WorldWideScience

Sample records for high throughput optimization

  1. Optimization and high-throughput screening of antimicrobial peptides.

    Science.gov (United States)

    Blondelle, Sylvie E; Lohner, Karl

    2010-01-01

    While a well-established process for lead compound discovery in for-profit companies, high-throughput screening is becoming more popular in basic and applied research settings in academia. The development of combinatorial libraries combined with easy and less expensive access to new technologies have greatly contributed to the implementation of high-throughput screening in academic laboratories. While such techniques were earlier applied to simple assays involving single targets or based on binding affinity, they have now been extended to more complex systems such as whole cell-based assays. In particular, the urgent need for new antimicrobial compounds that would overcome the rapid rise of drug-resistant microorganisms, where multiple target assays or cell-based assays are often required, has forced scientists to focus onto high-throughput technologies. Based on their existence in natural host defense systems and their different mode of action relative to commercial antibiotics, antimicrobial peptides represent a new hope in discovering novel antibiotics against multi-resistant bacteria. The ease of generating peptide libraries in different formats has allowed a rapid adaptation of high-throughput assays to the search for novel antimicrobial peptides. Similarly, the availability nowadays of high-quantity and high-quality antimicrobial peptide data has permitted the development of predictive algorithms to facilitate the optimization process. This review summarizes the various library formats that lead to de novo antimicrobial peptide sequences as well as the latest structural knowledge and optimization processes aimed at improving the peptides selectivity.

  2. Optimizing transformations for automated, high throughput analysis of flow cytometry data.

    Science.gov (United States)

    Finak, Greg; Perez, Juan-Manuel; Weng, Andrew; Gottardo, Raphael

    2010-11-04

    In a high throughput setting, effective flow cytometry data analysis depends heavily on proper data preprocessing. While usual preprocessing steps of quality assessment, outlier removal, normalization, and gating have received considerable scrutiny from the community, the influence of data transformation on the output of high throughput analysis has been largely overlooked. Flow cytometry measurements can vary over several orders of magnitude, cell populations can have variances that depend on their mean fluorescence intensities, and may exhibit heavily-skewed distributions. Consequently, the choice of data transformation can influence the output of automated gating. An appropriate data transformation aids in data visualization and gating of cell populations across the range of data. Experience shows that the choice of transformation is data specific. Our goal here is to compare the performance of different transformations applied to flow cytometry data in the context of automated gating in a high throughput, fully automated setting. We examine the most common transformations used in flow cytometry, including the generalized hyperbolic arcsine, biexponential, linlog, and generalized Box-Cox, all within the BioConductor flowCore framework that is widely used in high throughput, automated flow cytometry data analysis. All of these transformations have adjustable parameters whose effects upon the data are non-intuitive for most users. By making some modelling assumptions about the transformed data, we develop maximum likelihood criteria to optimize parameter choice for these different transformations. We compare the performance of parameter-optimized and default-parameter (in flowCore) data transformations on real and simulated data by measuring the variation in the locations of cell populations across samples, discovered via automated gating in both the scatter and fluorescence channels. We find that parameter-optimized transformations improve visualization, reduce

  3. Optimizing transformations for automated, high throughput analysis of flow cytometry data

    Directory of Open Access Journals (Sweden)

    Weng Andrew

    2010-11-01

    Full Text Available Abstract Background In a high throughput setting, effective flow cytometry data analysis depends heavily on proper data preprocessing. While usual preprocessing steps of quality assessment, outlier removal, normalization, and gating have received considerable scrutiny from the community, the influence of data transformation on the output of high throughput analysis has been largely overlooked. Flow cytometry measurements can vary over several orders of magnitude, cell populations can have variances that depend on their mean fluorescence intensities, and may exhibit heavily-skewed distributions. Consequently, the choice of data transformation can influence the output of automated gating. An appropriate data transformation aids in data visualization and gating of cell populations across the range of data. Experience shows that the choice of transformation is data specific. Our goal here is to compare the performance of different transformations applied to flow cytometry data in the context of automated gating in a high throughput, fully automated setting. We examine the most common transformations used in flow cytometry, including the generalized hyperbolic arcsine, biexponential, linlog, and generalized Box-Cox, all within the BioConductor flowCore framework that is widely used in high throughput, automated flow cytometry data analysis. All of these transformations have adjustable parameters whose effects upon the data are non-intuitive for most users. By making some modelling assumptions about the transformed data, we develop maximum likelihood criteria to optimize parameter choice for these different transformations. Results We compare the performance of parameter-optimized and default-parameter (in flowCore data transformations on real and simulated data by measuring the variation in the locations of cell populations across samples, discovered via automated gating in both the scatter and fluorescence channels. We find that parameter-optimized

  4. High-throughput characterization methods for lithium batteries

    Directory of Open Access Journals (Sweden)

    Yingchun Lyu

    2017-09-01

    Full Text Available The development of high-performance lithium ion batteries requires the discovery of new materials and the optimization of key components. By contrast with traditional one-by-one method, high-throughput method can synthesize and characterize a large number of compositionally varying samples, which is able to accelerate the pace of discovery, development and optimization process of materials. Because of rapid progress in thin film and automatic control technologies, thousands of compounds with different compositions could be synthesized rapidly right now, even in a single experiment. However, the lack of rapid or combinatorial characterization technologies to match with high-throughput synthesis methods, limit the application of high-throughput technology. Here, we review a series of representative high-throughput characterization methods used in lithium batteries, including high-throughput structural and electrochemical characterization methods and rapid measuring technologies based on synchrotron light sources.

  5. On the optimal trimming of high-throughput mRNA sequence data

    Directory of Open Access Journals (Sweden)

    Matthew D MacManes

    2014-01-01

    Full Text Available The widespread and rapid adoption of high-throughput sequencing technologies has afforded researchers the opportunity to gain a deep understanding of genome level processes that underlie evolutionary change, and perhaps more importantly, the links between genotype and phenotype. In particular, researchers interested in functional biology and adaptation have used these technologies to sequence mRNA transcriptomes of specific tissues, which in turn are often compared to other tissues, or other individuals with different phenotypes. While these techniques are extremely powerful, careful attention to data quality is required. In particular, because high-throughput sequencing is more error-prone than traditional Sanger sequencing, quality trimming of sequence reads should be an important step in all data processing pipelines. While several software packages for quality trimming exist, no general guidelines for the specifics of trimming have been developed. Here, using empirically derived sequence data, I provide general recommendations regarding the optimal strength of trimming, specifically in mRNA-Seq studies. Although very aggressive quality trimming is common, this study suggests that a more gentle trimming, specifically of those nucleotides whose Phred score < 2 or < 5, is optimal for most studies across a wide variety of metrics.

  6. High-throughput crystal-optimization strategies in the South Paris Yeast Structural Genomics Project: one size fits all?

    Science.gov (United States)

    Leulliot, Nicolas; Trésaugues, Lionel; Bremang, Michael; Sorel, Isabelle; Ulryck, Nathalie; Graille, Marc; Aboulfath, Ilham; Poupon, Anne; Liger, Dominique; Quevillon-Cheruel, Sophie; Janin, Joël; van Tilbeurgh, Herman

    2005-06-01

    Crystallization has long been regarded as one of the major bottlenecks in high-throughput structural determination by X-ray crystallography. Structural genomics projects have addressed this issue by using robots to set up automated crystal screens using nanodrop technology. This has moved the bottleneck from obtaining the first crystal hit to obtaining diffraction-quality crystals, as crystal optimization is a notoriously slow process that is difficult to automatize. This article describes the high-throughput optimization strategies used in the Yeast Structural Genomics project, with selected successful examples.

  7. Selection and optimization of hits from a high-throughput phenotypic screen against Trypanosoma cruzi.

    Science.gov (United States)

    Keenan, Martine; Alexander, Paul W; Chaplin, Jason H; Abbott, Michael J; Diao, Hugo; Wang, Zhisen; Best, Wayne M; Perez, Catherine J; Cornwall, Scott M J; Keatley, Sarah K; Thompson, R C Andrew; Charman, Susan A; White, Karen L; Ryan, Eileen; Chen, Gong; Ioset, Jean-Robert; von Geldern, Thomas W; Chatelain, Eric

    2013-10-01

    Inhibitors of Trypanosoma cruzi with novel mechanisms of action are urgently required to diversify the current clinical and preclinical pipelines. Increasing the number and diversity of hits available for assessment at the beginning of the discovery process will help to achieve this aim. We report the evaluation of multiple hits generated from a high-throughput screen to identify inhibitors of T. cruzi and from these studies the discovery of two novel series currently in lead optimization. Lead compounds from these series potently and selectively inhibit growth of T. cruzi in vitro and the most advanced compound is orally active in a subchronic mouse model of T. cruzi infection. High-throughput screening of novel compound collections has an important role to play in diversifying the trypanosomatid drug discovery portfolio. A new T. cruzi inhibitor series with good drug-like properties and promising in vivo efficacy has been identified through this process.

  8. High-throughput optimization by statistical designs: example with rat liver slices cryopreservation.

    Science.gov (United States)

    Martin, H; Bournique, B; Blanchi, B; Lerche-Langrand, C

    2003-08-01

    The purpose of this study was to optimize cryopreservation conditions of rat liver slices in a high-throughput format, with focus on reproducibility. A statistical design of 32 experiments was performed and intracellular lactate dehydrogenase (LDHi) activity and antipyrine (AP) metabolism were evaluated as biomarkers. At freezing, modified University of Wisconsin solution was better than Williams'E medium, and pure dimethyl sulfoxide was better than a cryoprotectant mixture. The best cryoprotectant concentrations were 10% for LDHi and 20% for AP metabolism. Fetal calf serum could be used at 50 or 80%, and incubation of slices with the cryoprotectant could last 10 or 20 min. At thawing, 42 degrees C was better than 22 degrees C. After thawing, 1h was better than 3h of preculture. Cryopreservation increased the interslice variability of the biomarkers. After cryopreservation, LDHi and AP metabolism levels were up to 84 and 80% of fresh values. However, these high levels were not reproducibly achieved. Two factors involved in the day-to-day variability of LDHi were identified: the incubation time with the cryoprotectant and the preculture time. In conclusion, the statistical design was very efficient to quickly determine optimized conditions by simultaneously measuring the role of numerous factors. The cryopreservation procedure developed appears suitable for qualitative metabolic profiling studies.

  9. Optimization of throughput in semipreparative chiral liquid chromatography using stacked injection.

    Science.gov (United States)

    Taheri, Mohammadreza; Fotovati, Mohsen; Hosseini, Seyed-Kiumars; Ghassempour, Alireza

    2017-10-01

    An interesting mode of chromatography for preparation of pure enantiomers from pure samples is the method of stacked injection as a pseudocontinuous procedure. Maximum throughput and minimal production costs can be achieved by the use of total chiral column length in this mode of chromatography. To maximize sample loading, often touching bands of the two enantiomers is automatically achieved. Conventional equations show direct correlation between touching-band loadability and the selectivity factor of two enantiomers. The important question for one who wants to obtain the highest throughput is "How to optimize different factors including selectivity, resolution, run time, and loading of the sample in order to save time without missing the touching-band resolution?" To answer this question, tramadol and propranolol were separated on cellulose 3,5-dimethyl phenyl carbamate, as two pure racemic mixtures with low and high solubilities in mobile phase, respectively. The mobile phase composition consisted of n-hexane solvent with alcohol modifier and diethylamine as the additive. A response surface methodology based on central composite design was used to optimize separation factors against the main responses. According to the stacked injection properties, two processes were investigated for maximizing throughput: one with a poorly soluble and another with a highly soluble racemic mixture. For each case, different optimization possibilities were inspected. It was revealed that resolution is a crucial response for separations of this kind. Peak area and run time are two critical parameters in optimization of stacked injection for binary mixtures which have low solubility in the mobile phase. © 2017 Wiley Periodicals, Inc.

  10. High-throughput optical system for HDES hyperspectral imager

    Science.gov (United States)

    Václavík, Jan; Melich, Radek; Pintr, Pavel; Pleštil, Jan

    2015-01-01

    Affordable, long-wave infrared hyperspectral imaging calls for use of an uncooled FPA with high-throughput optics. This paper describes the design of the optical part of a stationary hyperspectral imager in a spectral range of 7-14 um with a field of view of 20°×10°. The imager employs a push-broom method made by a scanning mirror. High throughput and a demand for simplicity and rigidity led to a fully refractive design with highly aspheric surfaces and off-axis positioning of the detector array. The design was optimized to exploit the machinability of infrared materials by the SPDT method and a simple assemblage.

  11. High-throughput transformation of Saccharomyces cerevisiae using liquid handling robots.

    Directory of Open Access Journals (Sweden)

    Guangbo Liu

    Full Text Available Saccharomyces cerevisiae (budding yeast is a powerful eukaryotic model organism ideally suited to high-throughput genetic analyses, which time and again has yielded insights that further our understanding of cell biology processes conserved in humans. Lithium Acetate (LiAc transformation of yeast with DNA for the purposes of exogenous protein expression (e.g., plasmids or genome mutation (e.g., gene mutation, deletion, epitope tagging is a useful and long established method. However, a reliable and optimized high throughput transformation protocol that runs almost no risk of human error has not been described in the literature. Here, we describe such a method that is broadly transferable to most liquid handling high-throughput robotic platforms, which are now commonplace in academic and industry settings. Using our optimized method, we are able to comfortably transform approximately 1200 individual strains per day, allowing complete transformation of typical genomic yeast libraries within 6 days. In addition, use of our protocol for gene knockout purposes also provides a potentially quicker, easier and more cost-effective approach to generating collections of double mutants than the popular and elegant synthetic genetic array methodology. In summary, our methodology will be of significant use to anyone interested in high throughput molecular and/or genetic analysis of yeast.

  12. High-throughput theoretical design of lithium battery materials

    International Nuclear Information System (INIS)

    Ling Shi-Gang; Gao Jian; Xiao Rui-Juan; Chen Li-Quan

    2016-01-01

    The rapid evolution of high-throughput theoretical design schemes to discover new lithium battery materials is reviewed, including high-capacity cathodes, low-strain cathodes, anodes, solid state electrolytes, and electrolyte additives. With the development of efficient theoretical methods and inexpensive computers, high-throughput theoretical calculations have played an increasingly important role in the discovery of new materials. With the help of automatic simulation flow, many types of materials can be screened, optimized and designed from a structural database according to specific search criteria. In advanced cell technology, new materials for next generation lithium batteries are of great significance to achieve performance, and some representative criteria are: higher energy density, better safety, and faster charge/discharge speed. (topical review)

  13. High throughput 16S rRNA gene amplicon sequencing

    DEFF Research Database (Denmark)

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

    S rRNA gene amplicon sequencing has been developed over the past few years and is now ready to use for more comprehensive studies related to plant operation and optimization thanks to short analysis time, low cost, high throughput, and high taxonomic resolution. In this study we show how 16S r......RNA gene amplicon sequencing can be used to reveal factors of importance for the operation of full-scale nutrient removal plants related to settling problems and floc properties. Using optimized DNA extraction protocols, indexed primers and our in-house Illumina platform, we prepared multiple samples...... be correlated to the presence of the species that are regarded as “strong” and “weak” floc formers. In conclusion, 16S rRNA gene amplicon sequencing provides a high throughput approach for a rapid and cheap community profiling of activated sludge that in combination with multivariate statistics can be used...

  14. A high-throughput reactor system for optimization of Mo–V–Nb mixed oxide catalyst composition in ethane ODH

    KAUST Repository

    Zhu, Haibo; Laveille, Paco; Rosenfeld, Devon C.; Hedhili, Mohamed N.; Basset, Jean-Marie

    2015-01-01

    75 Mo-V-Nb mixed oxide catalysts with a broad range of compositions were prepared by a simple evaporation method, and were screened for the ethane oxidative dehydrogenation (ODH) reaction. The compositions of these 75 catalysts were systematically changed by varying the Nb loading, and the Mo/V molar ratio. Characterization by XRD, XPS, H2-TPR and SEM revealed that an intimate structure is formed among the 3 components. The strong interaction among different components leads to the formation of a new phase or an "intimate structure". The dependency of conversion and selectivity on the catalyst composition was clearly demonstrated from the results of high-throughput testing. The optimized Mo-V-Nb molar composition was confirmed to be composed of a Nb content of 4-8%, a Mo content of 70-83%, and a V content of 12-25%. The enhanced catalytic performance of the mixed oxides is obviously due to the synergistic effects of the different components. The optimized compositions for ethane ODH revealed in our high-throughput tests and the structural information provided by our characterization studies can serve as the starting point for future efforts to improve the catalytic performance of Mo-V-Nb oxides. This journal is © The Royal Society of Chemistry.

  15. High throughput protein production screening

    Science.gov (United States)

    Beernink, Peter T [Walnut Creek, CA; Coleman, Matthew A [Oakland, CA; Segelke, Brent W [San Ramon, CA

    2009-09-08

    Methods, compositions, and kits for the cell-free production and analysis of proteins are provided. The invention allows for the production of proteins from prokaryotic sequences or eukaryotic sequences, including human cDNAs using PCR and IVT methods and detecting the proteins through fluorescence or immunoblot techniques. This invention can be used to identify optimized PCR and WT conditions, codon usages and mutations. The methods are readily automated and can be used for high throughput analysis of protein expression levels, interactions, and functional states.

  16. High throughput materials research and development for lithium ion batteries

    Directory of Open Access Journals (Sweden)

    Parker Liu

    2017-09-01

    Full Text Available Development of next generation batteries requires a breakthrough in materials. Traditional one-by-one method, which is suitable for synthesizing large number of sing-composition material, is time-consuming and costly. High throughput and combinatorial experimentation, is an effective method to synthesize and characterize huge amount of materials over a broader compositional region in a short time, which enables to greatly speed up the discovery and optimization of materials with lower cost. In this work, high throughput and combinatorial materials synthesis technologies for lithium ion battery research are discussed, and our efforts on developing such instrumentations are introduced.

  17. Machine learning in computational biology to accelerate high-throughput protein expression

    DEFF Research Database (Denmark)

    Sastry, Anand; Monk, Jonathan M.; Tegel, Hanna

    2017-01-01

    and machine learning identifies protein properties that hinder the HPA high-throughput antibody production pipeline. We predict protein expression and solubility with accuracies of 70% and 80%, respectively, based on a subset of key properties (aromaticity, hydropathy and isoelectric point). We guide...... the selection of protein fragments based on these characteristics to optimize high-throughput experimentation. Availability and implementation: We present the machine learning workflow as a series of IPython notebooks hosted on GitHub (https://github.com/SBRG/Protein_ML). The workflow can be used as a template...

  18. High Throughput Facility

    Data.gov (United States)

    Federal Laboratory Consortium — Argonne?s high throughput facility provides highly automated and parallel approaches to material and materials chemistry development. The facility allows scientists...

  19. High-throughput fractionation of human plasma for fast enrichment of low- and high-abundance proteins.

    Science.gov (United States)

    Breen, Lucas; Cao, Lulu; Eom, Kirsten; Srajer Gajdosik, Martina; Camara, Lila; Giacometti, Jasminka; Dupuy, Damian E; Josic, Djuro

    2012-05-01

    Fast, cost-effective and reproducible isolation of IgM from plasma is invaluable to the study of IgM and subsequent understanding of the human immune system. Additionally, vast amounts of information regarding human physiology and disease can be derived from analysis of the low abundance proteome of the plasma. In this study, methods were optimized for both the high-throughput isolation of IgM from human plasma, and the high-throughput isolation and fractionation of low abundance plasma proteins. To optimize the chromatographic isolation of IgM from human plasma, many variables were examined including chromatography resin, mobile phases, and order of chromatographic separations. Purification of IgM was achieved most successfully through isolation of immunoglobulin from human plasma using Protein A chromatography with a specific resin followed by subsequent fractionation using QA strong anion exchange chromatography. Through these optimization experiments, an additional method was established to prepare plasma for analysis of low abundance proteins. This method involved chromatographic depletion of high-abundance plasma proteins and reduction of plasma proteome complexity through further chromatographic fractionation. Purification of IgM was achieved with high purity as confirmed by SDS-PAGE and IgM-specific immunoblot. Isolation and fractionation of low abundance protein was also performed successfully, as confirmed by SDS-PAGE and mass spectrometry analysis followed by label-free quantitative spectral analysis. The level of purity of the isolated IgM allows for further IgM-specific analysis of plasma samples. The developed fractionation scheme can be used for high throughput screening of human plasma in order to identify low and high abundance proteins as potential prognostic and diagnostic disease biomarkers.

  20. High-throughput continuous cryopump

    International Nuclear Information System (INIS)

    Foster, C.A.

    1986-01-01

    A cryopump with a unique method of regeneration which allows continuous operation at high throughput has been constructed and tested. Deuterium was pumped continuously at a throughput of 30 Torr.L/s at a speed of 2000 L/s and a compression ratio of 200. Argon was pumped at a throughput of 60 Torr.L/s at a speed of 1275 L/s. To produce continuous operation of the pump, a method of regeneration that does not thermally cycle the pump is employed. A small chamber (the ''snail'') passes over the pumping surface and removes the frost from it either by mechanical action with a scraper or by local heating. The material removed is topologically in a secondary vacuum system with low conductance into the primary vacuum; thus, the exhaust can be pumped at pressures up to an effective compression ratio determined by the ratio of the pumping speed to the leakage conductance of the snail. The pump, which is all-metal-sealed and dry and which regenerates every 60 s, would be an ideal system for pumping tritium. Potential fusion applications are for mpmp limiters, for repeating pneumatic pellet injection lines, and for the centrifuge pellet injector spin tank, all of which will require pumping tritium at high throughput. Industrial applications requiring ultraclean pumping of corrosive gases at high throughput, such as the reactive ion etch semiconductor process, may also be feasible

  1. High-throughput micro-scale cultivations and chromatography modeling: Powerful tools for integrated process development.

    Science.gov (United States)

    Baumann, Pascal; Hahn, Tobias; Hubbuch, Jürgen

    2015-10-01

    Upstream processes are rather complex to design and the productivity of cells under suitable cultivation conditions is hard to predict. The method of choice for examining the design space is to execute high-throughput cultivation screenings in micro-scale format. Various predictive in silico models have been developed for many downstream processes, leading to a reduction of time and material costs. This paper presents a combined optimization approach based on high-throughput micro-scale cultivation experiments and chromatography modeling. The overall optimized system must not necessarily be the one with highest product titers, but the one resulting in an overall superior process performance in up- and downstream. The methodology is presented in a case study for the Cherry-tagged enzyme Glutathione-S-Transferase from Escherichia coli SE1. The Cherry-Tag™ (Delphi Genetics, Belgium) which can be fused to any target protein allows for direct product analytics by simple VIS absorption measurements. High-throughput cultivations were carried out in a 48-well format in a BioLector micro-scale cultivation system (m2p-Labs, Germany). The downstream process optimization for a set of randomly picked upstream conditions producing high yields was performed in silico using a chromatography modeling software developed in-house (ChromX). The suggested in silico-optimized operational modes for product capturing were validated subsequently. The overall best system was chosen based on a combination of excellent up- and downstream performance. © 2015 Wiley Periodicals, Inc.

  2. High-throughput screening assay used in pharmacognosy: Selection, optimization and validation of methods of enzymatic inhibition by UV-visible spectrophotometry

    Directory of Open Access Journals (Sweden)

    Graciela Granados-Guzmán

    2014-02-01

    Full Text Available In research laboratories of both organic synthesis and extraction of natural products, every day a lot of products that can potentially introduce some biological activity are obtained. Therefore it is necessary to have in vitro assays, which provide reliable information for further evaluation in in vivo systems. From this point of view, in recent years has intensified the use of high-throughput screening assays. Such trials should be optimized and validated for accurate and precise results, i.e. reliable. The present review addresses the steps needed to develop and validate bioanalytical methods, emphasizing UV-Visible spectrophotometry as detection system. Particularly focuses on the selection of the method, the optimization to determine the best experimental conditions, validation, implementation of optimized and validated method to real samples, and finally maintenance and possible transfer it to a new laboratory.

  3. The JCSG high-throughput structural biology pipeline

    International Nuclear Information System (INIS)

    Elsliger, Marc-André; Deacon, Ashley M.; Godzik, Adam; Lesley, Scott A.; Wooley, John; Wüthrich, Kurt; Wilson, Ian A.

    2010-01-01

    The Joint Center for Structural Genomics high-throughput structural biology pipeline has delivered more than 1000 structures to the community over the past ten years and has made a significant contribution to the overall goal of the NIH Protein Structure Initiative (PSI) of expanding structural coverage of the protein universe. The Joint Center for Structural Genomics high-throughput structural biology pipeline has delivered more than 1000 structures to the community over the past ten years. The JCSG has made a significant contribution to the overall goal of the NIH Protein Structure Initiative (PSI) of expanding structural coverage of the protein universe, as well as making substantial inroads into structural coverage of an entire organism. Targets are processed through an extensive combination of bioinformatics and biophysical analyses to efficiently characterize and optimize each target prior to selection for structure determination. The pipeline uses parallel processing methods at almost every step in the process and can adapt to a wide range of protein targets from bacterial to human. The construction, expansion and optimization of the JCSG gene-to-structure pipeline over the years have resulted in many technological and methodological advances and developments. The vast number of targets and the enormous amounts of associated data processed through the multiple stages of the experimental pipeline required the development of variety of valuable resources that, wherever feasible, have been converted to free-access web-based tools and applications

  4. Predictive Power of Machine Learning for Optimizing Solar Water Heater Performance: The Potential Application of High-Throughput Screening

    Directory of Open Access Journals (Sweden)

    Hao Li

    2017-01-01

    Full Text Available Predicting the performance of solar water heater (SWH is challenging due to the complexity of the system. Fortunately, knowledge-based machine learning can provide a fast and precise prediction method for SWH performance. With the predictive power of machine learning models, we can further solve a more challenging question: how to cost-effectively design a high-performance SWH? Here, we summarize our recent studies and propose a general framework of SWH design using a machine learning-based high-throughput screening (HTS method. Design of water-in-glass evacuated tube solar water heater (WGET-SWH is selected as a case study to show the potential application of machine learning-based HTS to the design and optimization of solar energy systems.

  5. Performance Evaluation of IEEE 802.11ah Networks With High-Throughput Bidirectional Traffic.

    Science.gov (United States)

    Šljivo, Amina; Kerkhove, Dwight; Tian, Le; Famaey, Jeroen; Munteanu, Adrian; Moerman, Ingrid; Hoebeke, Jeroen; De Poorter, Eli

    2018-01-23

    So far, existing sub-GHz wireless communication technologies focused on low-bandwidth, long-range communication with large numbers of constrained devices. Although these characteristics are fine for many Internet of Things (IoT) applications, more demanding application requirements could not be met and legacy Internet technologies such as Transmission Control Protocol/Internet Protocol (TCP/IP) could not be used. This has changed with the advent of the new IEEE 802.11ah Wi-Fi standard, which is much more suitable for reliable bidirectional communication and high-throughput applications over a wide area (up to 1 km). The standard offers great possibilities for network performance optimization through a number of physical- and link-layer configurable features. However, given that the optimal configuration parameters depend on traffic patterns, the standard does not dictate how to determine them. Such a large number of configuration options can lead to sub-optimal or even incorrect configurations. Therefore, we investigated how two key mechanisms, Restricted Access Window (RAW) grouping and Traffic Indication Map (TIM) segmentation, influence scalability, throughput, latency and energy efficiency in the presence of bidirectional TCP/IP traffic. We considered both high-throughput video streaming traffic and large-scale reliable sensing traffic and investigated TCP behavior in both scenarios when the link layer introduces long delays. This article presents the relations between attainable throughput per station and attainable number of stations, as well as the influence of RAW, TIM and TCP parameters on both. We found that up to 20 continuously streaming IP-cameras can be reliably connected via IEEE 802.11ah with a maximum average data rate of 160 kbps, whereas 10 IP-cameras can achieve average data rates of up to 255 kbps over 200 m. Up to 6960 stations transmitting every 60 s can be connected over 1 km with no lost packets. The presented results enable the fine tuning

  6. Quantitative description on structure-property relationships of Li-ion battery materials for high-throughput computations

    Science.gov (United States)

    Wang, Youwei; Zhang, Wenqing; Chen, Lidong; Shi, Siqi; Liu, Jianjun

    2017-12-01

    Li-ion batteries are a key technology for addressing the global challenge of clean renewable energy and environment pollution. Their contemporary applications, for portable electronic devices, electric vehicles, and large-scale power grids, stimulate the development of high-performance battery materials with high energy density, high power, good safety, and long lifetime. High-throughput calculations provide a practical strategy to discover new battery materials and optimize currently known material performances. Most cathode materials screened by the previous high-throughput calculations cannot meet the requirement of practical applications because only capacity, voltage and volume change of bulk were considered. It is important to include more structure-property relationships, such as point defects, surface and interface, doping and metal-mixture and nanosize effects, in high-throughput calculations. In this review, we established quantitative description of structure-property relationships in Li-ion battery materials by the intrinsic bulk parameters, which can be applied in future high-throughput calculations to screen Li-ion battery materials. Based on these parameterized structure-property relationships, a possible high-throughput computational screening flow path is proposed to obtain high-performance battery materials.

  7. Combinatorial materials synthesis and high-throughput screening: an integrated materials chip approach to mapping phase diagrams and discovery and optimization of functional materials.

    Science.gov (United States)

    Xiang, X D

    Combinatorial materials synthesis methods and high-throughput evaluation techniques have been developed to accelerate the process of materials discovery and optimization and phase-diagram mapping. Analogous to integrated circuit chips, integrated materials chips containing thousands of discrete different compositions or continuous phase diagrams, often in the form of high-quality epitaxial thin films, can be fabricated and screened for interesting properties. Microspot x-ray method, various optical measurement techniques, and a novel evanescent microwave microscope have been used to characterize the structural, optical, magnetic, and electrical properties of samples on the materials chips. These techniques are routinely used to discover/optimize and map phase diagrams of ferroelectric, dielectric, optical, magnetic, and superconducting materials.

  8. Determining the optimal size of small molecule mixtures for high throughput NMR screening

    International Nuclear Information System (INIS)

    Mercier, Kelly A.; Powers, Robert

    2005-01-01

    High-throughput screening (HTS) using NMR spectroscopy has become a common component of the drug discovery effort and is widely used throughout the pharmaceutical industry. NMR provides additional information about the nature of small molecule-protein interactions compared to traditional HTS methods. In order to achieve comparable efficiency, small molecules are often screened as mixtures in NMR-based assays. Nevertheless, an analysis of the efficiency of mixtures and a corresponding determination of the optimum mixture size (OMS) that minimizes the amount of material and instrumentation time required for an NMR screen has been lacking. A model for calculating OMS based on the application of the hypergeometric distribution function to determine the probability of a 'hit' for various mixture sizes and hit rates is presented. An alternative method for the deconvolution of large screening mixtures is also discussed. These methods have been applied in a high-throughput NMR screening assay using a small, directed library

  9. A rapid enzymatic assay for high-throughput screening of adenosine-producing strains

    Science.gov (United States)

    Dong, Huina; Zu, Xin; Zheng, Ping; Zhang, Dawei

    2015-01-01

    Adenosine is a major local regulator of tissue function and industrially useful as precursor for the production of medicinal nucleoside substances. High-throughput screening of adenosine overproducers is important for industrial microorganism breeding. An enzymatic assay of adenosine was developed by combined adenosine deaminase (ADA) with indophenol method. The ADA catalyzes the cleavage of adenosine to inosine and NH3, the latter can be accurately determined by indophenol method. The assay system was optimized to deliver a good performance and could tolerate the addition of inorganic salts and many nutrition components to the assay mixtures. Adenosine could be accurately determined by this assay using 96-well microplates. Spike and recovery tests showed that this assay can accurately and reproducibly determine increases in adenosine in fermentation broth without any pretreatment to remove proteins and potentially interfering low-molecular-weight molecules. This assay was also applied to high-throughput screening for high adenosine-producing strains. The high selectivity and accuracy of the ADA assay provides rapid and high-throughput analysis of adenosine in large numbers of samples. PMID:25580842

  10. The high throughput biomedicine unit at the institute for molecular medicine Finland: high throughput screening meets precision medicine.

    Science.gov (United States)

    Pietiainen, Vilja; Saarela, Jani; von Schantz, Carina; Turunen, Laura; Ostling, Paivi; Wennerberg, Krister

    2014-05-01

    The High Throughput Biomedicine (HTB) unit at the Institute for Molecular Medicine Finland FIMM was established in 2010 to serve as a national and international academic screening unit providing access to state of the art instrumentation for chemical and RNAi-based high throughput screening. The initial focus of the unit was multiwell plate based chemical screening and high content microarray-based siRNA screening. However, over the first four years of operation, the unit has moved to a more flexible service platform where both chemical and siRNA screening is performed at different scales primarily in multiwell plate-based assays with a wide range of readout possibilities with a focus on ultraminiaturization to allow for affordable screening for the academic users. In addition to high throughput screening, the equipment of the unit is also used to support miniaturized, multiplexed and high throughput applications for other types of research such as genomics, sequencing and biobanking operations. Importantly, with the translational research goals at FIMM, an increasing part of the operations at the HTB unit is being focused on high throughput systems biological platforms for functional profiling of patient cells in personalized and precision medicine projects.

  11. An Overlay Architecture for Throughput Optimal Multipath Routing

    Science.gov (United States)

    2017-01-14

    maximum throughput. Finally, we propose a threshold-based policy (BP-T) and a heuristic policy (OBP), which dynamically control traffic bifurcations...network stability region is available . Second, given any subset of nodes that are controllable, we also wish to develop an optimal routing policy that...case when tunnels do not overlap. We also develop a heuristic overlay control policy for use on general topologies, and show through simulation that

  12. High-Throughput Method for Strontium Isotope Analysis by Multi-Collector-Inductively Coupled Plasma-Mass Spectrometer

    Energy Technology Data Exchange (ETDEWEB)

    Wall, Andrew J. [National Energy Technology Lab. (NETL), Pittsburgh, PA, (United States); Capo, Rosemary C. [Univ. of Pittsburgh, PA (United States); Stewart, Brian W. [Univ. of Pittsburgh, PA (United States); Phan, Thai T. [Univ. of Pittsburgh, PA (United States); Jain, Jinesh C. [National Energy Technology Lab. (NETL), Pittsburgh, PA, (United States); Hakala, Alexandra [National Energy Technology Lab. (NETL), Pittsburgh, PA, (United States); Guthrie, George D. [National Energy Technology Lab. (NETL), Pittsburgh, PA, (United States)

    2016-09-22

    This technical report presents the details of the Sr column configuration and the high-throughput Sr separation protocol. Data showing the performance of the method as well as the best practices for optimizing Sr isotope analysis by MC-ICP-MS is presented. Lastly, this report offers tools for data handling and data reduction of Sr isotope results from the Thermo Scientific Neptune software to assist in data quality assurance, which help avoid issues of data glut associated with high sample throughput rapid analysis.

  13. High-Throughput Method for Strontium Isotope Analysis by Multi-Collector-Inductively Coupled Plasma-Mass Spectrometer

    Energy Technology Data Exchange (ETDEWEB)

    Hakala, Jacqueline Alexandra [National Energy Technology Lab. (NETL), Morgantown, WV (United States)

    2016-11-22

    This technical report presents the details of the Sr column configuration and the high-throughput Sr separation protocol. Data showing the performance of the method as well as the best practices for optimizing Sr isotope analysis by MC-ICP-MS is presented. Lastly, this report offers tools for data handling and data reduction of Sr isotope results from the Thermo Scientific Neptune software to assist in data quality assurance, which help avoid issues of data glut associated with high sample throughput rapid analysis.

  14. High-throughput microplate technique for enzymatic hydrolysis of lignocellulosic biomass.

    Science.gov (United States)

    Chundawat, Shishir P S; Balan, Venkatesh; Dale, Bruce E

    2008-04-15

    Several factors will influence the viability of a biochemical platform for manufacturing lignocellulosic based fuels and chemicals, for example, genetically engineering energy crops, reducing pre-treatment severity, and minimizing enzyme loading. Past research on biomass conversion has focused largely on acid based pre-treatment technologies that fractionate lignin and hemicellulose from cellulose. However, for alkaline based (e.g., AFEX) and other lower severity pre-treatments it becomes critical to co-hydrolyze cellulose and hemicellulose using an optimized enzyme cocktail. Lignocellulosics are appropriate substrates to assess hydrolytic activity of enzyme mixtures compared to conventional unrealistic substrates (e.g., filter paper, chromogenic, and fluorigenic compounds) for studying synergistic hydrolysis. However, there are few, if any, high-throughput lignocellulosic digestibility analytical platforms for optimizing biomass conversion. The 96-well Biomass Conversion Research Lab (BCRL) microplate method is a high-throughput assay to study digestibility of lignocellulosic biomass as a function of biomass composition, pre-treatment severity, and enzyme composition. The most suitable method for delivering milled biomass to the microplate was through multi-pipetting slurry suspensions. A rapid bio-enzymatic, spectrophotometric assay was used to determine fermentable sugars. The entire procedure was automated using a robotic pipetting workstation. Several parameters that affect hydrolysis in the microplate were studied and optimized (i.e., particle size reduction, slurry solids concentration, glucan loading, mass transfer issues, and time period for hydrolysis). The microplate method was optimized for crystalline cellulose (Avicel) and ammonia fiber expansion (AFEX) pre-treated corn stover. Copyright 2008 Wiley Periodicals, Inc.

  15. Roche genome sequencer FLX based high-throughput sequencing of ancient DNA

    DEFF Research Database (Denmark)

    Alquezar-Planas, David E; Fordyce, Sarah Louise

    2012-01-01

    Since the development of so-called "next generation" high-throughput sequencing in 2005, this technology has been applied to a variety of fields. Such applications include disease studies, evolutionary investigations, and ancient DNA. Each application requires a specialized protocol to ensure...... that the data produced is optimal. Although much of the procedure can be followed directly from the manufacturer's protocols, the key differences lie in the library preparation steps. This chapter presents an optimized protocol for the sequencing of fossil remains and museum specimens, commonly referred...

  16. High throughput sample processing and automated scoring

    Directory of Open Access Journals (Sweden)

    Gunnar eBrunborg

    2014-10-01

    Full Text Available The comet assay is a sensitive and versatile method for assessing DNA damage in cells. In the traditional version of the assay, there are many manual steps involved and few samples can be treated in one experiment. High throughput modifications have been developed during recent years, and they are reviewed and discussed. These modifications include accelerated scoring of comets; other important elements that have been studied and adapted to high throughput are cultivation and manipulation of cells or tissues before and after exposure, and freezing of treated samples until comet analysis and scoring. High throughput methods save time and money but they are useful also for other reasons: large-scale experiments may be performed which are otherwise not practicable (e.g., analysis of many organs from exposed animals, and human biomonitoring studies, and automation gives more uniform sample treatment and less dependence on operator performance. The high throughput modifications now available vary largely in their versatility, capacity, complexity and costs. The bottleneck for further increase of throughput appears to be the scoring.

  17. High Throughput Neuro-Imaging Informatics

    Directory of Open Access Journals (Sweden)

    Michael I Miller

    2013-12-01

    Full Text Available This paper describes neuroinformatics technologies at 1 mm anatomical scale based on high throughput 3D functional and structural imaging technologies of the human brain. The core is an abstract pipeline for converting functional and structural imagery into their high dimensional neuroinformatic representations index containing O(E3-E4 discriminating dimensions. The pipeline is based on advanced image analysis coupled to digital knowledge representations in the form of dense atlases of the human brain at gross anatomical scale. We demonstrate the integration of these high-dimensional representations with machine learning methods, which have become the mainstay of other fields of science including genomics as well as social networks. Such high throughput facilities have the potential to alter the way medical images are stored and utilized in radiological workflows. The neuroinformatics pipeline is used to examine cross-sectional and personalized analyses of neuropsychiatric illnesses in clinical applications as well as longitudinal studies. We demonstrate the use of high throughput machine learning methods for supporting (i cross-sectional image analysis to evaluate the health status of individual subjects with respect to the population data, (ii integration of image and non-image information for diagnosis and prognosis.

  18. Temperature-programmed technique accompanied with high-throughput methodology for rapidly searching the optimal operating temperature of MOX gas sensors.

    Science.gov (United States)

    Zhang, Guozhu; Xie, Changsheng; Zhang, Shunping; Zhao, Jianwei; Lei, Tao; Zeng, Dawen

    2014-09-08

    A combinatorial high-throughput temperature-programmed method to obtain the optimal operating temperature (OOT) of gas sensor materials is demonstrated here for the first time. A material library consisting of SnO2, ZnO, WO3, and In2O3 sensor films was fabricated by screen printing. Temperature-dependent conductivity curves were obtained by scanning this gas sensor library from 300 to 700 K in different atmospheres (dry air, formaldehyde, carbon monoxide, nitrogen dioxide, toluene and ammonia), giving the OOT of each sensor formulation as a function of the carrier and analyte gases. A comparative study of the temperature-programmed method and a conventional method showed good agreement in measured OOT.

  19. Toward reliable and repeatable automated STEM-EDS metrology with high throughput

    Science.gov (United States)

    Zhong, Zhenxin; Donald, Jason; Dutrow, Gavin; Roller, Justin; Ugurlu, Ozan; Verheijen, Martin; Bidiuk, Oleksii

    2018-03-01

    New materials and designs in complex 3D architectures in logic and memory devices have raised complexity in S/TEM metrology. In this paper, we report about a newly developed, automated, scanning transmission electron microscopy (STEM) based, energy dispersive X-ray spectroscopy (STEM-EDS) metrology method that addresses these challenges. Different methodologies toward repeatable and efficient, automated STEM-EDS metrology with high throughput are presented: we introduce the best known auto-EDS acquisition and quantification methods for robust and reliable metrology and present how electron exposure dose impacts the EDS metrology reproducibility, either due to poor signalto-noise ratio (SNR) at low dose or due to sample modifications at high dose conditions. Finally, we discuss the limitations of the STEM-EDS metrology technique and propose strategies to optimize the process both in terms of throughput and metrology reliability.

  20. High throughput on-chip analysis of high-energy charged particle tracks using lensfree imaging

    Energy Technology Data Exchange (ETDEWEB)

    Luo, Wei; Shabbir, Faizan; Gong, Chao; Gulec, Cagatay; Pigeon, Jeremy; Shaw, Jessica; Greenbaum, Alon; Tochitsky, Sergei; Joshi, Chandrashekhar [Electrical Engineering Department, University of California, Los Angeles, California 90095 (United States); Ozcan, Aydogan, E-mail: ozcan@ucla.edu [Electrical Engineering Department, University of California, Los Angeles, California 90095 (United States); Bioengineering Department, University of California, Los Angeles, California 90095 (United States); California NanoSystems Institute (CNSI), University of California, Los Angeles, California 90095 (United States)

    2015-04-13

    We demonstrate a high-throughput charged particle analysis platform, which is based on lensfree on-chip microscopy for rapid ion track analysis using allyl diglycol carbonate, i.e., CR-39 plastic polymer as the sensing medium. By adopting a wide-area opto-electronic image sensor together with a source-shifting based pixel super-resolution technique, a large CR-39 sample volume (i.e., 4 cm × 4 cm × 0.1 cm) can be imaged in less than 1 min using a compact lensfree on-chip microscope, which detects partially coherent in-line holograms of the ion tracks recorded within the CR-39 detector. After the image capture, using highly parallelized reconstruction and ion track analysis algorithms running on graphics processing units, we reconstruct and analyze the entire volume of a CR-39 detector within ∼1.5 min. This significant reduction in the entire imaging and ion track analysis time not only increases our throughput but also allows us to perform time-resolved analysis of the etching process to monitor and optimize the growth of ion tracks during etching. This computational lensfree imaging platform can provide a much higher throughput and more cost-effective alternative to traditional lens-based scanning optical microscopes for ion track analysis using CR-39 and other passive high energy particle detectors.

  1. Machine learning in computational biology to accelerate high-throughput protein expression.

    Science.gov (United States)

    Sastry, Anand; Monk, Jonathan; Tegel, Hanna; Uhlen, Mathias; Palsson, Bernhard O; Rockberg, Johan; Brunk, Elizabeth

    2017-08-15

    The Human Protein Atlas (HPA) enables the simultaneous characterization of thousands of proteins across various tissues to pinpoint their spatial location in the human body. This has been achieved through transcriptomics and high-throughput immunohistochemistry-based approaches, where over 40 000 unique human protein fragments have been expressed in E. coli. These datasets enable quantitative tracking of entire cellular proteomes and present new avenues for understanding molecular-level properties influencing expression and solubility. Combining computational biology and machine learning identifies protein properties that hinder the HPA high-throughput antibody production pipeline. We predict protein expression and solubility with accuracies of 70% and 80%, respectively, based on a subset of key properties (aromaticity, hydropathy and isoelectric point). We guide the selection of protein fragments based on these characteristics to optimize high-throughput experimentation. We present the machine learning workflow as a series of IPython notebooks hosted on GitHub (https://github.com/SBRG/Protein_ML). The workflow can be used as a template for analysis of further expression and solubility datasets. ebrunk@ucsd.edu or johanr@biotech.kth.se. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  2. Chromatographic Monoliths for High-Throughput Immunoaffinity Isolation of Transferrin from Human Plasma

    Directory of Open Access Journals (Sweden)

    Irena Trbojević-Akmačić

    2016-06-01

    Full Text Available Changes in protein glycosylation are related to different diseases and have a potential as diagnostic and prognostic disease biomarkers. Transferrin (Tf glycosylation changes are common marker for congenital disorders of glycosylation. However, biological interindividual variability of Tf N-glycosylation and genes involved in glycosylation regulation are not known. Therefore, high-throughput Tf isolation method and large scale glycosylation studies are needed in order to address these questions. Due to their unique chromatographic properties, the use of chromatographic monoliths enables very fast analysis cycle, thus significantly increasing sample preparation throughput. Here, we are describing characterization of novel immunoaffinity-based monolithic columns in a 96-well plate format for specific high-throughput purification of human Tf from blood plasma. We optimized the isolation and glycan preparation procedure for subsequent ultra performance liquid chromatography (UPLC analysis of Tf N-glycosylation and managed to increase the sensitivity for approximately three times compared to initial experimental conditions, with very good reproducibility. This work is licensed under a Creative Commons Attribution 4.0 International License.

  3. MetaUniDec: High-Throughput Deconvolution of Native Mass Spectra

    Science.gov (United States)

    Reid, Deseree J.; Diesing, Jessica M.; Miller, Matthew A.; Perry, Scott M.; Wales, Jessica A.; Montfort, William R.; Marty, Michael T.

    2018-04-01

    The expansion of native mass spectrometry (MS) methods for both academic and industrial applications has created a substantial need for analysis of large native MS datasets. Existing software tools are poorly suited for high-throughput deconvolution of native electrospray mass spectra from intact proteins and protein complexes. The UniDec Bayesian deconvolution algorithm is uniquely well suited for high-throughput analysis due to its speed and robustness but was previously tailored towards individual spectra. Here, we optimized UniDec for deconvolution, analysis, and visualization of large data sets. This new module, MetaUniDec, centers around a hierarchical data format 5 (HDF5) format for storing datasets that significantly improves speed, portability, and file size. It also includes code optimizations to improve speed and a new graphical user interface for visualization, interaction, and analysis of data. To demonstrate the utility of MetaUniDec, we applied the software to analyze automated collision voltage ramps with a small bacterial heme protein and large lipoprotein nanodiscs. Upon increasing collisional activation, bacterial heme-nitric oxide/oxygen binding (H-NOX) protein shows a discrete loss of bound heme, and nanodiscs show a continuous loss of lipids and charge. By using MetaUniDec to track changes in peak area or mass as a function of collision voltage, we explore the energetic profile of collisional activation in an ultra-high mass range Orbitrap mass spectrometer. [Figure not available: see fulltext.

  4. Optimal throughput for cognitive radio with energy harvesting in fading wireless channel.

    Science.gov (United States)

    Vu-Van, Hiep; Koo, Insoo

    2014-01-01

    Energy resource management is a crucial problem of a device with a finite capacity battery. In this paper, cognitive radio is considered to be a device with an energy harvester that can harvest energy from a non-RF energy resource while performing other actions of cognitive radio. Harvested energy will be stored in a finite capacity battery. At the start of the time slot of cognitive radio, the radio needs to determine if it should remain silent or carry out spectrum sensing based on the idle probability of the primary user and the remaining energy in order to maximize the throughput of the cognitive radio system. In addition, optimal sensing energy and adaptive transmission power control are also investigated in this paper to effectively utilize the limited energy of cognitive radio. Finding an optimal approach is formulated as a partially observable Markov decision process. The simulation results show that the proposed optimal decision scheme outperforms the myopic scheme in which current throughput is only considered when making a decision.

  5. High Throughput Transcriptomics @ USEPA (Toxicology ...

    Science.gov (United States)

    The ideal chemical testing approach will provide complete coverage of all relevant toxicological responses. It should be sensitive and specific It should identify the mechanism/mode-of-action (with dose-dependence). It should identify responses relevant to the species of interest. Responses should ideally be translated into tissue-, organ-, and organism-level effects. It must be economical and scalable. Using a High Throughput Transcriptomics platform within US EPA provides broader coverage of biological activity space and toxicological MOAs and helps fill the toxicological data gap. Slide presentation at the 2016 ToxForum on using High Throughput Transcriptomics at US EPA for broader coverage biological activity space and toxicological MOAs.

  6. Application of ToxCast High-Throughput Screening and ...

    Science.gov (United States)

    Slide presentation at the SETAC annual meeting on High-Throughput Screening and Modeling Approaches to Identify Steroidogenesis Distruptors Slide presentation at the SETAC annual meeting on High-Throughput Screening and Modeling Approaches to Identify Steroidogenssis Distruptors

  7. Improvement of IBAD-MgO texturing for high throughput of buffered substrate

    International Nuclear Information System (INIS)

    Ito, T.; Takahashi, Y.; Matsuse, K.; Kuriki, R.; Tokumaru, M.; Yoshizumi, M.; Izumi, T.

    2011-01-01

    The requirements from the market on two important factors of performance and cost need to be satisfied for commercialization of the coated conductors. Highly biaxially grain texturing with high production rate should be realized from the perspective of buffer layers processing. IBAD-MgO process is one of the major techniques which are possible to satisfy those requirements. The structure of our buffered substrate is IBS-GZO/IBAD-MgO/RFsputter-LaMnO 3 /PLD-CeO 2 . The PLD-CeO 2 process is the rate limiting and cost dominant one in this architecture. It is proposed that the self-texturing CeO 2 layer thickness could be reduced by optimization of the MgO processing due to higher MgO texturing and/or effective growth of self-texturing CeO 2 . Influence of the IBAD beam conditions and deposition time has been studied to optimize the IBAD conditions. Optimized IBAD conditions were decided from the viewpoints of in-plane grain texturing and the stability to obtain high texturing on fabrication. The Δφ value of CeO 2 layer was improved from 4-5 o to 3-3.5 o by the optimization. This buffered substrate gave high and uniform I c values of 524-565 A/cm-width for 50 m long GdBCO (1.5 μm) tape, indicating uniform distribution of Δφ(CeO 2 ). This improvement of Δφ(CeO 2 ) enables to reduce the CeO 2 thickness down to 300 nm without making Δφ(CeO 2 ) > 5 o , which improves CeO 2 throughput from 10 m/h to 30 m/h. A 50 m long patch sample showed more uniform Δφ distribution around 4 o even by high speed of 30 m/h as CeO 2 through-put. Highly and uniformly textured CeO 2 buffered substrate was obtained in 100 m long cost-effectively by optimization of IBAD-MgO processing.

  8. A high throughput DNA extraction method with high yield and quality

    Directory of Open Access Journals (Sweden)

    Xin Zhanguo

    2012-07-01

    Full Text Available Abstract Background Preparation of large quantity and high quality genomic DNA from a large number of plant samples is a major bottleneck for most genetic and genomic analyses, such as, genetic mapping, TILLING (Targeting Induced Local Lesion IN Genome, and next-generation sequencing directly from sheared genomic DNA. A variety of DNA preparation methods and commercial kits are available. However, they are either low throughput, low yield, or costly. Here, we describe a method for high throughput genomic DNA isolation from sorghum [Sorghum bicolor (L. Moench] leaves and dry seeds with high yield, high quality, and affordable cost. Results We developed a high throughput DNA isolation method by combining a high yield CTAB extraction method with an improved cleanup procedure based on MagAttract kit. The method yielded large quantity and high quality DNA from both lyophilized sorghum leaves and dry seeds. The DNA yield was improved by nearly 30 fold with 4 times less consumption of MagAttract beads. The method can also be used in other plant species, including cotton leaves and pine needles. Conclusion A high throughput system for DNA extraction from sorghum leaves and seeds was developed and validated. The main advantages of the method are low cost, high yield, high quality, and high throughput. One person can process two 96-well plates in a working day at a cost of $0.10 per sample of magnetic beads plus other consumables that other methods will also need.

  9. Complexity optimization and high-throughput low-latency hardware implementation of a multi-electrode spike-sorting algorithm.

    Science.gov (United States)

    Dragas, Jelena; Jackel, David; Hierlemann, Andreas; Franke, Felix

    2015-03-01

    Reliable real-time low-latency spike sorting with large data throughput is essential for studies of neural network dynamics and for brain-machine interfaces (BMIs), in which the stimulation of neural networks is based on the networks' most recent activity. However, the majority of existing multi-electrode spike-sorting algorithms are unsuited for processing high quantities of simultaneously recorded data. Recording from large neuronal networks using large high-density electrode sets (thousands of electrodes) imposes high demands on the data-processing hardware regarding computational complexity and data transmission bandwidth; this, in turn, entails demanding requirements in terms of chip area, memory resources and processing latency. This paper presents computational complexity optimization techniques, which facilitate the use of spike-sorting algorithms in large multi-electrode-based recording systems. The techniques are then applied to a previously published algorithm, on its own, unsuited for large electrode set recordings. Further, a real-time low-latency high-performance VLSI hardware architecture of the modified algorithm is presented, featuring a folded structure capable of processing the activity of hundreds of neurons simultaneously. The hardware is reconfigurable “on-the-fly” and adaptable to the nonstationarities of neuronal recordings. By transmitting exclusively spike time stamps and/or spike waveforms, its real-time processing offers the possibility of data bandwidth and data storage reduction.

  10. High-Throughput Analysis and Automation for Glycomics Studies.

    Science.gov (United States)

    Shubhakar, Archana; Reiding, Karli R; Gardner, Richard A; Spencer, Daniel I R; Fernandes, Daryl L; Wuhrer, Manfred

    This review covers advances in analytical technologies for high-throughput (HTP) glycomics. Our focus is on structural studies of glycoprotein glycosylation to support biopharmaceutical realization and the discovery of glycan biomarkers for human disease. For biopharmaceuticals, there is increasing use of glycomics in Quality by Design studies to help optimize glycan profiles of drugs with a view to improving their clinical performance. Glycomics is also used in comparability studies to ensure consistency of glycosylation both throughout product development and between biosimilars and innovator drugs. In clinical studies there is as well an expanding interest in the use of glycomics-for example in Genome Wide Association Studies-to follow changes in glycosylation patterns of biological tissues and fluids with the progress of certain diseases. These include cancers, neurodegenerative disorders and inflammatory conditions. Despite rising activity in this field, there are significant challenges in performing large scale glycomics studies. The requirement is accurate identification and quantitation of individual glycan structures. However, glycoconjugate samples are often very complex and heterogeneous and contain many diverse branched glycan structures. In this article we cover HTP sample preparation and derivatization methods, sample purification, robotization, optimized glycan profiling by UHPLC, MS and multiplexed CE, as well as hyphenated techniques and automated data analysis tools. Throughout, we summarize the advantages and challenges with each of these technologies. The issues considered include reliability of the methods for glycan identification and quantitation, sample throughput, labor intensity, and affordability for large sample numbers.

  11. Optimized high-throughput microRNA expression profiling provides novel biomarker assessment of clinical prostate and breast cancer biopsies

    Directory of Open Access Journals (Sweden)

    Fedele Vita

    2006-06-01

    Full Text Available Abstract Background Recent studies indicate that microRNAs (miRNAs are mechanistically involved in the development of various human malignancies, suggesting that they represent a promising new class of cancer biomarkers. However, previously reported methods for measuring miRNA expression consume large amounts of tissue, prohibiting high-throughput miRNA profiling from typically small clinical samples such as excision or core needle biopsies of breast or prostate cancer. Here we describe a novel combination of linear amplification and labeling of miRNA for highly sensitive expression microarray profiling requiring only picogram quantities of purified microRNA. Results Comparison of microarray and qRT-PCR measured miRNA levels from two different prostate cancer cell lines showed concordance between the two platforms (Pearson correlation R2 = 0.81; and extension of the amplification, labeling and microarray platform was successfully demonstrated using clinical core and excision biopsy samples from breast and prostate cancer patients. Unsupervised clustering analysis of the prostate biopsy microarrays separated advanced and metastatic prostate cancers from pooled normal prostatic samples and from a non-malignant precursor lesion. Unsupervised clustering of the breast cancer microarrays significantly distinguished ErbB2-positive/ER-negative, ErbB2-positive/ER-positive, and ErbB2-negative/ER-positive breast cancer phenotypes (Fisher exact test, p = 0.03; as well, supervised analysis of these microarray profiles identified distinct miRNA subsets distinguishing ErbB2-positive from ErbB2-negative and ER-positive from ER-negative breast cancers, independent of other clinically important parameters (patient age; tumor size, node status and proliferation index. Conclusion In sum, these findings demonstrate that optimized high-throughput microRNA expression profiling offers novel biomarker identification from typically small clinical samples such as breast

  12. Design of a High-Throughput Biological Crystallography Beamline for Superconducting Wiggler

    International Nuclear Information System (INIS)

    Tseng, P.C.; Chang, C.H.; Fung, H.S.; Ma, C.I.; Huang, L.J.; Jean, Y.C.; Song, Y.F.; Huang, Y.S.; Tsang, K.L.; Chen, C.T.

    2004-01-01

    We are constructing a high-throughput biological crystallography beamline BL13B, which utilizes the radiation generated from a 3.2 Tesla, 32-pole superconducting multipole wiggler, for multi-wavelength anomalous diffraction (MAD), single-wavelength anomalous diffraction (SAD), and other related experiments. This beamline is a standard double crystal monochromator (DCM) x-ray beamline equipped with a collimating mirror (CM) and a focusing mirror (FM). Both the CM and FM are one meter long and made of Si substrate, and the CM is side-cooled by water. Based on detailed thermal analysis, liquid nitrogen (LN2) cooling for both crystals of the DCM has been adopted to optimize the energy resolution and photon beam throughput. This beamline will deliver, through a 100 μm diameter pinhole, photon flux of greater than 1011 photons/sec in the energy range from 6.5 keV to 19 keV, which is comparable to existing protein crystallography beamlines from bending magnet source at high energy storage rings

  13. Optimizing the Energy and Throughput of a Water-Quality Monitoring System.

    Science.gov (United States)

    Olatinwo, Segun O; Joubert, Trudi-H

    2018-04-13

    This work presents a new approach to the maximization of energy and throughput in a wireless sensor network (WSN), with the intention of applying the approach to water-quality monitoring. Water-quality monitoring using WSN technology has become an interesting research area. Energy scarcity is a critical issue that plagues the widespread deployment of WSN systems. Different power supplies, harvesting energy from sustainable sources, have been explored. However, when energy-efficient models are not put in place, energy harvesting based WSN systems may experience an unstable energy supply, resulting in an interruption in communication, and low system throughput. To alleviate these problems, this paper presents the joint maximization of the energy harvested by sensor nodes and their information-transmission rate using a sum-throughput technique. A wireless information and power transfer (WIPT) method is considered by harvesting energy from dedicated radio frequency sources. Due to the doubly near-far condition that confronts WIPT systems, a new WIPT system is proposed to improve the fairness of resource utilization in the network. Numerical simulation results are presented to validate the mathematical formulations for the optimization problem, which maximize the energy harvested and the overall throughput rate. Defining the performance metrics of achievable throughput and fairness in resource sharing, the proposed WIPT system outperforms an existing state-of-the-art WIPT system, with the comparison based on numerical simulations of both systems. The improved energy efficiency of the proposed WIPT system contributes to addressing the problem of energy scarcity.

  14. Meeting Report: High-Throughput Technologies for In Vivo Imaging Agents

    Directory of Open Access Journals (Sweden)

    Robert J. Gillies

    2005-04-01

    Full Text Available Combinatorial chemistry and high-throughput screening have become standard tools for discovering new drug candidates with suitable pharmacological properties. Now, those same technologies are starting to be applied to the problem of discovering novel in vivo imaging agents. Important differences in the biological and pharmacological properties needed for imaging agents, compared to those for a therapeutic agent, require new screening methods that emphasize those characteristics, such as optimized residence time and tissue specificity, that make for a good imaging agent candidate.

  15. High Throughput PBTK: Open-Source Data and Tools for ...

    Science.gov (United States)

    Presentation on High Throughput PBTK at the PBK Modelling in Risk Assessment meeting in Ispra, Italy Presentation on High Throughput PBTK at the PBK Modelling in Risk Assessment meeting in Ispra, Italy

  16. Optimization of a micro-scale, high throughput process development tool and the demonstration of comparable process performance and product quality with biopharmaceutical manufacturing processes.

    Science.gov (United States)

    Evans, Steven T; Stewart, Kevin D; Afdahl, Chris; Patel, Rohan; Newell, Kelcy J

    2017-07-14

    In this paper, we discuss the optimization and implementation of a high throughput process development (HTPD) tool that utilizes commercially available micro-liter sized column technology for the purification of multiple clinically significant monoclonal antibodies. Chromatographic profiles generated using this optimized tool are shown to overlay with comparable profiles from the conventional bench-scale and clinical manufacturing scale. Further, all product quality attributes measured are comparable across scales for the mAb purifications. In addition to supporting chromatography process development efforts (e.g., optimization screening), comparable product quality results at all scales makes this tool is an appropriate scale model to enable purification and product quality comparisons of HTPD bioreactors conditions. The ability to perform up to 8 chromatography purifications in parallel with reduced material requirements per run creates opportunities for gathering more process knowledge in less time. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  17. Optimizing the Energy and Throughput of a Water-Quality Monitoring System

    Directory of Open Access Journals (Sweden)

    Segun O. Olatinwo

    2018-04-01

    Full Text Available This work presents a new approach to the maximization of energy and throughput in a wireless sensor network (WSN, with the intention of applying the approach to water-quality monitoring. Water-quality monitoring using WSN technology has become an interesting research area. Energy scarcity is a critical issue that plagues the widespread deployment of WSN systems. Different power supplies, harvesting energy from sustainable sources, have been explored. However, when energy-efficient models are not put in place, energy harvesting based WSN systems may experience an unstable energy supply, resulting in an interruption in communication, and low system throughput. To alleviate these problems, this paper presents the joint maximization of the energy harvested by sensor nodes and their information-transmission rate using a sum-throughput technique. A wireless information and power transfer (WIPT method is considered by harvesting energy from dedicated radio frequency sources. Due to the doubly near–far condition that confronts WIPT systems, a new WIPT system is proposed to improve the fairness of resource utilization in the network. Numerical simulation results are presented to validate the mathematical formulations for the optimization problem, which maximize the energy harvested and the overall throughput rate. Defining the performance metrics of achievable throughput and fairness in resource sharing, the proposed WIPT system outperforms an existing state-of-the-art WIPT system, with the comparison based on numerical simulations of both systems. The improved energy efficiency of the proposed WIPT system contributes to addressing the problem of energy scarcity.

  18. High-throughput design of low-activation, high-strength creep-resistant steels for nuclear-reactor applications

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Qi; Zwaag, Sybrand van der [Novel Aerospace Materials Group, Faculty of Aerospace Engineering, Delft University of Technology, Kluyverweg 1, 2629 HS, Delft (Netherlands); Xu, Wei, E-mail: xuwei@ral.neu.edu.cn [State Key Laboratory of Rolling and Automation, Northeastern University, 110819, Shenyang (China); Novel Aerospace Materials Group, Faculty of Aerospace Engineering, Delft University of Technology, Kluyverweg 1, 2629 HS, Delft (Netherlands)

    2016-02-15

    Reduced-activation ferritic/martensitic steels are prime candidate materials for structural applications in nuclear power reactors. However, their creep strength is much lower than that of creep-resistant steel developed for conventional fossil-fired power plants as alloying elements with a high neutron activation cannot be used. To improve the creep strength and to maintain a low activation, a high-throughput computational alloy design model coupling thermodynamics, precipitate-coarsening kinetics and an optimization genetic algorithm, is developed. Twelve relevant alloying elements with either low or high activation are considered simultaneously. The activity levels at 0–10 year after the end of irradiation are taken as optimization parameter. The creep-strength values (after exposure for 10 years at 650 °C) are estimated on the basis of the solid-solution strengthening and the precipitation hardening (taking into account precipitate coarsening). Potential alloy compositions leading to a high austenite fraction or a high percentage of undesirable second phase particles are rejected automatically in the optimization cycle. The newly identified alloys have a much higher precipitation hardening and solid-solution strengthening at the same activity level as existing reduced-activation ferritic/martensitic steels.

  19. High-Throughput Next-Generation Sequencing of Polioviruses

    Science.gov (United States)

    Montmayeur, Anna M.; Schmidt, Alexander; Zhao, Kun; Magaña, Laura; Iber, Jane; Castro, Christina J.; Chen, Qi; Henderson, Elizabeth; Ramos, Edward; Shaw, Jing; Tatusov, Roman L.; Dybdahl-Sissoko, Naomi; Endegue-Zanga, Marie Claire; Adeniji, Johnson A.; Oberste, M. Steven; Burns, Cara C.

    2016-01-01

    ABSTRACT The poliovirus (PV) is currently targeted for worldwide eradication and containment. Sanger-based sequencing of the viral protein 1 (VP1) capsid region is currently the standard method for PV surveillance. However, the whole-genome sequence is sometimes needed for higher resolution global surveillance. In this study, we optimized whole-genome sequencing protocols for poliovirus isolates and FTA cards using next-generation sequencing (NGS), aiming for high sequence coverage, efficiency, and throughput. We found that DNase treatment of poliovirus RNA followed by random reverse transcription (RT), amplification, and the use of the Nextera XT DNA library preparation kit produced significantly better results than other preparations. The average viral reads per total reads, a measurement of efficiency, was as high as 84.2% ± 15.6%. PV genomes covering >99 to 100% of the reference length were obtained and validated with Sanger sequencing. A total of 52 PV genomes were generated, multiplexing as many as 64 samples in a single Illumina MiSeq run. This high-throughput, sequence-independent NGS approach facilitated the detection of a diverse range of PVs, especially for those in vaccine-derived polioviruses (VDPV), circulating VDPV, or immunodeficiency-related VDPV. In contrast to results from previous studies on other viruses, our results showed that filtration and nuclease treatment did not discernibly increase the sequencing efficiency of PV isolates. However, DNase treatment after nucleic acid extraction to remove host DNA significantly improved the sequencing results. This NGS method has been successfully implemented to generate PV genomes for molecular epidemiology of the most recent PV isolates. Additionally, the ability to obtain full PV genomes from FTA cards will aid in facilitating global poliovirus surveillance. PMID:27927929

  20. Improvement of IBAD-MgO texturing for high throughput of buffered substrate

    Energy Technology Data Exchange (ETDEWEB)

    Ito, T., E-mail: t-ito@istec.or.jp [Superconductivity Research Laboratory, ISTEC, 1-10-13, Shinonome, Koto-ku, Tokyo 135-0062 (Japan); Takahashi, Y.; Matsuse, K.; Kuriki, R.; Tokumaru, M.; Yoshizumi, M.; Izumi, T. [Superconductivity Research Laboratory, ISTEC, 1-10-13, Shinonome, Koto-ku, Tokyo 135-0062 (Japan)

    2011-11-15

    The requirements from the market on two important factors of performance and cost need to be satisfied for commercialization of the coated conductors. Highly biaxially grain texturing with high production rate should be realized from the perspective of buffer layers processing. IBAD-MgO process is one of the major techniques which are possible to satisfy those requirements. The structure of our buffered substrate is IBS-GZO/IBAD-MgO/RFsputter-LaMnO{sub 3}/PLD-CeO{sub 2}. The PLD-CeO{sub 2} process is the rate limiting and cost dominant one in this architecture. It is proposed that the self-texturing CeO{sub 2} layer thickness could be reduced by optimization of the MgO processing due to higher MgO texturing and/or effective growth of self-texturing CeO{sub 2}. Influence of the IBAD beam conditions and deposition time has been studied to optimize the IBAD conditions. Optimized IBAD conditions were decided from the viewpoints of in-plane grain texturing and the stability to obtain high texturing on fabrication. The {Delta}{phi} value of CeO{sub 2} layer was improved from 4-5{sup o} to 3-3.5{sup o} by the optimization. This buffered substrate gave high and uniform I{sub c} values of 524-565 A/cm-width for 50 m long GdBCO (1.5 {mu}m) tape, indicating uniform distribution of {Delta}{phi}(CeO{sub 2}). This improvement of {Delta}{phi}(CeO{sub 2}) enables to reduce the CeO{sub 2} thickness down to 300 nm without making {Delta}{phi}(CeO{sub 2}) > 5{sup o}, which improves CeO{sub 2} throughput from 10 m/h to 30 m/h. A 50 m long patch sample showed more uniform {Delta}{phi} distribution around 4{sup o} even by high speed of 30 m/h as CeO{sub 2} through-put. Highly and uniformly textured CeO{sub 2} buffered substrate was obtained in 100 m long cost-effectively by optimization of IBAD-MgO processing.

