Latest performance of ArF immersion scanner NSR-S630D for high-volume manufacturing for 7nm node

NASA Astrophysics Data System (ADS)

Funatsu, Takayuki; Uehara, Yusaku; Hikida, Yujiro; Hayakawa, Akira; Ishiyama, Satoshi; Hirayama, Toru; Kono, Hirotaka; Shirata, Yosuke; Shibazaki, Yuichi

2015-03-01

In order to achieve stable operation in cutting-edge semiconductor manufacturing, Nikon has developed NSR-S630D with extremely accurate overlay while maintaining throughput in various conditions resembling a real production environment. In addition, NSR-S630D has been equipped with enhanced capabilities to maintain long-term overlay stability and user interface improvement all due to our newly developed application software platform. In this paper, we describe the most recent S630D performance in various conditions similar to real productions. In a production environment, superior overlay accuracy with high dose conditions and high throughput are often required; therefore, we have performed several experiments with high dose conditions to demonstrate NSR's thermal aberration capabilities in order to achieve world class overlay performance. Furthermore, we will introduce our new software that enables long term overlay performance.

The Stanford Automated Mounter: Enabling High-Throughput Protein Crystal Screening at SSRL

DOE Office of Scientific and Technical Information (OSTI.GOV)

Smith, C.A.; Cohen, A.E.

2009-05-26

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 screenedmore » 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.« less

Neutron reflecting supermirror structure

DOEpatents

Wood, J.L.

1992-12-01

An improved neutron reflecting supermirror structure comprising a plurality of stacked sets of bilayers of neutron reflecting materials. The improved neutron reflecting supermirror structure is adapted to provide extremely good performance at high incidence angles, i.e. up to four time the critical angle of standard neutron mirror structures. The reflection of neutrons striking the supermirror structure at a high critical angle provides enhanced neutron throughput, and hence more efficient and economical use of neutron sources. 2 figs.

Neutron reflecting supermirror structure

DOEpatents

Wood, James L.

1992-01-01

An improved neutron reflecting supermirror structure comprising a plurality of stacked sets of bilayers of neutron reflecting materials. The improved neutron reflecting supermirror structure is adapted to provide extremely good performance at high incidence angles, i.e. up to four time the critical angle of standard neutron mirror structures. The reflection of neutrons striking the supermirror structure at a high critical angle provides enhanced neutron throughput, and hence more efficient and economical use of neutron sources.

High-throughput on-chip in vivo neural regeneration studies using femtosecond laser nano-surgery and microfluidics

NASA Astrophysics Data System (ADS)

Rohde, Christopher B.; Zeng, Fei; Gilleland, Cody; Samara, Chrysanthi; Yanik, Mehmet F.

2009-02-01

In recent years, the advantages of using small invertebrate animals as model systems for human disease have become increasingly apparent and have resulted in three Nobel Prizes in medicine or chemistry during the last six years for studies conducted on the nematode Caenorhabditis elegans (C. elegans). The availability of a wide array of species-specific genetic techniques, along with the transparency of the worm and its ability to grow in minute volumes make C. elegans an extremely powerful model organism. We present a suite of technologies for complex high-throughput whole-animal genetic and drug screens. We demonstrate a high-speed microfluidic sorter that can isolate and immobilize C. elegans in a well-defined geometry, an integrated chip containing individually addressable screening chambers for incubation and exposure of individual animals to biochemical compounds, and a device for delivery of compound libraries in standard multiwell plates to microfluidic devices. The immobilization stability obtained by these devices is comparable to that of chemical anesthesia and the immobilization process does not affect lifespan, progeny production, or other aspects of animal health. The high-stability enables the use of a variety of key optical techniques. We use this to demonstrate femtosecond-laser nanosurgery and three-dimensional multiphoton microscopy. Used alone or in various combinations these devices facilitate a variety of high-throughput assays using whole animals, including mutagenesis and RNAi and drug screens at subcellular resolution, as well as high-throughput high-precision manipulations such as femtosecond-laser nanosurgery for large-scale in vivo neural degeneration and regeneration studies.

Large-scale protein-protein interactions detection by integrating big biosensing data with computational model.

PubMed

You, Zhu-Hong; Li, Shuai; Gao, Xin; Luo, Xin; Ji, Zhen

2014-01-01

Protein-protein interactions are the basis of biological functions, and studying these interactions on a molecular level is of crucial importance for understanding the functionality of a living cell. During the past decade, biosensors have emerged as an important tool for the high-throughput identification of proteins and their interactions. However, the high-throughput experimental methods for identifying PPIs are both time-consuming and expensive. On the other hand, high-throughput PPI data are often associated with high false-positive and high false-negative rates. Targeting at these problems, we propose a method for PPI detection by integrating biosensor-based PPI data with a novel computational model. This method was developed based on the algorithm of extreme learning machine combined with a novel representation of protein sequence descriptor. When performed on the large-scale human protein interaction dataset, the proposed method achieved 84.8% prediction accuracy with 84.08% sensitivity at the specificity of 85.53%. We conducted more extensive experiments to compare the proposed method with the state-of-the-art techniques, support vector machine. The achieved results demonstrate that our approach is very promising for detecting new PPIs, and it can be a helpful supplement for biosensor-based PPI data detection.

Revealing complex function, process and pathway interactions with high-throughput expression and biological annotation data.

PubMed

Singh, Nitesh Kumar; Ernst, Mathias; Liebscher, Volkmar; Fuellen, Georg; Taher, Leila

2016-10-20

The biological relationships both between and within the functions, processes and pathways that operate within complex biological systems are only poorly characterized, making the interpretation of large scale gene expression datasets extremely challenging. Here, we present an approach that integrates gene expression and biological annotation data to identify and describe the interactions between biological functions, processes and pathways that govern a phenotype of interest. The product is a global, interconnected network, not of genes but of functions, processes and pathways, that represents the biological relationships within the system. We validated our approach on two high-throughput expression datasets describing organismal and organ development. Our findings are well supported by the available literature, confirming that developmental processes and apoptosis play key roles in cell differentiation. Furthermore, our results suggest that processes related to pluripotency and lineage commitment, which are known to be critical for development, interact mainly indirectly, through genes implicated in more general biological processes. Moreover, we provide evidence that supports the relevance of cell spatial organization in the developing liver for proper liver function. Our strategy can be viewed as an abstraction that is useful to interpret high-throughput data and devise further experiments.

Quantitative secondary electron imaging for work function extraction at atomic level and layer identification of graphene

PubMed Central

Zhou, Yangbo; Fox, Daniel S; Maguire, Pierce; O’Connell, Robert; Masters, Robert; Rodenburg, Cornelia; Wu, Hanchun; Dapor, Maurizio; Chen, Ying; Zhang, Hongzhou

2016-01-01

Two-dimensional (2D) materials usually have a layer-dependent work function, which require fast and accurate detection for the evaluation of their device performance. A detection technique with high throughput and high spatial resolution has not yet been explored. Using a scanning electron microscope, we have developed and implemented a quantitative analytical technique which allows effective extraction of the work function of graphene. This technique uses the secondary electron contrast and has nanometre-resolved layer information. The measurement of few-layer graphene flakes shows the variation of work function between graphene layers with a precision of less than 10 meV. It is expected that this technique will prove extremely useful for researchers in a broad range of fields due to its revolutionary throughput and accuracy. PMID:26878907

Microbial community analysis of the hypersaline water of the Dead Sea using high-throughput amplicon sequencing.

PubMed

Jacob, Jacob H; Hussein, Emad I; Shakhatreh, Muhamad Ali K; Cornelison, Christopher T

2017-10-01

Amplicon sequencing using next-generation technology (bTEFAP ® ) has been utilized in describing the diversity of Dead Sea microbiota. The investigated area is a well-known salt lake in the western part of Jordan found in the lowest geographical location in the world (more than 420 m below sea level) and characterized by extreme salinity (approximately, 34%) in addition to other extreme conditions (low pH, unique ionic composition different from sea water). DNA was extracted from Dead Sea water. A total of 314,310 small subunit RNA (SSU rRNA) sequences were parsed, and 288,452 sequences were then clustered. For alpha diversity analysis, sample was rarefied to 3,000 sequences. The Shannon-Wiener index curve plot reached a plateau at approximately 3,000 sequences indicating that sequencing depth was sufficient to capture the full scope of microbial diversity. Archaea was found to be dominating the sequences (52%), whereas Bacteria constitute 45% of the sequences. Altogether, prokaryotic sequences (which constitute 97% of all sequences) were found to predominate. The findings expand on previous studies by using high-throughput amplicon sequencing to describe the microbial community in an environment which in recent years has been shown to hide some interesting diversity. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

Robust ridge regression estimators for nonlinear models with applications to high throughput screening assay data.

PubMed

Lim, Changwon

2015-03-30

Nonlinear regression is often used to evaluate the toxicity of a chemical or a drug by fitting data from a dose-response study. Toxicologists and pharmacologists may draw a conclusion about whether a chemical is toxic by testing the significance of the estimated parameters. However, sometimes the null hypothesis cannot be rejected even though the fit is quite good. One possible reason for such cases is that the estimated standard errors of the parameter estimates are extremely large. In this paper, we propose robust ridge regression estimation procedures for nonlinear models to solve this problem. The asymptotic properties of the proposed estimators are investigated; in particular, their mean squared errors are derived. The performances of the proposed estimators are compared with several standard estimators using simulation studies. The proposed methodology is also illustrated using high throughput screening assay data obtained from the National Toxicology Program. Copyright © 2014 John Wiley & Sons, Ltd.

High-Throughput Fabrication of Ultradense Annular Nanogap Arrays for Plasmon-Enhanced Spectroscopy.

PubMed

Cai, Hongbing; Meng, Qiushi; Zhao, Hui; Li, Mingling; Dai, Yanmeng; Lin, Yue; Ding, Huaiyi; Pan, Nan; Tian, Yangchao; Luo, Yi; Wang, Xiaoping

2018-06-13

The confinement of light into nanometer-sized metallic nanogaps can lead to an extremely high field enhancement, resulting in dramatically enhanced absorption, emission, and surface-enhanced Raman scattering (SERS) of molecules embedded in nanogaps. However, low-cost, high-throughput, and reliable fabrication of ultra-high-dense nanogap arrays with precise control of the gap size still remains a challenge. Here, by combining colloidal lithography and atomic layer deposition technique, a reproducible method for fabricating ultra-high-dense arrays of hexagonal close-packed annular nanogaps over large areas is demonstrated. The annular nanogap arrays with a minimum diameter smaller than 100 nm and sub-1 nm gap width have been produced, showing excellent SERS performance with a typical enhancement factor up to 3.1 × 10 6 and a detection limit of 10 -11 M. Moreover, it can also work as a high-quality field enhancement substrate for studying two-dimensional materials, such as MoSe 2 . Our method provides an attractive approach to produce controllable nanogaps for enhanced light-matter interaction at the nanoscale.

Improving Hierarchical Models Using Historical Data with Applications in High-Throughput Genomics Data Analysis.

PubMed

Li, Ben; Li, Yunxiao; Qin, Zhaohui S

2017-06-01

Modern high-throughput biotechnologies such as microarray and next generation sequencing produce a massive amount of information for each sample assayed. However, in a typical high-throughput experiment, only limited amount of data are observed for each individual feature, thus the classical 'large p , small n ' problem. Bayesian hierarchical model, capable of borrowing strength across features within the same dataset, has been recognized as an effective tool in analyzing such data. However, the shrinkage effect, the most prominent feature of hierarchical features, can lead to undesirable over-correction for some features. In this work, we discuss possible causes of the over-correction problem and propose several alternative solutions. Our strategy is rooted in the fact that in the Big Data era, large amount of historical data are available which should be taken advantage of. Our strategy presents a new framework to enhance the Bayesian hierarchical model. Through simulation and real data analysis, we demonstrated superior performance of the proposed strategy. Our new strategy also enables borrowing information across different platforms which could be extremely useful with emergence of new technologies and accumulation of data from different platforms in the Big Data era. Our method has been implemented in R package "adaptiveHM", which is freely available from https://github.com/benliemory/adaptiveHM.

Improving Hierarchical Models Using Historical Data with Applications in High-Throughput Genomics Data Analysis

PubMed Central

Li, Ben; Li, Yunxiao; Qin, Zhaohui S.

2016-01-01

Modern high-throughput biotechnologies such as microarray and next generation sequencing produce a massive amount of information for each sample assayed. However, in a typical high-throughput experiment, only limited amount of data are observed for each individual feature, thus the classical ‘large p, small n’ problem. Bayesian hierarchical model, capable of borrowing strength across features within the same dataset, has been recognized as an effective tool in analyzing such data. However, the shrinkage effect, the most prominent feature of hierarchical features, can lead to undesirable over-correction for some features. In this work, we discuss possible causes of the over-correction problem and propose several alternative solutions. Our strategy is rooted in the fact that in the Big Data era, large amount of historical data are available which should be taken advantage of. Our strategy presents a new framework to enhance the Bayesian hierarchical model. Through simulation and real data analysis, we demonstrated superior performance of the proposed strategy. Our new strategy also enables borrowing information across different platforms which could be extremely useful with emergence of new technologies and accumulation of data from different platforms in the Big Data era. Our method has been implemented in R package “adaptiveHM”, which is freely available from https://github.com/benliemory/adaptiveHM. PMID:28919931

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

PubMed

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.

A compact disk-like centrifugal microfluidic system for high-throughput nanoliter-scale protein crystallization screening.

PubMed

Li, Gang; Chen, Qiang; Li, Junjun; Hu, Xiaojian; Zhao, Jianlong

2010-06-01

A centrifuge-based microfluidic system has been developed that enables automated high-throughput and low-volume protein crystallizations. In this system, protein solution was automatically and accurately metered and dispensed into nanoliter-sized multiple reaction chambers, and it was mixed with various types of precipitants using a combination of capillary effect and centrifugal force. It has the advantages of simple fabrication, easy operation, and extremely low waste. To demonstrate the feasibility of this system, we constructed a chip containing 24 units and used it to perform lysozyme and cyan fluorescent protein (CyPet) crystallization trials. The results demonstrate that high-quality crystals can be grown and harvested from such a nanoliter-volume microfluidic system. Compared to other microfluidic technologies for protein crystallization, this microfluidic system allows zero waste, simple structure and convenient operation, which suggests that our microfluidic disk can be applied not only to protein crystallization, but also to the miniaturization of various biochemical reactions requiring precise nanoscale control.

Development of a High-Throughput Microwave Imaging System for Concealed Weapons Detection

DTIC Science & Technology

2016-07-15

hardware. Index Terms—Microwave imaging, multistatic radar, Fast Fourier Transform (FFT). I. INTRODUCTION Near-field microwave imaging is a non-ionizing...configuration, but its computational demands are extreme. Fast Fourier Transform (FFT) imaging has long been used to efficiently construct images sampled with...Simulated image of 25 point scatterers imaged at range 1.5m, with array layout depicted in Fig. 3. Left: image formed with Equation (5) ( Fourier

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wei, D. T.

Ion beam interference coating (IBIC) is a sputter-deposition process for multiple layers of optical thin films employing a Kaufman gun. It has achieved coatings of extremely low optical loss and high mechanical strength. It has many potential applications for a wide spectral range. This coating process is described in terms of principle, fabrication procedure, and optical measurements. Some discussions follow the history and outlooks of IBIC with emphasis on how to achieve low loss and on the throughput improvements.

Fabrication of wafer-scale nanopatterned sapphire substrate through phase separation lithography

NASA Astrophysics Data System (ADS)

Guo, Xu; Ni, Mengyang; Zhuang, Zhe; Dai, Jiangping; Wu, Feixiang; Cui, Yushuang; Yuan, Changsheng; Ge, Haixiong; Chen, Yanfeng

2016-04-01

A phase separation lithography (PSL) based on polymer blend provides an extremely simple, low-cost, and high-throughput way to fabricate wafer-scale disordered nanopatterns. This method was introduced to fabricate nanopatterned sapphire substrates (NPSSs) for GaN-based light-emitting diodes (LEDs). The PSL process only involved in spin-coating of polystyrene (PS)/polyethylene glycol (PEG) polymer blend on sapphire substrate and followed by a development with deionized water to remove PEG moiety. The PS nanoporous network was facilely obtained, and the structural parameters could be effectively tuned by controlling the PS/PEG weight ratio of the spin-coating solution. 2-in. wafer-scale NPSSs were conveniently achieved through the PS nanoporous network in combination with traditional nanofabrication methods, such as O2 reactive ion etching (RIE), e-beam evaporation deposition, liftoff, and chlorine-based RIE. In order to investigate the performance of such NPSSs, typical blue LEDs with emission wavelengths of ~450 nm were grown on the NPSS and a flat sapphire substrate (FSS) by metal-organic chemical vapor deposition, respectively. The integral photoluminescence (PL) intensity of the NPSS LED was enhanced by 32.3 % compared to that of the FSS-LED. The low relative standard deviation of 4.7 % for PL mappings of NPSS LED indicated the high uniformity of PL data across the whole 2-in. wafer. Extremely simple, low cost, and high throughput of the process and the ability to fabricate at the wafer scale make PSL a potential method for production of nanopatterned sapphire substrates.

Power laws and extreme values in antibody repertoires

NASA Astrophysics Data System (ADS)

Boyer, Sebastien; Biswas, Dipanwita; Scaramozzino, Natale; Kumar, Ananda Soshee; Nizak, Clément; Rivoire, Olivier

2015-03-01

Evolution by natural selection involves the succession of three steps: mutations, selection and proliferation. We are interested in describing and characterizing the result of selection over a population of many variants. After selection, this population will be dominated by the few best variants, with highest propensity to be selected, or highest ``selectivity.'' We ask the following question: how is the selectivity of the best variants distributed in the population? Extreme value theory, which characterizes the extreme tail of probability distributions in terms of a few universality class, has been proposed to describe it. To test this proposition and identify the relevant universality class, we performed quantitative in vitro experimental selections of libraries of >105 antibodies using the technique of phage display. Data obtained by high-throughput sequencing allows us to fit the selectivity distribution over more than two decades. In most experiments, the results show a striking power law for the selectivity distribution of the top antibodies, consistent with extreme value theory.

An Investigation of DC-DC Converter Power Density Using Si and SiC MOSFETS

DTIC Science & Technology

2010-05-07

submarine or small surface combatant, volumetric constraints quickly become extremely prohibitive. Dedicating generators for high power loads takes...thermal compounds were applied to the MOSFET-heat sink interface. For the Si APT26F120B2, MG Chemicals TC-450ML thermal epoxy was used to connect the... submarines , bus converter modules must be made optimally power dense in order to decrease volumetric requirements of the modules for a rated throughput

High-throughput screening of nanoparticle catalysts made by flame spray pyrolysis as hydrocarbon/NO oxidation catalysts.

PubMed

Weidenhof, B; Reiser, M; Stöwe, K; Maier, W F; Kim, M; Azurdia, J; Gulari, E; Seker, E; Barks, A; Laine, R M

2009-07-08

We describe here the use of liquid-feed flame spray pyrolysis (LF-FSP) to produce high surface area, nonporous, mixed-metal oxide nanopowders that were subsequently subjected to high-throughput screening to assess a set of materials for deNO(x) catalysis and hydrocarbon combustion. We were able to easily screen some 40 LF-FSP produced materials. LF-FSP produces nanopowders that very often consist of kinetic rather than thermodynamic phases. Such materials are difficult to access or are completely inaccessible via traditional catalyst preparation methods. Indeed, our studies identified a set of Ce(1-x)Zr(x)O(2) and Al(2)O(3)-Ce(1-x)Zr(x)O(2) nanopowders that offer surprisingly good activities for both NO(x) reduction and propane/propene oxidation both in high-throughput screening and in continuous flow catalytic studies. All of these catalysts offer activities comparable to traditional Pt/Al(2)O(3) catalysts but without Pt. Thus, although Pt-free, they are quite active for several extremely important emission control reactions, especially considering that these are only first generation materials. Indeed, efforts to dope the active catalysts with Pt actually led to lower catalytic activities. Thus the potential exists to completely change the materials used in emission control devices, especially for high-temperature reactions as these materials have already been exposed to 1500 degrees C; however, much research must be done before this potential is verified.

Building blocks for the development of an interface for high-throughput thin layer chromatography/ambient mass spectrometric analysis: a green methodology.

PubMed

Cheng, Sy-Chyi; Huang, Min-Zong; Wu, Li-Chieh; Chou, Chih-Chiang; Cheng, Chu-Nian; Jhang, Siou-Sian; Shiea, Jentaie

2012-07-17

Interfacing thin layer chromatography (TLC) with ambient mass spectrometry (AMS) has been an important area of analytical chemistry because of its capability to rapidly separate and characterize the chemical compounds. In this study, we have developed a high-throughput TLC-AMS system using building blocks to deal, deliver, and collect the TLC plate through an electrospray-assisted laser desorption ionization (ELDI) source. This is the first demonstration of the use of building blocks to construct and test the TLC-MS interfacing system. With the advantages of being readily available, cheap, reusable, and extremely easy to modify without consuming any material or reagent, the use of building blocks to develop the TLC-AMS interface is undoubtedly a green methodology. The TLC plate delivery system consists of a storage box, plate dealing component, conveyer, light sensor, and plate collecting box. During a TLC-AMS analysis, the TLC plate was sent to the conveyer from a stack of TLC plates placed in the storage box. As the TLC plate passed through the ELDI source, the chemical compounds separated on the plate would be desorbed by laser desorption and subsequently postionized by electrospray ionization. The samples, including a mixture of synthetic dyes and extracts of pharmaceutical drugs, were analyzed to demonstrate the capability of this TLC-ELDI/MS system for high-throughput analysis.

Bioinformatics Education—Perspectives and Challenges out of Africa

PubMed Central

Adebiyi, Ezekiel F.; Alzohairy, Ahmed M.; Everett, Dean; Ghedira, Kais; Ghouila, Amel; Kumuthini, Judit; Mulder, Nicola J.; Panji, Sumir; Patterton, Hugh-G.

2015-01-01

The discipline of bioinformatics has developed rapidly since the complete sequencing of the first genomes in the 1990s. The development of many high-throughput techniques during the last decades has ensured that bioinformatics has grown into a discipline that overlaps with, and is required for, the modern practice of virtually every field in the life sciences. This has placed a scientific premium on the availability of skilled bioinformaticians, a qualification that is extremely scarce on the African continent. The reasons for this are numerous, although the absence of a skilled bioinformatician at academic institutions to initiate a training process and build sustained capacity seems to be a common African shortcoming. This dearth of bioinformatics expertise has had a knock-on effect on the establishment of many modern high-throughput projects at African institutes, including the comprehensive and systematic analysis of genomes from African populations, which are among the most genetically diverse anywhere on the planet. Recent funding initiatives from the National Institutes of Health and the Wellcome Trust are aimed at ameliorating this shortcoming. In this paper, we discuss the problems that have limited the establishment of the bioinformatics field in Africa, as well as propose specific actions that will help with the education and training of bioinformaticians on the continent. This is an absolute requirement in anticipation of a boom in high-throughput approaches to human health issues unique to data from African populations. PMID:24990350

QSRA: a quality-value guided de novo short read assembler.

PubMed

Bryant, Douglas W; Wong, Weng-Keen; Mockler, Todd C

2009-02-24

New rapid high-throughput sequencing technologies have sparked the creation of a new class of assembler. Since all high-throughput sequencing platforms incorporate errors in their output, short-read assemblers must be designed to account for this error while utilizing all available data. We have designed and implemented an assembler, Quality-value guided Short Read Assembler, created to take advantage of quality-value scores as a further method of dealing with error. Compared to previous published algorithms, our assembler shows significant improvements not only in speed but also in output quality. QSRA generally produced the highest genomic coverage, while being faster than VCAKE. QSRA is extremely competitive in its longest contig and N50/N80 contig lengths, producing results of similar quality to those of EDENA and VELVET. QSRA provides a step closer to the goal of de novo assembly of complex genomes, improving upon the original VCAKE algorithm by not only drastically reducing runtimes but also increasing the viability of the assembly algorithm through further error handling capabilities.

Enabling inspection solutions for future mask technologies through the development of massively parallel E-Beam inspection

NASA Astrophysics Data System (ADS)

Malloy, Matt; Thiel, Brad; Bunday, Benjamin D.; Wurm, Stefan; Jindal, Vibhu; Mukhtar, Maseeh; Quoi, Kathy; Kemen, Thomas; Zeidler, Dirk; Eberle, Anna Lena; Garbowski, Tomasz; Dellemann, Gregor; Peters, Jan Hendrik

2015-09-01

The new device architectures and materials being introduced for sub-10nm manufacturing, combined with the complexity of multiple patterning and the need for improved hotspot detection strategies, have pushed current wafer inspection technologies to their limits. In parallel, gaps in mask inspection capability are growing as new generations of mask technologies are developed to support these sub-10nm wafer manufacturing requirements. In particular, the challenges associated with nanoimprint and extreme ultraviolet (EUV) mask inspection require new strategies that enable fast inspection at high sensitivity. The tradeoffs between sensitivity and throughput for optical and e-beam inspection are well understood. Optical inspection offers the highest throughput and is the current workhorse of the industry for both wafer and mask inspection. E-beam inspection offers the highest sensitivity but has historically lacked the throughput required for widespread adoption in the manufacturing environment. It is unlikely that continued incremental improvements to either technology will meet tomorrow's requirements, and therefore a new inspection technology approach is required; one that combines the high-throughput performance of optical with the high-sensitivity capabilities of e-beam inspection. To support the industry in meeting these challenges SUNY Poly SEMATECH has evaluated disruptive technologies that can meet the requirements for high volume manufacturing (HVM), for both the wafer fab [1] and the mask shop. Highspeed massively parallel e-beam defect inspection has been identified as the leading candidate for addressing the key gaps limiting today's patterned defect inspection techniques. As of late 2014 SUNY Poly SEMATECH completed a review, system analysis, and proof of concept evaluation of multiple e-beam technologies for defect inspection. A champion approach has been identified based on a multibeam technology from Carl Zeiss. This paper includes a discussion on the need for high-speed e-beam inspection and then provides initial imaging results from EUV masks and wafers from 61 and 91 beam demonstration systems. Progress towards high resolution and consistent intentional defect arrays (IDA) is also shown.

GROMACS 4.5: a high-throughput and highly parallel open source molecular simulation toolkit

PubMed Central

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

2013-01-01

Motivation: 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. Results: 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. Availability: GROMACS is an open source and free software available from http://www.gromacs.org. Contact: erik.lindahl@scilifelab.se Supplementary information: Supplementary data are available at Bioinformatics online. PMID:23407358

Transparent Nanopore Cavity Arrays Enable Highly Parallelized Optical Studies of Single Membrane Proteins on Chip.

PubMed

Diederichs, Tim; Nguyen, Quoc Hung; Urban, Michael; Tampé, Robert; Tornow, Marc

2018-06-13

Membrane proteins involved in transport processes are key targets for pharmaceutical research and industry. Despite continuous improvements and new developments in the field of electrical readouts for the analysis of transport kinetics, a well-suited methodology for high-throughput characterization of single transporters with nonionic substrates and slow turnover rates is still lacking. Here, we report on a novel architecture of silicon chips with embedded nanopore microcavities, based on a silicon-on-insulator technology for high-throughput optical readouts. Arrays containing more than 14 000 inverted-pyramidal cavities of 50 femtoliter volumes and 80 nm circular pore openings were constructed via high-resolution electron-beam lithography in combination with reactive ion etching and anisotropic wet etching. These cavities feature both, an optically transparent bottom and top cap. Atomic force microscopy analysis reveals an overall extremely smooth chip surface, particularly in the vicinity of the nanopores, which exhibits well-defined edges. Our unprecedented transparent chip design provides parallel and independent fluorescent readout of both cavities and buffer reservoir for unbiased single-transporter recordings. Spreading of large unilamellar vesicles with efficiencies up to 96% created nanopore-supported lipid bilayers, which are stable for more than 1 day. A high lipid mobility in the supported membrane was determined by fluorescent recovery after photobleaching. Flux kinetics of α-hemolysin were characterized at single-pore resolution with a rate constant of 0.96 ± 0.06 × 10 -3 s -1 . Here, we deliver an ideal chip platform for pharmaceutical research, which features high parallelism and throughput, synergistically combined with single-transporter resolution.

The ecology and diversity of microbial eukaryotes in geothermal springs.

PubMed

Oliverio, Angela M; Power, Jean F; Washburne, Alex; Cary, S Craig; Stott, Matthew B; Fierer, Noah

2018-04-16

Decades of research into the Bacteria and Archaea living in geothermal spring ecosystems have yielded great insight into the diversity of life and organismal adaptations to extreme environmental conditions. Surprisingly, while microbial eukaryotes (protists) are also ubiquitous in many environments, their diversity across geothermal springs has mostly been ignored. We used high-throughput sequencing to illuminate the diversity and structure of microbial eukaryotic communities found in 160 geothermal springs with broad ranges in temperature and pH across the Taupō Volcanic Zone in New Zealand. Protistan communities were moderately predictable in composition and varied most strongly across gradients in pH and temperature. Moreover, this variation mirrored patterns observed for bacterial and archaeal communities across the same spring samples, highlighting that there are similar ecological constraints across the tree of life. While extreme pH values were associated with declining protist diversity, high temperature springs harbored substantial amounts of protist diversity. Although protists are often overlooked in geothermal springs and other extreme environments, our results indicate that such environments can host distinct and diverse protistan communities.

In silico modelling of directed evolution: Implications for experimental design and stepwise evolution.

