometric detection.

    PubMed

    Grimley, Everett; Turner, Nicole; Newell, Clayton; Simpkins, Cuthbert; Rodriguez, Juan

    2011-06-01

    Quantitation of dissolved gases in blood or in other biological media is essential for understanding the dynamics of metabolic processes. Current detection techniques, while enabling rapid and convenient assessment of dissolved gases, provide only direct information on the partial pressure of gases dissolved in the aqueous fraction of the fluid. The more relevant quantity known as gas content, which refers to the total amount of the gas in all fractions of the sample, can be inferred from those partial pressures, but only indirectly through mathematical modeling. Here we describe a simple mass spectrometric technique for rapid and direct quantitation of gas content for a wide range of gases. The technique is based on a mass spectrometer detector that continuously monitors gases that are rapidly extracted from samples injected into a purge vessel. The accuracy and sample processing speed of the system is demonstrated with experiments that reproduce within minutes literature values for the solubility of various gases in water. The capability of the technique is further demonstrated through accurate determination of O(2) content in a lipid emulsion and in whole blood, using as little as 20 μL of sample. The approach to gas content quantitation described here should greatly expand the range of animals and conditions that may be used in studies of metabolic gas exchange, and facilitate the development of artificial oxygen carriers and resuscitation fluids.

  14. Novel image processing approach to detect malaria

    NASA Astrophysics Data System (ADS)

    Mas, David; Ferrer, Belen; Cojoc, Dan; Finaurini, Sara; Mico, Vicente; Garcia, Javier; Zalevsky, Zeev

    2015-09-01

    In this paper we present a novel image processing algorithm providing good preliminary capabilities for in vitro detection of malaria. The proposed concept is based upon analysis of the temporal variation of each pixel. Changes in dark pixels mean that inter cellular activity happened, indicating the presence of the malaria parasite inside the cell. Preliminary experimental results involving analysis of red blood cells being either healthy or infected with malaria parasites, validated the potential benefit of the proposed numerical approach.

  15. Glucose encapsulating liposome for signal amplification for quantitative detection of biomarkers with glucometer readout.

    PubMed

    Zhao, Yuting; Du, Dan; Lin, Yuehe

    2015-10-15

    A new technology was developed to quantitatively detect a broad range of disease biomarkers and proven to be portable, economical, and conveniently accessible. Measurements were performed based on releasing encapsulated glucose from antibody-tagged liposomes and subsequently detecting the released glucose using a commercial personal glucose meter (GM). The innovative aspect of this approach lies in the quantification of target biomarkers through the detection of glucose, thus expanding the applicability of the GM by broadening the range of target biomarkers instead of detecting only one analyte, glucose. Because of the bilayer membrane of liposomes, which can accommodate tens of thousands of glucose molecules, the sensitivity was greatly enhanced by using glucose encapsulating liposomes as a signal output and an amplifier. Here, the model analyte, protein 53 phosphorylated on Serine 15 (phospho-p53(15)), was captured by primary antibodies bound on magnetic Fe3O4 nanoparticles and then recognized by reporting antibodies conjugated to glucose encapsulating liposomes. Finally, the target phospho-p53(15) was detected by lysing the bound liposomes to release the encapsulated glucose (4 × 10(5) glucose molecules per liposome), which is detected with the GM. This approach was demonstrated to be a universal technology that can be easily produced to quantify a wide variety of biomarkers in medical diagnostics, food safety, public health, and environmental monitoring. In the near future, it is expected that these sensors, in combination with a portable GM, can be used in many fields such as physicians' laboratories, hospitals and the common household. PMID:26005847

  16. [Research progress of real-time quantitative PCR method for group A rotavirus detection].

    PubMed

    Guo, Yan-Qing; Li, Dan-Di; Duan, Zhao-Jun

    2013-11-01

    Group A rotavirus is one of the most significant etiological agents which causes acute gastroenteritis among infants and young children worldwide. So far, several method which includes electron microscopy (EM), enzyme immunoassay (EIA), reverse transcription-polymerase chain reaction (RT-PCR)and Real-time Quantitative PCR has been established for the detection of rotavirus. Compared with other methods, Real-time quantitative PCR have advantages in specificity, sensitivity, genotyping and quantitative accuracy. This article shows a overview of the application of real-time quantitative PCR technique to detecte group A rotavirus.

  17. Surface-enhanced Raman scattering for quantitative detection of ethyl carbamate in alcoholic beverages.

    PubMed

    Yang, Danting; Zhou, Haibo; Ying, Yibin; Niessner, Reinhard; Haisch, Christoph

    2013-11-01

    Ethyl carbamate, a by-product of fermentation and storage with widespread occurrence in fermented food and alcoholic beverages, is a compound potentially toxic to humans. In this work, a new approach for quantitative detection of ethyl carbamate in alcoholic beverages, based on surface-enhanced Raman scattering (SERS), is reported. Individual silver-coated gold nanoparticle colloids are used as SERS amplifiers, yielding high Raman enhancement of ethyl carbamate in three kinds of alcoholic beverages (vodka, Obstler, and white rum). The characteristic band at 1,003 cm(-1), which is the strongest and best reproducible peak in the SERS spectra, was used for quantitative evaluation of ethyl carbamate. The limit of detection, which corresponds to a signal-to-noise ratio of 3, was 9.0 × 10(-9) M (0.8 μg · L(-1)), 1.3 × 10(-7) M (11.6 μg · L(-1)), and 7.8 × 10(-8) M (6.9 μg · L(-1)), respectively. Surface-enhanced Raman spectroscopy offers great practical potential for the in situ assessment and identification of ethyl carbamate in the alcoholic beverage industry.

  18. [Rapid quantitative detection of sulfate reducing bacteria in oil field].

    PubMed

    Wei, Li; Ma, Fang; Wang, Ji-Hua; Zhao, Li-Jun

    2007-02-01

    It take long time and high cost to measure sulfate reducing bacteria (SRB) in wastewater of oil field. A rapid quantitative method was developed by combining polymerase chain reaction(PCR) and most probable number (MPN) to measure sulfate reducing bacteria (SRB) in wastewater of oil field. The bacterium solution was directly prepared from wastewater for PCR amplification, which ensured quantitative accuracy. Reaction system and amplification condition were designed using universal primers DSR1F and DSR5R of dissimilatory sulfite reductase in SRB. The result show that the accuracy of this method is two magnitude higher than that of MPN. The whole measuring process take 3 - 4 hours and the reproducibility of this method is extremely stable, being fit to practical process.

  19. Mapping quantitative trait loci in selected breeding populations: A segregation distortion approach.

    PubMed

    Cui, Y; Zhang, F; Xu, J; Li, Z; Xu, S

    2015-12-01

    Quantitative trait locus (QTL) mapping is often conducted in line-crossing experiments where a sample of individuals is randomly selected from a pool of all potential progeny. QTLs detected from such an experiment are important for us to understand the genetic mechanisms governing a complex trait, but may not be directly relevant to plant breeding if they are not detected from the breeding population where selection is targeting for. QTLs segregating in one population may not necessarily segregate in another population. To facilitate marker-assisted selection, QTLs must be detected from the very population which the selection is targeting. However, selected breeding populations often have depleted genetic variation with small population sizes, resulting in low power in detecting useful QTLs. On the other hand, if selection is effective, loci controlling the selected trait will deviate from the expected Mendelian segregation ratio. In this study, we proposed to detect QTLs in selected breeding populations via the detection of marker segregation distortion in either a single population or multiple populations using the same selection scheme. Simulation studies showed that QTL can be detected in strong selected populations with selected population sizes as small as 25 plants. We applied the new method to detect QTLs in two breeding populations of rice selected for high grain yield. Seven QTLs were identified, four of which have been validated in advanced generations in a follow-up study. Cloned genes in the vicinity of the four QTLs were also reported in the literatures. This mapping-by-selection approach provides a new avenue for breeders to improve breeding progress. The new method can be applied to breeding programs not only in rice but also in other agricultural species including crops, trees and animals.

  20. Resolving the Quantitative-Qualitative Dilemma: A Critical Realist Approach

    ERIC Educational Resources Information Center

    Scott, David

    2007-01-01

    The philosophical issues underpinning the quantitative-qualitative divide in educational research are examined. Three types of argument which support a resolution are considered: pragmatism, false duality and warranty through triangulation. In addition a number of proposed strategies--alignment, sequencing, translation and triangulation--are…

  1. A Quantitative Approach to the Design of School Bus Routes.

    ERIC Educational Resources Information Center

    Tracz, George S.

    A number of factors--including the reorganization of school administrative structures, the availability of new technology, increased competition among groups for limited resources, and changing patterns of communication--suggest an increased need for quantitative analysis in the school district decision-making process. One area of school…

  2. Improving Library Performance: Quantitative Approaches to Library Planning.

    ERIC Educational Resources Information Center

    Webster, Duane E.

    The use of analytical models and quantitative methods for both short- and long-range problem solving offer library managers an excellent opportunity to improve and rationalize decision-making for strategic and organizational planning. The first step is to identify the problems confronting the library and understand its current capabilities.…

  3. Detecting Gene-Environment Interactions for a Quantitative Trait in a Genome-Wide Association Study.

    PubMed

    Zhang, Pingye; Lewinger, Juan Pablo; Conti, David; Morrison, John L; Gauderman, W James

    2016-07-01

    A genome-wide association study (GWAS) typically is focused on detecting marginal genetic effects. However, many complex traits are likely to be the result of the interplay of genes and environmental factors. These SNPs may have a weak marginal effect and thus unlikely to be detected from a scan of marginal effects, but may be detectable in a gene-environment (G × E) interaction analysis. However, a genome-wide interaction scan (GWIS) using a standard test of G × E interaction is known to have low power, particularly when one corrects for testing multiple SNPs. Two 2-step methods for GWIS have been previously proposed, aimed at improving efficiency by prioritizing SNPs most likely to be involved in a G × E interaction using a screening step. For a quantitative trait, these include a method that screens on marginal effects [Kooperberg and Leblanc, 2008] and a method that screens on variance heterogeneity by genotype [Paré et al., 2010] In this paper, we show that the Paré et al. approach has an inflated false-positive rate in the presence of an environmental marginal effect, and we propose an alternative that remains valid. We also propose a novel 2-step approach that combines the two screening approaches, and provide simulations demonstrating that the new method can outperform other GWIS approaches. Application of this method to a G × Hispanic-ethnicity scan for childhood lung function reveals a SNP near the MARCO locus that was not identified by previous marginal-effect scans. PMID:27230133

  4. Quantitative Raman Spectroscopy when the Signal-to-Noise is Below the Limit of Quantitation due to Fluorescence Interference: Advantages of a Moving Window Sequentially Shifted Excitation Approach.

    PubMed

    Marshall, Sarah; Cooper, John B

    2016-09-01

    Raman spectroscopy is a useful analytical tool. However, its application is often limited because shot noise from fluorescence obscures the Raman signal. In such cases, quantitative analysis is not possible when the signal-to-noise ratio (SNR) drops below two. A method is described for performing quantitative Raman spectroscopy that not only removes fluorescence backgrounds, but also results in a significant improvement in the SNR. The Raman data is extracted using a moving window sequentially shifted excitation algorithm. To demonstrate the capabilities of the method, a binary mixture of two analytes at varying concentrations is quantified in the presence of a highly fluorescent dye. Linear calibration plots were constructed and validated for the binary model using individual Raman peaks with SNR ranging from 0.073-12.6; r(2) values are greater than 0.96 in all cases, with all but the weakest peaks yielding values greater than 0.997. The presented method demonstrates a universal and autonomous approach for the quantitative analysis of highly fluorescent samples via Raman spectroscopy. The lower limit on the SNR ratio for quantitative Raman analysis with the described method is 0.1. In order to assess the effectiveness of the presented method, the entire set of experiments was also processed using the more common shifted excitation Raman difference spectroscopy (SERDS) approach. The advantages of the proposed method over SERDS are demonstrated for both the detection limit and the SNR of the processed spectra. PMID:27613308

  5. Quantitative Detection of Pharmaceuticals Using a Combination of Paper Microfluidics and Wavelength Modulated Raman Spectroscopy

    PubMed Central

    Craig, Derek; Mazilu, Michael; Dholakia, Kishan

    2015-01-01

    Raman spectroscopy has proven to be an indispensable technique for the identification of various types of analytes due to the fingerprint vibration spectrum obtained. Paper microfluidics has also emerged as a low cost, easy to fabricate and portable approach for point of care testing. However, due to inherent background fluorescence, combining Raman spectroscopy with paper microfluidics is to date an unmet challenge in the absence of using surface enhanced mechanisms. We describe the first use of wavelength modulated Raman spectroscopy (WMRS) for analysis on a paper microfluidics platform. This study demonstrates the ability to suppress the background fluorescence of the paper using WMRS and the subsequent implementation of this technique for pharmaceutical analysis. The results of this study demonstrate that it is possible to discriminate between both paracetamol and ibuprofen, whilst, also being able to detect the presence of each analyte quantitatively at nanomolar concentrations. PMID:25938464

  6. Quantitative detection of pharmaceuticals using a combination of paper microfluidics and wavelength modulated Raman spectroscopy.

    PubMed

    Craig, Derek; Mazilu, Michael; Dholakia, Kishan

    2015-01-01

    Raman spectroscopy has proven to be an indispensable technique for the identification of various types of analytes due to the fingerprint vibration spectrum obtained. Paper microfluidics has also emerged as a low cost, easy to fabricate and portable approach for point of care testing. However, due to inherent background fluorescence, combining Raman spectroscopy with paper microfluidics is to date an unmet challenge in the absence of using surface enhanced mechanisms. We describe the first use of wavelength modulated Raman spectroscopy (WMRS) for analysis on a paper microfluidics platform. This study demonstrates the ability to suppress the background fluorescence of the paper using WMRS and the subsequent implementation of this technique for pharmaceutical analysis. The results of this study demonstrate that it is possible to discriminate between both paracetamol and ibuprofen, whilst, also being able to detect the presence of each analyte quantitatively at nanomolar concentrations.

  7. Groundtruth approach to accurate quantitation of fluorescence microarrays

    SciTech Connect

    Mascio-Kegelmeyer, L; Tomascik-Cheeseman, L; Burnett, M S; van Hummelen, P; Wyrobek, A J

    2000-12-01

    To more accurately measure fluorescent signals from microarrays, we calibrated our acquisition and analysis systems by using groundtruth samples comprised of known quantities of red and green gene-specific DNA probes hybridized to cDNA targets. We imaged the slides with a full-field, white light CCD imager and analyzed them with our custom analysis software. Here we compare, for multiple genes, results obtained with and without preprocessing (alignment, color crosstalk compensation, dark field subtraction, and integration time). We also evaluate the accuracy of various image processing and analysis techniques (background subtraction, segmentation, quantitation and normalization). This methodology calibrates and validates our system for accurate quantitative measurement of microarrays. Specifically, we show that preprocessing the images produces results significantly closer to the known ground-truth for these samples.

  8. Quantitative architectural analysis: a new approach to cortical mapping.

    PubMed

    Schleicher, Axel; Morosan, Patricia; Amunts, Katrin; Zilles, Karl

    2009-11-01

    Results from functional imaging studies are often still interpreted using the classical architectonic brain maps of Brodmann and his successors. One obvious weakness in traditional, architectural mapping is the subjective nature of localizing borders between cortical areas by means of a purely visual, microscopical examination of histological specimens. To overcome this limitation, objective mapping procedures based on quantitative cytoarchitecture have been generated. As a result, new maps for various species including man were established. In our contribution, principles of quantitative cytoarchitecture and algorithm-based cortical mapping are described for a cytoarchitectural parcellation of the human auditory cortex. Defining cortical borders based on quantified changes in cortical lamination is the decisive step towards a novel, highly improved probabilistic brain atlas.

  9. An electrochemical immunosensor for quantitative detection of ficolin-3

    NASA Astrophysics Data System (ADS)

    San, Lili; Zeng, Dongdong; Song, Shiping; Zuo, Xiaolei; Zhang, Huan; Wang, Chenguang; Wu, Jiarui; Mi, Xianqiang

    2016-06-01

    Diabetes mellitus (DM) is one of the most common metabolic disorders in the world, of which more than 90% is type-2 diabetes mellitus (T2DM). There is a rather urgent need for reliable, sensitive and quick detection techniques in clinical application of T2DM. Ficolin-3 is a potential biomarker of T2DM, because serum ficolin-3 levels are associated with insulin resistance and predict the incidence of T2DM. Herein, a sandwich-type electrochemical immunosensor was developed for the detection of ficolin-3 in human serum. Cyclic voltammetry and the amperometric current versus time were used to characterize the performance of the immunosensor. Under optimal conditions, the detection limitation of ficolin-3 was 100 ng ml–1 and the linear dynamic range was between 2 and 50 μg ml–1. The method has ideal accuracy, excellent stability and selectivity and has wide application prospects in clinical research.

  10. An electrochemical immunosensor for quantitative detection of ficolin-3

    NASA Astrophysics Data System (ADS)

    San, Lili; Zeng, Dongdong; Song, Shiping; Zuo, Xiaolei; Zhang, Huan; Wang, Chenguang; Wu, Jiarui; Mi, Xianqiang

    2016-06-01

    Diabetes mellitus (DM) is one of the most common metabolic disorders in the world, of which more than 90% is type-2 diabetes mellitus (T2DM). There is a rather urgent need for reliable, sensitive and quick detection techniques in clinical application of T2DM. Ficolin-3 is a potential biomarker of T2DM, because serum ficolin-3 levels are associated with insulin resistance and predict the incidence of T2DM. Herein, a sandwich-type electrochemical immunosensor was developed for the detection of ficolin-3 in human serum. Cyclic voltammetry and the amperometric current versus time were used to characterize the performance of the immunosensor. Under optimal conditions, the detection limitation of ficolin-3 was 100 ng ml-1 and the linear dynamic range was between 2 and 50 μg ml-1. The method has ideal accuracy, excellent stability and selectivity and has wide application prospects in clinical research.

  11. Tools for the quantitative analysis of sedimentation boundaries detected by fluorescence optical analytical ultracentrifugation.

    PubMed

    Zhao, Huaying; Casillas, Ernesto; Shroff, Hari; Patterson, George H; Schuck, Peter

    2013-01-01

    Fluorescence optical detection in sedimentation velocity analytical ultracentrifugation allows the study of macromolecules at nanomolar concentrations and below. This has significant promise, for example, for the study of systems of high-affinity protein interactions. Here we describe adaptations of the direct boundary modeling analysis approach implemented in the software SEDFIT that were developed to accommodate unique characteristics of the confocal fluorescence detection system. These include spatial gradients of signal intensity due to scanner movements out of the plane of rotation, temporal intensity drifts due to instability of the laser and fluorophores, and masking of the finite excitation and detection cone by the sample holder. In an extensive series of experiments with enhanced green fluorescent protein ranging from low nanomolar to low micromolar concentrations, we show that the experimental data provide sufficient information to determine the parameters required for first-order approximation of the impact of these effects on the recorded data. Systematic deviations of fluorescence optical sedimentation velocity data analyzed using conventional sedimentation models developed for absorbance and interference optics are largely removed after these adaptations, resulting in excellent fits that highlight the high precision of fluorescence sedimentation velocity data, thus allowing a more detailed quantitative interpretation of the signal boundaries that is otherwise not possible for this system.

  12. RNA-seq analysis for detecting quantitative trait-associated genes

    PubMed Central

    Seo, Minseok; Kim, Kwondo; Yoon, Joon; Jeong, Jin Young; Lee, Hyun-Jeong; Cho, Seoae; Kim, Heebal

    2016-01-01

    Many recent RNA-seq studies were focused mainly on detecting the differentially expressed genes (DEGs) between two or more conditions. In contrast, only a few attempts have been made to detect genes associated with quantitative traits, such as obesity index and milk yield, on RNA-seq experiment with large number of biological replicates. This study illustrates the linear model application on trait associated genes (TAGs) detection in two real RNA-seq datasets: 89 replicated human obesity related data and 21 replicated Holsteins’ milk production related RNA-seq data. Based on these two datasets, the performance between suggesting methods, such as ordinary regression and robust regression, and existing methods: DESeq2 and Voom, were compared. The results indicate that suggesting methods have much lower false discoveries compared to the precedent two group comparisons based approaches in our simulation study and qRT-PCR experiment. In particular, the robust regression outperforms existing DEG finding method as well as ordinary regression in terms of precision. Given the current trend in RNA-seq pricing, we expect our methods to be successfully applied in various RNA-seq studies with numerous biological replicates that handle continuous response traits. PMID:27071914

  13. Optical digital coherent detection technology enabled flexible and ultra-fast quantitative phase imaging.

    PubMed

    Feng, Yuan-Hua; Lu, Xing; Song, Lu; Guo, Xiaojie; Wang, Yawei; Zhu, Linyan; Sui, Qi; Li, Jianping; Shi, Kebin; Li, Zhaohui

    2016-07-25

    Quantitative phase imaging has been an important labeling-free microscopy modality for many biomedical and material science applications. In which, ultra-fast quantitative phase imaging is indispensable for dynamic or transient characteristics analysis. Conventional wide field optical interferometry is a common scheme for quantitative phase imaging, while its data acquisition rate is usually hindered by the frame rate of arrayed detector. By utilizing novel balanced-photo-detector based digital optics coherent detection techniques, we report on a method of constructing ultra-fast quantitative phase microscopy at the line-scan rate of 100 MHz with ~2 μm spatial resolution. PMID:27464166

  14. Quantitative approaches to uncover physical mechanisms of tissue morphogenesis

    PubMed Central

    Gleghorn, Jason P.; Manivannan, Sriram; Nelson, Celeste M.

    2013-01-01

    Morphogenesis, the creation of tissue and organ architecture, is a series of complex and dynamic processes driven by genetic programs, microenvironmental cues, and intercellular interactions. Elucidating the physical mechanisms that generate tissue form is key to understanding development, disease, and the strategies needed for regenerative therapies. Advancements in imaging technologies, genetic recombination techniques, laser ablation, and microfabricated tissue models have enabled quantitative descriptions of the cellular motions and tissue deformations and stresses with unprecedented temporal and spatial resolution. Using these data synergistically with increasingly more sophisticated physical, mathematical, and computational models will unveil the physical mechanisms that drive morphogenesis. PMID:23647971

  15. Quantitative and qualitative approaches to identifying migration chronology in a continental migrant

    USGS Publications Warehouse

    Beatty, William S.; Kesler, Dylan C.; Webb, Elisabeth B.; Raedeke, Andrew H.; Naylor, Luke W.; Humburg, Dale D.

    2013-01-01

    The degree to which extrinsic factors influence migration chronology in North American waterfowl has not been quantified, particularly for dabbling ducks. Previous studies have examined waterfowl migration using various methods, however, quantitative approaches to define avian migration chronology over broad spatio-temporal scales are limited, and the implications for using different approaches have not been assessed. We used movement data from 19 female adult mallards (Anas platyrhynchos) equipped with solar-powered global positioning system satellite transmitters to evaluate two individual level approaches for quantifying migration chronology. The first approach defined migration based on individual movements among geopolitical boundaries (state, provincial, international), whereas the second method modeled net displacement as a function of time using nonlinear models. Differences in migration chronologies identified by each of the approaches were examined with analysis of variance. The geopolitical method identified mean autumn migration midpoints at 15 November 2010 and 13 November 2011, whereas the net displacement method identified midpoints at 15 November 2010 and 14 November 2011. The mean midpoints for spring migration were 3 April 2011 and 20 March 2012 using the geopolitical method and 31 March 2011 and 22 March 2012 using the net displacement method. The duration, initiation date, midpoint, and termination date for both autumn and spring migration did not differ between the two individual level approaches. Although we did not detect differences in migration parameters between the different approaches, the net displacement metric offers broad potential to address questions in movement ecology for migrating species. Ultimately, an objective definition of migration chronology will allow researchers to obtain a comprehensive understanding of the extrinsic factors that drive migration at the individual and population levels. As a result, targeted

  16. Optimization of Quantitative PCR Methods for Enteropathogen Detection.

    PubMed

    Liu, Jie; Gratz, Jean; Amour, Caroline; Nshama, Rosemary; Walongo, Thomas; Maro, Athanasia; Mduma, Esto; Platts-Mills, James; Boisen, Nadia; Nataro, James; Haverstick, Doris M; Kabir, Furqan; Lertsethtakarn, Paphavee; Silapong, Sasikorn; Jeamwattanalert, Pimmada; Bodhidatta, Ladaporn; Mason, Carl; Begum, Sharmin; Haque, Rashidul; Praharaj, Ira; Kang, Gagandeep; Houpt, Eric R

    2016-01-01

    Detection and quantification of enteropathogens in stool specimens is useful for diagnosing the cause of diarrhea but is technically challenging. Here we evaluate several important determinants of quantification: specimen collection, nucleic acid extraction, and extraction and amplification efficiency. First, we evaluate the molecular detection and quantification of pathogens in rectal swabs versus stool, using paired flocked rectal swabs and whole stool collected from 129 children hospitalized with diarrhea in Tanzania. Swabs generally yielded a higher quantification cycle (Cq) (average 29.7, standard deviation 3.5 vs. 25.3 ± 2.9 from stool, P<0.001) but were still able to detect 80% of pathogens with a Cq < 30 in stool. Second, a simplified total nucleic acid (TNA) extraction procedure was compared to separate DNA and RNA extractions and showed 92% (318/344) sensitivity and 98% (951/968) specificity, with no difference in Cq value for the positive results (ΔCq(DNA+RNA-TNA) = -0.01 ± 1.17, P = 0.972, N = 318). Third, we devised a quantification scheme that adjusts pathogen quantity to the specimen's extraction and amplification efficiency, and show that this better estimates the quantity of spiked specimens than the raw target Cq. In sum, these methods for enteropathogen quantification, stool sample collection, and nucleic acid extraction will be useful for laboratories studying enteric disease. PMID:27336160

  17. Optimization of Quantitative PCR Methods for Enteropathogen Detection

    PubMed Central

    Liu, Jie; Gratz, Jean; Amour, Caroline; Nshama, Rosemary; Walongo, Thomas; Maro, Athanasia; Mduma, Esto; Platts-Mills, James; Boisen, Nadia; Nataro, James; Haverstick, Doris M.; Kabir, Furqan; Lertsethtakarn, Paphavee; Silapong, Sasikorn; Jeamwattanalert, Pimmada; Bodhidatta, Ladaporn; Mason, Carl; Begum, Sharmin; Haque, Rashidul; Praharaj, Ira; Kang, Gagandeep; Houpt, Eric R.

    2016-01-01

    Detection and quantification of enteropathogens in stool specimens is useful for diagnosing the cause of diarrhea but is technically challenging. Here we evaluate several important determinants of quantification: specimen collection, nucleic acid extraction, and extraction and amplification efficiency. First, we evaluate the molecular detection and quantification of pathogens in rectal swabs versus stool, using paired flocked rectal swabs and whole stool collected from 129 children hospitalized with diarrhea in Tanzania. Swabs generally yielded a higher quantification cycle (Cq) (average 29.7, standard deviation 3.5 vs. 25.3 ± 2.9 from stool, P<0.001) but were still able to detect 80% of pathogens with a Cq < 30 in stool. Second, a simplified total nucleic acid (TNA) extraction procedure was compared to separate DNA and RNA extractions and showed 92% (318/344) sensitivity and 98% (951/968) specificity, with no difference in Cq value for the positive results (ΔCq(DNA+RNA-TNA) = -0.01 ± 1.17, P = 0.972, N = 318). Third, we devised a quantification scheme that adjusts pathogen quantity to the specimen’s extraction and amplification efficiency, and show that this better estimates the quantity of spiked specimens than the raw target Cq. In sum, these methods for enteropathogen quantification, stool sample collection, and nucleic acid extraction will be useful for laboratories studying enteric disease. PMID:27336160

  18. Dual core quantum dots for highly quantitative ratiometric detection of trypsin activity in cystic fibrosis patients

    NASA Astrophysics Data System (ADS)

    Castelló Serrano, Iván; Stoica, Georgiana; Matas Adams, Alba; Palomares, Emilio

    2014-10-01

    We present herein two colour encoded silica nanospheres (2nanoSi) for the fluorescence quantitative ratiometric determination of trypsin in humans. Current detection methods for cystic fibrosis diagnosis are slow, costly and suffer from false positives. The 2nanoSi proved to be a highly sensitive, fast (minutes), and single-step approach nanosensor for the screening and diagnosis of cystic fibrosis, allowing the quantification of trypsin concentrations in a wide range relevant for clinical applications (25-350 μg L-1). Furthermore, as trypsin is directly related to the development of cystic fibrosis (CF), different human genotypes, i.e. CF homozygotic, CF heterozygotic, and unaffected, respectively, can be determined using our 2nanoSi nanospheres. We anticipate the 2nanoSi system to be a starting point for non-invasive, easy-to-use and cost effective ratiometric fluorescent biomarkers for recessive genetic diseases like human cystic fibrosis. In a screening program in which the goal is to detect disease and also the carrier status, early diagnosis could be of great help.We present herein two colour encoded silica nanospheres (2nanoSi) for the fluorescence quantitative ratiometric determination of trypsin in humans. Current detection methods for cystic fibrosis diagnosis are slow, costly and suffer from false positives. The 2nanoSi proved to be a highly sensitive, fast (minutes), and single-step approach nanosensor for the screening and diagnosis of cystic fibrosis, allowing the quantification of trypsin concentrations in a wide range relevant for clinical applications (25-350 μg L-1). Furthermore, as trypsin is directly related to the development of cystic fibrosis (CF), different human genotypes, i.e. CF homozygotic, CF heterozygotic, and unaffected, respectively, can be determined using our 2nanoSi nanospheres. We anticipate the 2nanoSi system to be a starting point for non-invasive, easy-to-use and cost effective ratiometric fluorescent biomarkers for

  19. Detection of cardiomyopathy in an animal model using quantitative autoradiography

    SciTech Connect

    Kubota, K.; Som, P.; Oster, Z.H.; Brill, A.B.; Goodman, M.M.; Knapp, F.F. Jr.; Atkins, H.L.; Sole, M.J.

    1988-10-01

    A fatty acid analog (15-p-iodophenyl)-3,3 dimethyl-pentadecanoic acid (DMIPP) was studied in cardiomyopathic (CM) and normal age-matched Syrian hamsters. Dual tracer quantitative wholebody autoradiography (QARG) with DMIPP and 2-(/sup 14/C(U))-2-deoxy-2-fluoro-D-glucose (FDG) or with FDG and /sup 201/Tl enabled comparison of the uptake of a fatty acid and a glucose analog with the blood flow. These comparisons were carried out at the onset and mid-stage of the disease before congestive failure developed. Groups of CM and normal animals were treated with verapamil from the age of 26 days, before the onset of the disease for 41 days. In CM hearts, areas of decreased DMIPP uptake were seen. These areas were much larger than the decrease in uptake of FDG or /sup 201/Tl. In early CM only minimal changes in FDG or /sup 201/Tl uptake were observed as compared to controls. Treatment of CM-prone animals with verapamil prevented any changes in DMIPP, FDG, or /sup 201/Tl uptake. DMIPP seems to be a more sensitive indicator of early cardiomyopathic changes as compared to /sup 201/Tl or FDG. The trial of DMIPP and SPECT in the diagnosis of human disease, as well as for monitoring the effects of drugs which may prevent it seems to be warranted.

  20. A Quantitative Proteomic Approach to Prion Disease Biomarker Research: Delving into the Glycoproteome

    PubMed Central

    Wei, Xin; Herbst, Allen; Ma, Di; Aiken, Judd; Li, Lingjun

    2011-01-01

    Mass spectrometry (MS) – based proteomic approaches have evolved as powerful tools for the discovery of biomarkers. However, the identification of potential protein biomarkers from biofluid samples is challenging because of the limited dynamic range of detection. Currently there is a lack of sensitive and reliable pre-mortem diagnostic test for prion diseases. Here, we describe the use of a combined MS-based approach for biomarker discovery in prion diseases from mouse plasma samples. To overcome the limited dynamic range of detection and sample complexity of plasma samples, we used lectin affinity chromatography and multi-dimensional separations to enrich and isolate glycoproteins at low abundance. Relative quantitation of a panel of proteins was obtained by a combination of isotopic labeling and validated by spectral counting. Overall 708 proteins were identified, 53 of which showed more than 2-fold increase in concentration whereas 58 exhibited more than 2-fold decrease. A few of the potential candidate markers were previously associated with prion or other neurodegenerative diseases. PMID:21469646

  1. A novel quantitative PCR assay for the detection of Streptococcus pneumoniae using the competence regulator gene target comX.

    PubMed

    Habets, Marrit N; Cremers, Amelieke J H; Bos, Martine P; Savelkoul, Paul; Eleveld, Marc J; Meis, Jacques F; Hermans, Peter W M; Melchers, Willem J; de Jonge, Marien I; Diavatopoulos, Dimitri A

    2016-02-01

    Streptococcus pneumoniae is responsible for an estimated 1.6 million deaths worldwide every year. While rapid detection and timely treatment with appropriate antibiotics is preferred, this is often difficult due to the amount of time that detection with blood cultures takes. In this study, a novel quantitative PCR assay for the detection of Streptococcus pneumoniae was developed. To identify novel targets, we analysed the pneumococcal genome for unique, repetitive DNA sequences. This approach identified comX, which is conserved and present in duplicate copies in Streptococcus pneumoniae but not in other bacterial species. Comparison with lytA, the current 'gold standard' for detection by quantitative PCR, demonstrated an analytic specificity of 100% for both assays on a panel of 10 pneumococcal and 18 non-pneumococcal isolates, but a reduction of 3.5 quantitation cycle values (± 0.23 sem), resulting in an increased analytical detection rate of comX. We validated our assay on DNA extracted from the serum of 30 bacteraemic patients who were blood culture positive for Streptococcus pneumoniae and 51 serum samples that were culture positive for other bacteria. This resulted in a similar clinical sensitivity between the comX and lytA assays (47%) and in a diagnostic specificity of 98.2 and 100% for the lytA and comX assays, respectively. In conclusion, we have developed a novel quantitative PCR assay with increased analytical sensitivity for the detection of Streptococcus pneumoniae, which may be used to develop a rapid bedside test for the direct detection of Streptococcus pneumoniae in clinical specimens.

  2. The conceptual approach to quantitative modeling of guard cells.

    PubMed

    Blatt, Michael R; Hills, Adrian; Chen, Zhong-Hua; Wang, Yizhou; Papanatsiou, Maria; Lew, Vigilio L

    2013-01-01

    Much of the 70% of global water usage associated with agriculture passes through stomatal pores of plant leaves. The guard cells, which regulate these pores, thus have a profound influence on photosynthetic carbon assimilation and water use efficiency of plants. We recently demonstrated how quantitative mathematical modeling of guard cells with the OnGuard modeling software yields detail sufficient to guide phenotypic and mutational analysis. This advance represents an all-important step toward applications in directing "reverse-engineering" of guard cell function for improved water use efficiency and carbon assimilation. OnGuard is nonetheless challenging for those unfamiliar with a modeler's way of thinking. In practice, each model construct represents a hypothesis under test, to be discarded, validated or refined by comparisons between model predictions and experimental results. The few guidelines set out here summarize the standard and logical starting points for users of the OnGuard software.

  3. Analytical bioconjugates, aptamers, enable specific quantitative detection of Listeria monocytogenes.

    PubMed

    Lee, Sang-Hee; Ahn, Ji-Young; Lee, Kyeong-Ah; Um, Hyun-Ju; Sekhon, Simranjeet Singh; Sun Park, Tae; Min, Jiho; Kim, Yang-Hoon

    2015-06-15

    As a major human pathogen in the Listeria genus, Listeria monocytogenes causes the bacterial disease listeriosis, which is a serious infection caused by eating food contaminated with the bacteria. We have developed an aptamer-based sandwich assay (ABSA) platform that demonstrates a promising potential for use in pathogen detection using aptamers as analytical bioconjugates. The whole-bacteria SELEX (WB-SELEX) strategy was adopted to generate aptamers with high affinity and specificity against live L. monocytogenes. Of the 35 aptamer candidates tested, LMCA2 and LMCA26 reacted to L. monocytogenes with high binding, and were consequently chosen as sensing probes. The ABSA platform can significantly enhance the sensitivity by employing a very specific aptamer pair for the sandwich complex. The ABSA platform exhibited a linear response over a wide concentration range of L. monocytogenes from 20 to 2×10(6) CFU per mL and was closely correlated with the following relationship: y=9533.3x+1542.3 (R(2)=0.99). Our proposed ABSA platform also provided excellent specificity for the tests to distinguish L. monocytogenes from other Listeria species and other bacterial genera (3 Listeria spp., 4 Salmonella spp., 2 Vibrio spp., 3 Escherichia coli and 3 Shigella spp.). Improvements in the sensitivity and specificity have not only facilitated the reliable detection of L. monocytogenes at extremely low concentrations, but also allowed for the development of a 96-well plate-based routine assay platform for multivalent diagnostics.

  4. Quantitative detection of bovine and porcine gelatin difference using surface plasmon resonance based biosensor

    NASA Astrophysics Data System (ADS)

    Wardani, Devy P.; Arifin, Muhammad; Suharyadi, Edi; Abraha, Kamsul

    2015-05-01

    Gelatin is a biopolymer derived from collagen that is widely used in food and pharmaceutical products. Due to some religion restrictions and health issues regarding the gelatin consumption which is extracted from certain species, it is necessary to establish a robust, reliable, sensitive and simple quantitative method to detect gelatin from different parent collagen species. To the best of our knowledge, there has not been a gelatin differentiation method based on optical sensor that could detect gelatin from different species quantitatively. Surface plasmon resonance (SPR) based biosensor is known to be a sensitive, simple and label free optical method for detecting biomaterials that is able to do quantitative detection. Therefore, we have utilized SPR-based biosensor to detect the differentiation between bovine and porcine gelatin in various concentration, from 0% to 10% (w/w). Here, we report the ability of SPR-based biosensor to detect difference between both gelatins, its sensitivity toward the gelatin concentration change, its reliability and limit of detection (LOD) and limit of quantification (LOQ) of the sensor. The sensor's LOD and LOQ towards bovine gelatin concentration are 0.38% and 1.26% (w/w), while towards porcine gelatin concentration are 0.66% and 2.20% (w/w), respectively. The results show that SPR-based biosensor is a promising tool for detecting gelatin from different raw materials quantitatively.

  5. New quantitative detection of pathogens in heterogeneous environmental samples

    NASA Astrophysics Data System (ADS)

    Lee, Eun-Hee; Wang, Xiaofang; Mitchell, Kristi; Chae, Seon-Ha; Son, Ahjeong

    2015-04-01

    Quantum dots and magnetic beads based genomic assay (NanoGene assay) has been developed for sensitive and inhibition resistant gene quantification to achieve in-situ bacteria monitoring in environmental samples. In this study, eaeA gene of pathogenic E. coli O157:H7 was quantified. The result demonstrated the excellent sensitivity (i.e., limit of detection: 87 gene copies for dsDNA and 890 zeptomolar for ssDNA) in the presence of nonspecific microbial populations (Kim et al., 2010; 2011a). The feasibility of the developed gene quantification for non-laboratory environment usage (in-situ use) was investigated. Therefore, DNA hybridization was achieved at ambient temperature and minimum agitation, and the analysis was completed within hours. Most importantly, the NanoGene assay demonstrated the resistance to the presence of naturally occurring inhibitors (humic acids, cations) and residual reagents (surfactants, alcohols) from DNA extraction (Kim et al., 2011b). The assay was also applied to humic acids laden soils (7 types of soils with various amount of organic matters) and successfully quantified 105 to 108 CFU of E. coli O157:H7 per gram soil (R2 = 0.99). The results indicate that the presented NanoGene assay is suitable for further development as an in-situ bacteria monitoring method for working with heterogeneous environmental samples (Wang et al., 2013). Another aspect of the method is to transform the NanoGene assay into a portable device that can be used as a pathogenic bacteria detector in environment. The project consisted of the first inline fluidic components development and characterization as well as the first integration effort on a briefcase platform for the in-situ pathogen detection system (IPDS) (Mitchell et al., 2014). Our long term vision is to further miniaturize the briefcase platform implementation of the IPDS and to commercialize the handheld version of the IPDS.

  6. Quantitative detection of Aspergillus spp. by real-time nucleic acid sequence-based amplification.

    PubMed

    Zhao, Yanan; Perlin, David S

    2013-01-01

    Rapid and quantitative detection of Aspergillus from clinical samples may facilitate an early diagnosis of invasive pulmonary aspergillosis (IPA). As nucleic acid-based detection is a viable option, we demonstrate that Aspergillus burdens can be rapidly and accurately detected by a novel real-time nucleic acid assay other than qPCR by using the combination of nucleic acid sequence-based amplification (NASBA) and the molecular beacon (MB) technology. Here, we detail a real-time NASBA assay to determine quantitative Aspergillus burdens in lungs and bronchoalveolar lavage (BAL) fluids of rats with experimental IPA.

  7. Furfural as a marker of cellulose degradation. A quantitative approach

    NASA Astrophysics Data System (ADS)

    Łojewski, Tomasz; Sawoszczuk, Tomasz; Łagan, Janusz Marek; Zięba, Katarzyna; Barański, Andrzej; Łojewska, Joanna

    2010-09-01

    Non-destructive methods of sampling during the physicochemical studies of historical objects such as old books and manuscripts seem to be an obvious choice. Since furfural has been shown to be one of the most abundant gaseous products of cellulose degradation, it can be considered as a convenient marker of degradation progress. The number of quantitative data concerning correlations between the emission of furfural and physicochemical and mechanical properties of paper is rather scarce in the literature. In the present studies, a model paper containing more than 99% of cellulose was aged inside closed vials at 90°C. Gaseous products of paper degradation were measured using sorption tubes filled with Tenax TA sorbent and GC-MS. The method has proved to be sufficiently sensitive for measuring furfural emission not only in accelerated degradation at 90°C but also during natural ageing of paper at room temperature even in relatively short time intervals of 2-28 days. The correlations between furfural emission and polymerization degree, pH, color, tear index, number of double folds and breaking length have been statistically confirmed at confidence level α=0.001. Basing on them it was possible to estimate the number of broken glycosidic bonds per one molecule of furfural formed during degradation—we found a value equal to 9.2.

  8. Drosophila wing modularity revisited through a quantitative genetic approach.

    PubMed

    Muñoz-Muñoz, Francesc; Carreira, Valeria Paula; Martínez-Abadías, Neus; Ortiz, Victoria; González-José, Rolando; Soto, Ignacio M

    2016-07-01

    To predict the response of complex morphological structures to selection it is necessary to know how the covariation among its different parts is organized. Two key features of covariation are modularity and integration. The Drosophila wing is currently considered a fully integrated structure. Here, we study the patterns of integration of the Drosophila wing and test the hypothesis of the wing being divided into two modules along the proximo-distal axis, as suggested by developmental, biomechanical, and evolutionary evidence. To achieve these goals we perform a multilevel analysis of covariation combining the techniques of geometric morphometrics and quantitative genetics. Our results indicate that the Drosophila wing is indeed organized into two main modules, the wing base and the wing blade. The patterns of integration and modularity were highly concordant at the phenotypic, genetic, environmental, and developmental levels. Besides, we found that modularity at the developmental level was considerably higher than modularity at other levels, suggesting that in the Drosophila wing direct developmental interactions are major contributors to total phenotypic shape variation. We propose that the precise time at which covariance-generating developmental processes occur and/or the magnitude of variation that they produce favor proximo-distal, rather than anterior-posterior, modularity in the Drosophila wing.

  9. Biological evolution of replicator systems: towards a quantitative approach.

    PubMed

    Martin, Osmel; Horvath, J E

    2013-04-01

    The aim of this work is to study the features of a simple replicator chemical model of the relation between kinetic stability and entropy production under the action of external perturbations. We quantitatively explore the different paths leading to evolution in a toy model where two independent replicators compete for the same substrate. To do that, the same scenario described originally by Pross (J Phys Org Chem 17:312-316, 2004) is revised and new criteria to define the kinetic stability are proposed. Our results suggest that fast replicator populations are continually favored by the effects of strong stochastic environmental fluctuations capable to determine the global population, the former assumed to be the only acting evolution force. We demonstrate that the process is continually driven by strong perturbations only, and that population crashes may be useful proxies for these catastrophic environmental fluctuations. As expected, such behavior is particularly enhanced under very large scale perturbations, suggesting a likely dynamical footprint in the recovery patterns of new species after mass extinction events in the Earth's geological past. Furthermore, the hypothesis that natural selection always favors the faster processes may give theoretical support to different studies that claim the applicability of maximum principles like the Maximum Metabolic Flux (MMF) or Maximum Entropy Productions Principle (MEPP), seen as the main goal of biological evolution.

  10. Drosophila wing modularity revisited through a quantitative genetic approach.

    PubMed

    Muñoz-Muñoz, Francesc; Carreira, Valeria Paula; Martínez-Abadías, Neus; Ortiz, Victoria; González-José, Rolando; Soto, Ignacio M

    2016-07-01

    To predict the response of complex morphological structures to selection it is necessary to know how the covariation among its different parts is organized. Two key features of covariation are modularity and integration. The Drosophila wing is currently considered a fully integrated structure. Here, we study the patterns of integration of the Drosophila wing and test the hypothesis of the wing being divided into two modules along the proximo-distal axis, as suggested by developmental, biomechanical, and evolutionary evidence. To achieve these goals we perform a multilevel analysis of covariation combining the techniques of geometric morphometrics and quantitative genetics. Our results indicate that the Drosophila wing is indeed organized into two main modules, the wing base and the wing blade. The patterns of integration and modularity were highly concordant at the phenotypic, genetic, environmental, and developmental levels. Besides, we found that modularity at the developmental level was considerably higher than modularity at other levels, suggesting that in the Drosophila wing direct developmental interactions are major contributors to total phenotypic shape variation. We propose that the precise time at which covariance-generating developmental processes occur and/or the magnitude of variation that they produce favor proximo-distal, rather than anterior-posterior, modularity in the Drosophila wing. PMID:27272402

  11. Calculation of measurement uncertainty in quantitative analysis of genetically modified organisms using intermediate precision--a practical approach.

    PubMed

    Zel, Jana; Gruden, Kristina; Cankar, Katarina; Stebih, Dejan; Blejec, Andrej

    2007-01-01

    Quantitative characterization of nucleic acids is becoming a frequently used method in routine analysis of biological samples, one use being the detection of genetically modified organisms (GMOs). Measurement uncertainty is an important factor to be considered in these analyses, especially where precise thresholds are set in regulations. Intermediate precision, defined as a measure between repeatability and reproducibility, is a parameter describing the real situation in laboratories dealing with quantitative aspects of molecular biology methods. In this paper, we describe the top-down approach to calculating measurement uncertainty, using intermediate precision, in routine GMO testing of food and feed samples. We illustrate its practicability in defining compliance of results with regulations. The method described is also applicable to other molecular methods for a variety of laboratory diagnostics where quantitative characterization of nucleic acids is needed.

  12. Toxicity screening of metals with special reference to quantitative approach.

    PubMed

    Ghosh, S K; Doctor, P B; Derasari, Anuradha; Amin, R J

    2004-01-01

    A series of metals Cr(6+), Al(3+), Cd(2+), Pb(2+), Cu(2+), Zn(2+), and Hg(2+) were tested in three systems--Microtox, Motility Test, and Growth Zone Inhibition Test. Toxicity endpoint of each metal was variable from system to system. Of the three systems, Microtox was the most sensitive system. In this system, Hg(2+) reacted as the most toxic element having EC(50) value 0.08 mg/L while Cd(2+) and Cr(6+) were least toxic with their EC(50) values 19.4 and 21.0 mg/L respectively. MEC(90) value of Motility Test was always needed more concentrations of toxicant in comparison to other systems. In comparison to Microtox, ten times more concentration of Hg(2+) (1.4 mg/L) was required to find out its MEC(90) value. Growth Zone Inhibition Test was very simple method from handling point of view. The usual practice of evaluation of toxicity screening in this system is either qualitatively or semi-qualitatively. Hence a study was designed to establish a quantitative technique, Growth Inhibition Test, as an alternative to this test using the same sensor organism B. cereus, which allows determination of MAC as well as MIC. MIC for Hg(2+) was found to be 0.03 mg/L in Growth Inhibition Test while the same element was needed more concentration (1.0 mg/L) in the case of Growth Zone Inhibition test to produce halo. However, all these systems including Growth Inhibition Test showed Hg(2+) was the most toxic element.

  13. Introduction to Focus Issue: Quantitative Approaches to Genetic Networks

    NASA Astrophysics Data System (ADS)

    Albert, Réka; Collins, James J.; Glass, Leon

    2013-06-01

    All cells of living organisms contain similar genetic instructions encoded in the organism's DNA. In any particular cell, the control of the expression of each different gene is regulated, in part, by binding of molecular complexes to specific regions of the DNA. The molecular complexes are composed of protein molecules, called transcription factors, combined with various other molecules such as hormones and drugs. Since transcription factors are coded by genes, cellular function is partially determined by genetic networks. Recent research is making large strides to understand both the structure and the function of these networks. Further, the emerging discipline of synthetic biology is engineering novel gene circuits with specific dynamic properties to advance both basic science and potential practical applications. Although there is not yet a universally accepted mathematical framework for studying the properties of genetic networks, the strong analogies between the activation and inhibition of gene expression and electric circuits suggest frameworks based on logical switching circuits. This focus issue provides a selection of papers reflecting current research directions in the quantitative analysis of genetic networks. The work extends from molecular models for the binding of proteins, to realistic detailed models of cellular metabolism. Between these extremes are simplified models in which genetic dynamics are modeled using classical methods of systems engineering, Boolean switching networks, differential equations that are continuous analogues of Boolean switching networks, and differential equations in which control is based on power law functions. The mathematical techniques are applied to study: (i) naturally occurring gene networks in living organisms including: cyanobacteria, Mycoplasma genitalium, fruit flies, immune cells in mammals; (ii) synthetic gene circuits in Escherichia coli and yeast; and (iii) electronic circuits modeling genetic networks

  14. Ecohydrology of agroecosystems: quantitative approaches towards sustainable irrigation.

    PubMed

    Vico, Giulia; Porporato, Amilcare

    2015-02-01

    Irrigation represents one of the main strategies to enhance and stabilize agricultural productivity, by mitigating the effects of rainfall vagaries. In the face of the projected growth in population and in biofuel demands, as well as shifts in climate and dietary habits, a more sustainable management of water resources in agroecosystems is needed. The field of ecohydrology, traditionally focusing on natural ecosystems, has the potential to offer the necessary quantitative tools to assess and compare agricultural enterprises across climates, soil types, crops, and irrigation strategies, accounting for the unpredictability of the hydro-climatic forcing. Here, agricultural sustainability and productivity are assessed with reference to water productivity (defined as the ratio between yield and total supplied water), yields, water requirements, and their variability-a crucial element for food security and resource allocation planning. These synthetic indicators are quantified by means of a probabilistic description of the soil water balance and crop development. The model results allow the interpretation of patterns of water productivity observed in Zea mays (maize) and Triticum aestivum (wheat), grown under a variety of soils, climates, and irrigation strategies. Employing the same modeling framework, the impact of rainfall pattern and irrigation strategy on yield and water requirements is further explored. The obtained standard deviations of yield and water requirements suggest the existence of a nonlinear tradeoff between yield stabilization and variability of water requirements, which in turn is strongly impacted by irrigation strategy. Moreover, intermediate rainfall amounts are associated to the highest variability in yields and irrigation requirements, although allowing the maximum water productivity. The existence of these tradeoffs between productivity, reliability, and sustainability poses a problem for water management, in particular in mesic climates.

  15. Impacts of using inbred animals in studies for detection of quantitative trait loci.

    PubMed

    Freyer, G; Vukasinovic, N; Cassell, B

    2009-02-01

    Effects of utilizing inbred and noninbred family structures in experiments for detection of quantitative trait loci (QTL) were compared in this simulation study. Simulations were based on a general pedigree design originating from 2 unrelated sires. A variance component approach of mapping QTL was applied to simulated data that reflected common family structures from dairy populations. Five different family structures were considered: FS0 without inbreeding, FS1 with an inbred sire from an aunt-nephew mating, FS2 with an inbred sire originating from a half-sib mating, FS3 and FS4 based on FS2 but containing an increased number of offspring of the inbred sire (FS3), and another extremely inbred sire with its final offspring (FS4). Sixty replicates each of the 5 family structures in 2 simulation scenarios each were analyzed to provide a praxis-like situation of QTL analysis. The largest proportion of QTL position estimates within the correct interval of 3 cM, best test statistic profiles and the smallest average bias were obtained from the pedigrees described by FS4 and FS2. The approach does not depend on the kind and number of genetic markers. Inbreeding is not a recommended practice for commercial dairy production because of possible inbreeding depression, but inbred animals and their offspring that already exist could be advantageous for QTL mapping, because of reduced genetic variance in inbred parents.

  16. Quantitative detection and differentiation of free-living amoeba species using SYBR green-based real-time PCR melting curve analysis.

    PubMed

    Behets, Jonas; Declerck, Priscilla; Delaedt, Yasmine; Verelst, Lieve; Ollevier, Frans

    2006-12-01

    Real-time polymerase chain reaction melting curve analysis (MCA) allows differentiation of several free-living amoebae species. Distinctive characteristics were found for Naegleria fowleri, N. lovaniensis, N. australiensis, N. gruberi, Hartmanella vermiformis, and Willaertia magna. Species specificity of the amplicons was confirmed using agarose gel electrophoresis and sequence-based approaches. Amplification efficiency ranged from 91% to 98%, indicating the quantitative potential of the assay. This MCA approach can be used for quantitative detection of free-living amoebae after cultivation but also as a culture-independent detection method.

  17. Quantitative hopanoid analysis enables robust pattern detection and comparison between laboratories.

    PubMed

    Wu, C-H; Kong, L; Bialecka-Fornal, M; Park, S; Thompson, A L; Kulkarni, G; Conway, S J; Newman, D K

    2015-07-01

    external diplopterol standards. These new quantitative approaches permit meaningful comparisons between studies, allowing more accurate hopanoid pattern detection in both laboratory and environmental samples. PMID:25865768

  18. Quantitative hopanoid analysis enables robust pattern detection and comparison between laboratories

    PubMed Central

    Wu, C-H; Kong, L; Bialecka-Fornal, M; Park, S; Thompson, A L; Kulkarni, G; Conway, S J; Newman, D K

    2015-01-01

    external diplopterol standards. These new quantitative approaches permit meaningful comparisons between studies, allowing more accurate hopanoid pattern detection in both laboratory and environmental samples. PMID:25865768

  19. A Discriminative Approach to EEG Seizure Detection

    PubMed Central

    Johnson, Ashley N.; Sow, Daby; Biem, Alain

    2011-01-01

    Seizures are abnormal sudden discharges in the brain with signatures represented in electroencephalograms (EEG). The efficacy of the application of speech processing techniques to discriminate between seizure and non-seizure states in EEGs is reported. The approach accounts for the challenges of unbalanced datasets (seizure and non-seizure), while also showing a system capable of real-time seizure detection. The Minimum Classification Error (MCE) algorithm, which is a discriminative learning algorithm with wide-use in speech processing, is applied and compared with conventional classification techniques that have already been applied to the discrimination between seizure and non-seizure states in the literature. The system is evaluated on 22 pediatric patients multi-channel EEG recordings. Experimental results show that the application of speech processing techniques and MCE compare favorably with conventional classification techniques in terms of classification performance, while requiring less computational overhead. The results strongly suggests the possibility of deploying the designed system at the bedside. PMID:22195192

  20. Quaternary coastal evolution of Oman (Arabian Peninsula) - a quantitative approach

    NASA Astrophysics Data System (ADS)

    Hoffmann, G.; Rupprechter, M.; Roepert, A.; Quraishi, K. Al; Balushi, N. Al; Grützner, C.; Reicherter, K.

    2012-04-01

    The paper reviews the Quaternary coastal evolution of Oman. Emphasise is put on quantifying the different forcing factors. The plate tectonic setting, the Quaternary climate evolution, the sea-level history and the impact of natural hazards are identified as key factors of coastal evolution. The Arabian Plate is characterized by a northward movement forming a continent-continent collision zone in the west and the Makran Subduction Zone in the east. As a result differential land movement is observable in Oman. The Quaternary climate evolution is well understood. Besides other proxies notably spelothems and aeolian deposits allow to draw a consistent picture. It is understood that changes in the position of the intertropical convergence zone result in intensity-changes of the summer monsoon. These changes are related to global atmospheric circulation patterns. Data on the sea-level history are sparse; despite general assumptions of a sea-level lowstand, correlating with the last glacial maximum, resulting in terrestrial conditions within the Arabian Gulf. Furthermore, a mid-Holocene sea level highstand in the range of +2m is documented in several locations. The coastlines of Oman are affected by tsunami and hurricanes. However, almost no instrumental or historical data on the impact of such natural hazards are available due to the isolation of the country in the past. Several Quaternary deposits have been investigated in a reconnaissance survey. There is sound geological evidence for a tsunami to have affected the coastline in 1945, with the possibility of older tsunami events being also recorded in the geological record. There is strong evidence of differential land movement along the coastline; locally indicated by marine terraces in elevations of up to 400m (Rupprechter at al. 2012). By quantifying the differential land movement for numerous sites, the sea-level history will be revealed. Ultimately the data will be utilized to form the base of a modeling approach

  1. A quantitative approach to identifying predators from nest remains

    USGS Publications Warehouse

    Anthony, R.M.; Grand, J.B.; Fondell, T.F.; Manly, B.F.

    2004-01-01

    Nesting success of Dusky Canada Geese (Branta canadensis occidentalis) has declined greatly since a major earthquake affected southern Alaska in 1964. To identify nest predators, we collected predation data at goose nests and photographs of predators at natural nests containing artificial eggs in 1997-2000. To document feeding behavior by nest predators, we compiled the evidence from destroyed nests with known predators on our study site and from previous studies. We constructed a profile for each predator group and compared the evidence from 895 nests with unknown predators to our predator profiles using mixture-model analysis. This analysis indicated that 72% of destroyed nests were depredated by Bald Eagles and 13% by brown bears, and also yielded the probability that each nest was correctly assigned to a predator group based on model fit. Model testing using simulations indicated that the proportion estimated for eagle predation was unbiased and the proportion for bear predation was slightly overestimated. This approach may have application whenever there are adequate data on nests destroyed by known predators and predators exhibit different feeding behavior at nests.