  1. High throughput screening method for assessing heterogeneity of microorganisms

    NARCIS (Netherlands)

    Ingham, C.J.; Sprenkels, A.J.; van Hylckama Vlieg, J.E.T.; Bomer, Johan G.; de Vos, W.M.; van den Berg, Albert

    2006-01-01

    The invention relates to the field of microbiology. Provided is a method which is particularly powerful for High Throughput Screening (HTS) purposes. More specific a high throughput method for determining heterogeneity or interactions of microorganisms is provided.

  2. Novel high-throughput cell-based hybridoma screening methodology using the Celigo Image Cytometer.

    Science.gov (United States)

    Zhang, Haohai; Chan, Leo Li-Ying; Rice, William; Kassam, Nasim; Longhi, Maria Serena; Zhao, Haitao; Robson, Simon C; Gao, Wenda; Wu, Yan

    2017-08-01

    Hybridoma screening is a critical step for antibody discovery, which necessitates prompt identification of potential clones from hundreds to thousands of hybridoma cultures against the desired immunogen. Technical issues associated with ELISA- and flow cytometry-based screening limit accuracy and diminish high-throughput capability, increasing time and cost. Conventional ELISA screening with coated antigen is also impractical for difficult-to-express hydrophobic membrane antigens or multi-chain protein complexes. Here, we demonstrate novel high-throughput screening methodology employing the Celigo Image Cytometer, which avoids nonspecific signals by contrasting antibody binding signals directly on living cells, with and without recombinant antigen expression. The image cytometry-based high-throughput screening method was optimized by detecting the binding of hybridoma supernatants to the recombinant antigen CD39 expressed on Chinese hamster ovary (CHO) cells. Next, the sensitivity of the image cytometer was demonstrated by serial dilution of purified CD39 antibody. Celigo was used to measure antibody affinities of commercial and in-house antibodies to membrane-bound CD39. This cell-based screening procedure can be completely accomplished within one day, significantly improving throughput and efficiency of hybridoma screening. Furthermore, measuring direct antibody binding to living cells eliminated both false positive and false negative hits. The image cytometry method was highly sensitive and versatile, and could detect positive antibody in supernatants at concentrations as low as ~5ng/mL, with concurrent K d binding affinity coefficient determination. We propose that this screening method will greatly facilitate antibody discovery and screening technologies. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. High throughput nanoimprint lithography for semiconductor memory applications

    Science.gov (United States)

    Ye, Zhengmao; Zhang, Wei; Khusnatdinov, Niyaz; Stachowiak, Tim; Irving, J. W.; Longsine, Whitney; Traub, Matthew; Fletcher, Brian; Liu, Weijun

    2017-03-01

    Imprint lithography is a promising technology for replication of nano-scale features. For semiconductor device applications, Canon deposits a low viscosity resist on a field by field basis using jetting technology. A patterned mask is lowered into the resist fluid which then quickly flows into the relief patterns in the mask by capillary action. Following this filling step, the resist is crosslinked under UV radiation, and then the mask is removed, leaving a patterned resist on the substrate. There are two critical components to meeting throughput requirements for imprint lithography. Using a similar approach to what is already done for many deposition and etch processes, imprint stations can be clustered to enhance throughput. The FPA-1200NZ2C is a four station cluster system designed for high volume manufacturing. For a single station, throughput includes overhead, resist dispense, resist fill time (or spread time), exposure and separation. Resist exposure time and mask/wafer separation are well understood processing steps with typical durations on the order of 0.10 to 0.20 seconds. To achieve a total process throughput of 17 wafers per hour (wph) for a single station, it is necessary to complete the fluid fill step in 1.2 seconds. For a throughput of 20 wph, fill time must be reduced to only one 1.1 seconds. There are several parameters that can impact resist filling. Key parameters include resist drop volume (smaller is better), system controls (which address drop spreading after jetting), Design for Imprint or DFI (to accelerate drop spreading) and material engineering (to promote wetting between the resist and underlying adhesion layer). In addition, it is mandatory to maintain fast filling, even for edge field imprinting. In this paper, we address the improvements made in all of these parameters to first enable a 1.20 second filling process for a device like pattern and have demonstrated this capability for both full fields and edge fields. Non

  4. High-throughput sample adaptive offset hardware architecture for high-efficiency video coding

    Science.gov (United States)

    Zhou, Wei; Yan, Chang; Zhang, Jingzhi; Zhou, Xin

    2018-03-01

    A high-throughput hardware architecture for a sample adaptive offset (SAO) filter in the high-efficiency video coding video coding standard is presented. First, an implementation-friendly and simplified bitrate estimation method of rate-distortion cost calculation is proposed to reduce the computational complexity in the mode decision of SAO. Then, a high-throughput VLSI architecture for SAO is presented based on the proposed bitrate estimation method. Furthermore, multiparallel VLSI architecture for in-loop filters, which integrates both deblocking filter and SAO filter, is proposed. Six parallel strategies are applied in the proposed in-loop filters architecture to improve the system throughput and filtering speed. Experimental results show that the proposed in-loop filters architecture can achieve up to 48% higher throughput in comparison with prior work. The proposed architecture can reach a high-operating clock frequency of 297 MHz with TSMC 65-nm library and meet the real-time requirement of the in-loop filters for 8 K × 4 K video format at 132 fps.

  5. In-Field High-Throughput Phenotyping of Cotton Plant Height Using LiDAR

    OpenAIRE

    Shangpeng Sun; Changying Li; Andrew H. Paterson

    2017-01-01

    A LiDAR-based high-throughput phenotyping (HTP) system was developed for cotton plant phenotyping in the field. The HTP system consists of a 2D LiDAR and an RTK-GPS mounted on a high clearance tractor. The LiDAR scanned three rows of cotton plots simultaneously from the top and the RTK-GPS was used to provide the spatial coordinates of the point cloud during data collection. Configuration parameters of the system were optimized to ensure the best data quality. A height profile for each plot w...

  6. A Fully Automated High-Throughput Flow Cytometry Screening System Enabling Phenotypic Drug Discovery.

    Science.gov (United States)

    Joslin, John; Gilligan, James; Anderson, Paul; Garcia, Catherine; Sharif, Orzala; Hampton, Janice; Cohen, Steven; King, Miranda; Zhou, Bin; Jiang, Shumei; Trussell, Christopher; Dunn, Robert; Fathman, John W; Snead, Jennifer L; Boitano, Anthony E; Nguyen, Tommy; Conner, Michael; Cooke, Mike; Harris, Jennifer; Ainscow, Ed; Zhou, Yingyao; Shaw, Chris; Sipes, Dan; Mainquist, James; Lesley, Scott

    2018-05-01

    The goal of high-throughput screening is to enable screening of compound libraries in an automated manner to identify quality starting points for optimization. This often involves screening a large diversity of compounds in an assay that preserves a connection to the disease pathology. Phenotypic screening is a powerful tool for drug identification, in that assays can be run without prior understanding of the target and with primary cells that closely mimic the therapeutic setting. Advanced automation and high-content imaging have enabled many complex assays, but these are still relatively slow and low throughput. To address this limitation, we have developed an automated workflow that is dedicated to processing complex phenotypic assays for flow cytometry. The system can achieve a throughput of 50,000 wells per day, resulting in a fully automated platform that enables robust phenotypic drug discovery. Over the past 5 years, this screening system has been used for a variety of drug discovery programs, across many disease areas, with many molecules advancing quickly into preclinical development and into the clinic. This report will highlight a diversity of approaches that automated flow cytometry has enabled for phenotypic drug discovery.

  7. The protein crystallography beamline BW6 at DORIS - automatic operation and high-throughput data collection

    CERN Document Server

    Blume, H; Bourenkov, G P; Kosciesza, D; Bartunik, H D

    2001-01-01

    The wiggler beamline BW6 at DORIS has been optimized for de-novo solution of protein structures on the basis of MAD phasing. Facilities for automatic data collection, rapid data transfer and storage, and online processing have been developed which provide adequate conditions for high-throughput applications, e.g., in structural genomics.

  8. High Throughput Determinations of Critical Dosing Parameters (IVIVE workshop)

    Science.gov (United States)

    High throughput toxicokinetics (HTTK) is an approach that allows for rapid estimations of TK for hundreds of environmental chemicals. HTTK-based reverse dosimetry (i.e, reverse toxicokinetics or RTK) is used in order to convert high throughput in vitro toxicity screening (HTS) da...

  9. High throughput nanostructure-initiator mass spectrometry screening of microbial growth conditions for maximal β-glucosidase production.

    Science.gov (United States)

    Cheng, Xiaoliang; Hiras, Jennifer; Deng, Kai; Bowen, Benjamin; Simmons, Blake A; Adams, Paul D; Singer, Steven W; Northen, Trent R

    2013-01-01

    Production of biofuels via enzymatic hydrolysis of complex plant polysaccharides is a subject of intense global interest. Microbial communities are known to express a wide range of enzymes necessary for the saccharification of lignocellulosic feedstocks and serve as a powerful reservoir for enzyme discovery. However, the growth temperature and conditions that yield high cellulase activity vary widely, and the throughput to identify optimal conditions has been limited by the slow handling and conventional analysis. A rapid method that uses small volumes of isolate culture to resolve specific enzyme activity is needed. In this work, a high throughput nanostructure-initiator mass spectrometry (NIMS)-based approach was developed for screening a thermophilic cellulolytic actinomycete, Thermobispora bispora, for β-glucosidase production under various growth conditions. Media that produced high β-glucosidase activity were found to be I/S + glucose or microcrystalline cellulose (MCC), Medium 84 + rolled oats, and M9TE + MCC at 45°C. Supernatants of cell cultures grown in M9TE + 1% MCC cleaved 2.5 times more substrate at 45°C than at all other temperatures. While T. bispora is reported to grow optimally at 60°C in Medium 84 + rolled oats and M9TE + 1% MCC, approximately 40% more conversion was observed at 45°C. This high throughput NIMS approach may provide an important tool in discovery and characterization of enzymes from environmental microbes for industrial and biofuel applications.

  10. High throughput nanostructure-initiator mass spectrometry screening of microbial growth conditions for maximal β-glucosidase production

    Directory of Open Access Journals (Sweden)

    Xiaoliang eCheng

    2013-12-01

    Full Text Available Production of biofuels via enzymatic hydrolysis of complex plant polysaccharides is a subject of intense global interest. Microbial communities are known to express a wide range of enzymes necessary for the saccharification of lignocellulosic feedstocks and serve as a powerful reservoir for enzyme discovery. However, the growth temperature and conditions that yield high cellulase activity vary widely, and the throughput to identify optimal conditions has been limited by the slow handling and conventional analysis. A rapid method that uses small volumes of isolate culture to resolve specific enzyme activity is needed. In this work, a high throughput nanostructure-initiator mass spectrometry (NIMS based approach was developed for screening a thermophilic cellulolytic actinomycete, Thermobispora bispora, for β-glucosidase production under various growth conditions. Media that produced high β-glucosidase activity were found to be I/S + glucose or microcrystalline cellulose (MCC, Medium 84 + rolled oats, and M9TE + MCC at 45 °C. Supernatants of cell cultures grown in M9TE + 1% MCC cleaved 2.5 times more substrate at 45 °C than at all other temperatures. While T. bispora is reported to grow optimally at 60 °C in Medium 84 + rolled oats and M9TE + 1% MCC, approximately 40% more conversion was observed at 45 °C. This high throughput NIMS approach may provide an important tool in discovery and characterization of enzymes from environmental microbes for industrial and biofuel applications.

  11. Applications of ambient mass spectrometry in high-throughput screening.

    Science.gov (United States)

    Li, Li-Ping; Feng, Bao-Sheng; Yang, Jian-Wang; Chang, Cui-Lan; Bai, Yu; Liu, Hu-Wei

    2013-06-07

    The development of rapid screening and identification techniques is of great importance for drug discovery, doping control, forensic identification, food safety and quality control. Ambient mass spectrometry (AMS) allows rapid and direct analysis of various samples in open air with little sample preparation. Recently, its applications in high-throughput screening have been in rapid progress. During the past decade, various ambient ionization techniques have been developed and applied in high-throughput screening. This review discusses typical applications of AMS, including DESI (desorption electrospray ionization), DART (direct analysis in real time), EESI (extractive electrospray ionization), etc., in high-throughput screening (HTS).

  12. High-throughput screening (HTS) and modeling of the retinoid ...

    Science.gov (United States)

    Presentation at the Retinoids Review 2nd workshop in Brussels, Belgium on the application of high throughput screening and model to the retinoid system Presentation at the Retinoids Review 2nd workshop in Brussels, Belgium on the application of high throughput screening and model to the retinoid system

  13. High-Throughput Lipolysis in 96-Well Plates for Rapid Screening of Lipid-Based Drug Delivery Systems

    DEFF Research Database (Denmark)

    Mosgaard, Mette D; Sassene, Philip J; Mu, Huiling

    2017-01-01

    The high-throughput in vitro intestinal lipolysis model (HTP) applicable for rapid and low-scale screening of lipid-based drug delivery systems (LbDDSs) was optimized and adjusted as to be conducted in 96-well plates (HTP-96). Three different LbDDSs (I-III) loaded with danazol or cinnarizine were...

  14. Applications of high-throughput clonogenic survival assays in high-LET particle microbeams

    Directory of Open Access Journals (Sweden)

    Antonios eGeorgantzoglou

    2016-01-01

    Full Text Available Charged particle therapy is increasingly becoming a valuable tool in cancer treatment, mainly due to the favorable interaction of particle radiation with matter. Its application is still limited due, in part, to lack of data regarding the radiosensitivity of certain cell lines to this radiation type, especially to high-LET particles. From the earliest days of radiation biology, the clonogenic survival assay has been used to provide radiation response data. This method produces reliable data but it is not optimized for high-throughput microbeam studies with high-LET radiation where high levels of cell killing lead to a very low probability of maintaining cells’ clonogenic potential. A new method, therefore, is proposed in this paper, which could potentially allow these experiments to be conducted in a high-throughput fashion. Cells are seeded in special polypropylene dishes and bright-field illumination provides cell visualization. Digital images are obtained and cell detection is applied based on corner detection, generating individual cell targets as x-y points. These points in the dish are then irradiated individually by a micron field size high-LET microbeam. Post-irradiation, time-lapse imaging follows cells’ response. All irradiated cells are tracked by linking trajectories in all time-frames, based on finding their nearest position. Cell divisions are detected based on cell appearance and individual cell temporary corner density. The number of divisions anticipated is low due to the high probability of cell killing from high-LET irradiation. Survival curves are produced based on cell’s capacity to divide at least 4-5 times. The process is repeated for a range of doses of radiation. Validation shows the efficiency of the proposed cell detection and tracking method in finding cell divisions.

  15. Applications of High-Throughput Clonogenic Survival Assays in High-LET Particle Microbeams.

    Science.gov (United States)

    Georgantzoglou, Antonios; Merchant, Michael J; Jeynes, Jonathan C G; Mayhead, Natalie; Punia, Natasha; Butler, Rachel E; Jena, Rajesh

    2015-01-01

    Charged particle therapy is increasingly becoming a valuable tool in cancer treatment, mainly due to the favorable interaction of particle radiation with matter. Its application is still limited due, in part, to lack of data regarding the radiosensitivity of certain cell lines to this radiation type, especially to high-linear energy transfer (LET) particles. From the earliest days of radiation biology, the clonogenic survival assay has been used to provide radiation response data. This method produces reliable data but it is not optimized for high-throughput microbeam studies with high-LET radiation where high levels of cell killing lead to a very low probability of maintaining cells' clonogenic potential. A new method, therefore, is proposed in this paper, which could potentially allow these experiments to be conducted in a high-throughput fashion. Cells are seeded in special polypropylene dishes and bright-field illumination provides cell visualization. Digital images are obtained and cell detection is applied based on corner detection, generating individual cell targets as x-y points. These points in the dish are then irradiated individually by a micron field size high-LET microbeam. Post-irradiation, time-lapse imaging follows cells' response. All irradiated cells are tracked by linking trajectories in all time-frames, based on finding their nearest position. Cell divisions are detected based on cell appearance and individual cell temporary corner density. The number of divisions anticipated is low due to the high probability of cell killing from high-LET irradiation. Survival curves are produced based on cell's capacity to divide at least four to five times. The process is repeated for a range of doses of radiation. Validation shows the efficiency of the proposed cell detection and tracking method in finding cell divisions.

  16. High-throughput scoring of seed germination

    NARCIS (Netherlands)

    Ligterink, Wilco; Hilhorst, Henk W.M.

    2017-01-01

    High-throughput analysis of seed germination for phenotyping large genetic populations or mutant collections is very labor intensive and would highly benefit from an automated setup. Although very often used, the total germination percentage after a nominated period of time is not very

  17. Throughput-Based Traffic Steering in LTE-Advanced HetNet Deployments

    DEFF Research Database (Denmark)

    Gimenez, Lucas Chavarria; Kovacs, Istvan Z.; Wigard, Jeroen

    2015-01-01

    The objective of this paper is to propose traffic steering solutions that aim at optimizing the end-user throughput. Two different implementations of an active mode throughput-based traffic steering algorithm for Heterogeneous Networks (HetNet) are introduced. One that always forces handover of t...... throughput is generally higher, reaching values of 36% and 18% for the medium- and high-load conditions....

  18. eRNA: a graphic user interface-based tool optimized for large data analysis from high-throughput RNA sequencing.

    Science.gov (United States)

    Yuan, Tiezheng; Huang, Xiaoyi; Dittmar, Rachel L; Du, Meijun; Kohli, Manish; Boardman, Lisa; Thibodeau, Stephen N; Wang, Liang

    2014-03-05

    RNA sequencing (RNA-seq) is emerging as a critical approach in biological research. However, its high-throughput advantage is significantly limited by the capacity of bioinformatics tools. The research community urgently needs user-friendly tools to efficiently analyze the complicated data generated by high throughput sequencers. We developed a standalone tool with graphic user interface (GUI)-based analytic modules, known as eRNA. The capacity of performing parallel processing and sample management facilitates large data analyses by maximizing hardware usage and freeing users from tediously handling sequencing data. The module miRNA identification" includes GUIs for raw data reading, adapter removal, sequence alignment, and read counting. The module "mRNA identification" includes GUIs for reference sequences, genome mapping, transcript assembling, and differential expression. The module "Target screening" provides expression profiling analyses and graphic visualization. The module "Self-testing" offers the directory setups, sample management, and a check for third-party package dependency. Integration of other GUIs including Bowtie, miRDeep2, and miRspring extend the program's functionality. eRNA focuses on the common tools required for the mapping and quantification analysis of miRNA-seq and mRNA-seq data. The software package provides an additional choice for scientists who require a user-friendly computing environment and high-throughput capacity for large data analysis. eRNA is available for free download at https://sourceforge.net/projects/erna/?source=directory.

  19. 20180311 - High Throughput Transcriptomics: From screening to pathways (SOT 2018)

    Science.gov (United States)

    The EPA ToxCast effort has screened thousands of chemicals across hundreds of high-throughput in vitro screening assays. The project is now leveraging high-throughput transcriptomic (HTTr) technologies to substantially expand its coverage of biological pathways. The first HTTr sc...

  20. High throughput label-free platform for statistical bio-molecular sensing

    DEFF Research Database (Denmark)

    Bosco, Filippo; Hwu, En-Te; Chen, Ching-Hsiu

    2011-01-01

    Sensors are crucial in many daily operations including security, environmental control, human diagnostics and patient monitoring. Screening and online monitoring require reliable and high-throughput sensing. We report on the demonstration of a high-throughput label-free sensor platform utilizing...

  1. In-situ nanoelectrospray for high-throughput screening of enzymes and real-time monitoring of reactions.

    Science.gov (United States)

    Yang, Yuhan; Han, Feifei; Ouyang, Jin; Zhao, Yunling; Han, Juan; Na, Na

    2016-01-01

    The in-situ and high-throughput evaluation of enzymes and real-time monitoring of enzyme catalyzed reactions in liquid phase is quite significant in the catalysis industry. In-situ nanoelectrospray, the direct sampling and ionization method for mass spectrometry, has been applied for high-throughput evaluation of enzymes, as well as the on-line monitoring of reactions. Simply inserting a capillary into a liquid system with high-voltage applied, analytes in liquid reaction system can be directly ionized at the capillary tip with small volume consumption. With no sample pre-treatment or injection procedure, different analytes such as saccharides, amino acids, alkaloids, peptides and proteins can be rapidly and directly extracted from liquid phase and ionized at the capillary tip. Taking irreversible transesterification reaction of vinyl acetate and ethanol as an example, this technique has been used for the high-throughput evaluation of enzymes, fast optimizations, as well as real-time monitoring of reaction catalyzed by different enzymes. In addition, it is even softer than traditional electrospray ionization. The present method can also be used for the monitoring of other homogenous and heterogeneous reactions in liquid phases, which will show potentials in the catalysis industry. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. High-throughput sequence alignment using Graphics Processing Units

    Directory of Open Access Journals (Sweden)

    Trapnell Cole

    2007-12-01

    Full Text Available Abstract Background The recent availability of new, less expensive high-throughput DNA sequencing technologies has yielded a dramatic increase in the volume of sequence data that must be analyzed. These data are being generated for several purposes, including genotyping, genome resequencing, metagenomics, and de novo genome assembly projects. Sequence alignment programs such as MUMmer have proven essential for analysis of these data, but researchers will need ever faster, high-throughput alignment tools running on inexpensive hardware to keep up with new sequence technologies. Results This paper describes MUMmerGPU, an open-source high-throughput parallel pairwise local sequence alignment program that runs on commodity Graphics Processing Units (GPUs in common workstations. MUMmerGPU uses the new Compute Unified Device Architecture (CUDA from nVidia to align multiple query sequences against a single reference sequence stored as a suffix tree. By processing the queries in parallel on the highly parallel graphics card, MUMmerGPU achieves more than a 10-fold speedup over a serial CPU version of the sequence alignment kernel, and outperforms the exact alignment component of MUMmer on a high end CPU by 3.5-fold in total application time when aligning reads from recent sequencing projects using Solexa/Illumina, 454, and Sanger sequencing technologies. Conclusion MUMmerGPU is a low cost, ultra-fast sequence alignment program designed to handle the increasing volume of data produced by new, high-throughput sequencing technologies. MUMmerGPU demonstrates that even memory-intensive applications can run significantly faster on the relatively low-cost GPU than on the CPU.

  3. High throughput imaging cytometer with acoustic focussing.

    Science.gov (United States)

    Zmijan, Robert; Jonnalagadda, Umesh S; Carugo, Dario; Kochi, Yu; Lemm, Elizabeth; Packham, Graham; Hill, Martyn; Glynne-Jones, Peter

    2015-10-31

    We demonstrate an imaging flow cytometer that uses acoustic levitation to assemble cells and other particles into a sheet structure. This technique enables a high resolution, low noise CMOS camera to capture images of thousands of cells with each frame. While ultrasonic focussing has previously been demonstrated for 1D cytometry systems, extending the technology to a planar, much higher throughput format and integrating imaging is non-trivial, and represents a significant jump forward in capability, leading to diagnostic possibilities not achievable with current systems. A galvo mirror is used to track the images of the moving cells permitting exposure times of 10 ms at frame rates of 50 fps with motion blur of only a few pixels. At 80 fps, we demonstrate a throughput of 208 000 beads per second. We investigate the factors affecting motion blur and throughput, and demonstrate the system with fluorescent beads, leukaemia cells and a chondrocyte cell line. Cells require more time to reach the acoustic focus than beads, resulting in lower throughputs; however a longer device would remove this constraint.

  4. High-Throughput Screening of a Luciferase Reporter of Gene Silencing on the Inactive X Chromosome.

    Science.gov (United States)

    Keegan, Alissa; Plath, Kathrin; Damoiseaux, Robert

    2018-01-01

    Assays of luciferase gene activity are a sensitive and quantitative reporter system suited to high-throughput screening. We adapted a luciferase assay to a screening strategy for identifying factors that reactivate epigenetically silenced genes. This epigenetic luciferase reporter is subject to endogenous gene silencing mechanisms on the inactive X chromosome (Xi) in primary mouse cells and thus captures the multilayered nature of chromatin silencing in development. Here, we describe the optimization of an Xi-linked luciferase reactivation assay in 384-well format and adaptation of the assay for high-throughput siRNA and chemical screening. Xi-luciferase reactivation screening has applications in stem cell biology and cancer therapy. We have used the approach described here to identify chromatin-modifying proteins and to identify drug combinations that enhance the gene reactivation activity of the DNA demethylating drug 5-aza-2'-deoxycytidine.

  5. High-throughput GPU-based LDPC decoding

    Science.gov (United States)

    Chang, Yang-Lang; Chang, Cheng-Chun; Huang, Min-Yu; Huang, Bormin

    2010-08-01

    Low-density parity-check (LDPC) code is a linear block code known to approach the Shannon limit via the iterative sum-product algorithm. LDPC codes have been adopted in most current communication systems such as DVB-S2, WiMAX, WI-FI and 10GBASE-T. LDPC for the needs of reliable and flexible communication links for a wide variety of communication standards and configurations have inspired the demand for high-performance and flexibility computing. Accordingly, finding a fast and reconfigurable developing platform for designing the high-throughput LDPC decoder has become important especially for rapidly changing communication standards and configurations. In this paper, a new graphic-processing-unit (GPU) LDPC decoding platform with the asynchronous data transfer is proposed to realize this practical implementation. Experimental results showed that the proposed GPU-based decoder achieved 271x speedup compared to its CPU-based counterpart. It can serve as a high-throughput LDPC decoder.

  6. Evaluating High Throughput Toxicokinetics and Toxicodynamics for IVIVE (WC10)

    Science.gov (United States)

    High-throughput screening (HTS) generates in vitro data for characterizing potential chemical hazard. TK models are needed to allow in vitro to in vivo extrapolation (IVIVE) to real world situations. The U.S. EPA has created a public tool (R package “httk” for high throughput tox...

  7. Optimization of cell line development in the GS-CHO expression system using a high-throughput, single cell-based clone selection system.

    Science.gov (United States)

    Nakamura, Tsuyoshi; Omasa, Takeshi

    2015-09-01

    Therapeutic antibodies are commonly produced by high-expressing, clonal and recombinant Chinese hamster ovary (CHO) cell lines. Currently, CHO cells dominate as a commercial production host because of their ease of use, established regulatory track record, and safety profile. CHO-K1SV is a suspension, protein-free-adapted CHO-K1-derived cell line employing the glutamine synthetase (GS) gene expression system (GS-CHO expression system). The selection of high-producing mammalian cell lines is a crucial step in process development for the production of therapeutic antibodies. In general, cloning by the limiting dilution method is used to isolate high-producing monoclonal CHO cells. However, the limiting dilution method is time consuming and has a low probability of monoclonality. To minimize the duration and increase the probability of obtaining high-producing clones with high monoclonality, an automated single cell-based clone selector, the ClonePix FL system, is available. In this study, we applied the high-throughput ClonePix FL system for cell line development using CHO-K1SV cells and investigated efficient conditions for single cell-based clone selection. CHO-K1SV cell growth at the pre-picking stage was improved by optimizing the formulation of semi-solid medium. The efficiency of picking and cell growth at the post-picking stage was improved by optimization of the plating time without decreasing the diversity of clones. The conditions for selection, including the medium formulation, were the most important factors for the single cell-based clone selection system to construct a high-producing CHO cell line. Copyright © 2015 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  8. Turbulent flow chromatography TFC-tandem mass spectrometry supporting in vitro/vivo studies of NCEs in high throughput fashion.

    Science.gov (United States)

    Verdirame, Maria; Veneziano, Maria; Alfieri, Anna; Di Marco, Annalise; Monteagudo, Edith; Bonelli, Fabio

    2010-03-11

    Turbulent Flow Chromatography (TFC) is a powerful approach for on-line extraction in bioanalytical studies. It improves sensitivity and reduces sample preparation time, two factors that are of primary importance in drug discovery. In this paper the application of the ARIA system to the analytical support of in vivo pharmacokinetics (PK) and in vitro drug metabolism studies is described, with an emphasis in high throughput optimization. For PK studies, a comparison between acetonitrile plasma protein precipitation (APPP) and TFC was carried out. Our optimized TFC methodology gave better S/N ratios and lower limit of quantification (LOQ) than conventional procedures. A robust and high throughput analytical method to support hepatocyte metabolic stability screening of new chemical entities was developed by hyphenation of TFC with mass spectrometry. An in-loop dilution injection procedure was implemented to overcome one of the main issues when using TFC, that is the early elution of hydrophilic compounds that renders low recoveries. A comparison between off-line solid phase extraction (SPE) and TFC was also carried out, and recovery, sensitivity (LOQ), matrix effect and robustness were evaluated. The use of two parallel columns in the configuration of the system provided a further increase of the throughput. Copyright 2009 Elsevier B.V. All rights reserved.

  9. High-throughput combinatorial chemical bath deposition: The case of doping Cu (In, Ga) Se film with antimony

    Science.gov (United States)

    Yan, Zongkai; Zhang, Xiaokun; Li, Guang; Cui, Yuxing; Jiang, Zhaolian; Liu, Wen; Peng, Zhi; Xiang, Yong

    2018-01-01

    The conventional methods for designing and preparing thin film based on wet process remain a challenge due to disadvantages such as time-consuming and ineffective, which hinders the development of novel materials. Herein, we present a high-throughput combinatorial technique for continuous thin film preparation relied on chemical bath deposition (CBD). The method is ideally used to prepare high-throughput combinatorial material library with low decomposition temperatures and high water- or oxygen-sensitivity at relatively high-temperature. To check this system, a Cu(In, Ga)Se (CIGS) thin films library doped with 0-19.04 at.% of antimony (Sb) was taken as an example to evaluate the regulation of varying Sb doping concentration on the grain growth, structure, morphology and electrical properties of CIGS thin film systemically. Combined with the Energy Dispersive Spectrometer (EDS), X-ray Photoelectron Spectroscopy (XPS), automated X-ray Diffraction (XRD) for rapid screening and Localized Electrochemical Impedance Spectroscopy (LEIS), it was confirmed that this combinatorial high-throughput system could be used to identify the composition with the optimal grain orientation growth, microstructure and electrical properties systematically, through accurately monitoring the doping content and material composition. According to the characterization results, a Sb2Se3 quasi-liquid phase promoted CIGS film-growth model has been put forward. In addition to CIGS thin film reported here, the combinatorial CBD also could be applied to the high-throughput screening of other sulfide thin film material systems.

  10. Optical tools for high-throughput screening of abrasion resistance of combinatorial libraries of organic coatings

    Science.gov (United States)

    Potyrailo, Radislav A.; Chisholm, Bret J.; Olson, Daniel R.; Brennan, Michael J.; Molaison, Chris A.

    2002-02-01

    Design, validation, and implementation of an optical spectroscopic system for high-throughput analysis of combinatorially developed protective organic coatings are reported. Our approach replaces labor-intensive coating evaluation steps with an automated system that rapidly analyzes 8x6 arrays of coating elements that are deposited on a plastic substrate. Each coating element of the library is 10 mm in diameter and 2 to 5 micrometers thick. Performance of coatings is evaluated with respect to their resistance to wear abrasion because this parameter is one of the primary considerations in end-use applications. Upon testing, the organic coatings undergo changes that are impossible to quantitatively predict using existing knowledge. Coatings are abraded using industry-accepted abrasion test methods at single-or multiple-abrasion conditions, followed by high- throughput analysis of abrasion-induced light scatter. The developed automated system is optimized for the analysis of diffusively scattered light that corresponds to 0 to 30% haze. System precision of 0.1 to 2.5% relative standard deviation provides capability for the reliable ranking of coatings performance. While the system was implemented for high-throughput screening of combinatorially developed organic protective coatings for automotive applications, it can be applied to a variety of other applications where materials ranking can be achieved using optical spectroscopic tools.

  11. High-throughput genotyping of single nucleotide polymorphisms with rolling circle amplification

    Directory of Open Access Journals (Sweden)

    Sun Zhenyu

    2001-08-01

    Full Text Available Abstract Background Single nucleotide polymorphisms (SNPs are the foundation of powerful complex trait and pharmacogenomic analyses. The availability of large SNP databases, however, has emphasized a need for inexpensive SNP genotyping methods of commensurate simplicity, robustness, and scalability. We describe a solution-based, microtiter plate method for SNP genotyping of human genomic DNA. The method is based upon allele discrimination by ligation of open circle probes followed by rolling circle amplification of the signal using fluorescent primers. Only the probe with a 3' base complementary to the SNP is circularized by ligation. Results SNP scoring by ligation was optimized to a 100,000 fold discrimination against probe mismatched to the SNP. The assay was used to genotype 10 SNPs from a set of 192 genomic DNA samples in a high-throughput format. Assay directly from genomic DNA eliminates the need to preamplify the target as done for many other genotyping methods. The sensitivity of the assay was demonstrated by genotyping from 1 ng of genomic DNA. We demonstrate that the assay can detect a single molecule of the circularized probe. Conclusions Compatibility with homogeneous formats and the ability to assay small amounts of genomic DNA meets the exacting requirements of automated, high-throughput SNP scoring.

  12. The Protein Maker: an automated system for high-throughput parallel purification

    International Nuclear Information System (INIS)

    Smith, Eric R.; Begley, Darren W.; Anderson, Vanessa; Raymond, Amy C.; Haffner, Taryn E.; Robinson, John I.; Edwards, Thomas E.; Duncan, Natalie; Gerdts, Cory J.; Mixon, Mark B.; Nollert, Peter; Staker, Bart L.; Stewart, Lance J.

    2011-01-01

    The Protein Maker instrument addresses a critical bottleneck in structural genomics by allowing automated purification and buffer testing of multiple protein targets in parallel with a single instrument. Here, the use of this instrument to (i) purify multiple influenza-virus proteins in parallel for crystallization trials and (ii) identify optimal lysis-buffer conditions prior to large-scale protein purification is described. The Protein Maker is an automated purification system developed by Emerald BioSystems for high-throughput parallel purification of proteins and antibodies. This instrument allows multiple load, wash and elution buffers to be used in parallel along independent lines for up to 24 individual samples. To demonstrate its utility, its use in the purification of five recombinant PB2 C-terminal domains from various subtypes of the influenza A virus is described. Three of these constructs crystallized and one diffracted X-rays to sufficient resolution for structure determination and deposition in the Protein Data Bank. Methods for screening lysis buffers for a cytochrome P450 from a pathogenic fungus prior to upscaling expression and purification are also described. The Protein Maker has become a valuable asset within the Seattle Structural Genomics Center for Infectious Disease (SSGCID) and hence is a potentially valuable tool for a variety of high-throughput protein-purification applications

  13. High Performance Computing Modernization Program Kerberos Throughput Test Report

    Science.gov (United States)

    2017-10-26

    Naval Research Laboratory Washington, DC 20375-5320 NRL/MR/5524--17-9751 High Performance Computing Modernization Program Kerberos Throughput Test ...NUMBER 5d. PROJECT NUMBER 5e. TASK NUMBER 5f. WORK UNIT NUMBER 2. REPORT TYPE1. REPORT DATE (DD-MM-YYYY) 4. TITLE AND SUBTITLE 6. AUTHOR(S) 8. PERFORMING...PAGE 18. NUMBER OF PAGES 17. LIMITATION OF ABSTRACT High Performance Computing Modernization Program Kerberos Throughput Test Report Daniel G. Gdula* and

  14. High-Throughput Block Optical DNA Sequence Identification.

    Science.gov (United States)

    Sagar, Dodderi Manjunatha; Korshoj, Lee Erik; Hanson, Katrina Bethany; Chowdhury, Partha Pratim; Otoupal, Peter Britton; Chatterjee, Anushree; Nagpal, Prashant

    2018-01-01

    Optical techniques for molecular diagnostics or DNA sequencing generally rely on small molecule fluorescent labels, which utilize light with a wavelength of several hundred nanometers for detection. Developing a label-free optical DNA sequencing technique will require nanoscale focusing of light, a high-throughput and multiplexed identification method, and a data compression technique to rapidly identify sequences and analyze genomic heterogeneity for big datasets. Such a method should identify characteristic molecular vibrations using optical spectroscopy, especially in the "fingerprinting region" from ≈400-1400 cm -1 . Here, surface-enhanced Raman spectroscopy is used to demonstrate label-free identification of DNA nucleobases with multiplexed 3D plasmonic nanofocusing. While nanometer-scale mode volumes prevent identification of single nucleobases within a DNA sequence, the block optical technique can identify A, T, G, and C content in DNA k-mers. The content of each nucleotide in a DNA block can be a unique and high-throughput method for identifying sequences, genes, and other biomarkers as an alternative to single-letter sequencing. Additionally, coupling two complementary vibrational spectroscopy techniques (infrared and Raman) can improve block characterization. These results pave the way for developing a novel, high-throughput block optical sequencing method with lossy genomic data compression using k-mer identification from multiplexed optical data acquisition. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. High-Throughput Scoring of Seed Germination.

    Science.gov (United States)

    Ligterink, Wilco; Hilhorst, Henk W M

    2017-01-01

    High-throughput analysis of seed germination for phenotyping large genetic populations or mutant collections is very labor intensive and would highly benefit from an automated setup. Although very often used, the total germination percentage after a nominated period of time is not very informative as it lacks information about start, rate, and uniformity of germination, which are highly indicative of such traits as dormancy, stress tolerance, and seed longevity. The calculation of cumulative germination curves requires information about germination percentage at various time points. We developed the GERMINATOR package: a simple, highly cost-efficient, and flexible procedure for high-throughput automatic scoring and evaluation of germination that can be implemented without the use of complex robotics. The GERMINATOR package contains three modules: (I) design of experimental setup with various options to replicate and randomize samples; (II) automatic scoring of germination based on the color contrast between the protruding radicle and seed coat on a single image; and (III) curve fitting of cumulative germination data and the extraction, recap, and visualization of the various germination parameters. GERMINATOR is a freely available package that allows the monitoring and analysis of several thousands of germination tests, several times a day by a single person.

  16. High throughput salt separation from uranium deposits

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, S.W.; Park, K.M.; Kim, J.G.; Kim, I.T.; Park, S.B., E-mail: swkwon@kaeri.re.kr [Korea Atomic Energy Research Inst. (Korea, Republic of)

    2014-07-01

    It is very important to increase the throughput of the salt separation system owing to the high uranium content of spent nuclear fuel and high salt fraction of uranium dendrites in pyroprocessing. Multilayer porous crucible system was proposed to increase a throughput of the salt distiller in this study. An integrated sieve-crucible assembly was also investigated for the practical use of the porous crucible system. The salt evaporation behaviors were compared between the conventional nonporous crucible and the porous crucible. Two step weight reductions took place in the porous crucible, whereas the salt weight reduced only at high temperature by distillation in a nonporous crucible. The first weight reduction in the porous crucible was caused by the liquid salt penetrated out through the perforated crucible during the temperature elevation until the distillation temperature. Multilayer porous crucibles have a benefit to expand the evaporation surface area. (author)

  17. High throughput, low set-up time reconfigurable linear feedback shift registers

    NARCIS (Netherlands)

    Nas, R.J.M.; Berkel, van C.H.

    2010-01-01

    This paper presents a hardware design for a scalable, high throughput, configurable LFSR. High throughput is achieved by producing L consecutive outputs per clock cycle with a clock cycle period that, for practical cases, increases only logarithmically with the block size L and the length of the

  18. High-throughput epitope identification for snakebite antivenom

    DEFF Research Database (Denmark)

    Engmark, Mikael; De Masi, Federico; Laustsen, Andreas Hougaard

    Insight into the epitopic recognition pattern for polyclonal antivenoms is a strong tool for accurate prediction of antivenom cross-reactivity and provides a basis for design of novel antivenoms. In this work, a high-throughput approach was applied to characterize linear epitopes in 966 individua...... toxins from pit vipers (Crotalidae) using the ICP Crotalidae antivenom. Due to an abundance of snake venom metalloproteinases and phospholipase A2s in the venoms used for production of the investigated antivenom, this study focuses on these toxin families.......Insight into the epitopic recognition pattern for polyclonal antivenoms is a strong tool for accurate prediction of antivenom cross-reactivity and provides a basis for design of novel antivenoms. In this work, a high-throughput approach was applied to characterize linear epitopes in 966 individual...

  19. Formation of Linear Gradient of Antibiotics on Microfluidic Chips for High-throughput Antibiotic Susceptibility Testing

    Science.gov (United States)

    Kim, Seunggyu; Lee, Seokhun; Jeon, Jessie S.

    2017-11-01

    To determine the most effective antimicrobial treatments of infectious pathogen, high-throughput antibiotic susceptibility test (AST) is critically required. However, the conventional AST requires at least 16 hours to reach the minimum observable population. Therefore, we developed a microfluidic system that allows maintenance of linear antibiotic concentration and measurement of local bacterial density. Based on the Stokes-Einstein equation, the flow rate in the microchannel was optimized so that linearization was achieved within 10 minutes, taking into account the diffusion coefficient of each antibiotic in the agar gel. As a result, the minimum inhibitory concentration (MIC) of each antibiotic against P. aeruginosa could be immediately determined 6 hours after treatment of the linear antibiotic concentration. In conclusion, our system proved the efficacy of a high-throughput AST platform through MIC comparison with Clinical and Laboratory Standards Institute (CLSI) range of antibiotics. This work was supported by the Climate Change Research Hub (Grant No. N11170060) of the KAIST and by the Brain Korea 21 Plus project.

  20. Towards a high throughput droplet-based agglutination assay

    KAUST Repository

    Kodzius, Rimantas; Castro, David; Foulds, Ian G.

    2013-01-01

    This work demonstrates the detection method for a high throughput droplet based agglutination assay system. Using simple hydrodynamic forces to mix and aggregate functionalized microbeads we avoid the need to use magnetic assistance or mixing structures. The concentration of our target molecules was estimated by agglutination strength, obtained through optical image analysis. Agglutination in droplets was performed with flow rates of 150 µl/min and occurred in under a minute, with potential to perform high-throughput measurements. The lowest target concentration detected in droplet microfluidics was 0.17 nM, which is three orders of magnitude more sensitive than a conventional card based agglutination assay.

  1. Towards a high throughput droplet-based agglutination assay

    KAUST Repository

    Kodzius, Rimantas

    2013-10-22

    This work demonstrates the detection method for a high throughput droplet based agglutination assay system. Using simple hydrodynamic forces to mix and aggregate functionalized microbeads we avoid the need to use magnetic assistance or mixing structures. The concentration of our target molecules was estimated by agglutination strength, obtained through optical image analysis. Agglutination in droplets was performed with flow rates of 150 µl/min and occurred in under a minute, with potential to perform high-throughput measurements. The lowest target concentration detected in droplet microfluidics was 0.17 nM, which is three orders of magnitude more sensitive than a conventional card based agglutination assay.

  2. High-throughput screening of small molecule libraries using SAMDI mass spectrometry.

    Science.gov (United States)

    Gurard-Levin, Zachary A; Scholle, Michael D; Eisenberg, Adam H; Mrksich, Milan

    2011-07-11

    High-throughput screening is a common strategy used to identify compounds that modulate biochemical activities, but many approaches depend on cumbersome fluorescent reporters or antibodies and often produce false-positive hits. The development of "label-free" assays addresses many of these limitations, but current approaches still lack the throughput needed for applications in drug discovery. This paper describes a high-throughput, label-free assay that combines self-assembled monolayers with mass spectrometry, in a technique called SAMDI, as a tool for screening libraries of 100,000 compounds in one day. This method is fast, has high discrimination, and is amenable to a broad range of chemical and biological applications.

  3. Status Of The Development Of A Thin Foil High Throughput X-Ray Telescope For The Soviet Spectrum X-Gamma Mission

    DEFF Research Database (Denmark)

    WESTERGAARD, NJ; BYRNAK, BP; Christensen, Finn Erland

    1989-01-01

    modification of this design is optimized with respect to high energy throughput of the telescope. The mechanical design and the status of the surface preparation technologies are described. Various X-ray and optical test facilities for the measurement of surface roughness, "orange peel", and figure errors...

  4. Preliminary High-Throughput Metagenome Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Dusheyko, Serge; Furman, Craig; Pangilinan, Jasmyn; Shapiro, Harris; Tu, Hank

    2007-03-26

    Metagenome data sets present a qualitatively different assembly problem than traditional single-organism whole-genome shotgun (WGS) assembly. The unique aspects of such projects include the presence of a potentially large number of distinct organisms and their representation in the data set at widely different fractions. In addition, multiple closely related strains could be present, which would be difficult to assemble separately. Failure to take these issues into account can result in poor assemblies that either jumble together different strains or which fail to yield useful results. The DOE Joint Genome Institute has sequenced a number of metagenomic projects and plans to considerably increase this number in the coming year. As a result, the JGI has a need for high-throughput tools and techniques for handling metagenome projects. We present the techniques developed to handle metagenome assemblies in a high-throughput environment. This includes a streamlined assembly wrapper, based on the JGI?s in-house WGS assembler, Jazz. It also includes the selection of sensible defaults targeted for metagenome data sets, as well as quality control automation for cleaning up the raw results. While analysis is ongoing, we will discuss preliminary assessments of the quality of the assembly results (http://fames.jgi-psf.org).

  5. High-throughput STR analysis for DNA database using direct PCR.

    Science.gov (United States)

    Sim, Jeong Eun; Park, Su Jeong; Lee, Han Chul; Kim, Se-Yong; Kim, Jong Yeol; Lee, Seung Hwan

    2013-07-01

    Since the Korean criminal DNA database was launched in 2010, we have focused on establishing an automated DNA database profiling system that analyzes short tandem repeat loci in a high-throughput and cost-effective manner. We established a DNA database profiling system without DNA purification using a direct PCR buffer system. The quality of direct PCR procedures was compared with that of conventional PCR system under their respective optimized conditions. The results revealed not only perfect concordance but also an excellent PCR success rate, good electropherogram quality, and an optimal intra/inter-loci peak height ratio. In particular, the proportion of DNA extraction required due to direct PCR failure could be minimized to <3%. In conclusion, the newly developed direct PCR system can be adopted for automated DNA database profiling systems to replace or supplement conventional PCR system in a time- and cost-saving manner. © 2013 American Academy of Forensic Sciences Published 2013. This article is a U.S. Government work and is in the public domain in the U.S.A.

  6. Optimization of Monte Carlo particle transport parameters and validation of a novel high throughput experimental setup to measure the biological effects of particle beams.

    Science.gov (United States)

    Patel, Darshana; Bronk, Lawrence; Guan, Fada; Peeler, Christopher R; Brons, Stephan; Dokic, Ivana; Abdollahi, Amir; Rittmüller, Claudia; Jäkel, Oliver; Grosshans, David; Mohan, Radhe; Titt, Uwe

    2017-11-01

    Accurate modeling of the relative biological effectiveness (RBE) of particle beams requires increased systematic in vitro studies with human cell lines with care towards minimizing uncertainties in biologic assays as well as physical parameters. In this study, we describe a novel high-throughput experimental setup and an optimized parameterization of the Monte Carlo (MC) simulation technique that is universally applicable for accurate determination of RBE of clinical ion beams. Clonogenic cell-survival measurements on a human lung cancer cell line (H460) are presented using proton irradiation. Experiments were performed at the Heidelberg Ion Therapy Center (HIT) with support from the Deutsches Krebsforschungszentrum (DKFZ) in Heidelberg, Germany using a mono-energetic horizontal proton beam. A custom-made variable range selector was designed for the horizontal beam line using the Geant4 MC toolkit. This unique setup enabled a high-throughput clonogenic assay investigation of multiple, well defined dose and linear energy transfer (LETs) per irradiation for human lung cancer cells (H460) cultured in a 96-well plate. Sensitivity studies based on application of different physics lists in conjunction with different electromagnetic constructors and production threshold values to the MC simulations were undertaken for accurate assessment of the calculated dose and the dose-averaged LET (LET d ). These studies were extended to helium and carbon ion beams. Sensitivity analysis of the MC parameterization revealed substantial dependence of the dose and LET d values on both the choice of physics list and the production threshold values. While the dose and LET d calculations using FTFP_BERT_LIV, FTFP_BERT_EMZ, FTFP_BERT_PEN and QGSP_BIC_EMY physics lists agree well with each other for all three ions, they show large differences when compared to the FTFP_BERT physics list with the default electromagnetic constructor. For carbon ions, the dose corresponding to the largest LET d

  7. A high-throughput multiplex method adapted for GMO detection.

    Science.gov (United States)

    Chaouachi, Maher; Chupeau, Gaëlle; Berard, Aurélie; McKhann, Heather; Romaniuk, Marcel; Giancola, Sandra; Laval, Valérie; Bertheau, Yves; Brunel, Dominique

    2008-12-24

    A high-throughput multiplex assay for the detection of genetically modified organisms (GMO) was developed on the basis of the existing SNPlex method designed for SNP genotyping. This SNPlex assay allows the simultaneous detection of up to 48 short DNA sequences (approximately 70 bp; "signature sequences") from taxa endogenous reference genes, from GMO constructions, screening targets, construct-specific, and event-specific targets, and finally from donor organisms. This assay avoids certain shortcomings of multiplex PCR-based methods already in widespread use for GMO detection. The assay demonstrated high specificity and sensitivity. The results suggest that this assay is reliable, flexible, and cost- and time-effective for high-throughput GMO detection.

  8. High throughput two-step ultrasonic spray deposited CH3NH3PbI3 thin film layer for solar cell application

    Science.gov (United States)

    Lan, Ding-Hung; Hong, Shao-Huan; Chou, Li-Hui; Wang, Xiao-Feng; Liu, Cheng-Liang

    2018-06-01

    Organometal halide perovskite materials have demonstrated tremendous advances in the photovoltaic field recently because of their advantageous features of simple fabrication and high power conversion efficiency. To meet the high demand for high throughput and cost-effective, we present a wet process method that enables the probing of the parameters for perovskite layer deposition through two-step sequential ultrasonic spray-coating. This paper describes a detailed investigation on the effects of modification of spray precursor solution (PbI2 and CH3NH3I precursor concentration and solvents used) and post-annealing condition (temperature and time), which can be performed to create optimal film quality as well as improve device efficiency. Through the systematic optimization, the inverted planar perovskite solar cells show the reproducible photovoltaic properties with best power conversion efficiency (PCE) of 10.40% and average PCE of 9.70 ± 0.40%. A continuous spray-coating technique for rapid fabrication of total 16 pieces of perovskite films was demonstrated for providing a viable alternative for the high throughput production of the perovskite solar cells.

  9. High efficient plastic solar cells fabricated with a high-throughput gravure printing method

    Energy Technology Data Exchange (ETDEWEB)

    Kopola, P.; Jin, H.; Tuomikoski, M.; Maaninen, A.; Hast, J. [VTT, Kaitovaeylae 1, FIN-90571 Oulu (Finland); Aernouts, T. [IMEC, Organic PhotoVoltaics, Polymer and Molecular Electronics, Kapeldreef 75, B-3001 Leuven (Belgium); Guillerez, S. [CEA-INES RDI, 50 Avenue Du Lac Leman, 73370 Le Bourget Du Lac (France)

    2010-10-15

    We report on polymer-based solar cells prepared by the high-throughput roll-to-roll gravure printing method. The engravings of the printing plate, along with process parameters like printing speed and ink properties, are studied to optimise the printability of the photoactive as well as the hole transport layer. For the hole transport layer, the focus is on testing different formulations to produce thorough wetting of the indium-tin-oxide (ITO) substrate. The challenge for the photoactive layer is to form a uniform layer with optimal nanomorphology in the poly-3-hexylthiophene (P3HT) and [6,6]-phenyl-C61-butyric acid methyl ester (PCBM) blend. This results in a power conversion efficiency of 2.8% under simulated AM1.5G solar illumination for a solar cell device with gravure-printed hole transport and a photoactive layer. (author)

  10. High Throughput Analysis of Photocatalytic Water Purification

    NARCIS (Netherlands)

    Sobral Romao, J.I.; Baiao Barata, David; Habibovic, Pamela; Mul, Guido; Baltrusaitis, Jonas

    2014-01-01

    We present a novel high throughput photocatalyst efficiency assessment method based on 96-well microplates and UV-Vis spectroscopy. We demonstrate the reproducibility of the method using methyl orange (MO) decomposition, and compare kinetic data obtained with those provided in the literature for

  11. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale L

    2005-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  12. High Throughput Synthesis and Screening for Agents Inhibiting Androgen Receptor Mediated Gene Transcription

    National Research Council Canada - National Science Library

    Boger, Dale

    2004-01-01

    .... This entails the high throughput synthesis of DNA binding agents related to distamycin, their screening for binding to androgen response elements using a new high throughput DNA binding screen...

  13. A CRISPR CASe for High-Throughput Silencing

    Directory of Open Access Journals (Sweden)

    Jacob eHeintze

    2013-10-01

    Full Text Available Manipulation of gene expression on a genome-wide level is one of the most important systematic tools in the post-genome era. Such manipulations have largely been enabled by expression cloning approaches using sequence-verified cDNA libraries, large-scale RNA interference libraries (shRNA or siRNA and zinc finger nuclease technologies. More recently, the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated (Cas9-mediated gene editing technology has been described that holds great promise for future use of this technology in genomic manipulation. It was suggested that the CRISPR system has the potential to be used in high-throughput, large-scale loss of function screening. Here we discuss some of the challenges in engineering of CRISPR/Cas genomic libraries and some of the aspects that need to be addressed in order to use this technology on a high-throughput scale.

  14. High-Throughput Screening of the Asymmetric Decarboxylative Alkylation Reaction of Enolate-Stabilized Enol Carbonates

    KAUST Repository

    Stoltz, Brian

    2010-06-14

    The use of high-throughput screening allowed for the optimization of reaction conditions for the palladium-catalyzed asymmetric decarboxylative alkylation reaction of enolate-stabilized enol carbonates. Changing to a non-polar reaction solvent and to an electron-deficient PHOX derivative as ligand from our standard reaction conditions improved the enantioselectivity for the alkylation of a ketal-protected,1,3-diketone-derived enol carbonate from 28% ee to 84% ee. Similar improvements in enantioselectivity were seen for a β-keto-ester derived- and an α-phenyl cyclohexanone-derived enol carbonate.

  15. High-Throughput Screening of the Asymmetric Decarboxylative Alkylation Reaction of Enolate-Stabilized Enol Carbonates

    KAUST Repository

    Stoltz, Brian; McDougal, Nolan; Virgil, Scott

    2010-01-01

    The use of high-throughput screening allowed for the optimization of reaction conditions for the palladium-catalyzed asymmetric decarboxylative alkylation reaction of enolate-stabilized enol carbonates. Changing to a non-polar reaction solvent and to an electron-deficient PHOX derivative as ligand from our standard reaction conditions improved the enantioselectivity for the alkylation of a ketal-protected,1,3-diketone-derived enol carbonate from 28% ee to 84% ee. Similar improvements in enantioselectivity were seen for a β-keto-ester derived- and an α-phenyl cyclohexanone-derived enol carbonate.

  16. Evaluation of a pooled strategy for high-throughput sequencing of cosmid clones from metagenomic libraries.

    Science.gov (United States)

    Lam, Kathy N; Hall, Michael W; Engel, Katja; Vey, Gregory; Cheng, Jiujun; Neufeld, Josh D; Charles, Trevor C

    2014-01-01

    High-throughput sequencing methods have been instrumental in the growing field of metagenomics, with technological improvements enabling greater throughput at decreased costs. Nonetheless, the economy of high-throughput sequencing cannot be fully leveraged in the subdiscipline of functional metagenomics. In this area of research, environmental DNA is typically cloned to generate large-insert libraries from which individual clones are isolated, based on specific activities of interest. Sequence data are required for complete characterization of such clones, but the sequencing of a large set of clones requires individual barcode-based sample preparation; this can become costly, as the cost of clone barcoding scales linearly with the number of clones processed, and thus sequencing a large number of metagenomic clones often remains cost-prohibitive. We investigated a hybrid Sanger/Illumina pooled sequencing strategy that omits barcoding altogether, and we evaluated this strategy by comparing the pooled sequencing results to reference sequence data obtained from traditional barcode-based sequencing of the same set of clones. Using identity and coverage metrics in our evaluation, we show that pooled sequencing can generate high-quality sequence data, without producing problematic chimeras. Though caveats of a pooled strategy exist and further optimization of the method is required to improve recovery of complete clone sequences and to avoid circumstances that generate unrecoverable clone sequences, our results demonstrate that pooled sequencing represents an effective and low-cost alternative for sequencing large sets of metagenomic clones.

  17. High throughput route selection in multi-rate wireless mesh networks

    Institute of Scientific and Technical Information of China (English)

    WEI Yi-fei; GUO Xiang-li; SONG Mei; SONG Jun-de

    2008-01-01

    Most existing Ad-hoc routing protocols use the shortest path algorithm with a hop count metric to select paths. It is appropriate in single-rate wireless networks, but has a tendency to select paths containing long-distance links that have low data rates and reduced reliability in multi-rate networks. This article introduces a high throughput routing algorithm utilizing the multi-rate capability and some mesh characteristics in wireless fidelity (WiFi) mesh networks. It uses the medium access control (MAC) transmission time as the routing metric, which is estimated by the information passed up from the physical layer. When the proposed algorithm is adopted, the Ad-hoc on-demand distance vector (AODV) routing can be improved as high throughput AODV (HT-AODV). Simulation results show that HT-AODV is capable of establishing a route that has high data-rate, short end-to-end delay and great network throughput.

  18. Alginate Immobilization of Metabolic Enzymes (AIME) for High-Throughput Screening Assays (SOT)

    Science.gov (United States)

    Alginate Immobilization of Metabolic Enzymes (AIME) for High-Throughput Screening Assays DE DeGroot, RS Thomas, and SO SimmonsNational Center for Computational Toxicology, US EPA, Research Triangle Park, NC USAThe EPA’s ToxCast program utilizes a wide variety of high-throughput s...

  19. A high-throughput sample preparation method for cellular proteomics using 96-well filter plates.

    Science.gov (United States)

    Switzar, Linda; van Angeren, Jordy; Pinkse, Martijn; Kool, Jeroen; Niessen, Wilfried M A

    2013-10-01

    A high-throughput sample preparation protocol based on the use of 96-well molecular weight cutoff (MWCO) filter plates was developed for shotgun proteomics of cell lysates. All sample preparation steps, including cell lysis, buffer exchange, protein denaturation, reduction, alkylation and proteolytic digestion are performed in a 96-well plate format, making the platform extremely well suited for processing large numbers of samples and directly compatible with functional assays for cellular proteomics. In addition, the usage of a single plate for all sample preparation steps following cell lysis reduces potential samples losses and allows for automation. The MWCO filter also enables sample concentration, thereby increasing the overall sensitivity, and implementation of washing steps involving organic solvents, for example, to remove cell membranes constituents. The optimized protocol allowed for higher throughput with improved sensitivity in terms of the number of identified cellular proteins when compared to an established protocol employing gel-filtration columns. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. High-throughput phenotyping and genomic selection: the frontiers of crop breeding converge.

    Science.gov (United States)

    Cabrera-Bosquet, Llorenç; Crossa, José; von Zitzewitz, Jarislav; Serret, María Dolors; Araus, José Luis

    2012-05-01

    Genomic selection (GS) and high-throughput phenotyping have recently been captivating the interest of the crop breeding community from both the public and private sectors world-wide. Both approaches promise to revolutionize the prediction of complex traits, including growth, yield and adaptation to stress. Whereas high-throughput phenotyping may help to improve understanding of crop physiology, most powerful techniques for high-throughput field phenotyping are empirical rather than analytical and comparable to genomic selection. Despite the fact that the two methodological approaches represent the extremes of what is understood as the breeding process (phenotype versus genome), they both consider the targeted traits (e.g. grain yield, growth, phenology, plant adaptation to stress) as a black box instead of dissecting them as a set of secondary traits (i.e. physiological) putatively related to the target trait. Both GS and high-throughput phenotyping have in common their empirical approach enabling breeders to use genome profile or phenotype without understanding the underlying biology. This short review discusses the main aspects of both approaches and focuses on the case of genomic selection of maize flowering traits and near-infrared spectroscopy (NIRS) and plant spectral reflectance as high-throughput field phenotyping methods for complex traits such as crop growth and yield. © 2012 Institute of Botany, Chinese Academy of Sciences.

  1. High-throughput single nucleotide polymorphism genotyping using nanofluidic Dynamic Arrays

    Directory of Open Access Journals (Sweden)

    Crenshaw Andrew

    2009-01-01

    Full Text Available Abstract Background Single nucleotide polymorphisms (SNPs have emerged as the genetic marker of choice for mapping disease loci and candidate gene association studies, because of their high density and relatively even distribution in the human genomes. There is a need for systems allowing medium multiplexing (ten to hundreds of SNPs with high throughput, which can efficiently and cost-effectively generate genotypes for a very large sample set (thousands of individuals. Methods that are flexible, fast, accurate and cost-effective are urgently needed. This is also important for those who work on high throughput genotyping in non-model systems where off-the-shelf assays are not available and a flexible platform is needed. Results We demonstrate the use of a nanofluidic Integrated Fluidic Circuit (IFC - based genotyping system for medium-throughput multiplexing known as the Dynamic Array, by genotyping 994 individual human DNA samples on 47 different SNP assays, using nanoliter volumes of reagents. Call rates of greater than 99.5% and call accuracies of greater than 99.8% were achieved from our study, which demonstrates that this is a formidable genotyping platform. The experimental set up is very simple, with a time-to-result for each sample of about 3 hours. Conclusion Our results demonstrate that the Dynamic Array is an excellent genotyping system for medium-throughput multiplexing (30-300 SNPs, which is simple to use and combines rapid throughput with excellent call rates, high concordance and low cost. The exceptional call rates and call accuracy obtained may be of particular interest to those working on validation and replication of genome- wide- association (GWA studies.

  2. HTTK: R Package for High-Throughput Toxicokinetics

    Science.gov (United States)

    Thousands of chemicals have been profiled by high-throughput screening programs such as ToxCast and Tox21; these chemicals are tested in part because most of them have limited or no data on hazard, exposure, or toxicokinetics. Toxicokinetic models aid in predicting tissue concent...

  3. A high-throughput, multi-channel photon-counting detector with picosecond timing

    CERN Document Server

    Lapington, J S; Miller, G M; Ashton, T J R; Jarron, P; Despeisse, M; Powolny, F; Howorth, J; Milnes, J

    2009-01-01

    High-throughput photon counting with high time resolution is a niche application area where vacuum tubes can still outperform solid-state devices. Applications in the life sciences utilizing time-resolved spectroscopies, particularly in the growing field of proteomics, will benefit greatly from performance enhancements in event timing and detector throughput. The HiContent project is a collaboration between the University of Leicester Space Research Centre, the Microelectronics Group at CERN, Photek Ltd., and end-users at the Gray Cancer Institute and the University of Manchester. The goal is to develop a detector system specifically designed for optical proteomics, capable of high content (multi-parametric) analysis at high throughput. The HiContent detector system is being developed to exploit this niche market. It combines multi-channel, high time resolution photon counting in a single miniaturized detector system with integrated electronics. The combination of enabling technologies; small pore microchanne...

  4. Space Link Extension Protocol Emulation for High-Throughput, High-Latency Network Connections

    Science.gov (United States)

    Tchorowski, Nicole; Murawski, Robert

    2014-01-01

    New space missions require higher data rates and new protocols to meet these requirements. These high data rate space communication links push the limitations of not only the space communication links, but of the ground communication networks and protocols which forward user data to remote ground stations (GS) for transmission. The Consultative Committee for Space Data Systems, (CCSDS) Space Link Extension (SLE) standard protocol is one protocol that has been proposed for use by the NASA Space Network (SN) Ground Segment Sustainment (SGSS) program. New protocol implementations must be carefully tested to ensure that they provide the required functionality, especially because of the remote nature of spacecraft. The SLE protocol standard has been tested in the NASA Glenn Research Center's SCENIC Emulation Lab in order to observe its operation under realistic network delay conditions. More specifically, the delay between then NASA Integrated Services Network (NISN) and spacecraft has been emulated. The round trip time (RTT) delay for the continental NISN network has been shown to be up to 120ms; as such the SLE protocol was tested with network delays ranging from 0ms to 200ms. Both a base network condition and an SLE connection were tested with these RTT delays, and the reaction of both network tests to the delay conditions were recorded. Throughput for both of these links was set at 1.2Gbps. The results will show that, in the presence of realistic network delay, the SLE link throughput is significantly reduced while the base network throughput however remained at the 1.2Gbps specification. The decrease in SLE throughput has been attributed to the implementation's use of blocking calls. The decrease in throughput is not acceptable for high data rate links, as the link requires constant data a flow in order for spacecraft and ground radios to stay synchronized, unless significant data is queued a the ground station. In cases where queuing the data is not an option

  5. Engineering a vitamin B12 high-throughput screening system by riboswitch sensor in Sinorhizobium meliloti.

    Science.gov (United States)

    Cai, Yingying; Xia, Miaomiao; Dong, Huina; Qian, Yuan; Zhang, Tongcun; Zhu, Beiwei; Wu, Jinchuan; Zhang, Dawei

    2018-05-11

    As a very important coenzyme in the cell metabolism, Vitamin B 12 (cobalamin, VB 12 ) has been widely used in food and medicine fields. The complete biosynthesis of VB 12 requires approximately 30 genes, but overexpression of these genes did not result in expected increase of VB 12 production. High-yield VB 12 -producing strains are usually obtained by mutagenesis treatments, thus developing an efficient screening approach is urgently needed. By the help of engineered strains with varied capacities of VB 12 production, a riboswitch library was constructed and screened, and the btuB element from Salmonella typhimurium was identified as the best regulatory device. A flow cytometry high-throughput screening system was developed based on the btuB riboswitch with high efficiency to identify positive mutants. Mutation of Sinorhizobium meliloti (S. meliloti) was optimized using the novel mutation technique of atmospheric and room temperature plasma (ARTP). Finally, the mutant S. meliloti MC5-2 was obtained and considered as a candidate for industrial applications. After 7 d's cultivation on a rotary shaker at 30 °C, the VB 12 titer of S. meliloti MC5-2 reached 156 ± 4.2 mg/L, which was 21.9% higher than that of the wild type strain S. meliloti 320 (128 ± 3.2 mg/L). The genome of S. meliloti MC5-2 was sequenced, and gene mutations were identified and analyzed. To our knowledge, it is the first time that a riboswitch element was used in S. meliloti. The flow cytometry high-throughput screening system was successfully developed and a high-yield VB 12 producing strain was obtained. The identified and analyzed gene mutations gave useful information for developing high-yield strains by metabolic engineering. Overall, this work provides a useful high-throughput screening method for developing high VB 12 -yield strains.

  6. Development of a Rapid Fluorescence-Based High-Throughput Screening Assay to Identify Novel Kynurenine 3-Monooxygenase Inhibitor Scaffolds.

    Science.gov (United States)

    Jacobs, K R; Guillemin, G J; Lovejoy, D B

    2018-02-01

    Kynurenine 3-monooxygenase (KMO) is a well-validated therapeutic target for the treatment of neurodegenerative diseases, including Alzheimer's disease (AD) and Huntington's disease (HD). This work reports a facile fluorescence-based KMO assay optimized for high-throughput screening (HTS) that achieves a throughput approximately 20-fold higher than the fastest KMO assay currently reported. The screen was run with excellent performance (average Z' value of 0.80) from 110,000 compounds across 341 plates and exceeded all statistical parameters used to describe a robust HTS assay. A subset of molecules was selected for validation by ultra-high-performance liquid chromatography, resulting in the confirmation of a novel hit with an IC 50 comparable to that of the well-described KMO inhibitor Ro-61-8048. A medicinal chemistry program is currently underway to further develop our novel KMO inhibitor scaffolds.

  7. Optimizing SIEM Throughput on the Cloud Using Parallelization.

    Science.gov (United States)

    Alam, Masoom; Ihsan, Asif; Khan, Muazzam A; Javaid, Qaisar; Khan, Abid; Manzoor, Jawad; Akhundzada, Adnan; Khan, Muhammad Khurram; Farooq, Sajid

    2016-01-01

    Processing large amounts of data in real time for identifying security issues pose several performance challenges, especially when hardware infrastructure is limited. Managed Security Service Providers (MSSP), mostly hosting their applications on the Cloud, receive events at a very high rate that varies from a few hundred to a couple of thousand events per second (EPS). It is critical to process this data efficiently, so that attacks could be identified quickly and necessary response could be initiated. This paper evaluates the performance of a security framework OSTROM built on the Esper complex event processing (CEP) engine under a parallel and non-parallel computational framework. We explain three architectures under which Esper can be used to process events. We investigated the effect on throughput, memory and CPU usage in each configuration setting. The results indicate that the performance of the engine is limited by the number of events coming in rather than the queries being processed. The architecture where 1/4th of the total events are submitted to each instance and all the queries are processed by all the units shows best results in terms of throughput, memory and CPU usage.