PubMed

Wedge, David C; Rowe, William; Kell, Douglas B; Knowles, Joshua

2009-03-07

We model the process of directed evolution (DE) in silico using genetic algorithms. Making use of the NK fitness landscape model, we analyse the effects of mutation rate, crossover and selection pressure on the performance of DE. A range of values of K, the epistatic interaction of the landscape, are considered, and high- and low-throughput modes of evolution are compared. Our findings suggest that for runs of or around ten generations' duration-as is typical in DE-there is little difference between the way in which DE needs to be configured in the high- and low-throughput regimes, nor across different degrees of landscape epistasis. In all cases, a high selection pressure (but not an extreme one) combined with a moderately high mutation rate works best, while crossover provides some benefit but only on the less rugged landscapes. These genetic algorithms were also compared with a "model-based approach" from the literature, which uses sequential fixing of the problem parameters based on fitting a linear model. Overall, we find that purely evolutionary techniques fare better than do model-based approaches across all but the smoothest landscapes.

The Extreme Ultraviolet Explorer - Optics fabrication and performance

NASA Technical Reports Server (NTRS)

Green, J.; Finley, D.; Bowyer, S.; Malina, R. F.

1986-01-01

The fabrication methods, testing and evaluation techniques, and performance results are presented for the mirrors for the Extreme Ultraviolet Explorer (EUVE). The finest mirror produced to date has a measured half energy width of 8 arcsec at optical wavelengths. With a polished nickel surface, the telescope throughput was 35 percent at 44 A and 60 percent at 256 A. The surface roughness is 20 A rms.

PGAS in-memory data processing for the Processing Unit of the Upgraded Electronics of the Tile Calorimeter of the ATLAS Detector

NASA Astrophysics Data System (ADS)

Ohene-Kwofie, Daniel; Otoo, Ekow

2015-10-01

The ATLAS detector, operated at the Large Hadron Collider (LHC) records proton-proton collisions at CERN every 50ns resulting in a sustained data flow up to PB/s. The upgraded Tile Calorimeter of the ATLAS experiment will sustain about 5PB/s of digital throughput. These massive data rates require extremely fast data capture and processing. Although there has been a steady increase in the processing speed of CPU/GPGPU assembled for high performance computing, the rate of data input and output, even under parallel I/O, has not kept up with the general increase in computing speeds. The problem then is whether one can implement an I/O subsystem infrastructure capable of meeting the computational speeds of the advanced computing systems at the petascale and exascale level. We propose a system architecture that leverages the Partitioned Global Address Space (PGAS) model of computing to maintain an in-memory data-store for the Processing Unit (PU) of the upgraded electronics of the Tile Calorimeter which is proposed to be used as a high throughput general purpose co-processor to the sROD of the upgraded Tile Calorimeter. The physical memory of the PUs are aggregated into a large global logical address space using RDMA- capable interconnects such as PCI- Express to enhance data processing throughput.

Towards roll-to-roll manufacturing of polymer photonic devices

NASA Astrophysics Data System (ADS)

Subbaraman, Harish; Lin, Xiaohui; Ling, Tao; Guo, L. Jay; Chen, Ray T.

2014-03-01

Traditionally, polymer photonic devices are fabricated using clean-room processes such as photolithography, e-beam lithography, reactive ion etching (RIE) and lift-off methods etc, which leads to long fabrication time, low throughput and high cost. We have utilized a novel process for fabricating polymer photonic devices using a combination of imprinting and ink jet printing methods, which provides high throughput on a variety of rigid and flexible substrates with low cost. We discuss the manufacturing challenges that need to be overcome in order to realize true implementation of roll-to-roll manufacturing of flexible polymer photonic systems. Several metrology and instrumentation challenges involved such as availability of particulate-free high quality substrate, development and implementation of high-speed in-line and off-line inspection and diagnostic tools with adaptive control for patterned and unpatterned material films, development of reliable hardware, etc need to be addressed and overcome in order to realize a successful manufacturing process. Due to extreme resolution requirements compared to print media, the burden of software and hardware tools on the throughput also needs to be carefully determined. Moreover, the effect of web wander and variations in web speed need to accurately be determined in the design of the system hardware and software. In this paper, we show the realization of solutions for few challenges, and utilizing these solutions for developing a high-rate R2R dual stage ink-jet printer that can provide alignment accuracy of <10μm at a web speed of 5m/min. The development of a roll-to-roll manufacturing system for polymer photonic systems opens limitless possibilities for the deployment of high performance components in a variety of applications including communication, sensing, medicine, agriculture, energy, lighting etc.

Uprated fine guidance sensor study

NASA Technical Reports Server (NTRS)

1984-01-01

Future orbital observatories will require star trackers of extremely high precision. These sensors must maintain high pointing accuracy and pointing stability simultaneously with a low light level signal from a guide star. To establish the fine guidance sensing requirements and to evaluate candidate fine guidance sensing concepts, the Space Telescope Optical Telescope Assembly was used as the reference optical system. The requirements review was separated into three areas: Optical Telescope Assembly (OTA), Fine Guidance Sensing and astrometry. The results show that the detectors should be installed directly onto the focal surface presented by the optics. This would maximize throughput and minimize point stability error by not incoporating any additional optical elements.

Neutron reflecting supermirror structure

DOEpatents

Wood, James L.

1992-01-01

An improved neutron reflecting supermirror structure comprising a plurality of stacked sets of bilayers of neutron reflecting materials. The improved neutron reflecting supermirror structure is adapted to provide extremely good performance at high incidence angles, i.e. up to four time the critical angle of standard neutron mirror structures. The reflection of neutrons striking the supermirror structure at a high critical angle provides enhanced neutron throughput, and hence more efficient and economical use of neutron sources. One layer of each set of bilayers consist of titanium, and the second layer of each set of bilayers consist of an alloy of nickel with carbon interstitially present in the nickel alloy.

Laissez-Faire : Fully Asymmetric Backscatter Communication

PubMed Central

Hu, Pan; Zhang, Pengyu; Ganesan, Deepak

2016-01-01

Backscatter provides dual-benefits of energy harvesting and low-power communication, making it attractive to a broad class of wireless sensors. But the design of a protocol that enables extremely power-efficient radios for harvesting-based sensors as well as high-rate data transfer for data-rich sensors presents a conundrum. In this paper, we present a new fully asymmetric backscatter communication protocol where nodes blindly transmit data as and when they sense. This model enables fully flexible node designs, from extraordinarily power-efficient backscatter radios that consume barely a few micro-watts to high-throughput radios that can stream at hundreds of Kbps while consuming a paltry tens of micro-watts. The challenge, however, lies in decoding concurrent streams at the reader, which we achieve using a novel combination of time-domain separation of interleaved signal edges, and phase-domain separation of colliding transmissions. We provide an implementation of our protocol, LF-Backscatter, and show that it can achieve an order of magnitude or more improvement in throughput, latency and power over state-of-art alternatives. PMID:28286885

Microextraction by packed sorbent: an emerging, selective and high-throughput extraction technique in bioanalysis.

PubMed

Pereira, Jorge; Câmara, José S; Colmsjö, Anders; Abdel-Rehim, Mohamed

2014-06-01

Sample preparation is an important analytical step regarding the isolation and concentration of desired components from complex matrices and greatly influences their reliable and accurate analysis and data quality. It is the most labor-intensive and error-prone process in analytical methodology and, therefore, may influence the analytical performance of the target analytes quantification. Many conventional sample preparation methods are relatively complicated, involving time-consuming procedures and requiring large volumes of organic solvents. Recent trends in sample preparation include miniaturization, automation, high-throughput performance, on-line coupling with analytical instruments and low-cost operation through extremely low volume or no solvent consumption. Micro-extraction techniques, such as micro-extraction by packed sorbent (MEPS), have these advantages over the traditional techniques. This paper gives an overview of MEPS technique, including the role of sample preparation in bioanalysis, the MEPS description namely MEPS formats (on- and off-line), sorbents, experimental and protocols, factors that affect the MEPS performance, and the major advantages and limitations of MEPS compared with other sample preparation techniques. We also summarize MEPS recent applications in bioanalysis. Copyright © 2014 John Wiley & Sons, Ltd.

Advances in single-cell experimental design made possible by automated imaging platforms with feedback through segmentation.

PubMed

Crick, Alex J; Cammarota, Eugenia; Moulang, Katie; Kotar, Jurij; Cicuta, Pietro

2015-01-01

Live optical microscopy has become an essential tool for studying the dynamical behaviors and variability of single cells, and cell-cell interactions. However, experiments and data analysis in this area are often extremely labor intensive, and it has often not been achievable or practical to perform properly standardized experiments on a statistically viable scale. We have addressed this challenge by developing automated live imaging platforms, to help standardize experiments, increasing throughput, and unlocking previously impossible ones. Our real-time cell tracking programs communicate in feedback with microscope and camera control software, and they are highly customizable, flexible, and efficient. As examples of our current research which utilize these automated platforms, we describe two quite different applications: egress-invasion interactions of malaria parasites and red blood cells, and imaging of immune cells which possess high motility and internal dynamics. The automated imaging platforms are able to track a large number of motile cells simultaneously, over hours or even days at a time, greatly increasing data throughput and opening up new experimental possibilities. Copyright © 2015 Elsevier Inc. All rights reserved.

Whole Wiskott‑Aldrich syndrome protein gene deletion identified by high throughput sequencing.

PubMed

He, Xiangling; Zou, Runying; Zhang, Bing; You, Yalan; Yang, Yang; Tian, Xin

2017-11-01

Wiskott‑Aldrich syndrome (WAS) is a rare X‑linked recessive immunodeficiency disorder, characterized by thrombocytopenia, small platelets, eczema and recurrent infections associated with increased risk of autoimmunity and malignancy disorders. Mutations in the WAS protein (WASP) gene are responsible for WAS. To date, WASP mutations, including missense/nonsense, splicing, small deletions, small insertions, gross deletions, and gross insertions have been identified in patients with WAS. In addition, WASP‑interacting proteins are suspected in patients with clinical features of WAS, in whom the WASP gene sequence and mRNA levels are normal. The present study aimed to investigate the application of next generation sequencing in definitive diagnosis and clinical therapy for WAS. A 5 month‑old child with WAS who displayed symptoms of thrombocytopenia was examined. Whole exome sequence analysis of genomic DNA showed that the coverage and depth of WASP were extremely low. Quantitative polymerase chain reaction indicated total WASP gene deletion in the proband. In conclusion, high throughput sequencing is useful for the verification of WAS on the genetic profile, and has implications for family planning guidance and establishment of clinical programs.

Evaluating reproducibility of differential expression discoveries in microarray studies by considering correlated molecular changes.

PubMed

Zhang, Min; Zhang, Lin; Zou, Jinfeng; Yao, Chen; Xiao, Hui; Liu, Qing; Wang, Jing; Wang, Dong; Wang, Chenguang; Guo, Zheng

2009-07-01

According to current consistency metrics such as percentage of overlapping genes (POG), lists of differentially expressed genes (DEGs) detected from different microarray studies for a complex disease are often highly inconsistent. This irreproducibility problem also exists in other high-throughput post-genomic areas such as proteomics and metabolism. A complex disease is often characterized with many coordinated molecular changes, which should be considered when evaluating the reproducibility of discovery lists from different studies. We proposed metrics percentage of overlapping genes-related (POGR) and normalized POGR (nPOGR) to evaluate the consistency between two DEG lists for a complex disease, considering correlated molecular changes rather than only counting gene overlaps between the lists. Based on microarray datasets of three diseases, we showed that though the POG scores for DEG lists from different studies for each disease are extremely low, the POGR and nPOGR scores can be rather high, suggesting that the apparently inconsistent DEG lists may be highly reproducible in the sense that they are actually significantly correlated. Observing different discovery results for a disease by the POGR and nPOGR scores will obviously reduce the uncertainty of the microarray studies. The proposed metrics could also be applicable in many other high-throughput post-genomic areas.

An Ultra-High Fluorescence Enhancement and High Throughput Assay for Revealing Expression and Internalization of Chemokine Receptor CXCR4.

PubMed

He, Hua; Wang, Xiaojuan; Cheng, Tiantian; Xia, Yongqing; Lao, Jun; Ge, Baosheng; Ren, Hao; Khan, Naseer Ullah; Huang, Fang

2016-04-18

Revealing chemokine receptor CXCR4 expression, distribution, and internalization levels in different cancers helps to evaluate cancer progression or prognosis and to set personalized treatment strategy. We here describe a sensitive and high-throughput immunoassay for determining CXCR4 expression and distribution in cancer cells. The assay is accessible to a wide range of users in an ordinary lab only by dip-coating poly(styrene-co-N-isopropylacrylamide) spheres on the glass substrate. The self- assembled spheres form three-dimensional photonic colloidal crystals which enhance the fluorescence of CF647 and Alexa Fluor 647 by a factor of up to 1000. CXCR4 in cells is detected by using the sandwich immunoassay, where the primary antibody recognizes CXCR4 and the secondary antibody is labeled with CF647. With the newly established assay, we quantified the total expression of CXCR4, its distribution on the cell membrane and cytoplasm, and revealed their internalization level upon SDF-1α activation in various cancer cells, even for those with extremely low expression level. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Fabrication of Single, Vertically Aligned Carbon Nanotubes in 3D Nanoscale Architectures

NASA Technical Reports Server (NTRS)

Kaul, Anupama B.; Megerian, Krikor G.; Von Allmen, Paul A.; Baron, Richard L.

2010-01-01

Plasma-enhanced chemical vapor deposition (PECVD) and high-throughput manufacturing techniques for integrating single, aligned carbon nanotubes (CNTs) into novel 3D nanoscale architectures have been developed. First, the PECVD growth technique ensures excellent alignment of the tubes, since the tubes align in the direction of the electric field in the plasma as they are growing. Second, the tubes generated with this technique are all metallic, so their chirality is predetermined, which is important for electronic applications. Third, a wafer-scale manufacturing process was developed that is high-throughput and low-cost, and yet enables the integration of just single, aligned tubes with nanoscale 3D architectures with unprecedented placement accuracy and does not rely on e-beam lithography. Such techniques should lend themselves to the integration of PECVD grown tubes for applications ranging from interconnects, nanoelectromechanical systems (NEMS), sensors, bioprobes, or other 3D electronic devices. Chemically amplified polyhydroxystyrene-resin-based deep UV resists were used in conjunction with excimer laser-based (lambda = 248 nm) step-and-repeat lithography to form Ni catalyst dots = 300 nm in diameter that nucleated single, vertically aligned tubes with high yield using dc PECVD growth. This is the first time such chemically amplified resists have been used, resulting in the nucleation of single, vertically aligned tubes. In addition, novel 3D nanoscale architectures have been created using topdown techniques that integrate single, vertically aligned tubes. These were enabled by implementing techniques that use deep-UV chemically amplified resists for small-feature-size resolution; optical lithography units that allow unprecedented control over layer-to-layer registration; and ICP (inductively coupled plasma) etching techniques that result in near-vertical, high-aspect-ratio, 3D nanoscale architectures, in conjunction with the use of materials that are structurally and chemically compatible with the high-temperature synthesis of the PECVD-grown tubes. The techniques offer a wafer-scale process solution for integrating single PECVD-grown nanotubes into novel architectures that should accelerate their integration in 3D electronics in general. NASA can directly benefit from this technology for its extreme-environment planetary missions. Current Si transistors are inherently more susceptible to high radiation, and do not tolerate extremes in temperature. These novel 3D nanoscale architectures can form the basis for NEMS switches that are inherently less susceptible to radiation or to thermal extremes.

Modeling high-efficiency extreme ultraviolet etched multilayer phase-shift masks

NASA Astrophysics Data System (ADS)

Sherwin, Stuart; Neureuther, Andrew; Naulleau, Patrick

2017-10-01

Achieving high-throughput extreme ultraviolet (EUV) patterning remains a major challenge due to low source power; phase-shift masks can help solve this challenge for dense features near the resolution limit by creating brighter images than traditional absorber masks when illuminated with the same source power. We explore applications of etched multilayer phase-shift masks for EUV lithography, both in the current-generation 0.33 NA and next-generation 0.55 NA systems. We derive analytic formulas for the thin-mask throughput gains, which are 2.42× for lines and spaces and 5.86× for contacts compared with an absorber mask with dipole and quadrupole illumination, respectively. Using rigorous finite-difference time-domain simulations, we quantify variations in these gains by pitch and orientation, finding 87% to 113% of the thin-mask value for lines and spaces and a 91% to 99% for contacts. We introduce an edge placement error metric, which accounts for CD errors, relative feature motion, and telecentricity errors, and use this metric both to optimize mask designs for individual features and to explore which features can be printed on the same mask. Furthermore, we find that although partial coherence shrinks the process window, at an achievable sigma of 0.2 we obtain a depth of focus of 340 nm and an exposure latitude of 39.2%, suggesting that partial coherence will not limit the feasibility of this technology. Finally, we show that many problems such as sensitivity to etch uniformity can be greatly mitigated using a central obscuration in the imaging pupil.

Rapid detection of single bacteria in unprocessed blood using Integrated Comprehensive Droplet Digital Detection

PubMed Central

Kang, Dong-Ku; Ali, M. Monsur; Zhang, Kaixiang; Huang, Susan S.; Peterson, Ellena; Digman, Michelle A.; Gratton, Enrico; Zhao, Weian

2014-01-01

Blood stream infection or sepsis is a major health problem worldwide, with extremely high mortality, which is partly due to the inability to rapidly detect and identify bacteria in the early stages of infection. Here we present a new technology termed ‘Integrated Comprehensive Droplet Digital Detection’ (IC 3D) that can selectively detect bacteria directly from milliliters of diluted blood at single-cell sensitivity in a one-step, culture- and amplification-free process within 1.5–4 h. The IC 3D integrates real-time, DNAzyme-based sensors, droplet microencapsulation and a high-throughput 3D particle counter system. Using Escherichia coli as a target, we demonstrate that the IC 3D can provide absolute quantification of both stock and clinical isolates of E. coli in spiked blood within a broad range of extremely low concentration from 1 to 10,000 bacteria per ml with exceptional robustness and limit of detection in the single digit regime. PMID:25391809

A quantitative and high-throughput assay of human papillomavirus DNA replication.

PubMed

Gagnon, David; Fradet-Turcotte, Amélie; Archambault, Jacques

2015-01-01

Replication of the human papillomavirus (HPV) double-stranded DNA genome is accomplished by the two viral proteins E1 and E2 in concert with host DNA replication factors. HPV DNA replication is an established model of eukaryotic DNA replication and a potential target for antiviral therapy. Assays to measure the transient replication of HPV DNA in transfected cells have been developed, which rely on a plasmid carrying the viral origin of DNA replication (ori) together with expression vectors for E1 and E2. Replication of the ori-plasmid is typically measured by Southern blotting or PCR analysis of newly replicated DNA (i.e., DpnI digested DNA) several days post-transfection. Although extremely valuable, these assays have been difficult to perform in a high-throughput and quantitative manner. Here, we describe a modified version of the transient DNA replication assay that circumvents these limitations by incorporating a firefly luciferase expression cassette in cis of the ori. Replication of this ori-plasmid by E1 and E2 results in increased levels of firefly luciferase activity that can be accurately quantified and normalized to those of Renilla luciferase expressed from a control plasmid, thus obviating the need for DNA extraction, digestion, and analysis. We provide a detailed protocol for performing the HPV type 31 DNA replication assay in a 96-well plate format suitable for small-molecule screening and EC50 determinations. The quantitative and high-throughput nature of the assay should greatly facilitate the study of HPV DNA replication and the identification of inhibitors thereof.

High-throughput genotyping for species identification and diversity assessment in germplasm collections.

PubMed

Mason, Annaliese S; Zhang, Jing; Tollenaere, Reece; Vasquez Teuber, Paula; Dalton-Morgan, Jessica; Hu, Liyong; Yan, Guijun; Edwards, David; Redden, Robert; Batley, Jacqueline

2015-09-01

Germplasm collections provide an extremely valuable resource for breeders and researchers. However, misclassification of accessions by species often hinders the effective use of these collections. We propose that use of high-throughput genotyping tools can provide a fast, efficient and cost-effective way of confirming species in germplasm collections, as well as providing valuable genetic diversity data. We genotyped 180 Brassicaceae samples sourced from the Australian Grains Genebank across the recently released Illumina Infinium Brassica 60K SNP array. Of these, 76 were provided on the basis of suspected misclassification and another 104 were sourced independently from the germplasm collection. Presence of the A- and C-genomes combined with principle components analysis clearly separated Brassica rapa, B. oleracea, B. napus, B. carinata and B. juncea samples into distinct species groups. Several lines were further validated using chromosome counts. Overall, 18% of samples (32/180) were misclassified on the basis of species. Within these 180 samples, 23/76 (30%) supplied on the basis of suspected misclassification were misclassified, and 9/105 (9%) of the samples randomly sourced from the Australian Grains Genebank were misclassified. Surprisingly, several individuals were also found to be the product of interspecific hybridization events. The SNP (single nucleotide polymorphism) array proved effective at confirming species, and provided useful information related to genetic diversity. As similar genomic resources become available for different crops, high-throughput molecular genotyping will offer an efficient and cost-effective method to screen germplasm collections worldwide, facilitating more effective use of these valuable resources by breeders and researchers. © 2015 John Wiley & Sons Ltd.

Flip the tip: an automated, high quality, cost-effective patch clamp screen.

PubMed

Lepple-Wienhues, Albrecht; Ferlinz, Klaus; Seeger, Achim; Schäfer, Arvid

2003-01-01

The race for creating an automated patch clamp has begun. Here, we present a novel technology to produce true gigaseals and whole cell preparations at a high rate. Suspended cells are flushed toward the tip of glass micropipettes. Seal, whole-cell break-in, and pipette/liquid handling are fully automated. Extremely stable seals and access resistance guarantee high recording quality. Data obtained from different cell types sealed inside pipettes show long-term stability, voltage clamp and seal quality, as well as block by compounds in the pM range. A flexible array of independent electrode positions minimizes consumables consumption at maximal throughput. Pulled micropipettes guarantee a proven gigaseal substrate with ultra clean and smooth surface at low cost.

Forged Under the Sun: Life and Art of Extremophiles from Andean Lakes.

PubMed

Albarracín, Virginia Helena; Gärtner, Wolfgang; Farias, María Eugenia

2016-01-01

High-altitude Andean lakes (HAAL) are a treasure chest for microbiological research in South America. Their indigenous microbial communities are exposed to extremely high UV irradiation and to multiple chemical extremes (Arsenic, high salt content, alkalinity). Microbes are found both, free-living or associated into microbial mats with different degrees of mineralization and lithification, including unique modern stromatolites located at 3570 m above sea level. Characterization of these polyextremophilic microbes began only recently, employing morphological and phylogenetic methods as well as high-throughput sequencing and proteomics approach. Aside from providing a general overview on microbial communities, special attention is given to various survival strategies; HAAL's microbes present a complex system of shared genetic and physiological mechanisms (UV-resistome) based on UV photoreceptors and stress sensors with their corresponding response regulators, UV avoidance and protection strategies, damage tolerance and UV damage repair. Molecular information will be provided for what is, so far the most studied HAAL molecule, a CPD-Class I photolyase from Acinetobacter Ver3 (Laguna Verde, 4400 m). This work further proposes some strategies that make an appeal for the preservation of HAAL, a highly fragile environment that offers promising and ample research possibilities. © 2015 The American Society of Photobiology.

Characterization of Extremely Lightweight Intrusion Detection (ELIDe) Power Utilization with Varying Throughput and Payload Sizes

DTIC Science & Technology

2015-09-01

Extremely Lightweight Intrusion Detection (ELIDe) algorithm on an Android -based mobile device. Our results show that the hashing and inner product...approximately 2.5 megabits per second (assuming a normal distribution of packet sizes) with no significant packet loss. 15. SUBJECT TERMS ELIDe, Android , pcap...system (OS). To run ELIDe, the current version was ported for use on Android .4 2.1 Mobile Device After ELIDe was ported to the Android mobile

Strategic and Operational Plan for Integrating Transcriptomics ...

EPA Pesticide Factsheets

Plans for incorporating high throughput transcriptomics into the current high throughput screening activities at NCCT; the details are in the attached slide presentation presentation on plans for incorporating high throughput transcriptomics into the current high throughput screening activities at NCCT, given at the OECD meeting on June 23, 2016

High-Throughput Experimental Approach Capabilities | Materials Science |

Science.gov Websites

NREL High-Throughput Experimental Approach Capabilities High-Throughput Experimental Approach by yellow and is for materials in the upper right sector. NREL's high-throughput experimental ,Te) and oxysulfide sputtering Combi-5: Nitrides and oxynitride sputtering We also have several non

A complex multi-notch astronomical filter to suppress the bright infrared sky.

PubMed

Bland-Hawthorn, J; Ellis, S C; Leon-Saval, S G; Haynes, R; Roth, M M; Löhmannsröben, H-G; Horton, A J; Cuby, J-G; Birks, T A; Lawrence, J S; Gillingham, P; Ryder, S D; Trinh, C

2011-12-06

A long-standing and profound problem in astronomy is the difficulty in obtaining deep near-infrared observations due to the extreme brightness and variability of the night sky at these wavelengths. A solution to this problem is crucial if we are to obtain the deepest possible observations of the early Universe, as redshifted starlight from distant galaxies appears at these wavelengths. The atmospheric emission between 1,000 and 1,800 nm arises almost entirely from a forest of extremely bright, very narrow hydroxyl emission lines that varies on timescales of minutes. The astronomical community has long envisaged the prospect of selectively removing these lines, while retaining high throughput between them. Here we demonstrate such a filter for the first time, presenting results from the first on-sky tests. Its use on current 8 m telescopes and future 30 m telescopes will open up many new research avenues in the years to come.

Introducing Bayesian thinking to high-throughput screening for false-negative rate estimation.

PubMed

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.

Carbon contamination topography analysis of EUV masks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fan, Y.-J.; Yankulin, L.; Thomas, P.

2010-03-12

The impact of carbon contamination on extreme ultraviolet (EUV) masks is significant due to throughput loss and potential effects on imaging performance. Current carbon contamination research primarily focuses on the lifetime of the multilayer surfaces, determined by reflectivity loss and reduced throughput in EUV exposure tools. However, contamination on patterned EUV masks can cause additional effects on absorbing features and the printed images, as well as impacting the efficiency of cleaning process. In this work, several different techniques were used to determine possible contamination topography. Lithographic simulations were also performed and the results compared with the experimental data.

Experimental station for ultrafast extreme ultraviolet spectroscopy for non-equilibrium dynamics in warm dense matter

NASA Astrophysics Data System (ADS)

Lee, Jong-won; Geng, Xiaotao; Jung, Jae Hyung; Cho, Min Sang; Yang, Seong Hyeok; Jo, Jawon; Lee, Chang-lyoul; Cho, Byoung Ick; Kim, Dong-Eon

2018-07-01

Recent interest in highly excited matter generated by intense femtosecond laser pulses has led to experimental methods that directly investigate ultrafast non-equilibrium electronic and structural dynamics. We present a tabletop experimental station for the extreme ultraviolet (EUV) spectroscopy used to trace L-edge dynamics in warm dense aluminum with a temporal resolution of a hundred femtoseconds. The system consists of the EUV probe generation part via a high-order harmonic generation process of femtosecond laser pulses with atomic clusters, a beamline with high-throughput optics and a sample-refreshment system of nano-foils utilizing the full repetition rate of the probe, and a flat-field EUV spectrograph. With the accumulation of an order of a hundred shots, a clear observation of the change in the aluminum L-shell absorption was achieved with a temporal resolution of 90 fs in a 600-fs window. The signature of a non-equilibrium electron distribution over a 10-eV range and its evolution to a 1-eV Fermi distribution are observed. This demonstrates the capability of this apparatus to capture the non-equilibrium electron-hole dynamics in highly excited warm dense matter conditions.

Ribosomal Binding Site Switching: An Effective Strategy for High-Throughput Cloning Constructions

PubMed Central

Li, Yunlong; Zhang, Yong; Lu, Pei; Rayner, Simon; Chen, Shiyun

2012-01-01

Direct cloning of PCR fragments by TA cloning or blunt end ligation are two simple methods which would greatly benefit high-throughput (HTP) cloning constructions if the efficiency can be improved. In this study, we have developed a ribosomal binding site (RBS) switching strategy for direct cloning of PCR fragments. RBS is an A/G rich region upstream of the translational start codon and is essential for gene expression. Change from A/G to T/C in the RBS blocks its activity and thereby abolishes gene expression. Based on this property, we introduced an inactive RBS upstream of a selectable marker gene, and designed a fragment insertion site within this inactive RBS. Forward and reverse insertions of specifically tailed fragments will respectively form an active and inactive RBS, thus all background from vector self-ligation and fragment reverse insertions will be eliminated due to the non-expression of the marker gene. The effectiveness of our strategy for TA cloning and blunt end ligation are confirmed. Application of this strategy to gene over-expression, a bacterial two-hybrid system, a bacterial one-hybrid system, and promoter bank construction are also verified. The advantages of this simple procedure, together with its low cost and high efficiency, makes our strategy extremely useful in HTP cloning constructions. PMID:23185557

Microfluidic system for high throughput characterisation of echogenic particles.

PubMed

Rademeyer, Paul; Carugo, Dario; Lee, Jeong Yu; Stride, Eleanor

2015-01-21

Echogenic particles, such as microbubbles and volatile liquid micro/nano droplets, have shown considerable potential in a variety of clinical diagnostic and therapeutic applications. The accurate prediction of their response to ultrasound excitation is however extremely challenging, and this has hindered the optimisation of techniques such as quantitative ultrasound imaging and targeted drug delivery. Existing characterisation techniques, such as ultra-high speed microscopy provide important insights, but suffer from a number of limitations; most significantly difficulty in obtaining large data sets suitable for statistical analysis and the need to physically constrain the particles, thereby altering their dynamics. Here a microfluidic system is presented that overcomes these challenges to enable the measurement of single echogenic particle response to ultrasound excitation. A co-axial flow focusing device is used to direct a continuous stream of unconstrained particles through the combined focal region of an ultrasound transducer and a laser. Both the optical and acoustic scatter from individual particles are then simultaneously recorded. Calibration of the device and example results for different types of echogenic particle are presented, demonstrating a high throughput of up to 20 particles per second and the ability to resolve changes in particle radius down to 0.1 μm with an uncertainty of less than 3%.