  2. Application of Person-Centered Approaches to Critical Quantitative Research: Exploring Inequities in College Financing Strategies

    ERIC Educational Resources Information Center

    Malcom-Piqueux, Lindsey

    2014-01-01

    This chapter discusses the utility of person-centered approaches to critical quantitative researchers. These techniques, which identify groups of individuals who share similar attributes, experiences, or outcomes, are contrasted with more commonly used variable-centered approaches. An illustrative example of a latent class analysis of the college…

  3. Liquid crystal-based sensors for selective and quantitative detection of nitrogen dioxide

    PubMed Central

    Sen, Avijit; Kupcho, Kurt A.; Grinwald, Bart A.; VanTreeck, Heidi J.; Acharya, Bharat R.

    2013-01-01

    A highly sensitive nitrogen dioxide (NO2) sensor based on orientational transition of a thin film of liquid crystal (LC) supported on a gold surface is reported. Transport of NO2 molecules through the LC film to the LC-gold interface induces an orientation transition in the LC film. The dynamic behavior of the sensor response exhibits a concentration-dependent response rate that is employed to generate an algorithm for quantitative determination of unknown concentrations. Sensitive, selective and reversible detection with minimal effects of environmental fluctuations suggest that these sensors can be used for quantitative NO2 detection for a number of applications. PMID:23526230

  4. Liquid crystal-based sensors for selective and quantitative detection of nitrogen dioxide.

    PubMed

    Sen, Avijit; Kupcho, Kurt A; Grinwald, Bart A; Vantreeck, Heidi J; Acharya, Bharat R

    2013-03-01

    A highly sensitive nitrogen dioxide (NO2) sensor based on orientational transition of a thin film of liquid crystal (LC) supported on a gold surface is reported. Transport of NO2 molecules through the LC film to the LC-gold interface induces an orientation transition in the LC film. The dynamic behavior of the sensor response exhibits a concentration-dependent response rate that is employed to generate an algorithm for quantitative determination of unknown concentrations. Sensitive, selective and reversible detection with minimal effects of environmental fluctuations suggest that these sensors can be used for quantitative NO2 detection for a number of applications. PMID:23526230

  5. A quantitative epigenetic approach for the assessment of cigarette consumption

    PubMed Central

    Philibert, Robert; Hollenbeck, Nancy; Andersen, Eleanor; Osborn, Terry; Gerrard, Meg; Gibbons, Frederick X.; Wang, Kai

    2015-01-01

    Smoking is the largest preventable cause of morbidity and mortality in the world. Despite the development of numerous preventive and treatment interventions, the rate of daily smoking in the United States is still approximately 22%. Effective psychosocial interventions and pharmacologic agents exist for the prevention and treatment of smoking. Unfortunately, both approaches are hindered by our inability to accurately quantify amount of cigarette consumption from the point of initial experimentation to the point of total dependency. Recently, we and others have demonstrated that smoking is associated with genome-wide changes in DNA methylation. However, whether this advance in basic science can be employed as a reliable assay that is useful for clinical diagnosis and treatment has not been shown. In this communication, we determine the sensitivity and specificity of five of the most consistently replicated CpG loci with respect to smoking status using data from a publically available dataset. We show that methylation status at a CpG locus in the aryl hydrocarbon receptor repressor, cg05575921, is both sensitive and specific for smoking status in adults with a receiver operated curve characteristic area under the curve of 0.99. Given recent demonstrations that methylation at this locus reflects both intensity of smoking and the degree of smoking cessation, we conclude that a methylation-based diagnostic at this locus could have a prominent role in understanding the impact of new products, such as e-cigarettes on initiation of cigarette smoking among adolescents, while improving the prevention and treatment of smoking, and smoking related disorders. PMID:26082730

  6. Quantitative molecular detection of putative periodontal pathogens in clinically healthy and periodontally diseased subjects.

    PubMed

    Göhler, André; Hetzer, Adrian; Holtfreter, Birte; Geisel, Marie Henrike; Schmidt, Carsten Oliver; Steinmetz, Ivo; Kocher, Thomas

    2014-01-01

    Periodontitis is a multi-microbial oral infection with high prevalence among adults. Putative oral pathogens are commonly found in periodontally diseased individuals. However, these organisms can be also detected in the oral cavity of healthy subjects. This leads to the hypothesis, that alterations in the proportion of these organisms relative to the total amount of oral microorganisms, namely their abundance, rather than their simple presence might be important in the transition from health to disease. Therefore, we developed a quantitative molecular method to determine the abundance of various oral microorganisms and the portion of bacterial and archaeal nucleic acid relative to the total nucleic acid extracted from individual samples. We applied quantitative real-time PCRs targeting single-copy genes of periodontal bacteria and 16S-rRNA genes of Bacteria and Archaea. Testing tongue scrapings of 88 matched pairs of periodontally diseased and healthy subjects revealed a significantly higher abundance of P. gingivalis and a higher total bacterial abundance in diseased subjects. In fully adjusted models the risk of being periodontally diseased was significantly higher in subjects with high P. gingivalis and total bacterial abundance. Interestingly, we found that moderate abundances of A. actinomycetemcomitans were associated with reduced risk for periodontal disease compared to subjects with low abundances, whereas for high abundances, this protective effect leveled off. Moderate archaeal abundances were health associated compared to subjects with low abundances. In conclusion, our methodological approach unraveled associations of the oral flora with periodontal disease, which would have gone undetected if only qualitative data had been determined.

  7. Quantitative approaches for assessment of white matter hyperintensities in elderly populations

    PubMed Central

    Brickman, Adam M.; Sneed, Joel R.; Provenzano, Frank A.; Garcon, Ernst; Johnert, Lauren; Muraskin, Jordan; Yeung, Lok-Kin; Zimmerman, Molly E.; Roose, Steven P.

    2011-01-01

    White matter hyperintensities (WMH) are areas of increased signal on T2-weighted magnetic resonance imaging (MRI), including fluid attenuated inverse recovery sequences. Total and regional WMH burden (i.e., volume or severity) has been associated with myriad cognitive, neurological, and psychiatric conditions among older adults. In the current report, we illustrate two approaches to quantify periventricular, deep, and total WMH and examine their reliability and criterion validity among 28 elderly patients enrolled in a depression treatment trial. The first approach, an operator-driven quantitative approach, involves visual inspection of individual MRI scans and manual labeling using a three-step series of procedures. The second approach, a fully automated quantitative approach, uses a processing stream that involves image segmentation, voxel intensity thresholding, and seed growing to label WMH and calculate their volume automatically. There was good agreement in WMH quantification between the two approaches (Cronbach’s alpha values from 0.835 to 0.968). Further, severity of WMH was significantly associated with worse depression and increased age, and these associations did not differ significantly between the two quantification approaches. We provide evidence for good reliability and criterion validity for two approaches for WMH volume determination. The operator-driven approach may be better suited for smaller studies with highly trained raters, whereas the fully automated quantitative approach may be more appropriate for larger, high-throughput studies. PMID:21680159

  8. Surface-Enhanced Raman Scattering Active Plasmonic Nanoparticles with Ultrasmall Interior Nanogap for Multiplex Quantitative Detection and Cancer Cell Imaging.

    PubMed

    Li, Jiuxing; Zhu, Zhi; Zhu, Bingqing; Ma, Yanli; Lin, Bingqian; Liu, Rudi; Song, Yanling; Lin, Hui; Tu, Song; Yang, Chaoyong

    2016-08-01

    Due to its large enhancement effect, nanostructure-based surface-enhanced Raman scattering (SERS) technology had been widely applied for bioanalysis and cell imaging. However, most SERS nanostructures suffer from poor signal reproducibility, which hinders the application of SERS nanostructures in quantitative detection. We report an etching-assisted approach to synthesize SERS-active plasmonic nanoparticles with 1 nm interior nanogap for multiplex quantitative detection and cancer cell imaging. Raman dyes and methoxy poly(ethylene glycol) thiol (mPEG-SH) were attached to gold nanoparticles (AuNPs) to prepare gold cores. Next, Ag atoms were deposited on gold cores in the presence of Pluronic F127 to form a Ag shell. HAuCl4 was used to etch the Ag shell and form an interior nanogap in Au@AgAuNPs, leading to increased Raman intensity of dyes. SERS intensity distribution of Au@AgAuNPs was found to be more uniform than that of aggregated AuNPs. Finally, Au@AgAuNPs were used for multiplex quantitative detection and cancer cell imaging. With the advantages of simple and rapid preparation of Au@AgAuNPs with highly uniform, stable, and reproducible Raman intensity, the method reported here will widen the applications of SERS-active nanoparticles in diagnostics and imaging. PMID:27385563

  9. Tomographic thallium-201 myocardial perfusion scintigrams after maximal coronary artery vasodilation with intravenous dipyridamole: comparison of qualitative and quantitative approaches

    SciTech Connect

    Francisco, D.A.; Collins, S.M.; Go, R.T.; Ehrhardt, J.C.; Van Kirk, O.C.; Marcus, M.L.

    1982-08-01

    Eighty-six patients had thallium-201 (/sup 201/Tl) myocardial perfusion scintigrams after intense coronary artery dilation with i.v. dipyridamole. Tomographic and planar /sup 201/Tl scintigrams were obtained in each patient. Tomographic scintigrams were interpreted using quantitative or visual criteria; planar scintigrams were assessed using visual criteria only. When visual criteria were used, interobserver variability was 40% for tomographic scintigrams and 44% for planar scintigrams. In the 24 patients with normal or nonsignificant CAD, quantitative analysis of the tomograms (range approach) indicated that one of 24 (4%) had a positive image (specificity 96%%); in contrast, when visual criteria were used to interpret the tomographic or planar /sup 201/Tl scintigrams, eight of 24 (33%) had positive scintigrams (specificity 67%). In the 51 abnormal patients, the sensitivity of detecting CAD was 46 of 51 (90%) for tomographic scintigrams interpreted quantitatively, 39 of 51 (76%) for tomographic scintigrams interpreted visually and 41 of 51 (80%) for planar scintigrams assessed visually. The tomographic imaging procedure (quantitative interpretation) also demonstrated a high sensitivity (89%) and specificity (100%) in 28 patients (10 normal and 18 CAD), with a clinical diagnosis of unstable angina pectoris. Overall, the predictive accuracy of an abnormal scintigram with quantitative tomographic imaging (98%) was significantly better (p<0.05) than either qualitative planar or pinhole imaging. (JMT)

  10. Ranking the cost of environmental regulation among the several states: A quantitative approach

    NASA Astrophysics Data System (ADS)

    Blackstock, Robert Curtis, Jr.

    This dissertation uses a hedonic approach to measure the stringency of environmental regulation among the several States. Unlike other works which use a generalized approach, this dissertation concentrates on the production of SO2 emissions by the electricity producing industry. Using a quantitative rather than a normative approach, twenty-eight States are summarily ranked from the most, to the least stringent in the passing and subsequent enforcement of environmental regulations.

  11. QUANTITATIVE VS. CONVENTIONAL PCR FOR DETECTION OF HUMAN ADENOVIRUSES IN WATER AND SEDIMENT SAMPLES

    PubMed Central

    STAGGEMEIER, Rodrigo; BORTOLUZZI, Marina; HECK, Tatiana Moraes da Silva; SPILKI, Fernando Rosado; ALMEIDA, Sabrina Esteves de Matos

    2015-01-01

    SUMMARY Human Adenoviruses (HAdV) are notably resistant in the environment. These agents may serve as effective indicators of fecal contamination, and may act as causative agents of a number of different diseases in human beings. Conventional polymerase chain reaction (PCR) and, more recently, quantitative PCR (qPCR) are widely used for detection of viral agents in environmental matrices. In the present study PCR and SYBR(r)Green qPCR assays were compared for detection of HAdV in water (55) and sediments (20) samples of spring and artesian wells, ponds and streams, collected from dairy farms. By the quantitative methodology HAdV were detected in 87.3% of the water samples and 80% of the sediments, while by the conventional PCR 47.3% and 35% were detected in water samples and sediments, respectively. PMID:26422153

  12. QUANTITATIVE VS. CONVENTIONAL PCR FOR DETECTION OF HUMAN ADENOVIRUSES IN WATER AND SEDIMENT SAMPLES.

    PubMed

    Staggemeier, Rodrigo; Bortoluzzi, Marina; Heck, Tatiana Moraes da Silva; Spilki, Fernando Rosado; Almeida, Sabrina Esteves de Matos

    2015-01-01

    Human Adenoviruses (HAdV) are notably resistant in the environment. These agents may serve as effective indicators of fecal contamination, and may act as causative agents of a number of different diseases in human beings. Conventional polymerase chain reaction (PCR) and, more recently, quantitative PCR (qPCR) are widely used for detection of viral agents in environmental matrices. In the present study PCR and SYBR(r)Green qPCR assays were compared for detection of HAdV in water (55) and sediments (20) samples of spring and artesian wells, ponds and streams, collected from dairy farms. By the quantitative methodology HAdV were detected in 87.3% of the water samples and 80% of the sediments, while by the conventional PCR 47.3% and 35% were detected in water samples and sediments, respectively.

  13. Measurement issues associated with quantitative molecular biology analysis of complex food matrices for the detection of food fraud.

    PubMed

    Burns, Malcolm; Wiseman, Gordon; Knight, Angus; Bramley, Peter; Foster, Lucy; Rollinson, Sophie; Damant, Andrew; Primrose, Sandy

    2016-01-01

    Following a report on a significant amount of horse DNA being detected in a beef burger product on sale to the public at a UK supermarket in early 2013, the Elliott report was published in 2014 and contained a list of recommendations for helping ensure food integrity. One of the recommendations included improving laboratory testing capacity and capability to ensure a harmonised approach for testing for food authenticity. Molecular biologists have developed exquisitely sensitive methods based on the polymerase chain reaction (PCR) or mass spectrometry for detecting the presence of particular nucleic acid or peptide/protein sequences. These methods have been shown to be specific and sensitive in terms of lower limits of applicability, but they are largely qualitative in nature. Historically, the conversion of these qualitative techniques into reliable quantitative methods has been beset with problems even when used on relatively simple sample matrices. When the methods are applied to complex sample matrices, as found in many foods, the problems are magnified resulting in a high measurement uncertainty associated with the result which may mean that the assay is not fit for purpose. However, recent advances in the technology and the understanding of molecular biology approaches have further given rise to the re-assessment of these methods for their quantitative potential. This review focuses on important issues for consideration when validating a molecular biology assay and the various factors that can impact on the measurement uncertainty of a result associated with molecular biology approaches used in detection of food fraud, with a particular focus on quantitative PCR-based and proteomics assays.

  14. Measurement issues associated with quantitative molecular biology analysis of complex food matrices for the detection of food fraud.

    PubMed

    Burns, Malcolm; Wiseman, Gordon; Knight, Angus; Bramley, Peter; Foster, Lucy; Rollinson, Sophie; Damant, Andrew; Primrose, Sandy

    2016-01-01

    Following a report on a significant amount of horse DNA being detected in a beef burger product on sale to the public at a UK supermarket in early 2013, the Elliott report was published in 2014 and contained a list of recommendations for helping ensure food integrity. One of the recommendations included improving laboratory testing capacity and capability to ensure a harmonised approach for testing for food authenticity. Molecular biologists have developed exquisitely sensitive methods based on the polymerase chain reaction (PCR) or mass spectrometry for detecting the presence of particular nucleic acid or peptide/protein sequences. These methods have been shown to be specific and sensitive in terms of lower limits of applicability, but they are largely qualitative in nature. Historically, the conversion of these qualitative techniques into reliable quantitative methods has been beset with problems even when used on relatively simple sample matrices. When the methods are applied to complex sample matrices, as found in many foods, the problems are magnified resulting in a high measurement uncertainty associated with the result which may mean that the assay is not fit for purpose. However, recent advances in the technology and the understanding of molecular biology approaches have further given rise to the re-assessment of these methods for their quantitative potential. This review focuses on important issues for consideration when validating a molecular biology assay and the various factors that can impact on the measurement uncertainty of a result associated with molecular biology approaches used in detection of food fraud, with a particular focus on quantitative PCR-based and proteomics assays. PMID:26631264

  15. Quantitative and qualitative approaches in the study of poverty and adolescent development: separation or integration?

    PubMed

    Leung, Janet T Y; Shek, Daniel T L

    2011-01-01

    This paper examines the use of quantitative and qualitative approaches to study the impact of economic disadvantage on family processes and adolescent development. Quantitative research has the merits of objectivity, good predictive and explanatory power, parsimony, precision and sophistication of analysis. Qualitative research, in contrast, provides a detailed, holistic, in-depth understanding of social reality and allows illumination of new insights. With the pragmatic considerations of methodological appropriateness, design flexibility, and situational responsiveness in responding to the research inquiry, a mixed methods approach could be a possibility of integrating quantitative and qualitative approaches and offers an alternative strategy to study the impact of economic disadvantage on family processes and adolescent development. PMID:21870673

  16. QUANTITATIVE DETECTION OF ENVIRONMENTALLY IMPORTANT DYES USING DIODE LASER/FIBER-OPTIC RAMAN

    EPA Science Inventory

    A compact diode laser/fiber-optic Raman spectrometer is used for quantitative detection of environmentally important dyes. This system is based on diode laser excitation at 782 mm, fiber optic probe technology, an imaging spectrometer, and state-of-the-art scientific CCD camera. ...

  17. NAIMA: target amplification strategy allowing quantitative on-chip detection of GMOs.

    PubMed

    Morisset, Dany; Dobnik, David; Hamels, Sandrine; Zel, Jana; Gruden, Kristina

    2008-10-01

    We have developed a novel multiplex quantitative DNA-based target amplification method suitable for sensitive, specific and quantitative detection on microarray. This new method named NASBA Implemented Microarray Analysis (NAIMA) was applied to GMO detection in food and feed, but its application can be extended to all fields of biology requiring simultaneous detection of low copy number DNA targets. In a first step, the use of tailed primers allows the multiplex synthesis of template DNAs in a primer extension reaction. A second step of the procedure consists of transcription-based amplification using universal primers. The cRNA product is further on directly ligated to fluorescent dyes labelled 3DNA dendrimers allowing signal amplification and hybridized without further purification on an oligonucleotide probe-based microarray for multiplex detection. Two triplex systems have been applied to test maize samples containing several transgenic lines, and NAIMA has shown to be sensitive down to two target copies and to provide quantitative data on the transgenic contents in a range of 0.1-25%. Performances of NAIMA are comparable to singleplex quantitative real-time PCR. In addition, NAIMA amplification is faster since 20 min are sufficient to achieve full amplification. PMID:18710880

  18. Quantitative detection of Listeria monocytogenes in biofilms by real-time PCR.

    PubMed

    Guilbaud, Morgan; de Coppet, Pierre; Bourion, Fabrice; Rachman, Cinta; Prévost, Hervé; Dousset, Xavier

    2005-04-01

    A quantitative method based on a real-time PCR assay to enumerate Listeria monocytogenes in biofilms was developed. The specificity for L. monocytogenes of primers targeting the listeriolysin gene was demonstrated using a SYBR Green I real-time PCR assay. The number of L. monocytogenes detected growing in biofilms was 6 x 10(2) CFU/cm2.

  19. Quantitative PCR for Detection and Enumeration of Genetic Markers of Bovine Fecal Pollution

    EPA Science Inventory

    Accurate assessment of health risks associated with bovine (cattle) fecal pollution requires a reliable host-specific genetic marker and a rapid quantification method. We report the development of quantitative PCR assays for the detection of two recently described cow feces-spec...

  20. NAIMA: target amplification strategy allowing quantitative on-chip detection of GMOs.

    PubMed

    Morisset, Dany; Dobnik, David; Hamels, Sandrine; Zel, Jana; Gruden, Kristina

    2008-10-01

    We have developed a novel multiplex quantitative DNA-based target amplification method suitable for sensitive, specific and quantitative detection on microarray. This new method named NASBA Implemented Microarray Analysis (NAIMA) was applied to GMO detection in food and feed, but its application can be extended to all fields of biology requiring simultaneous detection of low copy number DNA targets. In a first step, the use of tailed primers allows the multiplex synthesis of template DNAs in a primer extension reaction. A second step of the procedure consists of transcription-based amplification using universal primers. The cRNA product is further on directly ligated to fluorescent dyes labelled 3DNA dendrimers allowing signal amplification and hybridized without further purification on an oligonucleotide probe-based microarray for multiplex detection. Two triplex systems have been applied to test maize samples containing several transgenic lines, and NAIMA has shown to be sensitive down to two target copies and to provide quantitative data on the transgenic contents in a range of 0.1-25%. Performances of NAIMA are comparable to singleplex quantitative real-time PCR. In addition, NAIMA amplification is faster since 20 min are sufficient to achieve full amplification.

  1. NAIMA: target amplification strategy allowing quantitative on-chip detection of GMOs

    PubMed Central

    Morisset, Dany; Dobnik, David; Hamels, Sandrine; Žel, Jana; Gruden, Kristina

    2008-01-01

    We have developed a novel multiplex quantitative DNA-based target amplification method suitable for sensitive, specific and quantitative detection on microarray. This new method named NASBA Implemented Microarray Analysis (NAIMA) was applied to GMO detection in food and feed, but its application can be extended to all fields of biology requiring simultaneous detection of low copy number DNA targets. In a first step, the use of tailed primers allows the multiplex synthesis of template DNAs in a primer extension reaction. A second step of the procedure consists of transcription-based amplification using universal primers. The cRNA product is further on directly ligated to fluorescent dyes labelled 3DNA dendrimers allowing signal amplification and hybridized without further purification on an oligonucleotide probe-based microarray for multiplex detection. Two triplex systems have been applied to test maize samples containing several transgenic lines, and NAIMA has shown to be sensitive down to two target copies and to provide quantitative data on the transgenic contents in a range of 0.1–25%. Performances of NAIMA are comparable to singleplex quantitative real-time PCR. In addition, NAIMA amplification is faster since 20 min are sufficient to achieve full amplification. PMID:18710880

  2. Single Laboratory Comparison of Quantitative Real-time PCR Assays for the Detection of Fecal Pollution

    EPA Science Inventory

    There are numerous quantitative real-time PCR (qPCR) assays available to detect and enumerate fecal pollution in ambient waters. Each assay employs distinct primers and probes that target different rRNA genes and microorganisms leading to potential variations in concentration es...

  3. Highly sensitive qualitative and quantitative detection of reverse transcriptase activity: optimization, validation, and comparative analysis with other detection systems.

    PubMed

    Yamamoto, S; Folks, T M; Heneine, W

    1996-09-01

    An ultra-sensitive assay for reverse transcriptase (RT) activity called Amp-RT has been developed. An in vitro transcribed heteropolymeric RNA sequence was used as a template, and polymerase chain reaction (PCR) amplification with Southern-blot hybridization served as a detection system for the cDNA product of the reaction. Titration of Mg2+ and Mn2+ concentrations using the human immunodeficiency virus type 1 (HIV-1) and the human T lymphotropic virus type 1 (HTLV-I), respectively, showed optimal assay reactivity for both viruses at 2-20 mM of Mg2+. Analysis of density banded HIV-1 showed that the peak RT activity of the assay was associated with the fractions consistent with retrovirus particles. The sensitivity of Amp-RT was also compared with that of three conventional RT assays by using seven different retroviruses including HIV-1, simian immunodeficiency virus (SIV), caprine arthritis-encephalitis virus (CAEV), HTLV-I and HTLV-II, simian retrovirus type 2 (SRV-2), and gibbon ape leukemia virus (GALV). HTLV-I, HTLV-II, and GALV could not be detected by the three conventional RT assays. Amp-RT was able to detect all these viruses in 10(1)-10(3)-fold dilutions. Similarly, Amp-RT was found to be 10(3)-10(6)-fold more sensitive than the other RT assays in detecting HIV-1, SIV< or CAEV. Culture supernatants from uninfected cell lines were all Amp-RT negative. A quantitative Amp-RT assay was also developed by using recombinant HIV-1 RT and signal quantitation. The assay was found to have a 5 log linear range, and therefore, provides a useful tool for quantitating RT and retroviruses. Amp-RT offers a sensitive generic tool for the qualitative and quantitative detection of known and unknown retroviruses.

  4. Application of quantitative PCR for the detection of microorganisms in water.

    PubMed

    Botes, Marelize; de Kwaadsteniet, Michéle; Cloete, Thomas Eugene

    2013-01-01

    The occurrence of microorganisms in water due to contamination is a health risk and control thereof is a necessity. Conventional detection methods may be misleading and do not provide rapid results allowing for immediate action. The quantitative polymerase chain reaction (qPCR) method has proven to be an effective tool to detect and quantify microorganisms in water within a few hours. Quantitative PCR assays have recently been developed for the detection of specific adeno- and polyomaviruses, bacteria and protozoa in different water sources. The technique is highly sensitive and able to detect low numbers of microorganisms. Quantitative PCR can be applied for microbial source tracking in water sources, to determine the efficiency of water and wastewater treatment plants and act as a tool for risk assessment. Different qPCR assays exist depending on whether an internal control is used or whether measurements are taken at the end of the PCR reaction (end-point qPCR) or in the exponential phase (real-time qPCR). Fluorescent probes are used in the PCR reaction to hybridise within the target sequence to generate a signal and, together with specialised systems, quantify the amount of PCR product. Quantitative reverse transcription polymerase chain reaction (q-RT-PCR) is a more sensitive technique that detects low copy number RNA and can be applied to detect, e.g. enteric viruses and viable microorganisms in water, and measure specific gene expression. There is, however, a need to standardise qPCR protocols if this technique is to be used as an analytical diagnostic tool for routine monitoring. This review focuses on the application of qPCR in the detection of microorganisms in water.

  5. Quantitative risk assessment & leak detection criteria for a subsea oil export pipeline

    NASA Astrophysics Data System (ADS)

    Zhang, Fang-Yuan; Bai, Yong; Badaruddin, Mohd Fauzi; Tuty, Suhartodjo

    2009-06-01

    A quantitative risk assessment (QRA) based on leak detection criteria (LDC) for the design of a proposed subsea oil export pipeline is presented in this paper. The objective of this QRA/LDC study was to determine if current leak detection methodologies were sufficient, based on QRA results, while excluding the use of statistical leak detection; if not, an appropriate LDC for the leak detection system would need to be established. The famous UK PARLOC database was used for the calculation of pipeline failure rates, and the software POSVCM from MMS was used for oil spill simulations. QRA results revealed that the installation of a statistically based leak detection system (LDS) can significantly reduce time to leak detection, thereby mitigating the consequences of leakage. A sound LDC has been defined based on QRA study results and comments from various LDS vendors to assist the emergency response team (ERT) to quickly identify and locate leakage and employ the most effective measures to contain damage.

  6. Qualitative and Quantitative Detection of Botulinum Neurotoxins from Complex Matrices: Results of the First International Proficiency Test.

    PubMed

    Worbs, Sylvia; Fiebig, Uwe; Zeleny, Reinhard; Schimmel, Heinz; Rummel, Andreas; Luginbühl, Werner; Dorner, Brigitte G

    2015-12-01

    In the framework of the EU project EQuATox, a first international proficiency test (PT) on the detection and quantification of botulinum neurotoxins (BoNT) was conducted. Sample materials included BoNT serotypes A, B and E spiked into buffer, milk, meat extract and serum. Different methods were applied by the participants combining different principles of detection, identification and quantification. Based on qualitative assays, 95% of all results reported were correct. Successful strategies for BoNT detection were based on a combination of complementary immunological, MS-based and functional methods or on suitable functional in vivo/in vitro approaches (mouse bioassay, hemidiaphragm assay and Endopep-MS assay). Quantification of BoNT/A, BoNT/B and BoNT/E was performed by 48% of participating laboratories. It turned out that precise quantification of BoNT was difficult, resulting in a substantial scatter of quantitative data. This was especially true for results obtained by the mouse bioassay which is currently considered as "gold standard" for BoNT detection. The results clearly demonstrate the urgent need for certified BoNT reference materials and the development of methods replacing animal testing. In this context, the BoNT PT provided the valuable information that both the Endopep-MS assay and the hemidiaphragm assay delivered quantitative results superior to the mouse bioassay. PMID:26703724

  7. Qualitative and Quantitative Detection of Botulinum Neurotoxins from Complex Matrices: Results of the First International Proficiency Test

    PubMed Central

    Worbs, Sylvia; Fiebig, Uwe; Zeleny, Reinhard; Schimmel, Heinz; Rummel, Andreas; Luginbühl, Werner; Dorner, Brigitte G.

    2015-01-01

    In the framework of the EU project EQuATox, a first international proficiency test (PT) on the detection and quantification of botulinum neurotoxins (BoNT) was conducted. Sample materials included BoNT serotypes A, B and E spiked into buffer, milk, meat extract and serum. Different methods were applied by the participants combining different principles of detection, identification and quantification. Based on qualitative assays, 95% of all results reported were correct. Successful strategies for BoNT detection were based on a combination of complementary immunological, MS-based and functional methods or on suitable functional in vivo/in vitro approaches (mouse bioassay, hemidiaphragm assay and Endopep-MS assay). Quantification of BoNT/A, BoNT/B and BoNT/E was performed by 48% of participating laboratories. It turned out that precise quantification of BoNT was difficult, resulting in a substantial scatter of quantitative data. This was especially true for results obtained by the mouse bioassay which is currently considered as “gold standard” for BoNT detection. The results clearly demonstrate the urgent need for certified BoNT reference materials and the development of methods replacing animal testing. In this context, the BoNT PT provided the valuable information that both the Endopep-MS assay and the hemidiaphragm assay delivered quantitative results superior to the mouse bioassay. PMID:26703724

  8. Qualitative and Quantitative Detection of Botulinum Neurotoxins from Complex Matrices: Results of the First International Proficiency Test.

    PubMed

    Worbs, Sylvia; Fiebig, Uwe; Zeleny, Reinhard; Schimmel, Heinz; Rummel, Andreas; Luginbühl, Werner; Dorner, Brigitte G

    2015-11-26

    In the framework of the EU project EQuATox, a first international proficiency test (PT) on the detection and quantification of botulinum neurotoxins (BoNT) was conducted. Sample materials included BoNT serotypes A, B and E spiked into buffer, milk, meat extract and serum. Different methods were applied by the participants combining different principles of detection, identification and quantification. Based on qualitative assays, 95% of all results reported were correct. Successful strategies for BoNT detection were based on a combination of complementary immunological, MS-based and functional methods or on suitable functional in vivo/in vitro approaches (mouse bioassay, hemidiaphragm assay and Endopep-MS assay). Quantification of BoNT/A, BoNT/B and BoNT/E was performed by 48% of participating laboratories. It turned out that precise quantification of BoNT was difficult, resulting in a substantial scatter of quantitative data. This was especially true for results obtained by the mouse bioassay which is currently considered as "gold standard" for BoNT detection. The results clearly demonstrate the urgent need for certified BoNT reference materials and the development of methods replacing animal testing. In this context, the BoNT PT provided the valuable information that both the Endopep-MS assay and the hemidiaphragm assay delivered quantitative results superior to the mouse bioassay.

  9. A New Approach for Quantitative Evaluation of Ultrasonic Wave Attenuation in Composites

    NASA Astrophysics Data System (ADS)

    Ni, Qing-Qing; Li, Ran; Xia, Hong

    2016-06-01

    When ultrasonic waves propagate in composite materials, the propagation behaviors result from the combination effects of various factors, such as material anisotropy and viscoelastic property, internal microstructure and defects, incident wave characteristics and interface condition between composite components. It is essential to make it clear how these factors affect the ultrasonic wave propagation and attenuation characteristics, and how they mutually interact on each other. In the present paper, based on a newly developed time-domain finite element analysis code, PZflex, a unique approach for clarifying the detailed influence mechanism of aforementioned factors is proposed, in which each attenuation component can be extracted from the overall attenuation and analyzed respectively. By taking into consideration the interrelation between each individual attenuation component, the variation behaviors of each component and internal dynamic stress distribution against material anisotropy and matrix viscosity are separately and quantitatively evaluated. From the detailed analysis results of each attenuation component, the energy dissipation at interface is a major component in ultrasonic wave attenuation characteristics, which can provide a maximum contribution rate of 68.2 % to the overall attenuation, and each attenuation component is closely related to the material anisotropy and viscoelasticity. The results clarify the correlation between ultrasonic wave propagation characteristics and material viscoelastic properties, which will be useful in the further development of ultrasonic technology in defect detection.

  10. Quantitative PCR for detection of DNA damage in mitochondrial DNA of the fission yeast Schizosaccharomyces pombe.

    PubMed

    Senoo, Takanori; Yamanaka, Mayumi; Nakamura, Atori; Terashita, Tomoki; Kawano, Shinji; Ikeda, Shogo

    2016-08-01

    Quantitative polymerase chain reaction (QPCR) has been employed to detect DNA damage and repair in mitochondrial DNA (mtDNA) of human and several model organisms. The assay also permits the quantitation of relative mtDNA copy number in cells. Here, we developed the QPCR assay primers and reaction conditions for the fission yeast Schizosaccharomyces pombe, an important model of eukaryote biology, not previously described. Under these conditions, long targets (approximately 10kb) in mtDNA were quantitatively amplified using 0.1ng of crude DNA templates without isolation of mitochondria and mtDNA. Quantitative detection of oxidative DNA damage in mtDNA was illustrated by using a DNA template irradiated with UVA in the presence of riboflavin. The damage to mtDNA in S. pombe cells treated with hydrogen peroxide and paraquat was also quantitatively measured. Finally, we found that mtDNA copy number in S. pombe cells increased after transition into a stationary phase and that the damage to mtDNA due to endogenous cellular processes accumulated during chronological aging.

  11. Qualitative and quantitative detection of agricultural microorganisms expressing iturin and mop cyclase in soils.

    PubMed

    Kim, Sung Eun; Moon, Jae Sun; Choi, Won Sik; Lee, Eun Na; Lee, Sang Han; Kim, Sung Uk

    2010-12-22

    The environmental release of genetically engineered microorganisms (GEMs) to improve agriculture or remediate environmental hazards has raised concern over the fate of the organisms and their engineered genes. To detect the microorganisms released into the environment at the molecular level, Bacillus subtilis KB producing iturin and Pseudomonas fluorescens MX1 carrying the moc (mannityl opine catabolism) region from the Agrobacterium tumefaciens were employed as model microorganisms. Using specific fusion primers and the TaqMan probes, qualitative and quantitative detections of the model organisms by PCR and real-time PCR were conducted employing a small-scale soil-core device and pots during the six month period. The data indicate that the model bacteria can be easily detected by qualitative and quantitative methods in the test systems employed, and they do not give significant impacts on the other bacteria in soils on the Southern blotting analysis, although long-term observation may be needed.

  12. Detection of nonauthorized genetically modified organisms using differential quantitative polymerase chain reaction: application to 35S in maize.

    PubMed

    Cankar, Katarina; Chauvensy-Ancel, Valérie; Fortabat, Marie-Noelle; Gruden, Kristina; Kobilinsky, André; Zel, Jana; Bertheau, Yves

    2008-05-15

    Detection of nonauthorized genetically modified organisms (GMOs) has always presented an analytical challenge because the complete sequence data needed to detect them are generally unavailable although sequence similarity to known GMOs can be expected. A new approach, differential quantitative polymerase chain reaction (PCR), for detection of nonauthorized GMOs is presented here. This method is based on the presence of several common elements (e.g., promoter, genes of interest) in different GMOs. A statistical model was developed to study the difference between the number of molecules of such a common sequence and the number of molecules identifying the approved GMO (as determined by border-fragment-based PCR) and the donor organism of the common sequence. When this difference differs statistically from zero, the presence of a nonauthorized GMO can be inferred. The interest and scope of such an approach were tested on a case study of different proportions of genetically modified maize events, with the P35S promoter as the Cauliflower Mosaic Virus common sequence. The presence of a nonauthorized GMO was successfully detected in the mixtures analyzed and in the presence of (donor organism of P35S promoter). This method could be easily transposed to other common GMO sequences and other species and is applicable to other detection areas such as microbiology.

  13. Detection of nonauthorized genetically modified organisms using differential quantitative polymerase chain reaction: application to 35S in maize.

    PubMed

    Cankar, Katarina; Chauvensy-Ancel, Valérie; Fortabat, Marie-Noelle; Gruden, Kristina; Kobilinsky, André; Zel, Jana; Bertheau, Yves

    2008-05-15

    Detection of nonauthorized genetically modified organisms (GMOs) has always presented an analytical challenge because the complete sequence data needed to detect them are generally unavailable although sequence similarity to known GMOs can be expected. A new approach, differential quantitative polymerase chain reaction (PCR), for detection of nonauthorized GMOs is presented here. This method is based on the presence of several common elements (e.g., promoter, genes of interest) in different GMOs. A statistical model was developed to study the difference between the number of molecules of such a common sequence and the number of molecules identifying the approved GMO (as determined by border-fragment-based PCR) and the donor organism of the common sequence. When this difference differs statistically from zero, the presence of a nonauthorized GMO can be inferred. The interest and scope of such an approach were tested on a case study of different proportions of genetically modified maize events, with the P35S promoter as the Cauliflower Mosaic Virus common sequence. The presence of a nonauthorized GMO was successfully detected in the mixtures analyzed and in the presence of (donor organism of P35S promoter). This method could be easily transposed to other common GMO sequences and other species and is applicable to other detection areas such as microbiology. PMID:18346452

  14. Statistical modeling approach for detecting generalized synchronization

    NASA Astrophysics Data System (ADS)

    Schumacher, Johannes; Haslinger, Robert; Pipa, Gordon

    2012-05-01

    Detecting nonlinear correlations between time series presents a hard problem for data analysis. We present a generative statistical modeling method for detecting nonlinear generalized synchronization. Truncated Volterra series are used to approximate functional interactions. The Volterra kernels are modeled as linear combinations of basis splines, whose coefficients are estimated via l1 and l2 regularized maximum likelihood regression. The regularization manages the high number of kernel coefficients and allows feature selection strategies yielding sparse models. The method's performance is evaluated on different coupled chaotic systems in various synchronization regimes and analytical results for detecting m:n phase synchrony are presented. Experimental applicability is demonstrated by detecting nonlinear interactions between neuronal local field potentials recorded in different parts of macaque visual cortex.

  15. Detection of Prostate Cancer: Quantitative Multiparametric MR Imaging Models Developed Using Registered Correlative Histopathology.

    PubMed

    Metzger, Gregory J; Kalavagunta, Chaitanya; Spilseth, Benjamin; Bolan, Patrick J; Li, Xiufeng; Hutter, Diane; Nam, Jung W; Johnson, Andrew D; Henriksen, Jonathan C; Moench, Laura; Konety, Badrinath; Warlick, Christopher A; Schmechel, Stephen C; Koopmeiners, Joseph S

    2016-06-01

    Purpose To develop multiparametric magnetic resonance (MR) imaging models to generate a quantitative, user-independent, voxel-wise composite biomarker score (CBS) for detection of prostate cancer by using coregistered correlative histopathologic results, and to compare performance of CBS-based detection with that of single quantitative MR imaging parameters. Materials and Methods Institutional review board approval and informed consent were obtained. Patients with a diagnosis of prostate cancer underwent multiparametric MR imaging before surgery for treatment. All MR imaging voxels in the prostate were classified as cancer or noncancer on the basis of coregistered histopathologic data. Predictive models were developed by using more than one quantitative MR imaging parameter to generate CBS maps. Model development and evaluation of quantitative MR imaging parameters and CBS were performed separately for the peripheral zone and the whole gland. Model accuracy was evaluated by using the area under the receiver operating characteristic curve (AUC), and confidence intervals were calculated with the bootstrap procedure. The improvement in classification accuracy was evaluated by comparing the AUC for the multiparametric model and the single best-performing quantitative MR imaging parameter at the individual level and in aggregate. Results Quantitative T2, apparent diffusion coefficient (ADC), volume transfer constant (K(trans)), reflux rate constant (kep), and area under the gadolinium concentration curve at 90 seconds (AUGC90) were significantly different between cancer and noncancer voxels (P < .001), with ADC showing the best accuracy (peripheral zone AUC, 0.82; whole gland AUC, 0.74). Four-parameter models demonstrated the best performance in both the peripheral zone (AUC, 0.85; P = .010 vs ADC alone) and whole gland (AUC, 0.77; P = .043 vs ADC alone). Individual-level analysis showed statistically significant improvement in AUC in 82% (23 of 28) and 71% (24 of 34

  16. Fraud detection in medicare claims: A multivariate outlier detection approach

    SciTech Connect

    Burr, T.; Hale, C.; Kantor, M.

    1997-04-01

    We apply traditional and customized multivariate outlier detection methods to detect fraud in medicare claims. We use two sets of 11 derived features, and one set of the 22 combined features. The features are defined so that fraudulent medicare providers should tend to have larger features values than non-fraudulent providers. Therefore we have an apriori direction ({open_quotes}large values{close_quotes}) in high dimensional feature space to search for the multivariate outliers. We focus on three issues: (1) outlier masking (Example: the presence of one outlier can make it difficult to detect a second outlier), (2) the impact of having an apriori direction to search for fraud, and (3) how to compare our detection methods. Traditional methods include Mahalanobis distances, (with and without dimension reduction), k-nearest neighbor, and density estimation methods. Some methods attempt to mitigate the outlier masking problem (for example: minimum volume ellipsoid covariance estimator). Customized methods include ranking methods (such as Spearman rank ordering) that exploit the {open_quotes}large is suspicious{close_quotes} notion. No two methods agree completely which providers are most suspicious so we present ways to compare our methods. One comparison method uses a list of known-fraudulent providers. All comparison methods restrict attention to the most suspicious providers.

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

    NASA Astrophysics Data System (ADS)

    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

  18. Qualitative and Quantitative Approaches to the Study of Poverty: Taming the Tensions and Appreciating the Complementarities

    ERIC Educational Resources Information Center

    Balarabe Kura, Sulaiman Y.

    2012-01-01

    There is a germane relationship between qualitative and quantitative approaches to social science research. The relationship is empirically and theoretically demonstrated by poverty researchers. The study of poverty, as argued in this article, is a study of both numbers and contextualities. This article provides a general overview of qualitative…

  19. A Novel Approach to Teach the Generation of Bioelectrical Potentials from a Descriptive and Quantitative Perspective

    ERIC Educational Resources Information Center

    Rodriguez-Falces, Javier

    2013-01-01

    In electrophysiology studies, it is becoming increasingly common to explain experimental observations using both descriptive methods and quantitative approaches. However, some electrophysiological phenomena, such as the generation of extracellular potentials that results from the propagation of the excitation source along the muscle fiber, are…

  20. Pro-Social Behavior Amongst Students of Tertiary Institutions: An Explorative and a Quantitative Approach

    ERIC Educational Resources Information Center

    Quain, Samuel; Yidana, Xiaaba Dantallah; Ambotumah, Bernard Baba; Mensah-Livivnstone, Ike Joe Nii Annang

    2016-01-01

    The purpose of this paper was to explore antecedents of pro-social behavior amongst university students, using a private university as a case study. Following an explorative research, the study was guided by some theories relating to the phenomenon, focusing on gender and location factors. A quantitative approach was used in the follow up to the…

  1. A Quantitative Corpus-Based Approach to English Spatial Particles: Conceptual Symmetry and Its Pedagogical Implications

    ERIC Educational Resources Information Center

    Chen, Alvin Cheng-Hsien

    2014-01-01

    The present study aims to investigate how conceptual symmetry plays a role in the use of spatial particles in English and to further examine its pedagogical implications via a corpus-based evaluation of the course books in senior high schools in Taiwan. More specifically, we adopt a quantitative corpus-based approach to investigate whether bipolar…

  2. Poem Generator: A Comparative Quantitative Evaluation of a Microworlds-Based Learning Approach for Teaching English

    ERIC Educational Resources Information Center

    Jenkins, Craig

    2015-01-01

    This paper is a comparative quantitative evaluation of an approach to teaching poetry in the subject domain of English that employs a "guided discovery" pedagogy using computer-based microworlds. It uses a quasi-experimental design in order to measure performance gains in computational thinking and poetic thinking following a…

  3. Rapid and quantitative detection of C-reactive protein based on quantum dots and immunofiltration assay

    PubMed Central

    Zhang, Pengfei; Bao, Yan; Draz, Mohamed Shehata; Lu, Huiqi; Liu, Chang; Han, Huanxing

    2015-01-01

    Convenient and rapid immunofiltration assays (IFAs) enable on-site “yes” or “no” determination of disease markers. However, traditional IFAs are commonly qualitative or semi-quantitative and are very limited for the efficient testing of samples in field diagnostics. Here, we overcome these limitations by developing a quantum dots (QDs)-based fluorescent IFA for the quantitative detection of C-reactive proteins (CRP). CRP, the well-known diagnostic marker for acute viral and bacterial infections, was used as a model analyte to demonstrate performance and sensitivity of our developed QDs-based IFA. QDs capped with both polyethylene glycol (PEG) and glutathione were used as fluorescent labels for our IFAs. The presence of the surface PEG layer, which reduced the non-specific protein interactions, in conjunction with the inherent optical properties of QDs, resulted in lower background signal, increased sensitivity, and ability to detect CRP down to 0.79 mg/L with only 5 µL serum sample. In addition, the developed assay is simple, fast and can quantitatively detect CRP with a detection limit up to 200 mg/L. Clinical test results of our QD-based IFA are well correlated with the traditional latex enhance immune-agglutination aggregation. The proposed QD-based fluorescent IFA is very promising, and potentially will be adopted for multiplexed immunoassay and in field point-of-care test. PMID:26491289

  4. Broad-spectrum detection and quantitation methods of Soil-borne cereal mosaic virus isolates.

    PubMed

    Vaïanopoulos, Céline; Legrève, Anne; Moreau, Virginie; Bragard, Claude

    2009-08-01

    A broad-spectrum reverse transcription-polymerase chain reaction (RT-PCR) protocol was developed for detecting Soil-borne cereal mosaic virus (SBCMV) isolates, responsible for mosaic diseases in Europe, using primers targeting the highly conserved 3'-untranslated region of RNA-1 and RNA-2 of SBCMV. The 3'-end region is a privileged target for the detection of a wide range of isolates, because of sequence conservation, of the tRNA-like structure, the major role in viral replication and the signal amplification due to the presence of numerous genomic and subgenomic RNAs. The primers were also designed for virus quantitation using real-time RT-PCR with SYBR-Green chemistry. No cross-reaction with Wheat spindle streak mosaic virus, frequently associated with SBCMV, was observed. The use of RT-PCR and real-time quantitative RT-PCR allowed a more sensitive detection and quantitation of SBCMV to be made than was the case with ELISA. The methods enabled European isolates of SBCMV from Belgium, France, Germany, Italy and the UK to be detected and quantified. Real-time RT-PCR represents a new tool for comparing soil inoculum potential as well as cultivar resistance to SBCMV.

  5. Rapid and quantitative detection of C-reactive protein based on quantum dots and immunofiltration assay.

    PubMed

    Zhang, Pengfei; Bao, Yan; Draz, Mohamed Shehata; Lu, Huiqi; Liu, Chang; Han, Huanxing

    2015-01-01

    Convenient and rapid immunofiltration assays (IFAs) enable on-site "yes" or "no" determination of disease markers. However, traditional IFAs are commonly qualitative or semi-quantitative and are very limited for the efficient testing of samples in field diagnostics. Here, we overcome these limitations by developing a quantum dots (QDs)-based fluorescent IFA for the quantitative detection of C-reactive proteins (CRP). CRP, the well-known diagnostic marker for acute viral and bacterial infections, was used as a model analyte to demonstrate performance and sensitivity of our developed QDs-based IFA. QDs capped with both polyethylene glycol (PEG) and glutathione were used as fluorescent labels for our IFAs. The presence of the surface PEG layer, which reduced the non-specific protein interactions, in conjunction with the inherent optical properties of QDs, resulted in lower background signal, increased sensitivity, and ability to detect CRP down to 0.79 mg/L with only 5 µL serum sample. In addition, the developed assay is simple, fast and can quantitatively detect CRP with a detection limit up to 200 mg/L. Clinical test results of our QD-based IFA are well correlated with the traditional latex enhance immune-agglutination aggregation. The proposed QD-based fluorescent IFA is very promising, and potentially will be adopted for multiplexed immunoassay and in field point-of-care test.

  6. Object-Oriented Change Detection Based on Multi-Scale Approach

    NASA Astrophysics Data System (ADS)

    Jia, Yonghong; Zhou, Mingting; Jinshan, Ye

    2016-06-01

    The change detection of remote sensing images means analysing the change information quantitatively and recognizing the change types of the surface coverage data in different time phases. With the appearance of high resolution remote sensing image, object-oriented change detection method arises at this historic moment. In this paper, we research multi-scale approach for high resolution images, which includes multi-scale segmentation, multi-scale feature selection and multi-scale classification. Experimental results show that this method has a stronger advantage than the traditional single-scale method of high resolution remote sensing image change detection.

  7. Noninvasive imaging of hemoglobin concentration and oxygen saturation for detection of osteoarthritis in the finger joints using multispectral three-dimensional quantitative photoacoustic tomography

    NASA Astrophysics Data System (ADS)

    Sun, Yao; Sobel, Eric; Jiang, Huabei

    2013-05-01

    We present quantitative imaging of hemoglobin concentration and oxygen saturation in in vivo finger joints and evaluate the feasibility of detecting osteoarthritis (OA) in the hand using three-dimensional (3D) multispectral quantitative photoacoustic tomography (3D qPAT). The results show that both the anatomical structures and quantitative chromophore concentrations (oxy-hemoglobin and deoxy-hemoglobin) of different joint tissues (hard phalanges and soft cartilage/synovial fluid between phalanges) can be imaged in vivo with the multispectral 3D qPAT. Enhanced hemoglobin concentrations and dropped oxygen saturations in osteoarthritic phalanges and soft joint tissues in joint cavities have been observed. This study indicates that the multispectral 3D qPAT is a promising approach to detect the angiogenesis and hypoxia associated with OA disease and a potential clinical tool for early OA detection in the finger joints.

  8. A combined algorithm for T-wave alternans qualitative detection and quantitative measurement

    PubMed Central

    2013-01-01

    Background T-wave alternans (TWA) provides a noninvasive and clinically useful marker for the risk of sudden cardiac death (SCD). Current most widely used TWA detection algorithms work in two different domains: time and frequency. The disadvantage of the spectral analytical techniques is that they treat the alternans signal as a stationary wave with a constant amplitude and a phase. They cannot detect non-stationary characteristics of the signal. The temporal domain methods are sensitive to the alignment of the T-waves. In this study, we sought to develop a robust combined algorithm (CA) to assess T-wave alternans, which can qualitatively detect and quantitatively measure TWA in time domain. Methods The T wave sequences were extracted and the total energy of each T wave within the specified time-frequency region was calculated. The rank-sum test was applied to the ranked energy sequences of T waves to detect TWA qualitatively. The ECG containing TWA was quantitatively analyzed with correlation method. Results Simulation test result proved a mean sensitivity of 91.2% in detecting TWA, and for the SNR not less than 30 dB, the accuracy rate of detection achieved 100%. The clinical data experiment showed that the results from this method vs. spectral method had the correlation coefficients of 0.96. Conclusions A novel TWA analysis algorithm utilizing the wavelet transform and correlation technique is presented in this paper. TWAs are not only correctly detected qualitatively in frequency domain by energy value of T waves, but the alternans frequency and amplitude in temporal domain are measured quantitatively. PMID:23311454

  9. Quantitative detection of trace explosive vapors by programmed temperature desorption gas chromatography-electron capture detector.

    PubMed

    Field, Christopher R; Lubrano, Adam; Woytowitz, Morgan; Giordano, Braden C; Rose-Pehrsson, Susan L

    2014-07-25

    The direct liquid deposition of solution standards onto sorbent-filled thermal desorption tubes is used for the quantitative analysis of trace explosive vapor samples. The direct liquid deposition method yields a higher fidelity between the analysis of vapor samples and the analysis of solution standards than using separate injection methods for vapors and solutions, i.e., samples collected on vapor collection tubes and standards prepared in solution vials. Additionally, the method can account for instrumentation losses, which makes it ideal for minimizing variability and quantitative trace chemical detection. Gas chromatography with an electron capture detector is an instrumentation configuration sensitive to nitro-energetics, such as TNT and RDX, due to their relatively high electron affinity. However, vapor quantitation of these compounds is difficult without viable vapor standards. Thus, we eliminate the requirement for vapor standards by combining the sensitivity of the instrumentation with a direct liquid deposition protocol to analyze trace explosive vapor samples.

  10. Quantitative detection of trace explosive vapors by programmed temperature desorption gas chromatography-electron capture detector.

    PubMed

    Field, Christopher R; Lubrano, Adam; Woytowitz, Morgan; Giordano, Braden C; Rose-Pehrsson, Susan L

    2014-01-01

    The direct liquid deposition of solution standards onto sorbent-filled thermal desorption tubes is used for the quantitative analysis of trace explosive vapor samples. The direct liquid deposition method yields a higher fidelity between the analysis of vapor samples and the analysis of solution standards than using separate injection methods for vapors and solutions, i.e., samples collected on vapor collection tubes and standards prepared in solution vials. Additionally, the method can account for instrumentation losses, which makes it ideal for minimizing variability and quantitative trace chemical detection. Gas chromatography with an electron capture detector is an instrumentation configuration sensitive to nitro-energetics, such as TNT and RDX, due to their relatively high electron affinity. However, vapor quantitation of these compounds is difficult without viable vapor standards. Thus, we eliminate the requirement for vapor standards by combining the sensitivity of the instrumentation with a direct liquid deposition protocol to analyze trace explosive vapor samples. PMID:25145416

  11. Development of a quantitative fluorescence single primer isothermal amplification-based method for the detection of Salmonella.

    PubMed

    Wang, Jianchang; Li, Rui; Hu, Lianxia; Sun, Xiaoxia; Wang, Jinfeng; Li, Jing

    2016-02-16

    Food-borne disease caused by Salmonella has long been, and continues to be, an important global public health problem, necessitating rapid and accurate detection of Salmonella in food. Real time PCR is the most recently developed approach for Salmonella detection. Single primer isothermal amplification (SPIA), a novel gene amplification technique, has emerged as an attractive microbiological testing method. SPIA is performed under a constant temperature, eliminating the need for an expensive thermo-cycler. In addition, SPIA reactions can be accomplished in 30 min, faster than real time PCR that usually takes over 2h. We developed a quantitative fluorescence SPIA-based method for the detection of Salmonella. Using Salmonella Typhimurium genomic DNA as template and a primer targeting Salmonella invA gene, we showed the detection limit of SPIA was 2.0 × 10(1)fg DNA. Its successful amplification of different serotypic Salmonella genomic DNA but not non-Salmonella bacterial DNA demonstrated the specificity of SPIA. Furthermore, this method was validated with artificially contaminated beef. In conclusion, we showed high sensitivity and specificity of SPIA in the detection of Salmonella, comparable to real time PCR. In addition, SPIA is faster and more cost-effective (non-use of expensive cyclers), making it a potential alternative for field detection of Salmonella in resource-limited settings that are commonly encountered in developing countries.

  12. Approaches to mycotoxin detection using biosensors

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The number of toxins of concern has continued to rise as emerging toxins have taken on new significance and as interest has increased in detecting metabolites of established toxins (including masked mycotoxins). Of course while the desire exists to monitor for more compounds, resources for such moni...

  13. A Pedagogical Approach to Detective Fiction

    ERIC Educational Resources Information Center

    Reyes-Torres, Agustín

    2011-01-01

    One of the main concerns when teaching a foreign language is how to encourage students to read and become interested in its literature. This article presents detective fiction as a pedagogical tool that provides the key elements to make it appealing for young readers. In this way, the mystery, the action and the suspense in the story; the figure…

  14. Identification of recognition residues for ligation-based detection and quantitation of pseudouridine and N6-methyladenosine.

    PubMed

    Dai, Qing; Fong, Robert; Saikia, Mridusmita; Stephenson, David; Yu, Yi-tao; Pan, Tao; Piccirilli, Joseph A

    2007-01-01

    Over 100 chemical types of RNA modifications have been identified in thousands of sites in all three domains of life. Recent data suggest that modifications function synergistically to mediate biological function, and that cells may coordinately modulate modification levels for regulatory purposes. However, this area of RNA biology remains largely unexplored due to the lack of robust, high-throughput methods to quantify the extent of modification at specific sites. Recently, we developed a facile enzymatic ligation-based method for detection and quantitation of methylated 2'-hydroxyl groups within RNA. Here we exploit the principles of molecular recognition and nucleic acid chemistry to establish the experimental parameters for ligation-based detection and quantitation of pseudouridine (Psi) and N6-methyladenosine (m6A), two abundant modifications in eukaryotic rRNA/tRNA and mRNA, respectively. Detection of pseudouridylation at several sites in the large subunit rRNA derived from yeast demonstrates the feasibility of the approach for analysis of pseudouridylation in biological RNA samples. PMID:17881375

  15. Quantitative direct probe method for the detection of parvovirus B19.

    PubMed

    Boggino, H; Payne, D A

    2000-01-01

    Parvovirus B19 infection is associated with anemia and spontaneous abortions. While many qualitative assays are available, a few molecular-based quantitative methods have been described. This study reports the development and optimization of a quantitative direct-probe method for the detection of Parvovirus B19 DNA. Different concentrations of RNA probes were used to identify the optimal conditions for hybridizing to the target DNA. Detection of DNA was linear between concentrations of 2 ng/ml to 200 pg/ml. Because this method requires no enzymatic amplification, it is not susceptible to amplifier contamination or enzymatic inhibitors, and it can be applied to serum samples or paraffin-embedded tissue.

  16. Quantitative surface enhanced Raman scattering detection based on the ``sandwich'' structure substrate

    NASA Astrophysics Data System (ADS)

    Zhang, Junmeng; Qu, Shengchun; Zhang, Lisheng; Tang, Aiwei; Wang, Zhanguo

    2011-08-01

    A sandwich structured substrate was designed for quantitative molecular detection using surface enhanced Raman scattering (SERS), in which the probe molecule was sandwiched between silver nanoparticles (SNPs) and silver nanoarrays. The SNPs was prepared using Lee-Meisel method, and the silver nanoarrays was fabricated on porous anodic aluminum oxide (AAO) using electrodepositing method. The SERS studies show that the sandwich structured substrate exhibits good stability and reproducibility, and the detection sensitivity of Rhodamine 6G (R6G) and Melamine can respectively reach up to 10 -19 M and 10 -9 M, which is improved greatly as compared to other SERS substrates. The improved SERS sensitivity is closely associated with the stronger electromagnetic field enhancement, which stems from localized surface plasmon (LSP) coupling between the two silver nanostructures. Furthermore, the SERS intensity increased almost linearly as the mother concentration increased, which indicates that such a sandwich structure may be used as a good SERS substrate for quantitative analysis.