  8. Optimizing SIEM Throughput on the Cloud Using Parallelization.

    Directory of Open Access Journals (Sweden)

    Masoom Alam

    Full Text Available Processing large amounts of data in real time for identifying security issues pose several performance challenges, especially when hardware infrastructure is limited. Managed Security Service Providers (MSSP, mostly hosting their applications on the Cloud, receive events at a very high rate that varies from a few hundred to a couple of thousand events per second (EPS. It is critical to process this data efficiently, so that attacks could be identified quickly and necessary response could be initiated. This paper evaluates the performance of a security framework OSTROM built on the Esper complex event processing (CEP engine under a parallel and non-parallel computational framework. We explain three architectures under which Esper can be used to process events. We investigated the effect on throughput, memory and CPU usage in each configuration setting. The results indicate that the performance of the engine is limited by the number of events coming in rather than the queries being processed. The architecture where 1/4th of the total events are submitted to each instance and all the queries are processed by all the units shows best results in terms of throughput, memory and CPU usage.

  9. High throughput automated microbial bioreactor system used for clone selection and rapid scale-down process optimization.

    Science.gov (United States)

    Velez-Suberbie, M Lourdes; Betts, John P J; Walker, Kelly L; Robinson, Colin; Zoro, Barney; Keshavarz-Moore, Eli

    2018-01-01

    High throughput automated fermentation systems have become a useful tool in early bioprocess development. In this study, we investigated a 24 x 15 mL single use microbioreactor system, ambr 15f, designed for microbial culture. We compared the fed-batch growth and production capabilities of this system for two Escherichia coli strains, BL21 (DE3) and MC4100, and two industrially relevant molecules, hGH and scFv. In addition, different carbon sources were tested using bolus, linear or exponential feeding strategies, showing the capacity of the ambr 15f system to handle automated feeding. We used power per unit volume (P/V) as a scale criterion to compare the ambr 15f with 1 L stirred bioreactors which were previously scaled-up to 20 L with a different biological system, thus showing a potential 1,300 fold scale comparability in terms of both growth and product yield. By exposing the cells grown in the ambr 15f system to a level of shear expected in an industrial centrifuge, we determined that the cells are as robust as those from a bench scale bioreactor. These results provide evidence that the ambr 15f system is an efficient high throughput microbial system that can be used for strain and molecule selection as well as rapid scale-up. © 2017 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers Biotechnol. Prog., 34:58-68, 2018. © 2017 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers.

  10. High-throughput screening to identify inhibitors of lysine demethylases.

    Science.gov (United States)

    Gale, Molly; Yan, Qin

    2015-01-01

    Lysine demethylases (KDMs) are epigenetic regulators whose dysfunction is implicated in the pathology of many human diseases including various types of cancer, inflammation and X-linked intellectual disability. Particular demethylases have been identified as promising therapeutic targets, and tremendous efforts are being devoted toward developing suitable small-molecule inhibitors for clinical and research use. Several High-throughput screening strategies have been developed to screen for small-molecule inhibitors of KDMs, each with advantages and disadvantages in terms of time, cost, effort, reliability and sensitivity. In this Special Report, we review and evaluate the High-throughput screening methods utilized for discovery of novel small-molecule KDM inhibitors.

  11. High-throughput approach to the catalytic combustion of diesel soot

    Energy Technology Data Exchange (ETDEWEB)

    Iojoiu, Eduard Emil; Bassou, Badr; Guilhaume, Nolven; Farrusseng, David; Desmartin-Chomel, Arnold; Bianchi, Daniel; Mirodatos, Claude [Institut de recherches sur la catalyse et l' environnement de Lyon IRCELYON, UMR5256 CNRS Universite Lyon 1, 2 avenue Albert Einstein, F-69626 Villeurbanne Cedex (France); Lombaert, Karine [Renault, Diesel Innovative Catalytic Materials, Direction de l' Ingenierie Materiaux, 1 Allee Cornuel, 91510 Lardy (France)

    2008-08-30

    A methodology for the evaluation of diesel soot oxidation catalysts by high-throughput (HT) screening was developed. The optimal experimental conditions (soot amount, catalyst/soot ratio, type of contact, composition and flow rate of gas reactants) ensuring a reliable and reproducible detection of light-off temperatures in a 16 parallel channels reactor were set up. The temperature profile measured in the catalyst/soot bed under TPO conditions when the exothermic combustion of soot takes place was shown to provide an accurate measurement of the ignition. Its reproducibility and relevance were checked. The results obtained with a reference noble metal free catalyst (La{sub 0.8}Cr{sub 0.8}Li{sub 0.2}O{sub 3} perovskite) agree very well with literature data. Qualitative mechanistic features could be derived from these experiments, stressing the likely limiting step of oxygen transfer from catalyst surface to soot particulates to ignite the soot combustion. Ceria material was shown to be more appropriate than perovskite one. From an HT screening of a large diverse library (over 100 mixed oxides catalysts) under optimized conditions, about 10 new formulations were found to perform better than selected noble metal free reference materials. (author)

  12. High throughput production of mouse monoclonal antibodies using antigen microarrays

    DEFF Research Database (Denmark)

    De Masi, Federico; Chiarella, P.; Wilhelm, H.

    2005-01-01

    Recent advances in proteomics research underscore the increasing need for high-affinity monoclonal antibodies, which are still generated with lengthy, low-throughput antibody production techniques. Here we present a semi-automated, high-throughput method of hybridoma generation and identification....... Monoclonal antibodies were raised to different targets in single batch runs of 6-10 wk using multiplexed immunisations, automated fusion and cell-culture, and a novel antigen-coated microarray-screening assay. In a large-scale experiment, where eight mice were immunized with ten antigens each, we generated...

  13. High throughput electrospinning of high-quality nanofibers via an aluminum disk spinneret

    Science.gov (United States)

    Zheng, Guokuo

    In this work, a simple and efficient needleless high throughput electrospinning process using an aluminum disk spinneret with 24 holes is described. Electrospun mats produced by this setup consisted of fine fibers (nano-sized) of the highest quality while the productivity (yield) was many times that obtained from conventional single-needle electrospinning. The goal was to produce scaled-up amounts of the same or better quality nanofibers under variable concentration, voltage, and the working distance than those produced with the single needle lab setting. The fiber mats produced were either polymer or ceramic (such as molybdenum trioxide nanofibers). Through experimentation the optimum process conditions were defined to be: 24 kilovolt, a distance to collector of 15cm. More diluted solutions resulted in smaller diameter fibers. Comparing the morphologies of the nanofibers of MoO3 produced by both the traditional and the high throughput set up it was found that they were very similar. Moreover, the nanofibers production rate is nearly 10 times than that of traditional needle electrospinning. Thus, the high throughput process has the potential to become an industrial nanomanufacturing process and the materials processed by it may be used as filtration devices, in tissue engineering, and as sensors.

  14. High throughput protease profiling comprehensively defines active site specificity for thrombin and ADAMTS13.

    Science.gov (United States)

    Kretz, Colin A; Tomberg, Kärt; Van Esbroeck, Alexander; Yee, Andrew; Ginsburg, David

    2018-02-12

    We have combined random 6 amino acid substrate phage display with high throughput sequencing to comprehensively define the active site specificity of the serine protease thrombin and the metalloprotease ADAMTS13. The substrate motif for thrombin was determined by >6,700 cleaved peptides, and was highly concordant with previous studies. In contrast, ADAMTS13 cleaved only 96 peptides (out of >10 7 sequences), with no apparent consensus motif. However, when the hexapeptide library was substituted into the P3-P3' interval of VWF73, an exosite-engaging substrate of ADAMTS13, 1670 unique peptides were cleaved. ADAMTS13 exhibited a general preference for aliphatic amino acids throughout the P3-P3' interval, except at P2 where Arg was tolerated. The cleaved peptides assembled into a motif dominated by P3 Leu, and bulky aliphatic residues at P1 and P1'. Overall, the P3-P2' amino acid sequence of von Willebrand Factor appears optimally evolved for ADAMTS13 recognition. These data confirm the critical role of exosite engagement for substrates to gain access to the active site of ADAMTS13, and define the substrate recognition motif for ADAMTS13. Combining substrate phage display with high throughput sequencing is a powerful approach for comprehensively defining the active site specificity of proteases.

  15. Solion ion source for high-efficiency, high-throughput solar cell manufacturing

    Energy Technology Data Exchange (ETDEWEB)

    Koo, John, E-mail: john-koo@amat.com; Binns, Brant; Miller, Timothy; Krause, Stephen; Skinner, Wesley; Mullin, James [Applied Materials, Inc., Varian Semiconductor Equipment Business Unit, 35 Dory Road, Gloucester, Massachusetts 01930 (United States)

    2014-02-15

    In this paper, we introduce the Solion ion source for high-throughput solar cell doping. As the source power is increased to enable higher throughput, negative effects degrade the lifetime of the plasma chamber and the extraction electrodes. In order to improve efficiency, we have explored a wide range of electron energies and determined the conditions which best suit production. To extend the lifetime of the source we have developed an in situ cleaning method using only existing hardware. With these combinations, source life-times of >200 h for phosphorous and >100 h for boron ion beams have been achieved while maintaining 1100 cell-per-hour production.

  16. High throughput integrated thermal characterization with non-contact optical calorimetry

    Science.gov (United States)

    Hou, Sichao; Huo, Ruiqing; Su, Ming

    2017-10-01

    Commonly used thermal analysis tools such as calorimeter and thermal conductivity meter are separated instruments and limited by low throughput, where only one sample is examined each time. This work reports an infrared based optical calorimetry with its theoretical foundation, which is able to provide an integrated solution to characterize thermal properties of materials with high throughput. By taking time domain temperature information of spatially distributed samples, this method allows a single device (infrared camera) to determine the thermal properties of both phase change systems (melting temperature and latent heat of fusion) and non-phase change systems (thermal conductivity and heat capacity). This method further allows these thermal properties of multiple samples to be determined rapidly, remotely, and simultaneously. In this proof-of-concept experiment, the thermal properties of a panel of 16 samples including melting temperatures, latent heats of fusion, heat capacities, and thermal conductivities have been determined in 2 min with high accuracy. Given the high thermal, spatial, and temporal resolutions of the advanced infrared camera, this method has the potential to revolutionize the thermal characterization of materials by providing an integrated solution with high throughput, high sensitivity, and short analysis time.

  17. Filtering high-throughput protein-protein interaction data using a combination of genomic features

    Directory of Open Access Journals (Sweden)

    Patil Ashwini

    2005-04-01

    Full Text Available Abstract Background Protein-protein interaction data used in the creation or prediction of molecular networks is usually obtained from large scale or high-throughput experiments. This experimental data is liable to contain a large number of spurious interactions. Hence, there is a need to validate the interactions and filter out the incorrect data before using them in prediction studies. Results In this study, we use a combination of 3 genomic features – structurally known interacting Pfam domains, Gene Ontology annotations and sequence homology – as a means to assign reliability to the protein-protein interactions in Saccharomyces cerevisiae determined by high-throughput experiments. Using Bayesian network approaches, we show that protein-protein interactions from high-throughput data supported by one or more genomic features have a higher likelihood ratio and hence are more likely to be real interactions. Our method has a high sensitivity (90% and good specificity (63%. We show that 56% of the interactions from high-throughput experiments in Saccharomyces cerevisiae have high reliability. We use the method to estimate the number of true interactions in the high-throughput protein-protein interaction data sets in Caenorhabditis elegans, Drosophila melanogaster and Homo sapiens to be 27%, 18% and 68% respectively. Our results are available for searching and downloading at http://helix.protein.osaka-u.ac.jp/htp/. Conclusion A combination of genomic features that include sequence, structure and annotation information is a good predictor of true interactions in large and noisy high-throughput data sets. The method has a very high sensitivity and good specificity and can be used to assign a likelihood ratio, corresponding to the reliability, to each interaction.

  18. Standardized Method for High-throughput Sterilization of Arabidopsis Seeds.

    Science.gov (United States)

    Lindsey, Benson E; Rivero, Luz; Calhoun, Chistopher S; Grotewold, Erich; Brkljacic, Jelena

    2017-10-17

    Arabidopsis thaliana (Arabidopsis) seedlings often need to be grown on sterile media. This requires prior seed sterilization to prevent the growth of microbial contaminants present on the seed surface. Currently, Arabidopsis seeds are sterilized using two distinct sterilization techniques in conditions that differ slightly between labs and have not been standardized, often resulting in only partially effective sterilization or in excessive seed mortality. Most of these methods are also not easily scalable to a large number of seed lines of diverse genotypes. As technologies for high-throughput analysis of Arabidopsis continue to proliferate, standardized techniques for sterilizing large numbers of seeds of different genotypes are becoming essential for conducting these types of experiments. The response of a number of Arabidopsis lines to two different sterilization techniques was evaluated based on seed germination rate and the level of seed contamination with microbes and other pathogens. The treatments included different concentrations of sterilizing agents and times of exposure, combined to determine optimal conditions for Arabidopsis seed sterilization. Optimized protocols have been developed for two different sterilization methods: bleach (liquid-phase) and chlorine (Cl2) gas (vapor-phase), both resulting in high seed germination rates and minimal microbial contamination. The utility of these protocols was illustrated through the testing of both wild type and mutant seeds with a range of germination potentials. Our results show that seeds can be effectively sterilized using either method without excessive seed mortality, although detrimental effects of sterilization were observed for seeds with lower than optimal germination potential. In addition, an equation was developed to enable researchers to apply the standardized chlorine gas sterilization conditions to airtight containers of different sizes. The protocols described here allow easy, efficient, and

  19. MIPHENO: Data normalization for high throughput metabolic analysis.

    Science.gov (United States)

    High throughput methodologies such as microarrays, mass spectrometry and plate-based small molecule screens are increasingly used to facilitate discoveries from gene function to drug candidate identification. These large-scale experiments are typically carried out over the course...

  20. High-Throughput Thermodynamic Modeling and Uncertainty Quantification for ICME

    Science.gov (United States)

    Otis, Richard A.; Liu, Zi-Kui

    2017-05-01

    One foundational component of the integrated computational materials engineering (ICME) and Materials Genome Initiative is the computational thermodynamics based on the calculation of phase diagrams (CALPHAD) method. The CALPHAD method pioneered by Kaufman has enabled the development of thermodynamic, atomic mobility, and molar volume databases of individual phases in the full space of temperature, composition, and sometimes pressure for technologically important multicomponent engineering materials, along with sophisticated computational tools for using the databases. In this article, our recent efforts will be presented in terms of developing new computational tools for high-throughput modeling and uncertainty quantification based on high-throughput, first-principles calculations and the CALPHAD method along with their potential propagations to downstream ICME modeling and simulations.

  1. Controlling high-throughput manufacturing at the nano-scale

    Science.gov (United States)

    Cooper, Khershed P.

    2013-09-01

    Interest in nano-scale manufacturing research and development is growing. The reason is to accelerate the translation of discoveries and inventions of nanoscience and nanotechnology into products that would benefit industry, economy and society. Ongoing research in nanomanufacturing is focused primarily on developing novel nanofabrication techniques for a variety of applications—materials, energy, electronics, photonics, biomedical, etc. Our goal is to foster the development of high-throughput methods of fabricating nano-enabled products. Large-area parallel processing and highspeed continuous processing are high-throughput means for mass production. An example of large-area processing is step-and-repeat nanoimprinting, by which nanostructures are reproduced again and again over a large area, such as a 12 in wafer. Roll-to-roll processing is an example of continuous processing, by which it is possible to print and imprint multi-level nanostructures and nanodevices on a moving flexible substrate. The big pay-off is high-volume production and low unit cost. However, the anticipated cost benefits can only be realized if the increased production rate is accompanied by high yields of high quality products. To ensure product quality, we need to design and construct manufacturing systems such that the processes can be closely monitored and controlled. One approach is to bring cyber-physical systems (CPS) concepts to nanomanufacturing. CPS involves the control of a physical system such as manufacturing through modeling, computation, communication and control. Such a closely coupled system will involve in-situ metrology and closed-loop control of the physical processes guided by physics-based models and driven by appropriate instrumentation, sensing and actuation. This paper will discuss these ideas in the context of controlling high-throughput manufacturing at the nano-scale.

  2. High-Throughput Analysis and Automation for Glycomics Studies

    NARCIS (Netherlands)

    Shubhakar, A.; Reiding, K.R.; Gardner, R.A.; Spencer, D.I.R.; Fernandes, D.L.; Wuhrer, M.

    2015-01-01

    This review covers advances in analytical technologies for high-throughput (HTP) glycomics. Our focus is on structural studies of glycoprotein glycosylation to support biopharmaceutical realization and the discovery of glycan biomarkers for human disease. For biopharmaceuticals, there is increasing

  3. High-Throughput Cloning and Expression Library Creation for Functional Proteomics

    Science.gov (United States)

    Festa, Fernanda; Steel, Jason; Bian, Xiaofang; Labaer, Joshua

    2013-01-01

    The study of protein function usually requires the use of a cloned version of the gene for protein expression and functional assays. This strategy is particular important when the information available regarding function is limited. The functional characterization of the thousands of newly identified proteins revealed by genomics requires faster methods than traditional single gene experiments, creating the need for fast, flexible and reliable cloning systems. These collections of open reading frame (ORF) clones can be coupled with high-throughput proteomics platforms, such as protein microarrays and cell-based assays, to answer biological questions. In this tutorial we provide the background for DNA cloning, discuss the major high-throughput cloning systems (Gateway® Technology, Flexi® Vector Systems, and Creator™ DNA Cloning System) and compare them side-by-side. We also report an example of high-throughput cloning study and its application in functional proteomics. This Tutorial is part of the International Proteomics Tutorial Programme (IPTP12). Details can be found at http://www.proteomicstutorials.org. PMID:23457047

  4. High-throughput bioinformatics with the Cyrille2 pipeline system

    Directory of Open Access Journals (Sweden)

    de Groot Joost CW

    2008-02-01

    Full Text Available Abstract Background Modern omics research involves the application of high-throughput technologies that generate vast volumes of data. These data need to be pre-processed, analyzed and integrated with existing knowledge through the use of diverse sets of software tools, models and databases. The analyses are often interdependent and chained together to form complex workflows or pipelines. Given the volume of the data used and the multitude of computational resources available, specialized pipeline software is required to make high-throughput analysis of large-scale omics datasets feasible. Results We have developed a generic pipeline system called Cyrille2. The system is modular in design and consists of three functionally distinct parts: 1 a web based, graphical user interface (GUI that enables a pipeline operator to manage the system; 2 the Scheduler, which forms the functional core of the system and which tracks what data enters the system and determines what jobs must be scheduled for execution, and; 3 the Executor, which searches for scheduled jobs and executes these on a compute cluster. Conclusion The Cyrille2 system is an extensible, modular system, implementing the stated requirements. Cyrille2 enables easy creation and execution of high throughput, flexible bioinformatics pipelines.

  5. Structural, dielectric and ferroelectric properties of (Bi,Na)TiO3–BaTiO3 system studied by high throughput screening

    International Nuclear Information System (INIS)

    Hayden, Brian E.; Yakovlev, Sergey

    2016-01-01

    Thin-film materials libraries of the Bi 2 O 3 –Na 2 O–TiO 2 –BaO system in a broad composition range have been deposited in ultra-high vacuum from elemental evaporation sources and an oxygen plasma source. A high throughput approach was used for systematic compositional and structural characterization and the screening of the dielectric and ferroelectric properties. The perovskite (Bi,Na)TiO 3 –BaTiO 3 phase with a Ba concentration near the morphotropic phase boundary (ca. 6 at.%) exhibited a relative dielectric permittivity of 180, a loss tangent of 0.04 and remnant polarization of 19 μC/cm 2 . Compared to published data, observed remnant polarization is close to that known for epitaxially grown films but higher than the values reported for polycrystalline films. The high throughput methodology and systematic nature of the study allowed us to establish the composition boundaries of the phase with optimal dielectric and ferroelectric characteristics. - Highlights: • Bi 2 O 3 –Na 2 O–TiO 2 –BaO high throughput materials library was deposited using PVD method. • Materials were processed from individual molecular beam epitaxy sources of elements. • High throughput approach was used for structural, dielectric and ferroelectric study. • Composition boundaries of perovskite compounds with optimum properties are reported.

  6. Joint Throughput Maximization and Fair Uplink Transmission Scheduling in CDMA Systems

    Directory of Open Access Journals (Sweden)

    Symeon Papavassiliou

    2009-01-01

    Full Text Available We study the fundamental problem of optimal transmission scheduling in a code-division multiple-access wireless system in order to maximize the uplink system throughput, while satisfying the users quality-of-service (QoS requirements and maintaining fairness among them. The corresponding problem is expressed as a weighted throughput maximization problem, under certain power and QoS constraints, where the weights are the control parameters reflecting the fairness constraints. With the introduction of the power index capacity, it is shown that this optimization problem can be converted into a binary knapsack problem, where all the corresponding constraints are replaced by the power index capacities at some certain system power index. A two-step approach is followed to obtain the optimal solution. First, a simple method is proposed to find the optimal set of users to receive service for a given fixed target system load, and then the optimal solution is obtained as a global search within a certain range. Furthermore, a stochastic approximation method is presented to effectively identify the required control parameters. The performance evaluation reveals the advantages of our proposed policy over other existing ones and confirms that it achieves very high throughput while maintains fairness among the users, under different channel conditions and requirements.

  7. Towards sensitive, high-throughput, biomolecular assays based on fluorescence lifetime

    Science.gov (United States)

    Ioanna Skilitsi, Anastasia; Turko, Timothé; Cianfarani, Damien; Barre, Sophie; Uhring, Wilfried; Hassiepen, Ulrich; Léonard, Jérémie

    2017-09-01

    Time-resolved fluorescence detection for robust sensing of biomolecular interactions is developed by implementing time-correlated single photon counting in high-throughput conditions. Droplet microfluidics is used as a promising platform for the very fast handling of low-volume samples. We illustrate the potential of this very sensitive and cost-effective technology in the context of an enzymatic activity assay based on fluorescently-labeled biomolecules. Fluorescence lifetime detection by time-correlated single photon counting is shown to enable reliable discrimination between positive and negative control samples at a throughput as high as several hundred samples per second.

  8. High-Throughput Combinatorial Development of High-Entropy Alloys For Light-Weight Structural Applications

    Energy Technology Data Exchange (ETDEWEB)

    Van Duren, Jeroen K [Intermolecular, Inc., San Jose, CA (United States); Koch, Carl [North Carolina State Univ., Raleigh, NC (United States); Luo, Alan [The Ohio State Univ., Columbus, OH (United States); Sample, Vivek [Arconic, Pittsburgh, PA (United States); Sachdev, Anil [General Motors, Detroit, MI (United States)

    2017-12-29

    on Al-Cr-Fe-Ni, shows compressive strain >10% and specific compressive yield strength of 229 MPa x cc/g, yet does not show ductility in tensile tests due to cleavage. When replacing Cr in Al-Cr-Fe-based 4- and 5-element LDHEA with Mn, hardness drops 2x. Combined with compression test results, including those on the ternaries Al-Cr-Fe and Al-Mn-Fe suggest that Al-Mn-Fe-based LDHEA are still worth pursuing. These initial results only represent one compressive stress-strain curve per composition without any property optimization. As such, reproducibility needs to be followed by optimization to show their full potential. When including Li, Mg, and Zn, single-phase Li-Mg-Al-Ti-Zn LDHEA has been found with a specific ultimate compressive strength of 289MPa x cc/g. Al-Ti-Mn-Zn showed a specific ultimate compressive strength of 73MPa x cc/g. These initial results after hot isostatic pressing (HIP) of the ball-milled powders represent the lower end of what is possible, since no secondary processing (e.g. extrusion) has been performed to optimize strength and ductility. Compositions for multi-phase (e.g. dual-phase) LDHEA were identified largely by automated searches through CALPHAD databases, while screening for large face-centered-cubic (FCC) volume fractions, followed by experimental verification. This resulted in several new alloys. Li-Mg-Al-Mn-Fe and Mg-Mn-Fe-Co ball-milled powders upon HIP show specific ultimate compressive strengths of 198MPa x cc/g and 45MPa x cc/g, respectively. Several malleable quarternary Al-Zn-based alloys have been found upon arc/induction melting, yet with limited specific compressive yield strength (<75 MPa x cc/g). These initial results are all without any optimization for strength and/or ductility. High-throughput experimentation allowed us to triple the existing experimental HEA database as published in the past 10 years in less than 2 years which happened at a rate 10x higher than previous methods. Furthermore, we showed that high-throughput

  9. High-throughput cloning and expression in recalcitrant bacteria

    NARCIS (Netherlands)

    Geertsma, Eric R.; Poolman, Bert

    We developed a generic method for high-throughput cloning in bacteria that are less amenable to conventional DNA manipulations. The method involves ligation-independent cloning in an intermediary Escherichia coli vector, which is rapidly converted via vector-backbone exchange (VBEx) into an

  10. A high-throughput, multi-channel photon-counting detector with picosecond timing

    Science.gov (United States)

    Lapington, J. S.; Fraser, G. W.; Miller, G. M.; Ashton, T. J. R.; Jarron, P.; Despeisse, M.; Powolny, F.; Howorth, J.; Milnes, J.

    2009-06-01

    High-throughput photon counting with high time resolution is a niche application area where vacuum tubes can still outperform solid-state devices. Applications in the life sciences utilizing time-resolved spectroscopies, particularly in the growing field of proteomics, will benefit greatly from performance enhancements in event timing and detector throughput. The HiContent project is a collaboration between the University of Leicester Space Research Centre, the Microelectronics Group at CERN, Photek Ltd., and end-users at the Gray Cancer Institute and the University of Manchester. The goal is to develop a detector system specifically designed for optical proteomics, capable of high content (multi-parametric) analysis at high throughput. The HiContent detector system is being developed to exploit this niche market. It combines multi-channel, high time resolution photon counting in a single miniaturized detector system with integrated electronics. The combination of enabling technologies; small pore microchannel plate devices with very high time resolution, and high-speed multi-channel ASIC electronics developed for the LHC at CERN, provides the necessary building blocks for a high-throughput detector system with up to 1024 parallel counting channels and 20 ps time resolution. We describe the detector and electronic design, discuss the current status of the HiContent project and present the results from a 64-channel prototype system. In the absence of an operational detector, we present measurements of the electronics performance using a pulse generator to simulate detector events. Event timing results from the NINO high-speed front-end ASIC captured using a fast digital oscilloscope are compared with data taken with the proposed electronic configuration which uses the multi-channel HPTDC timing ASIC.

  11. A high-throughput, multi-channel photon-counting detector with picosecond timing

    International Nuclear Information System (INIS)

    Lapington, J.S.; Fraser, G.W.; Miller, G.M.; Ashton, T.J.R.; Jarron, P.; Despeisse, M.; Powolny, F.; Howorth, J.; Milnes, J.

    2009-01-01

    High-throughput photon counting with high time resolution is a niche application area where vacuum tubes can still outperform solid-state devices. Applications in the life sciences utilizing time-resolved spectroscopies, particularly in the growing field of proteomics, will benefit greatly from performance enhancements in event timing and detector throughput. The HiContent project is a collaboration between the University of Leicester Space Research Centre, the Microelectronics Group at CERN, Photek Ltd., and end-users at the Gray Cancer Institute and the University of Manchester. The goal is to develop a detector system specifically designed for optical proteomics, capable of high content (multi-parametric) analysis at high throughput. The HiContent detector system is being developed to exploit this niche market. It combines multi-channel, high time resolution photon counting in a single miniaturized detector system with integrated electronics. The combination of enabling technologies; small pore microchannel plate devices with very high time resolution, and high-speed multi-channel ASIC electronics developed for the LHC at CERN, provides the necessary building blocks for a high-throughput detector system with up to 1024 parallel counting channels and 20 ps time resolution. We describe the detector and electronic design, discuss the current status of the HiContent project and present the results from a 64-channel prototype system. In the absence of an operational detector, we present measurements of the electronics performance using a pulse generator to simulate detector events. Event timing results from the NINO high-speed front-end ASIC captured using a fast digital oscilloscope are compared with data taken with the proposed electronic configuration which uses the multi-channel HPTDC timing ASIC.

  12. Fluorescence-based high-throughput screening of dicer cleavage activity.

    Science.gov (United States)

    Podolska, Katerina; Sedlak, David; Bartunek, Petr; Svoboda, Petr

    2014-03-01

    Production of small RNAs by ribonuclease III Dicer is a key step in microRNA and RNA interference pathways, which employ Dicer-produced small RNAs as sequence-specific silencing guides. Further studies and manipulations of microRNA and RNA interference pathways would benefit from identification of small-molecule modulators. Here, we report a study of a fluorescence-based in vitro Dicer cleavage assay, which was adapted for high-throughput screening. The kinetic assay can be performed under single-turnover conditions (35 nM substrate and 70 nM Dicer) in a small volume (5 µL), which makes it suitable for high-throughput screening in a 1536-well format. As a proof of principle, a small library of bioactive compounds was analyzed, demonstrating potential of the assay.

  13. Repurposing a Benchtop Centrifuge for High-Throughput Single-Molecule Force Spectroscopy.

    Science.gov (United States)

    Yang, Darren; Wong, Wesley P

    2018-01-01

    We present high-throughput single-molecule manipulation using a benchtop centrifuge, overcoming limitations common in other single-molecule approaches such as high cost, low throughput, technical difficulty, and strict infrastructure requirements. An inexpensive and compact Centrifuge Force Microscope (CFM) adapted to a commercial centrifuge enables use by nonspecialists, and integration with DNA nanoswitches facilitates both reliable measurements and repeated molecular interrogation. Here, we provide detailed protocols for constructing the CFM, creating DNA nanoswitch samples, and carrying out single-molecule force measurements.

  14. A high-throughput liquid bead array-based screening technology for Bt presence in GMO manipulation.

    Science.gov (United States)

    Fu, Wei; Wang, Huiyu; Wang, Chenguang; Mei, Lin; Lin, Xiangmei; Han, Xueqing; Zhu, Shuifang

    2016-03-15

    The number of species and planting areas of genetically modified organisms (GMOs) has been rapidly developed during the past ten years. For the purpose of GMO inspection, quarantine and manipulation, we have now devised a high-throughput Bt-based GMOs screening method based on the liquid bead array. This novel method is based on the direct competitive recognition between biotinylated antibodies and beads-coupled antigens, searching for Bt presence in samples if it contains Bt Cry1 Aa, Bt Cry1 Ab, Bt Cry1 Ac, Bt Cry1 Ah, Bt Cry1 B, Bt Cry1 C, Bt Cry1 F, Bt Cry2 A, Bt Cry3 or Bt Cry9 C. Our method has a wide GMO species coverage so that more than 90% of the whole commercialized GMO species can be identified throughout the world. Under our optimization, specificity, sensitivity, repeatability and availability validation, the method shows a high specificity and 10-50 ng/mL sensitivity of quantification. We then assessed more than 1800 samples in the field and food market to prove capacity of our method in performing a high throughput screening work for GMO manipulation. Our method offers an applicant platform for further inspection and research on GMO plants. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Evaluation of Capacity on a High Throughput Vol-oxidizer for Operability

    International Nuclear Information System (INIS)

    Kim, Young Hwan; Park, Geun Il; Lee, Jung Won; Jung, Jae Hoo; Kim, Ki Ho; Lee, Yong Soon; Lee, Do Youn; Kim, Su Sung

    2010-01-01

    KAERI is developing a pyro-process. As a piece of process equipment, a high throughput vol-oxidizer which can handle a several tens kg HM/batch was developed to supply U 3 O 8 powders to an electrolytic reduction(ER) reactor. To increase the reduction yield, UO 2 pellets should be converted into uniform powders. In this paper, we aim at the evaluation of a high throughput vol-oxidizer for operability. The evaluation consisted of 3 targets, a mechanical motion test, a heating test and hull separation test. In order to test a high throughput vol-oxidizer, By using a control system, mechanical motion tests of the vol-oxidizer were conducted, and heating rates were analyzed. Also the separation tests of hulls for recovery rate were conducted. The test results of the vol-oxidizer are going to be applied for operability. A study on the characteristics of the volatile gas produced during a vol-oxidation process is not included in this study

  16. Fun with High Throughput Toxicokinetics (CalEPA webinar)

    Science.gov (United States)

    Thousands of chemicals have been profiled by high-throughput screening (HTS) programs such as ToxCast and Tox21. These chemicals are tested in part because there are limited or no data on hazard, exposure, or toxicokinetics (TK). TK models aid in predicting tissue concentrations ...

  17. New developments of RNAi in Paracoccidioides brasiliensis: prospects for high-throughput, genome-wide, functional genomics.

    Directory of Open Access Journals (Sweden)

    Tercio Goes

    2014-10-01

    Full Text Available The Fungal Genome Initiative of the Broad Institute, in partnership with the Paracoccidioides research community, has recently sequenced the genome of representative isolates of this human-pathogen dimorphic fungus: Pb18 (S1, Pb03 (PS2 and Pb01. The accomplishment of future high-throughput, genome-wide, functional genomics will rely upon appropriate molecular tools and straightforward techniques to streamline the generation of stable loss-of-function phenotypes. In the past decades, RNAi has emerged as the most robust genetic technique to modulate or to suppress gene expression in diverse eukaryotes, including fungi. These molecular tools and techniques, adapted for RNAi, were up until now unavailable for P. brasiliensis.In this paper, we report Agrobacterium tumefaciens mediated transformation of yeast cells for high-throughput applications with which higher transformation frequencies of 150±24 yeast cell transformants per 1×106 viable yeast cells were obtained. Our approach is based on a bifunctional selective marker fusion protein consisted of the Streptoalloteichus hindustanus bleomycin-resistance gene (Shble and the intrinsically fluorescent monomeric protein mCherry which was codon-optimized for heterologous expression in P. brasiliensis. We also report successful GP43 gene knock-down through the expression of intron-containing hairpin RNA (ihpRNA from a Gateway-adapted cassette (cALf which was purpose-built for gene silencing in a high-throughput manner. Gp43 transcript levels were reduced by 73.1±22.9% with this approach.We have a firm conviction that the genetic transformation technique and the molecular tools herein described will have a relevant contribution in future Paracoccidioides spp. functional genomics research.

  18. High throughput experimentation for the discovery of new catalysts

    International Nuclear Information System (INIS)

    Thomson, S.; Hoffmann, C.; Johann, T.; Wolf, A.; Schmidt, H.-W.; Farrusseng, D.; Schueth, F.

    2002-01-01

    Full text: The use of combinatorial chemistry to obtain new materials has been developed extensively by the pharmaceutical and biochemical industries, but such approaches have been slow to impact on the field of heterogeneous catalysis. The reasons for this lie in with difficulties associated in the synthesis, characterisation and determination of catalytic properties of such materials. In many synthetic and catalytic reactions, the conditions used are difficult to emulate using High Throughput Experimentation (HTE). Furthermore, the ability to screen these catalysts simultaneously in real time, requires the development and/or modification of characterisation methods. Clearly, there is a need for both high throughput synthesis and screening of new and novel reactions, and we describe several new concepts that help to achieve these goals. Although such problems have impeded the development of combinatorial catalysis, the fact remains that many highly attractive processes still exist for which no suitable catalysts have been developed. The ability to decrease the tiFme needed to evaluate catalyst is therefore essential and this makes the use of high throughput techniques highly desirable. In this presentation we will describe the synthesis, catalytic testing, and novel screening methods developed at the Max Planck Institute. Automated synthesis procedures, performed by the use of a modified Gilson pipette robot, will be described, as will the development of two 16 and 49 sample fixed bed reactors and two 25 and 29 sample three phase reactors for catalytic testing. We will also present new techniques for the characterisation of catalysts and catalytic products using standard IR microscopy and infrared focal plane array detection, respectively

  19. A High-Throughput, High-Accuracy System-Level Simulation Framework for System on Chips

    Directory of Open Access Journals (Sweden)

    Guanyi Sun

    2011-01-01

    Full Text Available Today's System-on-Chips (SoCs design is extremely challenging because it involves complicated design tradeoffs and heterogeneous design expertise. To explore the large solution space, system architects have to rely on system-level simulators to identify an optimized SoC architecture. In this paper, we propose a system-level simulation framework, System Performance Simulation Implementation Mechanism, or SPSIM. Based on SystemC TLM2.0, the framework consists of an executable SoC model, a simulation tool chain, and a modeling methodology. Compared with the large body of existing research in this area, this work is aimed at delivering a high simulation throughput and, at the same time, guaranteeing a high accuracy on real industrial applications. Integrating the leading TLM techniques, our simulator can attain a simulation speed that is not slower than that of the hardware execution by a factor of 35 on a set of real-world applications. SPSIM incorporates effective timing models, which can achieve a high accuracy after hardware-based calibration. Experimental results on a set of mobile applications proved that the difference between the simulated and measured results of timing performance is within 10%, which in the past can only be attained by cycle-accurate models.

  20. Modeling Steroidogenesis Disruption Using High-Throughput ...

    Science.gov (United States)

    Environmental chemicals can elicit endocrine disruption by altering steroid hormone biosynthesis and metabolism (steroidogenesis) causing adverse reproductive and developmental effects. Historically, a lack of assays resulted in few chemicals having been evaluated for effects on steroidogenesis. The steroidogenic pathway is a series of hydroxylation and dehydrogenation steps carried out by CYP450 and hydroxysteroid dehydrogenase enzymes, yet the only enzyme in the pathway for which a high-throughput screening (HTS) assay has been developed is aromatase (CYP19A1), responsible for the aromatization of androgens to estrogens. Recently, the ToxCast HTS program adapted the OECD validated H295R steroidogenesis assay using human adrenocortical carcinoma cells into a high-throughput model to quantitatively assess the concentration-dependent (0.003-100 µM) effects of chemicals on 10 steroid hormones including progestagens, androgens, estrogens and glucocorticoids. These results, in combination with two CYP19A1 inhibition assays, comprise a large dataset amenable to clustering approaches supporting the identification and characterization of putative mechanisms of action (pMOA) for steroidogenesis disruption. In total, 514 chemicals were tested in all CYP19A1 and steroidogenesis assays. 216 chemicals were identified as CYP19A1 inhibitors in at least one CYP19A1 assay. 208 of these chemicals also altered hormone levels in the H295R assay, suggesting 96% sensitivity in the

  1. Fully Automated Electro Membrane Extraction Autosampler for LC-MS Systems Allowing Soft Extractions for High-Throughput Applications

    DEFF Research Database (Denmark)

    Fuchs, David; Pedersen-Bjergaard, Stig; Jensen, Henrik

    2016-01-01

    was optimized for soft extraction of analytes and high sample throughput. Further, it was demonstrated that by flushing the EME-syringe with acidic wash buffer and reverting the applied electric potential, carry-over between samples can be reduced to below 1%. Performance of the system was characterized (RSD......, a complete analytical workflow of purification, separation, and analysis of sample could be achieved within only 5.5 min. With the developed system large sequences of samples could be analyzed in a completely automated manner. This high degree of automation makes the developed EME-autosampler a powerful tool...

  2. Towards low-delay and high-throughput cognitive radio vehicular networks

    Directory of Open Access Journals (Sweden)

    Nada Elgaml

    2017-12-01

    Full Text Available Cognitive Radio Vehicular Ad-hoc Networks (CR-VANETs exploit cognitive radios to allow vehicles to access the unused channels in their radio environment. Thus, CR-VANETs do not only suffer the traditional CR problems, especially spectrum sensing, but also suffer new challenges due to the highly dynamic nature of VANETs. In this paper, we present a low-delay and high-throughput radio environment assessment scheme for CR-VANETs that can be easily incorporated with the IEEE 802.11p standard developed for VANETs. Simulation results show that the proposed scheme significantly reduces the time to get the radio environment map and increases the CR-VANET throughput.

  3. High throughput reaction screening using desorption electrospray ionization mass spectrometry.

    Science.gov (United States)

    Wleklinski, Michael; Loren, Bradley P; Ferreira, Christina R; Jaman, Zinia; Avramova, Larisa; Sobreira, Tiago J P; Thompson, David H; Cooks, R Graham

    2018-02-14

    We report the high throughput analysis of reaction mixture arrays using methods and data handling routines that were originally developed for biological tissue imaging. Desorption electrospray ionization (DESI) mass spectrometry (MS) is applied in a continuous on-line process at rates that approach 10 4 reactions per h at area densities of up to 1 spot per mm 2 (6144 spots per standard microtiter plate) with the sprayer moving at ca. 10 4 microns per s. Data are analyzed automatically by MS using in-house software to create ion images of selected reagents and products as intensity plots in standard array format. Amine alkylation reactions were used to optimize the system performance on PTFE membrane substrates using methanol as the DESI spray/analysis solvent. Reaction times can be screening of processes like N -alkylation and Suzuki coupling reactions as reported herein. Products and by-products were confirmed by on-line MS/MS upon rescanning of the array.

  4. Reverse Phase Protein Arrays for High-throughput Toxicity Screening

    DEFF Research Database (Denmark)

    Pedersen, Marlene Lemvig; Block, Ines; List, Markus

    High-throughput screening is extensively applied for identification of drug targets and drug discovery and recently it found entry into toxicity testing. Reverse phase protein arrays (RPPAs) are used widespread for quantification of protein markers. We reasoned that RPPAs also can be utilized...... beneficially in automated high-throughput toxicity testing. An advantage of using RPPAs is that, in addition to the baseline toxicity readout, they allow testing of multiple markers of toxicity, such as inflammatory responses, which do not necessarily cumulate in cell death. We used transfection of si......RNAs with known killing effects as a model system to demonstrate that RPPA-based protein quantification can serve as substitute readout of cell viability, hereby reliably reflecting toxicity. In terms of automation, cell exposure, protein harvest, serial dilution and sample reformatting were performed using...

  5. Development of automatic image analysis methods for high-throughput and high-content screening

    NARCIS (Netherlands)

    Di, Zi

    2013-01-01

    This thesis focuses on the development of image analysis methods for ultra-high content analysis of high-throughput screens where cellular phenotype responses to various genetic or chemical perturbations that are under investigation. Our primary goal is to deliver efficient and robust image analysis

  6. Quantitative in vitro-to-in vivo extrapolation in a high-throughput environment

    International Nuclear Information System (INIS)

    Wetmore, Barbara A.

    2015-01-01

    High-throughput in vitro toxicity screening provides an efficient way to identify potential biological targets for environmental and industrial chemicals while conserving limited testing resources. However, reliance on the nominal chemical concentrations in these in vitro assays as an indicator of bioactivity may misrepresent potential in vivo effects of these chemicals due to differences in clearance, protein binding, bioavailability, and other pharmacokinetic factors. Development of high-throughput in vitro hepatic clearance and protein binding assays and refinement of quantitative in vitro-to-in vivo extrapolation (QIVIVE) methods have provided key tools to predict xenobiotic steady state pharmacokinetics. Using a process known as reverse dosimetry, knowledge of the chemical steady state behavior can be incorporated with HTS data to determine the external in vivo oral exposure needed to achieve internal blood concentrations equivalent to those eliciting bioactivity in the assays. These daily oral doses, known as oral equivalents, can be compared to chronic human exposure estimates to assess whether in vitro bioactivity would be expected at the dose-equivalent level of human exposure. This review will describe the use of QIVIVE methods in a high-throughput environment and the promise they hold in shaping chemical testing priorities and, potentially, high-throughput risk assessment strategies

  7. Space Link Extension (SLE) Emulation for High-Throughput Network Communication

    Science.gov (United States)

    Murawski, Robert W.; Tchorowski, Nicole; Golden, Bert

    2014-01-01

    As the data rate requirements for space communications increases, significant stress is placed not only on the wireless satellite communication links, but also on the ground networks which forward data from end-users to remote ground stations. These wide area network (WAN) connections add delay and jitter to the end-to-end satellite communication link, effects which can have significant impacts on the wireless communication link. It is imperative that any ground communication protocol can react to these effects such that the ground network does not become a bottleneck in the communication path to the satellite. In this paper, we present our SCENIC Emulation Lab testbed which was developed to test the CCSDS SLE protocol implementations proposed for use on future NASA communication networks. Our results show that in the presence of realistic levels of network delay, high-throughput SLE communication links can experience significant data rate throttling. Based on our observations, we present some insight into why this data throttling happens, and trace the probable issue back to non-optimal blocking communication which is sup-ported by the CCSDS SLE API recommended practices. These issues were presented as well to the SLE implementation developers which, based on our reports, developed a new release for SLE which we show fixes the SLE blocking issue and greatly improves the protocol throughput. In this paper, we also discuss future developments for our end-to-end emulation lab and how these improvements can be used to develop and test future space communication technologies.

  8. Quality control methodology for high-throughput protein-protein interaction screening.

    Science.gov (United States)

    Vazquez, Alexei; Rual, Jean-François; Venkatesan, Kavitha

    2011-01-01

    Protein-protein interactions are key to many aspects of the cell, including its cytoskeletal structure, the signaling processes in which it is involved, or its metabolism. Failure to form protein complexes or signaling cascades may sometimes translate into pathologic conditions such as cancer or neurodegenerative diseases. The set of all protein interactions between the proteins encoded by an organism constitutes its protein interaction network, representing a scaffold for biological function. Knowing the protein interaction network of an organism, combined with other sources of biological information, can unravel fundamental biological circuits and may help better understand the molecular basics of human diseases. The protein interaction network of an organism can be mapped by combining data obtained from both low-throughput screens, i.e., "one gene at a time" experiments and high-throughput screens, i.e., screens designed to interrogate large sets of proteins at once. In either case, quality controls are required to deal with the inherent imperfect nature of experimental assays. In this chapter, we discuss experimental and statistical methodologies to quantify error rates in high-throughput protein-protein interactions screens.

  9. High-throughput screening to enhance oncolytic virus immunotherapy

    Directory of Open Access Journals (Sweden)

    Allan KJ

    2016-04-01

    Full Text Available KJ Allan,1,2 David F Stojdl,1–3 SL Swift1 1Children’s Hospital of Eastern Ontario (CHEO Research Institute, 2Department of Biology, Microbiology and Immunology, 3Department of Pediatrics, University of Ottawa, Ottawa, ON, Canada Abstract: High-throughput screens can rapidly scan and capture large amounts of information across multiple biological parameters. Although many screens have been designed to uncover potential new therapeutic targets capable of crippling viruses that cause disease, there have been relatively few directed at improving the efficacy of viruses that are used to treat disease. Oncolytic viruses (OVs are biotherapeutic agents with an inherent specificity for treating malignant disease. Certain OV platforms – including those based on herpes simplex virus, reovirus, and vaccinia virus – have shown success against solid tumors in advanced clinical trials. Yet, many of these OVs have only undergone minimal engineering to solidify tumor specificity, with few extra modifications to manipulate additional factors. Several aspects of the interaction between an OV and a tumor-bearing host have clear value as targets to improve therapeutic outcomes. At the virus level, these include delivery to the tumor, infectivity, productivity, oncolysis, bystander killing, spread, and persistence. At the host level, these include engaging the immune system and manipulating the tumor microenvironment. Here, we review the chemical- and genome-based high-throughput screens that have been performed to manipulate such parameters during OV infection and analyze their impact on therapeutic efficacy. We further explore emerging themes that represent key areas of focus for future research. Keywords: oncolytic, virus, screen, high-throughput, cancer, chemical, genomic, immunotherapy

  10. High-throughput machining using high average power ultrashort pulse lasers and ultrafast polygon scanner

    Science.gov (United States)

    Schille, Joerg; Schneider, Lutz; Streek, André; Kloetzer, Sascha; Loeschner, Udo

    2016-03-01

    In this paper, high-throughput ultrashort pulse laser machining is investigated on various industrial grade metals (Aluminium, Copper, Stainless steel) and Al2O3 ceramic at unprecedented processing speeds. This is achieved by using a high pulse repetition frequency picosecond laser with maximum average output power of 270 W in conjunction with a unique, in-house developed two-axis polygon scanner. Initially, different concepts of polygon scanners are engineered and tested to find out the optimal architecture for ultrafast and precision laser beam scanning. Remarkable 1,000 m/s scan speed is achieved on the substrate, and thanks to the resulting low pulse overlap, thermal accumulation and plasma absorption effects are avoided at up to 20 MHz pulse repetition frequencies. In order to identify optimum processing conditions for efficient high-average power laser machining, the depths of cavities produced under varied parameter settings are analyzed and, from the results obtained, the characteristic removal values are specified. The maximum removal rate is achieved as high as 27.8 mm3/min for Aluminium, 21.4 mm3/min for Copper, 15.3 mm3/min for Stainless steel and 129.1 mm3/min for Al2O3 when full available laser power is irradiated at optimum pulse repetition frequency.

  11. Microscale High-Throughput Experimentation as an Enabling Technology in Drug Discovery: Application in the Discovery of (Piperidinyl)pyridinyl-1H-benzimidazole Diacylglycerol Acyltransferase 1 Inhibitors.

    Science.gov (United States)

    Cernak, Tim; Gesmundo, Nathan J; Dykstra, Kevin; Yu, Yang; Wu, Zhicai; Shi, Zhi-Cai; Vachal, Petr; Sperbeck, Donald; He, Shuwen; Murphy, Beth Ann; Sonatore, Lisa; Williams, Steven; Madeira, Maria; Verras, Andreas; Reiter, Maud; Lee, Claire Heechoon; Cuff, James; Sherer, Edward C; Kuethe, Jeffrey; Goble, Stephen; Perrotto, Nicholas; Pinto, Shirly; Shen, Dong-Ming; Nargund, Ravi; Balkovec, James; DeVita, Robert J; Dreher, Spencer D

    2017-05-11

    Miniaturization and parallel processing play an important role in the evolution of many technologies. We demonstrate the application of miniaturized high-throughput experimentation methods to resolve synthetic chemistry challenges on the frontlines of a lead optimization effort to develop diacylglycerol acyltransferase (DGAT1) inhibitors. Reactions were performed on ∼1 mg scale using glass microvials providing a miniaturized high-throughput experimentation capability that was used to study a challenging S N Ar reaction. The availability of robust synthetic chemistry conditions discovered in these miniaturized investigations enabled the development of structure-activity relationships that ultimately led to the discovery of soluble, selective, and potent inhibitors of DGAT1.

  12. High throughput electrophysiology: new perspectives for ion channel drug discovery

    DEFF Research Database (Denmark)

    Willumsen, Niels J; Bech, Morten; Olesen, Søren-Peter

    2003-01-01

    Proper function of ion channels is crucial for all living cells. Ion channel dysfunction may lead to a number of diseases, so-called channelopathies, and a number of common diseases, including epilepsy, arrhythmia, and type II diabetes, are primarily treated by drugs that modulate ion channels....... A cornerstone in current drug discovery is high throughput screening assays which allow examination of the activity of specific ion channels though only to a limited extent. Conventional patch clamp remains the sole technique with sufficiently high time resolution and sensitivity required for precise and direct...... characterization of ion channel properties. However, patch clamp is a slow, labor-intensive, and thus expensive, technique. New techniques combining the reliability and high information content of patch clamping with the virtues of high throughput philosophy are emerging and predicted to make a number of ion...

  13. High throughput screening of starch structures using carbohydrate microarrays

    DEFF Research Database (Denmark)

    Tanackovic, Vanja; Rydahl, Maja Gro; Pedersen, Henriette Lodberg

    2016-01-01

    In this study we introduce the starch-recognising carbohydrate binding module family 20 (CBM20) from Aspergillus niger for screening biological variations in starch molecular structure using high throughput carbohydrate microarray technology. Defined linear, branched and phosphorylated...

  14. Achieving high data throughput in research networks

    International Nuclear Information System (INIS)

    Matthews, W.; Cottrell, L.

    2001-01-01

    After less than a year of operation, the BaBar experiment at SLAC has collected almost 100 million particle collision events in a database approaching 165TB. Around 20 TB of data has been exported via the Internet to the BaBar regional center at IN2P3 in Lyon, France, and around 40TB of simulated data has been imported from the Lawrence Livermore National Laboratory (LLNL). BaBar collaborators plan to double data collection each year and export a third of the data to IN2P3. So within a few years the SLAC OC3 (155 Mbps) connection will be fully utilized by file transfer to France alone. Upgrades to infrastructure is essential and detailed understanding of performance issues and the requirements for reliable high throughput transfers is critical. In this talk results from active and passive monitoring and direct measurements of throughput will be reviewed. Methods for achieving the ambitious requirements will be discussed

  15. Achieving High Data Throughput in Research Networks

    International Nuclear Information System (INIS)

    Matthews, W

    2004-01-01

    After less than a year of operation, the BaBar experiment at SLAC has collected almost 100 million particle collision events in a database approaching 165TB. Around 20 TB of data has been exported via the Internet to the BaBar regional center at IN2P3 in Lyon, France, and around 40TB of simulated data has been imported from the Lawrence Livermore National Laboratory (LLNL). BaBar collaborators plan to double data collection each year and export a third of the data to IN2P3. So within a few years the SLAC OC3 (155Mbps) connection will be fully utilized by file transfer to France alone. Upgrades to infrastructure is essential and detailed understanding of performance issues and the requirements for reliable high throughput transfers is critical. In this talk results from active and passive monitoring and direct measurements of throughput will be reviewed. Methods for achieving the ambitious requirements will be discussed

  16. A priori Considerations When Conducting High-Throughput Amplicon-Based Sequence Analysis

    Directory of Open Access Journals (Sweden)

    Aditi Sengupta

    2016-03-01

    Full Text Available Amplicon-based sequencing strategies that include 16S rRNA and functional genes, alongside “meta-omics” analyses of communities of microorganisms, have allowed researchers to pose questions and find answers to “who” is present in the environment and “what” they are doing. Next-generation sequencing approaches that aid microbial ecology studies of agricultural systems are fast gaining popularity among agronomy, crop, soil, and environmental science researchers. Given the rapid development of these high-throughput sequencing techniques, researchers with no prior experience will desire information about the best practices that can be used before actually starting high-throughput amplicon-based sequence analyses. We have outlined items that need to be carefully considered in experimental design, sampling, basic bioinformatics, sequencing of mock communities and negative controls, acquisition of metadata, and in standardization of reaction conditions as per experimental requirements. Not all considerations mentioned here may pertain to a particular study. The overall goal is to inform researchers about considerations that must be taken into account when conducting high-throughput microbial DNA sequencing and sequences analysis.

  17. Multiplexing a high-throughput liability assay to leverage efficiencies.

    Science.gov (United States)

    Herbst, John; Anthony, Monique; Stewart, Jeremy; Connors, David; Chen, Taosheng; Banks, Martyn; Petrillo, Edward W; Agler, Michele

    2009-06-01

    In order to identify potential cytochrome P-450 3A4 (drug-metabolizing enzyme) inducers at an early stage of the drug discovery process, a cell-based transactivation high-throughput luciferase reporter assay for the human pregnane X receptor (PXR) in HepG2 cells has been implemented and multiplexed with a viability end point for data interpretation, as part of a Lead Profiling portfolio of assays. As a routine part of Lead Profiling operations, assays are periodically evaluated for utility as well as for potential improvements in technology or process. We used a recent evaluation of our PXR-transactivation assay as a model for the application of Lean Thinking-based process analysis to lab-bench assay optimization and automation. This resulted in the development of a 384-well multiplexed homogeneous assay simultaneously detecting PXR transactivation and HepG2 cell cytotoxicity. In order to multiplex fluorescent and luminescent read-outs, modifications to each assay were necessary, which included optimization of multiple assay parameters such as cell density, plate type, and reagent concentrations. Subsequently, a set of compounds including known cytotoxic compounds and PXR inducers were used to validate the multiplexed assay. Results from the multiplexed assay correlate well with those from the singleplexed assay formats measuring PXR transactivation and viability separately. Implementation of the multiplexed assay for routine compound profiling provides improved data quality, sample conservation, cost savings, and resource efficiencies.

  18. Enzyme free cloning for high throughput gene cloning and expression

    NARCIS (Netherlands)

    de Jong, R.N.; Daniëls, M.; Kaptein, R.; Folkers, G.E.

    2006-01-01

    Structural and functional genomics initiatives significantly improved cloning methods over the past few years. Although recombinational cloning is highly efficient, its costs urged us to search for an alternative high throughput (HTP) cloning method. We implemented a modified Enzyme Free Cloning

  19. Comprehensive optical and data management infrastructure for high-throughput light-sheet microscopy of whole mouse brains.

    Science.gov (United States)

    Müllenbroich, M Caroline; Silvestri, Ludovico; Onofri, Leonardo; Costantini, Irene; Hoff, Marcel Van't; Sacconi, Leonardo; Iannello, Giulio; Pavone, Francesco S

    2015-10-01

    Comprehensive mapping and quantification of neuronal projections in the central nervous system requires high-throughput imaging of large volumes with microscopic resolution. To this end, we have developed a confocal light-sheet microscope that has been optimized for three-dimensional (3-D) imaging of structurally intact clarified whole-mount mouse brains. We describe the optical and electromechanical arrangement of the microscope and give details on the organization of the microscope management software. The software orchestrates all components of the microscope, coordinates critical timing and synchronization, and has been written in a versatile and modular structure using the LabVIEW language. It can easily be adapted and integrated to other microscope systems and has been made freely available to the light-sheet community. The tremendous amount of data routinely generated by light-sheet microscopy further requires novel strategies for data handling and storage. To complete the full imaging pipeline of our high-throughput microscope, we further elaborate on big data management from streaming of raw images up to stitching of 3-D datasets. The mesoscale neuroanatomy imaged at micron-scale resolution in those datasets allows characterization and quantification of neuronal projections in unsectioned mouse brains.

  20. High-Throughput Analysis of Enzyme Activities

    Energy Technology Data Exchange (ETDEWEB)

    Lu, Guoxin [Iowa State Univ., Ames, IA (United States)

    2007-01-01

    High-throughput screening (HTS) techniques have been applied to many research fields nowadays. Robot microarray printing technique and automation microtiter handling technique allows HTS performing in both heterogeneous and homogeneous formats, with minimal sample required for each assay element. In this dissertation, new HTS techniques for enzyme activity analysis were developed. First, patterns of immobilized enzyme on nylon screen were detected by multiplexed capillary system. The imaging resolution is limited by the outer diameter of the capillaries. In order to get finer images, capillaries with smaller outer diameters can be used to form the imaging probe. Application of capillary electrophoresis allows separation of the product from the substrate in the reaction mixture, so that the product doesn't have to have different optical properties with the substrate. UV absorption detection allows almost universal detection for organic molecules. Thus, no modifications of either the substrate or the product molecules are necessary. This technique has the potential to be used in screening of local distribution variations of specific bio-molecules in a tissue or in screening of multiple immobilized catalysts. Another high-throughput screening technique is developed by directly monitoring the light intensity of the immobilized-catalyst surface using a scientific charge-coupled device (CCD). Briefly, the surface of enzyme microarray is focused onto a scientific CCD using an objective lens. By carefully choosing the detection wavelength, generation of product on an enzyme spot can be seen by the CCD. Analyzing the light intensity change over time on an enzyme spot can give information of reaction rate. The same microarray can be used for many times. Thus, high-throughput kinetic studies of hundreds of catalytic reactions are made possible. At last, we studied the fluorescence emission spectra of ADP and obtained the detection limits for ADP under three different

  1. A high-throughput microfluidic approach for 1000-fold leukocyte reduction of platelet-rich plasma

    Science.gov (United States)

    Xia, Hui; Strachan, Briony C.; Gifford, Sean C.; Shevkoplyas, Sergey S.

    2016-10-01

    Leukocyte reduction of donated blood products substantially reduces the risk of a number of transfusion-related complications. Current ‘leukoreduction’ filters operate by trapping leukocytes within specialized filtration material, while allowing desired blood components to pass through. However, the continuous release of inflammatory cytokines from the retained leukocytes, as well as the potential for platelet activation and clogging, are significant drawbacks of conventional ‘dead end’ filtration. To address these limitations, here we demonstrate our newly-developed ‘controlled incremental filtration’ (CIF) approach to perform high-throughput microfluidic removal of leukocytes from platelet-rich plasma (PRP) in a continuous flow regime. Leukocytes are separated from platelets within the PRP by progressively syphoning clarified PRP away from the concentrated leukocyte flowstream. Filtrate PRP collected from an optimally-designed CIF device typically showed a ~1000-fold (i.e. 99.9%) reduction in leukocyte concentration, while recovering >80% of the original platelets, at volumetric throughputs of ~1 mL/min. These results suggest that the CIF approach will enable users in many fields to now apply the advantages of microfluidic devices to particle separation, even for applications requiring macroscale flowrates.

  2. High-Throughput Dissection of AAV-Host Interactions: The Fast and the Curious.

    Science.gov (United States)

    Herrmann, Anne-Kathrin; Grimm, Dirk

    2018-05-18

    Over fifty years after its initial description, Adeno-associated virus (AAV) remains a most exciting but also most elusive study object in basic or applied virology. On the one hand, its simple structure not only facilitates investigations into virus biology, but combined with the availability of numerous natural AAV variants with distinct infection efficiency and specificity also makes AAV a preferred substrate for engineering of gene delivery vectors. On the other hand, it is striking to witness a recent flurry of reports that highlight and partially close persistent gaps in our understanding of AAV virus and vector biology. This is all the more perplexing considering that recombinant AAVs have already been used in >160 clinical trials and recently been commercialized as gene therapeutics. Here, we discuss a reason for these advances in AAV research, namely, the advent and application of powerful high-throughput technology for dissection of AAV-host interactions and optimization of AAV gene therapy vectors. As relevant examples, we focus on the discovery of (i) a "new" cellular AAV receptor, AAVR, (ii) host restriction factors for AAV entry, and (iii) AAV capsid determinants that mediate trafficking through the blood-brain barrier. While (i)/(ii) are prototypes of extra- or intracellular AAV host factors that were identified via high-throughput screenings, (iii) exemplifies the power of molecular evolution to investigate the virus itself. In the future, we anticipate that these and other key technologies will continue to accelerate the dissection of AAV biology and will yield a wealth of new designer viruses for clinical use. Copyright © 2018. Published by Elsevier Ltd.

  3. High throughput nonparametric probability density estimation.

    Science.gov (United States)

    Farmer, Jenny; Jacobs, Donald

    2018-01-01

    In high throughput applications, such as those found in bioinformatics and finance, it is important to determine accurate probability distribution functions despite only minimal information about data characteristics, and without using human subjectivity. Such an automated process for univariate data is implemented to achieve this goal by merging the maximum entropy method with single order statistics and maximum likelihood. The only required properties of the random variables are that they are continuous and that they are, or can be approximated as, independent and identically distributed. A quasi-log-likelihood function based on single order statistics for sampled uniform random data is used to empirically construct a sample size invariant universal scoring function. Then a probability density estimate is determined by iteratively improving trial cumulative distribution functions, where better estimates are quantified by the scoring function that identifies atypical fluctuations. This criterion resists under and over fitting data as an alternative to employing the Bayesian or Akaike information criterion. Multiple estimates for the probability density reflect uncertainties due to statistical fluctuations in random samples. Scaled quantile residual plots are also introduced as an effective diagnostic to visualize the quality of the estimated probability densities. Benchmark tests show that estimates for the probability density function (PDF) converge to the true PDF as sample size increases on particularly difficult test probability densities that include cases with discontinuities, multi-resolution scales, heavy tails, and singularities. These results indicate the method has general applicability for high throughput statistical inference.

  4. High-throughput full-automatic synchrotron-based tomographic microscopy

    International Nuclear Information System (INIS)

    Mader, Kevin; Marone, Federica; Hintermueller, Christoph; Mikuljan, Gordan; Isenegger, Andreas; Stampanoni, Marco

    2011-01-01

    At the TOMCAT (TOmographic Microscopy and Coherent rAdiology experimenTs) beamline of the Swiss Light Source with an energy range of 8-45 keV and voxel size from 0.37 (micro)m to 7.4 (micro)m, full tomographic datasets are typically acquired in 5 to 10 min. To exploit the speed of the system and enable high-throughput studies to be performed in a fully automatic manner, a package of automation tools has been developed. The samples are automatically exchanged, aligned, moved to the correct region of interest, and scanned. This task is accomplished through the coordination of Python scripts, a robot-based sample-exchange system, sample positioning motors and a CCD camera. The tools are suited for any samples that can be mounted on a standard SEM stub, and require no specific environmental conditions. Up to 60 samples can be analyzed at a time without user intervention. The throughput of the system is dependent on resolution, energy and sample size, but rates of four samples per hour have been achieved with 0.74 (micro)m voxel size at 17.5 keV. The maximum intervention-free scanning time is theoretically unlimited, and in practice experiments have been running unattended as long as 53 h (the average beam time allocation at TOMCAT is 48 h per user). The system is the first fully automated high-throughput tomography station: mounting samples, finding regions of interest, scanning and reconstructing can be performed without user intervention. The system also includes many features which accelerate and simplify the process of tomographic microscopy.

  5. Integrating prediction, provenance, and optimization into high energy workflows

    Energy Technology Data Exchange (ETDEWEB)

    Schram, M.; Bansal, V.; Friese, R. D.; Tallent, N. R.; Yin, J.; Barker, K. J.; Stephan, E.; Halappanavar, M.; Kerbyson, D. J.

    2017-10-01

    We propose a novel approach for efficient execution of workflows on distributed resources. The key components of this framework include: performance modeling to quantitatively predict workflow component behavior; optimization-based scheduling such as choosing an optimal subset of resources to meet demand and assignment of tasks to resources; distributed I/O optimizations such as prefetching; and provenance methods for collecting performance data. In preliminary results, these techniques improve throughput on a small Belle II workflow by 20%.

  6. High-throughput characterization for solar fuels materials discovery

    Science.gov (United States)

    Mitrovic, Slobodan; Becerra, Natalie; Cornell, Earl; Guevarra, Dan; Haber, Joel; Jin, Jian; Jones, Ryan; Kan, Kevin; Marcin, Martin; Newhouse, Paul; Soedarmadji, Edwin; Suram, Santosh; Xiang, Chengxiang; Gregoire, John; High-Throughput Experimentation Team

    2014-03-01

    In this talk I will present the status of the High-Throughput Experimentation (HTE) project of the Joint Center for Artificial Photosynthesis (JCAP). JCAP is an Energy Innovation Hub of the U.S. Department of Energy with a mandate to deliver a solar fuel generator based on an integrated photoelectrochemical cell (PEC). However, efficient and commercially viable catalysts or light absorbers for the PEC do not exist. The mission of HTE is to provide the accelerated discovery through combinatorial synthesis and rapid screening of material properties. The HTE pipeline also features high-throughput material characterization using x-ray diffraction and x-ray photoemission spectroscopy (XPS). In this talk I present the currently operating pipeline and focus on our combinatorial XPS efforts to build the largest free database of spectra from mixed-metal oxides, nitrides, sulfides and alloys. This work was performed at Joint Center for Artificial Photosynthesis, a DOE Energy Innovation Hub, supported through the Office of Science of the U.S. Department of Energy under Award No. DE-SC0004993.

  7. Fluorescent foci quantitation for high-throughput analysis

    Directory of Open Access Journals (Sweden)

    Elena Ledesma-Fernández

    2015-06-01

    Full Text Available A number of cellular proteins localize to discrete foci within cells, for example DNA repair proteins, microtubule organizing centers, P bodies or kinetochores. It is often possible to measure the fluorescence emission from tagged proteins within these foci as a surrogate for the concentration of that specific protein. We wished to develop tools that would allow quantitation of fluorescence foci intensities in high-throughput studies. As proof of principle we have examined the kinetochore, a large multi-subunit complex that is critical for the accurate segregation of chromosomes during cell division. Kinetochore perturbations lead to aneuploidy, which is a hallmark of cancer cells. Hence, understanding kinetochore homeostasis and regulation are important for a global understanding of cell division and genome integrity. The 16 budding yeast kinetochores colocalize within the nucleus to form a single focus. Here we have created a set of freely-available tools to allow high-throughput quantitation of kinetochore foci fluorescence. We use this ‘FociQuant’ tool to compare methods of kinetochore quantitation and we show proof of principle that FociQuant can be used to identify changes in kinetochore protein levels in a mutant that affects kinetochore function. This analysis can be applied to any protein that forms discrete foci in cells.