Monolithic Hydrogen Peroxide Catalyst Bed Development

NASA Technical Reports Server (NTRS)

Ponzo, J. B.

2003-01-01

With recent increased industry and government interest in rocket grade hydrogen peroxide as a viable propellant, significant effort has been expended to improve on earlier developments. This effort has been predominately centered in improving heterogeneous. typically catalyst beds; and homogeneous catalysts, which are typically solutions of catalytic substances. Heterogeneous catalyst beds have traditionally consisted of compressed wire screens plated with a catalytic substance, usually silver, and were used m many RCS applications (X-1, Mercury, and Centaur for example). Aerojet has devised a heterogeneous catalyst design that is monolithic (single piece), extremely compact, and has pressure drops equal to or less than traditional screen beds. The design consists of a bonded stack of very thin, photoetched metal plates, silver coated. This design leads to a high surface area per unit volume and precise flow area, resulting in high, stable, and repeatable performance. Very high throughputs have been demonstrated with 90% hydrogen peroxide. (0.60 lbm/s/sq in at 1775-175 psia) with no flooding of the catalyst bed. Bed life of over 900 seconds has also been demonstrated at throughputs of 0.60 lbm/s/sq in across varying chamber pressures. The monolithic design also exhibits good starting performance, short break-in periods, and will easily scale to various sizes.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sauter, Nicholas K., E-mail: nksauter@lbl.gov; Hattne, Johan; Grosse-Kunstleve, Ralf W.

The Computational Crystallography Toolbox (cctbx) is a flexible software platform that has been used to develop high-throughput crystal-screening tools for both synchrotron sources and X-ray free-electron lasers. Plans for data-processing and visualization applications are discussed, and the benefits and limitations of using graphics-processing units are evaluated. Current pixel-array detectors produce diffraction images at extreme data rates (of up to 2 TB h{sup −1}) that make severe demands on computational resources. New multiprocessing frameworks are required to achieve rapid data analysis, as it is important to be able to inspect the data quickly in order to guide the experiment in realmore » time. By utilizing readily available web-serving tools that interact with the Python scripting language, it was possible to implement a high-throughput Bragg-spot analyzer (cctbx.spotfinder) that is presently in use at numerous synchrotron-radiation beamlines. Similarly, Python interoperability enabled the production of a new data-reduction package (cctbx.xfel) for serial femtosecond crystallography experiments at the Linac Coherent Light Source (LCLS). Future data-reduction efforts will need to focus on specialized problems such as the treatment of diffraction spots on interleaved lattices arising from multi-crystal specimens. In these challenging cases, accurate modeling of close-lying Bragg spots could benefit from the high-performance computing capabilities of graphics-processing units.« less

A suite of MATLAB-based computational tools for automated analysis of COPAS Biosort data

PubMed Central

Morton, Elizabeth; Lamitina, Todd

2010-01-01

Complex Object Parametric Analyzer and Sorter (COPAS) devices are large-object, fluorescence-capable flow cytometers used for high-throughput analysis of live model organisms, including Drosophila melanogaster, Caenorhabditis elegans, and zebrafish. The COPAS is especially useful in C. elegans high-throughput genome-wide RNA interference (RNAi) screens that utilize fluorescent reporters. However, analysis of data from such screens is relatively labor-intensive and time-consuming. Currently, there are no computational tools available to facilitate high-throughput analysis of COPAS data. We used MATLAB to develop algorithms (COPAquant, COPAmulti, and COPAcompare) to analyze different types of COPAS data. COPAquant reads single-sample files, filters and extracts values and value ratios for each file, and then returns a summary of the data. COPAmulti reads 96-well autosampling files generated with the ReFLX adapter, performs sample filtering, graphs features across both wells and plates, performs some common statistical measures for hit identification, and outputs results in graphical formats. COPAcompare performs a correlation analysis between replicate 96-well plates. For many parameters, thresholds may be defined through a simple graphical user interface (GUI), allowing our algorithms to meet a variety of screening applications. In a screen for regulators of stress-inducible GFP expression, COPAquant dramatically accelerated data analysis and allowed us to rapidly move from raw data to hit identification. Because the COPAS file structure is standardized and our MATLAB code is freely available, our algorithms should be extremely useful for analysis of COPAS data from multiple platforms and organisms. The MATLAB code is freely available at our web site (www.med.upenn.edu/lamitinalab/downloads.shtml). PMID:20569218

ALFALFA DISCOVERY OF THE MOST METAL-POOR GAS-RICH GALAXY KNOWN: AGC 198691

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hirschauer, Alec S.; Salzer, John J.; Rhode, Katherine L., E-mail: ash@astro.indiana.edu, E-mail: slaz@astro.indiana.edu, E-mail: krhode@indiana.edu

We present spectroscopic observations of the nearby dwarf galaxy AGC 198691. This object is part of the Survey of H i in Extremely Low-Mass Dwarfs project, which is a multi-wavelength study of galaxies with H i masses in the range of 10{sup 6}–10{sup 7.2} M {sub ⊙}, discovered by the Arecibo Legacy Fast ALFA (ALFALFA) survey. We have obtained spectra of the lone H ii region in AGC 198691 with the new high-throughput KPNO Ohio State Multi-Object Spectrograph on the Mayall 4 m, as well as with the Blue Channel spectrograph on the MMT 6.5 m telescope. These observations enablemore » the measurement of the temperature-sensitive [O iii] λ 4363 line and hence the determination of a “direct” oxygen abundance for AGC 198691. We find this system to be an extremely metal-deficient (XMD) system with an oxygen abundance of 12+log(O/H) = 7.02 ± 0.03, making AGC 198691 the lowest-abundance star-forming galaxy known in the local universe. Two of the five lowest-abundance galaxies known have been discovered by the ALFALFA blind H i survey; this high yield of XMD galaxies represents a paradigm shift in the search for extremely metal-poor galaxies.« less

A world of opportunities with nanopore sequencing.

PubMed

Leggett, Richard M; Clark, Matthew D

2017-11-28

Oxford Nanopore Technologies' MinION sequencer was launched in pre-release form in 2014 and represents an exciting new sequencing paradigm. The device offers multi-kilobase reads and a streamed mode of operation that allows processing of reads as they are generated. Crucially, it is an extremely compact device that is powered from the USB port of a laptop computer, enabling it to be taken out of the lab and facilitating previously impossible in-field sequencing experiments to be undertaken. Many of the initial publications concerning the platform focused on provision of tools to access and analyse the new sequence formats and then demonstrating the assembly of microbial genomes. More recently, as throughput and accuracy have increased, it has been possible to begin work involving more complex genomes and metagenomes. With the release of the high-throughput GridION X5 and PromethION platforms, the sequencing of large genomes will become more cost efficient, and enable the leveraging of extremely long (>100 kb) reads for resolution of complex genomic structures. This review provides a brief overview of nanopore sequencing technology, describes the growing range of nanopore bioinformatics tools, and highlights some of the most influential publications that have emerged over the last 2 years. Finally, we look to the future and the potential the platform has to disrupt work in human, microbiome, and plant genomics. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Making big sense from big data in toxicology by read-across.

PubMed

Hartung, Thomas

2016-01-01

Modern information technologies have made big data available in safety sciences, i.e., extremely large data sets that may be analyzed only computationally to reveal patterns, trends and associations. This happens by (1) compilation of large sets of existing data, e.g., as a result of the European REACH regulation, (2) the use of omics technologies and (3) systematic robotized testing in a high-throughput manner. All three approaches and some other high-content technologies leave us with big data--the challenge is now to make big sense of these data. Read-across, i.e., the local similarity-based intrapolation of properties, is gaining momentum with increasing data availability and consensus on how to process and report it. It is predominantly applied to in vivo test data as a gap-filling approach, but can similarly complement other incomplete datasets. Big data are first of all repositories for finding similar substances and ensure that the available data is fully exploited. High-content and high-throughput approaches similarly require focusing on clusters, in this case formed by underlying mechanisms such as pathways of toxicity. The closely connected properties, i.e., structural and biological similarity, create the confidence needed for predictions of toxic properties. Here, a new web-based tool under development called REACH-across, which aims to support and automate structure-based read-across, is presented among others.

Pyrosequencing-based validation of a simple cell-suspension polymerase chain reaction assay for Campylobacter with application of high-processivity polymerase and novel internal amplification controls for rapid and specific detection.

PubMed

Oakley, Brian B; Line, J Eric; Berrang, Mark E; Johnson, Jessica M; Buhr, R Jeff; Cox, Nelson A; Hiett, Kelli L; Seal, Bruce S

2012-02-01

Although Campylobacter is an important food-borne human pathogen, there remains a lack of molecular diagnostic assays that are simple to use, cost-effective, and provide rapid results in research, clinical, or regulatory laboratories. Of the numerous Campylobacter assays that do exist, to our knowledge none has been empirically tested for specificity using high-throughput sequencing. Here we demonstrate the power of next-generation sequencing to determine the specificity of a widely cited Campylobacter-specific polymerase chain reaction (PCR) assay and describe a rapid method for direct cell suspension PCR to quickly and easily screen samples for Campylobacter. We present a specific protocol which eliminates the need for time-consuming and expensive genomic DNA extractions and, using a high-processivity polymerase, demonstrate conclusive screening of samples in <1 h. Pyrosequencing results show the assay to be extremely (>99%) sensitive, and spike-back experiments demonstrated a detection threshold of <10(2) CFU mL(-1). Additionally, we present 2 newly designed broad-range bacterial primer sets targeting the 23S rRNA gene that have wide applicability as internal amplification controls. Empirical testing of putative taxon-specific assays using high-throughput sequencing is an important validation step that is now financially feasible for research, regulatory, or clinical applications. Published by Elsevier Inc.

Printed Carbon Nanotube Electronics and Sensor Systems.

PubMed

Chen, Kevin; Gao, Wei; Emaminejad, Sam; Kiriya, Daisuke; Ota, Hiroki; Nyein, Hnin Yin Yin; Takei, Kuniharu; Javey, Ali

2016-06-01

Printing technologies offer large-area, high-throughput production capabilities for electronics and sensors on mechanically flexible substrates that can conformally cover different surfaces. These capabilities enable a wide range of new applications such as low-cost disposable electronics for health monitoring and wearables, extremely large format electronic displays, interactive wallpapers, and sensing arrays. Solution-processed carbon nanotubes have been shown to be a promising candidate for such printing processes, offering stable devices with high performance. Here, recent progress made in printed carbon nanotube electronics is discussed in terms of materials, processing, devices, and applications. Research challenges and opportunities moving forward from processing and system-level integration points of view are also discussed for enabling practical applications. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cancer cell classification with coherent diffraction imaging using an extreme ultraviolet radiation source

PubMed Central

Zürch, Michael; Foertsch, Stefan; Matzas, Mark; Pachmann, Katharina; Kuth, Rainer; Spielmann, Christian

2014-01-01

Abstract. In cancer treatment, it is highly desirable to classify single cancer cells in real time. The standard method is polymerase chain reaction requiring a substantial amount of resources and time. Here, we present an innovative approach for rapidly classifying different cell types: we measure the diffraction pattern of a single cell illuminated with coherent extreme ultraviolet (XUV) laser-generated radiation. These patterns allow distinguishing different breast cancer cell types in a subsequent step. Moreover, the morphology of the object can be retrieved from the diffraction pattern with submicron resolution. In a proof-of-principle experiment, we prepared single MCF7 and SKBR3 breast cancer cells on gold-coated silica slides. The output of a laser-driven XUV light source is focused onto a single unstained and unlabeled cancer cell. With the resulting diffraction pattern, we could clearly identify the different cell types. With an improved setup, it will not only be feasible to classify circulating tumor cells with a high throughput, but also to identify smaller objects such as bacteria or even viruses. PMID:26158049

3D Microfabrication Using Emulsion Mask Grayscale Photolithography Technique

NASA Astrophysics Data System (ADS)

Lee, Tze Pin; Mohamed, Khairudin

2016-02-01

Recently, the rapid development of technology such as biochips, microfluidic, micro-optical devices and micro-electromechanical-systems (MEMS) demands the capability to create complex design of three-dimensional (3D) microstructures. In order to create 3D microstructures, the traditional photolithography process often requires multiple photomasks to form 3D pattern from several stacked photoresist layers. This fabrication method is extremely time consuming, low throughput, costly and complicated to conduct for high volume manufacturing scale. On the other hand, next generation lithography such as electron beam lithography (EBL), focused ion beam lithography (FIB) and extreme ultraviolet lithography (EUV) are however too costly and the machines require expertise to setup. Therefore, the purpose of this study is to develop a simplified method in producing 3D microstructures using single grayscale emulsion mask technique. By using this grayscale fabrication method, microstructures of thickness as high as 500μm and as low as 20μm are obtained in a single photolithography exposure. Finally, the fabrication of 3D microfluidic channel has been demonstrated by using this grayscale photolithographic technique.

Recent advances in superhydrophobic surfaces and their relevance to biology and medicine.

PubMed

Ciasca, G; Papi, M; Businaro, L; Campi, G; Ortolani, M; Palmieri, V; Cedola, A; De Ninno, A; Gerardino, A; Maulucci, G; De Spirito, M

2016-02-04

By mimicking naturally occurring superhydrophobic surfaces, scientists can now realize artificial surfaces on which droplets of a few microliters of water are forced to assume an almost spherical shape and an extremely high contact angle. In recent decades, these surfaces have attracted much attention due to their technological applications for anti-wetting and self-cleaning materials. Very recently, researchers have shifted their interest to investigate whether superhydrophobic surfaces can be exploited to study biological systems. This research effort has stimulated the design and realization of new devices that allow us to actively organize, visualize and manipulate matter at both the microscale and nanoscale levels. Such precise control opens up wide applications in biomedicine, as it allows us to directly manipulate objects at the typical length scale of cells and macromolecules. This progress report focuses on recent biological and medical applications of superhydrophobicity. Particular regard is paid to those applications that involve the detection, manipulation and study of extremely small quantities of molecules, and to those that allow high throughput cell and biomaterial screening.

Cancer cell classification with coherent diffraction imaging using an extreme ultraviolet radiation source.

PubMed

Zürch, Michael; Foertsch, Stefan; Matzas, Mark; Pachmann, Katharina; Kuth, Rainer; Spielmann, Christian

2014-10-01

In cancer treatment, it is highly desirable to classify single cancer cells in real time. The standard method is polymerase chain reaction requiring a substantial amount of resources and time. Here, we present an innovative approach for rapidly classifying different cell types: we measure the diffraction pattern of a single cell illuminated with coherent extreme ultraviolet (XUV) laser-generated radiation. These patterns allow distinguishing different breast cancer cell types in a subsequent step. Moreover, the morphology of the object can be retrieved from the diffraction pattern with submicron resolution. In a proof-of-principle experiment, we prepared single MCF7 and SKBR3 breast cancer cells on gold-coated silica slides. The output of a laser-driven XUV light source is focused onto a single unstained and unlabeled cancer cell. With the resulting diffraction pattern, we could clearly identify the different cell types. With an improved setup, it will not only be feasible to classify circulating tumor cells with a high throughput, but also to identify smaller objects such as bacteria or even viruses.

X ray, extreme and far ultraviolet optical thin films for space applications

NASA Technical Reports Server (NTRS)

Zukic, Muamer; Torr, Douglas G.; Kim, Jongmin

1993-01-01

Far and extreme ultraviolet optical thin film filters find many uses in space astronomy, space astrophysics, and space aeronomy. Spacebased spectrographs are used for studying emission and absorption features of the earth, planets, sun, stars, and the interstellar medium. Most of these spectrographs use transmission or reflection filters. This requirement has prompted a search for selective filtering coatings with high throughput in the FUV and EUV spectral region. Important progress toward the development of thin film filters with improved efficiency and stability has been made in recent years. The goal for this field is the minimization of absorption to get high throughput and enhancement of wavelength selection. The Optical Aeronomy Laboratory (OAL) at the University of Alabama in Huntsville has recently developed the technology to determine optical constants of bulk and film materials for wavelengths extending from x-rays (0.1 nm) to the FUV (200 nm), and several materials have been identified that were used for designs of various optical devices which previously have been restricted to space application in the visible and near infrared. A new design concept called the Pi-multilayer was introduced and applied to the design of optical coatings for wavelengths extending from x-rays to the FUV. Section 3 of this report explains the Pi-multilayer approach and demonstrates its application for the design and fabrication of the FUV coatings. Two layer Pi-stacks have been utilized for the design of reflection filters in the EUV wavelength range from 70 - 100 nm. In order to eliminate losses due to the low reflection of the imaging optics and increase throughput and out-of-band rejection of the EUV instrumentation we introduced a self-filtering camera concept. In the FUV region, MgF2 and LiF crystals are known to be birefringent. Transmission polarizers and quarterwave retarders made of MgF2 or LiF crystals are commercially available but the performances are poor. New techniques for the design of the EUV and FUV polarizers and quarterwave retarders are described in Section 5. X- and gamma-ray detectors rely on a measurement of the electron which is effected when a ray interacts with matter. The design of an x- and gamma-ray telescope to operate in a particular region of the spectrum is, therefore, largely dictated by the mechanism through which the rays interact. Energy selection and the focusing of the incident high energy rays can be achieved with spectrally selective high reflective multilayers. The design and spectral performance of narrowband reflective x-ray Pi-multilayers are presented in section 6.

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

EPA Pesticide Factsheets

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

Real-Time Label-Free Surface Plasmon Resonance Biosensing with Gold Nanohole Arrays Fabricated by Nanoimprint Lithography

PubMed Central

Martinez-Perdiguero, Josu; Retolaza, Aritz; Otaduy, Deitze; Juarros, Aritz; Merino, Santos

2013-01-01

In this work we present a surface plasmon resonance sensor based on enhanced optical transmission through sub-wavelength nanohole arrays. This technique is extremely sensitive to changes in the refractive index of the surrounding medium which result in a modulation of the transmitted light. The periodic gold nanohole array sensors were fabricated by high-throughput thermal nanoimprint lithography. Square periodic arrays with sub-wavelength hole diameters were obtained and characterized. Using solutions with known refractive index, the array sensitivities were obtained. Finally, protein absorption was monitored in real-time demonstrating the label-free biosensing capabilities of the fabricated devices. PMID:24135989

Unlocking the Barite Paleoproductivity Proxy: Using a New Barite Extraction Method to Understand Productivity Trends During the Eocene Greenhouse

NASA Astrophysics Data System (ADS)

House, B. M.; Norris, R. D.

2017-12-01

The Early Eocene Climatic Optimum (EECO) around 50 Ma was a sustained period of extreme global warmth with ocean bottom water temperatures of up to 12° C. The marine biologic response to such climatic extremes is unclear, however, in part because proxies that integrate ecosystem-wide productivity signals are scarce. While the accumulation of marine barite (BaSO4) is one such proxy, its applicability has remained limited due to the difficulty in reliably quantifying barite. Discrete measurements of barite content in marine sediments are laborious, and indirect estimates provide unclear results. We have developed a fast, high-throughput method for reliable measurement of barite content that relies on selective extraction of barite rather than sample digestion and quantification of remaining barite. Tests of the new method reveal that it gives the expected results for a wide variety of sediment types and can quantitatively extract 10-100 times the amount of barite typically encountered in natural sediments. Altogether, our method provides an estimated ten-fold increase in analysis efficiency over current sample digestion methods and also works reliably on small ( 1 g or less) sediment samples. Furthermore, the instrumentation requirements of this method are minor, so samples can be analyzed in shipboard labs to generate real-time paleoproductivity records during coring expeditions. Because of the magnitude of throughput improvement, this new technique will permit the generation of large datasets needed to address previously intractable paleoclimate and paleoceanographic questions. One such question is how export productivity changes during climatic extremes. We used our new method to analyze globally distributed sediment cores to determine if the EECO represented a period of anomalous export productivity either due to higher rates of primary production or more vigorous heterotrophic metabolisms. An increase in export productivity could provide a mechanism for exiting periods of extreme warmth, and understanding the interplay between temperature, atmospheric CO2 levels, and export productivity during the EECO will help clarify how the marine biologic system functions as a whole.

Application of ToxCast High-Throughput Screening and ...

EPA Pesticide Factsheets

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

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

PubMed

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.

High Throughput Screening For Hazard and Risk of Environmental Contaminants

EPA Science Inventory

High throughput toxicity testing provides detailed mechanistic information on the concentration response of environmental contaminants in numerous potential toxicity pathways. High throughput screening (HTS) has several key advantages: (1) expense orders of magnitude less than an...

Cross-layer model design in wireless ad hoc networks for the Internet of Things.

PubMed

Yang, Xin; Wang, Ling; Xie, Jian; Zhang, Zhaolin

2018-01-01

Wireless ad hoc networks can experience extreme fluctuations in transmission traffic in the Internet of Things, which is widely used today. Currently, the most crucial issues requiring attention for wireless ad hoc networks are making the best use of low traffic periods, reducing congestion during high traffic periods, and improving transmission performance. To solve these problems, the present paper proposes a novel cross-layer transmission model based on decentralized coded caching in the physical layer and a content division multiplexing scheme in the media access control layer. Simulation results demonstrate that the proposed model effectively addresses these issues by substantially increasing the throughput and successful transmission rate compared to existing protocols without a negative influence on delay, particularly for large scale networks under conditions of highly contrasting high and low traffic periods.

Cross-layer model design in wireless ad hoc networks for the Internet of Things

PubMed Central

Wang, Ling; Xie, Jian; Zhang, Zhaolin

2018-01-01

Wireless ad hoc networks can experience extreme fluctuations in transmission traffic in the Internet of Things, which is widely used today. Currently, the most crucial issues requiring attention for wireless ad hoc networks are making the best use of low traffic periods, reducing congestion during high traffic periods, and improving transmission performance. To solve these problems, the present paper proposes a novel cross-layer transmission model based on decentralized coded caching in the physical layer and a content division multiplexing scheme in the media access control layer. Simulation results demonstrate that the proposed model effectively addresses these issues by substantially increasing the throughput and successful transmission rate compared to existing protocols without a negative influence on delay, particularly for large scale networks under conditions of highly contrasting high and low traffic periods. PMID:29734355

High Throughput Transcriptomics: From screening to pathways

EPA Science Inventory

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...

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

NASA Astrophysics Data System (ADS)

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.

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

PubMed

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

2017-01-01

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.

High Throughput Experimental Materials Database

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zakutayev, Andriy; Perkins, John; Schwarting, Marcus

The mission of the High Throughput Experimental Materials Database (HTEM DB) is to enable discovery of new materials with useful properties by releasing large amounts of high-quality experimental data to public. The HTEM DB contains information about materials obtained from high-throughput experiments at the National Renewable Energy Laboratory (NREL).

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

EPA Science Inventory

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...

Evaluation of Sequencing Approaches for High-Throughput Transcriptomics - (BOSC)

EPA Science Inventory

Whole-genome in vitro transcriptomics has shown the capability to identify mechanisms of action and estimates of potency for chemical-mediated effects in a toxicological framework, but with limited throughput and high cost. The generation of high-throughput global gene expression...

Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography

PubMed Central

Mueller, C.; Marx, A.; Epp, S. W.; Zhong, Y.; Kuo, A.; Balo, A. R.; Soman, J.; Schotte, F.; Lemke, H. T.; Owen, R. L.; Pai, E. F.; Pearson, A. R.; Olson, J. S.; Anfinrud, P. A.; Ernst, O. P.; Dwayne Miller, R. J.

2015-01-01

We present a crystallography chip enabling in situ room temperature crystallography at microfocus synchrotron beamlines and X-ray free-electron laser (X-FEL) sources. Compared to other in situ approaches, we observe extremely low background and high diffraction data quality. The chip design is robust and allows fast and efficient loading of thousands of small crystals. The ability to load a large number of protein crystals, at room temperature and with high efficiency, into prescribed positions enables high throughput automated serial crystallography with microfocus synchrotron beamlines. In addition, we demonstrate the application of this chip for femtosecond time-resolved serial crystallography at the Linac Coherent Light Source (LCLS, Menlo Park, California, USA). The chip concept enables multiple images to be acquired from each crystal, allowing differential detection of changes in diffraction intensities in order to obtain high signal-to-noise and fully exploit the time resolution capabilities of XFELs. PMID:26798825

Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography.

PubMed

Mueller, C; Marx, A; Epp, S W; Zhong, Y; Kuo, A; Balo, A R; Soman, J; Schotte, F; Lemke, H T; Owen, R L; Pai, E F; Pearson, A R; Olson, J S; Anfinrud, P A; Ernst, O P; Dwayne Miller, R J

2015-09-01

We present a crystallography chip enabling in situ room temperature crystallography at microfocus synchrotron beamlines and X-ray free-electron laser (X-FEL) sources. Compared to other in situ approaches, we observe extremely low background and high diffraction data quality. The chip design is robust and allows fast and efficient loading of thousands of small crystals. The ability to load a large number of protein crystals, at room temperature and with high efficiency, into prescribed positions enables high throughput automated serial crystallography with microfocus synchrotron beamlines. In addition, we demonstrate the application of this chip for femtosecond time-resolved serial crystallography at the Linac Coherent Light Source (LCLS, Menlo Park, California, USA). The chip concept enables multiple images to be acquired from each crystal, allowing differential detection of changes in diffraction intensities in order to obtain high signal-to-noise and fully exploit the time resolution capabilities of XFELs.

High-speed ultrafast laser machining with tertiary beam positioning (Conference Presentation)

NASA Astrophysics Data System (ADS)

Yang, Chuan; Zhang, Haibin

2017-03-01

For an industrial laser application, high process throughput and low average cost of ownership are critical to commercial success. Benefiting from high peak power, nonlinear absorption and small-achievable spot size, ultrafast lasers offer advantages of minimal heat affected zone, great taper and sidewall quality, and small via capability that exceeds the limits of their predecessors in via drilling for electronic packaging. In the past decade, ultrafast lasers have both grown in power and reduced in cost. For example, recently, disk and fiber technology have both shown stable operation in the 50W to 200W range, mostly at high repetition rate (beyond 500 kHz) that helps avoid detrimental nonlinear effects. However, to effectively and efficiently scale the throughput with the fast-growing power capability of the ultrafast lasers while keeping the beneficial laser-material interactions is very challenging, mainly because of the bottleneck imposed by the inertia-related acceleration limit and servo gain bandwidth when only stages and galvanometers are being used. On the other side, inertia-free scanning solutions like acoustic optics and electronic optical deflectors have small scan field, and therefore not suitable for large-panel processing. Our recent system developments combine stages, galvanometers, and AODs into a coordinated tertiary architecture for high bandwidth and meanwhile large field beam positioning. Synchronized three-level movements allow extremely fast local speed and continuous motion over the whole stage travel range. We present the via drilling results from such ultrafast system with up to 3MHz pulse to pulse random access, enabling high quality low cost ultrafast machining with emerging high average power laser sources.

Open Access, Retention and Throughput at the Central University of Technology

ERIC Educational Resources Information Center

de Beer, K. J.

2006-01-01

The most debatable question in higher education today is: Why first "open access" to promote massafication and now "capping" to restrict learner intake? (cf. SA Media Information 2004). Concerning the managing of this difficult and extremely sensitive issue, the Central University of Technology, Free State (CUT) has come a long…

Si-compatible cleaning process for graphene using low-density inductively coupled plasma.

PubMed

Lim, Yeong-Dae; Lee, Dae-Yeong; Shen, Tian-Zi; Ra, Chang-Ho; Choi, Jae-Young; Yoo, Won Jong

2012-05-22

We report a novel cleaning technique for few-layer graphene (FLG) by using inductively coupled plasma (ICP) of Ar with an extremely low plasma density of 3.5 × 10(8) cm(-3). It is known that conventional capacitively coupled plasma (CCP) treatments destroy the planar symmetry of FLG, giving rise to the generation of defects. However, ICP treatment with extremely low plasma density is able to remove polymer resist residues from FLG within 3 min at a room temperature of 300 K while retaining the carbon sp(2)-bonding of FLG. It is found that the carrier mobility and charge neutrality point of FLG are restored to their pristine defect-free state after the ICP treatment. Considering the application of graphene to silicon-based electronic devices, such a cleaning method can replace thermal vacuum annealing, electrical current annealing, and wet-chemical treatment due to its advantages of being a low-temperature, large-area, high-throughput, and Si-compatible process.

Hierarchy and extremes in selections from pools of randomized proteins

PubMed Central

Boyer, Sébastien; Biswas, Dipanwita; Kumar Soshee, Ananda; Scaramozzino, Natale; Nizak, Clément; Rivoire, Olivier

2016-01-01

Variation and selection are the core principles of Darwinian evolution, but quantitatively relating the diversity of a population to its capacity to respond to selection is challenging. Here, we examine this problem at a molecular level in the context of populations of partially randomized proteins selected for binding to well-defined targets. We built several minimal protein libraries, screened them in vitro by phage display, and analyzed their response to selection by high-throughput sequencing. A statistical analysis of the results reveals two main findings. First, libraries with the same sequence diversity but built around different “frameworks” typically have vastly different responses; second, the distribution of responses of the best binders in a library follows a simple scaling law. We show how an elementary probabilistic model based on extreme value theory rationalizes the latter finding. Our results have implications for designing synthetic protein libraries, estimating the density of functional biomolecules in sequence space, characterizing diversity in natural populations, and experimentally investigating evolvability (i.e., the potential for future evolution). PMID:26969726

Hierarchy and extremes in selections from pools of randomized proteins.