  17. Quantitative direct probe method for the detection of parvovirus B19.

    PubMed

    Boggino, H; Payne, D A

    2000-01-01

    Parvovirus B19 infection is associated with anemia and spontaneous abortions. While many qualitative assays are available, a few molecular-based quantitative methods have been described. This study reports the development and optimization of a quantitative direct-probe method for the detection of Parvovirus B19 DNA. Different concentrations of RNA probes were used to identify the optimal conditions for hybridizing to the target DNA. Detection of DNA was linear between concentrations of 2 ng/ml to 200 pg/ml. Because this method requires no enzymatic amplification, it is not susceptible to amplifier contamination or enzymatic inhibitors, and it can be applied to serum samples or paraffin-embedded tissue. PMID:10645984

  18. A medium hyperglycosylated podocalyxin enables noninvasive and quantitative detection of tumorigenic human pluripotent stem cells

    PubMed Central

    Tateno, Hiroaki; Onuma, Yasuko; Ito, Yuzuru; Hiemori, Keiko; Aiki, Yasuhiko; Shimizu, Madoka; Higuchi, Kumiko; Fukuda, Masakazu; Warashina, Masaki; Honda, Susumu; Asashima, Makoto; Hirabayashi, Jun

    2014-01-01

    While human pluripotent stem cells are attractive sources for cell-replacement therapies, a major concern remains regarding their tumorigenic potential. Thus, safety assessment of human pluripotent stem cell-based products in terms of tumorigenicity is critical. Previously we have identified a pluripotent stem cell-specific lectin probe rBC2LCN recognizing hyperglycosylated podocalyxin as a cell surface ligand. Here we demonstrate that hyperglycosylated podocalyxin is secreted from human pluripotent stem cells into cell culture supernatants. We establish a sandwich assay system, named the GlycoStem test, targeting the soluble hyperglycosylated podocalyxin using rBC2LCN. The GlycoStem test is sufficiently sensitive and quantitative to detect residual human pluripotent stem cells. This work provides a proof of concept for the noninvasive and quantitative detection of tumorigenic human pluripotent stem cells using cell culture supernatants. The developed method should increase the safety of human pluripotent stem cell-based cell therapies. PMID:24518842

  19. Quantitative detection of defects based on Markov-PCA-BP algorithm using pulsed infrared thermography technology

    NASA Astrophysics Data System (ADS)

    Tang, Qingju; Dai, Jingmin; Liu, Junyan; Liu, Chunsheng; Liu, Yuanlin; Ren, Chunping

    2016-07-01

    Quantitative detection of debonding defects' diameter and depth in TBCs has been carried out using pulsed infrared thermography technology. By combining principal component analysis with neural network theory, the Markov-PCA-BP algorithm was proposed. The principle and realization process of the proposed algorithm was described. In the prediction model, the principal components which can reflect most characteristics of the thermal wave signal were set as the input, and the defect depth and diameter was set as the output. The experimental data from pulsed infrared thermography tests of TBCs with flat bottom hole defects was selected as the training and testing sample. Markov-PCA-BP predictive system was arrived, based on which both the defect depth and diameter were identified accurately, which proved the effectiveness of the proposed method for quantitative detection of debonding defects in TBCs.

  20. Quantitative In Situ Detection of Phosphoproteins in Fixed Tissues Using Quantum Dot Technology

    PubMed Central

    Bodo, Juraj; Durkin, Lisa; Hsi, Eric D.

    2009-01-01

    Detection and quantitation of phosphoproteins (PPs) in fixed tissues will become increasingly important as additional inhibitors of protein kinases enter clinical use and new disease entities are defined by molecular changes affecting PP levels. We characterize fixation conditions suitable for accurate PP quantitation that are achievable in a clinical laboratory and illustrate the utility of in situ quantitation of PPs by quantum dot (QD) nanocrystals in two models: (1) a therapeutic model demonstrating effects of a targeted therapeutic (quantitative reduction of phospho-GSK3β) in xenografts treated with enzastaurin; and (2) a diagnostic model that identifies elevated levels of nuclear phospho-STAT5 in routine bone marrow biopsies from patients with acute myeloid leukemia based on the presence of the activating FLT3-ITD mutation. Finally, we document production of a well-characterized tissue microarray of widely available cell lines as a multilevel calibrator for validating numerous phosphoprotein assays. QD immunofluorescence is an ideal method for in situ quantitation of PPs in fixed samples, providing valuable cell type–specific and subcellular information about pathway activation in primary tissues. (J Histochem Cytochem 57:701–708, 2009) PMID:19332430

  1. Quantitative surface acoustic wave detection based on colloidal gold nanoparticles and their bioconjugates.

    PubMed

    Chiu, Chi-Shun; Gwo, Shangjr

    2008-05-01

    The immobilization scheme of monodispersed gold nanoparticles (10-nm diameter) on piezoelectric substrate surfaces using organosilane molecules as cross-linkers has been developed for lithium niobate (LiNbO3) and silicon oxide (SiO2)/gold-covered lithium tantalate (LiTaO3) of Rayleigh and guided shear horizontal- (guided SH) surface acoustic wave (SAW) sensors. In this study, comparative measurements of gold nanoparticle adsorption kinetics using high-resolution field-emission scanning electron microscopy and SAW sensors allow the frequency responses of SAW sensors to be quantitatively correlated with surface densities of adsorbed nanoparticles. Using this approach, gold nanoparticles are used as the "nanosized mass standards" to scale the mass loading in a wide dynamical range. Rayleigh-SAW and guided SH-SAW sensors are employed here to monitor the surface mass changes on the device surfaces in gas and liquid phases, respectively. The mass sensitivity ( approximately 20 Hz.cm2/ng) of Rayleigh-SAW device (fundamental oscillation frequency of 113.3 MHz in air) is more than 2 orders of magnitude higher than that of conventional 9-MHz quartz crystal microbalance sensors. Furthermore, in situ (aqueous solutions), real-time measurements of adsorption kinetics for both citrate-stabilized gold nanoparticles and DNA-gold nanoparticle conjugates are also demonstrated by guided SH-SAW (fundamental oscillation frequency of 121.3 MHz). By comparing frequency shifts between the adsorption cases of gold nanoparticles and DNA-gold nanoparticle conjugates, the average number of bound oligonucleotides per gold nanoparticle can also be determined. The high mass sensitivity ( approximately 6 Hz.cm2/ng) of guided SH-SAW sensors and successful detection of DNA-gold nanoparticle conjugates paves the way for real-time biosensing in liquids using nanoparticle-enhanced SAW devices. PMID:18363384

  2. Quantitative Magnetic Resonance Imaging Detects Changes in Meniscal Volume in Vivo After Partial Meniscectomy

    PubMed Central

    Bowers, Megan E.; Tung, Glenn A.; Oksendahl, Heidi L.; Hulstyn, Michael J.; Fadale, Paul D.; Machan, Jason T.; Fleming, Braden C.

    2010-01-01

    Background Quantifying changes in meniscal volume in vivo before and after partial meniscectomy (PM) could help elucidate the mechanisms involved in osteoarthritis development after meniscal injury and its surgical treatment. Purpose/Hypothesis To determine whether quantitative MRI (qMRI) could detect the immediate reduction in meniscal volume created by PM, while ruling out changes in unresected structures. We hypothesized that qMRI would be reliable for determining meniscal volume within the repeated images of unresected menisci. Additionally, we expected no significant difference in volume between the uninjured menisci of the injured knees and the same menisci of the uninjured knees. Study Design Controlled laboratory study. Methods Ten subjects with meniscal tears were evaluated with 3T MRI before and after arthroscopic PM. Manual segmentation was used to create models of the menisci and to determine the pre- and post-operative meniscal volumes for each subject. The responsiveness and reliability of qMRI for determining meniscal volume in vivo were evaluated using these measurements. We expected a decrease in volume of the resected menisci, but not in the uninjured menisci, after surgery. Results The mean pre-operative volume of the injured menisci was significantly greater than the mean post-operative volume (2896±277mm3 vs. 2480±277mm3; p=0.000). There was no significant difference between the mean pre- and post-operative volumes of the uninjured menisci (2687±256mm3 vs. 2694±256mm3; p=1.000). Conclusions Manual segmentation demonstrated a significant reduction in the volume of the surgically resected menisci after PM, but no significant change in the volume of unresected meniscal tissue, indicating that the manual segmentation method is responsive. Clinical Relevance This approach offers a novel, reliable method to study the relationship between the volume of meniscal tissue removed during PM and subsequent patient outcomes during long-term clinical

  3. Quantitative Systems Pharmacology Approaches Applied to Microphysiological Systems (MPS): Data Interpretation and Multi-MPS Integration

    PubMed Central

    Yu, J; Cilfone, NA; Large, EM; Sarkar, U; Wishnok, JS; Tannenbaum, SR; Hughes, DJ; Lauffenburger, DA; Griffith, LG; Stokes, CL; Cirit, M

    2015-01-01

    Our goal in developing Microphysiological Systems (MPS) technology is to provide an improved approach for more predictive preclinical drug discovery via a highly integrated experimental/computational paradigm. Success will require quantitative characterization of MPSs and mechanistic analysis of experimental findings sufficient to translate resulting insights from in vitro to in vivo. We describe herein a systems pharmacology approach to MPS development and utilization that incorporates more mechanistic detail than traditional pharmacokinetic/pharmacodynamic (PK/PD) models. A series of studies illustrates diverse facets of our approach. First, we demonstrate two case studies: a PK data analysis and an inflammation response––focused on a single MPS, the liver/immune MPS. Building on the single MPS modeling, a theoretical investigation of a four-MPS interactome then provides a quantitative way to consider several pharmacological concepts such as absorption, distribution, metabolism, and excretion in the design of multi-MPS interactome operation and experiments. PMID:26535159

  4. Differential plating medium for quantitative detection of histamine-producing bacteria.

    PubMed Central

    Niven, C F; Jeffrey, M B; Corlett, D A

    1981-01-01

    A histidine-containing agar medium has been devised for quantitative detection of histamine-producing bacteria that are alleged to be associated with scombroid fish poisoning outbreaks. The responsible bacteria produce a marked pH change in the agar, with attendant color change of pH indicator adjacent to the colonies, thus facilitating their recognition. Proteus morganii and Klebsiella pneumoniae were the two most common histidine-decarboxylating species isolated from scombroid fish and mahi mahi. PMID:7013698

  5. Simplified approach for quantitative digital holographic phase contrast imaging of living cells

    NASA Astrophysics Data System (ADS)

    Kemper, Björn; Vollmer, Angelika; Rommel, Christina E.; Schnekenburger, Jürgen; Bally, Gert Von

    2011-02-01

    Many interferometry-based quantitative phase contrast imaging techniques require a separately generated coherent reference wave. This results in a low phase stability and the demand for a precise adjustment of the intensity ratio between object and reference wave. To overcome these problems, the performance of a Michelson interferometer approach for digital holographic microscopy was analyzed that avoids a separately generated reference wave by superposition of different image areas. It is shown that this simplified arrangement yields improved phase stability. Furthermore, results from time-lapse investigations on living pancreas tumor cells demonstrate the capability of the method for reliable quantitative phase contrast imaging.

  6. Detecting qualitative interaction: a Bayesian approach.

    PubMed

    Bayman, Emine Ozgür; Chaloner, Kathryn; Cowles, Mary Kathryn

    2010-02-20

    Differences in treatment effects between centers in a multi-center trial may be important. These differences represent treatment by subgroup interaction. Peto defines qualitative interaction (QI) to occur when the simple treatment effect in one subgroup has a different sign than in another subgroup: this interaction is important. Interaction where the treatment effects are of the same sign in all subgroups is called quantitative and is often not important because the treatment recommendation is identical in all cases. A hierarchical model is used here with exchangeable mean responses to each treatment between subgroups. The posterior probability of QI and the corresponding Bayes factor are proposed as a diagnostic and as a test statistic. The model is motivated by two multi-center trials with binary responses. The frequentist power and size of the test using the Bayes factor are examined and compared with two other commonly used tests. The impact of imbalance between the sample sizes in each subgroup on power is examined, and the test based on the Bayes factor typically has better power for unbalanced designs, especially for small sample sizes. An exact test based on the Bayes factor is also suggested assuming the hierarchical model. The Bayes factor provides a concise summary of the evidence for or against QI. It is shown by example that it is easily adapted to summarize the evidence for 'clinically meaningful QI,' defined as the simple effects being of opposite signs and larger in absolute value than a minimal clinically meaningful effect.

  7. A DNA immunoprecipitation assay used in quantitative detection of in vitro DNA-protein complex binding.

    PubMed

    Kim, Min Young; Chae, Ji Hyung; Oh, Chang-Ho; Kim, Chul Geun

    2013-10-15

    To begin gene transcription, several transcription factors must bind to specific DNA sequences to form a complex via DNA-protein interactions. We established an in vitro method for specific and sensitive analyses of DNA-protein interactions based on a DNA immunoprecipitation (DIP) method. We verified the accuracy and efficiency of the DIP assay in quantitatively measuring DNA-protein binding using transcription factor CP2c as a model. With our DIP assay, we could detect specific interactions within a DNA-CP2c complex, with reproducible and quantitative binding values. In addition, we were able to effectively measure the changes in DNA-CP2c binding by the addition of a small molecule, FQI1 (factor quinolinone inhibitor 1), previously identified as a specific inhibitor of this binding. To identify a new regulator of DNA-CP2c binding, we analyzed several CP2c binding peptides and found that only one class of peptide severely inhibits DNA-CP2c binding. These data show that our DIP assay is very useful in quantitatively detecting the binding dynamics of DNA-protein complex. Because DNA-protein interaction is very dynamic in different cellular environments, our assay can be applied to the detection of active transcription factors, including promoter occupancy in normal and disease conditions. Moreover, it may be used to develop a targeted regulator of specific DNA-protein interaction.

  8. Validation of quantitative and qualitative methods for detecting allergenic ingredients in processed foods in Japan.

    PubMed

    Sakai, Shinobu; Adachi, Reiko; Akiyama, Hiroshi; Teshima, Reiko

    2013-06-19

    A labeling system for food allergenic ingredients was established in Japan in April 2002. To monitor the labeling, the Japanese government announced official methods for detecting allergens in processed foods in November 2002. The official methods consist of quantitative screening tests using enzyme-linked immunosorbent assays (ELISAs) and qualitative confirmation tests using Western blotting or polymerase chain reactions (PCR). In addition, the Japanese government designated 10 μg protein/g food (the corresponding allergenic ingredient soluble protein weight/food weight), determined by ELISA, as the labeling threshold. To standardize the official methods, the criteria for the validation protocol were described in the official guidelines. This paper, which was presented at the Advances in Food Allergen Detection Symposium, ACS National Meeting and Expo, San Diego, CA, Spring 2012, describes the validation protocol outlined in the official Japanese guidelines, the results of interlaboratory studies for the quantitative detection method (ELISA for crustacean proteins) and the qualitative detection method (PCR for shrimp and crab DNAs), and the reliability of the detection methods.

  9. Comparison of standard, quantitative and digital PCR in the detection of enterotoxigenic Bacteroides fragilis

    PubMed Central

    Purcell, Rachel V.; Pearson, John; Frizelle, Frank A.; Keenan, Jacqueline I.

    2016-01-01

    Gut colonization with enterotoxigenic Bacteroides fragilis (ETBF) appears to be associated with the development of colorectal cancer. However, differences in carriage rates are seen with various testing methods and sampling sites. We compared standard PCR, SYBR green and TaqMan quantitative PCR (qPCR) and digital PCR (dPCR) in detecting the B. fragilis toxin (bft) gene from cultured ETBF, and from matched luminal and faecal stool samples from 19 colorectal cancer patients. Bland-Altman analysis found that all three quantitative methods performed comparably in detecting bft from purified bacterial DNA, with the same limits of detection (<1 copy/μl). However, SYBR qPCR under-performed compared to TaqMan qPCR and dPCR in detecting bft in clinical stool samples; 13/38 samples were reported positive by SYBR, compared to 35 and 36 samples by TaqMan and dPCR, respectively. TaqMan qPCR and dPCR gave bft copy numbers that were 48-fold and 75-fold higher for the same samples than SYBR qPCR, respectively (p < 0.001). For samples that were bft-positive in both fecal and luminal stools, there was no difference in relative abundance between the sites, by any method tested. From our findings, we recommend the use of TaqMan qPCR as the preferred method to detect ETBF from clinical stool samples. PMID:27686415

  10. An ECL-PCR method for quantitative detection of point mutation

    NASA Astrophysics Data System (ADS)

    Zhu, Debin; Xing, Da; Shen, Xingyan; Chen, Qun; Liu, Jinfeng

    2005-04-01

    A new method for identification of point mutations was proposed. Polymerase chain reaction (PCR) amplification of a sequence from genomic DNA was followed by digestion with a kind of restriction enzyme, which only cut the wild-type amplicon containing its recognition site. Reaction products were detected by electrochemiluminescence (ECL) assay after adsorption of the resulting DNA duplexes to the solid phase. One strand of PCR products carries biotin to be bound on a streptavidin-coated microbead for sample selection. Another strand carries Ru(bpy)32+ (TBR) to react with tripropylamine (TPA) to emit light for ECL detection. The method was applied to detect a specific point mutation in H-ras oncogene in T24 cell line. The results show that the detection limit for H-ras amplicon is 100 fmol and the linear range is more than 3 orders of magnitude, thus, make quantitative analysis possible. The genotype can be clearly discriminated. Results of the study suggest that ECL-PCR is a feasible quantitative method for safe, sensitive and rapid detection of point mutation in human genes.

  11. Detecting initial orthostatic hypotension: a novel approach

    PubMed Central

    McJunkin, Brittain; Rose, Brandon; Amin, Om; Shah, Nirmita; Sharma, Sachin; Modi, Sujal; Kemper, Suzanne; Yousaf, Muhammad

    2015-01-01

    Our purpose, by modification of standard bedside tilt–testing, was to search for lesser known but important initial orthostatic hypotension (IOH), occurring transiently within the first 30 seconds of standing, heretofore only detectable with sophisticated continuous photoplethysmographic monitoring systems, not readily available in most medical facilities. In screened outpatients over 60 years of age, supine blood pressure (BP) parameters were recorded. To achieve readiness for immediate BP after standing, the cuff was re–inflated prior to standing, rather than after. Immediate, 1–, and 3–minute standing BPs were recorded. One hundred fifteen patients were studied (mean age, 71.1 years; 50.5% male). Eighteen (15.6%) had OH, of whom 14 (12.1%) had classical OH, and four (3.5%) had IOH. Early standing BP detection time was 20.1 ± 5.3 seconds. Immediate transient physiologic systolic BP decline was detected in non–OH (−8.8 ± 9.9 mm Hg; P < .0001). In contrast to classical OH (with lesser but persistent orthostatic BP decrements), IOH patients had immediate mean orthostatic systolic/diastolic BP change of −32.8 (±13.8) mm Hg/−14.0 (±8.5) mm Hg (P < .02), with recovery back to baseline by 1 minute. Two of the four IOH patients had pre–syncopal symptoms. For the first time, using standard inflation–deflation BP equipment, immediate transient standing physiologic BP decrement and IOH were demonstrated. This preliminary study confirms proof of principle that manual BP cuff inflation prior to standing may be useful and practical in diagnosing IOH, and may stimulate direct comparative studies with continuous monitoring systems. PMID:25816712

  12. Detecting initial orthostatic hypotension: a novel approach.

    PubMed

    McJunkin, Brittain; Rose, Brandon; Amin, Om; Shah, Nirmita; Sharma, Sachin; Modi, Sujal; Kemper, Suzanne; Yousaf, Muhammad

    2015-05-01

    Our purpose, by modification of standard bedside tilt-testing, was to search for lesser known but important initial orthostatic hypotension (IOH), occurring transiently within the first 30 seconds of standing, heretofore only detectable with sophisticated continuous photoplethysmographic monitoring systems, not readily available in most medical facilities. In screened outpatients over 60 years of age, supine blood pressure (BP) parameters were recorded. To achieve readiness for immediate BP after standing, the cuff was re-inflated prior to standing, rather than after. Immediate, 1-, and 3-minute standing BPs were recorded. One hundred fifteen patients were studied (mean age, 71.1 years; 50.5% male). Eighteen (15.6%) had OH, of whom 14 (12.1%) had classical OH, and four (3.5%) had IOH. Early standing BP detection time was 20.1 ± 5.3 seconds. Immediate transient physiologic systolic BP decline was detected in non-OH (-8.8 ± 9.9 mm Hg; P < .0001). In contrast to classical OH (with lesser but persistent orthostatic BP decrements), IOH patients had immediate mean orthostatic systolic/diastolic BP change of -32.8 (±13.8) mm Hg/-14.0 (±8.5) mm Hg (P < .02), with recovery back to baseline by 1 minute. Two of the four IOH patients had pre-syncopal symptoms. For the first time, using standard inflation-deflation BP equipment, immediate transient standing physiologic BP decrement and IOH were demonstrated. This preliminary study confirms proof of principle that manual BP cuff inflation prior to standing may be useful and practical in diagnosing IOH, and may stimulate direct comparative studies with continuous monitoring systems.

  13. A neural network approach to burst detection.

    PubMed

    Mounce, S R; Day, A J; Wood, A S; Khan, A; Widdop, P D; Machell, J

    2002-01-01

    This paper describes how hydraulic and water quality data from a distribution network may be used to provide a more efficient leakage management capability for the water industry. The research presented concerns the application of artificial neural networks to the issue of detection and location of leakage in treated water distribution systems. An architecture for an Artificial Neural Network (ANN) based system is outlined. The neural network uses time series data produced by sensors to directly construct an empirical model for predication and classification of leaks. Results are presented using data from an experimental site in Yorkshire Water's Keighley distribution system. PMID:11936639

  14. Design of an Evolutionary Approach for Intrusion Detection

    PubMed Central

    2013-01-01

    A novel evolutionary approach is proposed for effective intrusion detection based on benchmark datasets. The proposed approach can generate a pool of noninferior individual solutions and ensemble solutions thereof. The generated ensembles can be used to detect the intrusions accurately. For intrusion detection problem, the proposed approach could consider conflicting objectives simultaneously like detection rate of each attack class, error rate, accuracy, diversity, and so forth. The proposed approach can generate a pool of noninferior solutions and ensembles thereof having optimized trade-offs values of multiple conflicting objectives. In this paper, a three-phase, approach is proposed to generate solutions to a simple chromosome design in the first phase. In the first phase, a Pareto front of noninferior individual solutions is approximated. In the second phase of the proposed approach, the entire solution set is further refined to determine effective ensemble solutions considering solution interaction. In this phase, another improved Pareto front of ensemble solutions over that of individual solutions is approximated. The ensemble solutions in improved Pareto front reported improved detection results based on benchmark datasets for intrusion detection. In the third phase, a combination method like majority voting method is used to fuse the predictions of individual solutions for determining prediction of ensemble solution. Benchmark datasets, namely, KDD cup 1999 and ISCX 2012 dataset, are used to demonstrate and validate the performance of the proposed approach for intrusion detection. The proposed approach can discover individual solutions and ensemble solutions thereof with a good support and a detection rate from benchmark datasets (in comparison with well-known ensemble methods like bagging and boosting). In addition, the proposed approach is a generalized classification approach that is applicable to the problem of any field having multiple conflicting

  15. A metabolomics approach to identify and quantify the phytochemicals in watermelons by quantitative (1)HNMR.

    PubMed

    Jayaprakasha, G K; Patil, Bhimanagouda S

    2016-06-01

    Watermelon (Citrullus vulgaris) contains many health-promoting compounds, such as ascorbic acid, carotenoids, phenolic acids and amino acids including l-citrulline, arginine, and glutathione. Reported HPLC method for quantification of l-citrulline and sugars in watermelon involves, time-consuming sample preparation, post-column color development and detection with fluorescence and refractive index detectors. The present study describes development of a method to identify and quantify amino acids and sugars simultaneously from watermelon samples using quantitative proton NMR. Lyophilized watermelon samples (30-50mg) were extracted with deuterium oxide (D2O) by sonication and the centrifuged extract was directly used for quantification and identification with (1)HNMR. An external coaxial insert containing a 65µL of 0.012% 3-(trimethylsilyl) propionic-(2,2,3,3-d4) acid sodium salt (TSP-d4) in D2O was used as a quantitative reference. The levels of l-citrulline and sugars were measured in less than 6min. This rapid quantitation method was validated for specificity, linearity, accuracy, precision, reproducibility, and robustness. The limit of detection for l-citrulline was 38µg/mL and the limit of quantification was 71µg/mL; for sugars, the limits were 59-94µg/mL and 120µg/mL, respectively. This method can be used widely for confirmation and rapid quantitation of multiple compounds in large number of biological or breeding samples for routine analysis.

  16. A metabolomics approach to identify and quantify the phytochemicals in watermelons by quantitative (1)HNMR.

    PubMed

    Jayaprakasha, G K; Patil, Bhimanagouda S

    2016-06-01

    Watermelon (Citrullus vulgaris) contains many health-promoting compounds, such as ascorbic acid, carotenoids, phenolic acids and amino acids including l-citrulline, arginine, and glutathione. Reported HPLC method for quantification of l-citrulline and sugars in watermelon involves, time-consuming sample preparation, post-column color development and detection with fluorescence and refractive index detectors. The present study describes development of a method to identify and quantify amino acids and sugars simultaneously from watermelon samples using quantitative proton NMR. Lyophilized watermelon samples (30-50mg) were extracted with deuterium oxide (D2O) by sonication and the centrifuged extract was directly used for quantification and identification with (1)HNMR. An external coaxial insert containing a 65µL of 0.012% 3-(trimethylsilyl) propionic-(2,2,3,3-d4) acid sodium salt (TSP-d4) in D2O was used as a quantitative reference. The levels of l-citrulline and sugars were measured in less than 6min. This rapid quantitation method was validated for specificity, linearity, accuracy, precision, reproducibility, and robustness. The limit of detection for l-citrulline was 38µg/mL and the limit of quantification was 71µg/mL; for sugars, the limits were 59-94µg/mL and 120µg/mL, respectively. This method can be used widely for confirmation and rapid quantitation of multiple compounds in large number of biological or breeding samples for routine analysis. PMID:27130118

  17. Rapid and quantitative detection of C-reactive protein using quantum dots and immunochromatographic test strips

    PubMed Central

    Cheng, Xianglin; Pu, Xu; Jun, Pen; Zhu, XiaoBo; Zhu, Di; Chen, Ming

    2014-01-01

    Background Rapid immunochromatographic tests can detect disease markers in 10–15 minutes, which facilitates clinical diagnosis and treatment programs. However, most immunochromatographic tests employ gold nanoparticles as reporters, and these have only moderate sensitivity and act as qualitative methods for analyzing high biomarker concentrations. Methods In this study, we introduce quantum dots (QDs) as fluorescent probes and immunochromatographic strips to develop quantitative fluorescence point-of-care tests (QF-POCT) to analyze C-reactive protein (CRP) levels. Goat anti-rabbit IgG and rabbit IgG were used as control antibodies, and mouse monoclonal CRP antibody pairs were used for disease marker detection. One monoclonal CRP antibody was conjugated with QDs and served as a signal antibody, and the other monoclonal CRP antibody was dispensed onto the nitrocellulose membrane and served as a capturing antibody. In the presence of CRP, the fluorescence intensity of the monoclonal antibody-CRP-monoclonal antibody sandwich complex captured on the nitrocellulose membrane was determined using the fluorescence strip reader. Results QF-POCT assays could quantitatively analyze the concentration of CRP in 15 minutes had a detection limit of 0.25 mg/L, and had a wide detection linearity range (0.5–300 mg/L). The intra-assay and interassay coefficients of variation were 8.95% and 9.86% at 0.5 mg/L, 6.47% and 8.66% at 10 mg/L, and 6.81% and 9.10% at 60 mg/L, respectively. In a comparison between clinical samples, the results of this QD-based assay of CRP levels were significantly correlated with those of an Immulite 2000 assay (R=0.993, P<0.001). Conclusion Our results demonstrated that the QD-based immunochromatographic test is a rapid, sensitive, accurate, and quantitative method for the detection of disease biomarkers. PMID:25506215

  18. Application of Programmable Bio-Nano-Chip System for the Quantitative Detection of Drugs of Abuse in Oral Fluids*

    PubMed Central

    Christodoulides, Nicolaos; De La Garza, Richard; Simmons, Glennon W.; McRae, Michael P.; Wong, Jorge; Newton, Thomas F.; Smith, Regina; Mahoney, James J.; Hohenstein, Justin; Gomez, Sobeyda; Floriano, Pierre N.; Talavera, Humberto; Sloan, Daniel J.; Moody, David E.; Andrenyak, David M.; Kosten, Thomas R.; Haque, Ahmed; McDevitt, John T.

    2015-01-01

    Objective There is currently a gap in on-site drug of abuse monitoring. Current detection methods involve invasive sampling of blood and urine specimens, or collection of oral fluid, followed by qualitative screening tests using immunochromatographic cartridges. While remote laboratories then may provide confirmation and quantitative assessment of a presumptive positive, this instrumentation is expensive and decoupled from the initial sampling making the current drug-screening program inefficient and costly. The authors applied a noninvasive oral fluid sampling approach integrated with the in-development chip-based Programmable Bio-Nano-Chip (p-BNC) platform for the detection of drugs of abuse. Method The p-BNC assay methodology was applied for the detection of tetrahydrocannabinol, morphine, amphetamine, methamphetamine, cocaine, methadone and benzodiazepines, initially using spiked buffered samples and, ultimately, using oral fluid specimen collected from consented volunteers. Results Rapid (~10 minutes), sensitive detection (~ng/ml) and quantitation of 12 drugs of abuse was demonstrated on the p-BNC platform. Furthermore, the system provided visibility to time-course of select drug and metabolite profiles in oral fluids; for the drug cocaine, three regions of slope were observed that, when combined with concentration measurements from this and prior impairment studies, information about cocaine-induced impairment may be revealed. Conclusions This chip-based p-BNC detection modality has significant potential to be used in the future by law enforcement officers for roadside drug testing and to serve a variety of other settings, including outpatient and inpatient drug rehabilitation centers, emergency rooms, prisons, schools, and in the workplace. PMID:26048639

  19. An Energy-Based Three-Dimensional Segmentation Approach for the Quantitative Interpretation of Electron Tomograms

    PubMed Central

    Bartesaghi, Alberto; Sapiro, Guillermo; Subramaniam, Sriram

    2006-01-01

    Electron tomography allows for the determination of the three-dimensional structures of cells and tissues at resolutions significantly higher than that which is possible with optical microscopy. Electron tomograms contain, in principle, vast amounts of information on the locations and architectures of large numbers of subcellular assemblies and organelles. The development of reliable quantitative approaches for the analysis of features in tomograms is an important problem, and a challenging prospect due to the low signal-to-noise ratios that are inherent to biological electron microscopic images. This is, in part, a consequence of the tremendous complexity of biological specimens. We report on a new method for the automated segmentation of HIV particles and selected cellular compartments in electron tomograms recorded from fixed, plastic-embedded sections derived from HIV-infected human macrophages. Individual features in the tomogram are segmented using a novel robust algorithm that finds their boundaries as global minimal surfaces in a metric space defined by image features. The optimization is carried out in a transformed spherical domain with the center an interior point of the particle of interest, providing a proper setting for the fast and accurate minimization of the segmentation energy. This method provides tools for the semi-automated detection and statistical evaluation of HIV particles at different stages of assembly in the cells and presents opportunities for correlation with biochemical markers of HIV infection. The segmentation algorithm developed here forms the basis of the automated analysis of electron tomograms and will be especially useful given the rapid increases in the rate of data acquisition. It could also enable studies of much larger data sets, such as those which might be obtained from the tomographic analysis of HIV-infected cells from studies of large populations. PMID:16190467

  20. A Bayesian quantitative nondestructive evaluation (QNDE) approach to estimating remaining life of aging pressure vessels and piping*

    NASA Astrophysics Data System (ADS)

    Fong, J. T.; Filliben, J. J.; Heckert, N. A.; Guthrie, W. F.

    2013-01-01

    In this paper, we use a Bayesian quantitative nondestructive evaluation (QNDE) approach to estimating the remaining life of aging structures and components. Our approach depends on in-situ NDE measurements of detectable crack lengths and crack growth rates in a multi-crack region of an aging component as a basis for estimating the mean and standard deviation of its remaining life. We introduce a general theory of crack growth involving multiple cracks such that the mean and standard deviation of the initial crack lengths can be directly estimated from NDEmeasured crack length data over a period of several inspection intervals. A numerical example using synthetic NDE data for high strength steels is presented to illustrate this new methodology.

  1. A system coefficient approach for quantitative assessment of the solvent effects on membrane absorption from chemical mixtures.

    PubMed

    Xia, X R; Baynes, R E; Monteiro-Riviere, N A; Riviere, J E

    2007-01-01

    A system coefficient approach is proposed for quantitative assessment of the solvent effects on membrane absorption from chemical mixtures. The complicated molecular interactions are dissected into basic molecular interaction forces via Abraham's linear solvation energy relationship (LSER). The molecular interaction strengths of a chemical are represented by a set of solute descriptors, while those of a membrane/chemical mixture system are represented by a set of system coefficients. The system coefficients can be determined by using a set of probe compounds with known solute descriptors. Polydimethylsiloxane (PDMS) membrane-coated fibres and 32 probe compounds were used to demonstrate the proposed approach. When a solvent was added into the chemical mixture, the system coefficients were altered and detected by the system coefficient approach. The system coefficients of the PDMS/water system were (0.09, 0.49, -1.11, -2.36, -3.78, 3.50). When 25% ethanol was added into the PDMS/water system, the system coefficients were altered significantly (0.38, 0.41, -1.18, -2.07, -3.40, 2.81); and the solvent effect was quantitatively described by the changes in the system coefficients (0.29, -0.08, -0.07, 0.29, 0.38, -0.69). The LSER model adequately described the experimental data with a correlation coefficient (r(2)) of 0.995 and F-value of 1056 with p-value less than 0.0001.

  2. Multisensor neural network approach to mine detection

    NASA Astrophysics Data System (ADS)

    Iler, Amber L.; Marble, Jay A.; Rauss, Patrick J.

    2001-10-01

    A neural network is applied to data collected by the close-in detector for the Mine Hunter Killer (MHK) project with promising results. We use the ground penetrating radar (GPR) and metal detector to create three channels (two from the GPR) and train a basic, two layer (single hidden layer), feed-forward neural network. By experimenting with the number of hidden nodes and training goals, we were able to surpass the performance of the single sensors when we fused the three channels via our neural network and applied the trained net to different data. The fused sensors exceeded the best single sensor performance above 95 percent detection by providing a lower, but still high, false alarm rate. And though our three channel neural net worked best, we saw an increase in performance with fewer than three channels, as well.

  3. Quantitative subpixel spectral detection of targets in multispectral images. [terrestrial and planetary surfaces

    NASA Technical Reports Server (NTRS)

    Sabol, Donald E., Jr.; Adams, John B.; Smith, Milton O.

    1992-01-01

    The conditions that affect the spectral detection of target materials at the subpixel scale are examined. Two levels of spectral mixture analysis for determining threshold detection limits of target materials in a spectral mixture are presented, the cases where the target is detected as: (1) a component of a spectral mixture (continuum threshold analysis) and (2) residuals (residual threshold analysis). The results of these two analyses are compared under various measurement conditions. The examples illustrate the general approach that can be used for evaluating the spectral detectability of terrestrial and planetary targets at the subpixel scale.

  4. A textural approach based on Gabor functions for texture edge detection in ultrasound images.

    PubMed

    Chen, C M; Lu, H H; Han, K C

    2001-04-01

    Edge detection is an important, but difficult, step in quantitative ultrasound (US) image analysis. In this paper, we present a new textural approach for detecting a class of edges in US images; namely, the texture edges with a weak regional mean gray-level difference (RMGD) between adjacent regions. The proposed approach comprises a vision model-based texture edge detector using Gabor functions and a new texture-enhancement scheme. The experimental results on the synthetic edge images have shown that the performances of the four tested textural and nontextural edge detectors are about 20%-95% worse than that of the proposed approach. Moreover, the texture enhancement may improve the performance of the proposed texture edge detector by as much as 40%. The experiments on 20 clinical US images have shown that the proposed approach can find reasonable edges for real objects of interest with the performance of 0.4 +/- 0.08 in terms of the Pratt's figure.

  5. Detection of human brain tumor infiltration with quantitative stimulated Raman scattering microscopy.

    PubMed

    Ji, Minbiao; Lewis, Spencer; Camelo-Piragua, Sandra; Ramkissoon, Shakti H; Snuderl, Matija; Venneti, Sriram; Fisher-Hubbard, Amanda; Garrard, Mia; Fu, Dan; Wang, Anthony C; Heth, Jason A; Maher, Cormac O; Sanai, Nader; Johnson, Timothy D; Freudiger, Christian W; Sagher, Oren; Xie, Xiaoliang Sunney; Orringer, Daniel A

    2015-10-14

    Differentiating tumor from normal brain is a major barrier to achieving optimal outcome in brain tumor surgery. New imaging techniques for visualizing tumor margins during surgery are needed to improve surgical results. We recently demonstrated the ability of stimulated Raman scattering (SRS) microscopy, a nondestructive, label-free optical method, to reveal glioma infiltration in animal models. We show that SRS reveals human brain tumor infiltration in fresh, unprocessed surgical specimens from 22 neurosurgical patients. SRS detects tumor infiltration in near-perfect agreement with standard hematoxylin and eosin light microscopy (κ = 0.86). The unique chemical contrast specific to SRS microscopy enables tumor detection by revealing quantifiable alterations in tissue cellularity, axonal density, and protein/lipid ratio in tumor-infiltrated tissues. To ensure that SRS microscopic data can be easily used in brain tumor surgery, without the need for expert interpretation, we created a classifier based on cellularity, axonal density, and protein/lipid ratio in SRS images capable of detecting tumor infiltration with 97.5% sensitivity and 98.5% specificity. Quantitative SRS microscopy detects the spread of tumor cells, even in brain tissue surrounding a tumor that appears grossly normal. By accurately revealing tumor infiltration, quantitative SRS microscopy holds potential for improving the accuracy of brain tumor surgery.

  6. Validation of a quantitative PCR assay for detection and quantification of 'Candidatus Xenohaliotis californiensis'.

    PubMed

    Friedman, Carolyn S; Wight, Nate; Crosson, Lisa M; White, Samuel J; Strenge, Robyn M

    2014-04-01

    Withering syndrome (WS), a serious disease affecting abalone Haliotis spp., is caused by infection from an intracellular Rickettsia-like organism (WS-RLO). Diagnosis of the disease currently relies on a combination of histological examination and molecular methods (in situ hybridization, standard PCR, and sequence analysis). However, these techniques only provide a semi-quantitative assessment of bacterial load. We created a real-time quantitative PCR (qPCR) assay to specifically identify and enumerate bacterial loads of WS-RLO in abalone tissue, fecal, and seawater samples based on 16S rDNA gene copy numbers. The qPCR assay designed to detect DNA of the WS-RLO was validated according to standards set by the World Organisation for Animal Health. Standard curves derived from purified plasmid dilutions were linear across 7 logs of concentration, and efficiencies ranged from 90.2 to 97.4%. The limit of detection was 3 gene copies per reaction. Diagnostic sensitivity was 100% and specificity was 99.8%. The qPCR assay was robust, as evidenced by its high level of repeatability and reproducibility. This study has shown for the first time that WS-RLO DNA can be detected and quantified in abalone tissue, fecal, and seawater samples. The ability to detect and quantify RLO gene copies in a variety of materials will enable us to better understand transmission dynamics in both farmed and natural environments. PMID:24695238

  7. Detection of human brain tumor infiltration with quantitative stimulated Raman scattering microscopy

    PubMed Central

    Ji, Minbiao; Lewis, Spencer; Camelo-Piragua, Sandra; Ramkissoon, Shakti H.; Snuderl, Matija; Venneti, Sriram; Fisher-Hubbard, Amanda; Garrard, Mia; Fu, Dan; Wang, Anthony C.; Heth, Jason A.; Maher, Cormac O.; Sanai, Nader; Johnson, Timothy D.; Freudiger, Christian W.; Sagher, Oren; Xie, Xiaoliang Sunney; Orringer, Daniel A.

    2016-01-01

    Differentiating tumor from normal brain is a major barrier to achieving optimal outcome in brain tumor surgery. New imaging techniques for visualizing tumor margins during surgery are needed to improve surgical results. We recently demonstrated the ability of stimulated Raman scattering (SRS) microscopy, a non-destructive, label-free optical method, to reveal glioma infiltration in animal models. Here we show that SRS reveals human brain tumor infiltration in fresh, unprocessed surgical specimens from 22 neurosurgical patients. SRS detects tumor infiltration in near-perfect agreement with standard hematoxylin and eosin light microscopy (κ=0.86). The unique chemical contrast specific to SRS microscopy enables tumor detection by revealing quantifiable alterations in tissue cellularity, axonal density and protein:lipid ratio in tumor-infiltrated tissues. To ensure that SRS microscopic data can be easily used in brain tumor surgery, without the need for expert interpretation, we created a classifier based on cellularity, axonal density and protein:lipid ratio in SRS images capable of detecting tumor infiltration with 97.5% sensitivity and 98.5% specificity. Importantly, quantitative SRS microscopy detects the spread of tumor cells, even in brain tissue surrounding a tumor that appears grossly normal. By accurately revealing tumor infiltration, quantitative SRS microscopy holds potential for improving the accuracy of brain tumor surgery. PMID:26468325

  8. Quantitative Genetics and Functional–Structural Plant Growth Models: Simulation of Quantitative Trait Loci Detection for Model Parameters and Application to Potential Yield Optimization

    PubMed Central

    Letort, Véronique; Mahe, Paul; Cournède, Paul-Henry; de Reffye, Philippe; Courtois, Brigitte

    2008-01-01

    Background and Aims Prediction of phenotypic traits from new genotypes under untested environmental conditions is crucial to build simulations of breeding strategies to improve target traits. Although the plant response to environmental stresses is characterized by both architectural and functional plasticity, recent attempts to integrate biological knowledge into genetics models have mainly concerned specific physiological processes or crop models without architecture, and thus may prove limited when studying genotype × environment interactions. Consequently, this paper presents a simulation study introducing genetics into a functional–structural growth model, which gives access to more fundamental traits for quantitative trait loci (QTL) detection and thus to promising tools for yield optimization. Methods The GREENLAB model was selected as a reasonable choice to link growth model parameters to QTL. Virtual genes and virtual chromosomes were defined to build a simple genetic model that drove the settings of the species-specific parameters of the model. The QTL Cartographer software was used to study QTL detection of simulated plant traits. A genetic algorithm was implemented to define the ideotype for yield maximization based on the model parameters and the associated allelic combination. Key Results and Conclusions By keeping the environmental factors constant and using a virtual population with a large number of individuals generated by a Mendelian genetic model, results for an ideal case could be simulated. Virtual QTL detection was compared in the case of phenotypic traits – such as cob weight – and when traits were model parameters, and was found to be more accurate in the latter case. The practical interest of this approach is illustrated by calculating the parameters (and the corresponding genotype) associated with yield optimization of a GREENLAB maize model. The paper discusses the potentials of GREENLAB to represent environment × genotype

  9. Quantitation of HIV-1 by real-time amplification and detection

    NASA Astrophysics Data System (ADS)

    Jung, Paul M.; Yang, Naiquan; Kroeger, Paul E.

    1998-04-01

    A model assay for HIV-1 using a non-competitive internal standard in quantitative RT-PCR was coupled with real-time detection of both analyte and internal standard (IS) signals in a closed system. Real-time detection by the PE-ABI Prism 7700 relied on TaqMan probes specific for HIV and IS. The exogenous, non-competitive IS RNA was added in the same, known amount to a series of HIV RNA standards. The threshold cycle ratio from this internal standard calibration curve was used in the quantitation of HIV. Two configurations of reporter labels were compared. The HEX-HIV:FAM-IS system was the most precise, with nearly half-log discrimination over a range of 102 through 105 copies HIV-1 RNA. The FAM- HIV:HEX-IS system was less precise, but more sensitive and resistant to sample inhibition. The analysis of these signals and their impact on the range and precision of HIV quantitation is discussed. The design and synthesis of the fluorescently-labelled probes is also described.

  10. Calibration-free quantitation in microchip zone electrophoresis with conductivity detection.

    PubMed

    Noblitt, Scott D; Henry, Charles S

    2015-08-01

    The relationship between electrophoretic mobility and molar conductivity has previously led to speculation on achieving quantitation in zone electrophoresis without calibration curves when using conductivity detection. However, little work in this area has been pursued, possibly because of the breakdown of simple sensitivity-mobility relationships when working with partially protonated species. This topic is revisited with the aid of electrophoretic simulation software that produces facile predictions of analyte sensitivity relative to an internal standard. Calibration curve slopes for over 50 analyte/internal standard/BGE combinations were measured with both unbiased and electrokinetically biased injections using microchip electrophoresis with conductivity detection. The results were compared to theoretical expectations as computed with PeakMaster software. Good agreement was observed, with some systems being predicted with quantitative accuracy while others showed significant deviations. Some mechanisms that can lead to deviations from theory are demonstrated, but the causes for some discrepancies are still not understood. Overall, this work exhibits another useful application for simulation software, particularly for disposable devices where device-specific calibration curves cannot be collected. It also serves as quantitative validation for some outputs of PeakMaster simulation software.

  11. Slot immunoblot assay for detection and quantitation of periodontal disease-associated microorganisms in dental plaque.

    PubMed Central

    van Poperin, N; Lopatin, D E

    1991-01-01

    A rapid method for qualitative and quantitative detection of specific oral microorganisms from subgingival dental plaque is described. Plaque samples were suspended in phosphate-buffered saline containing protease inhibitors and 0.5% formaldehyde, briefly sonicated to disperse bacterial aggregates, and applied to nitrocellulose membranes in a slot blot manifold. Subsequent incubations with species-specific rabbit antibody and anti-rabbit antibody-alkaline phosphatase conjugate and development with BCIP-NBT substrate resulted in an easily discernible, permanent stain being deposited at the sample application site. Comparison with known concentrations of pure, cultured microorganisms applied to the same membranes permitted qualitative or semiquantitative plaque characterization by visual inspection. Analysis of the blots with a computer-linked flatbed scanner provided quantitative data on microbial content. The reproducibility of the results (standard error of the mean, less than 10%) obtained with slot immunoblotting greatly exceeded that of the results obtained with immunofluorescence analysis (standard error of the mean, greater than 57%). Because it is versatile, rapid, sensitive, reproducible, permanent, and relatively inexpensive, slot immunoblotting lends itself to use in large-scale investigations for the detection and quantitation of specific microbial species. PMID:1663511

  12. Development of an effective method for recovery of viral genomic RNA from environmental silty sediments for quantitative molecular detection.

    PubMed

    Miura, Takayuki; Masago, Yoshifumi; Sano, Daisuke; Omura, Tatsuo

    2011-06-01

    Nine approaches to recover viral RNA from environmental silty sediments were newly developed and compared to quantify RNA viruses in sediments using molecular methods. Four of the nine approaches employed direct procedures for extracting RNA from sediments (direct methods), and the remaining five approaches used indirect methods wherein viral particles were recovered before RNA extraction. A direct method using an SDS buffer with EDTA to lyse viral capsids in sediments, phenol-chloroform-isoamyl alcohol to extract RNA, isopropanol to concentrate RNA, and magnetic beads to purify RNA resulted in the highest rate of recovery (geometric mean of 11%, with a geometric standard deviation of 0.02; n = 7) of poliovirus 1 (PV1) inoculated in an environmental sediment sample. The direct method exhibiting the highest rate of PV1 recovery was applied to environmental sediment samples. One hundred eight sediment samples were collected from the Takagi River, Miyagi, Japan, and its estuary from November 2007 to April 2009, and the genomic RNAs of enterovirus and human norovirus in these samples were quantified by reverse transcription (RT)-quantitative PCR (qPCR). The human norovirus genome was detected in one sample collected at the bay, although its concentration was below the quantification limit. Meanwhile, the enterovirus genome was detected in two samples at the river mouth and river at concentrations of 8.6 × 10(2) and 2.4 × 10(2) copies/g (wet weight), respectively. This is the first report to obtain quantitative data for a human pathogenic virus in a river and in estuarine sediments using RT-qPCR.

  13. Automated Detection of P. falciparum Using Machine Learning Algorithms with Quantitative Phase Images of Unstained Cells.

    PubMed

    Park, Han Sang; Rinehart, Matthew T; Walzer, Katelyn A; Chi, Jen-Tsan Ashley; Wax, Adam

    2016-01-01

    Malaria detection through microscopic examination of stained blood smears is a diagnostic challenge that heavily relies on the expertise of trained microscopists. This paper presents an automated analysis method for detection and staging of red blood cells infected by the malaria parasite Plasmodium falciparum at trophozoite or schizont stage. Unlike previous efforts in this area, this study uses quantitative phase images of unstained cells. Erythrocytes are automatically segmented using thresholds of optical phase and refocused to enable quantitative comparison of phase images. Refocused images are analyzed to extract 23 morphological descriptors based on the phase information. While all individual descriptors are highly statistically different between infected and uninfected cells, each descriptor does not enable separation of populations at a level satisfactory for clinical utility. To improve the diagnostic capacity, we applied various machine learning techniques, including linear discriminant classification (LDC), logistic regression (LR), and k-nearest neighbor classification (NNC), to formulate algorithms that combine all of the calculated physical parameters to distinguish cells more effectively. Results show that LDC provides the highest accuracy of up to 99.7% in detecting schizont stage infected cells compared to uninfected RBCs. NNC showed slightly better accuracy (99.5%) than either LDC (99.0%) or LR (99.1%) for discriminating late trophozoites from uninfected RBCs. However, for early trophozoites, LDC produced the best accuracy of 98%. Discrimination of infection stage was less accurate, producing high specificity (99.8%) but only 45.0%-66.8% sensitivity with early trophozoites most often mistaken for late trophozoite or schizont stage and late trophozoite and schizont stage most often confused for each other. Overall, this methodology points to a significant clinical potential of using quantitative phase imaging to detect and stage malaria infection

  14. Automated Detection of P. falciparum Using Machine Learning Algorithms with Quantitative Phase Images of Unstained Cells

    PubMed Central

    Park, Han Sang; Rinehart, Matthew T.; Walzer, Katelyn A.; Chi, Jen-Tsan Ashley; Wax, Adam

    2016-01-01

    Malaria detection through microscopic examination of stained blood smears is a diagnostic challenge that heavily relies on the expertise of trained microscopists. This paper presents an automated analysis method for detection and staging of red blood cells infected by the malaria parasite Plasmodium falciparum at trophozoite or schizont stage. Unlike previous efforts in this area, this study uses quantitative phase images of unstained cells. Erythrocytes are automatically segmented using thresholds of optical phase and refocused to enable quantitative comparison of phase images. Refocused images are analyzed to extract 23 morphological descriptors based on the phase information. While all individual descriptors are highly statistically different between infected and uninfected cells, each descriptor does not enable separation of populations at a level satisfactory for clinical utility. To improve the diagnostic capacity, we applied various machine learning techniques, including linear discriminant classification (LDC), logistic regression (LR), and k-nearest neighbor classification (NNC), to formulate algorithms that combine all of the calculated physical parameters to distinguish cells more effectively. Results show that LDC provides the highest accuracy of up to 99.7% in detecting schizont stage infected cells compared to uninfected RBCs. NNC showed slightly better accuracy (99.5%) than either LDC (99.0%) or LR (99.1%) for discriminating late trophozoites from uninfected RBCs. However, for early trophozoites, LDC produced the best accuracy of 98%. Discrimination of infection stage was less accurate, producing high specificity (99.8%) but only 45.0%-66.8% sensitivity with early trophozoites most often mistaken for late trophozoite or schizont stage and late trophozoite and schizont stage most often confused for each other. Overall, this methodology points to a significant clinical potential of using quantitative phase imaging to detect and stage malaria infection

  15. Endotoxin Detection in Pharmaceuticals and Medical Devices with Kinetic-QCL, a Kinetic-Quantitative Chromogenic Limulus Amebocyte Lysate Assay.

    PubMed

    Berzofsky, Ronald N.

    1995-01-01

    The observation that endotoxin caused gelation in extracts of Limulus amebocytes has been expanded to the development of an in vitro kinetic, quantitative chromogenic LAL assay (Kinetic-QCL) for the detection of endotoxin in aqueous fluids. Within the last 15 years, the use of Limulus amebocyte lysate to detect and control the presence of pyrogenic substances in pharmaceuticals and medical devices has gained wide international acceptance. Both the United States and European Pharmacopoeias contain descriptions of and requirements for the LAL Bacterial Endotoxin Test. Both pharmacopoeias have begun to remove the rabbit pyrogen test requirement in a majority of drug monographs and have substituted endotoxin limits to be determined by LAL. The use of LAL has proved invaluable in controlling the level of endotoxin in finished product. The endotoxin contribution of raw materials and packaging material can be monitored as well. In-process testing at critical production steps can identify additional sources of endotoxin contamination, and depyrogenation processes can be validated by quantitating the degradation of endotoxin challenges. The speed, reproducibility, sensitivity, and economics of the Kinetic-QCL assay, in conjunction with the ppropriate equipment and software, over both the in vivo rabbit pyrogen test and the more traditional LAL gel-clot assay allow a more in-depth approach to the control of endotoxin in pharmaceuticals and medical devices.

  16. Quantitative evaluation of in vivo vital-dye fluorescence endoscopic imaging for the detection of Barrett's-associated neoplasia

    NASA Astrophysics Data System (ADS)

    Thekkek, Nadhi; Lee, Michelle H.; Polydorides, Alexandros D.; Rosen, Daniel G.; Anandasabapathy, Sharmila; Richards-Kortum, Rebecca

    2015-05-01

    Current imaging tools are associated with inconsistent sensitivity and specificity for detection of Barrett's-associated neoplasia. Optical imaging has shown promise in improving the classification of neoplasia in vivo. The goal of this pilot study was to evaluate whether in vivo vital dye fluorescence imaging (VFI) has the potential to improve the accuracy of early-detection of Barrett's-associated neoplasia. In vivo endoscopic VFI images were collected from 65 sites in 14 patients with confirmed Barrett's esophagus (BE), dysplasia, or esophageal adenocarcinoma using a modular video endoscope and a high-resolution microendoscope (HRME). Qualitative image features were compared to histology; VFI and HRME images show changes in glandular structure associated with neoplastic progression. Quantitative image features in VFI images were identified for objective image classification of metaplasia and neoplasia, and a diagnostic algorithm was developed using leave-one-out cross validation. Three image features extracted from VFI images were used to classify tissue as neoplastic or not with a sensitivity of 87.8% and a specificity of 77.6% (AUC=0.878). A multimodal approach incorporating VFI and HRME imaging can delineate epithelial changes present in Barrett's-associated neoplasia. Quantitative analysis of VFI images may provide a means for objective interpretation of BE during surveillance.

  17. Nanomolar colorimetric quantitative detection of Fe3 + and PPi with high selectivity

    NASA Astrophysics Data System (ADS)

    Li, Zhanxian; Li, Haixia; Shi, Caixia; Yu, Mingming; Wei, Liuhe; Ni, Zhonghai

    2016-04-01

    A novel rhodamine and 8-hydroxyquinoline-based derivative was synthesized, which is shown to act as a colorimetric chemosensor for Fe3 + in aqueous solution with high selectivity over various environmentally and biologically relevant metal ions and anions with a distinct color change from colorless to pink in very fast response time (< 1 min). Fe3 + can be detected quantitatively in the concentration range from 6.7 to 16 μM and the detection limit (LOD) on UV-vis response of the sensor can be as low as 15 nM. The 'in situ' prepared Fe3 + complex (1 ṡ Fe) showed high selectivity toward PPi against many common anions, and sensitivity (the LOD can be as low as 71 nM). In addition, both the chemosensor and the 'in situ' prepared Fe3 + complex are reusable for the detection of Fe3 + and PPi respectively.

  18. High resolution quantitative phase imaging of live cells with constrained optimization approach

    NASA Astrophysics Data System (ADS)

    Pandiyan, Vimal Prabhu; Khare, Kedar; John, Renu

    2016-03-01

    Quantitative phase imaging (QPI) aims at studying weakly scattering and absorbing biological specimens with subwavelength accuracy without any external staining mechanisms. Use of a reference beam at an angle is one of the necessary criteria for recording of high resolution holograms in most of the interferometric methods used for quantitative phase imaging. The spatial separation of the dc and twin images is decided by the reference beam angle and Fourier-filtered reconstructed image will have a very poor resolution if hologram is recorded below a minimum reference angle condition. However, it is always inconvenient to have a large reference beam angle while performing high resolution microscopy of live cells and biological specimens with nanometric features. In this paper, we treat reconstruction of digital holographic microscopy images as a constrained optimization problem with smoothness constraint in order to recover only complex object field in hologram plane even with overlapping dc and twin image terms. We solve this optimization problem by gradient descent approach iteratively and the smoothness constraint is implemented by spatial averaging with appropriate size. This approach will give excellent high resolution image recovery compared to Fourier filtering while keeping a very small reference angle. We demonstrate this approach on digital holographic microscopy of live cells by recovering the quantitative phase of live cells from a hologram recorded with nearly zero reference angle.

  19. [Self-perception of oral health and impact on quality of life among the elderly: a quantitative-qualitative approach].

    PubMed

    Haikal, Desirée Sant'Ana; Paula, Alfredo Maurício Batista de; Martins, Andrea Maria Eleutério de Barros Lima; Moreira, Allyson Nogueira; Ferreira, Efigênia Ferreira e

    2011-07-01

    A qualitative-quantitative approach was used in this study to obtain a clearer understanding of the relationship between self-perception, impact on quality of life and oral health among the elderly. Clinical examination and recorded interviews with objective and discursive questions were conducted with 45 institutionalized elderly people. Descriptive analyses of quantitative data were made. The interviews were transcribed and a systematic reading of the interviews was carried out selecting the components related to the categories under analysis. Photographic images of the oral clinical status were correlated with participants' speech. Quantitative analysis revealed: an average of 4.8 teeth; DMFT were 29.9; 57.7 % were toothless; 60% believed they did not need dental care; 75% suffered a great impact on quality of life due to oral health conditions, despite the fact that 67% evaluated their oral health positively. Underestimation of symptoms, lack of hope and resignation due to limitations regarding poor clinical status were detected. Most elderly people viewed such limitations as a consequence of aging and not as a problem that may be solved. This reality can be changed through information and guidance for elderly people.