  8. poolHiTS: A Shifted Transversal Design based pooling strategy for high-throughput drug screening

    Directory of Open Access Journals (Sweden)

    Woolf Peter J

    2008-05-01

    Full Text Available Abstract Background A key goal of drug discovery is to increase the throughput of small molecule screens without sacrificing screening accuracy. High-throughput screening (HTS in drug discovery involves testing a large number of compounds in a biological assay to identify active compounds. Normally, molecules from a large compound library are tested individually to identify the activity of each molecule. Usually a small number of compounds are found to be active, however the presence of false positive and negative testing errors suggests that this one-drug one-assay screening strategy can be significantly improved. Pooling designs are testing schemes that test mixtures of compounds in each assay, thereby generating a screen of the whole compound library in fewer tests. By repeatedly testing compounds in different combinations, pooling designs also allow for error-correction. These pooled designs, for specific experiment parameters, can be simply and efficiently created using the Shifted Transversal Design (STD pooling algorithm. However, drug screening contains a number of key constraints that require specific modifications if this pooling approach is to be useful for practical screen designs. Results In this paper, we introduce a pooling strategy called poolHiTS (Pooled High-Throughput Screening which is based on the STD algorithm. In poolHiTS, we implement a limit on the number of compounds that can be mixed in a single assay. In addition, we show that the STD-based pooling strategy is limited in the error-correction that it can achieve. Due to the mixing constraint, we show that it is more efficient to split a large library into smaller blocks of compounds, which are then tested using an optimized strategy repeated for each block. We package the optimal block selection algorithm into poolHiTS. The MATLAB codes for the poolHiTS algorithm and the corresponding decoding strategy are also provided. Conclusion We have produced a practical version

  9. High-throughput electrical characterization for robust overlay lithography control

    Science.gov (United States)

    Devender, Devender; Shen, Xumin; Duggan, Mark; Singh, Sunil; Rullan, Jonathan; Choo, Jae; Mehta, Sohan; Tang, Teck Jung; Reidy, Sean; Holt, Jonathan; Kim, Hyung Woo; Fox, Robert; Sohn, D. K.

    2017-03-01

    Realizing sensitive, high throughput and robust overlay measurement is a challenge in current 14nm and advanced upcoming nodes with transition to 300mm and upcoming 450mm semiconductor manufacturing, where slight deviation in overlay has significant impact on reliability and yield1). Exponentially increasing number of critical masks in multi-patterning lithoetch, litho-etch (LELE) and subsequent LELELE semiconductor processes require even tighter overlay specification2). Here, we discuss limitations of current image- and diffraction- based overlay measurement techniques to meet these stringent processing requirements due to sensitivity, throughput and low contrast3). We demonstrate a new electrical measurement based technique where resistance is measured for a macro with intentional misalignment between two layers. Overlay is quantified by a parabolic fitting model to resistance where minima and inflection points are extracted to characterize overlay control and process window, respectively. Analyses using transmission electron microscopy show good correlation between actual overlay performance and overlay obtained from fitting. Additionally, excellent correlation of overlay from electrical measurements to existing image- and diffraction- based techniques is found. We also discuss challenges of integrating electrical measurement based approach in semiconductor manufacturing from Back End of Line (BEOL) perspective. Our findings open up a new pathway for accessing simultaneous overlay as well as process window and margins from a robust, high throughput and electrical measurement approach.

  10. High-throughput Sequencing Based Immune Repertoire Study during Infectious Disease

    Directory of Open Access Journals (Sweden)

    Dongni Hou

    2016-08-01

    Full Text Available The selectivity of the adaptive immune response is based on the enormous diversity of T and B cell antigen-specific receptors. The immune repertoire, the collection of T and B cells with functional diversity in the circulatory system at any given time, is dynamic and reflects the essence of immune selectivity. In this article, we review the recent advances in immune repertoire study of infectious diseases that achieved by traditional techniques and high-throughput sequencing techniques. High-throughput sequencing techniques enable the determination of complementary regions of lymphocyte receptors with unprecedented efficiency and scale. This progress in methodology enhances the understanding of immunologic changes during pathogen challenge, and also provides a basis for further development of novel diagnostic markers, immunotherapies and vaccines.

  11. Green throughput taxation

    International Nuclear Information System (INIS)

    Bruvoll, A.; Ibenholt, K.

    1998-01-01

    According to optimal taxation theory, raw materials should be taxed to capture the embedded scarcity rent in their value. To reduce both natural resource use and the corresponding emissions, or the throughput in the economic system, the best policy may be a tax on material inputs. As a first approach to throughput taxation, this paper considers a tax on intermediates in the framework of a dynamic computable general equilibrium model with environmental feedbacks. To balance the budget, payroll taxes are reduced. As a result, welfare indicators as material consumption and leisure time consumption are reduced, while on the other hand all the environmental indicators improve. 27 refs

  12. High-Throughput Flow Cytometric Method for the Simultaneous Measurement of CAR-T Cell Characterization and Cytotoxicity against Solid Tumor Cell Lines.

    Science.gov (United States)

    Martinez, Emily M; Klebanoff, Samuel D; Secrest, Stephanie; Romain, Gabrielle; Haile, Samuel T; Emtage, Peter C R; Gilbert, Amy E

    2018-04-01

    High-throughput flow cytometry is an attractive platform for the analysis of adoptive cellular therapies such as chimeric antigen receptor T cell therapy (CAR-T) because it allows for the concurrent measurement of T cell-dependent cellular cytotoxicity (TDCC) and the functional characterization of engineered T cells with respect to percentage of CAR transduction, T cell phenotype, and measurement of T cell function such as activation in a single assay. The use of adherent tumor cell lines can be challenging in these flow-based assays. Here, we present the development of a high-throughput flow-based assay to measure TDCC for a CAR-T construct co-cultured with multiple adherent tumor cell lines. We describe optimal assay conditions (such as adherent cell dissociation techniques to minimize impact on cell viability) that result in robust cytotoxicity assays. In addition, we report on the concurrent use of T cell transduction and activation antibody panels (CD25) that provide further dissection of engineered T cell function. In conclusion, we present the development of a high-throughput flow cytometry method allowing for in vitro interrogation of solid tumor, targeting CAR-T cell-mediated cytotoxicity, CAR transduction, and engineered T cell characterization in a single assay.

  13. Subnuclear foci quantification using high-throughput 3D image cytometry

    Science.gov (United States)

    Wadduwage, Dushan N.; Parrish, Marcus; Choi, Heejin; Engelward, Bevin P.; Matsudaira, Paul; So, Peter T. C.

    2015-07-01

    Ionising radiation causes various types of DNA damages including double strand breaks (DSBs). DSBs are often recognized by DNA repair protein ATM which forms gamma-H2AX foci at the site of the DSBs that can be visualized using immunohistochemistry. However most of such experiments are of low throughput in terms of imaging and image analysis techniques. Most of the studies still use manual counting or classification. Hence they are limited to counting a low number of foci per cell (5 foci per nucleus) as the quantification process is extremely labour intensive. Therefore we have developed a high throughput instrumentation and computational pipeline specialized for gamma-H2AX foci quantification. A population of cells with highly clustered foci inside nuclei were imaged, in 3D with submicron resolution, using an in-house developed high throughput image cytometer. Imaging speeds as high as 800 cells/second in 3D were achieved by using HiLo wide-field depth resolved imaging and a remote z-scanning technique. Then the number of foci per cell nucleus were quantified using a 3D extended maxima transform based algorithm. Our results suggests that while most of the other 2D imaging and manual quantification studies can count only up to about 5 foci per nucleus our method is capable of counting more than 100. Moreover we show that 3D analysis is significantly superior compared to the 2D techniques.

  14. A high throughput array microscope for the mechanical characterization of biomaterials

    Science.gov (United States)

    Cribb, Jeremy; Osborne, Lukas D.; Hsiao, Joe Ping-Lin; Vicci, Leandra; Meshram, Alok; O'Brien, E. Tim; Spero, Richard Chasen; Taylor, Russell; Superfine, Richard

    2015-02-01

    In the last decade, the emergence of high throughput screening has enabled the development of novel drug therapies and elucidated many complex cellular processes. Concurrently, the mechanobiology community has developed tools and methods to show that the dysregulation of biophysical properties and the biochemical mechanisms controlling those properties contribute significantly to many human diseases. Despite these advances, a complete understanding of the connection between biomechanics and disease will require advances in instrumentation that enable parallelized, high throughput assays capable of probing complex signaling pathways, studying biology in physiologically relevant conditions, and capturing specimen and mechanical heterogeneity. Traditional biophysical instruments are unable to meet this need. To address the challenge of large-scale, parallelized biophysical measurements, we have developed an automated array high-throughput microscope system that utilizes passive microbead diffusion to characterize mechanical properties of biomaterials. The instrument is capable of acquiring data on twelve-channels simultaneously, where each channel in the system can independently drive two-channel fluorescence imaging at up to 50 frames per second. We employ this system to measure the concentration-dependent apparent viscosity of hyaluronan, an essential polymer found in connective tissue and whose expression has been implicated in cancer progression.

  15. High-throughput bioinformatics with the Cyrille2 pipeline system.

    NARCIS (Netherlands)

    Fiers, M.W.E.J.; Burgt, van der A.; Datema, E.; Groot, de J.C.W.; Ham, van R.C.H.J.

    2008-01-01

    Background - Modern omics research involves the application of high-throughput technologies that generate vast volumes of data. These data need to be pre-processed, analyzed and integrated with existing knowledge through the use of diverse sets of software tools, models and databases. The analyses

  16. High-throughput preparation and testing of ion-exchanged zeolites

    International Nuclear Information System (INIS)

    Janssen, K.P.F.; Paul, J.S.; Sels, B.F.; Jacobs, P.A.

    2007-01-01

    A high-throughput research platform was developed for the preparation and subsequent catalytic liquid-phase screening of ion-exchanged zeolites, for instance with regard to their use as heterogeneous catalysts. In this system aqueous solutions and other liquid as well as solid reagents are employed as starting materials and 24 samples are prepared on a library plate with a 4 x 6 layout. Volumetric dispensing of metal precursor solutions, weighing of zeolite and subsequent mixing/washing cycles of the starting materials and distributing reaction mixtures to the library plate are automatically performed by liquid and solid handlers controlled by a single common and easy-to-use programming software interface. The thus prepared materials are automatically contacted with reagent solutions, heated, stirred and sampled continuously using a modified liquid handling. The high-throughput platform is highly promising in enhancing synthesis of catalysts and their screening. In this paper the preparation of lanthanum-exchanged NaY zeolites (LaNaY) on the platform is reported, along with their use as catalyst for the conversion of renewables

  17. A Self-Reporting Photocatalyst for Online Fluorescence Monitoring of High Throughput RAFT Polymerization.

    Science.gov (United States)

    Yeow, Jonathan; Joshi, Sanket; Chapman, Robert; Boyer, Cyrille Andre Jean Marie

    2018-04-25

    Translating controlled/living radical polymerization (CLRP) from batch to the high throughput production of polymer libraries presents several challenges in terms of both polymer synthesis and characterization. Although recently there have been significant advances in the field of low volume, high throughput CLRP, techniques able to simultaneously monitor multiple polymerizations in an "online" manner have not yet been developed. Here, we report our discovery that 5,10,15,20-tetraphenyl-21H,23H-porphine zinc (ZnTPP) is a self-reporting photocatalyst that can mediate PET-RAFT polymerization as well as report on monomer conversion via changes in its fluorescence properties. This enables the use of a microplate reader to conduct high throughput "online" monitoring of PET-RAFT polymerizations performed directly in 384-well, low volume microtiter plates. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. High-throughput detection of ethanol-producing cyanobacteria in a microdroplet platform.

    Science.gov (United States)

    Abalde-Cela, Sara; Gould, Anna; Liu, Xin; Kazamia, Elena; Smith, Alison G; Abell, Chris

    2015-05-06

    Ethanol production by microorganisms is an important renewable energy source. Most processes involve fermentation of sugars from plant feedstock, but there is increasing interest in direct ethanol production by photosynthetic organisms. To facilitate this, a high-throughput screening technique for the detection of ethanol is required. Here, a method for the quantitative detection of ethanol in a microdroplet-based platform is described that can be used for screening cyanobacterial strains to identify those with the highest ethanol productivity levels. The detection of ethanol by enzymatic assay was optimized both in bulk and in microdroplets. In parallel, the encapsulation of engineered ethanol-producing cyanobacteria in microdroplets and their growth dynamics in microdroplet reservoirs were demonstrated. The combination of modular microdroplet operations including droplet generation for cyanobacteria encapsulation, droplet re-injection and pico-injection, and laser-induced fluorescence, were used to create this new platform to screen genetically engineered strains of cyanobacteria with different levels of ethanol production.

  19. High-throughput fragment screening by affinity LC-MS.

    Science.gov (United States)

    Duong-Thi, Minh-Dao; Bergström, Maria; Fex, Tomas; Isaksson, Roland; Ohlson, Sten

    2013-02-01

    Fragment screening, an emerging approach for hit finding in drug discovery, has recently been proven effective by its first approved drug, vemurafenib, for cancer treatment. Techniques such as nuclear magnetic resonance, surface plasmon resonance, and isothemal titration calorimetry, with their own pros and cons, have been employed for screening fragment libraries. As an alternative approach, screening based on high-performance liquid chromatography separation has been developed. In this work, we present weak affinity LC/MS as a method to screen fragments under high-throughput conditions. Affinity-based capillary columns with immobilized thrombin were used to screen a collection of 590 compounds from a fragment library. The collection was divided into 11 mixtures (each containing 35 to 65 fragments) and screened by MS detection. The primary screening was performed in 3500 fragments per day). Thirty hits were defined, which subsequently entered a secondary screening using an active site-blocked thrombin column for confirmation of specificity. One hit showed selective binding to thrombin with an estimated dissociation constant (K (D)) in the 0.1 mM range. This study shows that affinity LC/MS is characterized by high throughput, ease of operation, and low consumption of target and fragments, and therefore it promises to be a valuable method for fragment screening.

  20. High Throughput Multispectral Image Processing with Applications in Food Science.

    Directory of Open Access Journals (Sweden)

    Panagiotis Tsakanikas

    Full Text Available Recently, machine vision is gaining attention in food science as well as in food industry concerning food quality assessment and monitoring. Into the framework of implementation of Process Analytical Technology (PAT in the food industry, image processing can be used not only in estimation and even prediction of food quality but also in detection of adulteration. Towards these applications on food science, we present here a novel methodology for automated image analysis of several kinds of food products e.g. meat, vanilla crème and table olives, so as to increase objectivity, data reproducibility, low cost information extraction and faster quality assessment, without human intervention. Image processing's outcome will be propagated to the downstream analysis. The developed multispectral image processing method is based on unsupervised machine learning approach (Gaussian Mixture Models and a novel unsupervised scheme of spectral band selection for segmentation process optimization. Through the evaluation we prove its efficiency and robustness against the currently available semi-manual software, showing that the developed method is a high throughput approach appropriate for massive data extraction from food samples.

  1. High Throughput Multispectral Image Processing with Applications in Food Science.

    Science.gov (United States)

    Tsakanikas, Panagiotis; Pavlidis, Dimitris; Nychas, George-John

    2015-01-01

    Recently, machine vision is gaining attention in food science as well as in food industry concerning food quality assessment and monitoring. Into the framework of implementation of Process Analytical Technology (PAT) in the food industry, image processing can be used not only in estimation and even prediction of food quality but also in detection of adulteration. Towards these applications on food science, we present here a novel methodology for automated image analysis of several kinds of food products e.g. meat, vanilla crème and table olives, so as to increase objectivity, data reproducibility, low cost information extraction and faster quality assessment, without human intervention. Image processing's outcome will be propagated to the downstream analysis. The developed multispectral image processing method is based on unsupervised machine learning approach (Gaussian Mixture Models) and a novel unsupervised scheme of spectral band selection for segmentation process optimization. Through the evaluation we prove its efficiency and robustness against the currently available semi-manual software, showing that the developed method is a high throughput approach appropriate for massive data extraction from food samples.

  2. Structural, dielectric and ferroelectric properties of (Bi,Na)TiO{sub 3}–BaTiO{sub 3} system studied by high throughput screening

    Energy Technology Data Exchange (ETDEWEB)

    Hayden, Brian E. [Ilika Technologies Plc., Kenneth Dibben House, Enterprise Road, University of Southampton Science Park, Chilworth, Southampton SO16 7NS (United Kingdom); Department of Chemistry, University of Southampton, Highfield, Southampton SO17 1BJ (United Kingdom); Yakovlev, Sergey, E-mail: sergey.yakovlev@ilika.com [Ilika Technologies Plc., Kenneth Dibben House, Enterprise Road, University of Southampton Science Park, Chilworth, Southampton SO16 7NS (United Kingdom)

    2016-03-31

    Thin-film materials libraries of the Bi{sub 2}O{sub 3}–Na{sub 2}O–TiO{sub 2}–BaO system in a broad composition range have been deposited in ultra-high vacuum from elemental evaporation sources and an oxygen plasma source. A high throughput approach was used for systematic compositional and structural characterization and the screening of the dielectric and ferroelectric properties. The perovskite (Bi,Na)TiO{sub 3}–BaTiO{sub 3} phase with a Ba concentration near the morphotropic phase boundary (ca. 6 at.%) exhibited a relative dielectric permittivity of 180, a loss tangent of 0.04 and remnant polarization of 19 μC/cm{sup 2}. Compared to published data, observed remnant polarization is close to that known for epitaxially grown films but higher than the values reported for polycrystalline films. The high throughput methodology and systematic nature of the study allowed us to establish the composition boundaries of the phase with optimal dielectric and ferroelectric characteristics. - Highlights: • Bi{sub 2}O{sub 3}–Na{sub 2}O–TiO{sub 2}–BaO high throughput materials library was deposited using PVD method. • Materials were processed from individual molecular beam epitaxy sources of elements. • High throughput approach was used for structural, dielectric and ferroelectric study. • Composition boundaries of perovskite compounds with optimum properties are reported.

  3. Multiplex enrichment quantitative PCR (ME-qPCR): a high-throughput, highly sensitive detection method for GMO identification.

    Science.gov (United States)

    Fu, Wei; Zhu, Pengyu; Wei, Shuang; Zhixin, Du; Wang, Chenguang; Wu, Xiyang; Li, Feiwu; Zhu, Shuifang

    2017-04-01

    Among all of the high-throughput detection methods, PCR-based methodologies are regarded as the most cost-efficient and feasible methodologies compared with the next-generation sequencing or ChIP-based methods. However, the PCR-based methods can only achieve multiplex detection up to 15-plex due to limitations imposed by the multiplex primer interactions. The detection throughput cannot meet the demands of high-throughput detection, such as SNP or gene expression analysis. Therefore, in our study, we have developed a new high-throughput PCR-based detection method, multiplex enrichment quantitative PCR (ME-qPCR), which is a combination of qPCR and nested PCR. The GMO content detection results in our study showed that ME-qPCR could achieve high-throughput detection up to 26-plex. Compared to the original qPCR, the Ct values of ME-qPCR were lower for the same group, which showed that ME-qPCR sensitivity is higher than the original qPCR. The absolute limit of detection for ME-qPCR could achieve levels as low as a single copy of the plant genome. Moreover, the specificity results showed that no cross-amplification occurred for irrelevant GMO events. After evaluation of all of the parameters, a practical evaluation was performed with different foods. The more stable amplification results, compared to qPCR, showed that ME-qPCR was suitable for GMO detection in foods. In conclusion, ME-qPCR achieved sensitive, high-throughput GMO detection in complex substrates, such as crops or food samples. In the future, ME-qPCR-based GMO content identification may positively impact SNP analysis or multiplex gene expression of food or agricultural samples. Graphical abstract For the first-step amplification, four primers (A, B, C, and D) have been added into the reaction volume. In this manner, four kinds of amplicons have been generated. All of these four amplicons could be regarded as the target of second-step PCR. For the second-step amplification, three parallels have been taken for

  4. A gas trapping method for high-throughput metabolic experiments.

    Science.gov (United States)

    Krycer, James R; Diskin, Ciana; Nelson, Marin E; Zeng, Xiao-Yi; Fazakerley, Daniel J; James, David E

    2018-01-01

    Research into cellular metabolism has become more high-throughput, with typical cell-culture experiments being performed in multiwell plates (microplates). This format presents a challenge when trying to collect gaseous products, such as carbon dioxide (CO2), which requires a sealed environment and a vessel separate from the biological sample. To address this limitation, we developed a gas trapping protocol using perforated plastic lids in sealed cell-culture multiwell plates. We used this trap design to measure CO2 production from glucose and fatty acid metabolism, as well as hydrogen sulfide production from cysteine-treated cells. Our data clearly show that this gas trap can be applied to liquid and solid gas-collection media and can be used to study gaseous product generation by both adherent cells and cells in suspension. Since our gas traps can be adapted to multiwell plates of various sizes, they present a convenient, cost-effective solution that can accommodate the trend toward high-throughput measurements in metabolic research.

  5. High-throughput technology for novel SO2 oxidation catalysts

    International Nuclear Information System (INIS)

    Loskyll, Jonas; Stoewe, Klaus; Maier, Wilhelm F

    2011-01-01

    We review the state of the art and explain the need for better SO 2 oxidation catalysts for the production of sulfuric acid. A high-throughput technology has been developed for the study of potential catalysts in the oxidation of SO 2 to SO 3 . High-throughput methods are reviewed and the problems encountered with their adaptation to the corrosive conditions of SO 2 oxidation are described. We show that while emissivity-corrected infrared thermography (ecIRT) can be used for primary screening, it is prone to errors because of the large variations in the emissivity of the catalyst surface. UV-visible (UV-Vis) spectrometry was selected instead as a reliable analysis method of monitoring the SO 2 conversion. Installing plain sugar absorbents at reactor outlets proved valuable for the detection and quantitative removal of SO 3 from the product gas before the UV-Vis analysis. We also overview some elements used for prescreening and those remaining after the screening of the first catalyst generations. (topical review)

  6. Turbocharged molecular discovery of OLED emitters: from high-throughput quantum simulation to highly efficient TADF devices

    Science.gov (United States)

    Gómez-Bombarelli, Rafael; Aguilera-Iparraguirre, Jorge; Hirzel, Timothy D.; Ha, Dong-Gwang; Einzinger, Markus; Wu, Tony; Baldo, Marc A.; Aspuru-Guzik, Alán.

    2016-09-01

    Discovering new OLED emitters requires many experiments to synthesize candidates and test performance in devices. Large scale computer simulation can greatly speed this search process but the problem remains challenging enough that brute force application of massive computing power is not enough to successfully identify novel structures. We report a successful High Throughput Virtual Screening study that leveraged a range of methods to optimize the search process. The generation of candidate structures was constrained to contain combinatorial explosion. Simulations were tuned to the specific problem and calibrated with experimental results. Experimentalists and theorists actively collaborated such that experimental feedback was regularly utilized to update and shape the computational search. Supervised machine learning methods prioritized candidate structures prior to quantum chemistry simulation to prevent wasting compute on likely poor performers. With this combination of techniques, each multiplying the strength of the search, this effort managed to navigate an area of molecular space and identify hundreds of promising OLED candidate structures. An experimentally validated selection of this set shows emitters with external quantum efficiencies as high as 22%.

  7. Use and optimization of a dual-flowrate loading strategy to maximize throughput in protein-a affinity chromatography.

    Science.gov (United States)

    Ghose, Sanchayita; Nagrath, Deepak; Hubbard, Brian; Brooks, Clayton; Cramer, Steven M

    2004-01-01

    The effect of an alternate strategy employing two different flowrates during loading was explored as a means of increasing system productivity in Protein-A chromatography. The effect of such a loading strategy was evaluated using a chromatographic model that was able to accurately predict experimental breakthrough curves for this Protein-A system. A gradient-based optimization routine is carried out to establish the optimal loading conditions (initial and final flowrates and switching time). The two-step loading strategy (using a higher flowrate during the initial stages followed by a lower flowrate) was evaluated for an Fc-fusion protein and was found to result in significant improvements in process throughput. In an extension of this optimization routine, dynamic loading capacity and productivity were simultaneously optimized using a weighted objective function, and this result was compared to that obtained with the single flowrate. Again, the dual-flowrate strategy was found to be superior.

  8. Integration of an In Situ MALDI-Based High-Throughput Screening Process: A Case Study with Receptor Tyrosine Kinase c-MET.

    Science.gov (United States)

    Beeman, Katrin; Baumgärtner, Jens; Laubenheimer, Manuel; Hergesell, Karlheinz; Hoffmann, Martin; Pehl, Ulrich; Fischer, Frank; Pieck, Jan-Carsten

    2017-12-01

    Mass spectrometry (MS) is known for its label-free detection of substrates and products from a variety of enzyme reactions. Recent hardware improvements have increased interest in the use of matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS for high-throughput drug discovery. Despite interest in this technology, several challenges remain and must be overcome before MALDI-MS can be integrated as an automated "in-line reader" for high-throughput drug discovery. Two such hurdles include in situ sample processing and deposition, as well as integration of MALDI-MS for enzymatic screening assays that usually contain high levels of MS-incompatible components. Here we adapt our c-MET kinase assay to optimize for MALDI-MS compatibility and test its feasibility for compound screening. The pros and cons of the Echo (Labcyte) as a transfer system for in situ MALDI-MS sample preparation are discussed. We demonstrate that this method generates robust data in a 1536-grid format. We use the MALDI-MS to directly measure the ratio of c-MET substrate and phosphorylated product to acquire IC50 curves and demonstrate that the pharmacology is unaffected. The resulting IC50 values correlate well between the common label-based capillary electrophoresis and the label-free MALDI-MS detection method. We predict that label-free MALDI-MS-based high-throughput screening will become increasingly important and more widely used for drug discovery.

  9. Comprehensive evaluation and optimization of amplicon library preparation methods for high-throughput antibody sequencing.

    Science.gov (United States)

    Menzel, Ulrike; Greiff, Victor; Khan, Tarik A; Haessler, Ulrike; Hellmann, Ina; Friedensohn, Simon; Cook, Skylar C; Pogson, Mark; Reddy, Sai T

    2014-01-01

    High-throughput sequencing (HTS) of antibody repertoire libraries has become a powerful tool in the field of systems immunology. However, numerous sources of bias in HTS workflows may affect the obtained antibody repertoire data. A crucial step in antibody library preparation is the addition of short platform-specific nucleotide adapter sequences. As of yet, the impact of the method of adapter addition on experimental library preparation and the resulting antibody repertoire HTS datasets has not been thoroughly investigated. Therefore, we compared three standard library preparation methods by performing Illumina HTS on antibody variable heavy genes from murine antibody-secreting cells. Clonal overlap and rank statistics demonstrated that the investigated methods produced equivalent HTS datasets. PCR-based methods were experimentally superior to ligation with respect to speed, efficiency, and practicality. Finally, using a two-step PCR based method we established a protocol for antibody repertoire library generation, beginning from inputs as low as 1 ng of total RNA. In summary, this study represents a major advance towards a standardized experimental framework for antibody HTS, thus opening up the potential for systems-based, cross-experiment meta-analyses of antibody repertoires.

  10. High-throughput screening of carbohydrate-degrading enzymes using novel insoluble chromogenic substrate assay kits

    DEFF Research Database (Denmark)

    Schückel, Julia; Kracun, Stjepan Kresimir; Willats, William George Tycho

    2016-01-01

    for this is that advances in genome and transcriptome sequencing, together with associated bioinformatics tools allow for rapid identification of candidate CAZymes, but technology for determining an enzyme's biochemical characteristics has advanced more slowly. To address this technology gap, a novel high-throughput assay...... CPH and ICB substrates are provided in a 96-well high-throughput assay system. The CPH substrates can be made in four different colors, enabling them to be mixed together and thus increasing assay throughput. The protocol describes a 96-well plate assay and illustrates how this assay can be used...... for screening the activities of enzymes, enzyme cocktails, and broths....

  11. High-Throughput Quantification of SH2 Domain-Phosphopeptide Interactions with Cellulose-Peptide Conjugate Microarrays.

    Science.gov (United States)

    Engelmann, Brett W

    2017-01-01

    The Src Homology 2 (SH2) domain family primarily recognizes phosphorylated tyrosine (pY) containing peptide motifs. The relative affinity preferences among competing SH2 domains for phosphopeptide ligands define "specificity space," and underpins many functional pY mediated interactions within signaling networks. The degree of promiscuity exhibited and the dynamic range of affinities supported by individual domains or phosphopeptides is best resolved by a carefully executed and controlled quantitative high-throughput experiment. Here, I describe the fabrication and application of a cellulose-peptide conjugate microarray (CPCMA) platform to the quantitative analysis of SH2 domain specificity space. Included herein are instructions for optimal experimental design with special attention paid to common sources of systematic error, phosphopeptide SPOT synthesis, microarray fabrication, analyte titrations, data capture, and analysis.

  12. Integrated Automation of High-Throughput Screening and Reverse Phase Protein Array Sample Preparation

    DEFF Research Database (Denmark)

    Pedersen, Marlene Lemvig; Block, Ines; List, Markus

    into automated robotic high-throughput screens, which allows subsequent protein quantification. In this integrated solution, samples are directly forwarded to automated cell lysate preparation and preparation of dilution series, including reformatting to a protein spotter-compatible format after the high......-throughput screening. Tracking of huge sample numbers and data analysis from a high-content screen to RPPAs is accomplished via MIRACLE, a custom made software suite developed by us. To this end, we demonstrate that the RPPAs generated in this manner deliver reliable protein readouts and that GAPDH and TFR levels can...

  13. High-throughput screening of filamentous fungi using nanoliter-range droplet-based microfluidics

    Science.gov (United States)

    Beneyton, Thomas; Wijaya, I. Putu Mahendra; Postros, Prexilia; Najah, Majdi; Leblond, Pascal; Couvent, Angélique; Mayot, Estelle; Griffiths, Andrew D.; Drevelle, Antoine

    2016-06-01

    Filamentous fungi are an extremely important source of industrial enzymes because of their capacity to secrete large quantities of proteins. Currently, functional screening of fungi is associated with low throughput and high costs, which severely limits the discovery of novel enzymatic activities and better production strains. Here, we describe a nanoliter-range droplet-based microfluidic system specially adapted for the high-throughput sceening (HTS) of large filamentous fungi libraries for secreted enzyme activities. The platform allowed (i) compartmentalization of single spores in ~10 nl droplets, (ii) germination and mycelium growth and (iii) high-throughput sorting of fungi based on enzymatic activity. A 104 clone UV-mutated library of Aspergillus niger was screened based on α-amylase activity in just 90 minutes. Active clones were enriched 196-fold after a single round of microfluidic HTS. The platform is a powerful tool for the development of new production strains with low cost, space and time footprint and should bring enormous benefit for improving the viability of biotechnological processes.

  14. The application of the high throughput sequencing technology in the transposable elements.

    Science.gov (United States)

    Liu, Zhen; Xu, Jian-hong

    2015-09-01

    High throughput sequencing technology has dramatically improved the efficiency of DNA sequencing, and decreased the costs to a great extent. Meanwhile, this technology usually has advantages of better specificity, higher sensitivity and accuracy. Therefore, it has been applied to the research on genetic variations, transcriptomics and epigenomics. Recently, this technology has been widely employed in the studies of transposable elements and has achieved fruitful results. In this review, we summarize the application of high throughput sequencing technology in the fields of transposable elements, including the estimation of transposon content, preference of target sites and distribution, insertion polymorphism and population frequency, identification of rare copies, transposon horizontal transfers as well as transposon tagging. We also briefly introduce the major common sequencing strategies and algorithms, their advantages and disadvantages, and the corresponding solutions. Finally, we envision the developing trends of high throughput sequencing technology, especially the third generation sequencing technology, and its application in transposon studies in the future, hopefully providing a comprehensive understanding and reference for related scientific researchers.

  15. Chemiluminescence analyzer of NOx as a high-throughput screening tool in selective catalytic reduction of NO

    International Nuclear Information System (INIS)

    Oh, Kwang Seok; Woo, Seong Ihl

    2011-01-01

    A chemiluminescence-based analyzer of NO x gas species has been applied for high-throughput screening of a library of catalytic materials. The applicability of the commercial NO x analyzer as a rapid screening tool was evaluated using selective catalytic reduction of NO gas. A library of 60 binary alloys composed of Pt and Co, Zr, La, Ce, Fe or W on Al 2 O 3 substrate was tested for the efficiency of NO x removal using a home-built 64-channel parallel and sequential tubular reactor. The NO x concentrations measured by the NO x analyzer agreed well with the results obtained using micro gas chromatography for a reference catalyst consisting of 1 wt% Pt on γ-Al 2 O 3 . Most alloys showed high efficiency at 275 °C, which is typical of Pt-based catalysts for selective catalytic reduction of NO. The screening with NO x analyzer allowed to select Pt-Ce (X) (X=1–3) and Pt–Fe (2) as the optimal catalysts for NO x removal: 73% NO x conversion was achieved with the Pt–Fe (2) alloy, which was much better than the results for the reference catalyst and the other library alloys. This study demonstrates a sequential high-throughput method of practical evaluation of catalysts for the selective reduction of NO.

  16. High-Throughput Cancer Cell Sphere Formation for 3D Cell Culture.

    Science.gov (United States)

    Chen, Yu-Chih; Yoon, Euisik

    2017-01-01

    Three-dimensional (3D) cell culture is critical in studying cancer pathology and drug response. Though 3D cancer sphere culture can be performed in low-adherent dishes or well plates, the unregulated cell aggregation may skew the results. On contrary, microfluidic 3D culture can allow precise control of cell microenvironments, and provide higher throughput by orders of magnitude. In this chapter, we will look into engineering innovations in a microfluidic platform for high-throughput cancer cell sphere formation and review the implementation methods in detail.

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

    Science.gov (United States)

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

    2016-01-01

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

  18. High-throughput shotgun lipidomics by quadrupole time-of-flight mass spectrometry

    DEFF Research Database (Denmark)

    Ståhlman, Marcus; Ejsing, Christer S.; Tarasov, Kirill

    2009-01-01

    Technological advances in mass spectrometry and meticulous method development have produced several shotgun lipidomic approaches capable of characterizing lipid species by direct analysis of total lipid extracts. Shotgun lipidomics by hybrid quadrupole time-of-flight mass spectrometry allows...... the absolute quantification of hundreds of molecular glycerophospholipid species, glycerolipid species, sphingolipid species and sterol lipids. Future applications in clinical cohort studies demand detailed lipid molecule information and the application of high-throughput lipidomics platforms. In this review...... we describe a novel high-throughput shotgun lipidomic platform based on 96-well robot-assisted lipid extraction, automated sample infusion by mircofluidic-based nanoelectrospray ionization, and quantitative multiple precursor ion scanning analysis on a quadrupole time-of-flight mass spectrometer...

  19. High-throughput density functional calculations to optimize properties and interfacial chemistry of piezoelectric materials

    Science.gov (United States)

    Barr, Jordan A.; Lin, Fang-Yin; Ashton, Michael; Hennig, Richard G.; Sinnott, Susan B.

    2018-02-01

    High-throughput density functional theory calculations are conducted to search through 1572 A B O3 compounds to find a potential replacement material for lead zirconate titanate (PZT) that exhibits the same excellent piezoelectric properties as PZT and lacks both its use of the toxic element lead (Pb) and the formation of secondary alloy phases with platinum (Pt) electrodes. The first screening criterion employed a search through the Materials Project database to find A -B combinations that do not form ternary compounds with Pt. The second screening criterion aimed to eliminate potential candidates through first-principles calculations of their electronic structure, in which compounds with a band gap of 0.25 eV or higher were retained. Third, thermodynamic stability calculations were used to compare the candidates in a Pt environment to compounds already calculated to be stable within the Materials Project. Formation energies below or equal to 100 meV/atom were considered to be thermodynamically stable. The fourth screening criterion employed lattice misfit to identify those candidate perovskites that have low misfit with the Pt electrode and high misfit of potential secondary phases that can be formed when Pt alloys with the different A and B components. To aid in the final analysis, dynamic stability calculations were used to determine those perovskites that have dynamic instabilities that favor the ferroelectric distortion. Analysis of the data finds three perovskites warranting further investigation: CsNb O3 , RbNb O3 , and CsTa O3 .

  20. Infra-red thermography for high throughput field phenotyping in Solanum tuberosum.

    Directory of Open Access Journals (Sweden)

    Ankush Prashar

    Full Text Available The rapid development of genomic technology has made high throughput genotyping widely accessible but the associated high throughput phenotyping is now the major limiting factor in genetic analysis of traits. This paper evaluates the use of thermal imaging for the high throughput field phenotyping of Solanum tuberosum for differences in stomatal behaviour. A large multi-replicated trial of a potato mapping population was used to investigate the consistency in genotypic rankings across different trials and across measurements made at different times of day and on different days. The results confirmed a high degree of consistency between the genotypic rankings based on relative canopy temperature on different occasions. Genotype discrimination was enhanced both through normalising data by expressing genotype temperatures as differences from image means and through the enhanced replication obtained by using overlapping images. A Monte Carlo simulation approach was used to confirm the magnitude of genotypic differences that it is possible to discriminate. The results showed a clear negative association between canopy temperature and final tuber yield for this population, when grown under ample moisture supply. We have therefore established infrared thermography as an easy, rapid and non-destructive screening method for evaluating large population trials for genetic analysis. We also envisage this approach as having great potential for evaluating plant response to stress under field conditions.

  1. High-throughput purification of recombinant proteins using self-cleaving intein tags.

    Science.gov (United States)

    Coolbaugh, M J; Shakalli Tang, M J; Wood, D W

    2017-01-01

    High throughput methods for recombinant protein production using E. coli typically involve the use of affinity tags for simple purification of the protein of interest. One drawback of these techniques is the occasional need for tag removal before study, which can be hard to predict. In this work, we demonstrate two high throughput purification methods for untagged protein targets based on simple and cost-effective self-cleaving intein tags. Two model proteins, E. coli beta-galactosidase (βGal) and superfolder green fluorescent protein (sfGFP), were purified using self-cleaving versions of the conventional chitin-binding domain (CBD) affinity tag and the nonchromatographic elastin-like-polypeptide (ELP) precipitation tag in a 96-well filter plate format. Initial tests with shake flask cultures confirmed that the intein purification scheme could be scaled down, with >90% pure product generated in a single step using both methods. The scheme was then validated in a high throughput expression platform using 24-well plate cultures followed by purification in 96-well plates. For both tags and with both target proteins, the purified product was consistently obtained in a single-step, with low well-to-well and plate-to-plate variability. This simple method thus allows the reproducible production of highly pure untagged recombinant proteins in a convenient microtiter plate format. Copyright © 2016 Elsevier Inc. All rights reserved.

  2. High-throughput screening in niche-based assay identifies compounds to target preleukemic stem cells

    Science.gov (United States)

    Gerby, Bastien; Veiga, Diogo F.T.; Krosl, Jana; Nourreddine, Sami; Ouellette, Julianne; Haman, André; Lavoie, Geneviève; Fares, Iman; Tremblay, Mathieu; Litalien, Véronique; Ottoni, Elizabeth; Geoffrion, Dominique; Maddox, Paul S.; Chagraoui, Jalila; Hébert, Josée; Sauvageau, Guy; Kwok, Benjamin H.; Roux, Philippe P.

    2016-01-01

    Current chemotherapies for T cell acute lymphoblastic leukemia (T-ALL) efficiently reduce tumor mass. Nonetheless, disease relapse attributed to survival of preleukemic stem cells (pre-LSCs) is associated with poor prognosis. Herein, we provide direct evidence that pre-LSCs are much less chemosensitive to existing chemotherapy drugs than leukemic blasts because of a distinctive lower proliferative state. Improving therapies for T-ALL requires the development of strategies to target pre-LSCs that are absolutely dependent on their microenvironment. Therefore, we designed a robust protocol for high-throughput screening of compounds that target primary pre-LSCs maintained in a niche-like environment, on stromal cells that were engineered for optimal NOTCH1 activation. The multiparametric readout takes into account the intrinsic complexity of primary cells in order to specifically monitor pre-LSCs, which were induced here by the SCL/TAL1 and LMO1 oncogenes. We screened a targeted library of compounds and determined that the estrogen derivative 2-methoxyestradiol (2-ME2) disrupted both cell-autonomous and non–cell-autonomous pathways. Specifically, 2-ME2 abrogated pre-LSC viability and self-renewal activity in vivo by inhibiting translation of MYC, a downstream effector of NOTCH1, and preventing SCL/TAL1 activity. In contrast, normal hematopoietic stem/progenitor cells remained functional. These results illustrate how recapitulating tissue-like properties of primary cells in high-throughput screening is a promising avenue for innovation in cancer chemotherapy. PMID:27797342

  3. High Throughput Microplate Respiratory Measurements Using Minimal Quantities Of Isolated Mitochondria

    Science.gov (United States)

    Rogers, George W.; Brand, Martin D.; Petrosyan, Susanna; Ashok, Deepthi; Elorza, Alvaro A.; Ferrick, David A.; Murphy, Anne N.

    2011-01-01

    Recently developed technologies have enabled multi-well measurement of O2 consumption, facilitating the rate of mitochondrial research, particularly regarding the mechanism of action of drugs and proteins that modulate metabolism. Among these technologies, the Seahorse XF24 Analyzer was designed for use with intact cells attached in a monolayer to a multi-well tissue culture plate. In order to have a high throughput assay system in which both energy demand and substrate availability can be tightly controlled, we have developed a protocol to expand the application of the XF24 Analyzer to include isolated mitochondria. Acquisition of optimal rates requires assay conditions that are unexpectedly distinct from those of conventional polarography. The optimized conditions, derived from experiments with isolated mouse liver mitochondria, allow multi-well assessment of rates of respiration and proton production by mitochondria attached to the bottom of the XF assay plate, and require extremely small quantities of material (1–10 µg of mitochondrial protein per well). Sequential measurement of basal, State 3, State 4, and uncoupler-stimulated respiration can be made in each well through additions of reagents from the injection ports. We describe optimization and validation of this technique using isolated mouse liver and rat heart mitochondria, and apply the approach to discover that inclusion of phosphatase inhibitors in the preparation of the heart mitochondria results in a specific decrease in rates of Complex I-dependent respiration. We believe this new technique will be particularly useful for drug screening and for generating previously unobtainable respiratory data on small mitochondrial samples. PMID:21799747

  4. High throughput microplate respiratory measurements using minimal quantities of isolated mitochondria.

    Directory of Open Access Journals (Sweden)

    George W Rogers

    Full Text Available Recently developed technologies have enabled multi-well measurement of O(2 consumption, facilitating the rate of mitochondrial research, particularly regarding the mechanism of action of drugs and proteins that modulate metabolism. Among these technologies, the Seahorse XF24 Analyzer was designed for use with intact cells attached in a monolayer to a multi-well tissue culture plate. In order to have a high throughput assay system in which both energy demand and substrate availability can be tightly controlled, we have developed a protocol to expand the application of the XF24 Analyzer to include isolated mitochondria. Acquisition of optimal rates requires assay conditions that are unexpectedly distinct from those of conventional polarography. The optimized conditions, derived from experiments with isolated mouse liver mitochondria, allow multi-well assessment of rates of respiration and proton production by mitochondria attached to the bottom of the XF assay plate, and require extremely small quantities of material (1-10 µg of mitochondrial protein per well. Sequential measurement of basal, State 3, State 4, and uncoupler-stimulated respiration can be made in each well through additions of reagents from the injection ports. We describe optimization and validation of this technique using isolated mouse liver and rat heart mitochondria, and apply the approach to discover that inclusion of phosphatase inhibitors in the preparation of the heart mitochondria results in a specific decrease in rates of Complex I-dependent respiration. We believe this new technique will be particularly useful for drug screening and for generating previously unobtainable respiratory data on small mitochondrial samples.

  5. CrossCheck: an open-source web tool for high-throughput screen data analysis.

    Science.gov (United States)

    Najafov, Jamil; Najafov, Ayaz

    2017-07-19

    Modern high-throughput screening methods allow researchers to generate large datasets that potentially contain important biological information. However, oftentimes, picking relevant hits from such screens and generating testable hypotheses requires training in bioinformatics and the skills to efficiently perform database mining. There are currently no tools available to general public that allow users to cross-reference their screen datasets with published screen datasets. To this end, we developed CrossCheck, an online platform for high-throughput screen data analysis. CrossCheck is a centralized database that allows effortless comparison of the user-entered list of gene symbols with 16,231 published datasets. These datasets include published data from genome-wide RNAi and CRISPR screens, interactome proteomics and phosphoproteomics screens, cancer mutation databases, low-throughput studies of major cell signaling mediators, such as kinases, E3 ubiquitin ligases and phosphatases, and gene ontological information. Moreover, CrossCheck includes a novel database of predicted protein kinase substrates, which was developed using proteome-wide consensus motif searches. CrossCheck dramatically simplifies high-throughput screen data analysis and enables researchers to dig deep into the published literature and streamline data-driven hypothesis generation. CrossCheck is freely accessible as a web-based application at http://proteinguru.com/crosscheck.

  6. Simultaneous measurements of auto-immune and infectious disease specific antibodies using a high throughput multiplexing tool.

    Directory of Open Access Journals (Sweden)

    Atul Asati

    Full Text Available Considering importance of ganglioside antibodies as biomarkers in various immune-mediated neuropathies and neurological disorders, we developed a high throughput multiplexing tool for the assessment of gangliosides-specific antibodies based on Biolpex/Luminex platform. In this report, we demonstrate that the ganglioside high throughput multiplexing tool is robust, highly specific and demonstrating ∼100-fold higher concentration sensitivity for IgG detection than ELISA. In addition to the ganglioside-coated array, the high throughput multiplexing tool contains beads coated with influenza hemagglutinins derived from H1N1 A/Brisbane/59/07 and H1N1 A/California/07/09 strains. Influenza beads provided an added advantage of simultaneous detection of ganglioside- and influenza-specific antibodies, a capacity important for the assay of both infectious antigen-specific and autoimmune antibodies following vaccination or disease. Taken together, these results support the potential adoption of the ganglioside high throughput multiplexing tool for measuring ganglioside antibodies in various neuropathic and neurological disorders.

  7. Simulation Modeling to Compare High-Throughput, Low-Iteration Optimization Strategies for Metabolic Engineering.

    Science.gov (United States)

    Heinsch, Stephen C; Das, Siba R; Smanski, Michael J

    2018-01-01

    Increasing the final titer of a multi-gene metabolic pathway can be viewed as a multivariate optimization problem. While numerous multivariate optimization algorithms exist, few are specifically designed to accommodate the constraints posed by genetic engineering workflows. We present a strategy for optimizing expression levels across an arbitrary number of genes that requires few design-build-test iterations. We compare the performance of several optimization algorithms on a series of simulated expression landscapes. We show that optimal experimental design parameters depend on the degree of landscape ruggedness. This work provides a theoretical framework for designing and executing numerical optimization on multi-gene systems.

  8. Modular high-throughput test stand for versatile screening of thin-film materials libraries

    International Nuclear Information System (INIS)

    Thienhaus, Sigurd; Hamann, Sven; Ludwig, Alfred

    2011-01-01

    Versatile high-throughput characterization tools are required for the development of new materials using combinatorial techniques. Here, we describe a modular, high-throughput test stand for the screening of thin-film materials libraries, which can carry out automated electrical, magnetic and magnetoresistance measurements in the temperature range of −40 to 300 °C. As a proof of concept, we measured the temperature-dependent resistance of Fe–Pd–Mn ferromagnetic shape-memory alloy materials libraries, revealing reversible martensitic transformations and the associated transformation temperatures. Magneto-optical screening measurements of a materials library identify ferromagnetic samples, whereas resistivity maps support the discovery of new phases. A distance sensor in the same setup allows stress measurements in materials libraries deposited on cantilever arrays. A combination of these methods offers a fast and reliable high-throughput characterization technology for searching for new materials. Using this approach, a composition region has been identified in the Fe–Pd–Mn system that combines ferromagnetism and martensitic transformation.

  9. Spectrophotometric Enzyme Assays for High-Throughput Screening

    Directory of Open Access Journals (Sweden)

    Jean-Louis Reymond

    2004-01-01

    Full Text Available This paper reviews high-throughput screening enzyme assays developed in our laboratory over the last ten years. These enzyme assays were initially developed for the purpose of discovering catalytic antibodies by screening cell culture supernatants, but have proved generally useful for testing enzyme activities. Examples include TLC-based screening using acridone-labeled substrates, fluorogenic assays based on the β-elimination of umbelliferone or nitrophenol, and indirect assays such as the back-titration method with adrenaline and the copper-calcein fluorescence assay for aminoacids.

  10. HT-COMET: a novel automated approach for high throughput assessment of human sperm chromatin quality

    Science.gov (United States)

    Albert, Océane; Reintsch, Wolfgang E.; Chan, Peter; Robaire, Bernard

    2016-01-01

    STUDY QUESTION Can we make the comet assay (single-cell gel electrophoresis) for human sperm a more accurate and informative high throughput assay? SUMMARY ANSWER We developed a standardized automated high throughput comet (HT-COMET) assay for human sperm that improves its accuracy and efficiency, and could be of prognostic value to patients in the fertility clinic. WHAT IS KNOWN ALREADY The comet assay involves the collection of data on sperm DNA damage at the level of the single cell, allowing the use of samples from severe oligozoospermic patients. However, this makes comet scoring a low throughput procedure that renders large cohort analyses tedious. Furthermore, the comet assay comes with an inherent vulnerability to variability. Our objective is to develop an automated high throughput comet assay for human sperm that will increase both its accuracy and efficiency. STUDY DESIGN, SIZE, DURATION The study comprised two distinct components: a HT-COMET technical optimization section based on control versus DNAse treatment analyses (n = 3–5), and a cross-sectional study on 123 men presenting to a reproductive center with sperm concentrations categorized as severe oligozoospermia, oligozoospermia or normozoospermia. PARTICIPANTS/MATERIALS, SETTING, METHODS Sperm chromatin quality was measured using the comet assay: on classic 2-well slides for software comparison; on 96-well slides for HT-COMET optimization; after exposure to various concentrations of a damage-inducing agent, DNAse, using HT-COMET; on 123 subjects with different sperm concentrations using HT-COMET. Data from the 123 subjects were correlated to classic semen quality parameters and plotted as single-cell data in individual DNA damage profiles. MAIN RESULTS AND THE ROLE OF CHANCE We have developed a standard automated HT-COMET procedure for human sperm. It includes automated scoring of comets by a fully integrated high content screening setup that compares well with the most commonly used semi

  11. High-resolution and high-throughput multichannel Fourier transform spectrometer with two-dimensional interferogram warping compensation

    Science.gov (United States)

    Watanabe, A.; Furukawa, H.

    2018-04-01

    The resolution of multichannel Fourier transform (McFT) spectroscopy is insufficient for many applications despite its extreme advantage of high throughput. We propose an improved configuration to realise both performance using a two-dimensional area sensor. For the spectral resolution, we obtained the interferogram of a larger optical path difference by shifting the area sensor without altering any optical components. The non-linear phase error of the interferometer was successfully corrected using a phase-compensation calculation. Warping compensation was also applied to realise a higher throughput to accumulate the signal between vertical pixels. Our approach significantly improved the resolution and signal-to-noise ratio by factors of 1.7 and 34, respectively. This high-resolution and high-sensitivity McFT spectrometer will be useful for detecting weak light signals such as those in non-invasive diagnosis.

  12. Computational tools for high-throughput discovery in biology

    OpenAIRE

    Jones, Neil Christopher

    2007-01-01

    High throughput data acquisition technology has inarguably transformed the landscape of the life sciences, in part by making possible---and necessary---the computational disciplines of bioinformatics and biomedical informatics. These fields focus primarily on developing tools for analyzing data and generating hypotheses about objects in nature, and it is in this context that we address three pressing problems in the fields of the computational life sciences which each require computing capaci...

  13. Some Aspects of Crystal Centering During X-ray High-throughput Protein Crystallography Experiment

    Science.gov (United States)

    Gaponov, Yu. A.; Matsugaki, N.; Sasajima, K.; Igarashi, N.; Wakatsuki, S.

    A set of algorithms and procedures of a crystal loop centering during X-ray high-throughput protein crystallography experiment has been designed and developed. A simple algorithm of the crystal loop detection and preliminary recognition has been designed and developed. The crystal loop detection algorithm is based on finding out the crystal loop ending point (opposite to the crystal loop pin) using image cross section (digital image column) profile analysis. The crystal loop preliminary recognition procedure is based on finding out the crystal loop sizes and position using image cross section profile analysis. The crystal loop fine recognition procedure based on Hooke-Jeeves pattern search method with an ellipse as a fitting pattern has been designed and developed. The procedure of restoring missing coordinate of the crystal loop is described. Based on developed algorithms and procedures the optimal auto-centering procedure has been designed and developed. A procedure of optimal manual crystal centering (Two Clicks Procedure) has been designed and developed. Developed procedures have been integrated into control software system PCCS installed at crystallography beamlines Photon Factory BL5A and PF-AR NW12, KEK.

  14. A pocket device for high-throughput optofluidic holographic microscopy

    Science.gov (United States)

    Mandracchia, B.; Bianco, V.; Wang, Z.; Paturzo, M.; Bramanti, A.; Pioggia, G.; Ferraro, P.

    2017-06-01

    Here we introduce a compact holographic microscope embedded onboard a Lab-on-a-Chip (LoC) platform. A wavefront division interferometer is realized by writing a polymer grating onto the channel to extract a reference wave from the object wave impinging the LoC. A portion of the beam reaches the samples flowing along the channel path, carrying their information content to the recording device, while one of the diffraction orders from the grating acts as an off-axis reference wave. Polymeric micro-lenses are delivered forward the chip by Pyro-ElectroHydroDynamic (Pyro-EHD) inkjet printing techniques. Thus, all the required optical components are embedded onboard a pocket device, and fast, non-iterative, reconstruction algorithms can be used. We use our device in combination with a novel high-throughput technique, named Space-Time Digital Holography (STDH). STDH exploits the samples motion inside microfluidic channels to obtain a synthetic hologram, mapped in a hybrid space-time domain, and with intrinsic useful features. Indeed, a single Linear Sensor Array (LSA) is sufficient to build up a synthetic representation of the entire experiment (i.e. the STDH) with unlimited Field of View (FoV) along the scanning direction, independently from the magnification factor. The throughput of the imaging system is dramatically increased as STDH provides unlimited FoV, refocusable imaging of samples inside the liquid volume with no need for hologram stitching. To test our embedded STDH microscopy module, we counted, imaged and tracked in 3D with high-throughput red blood cells moving inside the channel volume under non ideal flow conditions.

  15. A comparison of high-throughput techniques for assaying circadian rhythms in plants.

    Science.gov (United States)

    Tindall, Andrew J; Waller, Jade; Greenwood, Mark; Gould, Peter D; Hartwell, James; Hall, Anthony

    2015-01-01

    Over the last two decades, the development of high-throughput techniques has enabled us to probe the plant circadian clock, a key coordinator of vital biological processes, in ways previously impossible. With the circadian clock increasingly implicated in key fitness and signalling pathways, this has opened up new avenues for understanding plant development and signalling. Our tool-kit has been constantly improving through continual development and novel techniques that increase throughput, reduce costs and allow higher resolution on the cellular and subcellular levels. With circadian assays becoming more accessible and relevant than ever to researchers, in this paper we offer a review of the techniques currently available before considering the horizons in circadian investigation at ever higher throughputs and resolutions.

  16. High-Throughput Tabular Data Processor - Platform independent graphical tool for processing large data sets.

    Science.gov (United States)

    Madanecki, Piotr; Bałut, Magdalena; Buckley, Patrick G; Ochocka, J Renata; Bartoszewski, Rafał; Crossman, David K; Messiaen, Ludwine M; Piotrowski, Arkadiusz

    2018-01-01

    High-throughput technologies generate considerable amount of data which often requires bioinformatic expertise to analyze. Here we present High-Throughput Tabular Data Processor (HTDP), a platform independent Java program. HTDP works on any character-delimited column data (e.g. BED, GFF, GTF, PSL, WIG, VCF) from multiple text files and supports merging, filtering and converting of data that is produced in the course of high-throughput experiments. HTDP can also utilize itemized sets of conditions from external files for complex or repetitive filtering/merging tasks. The program is intended to aid global, real-time processing of large data sets using a graphical user interface (GUI). Therefore, no prior expertise in programming, regular expression, or command line usage is required of the user. Additionally, no a priori assumptions are imposed on the internal file composition. We demonstrate the flexibility and potential of HTDP in real-life research tasks including microarray and massively parallel sequencing, i.e. identification of disease predisposing variants in the next generation sequencing data as well as comprehensive concurrent analysis of microarray and sequencing results. We also show the utility of HTDP in technical tasks including data merge, reduction and filtering with external criteria files. HTDP was developed to address functionality that is missing or rudimentary in other GUI software for processing character-delimited column data from high-throughput technologies. Flexibility, in terms of input file handling, provides long term potential functionality in high-throughput analysis pipelines, as the program is not limited by the currently existing applications and data formats. HTDP is available as the Open Source software (https://github.com/pmadanecki/htdp).

  17. COMPUTER APPROACHES TO WHEAT HIGH-THROUGHPUT PHENOTYPING

    Directory of Open Access Journals (Sweden)

    Afonnikov D.

    2012-08-01

    Full Text Available The growing need for rapid and accurate approaches for large-scale assessment of phenotypic characters in plants becomes more and more obvious in the studies looking into relationships between genotype and phenotype. This need is due to the advent of high throughput methods for analysis of genomes. Nowadays, any genetic experiment involves data on thousands and dozens of thousands of plants. Traditional ways of assessing most phenotypic characteristics (those with reliance on the eye, the touch, the ruler are little effective on samples of such sizes. Modern approaches seek to take advantage of automated phenotyping, which warrants a much more rapid data acquisition, higher accuracy of the assessment of phenotypic features, measurement of new parameters of these features and exclusion of human subjectivity from the process. Additionally, automation allows measurement data to be rapidly loaded into computer databases, which reduces data processing time.In this work, we present the WheatPGE information system designed to solve the problem of integration of genotypic and phenotypic data and parameters of the environment, as well as to analyze the relationships between the genotype and phenotype in wheat. The system is used to consolidate miscellaneous data on a plant for storing and processing various morphological traits and genotypes of wheat plants as well as data on various environmental factors. The system is available at www.wheatdb.org. Its potential in genetic experiments has been demonstrated in high-throughput phenotyping of wheat leaf pubescence.

  18. High-throughput selection for cellulase catalysts using chemical complementation.

    Science.gov (United States)

    Peralta-Yahya, Pamela; Carter, Brian T; Lin, Hening; Tao, Haiyan; Cornish, Virginia W

    2008-12-24

    Efficient enzymatic hydrolysis of lignocellulosic material remains one of the major bottlenecks to cost-effective conversion of biomass to ethanol. Improvement of glycosylhydrolases, however, is limited by existing medium-throughput screening technologies. Here, we report the first high-throughput selection for cellulase catalysts. This selection was developed by adapting chemical complementation to provide a growth assay for bond cleavage reactions. First, a URA3 counter selection was adapted to link chemical dimerizer activated gene transcription to cell death. Next, the URA3 counter selection was shown to detect cellulase activity based on cleavage of a tetrasaccharide chemical dimerizer substrate and decrease in expression of the toxic URA3 reporter. Finally, the utility of the cellulase selection was assessed by isolating cellulases with improved activity from a cellulase library created by family DNA shuffling. This application provides further evidence that chemical complementation can be readily adapted to detect different enzymatic activities for important chemical transformations for which no natural selection exists. Because of the large number of enzyme variants that selections can now test as compared to existing medium-throughput screens for cellulases, this assay has the potential to impact the discovery of improved cellulases and other glycosylhydrolases for biomass conversion from libraries of cellulases created by mutagenesis or obtained from natural biodiversity.

  19. Blood group genotyping: from patient to high-throughput donor screening.

    Science.gov (United States)

    Veldhuisen, B; van der Schoot, C E; de Haas, M

    2009-10-01

    Blood group antigens, present on the cell membrane of red blood cells and platelets, can be defined either serologically or predicted based on the genotypes of genes encoding for blood group antigens. At present, the molecular basis of many antigens of the 30 blood group systems and 17 human platelet antigens is known. In many laboratories, blood group genotyping assays are routinely used for diagnostics in cases where patient red cells cannot be used for serological typing due to the presence of auto-antibodies or after recent transfusions. In addition, DNA genotyping is used to support (un)-expected serological findings. Fetal genotyping is routinely performed when there is a risk of alloimmune-mediated red cell or platelet destruction. In case of patient blood group antigen typing, it is important that a genotyping result is quickly available to support the selection of donor blood, and high-throughput of the genotyping method is not a prerequisite. In addition, genotyping of blood donors will be extremely useful to obtain donor blood with rare phenotypes, for example lacking a high-frequency antigen, and to obtain a fully typed donor database to be used for a better matching between recipient and donor to prevent adverse transfusion reactions. Serological typing of large cohorts of donors is a labour-intensive and expensive exercise and hampered by the lack of sufficient amounts of approved typing reagents for all blood group systems of interest. Currently, high-throughput genotyping based on DNA micro-arrays is a very feasible method to obtain a large pool of well-typed blood donors. Several systems for high-throughput blood group genotyping are developed and will be discussed in this review.

  20. High-throughput assessment of context-dependent effects of chromatin proteins

    NARCIS (Netherlands)

    Brueckner, L. (Laura); Van Arensbergen, J. (Joris); Akhtar, W. (Waseem); L. Pagie (Ludo); B. van Steensel (Bas)

    2016-01-01

    textabstractBackground: Chromatin proteins control gene activity in a concerted manner. We developed a high-throughput assay to study the effects of the local chromatin environment on the regulatory activity of a protein of interest. The assay combines a previously reported multiplexing strategy

  1. High-throughput open source computational methods for genetics and genomics

    NARCIS (Netherlands)

    Prins, J.C.P.

    2015-01-01

    Biology is increasingly data driven by virtue of the development of high-throughput technologies, such as DNA and RNA sequencing. Computational biology and bioinformatics are scientific disciplines that cross-over between the disciplines of biology, informatics and statistics; which is clearly

  2. tcpl: The ToxCast Pipeline for High-Throughput Screening Data

    Science.gov (United States)

    Motivation: The large and diverse high-throughput chemical screening efforts carried out by the US EPAToxCast program requires an efficient, transparent, and reproducible data pipeline.Summary: The tcpl R package and its associated MySQL database provide a generalized platform fo...

  3. Combinatorial chemoenzymatic synthesis and high-throughput screening of sialosides.

    Science.gov (United States)

    Chokhawala, Harshal A; Huang, Shengshu; Lau, Kam; Yu, Hai; Cheng, Jiansong; Thon, Vireak; Hurtado-Ziola, Nancy; Guerrero, Juan A; Varki, Ajit; Chen, Xi

    2008-09-19

    Although the vital roles of structures containing sialic acid in biomolecular recognition are well documented, limited information is available on how sialic acid structural modifications, sialyl linkages, and the underlying glycan structures affect the binding or the activity of sialic acid-recognizing proteins and related downstream biological processes. A novel combinatorial chemoenzymatic method has been developed for the highly efficient synthesis of biotinylated sialosides containing different sialic acid structures and different underlying glycans in 96-well plates from biotinylated sialyltransferase acceptors and sialic acid precursors. By transferring the reaction mixtures to NeutrAvidin-coated plates and assaying for the yields of enzymatic reactions using lectins recognizing sialyltransferase acceptors but not the sialylated products, the biotinylated sialoside products can be directly used, without purification, for high-throughput screening to quickly identify the ligand specificity of sialic acid-binding proteins. For a proof-of-principle experiment, 72 biotinylated alpha2,6-linked sialosides were synthesized in 96-well plates from 4 biotinylated sialyltransferase acceptors and 18 sialic acid precursors using a one-pot three-enzyme system. High-throughput screening assays performed in NeutrAvidin-coated microtiter plates show that whereas Sambucus nigra Lectin binds to alpha2,6-linked sialosides with high promiscuity, human Siglec-2 (CD22) is highly selective for a number of sialic acid structures and the underlying glycans in its sialoside ligands.

  4. A high-throughput surface plasmon resonance biosensor based on differential interferometric imaging

    International Nuclear Information System (INIS)

    Wang, Daqian; Ding, Lili; Zhang, Wei; Zhang, Enyao; Yu, Xinglong; Luo, Zhaofeng; Ou, Huichao

    2012-01-01

    A new high-throughput surface plasmon resonance (SPR) biosensor based on differential interferometric imaging is reported. The two SPR interferograms of the sensing surface are imaged on two CCD cameras. The phase difference between the two interferograms is 180°. The refractive index related factor (RIRF) of the sensing surface is calculated from the two simultaneously acquired interferograms. The simulation results indicate that the RIRF exhibits a linear relationship with the refractive index of the sensing surface and is unaffected by the noise, drift and intensity distribution of the light source. The affinity and kinetic information can be extracted in real time from continuously acquired RIRF distributions. The results of refractometry experiments show that the dynamic detection range of SPR differential interferometric imaging system can be over 0.015 refractive index unit (RIU). High refractive index resolution is down to 0.45 RU (1 RU = 1 × 10 −6 RIU). Imaging and protein microarray experiments demonstrate the ability of high-throughput detection. The aptamer experiments demonstrate that the SPR sensor based on differential interferometric imaging has a great capability to be implemented for high-throughput aptamer kinetic evaluation. These results suggest that this biosensor has the potential to be utilized in proteomics and drug discovery after further improvement. (paper)

  5. 40 CFR Table 3 to Subpart Eeee of... - Operating Limits-High Throughput Transfer Racks

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 12 2010-07-01 2010-07-01 true Operating Limits-High Throughput Transfer Racks 3 Table 3 to Subpart EEEE of Part 63 Protection of Environment ENVIRONMENTAL PROTECTION... Throughput Transfer Racks As stated in § 63.2346(e), you must comply with the operating limits for existing...

  6. Management of High-Throughput DNA Sequencing Projects: Alpheus.

    Science.gov (United States)

    Miller, Neil A; Kingsmore, Stephen F; Farmer, Andrew; Langley, Raymond J; Mudge, Joann; Crow, John A; Gonzalez, Alvaro J; Schilkey, Faye D; Kim, Ryan J; van Velkinburgh, Jennifer; May, Gregory D; Black, C Forrest; Myers, M Kathy; Utsey, John P; Frost, Nicholas S; Sugarbaker, David J; Bueno, Raphael; Gullans, Stephen R; Baxter, Susan M; Day, Steve W; Retzel, Ernest F

    2008-12-26

    High-throughput DNA sequencing has enabled systems biology to begin to address areas in health, agricultural and basic biological research. Concomitant with the opportunities is an absolute necessity to manage significant volumes of high-dimensional and inter-related data and analysis. Alpheus is an analysis pipeline, database and visualization software for use with massively parallel DNA sequencing technologies that feature multi-gigabase throughput characterized by relatively short reads, such as Illumina-Solexa (sequencing-by-synthesis), Roche-454 (pyrosequencing) and Applied Biosystem's SOLiD (sequencing-by-ligation). Alpheus enables alignment to reference sequence(s), detection of variants and enumeration of sequence abundance, including expression levels in transcriptome sequence. Alpheus is able to detect several types of variants, including non-synonymous and synonymous single nucleotide polymorphisms (SNPs), insertions/deletions (indels), premature stop codons, and splice isoforms. Variant detection is aided by the ability to filter variant calls based on consistency, expected allele frequency, sequence quality, coverage, and variant type in order to minimize false positives while maximizing the identification of true positives. Alpheus also enables comparisons of genes with variants between cases and controls or bulk segregant pools. Sequence-based differential expression comparisons can be developed, with data export to SAS JMP Genomics for statistical analysis.

  7. High-throughput screening with micro-x-ray fluorescence

    International Nuclear Information System (INIS)

    Havrilla, George J.; Miller, Thomasin C.

    2005-01-01

    Micro-x-ray fluorescence (MXRF) is a useful characterization tool for high-throughput screening of combinatorial libraries. Due to the increasing threat of use of chemical warfare (CW) agents both in military actions and against civilians by terrorist extremists, there is a strong push to improve existing methods and develop means for the detection of a broad spectrum of CW agents in a minimal amount of time to increase national security. This paper describes a combinatorial high-throughput screening technique for CW receptor discovery to aid in sensor development. MXRF can screen materials for elemental composition at the mesoscale level (tens to hundreds of micrometers). The key aspect of this work is the use of commercial MXRF instrumentation coupled with the inherent heteroatom elements within the target molecules of the combinatorial reaction to provide rapid and specific identification of lead species. The method is demonstrated by screening an 11-mer oligopeptide library for selective binding of the degradation products of the nerve agent VX. The identified oligopeptides can be used as selective molecular receptors for sensor development. The MXRF screening method is nondestructive, requires minimal sample preparation or special tags for analysis, and the screening time depends on the desired sensitivity

  8. Label-free cell-cycle analysis by high-throughput quantitative phase time-stretch imaging flow cytometry

    Science.gov (United States)

    Mok, Aaron T. Y.; Lee, Kelvin C. M.; Wong, Kenneth K. Y.; Tsia, Kevin K.