PubMed

Boyer, Sébastien; Biswas, Dipanwita; Kumar Soshee, Ananda; Scaramozzino, Natale; Nizak, Clément; Rivoire, Olivier

2016-03-29

Variation and selection are the core principles of Darwinian evolution, but quantitatively relating the diversity of a population to its capacity to respond to selection is challenging. Here, we examine this problem at a molecular level in the context of populations of partially randomized proteins selected for binding to well-defined targets. We built several minimal protein libraries, screened them in vitro by phage display, and analyzed their response to selection by high-throughput sequencing. A statistical analysis of the results reveals two main findings. First, libraries with the same sequence diversity but built around different "frameworks" typically have vastly different responses; second, the distribution of responses of the best binders in a library follows a simple scaling law. We show how an elementary probabilistic model based on extreme value theory rationalizes the latter finding. Our results have implications for designing synthetic protein libraries, estimating the density of functional biomolecules in sequence space, characterizing diversity in natural populations, and experimentally investigating evolvability (i.e., the potential for future evolution).

Reversible Conversion of Dominant Polarity in Ambipolar Polymer/Graphene Oxide Hybrids

PubMed Central

Zhou, Ye; Han, Su-Ting; Sonar, Prashant; Ma, Xinlei; Chen, Jihua; Zheng, Zijian; Roy, V. A. L.

2015-01-01

The possibility to selectively modulate the charge carrier transport in semiconducting materials is extremely challenging for the development of high performance and low-power consuming logic circuits. Systematical control over the polarity (electrons and holes) in transistor based on solution processed layer by layer polymer/graphene oxide hybrid system has been demonstrated. The conversion degree of the polarity is well controlled and reversible by trapping the opposite carriers. Basically, an electron device is switched to be a hole only device or vice versa. Finally, a hybrid layer ambipolar inverter is demonstrated in which almost no leakage of opposite carrier is found. This hybrid material has wide range of applications in planar p-n junctions and logic circuits for high-throughput manufacturing of printed electronic circuits. PMID:25801827

High Throughput Determination of Critical Human Dosing Parameters (SOT)

EPA Science Inventory

High throughput toxicokinetics (HTTK) is a rapid approach that uses in vitro data to estimate TK for hundreds of environmental chemicals. Reverse dosimetry (i.e., reverse toxicokinetics or RTK) based on HTTK data converts high throughput in vitro toxicity screening (HTS) data int...

High Throughput Determinations of Critical Dosing Parameters (IVIVE workshop)

EPA Science Inventory

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...

Optimization of high-throughput nanomaterial developmental toxicity testing in zebrafish embryos

EPA Science Inventory

Nanomaterial (NM) developmental toxicities are largely unknown. With an extensive variety of NMs available, high-throughput screening methods may be of value for initial characterization of potential hazard. We optimized a zebrafish embryo test as an in vivo high-throughput assay...

A high-throughput method for the detection of homoeologous gene deletions in hexaploid wheat

PubMed Central

2010-01-01

Background Mutational inactivation of plant genes is an essential tool in gene function studies. Plants with inactivated or deleted genes may also be exploited for crop improvement if such mutations/deletions produce a desirable agronomical and/or quality phenotype. However, the use of mutational gene inactivation/deletion has been impeded in polyploid plant species by genetic redundancy, as polyploids contain multiple copies of the same genes (homoeologous genes) encoded by each of the ancestral genomes. Similar to many other crop plants, bread wheat (Triticum aestivum L.) is polyploid; specifically allohexaploid possessing three progenitor genomes designated as 'A', 'B', and 'D'. Recently modified TILLING protocols have been developed specifically for mutation detection in wheat. Whilst extremely powerful in detecting single nucleotide changes and small deletions, these methods are not suitable for detecting whole gene deletions. Therefore, high-throughput methods for screening of candidate homoeologous gene deletions are needed for application to wheat populations generated by the use of certain mutagenic agents (e.g. heavy ion irradiation) that frequently generate whole-gene deletions. Results To facilitate the screening for specific homoeologous gene deletions in hexaploid wheat, we have developed a TaqMan qPCR-based method that allows high-throughput detection of deletions in homoeologous copies of any gene of interest, provided that sufficient polymorphism (as little as a single nucleotide difference) amongst homoeologues exists for specific probe design. We used this method to identify deletions of individual TaPFT1 homoeologues, a wheat orthologue of the disease susceptibility and flowering regulatory gene PFT1 in Arabidopsis. This method was applied to wheat nullisomic-tetrasomic lines as well as other chromosomal deletion lines to locate the TaPFT1 gene to the long arm of chromosome 5. By screening of individual DNA samples from 4500 M2 mutant wheat lines generated by heavy ion irradiation, we detected multiple mutants with deletions of each TaPFT1 homoeologue, and confirmed these deletions using a CAPS method. We have subsequently designed, optimized, and applied this method for the screening of homoeologous deletions of three additional wheat genes putatively involved in plant disease resistance. Conclusions We have developed a method for automated, high-throughput screening to identify deletions of individual homoeologues of a wheat gene. This method is also potentially applicable to other polyploidy plants. PMID:21114819

Gene expression changes leading extreme alkaline tolerance in Amur ide (Leuciscus waleckii) inhabiting soda lake

PubMed Central

2013-01-01

Background Amur ide (Leuciscus waleckii) is an economically and ecologically important cyprinid species in Northern Asia. The Dali Nor population living in the soda lake Dali Nor can adapt the extremely high alkalinity, providing us a valuable material to understand the adaptation mechanism against extreme environmental stress in teleost. Results In this study, we generated high-throughput RNA-Seq data from three tissues gill, liver and kidney of L. waleckii living in the soda lake Dali Nor and the fresh water lake Ganggeng Nor, then performed parallel comparisons of three tissues. Our results showed that out of assembled 64,603 transcript contigs, 28,391 contigs had been assigned with a known function, corresponding to 20,371 unique protein accessions. We found 477, 2,761 and 3,376 differentially expressed genes (DEGs) in the gill, kidney, and liver, respectively, of Dali Nor population compared to Ganggeng Nor population with FDR ≤ 0.01and fold-change ≥ 2. Further analysis revealed that well-known functional categories of genes and signaling pathway, which are associated with stress response and extreme environment adaptation, have been significantly enriched, including the functional categories of “response to stimulus”, “transferase activity”, “transporter activity” and “oxidoreductase activity”, and signaling pathways of “mTOR signaling”, “EIF2 signaling”, “superpathway of cholesterol biosynthesis”. We also identified significantly DEGs encoding important modulators on stress adaptation and tolerance, including carbonic anhydrases, heat shock proteins, superoxide dismutase, glutathione S-transferases, aminopeptidase N, and aminotransferases. Conclusions Overall, this study demonstrated that transcriptome changes in L. waleckii played a role in adaptation to complicated environmental stress in the highly alkalized Dali Nor lake. The results set a foundation for further analyses on alkaline-responsive candidate genes, which help us understand teleost adaptation under extreme environmental stress and ultimately benefit future breeding for alkaline-tolerant fish strains. PMID:24094069

CORDIC-based digital signal processing (DSP) element for adaptive signal processing

NASA Astrophysics Data System (ADS)

Bolstad, Gregory D.; Neeld, Kenneth B.

1995-04-01

The High Performance Adaptive Weight Computation (HAWC) processing element is a CORDIC based application specific DSP element that, when connected in a linear array, can perform extremely high throughput (100s of GFLOPS) matrix arithmetic operations on linear systems of equations in real time. In particular, it very efficiently performs the numerically intense computation of optimal least squares solutions for large, over-determined linear systems. Most techniques for computing solutions to these types of problems have used either a hard-wired, non-programmable systolic array approach, or more commonly, programmable DSP or microprocessor approaches. The custom logic methods can be efficient, but are generally inflexible. Approaches using multiple programmable generic DSP devices are very flexible, but suffer from poor efficiency and high computation latencies, primarily due to the large number of DSP devices that must be utilized to achieve the necessary arithmetic throughput. The HAWC processor is implemented as a highly optimized systolic array, yet retains some of the flexibility of a programmable data-flow system, allowing efficient implementation of algorithm variations. This provides flexible matrix processing capabilities that are one to three orders of magnitude less expensive and more dense than the current state of the art, and more importantly, allows a realizable solution to matrix processing problems that were previously considered impractical to physically implement. HAWC has direct applications in RADAR, SONAR, communications, and image processing, as well as in many other types of systems.

Spatial tuning of acoustofluidic pressure nodes by altering net sonic velocity enables high-throughput, efficient cell sorting

DOE PAGES

Jung, Seung-Yong; Notton, Timothy; Fong, Erika; ...

2015-01-07

Particle sorting using acoustofluidics has enormous potential but widespread adoption has been limited by complex device designs and low throughput. Here, we report high-throughput separation of particles and T lymphocytes (600 μL min -1) by altering the net sonic velocity to reposition acoustic pressure nodes in a simple two-channel device. Finally, the approach is generalizable to other microfluidic platforms for rapid, high-throughput analysis.

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

PubMed Central

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

2017-01-01

Abstract 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. PMID:28458737

A Bright Future for Evolutionary Methods in Drug Design.

PubMed

Le, Tu C; Winkler, David A

2015-08-01

Most medicinal chemists understand that chemical space is extremely large, essentially infinite. Although high-throughput experimental methods allow exploration of drug-like space more rapidly, they are still insufficient to fully exploit the opportunities that such large chemical space offers. Evolutionary methods can synergistically blend automated synthesis and characterization methods with computational design to identify promising regions of chemical space more efficiently. We describe how evolutionary methods are implemented, and provide examples of published drug development research in which these methods have generated molecules with increased efficacy. We anticipate that evolutionary methods will play an important role in future drug discovery. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Application of laser-wakefield-based x-ray source to global food security issues

NASA Astrophysics Data System (ADS)

Kieffer, J. C.; Fourmaux, S.; Hallin, E.; Arnison, P.; Brereton, N.; Pitre, F.; Dixon, M.; Tran, N.

2017-05-01

We present the development of a high throughput phase contrast screening system based on LWFA Xray sources for plant imaging. We upgraded the INRS laser-betatron beam line and we illustrate its imaging potential through the innovative development of new tools for addressing issues relevant to global food security. This initiative, led by the Global Institute of Food Security (GIFS) at the U of Saskatchewan, aims to elucidate that part of the function that maps environmental inputs onto specific plant phenotypes. The prospect of correlating phenotypic expression with adaptation to environmental stresses will provide researchers with a new tool to assess breeding programs for crops meant to thrive under the climate extremes.

Acceleration of short and long DNA read mapping without loss of accuracy using suffix array.

PubMed

Tárraga, Joaquín; Arnau, Vicente; Martínez, Héctor; Moreno, Raul; Cazorla, Diego; Salavert-Torres, José; Blanquer-Espert, Ignacio; Dopazo, Joaquín; Medina, Ignacio

2014-12-01

HPG Aligner applies suffix arrays for DNA read mapping. This implementation produces a highly sensitive and extremely fast mapping of DNA reads that scales up almost linearly with read length. The approach presented here is faster (over 20× for long reads) and more sensitive (over 98% in a wide range of read lengths) than the current state-of-the-art mappers. HPG Aligner is not only an optimal alternative for current sequencers but also the only solution available to cope with longer reads and growing throughputs produced by forthcoming sequencing technologies. https://github.com/opencb/hpg-aligner. © The Author 2014. Published by Oxford University Press.

Genomic phenotyping by barcode sequencing broadly distinguishes between alkylating agents, oxidizing agents, and non-genotoxic agents, and reveals a role for aromatic amino acids in cellular recovery after quinone exposure.

PubMed

Svensson, J Peter; Quirós Pesudo, Laia; McRee, Siobhan K; Adeleye, Yeyejide; Carmichael, Paul; Samson, Leona D

2013-01-01

Toxicity screening of compounds provides a means to identify compounds harmful for human health and the environment. Here, we further develop the technique of genomic phenotyping to improve throughput while maintaining specificity. We exposed cells to eight different compounds that rely on different modes of action: four genotoxic alkylating (methyl methanesulfonate (MMS), N-Methyl-N-nitrosourea (MNU), N,N'-bis(2-chloroethyl)-N-nitroso-urea (BCNU), N-ethylnitrosourea (ENU)), two oxidizing (2-methylnaphthalene-1,4-dione (menadione, MEN), benzene-1,4-diol (hydroquinone, HYQ)), and two non-genotoxic (methyl carbamate (MC) and dimethyl sulfoxide (DMSO)) compounds. A library of S. cerevisiae 4,852 deletion strains, each identifiable by a unique genetic 'barcode', were grown in competition; at different time points the ratio between the strains was assessed by quantitative high throughput 'barcode' sequencing. The method was validated by comparison to previous genomic phenotyping studies and 90% of the strains identified as MMS-sensitive here were also identified as MMS-sensitive in a much lower throughput solid agar screen. The data provide profiles of proteins and pathways needed for recovery after both genotoxic and non-genotoxic compounds. In addition, a novel role for aromatic amino acids in the recovery after treatment with oxidizing agents was suggested. The role of aromatic acids was further validated; the quinone subgroup of oxidizing agents were extremely toxic in cells where tryptophan biosynthesis was compromised.

Genomic Phenotyping by Barcode Sequencing Broadly Distinguishes between Alkylating Agents, Oxidizing Agents, and Non-Genotoxic Agents, and Reveals a Role for Aromatic Amino Acids in Cellular Recovery after Quinone Exposure

PubMed Central

Svensson, J. Peter; Quirós Pesudo, Laia; McRee, Siobhan K.; Adeleye, Yeyejide; Carmichael, Paul; Samson, Leona D.

2013-01-01

Toxicity screening of compounds provides a means to identify compounds harmful for human health and the environment. Here, we further develop the technique of genomic phenotyping to improve throughput while maintaining specificity. We exposed cells to eight different compounds that rely on different modes of action: four genotoxic alkylating (methyl methanesulfonate (MMS), N-Methyl-N-nitrosourea (MNU), N,N′-bis(2-chloroethyl)-N-nitroso-urea (BCNU), N-ethylnitrosourea (ENU)), two oxidizing (2-methylnaphthalene-1,4-dione (menadione, MEN), benzene-1,4-diol (hydroquinone, HYQ)), and two non-genotoxic (methyl carbamate (MC) and dimethyl sulfoxide (DMSO)) compounds. A library of S. cerevisiae 4,852 deletion strains, each identifiable by a unique genetic ‘barcode’, were grown in competition; at different time points the ratio between the strains was assessed by quantitative high throughput ‘barcode’ sequencing. The method was validated by comparison to previous genomic phenotyping studies and 90% of the strains identified as MMS-sensitive here were also identified as MMS-sensitive in a much lower throughput solid agar screen. The data provide profiles of proteins and pathways needed for recovery after both genotoxic and non-genotoxic compounds. In addition, a novel role for aromatic amino acids in the recovery after treatment with oxidizing agents was suggested. The role of aromatic acids was further validated; the quinone subgroup of oxidizing agents were extremely toxic in cells where tryptophan biosynthesis was compromised. PMID:24040048

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

EPA Pesticide Factsheets

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

Evaluating High Throughput Toxicokinetics and Toxicodynamics for IVIVE (WC10)

EPA Science Inventory

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...

High-throughput RAD-SNP genotyping for characterization of sugar beet genotypes

USDA-ARS?s Scientific Manuscript database

High-throughput SNP genotyping provides a rapid way of developing resourceful set of markers for delineating the genetic architecture and for effective species discrimination. In the presented research, we demonstrate a set of 192 SNPs for effective genotyping in sugar beet using high-throughput mar...

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

EPA Science Inventory

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...

A quantitative literature-curated gold standard for kinase-substrate pairs

PubMed Central

2011-01-01

We describe the Yeast Kinase Interaction Database (KID, http://www.moseslab.csb.utoronto.ca/KID/), which contains high- and low-throughput data relevant to phosphorylation events. KID includes 6,225 low-throughput and 21,990 high-throughput interactions, from greater than 35,000 experiments. By quantitatively integrating these data, we identified 517 high-confidence kinase-substrate pairs that we consider a gold standard. We show that this gold standard can be used to assess published high-throughput datasets, suggesting that it will enable similar rigorous assessments in the future. PMID:21492431

High-Throughput Industrial Coatings Research at The Dow Chemical Company.

PubMed

Kuo, Tzu-Chi; Malvadkar, Niranjan A; Drumright, Ray; Cesaretti, Richard; Bishop, Matthew T

2016-09-12

At The Dow Chemical Company, high-throughput research is an active area for developing new industrial coatings products. Using the principles of automation (i.e., using robotic instruments), parallel processing (i.e., prepare, process, and evaluate samples in parallel), and miniaturization (i.e., reduce sample size), high-throughput tools for synthesizing, formulating, and applying coating compositions have been developed at Dow. In addition, high-throughput workflows for measuring various coating properties, such as cure speed, hardness development, scratch resistance, impact toughness, resin compatibility, pot-life, surface defects, among others have also been developed in-house. These workflows correlate well with the traditional coatings tests, but they do not necessarily mimic those tests. The use of such high-throughput workflows in combination with smart experimental designs allows accelerated discovery and commercialization.

Outlook for Development of High-throughput Cryopreservation for Small-bodied Biomedical Model Fishes★

PubMed Central

Tiersch, Terrence R.; Yang, Huiping; Hu, E.

2011-01-01

With the development of genomic research technologies, comparative genome studies among vertebrate species are becoming commonplace for human biomedical research. Fish offer unlimited versatility for biomedical research. Extensive studies are done using these fish models, yielding tens of thousands of specific strains and lines, and the number is increasing every day. Thus, high-throughput sperm cryopreservation is urgently needed to preserve these genetic resources. Although high-throughput processing has been widely applied for sperm cryopreservation in livestock for decades, application in biomedical model fishes is still in the concept-development stage because of the limited sample volumes and the biological characteristics of fish sperm. High-throughput processing in livestock was developed based on advances made in the laboratory and was scaled up for increased processing speed, capability for mass production, and uniformity and quality assurance. Cryopreserved germplasm combined with high-throughput processing constitutes an independent industry encompassing animal breeding, preservation of genetic diversity, and medical research. Currently, there is no specifically engineered system available for high-throughput of cryopreserved germplasm for aquatic species. This review is to discuss the concepts and needs for high-throughput technology for model fishes, propose approaches for technical development, and overview future directions of this approach. PMID:21440666

High-throughput measurements of biochemical responses using the plate::vision multimode 96 minilens array reader.

PubMed

Huang, Kuo-Sen; Mark, David; Gandenberger, Frank Ulrich

2006-01-01

The plate::vision is a high-throughput multimode reader capable of reading absorbance, fluorescence, fluorescence polarization, time-resolved fluorescence, and luminescence. Its performance has been shown to be quite comparable with other readers. When the reader is integrated into the plate::explorer, an ultrahigh-throughput screening system with event-driven software and parallel plate-handling devices, it becomes possible to run complicated assays with kinetic readouts in high-density microtiter plate formats for high-throughput screening. For the past 5 years, we have used the plate::vision and the plate::explorer to run screens and have generated more than 30 million data points. Their throughput, performance, and robustness have speeded up our drug discovery process greatly.

Reflective optical imaging system with balanced distortion

DOEpatents

Chapman, Henry N.; Hudyma, Russell M.; Shafer, David R.; Sweeney, Donald W.

1999-01-01

An optical system compatible with short wavelength (extreme ultraviolet) An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements comprise, in order from object to image, convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention enables the use of larger slit dimensions associated with ring field scanning optics, improves wafer throughput and allows higher semiconductor device density. The inventive optical system is characterized by reduced dynamic distortion because the static distortion is balanced across the slit width.

Adaptive and reliably acknowledged FSO communications

NASA Astrophysics Data System (ADS)

Fitz, Michael P.; Halford, Thomas R.; Kose, Cenk; Cromwell, Jonathan; Gordon, Steven

2015-05-01

Atmospheric turbulence causes the receive signal intensity on free space optical (FSO) communication links to vary over time. Scintillation fades can stymie connectivity for milliseconds at a time. To approach the information-theoretic limits of communication in such time-varying channels, it necessary to either code across extremely long blocks of data - thereby inducing unacceptable delays - or to vary the code rate according to the instantaneous channel conditions. We describe the design, laboratory testing, and over-the-air testing of an FSO modem that employs a protocol with adaptive coded modulation (ACM) and hybrid automatic repeat request. For links with fixed throughput, this protocol provides a 10dB reduction in the required received signal-to-noise ratio (SNR); for links with fixed range, this protocol provides the greater than a 3x increase in throughput. Independent U.S. Government tests demonstrate that our protocol effectively adapts the code rate to match the instantaneous channel conditions. The modem is able to provide throughputs in excess of 850 Mbps on links with ranges greater than 15 kilometers.

The Extreme Ultraviolet Normal Incidence Spectrograph (EUNIS)

NASA Technical Reports Server (NTRS)

Oegerle, William (Technical Monitor); Rabin, D.; Davila, J.; Thomas, R. J.; Engler, C.; Irish, S.; Keski-Kuha, R.; Novello, J.; Nowak, M.; Payne, L.;

TCP Throughput Profiles Using Measurements over Dedicated Connections

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rao, Nageswara S.; Liu, Qiang; Sen, Satyabrata

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, inmore » 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.« less

Enhancing high throughput toxicology - development of putative adverse outcome pathways linking US EPA ToxCast screening targets to relevant apical hazards.

EPA Science Inventory

High throughput toxicology programs, such as ToxCast and Tox21, have provided biological effects data for thousands of chemicals at multiple concentrations. Compared to traditional, whole-organism approaches, high throughput assays are rapid and cost-effective, yet they generall...

Evaluation of High-Throughput Chemical Exposure Models via Analysis of Matched Environmental and Biological Media Measurements

EPA Science Inventory

The U.S. EPA, under its ExpoCast program, is developing high-throughput near-field modeling methods to estimate human chemical exposure and to provide real-world context to high-throughput screening (HTS) hazard data. These novel modeling methods include reverse methods to infer ...

The development of a general purpose ARM-based processing unit for the ATLAS TileCal sROD

NASA Astrophysics Data System (ADS)

Cox, M. A.; Reed, R.; Mellado, B.

2015-01-01

After Phase-II upgrades in 2022, the data output from the LHC ATLAS Tile Calorimeter will increase significantly. ARM processors are common in mobile devices due to their low cost, low energy consumption and high performance. It is proposed that a cost-effective, high data throughput Processing Unit (PU) can be developed by using several consumer ARM processors in a cluster configuration to allow aggregated processing performance and data throughput while maintaining minimal software design difficulty for the end-user. This PU could be used for a variety of high-level functions on the high-throughput raw data such as spectral analysis and histograms to detect possible issues in the detector at a low level. High-throughput I/O interfaces are not typical in consumer ARM System on Chips but high data throughput capabilities are feasible via the novel use of PCI-Express as the I/O interface to the ARM processors. An overview of the PU is given and the results for performance and throughput testing of four different ARM Cortex System on Chips are presented.

[Current applications of high-throughput DNA sequencing technology in antibody drug research].

PubMed

Yu, Xin; Liu, Qi-Gang; Wang, Ming-Rong

2012-03-01

Since the publication of a high-throughput DNA sequencing technology based on PCR reaction was carried out in oil emulsions in 2005, high-throughput DNA sequencing platforms have been evolved to a robust technology in sequencing genomes and diverse DNA libraries. Antibody libraries with vast numbers of members currently serve as a foundation of discovering novel antibody drugs, and high-throughput DNA sequencing technology makes it possible to rapidly identify functional antibody variants with desired properties. Herein we present a review of current applications of high-throughput DNA sequencing technology in the analysis of antibody library diversity, sequencing of CDR3 regions, identification of potent antibodies based on sequence frequency, discovery of functional genes, and combination with various display technologies, so as to provide an alternative approach of discovery and development of antibody drugs.

Bayesian hierarchical modeling for subject-level response classification in peptide microarray immunoassays

PubMed Central

Imholte, Gregory; Gottardo, Raphael

2017-01-01

Summary The peptide microarray immunoassay simultaneously screens sample serum against thousands of peptides, determining the presence of antibodies bound to array probes. Peptide microarrays tiling immunogenic regions of pathogens (e.g. envelope proteins of a virus) are an important high throughput tool for querying and mapping antibody binding. Because of the assay’s many steps, from probe synthesis to incubation, peptide microarray data can be noisy with extreme outliers. In addition, subjects may produce different antibody profiles in response to an identical vaccine stimulus or infection, due to variability among subjects’ immune systems. We present a robust Bayesian hierarchical model for peptide microarray experiments, pepBayes, to estimate the probability of antibody response for each subject/peptide combination. Heavy-tailed error distributions accommodate outliers and extreme responses, and tailored random effect terms automatically incorporate technical effects prevalent in the assay. We apply our model to two vaccine trial datasets to demonstrate model performance. Our approach enjoys high sensitivity and specificity when detecting vaccine induced antibody responses. A simulation study shows an adaptive thresholding classification method has appropriate false discovery rate control with high sensitivity, and receiver operating characteristics generated on vaccine trial data suggest that pepBayes clearly separates responses from non-responses. PMID:27061097

Lessons from high-throughput protein crystallization screening: 10 years of practical experience

PubMed Central

JR, Luft; EH, Snell; GT, DeTitta

2011-01-01

Introduction X-ray crystallography provides the majority of our structural biological knowledge at a molecular level and in terms of pharmaceutical design is a valuable tool to accelerate discovery. It is the premier technique in the field, but its usefulness is significantly limited by the need to grow well-diffracting crystals. It is for this reason that high-throughput crystallization has become a key technology that has matured over the past 10 years through the field of structural genomics. Areas covered The authors describe their experiences in high-throughput crystallization screening in the context of structural genomics and the general biomedical community. They focus on the lessons learnt from the operation of a high-throughput crystallization screening laboratory, which to date has screened over 12,500 biological macromolecules. They also describe the approaches taken to maximize the success while minimizing the effort. Through this, the authors hope that the reader will gain an insight into the efficient design of a laboratory and protocols to accomplish high-throughput crystallization on a single-, multiuser-laboratory or industrial scale. Expert Opinion High-throughput crystallization screening is readily available but, despite the power of the crystallographic technique, getting crystals is still not a solved problem. High-throughput approaches can help when used skillfully; however, they still require human input in the detailed analysis and interpretation of results to be more successful. PMID:22646073

High-throughput screening based on label-free detection of small molecule microarrays

NASA Astrophysics Data System (ADS)

Zhu, Chenggang; Fei, Yiyan; Zhu, Xiangdong

2017-02-01

Based on small-molecule microarrays (SMMs) and oblique-incidence reflectivity difference (OI-RD) scanner, we have developed a novel high-throughput drug preliminary screening platform based on label-free monitoring of direct interactions between target proteins and immobilized small molecules. The screening platform is especially attractive for screening compounds against targets of unknown function and/or structure that are not compatible with functional assay development. In this screening platform, OI-RD scanner serves as a label-free detection instrument which is able to monitor about 15,000 biomolecular interactions in a single experiment without the need to label any biomolecule. Besides, SMMs serves as a novel format for high-throughput screening by immobilization of tens of thousands of different compounds on a single phenyl-isocyanate functionalized glass slide. Based on the high-throughput screening platform, we sequentially screened five target proteins (purified target proteins or cell lysate containing target protein) in high-throughput and label-free mode. We found hits for respective target protein and the inhibition effects for some hits were confirmed by following functional assays. Compared to traditional high-throughput screening assay, the novel high-throughput screening platform has many advantages, including minimal sample consumption, minimal distortion of interactions through label-free detection, multi-target screening analysis, which has a great potential to be a complementary screening platform in the field of drug discovery.

High-throughput analysis of yeast replicative aging using a microfluidic system

PubMed Central

Jo, Myeong Chan; Liu, Wei; Gu, Liang; Dang, Weiwei; Qin, Lidong

2015-01-01

Saccharomyces cerevisiae has been an important model for studying the molecular mechanisms of aging in eukaryotic cells. However, the laborious and low-throughput methods of current yeast replicative lifespan assays limit their usefulness as a broad genetic screening platform for research on aging. We address this limitation by developing an efficient, high-throughput microfluidic single-cell analysis chip in combination with high-resolution time-lapse microscopy. This innovative design enables, to our knowledge for the first time, the determination of the yeast replicative lifespan in a high-throughput manner. Morphological and phenotypical changes during aging can also be monitored automatically with a much higher throughput than previous microfluidic designs. We demonstrate highly efficient trapping and retention of mother cells, determination of the replicative lifespan, and tracking of yeast cells throughout their entire lifespan. Using the high-resolution and large-scale data generated from the high-throughput yeast aging analysis (HYAA) chips, we investigated particular longevity-related changes in cell morphology and characteristics, including critical cell size, terminal morphology, and protein subcellular localization. In addition, because of the significantly improved retention rate of yeast mother cell, the HYAA-Chip was capable of demonstrating replicative lifespan extension by calorie restriction. PMID:26170317

A trophic model of fringing coral reefs in Nanwan Bay, southern Taiwan suggests overfishing.