  20. Rapid Detection of Ceratocystis platani Inoculum by Quantitative Real-Time PCR Assay

    PubMed Central

    Ghelardini, Luisa; Belbahri, Lassaâd; Quartier, Marion; Santini, Alberto

    2013-01-01

    Ceratocystis platani is the causal agent of canker stain of plane trees, a lethal disease able to kill mature trees in one or two successive growing seasons. The pathogen is a quarantine organism and has a negative impact on anthropogenic and natural populations of plane trees. Contaminated sawdust produced during pruning and sanitation fellings can contribute to disease spread. The goal of this study was to design a rapid, real-time quantitative PCR assay to detect a C. platani airborne inoculum. Airborne inoculum traps (AITs) were placed in an urban setting in the city of Florence, Italy, where the disease was present. Primers and TaqMan minor groove binder (MGB) probes were designed to target cerato-platanin (CP) and internal transcribed spacer 2 (ITS2) genes. The detection limits of the assay were 0.05 pg/μl and 2 fg/μl of fungal DNA for CP and ITS, respectively. Pathogen detection directly from AITs demonstrated specificity and high sensitivity for C. platani, detecting DNA concentrations as low as 1.2 × 10−2 to 1.4 × 10−2 pg/μl, corresponding to ∼10 conidia per ml. Airborne inoculum traps were able to detect the C. platani inoculum within 200 m of the closest symptomatic infected plane tree. The combination of airborne trapping and real-time quantitative PCR assay provides a rapid and sensitive method for the specific detection of a C. platani inoculum. This technique may be used to identify the period of highest risk of pathogen spread in a site, thus helping disease management. PMID:23811499

  1. Molecular detection of Mikrocytos mackini in Pacific oysters using quantitative PCR.

    PubMed

    Polinski, Mark; Lowe, Geoff; Meyer, Gary; Corbeil, Serge; Colling, Axel; Caraguel, Charles; Abbott, Cathryn L

    2015-01-01

    Mikrocytos mackini is an internationally regulated pathogen and causative agent of Denman Island disease in Pacific oysters Crassostrea gigas. Recent phylogenetic breakthroughs have placed this parasite within a highly divergent and globally distributed eukaryotic lineage that has been designated a new taxonomic order, Mikrocytida. The discovery of this new radiation of parasites is accompanied by a heightened awareness of the many knowledge gaps that exist with respect to the general biology, epizootiology, and potential impact of mikrocytid parasites on hosts, ecosystems, and commercial fisheries. It has also highlighted current shortcomings regarding our ability to detect these organisms. In this study, we developed a species-specific, sensitive, and quantitative method for detecting M. mackini DNA from host tissues using probe-based real-time qPCR technology. A limit of sensitivity between 2 and 5 genome copy equivalents was achieved in a reaction matrix containing ≥ 40 ng/μL host gDNA without inhibition. This detection proved superior to existing methods based on conventional PCR, histology or gross pathology and is the first species-specific diagnostic test for M. mackini. Quantitative assessment of parasite DNA using this assay remained accurate to between 10 and 50 copies identifying that during infection, M. mackini DNA was significantly more prevalent in hemolymph, labial palp, and mid-body cross-sections compared to mantle or adductor muscle. DNA extracted from a mid-body cross-section also provided the highest likelihood for detection during diagnostic screening of infected oysters. Taken together, these findings provide strong analytical evidence for the adoption of qPCR as the new reference standard for detecting M. mackini and give preliminary insight into the distribution of the parasite within host tissues. Standardised operating methodologies for sample collection and qPCR testing are provided to aid in the international regulatory diagnosis of

  2. Food allergen detection methods: a coordinated approach.

    PubMed

    Goodwin, Philip R

    2004-01-01

    The levels (1-2%) and increasing severity of allergic responses to food in the adult population are well documented, as is the phenomenon of even higher (3-8%) and apparently increasing incidence in children, albeit that susceptibility decreases with age. Problematic foods include peanut, milk, eggs, tree nuts, and sesame, but the list is growing as awareness continues to rise. The amounts of such foods that can cause allergic reactions is difficult to gauge; however, the general consensus is that ingestion of low parts per million is sufficient to cause severe reactions in badly affected individuals. Symptoms can rapidly-within minutes-progress from minor discomfort to severe, even life-threatening anaphylactic shock in those worst affected. Given the combination of high incidence of atopy, potential severity of response, and apparently widespread instances of "hidden" allergens in the food supply, it is not surprising that this issue is increasingly subject to legislative and regulatory scrutiny. In order to assist in the control of allergen levels in foods to acceptable levels, analysts require a combination of test methods, each designed to produce accurate, timely, and cost-effective analytical information. Such information contributes significantly to Hazard Analysis Critical Control Point programs to determine food manufacturers' risk and improves the accuracy of monitoring and surveillance by food industry, commercial, and enforcement laboratories. Analysis thereby facilitates improvements in compliance with labeling laws with concomitant reductions in risks to atopic consumers. This article describes a combination of analytical approaches to fulfill the various needs of these 3 analytical communities.

  3. Rapid simultaneous detection and quantitation of infectious pancreatic necrosis virus (IPNV).

    PubMed

    Espinoza, Juan Carlos; Kuznar, Juan

    2002-08-01

    Infectious pancreatic necrosis virus (IPNV) is a pathogen of great concern in the salmon industry as well as in the environment. Taking advantage of the early immunofluorescent visualization of viral proteins in infected cells, a titration method was developed. At 16 h p.i., fluorescent foci were visualized with a monoclonal antibody against VP3-structural protein of the virus. The counting of each fluorescent cell allows the quantitation of infection foci; titres expressed in fluorescent foci/ml were equivalent to plaque forming units (PFU)/ml. With slight modifications, the same method used to detect the virus in field samples, can be applied to estimate virus contents. Some of the samples used during the assays were obtained from routine screening procedures. The titres recorded from positive samples correlated well with the clinical condition of the fish. With this method, rapid diagnosis and quantitation may simultaneously be performed with the same tissue extract.

  4. Qualitative and Quantitative Assays for Detection and Characterization of Protein Antimicrobials.

    PubMed

    Farris, M Heath; Ford, Kara A; Doyle, Richard C

    2016-01-01

    Initial evaluations of large microbial libraries for potential producers of novel antimicrobial proteins require both qualitative and quantitative methods to screen for target enzymes prior to investing greater research effort and resources. The goal of this protocol is to demonstrate two complementary assays for conducting these initial evaluations. The microslide diffusion assay provides an initial or simple detection screen to enable the qualitative and rapid assessment of proteolytic activity against an array of both viable and heat-killed bacterial target substrates. As a counterpart, the increased sensitivity and reproducibility of the dye-release assay provides a quantitative platform for evaluating and comparing environmental influences affecting the hydrolytic activity of protein antimicrobials. The ability to label specific heat-killed cell culture substrates with Remazol brilliant blue R dye expands this capability to tailor the dye-release assay to characterize enzymatic activity of interest.

  5. Qualitative and Quantitative Assays for Detection and Characterization of Protein Antimicrobials

    PubMed Central

    Farris, M. Heath; Ford, Kara A.; Doyle, Richard C.

    2016-01-01

    Initial evaluations of large microbial libraries for potential producers of novel antimicrobial proteins require both qualitative and quantitative methods to screen for target enzymes prior to investing greater research effort and resources. The goal of this protocol is to demonstrate two complementary assays for conducting these initial evaluations. The microslide diffusion assay provides an initial or simple detection screen to enable the qualitative and rapid assessment of proteolytic activity against an array of both viable and heat-killed bacterial target substrates. As a counterpart, the increased sensitivity and reproducibility of the dye-release assay provides a quantitative platform for evaluating and comparing environmental influences affecting the hydrolytic activity of protein antimicrobials. The ability to label specific heat-killed cell culture substrates with Remazol brilliant blue R dye expands this capability to tailor the dye-release assay to characterize enzymatic activity of interest. PMID:27166738

  6. Photofragmentation approaches for the detection of polyatomic molecules.

    SciTech Connect

    Headrick, Jeffrey M.; Hoops, Alexandra A.; Kulp, Thomas J.; Bisson, Scott E.; Reichardt, Thomas A.; Farrow, Roger L.; Settersten, Thomas B.

    2010-05-01

    We review three photofragmentation detection approaches, describing the detection of (1) vapor-phase mercuric chloride by photofragment emission, (2) vapor-phase nitro-containing compounds by photofragmentation-ionization, and (3) surface-bound organophosphonate compounds by photofragmentation-laser-induced fluorescence.

  7. Target identification of natural and traditional medicines with quantitative chemical proteomics approaches.

    PubMed

    Wang, Jigang; Gao, Liqian; Lee, Yew Mun; Kalesh, Karunakaran A; Ong, Yong Siang; Lim, Jaehong; Jee, Joo-Eun; Sun, Hongyan; Lee, Su Seong; Hua, Zi-Chun; Lin, Qingsong

    2016-06-01

    Natural and traditional medicines, being a great source of drugs and drug leads, have regained wide interests due to the limited success of high-throughput screening of compound libraries in the past few decades and the recent technology advancement. Many drugs/bioactive compounds exert their functions through interaction with their protein targets, with more and more drugs showing their ability to target multiple proteins, thus target identification has an important role in drug discovery and biomedical research fields. Identifying drug targets not only furthers the understanding of the mechanism of action (MOA) of a drug but also reveals its potential therapeutic applications and adverse side effects. Chemical proteomics makes use of affinity chromatography approaches coupled with mass spectrometry to systematically identify small molecule-protein interactions. Although traditional affinity-based chemical proteomics approaches have made great progress in the identification of cellular targets and elucidation of MOAs of many bioactive molecules, nonspecific binding remains a major issue which may reduce the accuracy of target identification and may hamper the drug development process. Recently, quantitative proteomics approaches, namely, metabolic labeling, chemical labeling, or label-free approaches, have been implemented in target identification to overcome such limitations. In this review, we will summarize and discuss the recent advances in the application of various quantitative chemical proteomics approaches for the identification of targets of natural and traditional medicines. PMID:26808165

  8. Event-specific quantitative detection of nine genetically modified maizes using one novel standard reference molecule.

    PubMed

    Yang, Litao; Guo, Jinchao; Pan, Aihu; Zhang, Haibo; Zhang, Kewei; Wang, Zhengming; Zhang, Dabing

    2007-01-10

    With the development of genetically modified organism (GMO) detection techniques, the Polymerase Chain Reaction (PCR) technique has been the mainstay for GMO detection, and real-time PCR is the most effective and important method for GMO quantification. An event-specific detection strategy based on the unique and specific integration junction sequences between the host plant genome DNA and the integrated gene is being developed for its high specificity. This study establishes the event-specific detection methods for TC1507 and CBH351 maizes. In addition, the event-specific TaqMan real-time PCR detection methods for another seven GM maize events (Bt11, Bt176, GA21, MON810, MON863, NK603, and T25) were systematically optimized and developed. In these PCR assays, the fluorescent quencher, TAMRA, was dyed on the T-base of the probe at the internal position to improve the intensity of the fluorescent signal. To overcome the difficulties in obtaining the certified reference materials of these GM maizes, one novel standard reference molecule containing all nine specific integration junction sequences of these GM maizes and the maize endogenous reference gene, zSSIIb, was constructed and used for quantitative analysis. The limits of detection of these methods were 20 copies for these different GM maizes, the limits of quantitation were about 20 copies, and the dynamic ranges for quantification were from 0.05 to 100% in 100 ng of DNA template. Furthermore, nine groups of the mixed maize samples of these nine GM maize events were quantitatively analyzed to evaluate the accuracy and precision. The accuracy expressed as bias varied from 0.67 to 28.00% for the nine tested groups of GM maize samples, and the precision expressed as relative standard deviations was from 0.83 to 26.20%. All of these indicated that the established event-specific real-time PCR detection systems and the reference molecule in this study are suitable for the identification and quantification of these GM

  9. Event-specific quantitative detection of nine genetically modified maizes using one novel standard reference molecule.

    PubMed

    Yang, Litao; Guo, Jinchao; Pan, Aihu; Zhang, Haibo; Zhang, Kewei; Wang, Zhengming; Zhang, Dabing

    2007-01-10

    With the development of genetically modified organism (GMO) detection techniques, the Polymerase Chain Reaction (PCR) technique has been the mainstay for GMO detection, and real-time PCR is the most effective and important method for GMO quantification. An event-specific detection strategy based on the unique and specific integration junction sequences between the host plant genome DNA and the integrated gene is being developed for its high specificity. This study establishes the event-specific detection methods for TC1507 and CBH351 maizes. In addition, the event-specific TaqMan real-time PCR detection methods for another seven GM maize events (Bt11, Bt176, GA21, MON810, MON863, NK603, and T25) were systematically optimized and developed. In these PCR assays, the fluorescent quencher, TAMRA, was dyed on the T-base of the probe at the internal position to improve the intensity of the fluorescent signal. To overcome the difficulties in obtaining the certified reference materials of these GM maizes, one novel standard reference molecule containing all nine specific integration junction sequences of these GM maizes and the maize endogenous reference gene, zSSIIb, was constructed and used for quantitative analysis. The limits of detection of these methods were 20 copies for these different GM maizes, the limits of quantitation were about 20 copies, and the dynamic ranges for quantification were from 0.05 to 100% in 100 ng of DNA template. Furthermore, nine groups of the mixed maize samples of these nine GM maize events were quantitatively analyzed to evaluate the accuracy and precision. The accuracy expressed as bias varied from 0.67 to 28.00% for the nine tested groups of GM maize samples, and the precision expressed as relative standard deviations was from 0.83 to 26.20%. All of these indicated that the established event-specific real-time PCR detection systems and the reference molecule in this study are suitable for the identification and quantification of these GM

  10. Metallothionein: structure/antigenicity and detection/quantitation in normal physiological fluids.

    PubMed Central

    Garvey, J S

    1984-01-01

    Recent experiments in the application of radioimmunoassay (RIA) in the detection and quantitation of metallothionein (MT) in human sera and urines demonstrate that it is possible to extend the lower limit of practical quantitation from the previous limit of 50-100 pg to 1 pg.RIA of normal sera indicates that the typical range of concentrations of MT is from less than 0.01 ng/mL to about 1 ng/mL, and that concentrations above 2 ng/mL should be considered abnormal. The typical range for normal urines is from less than 1 ng/mL to 10 ng/mL; concentrations above 10 ng/mL should be considered abnormal. A complementary assay, the enzyme-linked immunosorbent assay (ELISA), is under development. The ELISA is a competitive binding assay, detection and quantitation of MT being either by colorimetric or fluorimetric methods. The present useful range for MT quantitation in the ELISA is from about 50-50000 pg (fluorimetric) or 500-5000 pg (colorimetric). Recent experiments using the RIA have identified the principal antigenic determinants of vertebrate MTs as involving the immediate amino terminal residues (-MDPNC-) and the segment including residues 20-25 (-KCKECK- in human MT). Theoretical predictions of secondary structure based on hydrophilicity and sequence analysis indicate that the conformational profile is dominated by tetrapeptide candidates for beta turns (reverse turns) with 2-3 hexapeptide sequences being candidates for helical conformation and 4-5 short sequences (3-5 residues) being candidates for beta chain conformation. The helical candidates are predicted to be unstable and the analysis favors reverse turns for both determinants of vertebrate MT and a sequestered location for the joining region between clusters A and B. PMID:6203731

  11. Comparison of nine different real-time PCR chemistries for qualitative and quantitative applications in GMO detection.

    PubMed

    Buh Gasparic, Meti; Tengs, Torstein; La Paz, Jose Luis; Holst-Jensen, Arne; Pla, Maria; Esteve, Teresa; Zel, Jana; Gruden, Kristina

    2010-03-01

    Several techniques have been developed for detection and quantification of genetically modified organisms, but quantitative real-time PCR is by far the most popular approach. Among the most commonly used real-time PCR chemistries are TaqMan probes and SYBR green, but many other detection chemistries have also been developed. Because their performance has never been compared systematically, here we present an extensive evaluation of some promising chemistries: sequence-unspecific DNA labeling dyes (SYBR green), primer-based technologies (AmpliFluor, Plexor, Lux primers), and techniques involving double-labeled probes, comprising hybridization (molecular beacon) and hydrolysis (TaqMan, CPT, LNA, and MGB) probes, based on recently published experimental data. For each of the detection chemistries assays were included targeting selected loci. Real-time PCR chemistries were subsequently compared for their efficiency in PCR amplification and limits of detection and quantification. The overall applicability of the chemistries was evaluated, adding practicability and cost issues to the performance characteristics. None of the chemistries seemed to be significantly better than any other, but certain features favor LNA and MGB technology as good alternatives to TaqMan in quantification assays. SYBR green and molecular beacon assays can perform equally well but may need more optimization prior to use.

  12. Comparison of nine different real-time PCR chemistries for qualitative and quantitative applications in GMO detection.

    PubMed

    Buh Gasparic, Meti; Tengs, Torstein; La Paz, Jose Luis; Holst-Jensen, Arne; Pla, Maria; Esteve, Teresa; Zel, Jana; Gruden, Kristina

    2010-03-01

    Several techniques have been developed for detection and quantification of genetically modified organisms, but quantitative real-time PCR is by far the most popular approach. Among the most commonly used real-time PCR chemistries are TaqMan probes and SYBR green, but many other detection chemistries have also been developed. Because their performance has never been compared systematically, here we present an extensive evaluation of some promising chemistries: sequence-unspecific DNA labeling dyes (SYBR green), primer-based technologies (AmpliFluor, Plexor, Lux primers), and techniques involving double-labeled probes, comprising hybridization (molecular beacon) and hydrolysis (TaqMan, CPT, LNA, and MGB) probes, based on recently published experimental data. For each of the detection chemistries assays were included targeting selected loci. Real-time PCR chemistries were subsequently compared for their efficiency in PCR amplification and limits of detection and quantification. The overall applicability of the chemistries was evaluated, adding practicability and cost issues to the performance characteristics. None of the chemistries seemed to be significantly better than any other, but certain features favor LNA and MGB technology as good alternatives to TaqMan in quantification assays. SYBR green and molecular beacon assays can perform equally well but may need more optimization prior to use. PMID:20087729

  13. Methods for the Specific Detection and Quantitation of Amyloid-β Oligomers in Cerebrospinal Fluid.

    PubMed

    Schuster, Judith; Funke, Susanne Aileen

    2016-05-01

    Protein misfolding and aggregation are fundamental features of the majority of neurodegenerative diseases, like Alzheimer's disease (AD), Parkinson's disease, frontotemporal dementia, and prion diseases. Proteinaceous deposits in the brain of the patient, e.g., amyloid plaques consisting of the amyloid-β (Aβ) peptide and tangles composed of tau protein, are the hallmarks of AD. Soluble oligomers of Aβ and tau play a fundamental role in disease progression, and specific detection and quantification of the respective oligomeric proteins in cerebrospinal fluid may provide presymptomatically detectable biomarkers, paving the way for early diagnosis or even prognosis. Several studies on the development of techniques for the specific detection of Aβ oligomers were published, but some of the existing tools do not yet seem to be satisfactory, and the study results are contradicting. The detection of oligomers is challenging due to their polymorphous and unstable nature, their low concentration, and the presence of competing proteins and Aβ monomers in body fluids. Here, we present an overview of the current state of the development of methods for Aβ oligomer specific detection and quantitation. The methods are divided in the three subgroups: (i) enzyme linked immunosorbent assays (ELISA), (ii) methods for single oligomer detection, and (iii) others, which are mainly biosensor based methods. PMID:27163804

  14. Detection of Thielaviopsis basicola in soil with real-time quantitative PCR assays.

    PubMed

    Huang, Junli; Kang, Zhenhui

    2010-07-20

    Thielaviopsis basicola is a soil-borne fungus with a wide host range and a cosmopolitan distribution. It causes disease on many agricultural crops and in China it is the causal agent of black root rot on tobacco plant. Early diagnosis and detection of the pathogen in soil are critical to control this disease in field. The objective of this study was to develop sensitive and effective methods suitable for large-scale detection and quantification of T. basicola. Based on the nucleotide sequences of the internal transcribed spacer (ITS) regions of rDNA genes of Thielaviopsis spp, primers and TaqMan probe were designed specifically to amplify DNA from T. basicola and real-time, quantitative PCR (qPCR) assays were developed for rapid, specific and sensitive detection and quantification of T. basicola. It was sensitive with the detection limit of 100 fg microl(-1) genomic DNA of T. basicola in qPCR assays. By combining the qPCR assays with the efficient protocol to extract DNA from soil, it was possible to achieve real-time detection of T. basicola in soil in 4-5 h and the detection limit of 3 conidia per reaction in qPCR was recorded. The assays were applied to survey soils from tobacco fields in China for the presence of T. basicola and the analyses of naturally infested soil showed the reliability of the qPCR assays.

  15. Quantitative detection of promoter hypermethylation of multiple genes in the tumor, urine, and serum DNA of patients with renal cancer.

    PubMed

    Hoque, Mohammad Obaidul; Begum, Shahnaz; Topaloglu, Ozlem; Jeronimo, Carmen; Mambo, Elizabeth; Westra, William H; Califano, J A; Sidransky, David

    2004-08-01

    Aberrant promoter hypermethylation of several known or putative tumor suppressor genes occurs frequently during the pathogenesis of human cancers and is a promising marker for cancer detection. We investigated the feasibility of detecting aberrant DNA methylation in the urine and serum samples of renal cancer patients. We examined the tumor and the matched urine and serum DNA for aberrant methylation of nine gene promoters (CDH1, APC, MGMT, RASSF1A, GSTP1, p16, RAR-beta2, and ARF) from 17 patients with primary kidney cancer by quantitative fluorogenic real-time PCR. An additional 9 urine samples (total, 26) and 1 serum sample (total, 18) also were tested from renal cancer patients. Urine from 91 patients without genitourinary cancer and serum from 30 age-matched noncancer individuals were used as controls. Promoter hypermethylation of at least two of the genes studied was detected in 16 (94%) of 17 primary tumors. Aberrant methylation in urine and serum DNA generally was accompanied by methylation in the matched tumor samples. Urine samples from 91 control subjects without evidence of genitourinary cancer revealed no methylation of the MGMT, GSTP1, p16, and ARF genes, whereas methylation of RAR-beta2, RASSF1A, CDH1, APC, and TIMP3 was detected at low levels in a few control subjects. Overall, 23 (88%) of 26 urine samples and 12 (67%) of 18 serum samples from cancer patients were methylation positive for at least one of the genes tested. By combination of urine or serum analysis of renal cancer patients, hypermethylation was detected in 16 of 17 patients (94% sensitivity) with high specificity. Our findings suggest that promoter hypermethylation in urine or serum can be detected in the majority of renal cancer patients. This noninvasive high-throughput approach needs to be evaluated in large studies to assess its value in the early detection and surveillance of renal cancer.

  16. A new approach for quantitative analysis of L-phenylalanine using a novel semi-sandwich immunometric assay.

    PubMed

    Kubota, Kazuyuki; Mizukoshi, Toshimi; Miyano, Hiroshi

    2013-10-01

    Here, we describe a novel method for L-phenylalanine analysis using a sandwich-type immunometric assay approach for use as a new method for amino acid analysis. To overcome difficulties of the preparation of high-affinity and selectivity monoclonal antibodies against L-phenylalanine and the inability to use sandwich-type immunometric assays due to their small molecular weight, three procedures were examined. First, amino groups of L-phenylalanine were modified by "N-Fmoc-L-cysteine" (FC) residues and the derivative (FC-Phe) was used as a hapten. Immunization of mice with bovine serum albumin/FC-Phe conjugate successfully yielded specific monoclonal anti-FC-Phe antibodies. Second, a new derivatization reagent, "biotin linker conjugate of FC-Phe N-succinimidyl ester" (FC(Biotin)-NHS), was synthesized to convert L-phenylalanine to FC-(Biotin)-Phe as a hapten structure. The biotin moiety linked to the thiol group of cysteine formed a second binding site for streptavidin/horseradish peroxidase (HRP) conjugates for optical detection. Third, a new semi-sandwich-type immunometric assay was established using pre-derivatized L-phenylalanine, the monoclonal anti-FC-Phe antibody, and streptavidin/HRP conjugate (without second antibody). Using the new "semi-sandwich" immunometric assay system, a detection limit of 35 nM (60 amol per analysis) and a detection range of 0.1-20 μM were attained using a standard L-phenylalanine solution. Rat plasma samples were analyzed to test reliability. Intra-day assay precision was within 6% of the coefficient of variation; inter-day variation was 0.1%. The recovery rates were from 92.4 to 123.7%. This is the first report of the quantitative determination of L-phenylalanine using a reliable semi-sandwich immunometric assay approach and will be applicable to the quantitative determination of other amino acids.

  17. Competitive PCR-ELISA protocols for the quantitative and the standardized detection of viral genomes.

    PubMed

    Musiani, Monica; Gallinella, Giorgio; Venturoli, Simona; Zerbini, Marialuisa

    2007-01-01

    Competitive PCR-ELISA combines competitive PCR with an ELISA to allow quantitative detection of PCR products. It is based on the inclusion of an internal standard competitor molecule that is designed to differ from the target by a short sequence of nucleotides. Once such a competitor molecule has been designed and constructed, target and competitor sequences are concurrently PCR-amplified, before hybridization to two different specific probes and determination of their respective OD values by ELISA. The target can be quantified in relation to a titration curve of different dilutions of the competitor. The competitor can alternatively be used at a unique optimal concentration to allow for standardized detection of the target sequence. PCR-ELISA can be performed in 1 d in laboratories without access to a real-time PCR thermocycler. This technique is applied in diagnostics to monitor the course of infections and drug efficacy. Competitive PCR-ELISA protocols for the quantitative and for the standardized detection of parvovirus B19 are detailed here as an example of the technique.

  18. Simple, Rapid and Inexpensive Quantitative Fluorescent PCR Method for Detection of Microdeletion and Microduplication Syndromes

    PubMed Central

    Stofanko, Martin; Gonçalves-Dornelas, Higgor; Cunha, Pricila Silva; Pena, Heloísa B.; Vianna-Morgante, Angela M.; Pena, Sérgio Danilo Junho

    2013-01-01

    Because of economic limitations, the cost-effective diagnosis of patients affected with rare microdeletion or microduplication syndromes is a challenge in developing countries. Here we report a sensitive, rapid, and affordable detection method that we have called Microdeletion/Microduplication Quantitative Fluorescent PCR (MQF-PCR). Our procedure is based on the finding of genomic regions with high homology to segments of the critical microdeletion/microduplication region. PCR amplification of both using the same primer pair, establishes competitive kinetics and relative quantification of amplicons, as happens in microsatellite-based Quantitative Fluorescence PCR. We used patients with two common microdeletion syndromes, the Williams-Beuren syndrome (7q11.23 microdeletion) and the 22q11.2 microdeletion syndromes and discovered that MQF-PCR could detect both with 100% sensitivity and 100% specificity. Additionally, we demonstrated that the same principle could be reliably used for detection of microduplication syndromes, by using patients with the Lubs (MECP2 duplication) syndrome and the 17q11.2 microduplication involving the NF1 gene. We propose that MQF-PCR is a useful procedure for laboratory confirmation of the clinical diagnosis of microdeletion/microduplication syndromes, ideally suited for use in developing countries, but having general applicability as well. PMID:23620743

  19. Qualitative and quantitative approaches in the dose-response assessment of genotoxic carcinogens.

    PubMed

    Fukushima, Shoji; Gi, Min; Kakehashi, Anna; Wanibuchi, Hideki; Matsumoto, Michiharu

    2016-05-01

    Qualitative and quantitative approaches are important issues in field of carcinogenic risk assessment of the genotoxic carcinogens. Herein, we provide quantitative data on low-dose hepatocarcinogenicity studies for three genotoxic hepatocarcinogens: 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and N-nitrosodiethylamine (DEN). Hepatocarcinogenicity was examined by quantitative analysis of glutathione S-transferase placental form (GST-P) positive foci, which are the preneoplastic lesions in rat hepatocarcinogenesis and the endpoint carcinogenic marker in the rat liver medium-term carcinogenicity bioassay. We also examined DNA damage and gene mutations which occurred through the initiation stage of carcinogenesis. For the establishment of points of departure (PoD) from which the cancer-related risk can be estimated, we analyzed the above events by quantitative no-observed-effect level and benchmark dose approaches. MeIQx at low doses induced formation of DNA-MeIQx adducts; somewhat higher doses caused elevation of 8-hydroxy-2'-deoxyquanosine levels; at still higher doses gene mutations occurred; and the highest dose induced formation of GST-P positive foci. These data indicate that early genotoxic events in the pathway to carcinogenesis showed the expected trend of lower PoDs for earlier events in the carcinogenic process. Similarly, only the highest dose of IQ caused an increase in the number of GST-P positive foci in the liver, while IQ-DNA adduct formation was observed with low doses. Moreover, treatment with DEN at low doses had no effect on development of GST-P positive foci in the liver. These data on PoDs for the markers contribute to understand whether genotoxic carcinogens have a threshold for their carcinogenicity. The most appropriate approach to use in low dose-response assessment must be approved on the basis of scientific judgment.

  20. Performance of two real-time PCR assays for hepatitis B virus DNA detection and quantitation.

    PubMed

    Kania, Dramane; Ottomani, Laure; Meda, Nicolas; Peries, Marianne; Dujols, Pierre; Bolloré, Karine; Rénier, Wendy; Viljoen, Johannes; Ducos, Jacques; Van de Perre, Philippe; Tuaillon, Edouard

    2014-06-01

    In-house developed real-time PCR (qPCR) techniques could be useful conjunctives to the management of hepatitis B virus (HBV) infection in resource-limited settings with high prevalence. Two qPCR assays (qPCR1 and qPCR2), based on primers/probes targeting conserved regions of the X and S genes of HBV respectively, were evaluated using clinical samples of varying HBV genotypes, and compared to the commercial Roche Cobas AmpliPrep/Cobas TaqMan HBV Test v2.0. The lower detection limit (LDL) was established at 104 IU/ml for qPCR1, and 91 IU/ml for qPCR2. Good agreement and correlation were obtained between the Roche assay and both qPCR assays (r = 0.834 for qPCR1; and r = 0.870 for qPCR2). Differences in HBV DNA load of > 0.5 Log10 IU/ml between the Roche and the qPCR assays were found in 49/122 samples of qPCR1, and 35/122 samples of qPCR2. qPCR1 tended to underestimate HBV DNA quantity in samples with a low viral load and overestimate HBV DNA concentration in samples with a high viral load when compared to the Roche test. Both molecular tools that were developed, used on an open real-time PCR system, were reliable for HBV DNA detection and quantitation. The qPCR2 performed better than the qPCR1 and had the additional advantage of various HBV genotype detection and quantitation. This low cost quantitative HBV DNA PCR assay may be an alternative solution when implementing national programmes to diagnose, monitor and treat HBV infection in low- to middle-income countries where testing for HBV DNA is not available in governmental health programmes.

  1. Reliability of quantitative real-time PCR for bacterial detection in cystic fibrosis airway specimens.

    PubMed

    Zemanick, Edith T; Wagner, Brandie D; Sagel, Scott D; Stevens, Mark J; Accurso, Frank J; Harris, J Kirk

    2010-11-30

    The cystic fibrosis (CF) airway microbiome is complex; polymicrobial infections are common, and the presence of fastidious bacteria including anaerobes make culture-based diagnosis challenging. Quantitative real-time PCR (qPCR) offers a culture-independent method for bacterial quantification that may improve diagnosis of CF airway infections; however, the reliability of qPCR applied to CF airway specimens is unknown. We sought to determine the reliability of nine specific bacterial qPCR assays (total bacteria, three typical CF pathogens, and five anaerobes) applied to CF airway specimens. Airway and salivary specimens from clinically stable pediatric CF subjects were collected. Quantitative PCR assay repeatability was determined using triplicate reactions. Split-sample measurements were performed to measure variability introduced by DNA extraction. Results from qPCR were compared to standard microbial culture for Pseudomonas aeruginosa, Staphylococcus aureus, and Haemophilus influenzae, common pathogens in CF. We obtained 84 sputa, 47 oropharyngeal and 27 salivary specimens from 16 pediatric subjects with CF. Quantitative PCR detected bacterial DNA in over 97% of specimens. All qPCR assays were highly reproducible at quantities≥10(2) rRNA gene copies/reaction with coefficient of variation less than 20% for over 99% of samples. There was also excellent agreement between samples processed in duplicate. Anaerobic bacteria were highly prevalent and were detected in mean quantities similar to that of typical CF pathogens. Compared to a composite gold standard, qPCR and culture had variable sensitivities for detection of P. aeruginosa, S. aureus and H. influenzae from CF airway samples. By reliably quantifying fastidious airway bacteria, qPCR may improve our understanding of polymicrobial CF lung infections, progression of lung disease and ultimately improve antimicrobial treatments.

  2. Reliability of Quantitative Real-Time PCR for Bacterial Detection in Cystic Fibrosis Airway Specimens

    PubMed Central

    Zemanick, Edith T.; Wagner, Brandie D.; Sagel, Scott D.; Stevens, Mark J.; Accurso, Frank J.; Harris, J. Kirk

    2010-01-01

    The cystic fibrosis (CF) airway microbiome is complex; polymicrobial infections are common, and the presence of fastidious bacteria including anaerobes make culture-based diagnosis challenging. Quantitative real-time PCR (qPCR) offers a culture-independent method for bacterial quantification that may improve diagnosis of CF airway infections; however, the reliability of qPCR applied to CF airway specimens is unknown. We sought to determine the reliability of nine specific bacterial qPCR assays (total bacteria, three typical CF pathogens, and five anaerobes) applied to CF airway specimens. Airway and salivary specimens from clinically stable pediatric CF subjects were collected. Quantitative PCR assay repeatability was determined using triplicate reactions. Split-sample measurements were performed to measure variability introduced by DNA extraction. Results from qPCR were compared to standard microbial culture for Pseudomonas aeruginosa, Staphylococcus aureus, and Haemophilus influenzae, common pathogens in CF. We obtained 84 sputa, 47 oropharyngeal and 27 salivary specimens from 16 pediatric subjects with CF. Quantitative PCR detected bacterial DNA in over 97% of specimens. All qPCR assays were highly reproducible at quantities ≥102 rRNA gene copies/reaction with coefficient of variation less than 20% for over 99% of samples. There was also excellent agreement between samples processed in duplicate. Anaerobic bacteria were highly prevalent and were detected in mean quantities similar to that of typical CF pathogens. Compared to a composite gold standard, qPCR and culture had variable sensitivities for detection of P. aeruginosa, S. aureus and H. influenzae from CF airway samples. By reliably quantifying fastidious airway bacteria, qPCR may improve our understanding of polymicrobial CF lung infections, progression of lung disease and ultimately improve antimicrobial treatments. PMID:21152087

  3. Detection, monitoring, and quantitative analysis of wildfires with the BIRD satellite

    NASA Astrophysics Data System (ADS)

    Oertel, Dieter A.; Briess, Klaus; Lorenz, Eckehard; Skrbek, Wolfgang; Zhukov, Boris

    2004-02-01

    Increasing concern about environment and interest to avoid losses led to growing demands on space borne fire detection, monitoring and quantitative parameter estimation of wildfires. The global change research community intends to quantify the amount of gaseous and particulate matter emitted from vegetation fires, peat fires and coal seam fires. The DLR Institute of Space Sensor Technology and Planetary Exploration (Berlin-Adlershof) developed a small satellite called BIRD (Bi-spectral Infrared Detection) which carries a sensor package specially designed for fire detection. BIRD was launched as a piggy-back satellite on October 22, 2001 with ISRO"s Polar Satellite Launch Vehicle (PSLV). It is circling the Earth on a polar and sun-synchronous orbit at an altitude of 572 km and it is providing unique data for detailed analysis of high temperature events on Earth surface. The BIRD sensor package is dedicated for high resolution and reliable fire recognition. Active fire analysis is possible in the sub-pixel domain. The leading channel for fire detection and monitoring is the MIR channel at 3.8 μm. The rejection of false alarms is based on procedures using MIR/NIR (Middle Infra Red/Near Infra Red) and MIR/TIR (Middle Infra Red/Thermal Infra Red) radiance ratio thresholds. Unique results of BIRD wildfire detection and analysis over fire prone regions in Australia and Asia will be presented. BIRD successfully demonstrates innovative fire recognition technology for small satellites which permit to retrieve quantitative characteristics of active burning wildfires, such as the equivalent fire temperature, fire area, radiative energy release, fire front length and fire front strength.

  4. Quantitative detection of chemical compounds in human hair with coherent anti-Stokes Raman scattering microscopy

    NASA Astrophysics Data System (ADS)

    Zimmerley, Maxwell; Lin, Chia-Yu; Oertel, David C.; Marsh, Jennifer M.; Ward, Jimmie L.; Potma, Eric Olaf

    2009-07-01

    Coherent anti-Stokes Raman scattering (CARS) microscopy is used to determine the distribution and concentration of selected compounds in intact human hair. By generating images based on ratiometric CARS contrast, quantitative concentration maps of both water and externally applied d-glycine are produced in the cortex of human hair fibers. Both water and d-glycine are found to homogeneously distribute throughout the cortical regions of the hair. The ability to selectively detect molecular agents in hair fibers is of direct relevance to understanding the chemical and physical mechanisms that underlie the performance of hair-care products.

  5. A quantitative TaqMan PCR assay for the detection of Ureaplasma diversum.

    PubMed

    Marques, Lucas M; Amorim, Aline T; Martins, Hellen Braga; Rezende, Izadora Souza; Barbosa, Maysa Santos; Lobão, Tassia Neves; Campos, Guilherme B; Timenetsky, Jorge

    2013-12-27

    Ureaplasma diversum in veterinary studies is an undesirable microbe, which may cause infection in bulls and may result in seminal vesiculitis, balanopostitis, and alterations in spermatozoids, whereas in cows, it may cause placentitis, fetal alveolitis, abortion, and birth of weak calves. U. diversum is released through organic secretions, especially semen, preputial and vaginal mucus, conjunctival secretion, and milk. The aim of the present study was to develop a TaqMan probe, highly sensitive and specific quantitative PCR (qPCR) assay for the detection and quantification of U. diversum from genital swabs of bovines. Primers and probes specific to U. diversum 16S rRNA gene were designed. The specificity, detection limit, intra- and inter-assay variability of qPCR to detect this ureaplasma was compared with the results of the conventional PCR assay (cPCR). Swabs of vaginal mucus from 169 cows were tested. The qPCR assay detected as few as 10 copies of U. diversum and was 100-fold more sensitive than the cPCR. No cross-reactivity with other Mollicutes or eubacteria was observed. U. diversum was detected in 79 swabs (46.42%) by qPCR, while using cPCR it was detected in 42 (25%) samples. The difference in cPCR and qPCR ureaplasma detection between healthy and sick animals was not statistically significant. But the U. diversum load in samples from animals with genital disorders was higher than in healthy animals. The qPCR assay developed herein is highly sensitive and specific for the detection and quantification of U. diversum in vaginal bovine samples.

  6. Integrated genomics and molecular breeding approaches for dissecting the complex quantitative traits in crop plants.

    PubMed

    Kujur, Alice; Saxena, Maneesha S; Bajaj, Deepak; Laxmi; Parida, Swarup K

    2013-12-01

    The enormous population growth, climate change and global warming are now considered major threats to agriculture and world's food security. To improve the productivity and sustainability of agriculture, the development of highyielding and durable abiotic and biotic stress-tolerant cultivars and/climate resilient crops is essential. Henceforth, understanding the molecular mechanism and dissection of complex quantitative yield and stress tolerance traits is the prime objective in current agricultural biotechnology research. In recent years, tremendous progress has been made in plant genomics and molecular breeding research pertaining to conventional and next-generation whole genome, transcriptome and epigenome sequencing efforts, generation of huge genomic, transcriptomic and epigenomic resources and development of modern genomics-assisted breeding approaches in diverse crop genotypes with contrasting yield and abiotic stress tolerance traits. Unfortunately, the detailed molecular mechanism and gene regulatory networks controlling such complex quantitative traits is not yet well understood in crop plants. Therefore, we propose an integrated strategies involving available enormous and diverse traditional and modern -omics (structural, functional, comparative and epigenomics) approaches/resources and genomics-assisted breeding methods which agricultural biotechnologist can adopt/utilize to dissect and decode the molecular and gene regulatory networks involved in the complex quantitative yield and stress tolerance traits in crop plants. This would provide clues and much needed inputs for rapid selection of novel functionally relevant molecular tags regulating such complex traits to expedite traditional and modern marker-assisted genetic enhancement studies in target crop species for developing high-yielding stress-tolerant varieties.

  7. Gel-Based and Gel-Free Quantitative Proteomics Approaches at a Glance

    PubMed Central

    Abdallah, Cosette; Dumas-Gaudot, Eliane; Renaut, Jenny; Sergeant, Kjell

    2012-01-01

    Two-dimensional gel electrophoresis (2-DE) is widely applied and remains the method of choice in proteomics; however, pervasive 2-DE-related concerns undermine its prospects as a dominant separation technique in proteome research. Consequently, the state-of-the-art shotgun techniques are slowly taking over and utilising the rapid expansion and advancement of mass spectrometry (MS) to provide a new toolbox of gel-free quantitative techniques. When coupled to MS, the shotgun proteomic pipeline can fuel new routes in sensitive and high-throughput profiling of proteins, leading to a high accuracy in quantification. Although label-based approaches, either chemical or metabolic, gained popularity in quantitative proteomics because of the multiplexing capacity, these approaches are not without drawbacks. The burgeoning label-free methods are tag independent and suitable for all kinds of samples. The challenges in quantitative proteomics are more prominent in plants due to difficulties in protein extraction, some protein abundance in green tissue, and the absence of well-annotated and completed genome sequences. The goal of this perspective assay is to present the balance between the strengths and weaknesses of the available gel-based and -free methods and their application to plants. The latest trends in peptide fractionation amenable to MS analysis are as well discussed. PMID:23213324

  8. Quantitative PCR detection of Batrachochytrium dendrobatidis DNA from sediments and water

    USGS Publications Warehouse

    Kirshtein, J.D.; Anderson, C.W.; Wood, J.S.; Longcore, J.E.; Voytek, M.A.

    2007-01-01

    The fungal pathogen Batrachochytrium dendrobatidis (Bd) causes chytridiomycosis, a disease implicated in amphibian declines on 5 continents. Polymerase chain reaction (PCR) primer sets exist with which amphibians can be tested for this disease, and advances in sampling techniques allow non-invasive testing of animals. We developed filtering and PCR based quantitative methods by modifying existing PCR assays to detect Bd DNA in water and sediments, without the need for testing amphibians; we tested the methods at 4 field sites. The SYBR based assay using Boyle primers (SYBR/Boyle assay) and the Taqman based assay using Wood primers performed similarly with samples generated in the laboratory (Bd spiked filters), but the SYBR/Boyle assay detected Bd DNA in more field samples. We detected Bd DNA in water from 3 of 4 sites tested, including one pond historically negative for chytridiomycosis. Zoospore equivalents in sampled water ranged from 19 to 454 l-1 (nominal detection limit is 10 DNA copies, or about 0.06 zoospore). We did not detect DNA of Bd from sediments collected at any sites. Our filtering and amplification methods provide a new tool to investigate critical aspects of Bd in the environment. ?? Inter-Research 2007.

  9. Real-time quantitative PCR detection of Mycobacterium avium subspecies in meat products.

    PubMed

    Klanicova, B; Slana, I; Vondruskova, H; Kaevska, M; Pavlik, I

    2011-04-01

    The aim of this work was to examine various purchased meat products and to find out if any traces of Mycobacterium avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis could be detected in these samples. Analysis of the meat products (raw, cooked, and fermented) was performed using a real-time quantitative PCR (qPCR) method for the detection of specific insertion sequences: duplex qPCR for the detection of IS900 specific for M. avium subsp. paratuberculosis, and triplex qPCR for the detection of IS901 specific for Mycobacterium avium subsp. avium and IS 1245 specific for M. avium subsp. hominissuis. Of the 77 analyzed meat samples, 17 (22%) were found to contain M. avium subsp. paratuberculosis DNA, 4 (5%) samples contained Mycobacterium avium subsp. avium DNA, and in 12 (16%) samples M. avium subsp. hominissuis DNA was detected. The concentration of M. avium subsp. paratuberculosis and M. avium subsp. hominissuis DNA in some meat products exceeded 10(4) genomes per g. Culture examination of these mycobacterial subspecies was negative. By analyzing a range of meat products, we have provided evidence to support the hypothesis that M. avium is present in everyday commodities sold to the general public.

  10. Quantitative and simultaneous detection of four foodborne bacterial pathogens with a multi-channel SPR sensor.

    PubMed

    Taylor, Allen D; Ladd, Jon; Yu, Qiuming; Chen, Shengfu; Homola, Jirí; Jiang, Shaoyi

    2006-12-15

    We report the quantitative and simultaneous detection of four species of bacteria, Escherichia coli O157:H7, Salmonella choleraesuis serotype typhimurium, Listeria monocytogenes, and Campylobacter jejuni, using an eight-channel surface plasmon resonance (SPR) sensor based on wavelength division multiplexing. Detection curves showing SPR response versus analyte concentration were established for each species of bacteria in buffer at pH 7.4, apple juice at native pH 3.7, and apple juice at an adjusted pH of 7.4, as well as for a mixture containing all four species of bacteria in buffer. Control experiments were performed to show the non-fouling characteristics of the sensor surface as well as the specificity of the amplification antibodies used in this study. The limit of detection (LOD) for each of the four species of bacteria in the tested matrices ranges from 3.4 x 10(3) to 1.2 x 10(5) cfu/ml. Detection curves in buffer of an individual species of bacteria in a mixture of all four species of bacteria correlated well with detection curves of the individual species of bacteria alone. SPR responses were higher for bacteria in apple juice at pH 7.4 than in apple juice at pH 3.7. This difference in sensor response could be partly attributed to the pH dependence of antibody-antigen binding.

  11. Detection of earth-approaching asteroids in near real time

    NASA Technical Reports Server (NTRS)

    Rabinowitz, D. L.

    1991-01-01

    Computer software, called the Moving Object Detection Program (MODP), is described which detects earth-approaching asteroids in near real time. The software runs on a workstation linked to the output of the drift-scanning CCD camera of the Spacewatch Telescope. MOPD recognizes trailed images, detects motion, and accurately determines angular positions and rates of motion for moving objects in the scan images. The results are obtained a few seconds after the image signals are shifted out of the CCD. During 2 months of trial observations with this system, 304 asteroids were detected down to a limiting apparent magnitude for untrailed images of V = 20.5.

  12. Quantitative detection of astaxanthin and cantaxanthin in Atlantic salmon by resonance Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Ermakov, Igor V.; Ermakova, Maia R.; Gellermann, Werner

    2006-02-01

    Two major carotenoids species found in salmonids muscle tissues are astaxanthin and cantaxanthin. They are taken up from fish food and are responsible for the attractive red-orange color of salmon filet. Since carotenoids are powerful antioxidants and biomarkers of nutrient consumption, they are thought to indicate fish health and resistance to diseases in fish farm environments. Therefore, a rapid, accurate, quantitative optical technique for measuring carotenoid content in salmon tissues is of economic interest. We demonstrate the possibility of using fast, selective, quantitative detection of astaxanthin and cantaxanthin in salmon muscle tissues, employing resonance Raman spectroscopy. Analyzing strong Raman signals originating from the carbon-carbon double bond stretch vibrations of the carotenoid molecules under blue laser excitation, we are able to characterize quantitatively the concentrations of carotenoids in salmon muscle tissue. To validate the technique, we compared Raman data with absorption measurements of carotenoid extracts in acetone. A close correspondence was observed in absorption spectra for tissue extract in acetone and a pure astaxanthin solution. Raman results show a linear dependence between Raman and absorption data. The proposed technique holds promise as a method of rapid screening of carotenoid levels in fish muscle tissues and may be attractive for the fish farm industry to assess the dietary status of salmon, risk for infective diseases, and product quality control.

  13. Detecting Polychlorinated Biphenyls by Ah Receptor and Fluorescence Quantitative PCR with Exonuclease

    NASA Astrophysics Data System (ADS)

    Zhao, Xiaoxiang; Zhuang, Huisheng

    2010-11-01

    Tetrachlorobiphenyls as ligands were cultivated with goldfish, Ah receptors were extracted from the liver of goldfish and purified by hydroxyapatite. The complex of TCB ligands-receptors were analyzed by Surface Plasmon Resonance. DNA probes were amplified by PCR using Primers F1 and F2 with the DNA recognition site of responsive enhancer. DNA probes bound to the complex were not digested by exonuclease. The DNA that bound to the complex was quantified by real time PCR. A standard curve with TCB concentration to Ct values was obtained in the range of 10-12mol/L to 10-8 mol/L, according to TCB concentration in samples. The detection limit of the assay was below 10-12mol/L of TCB. Compared with HPLC, this assay is much more sensitive. These results suggest that fluorescence quantitative PCR with exonuclease by Ah receptors fits for detection of trace PCB.

  14. Detection and quantitation of resveratrol in tomato fruit (Lycopersicon esculentum Mill.).

    PubMed

    Ragab, Amr S; Van Fleet, Jennifer; Jankowski, Boris; Park, Joon-Hyun; Bobzin, Steven C

    2006-09-20

    Resveratrol is a stilbene phytoalexin well-known for its presence in grape, wine, and peanut. As a result of its antioxidant and chemopreventative properties, it has gained much attention as a functional food ingredient. A gas chromatography-mass spectrometry method for the detection of resveratrol, its 3-glucopyranoside piceid, and the cis isomers of both compounds has been developed and used to quantitate the levels of these compounds in the skin of commercially available tomato fruit (Lycopersicon esculentum Mill.). The resveratrol concentration remains relatively stable during fruit maturation, reaching a maximum concentration in the skin of 18.4 +/- 1.6 microg/g dry weight at 4 weeks postbreaker. No stilbenes were detected in the flesh of tomato fruit. PMID:16968079

  15. Establishment and assessment of two methods for quantitative detection of serum duck hepatitis B virus DNA

    PubMed Central

    Chen, Ya-Xi; Huang, Ai-Long; Qi, Zhen-Yuan; Guo, Shu-Hua

    2004-01-01

    AIM: To establish and assess the methods for quantitative detection of serum duck hepatitis B virus (DHBV) DNA by quantitative membrane hybridization using DHBV DNA probe labeled directly with alkaline phosphatase and fluorescence quantitative PCR (qPCR). METHODS: Probes of DHBV DNA labeled directly with alkaline phosphatase and chemiluminescent substrate CDP-star were used in this assay. DHBV DNA was detected by autoradiography, and then scanned by DNA dot-blot. In addition, three primers derived from DHBV DNA S gene were designed. Semi-nested primer was labeled by AmpliSensor. Standard curve of the positive standards of DHBV DNA was established after asymmetric preamplification, semi-nested amplification and on-line detection. Results from 100 samples detected separately by alkaline phosphatase direct-labeled DHBV DNA probe with dot-blot hybridization and digoxigenin-labeled DHBV DNA probe hybridization. Seventy samples of duck serum were tested by fluorescent qPCR and digoxigenin-labeled DHBV DNA probe in dot-blot hybridization assay and the correlation of results was analysed. RESULTS: Sensitivity of alkaline phosphatase direct-labeled DHBV DNA probe was 10 pg. The coincidence was 100% compared with digoxigenin-labeled DHBV DNA probe assay. After 30 cycles, amplification products showed two bands of about 180 bp and 70 bp by 20 g/L agarose gel electrophoresis. Concentration of amplification products was in direct proportion to the initial concentration of positive standards. The detection index was in direct proportion to the quantity of amplification products accumulated in the current cycle. The initial concentration of positive standards was in inverse proportion to the number of cycles needed for enough quantities of amplification products. Correlation coefficient of the results was (0.97, P < 0.01) between fluorescent qPCR and dot-blot hybridization. CONCLUSION: Alkaline phosphatase direct-labeled DHBV DNA probe in dot-blot hybridization and fluorescent q

  16. New approaches for the analysis of confluent cell layers with quantitative phase digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Pohl, L.; Kaiser, M.; Ketelhut, S.; Pereira, S.; Goycoolea, F.; Kemper, Björn

    2016-03-01

    Digital holographic microscopy (DHM) enables high resolution non-destructive inspection of technical surfaces and minimally-invasive label-free live cell imaging. However, the analysis of confluent cell layers represents a challenge as quantitative DHM phase images in this case do not provide sufficient information for image segmentation, determination of the cellular dry mass or calculation of the cell thickness. We present novel strategies for the analysis of confluent cell layers with quantitative DHM phase contrast utilizing a histogram based-evaluation procedure. The applicability of our approach is illustrated by quantification of drug induced cell morphology changes and it is shown that the method is capable to quantify reliable global morphology changes of confluent cell layers.

  17. The Appeal to Expert Opinion: Quantitative Support for a Bayesian Network Approach.

    PubMed

    Harris, Adam J L; Hahn, Ulrike; Madsen, Jens K; Hsu, Anne S

    2016-08-01

    The appeal to expert opinion is an argument form that uses the verdict of an expert to support a position or hypothesis. A previous scheme-based treatment of the argument form is formalized within a Bayesian network that is able to capture the critical aspects of the argument form, including the central considerations of the expert's expertise and trustworthiness. We propose this as an appropriate normative framework for the argument form, enabling the development and testing of quantitative predictions as to how people evaluate this argument, suggesting that such an approach might be beneficial to argumentation research generally. We subsequently present two experiments as an example of the potential for future research in this vein, demonstrating that participants' quantitative ratings of the convincingness of a proposition that has been supported with an appeal to expert opinion were broadly consistent with the predictions of the Bayesian model.

  18. Genetic algorithm based image binarization approach and its quantitative evaluation via pooling

    NASA Astrophysics Data System (ADS)

    Hu, Huijun; Liu, Ya; Liu, Maofu

    2015-12-01

    The binarized image is very critical to image visual feature extraction, especially shape feature, and the image binarization approaches have been attracted more attentions in the past decades. In this paper, the genetic algorithm is applied to optimizing the binarization threshold of the strip steel defect image. In order to evaluate our genetic algorithm based image binarization approach in terms of quantity, we propose the novel pooling based evaluation metric, motivated by information retrieval community, to avoid the lack of ground-truth binary image. Experimental results show that our genetic algorithm based binarization approach is effective and efficiency in the strip steel defect images and our quantitative evaluation metric on image binarization via pooling is also feasible and practical.

  19. Specific PCR and real-time PCR assays for detection and quantitation of 'Candidatus Phytoplasma phoenicium'.

    PubMed

    Jawhari, Maan; Abrahamian, Peter; Sater, Ali Abdel; Sobh, Hana; Tawidian, Patil; Abou-Jawdah, Yusuf

    2015-02-01

    Almond witches' broom (AlmWB) is a fast-spreading lethal disease of almond, peach and nectarine associated with 'Candidatus Phytoplasma phoenicium'. The development of PCR and quantitative real-time PCR (qPCR) assays for the sensitive and specific detection of the phytoplasma is of prime importance for early detection of 'Ca. P. phoenicium' and for epidemiological studies. The developed qPCR assay herein uses a TaqMan(®) probe labeled with Black Hole Quencher Plus. The specificity of the PCR and that of the qPCR detection protocols were tested on 17 phytoplasma isolates belonging to 11 phytoplasma 16S rRNA groups, on samples of almond, peach, nectarine, native plants and insects infected or uninfected with the phytoplasma. The developed assays showed high specificity against 'Ca. P. phoenicium' and no cross-reactivity against any other phytoplasma, plant or insect tested. The sensitivity of the developed PCR and qPCR assays was similar to the conventional nested PCR protocol using universal primers. The qPCR assay was further validated by quantitating AlmWB phytoplasma in different hosts, plant parts and potential insect vectors. The highest titers of 'Ca. P. phoenicium' were detected in the phloem tissues of stems and roots of almond and nectarine trees, where they averaged from 10(5) to 10(6) genomic units per nanogram of host DNA (GU/ng of DNA). The newly developed PCR and qPCR protocols are reliable, specific and sensitive methods that are easily applicable to high-throughput diagnosis of AlmWB in plants and insects and can be used for surveys of potential vectors and alternative hosts.

  20. Passive mass transport for direct and quantitative SERS detection using purified silica encapsulated metal nanoparticles

    NASA Astrophysics Data System (ADS)

    Shrestha, Binaya Kumar

    This thesis focuses on understanding implications of nanomaterial quality control and mass transport through internally etched silica coated nanoparticles for direct and quantitative molecular detection using surface enhanced Raman scattering (SERS). Prior to use, bare nanoparticles (partially or uncoated with silica) are removal using column chromatography to improve the quality of these nanomaterials and their SERS reproducibility. Separation of silica coated nanoparticles with two different diameters is achieved using Surfactant-free size exclusion chromatography with modest fractionation. Next, selective molecular transport is modeled and monitored using SERS and evaluated as a function of solution ionic strength, pH, and polarity. Molecular detection is achieved when the analytes first partition through the silica membrane then interact with the metal surface at short distances (i.e., less than 2 nm). The SERS intensities of unique molecular vibrational modes for a given molecule increases as the number of molecules that bind to the metal surface increases and are enhanced via both chemical and electromagnetic enhancement mechanisms as long as the vibrational mode has a component of polarizability tensor along the surface normal. SERS signals increase linearly with molecular concentration until the three-dimensional SERS-active volume is saturated with molecules. Implications of molecular orientation as well as surface selection rules on SERS intensities of molecular vibrational modes are studied to improve quantitative and reproducible SERS detection using internally etched Ag Au SiO2 nanoparticles. Using the unique vibrational modes, SERS intensities for p-aminothiophenol as a function of metal core compositions and plasmonics are studied. By understanding molecular transport mechanisms through internally etched silica matrices coated on metal nanoparticles, important experimental and materials design parameters are learned, which can be subsequently applied

  1. A Room Temperature Ultrasensitive Magnetoelectric Susceptometer for Quantitative Tissue Iron Detection

    PubMed Central

    Xi, Hao; Qian, Xiaoshi; Lu, Meng-Chien; Mei, Lei; Rupprecht, Sebastian; Yang, Qing X.; Zhang, Q. M.