    2018-02-01

    Biophysical properties of cells could complement and correlate biochemical markers to characterize a multitude of cellular states. Changes in cell size, dry mass and subcellular morphology, for instance, are relevant to cell-cycle progression which is prevalently evaluated by DNA-targeted fluorescence measurements. Quantitative-phase microscopy (QPM) is among the effective biophysical phenotyping tools that can quantify cell sizes and sub-cellular dry mass density distribution of single cells at high spatial resolution. However, limited camera frame rate and thus imaging throughput makes QPM incompatible with high-throughput flow cytometry - a gold standard in multiparametric cell-based assay. Here we present a high-throughput approach for label-free analysis of cell cycle based on quantitative-phase time-stretch imaging flow cytometry at a throughput of > 10,000 cells/s. Our time-stretch QPM system enables sub-cellular resolution even at high speed, allowing us to extract a multitude (at least 24) of single-cell biophysical phenotypes (from both amplitude and phase images). Those phenotypes can be combined to track cell-cycle progression based on a t-distributed stochastic neighbor embedding (t-SNE) algorithm. Using multivariate analysis of variance (MANOVA) discriminant analysis, cell-cycle phases can also be predicted label-free with high accuracy at >90% in G1 and G2 phase, and >80% in S phase. We anticipate that high throughput label-free cell cycle characterization could open new approaches for large-scale single-cell analysis, bringing new mechanistic insights into complex biological processes including diseases pathogenesis.

  9. High Throughput Plasma Water Treatment

    Science.gov (United States)

    Mujovic, Selman; Foster, John

    2016-10-01

    The troublesome emergence of new classes of micro-pollutants, such as pharmaceuticals and endocrine disruptors, poses challenges for conventional water treatment systems. In an effort to address these contaminants and to support water reuse in drought stricken regions, new technologies must be introduced. The interaction of water with plasma rapidly mineralizes organics by inducing advanced oxidation in addition to other chemical, physical and radiative processes. The primary barrier to the implementation of plasma-based water treatment is process volume scale up. In this work, we investigate a potentially scalable, high throughput plasma water reactor that utilizes a packed bed dielectric barrier-like geometry to maximize the plasma-water interface. Here, the water serves as the dielectric medium. High-speed imaging and emission spectroscopy are used to characterize the reactor discharges. Changes in methylene blue concentration and basic water parameters are mapped as a function of plasma treatment time. Experimental results are compared to electrostatic and plasma chemistry computations, which will provide insight into the reactor's operation so that efficiency can be assessed. Supported by NSF (CBET 1336375).

  10. Virtual high screening throughput and design of 14α-lanosterol ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-07-06

    Jul 6, 2009 ... Virtual high screening throughput and design of. 14α-lanosterol demethylase inhibitors against. Mycobacterium tuberculosis. Hildebert B. Maurice1*, Esther Tuarira1 and Kennedy Mwambete2. 1School of Pharmaceutical Sciences, Institute of Allied Health Sciences, Muhimbili University of Health and.

  11. PUFKEY: A High-Security and High-Throughput Hardware True Random Number Generator for Sensor Networks

    Directory of Open Access Journals (Sweden)

    Dongfang Li

    2015-10-01

    Full Text Available Random number generators (RNG play an important role in many sensor network systems and applications, such as those requiring secure and robust communications. In this paper, we develop a high-security and high-throughput hardware true random number generator, called PUFKEY, which consists of two kinds of physical unclonable function (PUF elements. Combined with a conditioning algorithm, true random seeds are extracted from the noise on the start-up pattern of SRAM memories. These true random seeds contain full entropy. Then, the true random seeds are used as the input for a non-deterministic hardware RNG to generate a stream of true random bits with a throughput as high as 803 Mbps. The experimental results show that the bitstream generated by the proposed PUFKEY can pass all standard national institute of standards and technology (NIST randomness tests and is resilient to a wide range of security attacks.

  12. PUFKEY: a high-security and high-throughput hardware true random number generator for sensor networks.

    Science.gov (United States)

    Li, Dongfang; Lu, Zhaojun; Zou, Xuecheng; Liu, Zhenglin

    2015-10-16

    Random number generators (RNG) play an important role in many sensor network systems and applications, such as those requiring secure and robust communications. In this paper, we develop a high-security and high-throughput hardware true random number generator, called PUFKEY, which consists of two kinds of physical unclonable function (PUF) elements. Combined with a conditioning algorithm, true random seeds are extracted from the noise on the start-up pattern of SRAM memories. These true random seeds contain full entropy. Then, the true random seeds are used as the input for a non-deterministic hardware RNG to generate a stream of true random bits with a throughput as high as 803 Mbps. The experimental results show that the bitstream generated by the proposed PUFKEY can pass all standard national institute of standards and technology (NIST) randomness tests and is resilient to a wide range of security attacks.

  13. Optimal Thawing of Cryopreserved Peripheral Blood Mononuclear Cells for Use in High-Throughput Human Immune Monitoring Studies

    Directory of Open Access Journals (Sweden)

    Ramu A. Subbramanian

    2012-07-01

    Full Text Available Cryopreserved peripheral blood mononuclear cells (PBMC constitute an important component of immune monitoring studies as they allow for efficient batch- testing of samples as well as for the validation and extension of original studies in the future. In this study, we systematically test the permutations of PBMC thawing practices commonly employed in the field and identify conditions that are high and low risk for the viability of PBMC and their functionality in downstream ELISPOT assays. The study identifies the addition of ice-chilled washing media to thawed cells at the same temperature as being a high risk practice, as it yields significantly lower viability and functionality of recovered PBMC when compared to warming the cryovials to 37 °C and adding a warm washing medium. We found thawed PBMC in cryovials could be kept up to 30 minutes at 37 °C in the presence of DMSO before commencement of washing, which surprisingly identifies exposure to DMSO as a low risk step during the thawing process. This latter finding is of considerable practical relevance since it permits batch-thawing of PBMC in high-throughput immune monitoring environments.

  14. Micellar Surfactant Association in the Presence of a Glucoside-based Amphiphile Detected via High-Throughput Small Angle X-ray Scattering

    Energy Technology Data Exchange (ETDEWEB)

    Stanic, Vesna [Brazilian Synchrotron Light Source, Campinas (Brazil); Broadbent, Charlotte [Columbia Univ., New York, NY (United States). Engineering Dept.; DiMasi, Elaine [Brookhaven National Lab. (BNL), Upton, NY (United States). Photon Sciences Division; Galleguillos, Ramiro [Lubrizol Advanced Materials, Cleveland, OH (United States); Woodward, Valerie [Lubrizol Advanced Materials, Cleveland, OH (United States)

    2016-11-14

    The interactions of mixtures of anionic and amphoteric surfactants with sugar amphiphiles were studied via high throughput small angle x-ray scattering (SAXS). The sugar amphiphile was composed of Caprate, Caprylate, and Oleate mixed ester of methyl glucoside, MeGCCO. Optimal surfactant interactions are sought which have desirable physical properties, which must be identified in a cost effective manner that can access the large phase space of possible molecular combinations. X-ray scattering patterns obtained via high throughput SAXS can probe a combinatorial sample space and reveal the incorporation of MeGCCO into the micelles and the molecular associations between surfactant molecules. Such data make it possible to efficiently assess the effects of the new amphiphiles in the formulation. A specific finding of this study is that formulations containing comparatively monodisperse and homogeneous surfactant mixtures can be reliably tuned by addition of NaCl, which swells the surfactant micelles with a monotonic dependence on salt concentration. In contrast, the presence of multiple different surfactants destroys clear correlations with NaCl concentration, even in otherwise similar series of formulations.

  15. Development of a high throughput single-particle screening for inorganic semiconductor nanorods as neural voltage sensor

    Science.gov (United States)

    Kuo, Yung; Park, Kyoungwon; Li, Jack; Ingargiola, Antonino; Park, Joonhyuck; Shvadchak, Volodymyr; Weiss, Shimon

    2017-08-01

    Monitoring membrane potential in neurons requires sensors with minimal invasiveness, high spatial and temporal (sub-ms) resolution, and large sensitivity for enabling detection of sub-threshold activities. While organic dyes and fluorescent proteins have been developed to possess voltage-sensing properties, photobleaching, cytotoxicity, low sensitivity, and low spatial resolution have obstructed further studies. Semiconductor nanoparticles (NPs), as prospective voltage sensors, have shown excellent sensitivity based on Quantum confined Stark effect (QCSE) at room temperature and at single particle level. Both theory and experiment have shown their voltage sensitivity can be increased significantly via material, bandgap, and structural engineering. Based on theoretical calculations, we synthesized one of the optimal candidates for voltage sensors: 12 nm type-II ZnSe/CdS nanorods (NRs), with an asymmetrically located seed. The voltage sensitivity and spectral shift were characterized in vitro using spectrally-resolved microscopy using electrodes grown by thin film deposition, which "sandwich" the NRs. We characterized multiple batches of such NRs and iteratively modified the synthesis to achieve higher voltage sensitivity (ΔF/F> 10%), larger spectral shift (>5 nm), better homogeneity, and better colloidal stability. Using a high throughput screening method, we were able to compare the voltage sensitivity of our NRs with commercial spherical quantum dots (QDs) with single particle statistics. Our method of high throughput screening with spectrally-resolved microscope also provides a versatile tool for studying single particles spectroscopy under field modulation.

  16. High-throughput verification of transcriptional starting sites by Deep-RACE

    DEFF Research Database (Denmark)

    Olivarius, Signe; Plessy, Charles; Carninci, Piero

    2009-01-01

    We present a high-throughput method for investigating the transcriptional starting sites of genes of interest, which we named Deep-RACE (Deep–rapid amplification of cDNA ends). Taking advantage of the latest sequencing technology, it allows the parallel analysis of multiple genes and is free...

  17. High-throughput microfluidics automated cytogenetic processing for effectively lowering biological process time and aid triage during radiation accidents

    International Nuclear Information System (INIS)

    Ramakumar, Adarsh

    2016-01-01

    Nuclear or radiation mass casualties require individual, rapid, and accurate dose-based triage of exposed subjects for cytokine therapy and supportive care, to save life. Radiation mass casualties will demand high-throughput individual diagnostic dose assessment for medical management of exposed subjects. Cytogenetic techniques are widely used for triage and definitive radiation biodosimetry. Prototype platform to demonstrate high-throughput microfluidic micro incubation to support the logistics of sample in miniaturized incubators from the site of accident to analytical labs has been developed. Efforts have been made, both at the level of developing concepts and advanced system for higher throughput in processing the samples and also implementing better and efficient methods of logistics leading to performance of lab-on-chip analyses. Automated high-throughput platform with automated feature extraction, storage, cross platform data linkage, cross platform validation and inclusion of multi-parametric biomarker approaches will provide the first generation high-throughput platform systems for effective medical management, particularly during radiation mass casualty events

  18. A High-Throughput Biological Calorimetry Core: Steps to Startup, Run, and Maintain a Multiuser Facility.

    Science.gov (United States)

    Yennawar, Neela H; Fecko, Julia A; Showalter, Scott A; Bevilacqua, Philip C

    2016-01-01

    Many labs have conventional calorimeters where denaturation and binding experiments are setup and run one at a time. While these systems are highly informative to biopolymer folding and ligand interaction, they require considerable manual intervention for cleaning and setup. As such, the throughput for such setups is limited typically to a few runs a day. With a large number of experimental parameters to explore including different buffers, macromolecule concentrations, temperatures, ligands, mutants, controls, replicates, and instrument tests, the need for high-throughput automated calorimeters is on the rise. Lower sample volume requirements and reduced user intervention time compared to the manual instruments have improved turnover of calorimetry experiments in a high-throughput format where 25 or more runs can be conducted per day. The cost and efforts to maintain high-throughput equipment typically demands that these instruments be housed in a multiuser core facility. We describe here the steps taken to successfully start and run an automated biological calorimetry facility at Pennsylvania State University. Scientists from various departments at Penn State including Chemistry, Biochemistry and Molecular Biology, Bioengineering, Biology, Food Science, and Chemical Engineering are benefiting from this core facility. Samples studied include proteins, nucleic acids, sugars, lipids, synthetic polymers, small molecules, natural products, and virus capsids. This facility has led to higher throughput of data, which has been leveraged into grant support, attracting new faculty hire and has led to some exciting publications. © 2016 Elsevier Inc. All rights reserved.

  19. Crystal Symmetry Algorithms in a High-Throughput Framework for Materials

    Science.gov (United States)

    Taylor, Richard

    The high-throughput framework AFLOW that has been developed and used successfully over the last decade is improved to include fully-integrated software for crystallographic symmetry characterization. The standards used in the symmetry algorithms conform with the conventions and prescriptions given in the International Tables of Crystallography (ITC). A standard cell choice with standard origin is selected, and the space group, point group, Bravais lattice, crystal system, lattice system, and representative symmetry operations are determined. Following the conventions of the ITC, the Wyckoff sites are also determined and their labels and site symmetry are provided. The symmetry code makes no assumptions on the input cell orientation, origin, or reduction and has been integrated in the AFLOW high-throughput framework for materials discovery by adding to the existing code base and making use of existing classes and functions. The software is written in object-oriented C++ for flexibility and reuse. A performance analysis and examination of the algorithms scaling with cell size and symmetry is also reported.

  20. Development of Control Applications for High-Throughput Protein Crystallography Experiments

    International Nuclear Information System (INIS)

    Gaponov, Yurii A.; Matsugaki, Naohiro; Honda, Nobuo; Sasajima, Kumiko; Igarashi, Noriyuki; Hiraki, Masahiko; Yamada, Yusuke; Wakatsuki, Soichi

    2007-01-01

    An integrated client-server control system (PCCS) with a unified relational database (PCDB) has been developed for high-throughput protein crystallography experiments on synchrotron beamlines. The major steps in protein crystallographic experiments (purification, crystallization, crystal harvesting, data collection, and data processing) are integrated into the software. All information necessary for performing protein crystallography experiments is stored in the PCDB database (except raw X-ray diffraction data, which is stored in the Network File Server). To allow all members of a protein crystallography group to participate in experiments, the system was developed as a multi-user system with secure network access based on TCP/IP secure UNIX sockets. Secure remote access to the system is possible from any operating system with X-terminal and SSH/X11 (Secure Shell with graphical user interface) support. Currently, the system covers the high-throughput X-ray data collection stages and is being commissioned at BL5A and NW12A (PF, PF-AR, KEK, Tsukuba, Japan)

  1. Target-dependent enrichment of virions determines the reduction of high-throughput sequencing in virus discovery.

    Directory of Open Access Journals (Sweden)

    Randi Holm Jensen

    Full Text Available Viral infections cause many different diseases stemming both from well-characterized viral pathogens but also from emerging viruses, and the search for novel viruses continues to be of great importance. High-throughput sequencing is an important technology for this purpose. However, viral nucleic acids often constitute a minute proportion of the total genetic material in a sample from infected tissue. Techniques to enrich viral targets in high-throughput sequencing have been reported, but the sensitivity of such methods is not well established. This study compares different library preparation techniques targeting both DNA and RNA with and without virion enrichment. By optimizing the selection of intact virus particles, both by physical and enzymatic approaches, we assessed the effectiveness of the specific enrichment of viral sequences as compared to non-enriched sample preparations by selectively looking for and counting read sequences obtained from shotgun sequencing. Using shotgun sequencing of total DNA or RNA, viral targets were detected at concentrations corresponding to the predicted level, providing a foundation for estimating the effectiveness of virion enrichment. Virion enrichment typically produced a 1000-fold increase in the proportion of DNA virus sequences. For RNA virions the gain was less pronounced with a maximum 13-fold increase. This enrichment varied between the different sample concentrations, with no clear trend. Despite that less sequencing was required to identify target sequences, it was not evident from our data that a lower detection level was achieved by virion enrichment compared to shotgun sequencing.

  2. Fluorescence-based high-throughput screening of dicer cleavage activity

    Czech Academy of Sciences Publication Activity Database

    Podolská, Kateřina; Sedlák, David; Bartůněk, Petr; Svoboda, Petr

    2014-01-01

    Roč. 19, č. 3 (2014), s. 417-426 ISSN 1087-0571 R&D Projects: GA ČR GA13-29531S; GA MŠk(CZ) LC06077; GA MŠk LM2011022 Grant - others:EMBO(DE) 1483 Institutional support: RVO:68378050 Keywords : Dicer * siRNA * high-throughput screening Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.423, year: 2014

  3. Optimization of wide-area ATM and local-area ethernet/FDDI network configurations for high-speed telemedicine communications employing NASA's ACTS.

    Science.gov (United States)

    McDermott, W R; Tri, J L; Mitchell, M P; Levens, S P; Wondrow, M A; Huie, L M; Khandheria, B K; Gilbert, B K

    1999-01-01

    A high data rate terrestrial and satellite network was implemented to transfer medical images and data. This article describes the a optimization of the workstations and switching equipment incorporated into the network. Topics discussed in this article include tuning of the network software, the configuration of the Sun Microsystems workstations, the FORE Systems asynchronous transfer mode switches, as well as the throughput results of two telemedicine experiments undertaken by Mayo's physician staff. The technical staff was successful in achieving the data throughput needed by the telemedicine software; particularly important was the proper determination of peak throughput and TCP window sizes to ensure optimum use of the resources available on the Sun Microsystems and Hewlett Packard workstations.

  4. High throughput electrophysiology: new perspectives for ion channel drug discovery

    DEFF Research Database (Denmark)

    Willumsen, Niels J; Bech, Morten; Olesen, Søren-Peter

    2003-01-01

    . A cornerstone in current drug discovery is high throughput screening assays which allow examination of the activity of specific ion channels though only to a limited extent. Conventional patch clamp remains the sole technique with sufficiently high time resolution and sensitivity required for precise and direct....... The introduction of new powerful HTS electrophysiological techniques is predicted to cause a revolution in ion channel drug discovery....

  5. High throughput diffractive multi-beam femtosecond laser processing using a spatial light modulator

    Energy Technology Data Exchange (ETDEWEB)

    Kuang Zheng [Laser Group, Department of Engineering, University of Liverpool Brownlow Street, Liverpool L69 3GQ (United Kingdom)], E-mail: z.kuang@liv.ac.uk; Perrie, Walter [Laser Group, Department of Engineering, University of Liverpool Brownlow Street, Liverpool L69 3GQ (United Kingdom); Leach, Jonathan [Department of Physics and Astronomy, University of Glasgow, Glasgow G12 8QQ (United Kingdom); Sharp, Martin; Edwardson, Stuart P. [Laser Group, Department of Engineering, University of Liverpool Brownlow Street, Liverpool L69 3GQ (United Kingdom); Padgett, Miles [Department of Physics and Astronomy, University of Glasgow, Glasgow G12 8QQ (United Kingdom); Dearden, Geoff; Watkins, Ken G. [Laser Group, Department of Engineering, University of Liverpool Brownlow Street, Liverpool L69 3GQ (United Kingdom)

    2008-12-30

    High throughput femtosecond laser processing is demonstrated by creating multiple beams using a spatial light modulator (SLM). The diffractive multi-beam patterns are modulated in real time by computer generated holograms (CGHs), which can be calculated by appropriate algorithms. An interactive LabVIEW program is adopted to generate the relevant CGHs. Optical efficiency at this stage is shown to be {approx}50% into first order beams and real time processing has been carried out at 50 Hz refresh rate. Results obtained demonstrate high precision surface micro-structuring on silicon and Ti6Al4V with throughput gain >1 order of magnitude.

  6. In-Field High-Throughput Phenotyping of Cotton Plant Height Using LiDAR

    Directory of Open Access Journals (Sweden)

    Shangpeng Sun

    2017-04-01

    Full Text Available A LiDAR-based high-throughput phenotyping (HTP system was developed for cotton plant phenotyping in the field. The HTP system consists of a 2D LiDAR and an RTK-GPS mounted on a high clearance tractor. The LiDAR scanned three rows of cotton plots simultaneously from the top and the RTK-GPS was used to provide the spatial coordinates of the point cloud during data collection. Configuration parameters of the system were optimized to ensure the best data quality. A height profile for each plot was extracted from the dense three dimensional point clouds; then the maximum height and height distribution of each plot were derived. In lab tests, single plants were scanned by LiDAR using 0.5° angular resolution and results showed an R2 value of 1.00 (RMSE = 3.46 mm in comparison to manual measurements. In field tests using the same angular resolution; the LiDAR-based HTP system achieved average R2 values of 0.98 (RMSE = 65 mm for cotton plot height estimation; compared to manual measurements. This HTP system is particularly useful for large field application because it provides highly accurate measurements; and the efficiency is greatly improved compared to similar studies using the side view scan.

  7. Modeling Disordered Materials with a High Throughput ab-initio Approach

    Science.gov (United States)

    2015-11-13

    Modeling Disordered Materials with a High Throughput ab - initio Approach Kesong Yang,1 Corey Oses,2 and Stefano Curtarolo3, 4 1Department of...J. Furthmüller, Efficient iterative schemes for ab initio total-energy calculations using a plane-wave basis set, Phys. Rev. B 54, 11169–11186 (1996

  8. A high-throughput pipeline for the design of real-time PCR signatures

    Directory of Open Access Journals (Sweden)

    Reifman Jaques

    2010-06-01

    Full Text Available Abstract Background Pathogen diagnostic assays based on polymerase chain reaction (PCR technology provide high sensitivity and specificity. However, the design of these diagnostic assays is computationally intensive, requiring high-throughput methods to identify unique PCR signatures in the presence of an ever increasing availability of sequenced genomes. Results We present the Tool for PCR Signature Identification (TOPSI, a high-performance computing pipeline for the design of PCR-based pathogen diagnostic assays. The TOPSI pipeline efficiently designs PCR signatures common to multiple bacterial genomes by obtaining the shared regions through pairwise alignments between the input genomes. TOPSI successfully designed PCR signatures common to 18 Staphylococcus aureus genomes in less than 14 hours using 98 cores on a high-performance computing system. Conclusions TOPSI is a computationally efficient, fully integrated tool for high-throughput design of PCR signatures common to multiple bacterial genomes. TOPSI is freely available for download at http://www.bhsai.org/downloads/topsi.tar.gz.

  9. High-throughput, temperature-controlled microchannel acoustophoresis device made with rapid prototyping

    DEFF Research Database (Denmark)

    Adams, Jonathan D; Ebbesen, Christian L.; Barnkob, Rune

    2012-01-01

    -slide format using low-cost, rapid-prototyping techniques. This high-throughput acoustophoresis chip (HTAC) utilizes a temperature-stabilized, standing ultrasonic wave, which imposes differential acoustic radiation forces that can separate particles according to size, density and compressibility. The device...

  10. A Functional High-Throughput Assay of Myelination in Vitro

    Science.gov (United States)

    2014-07-01

    Human induced pluripotent stem cells, hydrogels, 3D culture, electrophysiology, high-throughput assay 16. SECURITY CLASSIFICATION OF: 17...image the 3D rat dorsal root ganglion ( DRG ) cultures with sufficiently low background as to detect electrically-evoked depolarization events, as...of voltage-sensitive dyes. 8    We have made substantial progress in Task 4.1. We have fabricated neural fiber tracts from DRG explants and

  11. High-throughput anisotropic plasma etching of polyimide for MEMS

    International Nuclear Information System (INIS)

    Bliznetsov, Vladimir; Manickam, Anbumalar; Ranganathan, Nagarajan; Chen, Junwei

    2011-01-01

    This note describes a new high-throughput process of polyimide etching for the fabrication of MEMS devices with an organic sacrificial layer approach. Using dual frequency superimposed capacitively coupled plasma we achieved a vertical profile of polyimide with an etching rate as high as 3.5 µm min −1 . After the fabrication of vertical structures in a polyimide material, additional steps were performed to fabricate structural elements of MEMS by deposition of a SiO 2 layer and performing release etching of polyimide. (technical note)

  12. REDItools: high-throughput RNA editing detection made easy.

    Science.gov (United States)

    Picardi, Ernesto; Pesole, Graziano

    2013-07-15

    The reliable detection of RNA editing sites from massive sequencing data remains challenging and, although several methodologies have been proposed, no computational tools have been released to date. Here, we introduce REDItools a suite of python scripts to perform high-throughput investigation of RNA editing using next-generation sequencing data. REDItools are in python programming language and freely available at http://code.google.com/p/reditools/. ernesto.picardi@uniba.it or graziano.pesole@uniba.it Supplementary data are available at Bioinformatics online.

  13. Study on a digital pulse processing algorithm based on template-matching for high-throughput spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Wen, Xianfei; Yang, Haori

    2015-06-01

    A major challenge in utilizing spectroscopy techniques for nuclear safeguards is to perform high-resolution measurements at an ultra-high throughput rate. Traditionally, piled-up pulses are rejected to ensure good energy resolution. To improve throughput rate, high-pass filters are normally implemented to shorten pulses. However, this reduces signal-to-noise ratio and causes degradation in energy resolution. In this work, a pulse pile-up recovery algorithm based on template-matching was proved to be an effective approach to achieve high-throughput gamma ray spectroscopy. First, a discussion of the algorithm was given in detail. Second, the algorithm was then successfully utilized to process simulated piled-up pulses from a scintillator detector. Third, the algorithm was implemented to analyze high rate data from a NaI detector, a silicon drift detector and a HPGe detector. The promising results demonstrated the capability of this algorithm to achieve high-throughput rate without significant sacrifice in energy resolution. The performance of the template-matching algorithm was also compared with traditional shaping methods. - Highlights: • A detailed discussion on the template-matching algorithm was given. • The algorithm was tested on data from a NaI and a Si detector. • The algorithm was successfully implemented on high rate data from a HPGe detector. • The performance of the algorithm was compared with traditional shaping methods. • The advantage of the algorithm in active interrogation was discussed.

  14. Caveats and limitations of plate reader-based high-throughput kinetic measurements of intracellular calcium levels

    International Nuclear Information System (INIS)

    Heusinkveld, Harm J.; Westerink, Remco H.S.

    2011-01-01

    Calcium plays a crucial role in virtually all cellular processes, including neurotransmission. The intracellular Ca 2+ concentration ([Ca 2+ ] i ) is therefore an important readout in neurotoxicological and neuropharmacological studies. Consequently, there is an increasing demand for high-throughput measurements of [Ca 2+ ] i , e.g. using multi-well microplate readers, in hazard characterization, human risk assessment and drug development. However, changes in [Ca 2+ ] i are highly dynamic, thereby creating challenges for high-throughput measurements. Nonetheless, several protocols are now available for real-time kinetic measurement of [Ca 2+ ] i in plate reader systems, though the results of such plate reader-based measurements have been questioned. In view of the increasing use of plate reader systems for measurements of [Ca 2+ ] i a careful evaluation of current technologies is warranted. We therefore performed an extensive set of experiments, using two cell lines (PC12 and B35) and two fluorescent calcium-sensitive dyes (Fluo-4 and Fura-2), for comparison of a linear plate reader system with single cell fluorescence microscopy. Our data demonstrate that the use of plate reader systems for high-throughput real-time kinetic measurements of [Ca 2+ ] i is associated with many pitfalls and limitations, including erroneous sustained increases in fluorescence, limited sensitivity and lack of single cell resolution. Additionally, our data demonstrate that probenecid, which is often used to prevent dye leakage, effectively inhibits the depolarization-evoked increase in [Ca 2+ ] i . Overall, the data indicate that the use of current plate reader-based strategies for high-throughput real-time kinetic measurements of [Ca 2+ ] i is associated with caveats and limitations that require further investigation. - Research highlights: → The use of plate readers for high-throughput screening of intracellular Ca 2+ is associated with many pitfalls and limitations. → Single cell

  15. 3D material cytometry (3DMaC): a very high-replicate, high-throughput analytical method using microfabricated, shape-specific, cell-material niches.

    Science.gov (United States)

    Parratt, Kirsten; Jeong, Jenny; Qiu, Peng; Roy, Krishnendu

    2017-08-08

    Studying cell behavior within 3D material niches is key to understanding cell biology in health and diseases, and developing biomaterials for regenerative medicine applications. Current approaches to studying these cell-material niches have low throughput and can only analyze a few replicates per experiment resulting in reduced measurement assurance and analytical power. Here, we report 3D material cytometry (3DMaC), a novel high-throughput method based on microfabricated, shape-specific 3D cell-material niches and imaging cytometry. 3DMaC achieves rapid and highly multiplexed analyses of very high replicate numbers ("n" of 10 4 -10 6 ) of 3D biomaterial constructs. 3DMaC overcomes current limitations of low "n", low-throughput, and "noisy" assays, to provide rapid and simultaneous analyses of potentially hundreds of parameters in 3D biomaterial cultures. The method is demonstrated here for a set of 85 000 events containing twelve distinct cell-biomaterial micro-niches along with robust, customized computational methods for high-throughput analytics with potentially unprecedented statistical power.

  16. A robust robotic high-throughput antibody purification platform.

    Science.gov (United States)

    Schmidt, Peter M; Abdo, Michael; Butcher, Rebecca E; Yap, Min-Yin; Scotney, Pierre D; Ramunno, Melanie L; Martin-Roussety, Genevieve; Owczarek, Catherine; Hardy, Matthew P; Chen, Chao-Guang; Fabri, Louis J

    2016-07-15

    Monoclonal antibodies (mAbs) have become the fastest growing segment in the drug market with annual sales of more than 40 billion US$ in 2013. The selection of lead candidate molecules involves the generation of large repertoires of antibodies from which to choose a final therapeutic candidate. Improvements in the ability to rapidly produce and purify many antibodies in sufficient quantities reduces the lead time for selection which ultimately impacts on the speed with which an antibody may transition through the research stage and into product development. Miniaturization and automation of chromatography using micro columns (RoboColumns(®) from Atoll GmbH) coupled to an automated liquid handling instrument (ALH; Freedom EVO(®) from Tecan) has been a successful approach to establish high throughput process development platforms. Recent advances in transient gene expression (TGE) using the high-titre Expi293F™ system have enabled recombinant mAb titres of greater than 500mg/L. These relatively high protein titres reduce the volume required to generate several milligrams of individual antibodies for initial biochemical and biological downstream assays, making TGE in the Expi293F™ system ideally suited to high throughput chromatography on an ALH. The present publication describes a novel platform for purifying Expi293F™-expressed recombinant mAbs directly from cell-free culture supernatant on a Perkin Elmer JANUS-VariSpan ALH equipped with a plate shuttle device. The purification platform allows automated 2-step purification (Protein A-desalting/size exclusion chromatography) of several hundred mAbs per week. The new robotic method can purify mAbs with high recovery (>90%) at sub-milligram level with yields of up to 2mg from 4mL of cell-free culture supernatant. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Reverse Phase Protein Arrays for High-Throughput Protein Measurements in Mammospheres

    DEFF Research Database (Denmark)

    Pedersen, Marlene Lemvig; Block, Ines; List, Markus

    Protein Array (RPPA)-based readout format integrated into robotic siRNA screening. This technique would allow post-screening high-throughput quantification of protein changes. Recently, breast cancer stem cells (BCSCs) have attracted much attention, as a tumor- and metastasis-driving subpopulation...

  18. High Throughput WAN Data Transfer with Hadoop-based Storage

    Science.gov (United States)

    Amin, A.; Bockelman, B.; Letts, J.; Levshina, T.; Martin, T.; Pi, H.; Sfiligoi, I.; Thomas, M.; Wüerthwein, F.

    2011-12-01

    Hadoop distributed file system (HDFS) is becoming more popular in recent years as a key building block of integrated grid storage solution in the field of scientific computing. Wide Area Network (WAN) data transfer is one of the important data operations for large high energy physics experiments to manage, share and process datasets of PetaBytes scale in a highly distributed grid computing environment. In this paper, we present the experience of high throughput WAN data transfer with HDFS-based Storage Element. Two protocols, GridFTP and fast data transfer (FDT), are used to characterize the network performance of WAN data transfer.

  19. High Throughput WAN Data Transfer with Hadoop-based Storage

    International Nuclear Information System (INIS)

    Amin, A; Thomas, M; Bockelman, B; Letts, J; Martin, T; Pi, H; Sfiligoi, I; Wüerthwein, F; Levshina, T

    2011-01-01

    Hadoop distributed file system (HDFS) is becoming more popular in recent years as a key building block of integrated grid storage solution in the field of scientific computing. Wide Area Network (WAN) data transfer is one of the important data operations for large high energy physics experiments to manage, share and process datasets of PetaBytes scale in a highly distributed grid computing environment. In this paper, we present the experience of high throughput WAN data transfer with HDFS-based Storage Element. Two protocols, GridFTP and fast data transfer (FDT), are used to characterize the network performance of WAN data transfer.

  20. Development of novel, 384-well high-throughput assay panels for human drug transporters: drug interaction and safety assessment in support of discovery research.

    Science.gov (United States)

    Tang, Huaping; Shen, Ding Ren; Han, Yong-Hae; Kong, Yan; Balimane, Praveen; Marino, Anthony; Gao, Mian; Wu, Sophie; Xie, Dianlin; Soars, Matthew G; O'Connell, Jonathan C; Rodrigues, A David; Zhang, Litao; Cvijic, Mary Ellen

    2013-10-01

    Transporter proteins are known to play a critical role in affecting the overall absorption, distribution, metabolism, and excretion characteristics of drug candidates. In addition to efflux transporters (P-gp, BCRP, MRP2, etc.) that limit absorption, there has been a renewed interest in influx transporters at the renal (OATs, OCTs) and hepatic (OATPs, BSEP, NTCP, etc.) organ level that can cause significant clinical drug-drug interactions (DDIs). Several of these transporters are also critical for hepatobiliary disposition of bilirubin and bile acid/salts, and their inhibition is directly implicated in hepatic toxicities. Regulatory agencies took action to address transporter-mediated DDI with the goal of ensuring drug safety in the clinic and on the market. To meet regulatory requirements, advanced bioassay technology and automation solutions were implemented for high-throughput transporter screening to provide structure-activity relationship within lead optimization. To enhance capacity, several functional assay formats were miniaturized to 384-well throughput including novel fluorescence-based uptake and efflux inhibition assays using high-content image analysis as well as cell-based radioactive uptake and vesicle-based efflux inhibition assays. This high-throughput capability enabled a paradigm shift from studying transporter-related issues in the development space to identifying and dialing out these concerns early on in discovery for enhanced mechanism-based efficacy while circumventing DDIs and transporter toxicities.

  1. A Primer on High-Throughput Computing for Genomic Selection

    Directory of Open Access Journals (Sweden)

    Xiao-Lin eWu

    2011-02-01

    Full Text Available High-throughput computing (HTC uses computer clusters to solve advanced computational problems, with the goal of accomplishing high throughput over relatively long periods of time. In genomic selection, for example, a set of markers covering the entire genome is used to train a model based on known data, and the resulting model is used to predict the genetic merit of selection candidates. Sophisticated models are very computationally demanding and, with several traits to be evaluated sequentially, computing time is long and output is low. In this paper, we present scenarios and basic principles of how HTC can be used in genomic selection, implemented using various techniques from simple batch processing to pipelining in distributed computer clusters. Various scripting languages, such as shell scripting, Perl and R, are also very useful to devise pipelines. By pipelining, we can reduce total computing time and consequently increase throughput. In comparison to the traditional data processing pipeline residing on the central processors, performing general purpose computation on a graphics processing unit (GPU provide a new-generation approach to massive parallel computing in genomic selection. While the concept of HTC may still be new to many researchers in animal breeding, plant breeding, and genetics, HTC infrastructures have already been built in many institutions, such as the University of Wisconsin – Madison, which can be leveraged for genomic selection, in terms of central processing unit (CPU capacity, network connectivity, storage availability, and middleware connectivity. Exploring existing HTC infrastructures as well as general purpose computing environments will further expand our capability to meet increasing computing demands posed by unprecedented genomic data that we have today. We anticipate that HTC will impact genomic selection via better statistical models, faster solutions, and more competitive products (e.g., from design of

  2. Application of high-throughput sequencing in understanding human oral microbiome related with health and disease

    OpenAIRE

    Chen, Hui; Jiang, Wen

    2014-01-01

    The oral microbiome is one of most diversity habitat in the human body and they are closely related with oral health and disease. As the technique developing,, high throughput sequencing has become a popular approach applied for oral microbial analysis. Oral bacterial profiles have been studied to explore the relationship between microbial diversity and oral diseases such as caries and periodontal disease. This review describes the application of high-throughput sequencing for characterizati...

  3. PCR cycles above routine numbers do not compromise high-throughput DNA barcoding results.

    Science.gov (United States)

    Vierna, J; Doña, J; Vizcaíno, A; Serrano, D; Jovani, R

    2017-10-01

    High-throughput DNA barcoding has become essential in ecology and evolution, but some technical questions still remain. Increasing the number of PCR cycles above the routine 20-30 cycles is a common practice when working with old-type specimens, which provide little amounts of DNA, or when facing annealing issues with the primers. However, increasing the number of cycles can raise the number of artificial mutations due to polymerase errors. In this work, we sequenced 20 COI libraries in the Illumina MiSeq platform. Libraries were prepared with 40, 45, 50, 55, and 60 PCR cycles from four individuals belonging to four species of four genera of cephalopods. We found no relationship between the number of PCR cycles and the number of mutations despite using a nonproofreading polymerase. Moreover, even when using a high number of PCR cycles, the resulting number of mutations was low enough not to be an issue in the context of high-throughput DNA barcoding (but may still remain an issue in DNA metabarcoding due to chimera formation). We conclude that the common practice of increasing the number of PCR cycles should not negatively impact the outcome of a high-throughput DNA barcoding study in terms of the occurrence of point mutations.

  4. Advances in High Throughput Screening of Biomass Recalcitrance (Poster)

    Energy Technology Data Exchange (ETDEWEB)

    Turner, G. B.; Decker, S. R.; Tucker, M. P.; Law, C.; Doeppke, C.; Sykes, R. W.; Davis, M. F.; Ziebell, A.

    2012-06-01

    This was a poster displayed at the Symposium. Advances on previous high throughput screening of biomass recalcitrance methods have resulted in improved conversion and replicate precision. Changes in plate reactor metallurgy, improved preparation of control biomass, species-specific pretreatment conditions, and enzymatic hydrolysis parameters have reduced overall coefficients of variation to an average of 6% for sample replicates. These method changes have improved plate-to-plate variation of control biomass recalcitrance and improved confidence in sugar release differences between samples. With smaller errors plant researchers can have a higher degree of assurance more low recalcitrance candidates can be identified. Significant changes in plate reactor, control biomass preparation, pretreatment conditions and enzyme have significantly reduced sample and control replicate variability. Reactor plate metallurgy significantly impacts sugar release aluminum leaching into reaction during pretreatment degrades sugars and inhibits enzyme activity. Removal of starch and extractives significantly decreases control biomass variability. New enzyme formulations give more consistent and higher conversion levels, however required re-optimization for switchgrass. Pretreatment time and temperature (severity) should be adjusted to specific biomass types i.e. woody vs. herbaceous. Desalting of enzyme preps to remove low molecular weight stabilizers and improved conversion levels likely due to water activity impacts on enzyme structure and substrate interactions not attempted here due to need to continually desalt and validate precise enzyme concentration and activity.

  5. A high throughput mechanical screening device for cartilage tissue engineering.

    Science.gov (United States)

    Mohanraj, Bhavana; Hou, Chieh; Meloni, Gregory R; Cosgrove, Brian D; Dodge, George R; Mauck, Robert L

    2014-06-27

    Articular cartilage enables efficient and near-frictionless load transmission, but suffers from poor inherent healing capacity. As such, cartilage tissue engineering strategies have focused on mimicking both compositional and mechanical properties of native tissue in order to provide effective repair materials for the treatment of damaged or degenerated joint surfaces. However, given the large number design parameters available (e.g. cell sources, scaffold designs, and growth factors), it is difficult to conduct combinatorial experiments of engineered cartilage. This is particularly exacerbated when mechanical properties are a primary outcome, given the long time required for testing of individual samples. High throughput screening is utilized widely in the pharmaceutical industry to rapidly and cost-effectively assess the effects of thousands of compounds for therapeutic discovery. Here we adapted this approach to develop a high throughput mechanical screening (HTMS) system capable of measuring the mechanical properties of up to 48 materials simultaneously. The HTMS device was validated by testing various biomaterials and engineered cartilage constructs and by comparing the HTMS results to those derived from conventional single sample compression tests. Further evaluation showed that the HTMS system was capable of distinguishing and identifying 'hits', or factors that influence the degree of tissue maturation. Future iterations of this device will focus on reducing data variability, increasing force sensitivity and range, as well as scaling-up to even larger (96-well) formats. This HTMS device provides a novel tool for cartilage tissue engineering, freeing experimental design from the limitations of mechanical testing throughput. © 2013 Published by Elsevier Ltd.

  6. Recent advances in quantitative high throughput and high content data analysis.

    Science.gov (United States)

    Moutsatsos, Ioannis K; Parker, Christian N

    2016-01-01

    High throughput screening has become a basic technique with which to explore biological systems. Advances in technology, including increased screening capacity, as well as methods that generate multiparametric readouts, are driving the need for improvements in the analysis of data sets derived from such screens. This article covers the recent advances in the analysis of high throughput screening data sets from arrayed samples, as well as the recent advances in the analysis of cell-by-cell data sets derived from image or flow cytometry application. Screening multiple genomic reagents targeting any given gene creates additional challenges and so methods that prioritize individual gene targets have been developed. The article reviews many of the open source data analysis methods that are now available and which are helping to define a consensus on the best practices to use when analyzing screening data. As data sets become larger, and more complex, the need for easily accessible data analysis tools will continue to grow. The presentation of such complex data sets, to facilitate quality control monitoring and interpretation of the results will require the development of novel visualizations. In addition, advanced statistical and machine learning algorithms that can help identify patterns, correlations and the best features in massive data sets will be required. The ease of use for these tools will be important, as they will need to be used iteratively by laboratory scientists to improve the outcomes of complex analyses.

  7. A continuous high-throughput bioparticle sorter based on 3D traveling-wave dielectrophoresis.

    Science.gov (United States)

    Cheng, I-Fang; Froude, Victoria E; Zhu, Yingxi; Chang, Hsueh-Chia; Chang, Hsien-Chang

    2009-11-21

    We present a high throughput (maximum flow rate approximately 10 microl/min or linear velocity approximately 3 mm/s) continuous bio-particle sorter based on 3D traveling-wave dielectrophoresis (twDEP) at an optimum AC frequency of 500 kHz. The high throughput sorting is achieved with a sustained twDEP particle force normal to the continuous through-flow, which is applied over the entire chip by a single 3D electrode array. The design allows continuous fractionation of micron-sized particles into different downstream sub-channels based on differences in their twDEP mobility on both sides of the cross-over. Conventional DEP is integrated upstream to focus the particles into a single levitated queue to allow twDEP sorting by mobility difference and to minimize sedimentation and field-induced lysis. The 3D electrode array design minimizes the offsetting effect of nDEP (negative DEP with particle force towards regions with weak fields) on twDEP such that both forces increase monotonically with voltage to further increase the throughput. Effective focusing and separation of red blood cells from debris-filled heterogeneous samples are demonstrated, as well as size-based separation of poly-dispersed liposome suspensions into two distinct bands at 2.3 to 4.6 microm and 1.5 to 2.7 microm, at the highest throughput recorded in hand-held chips of 6 microl/min.

  8. High-throughput screening of microscale pitted substrate topographies for enhanced nonviral transfection efficiency in primary human fibroblasts

    DEFF Research Database (Denmark)

    Adler, Andrew F; Speidel, Alessondra T; Christoforou, Nicolas

    2011-01-01

    of microscale topographies, we have demonstrated an improvement in nonviral transfection efficiency for cells cultured on dense micropit patterns compared to smooth substrates, as verified with flow cytometry. A 25% increase in GFP(+) cells was observed independent of proliferation rate, accompanied by SEM....... Emerging literature has highlighted the influence of cell-topography interactions on modulation of many cell phenotypes, including protein expression and cytoskeletal behaviors implicated in endocytosis. Using high-throughput screening of primary human dermal fibroblasts cultured on a combinatorial library...... and confocal microscopy characterization to help explain the phenomenon qualitatively. This finding encourages researchers to investigate substrate topography as a new design consideration for the optimization of nonviral transfection systems....

  9. A cell-based high-throughput screening assay for radiation susceptibility using automated cell counting

    International Nuclear Information System (INIS)

    Hodzic, Jasmina; Dingjan, Ilse; Maas, Mariëlle JP; Meulen-Muileman, Ida H van der; Menezes, Renee X de; Heukelom, Stan; Verheij, Marcel; Gerritsen, Winald R; Geldof, Albert A; Triest, Baukelien van; Beusechem, Victor W van

    2015-01-01

    Radiotherapy is one of the mainstays in the treatment for cancer, but its success can be limited due to inherent or acquired resistance. Mechanisms underlying radioresistance in various cancers are poorly understood and available radiosensitizers have shown only modest clinical benefit. There is thus a need to identify new targets and drugs for more effective sensitization of cancer cells to irradiation. Compound and RNA interference high-throughput screening technologies allow comprehensive enterprises to identify new agents and targets for radiosensitization. However, the gold standard assay to investigate radiosensitivity of cancer cells in vitro, the colony formation assay (CFA), is unsuitable for high-throughput screening. We developed a new high-throughput screening method for determining radiation susceptibility. Fast and uniform irradiation of batches up to 30 microplates was achieved using a Perspex container and a clinically employed linear accelerator. The readout was done by automated counting of fluorescently stained nuclei using the Acumen eX3 laser scanning cytometer. Assay performance was compared to that of the CFA and the CellTiter-Blue homogeneous uniform-well cell viability assay. The assay was validated in a whole-genome siRNA library screening setting using PC-3 prostate cancer cells. On 4 different cancer cell lines, the automated cell counting assay produced radiation dose response curves that followed a linear-quadratic equation and that exhibited a better correlation to the results of the CFA than did the cell viability assay. Moreover, the cell counting assay could be used to detect radiosensitization by silencing DNA-PKcs or by adding caffeine. In a high-throughput screening setting, using 4 Gy irradiated and control PC-3 cells, the effects of DNA-PKcs siRNA and non-targeting control siRNA could be clearly discriminated. We developed a simple assay for radiation susceptibility that can be used for high-throughput screening. This will aid

  10. High-throughput sockets over RDMA for the Intel Xeon Phi coprocessor

    CERN Document Server

    Santogidis, Aram

    2017-01-01

    In this paper we describe the design, implementation and performance of Trans4SCIF, a user-level socket-like transport library for the Intel Xeon Phi coprocessor. Trans4SCIF library is primarily intended for high-throughput applications. It uses RDMA transfers over the native SCIF support, in a way that is transparent for the application, which has the illusion of using conventional stream sockets. We also discuss the integration of Trans4SCIF with the ZeroMQ messaging library, used extensively by several applications running at CERN. We show that this can lead to a substantial, up to 3x, increase of application throughput compared to the default TCP/IP transport option.

  11. Design and construction of a first-generation high-throughput integrated robotic molecular biology platform for bioenergy applications.

    Science.gov (United States)

    Hughes, Stephen R; Butt, Tauseef R; Bartolett, Scott; Riedmuller, Steven B; Farrelly, Philip

    2011-08-01

    The molecular biological techniques for plasmid-based assembly and cloning of gene open reading frames are essential for elucidating the function of the proteins encoded by the genes. High-throughput integrated robotic molecular biology platforms that have the capacity to rapidly clone and express heterologous gene open reading frames in bacteria and yeast and to screen large numbers of expressed proteins for optimized function are an important technology for improving microbial strains for biofuel production. The process involves the production of full-length complementary DNA libraries as a source of plasmid-based clones to express the desired proteins in active form for determination of their functions. Proteins that were identified by high-throughput screening as having desired characteristics are overexpressed in microbes to enable them to perform functions that will allow more cost-effective and sustainable production of biofuels. Because the plasmid libraries are composed of several thousand unique genes, automation of the process is essential. This review describes the design and implementation of an automated integrated programmable robotic workcell capable of producing complementary DNA libraries, colony picking, isolating plasmid DNA, transforming yeast and bacteria, expressing protein, and performing appropriate functional assays. These operations will allow tailoring microbial strains to use renewable feedstocks for production of biofuels, bioderived chemicals, fertilizers, and other coproducts for profitable and sustainable biorefineries. Published by Elsevier Inc.

  12. Patterning cell using Si-stencil for high-throughput assay

    KAUST Repository

    Wu, Jinbo

    2011-01-01

    In this communication, we report a newly developed cell pattering methodology by a silicon-based stencil, which exhibited advantages such as easy handling, reusability, hydrophilic surface and mature fabrication technologies. Cell arrays obtained by this method were used to investigate cell growth under a temperature gradient, which demonstrated the possibility of studying cell behavior in a high-throughput assay. This journal is © The Royal Society of Chemistry 2011.

  13. High-throughput diagnosis of potato cyst nematodes in soil samples.

    Science.gov (United States)

    Reid, Alex; Evans, Fiona; Mulholland, Vincent; Cole, Yvonne; Pickup, Jon

    2015-01-01

    Potato cyst nematode (PCN) is a damaging soilborne pest of potatoes which can cause major crop losses. In 2010, a new European Union directive (2007/33/EC) on the control of PCN came into force. Under the new directive, seed potatoes can only be planted on land which has been found to be free from PCN infestation following an official soil test. A major consequence of the new directive was the introduction of a new harmonized soil sampling rate resulting in a threefold increase in the number of samples requiring testing. To manage this increase with the same staffing resources, we have replaced the traditional diagnostic methods. A system has been developed for the processing of soil samples, extraction of DNA from float material, and detection of PCN by high-throughput real-time PCR. Approximately 17,000 samples are analyzed each year using this method. This chapter describes the high-throughput processes for the production of float material from soil samples, DNA extraction from the entire float, and subsequent detection and identification of PCN within these samples.

  14. A multilayer microdevice for cell-based high-throughput drug screening

    International Nuclear Information System (INIS)

    Liu, Chong; Wang, Lei; Li, Jingmin; Ding, Xiping; Chunyu, Li; Xu, Zheng; Wang, Qi

    2012-01-01

    A multilayer polydimethylsiloxane microdevice for cell-based high-throughput drug screening is described in this paper. This established microdevice was based on a modularization method and it integrated a drug/medium concentration gradient generator (CGG), pneumatic microvalves and a cell culture microchamber array. The CGG was able to generate five steps of linear concentrations with the same outlet flow rate. The medium/drug flowed through CGG and then into the pear-shaped cell culture microchambers vertically. This vertical perfusion mode was used to reduce the impact of the shear stress on the physiology of cells induced by the fluid flow in the microchambers. Pear-shaped microchambers with two arrays of miropillars at each outlet were adopted in this microdevice, which were beneficial to cell distribution. The chemotherapeutics Cisplatin (DDP)-induced Cisplatin-resistant cell line A549/DDP apoptotic experiments were performed well on this platform. The results showed that this novel microdevice could not only provide well-defined and stable conditions for cell culture, but was also useful for cell-based high-throughput drug screening with less reagents and time consumption. (paper)

  15. A High-throughput Selection for Cellulase Catalysts Using Chemical Complementation

    Science.gov (United States)

    Peralta-Yahya, Pamela; Carter, Brian T.; Lin, Hening; Tao, Haiyan; Cornish, Virginia W.

    2010-01-01

    Efficient enzymatic hydrolysis of lignocellulosic material remains one of the major bottlenecks to cost-effective conversion of biomass to ethanol. Improvement of glycosylhydrolases however is limited by existing medium-throughput screening technologies. Here, we report the first high-throughput selection for cellulase catalysts. This selection was developed by adapting chemical complementation to provide a growth assay for bond cleavage reactions. First, a URA3 counter selection was adapted to link chemical dimerizer activated gene transcription to cell death. Next, the URA3 counter selection was shown to detect cellulase activity based on cleavage of a tetrasaccharide chemical dimerizer substrate and decrease in expression of the toxic URA3 reporter. Finally, the utility of the cellulase selection was assessed by isolating cellulases with improved activity from a cellulase library created by family DNA shuffling. This application provides further evidence that chemical complementation can be readily adapted to detect different enzymatic activities for important chemical transformations for which no natural selection exists. Due to the large number of enzyme variants selections can test compared to existing medium-throughput screens for cellulases, this assay has the potential to impact the discovery of improved cellulases and other glycosylhydrolases for biomass conversion from libraries of cellulases created by mutagenesis or obtained from natural biodiversity. PMID:19053460

  16. An Automated High Throughput Proteolysis and Desalting Platform for Quantitative Proteomic Analysis

    Directory of Open Access Journals (Sweden)

    Albert-Baskar Arul

    2013-06-01

    Full Text Available Proteomics for biomarker validation needs high throughput instrumentation to analyze huge set of clinical samples for quantitative and reproducible analysis at a minimum time without manual experimental errors. Sample preparation, a vital step in proteomics plays a major role in identification and quantification of proteins from biological samples. Tryptic digestion a major check point in sample preparation for mass spectrometry based proteomics needs to be more accurate with rapid processing time. The present study focuses on establishing a high throughput automated online system for proteolytic digestion and desalting of proteins from biological samples quantitatively and qualitatively in a reproducible manner. The present study compares online protein digestion and desalting of BSA with conventional off-line (in-solution method and validated for real time sample for reproducibility. Proteins were identified using SEQUEST data base search engine and the data were quantified using IDEALQ software. The present study shows that the online system capable of handling high throughput samples in 96 well formats carries out protein digestion and peptide desalting efficiently in a reproducible and quantitative manner. Label free quantification showed clear increase of peptide quantities with increase in concentration with much linearity compared to off line method. Hence we would like to suggest that inclusion of this online system in proteomic pipeline will be effective in quantification of proteins in comparative proteomics were the quantification is really very crucial.

  17. A high throughput platform for understanding the influence of excipients on physical and chemical stability

    DEFF Research Database (Denmark)

    Raijada, Dhara; Cornett, Claus; Rantanen, Jukka

    2013-01-01

    The present study puts forward a miniaturized high-throughput platform to understand influence of excipient selection and processing on the stability of a given drug compound. Four model drugs (sodium naproxen, theophylline, amlodipine besylate and nitrofurantoin) and ten different excipients were...... for chemical degradation. The proposed high-throughput platform can be used during early drug development to simulate typical processing induced stress in a small scale and to understand possible phase transformation behaviour and influence of excipients on this....

  18. Fabrication of combinatorial nm-planar electrode array for high throughput evaluation of organic semiconductors

    International Nuclear Information System (INIS)

    Haemori, M.; Edura, T.; Tsutsui, K.; Itaka, K.; Wada, Y.; Koinuma, H.

    2006-01-01

    We have fabricated a combinatorial nm-planar electrode array by using photolithography and chemical mechanical polishing processes for high throughput electrical evaluation of organic devices. Sub-nm precision was achieved with respect to the average level difference between each pair of electrodes and a dielectric layer. The insulating property between the electrodes is high enough to measure I-V characteristics of organic semiconductors. Bottom-contact field-effect-transistors (FETs) of pentacene were fabricated on this electrode array by use of molecular beam epitaxy. It was demonstrated that the array could be used as a pre-patterned device substrate for high throughput screening of the electrical properties of organic semiconductors

  19. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    International Nuclear Information System (INIS)

    Cuttitta, Christina M.; Ericson, Daniel L.; Scalia, Alexander; Roessler, Christian G.; Teplitsky, Ella; Joshi, Karan; Campos, Olven; Agarwal, Rakhi; Allaire, Marc; Orville, Allen M.; Sweet, Robert M.; Soares, Alexei S.

    2015-01-01

    An acoustic high-throughput screening method is described for harvesting protein crystals and combining the protein crystals with chemicals such as a fragment library. Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s −1 ) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening

  20. Development of a High-Throughput Screen for Inhibitors of Epstein-Barr Virus EBNA1

    Science.gov (United States)

    Thompson, Scott; Messick, Troy; Schultz, David C.; Reichman, Melvin; Lieberman, Paul M.

    2012-01-01

    Latent infection with Epstein-Barr Virus (EBV) is a carcinogenic cofactor in several lymphoid and epithelial cell malignancies. At present, there are no small molecule inhibitors that specifically target EBV latent infection or latency-associated oncoproteins. EBNA1 is an EBV-encoded sequence-specific DNA-binding protein that is consistently expressed in EBV-associated tumors and required for stable maintenance of the viral genome in proliferating cells. EBNA1 is also thought to provide cell survival function in latently infected cells. In this work we describe the development of a biochemical high-throughput screening (HTS) method using a homogenous fluorescence polarization (FP) assay monitoring EBNA1 binding to its cognate DNA binding site. An FP-based counterscreen was developed using another EBV-encoded DNA binding protein, Zta, and its cognate DNA binding site. We demonstrate that EBNA1 binding to a fluorescent labeled DNA probe provides a robust assay with a Z-factor consistently greater than 0.6. A pilot screen of a small molecule library of ~14,000 compounds identified 3 structurally related molecules that selectively inhibit EBNA1, but not Zta. All three compounds had activity in a cell-based assay specific for the disruption of EBNA1 transcription repression function. One of the compounds was effective in reducing EBV genome copy number in Raji Burkitt lymphoma cells. These experiments provide a proof-of-concept that small molecule inhibitors of EBNA1 can be identified by biochemical high-throughput screening of compound libraries. Further screening in conjunction with medicinal chemistry optimization may provide a selective inhibitor of EBNA1 and EBV latent infection. PMID:20930215

  1. 40 CFR Table 9 to Subpart Eeee of... - Continuous Compliance With Operating Limits-High Throughput Transfer Racks

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 12 2010-07-01 2010-07-01 true Continuous Compliance With Operating Limits-High Throughput Transfer Racks 9 Table 9 to Subpart EEEE of Part 63 Protection of Environment...—Continuous Compliance With Operating Limits—High Throughput Transfer Racks As stated in §§ 63.2378(a) and (b...

  2. High-Throughput Quantification of Nanoparticle Degradation Using Computational Microscopy and Its Application to Drug Delivery Nanocapsules

    KAUST Repository

    Ray, Aniruddha

    2017-04-25

    Design and synthesis of degradable nanoparticles are very important in drug delivery and biosensing fields. Although accurate assessment of nanoparticle degradation rate would improve the characterization and optimization of drug delivery vehicles, current methods rely on estimating the size of the particles at discrete points over time using, for example, electron microscopy or dynamic light scattering (DLS), among other techniques, all of which have drawbacks and practical limitations. There is a significant need for a high-throughput and cost-effective technology to accurately monitor nanoparticle degradation as a function of time and using small amounts of sample. To address this need, here we present two different computational imaging-based methods for monitoring and quantification of nanoparticle degradation. The first method is suitable for discrete testing, where a computational holographic microscope is designed to track the size changes of protease-sensitive protein-core nanoparticles following degradation, by periodically sampling a subset of particles mixed with proteases. In the second method, a sandwich structure was utilized to observe, in real-time, the change in the properties of liquid nanolenses that were self-assembled around degrading nanoparticles, permitting continuous monitoring and quantification of the degradation process. These cost-effective holographic imaging based techniques enable high-throughput monitoring of the degradation of any type of nanoparticle, using an extremely small amount of sample volume that is at least 3 orders of magnitude smaller than what is required by, for example, DLS-based techniques.

  3. Meta-Analysis of High-Throughput Datasets Reveals Cellular Responses Following Hemorrhagic Fever Virus Infection

    Directory of Open Access Journals (Sweden)

    Gavin C. Bowick

    2011-05-01

    Full Text Available The continuing use of high-throughput assays to investigate cellular responses to infection is providing a large repository of information. Due to the large number of differentially expressed transcripts, often running into the thousands, the majority of these data have not been thoroughly investigated. Advances in techniques for the downstream analysis of high-throughput datasets are providing additional methods for the generation of additional hypotheses for further investigation. The large number of experimental observations, combined with databases that correlate particular genes and proteins with canonical pathways, functions and diseases, allows for the bioinformatic exploration of functional networks that may be implicated in replication or pathogenesis. Herein, we provide an example of how analysis of published high-throughput datasets of cellular responses to hemorrhagic fever virus infection can generate additional functional data. We describe enrichment of genes involved in metabolism, post-translational modification and cardiac damage; potential roles for specific transcription factors and a conserved involvement of a pathway based around cyclooxygenase-2. We believe that these types of analyses can provide virologists with additional hypotheses for continued investigation.

  4. High-throughput protein crystallization on the World Community Grid and the GPU

    International Nuclear Information System (INIS)

    Kotseruba, Yulia; Cumbaa, Christian A; Jurisica, Igor

    2012-01-01

    We have developed CPU and GPU versions of an automated image analysis and classification system for protein crystallization trial images from the Hauptman Woodward Institute's High-Throughput Screening lab. The analysis step computes 12,375 numerical features per image. Using these features, we have trained a classifier that distinguishes 11 different crystallization outcomes, recognizing 80% of all crystals, 94% of clear drops, 94% of precipitates. The computing requirements for this analysis system are large. The complete HWI archive of 120 million images is being processed by the donated CPU cycles on World Community Grid, with a GPU phase launching in early 2012. The main computational burden of the analysis is the measure of textural (GLCM) features within the image at multiple neighbourhoods, distances, and at multiple greyscale intensity resolutions. CPU runtime averages 4,092 seconds (single threaded) on an Intel Xeon, but only 65 seconds on an NVIDIA Tesla C2050. We report on the process of adapting the C++ code to OpenCL, optimized for multiple platforms.

  5. Two-dimensional materials from high-throughput computational exfoliation of experimentally known compounds

    Science.gov (United States)

    Mounet, Nicolas; Gibertini, Marco; Schwaller, Philippe; Campi, Davide; Merkys, Andrius; Marrazzo, Antimo; Sohier, Thibault; Castelli, Ivano Eligio; Cepellotti, Andrea; Pizzi, Giovanni; Marzari, Nicola

    2018-02-01

    Two-dimensional (2D) materials have emerged as promising candidates for next-generation electronic and optoelectronic applications. Yet, only a few dozen 2D materials have been successfully synthesized or exfoliated. Here, we search for 2D materials that can be easily exfoliated from their parent compounds. Starting from 108,423 unique, experimentally known 3D compounds, we identify a subset of 5,619 compounds that appear layered according to robust geometric and bonding criteria. High-throughput calculations using van der Waals density functional theory, validated against experimental structural data and calculated random phase approximation binding energies, further allowed the identification of 1,825 compounds that are either easily or potentially exfoliable. In particular, the subset of 1,036 easily exfoliable cases provides novel structural prototypes and simple ternary compounds as well as a large portfolio of materials to search from for optimal properties. For a subset of 258 compounds, we explore vibrational, electronic, magnetic and topological properties, identifying 56 ferromagnetic and antiferromagnetic systems, including half-metals and half-semiconductors.

  6. A Fully Automated High-Throughput Zebrafish Behavioral Ototoxicity Assay.

    Science.gov (United States)

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

    2017-08-01

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

  7. Combining high-throughput phenotyping and genome-wide association studies to reveal natural genetic variation in rice

    Science.gov (United States)

    Yang, Wanneng; Guo, Zilong; Huang, Chenglong; Duan, Lingfeng; Chen, Guoxing; Jiang, Ni; Fang, Wei; Feng, Hui; Xie, Weibo; Lian, Xingming; Wang, Gongwei; Luo, Qingming; Zhang, Qifa; Liu, Qian; Xiong, Lizhong

    2014-01-01

    Even as the study of plant genomics rapidly develops through the use of high-throughput sequencing techniques, traditional plant phenotyping lags far behind. Here we develop a high-throughput rice phenotyping facility (HRPF) to monitor 13 traditional agronomic traits and 2 newly defined traits during the rice growth period. Using genome-wide association studies (GWAS) of the 15 traits, we identify 141 associated loci, 25 of which contain known genes such as the Green Revolution semi-dwarf gene, SD1. Based on a performance evaluation of the HRPF and GWAS results, we demonstrate that high-throughput phenotyping has the potential to replace traditional phenotyping techniques and can provide valuable gene identification information. The combination of the multifunctional phenotyping tools HRPF and GWAS provides deep insights into the genetic architecture of important traits. PMID:25295980

  8. Ethoscopes: An open platform for high-throughput ethomics.

    Directory of Open Access Journals (Sweden)

    Quentin Geissmann

    2017-10-01

    Full Text Available Here, we present the use of ethoscopes, which are machines for high-throughput analysis of behavior in Drosophila and other animals. Ethoscopes provide a software and hardware solution that is reproducible and easily scalable. They perform, in real-time, tracking and profiling of behavior by using a supervised machine learning algorithm, are able to deliver behaviorally triggered stimuli to flies in a feedback-loop mode, and are highly customizable and open source. Ethoscopes can be built easily by using 3D printing technology and rely on Raspberry Pi microcomputers and Arduino boards to provide affordable and flexible hardware. All software and construction specifications are available at http://lab.gilest.ro/ethoscope.

  9. Correction of Microplate Data from High-Throughput Screening.

    Science.gov (United States)

    Wang, Yuhong; Huang, Ruili

    2016-01-01

    High-throughput screening (HTS) makes it possible to collect cellular response data from a large number of cell lines and small molecules in a timely and cost-effective manner. The errors and noises in the microplate-formatted data from HTS have unique characteristics, and they can be generally grouped into three categories: run-wise (temporal, multiple plates), plate-wise (background pattern, single plate), and well-wise (single well). In this chapter, we describe a systematic solution for identifying and correcting such errors and noises, mainly basing on pattern recognition and digital signal processing technologies.

  10. High-Throughput Network Communication with NetIO

    CERN Document Server

    Schumacher, J\\"orn; The ATLAS collaboration; Vandelli, Wainer

    2016-01-01

    HPC network technologies like Infiniband, TrueScale or OmniPath provide low-latency and high-throughput communication between hosts, which makes them attractive options for data-acquisition systems in large-scale high-energy physics experiments. Like HPC networks, DAQ networks are local and include a well specified number of systems. Unfortunately traditional network communication APIs for HPC clusters like MPI or PGAS target exclusively the HPC community and are not suited well for DAQ applications. It is possible to build distributed DAQ applications using low-level system APIs like Infiniband Verbs (and this has been done), but it requires a non negligible effort and expert knowledge. On the other hand, message services like 0MQ have gained popularity in the HEP community. Such APIs allow to build distributed applications with a high-level approach and provide good performance. Unfortunately their usage usually limits developers to TCP/IP-based networks. While it is possible to operate a TCP/IP stack on to...

  11. High-Throughput Printing Process for Flexible Electronics

    Science.gov (United States)

    Hyun, Woo Jin

    Printed electronics is an emerging field for manufacturing electronic devices with low cost and minimal material waste for a variety of applications including displays, distributed sensing, smart packaging, and energy management. Moreover, its compatibility with roll-to-roll production formats and flexible substrates is desirable for continuous, high-throughput production of flexible electronics. Despite the promise, however, the roll-to-roll production of printed electronics is quite challenging due to web movement hindering accurate ink registration and high-fidelity printing. In this talk, I will present a promising strategy for roll-to-roll production using a novel printing process that we term SCALE (Self-aligned Capillarity-Assisted Lithography for Electronics). By utilizing capillarity of liquid inks on nano/micro-structured substrates, the SCALE process facilitates high-resolution and self-aligned patterning of electrically functional inks with greatly improved printing tolerance. I will show the fabrication of key building blocks (e.g. transistor, resistor, capacitor) for electronic circuits using the SCALE process on plastics.

  12. High-throughput screening of tick-borne pathogens in Europe

    DEFF Research Database (Denmark)

    Michelet, Lorraine; Delannoy, Sabine; Devillers, Elodie

    2014-01-01

    was conducted on 7050 Ixodes ricinus nymphs collected from France, Denmark, and the Netherlands using a powerful new high-throughput approach. This advanced methodology permitted the simultaneous detection of 25 bacterial, and 12 parasitic species (including; Borrelia, Anaplasma, Ehrlichia, Rickettsia......, Bartonella, Candidatus Neoehrlichia, Coxiella, Francisella, Babesia, and Theileria genus) across 94 samples. We successfully determined the prevalence of expected (Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Rickettsia helvetica, Candidatus Neoehrlichia mikurensis, Babesia divergens, Babesia...

  13. High Throughput Single-cell and Multiple-cell Micro-encapsulation

    OpenAIRE

    Lagus, Todd P.; Edd, Jon F.

    2012-01-01

    Microfluidic encapsulation methods have been previously utilized to capture cells in picoliter-scale aqueous, monodisperse drops, providing confinement from a bulk fluid environment with applications in high throughput screening, cytometry, and mass spectrometry. We describe a method to not only encapsulate single cells, but to repeatedly capture a set number of cells (here we demonstrate one- and two-cell encapsulation) to study both isolation and the interactions between cells in groups of ...

  14. Low Cost, High-Throughput 3-D Pulmonary Imager Using Hyperpolarized Contrast Agents and Low-Field MRI

    Science.gov (United States)

    2017-10-01

    greater gas polarizations and production amounts/ throughputs- benefiting in particular from the advent of com- pact, high-power, relatively low- cost ...Award Number: W81XWH-15-1-0271 TITLE: Low- Cost , High-Throughput 3-D Pulmonary Imager Using Hyperpolarized Contrast Agents and Low-Field MRI...DISTRIBUTION STATEMENT: Approved for Public Release; Distribution Unlimited The views, opinions and/or findings contained in this report are those of the

  15. High throughput octal alpha/gamma spectrometer for low level bioassay estimations

    International Nuclear Information System (INIS)

    Bhasin, B.D.; Shirke, S.H.; Suri, M.M.; Vaidya, P.P.; Ghodgaonkar, M.D.