PubMed

Liu, Pi-Jen; Shao, Kwang-Tsao; Jan, Rong-Quen; Fan, Tung-Yung; Wong, Saou-Lien; Hwang, Jiang-Shiou; Chen, Jen-Ping; Chen, Chung-Chi; Lin, Hsing-Juh

2009-09-01

Several coral reefs of Nanwan Bay, Taiwan have recently undergone shifts to macroalgal or sea anemone dominance. Thus, a mass-balance trophic model was constructed to analyze the structure and functioning of the food web. The fringing reef model was comprised of 18 compartments, with the highest trophic level of 3.45 for piscivorous fish. Comparative analyses with other reef models demonstrated that Nanwan Bay was similar to reefs with high fishery catches. While coral biomass was not lower, fish biomass was lower than those of reefs with high catches. Consequently, the sums of consumption and respiratory flows and total system throughput were also decreased. The Nanwan Bay model potentially suggests an overfished status in which the mean trophic level of the catch, matter cycling, and trophic transfer efficiency are extremely reduced.

The novel top-coat material for RLS trade-off reduction in EUVL

NASA Astrophysics Data System (ADS)

Onishi, Ryuji; Sakamoto, Rikimaru; Fujitani, Noriaki; Endo, Takafumi; Ho, Bang-ching

2012-03-01

For the next generation lithography (NGL), several technologies have been proposed to achieve the 22nm-node devices and beyond. Extreme ultraviolet (EUV) lithography is one of the candidates for the next generation lithography. In EUV light source development, low power is one of the critical issue because of the low throughput, and another issue is Out of Band (OoB) light existing in EUV light. OoB is concerned to be the cause of deterioration for the lithography performance. In order to avoid this critical issue, we focused on development of the resist top coat material with OoB absorption property as Out of Band Protection Layer (OBPL). We designed this material having high absorbance around 240nm wavelength and high transmittance for EUV light. And this material aimed to improve sensitivity, resolution and LWR performance.

Reversible conversion of dominant polarity in ambipolar polymer/graphene oxide hybrids

DOE PAGES

Zhou, Ye; Han, Su -Ting; Sonar, Prashant; ...

2015-03-24

The possibility to selectively modulate the charge carrier transport in semiconducting materials is extremely challenging for the development of high performance and low-power consuming logic circuits. Systematical control over the polarity (electrons and holes) in transistor based on solution processed layer by layer polymer/graphene oxide hybrid system has been demonstrated. The conversion degree of the polarity is well controlled and reversible by trapping the opposite carriers. Basically, an electron device is switched to be a hole only device or vice versa. Finally, a hybrid layer ambipolar inverter is demonstrated in which almost no leakage of opposite carrier is found. Wemore » conclude that this hybrid material has wide range of applications in planar p-n junctions and logic circuits for high-throughput manufacturing of printed electronic circuits.« less

Demonstration of lithography patterns using reflective e-beam direct write

NASA Astrophysics Data System (ADS)

Freed, Regina; Sun, Jeff; Brodie, Alan; Petric, Paul; McCord, Mark; Ronse, Kurt; Haspeslagh, Luc; Vereecke, Bart

2011-04-01

Traditionally, e-beam direct write lithography has been too slow for most lithography applications. E-beam direct write lithography has been used for mask writing rather than wafer processing since the maximum blur requirements limit column beam current - which drives e-beam throughput. To print small features and a fine pitch with an e-beam tool requires a sacrifice in processing time unless one significantly increases the total number of beams on a single writing tool. Because of the uncertainty with regards to the optical lithography roadmap beyond the 22 nm technology node, the semiconductor equipment industry is in the process of designing and testing e-beam lithography tools with the potential for high volume wafer processing. For this work, we report on the development and current status of a new maskless, direct write e-beam lithography tool which has the potential for high volume lithography at and below the 22 nm technology node. A Reflective Electron Beam Lithography (REBL) tool is being developed for high throughput electron beam direct write maskless lithography. The system is targeting critical patterning steps at the 22 nm node and beyond at a capital cost equivalent to conventional lithography. Reflective Electron Beam Lithography incorporates a number of novel technologies to generate and expose lithographic patterns with a throughput and footprint comparable to current 193 nm immersion lithography systems. A patented, reflective electron optic or Digital Pattern Generator (DPG) enables the unique approach. The Digital Pattern Generator is a CMOS ASIC chip with an array of small, independently controllable lens elements (lenslets), which act as an array of electron mirrors. In this way, the REBL system is capable of generating the pattern to be written using massively parallel exposure by ~1 million beams at extremely high data rates (~ 1Tbps). A rotary stage concept using a rotating platen carrying multiple wafers optimizes the writing strategy of the DPG to achieve the capability of high throughput for sparse pattern wafer levels. The lens elements on the DPG are fabricated at IMEC (Leuven, Belgium) under IMEC's CMORE program. The CMOS fabricated DPG contains ~ 1,000,000 lens elements, allowing for 1,000,000 individually controllable beamlets. A single lens element consists of 5 electrodes, each of which can be set at controlled voltage levels to either absorb or reflect the electron beam. A system using a linear movable stage and the DPG integrated into the electron optics module was used to expose patterns on device representative wafers. Results of these exposure tests are discussed.

Measuring molecular biomarkers in epidemiologic studies: laboratory techniques and biospecimen considerations.

PubMed

Erickson, Heidi S

2012-09-28

The future of personalized medicine depends on the ability to efficiently and rapidly elucidate a reliable set of disease-specific molecular biomarkers. High-throughput molecular biomarker analysis methods have been developed to identify disease risk, diagnostic, prognostic, and therapeutic targets in human clinical samples. Currently, high throughput screening allows us to analyze thousands of markers from one sample or one marker from thousands of samples and will eventually allow us to analyze thousands of markers from thousands of samples. Unfortunately, the inherent nature of current high throughput methodologies, clinical specimens, and cost of analysis is often prohibitive for extensive high throughput biomarker analysis. This review summarizes the current state of high throughput biomarker screening of clinical specimens applicable to genetic epidemiology and longitudinal population-based studies with a focus on considerations related to biospecimens, laboratory techniques, and sample pooling. Copyright © 2012 John Wiley & Sons, Ltd.

The Advanced Telescope for High Energy Astrophysics

NASA Astrophysics Data System (ADS)

Guainazzi, Matteo

2017-08-01

Athena (the Advanced Telescope for High Energy Astrophysics) is a next generation X-ray observatory currently under study by ESA for launch in 2028. Athena is designed to address the Hot and Energetic Universe science theme, which addresses two key questions: 1) How did ordinary matter evolve into the large scale structures we see today? 2) How do black holes grow and shape the Universe. To address these topics Athena employs an innovative X-ray telescope based on Silicon Pore Optics technology to deliver extremely light weight and high throughput, while retaining excellent angular resolution. The mirror can be adjusted to focus onto one of two focal place instruments: the X-ray Integral Field Unit (X-IFU) which provides spatially-resolved, high resolution spectroscopy, and the Wide Field Imager (WFI) which provides spectral imaging over a large field of view, as well as high time resolution and count rate tolerance. Athena is currently in Phase A and the study status will be reviewed, along with the scientific motivations behind the mission.

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

Code of Federal Regulations, 2010 CFR

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...

A Call for Nominations of Quantitative High-Throughput ...

EPA Pesticide Factsheets

The National Research Council of the United States National Academies of Science has recently released a document outlining a long-range vision and strategy for transforming toxicity testing from largely whole animal-based testing to one based on in vitro assays. “Toxicity Testing in the 21st Century: A Vision and a Strategy” advises a focus on relevant human toxicity pathway assays. Toxicity pathways are defined in the document as “Cellular response pathways that, when sufficiently perturbed, are expected to result in adverse health effects”. Results of such pathway screens would serve as a filter to drive selection of more specific, targeted testing that will complement and validate the pathway assays. In response to this report, the US EPA has partnered with two NIH organizations, the National Toxicology Program and the NIH Chemical Genomics Center (NCGC), in a program named Tox21. A major goal of this collaboration is to screen chemical libraries consisting of known toxicants, chemicals of environmental and occupational exposure concern, and human pharmaceuticals in cell-based pathway assays. Currently, approximately 3000 compounds (increasing to 9000 by the end of 2009) are being validated and screened in quantitative high-throughput (qHTS) format at the NCGC producing extensive concentration-response data for a diverse set of potential toxicity pathways. The Tox21 collaboration is extremely interested in accessing additional toxicity pathway assa

Application of a high-throughput relative chemical stability assay to screen therapeutic protein formulations by assessment of conformational stability and correlation to aggregation propensity.

PubMed

Rizzo, Joseph M; Shi, Shuai; Li, Yunsong; Semple, Andrew; Esposito, Jessica J; Yu, Shenjiang; Richardson, Daisy; Antochshuk, Valentyn; Shameem, Mohammed

2015-05-01

In this study, an automated high-throughput relative chemical stability (RCS) assay was developed in which various therapeutic proteins were assessed to determine stability based on the resistance to denaturation post introduction to a chaotrope titration. Detection mechanisms of both intrinsic fluorescence and near UV circular dichroism (near-UV CD) are demonstrated. Assay robustness was investigated by comparing multiple independent assays and achieving r(2) values >0.95 for curve overlays. The complete reversibility of the assay was demonstrated by intrinsic fluorescence, near-UV CD, and biologic potency. To highlight the method utility, we compared the RCS assay with differential scanning calorimetry and dynamic scanning fluorimetry methodologies. Utilizing C1/2 values obtained from the RCS assay, formulation rank-ordering of 12 different mAb formulations was performed. The prediction of long-term stability on protein aggregation is obtained by demonstrating a good correlation with an r(2) of 0.83 between RCS and empirical aggregation propensity data. RCS promises to be an extremely useful tool to aid in candidate formulation development efforts based on the complete reversibility of the method to allow for multiple assessments without protein loss and the strong correlation between the C1/2 data obtained and accelerated stability under stressed conditions. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

Accelerating the design of solar thermal fuel materials through high throughput simulations.

PubMed

Liu, Yun; Grossman, Jeffrey C

2014-12-10

Solar thermal fuels (STF) store the energy of sunlight, which can then be released later in the form of heat, offering an emission-free and renewable solution for both solar energy conversion and storage. However, this approach is currently limited by the lack of low-cost materials with high energy density and high stability. In this Letter, we present an ab initio high-throughput computational approach to accelerate the design process and allow for searches over a broad class of materials. The high-throughput screening platform we have developed can run through large numbers of molecules composed of earth-abundant elements and identifies possible metastable structures of a given material. Corresponding isomerization enthalpies associated with the metastable structures are then computed. Using this high-throughput simulation approach, we have discovered molecular structures with high isomerization enthalpies that have the potential to be new candidates for high-energy density STF. We have also discovered physical principles to guide further STF materials design through structural analysis. More broadly, our results illustrate the potential of using high-throughput ab initio simulations to design materials that undergo targeted structural transitions.

The combination of gas-phase fluorophore technology and automation to enable high-throughput analysis of plant respiration.

PubMed

Scafaro, Andrew P; Negrini, A Clarissa A; O'Leary, Brendan; Rashid, F Azzahra Ahmad; Hayes, Lucy; Fan, Yuzhen; Zhang, You; Chochois, Vincent; Badger, Murray R; Millar, A Harvey; Atkin, Owen K

2017-01-01

Mitochondrial respiration in the dark ( R dark ) is a critical plant physiological process, and hence a reliable, efficient and high-throughput method of measuring variation in rates of R dark is essential for agronomic and ecological studies. However, currently methods used to measure R dark in plant tissues are typically low throughput. We assessed a high-throughput automated fluorophore system of detecting multiple O 2 consumption rates. The fluorophore technique was compared with O 2 -electrodes, infrared gas analysers (IRGA), and membrane inlet mass spectrometry, to determine accuracy and speed of detecting respiratory fluxes. The high-throughput fluorophore system provided stable measurements of R dark in detached leaf and root tissues over many hours. High-throughput potential was evident in that the fluorophore system was 10 to 26-fold faster per sample measurement than other conventional methods. The versatility of the technique was evident in its enabling: (1) rapid screening of R dark in 138 genotypes of wheat; and, (2) quantification of rarely-assessed whole-plant R dark through dissection and simultaneous measurements of above- and below-ground organs. Variation in absolute R dark was observed between techniques, likely due to variation in sample conditions (i.e. liquid vs. gas-phase, open vs. closed systems), indicating that comparisons between studies using different measuring apparatus may not be feasible. However, the high-throughput protocol we present provided similar values of R dark to the most commonly used IRGA instrument currently employed by plant scientists. Together with the greater than tenfold increase in sample processing speed, we conclude that the high-throughput protocol enables reliable, stable and reproducible measurements of R dark on multiple samples simultaneously, irrespective of plant or tissue type.

Simultaneous Measurements of Auto-Immune and Infectious Disease Specific Antibodies Using a High Throughput Multiplexing Tool

PubMed Central

Asati, Atul; Kachurina, Olga; Kachurin, Anatoly

2012-01-01

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. PMID:22952605

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

NASA Astrophysics Data System (ADS)

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.

Genetics of Hearing Loss – Syndromic

PubMed Central

Koffler, Tal; Ushakov, Kathy; Avraham, Karen B.

2015-01-01

Synopsis Hearing loss (HL) is one of the most common birth defects in developed countries and is a diverse pathology with different classifications. One of these is based on the association with other clinical features, defined as syndromic hearing loss (SHL). Determining the etiology of the HL in these patients is extremely beneficial as it enables a personalized approach to caring for the individual. Early screening can further aid in optimal rehabilitation for a child’s development and growth. Pathogenic variants in forty-five genes, encoding proteins functioning as ion channels, transcription factors, molecular motors and more, are known to lead to eleven forms of SHL. The development of high-throughput sequencing technology is facilitating rapid and low-cost diagnostics for patients with SHL. PMID:26443487

High-throughput microfluidic device for rare cell isolation

NASA Astrophysics Data System (ADS)

Yang, Daniel; Leong, Serena; Lei, Andy; Sohn, Lydia L.

2015-06-01

Enumerating and analyzing circulating tumor cells (CTCs)—cells that have been shed from primary solid tumors—can potentially be used to determine patient prognosis and track the progression of disease. There is a great challenge to create an effective platform that can isolate these cells, as they are extremely rare: only 1-10 CTCs are present in a 7.5mL of a cancer patient's peripheral blood. We have developed a novel microfluidic system that can isolate CTC populations label free. Our system consists of a multistage separator that employs inertial migration to sort cells based on size. We demonstrate the feasibility of our device by sorting colloids that are comparable in size to red blood cells (RBCs) and CTCs.

High-Throughput Microfluidic Device for Rare Cell Isolation.

PubMed

Yang, Daniel; Leong, Serena; Lei, Andy; Sohn, Lydia L

2015-05-04

Enumerating and analyzing circulating tumor cells (CTCs)-cells that have been shed from primary solid tumors-can potentially be used to determine patient prognosis and track the progression of disease. There is a great challenge to create an effective platform that can isolate these cells, as they are extremely rare: only 1-10 CTCs are present in a 7.5mL of a cancer patient's peripheral blood. We have developed a novel microfluidic system that can isolate CTC populations label free. Our system consists of a multistage separator that employs inertial migration to sort cells based on size. We demonstrate the feasibility of our device by sorting colloids that are comparable in size to red blood cells (RBCs) and CTCs.

An extreme ultraviolet telescope with no soft X-ray response

NASA Technical Reports Server (NTRS)

Finley, David S.; Jelinsky, Patrick; Bowyer, Stuart; Malina, Roger F.

1986-01-01

While EUV grazing incidence telescopes of conventional design exhibit a substantial X-ray response as well as an extreme UV response, and existing bandpass filters for the transmission of radiation longward of 400 A also transmit soft X-rays, the grazing incidence telescope presented suppresses this soft X-ray throughput through the incorporation of a Wolter Schwarzschild Type II mirror with large graze angles. The desirable features of an EUV photometric survey telescope are retained. An instrument of this design will be flown on the EUE mission, in order to make a survey of the sky at wavelengths longer than 400 A.

High-Throughput Computation and the Applicability of Monte Carlo Integration in Fatigue Load Estimation of Floating Offshore Wind Turbines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Graf, Peter A.; Stewart, Gordon; Lackner, Matthew

Long-term fatigue loads for floating offshore wind turbines are hard to estimate because they require the evaluation of the integral of a highly nonlinear function over a wide variety of wind and wave conditions. Current design standards involve scanning over a uniform rectangular grid of metocean inputs (e.g., wind speed and direction and wave height and period), which becomes intractable in high dimensions as the number of required evaluations grows exponentially with dimension. Monte Carlo integration offers a potentially efficient alternative because it has theoretical convergence proportional to the inverse of the square root of the number of samples, whichmore » is independent of dimension. In this paper, we first report on the integration of the aeroelastic code FAST into NREL's systems engineering tool, WISDEM, and the development of a high-throughput pipeline capable of sampling from arbitrary distributions, running FAST on a large scale, and postprocessing the results into estimates of fatigue loads. Second, we use this tool to run a variety of studies aimed at comparing grid-based and Monte Carlo-based approaches with calculating long-term fatigue loads. We observe that for more than a few dimensions, the Monte Carlo approach can represent a large improvement in computational efficiency, but that as nonlinearity increases, the effectiveness of Monte Carlo is correspondingly reduced. The present work sets the stage for future research focusing on using advanced statistical methods for analysis of wind turbine fatigue as well as extreme loads.« less

Assessment of the cPAS-based BGISEQ-500 platform for metagenomic sequencing.

PubMed

Fang, Chao; Zhong, Huanzi; Lin, Yuxiang; Chen, Bing; Han, Mo; Ren, Huahui; Lu, Haorong; Luber, Jacob M; Xia, Min; Li, Wangsheng; Stein, Shayna; Xu, Xun; Zhang, Wenwei; Drmanac, Radoje; Wang, Jian; Yang, Huanming; Hammarström, Lennart; Kostic, Aleksandar D; Kristiansen, Karsten; Li, Junhua

2018-03-01

More extensive use of metagenomic shotgun sequencing in microbiome research relies on the development of high-throughput, cost-effective sequencing. Here we present a comprehensive evaluation of the performance of the new high-throughput sequencing platform BGISEQ-500 for metagenomic shotgun sequencing and compare its performance with that of 2 Illumina platforms. Using fecal samples from 20 healthy individuals, we evaluated the intra-platform reproducibility for metagenomic sequencing on the BGISEQ-500 platform in a setup comprising 8 library replicates and 8 sequencing replicates. Cross-platform consistency was evaluated by comparing 20 pairwise replicates on the BGISEQ-500 platform vs the Illumina HiSeq 2000 platform and the Illumina HiSeq 4000 platform. In addition, we compared the performance of the 2 Illumina platforms against each other. By a newly developed overall accuracy quality control method, an average of 82.45 million high-quality reads (96.06% of raw reads) per sample, with 90.56% of bases scoring Q30 and above, was obtained using the BGISEQ-500 platform. Quantitative analyses revealed extremely high reproducibility between BGISEQ-500 intra-platform replicates. Cross-platform replicates differed slightly more than intra-platform replicates, yet a high consistency was observed. Only a low percentage (2.02%-3.25%) of genes exhibited significant differences in relative abundance comparing the BGISEQ-500 and HiSeq platforms, with a bias toward genes with higher GC content being enriched on the HiSeq platforms. Our study provides the first set of performance metrics for human gut metagenomic sequencing data using BGISEQ-500. The high accuracy and technical reproducibility confirm the applicability of the new platform for metagenomic studies, though caution is still warranted when combining metagenomic data from different platforms.

Acoustic separation of circulating tumor cells

PubMed Central

Li, Peng; Mao, Zhangming; Peng, Zhangli; Zhou, Lanlan; Chen, Yuchao; Huang, Po-Hsun; Truica, Cristina I.; Drabick, Joseph J.; El-Deiry, Wafik S.; Dao, Ming; Suresh, Subra; Huang, Tony Jun

2015-01-01

Circulating tumor cells (CTCs) are important targets for cancer biology studies. To further elucidate the role of CTCs in cancer metastasis and prognosis, effective methods for isolating extremely rare tumor cells from peripheral blood must be developed. Acoustic-based methods, which are known to preserve the integrity, functionality, and viability of biological cells using label-free and contact-free sorting, have thus far not been successfully developed to isolate rare CTCs using clinical samples from cancer patients owing to technical constraints, insufficient throughput, and lack of long-term device stability. In this work, we demonstrate the development of an acoustic-based microfluidic device that is capable of high-throughput separation of CTCs from peripheral blood samples obtained from cancer patients. Our method uses tilted-angle standing surface acoustic waves. Parametric numerical simulations were performed to design optimum device geometry, tilt angle, and cell throughput that is more than 20 times higher than previously possible for such devices. We first validated the capability of this device by successfully separating low concentrations (∼100 cells/mL) of a variety of cancer cells from cell culture lines from WBCs with a recovery rate better than 83%. We then demonstrated the isolation of CTCs in blood samples obtained from patients with breast cancer. Our acoustic-based separation method thus offers the potential to serve as an invaluable supplemental tool in cancer research, diagnostics, drug efficacy assessment, and therapeutics owing to its excellent biocompatibility, simple design, and label-free automated operation while offering the capability to isolate rare CTCs in a viable state. PMID:25848039

Acoustic separation of circulating tumor cells.

PubMed

Li, Peng; Mao, Zhangming; Peng, Zhangli; Zhou, Lanlan; Chen, Yuchao; Huang, Po-Hsun; Truica, Cristina I; Drabick, Joseph J; El-Deiry, Wafik S; Dao, Ming; Suresh, Subra; Huang, Tony Jun

2015-04-21

Circulating tumor cells (CTCs) are important targets for cancer biology studies. To further elucidate the role of CTCs in cancer metastasis and prognosis, effective methods for isolating extremely rare tumor cells from peripheral blood must be developed. Acoustic-based methods, which are known to preserve the integrity, functionality, and viability of biological cells using label-free and contact-free sorting, have thus far not been successfully developed to isolate rare CTCs using clinical samples from cancer patients owing to technical constraints, insufficient throughput, and lack of long-term device stability. In this work, we demonstrate the development of an acoustic-based microfluidic device that is capable of high-throughput separation of CTCs from peripheral blood samples obtained from cancer patients. Our method uses tilted-angle standing surface acoustic waves. Parametric numerical simulations were performed to design optimum device geometry, tilt angle, and cell throughput that is more than 20 times higher than previously possible for such devices. We first validated the capability of this device by successfully separating low concentrations (∼100 cells/mL) of a variety of cancer cells from cell culture lines from WBCs with a recovery rate better than 83%. We then demonstrated the isolation of CTCs in blood samples obtained from patients with breast cancer. Our acoustic-based separation method thus offers the potential to serve as an invaluable supplemental tool in cancer research, diagnostics, drug efficacy assessment, and therapeutics owing to its excellent biocompatibility, simple design, and label-free automated operation while offering the capability to isolate rare CTCs in a viable state.

High throughput light absorber discovery, Part 1: An algorithm for automated tauc analysis

DOE PAGES

Suram, Santosh K.; Newhouse, Paul F.; Gregoire, John M.

2016-09-23

High-throughput experimentation provides efficient mapping of composition-property relationships, and its implementation for the discovery of optical materials enables advancements in solar energy and other technologies. In a high throughput pipeline, automated data processing algorithms are often required to match experimental throughput, and we present an automated Tauc analysis algorithm for estimating band gap energies from optical spectroscopy data. The algorithm mimics the judgment of an expert scientist, which is demonstrated through its application to a variety of high throughput spectroscopy data, including the identification of indirect or direct band gaps in Fe 2O 3, Cu 2V 2O 7, and BiVOmore » 4. Here, the applicability of the algorithm to estimate a range of band gap energies for various materials is demonstrated by a comparison of direct-allowed band gaps estimated by expert scientists and by automated algorithm for 60 optical spectra.« less

Ultra-High Throughput Synthesis of Nanoparticles with Homogeneous Size Distribution Using a Coaxial Turbulent Jet Mixer

PubMed Central

2015-01-01

High-throughput production of nanoparticles (NPs) with controlled quality is critical for their clinical translation into effective nanomedicines for diagnostics and therapeutics. Here we report a simple and versatile coaxial turbulent jet mixer that can synthesize a variety of NPs at high throughput up to 3 kg/d, while maintaining the advantages of homogeneity, reproducibility, and tunability that are normally accessible only in specialized microscale mixing devices. The device fabrication does not require specialized machining and is easy to operate. As one example, we show reproducible, high-throughput formulation of siRNA-polyelectrolyte polyplex NPs that exhibit effective gene knockdown but exhibit significant dependence on batch size when formulated using conventional methods. The coaxial turbulent jet mixer can accelerate the development of nanomedicines by providing a robust and versatile platform for preparation of NPs at throughputs suitable for in vivo studies, clinical trials, and industrial-scale production. PMID:24824296

Characterization of matrix effects in developing rugged high-throughput LC-MS/MS methods for bioanalysis.

PubMed

Li, Fumin; Wang, Jun; Jenkins, Rand

2016-05-01

There is an ever-increasing demand for high-throughput LC-MS/MS bioanalytical assays to support drug discovery and development. Matrix effects of sofosbuvir (protonated) and paclitaxel (sodiated) were thoroughly evaluated using high-throughput chromatography (defined as having a run time ≤1 min) under 14 elution conditions with extracts from protein precipitation, liquid-liquid extraction and solid-phase extraction. A slight separation, in terms of retention time, between underlying matrix components and sofosbuvir/paclitaxel can greatly alleviate matrix effects. High-throughput chromatography, with proper optimization, can provide rapid and effective chromatographic separation under 1 min to alleviate matrix effects and enhance assay ruggedness for regulated bioanalysis.

Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mueller, C.; Marx, A.; Epp, S. W.

We present a crystallography chip enabling in situ room temperature crystallography at microfocus synchrotron beamlines and X-ray free-electron laser (X-FEL) sources. Compared to other in situ approaches, we observe extremely low background and high diffraction data quality. The chip design is robust and allows fast and efficient loading of thousands of small crystals. The ability to load a large number of protein crystals, at room temperature and with high efficiency, into prescribed positions enables high throughput automated serial crystallography with microfocus synchrotron beamlines. In addition, we demonstrate the application of this chip for femtosecond time-resolved serial crystallography at the Linacmore » Coherent Light Source (LCLS, Menlo Park, California, USA). As a result, the chip concept enables multiple images to be acquired from each crystal, allowing differential detection of changes in diffraction intensities in order to obtain high signal-to-noise and fully exploit the time resolution capabilities of XFELs.« less

Fixed target matrix for femtosecond time-resolved and in situ serial micro-crystallography

DOE PAGES

Mueller, C.; Marx, A.; Epp, S. W.; ...

2015-08-18

We present a crystallography chip enabling in situ room temperature crystallography at microfocus synchrotron beamlines and X-ray free-electron laser (X-FEL) sources. Compared to other in situ approaches, we observe extremely low background and high diffraction data quality. The chip design is robust and allows fast and efficient loading of thousands of small crystals. The ability to load a large number of protein crystals, at room temperature and with high efficiency, into prescribed positions enables high throughput automated serial crystallography with microfocus synchrotron beamlines. In addition, we demonstrate the application of this chip for femtosecond time-resolved serial crystallography at the Linacmore » Coherent Light Source (LCLS, Menlo Park, California, USA). As a result, the chip concept enables multiple images to be acquired from each crystal, allowing differential detection of changes in diffraction intensities in order to obtain high signal-to-noise and fully exploit the time resolution capabilities of XFELs.« less

High throughput system for magnetic manipulation of cells, polymers, and biomaterials

PubMed Central

Spero, Richard Chasen; Vicci, Leandra; Cribb, Jeremy; Bober, David; Swaminathan, Vinay; O’Brien, E. Timothy; Rogers, Stephen L.; Superfine, R.

2008-01-01

In the past decade, high throughput screening (HTS) has changed the way biochemical assays are performed, but manipulation and mechanical measurement of micro- and nanoscale systems have not benefited from this trend. Techniques using microbeads (particles ∼0.1–10 μm) show promise for enabling high throughput mechanical measurements of microscopic systems. We demonstrate instrumentation to magnetically drive microbeads in a biocompatible, multiwell magnetic force system. It is based on commercial HTS standards and is scalable to 96 wells. Cells can be cultured in this magnetic high throughput system (MHTS). The MHTS can apply independently controlled forces to 16 specimen wells. Force calibrations demonstrate forces in excess of 1 nN, predicted force saturation as a function of pole material, and powerlaw dependence of F∼r−2.7±0.1. We employ this system to measure the stiffness of SR2+ Drosophila cells. MHTS technology is a key step toward a high throughput screening system for micro- and nanoscale biophysical experiments. PMID:19044357

Carbohydrate Microarray Technology Applied to High-Throughput Mapping of Plant Cell Wall Glycans Using Comprehensive Microarray Polymer Profiling (CoMPP).

PubMed

Kračun, Stjepan Krešimir; Fangel, Jonatan Ulrik; Rydahl, Maja Gro; Pedersen, Henriette Lodberg; Vidal-Melgosa, Silvia; Willats, William George Tycho

2017-01-01

Cell walls are an important feature of plant cells and a major component of the plant glycome. They have both structural and physiological functions and are critical for plant growth and development. The diversity and complexity of these structures demand advanced high-throughput techniques to answer questions about their structure, functions and roles in both fundamental and applied scientific fields. Microarray technology provides both the high-throughput and the feasibility aspects required to meet that demand. In this chapter, some of the most recent microarray-based techniques relating to plant cell walls are described together with an overview of related contemporary techniques applied to carbohydrate microarrays and their general potential in glycoscience. A detailed experimental procedure for high-throughput mapping of plant cell wall glycans using the comprehensive microarray polymer profiling (CoMPP) technique is included in the chapter and provides a good example of both the robust and high-throughput nature of microarrays as well as their applicability to plant glycomics.

Identification of functional modules using network topology and high-throughput data.