    2016-01-01

    Iron is a trace mineral that plays a vital role in the human body. However, absorbing and accumulating excessive iron in body organs (iron overload) can damage or even destroy an organ. Even after many decades of research, progress on the development of noninvasive and low-cost tissue iron detection methods is very limited. Here we report a recent advance in a room-temperature ultrasensitive biomagnetic susceptometer for quantitative tissue iron detection. The biomagnetic susceptometer exploits recent advances in the magnetoelectric (ME) composite sensors that exhibit an ultrahigh AC magnetic sensitivity under the presence of a strong DC magnetic field. The first order gradiometer based on piezoelectric and magnetostrictive laminate (ME composite) structure shows an equivalent magnetic noise of 0.99 nT/rt Hz at 1 Hz in the presence of a DC magnetic field of 0.1 Tesla and a great common mode noise rejection ability. A prototype magnetoelectric liver susceptometry has been demonstrated with liver phantoms. The results indicate its output signals to be linearly responsive to iron concentrations from normal iron dose (0.05 mg Fe/g liver phantom) to 5 mg Fe/g liver phantom iron overload (100X overdose). The results here open up many innovative possibilities for compact-size, portable, cost-affordable, and room-temperature operated medical systems for quantitative determinations of tissue iron. PMID:27465206

  2. Immunoelectrophoresis employing avian antisera for the detection and quantitation of Pasteurella multocida antigens.

    PubMed

    McKinney, K L; Rimler, R B

    1981-01-01

    Immunoelectrophoresis with various buffer systems at high and low pH was examined for suitability to detect and quantitate Pasteurella multocida antigens with turkey or chicken anti-P. multocida sera. Counterimmunoelectrophoresis was used to develop a buffer system for one-dimensional, two-dimensional, and rocket immunoelectrophoresis. The effects of pH, buffer, and molarity on resolution of immunoprecipitates were determined; 0.05 M sodium acetate-acetic acid buffer at pH 5.6 was the most suitable buffer. This buffer could be used in counterimmunoelectrophoresis with turkey or chicken sera to detect minute amounts of P. multocida protein antigens (4.3 ng/test) or lipopolysaccharide (3.12 micrograms/test). One-dimensional immunoelectrophoresis with the acetate buffer system required treatment of the gels with a 17% NaCl solution to induce immunoprecipitation of P. multocida lipopolysaccharide. Other techniques using the acetate buffer system did not require the high salt treatment. In two-dimensional immunoelectrophoresis, antisera migrated in the second dimension at pH 8.6, but did not migrate at pH 5.6. Rocket immunoelectrophoresis with the acetate buffer system was effective for quantitating P. multocida antigens.

  3. In-vivo Tumor detection using diffusion reflection measurements of targeted gold nanorods - a quantitative study.

    PubMed

    Ankri, Rinat; Duadi, Hamootal; Motiei, Menachem; Fixler, Dror

    2012-03-01

    The ability to quantitatively and non-invasively detect nanoparticles has important implications on their development as an in-vivo cancer diagnostic tool. The Diffusion Reflection (DR) method is a simple, non-invasive imaging technique which has been proven useful for the investigation of tissue's optical parameters. In this study, Monte Carlo (MC) simulations, tissue-like phantom experiments and in-vivo measurements of the reflected light intensity from tumor bearing mice are presented. Following intravenous injection of antibody conjugated poly (ethylene glycol)-coated (PEGylated) gold nanorods (GNR) to tumor-bearing mice, accumulation of GNR in the tumor was clearly detected by the DR profile of the tumor. The ability of DR measurements to quantitate in-vivo the concentration of the GNR in the tumor was demonstrated and validated with Flame Atomic Absorption spectroscopy results. With GNR as absorbing contrast agents, DR has important potential applications in the image guided therapy of superficial tumors such as head and neck cancer, breast cancer and melanoma. PMID:22234916

  4. A Room Temperature Ultrasensitive Magnetoelectric Susceptometer for Quantitative Tissue Iron Detection.

    PubMed

    Xi, Hao; Qian, Xiaoshi; Lu, Meng-Chien; Mei, Lei; Rupprecht, Sebastian; Yang, Qing X; Zhang, Q M

    2016-01-01

    Iron is a trace mineral that plays a vital role in the human body. However, absorbing and accumulating excessive iron in body organs (iron overload) can damage or even destroy an organ. Even after many decades of research, progress on the development of noninvasive and low-cost tissue iron detection methods is very limited. Here we report a recent advance in a room-temperature ultrasensitive biomagnetic susceptometer for quantitative tissue iron detection. The biomagnetic susceptometer exploits recent advances in the magnetoelectric (ME) composite sensors that exhibit an ultrahigh AC magnetic sensitivity under the presence of a strong DC magnetic field. The first order gradiometer based on piezoelectric and magnetostrictive laminate (ME composite) structure shows an equivalent magnetic noise of 0.99 nT/rt Hz at 1 Hz in the presence of a DC magnetic field of 0.1 Tesla and a great common mode noise rejection ability. A prototype magnetoelectric liver susceptometry has been demonstrated with liver phantoms. The results indicate its output signals to be linearly responsive to iron concentrations from normal iron dose (0.05 mg Fe/g liver phantom) to 5 mg Fe/g liver phantom iron overload (100X overdose). The results here open up many innovative possibilities for compact-size, portable, cost-affordable, and room-temperature operated medical systems for quantitative determinations of tissue iron. PMID:27465206

  5. Quantitative single-molecule detection of protein based on DNA tetrahedron fluorescent nanolabels.

    PubMed

    Ding, Yongshun; Liu, Xingti; Zhu, Jing; Wang, Lei; Jiang, Wei

    2014-07-01

    A highly sensitive method for single-molecule quantitative detection of human IgG is presented by the employment of a new fluorescent nanolabel. In this method, fluorescent nanolabels were assembled by inserting SYBR Green I into DNA tetrahedron nanostructure. The bio-nanolabels were attached to the streptavidin-antihuman antibody by a specific reaction between biotin and streptavidin. The antibody was combined with the target antigen, human IgG, which was immobilized on the silanized glass subtrate surface. Finally, epi-fluorescence microscopy (EFM) coupled with an electron multiplying charge-coupled device was employed for fluorescence imaging. The fluorescent spots corresponding to single protein molecule on images were counted and further used for the quantitative detection. It was found that the new nanolabel shows good photostability, biocompatiblity and exhibits no blinking compared to traditional labels like fluorescence dyes and quantum dot (QDs). In addition, the number of fluorescence spots on the images has a linear relationship with the concentration of human IgG in the range of 3.0×10(-14) to 1.0×10(-12)mol L(-1). What is more, this method showed an excellent specificity and a low matrix effect.

  6. Detection and Quantitation of Heavy Metal Ions on Bona Fide DVDs Using DNA Molecular Beacon Probes.

    PubMed

    Zhang, Lingling; Wong, Jessica X H; Li, Xiaochun; Li, Yunchao; Yu, Hua-Zhong

    2015-01-01

    A sensitive and cost-effective method for the simultaneous quantitation of trace amounts of Hg(2+) and Pb(2+) in real-world samples has been developed using DNA molecular beacon probes bound to bona fide digital video discs (DVDs). With specially designed T-rich or G-rich loop sequences, the detection is based on the strong T-Hg(2+)-T coordination chemistry of Hg(2+) and the formation of G-quadruplexes induced by Pb(2+), respectively. In particular, the presence of metal cations leads to hairpin opening and exposure of the terminal biotin moiety for binding nanogold-streptavidin conjugates. The recognition signal was subsequently enhanced by gold nanoparticle-promoted silver deposition, which leads to quantifiable digital signals upon reading with a standard computer drive. This method exhibits a wide response range and low detection limits for both Hg(2+) and Pb(2+). In addition, the quantitative determination of heavy metals in food products (e.g., rice samples) has been demonstrated and the method compares favorably with other optical sensors developed recently.

  7. A Room Temperature Ultrasensitive Magnetoelectric Susceptometer for Quantitative Tissue Iron Detection

    NASA Astrophysics Data System (ADS)

    Xi, Hao; Qian, Xiaoshi; Lu, Meng-Chien; Mei, Lei; Rupprecht, Sebastian; Yang, Qing X.; Zhang, Q. M.

    2016-07-01

    Iron is a trace mineral that plays a vital role in the human body. However, absorbing and accumulating excessive iron in body organs (iron overload) can damage or even destroy an organ. Even after many decades of research, progress on the development of noninvasive and low-cost tissue iron detection methods is very limited. Here we report a recent advance in a room-temperature ultrasensitive biomagnetic susceptometer for quantitative tissue iron detection. The biomagnetic susceptometer exploits recent advances in the magnetoelectric (ME) composite sensors that exhibit an ultrahigh AC magnetic sensitivity under the presence of a strong DC magnetic field. The first order gradiometer based on piezoelectric and magnetostrictive laminate (ME composite) structure shows an equivalent magnetic noise of 0.99 nT/rt Hz at 1 Hz in the presence of a DC magnetic field of 0.1 Tesla and a great common mode noise rejection ability. A prototype magnetoelectric liver susceptometry has been demonstrated with liver phantoms. The results indicate its output signals to be linearly responsive to iron concentrations from normal iron dose (0.05 mg Fe/g liver phantom) to 5 mg Fe/g liver phantom iron overload (100X overdose). The results here open up many innovative possibilities for compact-size, portable, cost-affordable, and room-temperature operated medical systems for quantitative determinations of tissue iron.

  8. A preamplification approach to GMO detection in processed foods.

    PubMed

    Del Gaudio, S; Cirillo, A; Di Bernardo, G; Galderisi, U; Cipollaro, M

    2010-03-01

    DNA is widely used as a target for GMO analysis because of its stability and high detectability. Real-time PCR is the method routinely used in most analytical laboratories due to its quantitative performance and great sensitivity. Accurate DNA detection and quantification is dependent on the specificity and sensitivity of the amplification protocol as well as on the quality and quantity of the DNA used in the PCR reaction. In order to enhance the sensitivity of real-time PCR and consequently expand the number of analyzable target genes, we applied a preamplification technique to processed foods where DNA can be present in low amounts and/or in degraded forms thereby affecting the reliability of qualitative and quantitative results. The preamplification procedure utilizes a pool of primers targeting genes of interest and is followed by real-time PCR reactions specific for each gene. An improvement of Ct values was found comparing preamplified vs. non-preamplified DNA. The strategy reported in the present study will be also applicable to other fields requiring quantitative DNA testing by real-time PCR.

  9. A preamplification approach to GMO detection in processed foods.

    PubMed

    Del Gaudio, S; Cirillo, A; Di Bernardo, G; Galderisi, U; Cipollaro, M

    2010-03-01

    DNA is widely used as a target for GMO analysis because of its stability and high detectability. Real-time PCR is the method routinely used in most analytical laboratories due to its quantitative performance and great sensitivity. Accurate DNA detection and quantification is dependent on the specificity and sensitivity of the amplification protocol as well as on the quality and quantity of the DNA used in the PCR reaction. In order to enhance the sensitivity of real-time PCR and consequently expand the number of analyzable target genes, we applied a preamplification technique to processed foods where DNA can be present in low amounts and/or in degraded forms thereby affecting the reliability of qualitative and quantitative results. The preamplification procedure utilizes a pool of primers targeting genes of interest and is followed by real-time PCR reactions specific for each gene. An improvement of Ct values was found comparing preamplified vs. non-preamplified DNA. The strategy reported in the present study will be also applicable to other fields requiring quantitative DNA testing by real-time PCR. PMID:19823811

  10. Silicon photonic microring resonators for quantitative cytokine detection and T-cell secretion analysis.

    PubMed

    Luchansky, Matthew S; Bailey, Ryan C

    2010-03-01

    The ability to perform multiple simultaneous protein biomarker measurements in complex media with picomolar sensitivity presents a large challenge to disease diagnostics and fundamental biological studies. Silicon photonic microring resonators represent a promising platform for real-time detection of biomolecules on account of their spectral sensitivity toward surface binding events between a target and antibody-modified microrings. For all refractive index-based sensing schemes, the mass of bound analytes, in combination with other factors such as antibody affinity and surface density, contributes to the observed signal and measurement sensitivity. Therefore, proteins that are simultaneously low in abundance and have a lower molecular weight are often challenging to detect. By employing a more massive secondary antibody to amplify the signal arising from the initial binding event, it is possible to improve both the sensitivity and the specificity of protein assays, allowing for quantitative sensing in complex sample matrices. Herein, a sandwich assay is used to detect the 15.5 kDa human cytokine interleukin-2 (IL-2) at concentrations down to 100 pg/mL (6.5 pM) and to quantitate unknown solution concentrations over a dynamic range spanning 2.5 orders of magnitude. This same sandwich assay is then used to monitor the temporal secretion profile of IL-2 from Jurkat T lymphocytes in serum-containing cell culture media in the presence of the entire Jurkat secretome. The same temporal secretion analysis is performed in parallel using a commercial ELISA, revealing similar IL-2 concentration profiles but superior precision for the microring resonator sensing platform. Furthermore, we demonstrate the generality of the sandwich assay methodology on the microring resonator platform for the analysis of any biomolecular target for which two high-affinity antibodies exist by detecting the approximately 8 kDa cytokine interleukin-8 (IL-8) with a limit of detection and dynamic

  11. Two approaches to improving mental health care: positivist/quantitative versus skill-based/qualitative.

    PubMed

    Luchins, Daniel

    2012-01-01

    The quality improvement model currently used in medicine and mental health was adopted from industry, where it developed out of early 20th-century efforts to apply a positivist/quantitative agenda to improving manufacturing. This article questions the application of this model to mental health care. It argues that (1) developing "operational definitions" for something as value-laden as "quality" risks conflating two realms, what we measure with what we value; (2) when measurements that are tied to individuals are aggregated to establish benchmarks and goals, unwarranted mathematical assumptions are made; (3) choosing clinical outcomes is problematic; (4) there is little relationship between process measures and clinical outcomes; and (5) since changes in quality indices do not relate to improved clinical care, management's reliance on such indices provides an illusory sense of control. An alternative model is the older, skill-based/qualitative approach to knowing, which relies on "implicit/ expert" knowledge. These two approaches offer a series of contrasts: quality versus excellence, competence versus expertise, management versus leadership, extrinsic versus intrinsic rewards. The article concludes that we need not totally dispense with the current quality improvement model, but rather should balance quantitative efforts with the older qualitative approach in a mixed methods model. PMID:23179033

  12. A Systematic Approach for Quantitative Analysis of Multidisciplinary Design Optimization Framework

    NASA Astrophysics Data System (ADS)

    Kim, Sangho; Park, Jungkeun; Lee, Jeong-Oog; Lee, Jae-Woo

    An efficient Multidisciplinary Design and Optimization (MDO) framework for an aerospace engineering system should use and integrate distributed resources such as various analysis codes, optimization codes, Computer Aided Design (CAD) tools, Data Base Management Systems (DBMS), etc. in a heterogeneous environment, and need to provide user-friendly graphical user interfaces. In this paper, we propose a systematic approach for determining a reference MDO framework and for evaluating MDO frameworks. The proposed approach incorporates two well-known methods, Analytic Hierarchy Process (AHP) and Quality Function Deployment (QFD), in order to provide a quantitative analysis of the qualitative criteria of MDO frameworks. Identification and hierarchy of the framework requirements and the corresponding solutions for the reference MDO frameworks, the general one and the aircraft oriented one were carefully investigated. The reference frameworks were also quantitatively identified using AHP and QFD. An assessment of three in-house frameworks was then performed. The results produced clear and useful guidelines for improvement of the in-house MDO frameworks and showed the feasibility of the proposed approach for evaluating an MDO framework without a human interference.

  13. Two approaches to improving mental health care: positivist/quantitative versus skill-based/qualitative.

    PubMed

    Luchins, Daniel

    2012-01-01

    The quality improvement model currently used in medicine and mental health was adopted from industry, where it developed out of early 20th-century efforts to apply a positivist/quantitative agenda to improving manufacturing. This article questions the application of this model to mental health care. It argues that (1) developing "operational definitions" for something as value-laden as "quality" risks conflating two realms, what we measure with what we value; (2) when measurements that are tied to individuals are aggregated to establish benchmarks and goals, unwarranted mathematical assumptions are made; (3) choosing clinical outcomes is problematic; (4) there is little relationship between process measures and clinical outcomes; and (5) since changes in quality indices do not relate to improved clinical care, management's reliance on such indices provides an illusory sense of control. An alternative model is the older, skill-based/qualitative approach to knowing, which relies on "implicit/ expert" knowledge. These two approaches offer a series of contrasts: quality versus excellence, competence versus expertise, management versus leadership, extrinsic versus intrinsic rewards. The article concludes that we need not totally dispense with the current quality improvement model, but rather should balance quantitative efforts with the older qualitative approach in a mixed methods model.

  14. Real-time PCR assay for rapid qualitative and quantitative detection of Entamoeba histolytica.

    PubMed

    Orosz, Erika; Perkátai, Katalin; Kapusinszky, Beatrix; Farkas, Agnes; Kucsera, István

    2012-12-01

    Simple real-time PCR assay with one set of primer and probe for rapid, sensitive qualitative and quantitative detection of Entamoeba histolytica has been used. Consensus sequences were used to amplify a species-specific region of the 16S rRNA gene, and fluorescence resonance energy transfer hybridization probes were used for detection in a LightCycler platform (Roche). The anchor probe sequence was designed to be a perfect match for the 16S rRNA gene of Entamoeba species, while the acceptor probe sequence was designed for Entamoeba histolytica, which allowed differentiation. The performed characteristics of the real-time PCR assay were compared with ELISA antigen and microscopical detection from 77 samples of individuals with suspected clinical diagnosis of imported E. histolytica infection. Stool and liver abscess pus samples were examined with analytical sensitivity of 5 parasites per PCR reaction. The melting curve means Tms (standard deviation) in clinical isolates were 54°C. The real-time assay was 100% sensitive and specific for differentiation of Entamoeba histolytica, compared with conventional ELISA or microscopy. This real-time PCR assay with melting curve analysis is rapid, and specific for the detection and differentiation of Entamoeba histolytica. The suitability for routine use of this assay in clinical diagnostic laboratories is discussed.

  15. A hybrid approach for efficient anomaly detection using metaheuristic methods

    PubMed Central

    Ghanem, Tamer F.; Elkilani, Wail S.; Abdul-kader, Hatem M.

    2014-01-01

    Network intrusion detection based on anomaly detection techniques has a significant role in protecting networks and systems against harmful activities. Different metaheuristic techniques have been used for anomaly detector generation. Yet, reported literature has not studied the use of the multi-start metaheuristic method for detector generation. This paper proposes a hybrid approach for anomaly detection in large scale datasets using detectors generated based on multi-start metaheuristic method and genetic algorithms. The proposed approach has taken some inspiration of negative selection-based detector generation. The evaluation of this approach is performed using NSL-KDD dataset which is a modified version of the widely used KDD CUP 99 dataset. The results show its effectiveness in generating a suitable number of detectors with an accuracy of 96.1% compared to other competitors of machine learning algorithms. PMID:26199752

  16. A hybrid approach for efficient anomaly detection using metaheuristic methods.

    PubMed

    Ghanem, Tamer F; Elkilani, Wail S; Abdul-Kader, Hatem M

    2015-07-01

    Network intrusion detection based on anomaly detection techniques has a significant role in protecting networks and systems against harmful activities. Different metaheuristic techniques have been used for anomaly detector generation. Yet, reported literature has not studied the use of the multi-start metaheuristic method for detector generation. This paper proposes a hybrid approach for anomaly detection in large scale datasets using detectors generated based on multi-start metaheuristic method and genetic algorithms. The proposed approach has taken some inspiration of negative selection-based detector generation. The evaluation of this approach is performed using NSL-KDD dataset which is a modified version of the widely used KDD CUP 99 dataset. The results show its effectiveness in generating a suitable number of detectors with an accuracy of 96.1% compared to other competitors of machine learning algorithms. PMID:26199752

  17. An analytical approach to quantitative reconstruction of non-uniform attenuated brain SPECT.

    PubMed

    Liang, Z; Ye, J; Harrington, D P

    1994-11-01

    An analytical approach to quantitative brain SPECT (single-photon-emission computed tomography) with non-uniform attenuation is developed. The approach formulates accurately the projection-transform equation as a summation of primary- and scatter-photon contributions. The scatter contribution can be estimated using the multiple-energy-window samples and removed from the primary-energy-window data by subtraction. The approach models the primary contribution as a convolution of the attenuated source and the detector-response kernel at a constant depth from the detector with the central-ray approximation. The attenuated Radon transform of the source can be efficiently deconvolved using the depth-frequency relation. The approach inverts exactly the attenuated Radon transform by Fourier transforms and series expansions. The performance of the analytical approach was studied for both uniform- and non-uniform-attenuation cases, and compared to the conventional FBP (filtered-backprojection) method by computer simulations. A patient brain X-ray image was acquired by a CT (computed-tomography) scanner and converted to the object-specific attenuation map for 140 keV energy. The mathematical Hoffman brain phantom was used to simulate the emission source and was resized such that it was completely surrounded by the skull of the CT attenuation map. The detector-response kernel was obtained from measurements of a point source at several depths in air from a parallel-hole collimator of a SPECT camera. The projection data were simulated from the object-specific attenuating source including the depth-dependent detector response. Quantitative improvement (>5%) in reconstructing the data was demonstrated with the nonuniform attenuation compensation, as compared to the uniform attenuation correction and the conventional FBP reconstruction. The commuting time was less than 5 min on an HP/730 desktop computer for an image array of 1282*32 from 128 projections of 128*32 size. PMID

  18. Development of one-step quantitative reverse transcription PCR for the rapid detection of flaviviruses

    PubMed Central

    2013-01-01

    Background The genus Flavivirus includes several pathogenic agents that cause severe illness in humans. Re-emergence of West Nile virus in Europe and continuous spread of certain flaviviruses such as dengue, yellow fever and Japanese encephalitis viruses represent a global danger to public health. Therefore, a rapid and accurate molecular method is required for diagnostics and epidemiological surveillance of flaviviruses. Methods A Pan-Flavi quantitative RT-PCR assay using a Locked-Nucleic Acid probe targeting the flavivirus NS5 gene was developed and optimized to detect a wide range of flaviviruses simultaneously. The specificity and sensitivity of the Pan-Flavi assay were tested using RNA of different flaviviruses and non-flaviviruses. Furthermore, the assay was compared directly to flavivirus species-specific assays for the ability to detect flaviviruses sensitively. Results Two degenerate primers and one Locked-Nucleic Acids probe were designed to amplify most of the flaviviruses. To increase the specificity and fluorescence signal of the Pan-Flavi assay for detection of yellow fever virus and dengue virus 4, additional primers and probes were included. Viral RNA of thirty different flaviviruses was detected, verifying the broad range specificity. The testing of this assay was successful, using standard plasmid and RNA dilutions of yellow fever virus vaccine strain, dengue virus 1 and tick-borne encephalitis virus, with a sensitivity limit of 10–100 genome copies/reaction. Also comparatively good results were achieved for detecting different flaviviruses by the Pan-Flavi assay when compared to the flavivirus species-specific assays. Conclusion The assay is rapid, broad-range flavivirus-specific and highly sensitive making it a valuable tool for rapid detection of flaviviruses in livestock samples, epidemiological studies or as useful complement to single flavivirus-specific assays for clinical diagnosis. PMID:23410000

  19. A change detection approach to moving object detection in low frame-rate video

    SciTech Connect

    Porter, Reid B; Harvey, Neal R; Theiler, James P

    2009-01-01

    Moving object detection is of significant interest in temporal image analysis since it is a first step in many object identification and tracking applications. A key component in almost all moving object detection algorithms is a pixel-level classifier, where each pixel is predicted to be either part of a moving object or part of the background. In this paper we investigate a change detection approach to the pixel-level classification problem and evaluate its impact on moving object detection. The change detection approach that we investigate was previously applied to multi-and hyper-spectral datasets, where images were typically taken several days, or months apart. In this paper, we apply the approach to low-frame rate (1-2 frames per second) video datasets.

  20. Quantitative detection of TUSC3 promoter methylation -a potential biomarker for prognosis in lung cancer

    PubMed Central

    Duppel, Uta; Woenckhaus, Matthias; Schulz, Christian; Merk, Johannes; Dietmaier, Wolfgang

    2016-01-01

    Aberrant promoter methylation of tumor relevant genes frequently occurs in early steps of carcinogenesis and during tumor progression. Epigenetic alterations could be used as potential biomarkers for early detection and for prediction of prognosis and therapy response in lung cancer. The present study quantitatively analyzed the methylation status of known and potential gatekeeper and tumor suppressor genes [O-6-methylguanine-DNA methyltransferase (MGMT), Ras association domain family member 1A (RASSF1A), Ras protein activator like 1 (RASAL1), programmed cell death 4 (PDCD4), metastasis suppressor 1 (MTSS1) and tumor suppressor candidate 3 (TUSC3)] in 42 lung cancers and in corresponding non-malignant bronchus and lung tissue using bisulfite-conversion independent methylation-quantification of endonuclease-resistant DNA (MethyQESD). Methylation status was associated with clinical and pathological parameters. No methylation was found in the promoter regions of PDCD4 and MTSS1 of either compartment. MGMT, RASSF1A and RASAL1 showed sporadic (up to 26.2%) promoter methylation. The promoter of TUSC3, however, was frequently methylated in the tumor (59.5%), benign bronchus (67.9%) and alveolar lung (31.0%) tissues from each tumor patient. The methylation status of TUSC3 was significantly associated with smaller tumor size (P=0.008) and a longer overall survival (P=0.013). Pooled blood DNA of healthy individuals did not show any methylation of either gene. Therefore, methylation of TUSC3 shows prognostic and pathobiological relevance in lung cancer. Furthermore, quantitative detection of TUSC3 promoter methylation appears to be a promising tool for early detection and prediction of prognosis in lung cancer. However, additional studies are required to confirm this finding. PMID:27698890

  1. Quantitative Detection of Trace Malachite Green in Aquiculture Water Samples by Extractive Electrospray Ionization Mass Spectrometry.

    PubMed

    Fang, Xiaowei; Yang, Shuiping; Chingin, Konstantin; Zhu, Liang; Zhang, Xinglei; Zhou, Zhiquan; Zhao, Zhanfeng

    2016-01-01

    Exposure to malachite green (MG) may pose great health risks to humans; thus, it is of prime importance to develop fast and robust methods to quantitatively screen the presence of malachite green in water. Herein the application of extractive electrospray ionization mass spectrometry (EESI-MS) has been extended to the trace detection of MG within lake water and aquiculture water, due to the intensive use of MG as a biocide in fisheries. This method has the advantage of obviating offline liquid-liquid extraction or tedious matrix separation prior to the measurement of malachite green in native aqueous medium. The experimental results indicate that the extrapolated detection limit for MG was ~3.8 μg·L(-1) (S/N = 3) in lake water samples and ~0.5 μg·L(-1) in ultrapure water under optimized experimental conditions. The signal intensity of MG showed good linearity over the concentration range of 10-1000 μg·L(-1). Measurement of practical water samples fortified with MG at 0.01, 0.1 and 1.0 mg·L(-1) gave a good validation of the established calibration curve. The average recoveries and relative standard deviation (RSD) of malachite green in lake water and Carassius carassius fish farm effluent water were 115% (6.64% RSD), 85.4% (9.17% RSD) and 96.0% (7.44% RSD), respectively. Overall, the established EESI-MS/MS method has been demonstrated suitable for sensitive and rapid (<2 min per sample) quantitative detection of malachite green in various aqueous media, indicating its potential for online real-time monitoring of real life samples.

  2. Enhanced detection of surface-associated bacteria in indoor environments by quantitative PCR.

    PubMed

    Buttner, M P; Cruz-Perez, P; Stetzenbach, L D

    2001-06-01

    Methods for detecting microorganisms on surfaces are needed to locate biocontamination sources and to relate surface and airborne concentrations. Research was conducted in an experimental room to evaluate surface sampling methods and quantitative PCR (QPCR) for enhanced detection of a target biocontaminant present on flooring materials. QPCR and culture analyses were used to quantitate Bacillus subtilis (Bacillus globigii) endospores on vinyl tile, commercial carpet, and new and soiled residential carpet with samples obtained by four surface sampling methods: a swab kit, a sponge swipe, a cotton swab, and a bulk method. The initial data showed that greater overall sensitivity was obtained with the QPCR than with culture analysis; however, the QPCR results for bulk samples from residential carpet were negative. The swab kit and the sponge swipe methods were then tested with two levels of background biological contamination consisting of Penicillium chrysogenum spores. The B. subtilis values obtained by the QPCR method were greater than those obtained by culture analysis. The differences between the QPCR and culture data were significant for the samples obtained with the swab kit for all flooring materials except soiled residential carpet and with the sponge swipe for commercial carpet. The QPCR data showed that there were no significant differences between the swab kit and sponge swipe sampling methods for any of the flooring materials. Inhibition of QPCR due solely to biological contamination of flooring materials was not evident. However, some degree of inhibition was observed with the soiled residential carpet, which may have been caused by the presence of abiotic contaminants, alone or in combination with biological contaminants. The results of this research demonstrate the ability of QPCR to enhance detection and enumeration of biocontaminants on surface materials and provide information concerning the comparability of currently available surface sampling

  3. Quantitative detection of TUSC3 promoter methylation -a potential biomarker for prognosis in lung cancer

    PubMed Central

    Duppel, Uta; Woenckhaus, Matthias; Schulz, Christian; Merk, Johannes; Dietmaier, Wolfgang

    2016-01-01

    Aberrant promoter methylation of tumor relevant genes frequently occurs in early steps of carcinogenesis and during tumor progression. Epigenetic alterations could be used as potential biomarkers for early detection and for prediction of prognosis and therapy response in lung cancer. The present study quantitatively analyzed the methylation status of known and potential gatekeeper and tumor suppressor genes [O-6-methylguanine-DNA methyltransferase (MGMT), Ras association domain family member 1A (RASSF1A), Ras protein activator like 1 (RASAL1), programmed cell death 4 (PDCD4), metastasis suppressor 1 (MTSS1) and tumor suppressor candidate 3 (TUSC3)] in 42 lung cancers and in corresponding non-malignant bronchus and lung tissue using bisulfite-conversion independent methylation-quantification of endonuclease-resistant DNA (MethyQESD). Methylation status was associated with clinical and pathological parameters. No methylation was found in the promoter regions of PDCD4 and MTSS1 of either compartment. MGMT, RASSF1A and RASAL1 showed sporadic (up to 26.2%) promoter methylation. The promoter of TUSC3, however, was frequently methylated in the tumor (59.5%), benign bronchus (67.9%) and alveolar lung (31.0%) tissues from each tumor patient. The methylation status of TUSC3 was significantly associated with smaller tumor size (P=0.008) and a longer overall survival (P=0.013). Pooled blood DNA of healthy individuals did not show any methylation of either gene. Therefore, methylation of TUSC3 shows prognostic and pathobiological relevance in lung cancer. Furthermore, quantitative detection of TUSC3 promoter methylation appears to be a promising tool for early detection and prediction of prognosis in lung cancer. However, additional studies are required to confirm this finding.

  4. Quantitative Detection of Trace Malachite Green in Aquiculture Water Samples by Extractive Electrospray Ionization Mass Spectrometry.

    PubMed

    Fang, Xiaowei; Yang, Shuiping; Chingin, Konstantin; Zhu, Liang; Zhang, Xinglei; Zhou, Zhiquan; Zhao, Zhanfeng

    2016-01-01

    Exposure to malachite green (MG) may pose great health risks to humans; thus, it is of prime importance to develop fast and robust methods to quantitatively screen the presence of malachite green in water. Herein the application of extractive electrospray ionization mass spectrometry (EESI-MS) has been extended to the trace detection of MG within lake water and aquiculture water, due to the intensive use of MG as a biocide in fisheries. This method has the advantage of obviating offline liquid-liquid extraction or tedious matrix separation prior to the measurement of malachite green in native aqueous medium. The experimental results indicate that the extrapolated detection limit for MG was ~3.8 μg·L(-1) (S/N = 3) in lake water samples and ~0.5 μg·L(-1) in ultrapure water under optimized experimental conditions. The signal intensity of MG showed good linearity over the concentration range of 10-1000 μg·L(-1). Measurement of practical water samples fortified with MG at 0.01, 0.1 and 1.0 mg·L(-1) gave a good validation of the established calibration curve. The average recoveries and relative standard deviation (RSD) of malachite green in lake water and Carassius carassius fish farm effluent water were 115% (6.64% RSD), 85.4% (9.17% RSD) and 96.0% (7.44% RSD), respectively. Overall, the established EESI-MS/MS method has been demonstrated suitable for sensitive and rapid (<2 min per sample) quantitative detection of malachite green in various aqueous media, indicating its potential for online real-time monitoring of real life samples. PMID:27529262

  5. Quantitative Detection of Trace Malachite Green in Aquiculture Water Samples by Extractive Electrospray Ionization Mass Spectrometry

    PubMed Central

    Fang, Xiaowei; Yang, Shuiping; Chingin, Konstantin; Zhu, Liang; Zhang, Xinglei; Zhou, Zhiquan; Zhao, Zhanfeng

    2016-01-01

    Exposure to malachite green (MG) may pose great health risks to humans; thus, it is of prime importance to develop fast and robust methods to quantitatively screen the presence of malachite green in water. Herein the application of extractive electrospray ionization mass spectrometry (EESI-MS) has been extended to the trace detection of MG within lake water and aquiculture water, due to the intensive use of MG as a biocide in fisheries. This method has the advantage of obviating offline liquid-liquid extraction or tedious matrix separation prior to the measurement of malachite green in native aqueous medium. The experimental results indicate that the extrapolated detection limit for MG was ~3.8 μg·L−1 (S/N = 3) in lake water samples and ~0.5 μg·L−1 in ultrapure water under optimized experimental conditions. The signal intensity of MG showed good linearity over the concentration range of 10–1000 μg·L−1. Measurement of practical water samples fortified with MG at 0.01, 0.1 and 1.0 mg·L−1 gave a good validation of the established calibration curve. The average recoveries and relative standard deviation (RSD) of malachite green in lake water and Carassius carassius fish farm effluent water were 115% (6.64% RSD), 85.4% (9.17% RSD) and 96.0% (7.44% RSD), respectively. Overall, the established EESI-MS/MS method has been demonstrated suitable for sensitive and rapid (<2 min per sample) quantitative detection of malachite green in various aqueous media, indicating its potential for online real-time monitoring of real life samples. PMID:27529262

  6. Chemical approaches to detect and analyze protein sulfenic acids

    PubMed Central

    Furdui, Cristina M.; Poole, Leslie B.

    2013-01-01

    Orchestration of many processes relying on intracellular signal transduction is recognized to require the generation of hydrogen peroxide as a second messenger, yet relatively few molecular details of how this oxidant acts to regulate protein function are currently understood. This review describes emerging chemical tools and approaches that can be applied to study protein oxidation in biological systems, with a particular emphasis on a key player in protein redox regulation, cysteine sulfenic acid. While sulfenic acids (within purified proteins or simple mixtures) are detectable by physical approaches like X-ray crystallography, nuclear magnetic resonance and mass spectrometry, the propensity of these moieties to undergo further modification in complex biological systems has necessitated the development of chemical probes, reporter groups and analytical approaches to allow for their selective detection and quantification. Provided is an overview of techniques that are currently available for the study of sulfenic acids, and some of the biologically meaningful data that have been collected using such approaches. PMID:24105931

  7. Chemical approaches to detect and analyze protein sulfenic acids.

    PubMed

    Furdui, Cristina M; Poole, Leslie B

    2014-01-01

    Orchestration of many processes relying on intracellular signal transduction is recognized to require the generation of hydrogen peroxide as a second messenger, yet relatively few molecular details of how this oxidant acts to regulate protein function are currently understood. This review describes emerging chemical tools and approaches that can be applied to study protein oxidation in biological systems, with a particular emphasis on a key player in protein redox regulation, cysteine sulfenic acid. While sulfenic acids (within purified proteins or simple mixtures) are detectable by physical approaches like X-ray crystallography, nuclear magnetic resonance and mass spectrometry, the propensity of these moieties to undergo further modification in complex biological systems has necessitated the development of chemical probes, reporter groups and analytical approaches to allow for their selective detection and quantification. Provided is an overview of techniques that are currently available for the study of sulfenic acids, and some of the biologically meaningful data that have been collected using such approaches.

  8. Testing process predictions of models of risky choice: a quantitative model comparison approach

    PubMed Central

    Pachur, Thorsten; Hertwig, Ralph; Gigerenzer, Gerd; Brandstätter, Eduard

    2013-01-01

    This article presents a quantitative model comparison contrasting the process predictions of two prominent views on risky choice. One view assumes a trade-off between probabilities and outcomes (or non-linear functions thereof) and the separate evaluation of risky options (expectation models). Another view assumes that risky choice is based on comparative evaluation, limited search, aspiration levels, and the forgoing of trade-offs (heuristic models). We derived quantitative process predictions for a generic expectation model and for a specific heuristic model, namely the priority heuristic (Brandstätter et al., 2006), and tested them in two experiments. The focus was on two key features of the cognitive process: acquisition frequencies (i.e., how frequently individual reasons are looked up) and direction of search (i.e., gamble-wise vs. reason-wise). In Experiment 1, the priority heuristic predicted direction of search better than the expectation model (although neither model predicted the acquisition process perfectly); acquisition frequencies, however, were inconsistent with both models. Additional analyses revealed that these frequencies were primarily a function of what Rubinstein (1988) called “similarity.” In Experiment 2, the quantitative model comparison approach showed that people seemed to rely more on the priority heuristic in difficult problems, but to make more trade-offs in easy problems. This finding suggests that risky choice may be based on a mental toolbox of strategies. PMID:24151472

  9. Gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of Japanese encephalitis virus

    NASA Astrophysics Data System (ADS)

    Huang, Su-Hua; Yang, Tsuey-Ching; Tsai, Ming-Hong; Tsai, I.-Shou; Lu, Huang-Chih; Chuang, Pei-Hsin; Wan, Lei; Lin, Ying-Ju; Lai, Chih-Ho; Lin, Cheng-Wen

    2008-10-01

    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.

  10. Lifting the Humic Veil: A Novel Approach to Quantitating Occluded Iron in Peat Porewater

    NASA Astrophysics Data System (ADS)

    Veverica, T. J.; Kane, E. S.; Marcarelli, A. M.; Green, S. A.

    2014-12-01

    Anaerobic conditions common in peatlands lead microbes to reduce alternative electron acceptors for respiration in the order of their energetic yield: nitrate, manganese, ferric iron (Fe[III]), sulfate, and CO2. Many studies of peatland respiration report high CO2 fluxes that cannot be completely explained by measured pathways of microbial metabolism: it appears that an unquantified pool of electron acceptors is driving respiration and the efflux of CO2. The notion that undetected iron reduction may be an important contributing factor to these fluxes has been proposed, but has yet to be addressed. Among the possible causes for this anomaly is the widespread use of colorimetric assays for iron quantitation and speciation. Peat porewater contains high concentrations of dissolved humic and fulvic acids that occlude Fe(III), blocking colorimetric complex formation, potentially leading to an underestimate of Fe(III) in these challenging analytical matrices. We evaluated a novel ionic liquid extraction method in a variety of temperate peatland porewater samples and compared these results to data acquired simultaneously using colorimetric methods. We found a >50% average improvement in accuracy for total iron quantitation using ionic liquid extraction. Moreover, ionic liquid extraction of peat porewater consistently recovered Fe(III) whereas colorimetric methods detected 50-100% less, suggesting that DOM bound Fe(III) is not reliably detected using colorimetric assays in these systems and may represent a previously unidentified pool of electron acceptors in peat porewater.

  11. Quantitative Detection of Combustion Species using Ultra-Violet Diode Lasers

    NASA Technical Reports Server (NTRS)

    Pilgrim, J. S.; Peterson, K. A.

    2001-01-01

    Southwest Sciences is developing a new microgravity combustion diagnostic based on UV diode lasers. The instrument will allow absolute concentration measurements of combustion species on a variety of microgravity combustion platforms including the Space Station. Our approach uses newly available room temperature UV diode lasers, thereby keeping the instrument compact, rugged and energy efficient. The feasibility of the technique was demonstrated by measurement of CH radicals in laboratory flames. Further progress in fabrication technology of UV diode lasers at shorter wavelengths and higher power will result in detection of transient species in the deeper UV. High sensitivity detection of combustion radicals is provided with wavelength modulation absorption spectroscopy.

  12. Quantitative structure carcinogenicity relationship for detecting structural alerts in nitroso-compounds

    SciTech Connect

    Helguera, Aliuska Morales; Gonzalez, Maykel Perez . E-mail: mpgonzalez76@yahoo.es; Cordeiro, Maria Natalia D.S.; Perez, Miguel Angel Cabrera

    2007-06-01

    Prevention of environmentally induced cancers is a major health problem of which solutions depend on the rapid and accurate screening of potential chemical hazards. Lately, theoretical approaches such as the one proposed here - Quantitative Structure-Activity Relationship (QSAR) - are increasingly used for assessing the risks of environmental chemicals, since they can markedly reduce costs, avoid animal testing, and speed up policy decisions. This paper reports a QSAR study based on the Topological Substructural Molecular Design (TOPS-MODE) approach, aiming at predicting the rodent carcinogenicity of a set of nitroso-compounds selected from the Carcinogenic Potency Data Base (CPDB). The set comprises nitrosoureas (14 chemicals), N-nitrosamines (18 chemicals) C-nitroso-compounds (1 chemical), nitrosourethane (1 chemical) and nitrosoguanidine (1 chemical), which have been bioassayed in male rat using gavage as the route of administration. Here we are especially concerned in gathering the role of both parameters on the carcinogenic activity of this family of compounds. First, the regression model was derived, upon removal of one identified nitrosamine outlier, and was able to account for more than 84% of the variance in the experimental activity. Second, the TOPS-MODE approach afforded the bond contributions - expressed as fragment contributions to the carcinogenic activity - that can be interpreted and provide tools for better understanding the mechanisms of carcinogenesis. Finally, and most importantly, we demonstrate the potentialities of this approach towards the recognition of structural alerts for carcinogenicity predictions.

  13. Modeling approaches for qualitative and semi-quantitative analysis of cellular signaling networks

    PubMed Central

    2013-01-01

    A central goal of systems biology is the construction of predictive models of bio-molecular networks. Cellular networks of moderate size have been modeled successfully in a quantitative way based on differential equations. However, in large-scale networks, knowledge of mechanistic details and kinetic parameters is often too limited to allow for the set-up of predictive quantitative models. Here, we review methodologies for qualitative and semi-quantitative modeling of cellular signal transduction networks. In particular, we focus on three different but related formalisms facilitating modeling of signaling processes with different levels of detail: interaction graphs, logical/Boolean networks, and logic-based ordinary differential equations (ODEs). Albeit the simplest models possible, interaction graphs allow the identification of important network properties such as signaling paths, feedback loops, or global interdependencies. Logical or Boolean models can be derived from interaction graphs by constraining the logical combination of edges. Logical models can be used to study the basic input–output behavior of the system under investigation and to analyze its qualitative dynamic properties by discrete simulations. They also provide a suitable framework to identify proper intervention strategies enforcing or repressing certain behaviors. Finally, as a third formalism, Boolean networks can be transformed into logic-based ODEs enabling studies on essential quantitative and dynamic features of a signaling network, where time and states are continuous. We describe and illustrate key methods and applications of the different modeling formalisms and discuss their relationships. In particular, as one important aspect for model reuse, we will show how these three modeling approaches can be combined to a modeling pipeline (or model hierarchy) allowing one to start with the simplest representation of a signaling network (interaction graph), which can later be refined to

  14. Modeling approaches for qualitative and semi-quantitative analysis of cellular signaling networks.

    PubMed

    Samaga, Regina; Klamt, Steffen

    2013-01-01

    A central goal of systems biology is the construction of predictive models of bio-molecular networks. Cellular networks of moderate size have been modeled successfully in a quantitative way based on differential equations. However, in large-scale networks, knowledge of mechanistic details and kinetic parameters is often too limited to allow for the set-up of predictive quantitative models.Here, we review methodologies for qualitative and semi-quantitative modeling of cellular signal transduction networks. In particular, we focus on three different but related formalisms facilitating modeling of signaling processes with different levels of detail: interaction graphs, logical/Boolean networks, and logic-based ordinary differential equations (ODEs). Albeit the simplest models possible, interaction graphs allow the identification of important network properties such as signaling paths, feedback loops, or global interdependencies. Logical or Boolean models can be derived from interaction graphs by constraining the logical combination of edges. Logical models can be used to study the basic input-output behavior of the system under investigation and to analyze its qualitative dynamic properties by discrete simulations. They also provide a suitable framework to identify proper intervention strategies enforcing or repressing certain behaviors. Finally, as a third formalism, Boolean networks can be transformed into logic-based ODEs enabling studies on essential quantitative and dynamic features of a signaling network, where time and states are continuous.We describe and illustrate key methods and applications of the different modeling formalisms and discuss their relationships. In particular, as one important aspect for model reuse, we will show how these three modeling approaches can be combined to a modeling pipeline (or model hierarchy) allowing one to start with the simplest representation of a signaling network (interaction graph), which can later be refined to logical

  15. A hybrid calibration-free/artificial neural networks approach to the quantitative analysis of LIBS spectra

    NASA Astrophysics Data System (ADS)

    D'Andrea, Eleonora; Pagnotta, Stefano; Grifoni, Emanuela; Legnaioli, Stefano; Lorenzetti, Giulia; Palleschi, Vincenzo; Lazzerini, Beatrice

    2015-03-01

    A `hybrid' method is proposed for the quantitative analysis of materials by LIBS, combining the precision of the calibration-free LIBS (CF-LIBS) algorithm with the quickness of artificial neural networks. The method allows the precise determination of the samples' composition even in the presence of relatively large laser fluctuations and matrix effects. To show the strength and robustness of this approach, a number of synthetic LIBS spectra of Cu-Ni binary alloys with different composition were computer-simulated, in correspondence of different plasma temperatures, electron number densities and ablated mass. The CF-LIBS/ANN approach here proposed demonstrated to be capable, after appropriate training, of `learning' the basic physical relations between the experimentally measured line intensities and the plasma parameters. Because of that the composition of the sample can be correctly determined, as in CF-LIBS measurements, but in a much shorter time.

  16. Quantitative detection of powdered activated carbon in wastewater treatment plant effluent by thermogravimetric analysis (TGA).

    PubMed

    Krahnstöver, Therese; Plattner, Julia; Wintgens, Thomas

    2016-09-15

    For the elimination of potentially harmful micropollutants, powdered activated carbon (PAC) adsorption is applied in many wastewater treatment plants (WWTP). This holds the risk of PAC leakage into the WWTP effluent and desorption of contaminants into natural water bodies. In order to assess a potential PAC leakage, PAC concentrations below several mg/L have to be detected in the WWTP effluent. None of the methods that are used for water analysis today are able to differentiate between activated carbon and solid background matrix. Thus, a selective, quantitative and easily applicable method is still needed for the detection of PAC residues in wastewater. In the present study, a method was developed to quantitatively measure the PAC content in wastewater by using filtration and thermogravimetric analysis (TGA), which is a well-established technique for the distinction between different solid materials. For the sample filtration, quartz filters with a temperature stability up to 950 °C were used. This allowed for sensitive and well reproducible measurements, as the TGA was not affected by the presence of the filter. The sample's mass fractions were calculated by integrating the mass decrease rate obtained by TGA in specific, clearly identifiable peak areas. A two-step TGA heating method consisting of N2 and O2 atmospheres led to a good differentiation between PAC and biological background matrix, thanks to the reduction of peak overlapping. A linear correlation was found between a sample's PAC content and the corresponding peak areas under N2 and O2, the sample volume and the solid mass separated by filtration. Based on these findings, various wastewater samples from different WWTPs were then analyzed by TGA with regard to their PAC content. It was found that, compared to alternative techniques such as measurement of turbidity or total suspended solids, the newly developed TGA method allows for a quantitative and selective detection of PAC concentrations down to 0

  17. Quantitative detection of powdered activated carbon in wastewater treatment plant effluent by thermogravimetric analysis (TGA).

    PubMed

    Krahnstöver, Therese; Plattner, Julia; Wintgens, Thomas

    2016-09-15

    For the elimination of potentially harmful micropollutants, powdered activated carbon (PAC) adsorption is applied in many wastewater treatment plants (WWTP). This holds the risk of PAC leakage into the WWTP effluent and desorption of contaminants into natural water bodies. In order to assess a potential PAC leakage, PAC concentrations below several mg/L have to be detected in the WWTP effluent. None of the methods that are used for water analysis today are able to differentiate between activated carbon and solid background matrix. Thus, a selective, quantitative and easily applicable method is still needed for the detection of PAC residues in wastewater. In the present study, a method was developed to quantitatively measure the PAC content in wastewater by using filtration and thermogravimetric analysis (TGA), which is a well-established technique for the distinction between different solid materials. For the sample filtration, quartz filters with a temperature stability up to 950 °C were used. This allowed for sensitive and well reproducible measurements, as the TGA was not affected by the presence of the filter. The sample's mass fractions were calculated by integrating the mass decrease rate obtained by TGA in specific, clearly identifiable peak areas. A two-step TGA heating method consisting of N2 and O2 atmospheres led to a good differentiation between PAC and biological background matrix, thanks to the reduction of peak overlapping. A linear correlation was found between a sample's PAC content and the corresponding peak areas under N2 and O2, the sample volume and the solid mass separated by filtration. Based on these findings, various wastewater samples from different WWTPs were then analyzed by TGA with regard to their PAC content. It was found that, compared to alternative techniques such as measurement of turbidity or total suspended solids, the newly developed TGA method allows for a quantitative and selective detection of PAC concentrations down to 0

  18. An axial approach to detection in capillary electrophoresis

    SciTech Connect

    Taylor, J.A.

    1993-05-01

    Our approach involves on-axis illumination of the compounds inside the capillary detection region and is applied to absorbance and fluorescence detection. Absorbance measurements were made by focussing an incident laser beam into one capillary end; by using signals collected over the entire length of analyte band, this enhances the analytical path length of conventional absorbance detection 60x. This instrument offers a 15x improvement in detection limits. Three fluorescence detection experiments are discussed, all of which involve insertion of an optical fiber into capillary. The first uses a high refractive index liquid phase to obtain total internal reflectance along capillary axis, this reducing light scatter. The second uses a charge-coupled device camera for simultaneous imaging of a capillary array (this may be useful in genome sequencing, etc.). The third is a study of fluid motion inside the capillary under pressure-driven and electroosmotic flow. The thesis is divided into four parts. Figs, tabs.

  19. Sequential Bayesian Detection: A Model-Based Approach

    SciTech Connect

    Sullivan, E J; Candy, J V

    2007-08-13

    Sequential detection theory has been known for a long time evolving in the late 1940's by Wald and followed by Middleton's classic exposition in the 1960's coupled with the concurrent enabling technology of digital computer systems and the development of sequential processors. Its development, when coupled to modern sequential model-based processors, offers a reasonable way to attack physics-based problems. In this chapter, the fundamentals of the sequential detection are reviewed from the Neyman-Pearson theoretical perspective and formulated for both linear and nonlinear (approximate) Gauss-Markov, state-space representations. We review the development of modern sequential detectors and incorporate the sequential model-based processors as an integral part of their solution. Motivated by a wealth of physics-based detection problems, we show how both linear and nonlinear processors can seamlessly be embedded into the sequential detection framework to provide a powerful approach to solving non-stationary detection problems.

  20. Sequential Bayesian Detection: A Model-Based Approach

    SciTech Connect

    Candy, J V

    2008-12-08

    Sequential detection theory has been known for a long time evolving in the late 1940's by Wald and followed by Middleton's classic exposition in the 1960's coupled with the concurrent enabling technology of digital computer systems and the development of sequential processors. Its development, when coupled to modern sequential model-based processors, offers a reasonable way to attack physics-based problems. In this chapter, the fundamentals of the sequential detection are reviewed from the Neyman-Pearson theoretical perspective and formulated for both linear and nonlinear (approximate) Gauss-Markov, state-space representations. We review the development of modern sequential detectors and incorporate the sequential model-based processors as an integral part of their solution. Motivated by a wealth of physics-based detection problems, we show how both linear and nonlinear processors can seamlessly be embedded into the sequential detection framework to provide a powerful approach to solving non-stationary detection problems.

  1. A novel approach to quantitating leukemia fusion transcripts by qRT-PCR without the need for standard curves.

    PubMed

    Schumacher, Jonathan A; Scott Reading, N; Szankasi, Philippe; Matynia, Anna P; Kelley, Todd W

    2015-08-01

    Acute myeloid leukemia patients with recurrent cytogenetic abnormalities including inv(16);CBFB-MYH11 and t(15;17);PML-RARA may be assessed by monitoring the levels of the corresponding abnormal fusion transcripts by quantitative reverse transcription-PCR (qRT-PCR). Such testing is important for evaluating the response to therapy and for the detection of early relapse. Existing qRT-PCR methods are well established and in widespread use in clinical laboratories but they are laborious and require the generation of standard curves. Here, we describe a new method to quantitate fusion transcripts in acute myeloid leukemia by qRT-PCR without the need for standard curves. Our approach uses a plasmid calibrator containing both a fusion transcript sequence and a reference gene sequence, representing a perfect normalized copy number (fusion transcript copy number/reference gene transcript copy number; NCN) of 1.0. The NCN of patient specimens can be calculated relative to that of the single plasmid calibrator using experimentally derived PCR efficiency values. We compared the data obtained using the plasmid calibrator method to commercially available assays using standard curves and found that the results obtained by both methods are comparable over a broad range of values with similar sensitivities. Our method has the advantage of simplicity and is therefore lower in cost and may be less subject to errors that may be introduced during the generation of standard curves.

  2. Quantitative analysis of electrically detected Ramsey fringes in P-doped Si

    NASA Astrophysics Data System (ADS)

    Greenland, P. T.; Matmon, G.; Villis, B. J.; Bowyer, E. T.; Li, Juerong; Murdin, B. N.; van der Meer, A. F. G.; Redlich, B.; Pidgeon, C. R.; Aeppli, G.

    2015-10-01

    This work describes detection of the laser preparation and subsequent coherent manipulation of the quantum states of orbital levels of donors in doped Si, by measuring the voltage drop across an irradiated Si sample. This electrical signal, which arises from thermal ionization of excited orbital states, and which is detected on a millisecond time scale by a voltmeter, leads to much more sensitive detection than can be had using optical methods, but has not before been quantitatively described from first principles. We present here a unified theory which relates the voltage drop across the sample to the wave function of the excited donors, and compare its predictions to experiments in which pairs of picosecond pulses from the Dutch free-electron laser FELIX are used to resonantly and coherently excite P donors in Si. Although the voltage drop varies on a millisecond time scale we are able to measure Ramsey oscillation of the excitation on a picosecond time scale, thus confirming that the donor wave function, and not just its excited state population, is crucial in determining the electrical signal. We are also able to extract the recombination rate coefficient to the ground state of the donor as well as the photoionization cross section of the excited state and phonon induced thermal ionization rate from the excited state. These quantities, which were previously of limited interest, are here shown to be important in the description of electrical detection, which, in our unoptimized configuration, is sensitive enough to enable us to detect the excitation of ˜107 donors.

  3. Computer-aided CT coronary artery stenosis detection: comparison with human reading and quantitative coronary angiography.

    PubMed

    Rief, Matthias; Kranz, Anisha; Hartmann, Lisa; Roehle, Robert; Laule, Michael; Dewey, Marc

    2014-12-01

    To evaluate computer-aided stenosis detection for computed tomography coronary angiography (CTA) in comparison with human reading and conventional coronary angiography (CCA) as the reference standard. 50 patients underwent CTA and CCA and out of these 44 were evaluable for computer-aided stenosis detection. The diagnostic performance of the software and of human reading were compared and quantitative coronary angiography (QCA) served as the reference standard for the detection of significant stenosis (>50 %). Overall, three readers with high (reader 1), intermediate (reader 2) and low (reader 3) experience in cardiac CT imaging performed the manual CTA evaluation on a commercially available workstation, whereas the automated software processed the datasets without any human interaction. The prevalence of coronary artery disease was 41 % (18/44) and QCA indicated significant stenosis (>50 %) in 33 coronary vessels. The automated software accurately diagnosed 18 individuals with significant coronary artery disease (CAD), and correctly ruled out CAD in 10 patients. In summary the sensitivity of computer-aided detection was 100 %/94 % (per-patient/per-vessel) and the specificity was 38 %/70 %, the positive predictive value (PPV) was 53 %/42 % and the negative predictive value (NPV) was 100 %/98 %. In comparison, reader 1-3 showed per-patient sensitivities of 100/94/89 %, specificities of 73/69/50 %, PPVs of 72/68/55 % and NPVs of 100/95/87 %. Computer-aided detection yields a high NPV that is comparable to more experienced human readers. However, PPV is rather low and in the range of an unexperienced reader.

  4. Quantitative approach to skin field cancerization using a nanoencapsulated photodynamic therapy agent: a pilot study

    PubMed Central

    Passos, Simone K; de Souza, Paulo EN; Soares, Priscila KP; Eid, Danglades RM; Primo, Fernando L; Tedesco, Antonio Cláudio; Lacava, Zulmira GM; Morais, Paulo C

    2013-01-01

    Background This paper introduces a new nanoformulation of 5-aminolevulinic acid (nano-ALA) as well as a novel quantitative approach towards evaluating field cancerization for actinic keratosis and/or skin photodamage. In this pilot study, we evaluated field cancerization using nano-ALA and methyl aminolevulinate (MAL), the latter being commercialized as Metvix®. Methods and results Photodynamic therapy was used for the treatment of patients with selected skin lesions, whereas the fluorescence of the corresponding photosensitizer was used to evaluate the time evolution of field cancerization in a quantitative way. Field cancerization was quantified using newly developed color image segmentation software. Using photodynamic therapy as the precancer skin treatment and the approach introduced herein for evaluation of fluorescent area, we found that the half-life of field cancerization reduction was 43.3 days and 34.3 days for nano-ALA and MAL, respectively. We also found that nano-ALA targeted about 45% more skin lesion areas than MAL. Further, we found the mean reduction in area of skin field cancerization was about 10% greater for nano-ALA than for MAL. Conclusion Although preliminary, our findings indicate that the efficacy of nano-ALA in treating skin field cancerization is higher than that of MAL. PMID:23450821

  5. Applying quantitative structure-activity relationship approaches to nanotoxicology: current status and future potential.