    1995-01-01

    The present paper describes the development of a high throughput octal alpha spectrometry system specially developed for the estimation of low levels of actinides in bioassay and environmental samples. The system processes simultaneously the outputs coming from eight independent detectors. It can be configured to simultaneously record low level alpha and gamma spectra. The high throughput is achieved by using a prioritised multiplexer router. The prioritised multiplexing and routing coupled with fast 8K ADC (conversion time 20 μsec) allow simultaneous acquisition of multiple spectra without any significant loss in counts. The dual (8K, 24bit) port memory facilitates easy online viewing of spectrum buildup. A menu driven user friendly software makes the operating system convenient to use. A specially developed software provides built-in routines for processing the spectra and estimating the isotopic activity. The interactive mode of software provides easy identification of isotopes compatible with the separation chemistry of different actinides. (author). 6 refs., 2 figs

  16. Large-scale DNA Barcode Library Generation for Biomolecule Identification in High-throughput Screens.

    Science.gov (United States)

    Lyons, Eli; Sheridan, Paul; Tremmel, Georg; Miyano, Satoru; Sugano, Sumio

    2017-10-24

    High-throughput screens allow for the identification of specific biomolecules with characteristics of interest. In barcoded screens, DNA barcodes are linked to target biomolecules in a manner allowing for the target molecules making up a library to be identified by sequencing the DNA barcodes using Next Generation Sequencing. To be useful in experimental settings, the DNA barcodes in a library must satisfy certain constraints related to GC content, homopolymer length, Hamming distance, and blacklisted subsequences. Here we report a novel framework to quickly generate large-scale libraries of DNA barcodes for use in high-throughput screens. We show that our framework dramatically reduces the computation time required to generate large-scale DNA barcode libraries, compared with a naїve approach to DNA barcode library generation. As a proof of concept, we demonstrate that our framework is able to generate a library consisting of one million DNA barcodes for use in a fragment antibody phage display screening experiment. We also report generating a general purpose one billion DNA barcode library, the largest such library yet reported in literature. Our results demonstrate the value of our novel large-scale DNA barcode library generation framework for use in high-throughput screening applications.

  17. High-throughput metabolic state analysis: The missing link in integrated functional genomics of yeasts

    DEFF Research Database (Denmark)

    Villas-Bôas, Silas Granato; Moxley, Joel. F; Åkesson, Mats Fredrik

    2005-01-01

    that achieve comparable throughput, effort and cost compared with DNA arrays. Our sample workup method enables simultaneous metabolite measurements throughout central carbon metabolism and amino acid biosynthesis, using a standard GC-MS platform that was optimized for this Purpose. As an implementation proof......-of-concept, we assayed metabolite levels in two yeast strains and two different environmental conditions in the context of metabolic pathway reconstruction. We demonstrate that these differential metabolite level data distinguish among sample types, such as typical metabolic fingerprinting or footprinting. More...

  18. High-throughput investigation of polymerization kinetics by online monitoring of GPC and GC

    NARCIS (Netherlands)

    Hoogenboom, R.; Fijten, M.W.M.; Abeln, C.H.; Schubert, U.S.

    2004-01-01

    Gel permeation chromatography (GPC) and gas chromatography (GC) were successfully introduced into a high-throughput workflow. The feasibility and limitations of online GPC with a high-speed column was evaluated by measuring polystyrene standards and comparison of the results with regular offline GPC

  19. ToxCast Workflow: High-throughput screening assay data processing, analysis and management (SOT)

    Science.gov (United States)

    US EPA’s ToxCast program is generating data in high-throughput screening (HTS) and high-content screening (HCS) assays for thousands of environmental chemicals, for use in developing predictive toxicity models. Currently the ToxCast screening program includes over 1800 unique c...

  20. Computational and statistical methods for high-throughput mass spectrometry-based PTM analysis

    DEFF Research Database (Denmark)

    Schwämmle, Veit; Vaudel, Marc

    2017-01-01

    Cell signaling and functions heavily rely on post-translational modifications (PTMs) of proteins. Their high-throughput characterization is thus of utmost interest for multiple biological and medical investigations. In combination with efficient enrichment methods, peptide mass spectrometry analy...

  1. Applications of high-throughput sequencing to chromatin structure and function in mammals

    OpenAIRE

    Dunham, Ian

    2009-01-01

    High-throughput DNA sequencing approaches have enabled direct interrogation of chromatin samples from mammalian cells. We are beginning to develop a genome-wide description of nuclear function during development, but further data collection, refinement, and integration are needed.

  2. Low-Cost, High-Throughput 3-D Pulmonary Imager Using Hyperpolarized Contrast Agents and Low-Field MRI

    Science.gov (United States)

    2017-10-01

    low- cost and high-throughput was a key element proposed for this project, which we believe will be of significant benefit to the patients suffering...Award Number: W81XWH-15-1-0272 TITLE: Low- Cost , High-Throughput 3-D Pulmonary Imager Using Hyperpolarized Contrast Agents and Low-Field MRI...STATEMENT: Approved for Public Release; Distribution Unlimited The views, opinions and/or findings contained in this report are those of the author(s

  3. Protocol: high throughput silica-based purification of RNA from Arabidopsis seedlings in a 96-well format

    OpenAIRE

    Salvo-Chirnside, Eliane; Kane, Steven; Kerr, Lorraine E

    2011-01-01

    Abstract The increasing popularity of systems-based approaches to plant research has resulted in a demand for high throughput (HTP) methods to be developed. RNA extraction from multiple samples in an experiment is a significant bottleneck in performing systems-level genomic studies. Therefore we have established a high throughput method of RNA extraction from Arabidopsis thaliana to facilitate gene expression studies in this widely used plant model. We present optimised manual and automated p...

  4. From Classical to High Throughput Screening Methods for Feruloyl Esterases: A Review.

    Science.gov (United States)

    Ramírez-Velasco, Lorena; Armendáriz-Ruiz, Mariana; Rodríguez-González, Jorge Alberto; Müller-Santos, Marcelo; Asaff-Torres, Ali; Mateos-Díaz, Juan Carlos

    2016-01-01

    Feruloyl esterases (FAEs) are a diverse group of hydrolases widely distributed in plants and microorganisms which catalyzes the cleavage and formation of ester bonds between plant cell wall polysaccharides and phenolic acids. FAEs have gained importance in biofuel, medicine and food industries due to their capability of acting on a large range of substrates for cleaving ester bonds and synthesizing highadded value molecules through esterification and transesterification reactions. During the past two decades extensive studies have been carried out on the production, characterization and classification of FAEs, however only a few reports of suitable High Throughput Screening assays for this kind of enzymes have been reported. This review is focused on a concise but complete revision of classical to High Throughput Screening methods for FAEs, highlighting its advantages and disadvantages, and finally suggesting future perspectives for this important research field.

  5. AELAS: Automatic ELAStic property derivations via high-throughput first-principles computation

    Science.gov (United States)

    Zhang, S. H.; Zhang, R. F.

    2017-11-01

    The elastic properties are fundamental and important for crystalline materials as they relate to other mechanical properties, various thermodynamic qualities as well as some critical physical properties. However, a complete set of experimentally determined elastic properties is only available for a small subset of known materials, and an automatic scheme for the derivations of elastic properties that is adapted to high-throughput computation is much demanding. In this paper, we present the AELAS code, an automated program for calculating second-order elastic constants of both two-dimensional and three-dimensional single crystal materials with any symmetry, which is designed mainly for high-throughput first-principles computation. Other derivations of general elastic properties such as Young's, bulk and shear moduli as well as Poisson's ratio of polycrystal materials, Pugh ratio, Cauchy pressure, elastic anisotropy and elastic stability criterion, are also implemented in this code. The implementation of the code has been critically validated by a lot of evaluations and tests on a broad class of materials including two-dimensional and three-dimensional materials, providing its efficiency and capability for high-throughput screening of specific materials with targeted mechanical properties. Program Files doi:http://dx.doi.org/10.17632/f8fwg4j9tw.1 Licensing provisions: BSD 3-Clause Programming language: Fortran Nature of problem: To automate the calculations of second-order elastic constants and the derivations of other elastic properties for two-dimensional and three-dimensional materials with any symmetry via high-throughput first-principles computation. Solution method: The space-group number is firstly determined by the SPGLIB code [1] and the structure is then redefined to unit cell with IEEE-format [2]. Secondly, based on the determined space group number, a set of distortion modes is automatically specified and the distorted structure files are generated

  6. Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy

    DEFF Research Database (Denmark)

    Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H

    2017-01-01

    To target bacterial pathogens that invade and proliferate inside host cells, it is necessary to design intervention strategies directed against bacterial attachment, cellular invasion and intracellular proliferation. We present an automated microscopy-based, fast, high-throughput method for analy...

  7. High-throughput screening of small-molecule adsorption in MOF-74

    Science.gov (United States)

    Thonhauser, T.; Canepa, P.

    2014-03-01

    Using high-throughput screening coupled with state-of-the-art van der Waals density functional theory, we investigate the adsorption properties of four important molecules, H2, CO2, CH4, and H2O in MOF-74-  with  = Be, Mg, Al, Ca, Sc, Ti, V, Cr, Mn, Fe, Co, Ni, Cu, Zn, Sr, Zr, Nb, Ru, Rh, Pd, La, W, Os, Ir, and Pt. We show that high-throughput techniques can aid in speeding up the development and refinement of effective materials for hydrogen storage, carbon capture, and gas separation. The exploration of the configurational adsorption space allows us to extract crucial information concerning, for example, the competition of water with CO2 for the adsorption binding sites. We find that only a few noble metals--Rh, Pd, Os, Ir, and Pt--favor the adsorption of CO2 and hence are potential candidates for effective carbon-capture materials. Our findings further reveal significant differences in the binding characteristics of H2, CO2, CH4, and H2O within the MOF structure, indicating that molecular blends can be successfully separated by these nano-porous materials. Supported by DOE DE-FG02-08ER46491.

  8. A primer on high-throughput computing for genomic selection.

    Science.gov (United States)

    Wu, Xiao-Lin; Beissinger, Timothy M; Bauck, Stewart; Woodward, Brent; Rosa, Guilherme J M; Weigel, Kent A; Gatti, Natalia de Leon; Gianola, Daniel

    2011-01-01

    High-throughput computing (HTC) uses computer clusters to solve advanced computational problems, with the goal of accomplishing high-throughput over relatively long periods of time. In genomic selection, for example, a set of markers covering the entire genome is used to train a model based on known data, and the resulting model is used to predict the genetic merit of selection candidates. Sophisticated models are very computationally demanding and, with several traits to be evaluated sequentially, computing time is long, and output is low. In this paper, we present scenarios and basic principles of how HTC can be used in genomic selection, implemented using various techniques from simple batch processing to pipelining in distributed computer clusters. Various scripting languages, such as shell scripting, Perl, and R, are also very useful to devise pipelines. By pipelining, we can reduce total computing time and consequently increase throughput. In comparison to the traditional data processing pipeline residing on the central processors, performing general-purpose computation on a graphics processing unit provide a new-generation approach to massive parallel computing in genomic selection. While the concept of HTC may still be new to many researchers in animal breeding, plant breeding, and genetics, HTC infrastructures have already been built in many institutions, such as the University of Wisconsin-Madison, which can be leveraged for genomic selection, in terms of central processing unit capacity, network connectivity, storage availability, and middleware connectivity. Exploring existing HTC infrastructures as well as general-purpose computing environments will further expand our capability to meet increasing computing demands posed by unprecedented genomic data that we have today. We anticipate that HTC will impact genomic selection via better statistical models, faster solutions, and more competitive products (e.g., from design of marker panels to realized

  9. Automated image alignment for 2D gel electrophoresis in a high-throughput proteomics pipeline.

    Science.gov (United States)

    Dowsey, Andrew W; Dunn, Michael J; Yang, Guang-Zhong

    2008-04-01

    The quest for high-throughput proteomics has revealed a number of challenges in recent years. Whilst substantial improvements in automated protein separation with liquid chromatography and mass spectrometry (LC/MS), aka 'shotgun' proteomics, have been achieved, large-scale open initiatives such as the Human Proteome Organization (HUPO) Brain Proteome Project have shown that maximal proteome coverage is only possible when LC/MS is complemented by 2D gel electrophoresis (2-DE) studies. Moreover, both separation methods require automated alignment and differential analysis to relieve the bioinformatics bottleneck and so make high-throughput protein biomarker discovery a reality. The purpose of this article is to describe a fully automatic image alignment framework for the integration of 2-DE into a high-throughput differential expression proteomics pipeline. The proposed method is based on robust automated image normalization (RAIN) to circumvent the drawbacks of traditional approaches. These use symbolic representation at the very early stages of the analysis, which introduces persistent errors due to inaccuracies in modelling and alignment. In RAIN, a third-order volume-invariant B-spline model is incorporated into a multi-resolution schema to correct for geometric and expression inhomogeneity at multiple scales. The normalized images can then be compared directly in the image domain for quantitative differential analysis. Through evaluation against an existing state-of-the-art method on real and synthetically warped 2D gels, the proposed analysis framework demonstrates substantial improvements in matching accuracy and differential sensitivity. High-throughput analysis is established through an accelerated GPGPU (general purpose computation on graphics cards) implementation. Supplementary material, software and images used in the validation are available at http://www.proteomegrid.org/rain/.

  10. Use of genotyping by sequencing data to develop a high-throughput and multifunctional SNP panel for conservation applications in Pacific lamprey.

    Science.gov (United States)

    Hess, Jon E; Campbell, Nathan R; Docker, Margaret F; Baker, Cyndi; Jackson, Aaron; Lampman, Ralph; McIlraith, Brian; Moser, Mary L; Statler, David P; Young, William P; Wildbill, Andrew J; Narum, Shawn R

    2015-01-01

    Next-generation sequencing data can be mined for highly informative single nucleotide polymorphisms (SNPs) to develop high-throughput genomic assays for nonmodel organisms. However, choosing a set of SNPs to address a variety of objectives can be difficult because SNPs are often not equally informative. We developed an optimal combination of 96 high-throughput SNP assays from a total of 4439 SNPs identified in a previous study of Pacific lamprey (Entosphenus tridentatus) and used them to address four disparate objectives: parentage analysis, species identification and characterization of neutral and adaptive variation. Nine of these SNPs are FST outliers, and five of these outliers are localized within genes and significantly associated with geography, run-timing and dwarf life history. Two of the 96 SNPs were diagnostic for two other lamprey species that were morphologically indistinguishable at early larval stages and were sympatric in the Pacific Northwest. The majority (85) of SNPs in the panel were highly informative for parentage analysis, that is, putatively neutral with high minor allele frequency across the species' range. Results from three case studies are presented to demonstrate the broad utility of this panel of SNP markers in this species. As Pacific lamprey populations are undergoing rapid decline, these SNPs provide an important resource to address critical uncertainties associated with the conservation and recovery of this imperiled species. © 2014 John Wiley & Sons Ltd.

  11. Novel Acoustic Loading of a Mass Spectrometer: Toward Next-Generation High-Throughput MS Screening.

    Science.gov (United States)

    Sinclair, Ian; Stearns, Rick; Pringle, Steven; Wingfield, Jonathan; Datwani, Sammy; Hall, Eric; Ghislain, Luke; Majlof, Lars; Bachman, Martin

    2016-02-01

    High-throughput, direct measurement of substrate-to-product conversion by label-free detection, without the need for engineered substrates or secondary assays, could be considered the "holy grail" of drug discovery screening. Mass spectrometry (MS) has the potential to be part of this ultimate screening solution, but is constrained by the limitations of existing MS sample introduction modes that cannot meet the throughput requirements of high-throughput screening (HTS). Here we report data from a prototype system (Echo-MS) that uses acoustic droplet ejection (ADE) to transfer femtoliter-scale droplets in a rapid, precise, and accurate fashion directly into the MS. The acoustic source can load samples into the MS from a microtiter plate at a rate of up to three samples per second. The resulting MS signal displays a very sharp attack profile and ions are detected within 50 ms of activation of the acoustic transducer. Additionally, we show that the system is capable of generating multiply charged ion species from simple peptides and large proteins. The combination of high speed and low sample volume has significant potential within not only drug discovery, but also other areas of the industry. © 2015 Society for Laboratory Automation and Screening.

  12. Novel method for the high-throughput processing of slides for the comet assay.

    Science.gov (United States)

    Karbaschi, Mahsa; Cooke, Marcus S

    2014-11-26

    Single cell gel electrophoresis (the comet assay), continues to gain popularity as a means of assessing DNA damage. However, the assay's low sample throughput and laborious sample workup procedure are limiting factors to its application. "Scoring", or individually determining DNA damage levels in 50 cells per treatment, is time-consuming, but with the advent of high-throughput scoring, the limitation is now the ability to process significant numbers of comet slides. We have developed a novel method by which multiple slides may be manipulated, and undergo electrophoresis, in batches of 25 rather than individually and, importantly, retains the use of standard microscope comet slides, which are the assay convention. This decreases assay time by 60%, and benefits from an electrophoresis tank with a substantially smaller footprint, and more uniform orientation of gels during electrophoresis. Our high-throughput variant of the comet assay greatly increases the number of samples analysed, decreases assay time, number of individual slide manipulations, reagent requirements and risk of damage to slides. The compact nature of the electrophoresis tank is of particular benefit to laboratories where bench space is at a premium. This novel approach is a significant advance on the current comet assay procedure.

  13. Robust, high-throughput solution structural analyses by small angle X-ray scattering (SAXS)

    Energy Technology Data Exchange (ETDEWEB)

    Hura, Greg L.; Menon, Angeli L.; Hammel, Michal; Rambo, Robert P.; Poole II, Farris L.; Tsutakawa, Susan E.; Jenney Jr, Francis E.; Classen, Scott; Frankel, Kenneth A.; Hopkins, Robert C.; Yang, Sungjae; Scott, Joseph W.; Dillard, Bret D.; Adams, Michael W. W.; Tainer, John A.

    2009-07-20

    We present an efficient pipeline enabling high-throughput analysis of protein structure in solution with small angle X-ray scattering (SAXS). Our SAXS pipeline combines automated sample handling of microliter volumes, temperature and anaerobic control, rapid data collection and data analysis, and couples structural analysis with automated archiving. We subjected 50 representative proteins, mostly from Pyrococcus furiosus, to this pipeline and found that 30 were multimeric structures in solution. SAXS analysis allowed us to distinguish aggregated and unfolded proteins, define global structural parameters and oligomeric states for most samples, identify shapes and similar structures for 25 unknown structures, and determine envelopes for 41 proteins. We believe that high-throughput SAXS is an enabling technology that may change the way that structural genomics research is done.

  14. High-Throughput Dietary Exposure Predictions for Chemical Migrants from Food Packaging Materials

    Science.gov (United States)

    United States Environmental Protection Agency researchers have developed a Stochastic Human Exposure and Dose Simulation High -Throughput (SHEDS-HT) model for use in prioritization of chemicals under the ExpoCast program. In this research, new methods were implemented in SHEDS-HT...

  15. Acoustic transfer of protein crystals from agarose pedestals to micromeshes for high-throughput screening

    Energy Technology Data Exchange (ETDEWEB)

    Cuttitta, Christina M. [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); The City University of New York, 2800 Victory Boulevard, Staten Island, NY 10314 (United States); Ericson, Daniel L. [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); University at Buffalo, SUNY, 12 Capen Hall, Buffalo, NY 14260 (United States); Scalia, Alexander [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Binghamton University, 4400 Vestal Parkway East, Binghamton, NY 11973-5000 (United States); Roessler, Christian G. [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Teplitsky, Ella [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Stony Brook University, Stony Brook, NY 11794-5215 (United States); Joshi, Karan [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); PEC University of Technology, Chandigarh (India); Campos, Olven [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Florida Atlantic University, 777 Glades Road, Boca Raton, FL 33414 (United States); Agarwal, Rakhi; Allaire, Marc [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Orville, Allen M. [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Brookhaven National Laboratory, Upton, NY 11973-5000 (United States); Sweet, Robert M.; Soares, Alexei S., E-mail: soares@bnl.gov [Brookhaven National Laboratory, Upton, NY 11973-5000 (United States)

    2015-01-01

    An acoustic high-throughput screening method is described for harvesting protein crystals and combining the protein crystals with chemicals such as a fragment library. Acoustic droplet ejection (ADE) is an emerging technology with broad applications in serial crystallography such as growing, improving and manipulating protein crystals. One application of this technology is to gently transfer crystals onto MiTeGen micromeshes with minimal solvent. Once mounted on a micromesh, each crystal can be combined with different chemicals such as crystal-improving additives or a fragment library. Acoustic crystal mounting is fast (2.33 transfers s{sup −1}) and all transfers occur in a sealed environment that is in vapor equilibrium with the mother liquor. Here, a system is presented to retain crystals near the ejection point and away from the inaccessible dead volume at the bottom of the well by placing the crystals on a concave agarose pedestal (CAP) with the same chemical composition as the crystal mother liquor. The bowl-shaped CAP is impenetrable to crystals. Consequently, gravity will gently move the crystals into the optimal location for acoustic ejection. It is demonstrated that an agarose pedestal of this type is compatible with most commercially available crystallization conditions and that protein crystals are readily transferred from the agarose pedestal onto micromeshes with no loss in diffraction quality. It is also shown that crystals can be grown directly on CAPs, which avoids the need to transfer the crystals from the hanging drop to a CAP. This technology has been used to combine thermolysin and lysozyme crystals with an assortment of anomalously scattering heavy atoms. The results point towards a fast nanolitre method for crystal mounting and high-throughput screening.

  16. Recent advances in high-throughput molecular marker identification for superficial and invasive bladder cancers

    DEFF Research Database (Denmark)

    Andersen, Lars Dyrskjøt; Zieger, Karsten; Ørntoft, Torben Falck

    2007-01-01

    individually contributed to the management of the disease. However, the development of high-throughput techniques for simultaneous assessment of a large number of markers has allowed classification of tumors into clinically relevant molecular subgroups beyond those possible by pathological classification. Here......Bladder cancer is the fifth most common neoplasm in industrialized countries. Due to frequent recurrences of the superficial form of this disease, bladder cancer ranks as one of the most common cancers. Despite the description of a large number of tumor markers for bladder cancers, none have......, we review the recent advances in high-throughput molecular marker identification for superficial and invasive bladder cancers....

  17. High-throughput screening of ionic conductivity in polymer membranes

    International Nuclear Information System (INIS)

    Zapata, Pedro; Basak, Pratyay; Carson Meredith, J.

    2009-01-01

    Combinatorial and high-throughput techniques have been successfully used for efficient and rapid property screening in multiple fields. The use of these techniques can be an advantageous new approach to assay ionic conductivity and accelerate the development of novel materials in research areas such as fuel cells. A high-throughput ionic conductivity (HTC) apparatus is described and applied to screening candidate polymer electrolyte membranes for fuel cell applications. The device uses a miniature four-point probe for rapid, automated point-to-point AC electrochemical impedance measurements in both liquid and humid air environments. The conductivity of Nafion 112 HTC validation standards was within 1.8% of the manufacturer's specification. HTC screening of 40 novel Kynar poly(vinylidene fluoride) (PVDF)/acrylic polyelectrolyte (PE) membranes focused on varying the Kynar type (5x) and PE composition (8x) using reduced sample sizes. Two factors were found to be significant in determining the proton conducting capacity: (1) Kynar PVDF series: membranes containing a particular Kynar PVDF type exhibited statistically identical mean conductivity as other membranes containing different Kynar PVDF types that belong to the same series or family. (2) Maximum effective amount of polyelectrolyte: increments in polyelectrolyte content from 55 wt% to 60 wt% showed no statistically significant effect in increasing conductivity. In fact, some membranes experienced a reduction in conductivity.

  18. Combining high-throughput phenotyping and genome-wide association studies to reveal natural genetic variation in rice

    OpenAIRE

    Yang, Wanneng; Guo, Zilong; Huang, Chenglong; Duan, Lingfeng; Chen, Guoxing; Jiang, Ni; Fang, Wei; Feng, Hui; Xie, Weibo; Lian, Xingming; Wang, Gongwei; Luo, Qingming; Zhang, Qifa; Liu, Qian; Xiong, Lizhong

    2014-01-01

    Even as the study of plant genomics rapidly develops through the use of high-throughput sequencing techniques, traditional plant phenotyping lags far behind. Here we develop a high-throughput rice phenotyping facility (HRPF) to monitor 13 traditional agronomic traits and 2 newly defined traits during the rice growth period. Using genome-wide association studies (GWAS) of the 15 traits, we identify 141 associated loci, 25 of which contain known genes such as the Green Revolution semi-dwarf gen...

  19. Creation of a small high-throughput screening facility.

    Science.gov (United States)

    Flak, Tod

    2009-01-01

    The creation of a high-throughput screening facility within an organization is a difficult task, requiring a substantial investment of time, money, and organizational effort. Major issues to consider include the selection of equipment, the establishment of data analysis methodologies, and the formation of a group having the necessary competencies. If done properly, it is possible to build a screening system in incremental steps, adding new pieces of equipment and data analysis modules as the need grows. Based upon our experience with the creation of a small screening service, we present some guidelines to consider in planning a screening facility.

  20. Bacterial Microcolonies in Gel Beads for High-Throughput Screening of Libraries in Synthetic Biology.

    Science.gov (United States)

    Duarte, José M; Barbier, Içvara; Schaerli, Yolanda

    2017-11-17

    Synthetic biologists increasingly rely on directed evolution to optimize engineered biological systems. Applying an appropriate screening or selection method for identifying the potentially rare library members with the desired properties is a crucial step for success in these experiments. Special challenges include substantial cell-to-cell variability and the requirement to check multiple states (e.g., being ON or OFF depending on the input). Here, we present a high-throughput screening method that addresses these challenges. First, we encapsulate single bacteria into microfluidic agarose gel beads. After incubation, they harbor monoclonal bacterial microcolonies (e.g., expressing a synthetic construct) and can be sorted according their fluorescence by fluorescence activated cell sorting (FACS). We determine enrichment rates and demonstrate that we can measure the average fluorescent signals of microcolonies containing phenotypically heterogeneous cells, obviating the problem of cell-to-cell variability. Finally, we apply this method to sort a pBAD promoter library at ON and OFF states.

  1. High Resolution Melting (HRM for High-Throughput Genotyping—Limitations and Caveats in Practical Case Studies

    Directory of Open Access Journals (Sweden)

    Marcin Słomka

    2017-11-01

    Full Text Available High resolution melting (HRM is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs. This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup.

  2. High Resolution Melting (HRM) for High-Throughput Genotyping—Limitations and Caveats in Practical Case Studies

    Science.gov (United States)

    Słomka, Marcin; Sobalska-Kwapis, Marta; Wachulec, Monika; Bartosz, Grzegorz

    2017-01-01

    High resolution melting (HRM) is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs). This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup. PMID:29099791

  3. High Resolution Melting (HRM) for High-Throughput Genotyping-Limitations and Caveats in Practical Case Studies.

    Science.gov (United States)

    Słomka, Marcin; Sobalska-Kwapis, Marta; Wachulec, Monika; Bartosz, Grzegorz; Strapagiel, Dominik

    2017-11-03

    High resolution melting (HRM) is a convenient method for gene scanning as well as genotyping of individual and multiple single nucleotide polymorphisms (SNPs). This rapid, simple, closed-tube, homogenous, and cost-efficient approach has the capacity for high specificity and sensitivity, while allowing easy transition to high-throughput scale. In this paper, we provide examples from our laboratory practice of some problematic issues which can affect the performance and data analysis of HRM results, especially with regard to reference curve-based targeted genotyping. We present those examples in order of the typical experimental workflow, and discuss the crucial significance of the respective experimental errors and limitations for the quality and analysis of results. The experimental details which have a decisive impact on correct execution of a HRM genotyping experiment include type and quality of DNA source material, reproducibility of isolation method and template DNA preparation, primer and amplicon design, automation-derived preparation and pipetting inconsistencies, as well as physical limitations in melting curve distinction for alternative variants and careful selection of samples for validation by sequencing. We provide a case-by-case analysis and discussion of actual problems we encountered and solutions that should be taken into account by researchers newly attempting HRM genotyping, especially in a high-throughput setup.

  4. Identifying Inhibitors of Inflammation: A Novel High-Throughput MALDI-TOF Screening Assay for Salt-Inducible Kinases (SIKs).

    Science.gov (United States)

    Heap, Rachel E; Hope, Anthony G; Pearson, Lesley-Anne; Reyskens, Kathleen M S E; McElroy, Stuart P; Hastie, C James; Porter, David W; Arthur, J Simon C; Gray, David W; Trost, Matthias

    2017-12-01

    Matrix-assisted laser desorption/ionization time-of-flight (MALDI TOF) mass spectrometry has become a promising alternative for high-throughput drug discovery as new instruments offer high speed, flexibility and sensitivity, and the ability to measure physiological substrates label free. Here we developed and applied high-throughput MALDI TOF mass spectrometry to identify inhibitors of the salt-inducible kinase (SIK) family, which are interesting drug targets in the field of inflammatory disease as they control production of the anti-inflammatory cytokine interleukin-10 (IL-10) in macrophages. Using peptide substrates in in vitro kinase assays, we can show that hit identification of the MALDI TOF kinase assay correlates with indirect ADP-Hunter kinase assays. Moreover, we can show that both techniques generate comparable IC 50 data for a number of hit compounds and known inhibitors of SIK kinases. We further take these inhibitors to a fluorescence-based cellular assay using the SIK activity-dependent translocation of CRTC3 into the nucleus, thereby providing a complete assay pipeline for the identification of SIK kinase inhibitors in vitro and in cells. Our data demonstrate that MALDI TOF mass spectrometry is fully applicable to high-throughput kinase screening, providing label-free data comparable to that of current high-throughput fluorescence assays.

  5. Label-free detection of cellular drug responses by high-throughput bright-field imaging and machine learning.

    Science.gov (United States)

    Kobayashi, Hirofumi; Lei, Cheng; Wu, Yi; Mao, Ailin; Jiang, Yiyue; Guo, Baoshan; Ozeki, Yasuyuki; Goda, Keisuke

    2017-09-29

    In the last decade, high-content screening based on multivariate single-cell imaging has been proven effective in drug discovery to evaluate drug-induced phenotypic variations. Unfortunately, this method inherently requires fluorescent labeling which has several drawbacks. Here we present a label-free method for evaluating cellular drug responses only by high-throughput bright-field imaging with the aid of machine learning algorithms. Specifically, we performed high-throughput bright-field imaging of numerous drug-treated and -untreated cells (N = ~240,000) by optofluidic time-stretch microscopy with high throughput up to 10,000 cells/s and applied machine learning to the cell images to identify their morphological variations which are too subtle for human eyes to detect. Consequently, we achieved a high accuracy of 92% in distinguishing drug-treated and -untreated cells without the need for labeling. Furthermore, we also demonstrated that dose-dependent, drug-induced morphological change from different experiments can be inferred from the classification accuracy of a single classification model. Our work lays the groundwork for label-free drug screening in pharmaceutical science and industry.

  6. A target-based high throughput screen yields Trypanosoma brucei hexokinase small molecule inhibitors with antiparasitic activity.

    Directory of Open Access Journals (Sweden)

    Elizabeth R Sharlow

    2010-04-01

    Full Text Available The parasitic protozoan Trypanosoma brucei utilizes glycolysis exclusively for ATP production during infection of the mammalian host. The first step in this metabolic pathway is mediated by hexokinase (TbHK, an enzyme essential to the parasite that transfers the gamma-phospho of ATP to a hexose. Here we describe the identification and confirmation of novel small molecule inhibitors of bacterially expressed TbHK1, one of two TbHKs expressed by T. brucei, using a high throughput screening assay.Exploiting optimized high throughput screening assay procedures, we interrogated 220,233 unique compounds and identified 239 active compounds from which ten small molecules were further characterized. Computation chemical cluster analyses indicated that six compounds were structurally related while the remaining four compounds were classified as unrelated or singletons. All ten compounds were approximately 20-17,000-fold more potent than lonidamine, a previously identified TbHK1 inhibitor. Seven compounds inhibited T. brucei blood stage form parasite growth (0.03

  7. A high throughput architecture for a low complexity soft-output demapping algorithm

    Science.gov (United States)

    Ali, I.; Wasenmüller, U.; Wehn, N.

    2015-11-01

    Iterative channel decoders such as Turbo-Code and LDPC decoders show exceptional performance and therefore they are a part of many wireless communication receivers nowadays. These decoders require a soft input, i.e., the logarithmic likelihood ratio (LLR) of the received bits with a typical quantization of 4 to 6 bits. For computing the LLR values from a received complex symbol, a soft demapper is employed in the receiver. The implementation cost of traditional soft-output demapping methods is relatively large in high order modulation systems, and therefore low complexity demapping algorithms are indispensable in low power receivers. In the presence of multiple wireless communication standards where each standard defines multiple modulation schemes, there is a need to have an efficient demapper architecture covering all the flexibility requirements of these standards. Another challenge associated with hardware implementation of the demapper is to achieve a very high throughput in double iterative systems, for instance, MIMO and Code-Aided Synchronization. In this paper, we present a comprehensive communication and hardware performance evaluation of low complexity soft-output demapping algorithms to select the best algorithm for implementation. The main goal of this work is to design a high throughput, flexible, and area efficient architecture. We describe architectures to execute the investigated algorithms. We implement these architectures on a FPGA device to evaluate their hardware performance. The work has resulted in a hardware architecture based on the figured out best low complexity algorithm delivering a high throughput of 166 Msymbols/second for Gray mapped 16-QAM modulation on Virtex-5. This efficient architecture occupies only 127 slice registers, 248 slice LUTs and 2 DSP48Es.

  8. Multilayer Porous Crucibles for the High Throughput Salt Separation from Uranium Deposits

    International Nuclear Information System (INIS)

    Kwon, S. W.; Park, K. M.; Kim, J. G.; Kim, I. T.; Seo, B. K.; Moon, J. G.

    2013-01-01

    Solid cathode processing is necessary to separate the salt from the cathode since the uranium deposit in a solid cathode contains electrolyte salt. A physical separation process, such as a distillation separation, is more attractive than a chemical or dissolution process because physical processes generate much less secondary process. Distillation process was employed for the cathode processsing due to the advantages of minimal generation of secondary waste, compact unit process, simple and low cost equipment. The basis for vacuum distillation separation is the difference in vapor pressures between salt and uranium. A solid cathode deposit is heated in a heating region and salt vaporizes, while nonvolatile uranium remains behind. It is very important to increase the throughput of the salt separation system owing to the high uranium content of spent nuclear fuel and high salt fraction of uranium dendrites. The evaporation rate of the LiCl-KCl eutectic salt in vacuum distiller is not so high to come up with the generation capacity of uranium dendrites in an electro-refiner. Therefore, a wide evaporation area or high distillation temperature is necessary for the successful salt separation. In this study, it was attempted to enlarge a throughput of the salt distiller with a multilayer porous crucibles for the separation of adhered salt in the uranium deposits generated from the electrorefiner. The feasibility of the porous crucibles was tested by the salt distillation experiments. In this study, the salt distiller with multilayer porous crucibles was proposed and the feasibility of liquid salt separation was examined to increase a throughput. It was found that the effective separation of salt from uranium deposits was possible by the multilayer porous crucibles

  9. Multilayer Porous Crucibles for the High Throughput Salt Separation from Uranium Deposits

    Energy Technology Data Exchange (ETDEWEB)

    Kwon, S. W.; Park, K. M.; Kim, J. G.; Kim, I. T.; Seo, B. K.; Moon, J. G. [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2013-05-15

    Solid cathode processing is necessary to separate the salt from the cathode since the uranium deposit in a solid cathode contains electrolyte salt. A physical separation process, such as a distillation separation, is more attractive than a chemical or dissolution process because physical processes generate much less secondary process. Distillation process was employed for the cathode processsing due to the advantages of minimal generation of secondary waste, compact unit process, simple and low cost equipment. The basis for vacuum distillation separation is the difference in vapor pressures between salt and uranium. A solid cathode deposit is heated in a heating region and salt vaporizes, while nonvolatile uranium remains behind. It is very important to increase the throughput of the salt separation system owing to the high uranium content of spent nuclear fuel and high salt fraction of uranium dendrites. The evaporation rate of the LiCl-KCl eutectic salt in vacuum distiller is not so high to come up with the generation capacity of uranium dendrites in an electro-refiner. Therefore, a wide evaporation area or high distillation temperature is necessary for the successful salt separation. In this study, it was attempted to enlarge a throughput of the salt distiller with a multilayer porous crucibles for the separation of adhered salt in the uranium deposits generated from the electrorefiner. The feasibility of the porous crucibles was tested by the salt distillation experiments. In this study, the salt distiller with multilayer porous crucibles was proposed and the feasibility of liquid salt separation was examined to increase a throughput. It was found that the effective separation of salt from uranium deposits was possible by the multilayer porous crucibles.

  10. Retrofit Strategies for Incorporating Xenobiotic Metabolism into High Throughput Screening Assays (EMGS)

    Science.gov (United States)

    The US EPA’s ToxCast program is designed to assess chemical perturbations of molecular and cellular endpoints using a variety of high-throughput screening (HTS) assays. However, existing HTS assays have limited or no xenobiotic metabolism which could lead to a mischaracterization...

  11. Dimensioning storage and computing clusters for efficient High Throughput Computing

    CERN Multimedia

    CERN. Geneva

    2012-01-01

    Scientific experiments are producing huge amounts of data, and they continue increasing the size of their datasets and the total volume of data. These data are then processed by researchers belonging to large scientific collaborations, with the Large Hadron Collider being a good example. The focal point of Scientific Data Centres has shifted from coping efficiently with PetaByte scale storage to deliver quality data processing throughput. The dimensioning of the internal components in High Throughput Computing (HTC) data centers is of crucial importance to cope with all the activities demanded by the experiments, both the online (data acceptance) and the offline (data processing, simulation and user analysis). This requires a precise setup involving disk and tape storage services, a computing cluster and the internal networking to prevent bottlenecks, overloads and undesired slowness that lead to losses cpu cycles and batch jobs failures. In this paper we point out relevant features for running a successful s...

  12. Dimensioning storage and computing clusters for efficient high throughput computing

    International Nuclear Information System (INIS)

    Accion, E; Bria, A; Bernabeu, G; Caubet, M; Delfino, M; Espinal, X; Merino, G; Lopez, F; Martinez, F; Planas, E

    2012-01-01

    Scientific experiments are producing huge amounts of data, and the size of their datasets and total volume of data continues increasing. These data are then processed by researchers belonging to large scientific collaborations, with the Large Hadron Collider being a good example. The focal point of scientific data centers has shifted from efficiently coping with PetaByte scale storage to deliver quality data processing throughput. The dimensioning of the internal components in High Throughput Computing (HTC) data centers is of crucial importance to cope with all the activities demanded by the experiments, both the online (data acceptance) and the offline (data processing, simulation and user analysis). This requires a precise setup involving disk and tape storage services, a computing cluster and the internal networking to prevent bottlenecks, overloads and undesired slowness that lead to losses cpu cycles and batch jobs failures. In this paper we point out relevant features for running a successful data storage and processing service in an intensive HTC environment.

  13. High-throughput gene expression profiling of memory differentiation in primary human T cells

    Directory of Open Access Journals (Sweden)

    Russell Kate

    2008-08-01

    Full Text Available Abstract Background The differentiation of naive T and B cells into memory lymphocytes is essential for immunity to pathogens. Therapeutic manipulation of this cellular differentiation program could improve vaccine efficacy and the in vitro expansion of memory cells. However, chemical screens to identify compounds that induce memory differentiation have been limited by 1 the lack of reporter-gene or functional assays that can distinguish naive and memory-phenotype T cells at high throughput and 2 a suitable cell-line representative of naive T cells. Results Here, we describe a method for gene-expression based screening that allows primary naive and memory-phenotype lymphocytes to be discriminated based on complex genes signatures corresponding to these differentiation states. We used ligation-mediated amplification and a fluorescent, bead-based detection system to quantify simultaneously 55 transcripts representing naive and memory-phenotype signatures in purified populations of human T cells. The use of a multi-gene panel allowed better resolution than any constituent single gene. The method was precise, correlated well with Affymetrix microarray data, and could be easily scaled up for high-throughput. Conclusion This method provides a generic solution for high-throughput differentiation screens in primary human T cells where no single-gene or functional assay is available. This screening platform will allow the identification of small molecules, genes or soluble factors that direct memory differentiation in naive human lymphocytes.

  14. Development of a high-throughput microscale cell disruption platform for Pichia pastoris in rapid bioprocess design.

    Science.gov (United States)

    Bláha, Benjamin A F; Morris, Stephen A; Ogonah, Olotu W; Maucourant, Sophie; Crescente, Vincenzo; Rosenberg, William; Mukhopadhyay, Tarit K

    2018-01-01

    The time and cost benefits of miniaturized fermentation platforms can only be gained by employing complementary techniques facilitating high-throughput at small sample volumes. Microbial cell disruption is a major bottleneck in experimental throughput and is often restricted to large processing volumes. Moreover, for rigid yeast species, such as Pichia pastoris, no effective high-throughput disruption methods exist. The development of an automated, miniaturized, high-throughput, noncontact, scalable platform based on adaptive focused acoustics (AFA) to disrupt P. pastoris and recover intracellular heterologous protein is described. Augmented modes of AFA were established by investigating vessel designs and a novel enzymatic pretreatment step. Three different modes of AFA were studied and compared to the performance high-pressure homogenization. For each of these modes of cell disruption, response models were developed to account for five different performance criteria. Using multiple responses not only demonstrated that different operating parameters are required for different response optima, with highest product purity requiring suboptimal values for other criteria, but also allowed for AFA-based methods to mimic large-scale homogenization processes. These results demonstrate that AFA-mediated cell disruption can be used for a wide range of applications including buffer development, strain selection, fermentation process development, and whole bioprocess integration. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 34:130-140, 2018. © 2017 American Institute of Chemical Engineers.

  15. High-Throughput Process Development for Biopharmaceuticals.

    Science.gov (United States)

    Shukla, Abhinav A; Rameez, Shahid; Wolfe, Leslie S; Oien, Nathan

    2017-11-14

    The ability to conduct multiple experiments in parallel significantly reduces the time that it takes to develop a manufacturing process for a biopharmaceutical. This is particularly significant before clinical entry, because process development and manufacturing are on the "critical path" for a drug candidate to enter clinical development. High-throughput process development (HTPD) methodologies can be similarly impactful during late-stage development, both for developing the final commercial process as well as for process characterization and scale-down validation activities that form a key component of the licensure filing package. This review examines the current state of the art for HTPD methodologies as they apply to cell culture, downstream purification, and analytical techniques. In addition, we provide a vision of how HTPD activities across all of these spaces can integrate to create a rapid process development engine that can accelerate biopharmaceutical drug development. Graphical Abstract.

  16. High Throughput Line-of-Sight MIMO Systems for Next Generation Backhaul Applications

    Science.gov (United States)

    Song, Xiaohang; Cvetkovski, Darko; Hälsig, Tim; Rave, Wolfgang; Fettweis, Gerhard; Grass, Eckhard; Lankl, Berthold

    2017-09-01

    The evolution to ultra-dense next generation networks requires a massive increase in throughput and deployment flexibility. Therefore, novel wireless backhaul solutions that can support these demands are needed. In this work we present an approach for a millimeter wave line-of-sight MIMO backhaul design, targeting transmission rates in the order of 100 Gbit/s. We provide theoretical foundations for the concept showcasing its potential, which are confirmed through channel measurements. Furthermore, we provide insights into the system design with respect to antenna array setup, baseband processing, synchronization, and channel equalization. Implementation in a 60 GHz demonstrator setup proves the feasibility of the system concept for high throughput backhauling in next generation networks.

  17. Operational evaluation of high-throughput community-based mass prophylaxis using Just-in-time training.

    Science.gov (United States)

    Spitzer, James D; Hupert, Nathaniel; Duckart, Jonathan; Xiong, Wei

    2007-01-01

    Community-based mass prophylaxis is a core public health operational competency, but staffing needs may overwhelm the local trained health workforce. Just-in-time (JIT) training of emergency staff and computer modeling of workforce requirements represent two complementary approaches to address this logistical problem. Multnomah County, Oregon, conducted a high-throughput point of dispensing (POD) exercise to test JIT training and computer modeling to validate POD staffing estimates. The POD had 84% non-health-care worker staff and processed 500 patients per hour. Post-exercise modeling replicated observed staff utilization levels and queue formation, including development and amelioration of a large medical evaluation queue caused by lengthy processing times and understaffing in the first half-hour of the exercise. The exercise confirmed the feasibility of using JIT training for high-throughput antibiotic dispensing clinics staffed largely by nonmedical professionals. Patient processing times varied over the course of the exercise, with important implications for both staff reallocation and future POD modeling efforts. Overall underutilization of staff revealed the opportunity for greater efficiencies and even higher future throughputs.

  18. Risk-based high-throughput chemical screening and prioritization using exposure models and in vitro bioactivity assays

    DEFF Research Database (Denmark)

    Shin, Hyeong-Moo; Ernstoff, Alexi; Arnot, Jon

    2015-01-01

    We present a risk-based high-throughput screening (HTS) method to identify chemicals for potential health concerns or for which additional information is needed. The method is applied to 180 organic chemicals as a case study. We first obtain information on how the chemical is used and identify....../oral contact, or dermal exposure. The method provides high-throughput estimates of exposure and important input for decision makers to identify chemicals of concern for further evaluation with additional information or more refined models....

  19. Life in the fast lane: high-throughput chemistry for lead generation and optimisation.

    Science.gov (United States)

    Hunter, D

    2001-01-01

    The pharmaceutical industry has come under increasing pressure due to regulatory restrictions on the marketing and pricing of drugs, competition, and the escalating costs of developing new drugs. These forces can be addressed by the identification of novel targets, reductions in the development time of new drugs, and increased productivity. Emphasis has been placed on identifying and validating new targets and on lead generation: the response from industry has been very evident in genomics and high throughput screening, where new technologies have been applied, usually coupled with a high degree of automation. The combination of numerous new potential biological targets and the ability to screen large numbers of compounds against many of these targets has generated the need for large diverse compound collections. To address this requirement, high-throughput chemistry has become an integral part of the drug discovery process. Copyright 2002 Wiley-Liss, Inc.

  20. High-throughput computational methods and software for quantitative trait locus (QTL) mapping

    NARCIS (Netherlands)

    Arends, Danny

    2014-01-01

    De afgelopen jaren zijn vele nieuwe technologieen zoals Tiling arrays en High throughput DNA sequencing een belangrijke rol gaan spelen binnen het onderzoeksveld van de systeem genetica. Voor onderzoekers is het extreem belangrijk om te begrijpen dat deze methodes hun manier van werken zullen gaan

  1. Insights into Sonogashira cross-coupling by high-throughput kinetics and descriptor modeling

    NARCIS (Netherlands)

    an der Heiden, M.R.; Plenio, H.; Immel, S.; Burello, E.; Rothenberg, G.; Hoefsloot, H.C.J.

    2008-01-01

    A method is presented for the high-throughput monitoring of reaction kinetics in homogeneous catalysis, running up to 25 coupling reactions in a single reaction vessel. This method is demonstrated and validated on the Sonogashira reaction, analyzing the kinetics for almost 500 coupling reactions.

  2. Development of rapid high throughput biodosimetry tools for radiological triage

    International Nuclear Information System (INIS)

    Balajee, Adayabalam S.; Escalona, Maria; Smith, Tammy; Ryan, Terri; Dainiak, Nicholas

    2018-01-01

    Accidental or intentional radiological or nuclear (R/N) disasters constitute a major threat around the globe that can affect several tens, hundreds and thousands of humans. Currently available cytogenetic biodosimeters are time consuming and laborious to perform making them impractical for triage scenarios. Therefore, it is imperative to develop high throughput techniques which will enable timely assessment of personalized dose for making an appropriate 'life-saving' clinical decision

  3. Intel: High Throughput Computing Collaboration: A CERN openlab / Intel collaboration

    CERN Multimedia

    CERN. Geneva

    2015-01-01

    The Intel/CERN High Throughput Computing Collaboration studies the application of upcoming Intel technologies to the very challenging environment of the LHC trigger and data-acquisition systems. These systems will need to transport and process many terabits of data every second, in some cases with tight latency constraints. Parallelisation and tight integration of accelerators and classical CPU via Intel's OmniPath fabric are the key elements in this project.

  4. Printing Proteins as Microarrays for High-Throughput Function Determination

    Science.gov (United States)

    MacBeath, Gavin; Schreiber, Stuart L.

    2000-09-01

    Systematic efforts are currently under way to construct defined sets of cloned genes for high-throughput expression and purification of recombinant proteins. To facilitate subsequent studies of protein function, we have developed miniaturized assays that accommodate extremely low sample volumes and enable the rapid, simultaneous processing of thousands of proteins. A high-precision robot designed to manufacture complementary DNA microarrays was used to spot proteins onto chemically derivatized glass slides at extremely high spatial densities. The proteins attached covalently to the slide surface yet retained their ability to interact specifically with other proteins, or with small molecules, in solution. Three applications for protein microarrays were demonstrated: screening for protein-protein interactions, identifying the substrates of protein kinases, and identifying the protein targets of small molecules.

  5. A high throughput biochemical fluorometric method for measuring lipid peroxidation in HDL.

    Directory of Open Access Journals (Sweden)

    Theodoros Kelesidis

    Full Text Available Current cell-based assays for determining the functional properties of high-density lipoproteins (HDL have limitations. We report here the development of a new, robust fluorometric cell-free biochemical assay that measures HDL lipid peroxidation (HDLox based on the oxidation of the fluorochrome Amplex Red. HDLox correlated with previously validated cell-based (r = 0.47, p<0.001 and cell-free assays (r = 0.46, p<0.001. HDLox distinguished dysfunctional HDL in established animal models of atherosclerosis and Human Immunodeficiency Virus (HIV patients. Using an immunoaffinity method for capturing HDL, we demonstrate the utility of this novel assay for measuring HDLox in a high throughput format. Furthermore, HDLox correlated significantly with measures of cardiovascular diseases including carotid intima media thickness (r = 0.35, p<0.01 and subendocardial viability ratio (r = -0.21, p = 0.05 and physiological parameters such as metabolic and anthropometric parameters (p<0.05. In conclusion, we report the development of a new fluorometric method that offers a reproducible and rapid means for determining HDL function/quality that is suitable for high throughput implementation.

  6. Introducing Bayesian thinking to high-throughput screening for false-negative rate estimation.

    Science.gov (United States)

    Wei, Xin; Gao, Lin; Zhang, Xiaolei; Qian, Hong; Rowan, Karen; Mark, David; Peng, Zhengwei; Huang, Kuo-Sen

    2013-10-01

    High-throughput screening (HTS) has been widely used to identify active compounds (hits) that bind to biological targets. Because of cost concerns, the comprehensive screening of millions of compounds is typically conducted without replication. Real hits that fail to exhibit measurable activity in the primary screen due to random experimental errors will be lost as false-negatives. Conceivably, the projected false-negative rate is a parameter that reflects screening quality. Furthermore, it can be used to guide the selection of optimal numbers of compounds for hit confirmation. Therefore, a method that predicts false-negative rates from the primary screening data is extremely valuable. In this article, we describe the implementation of a pilot screen on a representative fraction (1%) of the screening library in order to obtain information about assay variability as well as a preliminary hit activity distribution profile. Using this training data set, we then developed an algorithm based on Bayesian logic and Monte Carlo simulation to estimate the number of true active compounds and potential missed hits from the full library screen. We have applied this strategy to five screening projects. The results demonstrate that this method produces useful predictions on the numbers of false negatives.

  7. Benchmarking Ligand-Based Virtual High-Throughput Screening with the PubChem Database

    Directory of Open Access Journals (Sweden)

    Mariusz Butkiewicz

    2013-01-01

    Full Text Available With the rapidly increasing availability of High-Throughput Screening (HTS data in the public domain, such as the PubChem database, methods for ligand-based computer-aided drug discovery (LB-CADD have the potential to accelerate and reduce the cost of probe development and drug discovery efforts in academia. We assemble nine data sets from realistic HTS campaigns representing major families of drug target proteins for benchmarking LB-CADD methods. Each data set is public domain through PubChem and carefully collated through confirmation screens validating active compounds. These data sets provide the foundation for benchmarking a new cheminformatics framework BCL::ChemInfo, which is freely available for non-commercial use. Quantitative structure activity relationship (QSAR models are built using Artificial Neural Networks (ANNs, Support Vector Machines (SVMs, Decision Trees (DTs, and Kohonen networks (KNs. Problem-specific descriptor optimization protocols are assessed including Sequential Feature Forward Selection (SFFS and various information content measures. Measures of predictive power and confidence are evaluated through cross-validation, and a consensus prediction scheme is tested that combines orthogonal machine learning algorithms into a single predictor. Enrichments ranging from 15 to 101 for a TPR cutoff of 25% are observed.

  8. A High-Throughput SU-8Microfluidic Magnetic Bead Separator

    DEFF Research Database (Denmark)

    Bu, Minqiang; Christensen, T. B.; Smistrup, Kristian

    2007-01-01

    We present a novel microfluidic magnetic bead separator based on SU-8 fabrication technique for high through-put applications. The experimental results show that magnetic beads can be captured at an efficiency of 91 % and 54 % at flow rates of 1 mL/min and 4 mL/min, respectively. Integration...... of soft magnetic elements in the chip leads to a slightly higher capturing efficiency and a more uniform distribution of captured beads over the separation chamber than the system without soft magnetic elements....

  9. A high-throughput and quantitative method to assess the mutagenic potential of translesion DNA synthesis

    Science.gov (United States)

    Taggart, David J.; Camerlengo, Terry L.; Harrison, Jason K.; Sherrer, Shanen M.; Kshetry, Ajay K.; Taylor, John-Stephen; Huang, Kun; Suo, Zucai

    2013-01-01

    Cellular genomes are constantly damaged by endogenous and exogenous agents that covalently and structurally modify DNA to produce DNA lesions. Although most lesions are mended by various DNA repair pathways in vivo, a significant number of damage sites persist during genomic replication. Our understanding of the mutagenic outcomes derived from these unrepaired DNA lesions has been hindered by the low throughput of existing sequencing methods. Therefore, we have developed a cost-effective high-throughput short oligonucleotide sequencing assay that uses next-generation DNA sequencing technology for the assessment of the mutagenic profiles of translesion DNA synthesis catalyzed by any error-prone DNA polymerase. The vast amount of sequencing data produced were aligned and quantified by using our novel software. As an example, the high-throughput short oligonucleotide sequencing assay was used to analyze the types and frequencies of mutations upstream, downstream and at a site-specifically placed cis–syn thymidine–thymidine dimer generated individually by three lesion-bypass human Y-family DNA polymerases. PMID:23470999

  10. The French press: a repeatable and high-throughput approach to exercising zebrafish (Danio rerio).

    Science.gov (United States)

    Usui, Takuji; Noble, Daniel W A; O'Dea, Rose E; Fangmeier, Melissa L; Lagisz, Malgorzata; Hesselson, Daniel; Nakagawa, Shinichi

    2018-01-01

    Zebrafish are increasingly used as a vertebrate model organism for various traits including swimming performance, obesity and metabolism, necessitating high-throughput protocols to generate standardized phenotypic information. Here, we propose a novel and cost-effective method for exercising zebrafish, using a coffee plunger and magnetic stirrer. To demonstrate the use of this method, we conducted a pilot experiment to show that this simple system provides repeatable estimates of maximal swim performance (intra-class correlation [ICC] = 0.34-0.41) and observe that exercise training of zebrafish on this system significantly increases their maximum swimming speed. We propose this high-throughput and reproducible system as an alternative to traditional linear chamber systems for exercising zebrafish and similarly sized fishes.

  11. The University of Kansas High-Throughput Screening Laboratory. Part II: enabling collaborative drug-discovery partnerships through cutting-edge screening technology.

    Science.gov (United States)

    McDonald, Peter R; Roy, Anuradha; Chaguturu, Rathnam

    2011-07-01

    The University of Kansas High-Throughput Screening (KU HTS) core is a state-of-the-art drug-discovery facility with an entrepreneurial open-service policy, which provides centralized resources supporting public- and private-sector research initiatives. The KU HTS core was established in 2002 at the University of Kansas with support from an NIH grant and the state of Kansas. It collaborates with investigators from national and international academic, nonprofit and pharmaceutical organizations in executing HTS-ready assay development and screening of chemical libraries for target validation, probe selection, hit identification and lead optimization. This is part two of a contribution from the KU HTS laboratory.

  12. A High-Throughput Antibody-Based Microarray Typing Platform

    Directory of Open Access Journals (Sweden)

    Ashan Perera

    2013-05-01

    Full Text Available Many rapid methods have been developed for screening foods for the presence of pathogenic microorganisms. Rapid methods that have the additional ability to identify microorganisms via multiplexed immunological recognition have the potential for classification or typing of microbial contaminants thus facilitating epidemiological investigations that aim to identify outbreaks and trace back the contamination to its source. This manuscript introduces a novel, high throughput typing platform that employs microarrayed multiwell plate substrates and laser-induced fluorescence of the nucleic acid intercalating dye/stain SYBR Gold for detection of antibody-captured bacteria. The aim of this study was to use this platform for comparison of different sets of antibodies raised against the same pathogens as well as demonstrate its potential effectiveness for serotyping. To that end, two sets of antibodies raised against each of the “Big Six” non-O157 Shiga toxin-producing E. coli (STEC as well as E. coli O157:H7 were array-printed into microtiter plates, and serial dilutions of the bacteria were added and subsequently detected. Though antibody specificity was not sufficient for the development of an STEC serotyping method, the STEC antibody sets performed reasonably well exhibiting that specificity increased at lower capture antibody concentrations or, conversely, at lower bacterial target concentrations. The favorable results indicated that with sufficiently selective and ideally concentrated sets of biorecognition elements (e.g., antibodies or aptamers, this high-throughput platform can be used to rapidly type microbial isolates derived from food samples within ca. 80 min of total assay time. It can also potentially be used to detect the pathogens from food enrichments and at least serve as a platform for testing antibodies.

  13. GlycoExtractor: a web-based interface for high throughput processing of HPLC-glycan data.

    Science.gov (United States)

    Artemenko, Natalia V; Campbell, Matthew P; Rudd, Pauline M

    2010-04-05

    Recently, an automated high-throughput HPLC platform has been developed that can be used to fully sequence and quantify low concentrations of N-linked sugars released from glycoproteins, supported by an experimental database (GlycoBase) and analytical tools (autoGU). However, commercial packages that support the operation of HPLC instruments and data storage lack platforms for the extraction of large volumes of data. The lack of resources and agreed formats in glycomics is now a major limiting factor that restricts the development of bioinformatic tools and automated workflows for high-throughput HPLC data analysis. GlycoExtractor is a web-based tool that interfaces with a commercial HPLC database/software solution to facilitate the extraction of large volumes of processed glycan profile data (peak number, peak areas, and glucose unit values). The tool allows the user to export a series of sample sets to a set of file formats (XML, JSON, and CSV) rather than a collection of disconnected files. This approach not only reduces the amount of manual refinement required to export data into a suitable format for data analysis but also opens the field to new approaches for high-throughput data interpretation and storage, including biomarker discovery and validation and monitoring of online bioprocessing conditions for next generation biotherapeutics.

  14. High-throughput measurement of polymer film thickness using optical dyes

    Science.gov (United States)

    Grunlan, Jaime C.; Mehrabi, Ali R.; Ly, Tien

    2005-01-01

    Optical dyes were added to polymer solutions in an effort to create a technique for high-throughput screening of dry polymer film thickness. Arrays of polystyrene films, cast from a toluene solution, containing methyl red or solvent green were used to demonstrate the feasibility of this technique. Measurements of the peak visible absorbance of each film were converted to thickness using the Beer-Lambert relationship. These absorbance-based thickness calculations agreed within 10% of thickness measured using a micrometer for polystyrene films that were 10-50 µm. At these thicknesses it is believed that the absorbance values are actually more accurate. At least for this solvent-based system, thickness was shown to be accurately measured in a high-throughput manner that could potentially be applied to other equivalent systems. Similar water-based films made with poly(sodium 4-styrenesulfonate) dyed with malachite green oxalate or congo red did not show the same level of agreement with the micrometer measurements. Extensive phase separation between polymer and dye resulted in inflated absorbance values and calculated thickness that was often more than 25% greater than that measured with the micrometer. Only at thicknesses below 15 µm could reasonable accuracy be achieved for the water-based films.

  15. Multiplex High-Throughput Targeted Proteomic Assay To Identify Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Baud, Anna; Wessely, Frank; Mazzacuva, Francesca; McCormick, James; Camuzeaux, Stephane; Heywood, Wendy E; Little, Daniel; Vowles, Jane; Tuefferd, Marianne; Mosaku, Olukunbi; Lako, Majlinda; Armstrong, Lyle; Webber, Caleb; Cader, M Zameel; Peeters, Pieter; Gissen, Paul; Cowley, Sally A; Mills, Kevin

    2017-02-21

    Induced pluripotent stem cells have great potential as a human model system in regenerative medicine, disease modeling, and drug screening. However, their use in medical research is hampered by laborious reprogramming procedures that yield low numbers of induced pluripotent stem cells. For further applications in research, only the best, competent clones should be used. The standard assays for pluripotency are based on genomic approaches, which take up to 1 week to perform and incur significant cost. Therefore, there is a need for a rapid and cost-effective assay able to distinguish between pluripotent and nonpluripotent cells. Here, we describe a novel multiplexed, high-throughput, and sensitive peptide-based multiple reaction monitoring mass spectrometry assay, allowing for the identification and absolute quantitation of multiple core transcription factors and pluripotency markers. This assay provides simpler and high-throughput classification into either pluripotent or nonpluripotent cells in 7 min analysis while being more cost-effective than conventional genomic tests.

  16. Application of high-throughput DNA sequencing in phytopathology.

    Science.gov (United States)

    Studholme, David J; Glover, Rachel H; Boonham, Neil

    2011-01-01

    The new sequencing technologies are already making a big impact in academic research on medically important microbes and may soon revolutionize diagnostics, epidemiology, and infection control. Plant pathology also stands to gain from exploiting these opportunities. This manuscript reviews some applications of these high-throughput sequencing methods that are relevant to phytopathology, with emphasis on the associated computational and bioinformatics challenges and their solutions. Second-generation sequencing technologies have recently been exploited in genomics of both prokaryotic and eukaryotic plant pathogens. They are also proving to be useful in diagnostics, especially with respect to viruses. Copyright © 2011 by Annual Reviews. All rights reserved.

  17. High-throughput ab-initio dilute solute diffusion database.

    Science.gov (United States)

    Wu, Henry; Mayeshiba, Tam; Morgan, Dane

    2016-07-19

    We demonstrate automated generation of diffusion databases from high-throughput density functional theory (DFT) calculations. A total of more than 230 dilute solute diffusion systems in Mg, Al, Cu, Ni, Pd, and Pt host lattices have been determined using multi-frequency diffusion models. We apply a correction method for solute diffusion in alloys using experimental and simulated values of host self-diffusivity. We find good agreement with experimental solute diffusion data, obtaining a weighted activation barrier RMS error of 0.176 eV when excluding magnetic solutes in non-magnetic alloys. The compiled database is the largest collection of consistently calculated ab-initio solute diffusion data in the world.

  18. Development and validation of a 48-target analytical method for high-throughput monitoring of genetically modified organisms.

    Science.gov (United States)

    Li, Xiaofei; Wu, Yuhua; Li, Jun; Li, Yunjing; Long, Likun; Li, Feiwu; Wu, Gang

    2015-01-05

    The rapid increase in the number of genetically modified (GM) varieties has led to a demand for high-throughput methods to detect genetically modified organisms (GMOs). We describe a new dynamic array-based high throughput method to simultaneously detect 48 targets in 48 samples on a Fludigm system. The test targets included species-specific genes, common screening elements, most of the Chinese-approved GM events, and several unapproved events. The 48 TaqMan assays successfully amplified products from both single-event samples and complex samples with a GMO DNA amount of 0.05 ng, and displayed high specificity. To improve the sensitivity of detection, a preamplification step for 48 pooled targets was added to enrich the amount of template before performing dynamic chip assays. This dynamic chip-based method allowed the synchronous high-throughput detection of multiple targets in multiple samples. Thus, it represents an efficient, qualitative method for GMO multi-detection.

  19. MSP-HTPrimer: a high-throughput primer design tool to improve assay design for DNA methylation analysis in epigenetics.

    Science.gov (United States)

    Pandey, Ram Vinay; Pulverer, Walter; Kallmeyer, Rainer; Beikircher, Gabriel; Pabinger, Stephan; Kriegner, Albert; Weinhäusel, Andreas

    2016-01-01

    Bisulfite (BS) conversion-based and methylation-sensitive restriction enzyme (MSRE)-based PCR methods have been the most commonly used techniques for locus-specific DNA methylation analysis. However, both methods have advantages and limitations. Thus, an integrated approach would be extremely useful to quantify the DNA methylation status successfully with great sensitivity and specificity. Designing specific and optimized primers for target regions is the most critical and challenging step in obtaining the adequate DNA methylation results using PCR-based methods. Currently, no integrated, optimized, and high-throughput methylation-specific primer design software methods are available for both BS- and MSRE-based methods. Therefore an integrated, powerful, and easy-to-use methylation-specific primer design pipeline with great accuracy and success rate will be very useful. We have developed a new web-based pipeline, called MSP-HTPrimer, to design primers pairs for MSP, BSP, pyrosequencing, COBRA, and MSRE assays on both genomic strands. First, our pipeline converts all target sequences into bisulfite-treated templates for both forward and reverse strand and designs all possible primer pairs, followed by filtering for single nucleotide polymorphisms (SNPs) and known repeat regions. Next, each primer pairs are annotated with the upstream and downstream RefSeq genes, CpG island, and cut sites (for COBRA and MSRE). Finally, MSP-HTPrimer selects specific primers from both strands based on custom and user-defined hierarchical selection criteria. MSP-HTPrimer produces a primer pair summary output table in TXT and HTML format for display and UCSC custom tracks for resulting primer pairs in GTF format. MSP-HTPrimer is an integrated, web-based, and high-throughput pipeline and has no limitation on the number and size of target sequences and designs MSP, BSP, pyrosequencing, COBRA, and MSRE assays. It is the only pipeline, which automatically designs primers on both genomic

  20. High-throughput Transcriptome analysis, CAGE and beyond

    KAUST Repository

    Kodzius, Rimantas

    2008-11-25

    1. Current research - PhD work on discovery of new allergens - Postdoctoral work on Transcriptional Start Sites a) Tag based technologies allow higher throughput b) CAGE technology to define promoters c) CAGE data analysis to understand Transcription - Wo

  1. High-throughput Transcriptome analysis, CAGE and beyond

    KAUST Repository

    Kodzius, Rimantas

    2008-01-01

    1. Current research - PhD work on discovery of new allergens - Postdoctoral work on Transcriptional Start Sites a) Tag based technologies allow higher throughput b) CAGE technology to define promoters c) CAGE data analysis to understand Transcription - Wo

  2. Device Centric Throughput and QoS Optimization for IoTsin a Smart Building Using CRN-Techniques

    Directory of Open Access Journals (Sweden)

    Saleem Aslam

    2016-10-01

    Full Text Available The Internet of Things (IoT has gained an incredible importance in the communication and networking industry due to its innovative solutions and advantages in diverse domains. The IoT’ network is a network of smart physical objects: devices, vehicles, buildings, etc. The IoT has a number of applications ranging from smart home, smart surveillance to smart healthcare systems. Since IoT consists of various heterogeneous devices that exhibit different traffic patterns and expect different quality of service (QoS in terms of data rate, bit error rate and the stability index of the channel, therefore, in this paper, we formulated an optimization problem to assign channels to heterogeneous IoT devices within a smart building for the provisioning of their desired QoS. To solve this problem, a novel particle swarm optimization-based algorithm is proposed. Then, exhaustive simulations are carried out to evaluate the performance of the proposed algorithm. Simulation results demonstrate the supremacy of our proposed algorithm over the existing ones in terms of throughput, bit error rate and the stability index of the channel.

  3. OptoDyCE: Automated system for high-throughput all-optical dynamic cardiac electrophysiology

    Science.gov (United States)

    Klimas, Aleksandra; Yu, Jinzhu; Ambrosi, Christina M.; Williams, John C.; Bien, Harold; Entcheva, Emilia

    2016-02-01

    In the last two decades, market were due to cardiac toxicity, where unintended interactions with ion channels disrupt the heart's normal electrical function. Consequently, all new drugs must undergo preclinical testing for cardiac liability, adding to an already expensive and lengthy process. Recognition that proarrhythmic effects often result from drug action on multiple ion channels demonstrates a need for integrative and comprehensive measurements. Additionally, patient-specific therapies relying on emerging technologies employing stem-cell derived cardiomyocytes (e.g. induced pluripotent stem-cell-derived cardiomyocytes, iPSC-CMs) require better screening methods to become practical. However, a high-throughput, cost-effective approach for cellular cardiac electrophysiology has not been feasible. Optical techniques for manipulation and recording provide a contactless means of dynamic, high-throughput testing of cells and tissues. Here, we consider the requirements for all-optical electrophysiology for drug testing, and we implement and validate OptoDyCE, a fully automated system for all-optical cardiac electrophysiology. We demonstrate the high-throughput capabilities using multicellular samples in 96-well format by combining optogenetic actuation with simultaneous fast high-resolution optical sensing of voltage or intracellular calcium. The system can also be implemented using iPSC-CMs and other cell-types by delivery of optogenetic drivers, or through the modular use of dedicated light-sensitive somatic cells in conjunction with non-modified cells. OptoDyCE provides a truly modular and dynamic screening system, capable of fully-automated acquisition of high-content information integral for improved discovery and development of new drugs and biologics, as well as providing a means of better understanding of electrical disturbances in the heart.