PubMed

Ulitsky, Igor; Shamir, Ron

2007-01-26

With the advent of systems biology, biological knowledge is often represented today by networks. These include regulatory and metabolic networks, protein-protein interaction networks, and many others. At the same time, high-throughput genomics and proteomics techniques generate very large data sets, which require sophisticated computational analysis. Usually, separate and different analysis methodologies are applied to each of the two data types. An integrated investigation of network and high-throughput information together can improve the quality of the analysis by accounting simultaneously for topological network properties alongside intrinsic features of the high-throughput data. We describe a novel algorithmic framework for this challenge. We first transform the high-throughput data into similarity values, (e.g., by computing pairwise similarity of gene expression patterns from microarray data). Then, given a network of genes or proteins and similarity values between some of them, we seek connected sub-networks (or modules) that manifest high similarity. We develop algorithms for this problem and evaluate their performance on the osmotic shock response network in S. cerevisiae and on the human cell cycle network. We demonstrate that focused, biologically meaningful and relevant functional modules are obtained. In comparison with extant algorithms, our approach has higher sensitivity and higher specificity. We have demonstrated that our method can accurately identify functional modules. Hence, it carries the promise to be highly useful in analysis of high throughput data.

Stepping into the omics era: Opportunities and challenges for biomaterials science and engineering.

PubMed

Groen, Nathalie; Guvendiren, Murat; Rabitz, Herschel; Welsh, William J; Kohn, Joachim; de Boer, Jan

2016-04-01

The research paradigm in biomaterials science and engineering is evolving from using low-throughput and iterative experimental designs towards high-throughput experimental designs for materials optimization and the evaluation of materials properties. Computational science plays an important role in this transition. With the emergence of the omics approach in the biomaterials field, referred to as materiomics, high-throughput approaches hold the promise of tackling the complexity of materials and understanding correlations between material properties and their effects on complex biological systems. The intrinsic complexity of biological systems is an important factor that is often oversimplified when characterizing biological responses to materials and establishing property-activity relationships. Indeed, in vitro tests designed to predict in vivo performance of a given biomaterial are largely lacking as we are not able to capture the biological complexity of whole tissues in an in vitro model. In this opinion paper, we explain how we reached our opinion that converging genomics and materiomics into a new field would enable a significant acceleration of the development of new and improved medical devices. The use of computational modeling to correlate high-throughput gene expression profiling with high throughput combinatorial material design strategies would add power to the analysis of biological effects induced by material properties. We believe that this extra layer of complexity on top of high-throughput material experimentation is necessary to tackle the biological complexity and further advance the biomaterials field. In this opinion paper, we postulate that converging genomics and materiomics into a new field would enable a significant acceleration of the development of new and improved medical devices. The use of computational modeling to correlate high-throughput gene expression profiling with high throughput combinatorial material design strategies would add power to the analysis of biological effects induced by material properties. We believe that this extra layer of complexity on top of high-throughput material experimentation is necessary to tackle the biological complexity and further advance the biomaterials field. Copyright © 2016. Published by Elsevier Ltd.

A high-throughput AO/PI-based cell concentration and viability detection method using the Celigo image cytometry.

PubMed

Chan, Leo Li-Ying; Smith, Tim; Kumph, Kendra A; Kuksin, Dmitry; Kessel, Sarah; Déry, Olivier; Cribbes, Scott; Lai, Ning; Qiu, Jean

2016-10-01

To ensure cell-based assays are performed properly, both cell concentration and viability have to be determined so that the data can be normalized to generate meaningful and comparable results. Cell-based assays performed in immuno-oncology, toxicology, or bioprocessing research often require measuring of multiple samples and conditions, thus the current automated cell counter that uses single disposable counting slides is not practical for high-throughput screening assays. In the recent years, a plate-based image cytometry system has been developed for high-throughput biomolecular screening assays. In this work, we demonstrate a high-throughput AO/PI-based cell concentration and viability method using the Celigo image cytometer. First, we validate the method by comparing directly to Cellometer automated cell counter. Next, cell concentration dynamic range, viability dynamic range, and consistency are determined. The high-throughput AO/PI method described here allows for 96-well to 384-well plate samples to be analyzed in less than 7 min, which greatly reduces the time required for the single sample-based automated cell counter. In addition, this method can improve the efficiency for high-throughput screening assays, where multiple cell counts and viability measurements are needed prior to performing assays such as flow cytometry, ELISA, or simply plating cells for cell culture.

Accelerating the Design of Solar Thermal Fuel Materials through High Throughput Simulations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Y; Grossman, JC

2014-12-01

Solar thermal fuels (STF) store the energy of sunlight, which can then be released later in the form of heat, offering an emission-free and renewable solution for both solar energy conversion and storage. However, this approach is currently limited by the lack of low-cost materials with high energy density and high stability. In this Letter, we present an ab initio high-throughput computational approach to accelerate the design process and allow for searches over a broad class of materials. The high-throughput screening platform we have developed can run through large numbers of molecules composed of earth-abundant elements and identifies possible metastablemore » structures of a given material. Corresponding isomerization enthalpies associated with the metastable structures are then computed. Using this high-throughput simulation approach, we have discovered molecular structures with high isomerization enthalpies that have the potential to be new candidates for high-energy density STF. We have also discovered physical principles to guide further STF materials design through structural analysis. More broadly, our results illustrate the potential of using high-throughput ab initio simulations to design materials that undergo targeted structural transitions.« less

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

Code of Federal Regulations, 2010 CFR

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...

Accurate high-throughput structure mapping and prediction with transition metal ion FRET

PubMed Central

Yu, Xiaozhen; Wu, Xiongwu; Bermejo, Guillermo A.; Brooks, Bernard R.; Taraska, Justin W.

2013-01-01

Mapping the landscape of a protein’s conformational space is essential to understanding its functions and regulation. The limitations of many structural methods have made this process challenging for most proteins. Here, we report that transition metal ion FRET (tmFRET) can be used in a rapid, highly parallel screen, to determine distances from multiple locations within a protein at extremely low concentrations. The distances generated through this screen for the protein Maltose Binding Protein (MBP) match distances from the crystal structure to within a few angstroms. Furthermore, energy transfer accurately detects structural changes during ligand binding. Finally, fluorescence-derived distances can be used to guide molecular simulations to find low energy states. Our results open the door to rapid, accurate mapping and prediction of protein structures at low concentrations, in large complex systems, and in living cells. PMID:23273426

The STROBE-X Science Case: An Overview

NASA Astrophysics Data System (ADS)

Maccarone, Thomas J.; STROBE-X consortium

2018-01-01

STROBE-X is a proposed NASA Probe class mission aimed at the extremes of high throughput X-ray astronomy, making use of an 8 m^2 total collecting area, CCD-quality spectral resolution, and a state-of-the art wide field monitor with both very large instantaneous sky coverage (ideal for follow-up of LIGO events) and good intrinsic spectral and time resolution. The core goals are time domain astrophysics and high count spectroscopy. Its capabilities span a broad range of topics, including those traditional to X-ray timing missions, like understanding the equation of states of neutron stars, and the spin distributions and masses of neutron stars and stellar mass and supermassive black holes, and the rates, and detailed properties, of a variety of classes of X-ray transients; and also topics not traditionally studied by such missions such as the spectra of supernova remnants, comets and of clusters and groups of galaxies.

Gas dispersion measurements using a mobile Raman lidar system

NASA Technical Reports Server (NTRS)

Houston, J. D.; Brown, D. R.

1986-01-01

The exploitation of natural gas resources to supply energy demands has resulted in the need to engineer pipelines and plants capable of handling extremely high pressures and throughputs. Consequently, more attention has been directed to evaluating the consequences of releases of material whether accidental or deliberate in nature. An important aspect of assessing the consequences of a release is an understanding of how gas disperses in the atmosphere over a wide range of release and atmospheric conditions. The most cost effective way of providing such information is through the development and use of reliable theoretical prediction methods. The need for some form of remote sensing device was identified. The various possibilities studied led to the conclusion that LIDAR (Light Detection And Ranging) offered the most suitable method. The system designed and built is described, and its recent use in monitoring operational ventings from a high pressure transmission system is discussed.

2013 R&D 100 Award: 'SHIELD' protects NIF optics from harmful pulses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chou, Jason

In the past, it took as long as 12 hours to manually screen 48 critical checkpoints at the National Ignition Facility (NIF) for harmful laser pulses. The screening equipment had to be moved from point to point throughout a facility the size of three football fields. Now with a new technology, called Laser SHIELD (Screening at High-throughput to Identify Energetic Laser Distortion), and with the push of a button, the screening can be done in less than one second. Proper screening of pulses is critical for the operation of high-energy lasers to ensure that the laser does not exceed safemore » operating conditions for optics. The energetic beams of light are so powerful that, when left uncontrolled, they can shatter the extremely valuable glass inside the laser. If a harmful pulse is found, immediate adjustments can be made in order to protect the optics for the facility. Laser SHIELD is a custom-designed high-throughput screening system built from low-cost and commercially available components found in the telecommunications industry. Its all-fiber design makes it amenable to the unique needs of high-energy laser facilities, including routing to intricate pick-off locations, immunity to electromagnetic interference and low-loss transport (up to several kilometers). The technology offers several important benefits for NIF. First, the facility is able to fire more shots in less time-an efficiency that saves the facility millions of dollars each year. Second, high-energy lasers are more flexible to wavelength changes requested by target physicists. Third, by identifying harmful pulses before they damage the laser's optics, the facility potentially saves hundreds of thousands of dollars in maintenance costs each year.« less

2013 R&D 100 Award: 'SHIELD' protects NIF optics from harmful pulses

ScienceCinema

Chou, Jason

2018-02-13

In the past, it took as long as 12 hours to manually screen 48 critical checkpoints at the National Ignition Facility (NIF) for harmful laser pulses. The screening equipment had to be moved from point to point throughout a facility the size of three football fields. Now with a new technology, called Laser SHIELD (Screening at High-throughput to Identify Energetic Laser Distortion), and with the push of a button, the screening can be done in less than one second. Proper screening of pulses is critical for the operation of high-energy lasers to ensure that the laser does not exceed safe operating conditions for optics. The energetic beams of light are so powerful that, when left uncontrolled, they can shatter the extremely valuable glass inside the laser. If a harmful pulse is found, immediate adjustments can be made in order to protect the optics for the facility. Laser SHIELD is a custom-designed high-throughput screening system built from low-cost and commercially available components found in the telecommunications industry. Its all-fiber design makes it amenable to the unique needs of high-energy laser facilities, including routing to intricate pick-off locations, immunity to electromagnetic interference and low-loss transport (up to several kilometers). The technology offers several important benefits for NIF. First, the facility is able to fire more shots in less time-an efficiency that saves the facility millions of dollars each year. Second, high-energy lasers are more flexible to wavelength changes requested by target physicists. Third, by identifying harmful pulses before they damage the laser's optics, the facility potentially saves hundreds of thousands of dollars in maintenance costs each year.

Compound Transfer by Acoustic Droplet Ejection Promotes Quality and Efficiency in Ultra-High-Throughput Screening Campaigns.

PubMed

Dawes, Timothy D; Turincio, Rebecca; Jones, Steven W; Rodriguez, Richard A; Gadiagellan, Dhireshan; Thana, Peter; Clark, Kevin R; Gustafson, Amy E; Orren, Linda; Liimatta, Marya; Gross, Daniel P; Maurer, Till; Beresini, Maureen H

2016-02-01

Acoustic droplet ejection (ADE) as a means of transferring library compounds has had a dramatic impact on the way in which high-throughput screening campaigns are conducted in many laboratories. Two Labcyte Echo ADE liquid handlers form the core of the compound transfer operation in our 1536-well based ultra-high-throughput screening (uHTS) system. Use of these instruments has promoted flexibility in compound formatting in addition to minimizing waste and eliminating compound carryover. We describe the use of ADE for the generation of assay-ready plates for primary screening as well as for follow-up dose-response evaluations. Custom software has enabled us to harness the information generated by the ADE instrumentation. Compound transfer via ADE also contributes to the screening process outside of the uHTS system. A second fully automated ADE-based system has been used to augment the capacity of the uHTS system as well as to permit efficient use of previously picked compound aliquots for secondary assay evaluations. Essential to the utility of ADE in the high-throughput screening process is the high quality of the resulting data. Examples of data generated at various stages of high-throughput screening campaigns are provided. Advantages and disadvantages of the use of ADE in high-throughput screening are discussed. © 2015 Society for Laboratory Automation and Screening.

An Automated High-Throughput System to Fractionate Plant Natural Products for Drug Discovery

PubMed Central

Tu, Ying; Jeffries, Cynthia; Ruan, Hong; Nelson, Cynthia; Smithson, David; Shelat, Anang A.; Brown, Kristin M.; Li, Xing-Cong; Hester, John P.; Smillie, Troy; Khan, Ikhlas A.; Walker, Larry; Guy, Kip; Yan, Bing

2010-01-01

The development of an automated, high-throughput fractionation procedure to prepare and analyze natural product libraries for drug discovery screening is described. Natural products obtained from plant materials worldwide were extracted and first prefractionated on polyamide solid-phase extraction cartridges to remove polyphenols, followed by high-throughput automated fractionation, drying, weighing, and reformatting for screening and storage. The analysis of fractions with UPLC coupled with MS, PDA and ELSD detectors provides information that facilitates characterization of compounds in active fractions. Screening of a portion of fractions yielded multiple assay-specific hits in several high-throughput cellular screening assays. This procedure modernizes the traditional natural product fractionation paradigm by seamlessly integrating automation, informatics, and multimodal analytical interrogation capabilities. PMID:20232897

Cryogenic Fiber Optic Assemblies for Spaceflight Environments: Design, Manufacturing, Testing, and Integration

NASA Technical Reports Server (NTRS)

Thomes, W. Joe; Ott, Melanie N.; Chuska, Richard; Switzer, Robert; Onuma, Eleanya; Blair, Diana; Frese, Erich; Matyseck, Marc

2016-01-01

Fiber optic assemblies have been used on spaceflight missions for many years as an enabling technology for routing, transmitting, and detecting optical signals. Due to the overwhelming success of NASA in implementing fiber optic assemblies on spaceflight science-based instruments, system scientists increasingly request fibers that perform in extreme environments while still maintaining very high optical transmission, stability, and reliability. Many new applications require fiber optic assemblies that will operate down to cryogenic temperatures as low as 20 Kelvin. In order for the fiber assemblies to operate with little loss in optical throughput at these extreme temperatures requires a system level approach all the way from how the fiber assembly is manufactured to how it is held, routed, and integrated. The NASA Goddard Code 562 Photonics Group has been designing, manufacturing, testing, and integrating fiber optics for spaceflight and other high reliability applications for nearly 20 years. Design techniques and lessons learned over the years are consistently applied to developing new fiber optic assemblies that meet these demanding environments. System level trades, fiber assembly design methods, manufacturing, testing, and integration will be discussed. Specific recent examples of ground support equipment for the James Webb Space Telescope (JWST); the Ice, Cloud and Land Elevation Satellite-2 (ICESat-2); and others will be included.

Exploiting induced variation to dissect quantitative traits in barley.

PubMed

Druka, Arnis; Franckowiak, Jerome; Lundqvist, Udda; Bonar, Nicola; Alexander, Jill; Guzy-Wrobelska, Justyna; Ramsay, Luke; Druka, Ilze; Grant, Iain; Macaulay, Malcolm; Vendramin, Vera; Shahinnia, Fahimeh; Radovic, Slobodanka; Houston, Kelly; Harrap, David; Cardle, Linda; Marshall, David; Morgante, Michele; Stein, Nils; Waugh, Robbie

2010-04-01

The identification of genes underlying complex quantitative traits such as grain yield by means of conventional genetic analysis (positional cloning) requires the development of several large mapping populations. However, it is possible that phenotypically related, but more extreme, allelic variants generated by mutational studies could provide a means for more efficient cloning of QTLs (quantitative trait loci). In barley (Hordeum vulgare), with the development of high-throughput genome analysis tools, efficient genome-wide identification of genetic loci harbouring mutant alleles has recently become possible. Genotypic data from NILs (near-isogenic lines) that carry induced or natural variants of genes that control aspects of plant development can be compared with the location of QTLs to potentially identify candidate genes for development--related traits such as grain yield. As yield itself can be divided into a number of allometric component traits such as tillers per plant, kernels per spike and kernel size, mutant alleles that both affect these traits and are located within the confidence intervals for major yield QTLs may represent extreme variants of the underlying genes. In addition, the development of detailed comparative genomic models based on the alignment of a high-density barley gene map with the rice and sorghum physical maps, has enabled an informed prioritization of 'known function' genes as candidates for both QTLs and induced mutant genes.

Cryogenic fiber optic assemblies for spaceflight environments: design, manufacturing, testing, and integration

NASA Astrophysics Data System (ADS)

Thomes, W. Joe; Ott, Melanie N.; Chuska, Richard; Switzer, Robert; Onuma, Eleanya; Blair, Diana; Frese, Erich; Matyseck, Marc

2016-09-01

Fiber optic assemblies have been used on spaceflight missions for many years as an enabling technology for routing, transmitting, and detecting optical signals. Due to the overwhelming success of NASA in implementing fiber optic assemblies on spaceflight science-based instruments, system scientists increasingly request fibers that perform in extreme environments while still maintaining very high optical transmission, stability, and reliability. Many new applications require fiber optic assemblies that will operate down to cryogenic temperatures as low as 20 Kelvin. In order for the fiber assemblies to operate with little loss in optical throughput at these extreme temperatures requires a system level approach all the way from how the fiber assembly is manufactured to how it is held, routed, and integrated. The NASA Goddard Code 562 Photonics Group has been designing, manufacturing, testing, and integrating fiber optics for spaceflight and other high reliability applications for nearly 20 years. Design techniques and lessons learned over the years are consistently applied to developing new fiber optic assemblies that meet these demanding environments. System level trades, fiber assembly design methods, manufacturing, testing, and integration will be discussed. Specific recent examples of ground support equipment for the James Webb Space Telescope (JWST); the Ice, Cloud and Land Elevation Satellite-2 (ICESat- 2); and others will be included.

High-throughput measurements of the optical redox ratio using a commercial microplate reader.

PubMed

Cannon, Taylor M; Shah, Amy T; Walsh, Alex J; Skala, Melissa C

2015-01-01

There is a need for accurate, high-throughput, functional measures to gauge the efficacy of potential drugs in living cells. As an early marker of drug response in cells, cellular metabolism provides an attractive platform for high-throughput drug testing. Optical techniques can noninvasively monitor NADH and FAD, two autofluorescent metabolic coenzymes. The autofluorescent redox ratio, defined as the autofluorescence intensity of NADH divided by that of FAD, quantifies relative rates of cellular glycolysis and oxidative phosphorylation. However, current microscopy methods for redox ratio quantification are time-intensive and low-throughput, limiting their practicality in drug screening. Alternatively, high-throughput commercial microplate readers quickly measure fluorescence intensities for hundreds of wells. This study found that a commercial microplate reader can differentiate the receptor status of breast cancer cell lines (p < 0.05) based on redox ratio measurements without extrinsic contrast agents. Furthermore, microplate reader redox ratio measurements resolve response (p < 0.05) and lack of response (p > 0.05) in cell lines that are responsive and nonresponsive, respectively, to the breast cancer drug trastuzumab. These studies indicate that the microplate readers can be used to measure the redox ratio in a high-throughput manner and are sensitive enough to detect differences in cellular metabolism that are consistent with microscopy results.

Exploring the hydraulic fracturing parameter space: a novel high-pressure, high-throughput reactor system for investigating subsurface chemical transformations.

PubMed

Sumner, Andrew J; Plata, Desiree L

2018-02-21

Hydraulic fracturing coupled with horizontal drilling (HDHF) involves the deep-well injection of a fracturing fluid composed of diverse and numerous chemical additives designed to facilitate the release and collection of natural gas from shale plays. Analyses of flowback wastewaters have revealed organic contamination from both geogenic and anthropogenic sources. The additional detections of undisclosed halogenated chemicals suggest unintended in situ transformation of reactive additives, but the formation pathways for these are unclear in subsurface brines. To develop an efficient experimental framework for investigating the complex shale-well parameter space, we have reviewed and synthesized geospatial well data detailing temperature, pressure, pH, and halide ion values as well as industrial chemical disclosure and concentration data. Our findings showed subsurface conditions can reach pressures up to 4500 psi (310 bars) and temperatures up to 95 °C, while at least 588 unique chemicals have been disclosed by industry, including reactive oxidants and acids. Given the extreme conditions necessary to simulate the subsurface, we briefly highlighted existing geochemical reactor systems rated to the necessary pressures and temperatures, identifying throughput as a key limitation. In response, we designed and developed a custom reactor system capable of achieving 5000 psi (345 bars) and 90 °C at low cost with 15 individual reactors that are readily turned over. To demonstrate the system's throughput, we simultaneously tested 12 disclosed HDHF chemicals against a radical initiator compound in simulated subsurface conditions, ruling out a dozen potential transformation pathways in a single experiment. This review outlines the dynamic and diverse parameter range experienced by HDHF chemical additives and provides an optimized framework and novel reactor system for the methodical study of subsurface transformation pathways. Ultimately, enabling such studies will provide urgently needed clarity for water treatment downstream or releases to the environment.

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

PubMed Central

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

An image analysis toolbox for high-throughput C. elegans assays

PubMed Central

Wählby, Carolina; Kamentsky, Lee; Liu, Zihan H.; Riklin-Raviv, Tammy; Conery, Annie L.; O’Rourke, Eyleen J.; Sokolnicki, Katherine L.; Visvikis, Orane; Ljosa, Vebjorn; Irazoqui, Javier E.; Golland, Polina; Ruvkun, Gary; Ausubel, Frederick M.; Carpenter, Anne E.

2012-01-01

We present a toolbox for high-throughput screening of image-based Caenorhabditis elegans phenotypes. The image analysis algorithms measure morphological phenotypes in individual worms and are effective for a variety of assays and imaging systems. This WormToolbox is available via the open-source CellProfiler project and enables objective scoring of whole-animal high-throughput image-based assays of C. elegans for the study of diverse biological pathways relevant to human disease. PMID:22522656

High-throughput, image-based screening of pooled genetic variant libraries

PubMed Central

Emanuel, George; Moffitt, Jeffrey R.; Zhuang, Xiaowei

2018-01-01

Image-based, high-throughput screening of genetic perturbations will advance both biology and biotechnology. We report a high-throughput screening method that allows diverse genotypes and corresponding phenotypes to be imaged in numerous individual cells. We achieve genotyping by introducing barcoded genetic variants into cells and using massively multiplexed FISH to measure the barcodes. We demonstrated this method by screening mutants of the fluorescent protein YFAST, yielding brighter and more photostable YFAST variants. PMID:29083401

Experimental Design for Combinatorial and High Throughput Materials Development

NASA Astrophysics Data System (ADS)

Cawse, James N.

2002-12-01

In the past decade, combinatorial and high throughput experimental methods have revolutionized the pharmaceutical industry, allowing researchers to conduct more experiments in a week than was previously possible in a year. Now high throughput experimentation is rapidly spreading from its origins in the pharmaceutical world to larger industrial research establishments such as GE and DuPont, and even to smaller companies and universities. Consequently, researchers need to know the kinds of problems, desired outcomes, and appropriate patterns for these new strategies. Editor James Cawse's far-reaching study identifies and applies, with specific examples, these important new principles and techniques. Experimental Design for Combinatorial and High Throughput Materials Development progresses from methods that are now standard, such as gradient arrays, to mathematical developments that are breaking new ground. The former will be particularly useful to researchers entering the field, while the latter should inspire and challenge advanced practitioners. The book's contents are contributed by leading researchers in their respective fields. Chapters include: -High Throughput Synthetic Approaches for the Investigation of Inorganic Phase Space -Combinatorial Mapping of Polymer Blends Phase Behavior -Split-Plot Designs -Artificial Neural Networks in Catalyst Development -The Monte Carlo Approach to Library Design and Redesign This book also contains over 200 useful charts and drawings. Industrial chemists, chemical engineers, materials scientists, and physicists working in combinatorial and high throughput chemistry will find James Cawse's study to be an invaluable resource.

Deciphering the genomic targets of alkylating polyamide conjugates using high-throughput sequencing

PubMed Central

Chandran, Anandhakumar; Syed, Junetha; Taylor, Rhys D.; Kashiwazaki, Gengo; Sato, Shinsuke; Hashiya, Kaori; Bando, Toshikazu; Sugiyama, Hiroshi

2016-01-01

Chemically engineered small molecules targeting specific genomic sequences play an important role in drug development research. Pyrrole-imidazole polyamides (PIPs) are a group of molecules that can bind to the DNA minor-groove and can be engineered to target specific sequences. Their biological effects rely primarily on their selective DNA binding. However, the binding mechanism of PIPs at the chromatinized genome level is poorly understood. Herein, we report a method using high-throughput sequencing to identify the DNA-alkylating sites of PIP-indole-seco-CBI conjugates. High-throughput sequencing analysis of conjugate 2 showed highly similar DNA-alkylating sites on synthetic oligos (histone-free DNA) and on human genomes (chromatinized DNA context). To our knowledge, this is the first report identifying alkylation sites across genomic DNA by alkylating PIP conjugates using high-throughput sequencing. PMID:27098039

A standardized framework for accurate, high-throughput genotyping of recombinant and non-recombinant viral sequences.

PubMed

Alcantara, Luiz Carlos Junior; Cassol, Sharon; Libin, Pieter; Deforche, Koen; Pybus, Oliver G; Van Ranst, Marc; Galvão-Castro, Bernardo; Vandamme, Anne-Mieke; de Oliveira, Tulio

2009-07-01

Human immunodeficiency virus type-1 (HIV-1), hepatitis B and C and other rapidly evolving viruses are characterized by extremely high levels of genetic diversity. To facilitate diagnosis and the development of prevention and treatment strategies that efficiently target the diversity of these viruses, and other pathogens such as human T-lymphotropic virus type-1 (HTLV-1), human herpes virus type-8 (HHV8) and human papillomavirus (HPV), we developed a rapid high-throughput-genotyping system. The method involves the alignment of a query sequence with a carefully selected set of pre-defined reference strains, followed by phylogenetic analysis of multiple overlapping segments of the alignment using a sliding window. Each segment of the query sequence is assigned the genotype and sub-genotype of the reference strain with the highest bootstrap (>70%) and bootscanning (>90%) scores. Results from all windows are combined and displayed graphically using color-coded genotypes. The new Virus-Genotyping Tools provide accurate classification of recombinant and non-recombinant viruses and are currently being assessed for their diagnostic utility. They have incorporated into several HIV drug resistance algorithms including the Stanford (http://hivdb.stanford.edu) and two European databases (http://www.umcutrecht.nl/subsite/spread-programme/ and http://www.hivrdb.org.uk/) and have been successfully used to genotype a large number of sequences in these and other databases. The tools are a PHP/JAVA web application and are freely accessible on a number of servers including: http://bioafrica.mrc.ac.za/rega-genotype/html/, http://lasp.cpqgm.fiocruz.br/virus-genotype/html/, http://jose.med.kuleuven.be/genotypetool/html/.

Separation of rare oligodendrocyte progenitor cells from brain using a high-throughput multilayer thermoplastic-based microfluidic device.

PubMed

Didar, Tohid Fatanat; Li, Kebin; Veres, Teodor; Tabrizian, Maryam

2013-07-01

Despite the advances made in the field of regenerative medicine, the progress in cutting-edge technologies for separating target therapeutic cells are still at early stage of development. These cells are often rare, such as stem cells or progenitor cells that their overall properties should be maintained during the separation process for their subsequent application in regenerative medicine. This work, presents separation of oligodendrocyte progenitor cells (OPCs) from rat brain primary cultures using an integrated thermoplastic elastomeric (TPE)- based multilayer microfluidic device fabricated using hot-embossing technology. OPCs are frequently used in recovery, repair and regeneration of central nervous system after injuries. Indeed, their ability to differentiate in vitro into myelinating oligodendrocytes, are extremely important for myelin repair. OPCs form 5-10% of the glial cells population. The traditional macroscale techniques for OPCs separation require pre-processing of cells and/or multiple time consuming steps with low efficiency leading very often to alteration of their properties. The proposed methodology implies to separate OPCs based on their smaller size compared to other cells from the brain tissue mixture. Using aforementioned microfluidic chip embedded with a 5 μm membrane pore size and micropumping system, a separation efficiency more than 99% was achieved. This microchip was able to operate at flow rates up to 100 μl/min, capable of separating OPCs from a confluent 75 cm(2) cell culture flask in less than 10 min, which provides us with a high-throughput and highly efficient separation expected from any cell sorting techniques. Copyright © 2013 Elsevier Ltd. All rights reserved.

Nanoparticle–Film Plasmon Ruler Interrogated with Transmission Visible Spectroscopy

PubMed Central

2015-01-01

The widespread use of plasmonic nanorulers (PNRs) in sensing platforms has been plagued by technical challenges associated with the development of methods to fabricate precisely controlled nanostructures with high yield and characterize them with high throughput. We have previously shown that creating PNRs in a nanoparticle–film (NP–film) format enables the fabrication of an extremely large population of uniform PNRs with 100% yield using a self-assembly approach, which facilitates high-throughput PNR characterization using ensemble spectroscopic measurements and eliminates the need for expensive microscopy systems required by many other PNR platforms. We expand upon this prior work herein, showing that the NP–film PNR can be made compatible with aqueous sensing studies by adapting it for use in a transmission localized surface plasmon resonance spectroscopy format, where the coupled NP–film resonance responsible for the PNR signal is directly probed using an extinction measurement from a standard spectrophotometer. We designed slide holders that fit inside standard spectrophotometer cuvettes and position NP–film samples so that the coupled NP–film resonance can be detected in a collinear optical configuration. Once the NP–film PNR samples are cuvette-compatible, it is straightforward to calibrate the PNR in aqueous solution and use it to characterize dynamic, angstrom-scale distance changes resulting from pH-induced swelling of polyelectrolyte (PE) spacer layers as thin as 1 PE layer and also of a self-assembled monolayer of an amine-terminated alkanethiol. This development is an important step toward making PNR sensors more user-friendly and encouraging their widespread use in various sensing schemes. PMID:25541618

Development of rapid and sensitive high throughput pharmacologic assays for marine phycotoxins.