    PubMed

    Winkler, David A; Mombelli, Enrico; Pietroiusti, Antonio; Tran, Lang; Worth, Andrew; Fadeel, Bengt; McCall, Maxine J

    2013-11-01

    The potential (eco)toxicological hazard posed by engineered nanoparticles is a major scientific and societal concern since several industrial sectors (e.g. electronics, biomedicine, and cosmetics) are exploiting the innovative properties of nanostructures resulting in their large-scale production. Many consumer products contain nanomaterials and, given their complex life-cycle, it is essential to anticipate their (eco)toxicological properties in a fast and inexpensive way in order to mitigate adverse effects on human health and the environment. In this context, the application of the structure-toxicity paradigm to nanomaterials represents a promising approach. Indeed, according to this paradigm, it is possible to predict toxicological effects induced by chemicals on the basis of their structural similarity with chemicals for which toxicological endpoints have been previously measured. These structure-toxicity relationships can be quantitative or qualitative in nature and they can predict toxicological effects directly from the physicochemical properties of the entities (e.g. nanoparticles) of interest. Therefore, this approach can aid in prioritizing resources in toxicological investigations while reducing the ethical and monetary costs that are related to animal testing. The purpose of this review is to provide a summary of recent key advances in the field of QSAR modelling of nanomaterial toxicity, to identify the major gaps in research required to accelerate the use of quantitative structure-activity relationship (QSAR) methods, and to provide a roadmap for future research needed to achieve QSAR models useful for regulatory purposes. PMID:23165187

  6. A quantitative approach to assessing the efficacy of occupant protection programs: A case study from Montana.

    PubMed

    Manlove, Kezia; Stanley, Laura; Peck, Alyssa

    2015-10-01

    Quantitative evaluation of vehicle occupant protection programs is critical for ensuring efficient government resource allocation, but few methods exist for conducting evaluation across multiple programs simultaneously. Here we present an analysis of occupant protection efficacy in the state of Montana. This approach relies on seat belt compliance rates as measured by the National Occupant Protection Usage Survey (NOPUS). A hierarchical logistic regression model is used to estimate the impacts of four Montana Department of Transportation (MDT)-funded occupant protection programs used in the state of Montana, following adjustment for a suite of potential confounders. Activity from two programs, Buckle Up coalitions and media campaigns, are associated with increased seat belt use in Montana, whereas the impact of another program, Selective Traffic Enforcement, is potentially masked by other program activity. A final program, Driver's Education, is not associated with any shift in seat belt use. This method allows for a preliminary quantitative estimation of program impacts without requiring states to obtain any new seat belt use data. This approach provides states a preliminary look at program impacts, and a means for carefully planning future program allocation and investigation.

  7. Applying quantitative structure-activity relationship approaches to nanotoxicology: current status and future potential.

    PubMed

    Winkler, David A; Mombelli, Enrico; Pietroiusti, Antonio; Tran, Lang; Worth, Andrew; Fadeel, Bengt; McCall, Maxine J

    2013-11-01

    The potential (eco)toxicological hazard posed by engineered nanoparticles is a major scientific and societal concern since several industrial sectors (e.g. electronics, biomedicine, and cosmetics) are exploiting the innovative properties of nanostructures resulting in their large-scale production. Many consumer products contain nanomaterials and, given their complex life-cycle, it is essential to anticipate their (eco)toxicological properties in a fast and inexpensive way in order to mitigate adverse effects on human health and the environment. In this context, the application of the structure-toxicity paradigm to nanomaterials represents a promising approach. Indeed, according to this paradigm, it is possible to predict toxicological effects induced by chemicals on the basis of their structural similarity with chemicals for which toxicological endpoints have been previously measured. These structure-toxicity relationships can be quantitative or qualitative in nature and they can predict toxicological effects directly from the physicochemical properties of the entities (e.g. nanoparticles) of interest. Therefore, this approach can aid in prioritizing resources in toxicological investigations while reducing the ethical and monetary costs that are related to animal testing. The purpose of this review is to provide a summary of recent key advances in the field of QSAR modelling of nanomaterial toxicity, to identify the major gaps in research required to accelerate the use of quantitative structure-activity relationship (QSAR) methods, and to provide a roadmap for future research needed to achieve QSAR models useful for regulatory purposes.

  8. Gold Nanoparticle Labeling Based ICP-MS Detection/Measurement of Bacteria, and Their Quantitative Photothermal Destruction

    PubMed Central

    Lin, Yunfeng

    2015-01-01

    Bacteria such as Salmonella and E. coli present a great challenge in public health care in today’s society. Protection of public safety against bacterial contamination and rapid diagnosis of infection require simple and fast assays for the detection and elimination of bacterial pathogens. After utilizing Salmonella DT104 as an example bacterial strain for our investigation, we report a rapid and sensitive assay for the qualitative and quantitative detection of bacteria by using antibody affinity binding, popcorn shaped gold nanoparticle (GNPOPs) labeling, surfance enchanced Raman spectroscopy (SERS), and inductively coupled plasma mass spectrometry (ICP-MS) detection. For qualitative analysis, our assay can detect Salmonella within 10 min by Raman spectroscopy; for quantitative analysis, our assay has the ability to measure as few as 100 Salmonella DT104 in a 1 mL sample (100 CFU/mL) within 40 min. Based on the quantitative detection, we investigated the quantitative destruction of Salmonella DT104, and the assay’s photothermal efficiency in order to reduce the amount of GNPOPs in the assay to ultimately to eliminate any potential side effects/toxicity to the surrounding cells in vivo. Results suggest that our assay may serve as a promising candidate for qualitative and quantitative detection and elimination of a variety of bacterial pathogens. PMID:26417447

  9. Detection and identification of genotypes of Prototheca zopfii in clinical samples by quantitative PCR analysis.

    PubMed

    Onozaki, Masanobu; Makimura, Koichi; Satoh, Kazuo; Hasegawa, Atsuhiko

    2013-01-01

    In this study, a specific quantitative PCR system for the detection and identification of Prototheca zopfii genotypes was developed using a TaqMan(®) MGB probe and ResoLight dye. The P. zopfii-specific primers 18PZF1 and 18PZR1 were generated on the basis of the alignment of the small subunit ribosomal DNA domain base sequences of the genera Chlorella and Prototheca obtained from DDBJ/EMBL/GenBank, and the TaqMan(®) MGB probe PZP1 was designed corresponding to this amplification region. Analysis of the melting curves of the amplicons using ResoLight dye was able to differentiate between P. zopfii genotypes 1 and 2. The specificity of this detection system was examined using strains from a culture collection (28 strains) and clinical isolates (140 strains). The TaqMan(®) MGB probe amplicon was detected only in reference strains of P. zopfii (n = 12) and clinical isolates (n = 135). Ninety-two clinical specimens from cows with mastitis (36 samples) and healthy controls (56 samples) were also tested. All isolates from milk samples (n = 92) and clinical isolates (n = 135) were identified as P. zopfii genotype 2. PMID:24047735

  10. Rapid quantitative detection of Aeromonas hydrophila strains associated with disease outbreaks in catfish aquaculture.

    PubMed

    Griffin, Matt J; Goodwin, Andrew E; Merry, Gwenn E; Liles, Mark R; Williams, Malachi A; Ware, Cynthia; Waldbieser, Geoffrey C

    2013-07-01

    A new strain of Aeromonas hydrophila has been implicated in significant losses in farm-raised catfish. Outbreaks attributable to this new strain began in Alabama in the summer of 2009 and have spread to Arkansas and Mississippi in subsequent years. These outbreaks mostly afflicted market-sized fish and resulted in considerable losses in short periods of time. The present research was designed to develop an expeditious diagnostic procedure to detect the new strains of A. hydrophila due to the rapid onset and biosecurity concerns associated with this new disease. A discriminatory quantitative polymerase chain reaction assay was developed using gene sequences unique to the virulent strains identified in a related comparative genomic study. Using this assay, suspect colonies on a culture plate can be positively identified as the new strain within 2 hr. The assay is repeatable and reproducible with a linear dynamic range covering 8 orders of magnitude and a sensitivity of approximately 7 copies of target DNA in a 15-µl reaction. In addition, the assay is able to detect and quantify the virulent strain from catfish tissues (0.025 g), pond water (40 ml), and sediments (0.25 g) with a sensitivity limit of approximately 100 bacteria in a sample. This assay provides rapid discrimination between the new virulent strain and more common A. hydrophila and is useful for epidemiological studies involving the detection and quantification of the virulent strain in environmental samples and fish tissues.

  11. A miniaturized optoelectronic system for rapid quantitative label-free detection of harmful species in food

    NASA Astrophysics Data System (ADS)

    Raptis, Ioannis; Misiakos, Konstantinos; Makarona, Eleni; Salapatas, Alexandros; Petrou, Panagiota; Kakabakos, Sotirios; Botsialas, Athanasios; Jobst, Gerhard; Haasnoot, Willem; Fernandez-Alba, Amadeo; Lees, Michelle; Valamontes, Evangelos

    2016-03-01

    Optical biosensors have emerged in the past decade as the most promising candidates for portable, highly-sensitive bioanalytical systems that can be employed for in-situ measurements. In this work, a miniaturized optoelectronic system for rapid, quantitative, label-free detection of harmful species in food is presented. The proposed system has four distinctive features that can render to a powerful tool for the next generation of Point-of-Need applications, namely it accommodates the light sources and ten interferometric biosensors on a single silicon chip of a less-than-40mm2 footprint, each sensor can be individually functionalized for a specific target analyte, the encapsulation can be performed at the wafer-scale, and finally it exploits a new operation principle, Broad-band Mach-Zehnder Interferometry to ameliorate its analytical capabilities. Multi-analyte evaluation schemes for the simultaneous detection of harmful contaminants, such as mycotoxins, allergens and pesticides, proved that the proposed system is capable of detecting within short time these substances at concentrations below the limits imposed by regulatory authorities, rendering it to a novel tool for the near-future food safety applications.

  12. Quantitative analysis and detection of adulteration in pork using near-infrared spectroscopy

    NASA Astrophysics Data System (ADS)

    Fan, Yuxia; Cheng, Fang; Xie, Lijuan

    2010-04-01

    Authenticity is an important food quality criterion. Rapid methods for confirming authenticity or detecting adulteration are increasingly demanded by food processors and consumers. Near infrared (NIR) spectroscopy has been used to detect economic adulteration in pork . Pork samples were adulterated with liver and chicken in 10% increments. Prediction and quantitative analysis were done using raw data and pretreatment spectra. The optimal prediction result was achieved by partial least aquares(PLS) regression with standard normal variate(SNV) pretreatment for pork adulterated with liver samples, and the correlation coefficient(R value), the root mean square error of calibration(RMSEC) and the root mean square error of prediction (RMSEP) were 0.97706, 0.0673 and 0.0732, respectively. The best model for pork meat adulterated with chicken samples was obtained by PLS with the raw spectra, and the correlation coefficient(R value), RMSEP and RMSEC were 0.98614, 0.0525, and 0.122, respectively. The result shows that NIR technology can be successfully used to detect adulteration in pork meat adulterated with liver and chicken.

  13. [Detection of enteric pathogenic bacteria from surface waters by quantitative polymerase chain reaction (QPCR)].

    PubMed

    Liu, Yong-Jun; Zhang, Chong-Miao; Wang, Xiao-Chang; Lü, Ying-Jun; Zuo, Li-Li

    2008-05-01

    A rapid quantitative polymerase chain reaction (QPCR) analysis method with universal primers was developed to detect cell densities of the enteric pathogenic bacteria from 5 surface water of Xi'an City for 4 months continuously. And the detection results by QPCR method were compared with counts of coliforms colony-forming units (CFU) determined by membrane filter (MF) analysis. The results showed that QPCR method had an estimated 94% confidence, and detection limit was 2.7 Escherichia coli cells per sample in undiluted DNA extracts. For five surface waters (N = 60), the geometric mean of pathogenic bacteria concentration determined by QPCR was 2.2-5 times of corresponding coliform CFU determined by MF analysis. Using QPCR analysis, these geometric means of pathogenic bacteria concentration ranged from 25 CCE/100 mL to 67 000 CCE/100 mL. Using MF culture analysis, coliforms ranged from 3 CFU/100 mL to 45 000 CFU/100 mL. Regression analysis showed that there was a significant positive correlation between pathogenic bacteria determined by QPCR method and coliforms determined by MF method, the correlation coefficient (r) was 0.983.

  14. Icing detection from geostationary satellite data using machine learning approaches

    NASA Astrophysics Data System (ADS)

    Lee, J.; Ha, S.; Sim, S.; Im, J.

    2015-12-01

    Icing can cause a significant structural damage to aircraft during flight, resulting in various aviation accidents. Icing studies have been typically performed using two approaches: one is a numerical model-based approach and the other is a remote sensing-based approach. The model based approach diagnoses aircraft icing using numerical atmospheric parameters such as temperature, relative humidity, and vertical thermodynamic structure. This approach tends to over-estimate icing according to the literature. The remote sensing-based approach typically uses meteorological satellite/ground sensor data such as Geostationary Operational Environmental Satellite (GOES) and Dual-Polarization radar data. This approach detects icing areas by applying thresholds to parameters such as liquid water path and cloud optical thickness derived from remote sensing data. In this study, we propose an aircraft icing detection approach which optimizes thresholds for L1B bands and/or Cloud Optical Thickness (COT) from Communication, Ocean and Meteorological Satellite-Meteorological Imager (COMS MI) and newly launched Himawari-8 Advanced Himawari Imager (AHI) over East Asia. The proposed approach uses machine learning algorithms including decision trees (DT) and random forest (RF) for optimizing thresholds of L1B data and/or COT. Pilot Reports (PIREPs) from South Korea and Japan were used as icing reference data. Results show that RF produced a lower false alarm rate (1.5%) and a higher overall accuracy (98.8%) than DT (8.5% and 75.3%), respectively. The RF-based approach was also compared with the existing COMS MI and GOES-R icing mask algorithms. The agreements of the proposed approach with the existing two algorithms were 89.2% and 45.5%, respectively. The lower agreement with the GOES-R algorithm was possibly due to the high uncertainty of the cloud phase product from COMS MI.

  15. A paper/polymer hybrid microfluidic microplate for rapid quantitative detection of multiple disease biomarkers.

    PubMed

    Sanjay, Sharma T; Dou, Maowei; Sun, Jianjun; Li, XiuJun

    2016-01-01

    Enzyme linked immunosorbent assay (ELISA) is one of the most widely used laboratory disease diagnosis methods. However, performing ELISA in low-resource settings is limited by long incubation time, large volumes of precious reagents, and well-equipped laboratories. Herein, we developed a simple, miniaturized paper/PMMA (poly(methyl methacrylate)) hybrid microfluidic microplate for low-cost, high throughput, and point-of-care (POC) infectious disease diagnosis. The novel use of porous paper in flow-through microwells facilitates rapid antibody/antigen immobilization and efficient washing, avoiding complicated surface modifications. The top reagent delivery channels can simply transfer reagents to multiple microwells thus avoiding repeated manual pipetting and costly robots. Results of colorimetric ELISA can be observed within an hour by the naked eye. Quantitative analysis was achieved by calculating the brightness of images scanned by an office scanner. Immunoglobulin G (IgG) and Hepatitis B surface Antigen (HBsAg) were quantitatively analyzed with good reliability in human serum samples. Without using any specialized equipment, the limits of detection of 1.6 ng/mL for IgG and 1.3 ng/mL for HBsAg were achieved, which were comparable to commercial ELISA kits using specialized equipment. We envisage that this simple POC hybrid microplate can have broad applications in various bioassays, especially in resource-limited settings. PMID:27456979

  16. On-chip quantitative detection of pathogen genes by autonomous microfluidic PCR platform.

    PubMed

    Tachibana, Hiroaki; Saito, Masato; Shibuya, Shogo; Tsuji, Koji; Miyagawa, Nobuyuki; Yamanaka, Keiichiro; Tamiya, Eiichi

    2015-12-15

    Polymerase chain reaction (PCR)-based genetic testing has become a routine part of clinical diagnoses and food testing. In these fields, rapid, easy-to-use, and cost-efficient PCR chips are expected to be appeared for providing such testing on-site. In this study, a new autonomous disposable plastic microfluidic PCR chip was created, and was utilized for quantitative detection of pathogenic microorganisms. To control the capillary flow of the following solution in the PCR microchannel, a driving microchannel was newly designed behind the PCR microchannel. This allowed the effective PCR by simply dropping the PCR solution onto the inlet without any external pumps. In order to achieve disposability, injection-molded cyclo-olefin polymer (COP) of a cost-competitive plastic was used for the PCR chip. We discovered that coating the microchannel walls with non-ionic surfactant produced a suitable hydrophilic surface for driving the capillary flow through the 1250-mm long microchannel. As a result, quantitative real-time PCR with the lowest initial concentration of human, Escherichia coli (E. coli), and pathogenic E. coli O157 genomic DNA of 4, 0.0019, 0.031 pg/μl, respectively, was successfully achieved in less than 18 min. Our results indicate that the platform presented in this study provided a rapid, easy-to-use, and low-cost real-time PCR system that could be potentially used for on-site gene testing.

  17. A paper/polymer hybrid microfluidic microplate for rapid quantitative detection of multiple disease biomarkers

    PubMed Central

    Sanjay, Sharma T.; Dou, Maowei; Sun, Jianjun; Li, XiuJun

    2016-01-01

    Enzyme linked immunosorbent assay (ELISA) is one of the most widely used laboratory disease diagnosis methods. However, performing ELISA in low-resource settings is limited by long incubation time, large volumes of precious reagents, and well-equipped laboratories. Herein, we developed a simple, miniaturized paper/PMMA (poly(methyl methacrylate)) hybrid microfluidic microplate for low-cost, high throughput, and point-of-care (POC) infectious disease diagnosis. The novel use of porous paper in flow-through microwells facilitates rapid antibody/antigen immobilization and efficient washing, avoiding complicated surface modifications. The top reagent delivery channels can simply transfer reagents to multiple microwells thus avoiding repeated manual pipetting and costly robots. Results of colorimetric ELISA can be observed within an hour by the naked eye. Quantitative analysis was achieved by calculating the brightness of images scanned by an office scanner. Immunoglobulin G (IgG) and Hepatitis B surface Antigen (HBsAg) were quantitatively analyzed with good reliability in human serum samples. Without using any specialized equipment, the limits of detection of 1.6 ng/mL for IgG and 1.3 ng/mL for HBsAg were achieved, which were comparable to commercial ELISA kits using specialized equipment. We envisage that this simple POC hybrid microplate can have broad applications in various bioassays, especially in resource-limited settings. PMID:27456979

  18. Quantitative Phase Fraction Detection in Organic Photovoltaic Materials through EELS Imaging

    DOE PAGESBeta

    Dyck, Ondrej; Hu, Sheng; Das, Sanjib; Keum, Jong; Xiao, Kai; Khomami, Bamin; Duscher, Gerd

    2015-11-24

    Organic photovoltaic materials have recently seen intense interest from the research community. Improvements in device performance are occurring at an impressive rate; however, visualization of the active layer phase separation still remains a challenge. Our paper outlines the application of two electron energy-loss spectroscopic (EELS) imaging techniques that can complement and enhance current phase detection techniques. Specifically, the bulk plasmon peak position, often used to produce contrast between phases in energy filtered transmission electron microscopy (EFTEM), is quantitatively mapped across a sample cross section. One complementary spectrum image capturing the carbon and sulfur core loss edges is compared with themore » plasmon peak map and found to agree quite well, indicating that carbon and sulfur density differences between the two phases also allows phase discrimination. Additionally, an analytical technique for determining absolute atomic areal density is used to produce an absolute carbon and sulfur areal density map. We also show how these maps may be re-interpreted as a phase ratio map, giving quantitative information about the purity of the phases within the junction.« less

  19. Quantitative detection of nitric oxide in exhaled human breath by extractive electrospray ionization mass spectrometry

    NASA Astrophysics Data System (ADS)

    Pan, Susu; Tian, Yong; Li, Ming; Zhao, Jiuyan; Zhu, Lanlan; Zhang, Wei; Gu, Haiwei; Wang, Haidong; Shi, Jianbo; Fang, Xiang; Li, Penghui; Chen, Huanwen

    2015-03-01

    Exhaled nitric oxide (eNO) is a useful biomarker of various physiological conditions, including asthma and other pulmonary diseases. Herein a fast and sensitive analytical method has been developed for the quantitative detection of eNO based on extractive electrospray ionization mass spectrometry (EESI-MS). Exhaled NO molecules selectively reacted with 2-phenyl-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO) reagent, and eNO concentration was derived based on the EESI-MS response of 1-oxyl-2-phenyl-4, 4, 5, 5-tetramethylimidazoline (PTI) product. The method allowed quantification of eNO below ppb level (~0.02 ppbv) with a relative standard deviation (RSD) of 11.6%. In addition, eNO levels of 20 volunteers were monitored by EESI-MS over the time period of 10 hrs. Long-term eNO response to smoking a cigarette was recorded, and the observed time-dependent profile was discussed. This work extends the application of EESI-MS to small molecules (<30 Da) with low proton affinity and collision-induced dissociation efficiency, which are usually poorly visible by conventional ion trap mass spectrometers. Long-term quantitative profiling of eNO by EESI-MS opens new possibilities for the research of human metabolism and clinical diagnosis.

  20. A paper/polymer hybrid microfluidic microplate for rapid quantitative detection of multiple disease biomarkers

    NASA Astrophysics Data System (ADS)

    Sanjay, Sharma T.; Dou, Maowei; Sun, Jianjun; Li, Xiujun

    2016-07-01

    Enzyme linked immunosorbent assay (ELISA) is one of the most widely used laboratory disease diagnosis methods. However, performing ELISA in low-resource settings is limited by long incubation time, large volumes of precious reagents, and well-equipped laboratories. Herein, we developed a simple, miniaturized paper/PMMA (poly(methyl methacrylate)) hybrid microfluidic microplate for low-cost, high throughput, and point-of-care (POC) infectious disease diagnosis. The novel use of porous paper in flow-through microwells facilitates rapid antibody/antigen immobilization and efficient washing, avoiding complicated surface modifications. The top reagent delivery channels can simply transfer reagents to multiple microwells thus avoiding repeated manual pipetting and costly robots. Results of colorimetric ELISA can be observed within an hour by the naked eye. Quantitative analysis was achieved by calculating the brightness of images scanned by an office scanner. Immunoglobulin G (IgG) and Hepatitis B surface Antigen (HBsAg) were quantitatively analyzed with good reliability in human serum samples. Without using any specialized equipment, the limits of detection of 1.6 ng/mL for IgG and 1.3 ng/mL for HBsAg were achieved, which were comparable to commercial ELISA kits using specialized equipment. We envisage that this simple POC hybrid microplate can have broad applications in various bioassays, especially in resource-limited settings.

  1. Quantitative Phase Fraction Detection in Organic Photovoltaic Materials through EELS Imaging

    SciTech Connect

    Dyck, Ondrej; Hu, Sheng; Das, Sanjib; Keum, Jong; Xiao, Kai; Khomami, Bamin; Duscher, Gerd

    2015-11-24

    Organic photovoltaic materials have recently seen intense interest from the research community. Improvements in device performance are occurring at an impressive rate; however, visualization of the active layer phase separation still remains a challenge. Our paper outlines the application of two electron energy-loss spectroscopic (EELS) imaging techniques that can complement and enhance current phase detection techniques. Specifically, the bulk plasmon peak position, often used to produce contrast between phases in energy filtered transmission electron microscopy (EFTEM), is quantitatively mapped across a sample cross section. One complementary spectrum image capturing the carbon and sulfur core loss edges is compared with the plasmon peak map and found to agree quite well, indicating that carbon and sulfur density differences between the two phases also allows phase discrimination. Additionally, an analytical technique for determining absolute atomic areal density is used to produce an absolute carbon and sulfur areal density map. We also show how these maps may be re-interpreted as a phase ratio map, giving quantitative information about the purity of the phases within the junction.

  2. On-chip quantitative detection of pathogen genes by autonomous microfluidic PCR platform.

    PubMed

    Tachibana, Hiroaki; Saito, Masato; Shibuya, Shogo; Tsuji, Koji; Miyagawa, Nobuyuki; Yamanaka, Keiichiro; Tamiya, Eiichi

    2015-12-15

    Polymerase chain reaction (PCR)-based genetic testing has become a routine part of clinical diagnoses and food testing. In these fields, rapid, easy-to-use, and cost-efficient PCR chips are expected to be appeared for providing such testing on-site. In this study, a new autonomous disposable plastic microfluidic PCR chip was created, and was utilized for quantitative detection of pathogenic microorganisms. To control the capillary flow of the following solution in the PCR microchannel, a driving microchannel was newly designed behind the PCR microchannel. This allowed the effective PCR by simply dropping the PCR solution onto the inlet without any external pumps. In order to achieve disposability, injection-molded cyclo-olefin polymer (COP) of a cost-competitive plastic was used for the PCR chip. We discovered that coating the microchannel walls with non-ionic surfactant produced a suitable hydrophilic surface for driving the capillary flow through the 1250-mm long microchannel. As a result, quantitative real-time PCR with the lowest initial concentration of human, Escherichia coli (E. coli), and pathogenic E. coli O157 genomic DNA of 4, 0.0019, 0.031 pg/μl, respectively, was successfully achieved in less than 18 min. Our results indicate that the platform presented in this study provided a rapid, easy-to-use, and low-cost real-time PCR system that could be potentially used for on-site gene testing. PMID:26210470

  3. Detection of Differential Item Functioning Using the Lasso Approach

    ERIC Educational Resources Information Center

    Magis, David; Tuerlinckx, Francis; De Boeck, Paul

    2015-01-01

    This article proposes a novel approach to detect differential item functioning (DIF) among dichotomously scored items. Unlike standard DIF methods that perform an item-by-item analysis, we propose the "LR lasso DIF method": logistic regression (LR) model is formulated for all item responses. The model contains item-specific intercepts,…

  4. Detection of Turner Syndrome by quantitative PCR of SHOX and VAMP7 genes.

    PubMed

    Ibarra-Ramírez, Marisol; Zamudio-Osuna, Michelle Jesús; Campos-Acevedo, Luis Daniel; Gallardo-Blanco, Hugo Leonid; Cerda-Flores, Ricardo Martin; Rodríguez-Sánchez, Irám Pablo; Martínez-de-Villarreal, Laura Elia

    2015-02-01

    Turner Syndrome (TS) is an unfavorable genetic condition with a prevalence of 1:2500 in newborn girls. Prompt and effective diagnosis is very important to appropriately monitor the comorbidities. The aim of the present study was to propose a feasible and practical molecular diagnostic tool for newborn screening by quantifying the gene dosage of the SHOX, VAMP7, XIST, UBA1, and SRY genes by quantitative polymerase chain reaction (qPCR) in individuals with a diagnosis of complete X monosomy, as well as those with TS variants, and then compare the results to controls without chromosomal abnormalities. According to our results, the most useful markers for these chromosomal variants were the genes found in the pseudoautosomic regions 1 and 2 (PAR1 and PAR2), because differences in gene dosage (relative quantification) between groups were more evident in SHOX and VAMP7 gene expression. Therefore, we conclude that these markers are useful for early detection in aneuploidies involving sex chromosomes.

  5. Quantitative polymerase chain reaction (PCR) for detection of aquatic animal pathogens in a diagnostic laboratory setting

    USGS Publications Warehouse

    Purcell, Maureen K.; Getchell, Rodman G.; McClure, Carol A.; Weber, S.E.; Garver, Kyle A.

    2011-01-01

    Real-time, or quantitative, polymerase chain reaction (qPCR) is quickly supplanting other molecular methods for detecting the nucleic acids of human and other animal pathogens owing to the speed and robustness of the technology. As the aquatic animal health community moves toward implementing national diagnostic testing schemes, it will need to evaluate how qPCR technology should be employed. This review outlines the basic principles of qPCR technology, considerations for assay development, standards and controls, assay performance, diagnostic validation, implementation in the diagnostic laboratory, and quality assurance and control measures. These factors are fundamental for ensuring the validity of qPCR assay results obtained in the diagnostic laboratory setting.

  6. Quantitative analysis of serum methohexital by GLC using capillary column and nitrogen-selective detection.

    PubMed

    Le Normand, Y; De Villepoix, C; Athouel, A; Kergueris, M F; Bourin, M; Larousse, C

    1988-01-01

    A gas chromatographic method for routine quantitation of methohexital in plasma samples is reported. One-step extraction in organic phase, the use of a fused silica capillary column, and nitrogen-selective detection permit simple, precise, and sensitive determination of methohexital in plasma. A linear relationship is described between peak height ratio and methohexital concentrations ranging from 0.125 to 50.0 micrograms/ml (r = 0.998). The sensitivity limit of the assay was 6 ng/ml in plasma. No interfering peak was observed with numerous other drugs. The procedure was successfully applied to the determination of pharmacokinetic parameters of methohexital after IV administration or continuous infusion in a child and an adult.

  7. Quantitative carbon detector for enhanced detection of molecules in foods, pharmaceuticals, cosmetics, flavors, and fuels.

    PubMed

    Beach, Connor A; Krumm, Christoph; Spanjers, Charles S; Maduskar, Saurabh; Jones, Andrew J; Dauenhauer, Paul J

    2016-03-01

    Analysis of trace compounds, such as pesticides and other contaminants, within consumer products, fuels, and the environment requires quantification of increasingly complex mixtures of difficult-to-quantify compounds. Many compounds of interest are non-volatile and exhibit poor response in current gas chromatography and flame ionization systems. Here we show the reaction of trimethylsilylated chemical analytes to methane using a quantitative carbon detector (QCD; the Polyarc™ reactor) within a gas chromatograph (GC), thereby enabling enhanced detection (up to 10×) of highly functionalized compounds including carbohydrates, acids, drugs, flavorants, and pesticides. Analysis of a complex mixture of compounds shows that the GC-QCD method exhibits faster and more accurate analysis of complex mixtures commonly encountered in everyday products and the environment. PMID:26842653

  8. Pedestrian detection from thermal images: A sparse representation based approach

    NASA Astrophysics Data System (ADS)

    Qi, Bin; John, Vijay; Liu, Zheng; Mita, Seiichi

    2016-05-01

    Pedestrian detection, a key technology in computer vision, plays a paramount role in the applications of advanced driver assistant systems (ADASs) and autonomous vehicles. The objective of pedestrian detection is to identify and locate people in a dynamic environment so that accidents can be avoided. With significant variations introduced by illumination, occlusion, articulated pose, and complex background, pedestrian detection is a challenging task for visual perception. Different from visible images, thermal images are captured and presented with intensity maps based objects' emissivity, and thus have an enhanced spectral range to make human beings perceptible from the cool background. In this study, a sparse representation based approach is proposed for pedestrian detection from thermal images. We first adopted the histogram of sparse code to represent image features and then detect pedestrian with the extracted features in an unimodal and a multimodal framework respectively. In the unimodal framework, two types of dictionaries, i.e. joint dictionary and individual dictionary, are built by learning from prepared training samples. In the multimodal framework, a weighted fusion scheme is proposed to further highlight the contributions from features with higher separability. To validate the proposed approach, experiments were conducted to compare with three widely used features: Haar wavelets (HWs), histogram of oriented gradients (HOG), and histogram of phase congruency (HPC) as well as two classification methods, i.e. AdaBoost and support vector machine (SVM). Experimental results on a publicly available data set demonstrate the superiority of the proposed approach.

  9. Multistage optical smoke detection approach for smoke alarm systems

    NASA Astrophysics Data System (ADS)

    Nguyen, Truc Kim Thi; Kim, Jong-Myon

    2013-05-01

    We propose a novel multistage smoke detection algorithm based on inherent optical characteristics such as diffusion, color, and texture of smoke. Moving regions in a video frame are detected by an approximate median background subtraction method using the diffusion behavior of smoke. These moving regions are segmented by a fuzzy C-means (FCM) clustering algorithm that uses the hue and saturation components of moving pixels in the hue-saturation-intensity color space. A decision rule is used to select candidate smoke regions from smoke-colored FCM clusters. An object tracking approach is employed in the candidate smoke region to detect candidate smoke objects in the video frame, and image texture parameters are extracted from these objects using a gray level co-occurrence matrix (GLCM). The thirteen GLCM features are selected to constitute the feature vector by applying principal components analysis, resulting in high-accuracy smoke detection. Finally, a back propagation neural network is utilized as a classifier to discriminate smoke and nonsmoke using the selected feature vector. Experimental results using a standard experimental dataset of video clips demonstrate that the proposed approach outperforms state-of-the-art smoke detection approaches in terms of accuracy, making real-life implementation feasible.

  10. Accuracy, precision, and method detection limits of quantitative PCR for airborne bacteria and fungi.

    PubMed

    Hospodsky, Denina; Yamamoto, Naomichi; Peccia, Jordan

    2010-11-01

    Real-time quantitative PCR (qPCR) for rapid and specific enumeration of microbial agents is finding increased use in aerosol science. The goal of this study was to determine qPCR accuracy, precision, and method detection limits (MDLs) within the context of indoor and ambient aerosol samples. Escherichia coli and Bacillus atrophaeus vegetative bacterial cells and Aspergillus fumigatus fungal spores loaded onto aerosol filters were considered. Efficiencies associated with recovery of DNA from aerosol filters were low, and excluding these efficiencies in quantitative analysis led to underestimating the true aerosol concentration by 10 to 24 times. Precision near detection limits ranged from a 28% to 79% coefficient of variation (COV) for the three test organisms, and the majority of this variation was due to instrument repeatability. Depending on the organism and sampling filter material, precision results suggest that qPCR is useful for determining dissimilarity between two samples only if the true differences are greater than 1.3 to 3.2 times (95% confidence level at n = 7 replicates). For MDLs, qPCR was able to produce a positive response with 99% confidence from the DNA of five B. atrophaeus cells and less than one A. fumigatus spore. Overall MDL values that included sample processing efficiencies ranged from 2,000 to 3,000 B. atrophaeus cells per filter and 10 to 25 A. fumigatus spores per filter. Applying the concepts of accuracy, precision, and MDL to qPCR aerosol measurements demonstrates that sample processing efficiencies must be accounted for in order to accurately estimate bioaerosol exposure, provides guidance on the necessary statistical rigor required to understand significant differences among separate aerosol samples, and prevents undetected (i.e., nonquantifiable) values for true aerosol concentrations that may be significant.

  11. A new approach for DNA detection by SERRS.

    PubMed

    Faulds, Karen; Fruk, Ljiljana; Robson, David C; Thompson, David G; Enright, Alexis; Smith, W Ewen; Graham, Duncan

    2006-01-01

    A new approach for the detection of DNA using surface enhance resonance Raman scattering (SERRS) is reported. The majority of existing techniques use fluorescence spectroscopy with advanced probe design to provide information on the identity of specific DNA sequences down to single base resolution. A new approach to the labelling of DNA is discussed which uses Michael addition to couple thiolated DNA to dye labels specifically designed to attach to silver surfaces. When combined with existing strategies for sensitive detection of DNA using commercially available labels, a new class of biomolecular probe known as a SERRS Beacon was produced. The detection techniques of fluorescence and surface enhanced resonance Raman scattering (SERRS) are combined to give a sensitive and selective system for use in the development and creation of novel assays for specifically defined targets. It demonstrates improved potential for multiplexing analysis. PMID:16833121

  12. A new approach for DNA detection by SERRS.

    PubMed

    Faulds, Karen; Fruk, Ljiljana; Robson, David C; Thompson, David G; Enright, Alexis; Smith, W Ewen; Graham, Duncan

    2006-01-01

    A new approach for the detection of DNA using surface enhance resonance Raman scattering (SERRS) is reported. The majority of existing techniques use fluorescence spectroscopy with advanced probe design to provide information on the identity of specific DNA sequences down to single base resolution. A new approach to the labelling of DNA is discussed which uses Michael addition to couple thiolated DNA to dye labels specifically designed to attach to silver surfaces. When combined with existing strategies for sensitive detection of DNA using commercially available labels, a new class of biomolecular probe known as a SERRS Beacon was produced. The detection techniques of fluorescence and surface enhanced resonance Raman scattering (SERRS) are combined to give a sensitive and selective system for use in the development and creation of novel assays for specifically defined targets. It demonstrates improved potential for multiplexing analysis.

  13. Rapid detection and quantitation of Bluetongue virus (BTV) using a Molecular Beacon fluorescent probe assay.

    PubMed

    Orrù, Germano; Ferrando, Maria Laura; Meloni, Mauro; Liciardi, Manuele; Savini, Giovanni; De Santis, Paola

    2006-10-01

    Bluetongue virus (BTV) is the causative agent of Bluetongue (BT) disease in ruminant livestock and occurs almost worldwide between latitudes 35 degrees S and 50 degrees N; 24 serotypes of BTV are known of which 8 circulate periodically within parts of the Mediterranean Region. A fast (about 3.5 h) and versatile diagnostic procedure able to detect and quantify BTV-RNA, has been developed using a Molecular Beacon (MB) fluorescent probe; PCR primers were designed to target 91 bp within the NS3 conserved region of the viral RNA segment 10 (S10) and bracketed the MB fluorescence probe hybridisation site. The MB fluorescent probe was used to develop two Bluetongue serogroup-specific assays: a quantitative real time reverse transcriptase polymerase chain reaction (RT-PCR) and a traditional RT-PCR. These were tested using BTV-RNAs extracted from the blood and organs of BT-affected animals, and from virus isolate suspensions. The samples included ten serotypes (BTV-1-BTV-9 and BTV-16); of these, BTV serotypes -1, -2, -4, -9 and -16 have since 1998 been involved in the extensive outbreaks of BT across the Mediterranean Region. To evaluate the specificity and sensitivity of the MB probe, all positive samples (and negative controls) were tested using the developed quantitative real time RT-PCR and traditional RT-PCR assays. The former test had a detection limit of 10(3) cDNA molecules per reaction with a log-linear quantification range of up to 10(11) (R2 = 0.98), while the latter test was able to detect 500 cDNA-BTV molecules/PCR. The results show that the MB fluorescent probe is both rapid and versatile for the laboratory diagnosis of Bluetongue and for quantifying levels of viraemia in BTV-affected animals. An "in silico" comparison of the primers and MB fluorescent probe used in this study showed that it is possible to detect all 24 serotypes of BTV.

  14. The ghosts of departed quantities: approaches to dealing with observations below the limit of quantitation.

    PubMed

    Senn, Stephen; Holford, Nick; Hockey, Hans

    2012-12-30

    A common but not necessarily logical requirement in drug development is that a 'limit of quantitation' be set for chemical assays and that observations that fall below the limit should not be treated as real data but should be labelled as below the limit and set aside for special treatment. We examine five of seven approaches to analysing such data considered by Beal in 2001, concentrating in particular on two: one that treats the data as a truncated sample and another that treats them as a censored sample. In fact, using a pattern-mixture framework, one can show that the former consists of using the conditional distribution of the 'acceptable values' and the latter adds the information from the marginal mixing distribution. We illustrate these approaches with a real example, concentrating in particular on the two likelihood-based methods, provide various formulae that may be used to compare these and other approaches, check these formulae using simulations and make some recommendations as to which approach one should use. PMID:22825800

  15. Information fusion approach for detection of brain structures in MRI

    NASA Astrophysics Data System (ADS)

    Shademan, Azad; Soltanian-Zadeh, Hamid

    2002-05-01

    This paper presents an information fusion approach for automatic detection of mid-brain nuclei (caudate, putamen, globus pallidus, and thalamus) from MRI. The method is based on fusion of anatomical information, obtained from brain atlases and expert physicians, into MRI numerical information within a fuzzy framework, employed to model intrinsic uncertainty of problem. First step of this method is segmentation of brain tissues (gray matter, white matter, and cerebrospinal fluid). Physical landmarks such as inter-hemispheric plane alongside numerical information from segmentation step are then used to describe the nuclei. Each nucleus is defined according to a unique description according to physical landmarks and anatomical landmarks, most of which are the previously detected nuclei. Also, a detected nucleus in slice n serves as key landmark to detect same nucleus in slice n+1. These steps construct fuzzy decision maps. Overall decision is made after fusing all of decisions according to a fusion operator. This approach has been implemented to detect caudate, putamen, and thalamus from a sequence of axial T1-weighted brain MRI's. Our experience shows that final nuclei detection results are highly dependent upon primary tissue segmentation. The method is validated by comparing resultant nuclei volumes with those obtained using manual segmentation performed by expert physicians.

  16. Single-molecule tracing on a fluidic microchip for quantitative detection of low-abundance nucleic acids.

    PubMed

    Wang, Tza-Huei; Peng, Yahui; Zhang, Chunyang; Wong, Pak Kin; Ho, Chih-Ming

    2005-04-20

    Here, we report a method capable of quantitative detection of low-abundance DNA/RNA molecules by incorporating confocal fluorescence spectroscopy, molecular beacons, and a molecular-confinement microfluidic reactor. By using a combination of ac and dc fields via a trio of 3-D electrodes in the microreactor, we are able to precisely direct the transport of individual molecules to a minuscule laser-focused detection volume ( approximately 1 fL). A burst of fluorescence photons is detected whenever a molecular beacon-target hybrid flows through the detection region, and the amount of targets can be directly quantified according to the number of recorded single-molecule flow-through events. This assay consumes only attomoles of molecular probes and is able to quantitatively detect subpicomolar DNA targets. A measurement time of less than 2 min is sufficient to complete the detection.

  17. Detection of quantitative trait loci for growth and carcass composition in cattle.

    PubMed

    Casas, E; Shackelford, S D; Keele, J W; Koohmaraie, M; Smith, T P L; Stone, R T

    2003-12-01

    The objective of the present study was to detect quantitative trait loci for economically important traits in a family from a Bos indicus x Bos taurus sire. A Brahman x Hereford sire was used to develop a half-sib family (n = 547). The sire was mated to Bos taurus cows. Traits analyzed were birth (kg) and weaning weights (kg); hot carcass weight (kg); marbling score; longissimus area (cm2); USDA yield grade; estimated kidney, pelvic, and heart fat (%); fat thickness (cm); fat yield (%); and retail product yield (%). Meat tenderness was measured as Warner-Bratzler shear force (kg) at 3 and 14 d postmortem. Two hundred and thirty-eight markers were genotyped in 185 offspring. One hundred and thirty markers were used to genotype the remaining 362 offspring. A total of 312 markers were used in the final analysis. Seventy-four markers were common to both groups. Significant QTL (expected number of false-positives < 0.05) were observed for birth weight and longissimus area on chromosome 5, for longissimus area on chromosome 6, for retail product yield on chromosome 9, for birth weight on chromosome 21, and for marbling score on chromosome 23. Evidence suggesting (expected number of false-positives < 1) the presence of QTL was detected for several traits. Putative QTL for birth weight were detected on chromosomes 1, 2, and 3, and for weaning weight on chromosome 29. For hot carcass weight, QTL were detected on chromosomes 10, 18, and 29. Four QTL for yield grade were identified on chromosomes 2, 11, 14, and 19. Three QTL for fat thickness were detected on chromosomes 2, 3, 7, and 14. For marbling score, QTL were identified on chromosomes 3, 10, 14, and 27. Four QTL were identified for retail product yield on chromosomes 12, 18, 19, and 29. A QTL for estimated kidney, pelvic, and heart fat was detected on chromosome 15, and a QTL for meat tenderness measured as Warner-Bratzler shear force at 3 d postmortem was identified on chromosome 20. Two QTL were detected for meat

  18. Detection of quantitative trait loci for growth and carcass composition in cattle.

    PubMed

    Casas, E; Shackelford, S D; Keele, J W; Koohmaraie, M; Smith, T P L; Stone, R T

    2003-12-01

    The objective of the present study was to detect quantitative trait loci for economically important traits in a family from a Bos indicus x Bos taurus sire. A Brahman x Hereford sire was used to develop a half-sib family (n = 547). The sire was mated to Bos taurus cows. Traits analyzed were birth (kg) and weaning weights (kg); hot carcass weight (kg); marbling score; longissimus area (cm2); USDA yield grade; estimated kidney, pelvic, and heart fat (%); fat thickness (cm); fat yield (%); and retail product yield (%). Meat tenderness was measured as Warner-Bratzler shear force (kg) at 3 and 14 d postmortem. Two hundred and thirty-eight markers were genotyped in 185 offspring. One hundred and thirty markers were used to genotype the remaining 362 offspring. A total of 312 markers were used in the final analysis. Seventy-four markers were common to both groups. Significant QTL (expected number of false-positives < 0.05) were observed for birth weight and longissimus area on chromosome 5, for longissimus area on chromosome 6, for retail product yield on chromosome 9, for birth weight on chromosome 21, and for marbling score on chromosome 23. Evidence suggesting (expected number of false-positives < 1) the presence of QTL was detected for several traits. Putative QTL for birth weight were detected on chromosomes 1, 2, and 3, and for weaning weight on chromosome 29. For hot carcass weight, QTL were detected on chromosomes 10, 18, and 29. Four QTL for yield grade were identified on chromosomes 2, 11, 14, and 19. Three QTL for fat thickness were detected on chromosomes 2, 3, 7, and 14. For marbling score, QTL were identified on chromosomes 3, 10, 14, and 27. Four QTL were identified for retail product yield on chromosomes 12, 18, 19, and 29. A QTL for estimated kidney, pelvic, and heart fat was detected on chromosome 15, and a QTL for meat tenderness measured as Warner-Bratzler shear force at 3 d postmortem was identified on chromosome 20. Two QTL were detected for meat

  19. Detecting Long-Range Enhancer-Promoter Interactions by Quantitative Chromosome Conformation Capture.

    PubMed

    Deng, Wulan; Blobel, Gerd A

    2017-01-01

    Chromosome conformation capture (3C) technology and its derivatives are currently the primary methodologies measuring contacts among genomic elements. In fact, the lion share of what is currently known about chromosome folding is based on 3C-related approaches. For example, distal enhancers are commonly in physically proximity with their target genes, forming chromatin loops. Additional layers of chromatin organization have been described using 3C-based techniques, including topological domains (TADs) and sub-TADs. Finally, inter-chromosomal interactions have been reported although they are much less frequent. 3C is becoming increasingly widespread in its use for understanding genome organization. Here we provide a protocol for quantitative 3C using real-time PCR analysis, along with essential quality controls and normalization methods. PMID:27662870

  20. Quantitative Assessment of Detection Frequency for the INL Ambient Air Monitoring Network

    SciTech Connect

    A. Jeffrey Sondrup; Arthur S. Rood

    2014-11-01

    A quantitative assessment of the Idaho National Laboratory (INL) air monitoring network was performed using frequency of detection as the performance metric. The INL air monitoring network consists of 37 low-volume air samplers in 31 different locations. Twenty of the samplers are located on INL (onsite) and 17 are located off INL (offsite). Detection frequencies were calculated using both BEA and ESER laboratory minimum detectable activity (MDA) levels. The CALPUFF Lagrangian puff dispersion model, coupled with 1 year of meteorological data, was used to calculate time-integrated concentrations at sampler locations for a 1-hour release of unit activity (1 Ci) for every hour of the year. The unit-activity time-integrated concentration (TICu) values were calculated at all samplers for releases from eight INL facilities. The TICu values were then scaled and integrated for a given release quantity and release duration. All facilities modeled a ground-level release emanating either from the center of the facility or at a point where significant emissions are possible. In addition to ground-level releases, three existing stacks at the Advanced Test Reactor Complex, Idaho Nuclear Technology and Engineering Center, and Material and Fuels Complex were also modeled. Meteorological data from the 35 stations comprising the INL Mesonet network, data from the Idaho Falls Regional airport, upper air data from the Boise airport, and three-dimensional gridded data from the weather research forecasting model were used for modeling. Three representative radionuclides identified as key radionuclides in INL’s annual National Emission Standards for Hazardous Air Pollutants evaluations were considered for the frequency of detection analysis: Cs-137 (beta-gamma emitter), Pu-239 (alpha emitter), and Sr-90 (beta emitter). Source-specific release quantities were calculated for each radionuclide, such that the maximum inhalation dose at any publicly accessible sampler or the National

  1. Bayesian approach to the detection problem in gravitational wave astronomy

    SciTech Connect

    Littenberg, Tyson B.; Cornish, Neil J.

    2009-09-15

    The analysis of data from gravitational wave detectors can be divided into three phases: search, characterization, and evaluation. The evaluation of the detection--determining whether a candidate event is astrophysical in origin or some artifact created by instrument noise--is a crucial step in the analysis. The ongoing analyses of data from ground-based detectors employ a frequentist approach to the detection problem. A detection statistic is chosen, for which background levels and detection efficiencies are estimated from Monte Carlo studies. This approach frames the detection problem in terms of an infinite collection of trials, with the actual measurement corresponding to some realization of this hypothetical set. Here we explore an alternative, Bayesian approach to the detection problem, that considers prior information and the actual data in hand. Our particular focus is on the computational techniques used to implement the Bayesian analysis. We find that the parallel tempered Markov chain Monte Carlo (PTMCMC) algorithm is able to address all three phases of the analysis in a coherent framework. The signals are found by locating the posterior modes, the model parameters are characterized by mapping out the joint posterior distribution, and finally, the model evidence is computed by thermodynamic integration. As a demonstration, we consider the detection problem of selecting between models describing the data as instrument noise, or instrument noise plus the signal from a single compact galactic binary. The evidence ratios, or Bayes factors, computed by the PTMCMC algorithm are found to be in close agreement with those computed using a reversible jump Markov chain Monte Carlo algorithm.

  2. Determination of Detection Limits and Quantitation Limits for Compounds in a Database of GC/MS by FUMI Theory

    PubMed Central

    Nakashima, Shinya; Hayashi, Yuzuru

    2016-01-01

    The aim of this paper is to propose a stochastic method for estimating the detection limits (DLs) and quantitation limits (QLs) of compounds registered in a database of a GC/MS system and prove its validity with experiments. The approach described in ISO 11843 Part 7 is adopted here as an estimation means of DL and QL, and the decafluorotriphenylphosphine (DFTPP) tuning and retention time locking are carried out for adjusting the system. Coupled with the data obtained from the system adjustment experiments, the information (noise and signal of chromatograms and calibration curves) stored in the database is used for the stochastic estimation, dispensing with the repetition measurements. Of sixty-six pesticides, the DL values obtained by the ISO method were compared with those from the statistical approach and the correlation between them was observed to be excellent with the correlation coefficient of 0.865. The accuracy of the method proposed was also examined and concluded to be satisfactory as well. The samples used are commercial products of pesticides mixtures and the uncertainty from sample preparation processes is not taken into account. PMID:27162706

  3. Current and New Approaches in GMO Detection: Challenges and Solutions

    PubMed Central

    Fraiture, Marie-Alice; Herman, Philippe; Taverniers, Isabel; De Loose, Marc; Deforce, Dieter; Roosens, Nancy H.

    2015-01-01

    In many countries, genetically modified organisms (GMO) legislations have been established in order to guarantee the traceability of food/feed products on the market and to protect the consumer freedom of choice. Therefore, several GMO detection strategies, mainly based on DNA, have been developed to implement these legislations. Due to its numerous advantages, the quantitative PCR (qPCR) is the method of choice for the enforcement laboratories in GMO routine analysis. However, given the increasing number and diversity of GMO developed and put on the market around the world, some technical hurdles could be encountered with the qPCR technology, mainly owing to its inherent properties. To address these challenges, alternative GMO detection methods have been developed, allowing faster detections of single GM target (e.g., loop-mediated isothermal amplification), simultaneous detections of multiple GM targets (e.g., PCR capillary gel electrophoresis, microarray, and Luminex), more accurate quantification of GM targets (e.g., digital PCR), or characterization of partially known (e.g., DNA walking and Next Generation Sequencing (NGS)) or unknown (e.g., NGS) GMO. The benefits and drawbacks of these methods are discussed in this review. PMID:26550567

  4. Current and new approaches in GMO detection: challenges and solutions.

    PubMed

    Fraiture, Marie-Alice; Herman, Philippe; Taverniers, Isabel; De Loose, Marc; Deforce, Dieter; Roosens, Nancy H

    2015-01-01

    In many countries, genetically modified organisms (GMO) legislations have been established in order to guarantee the traceability of food/feed products on the market and to protect the consumer freedom of choice. Therefore, several GMO detection strategies, mainly based on DNA, have been developed to implement these legislations. Due to its numerous advantages, the quantitative PCR (qPCR) is the method of choice for the enforcement laboratories in GMO routine analysis. However, given the increasing number and diversity of GMO developed and put on the market around the world, some technical hurdles could be encountered with the qPCR technology, mainly owing to its inherent properties. To address these challenges, alternative GMO detection methods have been developed, allowing faster detections of single GM target (e.g., loop-mediated isothermal amplification), simultaneous detections of multiple GM targets (e.g., PCR capillary gel electrophoresis, microarray, and Luminex), more accurate quantification of GM targets (e.g., digital PCR), or characterization of partially known (e.g., DNA walking and Next Generation Sequencing (NGS)) or unknown (e.g., NGS) GMO. The benefits and drawbacks of these methods are discussed in this review. PMID:26550567

  5. Current and new approaches in GMO detection: challenges and solutions.

    PubMed

    Fraiture, Marie-Alice; Herman, Philippe; Taverniers, Isabel; De Loose, Marc; Deforce, Dieter; Roosens, Nancy H

    2015-01-01

    In many countries, genetically modified organisms (GMO) legislations have been established in order to guarantee the traceability of food/feed products on the market and to protect the consumer freedom of choice. Therefore, several GMO detection strategies, mainly based on DNA, have been developed to implement these legislations. Due to its numerous advantages, the quantitative PCR (qPCR) is the method of choice for the enforcement laboratories in GMO routine analysis. However, given the increasing number and diversity of GMO developed and put on the market around the world, some technical hurdles could be encountered with the qPCR technology, mainly owing to its inherent properties. To address these challenges, alternative GMO detection methods have been developed, allowing faster detections of single GM target (e.g., loop-mediated isothermal amplification), simultaneous detections of multiple GM targets (e.g., PCR capillary gel electrophoresis, microarray, and Luminex), more accurate quantification of GM targets (e.g., digital PCR), or characterization of partially known (e.g., DNA walking and Next Generation Sequencing (NGS)) or unknown (e.g., NGS) GMO. The benefits and drawbacks of these methods are discussed in this review.

  6. Clinical anatomy of the suprarenal arteries: a quantitative approach by aortography.

    PubMed

    Toni, R; Mosca, S; Favero, L; Ricci, S; Roversi, R; Toni, G; Vezzadini, P

    1988-01-01

    The angiographic visualization, arterial origin and mean diameter per age group (20-40, 41-60 years) of the suprarenal arterial vessels have been quantitatively investigated by aortography in 100 patients without suprarenal disease. Visualization of the various arteries was achieved in a percentage substantially comparable to the anatomic data of the literature, though with lower detection of the superior suprarenal vessels. A variable site of origin was present particularly for the superior and middle suprarenal vessels compared to the inferior suprarenal arteries and possible embryological reasons and clinical implications are discussed. Statistically significant differences were found in the mean diameter of each arterial vessel in all the subjects examined, thus substantiating the concept of differential arterial supply to various portions of the gland. Age-related changes were demonstrated in the right middle suprarenal artery, suggesting a predominant role of this vessel in the physiological adaptation of the blood supply to the gland with increasing age.

  7. A smartphone-readable barcode assay for the detection and quantitation of pesticide residues.

    PubMed

    Guo, Juan; Wong, Jessica X H; Cui, Caie; Li, Xiaochun; Yu, Hua-Zhong

    2015-08-21

    In this paper, we present a smartphone-readable barcode assay for the qualitative detection of methyl parathion residues, a toxic organophosphorus pesticide that is popularly used in agriculture worldwide. The detection principle is based on the irreversible inhibition of the enzymatic activity of acetylcholinesterase (AchE) by methyl parathion; AchE catalytically hydrolyzes acetylthiocholine iodine to thiocholine that in turn dissociates dithiobis-nitrobenzoate to produce a yellow product (deprotonated thio-nitrobenzoate). The yellow intensity of the product was confirmed to be inversely dependent on the concentration of the pesticide. We have designed a barcode-formatted assay chip by using a PDMS (polydimethylsiloxane) channel plate (as the reaction reservoir), situated under a printed partial barcode, to complete the whole barcode such that it can be directly read by a barcode scanning app installed on a smartphone. The app is able to qualitatively present the result of the pesticide test; the absence or a low concentration of methyl parathion results in the barcode reading as "-", identifying the test as negative for pesticides. Upon obtaining a positive result (the app reads a "+" character), the captured image can be further analyzed to quantitate the methyl parathion concentration in the sample. Besides the portability and simplicity, this mobile-app based colorimetric barcode assay compares favorably with the standard spectrophotometric method. PMID:26087169

  8. Quantitative lateral flow strip assays as User-Friendly Tools To Detect Biomarker Profiles For Leprosy

    PubMed Central

    van Hooij, Anouk; Tjon Kon Fat, Elisa M.; Richardus, Renate; van den Eeden, Susan J. F.; Wilson, Louis; de Dood, Claudia J.; Faber, Roel; Alam, Korshed; Richardus, Jan Hendrik; Corstjens, Paul L. A. M.; Geluk, Annemieke

    2016-01-01

    Leprosy is a debilitating, infectious disease caused by Mycobacterium leprae. Despite the availability of multidrug therapy, transmission is unremitting. Thus, early identification of M. leprae infection is essential to reduce transmission. The immune response to M. leprae is determined by host genetics, resulting in paucibacillary (PB) and multibacillary (MB) leprosy associated with dominant cellular or humoral immunity, respectively. This spectral pathology of leprosy compels detection of immunity to M. leprae to be based on multiple, diverse biomarkers. In this study we have applied quantitative user friendly lateral flow assays (LFAs) for four immune markers (anti-PGL-I antibodies, IL-10, CCL4 and IP-10) for whole blood samples from a longitudinal BCG vaccination field-trial in Bangladesh. Different biomarker profiles, in contrast to single markers, distinguished M. leprae infected from non-infected test groups, patients from household contacts (HHC) and endemic controls (EC), or MB from PB patients. The test protocol presented in this study merging detection of innate, adaptive cellular as well as humoral immunity, thus provides a convenient tool to measure specific biomarker profiles for M. leprae infection and leprosy utilizing a field-friendly technology. PMID:27682181

  9. Highly sensitive quantitative PCR for the detection and differentiation of Pseudogymnoascus destructans and other Pseudogymnoascus species.