  4. High-throughput materials discovery and development: breakthroughs and challenges in the mapping of the materials genome

    Science.gov (United States)

    Buongiorno Nardelli, Marco

    High-Throughput Quantum-Mechanics computation of materials properties by ab initio methods has become the foundation of an effective approach to materials design, discovery and characterization. This data driven approach to materials science currently presents the most promising path to the development of advanced technological materials that could solve or mitigate important social and economic challenges of the 21st century. In particular, the rapid proliferation of computational data on materials properties presents the possibility to complement and extend materials property databases where the experimental data is lacking and difficult to obtain. Enhanced repositories such as AFLOWLIB open novel opportunities for structure discovery and optimization, including uncovering of unsuspected compounds, metastable structures and correlations between various properties. The practical realization of these opportunities depends almost exclusively on the the design of efficient algorithms for electronic structure simulations of realistic material systems beyond the limitations of the current standard theories. In this talk, I will review recent progress in theoretical and computational tools, and in particular, discuss the development and validation of novel functionals within Density Functional Theory and of local basis representations for effective ab-initio tight-binding schemes. Marco Buongiorno Nardelli is a pioneer in the development of computational platforms for theory/data/applications integration rooted in his profound and extensive expertise in the design of electronic structure codes and in his vision for sustainable and innovative software development for high-performance materials simulations. His research activities range from the design and discovery of novel materials for 21st century applications in renewable energy, environment, nano-electronics and devices, the development of advanced electronic structure theories and high-throughput techniques in

  5. Nanosphere Templating Through Controlled Evaporation: A High Throughput Method For Building SERS Substrates

    Science.gov (United States)

    Alexander, Kristen; Hampton, Meredith; Lopez, Rene; Desimone, Joseph

    2009-03-01

    When a pair of noble metal nanoparticles are brought close together, the plasmonic properties of the pair (known as a ``dimer'') give rise to intense electric field enhancements in the interstitial gap. These fields present a simple yet exquisitely sensitive system for performing single molecule surface-enhanced Raman spectroscopy (SM-SERS). Problems associated with current fabrication methods of SERS-active substrates include reproducibility issues, high cost of production and low throughput. In this study, we present a novel method for the high throughput fabrication of high quality SERS substrates. Using a polymer templating technique followed by the placement of thiolated nanoparticles through meniscus force deposition, we are able to fabricate large arrays of identical, uniformly spaced dimers in a quick, reproducible manner. Subsequent theoretical and experimental studies have confirmed the strong dependence of the SERS enhancement on both substrate geometry (e.g. dimer size, shape and gap size) and the polarization of the excitation source.

  6. High-throughput metagenomic technologies for complex microbial community analysis: open and closed formats.

    Science.gov (United States)

    Zhou, Jizhong; He, Zhili; Yang, Yunfeng; Deng, Ye; Tringe, Susannah G; Alvarez-Cohen, Lisa

    2015-01-27

    Understanding the structure, functions, activities and dynamics of microbial communities in natural environments is one of the grand challenges of 21st century science. To address this challenge, over the past decade, numerous technologies have been developed for interrogating microbial communities, of which some are amenable to exploratory work (e.g., high-throughput sequencing and phenotypic screening) and others depend on reference genes or genomes (e.g., phylogenetic and functional gene arrays). Here, we provide a critical review and synthesis of the most commonly applied "open-format" and "closed-format" detection technologies. We discuss their characteristics, advantages, and disadvantages within the context of environmental applications and focus on analysis of complex microbial systems, such as those in soils, in which diversity is high and reference genomes are few. In addition, we discuss crucial issues and considerations associated with applying complementary high-throughput molecular technologies to address important ecological questions. Copyright © 2015 Zhou et al.

  7. Scheduling Algorithms for Maximizing Throughput with Zero-Forcing Beamforming in a MIMO Wireless System

    Science.gov (United States)

    Foronda, Augusto; Ohta, Chikara; Tamaki, Hisashi

    Dirty paper coding (DPC) is a strategy to achieve the region capacity of multiple input multiple output (MIMO) downlink channels and a DPC scheduler is throughput optimal if users are selected according to their queue states and current rates. However, DPC is difficult to implement in practical systems. One solution, zero-forcing beamforming (ZFBF) strategy has been proposed to achieve the same asymptotic sum rate capacity as that of DPC with an exhaustive search over the entire user set. Some suboptimal user group selection schedulers with reduced complexity based on ZFBF strategy (ZFBF-SUS) and proportional fair (PF) scheduling algorithm (PF-ZFBF) have also been proposed to enhance the throughput and fairness among the users, respectively. However, they are not throughput optimal, fairness and throughput decrease if each user queue length is different due to different users channel quality. Therefore, we propose two different scheduling algorithms: a throughput optimal scheduling algorithm (ZFBF-TO) and a reduced complexity scheduling algorithm (ZFBF-RC). Both are based on ZFBF strategy and, at every time slot, the scheduling algorithms have to select some users based on user channel quality, user queue length and orthogonality among users. Moreover, the proposed algorithms have to produce the rate allocation and power allocation for the selected users based on a modified water filling method. We analyze the schedulers complexity and numerical results show that ZFBF-RC provides throughput and fairness improvements compared to the ZFBF-SUS and PF-ZFBF scheduling algorithms.

  8. A production throughput forecasting system in an automated hard disk drive test operation using GRNN

    Energy Technology Data Exchange (ETDEWEB)

    Samattapapong, N.; Afzulpurkar, N.

    2016-07-01

    The goal of this paper is to develop a pragmatic system of a production throughput forecasting system for an automated test operation in a hard drive manufacturing plant. The accurate forecasting result is necessary for the management team to response to any changes in the production processes and the resources allocations. In this study, we design a production throughput forecasting system in an automated test operation in hard drive manufacturing plant. In the proposed system, consists of three main stages. In the first stage, a mutual information method was adopted for selecting the relevant inputs into the forecasting model. In the second stage, a generalized regression neural network (GRNN) was implemented in the forecasting model development phase. Finally, forecasting accuracy was improved by searching the optimal smoothing parameter which selected from comparisons result among three optimization algorithms: particle swarm optimization (PSO), unrestricted search optimization (USO) and interval halving optimization (IHO). The experimental result shows that (1) the developed production throughput forecasting system using GRNN is able to provide forecasted results close to actual values, and to projected the future trends of production throughput in an automated hard disk drive test operation; (2) An IHO algorithm performed as superiority appropriate optimization method than the other two algorithms. (3) Compared with current forecasting system in manufacturing, the results show that the proposed system’s performance is superior to the current system in prediction accuracy and suitable for real-world application. The production throughput volume is a key performance index of hard disk drive manufacturing systems that need to be forecast. Because of the production throughput forecasting result is useful information for management team to respond to any changing in production processes and resources allocation. However, a practically forecasting system for

  9. High-throughput in vivo genotoxicity testing: an automated readout system for the somatic mutation and recombination test (SMART.

    Directory of Open Access Journals (Sweden)

    Benoit Lombardot

    Full Text Available Genotoxicity testing is an important component of toxicity assessment. As illustrated by the European registration, evaluation, authorization, and restriction of chemicals (REACH directive, it concerns all the chemicals used in industry. The commonly used in vivo mammalian tests appear to be ill adapted to tackle the large compound sets involved, due to throughput, cost, and ethical issues. The somatic mutation and recombination test (SMART represents a more scalable alternative, since it uses Drosophila, which develops faster and requires less infrastructure. Despite these advantages, the manual scoring of the hairs on Drosophila wings required for the SMART limits its usage. To overcome this limitation, we have developed an automated SMART readout. It consists of automated imaging, followed by an image analysis pipeline that measures individual wing genotoxicity scores. Finally, we have developed a wing score-based dose-dependency approach that can provide genotoxicity profiles. We have validated our method using 6 compounds, obtaining profiles almost identical to those obtained from manual measures, even for low-genotoxicity compounds such as urethane. The automated SMART, with its faster and more reliable readout, fulfills the need for a high-throughput in vivo test. The flexible imaging strategy we describe and the analysis tools we provide should facilitate the optimization and dissemination of our methods.

  10. A method for high throughput bioelectrochemical research based on small scale microbial electrolysis cells

    KAUST Repository

    Call, Douglas F.; Logan, Bruce E.

    2011-01-01

    There is great interest in studying exoelectrogenic microorganisms, but existing methods can require expensive electrochemical equipment and specialized reactors. We developed a simple system for conducting high throughput bioelectrochemical

  11. High pressure inertial focusing for separating and concentrating bacteria at high throughput

    Science.gov (United States)

    Cruz, J.; Hooshmand Zadeh, S.; Graells, T.; Andersson, M.; Malmström, J.; Wu, Z. G.; Hjort, K.

    2017-08-01

    Inertial focusing is a promising microfluidic technology for concentration and separation of particles by size. However, there is a strong correlation of increased pressure with decreased particle size. Theory and experimental results for larger particles were used to scale down the phenomenon and find the conditions that focus 1 µm particles. High pressure experiments in robust glass chips were used to demonstrate the alignment. We show how the technique works for 1 µm spherical polystyrene particles and for Escherichia coli, not being harmful for the bacteria at 50 µl min-1. The potential to focus bacteria, simplicity of use and high throughput make this technology interesting for healthcare applications, where concentration and purification of a sample may be required as an initial step.

  12. Systems biology of bacterial nitrogen fixation: High-throughput technology and its integrative description with constraint-based modeling

    Directory of Open Access Journals (Sweden)

    Resendis-Antonio Osbaldo

    2011-07-01

    Full Text Available Abstract Background Bacterial nitrogen fixation is the biological process by which atmospheric nitrogen is uptaken by bacteroids located in plant root nodules and converted into ammonium through the enzymatic activity of nitrogenase. In practice, this biological process serves as a natural form of fertilization and its optimization has significant implications in sustainable agricultural programs. Currently, the advent of high-throughput technology supplies with valuable data that contribute to understanding the metabolic activity during bacterial nitrogen fixation. This undertaking is not trivial, and the development of computational methods useful in accomplishing an integrative, descriptive and predictive framework is a crucial issue to decoding the principles that regulated the metabolic activity of this biological process. Results In this work we present a systems biology description of the metabolic activity in bacterial nitrogen fixation. This was accomplished by an integrative analysis involving high-throughput data and constraint-based modeling to characterize the metabolic activity in Rhizobium etli bacteroids located at the root nodules of Phaseolus vulgaris (bean plant. Proteome and transcriptome technologies led us to identify 415 proteins and 689 up-regulated genes that orchestrate this biological process. Taking into account these data, we: 1 extended the metabolic reconstruction reported for R. etli; 2 simulated the metabolic activity during symbiotic nitrogen fixation; and 3 evaluated the in silico results in terms of bacteria phenotype. Notably, constraint-based modeling simulated nitrogen fixation activity in such a way that 76.83% of the enzymes and 69.48% of the genes were experimentally justified. Finally, to further assess the predictive scope of the computational model, gene deletion analysis was carried out on nine metabolic enzymes. Our model concluded that an altered metabolic activity on these enzymes induced

  13. EMBRYONIC VASCULAR DISRUPTION ADVERSE OUTCOMES: LINKING HIGH THROUGHPUT SIGNALING SIGNATURES WITH FUNCTIONAL CONSEQUENCES

    Science.gov (United States)

    Embryonic vascular disruption is an important adverse outcome pathway (AOP) given the knowledge that chemical disruption of early cardiovascular system development leads to broad prenatal defects. High throughput screening (HTS) assays provide potential building blocks for AOP d...

  14. Uplink SDMA with Limited Feedback: Throughput Scaling

    Directory of Open Access Journals (Sweden)

    Jeffrey G. Andrews

    2008-01-01

    Full Text Available Combined space division multiple access (SDMA and scheduling exploit both spatial multiplexing and multiuser diversity, increasing throughput significantly. Both SDMA and scheduling require feedback of multiuser channel sate information (CSI. This paper focuses on uplink SDMA with limited feedback, which refers to efficient techniques for CSI quantization and feedback. To quantify the throughput of uplink SDMA and derive design guidelines, the throughput scaling with system parameters is analyzed. The specific parameters considered include the numbers of users, antennas, and feedback bits. Furthermore, different SNR regimes and beamforming methods are considered. The derived throughput scaling laws are observed to change for different SNR regimes. For instance, the throughput scales logarithmically with the number of users in the high SNR regime but double logarithmically in the low SNR regime. The analysis of throughput scaling suggests guidelines for scheduling in uplink SDMA. For example, to maximize throughput scaling, scheduling should use the criterion of minimum quantization errors for the high SNR regime and maximum channel power for the low SNR regime.

  15. A high-throughput qPCR system for simultaneous quantitative detection of dairy Lactococcus lactis and Leuconostoc bacteriophages

    DEFF Research Database (Denmark)

    Muhammed, Musemma Kedir; Krych, Lukasz; Nielsen, Dennis Sandris

    2017-01-01

    simultaneous quantitative detection of Lc. lactis 936 (now SK1virus), P335, c2 (now C2virus) and Leuconostoc phage groups. Component assays are designed to have high efficiencies and nearly the same dynamic detection ranges, i.e., from 1.1 x 105 to 1.1 x 101 phage genomes per reaction, which corresponds to 9 x......Simultaneous quantitative detection of Lactococcus (Lc.) lactis and Leuconostoc species bacteriophages (phages) has not been reported in dairies using undefined mixed-strain DL-starters, probably due to the lack of applicable methods. We optimized a high-throughput qPCR system that allows...... 107 to 9 x 103 phage particles mL-1 without any additional up-concentrating steps. The amplification efficiencies of the corresponding assays were 100.1±2.6, 98.7±2.3, 101.0±2.3 and 96.2±6.2. The qPCR system was tested on samples obtained from a dairy plant that employed traditional mother...

  16. Crop 3D-a LiDAR based platform for 3D high-throughput crop phenotyping.

    Science.gov (United States)

    Guo, Qinghua; Wu, Fangfang; Pang, Shuxin; Zhao, Xiaoqian; Chen, Linhai; Liu, Jin; Xue, Baolin; Xu, Guangcai; Li, Le; Jing, Haichun; Chu, Chengcai

    2018-03-01

    With the growing population and the reducing arable land, breeding has been considered as an effective way to solve the food crisis. As an important part in breeding, high-throughput phenotyping can accelerate the breeding process effectively. Light detection and ranging (LiDAR) is an active remote sensing technology that is capable of acquiring three-dimensional (3D) data accurately, and has a great potential in crop phenotyping. Given that crop phenotyping based on LiDAR technology is not common in China, we developed a high-throughput crop phenotyping platform, named Crop 3D, which integrated LiDAR sensor, high-resolution camera, thermal camera and hyperspectral imager. Compared with traditional crop phenotyping techniques, Crop 3D can acquire multi-source phenotypic data in the whole crop growing period and extract plant height, plant width, leaf length, leaf width, leaf area, leaf inclination angle and other parameters for plant biology and genomics analysis. In this paper, we described the designs, functions and testing results of the Crop 3D platform, and briefly discussed the potential applications and future development of the platform in phenotyping. We concluded that platforms integrating LiDAR and traditional remote sensing techniques might be the future trend of crop high-throughput phenotyping.

  17. TCP Throughput Profiles Using Measurements over Dedicated Connections

    Energy Technology Data Exchange (ETDEWEB)

    Rao, Nageswara S. [ORNL; Liu, Qiang [ORNL; Sen, Satyabrata [ORNL; Towsley, Don [University of Massachusetts, Amherst; Vardoyan, Gayane [University of Massachusetts, Amherst; Kettimuthu, R. [Argonne National Laboratory (ANL); Foster, Ian [University of Chicago

    2017-06-01

    Wide-area data transfers in high-performance computing infrastructures are increasingly being carried over dynamically provisioned dedicated network connections that provide high capacities with no competing traffic. We present extensive TCP throughput measurements and time traces over a suite of physical and emulated 10 Gbps connections with 0-366 ms round-trip times (RTTs). Contrary to the general expectation, they show significant statistical and temporal variations, in addition to the overall dependencies on the congestion control mechanism, buffer size, and the number of parallel streams. We analyze several throughput profiles that have highly desirable concave regions wherein the throughput decreases slowly with RTTs, in stark contrast to the convex profiles predicted by various TCP analytical models. We present a generic throughput model that abstracts the ramp-up and sustainment phases of TCP flows, which provides insights into qualitative trends observed in measurements across TCP variants: (i) slow-start followed by well-sustained throughput leads to concave regions; (ii) large buffers and multiple parallel streams expand the concave regions in addition to improving the throughput; and (iii) stable throughput dynamics, indicated by a smoother Poincare map and smaller Lyapunov exponents, lead to wider concave regions. These measurements and analytical results together enable us to select a TCP variant and its parameters for a given connection to achieve high throughput with statistical guarantees.

  18. High-Throughput Non-destructive Phenotyping of Traits that Contribute to Salinity Tolerance in Arabidopsis thaliana

    KAUST Repository

    Awlia, Mariam

    2016-09-28

    Reproducible and efficient high-throughput phenotyping approaches, combined with advances in genome sequencing, are facilitating the discovery of genes affecting plant performance. Salinity tolerance is a desirable trait that can be achieved through breeding, where most have aimed at selecting for plants that perform effective ion exclusion from the shoots. To determine overall plant performance under salt stress, it is helpful to investigate several plant traits collectively in one experimental setup. Hence, we developed a quantitative phenotyping protocol using a high-throughput phenotyping system, with RGB and chlorophyll fluorescence (ChlF) imaging, which captures the growth, morphology, color and photosynthetic performance of Arabidopsis thaliana plants in response to salt stress. We optimized our salt treatment by controlling the soil-water content prior to introducing salt stress. We investigated these traits over time in two accessions in soil at 150, 100, or 50 mM NaCl to find that the plants subjected to 100 mM NaCl showed the most prominent responses in the absence of symptoms of severe stress. In these plants, salt stress induced significant changes in rosette area and morphology, but less prominent changes in rosette coloring and photosystem II efficiency. Clustering of ChlF traits with plant growth of nine accessions maintained at 100 mM NaCl revealed that in the early stage of salt stress, salinity tolerance correlated with non-photochemical quenching processes and during the later stage, plant performance correlated with quantum yield. This integrative approach allows the simultaneous analysis of several phenotypic traits. In combination with various genetic resources, the phenotyping protocol described here is expected to increase our understanding of plant performance and stress responses, ultimately identifying genes that improve plant performance in salt stress conditions.

  19. Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection.

    Science.gov (United States)

    Ernstsen, Christina L; Login, Frédéric H; Jensen, Helene H; Nørregaard, Rikke; Møller-Jensen, Jakob; Nejsum, Lene N

    2017-10-01

    Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacterial colonies in infected host cells (Detection and quantification of intracellular bacterial colonies by automated, high-throughput microscopy, Ernstsen et al., 2017 [1]). The infected cells were imaged with a 10× objective and number of intracellular bacterial colonies, their size distribution and the number of cell nuclei were automatically quantified using a spot detection-tool. The spot detection-output was exported to Excel, where data analysis was performed. In this article, micrographs and spot detection data are made available to facilitate implementation of the method.

  20. Multiple and high-throughput droplet reactions via combination of microsampling technique and microfluidic chip

    KAUST Repository

    Wu, Jinbo

    2012-11-20

    Microdroplets offer unique compartments for accommodating a large number of chemical and biological reactions in tiny volume with precise control. A major concern in droplet-based microfluidics is the difficulty to address droplets individually and achieve high throughput at the same time. Here, we have combined an improved cartridge sampling technique with a microfluidic chip to perform droplet screenings and aggressive reaction with minimal (nanoliter-scale) reagent consumption. The droplet composition, distance, volume (nanoliter to subnanoliter scale), number, and sequence could be precisely and digitally programmed through the improved sampling technique, while sample evaporation and cross-contamination are effectively eliminated. Our combined device provides a simple model to utilize multiple droplets for various reactions with low reagent consumption and high throughput. © 2012 American Chemical Society.

  1. Ultra-high-throughput screening of an in vitro-synthesized horseradish peroxidase displayed on microbeads using cell sorter.

    Directory of Open Access Journals (Sweden)

    Bo Zhu

    Full Text Available The C1a isoenzyme of horseradish peroxidase (HRP is an industrially important heme-containing enzyme that utilizes hydrogen peroxide to oxidize a wide variety of inorganic and organic compounds for practical applications, including synthesis of fine chemicals, medical diagnostics, and bioremediation. To develop a ultra-high-throughput screening system for HRP, we successfully produced active HRP in an Escherichia coli cell-free protein synthesis system, by adding disulfide bond isomerase DsbC and optimizing the concentrations of hemin and calcium ions and the temperature. The biosynthesized HRP was fused with a single-chain Cro (scCro DNA-binding tag at its N-terminal and C-terminal sites. The addition of the scCro-tag at both ends increased the solubility of the protein. Next, HRP and its fusion proteins were successfully synthesized in a water droplet emulsion by using hexadecane as the oil phase and SunSoft No. 818SK as the surfactant. HRP fusion proteins were displayed on microbeads attached with double-stranded DNA (containing the scCro binding sequence via scCro-DNA interactions. The activities of the immobilized HRP fusion proteins were detected with a tyramide-based fluorogenic assay using flow cytometry. Moreover, a model microbead library containing wild type hrp (WT and inactive mutant (MUT genes was screened using fluorescence-activated cell-sorting, thus efficiently enriching the WT gene from the 1:100 (WT:MUT library. The technique described here could serve as a novel platform for the ultra-high-throughput discovery of more useful HRP mutants and other heme-containing peroxidases.

  2. NMRbot: Python scripts enable high-throughput data collection on current Bruker BioSpin NMR spectrometers.

    Science.gov (United States)

    Clos, Lawrence J; Jofre, M Fransisca; Ellinger, James J; Westler, William M; Markley, John L

    2013-06-01

    To facilitate the high-throughput acquisition of nuclear magnetic resonance (NMR) experimental data on large sets of samples, we have developed a simple and straightforward automated methodology that capitalizes on recent advances in Bruker BioSpin NMR spectrometer hardware and software. Given the daunting challenge for non-NMR experts to collect quality spectra, our goal was to increase user accessibility, provide customized functionality, and improve the consistency and reliability of resultant data. This methodology, NMRbot, is encoded in a set of scripts written in the Python programming language accessible within the Bruker BioSpin TopSpin ™ software. NMRbot improves automated data acquisition and offers novel tools for use in optimizing experimental parameters on the fly. This automated procedure has been successfully implemented for investigations in metabolomics, small-molecule library profiling, and protein-ligand titrations on four Bruker BioSpin NMR spectrometers at the National Magnetic Resonance Facility at Madison. The investigators reported benefits from ease of setup, improved spectral quality, convenient customizations, and overall time savings.

  3. Laboratory Information Management Software for genotyping workflows: applications in high throughput crop genotyping

    Directory of Open Access Journals (Sweden)

    Prasanth VP

    2006-08-01

    Full Text Available Abstract Background With the advances in DNA sequencer-based technologies, it has become possible to automate several steps of the genotyping process leading to increased throughput. To efficiently handle the large amounts of genotypic data generated and help with quality control, there is a strong need for a software system that can help with the tracking of samples and capture and management of data at different steps of the process. Such systems, while serving to manage the workflow precisely, also encourage good laboratory practice by standardizing protocols, recording and annotating data from every step of the workflow. Results A laboratory information management system (LIMS has been designed and implemented at the International Crops Research Institute for the Semi-Arid Tropics (ICRISAT that meets the requirements of a moderately high throughput molecular genotyping facility. The application is designed as modules and is simple to learn and use. The application leads the user through each step of the process from starting an experiment to the storing of output data from the genotype detection step with auto-binning of alleles; thus ensuring that every DNA sample is handled in an identical manner and all the necessary data are captured. The application keeps track of DNA samples and generated data. Data entry into the system is through the use of forms for file uploads. The LIMS provides functions to trace back to the electrophoresis gel files or sample source for any genotypic data and for repeating experiments. The LIMS is being presently used for the capture of high throughput SSR (simple-sequence repeat genotyping data from the legume (chickpea, groundnut and pigeonpea and cereal (sorghum and millets crops of importance in the semi-arid tropics. Conclusion A laboratory information management system is available that has been found useful in the management of microsatellite genotype data in a moderately high throughput genotyping

  4. A high-throughput fluorescence resonance energy transfer (FRET)-based endothelial cell apoptosis assay and its application for screening vascular disrupting agents

    International Nuclear Information System (INIS)

    Zhu, Xiaoming; Fu, Afu; Luo, Kathy Qian

    2012-01-01

    Highlights: ► An endothelial cell apoptosis assay using FRET-based biosensor was developed. ► The fluorescence of the cells changed from green to blue during apoptosis. ► This method was developed into a high-throughput assay in 96-well plates. ► This assay was applied to screen vascular disrupting agents. -- Abstract: In this study, we developed a high-throughput endothelial cell apoptosis assay using a fluorescence resonance energy transfer (FRET)-based biosensor. After exposure to apoptotic inducer UV-irradiation or anticancer drugs such as paclitaxel, the fluorescence of the cells changed from green to blue. We developed this method into a high-throughput assay in 96-well plates by measuring the emission ratio of yellow fluorescent protein (YFP) to cyan fluorescent protein (CFP) to monitor the activation of a key protease, caspase-3, during apoptosis. The Z′ factor for this assay was above 0.5 which indicates that this assay is suitable for a high-throughput analysis. Finally, we applied this functional high-throughput assay for screening vascular disrupting agents (VDA) which could induce endothelial cell apoptosis from our in-house compounds library and dioscin was identified as a hit. As this assay allows real time and sensitive detection of cell apoptosis, it will be a useful tool for monitoring endothelial cell apoptosis in living cell situation and for identifying new VDA candidates via a high-throughput screening.

  5. Raman-Activated Droplet Sorting (RADS) for Label-Free High-Throughput Screening of Microalgal Single-Cells.

    Science.gov (United States)

    Wang, Xixian; Ren, Lihui; Su, Yetian; Ji, Yuetong; Liu, Yaoping; Li, Chunyu; Li, Xunrong; Zhang, Yi; Wang, Wei; Hu, Qiang; Han, Danxiang; Xu, Jian; Ma, Bo

    2017-11-21

    Raman-activated cell sorting (RACS) has attracted increasing interest, yet throughput remains one major factor limiting its broader application. Here we present an integrated Raman-activated droplet sorting (RADS) microfluidic system for functional screening of live cells in a label-free and high-throughput manner, by employing AXT-synthetic industrial microalga Haematococcus pluvialis (H. pluvialis) as a model. Raman microspectroscopy analysis of individual cells is carried out prior to their microdroplet encapsulation, which is then directly coupled to DEP-based droplet sorting. To validate the system, H. pluvialis cells containing different levels of AXT were mixed and underwent RADS. Those AXT-hyperproducing cells were sorted with an accuracy of 98.3%, an enrichment ratio of eight folds, and a throughput of ∼260 cells/min. Of the RADS-sorted cells, 92.7% remained alive and able to proliferate, which is equivalent to the unsorted cells. Thus, the RADS achieves a much higher throughput than existing RACS systems, preserves the vitality of cells, and facilitates seamless coupling with downstream manipulations such as single-cell sequencing and cultivation.

  6. High throughput screening of ligand binding to macromolecules using high resolution powder diffraction

    Science.gov (United States)

    Von Dreele, Robert B.; D'Amico, Kevin

    2006-10-31

    A process is provided for the high throughput screening of binding of ligands to macromolecules using high resolution powder diffraction data including producing a first sample slurry of a selected polycrystalline macromolecule material and a solvent, producing a second sample slurry of a selected polycrystalline macromolecule material, one or more ligands and the solvent, obtaining a high resolution powder diffraction pattern on each of said first sample slurry and the second sample slurry, and, comparing the high resolution powder diffraction pattern of the first sample slurry and the high resolution powder diffraction pattern of the second sample slurry whereby a difference in the high resolution powder diffraction patterns of the first sample slurry and the second sample slurry provides a positive indication for the formation of a complex between the selected polycrystalline macromolecule material and at least one of the one or more ligands.

  7. Quantitative high throughput analytics to support polysaccharide production process development.

    Science.gov (United States)

    Noyes, Aaron; Godavarti, Ranga; Titchener-Hooker, Nigel; Coffman, Jonathan; Mukhopadhyay, Tarit

    2014-05-19

    The rapid development of purification processes for polysaccharide vaccines is constrained by a lack of analytical tools current technologies for the measurement of polysaccharide recovery and process-related impurity clearance are complex, time-consuming, and generally not amenable to high throughput process development (HTPD). HTPD is envisioned to be central to the improvement of existing polysaccharide manufacturing processes through the identification of critical process parameters that potentially impact the quality attributes of the vaccine and to the development of de novo processes for clinical candidates, across the spectrum of downstream processing. The availability of a fast and automated analytics platform will expand the scope, robustness, and evolution of Design of Experiment (DOE) studies. This paper details recent advances in improving the speed, throughput, and success of in-process analytics at the micro-scale. Two methods, based on modifications of existing procedures, are described for the rapid measurement of polysaccharide titre in microplates without the need for heating steps. A simplification of a commercial endotoxin assay is also described that features a single measurement at room temperature. These assays, along with existing assays for protein and nucleic acids are qualified for deployment in the high throughput screening of polysaccharide feedstreams. Assay accuracy, precision, robustness, interference, and ease of use are assessed and described. In combination, these assays are capable of measuring the product concentration and impurity profile of a microplate of 96 samples in less than one day. This body of work relies on the evaluation of a combination of commercially available and clinically relevant polysaccharides to ensure maximum versatility and reactivity of the final assay suite. Together, these advancements reduce overall process time by up to 30-fold and significantly reduce sample volume over current practices. The

  8. Development and Optimization of an Alternative Electrospinning Process for High Throughput

    Science.gov (United States)

    Thoppey Muthuraman, Nagarajan

    This work is an investigation of the prospect of electrospinning from the simplest aperture-free system, a flat plate on which polymer solution is placed as droplets or undergoes a gravity-assisted flow. Nanofibers with a similar fiber diameter and diameter distribution were fabricated at similar voltages and working distances as that in an aperture-based system, however with much more flexibility to scale up the process and with no openings or nozzles that can clog. It is verified that the field gradient at the site of jet formation is important. In particular, it is shown that the relatively homogeneous electric field on the plate surface does not promote electrospinning as compared with the significantly more inhomogeneous field at the needle tip in the needle-plate configuration. However, the strong field gradient at the plate edge allows electrospinning from unconfined droplets of the polymer solution and formation of fibers with very similar diameters and diameter distributions as those fabricated by traditional needle electrospinning for the same polymer solution. Further it is also shown that this edge-plate methodology can be extended to systems with many "edges" and curved edges (such as those from a hollow cylinder) for massively-parallel electrospinning (that is, higher potential throughput). A detailed examination of the changes in fiber diameter, diameter distribution, and mat porosity is reported as a function of the electric field magnitude and geometry, and it is concluded that the process is quite stable over a range of experimental conditions. The connection between fiber properties and spinning conditions via changes in the length and duration of the linear region and the degree of whipping is discussed in the context of comparing edge-plate and needle-plate electrospinning. Not only do these results address issues specific to such a surface-based, parallel aperture-less electrospinning approach, they also continue to expand understanding of

  9. Image Harvest: an open-source platform for high-throughput plant image processing and analysis

    Science.gov (United States)

    Knecht, Avi C.; Campbell, Malachy T.; Caprez, Adam; Swanson, David R.; Walia, Harkamal

    2016-01-01

    High-throughput plant phenotyping is an effective approach to bridge the genotype-to-phenotype gap in crops. Phenomics experiments typically result in large-scale image datasets, which are not amenable for processing on desktop computers, thus creating a bottleneck in the image-analysis pipeline. Here, we present an open-source, flexible image-analysis framework, called Image Harvest (IH), for processing images originating from high-throughput plant phenotyping platforms. Image Harvest is developed to perform parallel processing on computing grids and provides an integrated feature for metadata extraction from large-scale file organization. Moreover, the integration of IH with the Open Science Grid provides academic researchers with the computational resources required for processing large image datasets at no cost. Image Harvest also offers functionalities to extract digital traits from images to interpret plant architecture-related characteristics. To demonstrate the applications of these digital traits, a rice (Oryza sativa) diversity panel was phenotyped and genome-wide association mapping was performed using digital traits that are used to describe different plant ideotypes. Three major quantitative trait loci were identified on rice chromosomes 4 and 6, which co-localize with quantitative trait loci known to regulate agronomically important traits in rice. Image Harvest is an open-source software for high-throughput image processing that requires a minimal learning curve for plant biologists to analyzephenomics datasets. PMID:27141917

  10. The Stanford Automated Mounter: Enabling High-Throughput Protein Crystal Screening at SSRL

    International Nuclear Information System (INIS)

    Smith, C.A.; Cohen, A.E.

    2009-01-01

    The macromolecular crystallography experiment lends itself perfectly to high-throughput technologies. The initial steps including the expression, purification, and crystallization of protein crystals, along with some of the later steps involving data processing and structure determination have all been automated to the point where some of the last remaining bottlenecks in the process have been crystal mounting, crystal screening, and data collection. At the Stanford Synchrotron Radiation Laboratory, a National User Facility that provides extremely brilliant X-ray photon beams for use in materials science, environmental science, and structural biology research, the incorporation of advanced robotics has enabled crystals to be screened in a true high-throughput fashion, thus dramatically accelerating the final steps. Up to 288 frozen crystals can be mounted by the beamline robot (the Stanford Auto-Mounting System) and screened for diffraction quality in a matter of hours without intervention. The best quality crystals can then be remounted for the collection of complete X-ray diffraction data sets. Furthermore, the entire screening and data collection experiment can be controlled from the experimenter's home laboratory by means of advanced software tools that enable network-based control of the highly automated beamlines.

  11. Adaptation to high throughput batch chromatography enhances multivariate screening.

    Science.gov (United States)

    Barker, Gregory A; Calzada, Joseph; Herzer, Sibylle; Rieble, Siegfried

    2015-09-01

    High throughput process development offers unique approaches to explore complex process design spaces with relatively low material consumption. Batch chromatography is one technique that can be used to screen chromatographic conditions in a 96-well plate. Typical batch chromatography workflows examine variations in buffer conditions or comparison of multiple resins in a given process, as opposed to the assessment of protein loading conditions in combination with other factors. A modification to the batch chromatography paradigm is described here where experimental planning, programming, and a staggered loading approach increase the multivariate space that can be explored with a liquid handling system. The iterative batch chromatography (IBC) approach is described, which treats every well in a 96-well plate as an individual experiment, wherein protein loading conditions can be varied alongside other factors such as wash and elution buffer conditions. As all of these factors are explored in the same experiment, the interactions between them are characterized and the number of follow-up confirmatory experiments is reduced. This in turn improves statistical power and throughput. Two examples of the IBC method are shown and the impact of the load conditions are assessed in combination with the other factors explored. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Development of Microfluidic Systems Enabling High-Throughput Single-Cell Protein Characterization

    OpenAIRE

    Fan, Beiyuan; Li, Xiufeng; Chen, Deyong; Peng, Hongshang; Wang, Junbo; Chen, Jian

    2016-01-01

    This article reviews recent developments in microfluidic systems enabling high-throughput characterization of single-cell proteins. Four key perspectives of microfluidic platforms are included in this review: (1) microfluidic fluorescent flow cytometry; (2) droplet based microfluidic flow cytometry; (3) large-array micro wells (microengraving); and (4) large-array micro chambers (barcode microchips). We examine the advantages and limitations of each technique and discuss future research oppor...

  13. Solid-phase cloning for high-throughput assembly of single and multiple DNA parts

    DEFF Research Database (Denmark)

    Lundqvist, Magnus; Edfors, Fredrik; Sivertsson, Åsa

    2015-01-01

    We describe solid-phase cloning (SPC) for high-throughput assembly of expression plasmids. Our method allows PCR products to be put directly into a liquid handler for capture and purification using paramagnetic streptavidin beads and conversion into constructs by subsequent cloning reactions. We ...

  14. A high-throughput method for GMO multi-detection using a microfluidic dynamic array

    NARCIS (Netherlands)

    Brod, F.C.A.; Dijk, van J.P.; Voorhuijzen, M.M.; Dinon, A.Z.; Guimarães, L.H.S.; Scholtens, I.M.J.; Arisi, A.C.M.; Kok, E.J.

    2014-01-01

    The ever-increasing production of genetically modified crops generates a demand for high-throughput DNAbased methods for the enforcement of genetically modified organisms (GMO) labelling requirements. The application of standard real-time PCR will become increasingly costly with the growth of the

  15. Defining the taxonomic domain of applicability for mammalian-based high-throughput screening assays

    Science.gov (United States)

    Cell-based high throughput screening (HTS) technologies are becoming mainstream in chemical safety evaluations. The US Environmental Protection Agency (EPA) Toxicity Forecaster (ToxCastTM) and the multi-agency Tox21 Programs have been at the forefront in advancing this science, m...

  16. High Throughput Preparation of Aligned Nanofibers Using an Improved Bubble-Electrospinning

    Directory of Open Access Journals (Sweden)

    Liang Yu

    2017-11-01

    Full Text Available An improved bubble-electrospinning, consisting of a cone shaped air nozzle, a copper solution reservoir connected directly to the power generator, and a high speed rotating copper wire drum as a collector, was presented successfully to obtain high throughput preparation of aligned nanofibers. The influences of drum rotation speed on morphology and properties of obtained nanofibers were explored and researched. The results showed that the alignment degree, diameter distribution, and properties of nanofibers were improved with the increase of the drum rotation speed.

  17. Droplet electrospray ionization mass spectrometry for high throughput screening for enzyme inhibitors.

    Science.gov (United States)

    Sun, Shuwen; Kennedy, Robert T

    2014-09-16

    High throughput screening (HTS) is important for identifying molecules with desired properties. Mass spectrometry (MS) is potentially powerful for label-free HTS due to its high sensitivity, speed, and resolution. Segmented flow, where samples are manipulated as droplets separated by an immiscible fluid, is an intriguing format for high throughput MS because it can be used to reliably and precisely manipulate nanoliter volumes and can be directly coupled to electrospray ionization (ESI) MS for rapid analysis. In this study, we describe a "MS Plate Reader" that couples standard multiwell plate HTS workflow to droplet ESI-MS. The MS plate reader can reformat 3072 samples from eight 384-well plates into nanoliter droplets segmented by an immiscible oil at 4.5 samples/s and sequentially analyze them by MS at 2 samples/s. Using the system, a label-free screen for cathepsin B modulators against 1280 chemicals was completed in 45 min with a high Z-factor (>0.72) and no false positives (24 of 24 hits confirmed). The assay revealed 11 structures not previously linked to cathepsin inhibition. For even larger scale screening, reformatting and analysis could be conducted simultaneously, which would enable more than 145,000 samples to be analyzed in 1 day.

  18. High-Throughput Proteomics Using High Efficiency Multiple-Capillary Liquid Chromatography With On-Line High-Performance ESI FTICR Mass Spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    Shen, Yufeng (BATTELLE (PACIFIC NW LAB)); Tolic, Nikola (BATTELLE (PACIFIC NW LAB)); Zhao, Rui (ASSOC WESTERN UNIVERSITY); Pasa Tolic, Ljiljana (BATTELLE (PACIFIC NW LAB)); Li, Lingjun (Illinois Univ Of-Urbana/Champa); Berger, Scott J.(ASSOC WESTERN UNIVERSITY); Harkewicz, Richard (BATTELLE (PACIFIC NW LAB)); Anderson, Gordon A.(BATTELLE (PACIFIC NW LAB)); Belov, Mikhail E.(BATTELLE (PACIFIC NW LAB)); Smith, Richard D.(BATTELLE (PACIFIC NW LAB))

    2000-12-01

    We report on the design and application of a high-efficiency multiple-capillary liquid chromatography (LC) system for high-throughput proteome analysis. The multiple-capillary LC system was operated at the pressure of 10,000 psi using commercial LC pumps to deliver the mobile phase and newly developed passive feedback valves to switch the mobile phase flow and introduce samples. The multiple-capillary LC system was composed of several serially connected dual-capillary column devices. The dual-capillary column approach was designed to eliminate the time delay for regeneration (or equilibrium) of the capillary column after its use under the mobile phase gradient condition (i.e. one capillary column was used in separation and the other was washed using mobile phase A). The serially connected dual-capillary columns and ESI sources were operated independently, and could be used for either''backup'' operation or with other mass spectrometer(s). This high-efficiency multiple-capillary LC system uses switching valves for all operations and is highly amenable to automation. The separations efficiency of dual-capillary column device, optimal capillary dimensions (column length and packed particle size), suitable mobile phases for electrospray, and the capillary re-generation were investigated. A high magnetic field (11.5 tesla) Fourier transform ion cyclotron resonance (FTICR) mass spectrometer was coupled on-line with this high-efficiency multiple-capillary LC system through an electrospray ionization source. The capillary LC provided a peak capacity of {approx}600, and the 2-D capillary LC-FTICR provided a combined resolving power of > 6 x 10 7 polypeptide isotopic distributions. For yeast cellular tryptic digests, > 100,000 polypeptides were typically detected, and {approx}1,000 proteins can be characterized in a single run.

  19. A bioimage informatics platform for high-throughput embryo phenotyping.

    Science.gov (United States)

    Brown, James M; Horner, Neil R; Lawson, Thomas N; Fiegel, Tanja; Greenaway, Simon; Morgan, Hugh; Ring, Natalie; Santos, Luis; Sneddon, Duncan; Teboul, Lydia; Vibert, Jennifer; Yaikhom, Gagarine; Westerberg, Henrik; Mallon, Ann-Marie

    2018-01-01

    High-throughput phenotyping is a cornerstone of numerous functional genomics projects. In recent years, imaging screens have become increasingly important in understanding gene-phenotype relationships in studies of cells, tissues and whole organisms. Three-dimensional (3D) imaging has risen to prominence in the field of developmental biology for its ability to capture whole embryo morphology and gene expression, as exemplified by the International Mouse Phenotyping Consortium (IMPC). Large volumes of image data are being acquired by multiple institutions around the world that encompass a range of modalities, proprietary software and metadata. To facilitate robust downstream analysis, images and metadata must be standardized to account for these differences. As an open scientific enterprise, making the data readily accessible is essential so that members of biomedical and clinical research communities can study the images for themselves without the need for highly specialized software or technical expertise. In this article, we present a platform of software tools that facilitate the upload, analysis and dissemination of 3D images for the IMPC. Over 750 reconstructions from 80 embryonic lethal and subviable lines have been captured to date, all of which are openly accessible at mousephenotype.org. Although designed for the IMPC, all software is available under an open-source licence for others to use and develop further. Ongoing developments aim to increase throughput and improve the analysis and dissemination of image data. Furthermore, we aim to ensure that images are searchable so that users can locate relevant images associated with genes, phenotypes or human diseases of interest. © The Author 2016. Published by Oxford University Press.

  20. Development on the High-throughput Vol-oxidizer for Decladding and Voloxidation of Spent Fuel Rod-cuts

    International Nuclear Information System (INIS)

    Kim, Young Hwang; Jung, Jae Hoo; Kim, Ki Ho; Park, Byung Buk; Lee, Hyo Jik; Kim, Sung Hyun; Park, Hee Sung; Lee, Jong Kwang; Kim, Ho Dong

    2009-12-01

    A high-throughput vol-oxidizer which can handle a several ten kg HM/batch is being developed to supply U 3 O 8 powders to an electrolytic reduction reactor in pyro-processing. At the first year step(2007), for enhancement of oxidation and recovery rate, we analyzed the mechanical and chemical methods, and devised the main mechanism with ball drop methods and rotary kiln type. Also, the main devices for oxidation and recovery of rod-cuts were designed by using the Solid Works and COSMOS program tools, and manufactured after thermal/mechanical analysis. In order to verify the main devices, simulation fuels(W 90%+SiO 2 10%) were manufactured and the main devices were tested for the oxidation and recovery rate of its. Here the expansion ratio of simulation fuel is similar to U 3 O 8 (2.7). At the second year step(2008), with the constant ration of rod-cuts volume and expansion ratio of U 3 O 8 (2.7), we produced a theoretical equation that can estimate the volume of rod-cuts according to a variation of their weight and lengths. We considered various materials such as ceramics and Ni-Cr, finally, the APM material which can constantly maintain against high temperature(1,200 .deg. C) and vacuum(1 torr) was selected and a vol-oxidizer was designed. At the third year step(2009), in order to manufacture a high-throughput vol-oxidizer, we have analyzed the vol-oxidizer for remote operability and maintainability, also the remote assembling and disassembling possibilities of the selected modules have been analyzed in terms of visibility, interference, approach, weight, and so on. We have presented final modular design and manufactured a high-throughput vol-oxidizer. Also, we have conducted the blank, heating(over 500 .deg. C) and hull separation test(capacity : 50 kg HM/batch, hull length 50mm) on the high-throughput vol-oxidizer. Also, these design technologies for the high-throughput vol-oxidizer will be utilized in the development of a more efficient vol-oxidizer with higher

  1. X-ray phase microtomography with a single grating for high-throughput investigations of biological tissue.

    Science.gov (United States)

    Zdora, Marie-Christine; Vila-Comamala, Joan; Schulz, Georg; Khimchenko, Anna; Hipp, Alexander; Cook, Andrew C; Dilg, Daniel; David, Christian; Grünzweig, Christian; Rau, Christoph; Thibault, Pierre; Zanette, Irene

    2017-02-01

    The high-throughput 3D visualisation of biological specimens is essential for studying diseases and developmental disorders. It requires imaging methods that deliver high-contrast, high-resolution volumetric information at short sample preparation and acquisition times. Here we show that X-ray phase-contrast tomography using a single grating can provide a powerful alternative to commonly employed techniques, such as high-resolution episcopic microscopy (HREM). We present the phase tomography of a mouse embryo in paraffin obtained with an X-ray single-grating interferometer at I13-2 Beamline at Diamond Light Source and discuss the results in comparison with HREM measurements. The excellent contrast and quantitative density information achieved non-destructively and without staining using a simple, robust setup make X-ray single-grating interferometry an optimum candidate for high-throughput imaging of biological specimens as an alternative for existing methods like HREM.

  2. DRABAL: novel method to mine large high-throughput screening assays using Bayesian active learning

    KAUST Repository

    Soufan, Othman; Ba Alawi, Wail; Afeef, Moataz A.; Essack, Magbubah; Kalnis, Panos; Bajic, Vladimir B.

    2016-01-01

    Mining high-throughput screening (HTS) assays is key for enhancing decisions in the area of drug repositioning and drug discovery. However, many challenges are encountered in the process of developing suitable and accurate methods

  3. Genecentric: a package to uncover graph-theoretic structure in high-throughput epistasis data.

    Science.gov (United States)

    Gallant, Andrew; Leiserson, Mark D M; Kachalov, Maxim; Cowen, Lenore J; Hescott, Benjamin J

    2013-01-18

    New technology has resulted in high-throughput screens for pairwise genetic interactions in yeast and other model organisms. For each pair in a collection of non-essential genes, an epistasis score is obtained, representing how much sicker (or healthier) the double-knockout organism will be compared to what would be expected from the sickness of the component single knockouts. Recent algorithmic work has identified graph-theoretic patterns in this data that can indicate functional modules, and even sets of genes that may occur in compensatory pathways, such as a BPM-type schema first introduced by Kelley and Ideker. However, to date, any algorithms for finding such patterns in the data were implemented internally, with no software being made publically available. Genecentric is a new package that implements a parallelized version of the Leiserson et al. algorithm (J Comput Biol 18:1399-1409, 2011) for generating generalized BPMs from high-throughput genetic interaction data. Given a matrix of weighted epistasis values for a set of double knock-outs, Genecentric returns a list of generalized BPMs that may represent compensatory pathways. Genecentric also has an extension, GenecentricGO, to query FuncAssociate (Bioinformatics 25:3043-3044, 2009) to retrieve GO enrichment statistics on generated BPMs. Python is the only dependency, and our web site provides working examples and documentation. We find that Genecentric can be used to find coherent functional and perhaps compensatory gene sets from high throughput genetic interaction data. Genecentric is made freely available for download under the GPLv2 from http://bcb.cs.tufts.edu/genecentric.

  4. Multispot single-molecule FRET: High-throughput analysis of freely diffusing molecules.

    Directory of Open Access Journals (Sweden)

    Antonino Ingargiola

    Full Text Available We describe an 8-spot confocal setup for high-throughput smFRET assays and illustrate its performance with two characteristic experiments. First, measurements on a series of freely diffusing doubly-labeled dsDNA samples allow us to demonstrate that data acquired in multiple spots in parallel can be properly corrected and result in measured sample characteristics consistent with those obtained with a standard single-spot setup. We then take advantage of the higher throughput provided by parallel acquisition to address an outstanding question about the kinetics of the initial steps of bacterial RNA transcription. Our real-time kinetic analysis of promoter escape by bacterial RNA polymerase confirms results obtained by a more indirect route, shedding additional light on the initial steps of transcription. Finally, we discuss the advantages of our multispot setup, while pointing potential limitations of the current single laser excitation design, as well as analysis challenges and their solutions.

  5. On Maximizing the Throughput of Packet Transmission under Energy Constraints.

    Science.gov (United States)

    Wu, Weiwei; Dai, Guangli; Li, Yan; Shan, Feng

    2018-06-23

    More and more Internet of Things (IoT) wireless devices have been providing ubiquitous services over the recent years. Since most of these devices are powered by batteries, a fundamental trade-off to be addressed is the depleted energy and the achieved data throughput in wireless data transmission. By exploiting the rate-adaptive capacities of wireless devices, most existing works on energy-efficient data transmission try to design rate-adaptive transmission policies to maximize the amount of transmitted data bits under the energy constraints of devices. Such solutions, however, cannot apply to scenarios where data packets have respective deadlines and only integrally transmitted data packets contribute. Thus, this paper introduces a notion of weighted throughput, which measures how much total value of data packets are successfully and integrally transmitted before their own deadlines. By designing efficient rate-adaptive transmission policies, this paper aims to make the best use of the energy and maximize the weighted throughput. What is more challenging but with practical significance, we consider the fading effect of wireless channels in both offline and online scenarios. In the offline scenario, we develop an optimal algorithm that computes the optimal solution in pseudo-polynomial time, which is the best possible solution as the problem undertaken is NP-hard. In the online scenario, we propose an efficient heuristic algorithm based on optimal properties derived for the optimal offline solution. Simulation results validate the efficiency of the proposed algorithm.

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

    International Nuclear Information System (INIS)

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

    2009-01-01

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

  7. High-throughput tri-colour flow cytometry technique to assess Plasmodium falciparum parasitaemia in bioassays

    DEFF Research Database (Denmark)

    Tiendrebeogo, Regis W; Adu, Bright; Singh, Susheel K

    2014-01-01

    BACKGROUND: Unbiased flow cytometry-based methods have become the technique of choice in many laboratories for high-throughput, accurate assessments of malaria parasites in bioassays. A method to quantify live parasites based on mitotracker red CMXRos was recently described but consistent...... distinction of early ring stages of Plasmodium falciparum from uninfected red blood cells (uRBC) remains a challenge. METHODS: Here, a high-throughput, three-parameter (tri-colour) flow cytometry technique based on mitotracker red dye, the nucleic acid dye coriphosphine O (CPO) and the leucocyte marker CD45...... for enumerating live parasites in bioassays was developed. The technique was applied to estimate the specific growth inhibition index (SGI) in the antibody-dependent cellular inhibition (ADCI) assay and compared to parasite quantification by microscopy and mitotracker red staining. The Bland-Altman analysis...

  8. Leveraging the Power of High Performance Computing for Next Generation Sequencing Data Analysis: Tricks and Twists from a High Throughput Exome Workflow

    Science.gov (United States)

    Wonczak, Stephan; Thiele, Holger; Nieroda, Lech; Jabbari, Kamel; Borowski, Stefan; Sinha, Vishal; Gunia, Wilfried; Lang, Ulrich; Achter, Viktor; Nürnberg, Peter

    2015-01-01

    Next generation sequencing (NGS) has been a great success and is now a standard method of research in the life sciences. With this technology, dozens of whole genomes or hundreds of exomes can be sequenced in rather short time, producing huge amounts of data. Complex bioinformatics analyses are required to turn these data into scientific findings. In order to run these analyses fast, automated workflows implemented on high performance computers are state of the art. While providing sufficient compute power and storage to meet the NGS data challenge, high performance computing (HPC) systems require special care when utilized for high throughput processing. This is especially true if the HPC system is shared by different users. Here, stability, robustness and maintainability are as important for automated workflows as speed and throughput. To achieve all of these aims, dedicated solutions have to be developed. In this paper, we present the tricks and twists that we utilized in the implementation of our exome data processing workflow. It may serve as a guideline for other high throughput data analysis projects using a similar infrastructure. The code implementing our solutions is provided in the supporting information files. PMID:25942438

  9. Generalized schemes for high throughput manipulation of the Desulfovibrio vulgaris Hildenborough genome

    Energy Technology Data Exchange (ETDEWEB)

    Chhabra, S.R.; Butland, G.; Elias, D.; Chandonia, J.-M.; Fok, V.; Juba, T.; Gorur, A.; Allen, S.; Leung, C.-M.; Keller, K.; Reveco, S.; Zane, G.; Semkiw, E.; Prathapam, R.; Gold, B.; Singer, M.; Ouellet, M.; Sazakal, E.; Jorgens, D.; Price, M.; Witkowska, E.; Beller, H.; Hazen, T.C.; Biggin, M.; Auer, M.; Wall, J.; Keasling, J.

    2011-07-15

    The ability to conduct advanced functional genomic studies of the thousands of sequenced bacteria has been hampered by the lack of available tools for making high- throughput chromosomal manipulations in a systematic manner that can be applied across diverse species. In this work, we highlight the use of synthetic biological tools to assemble custom suicide vectors with reusable and interchangeable DNA “parts” to facilitate chromosomal modification at designated loci. These constructs enable an array of downstream applications including gene replacement and creation of gene fusions with affinity purification or localization tags. We employed this approach to engineer chromosomal modifications in a bacterium that has previously proven difficult to manipulate genetically, Desulfovibrio vulgaris Hildenborough, to generate a library of over 700 strains. Furthermore, we demonstrate how these modifications can be used for examining metabolic pathways, protein-protein interactions, and protein localization. The ubiquity of suicide constructs in gene replacement throughout biology suggests that this approach can be applied to engineer a broad range of species for a diverse array of systems biological applications and is amenable to high-throughput implementation.

  10. New approach for high-throughput screening of drug activity on Plasmodium liver stages.

    NARCIS (Netherlands)

    Gego, A.; Silvie, O.; Franetich, J.F.; Farhati, K.; Hannoun, L.; Luty, A.J.F.; Sauerwein, R.W.; Boucheix, C.; Rubinstein, E.; Mazier, D.

    2006-01-01

    Plasmodium liver stages represent potential targets for antimalarial prophylactic drugs. Nevertheless, there is a lack of molecules active on these stages. We have now developed a new approach for the high-throughput screening of drug activity on Plasmodium liver stages in vitro, based on an

  11. Discovery of viruses and virus-like pathogens in pistachio using high-throughput sequencing

    Science.gov (United States)

    Pistachio (Pistacia vera L.) trees from the National Clonal Germplasm Repository (NCGR) and orchards in California were surveyed for viruses and virus-like agents by high-throughput sequencing (HTS). Analyses of 60 trees including clonal UCB-1 hybrid rootstock (P. atlantica × P. integerrima) identif...

  12. GROMACS 4.5: A high-throughput and highly parallel open source molecular simulation toolkit

    Energy Technology Data Exchange (ETDEWEB)

    Pronk, Sander [Science for Life Lab., Stockholm (Sweden); KTH Royal Institute of Technology, Stockholm (Sweden); Pall, Szilard [Science for Life Lab., Stockholm (Sweden); KTH Royal Institute of Technology, Stockholm (Sweden); Schulz, Roland [Univ. of Tennessee, Knoxville, TN (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Larsson, Per [Univ. of Virginia, Charlottesville, VA (United States); Bjelkmar, Par [Science for Life Lab., Stockholm (Sweden); Stockholm Univ., Stockholm (Sweden); Apostolov, Rossen [Science for Life Lab., Stockholm (Sweden); KTH Royal Institute of Technology, Stockholm (Sweden); Shirts, Michael R. [Univ. of Virginia, Charlottesville, VA (United States); Smith, Jeremy C. [Univ. of Tennessee, Knoxville, TN (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Kasson, Peter M. [Univ. of Virginia, Charlottesville, VA (United States); van der Spoel, David [Science for Life Lab., Stockholm (Sweden); Uppsala Univ., Uppsala (Sweden); Hess, Berk [Science for Life Lab., Stockholm (Sweden); KTH Royal Institute of Technology, Stockholm (Sweden); Lindahl, Erik [Science for Life Lab., Stockholm (Sweden); KTH Royal Institute of Technology, Stockholm (Sweden); Stockholm Univ., Stockholm (Sweden)

    2013-02-13

    In this study, molecular simulation has historically been a low-throughput technique, but faster computers and increasing amounts of genomic and structural data are changing this by enabling large-scale automated simulation of, for instance, many conformers or mutants of biomolecules with or without a range of ligands. At the same time, advances in performance and scaling now make it possible to model complex biomolecular interaction and function in a manner directly testable by experiment. These applications share a need for fast and efficient software that can be deployed on massive scale in clusters, web servers, distributed computing or cloud resources. As a result, we present a range of new simulation algorithms and features developed during the past 4 years, leading up to the GROMACS 4.5 software package. The software now automatically handles wide classes of biomolecules, such as proteins, nucleic acids and lipids, and comes with all commonly used force fields for these molecules built-in. GROMACS supports several implicit solvent models, as well as new free-energy algorithms, and the software now uses multithreading for efficient parallelization even on low-end systems, including windows-based workstations. Together with hand-tuned assembly kernels and state-of-the-art parallelization, this provides extremely high performance and cost efficiency for high-throughput as well as massively parallel simulations.

  13. Combining Amplification Typing of L1 Active Subfamilies (ATLAS) with High-Throughput Sequencing.

    Science.gov (United States)

    Rahbari, Raheleh; Badge, Richard M

    2016-01-01

    With the advent of new generations of high-throughput sequencing technologies, the catalog of human genome variants created by retrotransposon activity is expanding rapidly. However, despite these advances in describing L1 diversity and the fact that L1 must retrotranspose in the germline or prior to germline partitioning to be evolutionarily successful, direct assessment of de novo L1 retrotransposition in the germline or early embryogenesis has not been achieved for endogenous L1 elements. A direct study of de novo L1 retrotransposition into susceptible loci within sperm DNA (Freeman et al., Hum Mutat 32(8):978-988, 2011) suggested that the rate of L1 retrotransposition in the germline is much lower than previously estimated (ATLAS L1 display technique (Badge et al., Am J Hum Genet 72(4):823-838, 2003) to investigate de novo L1 retrotransposition in human genomes. In this chapter, we describe how we combined a high-coverage ATLAS variant with high-throughput sequencing, achieving 11-25× sequence depth per single amplicon, to study L1 retrotransposition in whole genome amplified (WGA) DNAs.

  14. A DNA fingerprinting procedure for ultra high-throughput genetic analysis of insects.

    Science.gov (United States)

    Schlipalius, D I; Waldron, J; Carroll, B J; Collins, P J; Ebert, P R

    2001-12-01

    Existing procedures for the generation of polymorphic DNA markers are not optimal for insect studies in which the organisms are often tiny and background molecular information is often non-existent. We have used a new high throughput DNA marker generation protocol called randomly amplified DNA fingerprints (RAF) to analyse the genetic variability in three separate strains of the stored grain pest, Rhyzopertha dominica. This protocol is quick, robust and reliable even though it requires minimal sample preparation, minute amounts of DNA and no prior molecular analysis of the organism. Arbitrarily selected oligonucleotide primers routinely produced approximately 50 scoreable polymorphic DNA markers, between individuals of three independent field isolates of R. dominica. Multivariate cluster analysis using forty-nine arbitrarily selected polymorphisms generated from a single primer reliably separated individuals into three clades corresponding to their geographical origin. The resulting clades were quite distinct, with an average genetic difference of 37.5 +/- 6.0% between clades and of 21.0 +/- 7.1% between individuals within clades. As a prelude to future gene mapping efforts, we have also assessed the performance of RAF under conditions commonly used in gene mapping. In this analysis, fingerprints from pooled DNA samples accurately and reproducibly reflected RAF profiles obtained from individual DNA samples that had been combined to create the bulked samples.

  15. A versatile, high through-put, bead-based phagocytosis assay for Plasmodium falciparum

    DEFF Research Database (Denmark)

    Lloyd, Yukie M.; Ngati, Elise P.; Salanti, Ali

    2017-01-01

    Antibody-mediated phagocytosis is an important immune effector mechanism against Plasmodium falciparum-infected erythrocytes (IE); however, current phagocytosis assays use IE collected from infected individuals or from in vitro cultures of P. falciparum, making them prone to high variation....... A simple, high-throughput flow cytometric assay was developed that uses THP-1 cells and fluorescent beads covalently-coupled with the malarial antigen VAR2CSA. The assay is highly repeatable, provides both the overall percent phagocytosis and semi-quantitates the number of antigen-coupled beads...

  16. Data for automated, high-throughput microscopy analysis of intracellular bacterial colonies using spot detection

    DEFF Research Database (Denmark)

    Ernstsen, Christina Lundgaard; Login, Frédéric H.; Jensen, Helene Halkjær

    2017-01-01

    Quantification of intracellular bacterial colonies is useful in strategies directed against bacterial attachment, subsequent cellular invasion and intracellular proliferation. An automated, high-throughput microscopy-method was established to quantify the number and size of intracellular bacteria...

  17. Evaluation of Simple and Inexpensive High-Throughput Methods for Phytic Acid Determination

    DEFF Research Database (Denmark)

    Raboy, Victor; Johnson, Amy; Bilyeu, Kristin

    2017-01-01

    High-throughput/low-cost/low-tech methods for phytic acid determination that are sufficiently accurate and reproducible would be of value in plant genetics, crop breeding and in the food and feed industries. Variants of two candidate methods, those described by Vaintraub and Lapteva (Anal Biochem...... and legume flours regardless of endogenous phytic acid levels or matrix constituents....

  18. Determining the optimal number of individual samples to pool for quantification of average herd levels of antimicrobial resistance genes in Danish pig herds using high-throughput qPCR

    DEFF Research Database (Denmark)

    Clasen, Julie; Mellerup, Anders; Olsen, John Elmerdahl

    2016-01-01

    The primary objective of this study was to determine the minimum number of individual fecal samples to pool together in order to obtain a representative sample for herd level quantification of antimicrobial resistance (AMR) genes in a Danish pig herd, using a novel high-throughput qPCR assay...

  19. PRIMEGENSw3: a web-based tool for high-throughput primer and probe design.

    Science.gov (United States)

    Kushwaha, Garima; Srivastava, Gyan Prakash; Xu, Dong

    2015-01-01

    Highly specific and efficient primer and probe design has been a major hurdle in many high-throughput techniques. Successful implementation of any PCR or probe hybridization technique depends on the quality of primers and probes used in terms of their specificity and cross-hybridization. Here we describe PRIMEGENSw3, a set of web-based utilities for high-throughput primer and probe design. These utilities allow users to select genomic regions and to design primer/probe for selected regions in an interactive, user-friendly, and automatic fashion. The system runs the PRIMEGENS algorithm in the back-end on the high-performance server with the stored genomic database or user-provided custom database for cross-hybridization check. Cross-hybridization is checked not only using BLAST but also by checking mismatch positions and energy calculation of potential hybridization hits. The results can be visualized online and also can be downloaded. The average success rate of primer design using PRIMEGENSw3 is ~90 %. The web server also supports primer design for methylated sequences, which is used in epigenetic studies. Stand-alone version of the software is also available for download at the website.

  20. “httk”: EPA’s Tool for High Throughput Toxicokinetics (CompTox CoP)

    Science.gov (United States)

    Thousands of chemicals have been pro?led by high-throughput screening programs such as ToxCast and Tox21; these chemicals are tested in part because most of them have limited or no data on hazard, exposure, or toxicokinetics. Toxicokinetic models aid in predicting tissue concentr...

  1. High-throughput analysis of endogenous fruit glycosyl hydrolases using a novel chromogenic hydrogel substrate assay

    DEFF Research Database (Denmark)

    Schückel, Julia; Kracun, Stjepan Kresimir; Lausen, Thomas Frederik

    2017-01-01

    A broad range of enzyme activities can be found in a wide range of different fruits and fruiting bodies but there is a lack of methods where many samples can be handled in a high-throughput and efficient manner. In particular, plant polysaccharide degrading enzymes – glycosyl hydrolases (GHs) play...... led to a more profound understanding of the importance of GH activity and regulation, current methods for determining glycosyl hydrolase activity are lacking in throughput and fail to keep up with data output from transcriptome research. Here we present the use of a versatile, easy...

  2. Ultrasensitive Single Fluorescence-Labeled Probe-Mediated Single Universal Primer-Multiplex-Droplet Digital Polymerase Chain Reaction for High-Throughput Genetically Modified Organism Screening.

    Science.gov (United States)

    Niu, Chenqi; Xu, Yuancong; Zhang, Chao; Zhu, Pengyu; Huang, Kunlun; Luo, Yunbo; Xu, Wentao

    2018-05-01

    As genetically modified (GM) technology develops and genetically modified organisms (GMOs) become more available, GMOs face increasing regulations and pressure to adhere to strict labeling guidelines. A singleplex detection method cannot perform the high-throughput analysis necessary for optimal GMO detection. Combining the advantages of multiplex detection and droplet digital polymerase chain reaction (ddPCR), a single universal primer-multiplex-ddPCR (SUP-M-ddPCR) strategy was proposed for accurate broad-spectrum screening and quantification. The SUP increases efficiency of the primers in PCR and plays an important role in establishing a high-throughput, multiplex detection method. Emerging ddPCR technology has been used for accurate quantification of nucleic acid molecules without a standard curve. Using maize as a reference point, four heterologous sequences ( 35S, NOS, NPTII, and PAT) were selected to evaluate the feasibility and applicability of this strategy. Surprisingly, these four genes cover more than 93% of the transgenic maize lines and serve as preliminary screening sequences. All screening probes were labeled with FAM fluorescence, which allows the signals from the samples with GMO content and those without to be easily differentiated. This fiveplex screening method is a new development in GMO screening. Utilizing an optimal amplification assay, the specificity, limit of detection (LOD), and limit of quantitation (LOQ) were validated. The LOD and LOQ of this GMO screening method were 0.1% and 0.01%, respectively, with a relative standard deviation (RSD) < 25%. This method could serve as an important tool for the detection of GM maize from different processed, commercially available products. Further, this screening method could be applied to other fields that require reliable and sensitive detection of DNA targets.

  3. Micropillar arrays as a high-throughput screening platform for therapeutics in multiple sclerosis.

    Science.gov (United States)

    Mei, Feng; Fancy, Stephen P J; Shen, Yun-An A; Niu, Jianqin; Zhao, Chao; Presley, Bryan; Miao, Edna; Lee, Seonok; Mayoral, Sonia R; Redmond, Stephanie A; Etxeberria, Ainhoa; Xiao, Lan; Franklin, Robin J M; Green, Ari; Hauser, Stephen L; Chan, Jonah R

    2014-08-01

    Functional screening for compounds that promote remyelination represents a major hurdle in the development of rational therapeutics for multiple sclerosis. Screening for remyelination is problematic, as myelination requires the presence of axons. Standard methods do not resolve cell-autonomous effects and are not suited for high-throughput formats. Here we describe a binary indicant for myelination using micropillar arrays (BIMA). Engineered with conical dimensions, micropillars permit resolution of the extent and length of membrane wrapping from a single two-dimensional image. Confocal imaging acquired from the base to the tip of the pillars allows for detection of concentric wrapping observed as 'rings' of myelin. The platform is formatted in 96-well plates, amenable to semiautomated random acquisition and automated detection and quantification. Upon screening 1,000 bioactive molecules, we identified a cluster of antimuscarinic compounds that enhance oligodendrocyte differentiation and remyelination. Our findings demonstrate a new high-throughput screening platform for potential regenerative therapeutics in multiple sclerosis.