PubMed

Van Dolah, F M; Finley, E L; Haynes, B L; Doucette, G J; Moeller, P D; Ramsdell, J S

1994-01-01

The lack of rapid, high throughput assays is a major obstacle to many aspects of research on marine phycotoxins. Here we describe the application of microplate scintillation technology to develop high throughput assays for several classes of marine phycotoxin based on their differential pharmacologic actions. High throughput "drug discovery" format microplate receptor binding assays developed for brevetoxins/ciguatoxins and for domoic acid are described. Analysis for brevetoxins/ciguatoxins is carried out by binding competition with [3H] PbTx-3 for site 5 on the voltage dependent sodium channel in rat brain synaptosomes. Analysis of domoic acid is based on binding competition with [3H] kainic acid for the kainate/quisqualate glutamate receptor using frog brain synaptosomes. In addition, a high throughput microplate 45Ca flux assay for determination of maitotoxins is described. These microplate assays can be completed within 3 hours, have sensitivities of less than 1 ng, and can analyze dozens of samples simultaneously. The assays have been demonstrated to be useful for assessing algal toxicity and for assay-guided purification of toxins, and are applicable to the detection of biotoxins in seafood.

High-Throughput/High-Content Screening Assays with Engineered Nanomaterials in ToxCast

EPA Science Inventory

High-throughput and high-content screens are attractive approaches for prioritizing nanomaterial hazards and informing targeted testing due to the impracticality of using traditional toxicological testing on the large numbers and varieties of nanomaterials. The ToxCast program a...

High-throughput protein concentration and buffer exchange: comparison of ultrafiltration and ammonium sulfate precipitation.

PubMed

Moore, Priscilla A; Kery, Vladimir

2009-01-01

High-throughput protein purification is a complex, multi-step process. There are several technical challenges in the course of this process that are not experienced when purifying a single protein. Among the most challenging are the high-throughput protein concentration and buffer exchange, which are not only labor-intensive but can also result in significant losses of purified proteins. We describe two methods of high-throughput protein concentration and buffer exchange: one using ammonium sulfate precipitation and one using micro-concentrating devices based on membrane ultrafiltration. We evaluated the efficiency of both methods on a set of 18 randomly selected purified proteins from Shewanella oneidensis. While both methods provide similar yield and efficiency, the ammonium sulfate precipitation is much less labor intensive and time consuming than the ultrafiltration.

Challenges and requirements of mask data processing for multi-beam mask writer

NASA Astrophysics Data System (ADS)

Choi, Jin; Lee, Dong Hyun; Park, Sinjeung; Lee, SookHyun; Tamamushi, Shuichi; Shin, In Kyun; Jeon, Chan Uk

2015-07-01

To overcome the resolution and throughput of current mask writer for advanced lithography technologies, the platform of e-beam writer have been evolved by the developments of hardware and software in writer. Especially, aggressive optical proximity correction (OPC) for unprecedented extension of optical lithography and the needs of low sensitivity resist for high resolution result in the limit of variable shaped beam writer which is widely used for mass production. The multi-beam mask writer is attractive candidate for photomask writing of sub-10nm device because of its high speed and the large degree of freedom which enable high dose and dose modulation for each pixel. However, the higher dose and almost unlimited appetite for dose modulation challenge the mask data processing (MDP) in aspects of extreme data volume and correction method. Here, we discuss the requirements of mask data processing for multi-beam mask writer and presents new challenges of the data format, data flow, and correction method for user and supplier MDP tool.

Development of the automated circulating tumor cell recovery system with microcavity array.

PubMed

Negishi, Ryo; Hosokawa, Masahito; Nakamura, Seita; Kanbara, Hisashige; Kanetomo, Masafumi; Kikuhara, Yoshihito; Tanaka, Tsuyoshi; Matsunaga, Tadashi; Yoshino, Tomoko

2015-05-15

Circulating tumor cells (CTCs) are well recognized as useful biomarker for cancer diagnosis and potential target of drug discovery for metastatic cancer. Efficient and precise recovery of extremely low concentrations of CTCs from blood has been required to increase the detection sensitivity. Here, an automated system equipped with a microcavity array (MCA) was demonstrated for highly efficient and reproducible CTC recovery. The use of MCA allows selective recovery of cancer cells from whole blood on the basis of differences in size between tumor and blood cells. Intra- and inter-assays revealed that the automated system achieved high efficiency and reproducibility equal to the assay manually performed by well-trained operator. Under optimized assay workflow, the automated system allows efficient and precise cell recovery for non-small cell lung cancer cells spiked in whole blood. The automated CTC recovery system will contribute to high-throughput analysis in the further clinical studies on large cohort of cancer patients. Copyright © 2014 Elsevier B.V. All rights reserved.

Miniaturized high throughput detection system for capillary array electrophoresis on chip with integrated light emitting diode array as addressed ring-shaped light source.

PubMed

Ren, Kangning; Liang, Qionglin; Mu, Xuan; Luo, Guoan; Wang, Yiming

2009-03-07

A novel miniaturized, portable fluorescence detection system for capillary array electrophoresis (CAE) on a microfluidic chip was developed, consisting of a scanning light-emitting diode (LED) light source and a single point photoelectric sensor. Without charge coupled detector (CCD), lens, fibers and moving parts, the system was extremely simplified. Pulsed driving of the LED significantly increased the sensitivity, and greatly reduced the power consumption and photobleaching effect. The highly integrated system was robust and easy to use. All the advantages realized the concept of a portable micro-total analysis system (micro-TAS), which could work on a single universal serial bus (USB) port. Compared with traditional CAE detecting systems, the current system could scan the radial capillary array with high scanning rate. An 8-channel CAE of fluorescein isothiocyanate (FITC) labeled arginine (Arg) on chip was demonstrated with this system, resulting in a limit of detection (LOD) of 640 amol.

The clinical impact of recent advances in LC-MS for cancer biomarker discovery and verification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Hui; Shi, Tujin; Qian, Wei-Jun

2015-12-04

Mass spectrometry-based proteomics has become an indispensable tool in biomedical research with broad applications ranging from fundamental biology, systems biology, and biomarker discovery. Recent advances in LC-MS have made it become a major technology in clinical applications, especially in cancer biomarker discovery and verification. To overcome the challenges associated with the analysis of clinical samples, such as extremely wide dynamic range of protein concentrations in biofluids and the need to perform high throughput and accurate quantification, significant efforts have been devoted to improve the overall performance of LC-MS bases clinical proteomics. In this review, we summarize the recent advances inmore » LC-MS in the aspect of cancer biomarker discovery and quantification, and discuss its potentials, limitations, and future perspectives.« less

Reflective optical imaging system

DOEpatents

Shafer, David R.

2000-01-01

An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements are characterized in order from object to image as convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention increases the slit dimensions associated with ringfield scanning optics, improves wafer throughput and allows higher semiconductor device density.

Reflective optical imaging method and circuit

DOEpatents

Shafer, David R.

2001-01-01

An optical system compatible with short wavelength (extreme ultraviolet) radiation comprising four reflective elements for projecting a mask image onto a substrate. The four optical elements are characterized in order from object to image as convex, concave, convex and concave mirrors. The optical system is particularly suited for step and scan lithography methods. The invention increases the slit dimensions associated with ringfield scanning optics, improves wafer throughput and allows higher semiconductor device density.

Extremely Lightweight Intrusion Detection (ELIDe)

DTIC Science & Technology

2013-12-01

devices that would be more commonly found in a dynamic tactical environment. As a point of reference, the Raspberry Pi single-chip computer (4) is...the ELIDe application onto a resource- constrained hardware platform more likely to be used in a mobile tactical network, and the Raspberry Pi was...chosen as that representative platform. ELIDe was successfully tested on a Raspberry Pi , its throughput was benchmarked at approximately 8.3 megabits

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

NASA Astrophysics Data System (ADS)

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.

SSR_pipeline: a bioinformatic infrastructure for identifying microsatellites from paired-end Illumina high-throughput DNA sequencing data

USGS Publications Warehouse

Miller, Mark P.; Knaus, Brian J.; Mullins, Thomas D.; Haig, Susan M.

2013-01-01

SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (e.g., microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains 3 analysis modules along with a fourth control module that can automate analyses of large volumes of data. The modules are used to 1) identify the subset of paired-end sequences that pass Illumina quality standards, 2) align paired-end reads into a single composite DNA sequence, and 3) identify sequences that possess microsatellites (both simple and compound) conforming to user-specified parameters. The microsatellite search algorithm is extremely efficient, and we have used it to identify repeats with motifs from 2 to 25bp in length. Each of the 3 analysis modules can also be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc.). We demonstrate use of the program with data from the brine fly Ephydra packardi (Diptera: Ephydridae) and provide empirical timing benchmarks to illustrate program performance on a common desktop computer environment. We further show that the Illumina platform is capable of identifying large numbers of microsatellites, even when using unenriched sample libraries and a very small percentage of the sequencing capacity from a single DNA sequencing run. All modules from SSR_pipeline are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, and Windows).

SSR_pipeline: a bioinformatic infrastructure for identifying microsatellites from paired-end Illumina high-throughput DNA sequencing data.

PubMed

Miller, Mark P; Knaus, Brian J; Mullins, Thomas D; Haig, Susan M

2013-01-01

SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (e.g., microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains 3 analysis modules along with a fourth control module that can automate analyses of large volumes of data. The modules are used to 1) identify the subset of paired-end sequences that pass Illumina quality standards, 2) align paired-end reads into a single composite DNA sequence, and 3) identify sequences that possess microsatellites (both simple and compound) conforming to user-specified parameters. The microsatellite search algorithm is extremely efficient, and we have used it to identify repeats with motifs from 2 to 25 bp in length. Each of the 3 analysis modules can also be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc.). We demonstrate use of the program with data from the brine fly Ephydra packardi (Diptera: Ephydridae) and provide empirical timing benchmarks to illustrate program performance on a common desktop computer environment. We further show that the Illumina platform is capable of identifying large numbers of microsatellites, even when using unenriched sample libraries and a very small percentage of the sequencing capacity from a single DNA sequencing run. All modules from SSR_pipeline are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, and Windows).

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

PubMed

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.

High throughput single cell counting in droplet-based microfluidics.

PubMed

Lu, Heng; Caen, Ouriel; Vrignon, Jeremy; Zonta, Eleonora; El Harrak, Zakaria; Nizard, Philippe; Baret, Jean-Christophe; Taly, Valérie

2017-05-02

Droplet-based microfluidics is extensively and increasingly used for high-throughput single-cell studies. However, the accuracy of the cell counting method directly impacts the robustness of such studies. We describe here a simple and precise method to accurately count a large number of adherent and non-adherent human cells as well as bacteria. Our microfluidic hemocytometer provides statistically relevant data on large populations of cells at a high-throughput, used to characterize cell encapsulation and cell viability during incubation in droplets.

High-Throughput Sequencing of Germline and Tumor From Men with Early-Onset Metastatic Prostate Cancer

DTIC Science & Technology

2016-12-01

AWARD NUMBER: W81XWH-13-1-0371 TITLE: High-Throughput Sequencing of Germline and Tumor From Men with Early- Onset Metastatic Prostate Cancer...DATES COVERED 30 Sep 2013 - 29 Sep 2016 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER High-Throughput Sequencing of Germline and Tumor From Men with...presenting with metastatic prostate cancer at a young age (before age 60 years). Whole exome sequencing identified a panel of germline variants that have

Blood parasites shape extreme major histocompatibility complex diversity in a migratory passerine.

PubMed

Biedrzycka, Aleksandra; Bielański, Wojciech; Ćmiel, Adam; Solarz, Wojciech; Zając, Tadeusz; Migalska, Magdalena; Sebastian, Alvaro; Westerdahl, Helena; Radwan, Jacek

2018-06-01

Pathogens are one of the main forces driving the evolution and maintenance of the highly polymorphic genes of the vertebrate major histocompatibility complex (MHC). Although MHC proteins are crucial in pathogen recognition, it is still poorly understood how pathogen-mediated selection promotes and maintains MHC diversity, and especially so in host species with highly duplicated MHC genes. Sedge warblers (Acrocephalus schoenobaenus) have highly duplicated MHC genes, and using data from high-throughput MHC genotyping, we were able to investigate to what extent avian malaria parasites explain temporal MHC class I supertype fluctuations in a long-term study population. We investigated infection status and infection intensities of two different strains of Haemoproteus, that is avian malaria parasites that are known to have significant fitness consequences in sedge warblers. We found that prevalence of avian malaria in carriers of specific MHC class I supertypes was a significant predictor of their frequency changes between years. This finding suggests that avian malaria infections partly drive the temporal fluctuations of the MHC class I supertypes. Furthermore, we found that individuals with a large number of different supertypes had higher resistance to avian malaria, but there was no evidence for an optimal MHC class I diversity. Thus, the two studied malaria parasite strains appear to select for a high MHC class I supertype diversity. Such selection may explain the maintenance of the extremely high number of MHC class I gene copies in sedge warblers and possibly also in other passerines where avian malaria is a common disease. © 2018 John Wiley & Sons Ltd.

Combining high-throughput MALDI-TOF mass spectrometry and isoelectric focusing gel electrophoresis for virtual 2D gel-based proteomics.

PubMed

Lohnes, Karen; Quebbemann, Neil R; Liu, Kate; Kobzeff, Fred; Loo, Joseph A; Ogorzalek Loo, Rachel R

2016-07-15

The virtual two-dimensional gel electrophoresis/mass spectrometry (virtual 2D gel/MS) technology combines the premier, high-resolution capabilities of 2D gel electrophoresis with the sensitivity and high mass accuracy of mass spectrometry (MS). Intact proteins separated by isoelectric focusing (IEF) gel electrophoresis are imaged from immobilized pH gradient (IPG) polyacrylamide gels (the first dimension of classic 2D-PAGE) by matrix-assisted laser desorption/ionization (MALDI) MS. Obtaining accurate intact masses from sub-picomole-level proteins embedded in 2D-PAGE gels or in IPG strips is desirable to elucidate how the protein of one spot identified as protein 'A' on a 2D gel differs from the protein of another spot identified as the same protein, whenever tryptic peptide maps fail to resolve the issue. This task, however, has been extremely challenging. Virtual 2D gel/MS provides access to these intact masses. Modifications to our matrix deposition procedure improve the reliability with which IPG gels can be prepared; the new procedure is described. Development of this MALDI MS imaging (MSI) method for high-throughput MS with integrated 'top-down' MS to elucidate protein isoforms from complex biological samples is described and it is demonstrated that a 4-cm IPG gel segment can now be imaged in approximately 5min. Gel-wide chemical and enzymatic methods with further interrogation by MALDI MS/MS provide identifications, sequence-related information, and post-translational/transcriptional modification information. The MSI-based virtual 2D gel/MS platform may potentially link the benefits of 'top-down' and 'bottom-up' proteomics. Copyright © 2016 Elsevier Inc. All rights reserved.

Self-leveling 2D DPN probe arrays

NASA Astrophysics Data System (ADS)

Haaheim, Jason R.; Val, Vadim; Solheim, Ed; Bussan, John; Fragala, J.; Nelson, Mike

2010-02-01

Dip Pen Nanolithography® (DPN®) is a direct write scanning probe-based technique which operates under ambient conditions, making it suitable to deposit a wide range of biological and inorganic materials. Precision nanoscale deposition is a fundamental requirement to advance nanoscale technology in commercial applications, and tailoring chemical composition and surface structure on the sub-100 nm scale benefits researchers in areas ranging from cell adhesion to cell-signaling and biomimetic membranes. These capabilities naturally suggest a "Desktop Nanofab" concept - a turnkey system that allows a non-expert user to rapidly create high resolution, scalable nanostructures drawing upon well-characterized ink and substrate pairings. In turn, this system is fundamentally supported by a portfolio of MEMS devices tailored for microfluidic ink delivery, directed placement of nanoscale materials, and cm2 tip arrays for high-throughput nanofabrication. Massively parallel two-dimensional nanopatterning is now commercially available via NanoInk's 2D nano PrintArray™, making DPN a high-throughput (>3×107 μm2 per hour), flexible and versatile method for precision nanoscale pattern formation. However, cm2 arrays of nanoscopic tips introduce the nontrivial problem of getting them all evenly touching the surface to ensure homogeneous deposition; this requires extremely precise leveling of the array. Herein, we describe how we have made the process simple by way of a selfleveling gimbal attachment, coupled with semi-automated software leveling routines which bring the cm^2 chip to within 0.002 degrees of co-planarity. This excellent co-planarity yields highly homogeneous features across a square centimeter, with <6% feature size standard deviation. We have engineered the devices to be easy to use, wire-free, and fully integrated with both of our patterning tools: the DPN 5000, and the NLP 2000.

'Enzyme Test Bench': A biochemical application of the multi-rate modeling

NASA Astrophysics Data System (ADS)

Rachinskiy, K.; Schultze, H.; Boy, M.; Büchs, J.

2008-11-01

In the expanding field of 'white biotechnology' enzymes are frequently applied to catalyze the biochemical reaction from a resource material to a valuable product. Evolutionary designed to catalyze the metabolism in any life form, they selectively accelerate complex reactions under physiological conditions. Modern techniques, such as directed evolution, have been developed to satisfy the increasing demand on enzymes. Applying these techniques together with rational protein design, we aim at improving of enzymes' activity, selectivity and stability. To tap the full potential of these techniques, it is essential to combine them with adequate screening methods. Nowadays a great number of high throughput colorimetric and fluorescent enzyme assays are applied to measure the initial enzyme activity with high throughput. However, the prediction of enzyme long term stability within short experiments is still a challenge. A new high throughput technique for enzyme characterization with specific attention to the long term stability, called 'Enzyme Test Bench', is presented. The concept of the Enzyme Test Bench consists of short term enzyme tests conducted under partly extreme conditions to predict the enzyme long term stability under moderate conditions. The technique is based on the mathematical modeling of temperature dependent enzyme activation and deactivation. Adapting the temperature profiles in sequential experiments by optimum non-linear experimental design, the long term deactivation effects can be purposefully accelerated and detected within hours. During the experiment the enzyme activity is measured online to estimate the model parameters from the obtained data. Thus, the enzyme activity and long term stability can be calculated as a function of temperature. The results of the characterization, based on micro liter format experiments of hours, are in good agreement with the results of long term experiments in 1L format. Thus, the new technique allows for both: the enzyme screening with regard to the long term stability and the choice of the optimal process temperature. The presented article gives a successful example for the application of multi-rate modeling, experimental design and parameter estimation within biochemical engineering. At the same time, it shows the limitations of the methods at the state of the art and addresses the current problems to the applied mathematics community.

High-throughput sequencing methods to study neuronal RNA-protein interactions.

PubMed

Ule, Jernej

2009-12-01

UV-cross-linking and RNase protection, combined with high-throughput sequencing, have provided global maps of RNA sites bound by individual proteins or ribosomes. Using a stringent purification protocol, UV-CLIP (UV-cross-linking and immunoprecipitation) was able to identify intronic and exonic sites bound by splicing regulators in mouse brain tissue. Ribosome profiling has been used to quantify ribosome density on budding yeast mRNAs under different environmental conditions. Post-transcriptional regulation in neurons requires high spatial and temporal precision, as is evident from the role of localized translational control in synaptic plasticity. It remains to be seen if the high-throughput methods can be applied quantitatively to study the dynamics of RNP (ribonucleoprotein) remodelling in specific neuronal populations during the neurodegenerative process. It is certain, however, that applications of new biochemical techniques followed by high-throughput sequencing will continue to provide important insights into the mechanisms of neuronal post-transcriptional regulation.

Lessons learned in the analysis of high-dimensional data in vaccinomics

PubMed Central

Oberg, Ann L.; McKinney, Brett A.; Schaid, Daniel J.; Pankratz, V. Shane; Kennedy, Richard B.; Poland, Gregory A.

2015-01-01

The field of vaccinology is increasingly moving toward the generation, analysis, and modeling of extremely large and complex high-dimensional datasets. We have used data such as these in the development and advancement of the field of vaccinomics to enable prediction of vaccine responses and to develop new vaccine candidates. However, the application of systems biology to what has been termed “big data,” or “high-dimensional data,” is not without significant challenges—chief among them a paucity of gold standard analysis and modeling paradigms with which to interpret the data. In this article, we relate some of the lessons we have learned over the last decade of working with high-dimensional, high-throughput data as applied to the field of vaccinomics. The value of such efforts, however, is ultimately to better understand the immune mechanisms by which protective and non-protective responses to vaccines are generated, and to use this information to support a personalized vaccinology approach in creating better, and safer, vaccines for the public health. PMID:25957070

Lessons learned in the analysis of high-dimensional data in vaccinomics.

PubMed

Oberg, Ann L; McKinney, Brett A; Schaid, Daniel J; Pankratz, V Shane; Kennedy, Richard B; Poland, Gregory A

2015-09-29

The field of vaccinology is increasingly moving toward the generation, analysis, and modeling of extremely large and complex high-dimensional datasets. We have used data such as these in the development and advancement of the field of vaccinomics to enable prediction of vaccine responses and to develop new vaccine candidates. However, the application of systems biology to what has been termed "big data," or "high-dimensional data," is not without significant challenges-chief among them a paucity of gold standard analysis and modeling paradigms with which to interpret the data. In this article, we relate some of the lessons we have learned over the last decade of working with high-dimensional, high-throughput data as applied to the field of vaccinomics. The value of such efforts, however, is ultimately to better understand the immune mechanisms by which protective and non-protective responses to vaccines are generated, and to use this information to support a personalized vaccinology approach in creating better, and safer, vaccines for the public health. Copyright © 2015 Elsevier Ltd. All rights reserved.

High-speed single-photon signaling for daytime QKD

NASA Astrophysics Data System (ADS)

Bienfang, Joshua; Restelli, Alessandro; Clark, Charles

2011-03-01

The distribution of quantum-generated cryptographic key at high throughputs can be critically limited by the performance of the systems' single-photon detectors. While noise and afterpulsing are considerations for all single-photon QKD systems, high-transmission rate systems also have critical detector timing-resolution and recovery time requirements. We present experimental results exploiting the high timing resolution and count-rate stability of modified single-photon avalanche diodes (SPADs) in our GHz QKD system operating over a 1.5 km free-space link that demonstrate the ability to apply extremely short temporal gates, enabling daytime free-space QKD with a 4% QBER. We also discuss recent advances in gating techniques for InGaAs SPADs that are suitable for high-speed fiber-based QKD. We present afterpulse-probability measurements that demonstrate the ability to support single-photon count rates above 100 MHz with low afterpulse probability. These results will benefit the design and characterization of free-space and fiber QKD systems. A. Restelli, J.C. Bienfang A. Mink, and C.W. Clark, IEEE J. Sel. Topics in Quant. Electron 16, 1084 (2010).

Evaluation of Compatibility of ToxCast High-Throughput/High-Content Screening Assays with Engineered Nanomaterials

EPA Science Inventory

High-throughput and high-content screens are attractive approaches for prioritizing nanomaterial hazards and informing targeted testing due to the impracticality of using traditional toxicological testing on the large numbers and varieties of nanomaterials. The ToxCast program a...

Droplet-based microfluidic high-throughput screening of heterologous enzymes secreted by the yeast Yarrowia lipolytica.

PubMed

Beneyton, Thomas; Thomas, Stéphane; Griffiths, Andrew D; Nicaud, Jean-Marc; Drevelle, Antoine; Rossignol, Tristan

2017-01-31

Droplet-based microfluidics is becoming an increasingly attractive alternative to microtiter plate techniques for enzymatic high-throughput screening (HTS), especially for exploring large diversities with lower time and cost footprint. In this case, the assayed enzyme has to be accessible to the substrate within the water-in-oil droplet by being ideally extracellular or displayed at the cell surface. However, most of the enzymes screened to date are expressed within the cytoplasm of Escherichia coli cells, which means that a lysis step must take place inside the droplets for enzyme activity to be assayed. Here, we take advantage of the excellent secretion abilities of the yeast Yarrowia lipolytica to describe a highly efficient expression system particularly suitable for the droplet-based microfluidic HTS. Five hydrolytic genes from Aspergillus niger genome were chosen and the corresponding five Yarrowia lipolytica producing strains were constructed. Each enzyme (endo-β-1,4-xylanase B and C; 1,4-β-cellobiohydrolase A; endoglucanase A; aspartic protease) was successfully overexpressed and secreted in an active form in the crude supernatant. A droplet-based microfluidic HTS system was developed to (a) encapsulate single yeast cells; (b) grow yeast in droplets; (c) inject the relevant enzymatic substrate; (d) incubate droplets on chip; (e) detect enzymatic activity; and (f) sort droplets based on enzymatic activity. Combining this integrated microfluidic platform with gene expression in Y. lipolytica results in remarkably low variability in the enzymatic activity at the single cell level within a given monoclonal population (<5%). Xylanase, cellobiohydrolase and protease activities were successfully assayed using this system. We then used the system to screen for thermostable variants of endo-β-1,4-xylanase C in error-prone PCR libraries. Variants displaying higher thermostable xylanase activities compared to the wild-type were isolated (up to 4.7-fold improvement). Yarrowia lipolytica was used to express fungal genes encoding hydrolytic enzymes of interest. We developed a successful droplet-based microfluidic platform for the high-throughput screening (10 5 strains/h) of Y. lipolytica based on enzyme secretion and activity. This approach provides highly efficient tools for the HTS of recombinant enzymatic activities. This should be extremely useful for discovering new biocatalysts via directed evolution or protein engineering approaches and should lead to major advances in microbial cell factory development.

SwellGel: an affinity chromatography technology for high-capacity and high-throughput purification of recombinant-tagged proteins.

PubMed

Draveling, C; Ren, L; Haney, P; Zeisse, D; Qoronfleh, M W

2001-07-01

The revolution in genomics and proteomics is having a profound impact on drug discovery. Today's protein scientist demands a faster, easier, more reliable way to purify proteins. A high capacity, high-throughput new technology has been developed in Perbio Sciences for affinity protein purification. This technology utilizes selected chromatography media that are dehydrated to form uniform aggregates. The SwellGel aggregates will instantly rehydrate upon addition of the protein sample, allowing purification and direct performance of multiple assays in a variety of formats. SwellGel technology has greater stability and is easier to handle than standard wet chromatography resins. The microplate format of this technology provides high-capacity, high-throughput features, recovering milligram quantities of protein suitable for high-throughput screening or biophysical/structural studies. Data will be presented applying SwellGel technology to recombinant 6x His-tagged protein and glutathione-S-transferase (GST) fusion protein purification. Copyright 2001 Academic Press.

Large-scale microfluidics providing high-resolution and high-throughput screening of Caenorhabditis elegans poly-glutamine aggregation model

NASA Astrophysics Data System (ADS)

Mondal, Sudip; Hegarty, Evan; Martin, Chris; Gökçe, Sertan Kutal; Ghorashian, Navid; Ben-Yakar, Adela

2016-10-01

Next generation drug screening could benefit greatly from in vivo studies, using small animal models such as Caenorhabditis elegans for hit identification and lead optimization. Current in vivo assays can operate either at low throughput with high resolution or with low resolution at high throughput. To enable both high-throughput and high-resolution imaging of C. elegans, we developed an automated microfluidic platform. This platform can image 15 z-stacks of ~4,000 C. elegans from 96 different populations using a large-scale chip with a micron resolution in 16 min. Using this platform, we screened ~100,000 animals of the poly-glutamine aggregation model on 25 chips. We tested the efficacy of ~1,000 FDA-approved drugs in improving the aggregation phenotype of the model and identified four confirmed hits. This robust platform now enables high-content screening of various C. elegans disease models at the speed and cost of in vitro cell-based assays.

ToxCast Dashboard

EPA Pesticide Factsheets

The ToxCast Dashboard helps users examine high-throughput assay data to inform chemical safety decisions. To date, it has data on over 9,000 chemicals and information from more than 1,000 high-throughput assay endpoint components.

RapidTox Dashboard

EPA Pesticide Factsheets

The ToxCast Dashboard helps users examine high-throughput assay data to inform chemical safety decisions. To date, it has data on over 9,000 chemicals and information from more than 1,000 high-throughput assay endpoint components.

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

PubMed Central

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

A high-quality annotated transcriptome of swine peripheral blood

USDA-ARS?s Scientific Manuscript database

Background: High throughput gene expression profiling assays of peripheral blood are widely used in biomedicine, as well as in animal genetics and physiology research. Accurate, comprehensive, and precise interpretation of such high throughput assays relies on well-characterized reference genomes an...

Industrializing electrophysiology: HT automated patch clamp on SyncroPatch® 96 using instant frozen cells.

PubMed

Polonchuk, Liudmila

2014-01-01

Patch-clamping is a powerful technique for investigating the ion channel function and regulation. However, its low throughput hampered profiling of large compound series in early drug development. Fortunately, automation has revolutionized the area of experimental electrophysiology over the past decade. Whereas the first automated patch-clamp instruments using the planar patch-clamp technology demonstrated rather a moderate throughput, few second-generation automated platforms recently launched by various companies have significantly increased ability to form a high number of high-resistance seals. Among them is SyncroPatch(®) 96 (Nanion Technologies GmbH, Munich, Germany), a fully automated giga-seal patch-clamp system with the highest throughput on the market. By recording from up to 96 cells simultaneously, the SyncroPatch(®) 96 allows to substantially increase throughput without compromising data quality. This chapter describes features of the innovative automated electrophysiology system and protocols used for a successful transfer of the established hERG assay to this high-throughput automated platform.