    PubMed

    Shuey, Megan M; Drees, Kevin P; Lindner, Daniel L; Keim, Paul; Foster, Jeffrey T

    2014-03-01

    White-nose syndrome is a fungal disease that has decimated bat populations across eastern North America. Identification of the etiologic agent, Pseudogymnoascus destructans (formerly Geomyces destructans), in environmental samples is essential to proposed management plans. A major challenge is the presence of closely related species, which are ubiquitous in many soils and cave sediments and often present in high abundance. We present a dual-probe real-time quantitative PCR assay capable of detecting and differentiating P. destructans from closely related fungi in environmental samples from North America. The assay, based on a single nucleotide polymorphism (SNP) specific to P. destructans, is capable of rapid low-level detection from various sampling media, including sediment, fecal samples, wing biopsy specimens, and skin swabs. This method is a highly sensitive, high-throughput method for identifying P. destructans, other Pseudogymnoascus spp., and Geomyces spp. in the environment, providing a fundamental component of research and risk assessment for addressing this disease, as well as other ecological and mycological work on related fungi. PMID:24375140

  10. Quantitative prediction of perceptual decisions during near-threshold fear detection

    NASA Astrophysics Data System (ADS)

    Pessoa, Luiz; Padmala, Srikanth

    2005-04-01

    A fundamental goal of cognitive neuroscience is to explain how mental decisions originate from basic neural mechanisms. The goal of the present study was to investigate the neural correlates of perceptual decisions in the context of emotional perception. To probe this question, we investigated how fluctuations in functional MRI (fMRI) signals were correlated with behavioral choice during a near-threshold fear detection task. fMRI signals predicted behavioral choice independently of stimulus properties and task accuracy in a network of brain regions linked to emotional processing: posterior cingulate cortex, medial prefrontal cortex, right inferior frontal gyrus, and left insula. We quantified the link between fMRI signals and behavioral choice in a whole-brain analysis by determining choice probabilities by means of signal-detection theory methods. Our results demonstrate that voxel-wise fMRI signals can reliably predict behavioral choice in a quantitative fashion (choice probabilities ranged from 0.63 to 0.78) at levels comparable to neuronal data. We suggest that the conscious decision that a fearful face has been seen is represented across a network of interconnected brain regions that prepare the organism to appropriately handle emotionally challenging stimuli and that regulate the associated emotional response. decision making | emotion | functional MRI

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

    PubMed

    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.

  12. Feasibility of the grandprogeny design for quantitative trait loci (QTL) detection in purebred beef cattle.

    PubMed

    Moody, D E; Pomp, D; Buchanan, D S

    1997-04-01

    The grandprogeny design (GPD) was developed for dairy cattle to use existing pedigreed populations for quantitative trait locus (QTL) detection. Marker genotypes of grandsires and sons are determined, and trait phenotypic data from grandprogeny are analyzed. The objective of this study was to investigate the potential application of GPD in purebred beef cattle populations. Pedigree structures of Angus (n = 123,319), Hereford (n = 107,778), Brangus (n = 14,449), and Gelbvieh (n = 8,114) sire evaluation reports were analyzed to identify potentially useful families. Power of QTL detection was calculated for a range of QTL effects (.1 to .5 SD) and two Type I error rates (.01 and .001). Reasonable power (> .75) could be achieved using GPD in Angus and Hereford for QTL having moderate effects (.3 SD) on weaning weight and large effects (.4 to .5 SD) on birth, yearling, and maternal weaning weights by genotyping 500 animals. Existing Gelbvieh and Brangus families useful for GPD were limited, and reasonable power could be expected only for QTL having large effects on weaning or birth weights. Although family structures suitable for GPD exist in purebred beef populations, large amounts of genotyping would be required to achieve reasonable power, and only QTL having moderate to large effects could be expected to be identified. PMID:9110205

  13. High-resolution mass spectrometry method for the detection, characterization and quantitation of pharmaceuticals in water.

    PubMed

    Pinhancos, Rebeca; Maass, Sara; Ramanathan, Dil M

    2011-11-01

    The presence of pharmaceuticals in drinking water is an emerging environmental concern. In most environmental testing laboratories, LC-MS/MS assays based on selected reaction monitoring are used as part of a battery of tests used to assure water quality. Although LC-MS/MS continues to be the best tool for detecting pharmaceuticals in water, the combined use of hybrid high-resolution mass spectrometry (HRMS) and ultrahigh pressure liquid chromatography (UHPLC) is starting to become a practical tool to study emerging environmental contaminants. The hybrid LTQ-orbitrap mass spectrometer is suitable for integrated quantitative and qualitative bioanalysis because of the following reasons: (1) the ability to collect full-scan HRMS spectra with scan speeds suitable for UHPLC separations, (2) routine measurement of mass with less than 5 ppm mass accuracy, (3) high mass resolving power, and (4) ability to perform on-the-fly polarity switching in the linear ion trap (LTQ). In the present work, we provide data demonstrating the application of UHPLC-LTQ-orbitrap for the detection, characterization and quantification of pharmaceuticals and their metabolites in drinking water.

  14. A quantitative minimally invasive assay for the detection of metals in the stratum corneum.

    PubMed

    Cullander, C; Jeske, S; Imbert, D; Grant, P G; Bench, G

    2000-03-01

    A quantitative, minimally invasive tape-stripping assay for the detection of metals on and in skin that also has application to the detection of metallic elements on dry surfaces (where human contact could occur) has been developed. This development included construction, using commercial products, of an approximately 25 microm thick, low-metal content tape suitable both for tape-stripping and elemental analysis. Individual tapes were sequentially applied to the skin surface and then removed, taking with them a sample of the dead outer layer of the skin (stratum corneum). Analysis of such tape strip samples by particle induced X-ray emission (PIXE)--a well-characterized, sensitive, analytical technique based on X-ray spectrometry--identified and accurately quantified the metals in the sample. The assay had elemental sensitivities of approximately 1 ng/cm2 for many metals and analysis of elemental contents could be performed in as little as 5 min. The feasibility of the assay for measuring metals in the stratum corneum was demonstrated on the forearms of healthy human volunteers. Samples from approximately half the subjects were found to contain zirconium, possibly arising from the use of roll-on antiperspirants. The assay has potential as a tool: (1) for risk assessment, (2) to identify exposure levels following possible contact with a hazardous metal, and (3) to determine the effectiveness of cleanup or removal measures. PMID:10719909

  15. A novel quantitative approach for evaluating contact mechanics of meniscal replacements.

    PubMed

    Linder-Ganz, E; Elsner, J J; Danino, A; Guilak, F; Shterling, A

    2010-02-01

    One of the functions of the meniscus is to distribute contact forces over the articular surfaces by increasing the joint contact areas. It is widely accepted that total/partial loss of the meniscus increases the risk of joint degeneration. A short-term method for evaluating whether degenerative arthritis can be prevented or not would be to determine if the peak pressure and contact area coverage of the tibial plateau (TP) in the knee are restored at the time of implantation. Although several published studies already utilized TP contact pressure measurements as an indicator for biomechanical performance of allograft menisci, there is a paucity of a quantitative method for evaluation of these parameters in situ with a single effective parameter. In the present study, we developed such a method and used it to assess the load distribution ability of various meniscal implant configurations in human cadaveric knees (n=3). Contact pressures under the intact meniscus were measured under compression (1200 N, 0 deg flexion). Next, total meniscectomy was performed and the protocol was repeated with meniscal implants. Resultant pressure maps were evaluated for the peak pressure value, total contact area, and its distribution pattern, all with respect to the natural meniscus output. Two other measures--implant-dislocation and implant-impingement on the ligaments--were also considered. If any of these occurred, the score was zeroed. The total implant score was based on an adjusted calculation of the aforementioned measures, where the natural meniscus score was always 100. Laboratory experiments demonstrated a good correlation between qualitative and quantitative evaluations of the same pressure map outputs, especially in cases where there were contradicting indications between different parameters. Overall, the proposed approach provides a novel, validated method for quantitative assessment of the biomechanical performance of meniscal implants, which can be used in various

  16. On-site semi-quantitative analysis for ammonium nitrate detection using digital image colourimetry.

    PubMed

    Choodum, Aree; Boonsamran, Pichapat; NicDaeid, Niamh; Wongniramaikul, Worawit

    2015-12-01

    Digital image colourimetry was successfully applied in the semi-quantitative analysis of ammonium nitrate using Griess's test with zinc reduction. A custom-built detection box was developed to enable reproducible lighting of samples, and was used with the built-in webcams of a netbook and an ultrabook for on-site detection. The webcams were used for colour imaging of chemical reaction products in the samples, while the netbook was used for on-site colour analysis. The analytical performance was compared to a commercial external webcam and a digital single-lens reflex (DSLR) camera. The relationship between Red-Green-Blue intensities and ammonium nitrate concentration was investigated. The green channel intensity (IG) was the most sensitive for the pink-violet products from ammonium nitrate that revealed a spectrometric absorption peak at 546 nm. A wide linear range (5 to 250 mgL⁻¹) with a high sensitivity was obtained with the built-in webcam of the ultrabook. A considerably lower detection limit (1.34 ± 0.05mgL⁻¹) was also obtained using the ultrabook, in comparison with the netbook (2.6 ± 0.2 mgL⁻¹), the external web cam (3.4 ± 0.1 mgL⁻¹) and the DSLR (8.0 ± 0.5 mgL⁻¹). The best inter-day precision (over 3 days) was obtained with the external webcam (0.40 to 1.34%RSD), while the netbook and the ultrabook had 0.52 to 3.62% and 1.25 to 4.99% RSDs, respectively. The relative errors were +3.6, +5.6 and -7.1%, on analysing standard ammonium nitrate solutions of known concentration using IG, for the ultrabook, the external webcam, and the netbook, respectively, while the DSLR gave -4.4% relative error. However, the IG of the pink-violet reaction product suffers from interference by soil, so that blank subtraction (|IG-IGblank| or |AG-AGblank|) is recommended for soil sample analysis. This method also gave very good accuracies of -0.11 to -5.61% for spiked soil samples and the results presented for five seized samples showed good correlations between

  17. On-site semi-quantitative analysis for ammonium nitrate detection using digital image colourimetry.

    PubMed

    Choodum, Aree; Boonsamran, Pichapat; NicDaeid, Niamh; Wongniramaikul, Worawit

    2015-12-01

    Digital image colourimetry was successfully applied in the semi-quantitative analysis of ammonium nitrate using Griess's test with zinc reduction. A custom-built detection box was developed to enable reproducible lighting of samples, and was used with the built-in webcams of a netbook and an ultrabook for on-site detection. The webcams were used for colour imaging of chemical reaction products in the samples, while the netbook was used for on-site colour analysis. The analytical performance was compared to a commercial external webcam and a digital single-lens reflex (DSLR) camera. The relationship between Red-Green-Blue intensities and ammonium nitrate concentration was investigated. The green channel intensity (IG) was the most sensitive for the pink-violet products from ammonium nitrate that revealed a spectrometric absorption peak at 546 nm. A wide linear range (5 to 250 mgL⁻¹) with a high sensitivity was obtained with the built-in webcam of the ultrabook. A considerably lower detection limit (1.34 ± 0.05mgL⁻¹) was also obtained using the ultrabook, in comparison with the netbook (2.6 ± 0.2 mgL⁻¹), the external web cam (3.4 ± 0.1 mgL⁻¹) and the DSLR (8.0 ± 0.5 mgL⁻¹). The best inter-day precision (over 3 days) was obtained with the external webcam (0.40 to 1.34%RSD), while the netbook and the ultrabook had 0.52 to 3.62% and 1.25 to 4.99% RSDs, respectively. The relative errors were +3.6, +5.6 and -7.1%, on analysing standard ammonium nitrate solutions of known concentration using IG, for the ultrabook, the external webcam, and the netbook, respectively, while the DSLR gave -4.4% relative error. However, the IG of the pink-violet reaction product suffers from interference by soil, so that blank subtraction (|IG-IGblank| or |AG-AGblank|) is recommended for soil sample analysis. This method also gave very good accuracies of -0.11 to -5.61% for spiked soil samples and the results presented for five seized samples showed good correlations between

  18. A support vector machine approach for detection of microcalcifications.

    PubMed

    El-Naqa, Issam; Yang, Yongyi; Wernick, Miles N; Galatsanos, Nikolas P; Nishikawa, Robert M

    2002-12-01

    In this paper, we investigate an approach based on support vector machines (SVMs) for detection of microcalcification (MC) clusters in digital mammograms, and propose a successive enhancement learning scheme for improved performance. SVM is a machine-learning method, based on the principle of structural risk minimization, which performs well when applied to data outside the training set. We formulate MC detection as a supervised-learning problem and apply SVM to develop the detection algorithm. We use the SVM to detect at each location in the image whether an MC is present or not. We tested the proposed method using a database of 76 clinical mammograms containing 1120 MCs. We use free-response receiver operating characteristic curves to evaluate detection performance, and compare the proposed algorithm with several existing methods. In our experiments, the proposed SVM framework outperformed all the other methods tested. In particular, a sensitivity as high as 94% was achieved by the SVM method at an error rate of one false-positive cluster per image. The ability of SVM to out perform several well-known methods developed for the widely studied problem of MC detection suggests that SVM is a promising technique for object detection in a medical imaging application. PMID:12588039

  19. The power to detect quantitative trait loci using resequenced, experimentally evolved populations of diploid, sexual organisms.

    PubMed

    Baldwin-Brown, James G; Long, Anthony D; Thornton, Kevin R

    2014-04-01

    A novel approach for dissecting complex traits is to experimentally evolve laboratory populations under a controlled environment shift, resequence the resulting populations, and identify single nucleotide polymorphisms (SNPs) and/or genomic regions highly diverged in allele frequency. To better understand the power and localization ability of such an evolve and resequence (E&R) approach, we carried out forward-in-time population genetics simulations of 1 Mb genomic regions under a large combination of experimental conditions, then attempted to detect significantly diverged SNPs. Our analysis indicates that the ability to detect differentiation between populations is primarily affected by selection coefficient, population size, number of replicate populations, and number of founding haplotypes. We estimate that E&R studies can detect and localize causative sites with 80% success or greater when the number of founder haplotypes is over 500, experimental populations are replicated at least 25-fold, population size is at least 1,000 diploid individuals, and the selection coefficient on the locus of interest is at least 0.1. More achievable experimental designs (less replicated, fewer founder haplotypes, smaller effective population size, and smaller selection coefficients) can have power of greater than 50% to identify a handful of SNPs of which one is likely causative. Similarly, in cases where s ≥ 0.2, less demanding experimental designs can yield high power.

  20. [A novel quantitative approach to study dynamic anaerobic process at micro scale].

    PubMed

    Zhang, Zhong-Liang; Wu, Jing; Jiang, Jian-Kai; Jiang, Jie; Li, Huai-Zhi

    2012-11-01

    Anaerobic digestion is attracting more and more interests because of its advantages such as low cost and recovery of clean energy etc. In order to overcome the drawbacks of the existed methods to study the dynamic anaerobic process, a novel microscopical quantitative approach at the granule level was developed combining both the microdevice and the quantitative image analysis techniques. This experiment displayed the process and characteristics of the gas production at static state for the first time and the results indicated that the method was of satisfactory repeatability. The gas production process at static state could be divided into three stages including rapid linear increasing stage, decelerated increasing stage and slow linear increasing stage. The rapid linear increasing stage was long and the biogas rate was high under high initial organic loading rate. The results showed that it was feasible to make the anaerobic process to be carried out in the microdevice; furthermore this novel method was reliable and could clearly display the dynamic process of the anaerobic reaction at the micro scale. The results are helpful to understand the anaerobic process.

  1. The new approach of polarimetric attenuation correction for improving radar quantitative precipitation estimation(QPE)

    NASA Astrophysics Data System (ADS)

    Gu, Ji-Young; Suk, Mi-Kyung; Nam, Kyung-Yeub; Ko, Jeong-Seok; Ryzhkov, Alexander

    2016-04-01

    To obtain high-quality radar quantitative precipitation estimation data, reliable radar calibration and efficient attenuation correction are very important. Because microwave radiation at shorter wavelength experiences strong attenuation in precipitation, accounting for this attenuation is the essential work at shorter wavelength radar. In this study, the performance of different attenuation/differential attenuation correction schemes at C band is tested for two strong rain events which occurred in central Oklahoma. And also, a new attenuation correction scheme (combination of self-consistency and hot-spot concept methodology) that separates relative contributions of strong convective cells and the rest of the storm to the path-integrated total and differential attenuation is among the algorithms explored. A quantitative use of weather radar measurement such as rainfall estimation relies on the reliable attenuation correction. We examined the impact of attenuation correction on estimates of rainfall in heavy rain events by using cross-checking with S-band radar measurements which are much less affected by attenuation and compared the storm rain totals obtained from the corrected Z and KDP and rain gages in these cases. This new approach can be utilized at shorter wavelength radars efficiently. Therefore, it is very useful to Weather Radar Center of Korea Meteorological Administration preparing X-band research dual Pol radar network.

  2. Climate change and dengue: a critical and systematic review of quantitative modelling approaches

    PubMed Central

    2014-01-01

    Background Many studies have found associations between climatic conditions and dengue transmission. However, there is a debate about the future impacts of climate change on dengue transmission. This paper reviewed epidemiological evidence on the relationship between climate and dengue with a focus on quantitative methods for assessing the potential impacts of climate change on global dengue transmission. Methods A literature search was conducted in October 2012, using the electronic databases PubMed, Scopus, ScienceDirect, ProQuest, and Web of Science. The search focused on peer-reviewed journal articles published in English from January 1991 through October 2012. Results Sixteen studies met the inclusion criteria and most studies showed that the transmission of dengue is highly sensitive to climatic conditions, especially temperature, rainfall and relative humidity. Studies on the potential impacts of climate change on dengue indicate increased climatic suitability for transmission and an expansion of the geographic regions at risk during this century. A variety of quantitative modelling approaches were used in the studies. Several key methodological issues and current knowledge gaps were identified through this review. Conclusions It is important to assemble spatio-temporal patterns of dengue transmission compatible with long-term data on climate and other socio-ecological changes and this would advance projections of dengue risks associated with climate change. PMID:24669859

  3. A quantitative approach for hydrological drought characterization in southwestern China using GRACE

    NASA Astrophysics Data System (ADS)

    Chao, Nengfang; Wang, Zhengtao; Jiang, Weiping; Chao, Dingbo

    2016-06-01

    A quantitative approach for hydrological drought characterization, based on non-seasonal water storage deficit data from NASA's Gravity Recovery and Climate Experiment (GRACE) satellite mission, is assessed. Non-seasonal storage deficit is the negative terrestrial water storage after deducting trend, acceleration and seasonal signals, and it is designated as a drought event when it persists for three or more continuous months. The non-seasonal water storage deficit is used for measuring the hydrological drought in southwestern China. It is found that this storage-deficit method clearly identifies hydrological drought onset, end and duration, and quantifies instantaneous severity, peak drought magnitude, and time to recovery. Moreover, it is found that severe droughts have frequently struck southwestern China in the past several decades, among which, the drought of 2011-2012 was the most severe; the duration was 10 months, the severity was -208.92 km3/month, and the time to recovery was 17 months. These results compare well with the National Climate Center of China drought databases, which signifies that the GRACE-based non-seasonal water storage deficit has a quantitative effect on hydrological drought characterization and provides an effective tool for researching droughts.

  4. Detection and segmentation of cell nuclei in virtual microscopy images: a minimum-model approach.

    PubMed

    Wienert, Stephan; Heim, Daniel; Saeger, Kai; Stenzinger, Albrecht; Beil, Michael; Hufnagl, Peter; Dietel, Manfred; Denkert, Carsten; Klauschen, Frederick

    2012-01-01

    Automated image analysis of cells and tissues has been an active research field in medical informatics for decades but has recently attracted increased attention due to developments in computer and microscopy hardware and the awareness that scientific and diagnostic pathology require novel approaches to perform objective quantitative analyses of cellular and tissue specimens. Model-based approaches use a priori information on cell shape features to obtain the segmentation, which may introduce a bias favouring the detection of cell nuclei only with certain properties. In this study we present a novel contour-based "minimum-model" cell detection and segmentation approach that uses minimal a priori information and detects contours independent of their shape. This approach avoids a segmentation bias with respect to shape features and allows for an accurate segmentation (precision = 0.908; recall = 0.859; validation based on ∼8000 manually-labeled cells) of a broad spectrum of normal and disease-related morphological features without the requirement of prior training.

  5. Decoding brain cancer dynamics: a quantitative histogram-based approach using temporal MRI

    NASA Astrophysics Data System (ADS)

    Zhou, Mu; Hall, Lawrence O.; Goldgof, Dmitry B.; Russo, Robin; Gillies, Robert J.; Gatenby, Robert A.

    2015-03-01

    Brain tumor heterogeneity remains a challenge for probing brain cancer evolutionary dynamics. In light of evolution, it is a priority to inspect the cancer system from a time-domain perspective since it explicitly tracks the dynamics of cancer variations. In this paper, we study the problem of exploring brain tumor heterogeneity from temporal clinical magnetic resonance imaging (MRI) data. Our goal is to discover evidence-based knowledge from such temporal imaging data, where multiple clinical MRI scans from Glioblastoma multiforme (GBM) patients are generated during therapy. In particular, we propose a quantitative histogram-based approach that builds a prediction model to measure the difference in histograms obtained from pre- and post-treatment. The study could significantly assist radiologists by providing a metric to identify distinctive patterns within each tumor, which is crucial for the goal of providing patient-specific treatments. We examine the proposed approach for a practical application - clinical survival group prediction. Experimental results show that our approach achieved 90.91% accuracy.

  6. In situ immune infrared fluorescent staining for detection and quantitation of bluetongue virus in Culicoides insect cell culture.

    PubMed

    Mecham, James O; Brown, Philip L; McHolland, Linda E

    2009-06-01

    Bluetongue virus (BTV) is transmitted to sheep, cattle and other ruminants by Culicoides spp. of biting midges. Cell lines have been developed from Culicoides sonorensis; however, techniques to detect and quantitate viable virus directly in these insect cells are lacking. In situ immune infrared fluorescent staining techniques were developed to visualize and quantitate BTV infection in Culicoides cell culture by both an endpoint titration and an agarose overlay fluorescent focus assay. Insect cell cultures infected with BTV were fixed, permeabilized and reacted with virus-specific monoclonal antibody and fluorescent-labeled secondary antibody. Virus replication in the infected cells was visualized and quantitated by measuring fluorescence with an infrared imager. The sensitivity of virus detection in insect cell culture using these techniques was comparable to or better than detection by standard techniques in vertebrate cell culture.

  7. Advances in Assays and Analytical Approaches for Botulinum Toxin Detection

    SciTech Connect

    Grate, Jay W.; Ozanich, Richard M.; Warner, Marvin G.; Bruckner-Lea, Cindy J.; Marks, James D.

    2010-08-04

    Methods to detect botulinum toxin, the most poisonous substance known, are reviewed. Current assays are being developed with two main objectives in mind: 1) to obtain sufficiently low detection limits to replace the mouse bioassay with an in vitro assay, and 2) to develop rapid assays for screening purposes that are as sensitive as possible while requiring an hour or less to process the sample an obtain the result. This review emphasizes the diverse analytical approaches and devices that have been developed over the last decade, while also briefly reviewing representative older immunoassays to provide background and context.

  8. Toward the conceptual and quantitative understanding of biosolids conditioning: the gel approach.

    PubMed

    Dursun, Derya; Dentel, Steven K

    2009-01-01

    Proper chemical conditioning of wastewater solids is crucial for both operational and economic reasons, but the process has defied satisfactory description to date, in either conceptual or quantitative terms. In this research, a new conceptual model of biosolids structure--likening it to a colloidal gel--was assessed as a means of interpreting conditioning mechanisms. The basis of the gel approach lies in the colligative properties that are altered by lowering of the solvent chemical potential by introducing a solute. Results indicate that inorganic conditioners form precipitates and complexes thus collapsing the gel network and forming particulates, whereas organic polymers lead to heterogeneous collapse with limited diffusion inside the gel. A gel model, based on the osmotic pressure, was found reasonably successful in defining the conditioning efficacy of biosolids. Beyond the model's fundamental value, these results validate a new way of understanding how conditioning and dewatering operate, which should help to improve the selection and optimization of these processes.

  9. Shotgun Approach for Quantitative Imaging of Phospholipids Using Nanospray Desorption Electrospray Ionization Mass Spectrometry

    SciTech Connect

    Lanekoff, Ingela T.; Thomas, Mathew; Laskin, Julia

    2014-02-04

    Mass spectrometry imaging (MSI) has been extensively used for determining spatial distributions of molecules in biological samples, and there is increasing interest in using MSI for quantification. Nanospray desorption electrospray ionization, or nano-DESI, is an ambient MSI technique where a solvent is used for localized extraction of molecules followed by nanoelectrospray ionization. Doping the nano-DESI solvent with carefully selected standards enables online quantification during MSI experiments. In this proof-of-principle study, we demonstrate this quantification approach can be extended to provide shotgun-like quantification of phospholipids in thin brain tissue sections. Specifically, two phosphatidylcholine (PC) standards were added to the nano-DESI solvent for simultaneous imaging and quantification of 22 PC species observed in nano-DESI MSI. Furthermore, by combining the quantitative data obtained in the individual pixels, we demonstrate quantification of these PC species in seven different regions of a rat brain tissue section.

  10. Serial Quantitative PCR Assay for Detection, Species Discrimination, and Quantification of Leishmania spp. in Human Samples▿

    PubMed Central

    Weirather, Jason L.; Jeronimo, Selma M. B.; Gautam, Shalini; Sundar, Shyam; Kang, Mitchell; Kurtz, Melissa A.; Haque, Rashidul; Schriefer, Albert; Talhari, Sinésio; Carvalho, Edgar M.; Donelson, John E.; Wilson, Mary E.

    2011-01-01

    The Leishmania species cause a variety of human disease syndromes. Methods for diagnosis and species differentiation are insensitive and many require invasive sampling. Although quantitative PCR (qPCR) methods are reported for leishmania detection, no systematic method to quantify parasites and determine the species in clinical specimens is established. We developed a serial qPCR strategy to identify and rapidly differentiate Leishmania species and quantify parasites in clinical or environmental specimens. SYBR green qPCR is mainly employed, with corresponding TaqMan assays for validation. The screening primers recognize kinetoplast minicircle DNA of all Leishmania species. Species identification employs further qPCR set(s) individualized for geographic regions, combining species-discriminating probes with melt curve analysis. The assay was sufficient to detect Leishmania parasites, make species determinations, and quantify Leishmania spp. in sera, cutaneous biopsy specimens, or cultured isolates from subjects from Bangladesh or Brazil with different forms of leishmaniasis. The multicopy kinetoplast DNA (kDNA) probes were the most sensitive and useful for quantification based on promastigote standard curves. To test their validity for quantification, kDNA copy numbers were compared between Leishmania species, isolates, and life stages using qPCR. Maxicircle and minicircle copy numbers differed up to 6-fold between Leishmania species, but the differences were smaller between strains of the same species. Amastigote and promastigote leishmania life stages retained similar numbers of kDNA maxi- or minicircles. Thus, serial qPCR is useful for leishmania detection and species determination and for absolute quantification when compared to a standard curve from the same Leishmania species. PMID:22042830

  11. [Selective detection of viable pathogenic bacteria in water using reverse transcription quantitative PCR].

    PubMed

    Lin, Yi-Wen; Li, Dan; Wu, Shu-Xu; He, Miao; Yang, Tian

    2012-11-01

    A reverse transcription q quantitative PCR (RT-qPCR) assay method was established, which can quantify the copy numbers of RNA in pathogenic bacteria of E. coli and Enterococcus faecium. The results showed that cDNA was generated with the RT-PCR reagents, target gene was quantified with the qPCR, the copy numbers of RNA were stable at about 1 copies x CFU(-1) for E. coli and 7.98 x 10(2) copies x CFU(-1) for Enterococcus faecium respectively during the stationary grow phase for the both indicator bacteria [E. coli (6-18 h) and Enterococcus faecium (10-38 h)]. The established RT-qPCR method can quantify the numbers of viable bacteria through detecting bacterial RNA targets. Through detecting the heat-treated E. coli and Enterococcus faecium by three methods (culture method, qPCR, RT-qPCR), we found that the qPCR and RT-qPCR can distinguish 1.43 lg copy non-viable E. coli and 2.5 lg copy non-viable Enterococcus faecium. These results indicated that the established methods could effectively distinguish viable bacteria from non-viable bacteria. Finally we used this method to evaluate the real effluents of the secondary sedimentation of wastewater treatment plant (WWTP), the results showed that the correlation coefficients (R2) between RT-qPCR and culture method were 0.930 (E. coli) and 0.948 (Enterococcus faecium), and this established RT-PCR method can rapidly detect viable pathogenic bacteria in genuine waters.

  12. DetectTLC: Automated Reaction Mixture Screening Utilizing Quantitative Mass Spectrometry Image Features

    NASA Astrophysics Data System (ADS)

    Kaddi, Chanchala D.; Bennett, Rachel V.; Paine, Martin R. L.; Banks, Mitchel D.; Weber, Arthur L.; Fernández, Facundo M.; Wang, May D.

    2016-02-01

    Full characterization of complex reaction mixtures is necessary to understand mechanisms, optimize yields, and elucidate secondary reaction pathways. Molecular-level information for species in such mixtures can be readily obtained by coupling mass spectrometry imaging (MSI) with thin layer chromatography (TLC) separations. User-guided investigation of imaging data for mixture components with known m/z values is generally straightforward; however, spot detection for unknowns is highly tedious, and limits the applicability of MSI in conjunction with TLC. To accelerate imaging data mining, we developed DetectTLC, an approach that automatically identifies m/z values exhibiting TLC spot-like regions in MS molecular images. Furthermore, DetectTLC can also spatially match m/z values for spots acquired during alternating high and low collision-energy scans, pairing product ions with precursors to enhance structural identification. As an example, DetectTLC is applied to the identification and structural confirmation of unknown, yet significant, products of abiotic pyrazinone and aminopyrazine nucleoside analog synthesis.

  13. DetectTLC: Automated Reaction Mixture Screening Utilizing Quantitative Mass Spectrometry Image Feature

    PubMed Central

    Kaddi, Chanchala D.; Bennett, Rachel V.; Paine, Martin R. L.; Banks, Mitchel D.; Weber, Arthur L.; Fernández, Facundo M.; Wang, May D.

    2016-01-01

    Full characterization of complex reaction mixtures is necessary to understand mechanisms, optimize yields, and elucidate secondary reaction pathways. Molecular-level information for species in such mixtures can be readily obtained by coupling mass spectrometry imaging (MSI) with thin layer chromatography (TLC) separations. User-guided investigation of imaging data for mixture components with known m/z values is generally straightforward; however, spot detection for unknowns is highly tedious, and limits the applicability of MSI in conjunction with TLC. To accelerate imaging data mining, we developed DetectTLC, an approach that automatically identifies m/z values exhibiting TLC spot-like regions in MS molecular images. Furthermore, DetectTLC can also spatially match m/z values for spots acquired during alternating high and low collision-energy scans, pairing product ions with precursors to enhance structural identification. As an example, DetectTLC is applied to the identification and structural confirmation of unknown, yet significant, products of abiotic pyrazinone and aminopyrazine nucleoside analog synthesis. PMID:26508443

  14. DetectTLC: Automated Reaction Mixture Screening Utilizing Quantitative Mass Spectrometry Image Features.

    PubMed

    Kaddi, Chanchala D; Bennett, Rachel V; Paine, Martin R L; Banks, Mitchel D; Weber, Arthur L; Fernández, Facundo M; Wang, May D

    2016-02-01

    Full characterization of complex reaction mixtures is necessary to understand mechanisms, optimize yields, and elucidate secondary reaction pathways. Molecular-level information for species in such mixtures can be readily obtained by coupling mass spectrometry imaging (MSI) with thin layer chromatography (TLC) separations. User-guided investigation of imaging data for mixture components with known m/z values is generally straightforward; however, spot detection for unknowns is highly tedious, and limits the applicability of MSI in conjunction with TLC. To accelerate imaging data mining, we developed DetectTLC, an approach that automatically identifies m/z values exhibiting TLC spot-like regions in MS molecular images. Furthermore, DetectTLC can also spatially match m/z values for spots acquired during alternating high and low collision-energy scans, pairing product ions with precursors to enhance structural identification. As an example, DetectTLC is applied to the identification and structural confirmation of unknown, yet significant, products of abiotic pyrazinone and aminopyrazine nucleoside analog synthesis. Graphical Abstract ᅟ.

  15. DetectTLC: Automated Reaction Mixture Screening Utilizing Quantitative Mass Spectrometry Image Features.

    PubMed

    Kaddi, Chanchala D; Bennett, Rachel V; Paine, Martin R L; Banks, Mitchel D; Weber, Arthur L; Fernández, Facundo M; Wang, May D

    2016-02-01

    Full characterization of complex reaction mixtures is necessary to understand mechanisms, optimize yields, and elucidate secondary reaction pathways. Molecular-level information for species in such mixtures can be readily obtained by coupling mass spectrometry imaging (MSI) with thin layer chromatography (TLC) separations. User-guided investigation of imaging data for mixture components with known m/z values is generally straightforward; however, spot detection for unknowns is highly tedious, and limits the applicability of MSI in conjunction with TLC. To accelerate imaging data mining, we developed DetectTLC, an approach that automatically identifies m/z values exhibiting TLC spot-like regions in MS molecular images. Furthermore, DetectTLC can also spatially match m/z values for spots acquired during alternating high and low collision-energy scans, pairing product ions with precursors to enhance structural identification. As an example, DetectTLC is applied to the identification and structural confirmation of unknown, yet significant, products of abiotic pyrazinone and aminopyrazine nucleoside analog synthesis. Graphical Abstract ᅟ. PMID:26508443

  16. Graphene-Semiconductor Catalytic Nanodiodes for Quantitative Detection of Hot Electrons Induced by a Chemical Reaction.

    PubMed

    Lee, Hyosun; Nedrygailov, Ievgen I; Lee, Young Keun; Lee, Changhwan; Choi, Hongkyw; Choi, Jin Sik; Choi, Choon-Gi; Park, Jeong Young

    2016-03-01

    Direct detection of hot electrons generated by exothermic surface reactions on nanocatalysts is an effective strategy to obtain insight into electronic excitation during chemical reactions. For this purpose, we fabricated a novel catalytic nanodiode based on a Schottky junction between a single layer of graphene and an n-type TiO2 layer that enables the detection of hot electron flows produced by hydrogen oxidation on Pt nanoparticles. By making a comparative analysis of data obtained from measuring the hot electron current (chemicurrent) and turnover frequency, we demonstrate that graphene's unique electronic structure and extraordinary material properties, including its atomically thin nature and ballistic electron transport, allow improved conductivity at the interface between the catalytic Pt nanoparticles and the support. Thereby, graphene-based nanodiodes offer an effective and facile way to approach the study of chemical energy conversion mechanisms in composite catalysts with carbon-based supports.

  17. Quantitative elemental detection of size-segregated particles using laser-induced breakdown spectroscopy

    NASA Astrophysics Data System (ADS)

    Wang, Zhen Zhen; Deguchi, Yoshihiro; Kuwahara, Masakazu; Taira, Takuya; Zhang, Xiao Bo; Yan, Jun Jie; Liu, Ji Ping; Watanabe, Hiroaki; Kurose, Ryoichi

    2013-09-01

    In order to simulate coal combustion and develop optimal and stable boiler control systems in real power plants, it is imperative to obtain the detailed information in coal combustion processes as well as to measure species contents in fly ash, which should be controlled and analyzed for enhancing boiler efficiency and reducing environmental pollution. The fly ash consists of oxides (SiO2, Al2O3, Fe2O3, CaO, and so on), unburned carbon, and other minor elements. Recently laser-induced breakdown spectroscopy (LIBS) technique has been applied to coal combustion and other industrial fields because of the fast response, high sensitivity, real-time and non-contact features. In these applications it is important to measure controlling factors without any sample preparation to maintain the real-time measurement feature. The relation between particle content and particle diameter is also one of the vital researches, because compositions of particles are dependent on their diameter. In this study, we have detected the contents of size-segregated particles using LIBS. Particles were classified by an Anderson cascade impactor and their contents were measured using the output of 1064 nm YAG laser, a spectrograph and an ICCD camera. The plasma conditions such as plasma temperature are dependent on the size of particles and these effects must be corrected to obtain quantitative information. The plasma temperature was corrected by the emission intensity ratio from the same atom. Using this correction method, the contents of particles can be measured quantitatively in fixed experimental parameters. This method was applied to coal and fly ash from a coal-fired burner to measure unburned carbon and other contents according to the particle diameter. The acquired results demonstrate that the LIBS technique is applicable to measure size-segregated particle contents in real time and this method is useful for the analysis of coal combustion and its control because of its sensitive and

  18. Allele-specific qRT-PCR demonstrates superior detection of single nucleotide polymorphisms as genetic markers for West Nile virus compared to Luminex® and quantitative sequencing.

    PubMed

    Worwa, Gabriella; Andrade, Christy C; Thiemann, Tara C; Park, Bborie; Maharaj, Payal D; Anishchenko, Michael; Brault, Aaron C; Reisen, William K

    2014-01-01

    To enable in vivo and in vitro competitive fitness comparisons among West Nile viruses (WNV), three reference viruses were marked genetically by site-directed mutagenesis with five synonymous nucleotide substitutions in the envelope gene region of the genome. Phenotypic neutrality of the mutants was assessed experimentally by competitive replication in cell culture and genetic stability of the substituted nucleotides was confirmed by direct sequencing. Luminex(®) technology, quantitative sequencing and quantitative RT-PCR (qRT-PCR) were compared in regard to specificity, sensitivity and accuracy for quantitation of wildtype and genetically marked viruses in mixed samples based on RNA obtained from samples of known viral titers. Although Luminex(®) technology and quantitative sequencing provided semi-quantitative or qualitative measurements, a sequence-specific primer extension approach using a specific reverse primer set in singleplex qRT-PCR demonstrated the best quantitation and specificity in the detection of RNA from wildtype and mutant viruses.

  19. Repertoire of novel sequence signatures for the detection of Candidatus Liberibacter asiaticus by quantitative real-time PCR

    PubMed Central

    2014-01-01

    Background Huanglongbing (HLB) or citrus greening is a devastating disease of citrus. The gram-negative bacterium Candidatus Liberibacter asiaticus (Las) belonging to the α-proteobacteria is responsible for HLB in North America as well as in Asia. Currently, there is no cure for this disease. Early detection and quarantine of Las-infected trees are important management strategies used to prevent HLB from invading HLB-free citrus producing regions. Quantitative real-time PCR (qRT-PCR) based molecular diagnostic assays have been routinely used in the detection and diagnosis of Las. The oligonucleotide primer pairs based on conserved genes or regions, which include 16S rDNA and the β-operon, have been widely employed in the detection of Las by qRT-PCR. The availability of whole genome sequence of Las now allows the design of primers beyond the conserved regions for the detection of Las explicitly. Results We took a complimentary approach by systematically screening the genes in a genome-wide fashion, to identify the unique signatures that are only present in Las by an exhaustive sequence based similarity search against the nucleotide sequence database. Our search resulted in 34 probable unique signatures. Furthermore, by designing the primer pair specific to the identified signatures, we showed that most of our primer sets are able to detect Las from the infected plant and psyllid materials collected from the USA and China by qRT-PCR. Overall, 18 primer pairs of the 34 are found to be highly specific to Las with no cross reactivity to the closely related species Ca. L. americanus (Lam) and Ca. L. africanus (Laf). Conclusions We have designed qRT-PCR primers based on Las specific genes. Among them, 18 are suitable for the detection of Las from Las-infected plant and psyllid samples. The repertoire of primers that we have developed and characterized in this study enhanced the qRT-PCR based molecular diagnosis of HLB. PMID:24533511

  20. Persistence of host-associated Bacteroidales gene markers and their quantitative detection in an urban and agricultural mixed prairie watershed.

    PubMed

    Tambalo, Dinah D; Fremaux, Bastien; Boa, Tyler; Yost, Christopher K

    2012-06-01

    Microbial source tracking is an emerging tool developed to protect water sources from faecal pollution. In this study, we evaluated the suitability of real time-quantitative PCR (qPCR) Taqman assays developed for detection of host-associated Bacteroidales markers in a prairie watershed. The qPCR primers and probes used in this study exhibited high accuracy (88-96% sensitivity and ≥ 99% host specificity) in detecting Bacteroidales spp. that are associated with faeces from humans, ruminants, bovines, and horses. The ruminant- and human-associated markers were also found in high concentrations within individual faecal samples, ranging from 3.4 to 7.3 log(10) marker copy numberg(-1) of individual host faeces. Following validation of host sensitivity and specificity, the host-associated Bacteroidales markers were detected in the Qu'Appelle Valley watershed of Saskatchewan, Canada which experiences a diversity of anthropogenic inputs. Concentrations of the ruminant marker were well-correlated with proximity to cattle operations and there was a correlation between the marker and Escherichia coli concentrations at these sites. Low concentrations of the human faecal marker were measured throughout the sampling sites, and may indicate a consistent influx of human faecal pollution into the watershed area. Persistence of each of the Bacteroidales host-associated marker was also studied in situ. The results indicated that the markers persist for shorter periods of time (99% decay in <8 days) compared with the conventional E. coli marker (99% decay in >15 days), suggesting they are effective at detecting recent faecal contamination events. The levels of Bacteroidales markers and E. coli counts did not correlate with the presence of the pathogenic bacteria, Salmonella spp. or Campylobacter spp. detected in the Qu'Appelle Valley. Collectively, the results obtained in this study demonstrated that the qPCR approach for detecting host-associated Bacteroidales spp. markers can be a

  1. Improved minimal residual disease detection by targeted quantitative polymerase chain reaction in Nucleophosmin 1 type a mutated acute myeloid leukemia.

    PubMed

    Pettersson, Louise; Levéen, Per; Axler, Olof; Dvorakova, Dana; Juliusson, Gunnar; Ehinger, Mats

    2016-10-01

    Multicolor flow cytometry (MFC) and real-time quantitative PCR (RQ-PCR) are important independent techniques to determine minimal residual disease (MRD) in acute myeloid leukemia (AML). MFC is the standard method, but may be unreliable. Therefore, MFC-based determination of MRD with an RQ-PCR-based approach targeting the nucleophosmin 1 (NPM1) type A mutation was set out to compare. Since most current NPM1 RQ-PCR MRD protocols suffer from clear definitions of quantifiability, we sought to define quantifiability in a reproducible and standardized manner. The limit of quantifiability of our RQ-PCR protocol for the NPM1 type A mutation varied between 0.002% and 0.04% residual leukemic cells depending on the features of the standard curve for each PCR experiment. The limit of detection was close to 0.001% leukemic cells. The limit of detection by MFC ranged from 0.01% to 1% depending on the phenotype of the leukemic cells as compared with non-leukemic bone marrow cells. Forty-five MRD samples from 15 patients using both NPM1 mutation specific RQ-PCR and MFC were analyzed. In 32 of the 45 samples (71%), an MRD-signal could be detected with RQ-PCR. A quantifiable NPM1 mutation signal was found in 15 samples (33%) (range 0.003%-2.6% leukemic cells). By contrast, only two follow-up samples (4%) showed residual leukemic cells (0.04% and 0.3%, respectively) by MFC. Thus, RQ-PCR of the NPM1 type A mutation was more sensitive and reliable than MFC for determination of MRD, which might have clinical implications. © 2016 Wiley Periodicals, Inc.

  2. Improved minimal residual disease detection by targeted quantitative polymerase chain reaction in Nucleophosmin 1 type a mutated acute myeloid leukemia.

    PubMed

    Pettersson, Louise; Levéen, Per; Axler, Olof; Dvorakova, Dana; Juliusson, Gunnar; Ehinger, Mats

    2016-10-01

    Multicolor flow cytometry (MFC) and real-time quantitative PCR (RQ-PCR) are important independent techniques to determine minimal residual disease (MRD) in acute myeloid leukemia (AML). MFC is the standard method, but may be unreliable. Therefore, MFC-based determination of MRD with an RQ-PCR-based approach targeting the nucleophosmin 1 (NPM1) type A mutation was set out to compare. Since most current NPM1 RQ-PCR MRD protocols suffer from clear definitions of quantifiability, we sought to define quantifiability in a reproducible and standardized manner. The limit of quantifiability of our RQ-PCR protocol for the NPM1 type A mutation varied between 0.002% and 0.04% residual leukemic cells depending on the features of the standard curve for each PCR experiment. The limit of detection was close to 0.001% leukemic cells. The limit of detection by MFC ranged from 0.01% to 1% depending on the phenotype of the leukemic cells as compared with non-leukemic bone marrow cells. Forty-five MRD samples from 15 patients using both NPM1 mutation specific RQ-PCR and MFC were analyzed. In 32 of the 45 samples (71%), an MRD-signal could be detected with RQ-PCR. A quantifiable NPM1 mutation signal was found in 15 samples (33%) (range 0.003%-2.6% leukemic cells). By contrast, only two follow-up samples (4%) showed residual leukemic cells (0.04% and 0.3%, respectively) by MFC. Thus, RQ-PCR of the NPM1 type A mutation was more sensitive and reliable than MFC for determination of MRD, which might have clinical implications. © 2016 Wiley Periodicals, Inc. PMID:27191933

  3. Quantitative real-time PCR assay for detection of Paenibacillus polymyxa using membrane-fusion protein-based primers.

    PubMed

    Cho, Min Seok; Park, Dong Suk; Lee, Jung Won; Chi, Hee Youn; Sohn, Soo-In; Jeon, Bong-Kyun; Ma, Jong-Beom

    2012-11-01

    Paenibacillus polymyxa is known to be a plant-growthpromoting rhizobacterium. The present study describes a quantitative polymerase chain reaction (qPCR) assay for the specific detection and quantitation of P. polymyxa using a primer pair based on the sequence of a membranefusion protein for the amplification of a 268 bp DNA fragment. This study reports that the qPCR-based method is applicable for the rapid and sensitive detection of P. polymyxa and can be used as an alternative method for agricultural soil monitoring.

  4. Approaching the Heisenberg Limit without Single-Particle Detection.

    PubMed

    Davis, Emily; Bentsen, Gregory; Schleier-Smith, Monika

    2016-02-01

    We propose an approach to quantum phase estimation that can attain precision near the Heisenberg limit without requiring single-particle-resolved state detection. We show that the "one-axis twisting" interaction, well known for generating spin squeezing in atomic ensembles, can also amplify the output signal of an entanglement-enhanced interferometer to facilitate readout. Applying this interaction-based readout to oversqueezed, non-Gaussian states yields a Heisenberg scaling in phase sensitivity, which persists in the presence of detection noise as large as the quantum projection noise of an unentangled ensemble. Even in dissipative implementations-e.g., employing light-mediated interactions in an optical cavity or Rydberg dressing-the method significantly relaxes the detection resolution required for spectroscopy beyond the standard quantum limit. PMID:26894711

  5. Approaching the Heisenberg Limit Without Single-Particle Detection

    NASA Astrophysics Data System (ADS)

    Bentsen, Gregory; Davis, Emily; Schleier-Smith, Monika

    2016-05-01

    Achieving Heisenberg-limited measurements with ensembles of more than a few particles remains a major outstanding challenge. The problem is two-fold: one must not only prepare a sufficiently sensitive state, but also be able to detect it. While it is commonly assumed that Heisenberg-limited measurement demands single-particle-resolved detection, we propose an alternative approach that bypasses this requirement. We show that the ``one-axis twisting'' interaction, well known for generating spin squeezing in atomic ensembles, can also amplify the output signal of an entanglement-enhanced interferometer to facilitate readout. Even in the presence of dissipation, the protocol significantly relaxes the detection resolution required for spectroscopy beyond the standard quantum limit, and achieves near-Heisenberg-limited precision in a √{ N}-times shorter evolution than is required to reach the GHZ state. AFOSR, NSF.

  6. Mammography mass detection: a multi-stage hybrid approach

    NASA Astrophysics Data System (ADS)

    Sahba, Nima; Tavakoli, Vahid; Ahmadian, Alireza; Giti, Masoumeh

    2009-02-01

    Here in this paper a combined method of pixel based and region based mass detection is proposed. In the first step, the background and pectoral muscle are filtered from mammography images and the image contrast is enhanced using an adaptive density weighted approach. Then, in a coarse level, suspected regions are extracted based on mathematical morphology and adaptive thresholding methods. Finally, to reduce the false positives produced in the coarse stage, a useful feature vector based on ranklet transform is obtained and fed into a support vector machine classifier to detect masses. MIAS (Mammographic Image Analysis Society) and Imam Hospital databases were used to evaluate the performance of the algorithm. The sensitivity and specificity of the proposed method are 74% and 91% respectively. The proposed algorithm shows a high degree of robustness in detecting masses of different shapes.

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

    PubMed

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

    2008-05-15

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

  8. An integrated pattern recognition approach for intrusion detection.

    PubMed

    Pandit, Amod; Stanley, R Joe; McMillin, Bruce

    2002-01-01

    Intrusion detection systems (IDS) attempt to address the vulnerability of computer-based systems for abuse by insiders and to penetration by outsiders. An IDS is required to examine an enormous amount of data generated by computer networks to assist in the abuse detection process. Thus, there is a need to develop automated tools that address these requirements to assist system operators in the detection of violations of existing security policies. In this research, an automated IDS is proposed for insider threats in a distributed system. The proposed IDS functions as an anomaly detector for insider system operations based on the analysis of the system's log files. The approach integrates dynamic programming and adaptive resonance theory (ART1) clustering. The integrated approach aligns sequences of log events with prototypical sequences of events for performing tasks and classifies the aligned sequences for intrusion detection. The system examined for this research is a Boots System for controlling the movement of boots from one place to another under specific security restrictions related to the boot orders. We present the proposed model, the results achieved and the analysis of an implemented prototype.

  9. DNA signature-based approaches for bacterial detection and identification.

    PubMed

    Albuquerque, Pedro; Mendes, Marta V; Santos, Catarina L; Moradas-Ferreira, Pedro; Tavares, Fernando

    2009-06-01

    During the late eighties, environmental microbiologists realized the potential of the polymerase chain reaction (PCR) for the design of innovative approaches to study microbial communities or to detect and identify microorganisms in diverse and complex environments. In contrast to long-established methods of cultivation-based microbial identification, PCR-based techniques allow for the identification of microorganisms regardless of their culturability. A large number of reports have been published that describe PCR-inspired methods, frequently complemented by sequencing or hybridization profiling, to infer taxonomic and clonal microbial diversity or to detect and identify microorganisms using taxa-specific genomic markers. Typing methods have been particularly useful for microbial ecology-driven studies; however, they are not suitable for diagnostic purposes, such as the detection of specific species, strains or clones. Recently, comprehensive reviews have been written describing the panoply of typing methods available and describing their advantages and limitations; however, molecular approaches for bacterial detection and identification were either not considered or only vaguely discussed. This review focuses on DNA-based methods for bacterial detection and identification, highlighting strategies for selecting taxa-specific loci and emphasizing the molecular techniques and emerging technological solutions for increasing the detection specificity and sensitivity. The massive and increasing number of available bacterial sequences in databases, together with already employed bioinformatics tools, hold promise of more reliable, fast and cost-effective methods for bacterial identification in a wide range of samples in coming years. This tendency will foster the validation and certification of these methods and their routine implementation by certified diagnostic laboratories.

  10. Developmental differences in auditory detection and localization of approaching vehicles.

    PubMed

    Barton, Benjamin K; Lew, Roger; Kovesdi, Casey; Cottrell, Nicholas D; Ulrich, Thomas

    2013-04-01

    Pedestrian safety is a significant problem in the United States, with thousands being injured each year. Multiple risk factors exist, but one poorly understood factor is pedestrians' ability to attend to vehicles using auditory cues. Auditory information in the pedestrian setting is increasing in importance with the growing number of quieter hybrid and all-electric vehicles on America's roadways that do not emit sound cues pedestrians expect from an approaching vehicle. Our study explored developmental differences in pedestrians' detection and localization of approaching vehicles. Fifty children ages 6-9 years, and 35 adults participated. Participants' performance varied significantly by age, and with increasing speed and direction of the vehicle's approach. Results underscore the importance of understanding children's and adults' use of auditory cues for pedestrian safety and highlight the need for further research.

  11. Developmental differences in auditory detection and localization of approaching vehicles.

    PubMed

    Barton, Benjamin K; Lew, Roger; Kovesdi, Casey; Cottrell, Nicholas D; Ulrich, Thomas

    2013-04-01

    Pedestrian safety is a significant problem in the United States, with thousands being injured each year. Multiple risk factors exist, but one poorly understood factor is pedestrians' ability to attend to vehicles using auditory cues. Auditory information in the pedestrian setting is increasing in importance with the growing number of quieter hybrid and all-electric vehicles on America's roadways that do not emit sound cues pedestrians expect from an approaching vehicle. Our study explored developmental differences in pedestrians' detection and localization of approaching vehicles. Fifty children ages 6-9 years, and 35 adults participated. Participants' performance varied significantly by age, and with increasing speed and direction of the vehicle's approach. Results underscore the importance of understanding children's and adults' use of auditory cues for pedestrian safety and highlight the need for further research. PMID:23357030

  12. Fault detection and diagnosis using neural network approaches

    NASA Technical Reports Server (NTRS)

    Kramer, Mark A.

    1992-01-01

    Neural networks can be used to detect and identify abnormalities in real-time process data. Two basic approaches can be used, the first based on training networks using data representing both normal and abnormal modes of process behavior, and the second based on statistical characterization of the normal mode only. Given data representative of process faults, radial basis function networks can effectively identify failures. This approach is often limited by the lack of fault data, but can be facilitated by process simulation. The second approach employs elliptical and radial basis function neural networks and other models to learn the statistical distributions of process observables under normal conditions. Analytical models of failure modes can then be applied in combination with the neural network models to identify faults. Special methods can be applied to compensate for sensor failures, to produce real-time estimation of missing or failed sensors based on the correlations codified in the neural network.

  13. Quantitative risk-based approach for improving water quality management in mining.

    PubMed

    Liu, Wenying; Moran, Chris J; Vink, Sue

    2011-09-01

    The potential environmental threats posed by freshwater withdrawal and mine water discharge are some of the main drivers for the mining industry to improve water management. The use of multiple sources of water supply and introducing water reuse into the mine site water system have been part of the operating philosophies employed by the mining industry to realize these improvements. However, a barrier to implementation of such good water management practices is concomitant water quality variation and the resulting impacts on the efficiency of mineral separation processes, and an increased environmental consequence of noncompliant discharge events. There is an increasing appreciation that conservative water management practices, production efficiency, and environmental consequences are intimately linked through the site water system. It is therefore essential to consider water management decisions and their impacts as an integrated system as opposed to dealing with each impact separately. This paper proposes an approach that could assist mine sites to manage water quality issues in a systematic manner at the system level. This approach can quantitatively forecast the risk related with water quality and evaluate the effectiveness of management strategies in mitigating the risk by quantifying implications for production and hence economic viability. PMID:21797262

  14. A Novel Multiparametric Approach to 3D Quantitative MRI of the Brain

    PubMed Central

    Palma, Giuseppe; Tedeschi, Enrico; Borrelli, Pasquale; Cocozza, Sirio; Russo, Carmela; Liu, Saifeng; Ye, Yongquan; Comerci, Marco; Alfano, Bruno; Salvatore, Marco; Haacke, E. Mark; Mancini, Marcello

    2015-01-01

    Magnetic Resonance properties of tissues can be quantified in several respects: relaxation processes, density of imaged nuclei, magnetism of environmental molecules, etc. In this paper, we propose a new comprehensive approach to obtain 3D high resolution quantitative maps of arbitrary body districts, mainly focusing on the brain. The theory presented makes it possible to map longitudinal (R1), pure transverse (R2) and free induction decay (R2*) rates, along with proton density (PD) and magnetic susceptibility (χ), from a set of fast acquisition sequences in steady-state that are highly insensitive to flow phenomena. A novel denoising scheme is described and applied to the acquired datasets to enhance the signal to noise ratio of the derived maps and an information theory approach compensates for biases from radio frequency (RF) inhomogeneities, if no direct measure of the RF field is available. Finally, the results obtained on sample brain scans of healthy controls and multiple sclerosis patients are presented and discussed. PMID:26284778

  15. Quantitative risk-based approach for improving water quality management in mining.

    PubMed

    Liu, Wenying; Moran, Chris J; Vink, Sue

    2011-09-01

    The potential environmental threats posed by freshwater withdrawal and mine water discharge are some of the main drivers for the mining industry to improve water management. The use of multiple sources of water supply and introducing water reuse into the mine site water system have been part of the operating philosophies employed by the mining industry to realize these improvements. However, a barrier to implementation of such good water management practices is concomitant water quality variation and the resulting impacts on the efficiency of mineral separation processes, and an increased environmental consequence of noncompliant discharge events. There is an increasing appreciation that conservative water management practices, production efficiency, and environmental consequences are intimately linked through the site water system. It is therefore essential to consider water management decisions and their impacts as an integrated system as opposed to dealing with each impact separately. This paper proposes an approach that could assist mine sites to manage water quality issues in a systematic manner at the system level. This approach can quantitatively forecast the risk related with water quality and evaluate the effectiveness of management strategies in mitigating the risk by quantifying implications for production and hence economic viability.

  16. Physical oncology: a bench-to-bedside quantitative and predictive approach.

    PubMed

    Frieboes, Hermann B; Chaplain, Mark A J; Thompson, Alastair M; Bearer, Elaine L; Lowengrub, John S; Cristini, Vittorio

    2011-01-15

    Cancer models relating basic science to clinical care in oncology may fail to address the nuances of tumor behavior and therapy, as in the case, discussed herein, of the complex multiscale dynamics leading to the often-observed enhanced invasiveness, paradoxically induced by the very antiangiogenic therapy designed to destroy the tumor. Studies would benefit from approaches that quantitatively link the multiple physical and temporal scales from molecule to tissue in order to offer outcome predictions for individual patients. Physical oncology is an approach that applies fundamental principles from the physical and biological sciences to explain certain cancer behaviors as observable characteristics arising from the underlying physical and biochemical events. For example, the transport of oxygen molecules through tissue affects phenotypic characteristics such as cell proliferation, apoptosis, and adhesion, which in turn underlie the patient-scale tumor growth and invasiveness. Our review of physical oncology illustrates how tumor behavior and treatment response may be a quantifiable function of marginally stable molecular and/or cellular conditions modulated by inhomogeneity. By incorporating patient-specific genomic, proteomic, metabolomic, and cellular data into multiscale physical models, physical oncology could complement current clinical practice through enhanced understanding of cancer behavior, thus potentially improving patient survival.