  4. Use of high-throughput mass spectrometry to elucidate host-pathogen interactions in Salmonella

    Energy Technology Data Exchange (ETDEWEB)

    Rodland, Karin D.; Adkins, Joshua N.; Ansong, Charles; Chowdhury, Saiful M.; Manes, Nathan P.; Shi, Liang; Yoon, Hyunjin; Smith, Richard D.; Heffron, Fred

    2008-12-01

    New improvements to mass spectrometry include increased sensitivity, improvements in analyzing the collected data, and most important, from the standpoint of this review, a much higher throughput allowing analysis of many samples in a single day. This short review describes how host-pathogen interactions can be dissected by mass spectrometry using Salmonella as a model system. The approach allowed direct identification of the majority of annotate Salmonella proteins, how expression changed under various in vitro growth conditions, and how this relates to virulence and expression within host cell cells. One of the most significant findings is that a very high percentage of the all annotated genes (>20%) are regulated post-transcriptionally. In addition, new and unexpected interactions have been identified for several Salmonella virulence regulators that involve protein-protein interactions suggesting additional functions of the regulator in coordinating virulence expression. Overall high throughput mass spectrometer provides a new view of pathogen-host interaction emphasizing the protein products and defining how protein interactions determine the outcome of infection.

  5. A high content, high throughput cellular thermal stability assay for measuring drug-target engagement in living cells.

    Science.gov (United States)

    Massey, Andrew J

    2018-01-01

    Determining and understanding drug target engagement is critical for drug discovery. This can be challenging within living cells as selective readouts are often unavailable. Here we describe a novel method for measuring target engagement in living cells based on the principle of altered protein thermal stabilization / destabilization in response to ligand binding. This assay (HCIF-CETSA) utilizes high content, high throughput single cell immunofluorescent detection to determine target protein levels following heating of adherent cells in a 96 well plate format. We have used target engagement of Chk1 by potent small molecule inhibitors to validate the assay. Target engagement measured by this method was subsequently compared to target engagement measured by two alternative methods (autophosphorylation and CETSA). The HCIF-CETSA method appeared robust and a good correlation in target engagement measured by this method and CETSA for the selective Chk1 inhibitor V158411 was observed. However, these EC50 values were 23- and 12-fold greater than the autophosphorylation IC50. The described method is therefore a valuable advance in the CETSA method allowing the high throughput determination of target engagement in adherent cells.

  6. High throughput single-cell and multiple-cell micro-encapsulation.

    Science.gov (United States)

    Lagus, Todd P; Edd, Jon F

    2012-06-15

    Microfluidic encapsulation methods have been previously utilized to capture cells in picoliter-scale aqueous, monodisperse drops, providing confinement from a bulk fluid environment with applications in high throughput screening, cytometry, and mass spectrometry. We describe a method to not only encapsulate single cells, but to repeatedly capture a set number of cells (here we demonstrate one- and two-cell encapsulation) to study both isolation and the interactions between cells in groups of controlled sizes. By combining drop generation techniques with cell and particle ordering, we demonstrate controlled encapsulation of cell-sized particles for efficient, continuous encapsulation. Using an aqueous particle suspension and immiscible fluorocarbon oil, we generate aqueous drops in oil with a flow focusing nozzle. The aqueous flow rate is sufficiently high to create ordering of particles which reach the nozzle at integer multiple frequencies of the drop generation frequency, encapsulating a controlled number of cells in each drop. For representative results, 9.9 μm polystyrene particles are used as cell surrogates. This study shows a single-particle encapsulation efficiency P(k=1) of 83.7% and a double-particle encapsulation efficiency P(k=2) of 79.5% as compared to their respective Poisson efficiencies of 39.3% and 33.3%, respectively. The effect of consistent cell and particle concentration is demonstrated to be of major importance for efficient encapsulation, and dripping to jetting transitions are also addressed. Continuous media aqueous cell suspensions share a common fluid environment which allows cells to interact in parallel and also homogenizes the effects of specific cells in measurements from the media. High-throughput encapsulation of cells into picoliter-scale drops confines the samples to protect drops from cross-contamination, enable a measure of cellular diversity within samples, prevent dilution of reagents and expressed biomarkers, and amplify

  7. Molecular classification of fatty liver by high-throughput profiling of protein post-translational modifications.

    Science.gov (United States)

    Urasaki, Yasuyo; Fiscus, Ronald R; Le, Thuc T

    2016-04-01

    We describe an alternative approach to classifying fatty liver by profiling protein post-translational modifications (PTMs) with high-throughput capillary isoelectric focusing (cIEF) immunoassays. Four strains of mice were studied, with fatty livers induced by different causes, such as ageing, genetic mutation, acute drug usage, and high-fat diet. Nutrient-sensitive PTMs of a panel of 12 liver metabolic and signalling proteins were simultaneously evaluated with cIEF immunoassays, using nanograms of total cellular protein per assay. Changes to liver protein acetylation, phosphorylation, and O-N-acetylglucosamine glycosylation were quantified and compared between normal and diseased states. Fatty liver tissues could be distinguished from one another by distinctive protein PTM profiles. Fatty liver is currently classified by morphological assessment of lipid droplets, without identifying the underlying molecular causes. In contrast, high-throughput profiling of protein PTMs has the potential to provide molecular classification of fatty liver. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  8. Accurate Classification of Protein Subcellular Localization from High-Throughput Microscopy Images Using Deep Learning

    Directory of Open Access Journals (Sweden)

    Tanel Pärnamaa

    2017-05-01

    Full Text Available High-throughput microscopy of many single cells generates high-dimensional data that are far from straightforward to analyze. One important problem is automatically detecting the cellular compartment where a fluorescently-tagged protein resides, a task relatively simple for an experienced human, but difficult to automate on a computer. Here, we train an 11-layer neural network on data from mapping thousands of yeast proteins, achieving per cell localization classification accuracy of 91%, and per protein accuracy of 99% on held-out images. We confirm that low-level network features correspond to basic image characteristics, while deeper layers separate localization classes. Using this network as a feature calculator, we train standard classifiers that assign proteins to previously unseen compartments after observing only a small number of training examples. Our results are the most accurate subcellular localization classifications to date, and demonstrate the usefulness of deep learning for high-throughput microscopy.

  9. Accurate Classification of Protein Subcellular Localization from High-Throughput Microscopy Images Using Deep Learning.

    Science.gov (United States)

    Pärnamaa, Tanel; Parts, Leopold

    2017-05-05

    High-throughput microscopy of many single cells generates high-dimensional data that are far from straightforward to analyze. One important problem is automatically detecting the cellular compartment where a fluorescently-tagged protein resides, a task relatively simple for an experienced human, but difficult to automate on a computer. Here, we train an 11-layer neural network on data from mapping thousands of yeast proteins, achieving per cell localization classification accuracy of 91%, and per protein accuracy of 99% on held-out images. We confirm that low-level network features correspond to basic image characteristics, while deeper layers separate localization classes. Using this network as a feature calculator, we train standard classifiers that assign proteins to previously unseen compartments after observing only a small number of training examples. Our results are the most accurate subcellular localization classifications to date, and demonstrate the usefulness of deep learning for high-throughput microscopy. Copyright © 2017 Parnamaa and Parts.

  10. Parallel workflow for high-throughput (>1,000 samples/day quantitative analysis of human insulin-like growth factor 1 using mass spectrometric immunoassay.

    Directory of Open Access Journals (Sweden)

    Paul E Oran

    Full Text Available Insulin-like growth factor 1 (IGF1 is an important biomarker for the management of growth hormone disorders. Recently there has been rising interest in deploying mass spectrometric (MS methods of detection for measuring IGF1. However, widespread clinical adoption of any MS-based IGF1 assay will require increased throughput and speed to justify the costs of analyses, and robust industrial platforms that are reproducible across laboratories. Presented here is an MS-based quantitative IGF1 assay with performance rating of >1,000 samples/day, and a capability of quantifying IGF1 point mutations and posttranslational modifications. The throughput of the IGF1 mass spectrometric immunoassay (MSIA benefited from a simplified sample preparation step, IGF1 immunocapture in a tip format, and high-throughput MALDI-TOF MS analysis. The Limit of Detection and Limit of Quantification of the resulting assay were 1.5 μg/L and 5 μg/L, respectively, with intra- and inter-assay precision CVs of less than 10%, and good linearity and recovery characteristics. The IGF1 MSIA was benchmarked against commercially available IGF1 ELISA via Bland-Altman method comparison test, resulting in a slight positive bias of 16%. The IGF1 MSIA was employed in an optimized parallel workflow utilizing two pipetting robots and MALDI-TOF-MS instruments synced into one-hour phases of sample preparation, extraction and MSIA pipette tip elution, MS data collection, and data processing. Using this workflow, high-throughput IGF1 quantification of 1,054 human samples was achieved in approximately 9 hours. This rate of assaying is a significant improvement over existing MS-based IGF1 assays, and is on par with that of the enzyme-based immunoassays. Furthermore, a mutation was detected in ∼1% of the samples (SNP: rs17884626, creating an A→T substitution at position 67 of the IGF1, demonstrating the capability of IGF1 MSIA to detect point mutations and posttranslational modifications.

  11. High-Throughput Screening Using Fourier-Transform Infrared Imaging

    Directory of Open Access Journals (Sweden)

    Erdem Sasmaz

    2015-06-01

    Full Text Available Efficient parallel screening of combinatorial libraries is one of the most challenging aspects of the high-throughput (HT heterogeneous catalysis workflow. Today, a number of methods have been used in HT catalyst studies, including various optical, mass-spectrometry, and gas-chromatography techniques. Of these, rapid-scanning Fourier-transform infrared (FTIR imaging is one of the fastest and most versatile screening techniques. Here, the new design of the 16-channel HT reactor is presented and test results for its accuracy and reproducibility are shown. The performance of the system was evaluated through the oxidation of CO over commercial Pd/Al2O3 and cobalt oxide nanoparticles synthesized with different reducer-reductant molar ratios, surfactant types, metal and surfactant concentrations, synthesis temperatures, and ramp rates.

  12. Image Harvest: an open-source platform for high-throughput plant image processing and analysis.

    Science.gov (United States)

    Knecht, Avi C; Campbell, Malachy T; Caprez, Adam; Swanson, David R; Walia, Harkamal

    2016-05-01

    High-throughput plant phenotyping is an effective approach to bridge the genotype-to-phenotype gap in crops. Phenomics experiments typically result in large-scale image datasets, which are not amenable for processing on desktop computers, thus creating a bottleneck in the image-analysis pipeline. Here, we present an open-source, flexible image-analysis framework, called Image Harvest (IH), for processing images originating from high-throughput plant phenotyping platforms. Image Harvest is developed to perform parallel processing on computing grids and provides an integrated feature for metadata extraction from large-scale file organization. Moreover, the integration of IH with the Open Science Grid provides academic researchers with the computational resources required for processing large image datasets at no cost. Image Harvest also offers functionalities to extract digital traits from images to interpret plant architecture-related characteristics. To demonstrate the applications of these digital traits, a rice (Oryza sativa) diversity panel was phenotyped and genome-wide association mapping was performed using digital traits that are used to describe different plant ideotypes. Three major quantitative trait loci were identified on rice chromosomes 4 and 6, which co-localize with quantitative trait loci known to regulate agronomically important traits in rice. Image Harvest is an open-source software for high-throughput image processing that requires a minimal learning curve for plant biologists to analyzephenomics datasets. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  13. High-throughput mouse genotyping using robotics automation.

    Science.gov (United States)

    Linask, Kaari L; Lo, Cecilia W

    2005-02-01

    The use of mouse models is rapidly expanding in biomedical research. This has dictated the need for the rapid genotyping of mutant mouse colonies for more efficient utilization of animal holding space. We have established a high-throughput protocol for mouse genotyping using two robotics workstations: a liquid-handling robot to assemble PCR and a microfluidics electrophoresis robot for PCR product analysis. This dual-robotics setup incurs lower start-up costs than a fully automated system while still minimizing human intervention. Essential to this automation scheme is the construction of a database containing customized scripts for programming the robotics workstations. Using these scripts and the robotics systems, multiple combinations of genotyping reactions can be assembled simultaneously, allowing even complex genotyping data to be generated rapidly with consistency and accuracy. A detailed protocol, database, scripts, and additional background information are available at http://dir.nhlbi.nih.gov/labs/ldb-chd/autogene/.

  14. Sources of PCR-induced distortions in high-throughput sequencing data sets

    Science.gov (United States)

    Kebschull, Justus M.; Zador, Anthony M.

    2015-01-01

    PCR permits the exponential and sequence-specific amplification of DNA, even from minute starting quantities. PCR is a fundamental step in preparing DNA samples for high-throughput sequencing. However, there are errors associated with PCR-mediated amplification. Here we examine the effects of four important sources of error—bias, stochasticity, template switches and polymerase errors—on sequence representation in low-input next-generation sequencing libraries. We designed a pool of diverse PCR amplicons with a defined structure, and then used Illumina sequencing to search for signatures of each process. We further developed quantitative models for each process, and compared predictions of these models to our experimental data. We find that PCR stochasticity is the major force skewing sequence representation after amplification of a pool of unique DNA amplicons. Polymerase errors become very common in later cycles of PCR but have little impact on the overall sequence distribution as they are confined to small copy numbers. PCR template switches are rare and confined to low copy numbers. Our results provide a theoretical basis for removing distortions from high-throughput sequencing data. In addition, our findings on PCR stochasticity will have particular relevance to quantification of results from single cell sequencing, in which sequences are represented by only one or a few molecules. PMID:26187991

  15. High throughput and accurate serum proteome profiling by integrated sample preparation technology and single-run data independent mass spectrometry analysis.

    Science.gov (United States)

    Lin, Lin; Zheng, Jiaxin; Yu, Quan; Chen, Wendong; Xing, Jinchun; Chen, Chenxi; Tian, Ruijun

    2018-03-01

    Mass spectrometry (MS)-based serum proteome analysis is extremely challenging due to its high complexity and dynamic range of protein abundances. Developing high throughput and accurate serum proteomic profiling approach capable of analyzing large cohorts is urgently needed for biomarker discovery. Herein, we report a streamlined workflow for fast and accurate proteomic profiling from 1μL of blood serum. The workflow combined an integrated technique for highly sensitive and reproducible sample preparation and a new data-independent acquisition (DIA)-based MS method. Comparing with standard data dependent acquisition (DDA) approach, the optimized DIA method doubled the number of detected peptides and proteins with better reproducibility. Without protein immunodepletion and prefractionation, the single-run DIA analysis enables quantitative profiling of over 300 proteins with 50min gradient time. The quantified proteins span more than five orders of magnitude of abundance range and contain over 50 FDA-approved disease markers. The workflow allowed us to analyze 20 serum samples per day, with about 358 protein groups per sample being identified. A proof-of-concept study on renal cell carcinoma (RCC) serum samples confirmed the feasibility of the workflow for large scale serum proteomic profiling and disease-related biomarker discovery. Blood serum or plasma is the predominant specimen for clinical proteomic studies while the analysis is extremely challenging for its high complexity. Many efforts had been made in the past for serum proteomics for maximizing protein identifications, whereas few have been concerned with throughput and reproducibility. Here, we establish a rapid, robust and high reproducible DIA-based workflow for streamlined serum proteomic profiling from 1μL serum. The workflow doesn't need protein depletion and pre-fractionation, while still being able to detect disease-relevant proteins accurately. The workflow is promising in clinical application

  16. High throughput second harmonic imaging for label-free biological applications

    KAUST Repository

    Macias Romero, Carlos; Didier, Marie E P; Jourdain, Pascal; Marquet, Pierre; Magistretti, Pierre J.; Tarun, Orly B.; Zubkovs, Vitalijs; Radenovic, Aleksandra; Roke, Sylvie

    2014-01-01

    Second harmonic generation (SHG) is inherently sensitive to the absence of spatial centrosymmetry, which can render it intrinsically sensitive to interfacial processes, chemical changes and electrochemical responses. Here, we seek to improve the imaging throughput of SHG microscopy by using a wide-field imaging scheme in combination with a medium-range repetition rate amplified near infrared femtosecond laser source and gated detection. The imaging throughput of this configuration is tested by measuring the optical image contrast for different image acquisition times of BaTiO3 nanoparticles in two different wide-field setups and one commercial point-scanning configuration. We find that the second harmonic imaging throughput is improved by 2-3 orders of magnitude compared to point-scan imaging. Capitalizing on this result, we perform low fluence imaging of (parts of) living mammalian neurons in culture.

  17. Machine Learning for High-Throughput Stress Phenotyping in Plants.

    Science.gov (United States)

    Singh, Arti; Ganapathysubramanian, Baskar; Singh, Asheesh Kumar; Sarkar, Soumik

    2016-02-01

    Advances in automated and high-throughput imaging technologies have resulted in a deluge of high-resolution images and sensor data of plants. However, extracting patterns and features from this large corpus of data requires the use of machine learning (ML) tools to enable data assimilation and feature identification for stress phenotyping. Four stages of the decision cycle in plant stress phenotyping and plant breeding activities where different ML approaches can be deployed are (i) identification, (ii) classification, (iii) quantification, and (iv) prediction (ICQP). We provide here a comprehensive overview and user-friendly taxonomy of ML tools to enable the plant community to correctly and easily apply the appropriate ML tools and best-practice guidelines for various biotic and abiotic stress traits. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. HTP-OligoDesigner: An Online Primer Design Tool for High-Throughput Gene Cloning and Site-Directed Mutagenesis.

    Science.gov (United States)

    Camilo, Cesar M; Lima, Gustavo M A; Maluf, Fernando V; Guido, Rafael V C; Polikarpov, Igor

    2016-01-01

    Following burgeoning genomic and transcriptomic sequencing data, biochemical and molecular biology groups worldwide are implementing high-throughput cloning and mutagenesis facilities in order to obtain a large number of soluble proteins for structural and functional characterization. Since manual primer design can be a time-consuming and error-generating step, particularly when working with hundreds of targets, the automation of primer design process becomes highly desirable. HTP-OligoDesigner was created to provide the scientific community with a simple and intuitive online primer design tool for both laboratory-scale and high-throughput projects of sequence-independent gene cloning and site-directed mutagenesis and a Tm calculator for quick queries.

  19. Real-time, high-throughput measurements of peptide-MHC-I dissociation using a scintillation proximity assay

    DEFF Research Database (Denmark)

    Harndahl, Mikkel; Rasmussen, Michael; Røder, Gustav Andreas

    2011-01-01

    and it is well suited for high-throughput screening. To exemplify this, we screened a panel of 384 high-affinity peptides binding to the MHC class I molecule, HLA-A*02:01, and observed the rates of dissociation that ranged from 0.1h to 46h depending on the peptide used.......Efficient presentation of peptide-MHC class I complexes to immune T cells depends upon stable peptide-MHC class I interactions. Theoretically, determining the rate of dissociation of a peptide-MHC class I complexes is straightforward; in practical terms, however, generating the accurate and closely...... timed data needed to determine the rate of dissociation is not simple. Ideally, one should use a homogenous assay involving an inexhaustible and label-free assay principle. Here, we present a homogenous, high-throughput peptide-MHC class I dissociation assay, which by and large fulfill these ideal...

  20. High-throughput characterization of film thickness in thin film materials libraries by digital holographic microscopy

    International Nuclear Information System (INIS)

    Lai Yiuwai; Hofmann, Martin R; Ludwig, Alfred; Krause, Michael; Savan, Alan; Thienhaus, Sigurd; Koukourakis, Nektarios

    2011-01-01

    A high-throughput characterization technique based on digital holography for mapping film thickness in thin-film materials libraries was developed. Digital holographic microscopy is used for fully automatic measurements of the thickness of patterned films with nanometer resolution. The method has several significant advantages over conventional stylus profilometry: it is contactless and fast, substrate bending is compensated, and the experimental setup is simple. Patterned films prepared by different combinatorial thin-film approaches were characterized to investigate and demonstrate this method. The results show that this technique is valuable for the quick, reliable and high-throughput determination of the film thickness distribution in combinatorial materials research. Importantly, it can also be applied to thin films that have been structured by shadow masking.

  1. High-throughput characterization of film thickness in thin film materials libraries by digital holographic microscopy.

    Science.gov (United States)

    Lai, Yiu Wai; Krause, Michael; Savan, Alan; Thienhaus, Sigurd; Koukourakis, Nektarios; Hofmann, Martin R; Ludwig, Alfred

    2011-10-01

    A high-throughput characterization technique based on digital holography for mapping film thickness in thin-film materials libraries was developed. Digital holographic microscopy is used for fully automatic measurements of the thickness of patterned films with nanometer resolution. The method has several significant advantages over conventional stylus profilometry: it is contactless and fast, substrate bending is compensated, and the experimental setup is simple. Patterned films prepared by different combinatorial thin-film approaches were characterized to investigate and demonstrate this method. The results show that this technique is valuable for the quick, reliable and high-throughput determination of the film thickness distribution in combinatorial materials research. Importantly, it can also be applied to thin films that have been structured by shadow masking.

  2. Gold-coated polydimethylsiloxane microwells for high-throughput electrochemiluminescence analysis of intracellular glucose at single cells.

    Science.gov (United States)

    Xia, Juan; Zhou, Junyu; Zhang, Ronggui; Jiang, Dechen; Jiang, Depeng

    2018-06-04

    In this communication, a gold-coated polydimethylsiloxane (PDMS) chip with cell-sized microwells was prepared through a stamping and spraying process that was applied directly for high-throughput electrochemiluminescence (ECL) analysis of intracellular glucose at single cells. As compared with the previous multiple-step fabrication of photoresist-based microwells on the electrode, the preparation process is simple and offers fresh electrode surface for higher luminescence intensity. More luminescence intensity was recorded from cell-retained microwells than that at the planar region among the microwells that was correlated with the content of intracellular glucose. The successful monitoring of intracellular glucose at single cells using this PDMS chip will provide an alternative strategy for high-throughput single-cell analysis. Graphical abstract ᅟ.

  3. Microengineering methods for cell-based microarrays and high-throughput drug-screening applications

    International Nuclear Information System (INIS)

    Xu Feng; Wu Jinhui; Wang Shuqi; Gurkan, Umut Atakan; Demirci, Utkan; Durmus, Naside Gozde

    2011-01-01

    Screening for effective therapeutic agents from millions of drug candidates is costly, time consuming, and often faces concerns due to the extensive use of animals. To improve cost effectiveness, and to minimize animal testing in pharmaceutical research, in vitro monolayer cell microarrays with multiwell plate assays have been developed. Integration of cell microarrays with microfluidic systems has facilitated automated and controlled component loading, significantly reducing the consumption of the candidate compounds and the target cells. Even though these methods significantly increased the throughput compared to conventional in vitro testing systems and in vivo animal models, the cost associated with these platforms remains prohibitively high. Besides, there is a need for three-dimensional (3D) cell-based drug-screening models which can mimic the in vivo microenvironment and the functionality of the native tissues. Here, we present the state-of-the-art microengineering approaches that can be used to develop 3D cell-based drug-screening assays. We highlight the 3D in vitro cell culture systems with live cell-based arrays, microfluidic cell culture systems, and their application to high-throughput drug screening. We conclude that among the emerging microengineering approaches, bioprinting holds great potential to provide repeatable 3D cell-based constructs with high temporal, spatial control and versatility.

  4. Microengineering methods for cell-based microarrays and high-throughput drug-screening applications

    Energy Technology Data Exchange (ETDEWEB)

    Xu Feng; Wu Jinhui; Wang Shuqi; Gurkan, Umut Atakan; Demirci, Utkan [Department of Medicine, Demirci Bio-Acoustic-MEMS in Medicine (BAMM) Laboratory, Center for Biomedical Engineering, Brigham and Women' s Hospital, Harvard Medical School, Boston, MA (United States); Durmus, Naside Gozde, E-mail: udemirci@rics.bwh.harvard.edu [School of Engineering and Division of Biology and Medicine, Brown University, Providence, RI (United States)

    2011-09-15

    Screening for effective therapeutic agents from millions of drug candidates is costly, time consuming, and often faces concerns due to the extensive use of animals. To improve cost effectiveness, and to minimize animal testing in pharmaceutical research, in vitro monolayer cell microarrays with multiwell plate assays have been developed. Integration of cell microarrays with microfluidic systems has facilitated automated and controlled component loading, significantly reducing the consumption of the candidate compounds and the target cells. Even though these methods significantly increased the throughput compared to conventional in vitro testing systems and in vivo animal models, the cost associated with these platforms remains prohibitively high. Besides, there is a need for three-dimensional (3D) cell-based drug-screening models which can mimic the in vivo microenvironment and the functionality of the native tissues. Here, we present the state-of-the-art microengineering approaches that can be used to develop 3D cell-based drug-screening assays. We highlight the 3D in vitro cell culture systems with live cell-based arrays, microfluidic cell culture systems, and their application to high-throughput drug screening. We conclude that among the emerging microengineering approaches, bioprinting holds great potential to provide repeatable 3D cell-based constructs with high temporal, spatial control and versatility.

  5. High-throughput differentiation of heparin from other glycosaminoglycans by pyrolysis mass spectrometry.

    Science.gov (United States)

    Nemes, Peter; Hoover, William J; Keire, David A

    2013-08-06

    Sensors with high chemical specificity and enhanced sample throughput are vital to screening food products and medical devices for chemical or biochemical contaminants that may pose a threat to public health. For example, the rapid detection of oversulfated chondroitin sulfate (OSCS) in heparin could prevent reoccurrence of heparin adulteration that caused hundreds of severe adverse events including deaths worldwide in 2007-2008. Here, rapid pyrolysis is integrated with direct analysis in real time (DART) mass spectrometry to rapidly screen major glycosaminoglycans, including heparin, chondroitin sulfate A, dermatan sulfate, and OSCS. The results demonstrate that, compared to traditional liquid chromatography-based analyses, pyrolysis mass spectrometry achieved at least 250-fold higher sample throughput and was compatible with samples volume-limited to about 300 nL. Pyrolysis yielded an abundance of fragment ions (e.g., 150 different m/z species), many of which were specific to the parent compound. Using multivariate and statistical data analysis models, these data enabled facile differentiation of the glycosaminoglycans with high throughput. After method development was completed, authentically contaminated samples obtained during the heparin crisis by the FDA were analyzed in a blinded manner for OSCS contamination. The lower limit of differentiation and detection were 0.1% (w/w) OSCS in heparin and 100 ng/μL (20 ng) OSCS in water, respectively. For quantitative purposes the linear dynamic range spanned approximately 3 orders of magnitude. Moreover, this chemical readout was successfully employed to find clues in the manufacturing history of the heparin samples that can be used for surveillance purposes. The presented technology and data analysis protocols are anticipated to be readily adaptable to other chemical and biochemical agents and volume-limited samples.

  6. Versatile High-Throughput Fluorescence Assay for Monitoring Cas9 Activity.

    Science.gov (United States)

    Seamon, Kyle J; Light, Yooli K; Saada, Edwin A; Schoeniger, Joseph S; Harmon, Brooke

    2018-06-05

    The RNA-guided DNA nuclease Cas9 is now widely used for the targeted modification of genomes of human cells and various organisms. Despite the extensive use of Clustered Regularly Interspaced Palindromic Repeats (CRISPR) systems for genome engineering and the rapid discovery and engineering of new CRISPR-associated nucleases, there are no high-throughput assays for measuring enzymatic activity. The current laboratory and future therapeutic uses of CRISPR technology have a significant risk of accidental exposure or clinical off-target effects, underscoring the need for therapeutically effective inhibitors of Cas9. Here, we develop a fluorescence assay for monitoring Cas9 nuclease activity and demonstrate its utility with S. pyogenes (Spy), S. aureus (Sau), and C. jejuni (Cje) Cas9. The assay was validated by quantitatively profiling the species specificity of published anti-CRISPR (Acr) proteins, confirming the reported inhibition of Spy Cas9 by AcrIIA4 and Cje Cas9 by AcrIIC1 and no inhibition of Sau Cas9 by either anti-CRISPR. To identify drug-like inhibitors, we performed a screen of 189 606 small molecules for inhibition of Spy Cas9. Of 437 hits (0.2% hit rate), six were confirmed as Cas9 inhibitors in a direct gel electrophoresis secondary assay. The high-throughput nature of this assay makes it broadly applicable for the discovery of additional Cas9 inhibitors or the characterization of Cas9 enzyme variants.

  7. High-Throughput and Low-Latency Network Communication with NetIO

    Science.gov (United States)

    Schumacher, Jörn; Plessl, Christian; Vandelli, Wainer

    2017-10-01

    HPC network technologies like Infiniband, TrueScale or OmniPath provide low- latency and high-throughput communication between hosts, which makes them attractive options for data-acquisition systems in large-scale high-energy physics experiments. Like HPC networks, DAQ networks are local and include a well specified number of systems. Unfortunately traditional network communication APIs for HPC clusters like MPI or PGAS exclusively target the HPC community and are not suited well for DAQ applications. It is possible to build distributed DAQ applications using low-level system APIs like Infiniband Verbs, but it requires a non-negligible effort and expert knowledge. At the same time, message services like ZeroMQ have gained popularity in the HEP community. They make it possible to build distributed applications with a high-level approach and provide good performance. Unfortunately, their usage usually limits developers to TCP/IP- based networks. While it is possible to operate a TCP/IP stack on top of Infiniband and OmniPath, this approach may not be very efficient compared to a direct use of native APIs. NetIO is a simple, novel asynchronous message service that can operate on Ethernet, Infiniband and similar network fabrics. In this paper the design and implementation of NetIO is presented and described, and its use is evaluated in comparison to other approaches. NetIO supports different high-level programming models and typical workloads of HEP applications. The ATLAS FELIX project [1] successfully uses NetIO as its central communication platform. The architecture of NetIO is described in this paper, including the user-level API and the internal data-flow design. The paper includes a performance evaluation of NetIO including throughput and latency measurements. The performance is compared against the state-of-the- art ZeroMQ message service. Performance measurements are performed in a lab environment with Ethernet and FDR Infiniband networks.

  8. Rapid 2,2'-bicinchoninic-based xylanase assay compatible with high throughput screening

    Science.gov (United States)

    William R. Kenealy; Thomas W. Jeffries

    2003-01-01

    High-throughput screening requires simple assays that give reliable quantitative results. A microplate assay was developed for reducing sugar analysis that uses a 2,2'-bicinchoninic-based protein reagent. Endo-1,4-â-D-xylanase activity against oat spelt xylan was detected at activities of 0.002 to 0.011 IU ml−1. The assay is linear for sugar...

  9. Theory and implementation of a very high throughput true random number generator in field programmable gate array

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yonggang, E-mail: wangyg@ustc.edu.cn; Hui, Cong; Liu, Chong; Xu, Chao [Department of Modern Physics, University of Science and Technology of China, Hefei 230026 (China)

    2016-04-15

    The contribution of this paper is proposing a new entropy extraction mechanism based on sampling phase jitter in ring oscillators to make a high throughput true random number generator in a field programmable gate array (FPGA) practical. Starting from experimental observation and analysis of the entropy source in FPGA, a multi-phase sampling method is exploited to harvest the clock jitter with a maximum entropy and fast sampling speed. This parametrized design is implemented in a Xilinx Artix-7 FPGA, where the carry chains in the FPGA are explored to realize the precise phase shifting. The generator circuit is simple and resource-saving, so that multiple generation channels can run in parallel to scale the output throughput for specific applications. The prototype integrates 64 circuit units in the FPGA to provide a total output throughput of 7.68 Gbps, which meets the requirement of current high-speed quantum key distribution systems. The randomness evaluation, as well as its robustness to ambient temperature, confirms that the new method in a purely digital fashion can provide high-speed high-quality random bit sequences for a variety of embedded applications.

  10. High Throughput System for Plant Height and Hyperspectral Measurement

    Science.gov (United States)

    Zhao, H.; Xu, L.; Jiang, H.; Shi, S.; Chen, D.

    2018-04-01

    Hyperspectral and three-dimensional measurement can obtain the intrinsic physicochemical properties and external geometrical characteristics of objects, respectively. Currently, a variety of sensors are integrated into a system to collect spectral and morphological information in agriculture. However, previous experiments were usually performed with several commercial devices on a single platform. Inadequate registration and synchronization among instruments often resulted in mismatch between spectral and 3D information of the same target. And narrow field of view (FOV) extends the working hours in farms. Therefore, we propose a high throughput prototype that combines stereo vision and grating dispersion to simultaneously acquire hyperspectral and 3D information.

  11. HIGH THROUGHPUT SYSTEM FOR PLANT HEIGHT AND HYPERSPECTRAL MEASUREMENT

    Directory of Open Access Journals (Sweden)

    H. Zhao

    2018-04-01

    Full Text Available Hyperspectral and three-dimensional measurement can obtain the intrinsic physicochemical properties and external geometrical characteristics of objects, respectively. Currently, a variety of sensors are integrated into a system to collect spectral and morphological information in agriculture. However, previous experiments were usually performed with several commercial devices on a single platform. Inadequate registration and synchronization among instruments often resulted in mismatch between spectral and 3D information of the same target. And narrow field of view (FOV extends the working hours in farms. Therefore, we propose a high throughput prototype that combines stereo vision and grating dispersion to simultaneously acquire hyperspectral and 3D information.

  12. ZNStress: a high-throughput drug screening protocol for identification of compounds modulating neuronal stress in the transgenic mutant sod1G93R zebrafish model of amyotrophic lateral sclerosis.

    Science.gov (United States)

    McGown, Alexander; Shaw, Dame Pamela J; Ramesh, Tennore

    2016-07-26

    Amyotrophic lateral sclerosis (ALS) is a lethal neurodegenerative disease with death on average within 2-3 years of symptom onset. Mutations in superoxide dismutase 1 (SOD1) have been identified to cause ALS. Riluzole, the only neuroprotective drug for ALS provides life extension of only 3 months on average. Thishighlights the need for compound screening in disease models to identify new neuroprotective therapies for this disease. Zebrafish is an emerging model system that is well suited for the study of diseasepathophysiology and also for high throughput (HT) drug screening. The mutant sod1 zebrafish model of ALS mimics the hallmark features of ALS. Using a fluorescence based readout of neuronal stress, we developed a high throughput (HT) screen to identify neuroprotective compounds. Here we show that the zebrafish screen is a robust system that can be used to rapidly screen thousands ofcompounds and also demonstrate that riluzole is capable of reducing neuronal stress in this model system. The screen shows optimal quality control, maintaining a high sensitivity and specificity withoutcompromising throughput. Most importantly, we demonstrate that many compounds previously failed in human clinical trials, showed no stress reducing activity in the zebrafish assay. We conclude that HT drug screening using a mutant sod1 zebrafish is a reliable model system which supplemented with secondary assays would be useful in identifying drugs with potential for neuroprotective efficacy in ALS.

  13. DESIGN OF LOW EPI AND HIGH THROUGHPUT CORDIC CELL TO IMPROVE THE PERFORMANCE OF MOBILE ROBOT

    Directory of Open Access Journals (Sweden)

    P. VELRAJKUMAR

    2014-04-01

    Full Text Available This paper mainly focuses on pass logic based design, which gives an low Energy Per Instruction (EPI and high throughput COrdinate Rotation Digital Computer (CORDIC cell for application of robotic exploration. The basic components of CORDIC cell namely register, multiplexer and proposed adder is designed using pass transistor logic (PTL design. The proposed adder is implemented in bit-parallel iterative CORDIC circuit whereas designed using DSCH2 VLSI CAD tool and their layouts are generated by Microwind 3 VLSI CAD tool. The propagation delay, area and power dissipation are calculated from the simulated results for proposed adder based CORDIC cell. The EPI, throughput and effect of temperature are calculated from generated layout. The output parameter of generated layout is analysed using BSIM4 advanced analyzer. The simulated result of the proposed adder based CORDIC circuit is compared with other adder based CORDIC circuits. From the analysis of these simulated results, it was found that the proposed adder based CORDIC circuit dissipates low power, gives faster response, low EPI and high throughput.

  14. SINA: accurate high-throughput multiple sequence alignment of ribosomal RNA genes.

    Science.gov (United States)

    Pruesse, Elmar; Peplies, Jörg; Glöckner, Frank Oliver

    2012-07-15

    In the analysis of homologous sequences, computation of multiple sequence alignments (MSAs) has become a bottleneck. This is especially troublesome for marker genes like the ribosomal RNA (rRNA) where already millions of sequences are publicly available and individual studies can easily produce hundreds of thousands of new sequences. Methods have been developed to cope with such numbers, but further improvements are needed to meet accuracy requirements. In this study, we present the SILVA Incremental Aligner (SINA) used to align the rRNA gene databases provided by the SILVA ribosomal RNA project. SINA uses a combination of k-mer searching and partial order alignment (POA) to maintain very high alignment accuracy while satisfying high throughput performance demands. SINA was evaluated in comparison with the commonly used high throughput MSA programs PyNAST and mothur. The three BRAliBase III benchmark MSAs could be reproduced with 99.3, 97.6 and 96.1 accuracy. A larger benchmark MSA comprising 38 772 sequences could be reproduced with 98.9 and 99.3% accuracy using reference MSAs comprising 1000 and 5000 sequences. SINA was able to achieve higher accuracy than PyNAST and mothur in all performed benchmarks. Alignment of up to 500 sequences using the latest SILVA SSU/LSU Ref datasets as reference MSA is offered at http://www.arb-silva.de/aligner. This page also links to Linux binaries, user manual and tutorial. SINA is made available under a personal use license.

  15. Morphology control in polymer blend fibers—a high throughput computing approach

    Science.gov (United States)

    Sesha Sarath Pokuri, Balaji; Ganapathysubramanian, Baskar

    2016-08-01

    Fibers made from polymer blends have conventionally enjoyed wide use, particularly in textiles. This wide applicability is primarily aided by the ease of manufacturing such fibers. More recently, the ability to tailor the internal morphology of polymer blend fibers by carefully designing processing conditions has enabled such fibers to be used in technologically relevant applications. Some examples include anisotropic insulating properties for heat and anisotropic wicking of moisture, coaxial morphologies for optical applications as well as fibers with high internal surface area for filtration and catalysis applications. However, identifying the appropriate processing conditions from the large space of possibilities using conventional trial-and-error approaches is a tedious and resource-intensive process. Here, we illustrate a high throughput computational approach to rapidly explore and characterize how processing conditions (specifically blend ratio and evaporation rates) affect the internal morphology of polymer blends during solvent based fabrication. We focus on a PS: PMMA system and identify two distinct classes of morphologies formed due to variations in the processing conditions. We subsequently map the processing conditions to the morphology class, thus constructing a ‘phase diagram’ that enables rapid identification of processing parameters for specific morphology class. We finally demonstrate the potential for time dependent processing conditions to get desired features of the morphology. This opens up the possibility of rational stage-wise design of processing pathways for tailored fiber morphology using high throughput computing.

  16. Laser-Induced Fluorescence Detection in High-Throughput Screening of Heterogeneous Catalysts and Single Cells Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Su, Hui [Iowa State Univ., Ames, IA (United States)

    2001-01-01

    Laser-induced fluorescence detection is one of the most sensitive detection techniques and it has found enormous applications in various areas. The purpose of this research was to develop detection approaches based on laser-induced fluorescence detection in two different areas, heterogeneous catalysts screening and single cell study. First, we introduced laser-induced imaging (LIFI) as a high-throughput screening technique for heterogeneous catalysts to explore the use of this high-throughput screening technique in discovery and study of various heterogeneous catalyst systems. This scheme is based on the fact that the creation or the destruction of chemical bonds alters the fluorescence properties of suitably designed molecules. By irradiating the region immediately above the catalytic surface with a laser, the fluorescence intensity of a selected product or reactant can be imaged by a charge-coupled device (CCD) camera to follow the catalytic activity as a function of time and space. By screening the catalytic activity of vanadium pentoxide catalysts in oxidation of naphthalene, we demonstrated LIFI has good detection performance and the spatial and temporal resolution needed for high-throughput screening of heterogeneous catalysts. The sample packing density can reach up to 250 x 250 subunits/cm2 for 40-μm wells. This experimental set-up also can screen solid catalysts via near infrared thermography detection.

  17. Laser-Induced Fluorescence Detection in High-Throughput Screening of Heterogeneous Catalysts and Single Cells Analysis

    International Nuclear Information System (INIS)

    Hui Su

    2001-01-01

    Laser-induced fluorescence detection is one of the most sensitive detection techniques and it has found enormous applications in various areas. The purpose of this research was to develop detection approaches based on laser-induced fluorescence detection in two different areas, heterogeneous catalysts screening and single cell study. First, we introduced laser-induced imaging (LIFI) as a high-throughput screening technique for heterogeneous catalysts to explore the use of this high-throughput screening technique in discovery and study of various heterogeneous catalyst systems. This scheme is based on the fact that the creation or the destruction of chemical bonds alters the fluorescence properties of suitably designed molecules. By irradiating the region immediately above the catalytic surface with a laser, the fluorescence intensity of a selected product or reactant can be imaged by a charge-coupled device (CCD) camera to follow the catalytic activity as a function of time and space. By screening the catalytic activity of vanadium pentoxide catalysts in oxidation of naphthalene, we demonstrated LIFI has good detection performance and the spatial and temporal resolution needed for high-throughput screening of heterogeneous catalysts. The sample packing density can reach up to 250 x 250 subunits/cm(sub 2) for 40-(micro)m wells. This experimental set-up also can screen solid catalysts via near infrared thermography detection

  18. GROMACS 4.5: a high-throughput and highly parallel open source molecular simulation toolkit.

    Science.gov (United States)

    Pronk, Sander; Páll, Szilárd; Schulz, Roland; Larsson, Per; Bjelkmar, Pär; Apostolov, Rossen; Shirts, Michael R; Smith, Jeremy C; Kasson, Peter M; van der Spoel, David; Hess, Berk; Lindahl, Erik

    2013-04-01

    Molecular simulation has historically been a low-throughput technique, but faster computers and increasing amounts of genomic and structural data are changing this by enabling large-scale automated simulation of, for instance, many conformers or mutants of biomolecules with or without a range of ligands. At the same time, advances in performance and scaling now make it possible to model complex biomolecular interaction and function in a manner directly testable by experiment. These applications share a need for fast and efficient software that can be deployed on massive scale in clusters, web servers, distributed computing or cloud resources. Here, we present a range of new simulation algorithms and features developed during the past 4 years, leading up to the GROMACS 4.5 software package. The software now automatically handles wide classes of biomolecules, such as proteins, nucleic acids and lipids, and comes with all commonly used force fields for these molecules built-in. GROMACS supports several implicit solvent models, as well as new free-energy algorithms, and the software now uses multithreading for efficient parallelization even on low-end systems, including windows-based workstations. Together with hand-tuned assembly kernels and state-of-the-art parallelization, this provides extremely high performance and cost efficiency for high-throughput as well as massively parallel simulations. GROMACS is an open source and free software available from http://www.gromacs.org. Supplementary data are available at Bioinformatics online.

  19. High-throughput Cloning and Expression of Integral Membrane Proteins in Escherichia coli

    Science.gov (United States)

    Bruni, Renato

    2014-01-01

    Recently, several structural genomics centers have been established and a remarkable number of three-dimensional structures of soluble proteins have been solved. For membrane proteins, the number of structures solved has been significantly trailing those for their soluble counterparts, not least because over-expression and purification of membrane proteins is a much more arduous process. By using high throughput technologies, a large number of membrane protein targets can be screened simultaneously and a greater number of expression and purification conditions can be employed, leading to a higher probability of successfully determining the structure of membrane proteins. This unit describes the cloning, expression and screening of membrane proteins using high throughput methodologies developed in our laboratory. Basic Protocol 1 deals with the cloning of inserts into expression vectors by ligation-independent cloning. Basic Protocol 2 describes the expression and purification of the target proteins on a miniscale. Lastly, for the targets that express at the miniscale, basic protocols 3 and 4 outline the methods employed for the expression and purification of targets at the midi-scale, as well as a procedure for detergent screening and identification of detergent(s) in which the target protein is stable. PMID:24510647

  20. High-throughput kinase assays with protein substrates using fluorescent polymer superquenching

    Directory of Open Access Journals (Sweden)

    Weatherford Wendy

    2005-05-01

    Full Text Available Abstract Background High-throughput screening is used by the pharmaceutical industry for identifying lead compounds that interact with targets of pharmacological interest. Because of the key role that aberrant regulation of protein phosphorylation plays in diseases such as cancer, diabetes and hypertension, kinases have become one of the main drug targets. With the exception of antibody-based assays, methods to screen for specific kinase activity are generally restricted to the use of small synthetic peptides as substrates. However, the use of natural protein substrates has the advantage that potential inhibitors can be detected that affect enzyme activity by binding to a site other than the catalytic site. We have previously reported a non-radioactive and non-antibody-based fluorescence quench assay for detection of phosphorylation or dephosphorylation using synthetic peptide substrates. The aim of this work is to develop an assay for detection of phosphorylation of chemically unmodified proteins based on this polymer superquenching platform. Results Using a modified QTL Lightspeed™ assay, phosphorylation of native protein was quantified by the interaction of the phosphorylated proteins with metal-ion coordinating groups co-located with fluorescent polymer deposited onto microspheres. The binding of phospho-protein inhibits a dye-labeled "tracer" peptide from associating to the phosphate-binding sites present on the fluorescent microspheres. The resulting inhibition of quench generates a "turn on" assay, in which the signal correlates with the phosphorylation of the substrate. The assay was tested on three different proteins: Myelin Basic Protein (MBP, Histone H1 and Phosphorylated heat- and acid-stable protein (PHAS-1. Phosphorylation of the proteins was detected by Protein Kinase Cα (PKCα and by the Interleukin -1 Receptor-associated Kinase 4 (IRAK4. Enzyme inhibition yielded IC50 values that were comparable to those obtained using

  1. High-throughput kinase assays with protein substrates using fluorescent polymer superquenching.

    Science.gov (United States)

    Rininsland, Frauke; Stankewicz, Casey; Weatherford, Wendy; McBranch, Duncan

    2005-05-31

    High-throughput screening is used by the pharmaceutical industry for identifying lead compounds that interact with targets of pharmacological interest. Because of the key role that aberrant regulation of protein phosphorylation plays in diseases such as cancer, diabetes and hypertension, kinases have become one of the main drug targets. With the exception of antibody-based assays, methods to screen for specific kinase activity are generally restricted to the use of small synthetic peptides as substrates. However, the use of natural protein substrates has the advantage that potential inhibitors can be detected that affect enzyme activity by binding to a site other than the catalytic site. We have previously reported a non-radioactive and non-antibody-based fluorescence quench assay for detection of phosphorylation or dephosphorylation using synthetic peptide substrates. The aim of this work is to develop an assay for detection of phosphorylation of chemically unmodified proteins based on this polymer superquenching platform. Using a modified QTL Lightspeed assay, phosphorylation of native protein was quantified by the interaction of the phosphorylated proteins with metal-ion coordinating groups co-located with fluorescent polymer deposited onto microspheres. The binding of phospho-protein inhibits a dye-labeled "tracer" peptide from associating to the phosphate-binding sites present on the fluorescent microspheres. The resulting inhibition of quench generates a "turn on" assay, in which the signal correlates with the phosphorylation of the substrate. The assay was tested on three different proteins: Myelin Basic Protein (MBP), Histone H1 and Phosphorylated heat- and acid-stable protein (PHAS-1). Phosphorylation of the proteins was detected by Protein Kinase Calpha (PKCalpha) and by the Interleukin -1 Receptor-associated Kinase 4 (IRAK4). Enzyme inhibition yielded IC50 values that were comparable to those obtained using peptide substrates. Statistical parameters that

  2. Developing a novel fiber optic fluorescence device for multiplexed high-throughput cytotoxic screening.

    Science.gov (United States)

    Lee, Dennis; Barnes, Stephen

    2010-01-01

    The need for new pharmacological agents is unending. Yet the drug discovery process has changed substantially over the past decade and continues to evolve in response to new technologies. There is presently a high demand to reduce discovery time by improving specific lab disciplines and developing new technology platforms in the area of cell-based assay screening. Here we present the developmental concept and early stage testing of the Ab-Sniffer, a novel fiber optic fluorescence device for high-throughput cytotoxicity screening using an immobilized whole cell approach. The fused silica fibers are chemically functionalized with biotin to provide interaction with fluorescently labeled, streptavidin functionalized alginate-chitosan microspheres. The microspheres are also functionalized with Concanavalin A to facilitate binding to living cells. By using lymphoma cells and rituximab in an adaptation of a well-known cytotoxicity protocol we demonstrate the utility of the Ab-Sniffer for functional screening of potential drug compounds rather than indirect, non-functional screening via binding assay. The platform can be extended to any assay capable of being tied to a fluorescence response including multiple target cells in each well of a multi-well plate for high-throughput screening.

  3. Automated cleaning and pre-processing of immunoglobulin gene sequences from high-throughput sequencing

    Directory of Open Access Journals (Sweden)

    Miri eMichaeli

    2012-12-01

    Full Text Available High throughput sequencing (HTS yields tens of thousands to millions of sequences that require a large amount of pre-processing work to clean various artifacts. Such cleaning cannot be performed manually. Existing programs are not suitable for immunoglobulin (Ig genes, which are variable and often highly mutated. This paper describes Ig-HTS-Cleaner (Ig High Throughput Sequencing Cleaner, a program containing a simple cleaning procedure that successfully deals with pre-processing of Ig sequences derived from HTS, and Ig-Indel-Identifier (Ig Insertion – Deletion Identifier, a program for identifying legitimate and artifact insertions and/or deletions (indels. Our programs were designed for analyzing Ig gene sequences obtained by 454 sequencing, but they are applicable to all types of sequences and sequencing platforms. Ig-HTS-Cleaner and Ig-Indel-Identifier have been implemented in Java and saved as executable JAR files, supported on Linux and MS Windows. No special requirements are needed in order to run the programs, except for correctly constructing the input files as explained in the text. The programs' performance has been tested and validated on real and simulated data sets.

  4. High-Throughput Tools for Characterization of Antibody Epitopes

    DEFF Research Database (Denmark)

    Christiansen, Anders

    mapping. In Chapter 1, it was examined whether combining phage display, a traditional epitope mapping approach, with HTS would improve the method. The developed approach was successfully used to map Ara h 1 epitopes in sera from patients with peanut allergy. Notably, the sera represented difficult...... proliferation advantages. Finally, in Chapter 4, a different emerging technology, next-generation peptide microarrays, was applied for epitope mapping of major peanut allergens using sera from allergic patients. New developments in the peptide microarray have enabled a greatly increased throughput....... In this study, these improvements were utilized to characterize epitopes at high resolution, i.e. determine the importance of each residue for antibody binding, for all major peanut allergens. Epitope reactivity among patients often converged on known epitope hotspots, however the binding patterns were somewhat...

  5. Mechanical Conversion for High-Throughput TEM Sample Preparation

    International Nuclear Information System (INIS)

    Kendrick, Anthony B; Moore, Thomas M; Zaykova-Feldman, Lyudmila

    2006-01-01

    This paper presents a novel method of direct mechanical conversion from lift-out sample to TEM sample holder. The lift-out sample is prepared in the FIB using the in-situ liftout Total Release TM method. The mechanical conversion is conducted using a mechanical press and one of a variety of TEM coupons, including coupons for both top-side and back-side thinning. The press joins a probe tip point with attached TEM sample to the sample coupon and separates the complete assembly as a 3mm diameter TEM grid, compatible with commercially available TEM sample holder rods. This mechanical conversion process lends itself well to the high through-put requirements of in-line process control and to materials characterization labs where instrument utilization and sample security are critically important

  6. A hybrid MAC protocol design for energy-efficient very-high-throughput millimeter wave, wireless sensor communication networks

    Science.gov (United States)

    Jian, Wei; Estevez, Claudio; Chowdhury, Arshad; Jia, Zhensheng; Wang, Jianxin; Yu, Jianguo; Chang, Gee-Kung

    2010-12-01

    This paper presents an energy-efficient Medium Access Control (MAC) protocol for very-high-throughput millimeter-wave (mm-wave) wireless sensor communication networks (VHT-MSCNs) based on hybrid multiple access techniques of frequency division multiplexing access (FDMA) and time division multiplexing access (TDMA). An energy-efficient Superframe for wireless sensor communication network employing directional mm-wave wireless access technologies is proposed for systems that require very high throughput, such as high definition video signals, for sensing, processing, transmitting, and actuating functions. Energy consumption modeling for each network element and comparisons among various multi-access technologies in term of power and MAC layer operations are investigated for evaluating the energy-efficient improvement of proposed MAC protocol.

  7. A High Throughput, 384-Well, Semi-Automated, Hepatocyte Intrinsic Clearance Assay for Screening New Molecular Entities in Drug Discovery.

    Science.gov (United States)

    Heinle, Lance; Peterkin, Vincent; de Morais, Sonia M; Jenkins, Gary J; Badagnani, Ilaria

    2015-01-01

    A high throughput, semi-automated clearance screening assay in hepatocytes was developed allowing a scientist to generate data for 96 compounds in one week. The 384-well format assay utilizes a Thermo Multidrop Combi and an optimized LC-MS/MS method. The previously reported LCMS/ MS method reduced the analytical run time by 3-fold, down to 1.2 min injection-to-injection. The Multidrop was able to deliver hepatocytes to 384-well plates with minimal viability loss. Comparison of results from the new 384-well and historical 24-well assays yielded a correlation of 0.95. In addition, results obtained for 25 marketed drugs with various metabolism pathways had a correlation of 0.75 when compared with literature values. Precision was maintained in the new format as 8 compounds tested in ≥39 independent experiments had coefficients of variation ≤21%. The ability to predict in vivo clearances using the new stability assay format was also investigated using 22 marketed drugs and 26 AbbVie compounds. Correction of intrinsic clearance values with binding to hepatocytes (in vitro data) and plasma (in vivo data) resulted in a higher in vitro to in vivo correlation when comparing 22 marketed compounds in human (0.80 vs 0.35) and 26 AbbVie Discovery compounds in rat (0.56 vs 0.17), demonstrating the importance of correcting for binding in clearance studies. This newly developed high throughput, semi-automated clearance assay allows for rapid screening of Discovery compounds to enable Structure Activity Relationship (SAR) analysis based on high quality hepatocyte stability data in sufficient quantity and quality to drive the next round of compound synthesis.

  8. A ground-up approach to High Throughput Cloud Computing in High-Energy Physics

    CERN Document Server

    AUTHOR|(INSPIRE)INSPIRE-00245123; Ganis, Gerardo; Bagnasco, Stefano

    The thesis explores various practical approaches in making existing High Throughput computing applications common in High Energy Physics work on cloud-provided resources, as well as opening the possibility for running new applications. The work is divided into two parts: firstly we describe the work done at the computing facility hosted by INFN Torino to entirely convert former Grid resources into cloud ones, eventually running Grid use cases on top along with many others in a more flexible way. Integration and conversion problems are duly described. The second part covers the development of solutions for automatizing the orchestration of cloud workers based on the load of a batch queue and the development of HEP applications based on ROOT's PROOF that can adapt at runtime to a changing number of workers.

  9. Statistical removal of background signals from high-throughput 1H NMR line-broadening ligand-affinity screens

    International Nuclear Information System (INIS)

    Worley, Bradley; Sisco, Nicholas J.; Powers, Robert

    2015-01-01

    NMR ligand-affinity screens are vital to drug discovery, are routinely used to screen fragment-based libraries, and used to verify chemical leads from high-throughput assays and virtual screens. NMR ligand-affinity screens are also a highly informative first step towards identifying functional epitopes of unknown proteins, as well as elucidating the biochemical functions of protein–ligand interaction at their binding interfaces. While simple one-dimensional 1 H NMR experiments are capable of indicating binding through a change in ligand line shape, they are plagued by broad, ill-defined background signals from protein 1 H resonances. We present an uncomplicated method for subtraction of protein background in high-throughput ligand-based affinity screens, and show that its performance is maximized when phase-scatter correction is applied prior to subtraction

  10. Digital imaging of root traits (DIRT): a high-throughput computing and collaboration platform for field-based root phenomics.

    Science.gov (United States)

    Das, Abhiram; Schneider, Hannah; Burridge, James; Ascanio, Ana Karine Martinez; Wojciechowski, Tobias; Topp, Christopher N; Lynch, Jonathan P; Weitz, Joshua S; Bucksch, Alexander

    2015-01-01

    Plant root systems are key drivers of plant function and yield. They are also under-explored targets to meet global food and energy demands. Many new technologies have been developed to characterize crop root system architecture (CRSA). These technologies have the potential to accelerate the progress in understanding the genetic control and environmental response of CRSA. Putting this potential into practice requires new methods and algorithms to analyze CRSA in digital images. Most prior approaches have solely focused on the estimation of root traits from images, yet no integrated platform exists that allows easy and intuitive access to trait extraction and analysis methods from images combined with storage solutions linked to metadata. Automated high-throughput phenotyping methods are increasingly used in laboratory-based efforts to link plant genotype with phenotype, whereas similar field-based studies remain predominantly manual low-throughput. Here, we present an open-source phenomics platform "DIRT", as a means to integrate scalable supercomputing architectures into field experiments and analysis pipelines. DIRT is an online platform that enables researchers to store images of plant roots, measure dicot and monocot root traits under field conditions, and share data and results within collaborative teams and the broader community. The DIRT platform seamlessly connects end-users with large-scale compute "commons" enabling the estimation and analysis of root phenotypes from field experiments of unprecedented size. DIRT is an automated high-throughput computing and collaboration platform for field based crop root phenomics. The platform is accessible at http://www.dirt.iplantcollaborative.org/ and hosted on the iPlant cyber-infrastructure using high-throughput grid computing resources of the Texas Advanced Computing Center (TACC). DIRT is a high volume central depository and high-throughput RSA trait computation platform for plant scientists working on crop roots

  11. HTTK R Package v1.4 - JSS Article on HTTK: R Package for High-Throughput Toxicokinetics

    Data.gov (United States)

    U.S. Environmental Protection Agency — httk: High-Throughput Toxicokinetics Functions and data tables for simulation and statistical analysis of chemical toxicokinetics ("TK") using data obtained from...

  12. High Throughput Measurement of Locomotor Sensitization to Volatilized Cocaine in Drosophila melanogaster.

    Science.gov (United States)

    Filošević, Ana; Al-Samarai, Sabina; Andretić Waldowski, Rozi

    2018-01-01

    Drosophila melanogaster can be used to identify genes with novel functional roles in neuronal plasticity induced by repeated consumption of addictive drugs. Behavioral sensitization is a relatively simple behavioral output of plastic changes that occur in the brain after repeated exposures to drugs of abuse. The development of screening procedures for genes that control behavioral sensitization has stalled due to a lack of high-throughput behavioral tests that can be used in genetically tractable organism, such as Drosophila . We have developed a new behavioral test, FlyBong, which combines delivery of volatilized cocaine (vCOC) to individually housed flies with objective quantification of their locomotor activity. There are two main advantages of FlyBong: it is high-throughput and it allows for comparisons of locomotor activity of individual flies before and after single or multiple exposures. At the population level, exposure to vCOC leads to transient and concentration-dependent increase in locomotor activity, representing sensitivity to an acute dose. A second exposure leads to further increase in locomotion, representing locomotor sensitization. We validate FlyBong by showing that locomotor sensitization at either the population or individual level is absent in the mutants for circadian genes period (per) , Clock (Clk) , and cycle (cyc) . The locomotor sensitization that is present in timeless (tim) and pigment dispersing factor (pdf) mutant flies is in large part not cocaine specific, but derived from increased sensitivity to warm air. Circadian genes are not only integral part of the neural mechanism that is required for development of locomotor sensitization, but in addition, they modulate the intensity of locomotor sensitization as a function of the time of day. Motor-activating effects of cocaine are sexually dimorphic and require a functional dopaminergic transporter. FlyBong is a new and improved method for inducing and measuring locomotor sensitization

  13. High Throughput Measurement of Locomotor Sensitization to Volatilized Cocaine in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Ana Filošević

    2018-02-01

    Full Text Available Drosophila melanogaster can be used to identify genes with novel functional roles in neuronal plasticity induced by repeated consumption of addictive drugs. Behavioral sensitization is a relatively simple behavioral output of plastic changes that occur in the brain after repeated exposures to drugs of abuse. The development of screening procedures for genes that control behavioral sensitization has stalled due to a lack of high-throughput behavioral tests that can be used in genetically tractable organism, such as Drosophila. We have developed a new behavioral test, FlyBong, which combines delivery of volatilized cocaine (vCOC to individually housed flies with objective quantification of their locomotor activity. There are two main advantages of FlyBong: it is high-throughput and it allows for comparisons of locomotor activity of individual flies before and after single or multiple exposures. At the population level, exposure to vCOC leads to transient and concentration-dependent increase in locomotor activity, representing sensitivity to an acute dose. A second exposure leads to further increase in locomotion, representing locomotor sensitization. We validate FlyBong by showing that locomotor sensitization at either the population or individual level is absent in the mutants for circadian genes period (per, Clock (Clk, and cycle (cyc. The locomotor sensitization that is present in timeless (tim and pigment dispersing factor (pdf mutant flies is in large part not cocaine specific, but derived from increased sensitivity to warm air. Circadian genes are not only integral part of the neural mechanism that is required for development of locomotor sensitization, but in addition, they modulate the intensity of locomotor sensitization as a function of the time of day. Motor-activating effects of cocaine are sexually dimorphic and require a functional dopaminergic transporter. FlyBong is a new and improved method for inducing and measuring locomotor

  14. Lessons we learned from high-throughput and top-down systems biology analyses about glioma stem cells.

    Science.gov (United States)

    Mock, Andreas; Chiblak, Sara; Herold-Mende, Christel

    2014-01-01

    A growing body of evidence suggests that glioma stem cells (GSCs) account for tumor initiation, therapy resistance, and the subsequent regrowth of gliomas. Thus, continuous efforts have been undertaken to further characterize this subpopulation of less differentiated tumor cells. Although we are able to enrich GSCs, we still lack a comprehensive understanding of GSC phenotypes and behavior. The advent of high-throughput technologies raised hope that incorporation of these newly developed platforms would help to tackle such questions. Since then a couple of comparative genome-, transcriptome- and proteome-wide studies on GSCs have been conducted giving new insights in GSC biology. However, lessons had to be learned in designing high-throughput experiments and some of the resulting conclusions fell short of expectations because they were performed on only a few GSC lines or at one molecular level instead of an integrative poly-omics approach. Despite these shortcomings, our knowledge of GSC biology has markedly expanded due to a number of survival-associated biomarkers as well as glioma-relevant signaling pathways and therapeutic targets being identified. In this article we review recent findings obtained by comparative high-throughput analyses of GSCs. We further summarize fundamental concepts of systems biology as well as its applications for glioma stem cell research.

  15. A High Throughput Model of Post-Traumatic Osteoarthritis using Engineered Cartilage Tissue Analogs

    Science.gov (United States)

    Mohanraj, Bhavana; Meloni, Gregory R.; Mauck, Robert L.; Dodge, George R.

    2014-01-01

    (1) Objective A number of in vitro models of post-traumatic osteoarthritis (PTOA) have been developed to study the effect of mechanical overload on the processes that regulate cartilage degeneration. While such frameworks are critical for the identification therapeutic targets, existing technologies are limited in their throughput capacity. Here, we validate a test platform for high-throughput mechanical injury incorporating engineered cartilage. (2) Method We utilized a high throughput mechanical testing platform to apply injurious compression to engineered cartilage and determined their strain and strain rate dependent responses to injury. Next, we validated this response by applying the same injury conditions to cartilage explants. Finally, we conducted a pilot screen of putative PTOA therapeutic compounds. (3) Results Engineered cartilage response to injury was strain dependent, with a 2-fold increase in GAG loss at 75% compared to 50% strain. Extensive cell death was observed adjacent to fissures, with membrane rupture corroborated by marked increases in LDH release. Testing of established PTOA therapeutics showed that pan-caspase inhibitor (ZVF) was effective at reducing cell death, while the amphiphilic polymer (P188) and the free-radical scavenger (NAC) reduced GAG loss as compared to injury alone. (4) Conclusions The injury response in this engineered cartilage model replicated key features of the response from cartilage explants, validating this system for application of physiologically relevant injurious compression. This study establishes a novel tool for the discovery of mechanisms governing cartilage injury, as well as a screening platform for the identification of new molecules for the treatment of PTOA. PMID:24999113

  16. High-throughput testing of terpenoid biosynthesis candidate genes using transient expression in Nicotiana benthamiana

    DEFF Research Database (Denmark)

    Bach, Søren Spanner; Bassard, Jean-Étienne André; Andersen-Ranberg, Johan

    2014-01-01

    To respond to the rapidly growing number of genes putatively involved in terpenoid metabolism, a robust high-throughput platform for functional testing is needed. An in planta expression system offers several advantages such as the capacity to produce correctly folded and active enzymes localized...

  17. SeqAPASS to evaluate conservation of high-throughput screening targets across non-mammalian species

    Science.gov (United States)

    Cell-based high-throughput screening (HTS) and computational technologies are being applied as tools for toxicity testing in the 21st century. The U.S. Environmental Protection Agency (EPA) embraced these technologies and created the ToxCast Program in 2007, which has served as a...

  18. Tiered High-Throughput Screening Approach to Identify ...

    Science.gov (United States)

    High-throughput screening (HTS) for potential thyroid–disrupting chemicals requires a system of assays to capture multiple molecular-initiating events (MIEs) that converge on perturbed thyroid hormone (TH) homeostasis. Screening for MIEs specific to TH-disrupting pathways is limited in the US EPA ToxCast screening assay portfolio. To fill one critical screening gap, the Amplex UltraRed-thyroperoxidase (AUR-TPO) assay was developed to identify chemicals that inhibit TPO, as decreased TPO activity reduces TH synthesis. The ToxCast Phase I and II chemical libraries, comprised of 1,074 unique chemicals, were initially screened using a single, high concentration to identify potential TPO inhibitors. Chemicals positive in the single concentration screen were retested in concentration-response. Due to high false positive rates typically observed with loss-of-signal assays such as AUR-TPO, we also employed two additional assays in parallel to identify possible sources of nonspecific assay signal loss, enabling stratification of roughly 300 putative TPO inhibitors based upon selective AUR-TPO activity. A cell-free luciferase inhibition assay was used to identify nonspecific enzyme inhibition among the putative TPO inhibitors, and a cytotoxicity assay using a human cell line was used to estimate the cellular tolerance limit. Additionally, the TPO inhibition activities of 150 chemicals were compared between the AUR-TPO and an orthogonal peroxidase oxidation assay using

  19. Generating information-rich high-throughput experimental materials genomes using functional clustering via multitree genetic programming and information theory.

    Science.gov (United States)

    Suram, Santosh K; Haber, Joel A; Jin, Jian; Gregoire, John M

    2015-04-13

    High-throughput experimental methodologies are capable of synthesizing, screening and characterizing vast arrays of combinatorial material libraries at a very rapid rate. These methodologies strategically employ tiered screening wherein the number of compositions screened decreases as the complexity, and very often the scientific information obtained from a screening experiment, increases. The algorithm used for down-selection of samples from higher throughput screening experiment to a lower throughput screening experiment is vital in achieving information-rich experimental materials genomes. The fundamental science of material discovery lies in the establishment of composition-structure-property relationships, motivating the development of advanced down-selection algorithms which consider the information value of the selected compositions, as opposed to simply selecting the best performing compositions from a high throughput experiment. Identification of property fields (composition regions with distinct composition-property relationships) in high throughput data enables down-selection algorithms to employ advanced selection strategies, such as the selection of representative compositions from each field or selection of compositions that span the composition space of the highest performing field. Such strategies would greatly enhance the generation of data-driven discoveries. We introduce an informatics-based clustering of composition-property functional relationships using a combination of information theory and multitree genetic programming concepts for identification of property fields in a composition library. We demonstrate our approach using a complex synthetic composition-property map for a 5 at. % step ternary library consisting of four distinct property fields and finally explore the application of this methodology for capturing relationships between composition and catalytic activity for the oxygen evolution reaction for 5429 catalyst compositions in a

  20. WormScan: a technique for high-throughput phenotypic analysis of Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Mark D Mathew

    Full Text Available BACKGROUND: There are four main phenotypes that are assessed in whole organism studies of Caenorhabditis elegans; mortality, movement, fecundity and size. Procedures have been developed that focus on the digital analysis of some, but not all of these phenotypes and may be limited by expense and limited throughput. We have developed WormScan, an automated image acquisition system that allows quantitative analysis of each of these four phenotypes on standard NGM plates seeded with E. coli. This system is very easy to implement and has the capacity to be used in high-throughput analysis. METHODOLOGY/PRINCIPAL FINDINGS: Our system employs a readily available consumer grade flatbed scanner. The method uses light stimulus from the scanner rather than physical stimulus to induce movement. With two sequential scans it is possible to quantify the induced phototactic response. To demonstrate the utility of the method, we measured the phenotypic response of C. elegans to phosphine gas exposure. We found that stimulation of movement by the light of the scanner was equivalent to physical stimulation for the determination of mortality. WormScan also provided a quantitative assessment of health for the survivors. Habituation from light stimulation of continuous scans was similar to habituation caused by physical stimulus. CONCLUSIONS/SIGNIFICANCE: There are existing systems for the automated phenotypic data collection of C. elegans. The specific advantages of our method over existing systems are high-throughput assessment of a greater range of phenotypic endpoints including determination of mortality and quantification of the mobility of survivors. Our system is also inexpensive and very easy to implement. Even though we have focused on demonstrating the usefulness of WormScan in toxicology, it can be used in a wide range of additional C. elegans studies including lifespan determination, development, pathology and behavior. Moreover, we have even adapted the