  17. Direct magnetic resonance detection of myelin and prospects for quantitative imaging of myelin density

    PubMed Central

    Wilhelm, Michael J.; Ong, Henry H.; Tsai, Ping-Huei; Hackney, David B.; Wehrli, Felix W.

    2012-01-01

    Magnetic resonance imaging has previously demonstrated its potential for indirectly mapping myelin density, either by relaxometric detection of myelin water or magnetization transfer. Here, we investigated whether myelin can be detected and possibly quantified directly. We identified the spectrum of myelin in the spinal cord in situ as well as in myelin lipids extracted via a sucrose gradient method, and investigated its spectral properties. High-resolution solution NMR spectroscopy showed the extract composition to be in agreement with myelin’s known chemical make-up. The 400-MHz 1H spectrum of the myelin extract, at 20 °C (room temperature) and 37 °C, consists of a narrow water resonance superimposed on a broad envelope shifted ∼3.5 ppm upfield, suggestive of long-chain methylene protons. Superimposed on this signal are narrow components resulting from functional groups matching the chemical shifts of the constituents making up myelin lipids. The spectrum could be modeled as a sum of super-Lorentzians with a T2* distribution covering a wide range of values (0.008–26 ms). Overall, there was a high degree of similarity between the spectral properties of extracted myelin lipids and those found in neural tissue. The normalized difference spectrum had the hallmarks of membrane proteins, not present in the myelin extract. Using 3D radially ramp-sampled proton MRI, with a combination of adiabatic inversion and echo subtraction, the feasibility of direct myelin imaging in situ is demonstrated. Last, the integrated signal from myelin suspensions is shown, both spectroscopically and by imaging, to scale with concentration, suggesting the potential for quantitative determination of myelin density. PMID:22628562

  18. Whole genome scan to detect quantitative trait loci for bovine milk protein composition.

    PubMed

    Schopen, G C B; Koks, P D; van Arendonk, J A M; Bovenhuis, H; Visker, M H P W

    2009-08-01

    The objective of this study was to perform a whole genome scan to detect quantitative trait loci (QTL) for milk protein composition in 849 Holstein-Friesian cows originating from seven sires. One morning milk sample was analysed for the major milk proteins using capillary zone electrophoresis. A genetic map was constructed with 1341 single nucleotide polymorphisms, covering 2829 centimorgans (cM) and 95% of the cattle genome. The chromosomal regions most significantly related to milk protein composition (P(genome) < 0.05) were found on Bos taurus autosomes (BTA) 6, 11 and 14. The QTL on BTA6 was found at about 80 cM, and affected alpha(S1)-casein, alpha(S2)-casein, beta-casein and kappa-casein. The QTL on BTA11 was found at 124 cM, and affected beta-lactoglobulin, and the QTL on BTA14 was found at 0 cM, and affected protein percentage. The proportion of phenotypic variance explained by the QTL was 3.6% for beta-casein and 7.9% for kappa-casein on BTA6, 28.3% for beta-lactoglobulin on BTA11, and 8.6% for protein percentage on BTA14. The QTL affecting alpha(S2)-casein on BTA6 and 17 showed a significant interaction. We investigated the extent to which the detected QTL affecting milk protein composition could be explained by known polymorphisms in beta-casein, kappa-casein, beta-lactoglobulin and DGAT1 genes. Correction for these polymorphisms decreased the proportion of phenotypic variance explained by the QTL previously found on BTA6, 11 and 14. Thus, several significant QTL affecting milk protein composition were found, of which some QTL could partially be explained by polymorphisms in milk protein genes.

  19. Fine mapping and detection of the causative mutation underlying Quantitative Trait Loci.

    PubMed

    Uleberg, E; Meuwissen, T H E

    2010-10-01

    The effect on power and precision of including the causative SNP amongst the investigated markers in Quantitative Trait Loci (QTL) mapping experiments was investigated. Three fine mapping methods were tested to see which was most efficient in finding the causative mutation: combined linkage and linkage disequilibrium mapping (LLD); association mapping (MARK); a combination of LLD and association mapping (LLDMARK). Two simulated data sets were analysed: in one set, the causative SNP was included amongst the markers, while in the other set the causative SNP was masked between markers. Including the causative SNP amongst the markers increased both precision and power in the analyses. For the LLD method the number of correctly positioned QTL increased from 17 for the analysis without the causative SNP to 77 for the analysis including the causative SNP. The likelihood of the data analysis increased from 3.4 to 13.3 likelihood units for the MARK method when the causative SNP was included. When the causative SNP was masked between the analysed markers, the LLD method was most efficient in detecting the correct QTL position, while the MARK method was most efficient when the causative SNP was included as a marker in the analysis. The LLDMARK method, combining association mapping and LLD, assumes a QTL as the null hypothesis (using LLD method) and tests whether the 'putative causative SNP' explains significantly more variance than a QTL in the region. Thus, if the putative causative SNP does not only give an Identical-By-Descent (IBD) signal, but also an Alike-In-State (AIS) signal, LLDMARK gives a positive likelihood ratio. LLDMARK detected less than half as many causative SNPs as the other methods, and also had a relatively high false discovery rate when the QTL effect was large. LLDMARK may however be more robust against spurious associations, because the regional IBD is largely corrected for by fitting a QTL effect in the null hypothesis model.

  20. Detecting Renal Allograft Inflammation Using Quantitative Urine Metabolomics and CXCL10

    PubMed Central

    Ho, Julie; Sharma, Atul; Mandal, Rupasri; Wishart, David S.; Wiebe, Chris; Storsley, Leroy; Karpinski, Martin; Gibson, Ian W.; Nickerson, Peter W.; Rush, David N.

    2016-01-01

    Background The goal of this study was to characterize urinary metabolomics for the noninvasive detection of cellular inflammation and to determine if adding urinary chemokine ligand 10 (CXCL10) improves the overall diagnostic discrimination. Methods Urines (n = 137) were obtained before biopsy in 113 patients with no (n = 66), mild (borderline or subclinical; n = 58), or severe (clinical; n = 13) rejection from a prospective cohort of adult renal transplant patients (n = 113). Targeted, quantitative metabolomics was performed with direct flow injection tandem mass spectrometry using multiple reaction monitoring (ABI 4000 Q-Trap). Urine CXCL10 was measured by enzyme-linked immunosorbent assay. A projection on latent structures discriminant analysis was performed and validated using leave-one-out cross-validation, and an optimal 2-component model developed. Chemokine ligand 10 area under the curve (AUC) was determined and net reclassification index and integrated discrimination index analyses were performed. Results PLS2 demonstrated that urinary metabolites moderately discriminated the 3 groups (Cohen κ, 0.601; 95% confidence interval [95% CI], 0.46-0.74; P < 0.001). Using binary classifiers, urinary metabolites and CXCL10 demonstrated an AUC of 0.81 (95% CI, 0.74-0.88) and 0.76 (95% CI, 0.68-0.84), respectively, and a combined AUC of 0.84 (95% CI, 0.78-0.91) for detecting alloimmune inflammation that was improved by net reclassification index and integrated discrimination index analyses. Urinary CXCL10 was the best univariate discriminator, followed by acylcarnitines and hexose. Conclusions Urinary metabolomics can noninvasively discriminate noninflamed renal allografts from those with subclinical and clinical inflammation, and the addition of urine CXCL10 had a modest but significant effect on overall diagnostic performance. These data suggest that urinary metabolomics and CXCL10 may be useful for noninvasive monitoring of alloimmune inflammation in renal

  1. Detection of Dental Fluorosis-Associated Quantitative Trait Loci on Mouse Chromosomes 2 and 11

    PubMed Central

    Everett, Eric T.; Yan, Dong; Weaver, Marjorie; Liu, Lixiang; Foroud, Tatiana; Martinez-Mier, E. Angeles

    2008-01-01

    Systemic exposure to greater than optimal fluoride (F) can lead to dental fluorosis (DF). Parental A/J (DF-susceptible) and 129P3/J (DF-resistant) inbred mice were used for histological studies and to generate F2 progeny. Mice were treated with 0 or 50 ppm F in their drinking water for 60 days. A clinical criterion (modified Thylstrup and Fejerskov categorical scale) was used to assess the severity of DF for each individual F2 animal. Parental strains were subjected to histological examination of maturing enamel. F treatment resulted in accumulation of amelogenins in the maturing enamel of A/J mice. Quantitative trait loci (QTL) detection was performed using phenotypic extreme F2 animals genotyped for 354 single nucleotide polymorphism-based markers distributed throughout the mouse genome followed by χ2 analysis. Significant evidence of association was observed on chromosomes 2 and 11 for a series of consecutive markers (p < 0.0001). Further analyses were performed to examine whether the phenotypic effects were found in both male and female F2 mice or whether there was evidence for gender-specific effects. Analyses performed using the markers on chromosomes 2 and 11 which were significant in the mixed-gender mice were also significant when analyses were limited to only the male or female mice. The QTL detected on chromosomes 2 and 11 which influence the variation in response to fluorosis have their effect in mice of both genders. Finally, the QTL in both chromosomes appear to have an additive effect. PMID:18701810

  2. Sensitive quantitative detection of commensal bacteria by rRNA-targeted reverse transcription-PCR.

    PubMed

    Matsuda, Kazunori; Tsuji, Hirokazu; Asahara, Takashi; Kado, Yukiko; Nomoto, Koji

    2007-01-01

    A sensitive rRNA-targeted reverse transcription-quantitative PCR (RT-qPCR) method was developed for exact and sensitive enumeration of subdominant bacterial populations. Using group- or species-specific primers for 16S or 23S rRNA, analytical curves were constructed for Escherichia coli, Enterococcus faecalis, Staphylococcus aureus, Clostridium perfringens, and Pseudomonas aeruginosa, and the threshold cycle value was found to be linear up to an RNA amount of 10(-3) cell per RT-PCR. The number of bacteria in culture was determined by RT-qPCR, and the results correlated well with the CFU count over the range from 10(0) to 10(5) CFU. The bacterial counts obtained by RT-qPCR were the same as the CFU counts irrespective of the growth phase in vitro, except for C. perfringens during starvation periods; the viable cell counts obtained by using a combination of 4',6-diamidino-2-phenylindole (DAPI) staining and SYTO9-propidium iodide double staining were in good agreement with the RT-qPCR counts rather than with the CFU counts. The RT-qPCR method could detect endogenous Enterobacteriaceae and P. aeruginosa in feces of hospitalized patients (n = 38) at a level of 10(3) cells per g of feces, and for enumeration of S. aureus or P. aeruginosa spiked into human peripheral blood, the lower detection limit for RT-qPCR quantification of the bacteria was 2 cells per ml of blood, suggesting that this method was equivalent to the conventional culture method. As only 5 h was needed for RT-qPCR quantification, we suggest that rRNA-targeted RT-qPCR assays provide a sensitive and convenient system for quantification of commensal bacteria and for examining their possible invasion of a host.

  3. Plasmonic liquid marbles: a miniature substrate-less SERS platform for quantitative and multiplex ultratrace molecular detection.

    PubMed

    Lee, Hiang Kwee; Lee, Yih Hong; Phang, In Yee; Wei, Jiaqi; Miao, Yue-E; Liu, Tianxi; Ling, Xing Yi

    2014-05-12

    Inspired by aphids, liquid marbles have been studied extensively and have found application as isolated microreactors, as micropumps, and in sensing. However, current liquid-marble-based sensing methodologies are limited to qualitative colorimetry-based detection. Herein we describe the fabrication of a plasmonic liquid marble as a substrate-less analytical platform which, when coupled with ultrasensitive SERS, enables simultaneous multiplex quantification and the identification of ultratrace analytes across separate phases. Our plasmonic liquid marble demonstrates excellent mechanical stability and is suitable for the quantitative examination of ultratrace analytes, with detection limits as low as 0.3 fmol, which corresponds to an analytical enhancement factor of 5×10(8). The results of our simultaneous detection scheme based on plasmonic liquid marbles and an aqueous-solid-organic interface quantitatively tally with those found for the individual detection of methylene blue and coumarin.

  4. A quantitative x-ray detection system for gold nanoparticle tumour biomarkers

    NASA Astrophysics Data System (ADS)

    Ricketts, K.; Castoldi, A.; Guazzoni, C.; Ozkan, C.; Christodoulou, C.; Gibson, A. P.; Royle, G. J.

    2012-09-01

    X-ray fluorescence techniques have proven beneficial for identifying and quantifying trace elements in biological tissues. A novel approach is being developed that employs x-ray fluorescence with an aim to locate heavy nanoparticles, such as gold, which are embedded into tissues. Such nanoparticles can be functionalized to act as markers for tumour characteristics to map the disease state, with the future aim of imaging them to inform cancer therapy regimes. The uptake of functionalized nanoparticles by cancer cells will also enable detection of small clusters of infiltrating cancer cells which are currently missed by commonly used imaging modalities. The novel system, consisting of an energy-resolving silicon drift detector with high spectral resolution, shows potential in both quantification of and sensitivity to nanoparticle concentrations typically found in tumours. A series of synchrotron measurements are presented; a linear relationship between fluorescence intensity and gold nanoparticle (GNP) concentration was found down to 0.005 mgAu ml-1, the detection limit of the system. Successful use of a bench-top source, suitable for possible future clinical use, is also demonstrated, and found not to degrade the detection limit or accuracy of the GNP concentration measurement. The achieved system sensitivity suggests possible future clinical usefulness in measuring tumour uptake in vivo, particularly in shallow tumour sites and small animals, in ex vivo tissue and in 3D in vitro research samples.

  5. Precision, limit of detection and range of quantitation in competitive ELISA.

    PubMed

    Hayashi, Yuzuru; Matsuda, Rieko; Maitani, Tamio; Imai, Kazuhiro; Nishimura, Waka; Ito, Katsutoshi; Maeda, Masako

    2004-03-01

    This paper develops a mathematical model for describing the within-plate variation as the RSD (relative standard deviation) of absorbance measurements in a wide concentration range in competitive ELISA and proposes a method for determining the limit of detection (LOD) and range of quantitation (ROQ). The ELISA for 17 alpha-hydroxyprogesterone is taken as an example. The theoretical RSD description involves analyte concentration as an independent variable and error sources as parameters which concern the pipetting and absorbance measurement. Our model can dispense with repeated experiments of real samples, but the error parameters should be determined experimentally. The theory is in good agreement with the experiments. The most influential error sources at low and high sample concentrations are shown to be the pipetting of a viscous solution of antiserum and the absorbance inherent to the wells of a plate, respectively. The LOD and ROQ are defined as the concentration with 30% RSD and the region with <10% RSD, respectively, and are found in the theoretical plot of the RSD of concentration estimates vs concentration.

  6. Utilization of triangle nanosilver to prepare spherical nanosilver and quantitatively detect trace titanium by SERS

    NASA Astrophysics Data System (ADS)

    Liu, Qingye; Wen, Guiqing; Zhang, Xinghui; Liang, Aihui; Jiang, Zhiliang

    2014-12-01

    The blue triangle nanosilver (BAgNP) sol was prepared by the two reducers of NaBH4 and H2O2. Using BAgNP as the precursor, a small spherical nanosilver (AgNP) sol in yellow was synthesized by addition of suitable amounts of X - ( X = Cl, Br, and I). The oxidization process of BAgNP to AgNP was studied in detail by resonance Rayleigh scattering (RRS), surface-enhanced Raman scattering (SERS), laser scattering, surface plasmon resonance (SPR) absorption, and microscope techniques. It has been observed that NaCl accelerated the oxidizing BAgNP to form AgNP, and an oxidizing mechanism and quasi-nanograting Raman-scattering enhanced mechanism were developed to explain the phenomena. Using the BAgNP sol as substrate and based on the catalysis of Ti(IV) on the BrO3 - oxidizing safranine T (ST) molecular probe with a strong SERS peak at 1,535 cm-1, a new catalytic SERS quantitative method was developed for the determination of 1.0 to 100 ng/mL Ti, with a detection limit of 0.4 ng/mL.

  7. Utilization of triangle nanosilver to prepare spherical nanosilver and quantitatively detect trace titanium by SERS.

    PubMed

    Liu, Qingye; Wen, Guiqing; Zhang, Xinghui; Liang, Aihui; Jiang, Zhiliang

    2014-01-01

    The blue triangle nanosilver (BAgNP) sol was prepared by the two reducers of NaBH4 and H2O2. Using BAgNP as the precursor, a small spherical nanosilver (AgNP) sol in yellow was synthesized by addition of suitable amounts of X (-) (X = Cl, Br, and I). The oxidization process of BAgNP to AgNP was studied in detail by resonance Rayleigh scattering (RRS), surface-enhanced Raman scattering (SERS), laser scattering, surface plasmon resonance (SPR) absorption, and microscope techniques. It has been observed that NaCl accelerated the oxidizing BAgNP to form AgNP, and an oxidizing mechanism and quasi-nanograting Raman-scattering enhanced mechanism were developed to explain the phenomena. Using the BAgNP sol as substrate and based on the catalysis of Ti(IV) on the BrO3 (-) oxidizing safranine T (ST) molecular probe with a strong SERS peak at 1,535 cm(-1), a new catalytic SERS quantitative method was developed for the determination of 1.0 to 100 ng/mL Ti, with a detection limit of 0.4 ng/mL.

  8. An electronic tongue for gliadins semi-quantitative detection in foodstuffs.

    PubMed

    Peres, António M; Dias, Luís G; Veloso, Ana C A; Meirinho, Sofia G; Morais, Jorge Sá; Machado, Adélio A S C

    2011-01-15

    An all-solid-state potentiometric electronic tongue with 36 polymeric membranes has been used for the first time to detect gliadins, which are primarily responsible for gluten intolerance in people suffering from celiac disease. A linear discriminant model, based on the signals of 11 polymeric membranes, selected from the 36 above using a stepwise procedure, was used to semi-quantitatively classify samples of a "Gluten-free" foodstuff (baby milked flour), previously contaminated with known amounts of gliadins (<10, 20-50 or >50mg/kg), as "Gluten-free", "Low-Gluten content" or "Gluten-containing". For this food matrix, the device had sensitivity towards gliadins of 1-2mg/kg and overall sensitivity and specificity of 77% and 78%, respectively. Moreover, the device never identified an ethanolic extract containing gliadins as "Gluten-free". Finally, the system also allowed distinguishing "Gluten-free" and "Gluten-containing" foodstuffs (15 foods, including breads, flours, baby milked flours, cookies and breakfast cereals) with an overall sensitivity and specificity greater than 83%, using the signals of only 4 selected polymeric membranes (selected using a stepwise procedure). Since only one "Gluten-containing" foodstuff was misclassified as "Gluten-free", the device could be used as a preliminary tool for quality control of foods for celiac patients.

  9. Sequence analysis and quantitative detection of Norwalk-like viruses in cultured oysters of China

    NASA Astrophysics Data System (ADS)

    Wang, Jun; Tang, Qingjuan; Yue, Zhiqin; Li, Zhaojie; Zhang, Jin; Xue, Changhu

    2008-05-01

    We isolated 4 Norwalk-like viruses (NLVs) contaminated oysters from 33 Chinese oysters collected from local commercial sources of Shandong Province. After amplification of the RNA-dependent RNA polymerase (RdRp) region of NLVs genomes with RT-PCR, the open reading frame 1 (ORF1) of the RdRp was sequenced and subjected to multiple-sequence alignment. The results showed that NLVs in the four isolates belong to genogroup II. The sequence comparison showed that the similarity between four Chinese oyster isolates were higher than 99.0%, which indicated that NLVs prevalent in close areas have high homogeneity in genome sequences. In addition, the most conserved sequences between diverse NLVs were used to design primers and TaqMan probes, then the real-time quantitative PCR assay was performed. According to the standard curve of GII NLVs, the original amounts (copies) of NLVs in positive patient’s fecal isolate, positive Japanese oyster isolate, and the Chinese oyster isolate were 8.9×108, 1.25×108 and 4.7×101 respectively. The detecting limit of NLVs was 1×101 copies. This study will be helpful for routine diagnosis of NLVs pathogens in foods and thus for avoiding food poisoning in the future.

  10. Fecal /sup 13/C analysis for the detection and quantitation of intestinal malabsorption

    SciTech Connect

    Schoeller, D.A.; Klein, P.D.; MacLean, W.C. Jr.; Watkins, J.B.; Van Santen, E.

    1981-03-01

    The use of /sup 14/CO/sub 2/ breath tests and fecal analyses for the detection and quantitation of intestinal malabsorption has been extensively documented in adult subjects. The use of stable isotopes has extended the range of breath test applications to include pediatric and obstetric subjects. Here we report a fecal /sup 13/C analysis that can be used in conjunction with /sup 13/CO/sub 2/ breath tests. Twenty-four-hour fecal samples were collected before and after the administration of a labeled substrate. The samples were homogenized and combusted to CO/sub 2/, and the /sup 13/C abundance was determined by high-precision, differential isotope ratio mass spectrometry. The isotopic variation between successive 24 hr fecal samples was 0.6 per thousand (0.0006 atom percent). This variation limited the sensitivity of the fecal analysis to 13 ..mu..mol of /sup 13/C label per mole of fecal carbon. Simultaneous cholyglycine /sup 13/CO/sub 2/ breath tests and fecal assays were performed in five children. One child with bacterial overgrowth had an abnormal breath test and a normal fecal test. Of three children with ileal dysfunction, only one had an abnormal breath test, whereas the fecal test was abnormal in all three. Both the breath test and fecal test were abnormal for a child who had undergone an ileal resection. Both tests were normal for a child with ulcerative colitis.

  11. Detecting outlier peptides in quantitative high-throughput mass spectrometry data.

    PubMed

    Erhard, Florian; Zimmer, Ralf

    2012-06-18

    Quantitative high-throughput mass spectrometry has become an established tool to measure relative gene expression proteome-wide. The output of such an experiment usually consists of a list of expression ratios (fold changes) for several thousand proteins between two conditions. However, we observed that individual peptide fold changes may show a significantly different behavior than other peptides from the same protein and that these differences cannot be explained by imprecise measurements. Such outlier peptides can be the consequence of several technical (misidentifications, misquantifications) or biological (post-translational modifications, differential regulation of isoforms) reasons. We developed a method to detect outlier peptides in mass spectrometry data which is able to delineate imprecise measurements from real outlier peptides with high accuracy when the true difference is as small as 1.4 fold. We applied our method to experimental data and investigated the different technical and biological effects that result in outlier peptides. Our method will assist future research to reduce technical bias and can help to identify genes with differentially regulated protein isoforms in high throughput mass spectrometry data. PMID:22483996

  12. Immunological Detection and Quantitation of Tryptophan Decarboxylase in Developing Catharanthus roseus Seedlings 1

    PubMed Central

    Fernandez, Jesus Alvarez; Owen, Terence G.; Kurz, Wolfgang G. W.; De Luca, Vincenzo

    1989-01-01

    l-Tryptophan decarboxylase (TDC) (EC 4.2.1.27) enzyme activity was induced in cell suspension cultures of Catharanthus roseus after treatment with a Pythium aphanidermatum elicitor preparation. The enzyme was extracted from lyophilized cells containing high levels of TDC and the protein was purified to homogeneity. The pure protein was used to produce highly specific polyclonal antibodies, and an enzyme-linked immunosorbent assay (ELISA) was developed to quantitate the level of TDC antigen during seedling development and in leaves of the mature plant. Western immunoblotting of proteins after SDS-PAGE with anti-TDC antibodies detected several immunoreactive proteins (40, 44, 54.8, 55, and 67 kilodaltons) which appeared at different stages during seedling development and in leaves of the mature plant. The major 54.8 and 55 kilodalton antigenic proteins in immunoblots appeared transiently between days 1 to 5 and 5 to 8 of seedling development, respectively. The 54.8 kilodalton protein was devoid of TDC enzyme activity, whereas the appearance of the 55 kilodalton protein coincided with the appearance of this decarboxylase activity. The minor immunoreactive proteins (40, 44, and 67 kilodaltons) appeared after day 5 of seedling development and in older leaves of the mature plant, and their relationship, if any, to TDC is presently unknown. Results suggest that the synthesis and degradation of TDC protein is highly regulated in Catharanthus roseus and that this regulation follows a preset developmental program. Images Figure 3 Figure 5 PMID:16667047

  13. SERS-based quantitative detection of ovarian cancer prognostic factor haptoglobin

    PubMed Central

    Perumal, Jayakumar; Balasundaram, Ghayathri; Mahyuddin, Aniza P; Choolani, Mahesh; Olivo, Malini

    2015-01-01

    Surface-enhanced Raman spectroscopy (SERS) is increasingly being used for biosensing because of its high sensitivity and low detection limit, which are made possible by the unique Raman ‘fingerprint’ spectra from the biomolecules. Here we propose a novel SERS method for the fast, sensitive, and reliable quantitative analysis of haptoglobin (Hp), an acute phase plasma glycoprotein that is widely gaining application as a prognostic ovarian cancer biomarker. We exploited the peroxidase activity of the hemoglobin–haptoglobin (Hb–Hp) complex formed by the selective and specific binding of Hp to free Hb to catalyze the reaction of 3,3′,5,5′-tetramethylbenzidine (TMB) substrate and hydrogen peroxide to result in the final product of strongly SERS-active TMB2+. We observed a linear increase in the SERS signal of TMB2+ with increasing concentrations of Hb–Hp complex from 50 nM to 34 μM. Based on this concentration-dependent SERS spectrum, we quantified Hp in clinical samples. We observed that our inference about the prognosis of the disease coincided with the histology data and that our method was much more sensitive than the enzyme-linked immunosorbent assay method. PMID:25834423

  14. Iridium-191 angiocardiography for the detection and quantitation of left-to-right shunting

    SciTech Connect

    Treves, S.; Cheng, C.; Samuel, A.; Lambrecht, R.; Babchyck, B.; Zimmerman, R.; Norwood, W.

    1980-12-01

    An osmium-191 ..-->.. iridium-191 generator that can deliver multiple doses of Ir-191m for first-pass radionuclide angiography has been developed. Iridium-191m has a physical half-life of 4.96 sec and decays with emission of 65-keV and 129-keV photons in 58 and 30% abundance, respectively. Using a gamma camera, Ir-191m radionuclide angiography was carried out, in dogs and ten patients, for the detection and quantitation of left-to-right shunting. In a one-year-old patient, 25 mCi of Ir-191m results in a whole-body radiation absorbed dose of 35 mrad. Multiple Ir-191m angiograms can be performed, seconds to minutes apart, without interference from background. The 15.4-day half-life of Os-191 permits transportation of the generator to centers far from the production facility. With the low radiation dose, high information density, and the ability to repeat studies with Ir-191m, clinical use of radionuclide angiography should be expanded.

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

    PubMed

    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 < 0.05). Our protocol is rapid, specific, sensitive, and provides a powerful tool for quantification of O. sinensis from soil.

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

    PubMed

    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 < 0.05). Our protocol is rapid, specific, sensitive, and provides a powerful tool for quantification of O. sinensis from soil. PMID:23540339

  17. A Wavelet-Based Approach to Fall Detection

    PubMed Central

    Palmerini, Luca; Bagalà, Fabio; Zanetti, Andrea; Klenk, Jochen; Becker, Clemens; Cappello, Angelo

    2015-01-01

    Falls among older people are a widely documented public health problem. Automatic fall detection has recently gained huge importance because it could allow for the immediate communication of falls to medical assistance. The aim of this work is to present a novel wavelet-based approach to fall detection, focusing on the impact phase and using a dataset of real-world falls. Since recorded falls result in a non-stationary signal, a wavelet transform was chosen to examine fall patterns. The idea is to consider the average fall pattern as the “prototype fall”.In order to detect falls, every acceleration signal can be compared to this prototype through wavelet analysis. The similarity of the recorded signal with the prototype fall is a feature that can be used in order to determine the difference between falls and daily activities. The discriminative ability of this feature is evaluated on real-world data. It outperforms other features that are commonly used in fall detection studies, with an Area Under the Curve of 0.918. This result suggests that the proposed wavelet-based feature is promising and future studies could use this feature (in combination with others considering different fall phases) in order to improve the performance of fall detection algorithms. PMID:26007719

  18. A wavelet-based approach to fall detection.

    PubMed

    Palmerini, Luca; Bagalà, Fabio; Zanetti, Andrea; Klenk, Jochen; Becker, Clemens; Cappello, Angelo

    2015-01-01

    Falls among older people are a widely documented public health problem. Automatic fall detection has recently gained huge importance because it could allow for the immediate communication of falls to medical assistance. The aim of this work is to present a novel wavelet-based approach to fall detection, focusing on the impact phase and using a dataset of real-world falls. Since recorded falls result in a non-stationary signal, a wavelet transform was chosen to examine fall patterns. The idea is to consider the average fall pattern as the "prototype fall".In order to detect falls, every acceleration signal can be compared to this prototype through wavelet analysis. The similarity of the recorded signal with the prototype fall is a feature that can be used in order to determine the difference between falls and daily activities. The discriminative ability of this feature is evaluated on real-world data. It outperforms other features that are commonly used in fall detection studies, with an Area Under the Curve of 0.918. This result suggests that the proposed wavelet-based feature is promising and future studies could use this feature (in combination with others considering different fall phases) in order to improve the performance of fall detection algorithms. PMID:26007719

  19. Quantitative signal detection for vaccines: effects of stratification, background and masking on GlaxoSmithKline's spontaneous reports database.

    PubMed

    Zeinoun, Ziad; Seifert, Harry; Verstraeten, Thomas

    2009-09-01

    Disproportionality analyses are increasingly gaining acceptance as signal detection tools for drug adverse events. Their application to vaccine adverse events has not been well evaluated. Disproportionality analyses based on the Multi-Item Gamma Poisson Shrinkage principle (MGPS) were applied to spontaneous adverse events reports for five vaccines with different reporting profiles. Sensitivity analyses were used to assess the potential impact of changing key parameters. We used the Company's in-house spontaneous adverse events database. We evaluated the impact of stratification, the comparator dataset and the potential for masking. We conducted a semi-quantitative assessment by comparing the changes in the disproportionality scores and the number of vaccine-event pairs that exceeded an arbitrary threshold as a measure of the impact of any of these choices. The results show that stratification by age and region has a significant impact. The effect of the comparator dataset was dependent on the vaccine being evaluated. The potential for a masking effect was only weakly noticeable. In conclusion, we opt to start with a conservative approach in which we will supplement our primary stratification against the vaccine database with unstratified analyses as well as analyses against the entire database. We recommend that similar studies be performed before introducing disproportionality analyses to a new vaccine adverse events database.

  20. Manufacture and Testing of a High Field Gradient Magnetic Fractionation System for Quantitative Detection of Plasmodium falciparum Gametocytes

    NASA Astrophysics Data System (ADS)

    Karl, Stephan; Woodward, Robert C.; Davis, Timothy M. E.; St. Pierre, Tim G.

    2010-12-01

    Plasmodium falciparum is the most dangerous of the human malaria parasite species and accounts for millions of clinical episodes of malaria each year in tropical countries. The pathogenicity of Plasmodium falciparum is a result of its ability to infect erythrocytes where it multiplies asexually over 48 h or develops into sexual forms known as gametocytes. If sufficient male and female gametocytes are taken up by a mosquito vector, it becomes infectious. Therefore, the presence and density of gametocytes in human blood is an important indicator of human-to-mosquito transmission of malaria. Recently, we have shown that high field gradient magnetic fractionation improves gametocyte detection in human blood samples. Here we present two important new developments. Firstly we introduce a quantitative approach to replace the previous qualitative method and, secondly, we describe a novel method that enables cost-effective production of the magnetic fractionation equipment required to carry out gametocyte quantification. We show that our custom-made magnetic fractionation equipment can deliver results with similar sensitivity and convenience but for a small fraction of the cost.

  1. Quantitative detection of enteroviruses in activated sludge by cell culture and real-time RT-PCR using paramagnetic capturing.

    PubMed

    Pusch, D; Ihle, St; Lebuhn, M; Graeber, I; López-Pila, J M

    2005-09-01

    We have compared in extracts of activated sludge the number of enteroviruses detectable with buffalo green monkey (BGM) cell-cultures versus the number of enteroviral genomes determined by reverse-transcription quantitative real-time PCR (RT-qPCR). In order to find conditions adequate for quantifying enteroviral RNA isolated from (waste)water we have investigated affinity capture of RNA with polystyrene beads (Dynabeads). The capture efficiency strongly depended on the genomic region chosen for the affinity binding. Capture of the RNA by its 3'-tail was most efficient (almost 100%); other regions within the genome yielded variable but lower results. Indirect capture (first hybridization of the RNA to the oligonucleotides, then attachment of the duplex molecules to the beads) was much more efficient than direct capture (attachment of the oligonucleotides to the beads first, then binding of the RNA), and resulted in RNA capture of maximally 60-80%. At least partly, this was due to incomplete hybridization of the RNA to the complementary oligonucleotides. No correlation was found between the number of cytopathic effects (CPE) determined by cell culture and the number of genomes quantified by RT-qPCR; RT-qPCR values were consistently much higher than the number of CPE. This points to overestimation of infectious enteroviruses by RT-qPCR and/or underestimation by the cell culture approach.

  2. Cytometric Approach for Detection of Encephalitozoon intestinalis, an Emergent Agent▿

    PubMed Central

    Barbosa, Joana; Rodrigues, Acácio Gonçalves; Pina-Vaz, Cidália

    2009-01-01

    Encephalitozoon intestinalis is responsible for intestinal disease in patients with AIDS and immunocompetent patients. The infectious form is a small spore that is resistant to water treatment procedures. Its detection is very important, but detection is very cumbersome and time-consuming. Our main objective was to develop and optimize a specific flow cytometric (FC) protocol for the detection of E. intestinalis in hospital tap water and human feces. To determine the optimal specific antibody (Microspor-FA) concentration, a known concentration of E. intestinalis spores (Waterborne, Inc.) was suspended in hospital tap water and stool specimens with different concentrations of Microspor-FA, and the tap water and stool specimens were incubated under different conditions. The sensitivity limit and specificity were also evaluated. To study spore infectivity, double staining with propidium iodide (PI) and Microspor-FA was undertaken. Distinct approaches for filtration and centrifugation of the stool specimens were used. E. intestinalis spores stained with 10 μg/ml of Microspor-FA at 25°C overnight provided the best results. The detection limit was 5 × 104 spores/ml, and good specificity was demonstrated. Simultaneous staining with Microspor-FA and PI ensured that the E. intestinalis spores were dead and therefore noninfectious. With the stool specimens, better spore recovery was observed with a saturated solution of NaCl and centrifugation at 1,500 × g for 15 min. A new approach for the detection of E. intestinalis from tap water or human feces that ensures that the spores are not viable is now available and represents an important step for the prevention of this threat to public health. PMID:19439525

  3. Enhanced Signal and Quantitative Detection of Anti-Interferon-Gamma Antibody by Using a Nanometer Biolinker

    PubMed Central

    Tsai, Pei-I; Lee, Adam Shih-Yuan; Lee, Shu-Sheng; Chung, Ming-Han; Liu, Meng-Wei; Lee, Chih-Kung

    2016-01-01

    For rapid screening and quantification of an antisera antibody, a nanometer bithiophene-based conductive biolinker can enhanced signal performance and can be used to verify the interaction of an anti-IFN-γ antibody with an IFN-γ protein. The experimental measurements take a generic approach which takes advantage of the functionality of thiophene-based linkers for biosensors. Effects associated with using bithiophene as a biolinker for surface plasmon resonance (SPR) spectroscopy are examined in this paper. By using an atomic force microscope (AFM), it was observed that the morphology of the bithiophene modified gold sensor surface became smoother than the original gold surface. We compared the response and concentration of the anti-IFN-γ antibody on a bithiophene-coated and dextran-coated biochip as well as on different thickness-modified surfaces under SPR relevant conditions. The results indicate that a response to IFN-γ molecules immobilized on a sensor using a bithiophene biolinker improved more than 8-fold when compared to that of a sensor using a dextran biolinker. Furthermore, the regeneration ability of the sensor surface shows good repeatability as only less than a 1% decrease was found after repeating the experimental work over 6 cycles. The characteristics provided us with a good platform for rapid screening, real-time monitoring and quantitative concentration of the autoimmune antibody activities. PMID:27459633

  4. Genetic variation detected by quantitative analysis of end-labeled genomic DNA fragments

    SciTech Connect

    Asakawa, Jun-Ichi; Kodaira, Mieko; Satoh, Chiyoko; Kuick, R.; Hanash, S.M.; Neel, J.V.

    1994-09-13

    The continuing efforts to evaluate specific human populations for altered germinal mutation rates would profit from more efficient and more specific approaches than those of the past. To this end, the authors have explored the potential usefulness of two-dimensional electrophoresis of DNA fragments obtained from restriction-enzyme-digested genomic DNA. This permits the analysis, on a single preparation, of {approx} 2000 DNA fragments varying in size from 1.0 to 5.0 kb in the first dimension and from 0.3 to 2.0 kb in the second dimension. To enter into a genetic analysis, these fragments must exhibit positional and quantitative stability. With respect to the latter, if spots are the product of only one fragment, the coefficient of variation of spot intensity should be approximately {le} 0.12. At present, 482 of the spots in preparations meet these standards. In an examination of preparations based on three Japanese mother/father/child trios, 43 of these 482 spots were found to exhibit variation that segregated within families according to mendelian principles. The authors have established the feasibility of cloning a variant fragment from such gels and establishing its nucleotide sequence. This technology should be highly efficient in monitoring for mutations resulting in loss/gain/rearrangement events in DNA fragments distributed throughout the genome. 17 refs., 2 figs., 2 tabs.

  5. Quantitative metabolic imaging using endogenous fluorescence to detect stem cell differentiation

    NASA Astrophysics Data System (ADS)

    Quinn, Kyle P.; Sridharan, Gautham V.; Hayden, Rebecca S.; Kaplan, David L.; Lee, Kyongbum; Georgakoudi, Irene

    2013-12-01

    The non-invasive high-resolution spatial mapping of cell metabolism within tissues could provide substantial advancements in assessing the efficacy of stem cell therapy and understanding tissue development. Here, using two-photon excited fluorescence microscopy, we elucidate the relationships among endogenous cell fluorescence, cell redox state, and the differentiation of human mesenchymal stem cells into adipogenic and osteoblastic lineages. Using liquid chromatography/mass spectrometry and quantitative PCR, we evaluate the sensitivity of an optical redox ratio of FAD/(NADH + FAD) to metabolic changes associated with stem cell differentiation. Furthermore, we probe the underlying physiological mechanisms, which relate a decrease in the redox ratio to the onset of differentiation. Because traditional assessments of stem cells and engineered tissues are destructive, time consuming, and logistically intensive, the development and validation of a non-invasive, label-free approach to defining the spatiotemporal patterns of cell differentiation can offer a powerful tool for rapid, high-content characterization of cell and tissue cultures.

  6. Quantitative detection of syntrophic fatty acid-degrading bacterial communities in methanogenic environments.

    PubMed

    Mathai, Prince P; Zitomer, Daniel H; Maki, James S

    2015-06-01

    In methanogenic habitats, volatile fatty acids (VFA), such as propionate and butyrate, are major intermediates in organic matter degradation. VFA are further metabolized to H(2), acetate and CO(2) by syntrophic fatty acid-degrading bacteria (SFAB) in association with methanogenic archaea. Despite their indispensable role in VFA degradation, little is known about SFAB abundance and their environmental distribution. To facilitate ecological studies, we developed four novel genus-specific quantitative PCR (qPCR) assays, with primer sets targeting known SFAB: Syntrophobacter, Smithella, Pelotomaculum and Syntrophomonas. Primer set specificity was confirmed using in silico and experimental (target controls, clone libraries and melt-curve analysis) approaches. These qPCR assays were applied to quantify SFAB in a variety of mesophilic methanogenic habitats, including a laboratory propionate enrichment culture, pilot- and full-scale anaerobic reactors, cow rumen, horse faeces, an experimental rice paddy soil, a bog stream and swamp sediments. The highest SFAB 16S rRNA gene copy numbers were found in the propionate enrichment culture and anaerobic reactors, followed by the bog stream and swamp sediment samples. In addition, it was observed that SFAB and methanogen abundance varied with reactor configuration and substrate identity. To our knowledge, this research represents the first comprehensive study to quantify SFAB in methanogenic habitats using qPCR-based methods. These molecular tools will help investigators better understand syntrophic microbial communities in engineered and natural environments.

  7. Real-time quantitative PCR detection of WT1 gene expression in children with AML: prognostic significance, correlation with disease status and residual disease detection by flow cytometry.

    PubMed

    Trka, J; Kalinová, M; Hrusák, O; Zuna, J; Krejcí, O; Madzo, J; Sedlácek, P; Vávra, V; Michalová, K; Jarosová, M; Starý, J

    2002-07-01

    The clinical significance of WT1 gene expression at diagnosis and during therapy of AML has not yet been resolved. We analysed WT1 expression at presentation in an unselected group of 47 childhood AML patients using real-time quantitative reverse-transcription PCR. We also showed that within the first 30 h following aspiration RQ-RT-PCR results were not influenced by transportation time. We observed lower levels of WT1 transcript in AML M5 (P = 0.0015); no association was found between expression levels and sex, initial leukocyte count and karyotype-based prognostic groups. There was significant correlation between very low WT1 expression at presentation and excellent outcome (EFS P = 0.0014). Combined analysis of WT1 levels, three-colour flow cytometry residual disease detection and the course of the disease in 222 samples from 28 children with AML showed remarkable correlation. Fourteen patients expressed high WT1 levels at presentation. In eight of them, who suffered relapse or did not reach complete remission, dynamics of WT1 levels clearly correlated with the disease status and residual disease by flow cytometry. We conclude that very low WT1 levels at presentation represent a good prognostic factor and that RQ-RT-PCR-based analysis of WT1 expression is a promising and rapid approach for monitoring of MRD in approximately half of paediatric AML patients.

  8. Quantitative real time polymerase chain reaction assays for the sensitive detection of Besnoitia besnoiti infection in cattle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bovine besnoitiosis, an economically important disease in cattle in many countries of Africa and Asia, has re-emerged in Europe. Sensitive and quantitative DNA detection methods are needed to determine whether serologically positive animals are infectious and to examine the role of vectors (e.g. ha...

  9. Single Laboratory Comparison of Quantitative Real-Time PCR Assays for the Detection of Human Fecal Pollution

    EPA Science Inventory

    There are numerous quantitative real-time PCR (qPCR) methods available to detect and enumerate human fecal pollution in ambient waters. Each assay employs distinct primers and/or probes and many target different genes and microorganisms leading to potential variations in method ...

  10. Single Laboratory Comparison of Quantitative Real-Time PCR Assays for the Detection of Human Fecal Pollution - Poster

    EPA Science Inventory

    There are numerous quantitative real-time PCR (qPCR) methods available to detect and enumerate human fecal pollution in ambient waters. Each assay employs distinct primers and/or probes and many target different genes and microorganisms leading to potential variations in method p...

  11. EVALUATION OF RAPID DNA EXTRACTION PROCEDURES FOR THE QUANTITATIVE DETECTION OF FUNGAL CELLS USING REAL TIME PCR ANALYSIS

    EPA Science Inventory

    The ease and rapidity of quantitative DNA sequence detection by real-time PCR instruments promises to make their use increasingly common for the microbial analysis many different types of environmental samples. To fully exploit the capabilities of these instruments, correspondin...

  12. Quantitative detection of perchlorate-reducing bacteria by real-time PCR targeting the perchlorate reductase gene.

    PubMed

    Nozawa-Inoue, Mamie; Jien, Mercy; Hamilton, Nicholas S; Stewart, Valley; Scow, Kate M; Hristova, Krassimira R

    2008-03-01

    A quantitative real-time PCR assay targeting the pcrA gene, encoding the catalytic subunit of perchlorate reductase, detected pcrA genes from perchlorate-reducing bacteria in three different genera and from soil microbial communities. Partial pcrA sequences indicated differences in the composition of perchlorate-reducing bacterial communities following exposure to different electron donors.

  13. Detection and quantitation of Solenopsis invicta virus-2 genomic and intermediary replicating viral RNA in fire ant workers and larvae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A quantitative real-time PCR (QPCR) method was developed to detect and quantify Solenopsis invicta virus-2 (SINV-2) infecting individual ants of Solenopsis invicta. The two-step method utilized a gene-specific oligonucleotide primer targeting an upstream region of the SINV-2 conserved domain of RNA...

  14. Early detection of Brucella canis via quantitative polymerase chain reaction analysis.

    PubMed

    Kauffman, L K; Bjork, J K; Gallup, J M; Boggiatto, P M; Bellaire, B H; Petersen, C A

    2014-02-01

    Canine brucellosis is a reportable zoonotic disease that can lead to canine reproductive losses and human infection through contact with infected urine or other genitourinary secretions. Although many locations require testing and euthanasia of positive dogs, current diagnosis is limited by the time required for seroconversion, for example, presence of B. canis-specific antibodies. The goal of this study was to determine the diagnostic ability of Brucella canis-specific quantitative polymerase chain reaction (qPCR) assay to detect B. canis in field samples prior to serological positivity for faster diagnosis and prevention of transmission within kennels or in households. Two kennels, one of which was located in the owner's home, were sampled following observation of suggestive clinical signs and positive serology of at least one dog. Specimens obtained were comparatively analysed via serology and qPCR analysis. 107 dogs were analysed for B. canis infection via qPCR: 105 via whole-blood samples, 65 via vaginal swab, six via urine and seven via genitourinary tract tissue taken at necropsy. Forty-five dogs were found to be infected with canine brucellosis via qPCR, of which 22 (48.89%) were seropositive. A statistically significant number (P = 0.0228) of qPCR-positive dogs, 5/25 (20.00%), seroconverted within a 30-day interval after initial serologic testing. As compared to serology, qPCR analysis of DNA from vaginal swabs had a sensitivity of 92.31% and specificity of 51.92%, and qPCR analysis of DNA from whole-blood samples had a sensitivity of 16.67% and specificity of 100%. B. canis outer membrane protein 25 DNA qPCR from non-invasive vaginal swab and urine samples provided early detection of B. canis infection in dogs prior to detection of antibodies. This assay provides a critical tool to decrease zoonotic spread of canine brucellosis, its associated clinical presentation(s), and emotional and economic repercussions.

  15. Outlier Detection In Linear Regression Using Standart Parity Space Approach

    NASA Astrophysics Data System (ADS)

    Mustafa Durdag, Utkan; Hekimoglu, Serif

    2013-04-01

    Despite all technological advancements, outliers may occur due to some mistakes in engineering measurements. Before estimation of unknown parameters, aforementioned outliers must be detected and removed from the measurements. There are two main outlier detection methods: the conventional tests based on least square approach (e.g. Baarda, Pope etc.) and the robust tests (e.g. Huber, Hampel etc.) are used to identify outliers in a set of measurement. Standart Parity Space Approach is one of the important model-based Fault Detection and Isolation (FDI) technique that usually uses in Control Engineering. In this study the standart parity space method is used for outlier detection in linear regression. Our main goal is to compare success of two approaches of standart parity space method and conventional tests in linear regression through the Monte Carlo simulation with each other. The least square estimation is the most common estimator as known and it minimizes the sum of squared residuals. In standart parity space approach to eliminate unknown vector, the measurement vector projected onto the left null space of the coefficient matrix. Thus, the orthogonal condition of parity vector is satisfied and only the effects of noise vector noticed. The residual vector is derived from two cases that one is absence of an outlier; the other is occurrence of an outlier. Its likelihood function is used for determining the detection decision function for global Test. Localization decision function is calculated for each column of parity matrix and the maximum one of these values is accepted as an outlier. There are some results obtained from two different intervals that one of them is between 3σ and 6σ (small outlier) the other one is between 6σ and 12σ (large outlier) for outlier generator when the number of unknown parameter is chosen 2 and 3. The measure success rates (MSR) of Baarda's method is better than the standart parity space method when the confidence intervals are

  16. A novel hybridization approach for detection of citrus viroids.

    PubMed

    Murcia, N; Serra, P; Olmos, A; Duran-Vila, N

    2009-04-01

    Citrus plants are natural hosts of several viroid species all belonging to the family Pospiviroidae. Previous attempts to detect viroids from field-grown species and cultivars yielded erratic results unless analyses were performed using Etrog citron a secondary bio-amplification host. To overcome the use of Etrog citron a number of RT-PCR approaches have been proposed with different degrees of success. Here we report the suitability of an easy to handle northern hybridization protocol for viroid detection of samples collected from field-grown citrus species and cultivars. The protocol involves: (i) Nucleic acid preparations from bark tissue samples collected from field-grown trees regardless of the growing season and storage conditions; (ii) Separation in 5% PAGE or 1% agarose, blotting to membrane and fixing; (iii) Hybridization with viroid-specific DIG-labelled probes and detection with anti-DIG-alkaline phosphatase conjugate and autoradiography with the CSPD substrate. The method has been tested with viroid-infected trees of sweet orange, lemon, mandarin, grapefruit, sour orange, Swingle citrumello, Tahiti lime and Mexican lime. This novel hybridization approach is extremely sensitive, easy to handle and shortens the time needed for reliable viroid indexing tests. The suitability of PCR generated DIG-labelled probes and the sensitivity achieved when the samples are separated and blotted from non-denaturing gels are discussed.

  17. New viruses in veterinary medicine, detected by metagenomic approaches.

    PubMed

    Belák, Sándor; Karlsson, Oskar E; Blomström, Anne-Lie; Berg, Mikael; Granberg, Fredrik

    2013-07-26

    In our world, which is faced today with exceptional environmental changes and dramatically intensifying globalisation, we are encountering challenges due to many new factors, including the emergence or re-emergence of novel, so far "unknown" infectious diseases. Although a broad arsenal of diagnostic methods is at our disposal, the majority of the conventional diagnostic tests is highly virus-specific or is targeted entirely towards a limited group of infectious agents. This specificity complicates or even hinders the detection of new or unexpected pathogens, such as new, emerging or re-emerging viruses or novel viral variants. The recently developed approaches of viral metagenomics provide an effective novel way to screen samples and detect viruses without previous knowledge of the infectious agent, thereby enabling a better diagnosis and disease control, in line with the "One World, One Health" principles (www.oneworldonehealth.org). Using metagenomic approaches, we have recently identified a broad variety of new viruses, such as novel bocaviruses, Torque Teno viruses, astroviruses, rotaviruses and kobuviruses in porcine disease syndromes, new virus variants in honeybee populations, as well as a range of other infectious agents in further host species. These findings indicate that the metagenomic detection of viral pathogens is becoming now a powerful, cultivation-independent, and useful novel diagnostic tool in veterinary diagnostic virology. PMID:23428379

  18. New viruses in veterinary medicine, detected by metagenomic approaches.

    PubMed

    Belák, Sándor; Karlsson, Oskar E; Blomström, Anne-Lie; Berg, Mikael; Granberg, Fredrik

    2013-07-26

    In our world, which is faced today with exceptional environmental changes and dramatically intensifying globalisation, we are encountering challenges due to many new factors, including the emergence or re-emergence of novel, so far "unknown" infectious diseases. Although a broad arsenal of diagnostic methods is at our disposal, the majority of the conventional diagnostic tests is highly virus-specific or is targeted entirely towards a limited group of infectious agents. This specificity complicates or even hinders the detection of new or unexpected pathogens, such as new, emerging or re-emerging viruses or novel viral variants. The recently developed approaches of viral metagenomics provide an effective novel way to screen samples and detect viruses without previous knowledge of the infectious agent, thereby enabling a better diagnosis and disease control, in line with the "One World, One Health" principles (www.oneworldonehealth.org). Using metagenomic approaches, we have recently identified a broad variety of new viruses, such as novel bocaviruses, Torque Teno viruses, astroviruses, rotaviruses and kobuviruses in porcine disease syndromes, new virus variants in honeybee populations, as well as a range of other infectious agents in further host species. These findings indicate that the metagenomic detection of viral pathogens is becoming now a powerful, cultivation-independent, and useful novel diagnostic tool in veterinary diagnostic virology.

  19. Whole-genome scan to detect quantitative trait loci associated with milk protein composition in 3 French dairy cattle breeds.

    PubMed

    Sanchez, M P; Govignon-Gion, A; Ferrand, M; Gelé, M; Pourchet, D; Amigues, Y; Fritz, S; Boussaha, M; Capitan, A; Rocha, D; Miranda, G; Martin, P; Brochard, M; Boichard, D

    2016-10-01

    In the context of the PhénoFinLait project, a genome-wide analysis was performed to detect quantitative trait loci (QTL) that affect milk protein composition estimated using mid-infrared spectrometry in the Montbéliarde (MO), Normande (NO), and Holstein (HO) French dairy cattle breeds. The 6 main milk proteins (α-lactalbumin, β-lactoglobulin, and αS1-, αS2-, β-, and κ-caseins) expressed as grams per 100g of milk (% of milk) or as grams per 100g of protein (% of protein) were estimated in 848,068 test-day milk samples from 156,660 cows. Genotyping was performed for 2,773 MO, 2,673 NO, and 2,208 HO cows using the Illumina BovineSNP50 BeadChip (Illumina Inc., San Diego, CA). Individual test-day records were adjusted for environmental effects and then averaged per cow to define the phenotypes analyzed. Quantitative trait loci detection was performed within each breed using a linkage disequilibrium and linkage analysis approach. A total of 39 genomic regions distributed on 20 of the 29 Bos taurus autosomes (BTA) were significantly associated with milk protein composition at a genome-wide level of significance in at least 1 of the 3 breeds. The 9 most significant QTL were located on BTA2 (133 Mbp), BTA6 (38, 47, and 87 Mbp), BTA11 (103 Mbp), BTA14 (1.8 Mbp), BTA20 (32 and 58 Mbp), and BTA29 (8 Mbp). The BTA6 (87 Mbp), BTA11, and BTA20 (58 Mbp) QTL were found in all 3 breeds, and they had highly significant effects on κ-casein, β-lactoglobulin, and α-lactalbumin, expressed as a percentage of protein, respectively. Each of these QTL explained between 13% (BTA14) and 51% (BTA11) of the genetic variance of the trait. Many other QTL regions were also identified in at least one breed. They were located on 14 additional chromosomes (1, 3, 4, 5, 7, 15, 17, 19, 21, 22, 24, 25, 26, and 27), and they explained 2 to 8% of the genetic variance of 1 or more protein composition traits. Concordance analyses, performed between QTL status and sequence-derived polymorphisms from

  20. Whole-genome scan to detect quantitative trait loci associated with milk protein composition in 3 French dairy cattle breeds.

    PubMed

    Sanchez, M P; Govignon-Gion, A; Ferrand, M; Gelé, M; Pourchet, D; Amigues, Y; Fritz, S; Boussaha, M; Capitan, A; Rocha, D; Miranda, G; Martin, P; Brochard, M; Boichard, D

    2016-10-01

    In the context of the PhénoFinLait project, a genome-wide analysis was performed to detect quantitative trait loci (QTL) that affect milk protein composition estimated using mid-infrared spectrometry in the Montbéliarde (MO), Normande (NO), and Holstein (HO) French dairy cattle breeds. The 6 main milk proteins (α-lactalbumin, β-lactoglobulin, and αS1-, αS2-, β-, and κ-caseins) expressed as grams per 100g of milk (% of milk) or as grams per 100g of protein (% of protein) were estimated in 848,068 test-day milk samples from 156,660 cows. Genotyping was performed for 2,773 MO, 2,673 NO, and 2,208 HO cows using the Illumina BovineSNP50 BeadChip (Illumina Inc., San Diego, CA). Individual test-day records were adjusted for environmental effects and then averaged per cow to define the phenotypes analyzed. Quantitative trait loci detection was performed within each breed using a linkage disequilibrium and linkage analysis approach. A total of 39 genomic regions distributed on 20 of the 29 Bos taurus autosomes (BTA) were significantly associated with milk protein composition at a genome-wide level of significance in at least 1 of the 3 breeds. The 9 most significant QTL were located on BTA2 (133 Mbp), BTA6 (38, 47, and 87 Mbp), BTA11 (103 Mbp), BTA14 (1.8 Mbp), BTA20 (32 and 58 Mbp), and BTA29 (8 Mbp). The BTA6 (87 Mbp), BTA11, and BTA20 (58 Mbp) QTL were found in all 3 breeds, and they had highly significant effects on κ-casein, β-lactoglobulin, and α-lactalbumin, expressed as a percentage of protein, respectively. Each of these QTL explained between 13% (BTA14) and 51% (BTA11) of the genetic variance of the trait. Many other QTL regions were also identified in at least one breed. They were located on 14 additional chromosomes (1, 3, 4, 5, 7, 15, 17, 19, 21, 22, 24, 25, 26, and 27), and they explained 2 to 8% of the genetic variance of 1 or more protein composition traits. Concordance analyses, performed between QTL status and sequence-derived polymorphisms from

  1. Quantitative detection of methanotrophs in soil by novel pmoA-targeted real-time PCR assays.

    PubMed

    Kolb, Steffen; Knief, Claudia; Stubner, Stephan; Conrad, Ralf

    2003-05-01

    Methane oxidation in soils is mostly accomplished by methanotrophic bacteria. Little is known about the abundance of methanotrophs in soils, since quantification by cultivation and microscopic techniques is cumbersome. Comparison of 16S ribosomal DNA and pmoA (alpha subunit of the particulate methane monooxygenase) phylogenetic trees showed good correlation and revealed five distinct groups of methanotrophs within the alpha and gamma subclasses of Proteobacteria: the Methylococcus group, the Methylobacter/Methylosarcina group, the Methylosinus group, the Methylocapsa group, and the forest clones group (a cluster of pmoA sequences retrieved from forest soils). We developed quantitative real-time PCR assays with SybrGreen for each of these five groups and for all methanotrophic bacteria by targeting the pmoA gene. Detection limits were between 10(1) and 10(2) target molecules per reaction for all assays. Real-time PCR analysis of soil samples spiked with cells of Methylococcus capsulatus, Methylomicrobium album, and Methylosinus trichosporium recovered almost all the added bacteria. Only the Methylosinus-specific assay recovered only 20% of added cells, possibly due to a lower lysis efficiency of type II methanotrophs. Analysis of the methanotrophic community structure in a flooded rice field soil showed (5.0 +/- 1.4) x 10(6) pmoA molecules g(-1) for all methanotrophs. The Methylosinus group was predominant (2.7 x 10(6) +/- 1.1 x 10(6) target molecules g(-1)). In addition, bacteria of the Methylobacter/Methylosarcina