HIGH THROUGHPUT ASSESSMENTS OF CONVENTIONAL AND ALTERNATIVE COMPOUNDS

EPA Science Inventory

High throughput approaches for quantifying chemical hazard, exposure, and sustainability have the potential to dramatically impact the pace and nature of risk assessments. Integrated evaluation strategies developed at the US EPA incorporate inherency,bioactivity,bioavailability, ...

GiNA, an efficient and high-throughput software for horticultural phenotyping

USDA-ARS?s Scientific Manuscript database

Traditional methods for trait phenotyping have been a bottleneck for research in many crop species due to their intensive labor, high cost, complex implementation, lack of reproducibility and propensity to subjective bias. Recently, multiple high-throughput phenotyping platforms have been developed,...

High-throughput quantification of hydroxyproline for determination of collagen.

PubMed

Hofman, Kathleen; Hall, Bronwyn; Cleaver, Helen; Marshall, Susan

2011-10-15

An accurate and high-throughput assay for collagen is essential for collagen research and development of collagen products. Hydroxyproline is routinely assayed to provide a measurement for collagen quantification. The time required for sample preparation using acid hydrolysis and neutralization prior to assay is what limits the current method for determining hydroxyproline. This work describes the conditions of alkali hydrolysis that, when combined with the colorimetric assay defined by Woessner, provide a high-throughput, accurate method for the measurement of hydroxyproline. Copyright © 2011 Elsevier Inc. All rights reserved.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wall, Andrew J.; Capo, Rosemary C.; Stewart, Brian W.

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.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hakala, Jacqueline Alexandra

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.

A Memory Efficient Network Encryption Scheme

NASA Astrophysics Data System (ADS)

El-Fotouh, Mohamed Abo; Diepold, Klaus

In this paper, we studied the two widely used encryption schemes in network applications. Shortcomings have been found in both schemes, as these schemes consume either more memory to gain high throughput or low memory with low throughput. The need has aroused for a scheme that has low memory requirements and in the same time possesses high speed, as the number of the internet users increases each day. We used the SSM model [1], to construct an encryption scheme based on the AES. The proposed scheme possesses high throughput together with low memory requirements.

HTP-NLP: A New NLP System for High Throughput Phenotyping.

PubMed

Schlegel, Daniel R; Crowner, Chris; Lehoullier, Frank; Elkin, Peter L

2017-01-01

Secondary use of clinical data for research requires a method to quickly process the data so that researchers can quickly extract cohorts. We present two advances in the High Throughput Phenotyping NLP system which support the aim of truly high throughput processing of clinical data, inspired by a characterization of the linguistic properties of such data. Semantic indexing to store and generalize partially-processed results and the use of compositional expressions for ungrammatical text are discussed, along with a set of initial timing results for the system.

Combined Effect of Random Transmit Power Control and Inter-Path Interference Cancellation on DS-CDMA Packet Mobile Communications

NASA Astrophysics Data System (ADS)

Kudoh, Eisuke; Ito, Haruki; Wang, Zhisen; Adachi, Fumiyuki

In mobile communication systems, high speed packet data services are demanded. In the high speed data transmission, throughput degrades severely due to severe inter-path interference (IPI). Recently, we proposed a random transmit power control (TPC) to increase the uplink throughput of DS-CDMA packet mobile communications. In this paper, we apply IPI cancellation in addition to the random TPC. We derive the numerical expression of the received signal-to-interference plus noise power ratio (SINR) and introduce IPI cancellation factor. We also derive the numerical expression of system throughput when IPI is cancelled ideally to compare with the Monte Carlo numerically evaluated system throughput. Then we evaluate, by Monte-Carlo numerical computation method, the combined effect of random TPC and IPI cancellation on the uplink throughput of DS-CDMA packet mobile communications.

Evaluating Rapid Models for High-Throughput Exposure Forecasting (SOT)

EPA Science Inventory

High throughput exposure screening models can provide quantitative predictions for thousands of chemicals; however these predictions must be systematically evaluated for predictive ability. Without the capability to make quantitative, albeit uncertain, forecasts of exposure, the ...

Solar fuels photoanode materials discovery by integrating high-throughput theory and experiment

DOE PAGES

Yan, Qimin; Yu, Jie; Suram, Santosh K.; ...

2017-03-06

The limited number of known low-band-gap photoelectrocatalytic materials poses a significant challenge for the generation of chemical fuels from sunlight. Here, using high-throughput ab initio theory with experiments in an integrated workflow, we find eight ternary vanadate oxide photoanodes in the target band-gap range (1.2-2.8 eV). Detailed analysis of these vanadate compounds reveals the key role of VO 4 structural motifs and electronic band-edge character in efficient photoanodes, initiating a genome for such materials and paving the way for a broadly applicable high-throughput-discovery and materials-by-design feedback loop. Considerably expanding the number of known photoelectrocatalysts for water oxidation, our study establishesmore » ternary metal vanadates as a prolific class of photoanodematerials for generation of chemical fuels from sunlight and demonstrates our high-throughput theory-experiment pipeline as a prolific approach to materials discovery.« less

Microfluidics for cell-based high throughput screening platforms - A review.

PubMed

Du, Guansheng; Fang, Qun; den Toonder, Jaap M J

2016-01-15

In the last decades, the basic techniques of microfluidics for the study of cells such as cell culture, cell separation, and cell lysis, have been well developed. Based on cell handling techniques, microfluidics has been widely applied in the field of PCR (Polymerase Chain Reaction), immunoassays, organ-on-chip, stem cell research, and analysis and identification of circulating tumor cells. As a major step in drug discovery, high-throughput screening allows rapid analysis of thousands of chemical, biochemical, genetic or pharmacological tests in parallel. In this review, we summarize the application of microfluidics in cell-based high throughput screening. The screening methods mentioned in this paper include approaches using the perfusion flow mode, the droplet mode, and the microarray mode. We also discuss the future development of microfluidic based high throughput screening platform for drug discovery. Copyright © 2015 Elsevier B.V. All rights reserved.

Solar fuels photoanode materials discovery by integrating high-throughput theory and experiment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yan, Qimin; Yu, Jie; Suram, Santosh K.

The limited number of known low-band-gap photoelectrocatalytic materials poses a significant challenge for the generation of chemical fuels from sunlight. Here, using high-throughput ab initio theory with experiments in an integrated workflow, we find eight ternary vanadate oxide photoanodes in the target band-gap range (1.2-2.8 eV). Detailed analysis of these vanadate compounds reveals the key role of VO 4 structural motifs and electronic band-edge character in efficient photoanodes, initiating a genome for such materials and paving the way for a broadly applicable high-throughput-discovery and materials-by-design feedback loop. Considerably expanding the number of known photoelectrocatalysts for water oxidation, our study establishesmore » ternary metal vanadates as a prolific class of photoanodematerials for generation of chemical fuels from sunlight and demonstrates our high-throughput theory-experiment pipeline as a prolific approach to materials discovery.« less

Fulfilling the promise of the materials genome initiative with high-throughput experimental methodologies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Green, Martin L.; Choi, C. L.; Hattrick-Simpers, J. R.

The Materials Genome Initiative, a national effort to introduce new materials into the market faster and at lower cost, has made significant progress in computational simulation and modeling of materials. To build on this progress, a large amount of experimental data for validating these models, and informing more sophisticated ones, will be required. High-throughput experimentation generates large volumes of experimental data using combinatorial materials synthesis and rapid measurement techniques, making it an ideal experimental complement to bring the Materials Genome Initiative vision to fruition. This paper reviews the state-of-the-art results, opportunities, and challenges in high-throughput experimentation for materials design. Asmore » a result, a major conclusion is that an effort to deploy a federated network of high-throughput experimental (synthesis and characterization) tools, which are integrated with a modern materials data infrastructure, is needed.« less

Development and Validation of an Automated High-Throughput System for Zebrafish In Vivo Screenings

PubMed Central

Virto, Juan M.; Holgado, Olaia; Diez, Maria; Izpisua Belmonte, Juan Carlos; Callol-Massot, Carles

2012-01-01

The zebrafish is a vertebrate model compatible with the paradigms of drug discovery. The small size and transparency of zebrafish embryos make them amenable for the automation necessary in high-throughput screenings. We have developed an automated high-throughput platform for in vivo chemical screenings on zebrafish embryos that includes automated methods for embryo dispensation, compound delivery, incubation, imaging and analysis of the results. At present, two different assays to detect cardiotoxic compounds and angiogenesis inhibitors can be automatically run in the platform, showing the versatility of the system. A validation of these two assays with known positive and negative compounds, as well as a screening for the detection of unknown anti-angiogenic compounds, have been successfully carried out in the system developed. We present a totally automated platform that allows for high-throughput screenings in a vertebrate organism. PMID:22615792

BiQ Analyzer HT: locus-specific analysis of DNA methylation by high-throughput bisulfite sequencing

PubMed Central

Lutsik, Pavlo; Feuerbach, Lars; Arand, Julia; Lengauer, Thomas; Walter, Jörn; Bock, Christoph

2011-01-01

Bisulfite sequencing is a widely used method for measuring DNA methylation in eukaryotic genomes. The assay provides single-base pair resolution and, given sufficient sequencing depth, its quantitative accuracy is excellent. High-throughput sequencing of bisulfite-converted DNA can be applied either genome wide or targeted to a defined set of genomic loci (e.g. using locus-specific PCR primers or DNA capture probes). Here, we describe BiQ Analyzer HT (http://biq-analyzer-ht.bioinf.mpi-inf.mpg.de/), a user-friendly software tool that supports locus-specific analysis and visualization of high-throughput bisulfite sequencing data. The software facilitates the shift from time-consuming clonal bisulfite sequencing to the more quantitative and cost-efficient use of high-throughput sequencing for studying locus-specific DNA methylation patterns. In addition, it is useful for locus-specific visualization of genome-wide bisulfite sequencing data. PMID:21565797

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

PubMed

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.

Fulfilling the promise of the materials genome initiative with high-throughput experimental methodologies

DOE PAGES

Green, Martin L.; Choi, C. L.; Hattrick-Simpers, J. R.; ...

2017-03-28

The Materials Genome Initiative, a national effort to introduce new materials into the market faster and at lower cost, has made significant progress in computational simulation and modeling of materials. To build on this progress, a large amount of experimental data for validating these models, and informing more sophisticated ones, will be required. High-throughput experimentation generates large volumes of experimental data using combinatorial materials synthesis and rapid measurement techniques, making it an ideal experimental complement to bring the Materials Genome Initiative vision to fruition. This paper reviews the state-of-the-art results, opportunities, and challenges in high-throughput experimentation for materials design. Asmore » a result, a major conclusion is that an effort to deploy a federated network of high-throughput experimental (synthesis and characterization) tools, which are integrated with a modern materials data infrastructure, is needed.« less

High Throughput Genotoxicity Profiling of the US EPA ToxCast Chemical Library

EPA Science Inventory

A key aim of the ToxCast project is to investigate modern molecular and genetic high content and high throughput screening (HTS) assays, along with various computational tools to supplement and perhaps replace traditional assays for evaluating chemical toxicity. Genotoxicity is a...

Stepping into the omics era: Opportunities and challenges for biomaterials science and engineering☆

PubMed Central

Rabitz, Herschel; Welsh, William J.; Kohn, Joachim; de Boer, Jan

2016-01-01

The research paradigm in biomaterials science and engineering is evolving from using low-throughput and iterative experimental designs towards high-throughput experimental designs for materials optimization and the evaluation of materials properties. Computational science plays an important role in this transition. With the emergence of the omics approach in the biomaterials field, referred to as materiomics, high-throughput approaches hold the promise of tackling the complexity of materials and understanding correlations between material properties and their effects on complex biological systems. The intrinsic complexity of biological systems is an important factor that is often oversimplified when characterizing biological responses to materials and establishing property-activity relationships. Indeed, in vitro tests designed to predict in vivo performance of a given biomaterial are largely lacking as we are not able to capture the biological complexity of whole tissues in an in vitro model. In this opinion paper, we explain how we reached our opinion that converging genomics and materiomics into a new field would enable a significant acceleration of the development of new and improved medical devices. The use of computational modeling to correlate high-throughput gene expression profiling with high throughput combinatorial material design strategies would add power to the analysis of biological effects induced by material properties. We believe that this extra layer of complexity on top of high-throughput material experimentation is necessary to tackle the biological complexity and further advance the biomaterials field. PMID:26876875

A high-throughput next-generation sequencing-based method for detecting the mutational fingerprint of carcinogens

PubMed Central

Besaratinia, Ahmad; Li, Haiqing; Yoon, Jae-In; Zheng, Albert; Gao, Hanlin; Tommasi, Stella

2012-01-01

Many carcinogens leave a unique mutational fingerprint in the human genome. These mutational fingerprints manifest as specific types of mutations often clustering at certain genomic loci in tumor genomes from carcinogen-exposed individuals. To develop a high-throughput method for detecting the mutational fingerprint of carcinogens, we have devised a cost-, time- and labor-effective strategy, in which the widely used transgenic Big Blue® mouse mutation detection assay is made compatible with the Roche/454 Genome Sequencer FLX Titanium next-generation sequencing technology. As proof of principle, we have used this novel method to establish the mutational fingerprints of three prominent carcinogens with varying mutagenic potencies, including sunlight ultraviolet radiation, 4-aminobiphenyl and secondhand smoke that are known to be strong, moderate and weak mutagens, respectively. For verification purposes, we have compared the mutational fingerprints of these carcinogens obtained by our newly developed method with those obtained by parallel analyses using the conventional low-throughput approach, that is, standard mutation detection assay followed by direct DNA sequencing using a capillary DNA sequencer. We demonstrate that this high-throughput next-generation sequencing-based method is highly specific and sensitive to detect the mutational fingerprints of the tested carcinogens. The method is reproducible, and its accuracy is comparable with that of the currently available low-throughput method. In conclusion, this novel method has the potential to move the field of carcinogenesis forward by allowing high-throughput analysis of mutations induced by endogenous and/or exogenous genotoxic agents. PMID:22735701

A high-throughput next-generation sequencing-based method for detecting the mutational fingerprint of carcinogens.

PubMed

Besaratinia, Ahmad; Li, Haiqing; Yoon, Jae-In; Zheng, Albert; Gao, Hanlin; Tommasi, Stella

2012-08-01

Many carcinogens leave a unique mutational fingerprint in the human genome. These mutational fingerprints manifest as specific types of mutations often clustering at certain genomic loci in tumor genomes from carcinogen-exposed individuals. To develop a high-throughput method for detecting the mutational fingerprint of carcinogens, we have devised a cost-, time- and labor-effective strategy, in which the widely used transgenic Big Blue mouse mutation detection assay is made compatible with the Roche/454 Genome Sequencer FLX Titanium next-generation sequencing technology. As proof of principle, we have used this novel method to establish the mutational fingerprints of three prominent carcinogens with varying mutagenic potencies, including sunlight ultraviolet radiation, 4-aminobiphenyl and secondhand smoke that are known to be strong, moderate and weak mutagens, respectively. For verification purposes, we have compared the mutational fingerprints of these carcinogens obtained by our newly developed method with those obtained by parallel analyses using the conventional low-throughput approach, that is, standard mutation detection assay followed by direct DNA sequencing using a capillary DNA sequencer. We demonstrate that this high-throughput next-generation sequencing-based method is highly specific and sensitive to detect the mutational fingerprints of the tested carcinogens. The method is reproducible, and its accuracy is comparable with that of the currently available low-throughput method. In conclusion, this novel method has the potential to move the field of carcinogenesis forward by allowing high-throughput analysis of mutations induced by endogenous and/or exogenous genotoxic agents.

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

PubMed

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.

Expedient Caution: Approximating Exposure and Dosimetry to Understand Chemical Risk (OSU EMT Research Day keynote presentation)

EPA Science Inventory

I describe research on high throughput exposure and toxicokinetics. These tools provide context for data generated by high throughput toxicity screening to allow risk-based prioritization of thousands of chemicals.

MIPHENO: Data normalization for high throughput metabolic analysis.

EPA Science Inventory

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...

High-Throughput Pharmacokinetics for Environmental Chemicals (SOT)

EPA Science Inventory

High throughput screening (HTS) promises to allow prioritization of thousands of environmental chemicals with little or no in vivo information. For bioactivity identified by HTS, toxicokinetic (TK) models are essential to predict exposure thresholds below which no significant bio...

High Throughput Computing Impact on Meta Genomics (Metagenomics Informatics Challenges Workshop: 10K Genomes at a Time)

ScienceCinema

Gore, Brooklin

2018-02-01

This presentation includes a brief background on High Throughput Computing, correlating gene transcription factors, optical mapping, genotype to phenotype mapping via QTL analysis, and current work on next gen sequencing.

Establishment of integrated protocols for automated high throughput kinetic chlorophyll fluorescence analyses.

PubMed

Tschiersch, Henning; Junker, Astrid; Meyer, Rhonda C; Altmann, Thomas

2017-01-01

Automated plant phenotyping has been established as a powerful new tool in studying plant growth, development and response to various types of biotic or abiotic stressors. Respective facilities mainly apply non-invasive imaging based methods, which enable the continuous quantification of the dynamics of plant growth and physiology during developmental progression. However, especially for plants of larger size, integrative, automated and high throughput measurements of complex physiological parameters such as photosystem II efficiency determined through kinetic chlorophyll fluorescence analysis remain a challenge. We present the technical installations and the establishment of experimental procedures that allow the integrated high throughput imaging of all commonly determined PSII parameters for small and large plants using kinetic chlorophyll fluorescence imaging systems (FluorCam, PSI) integrated into automated phenotyping facilities (Scanalyzer, LemnaTec). Besides determination of the maximum PSII efficiency, we focused on implementation of high throughput amenable protocols recording PSII operating efficiency (Φ PSII ). Using the presented setup, this parameter is shown to be reproducibly measured in differently sized plants despite the corresponding variation in distance between plants and light source that caused small differences in incident light intensity. Values of Φ PSII obtained with the automated chlorophyll fluorescence imaging setup correlated very well with conventionally determined data using a spot-measuring chlorophyll fluorometer. The established high throughput operating protocols enable the screening of up to 1080 small and 184 large plants per hour, respectively. The application of the implemented high throughput protocols is demonstrated in screening experiments performed with large Arabidopsis and maize populations assessing natural variation in PSII efficiency. The incorporation of imaging systems suitable for kinetic chlorophyll fluorescence analysis leads to a substantial extension of the feature spectrum to be assessed in the presented high throughput automated plant phenotyping platforms, thus enabling the simultaneous assessment of plant architectural and biomass-related traits and their relations to physiological features such as PSII operating efficiency. The implemented high throughput protocols are applicable to a broad spectrum of model and crop plants of different sizes (up to 1.80 m height) and architectures. The deeper understanding of the relation of plant architecture, biomass formation and photosynthetic efficiency has a great potential with respect to crop and yield improvement strategies.

Identifying drought adaptive traits in upland cotton using a proximal sensing cart for high-throughput phenotyping

USDA-ARS?s Scientific Manuscript database

Field-based high-throughput phenotyping is an emerging approach to characterize difficult, time-sensitive plant traits in relevant growing conditions. Proximal sensing carts have been developed as an alternative platform to more costly high-clearance tractors for phenotyping dynamic traits in the fi...

High-throughput profiling and analysis of plant responses over time to abiotic stress

USDA-ARS?s Scientific Manuscript database

Energy sorghum (Sorghum bicolor (L.) Moench) is a rapidly growing, high-biomass, annual crop prized for abiotic stress tolerance. Measuring genotype-by-environment (G x E) interactions remains a progress bottleneck. High throughput phenotyping within controlled environments has been proposed as a po...

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

EPA Science Inventory

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...

A high-throughput multiplex method adapted for GMO detection.

PubMed

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.

Use of big data in drug development for precision medicine

PubMed Central

Kim, Rosa S.; Goossens, Nicolas; Hoshida, Yujin

2016-01-01

Summary Drug development has been a costly and lengthy process with an extremely low success rate and lack of consideration of individual diversity in drug response and toxicity. Over the past decade, an alternative “big data” approach has been expanding at an unprecedented pace based on the development of electronic databases of chemical substances, disease gene/protein targets, functional readouts, and clinical information covering inter-individual genetic variations and toxicities. This paradigm shift has enabled systematic, high-throughput, and accelerated identification of novel drugs or repurposed indications of existing drugs for pathogenic molecular aberrations specifically present in each individual patient. The exploding interest from the information technology and direct-to-consumer genetic testing industries has been further facilitating the use of big data to achieve personalized Precision Medicine. Here we overview currently available resources and discuss future prospects. PMID:27430024

RIPiT-Seq: A high-throughput approach for footprinting RNA:protein complexes

PubMed Central

Singh, Guramrit; Ricci, Emiliano P.; Moore, Melissa J.

2013-01-01

Development of high-throughput approaches to map the RNA interaction sites of individual RNA binding proteins (RBPs) transcriptome-wide is rapidly transforming our understanding of post-transcriptional gene regulatory mechanisms. Here we describe a ribonucleoprotein (RNP) footprinting approach we recently developed for identifying occupancy sites of both individual RBPs and multi-subunit RNP complexes. RNA:protein immunoprecipitation in tandem (RIPiT) yields highly specific RNA footprints of cellular RNPs isolated via two sequential purifications; the resulting RNA footprints can then be identified by high-throughput sequencing (Seq). RIPiT-Seq is broadly applicable to all RBPs regardless of their RNA binding mode and thus provides a means to map the RNA binding sites of RBPs with poor inherent ultraviolet (UV) crosslinkability. Further, among current high-throughput approaches, RIPiT has the unique capacity to differentiate binding sites of RNPs with overlapping protein composition. It is therefore particularly suited for studying dynamic RNP assemblages whose composition evolves as gene expression proceeds. PMID:24096052

Development and validation of a 48-target analytical method for high-throughput monitoring of genetically modified organisms.

PubMed

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.

Development and Validation of A 48-Target Analytical Method for High-throughput Monitoring of Genetically Modified Organisms

PubMed Central

Li, Xiaofei; Wu, Yuhua; Li, Jun; Li, Yunjing; Long, Likun; Li, Feiwu; Wu, Gang

2015-01-01

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. PMID:25556930

Key Ecological Roles for Zoosporic True Fungi in Aquatic Habitats.

PubMed

Gleason, Frank H; Scholz, Bettina; Jephcott, Thomas G; van Ogtrop, Floris F; Henderson, Linda; Lilje, Osu; Kittelmann, Sandra; Macarthur, Deborah J

2017-03-01

The diversity and abundance of zoosporic true fungi have been analyzed recently using fungal sequence libraries and advances in molecular methods, such as high-throughput sequencing. This review focuses on four evolutionary primitive true fungal phyla: the Aphelidea, Chytridiomycota, Neocallimastigomycota, and Rosellida (Cryptomycota), most species of which are not polycentric or mycelial (filamentous), rather they tend to be primarily monocentric (unicellular). Zoosporic fungi appear to be both abundant and diverse in many aquatic habitats around the world, with abundance often exceeding other fungal phyla in these habitats, and numerous novel genetic sequences identified. Zoosporic fungi are able to survive extreme conditions, such as high and extremely low pH; however, more work remains to be done. They appear to have important ecological roles as saprobes in decomposition of particulate organic substrates, pollen, plant litter, and dead animals; as parasites of zooplankton and algae; as parasites of vertebrate animals (such as frogs); and as symbionts in the digestive tracts of mammals. Some chytrids cause economically important diseases of plants and animals. They regulate sizes of phytoplankton populations. Further metagenomics surveys of aquatic ecosystems are expected to enlarge our knowledge of the diversity of true zoosporic fungi. Coupled with studies on their functional ecology, we are moving closer to unraveling the role of zoosporic fungi in carbon cycling and the impact of climate change on zoosporic fungal populations.

MSP-HTPrimer: a high-throughput primer design tool to improve assay design for DNA methylation analysis in epigenetics.

PubMed

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 strands to increase the success rate. It is a standalone web-based pipeline, which is fully configured within a virtual machine and thus can be readily used without any configuration. We have experimentally validated primer pairs designed by our pipeline and shown a very high success rate of primer pairs: out of 66 BSP primer pairs, 63 were successfully validated without any further optimization step and using the same qPCR conditions. The MSP-HTPrimer pipeline is freely available from http://sourceforge.net/p/msp-htprimer.

High-Throughput Genetic Screen Reveals that Early Attachment and Biofilm Formation Are Necessary for Full Pyoverdine Production by Pseudomonas aeruginosa

PubMed Central

Kang, Donghoon; Kirienko, Natalia V.

2017-01-01

Pseudomonas aeruginosa is a re-emerging, multidrug-resistant, opportunistic pathogen that threatens the lives of immunocompromised patients, patients with cystic fibrosis, and those in critical care units. One of the most important virulence factors in this pathogen is the siderophore pyoverdine. Pyoverdine serves several critical roles during infection. Due to its extremely high affinity for ferric iron, pyoverdine gives the pathogen a significant advantage over the host in their competition for iron. In addition, pyoverdine can regulate the production of multiple bacterial virulence factors and perturb host mitochondrial homeostasis. Inhibition of pyoverdine biosynthesis decreases P. aeruginosa pathogenicity in multiple host models. To better understand the regulation of pyoverdine production, we developed a high-throughput genetic screen that uses the innate fluorescence of pyoverdine to identify genes necessary for its biosynthesis. A substantial number of hits showing severe impairment of pyoverdine production were in genes responsible for early attachment and biofilm formation. In addition to genetic disruption of biofilm, both physical and chemical perturbations also attenuated pyoverdine production. This regulatory relationship between pyoverdine and biofilm is particularly significant in the context of P. aeruginosa multidrug resistance, where the formation of biofilm is a key mechanism preventing access to antimicrobials and the immune system. Furthermore, we demonstrate that the biofilm inhibitor 2-amino-5,6-dimethylbenzimidazole effectively attenuates pyoverdine production and rescues Caenorhabditis elegans from P. aeruginosa-mediated pathogenesis. Our findings suggest that targeting biofilm formation in P. aeruginosa infections may have multiple therapeutic benefits and that employing an unbiased, systems biology-based approach may be useful for understanding the regulation of specific virulence factors and identifying novel anti-virulence therapeutics or new applications for existing therapies for P. aeruginosa infections. PMID:28928729

A high-throughput label-free nanoparticle analyser.

PubMed

Fraikin, Jean-Luc; Teesalu, Tambet; McKenney, Christopher M; Ruoslahti, Erkki; Cleland, Andrew N

2011-05-01

Synthetic nanoparticles and genetically modified viruses are used in a range of applications, but high-throughput analytical tools for the physical characterization of these objects are needed. Here we present a microfluidic analyser that detects individual nanoparticles and characterizes complex, unlabelled nanoparticle suspensions. We demonstrate the detection, concentration analysis and sizing of individual synthetic nanoparticles in a multicomponent mixture with sufficient throughput to analyse 500,000 particles per second. We also report the rapid size and titre analysis of unlabelled bacteriophage T7 in both salt solution and mouse blood plasma, using just ~1 × 10⁻⁶ l of analyte. Unexpectedly, in the native blood plasma we discover a large background of naturally occurring nanoparticles with a power-law size distribution. The high-throughput detection capability, scalable fabrication and simple electronics of this instrument make it well suited for diverse applications.

High Performance Computing Modernization Program Kerberos Throughput Test Report

DTIC Science & Technology

2017-10-26

functionality as Kerberos plugins. The pre -release production kit was used in these tests to compare against the current release kit. YubiKey support...HPCMP Kerberos Throughput Test Report 3 2. THROUGHPUT TESTING 2.1 Testing Components Throughput testing was done to determine the benefits of the pre ...both the current release kit and the pre -release production kit for a total of 378 individual tests in order to note any improvements. Based on work

Metabolomics Approach for Toxicity Screening of Volatile Substances

EPA Science Inventory

In 2007 the National Research Council envisioned the need for inexpensive, high throughput, cell based toxicity testing methods relevant to human health. High Throughput Screening (HTS) in vitro screening approaches have addressed these problems by using robotics. However, the ch...

AOPs & Biomarkers: Bridging High Throughput Screening and Regulatory Decision Making.

EPA Science Inventory

As high throughput screening (HTS) approaches play a larger role in toxicity testing, computational toxicology has emerged as a critical component in interpreting the large volume of data produced. Computational models for this purpose are becoming increasingly more sophisticated...

New High Throughput Methods to Estimate Chemical Exposure

EPA Science Inventory

EPA has made many recent advances in high throughput bioactivity testing. However, concurrent advances in rapid, quantitative prediction of human and ecological exposures have been lacking, despite the clear importance of both measures for a risk-based approach to prioritizing an...

Fully Bayesian Analysis of High-throughput Targeted Metabolomics Assays

EPA Science Inventory

High-throughput metabolomic assays that allow simultaneous targeted screening of hundreds of metabolites have recently become available in kit form. Such assays provide a window into understanding changes to biochemical pathways due to chemical exposure or disease, and are usefu...

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

PubMed

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.

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

NASA Astrophysics Data System (ADS)

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.

High Throughput Determination of Critical Human Dosing ...

EPA Pesticide Factsheets

High throughput toxicokinetics (HTTK) is a rapid approach that uses in vitro data to estimate TK for hundreds of environmental chemicals. Reverse dosimetry (i.e., reverse toxicokinetics or RTK) based on HTTK data converts high throughput in vitro toxicity screening (HTS) data into predicted human equivalent doses that can be linked with biologically relevant exposure scenarios. Thus, HTTK provides essential data for risk prioritization for thousands of chemicals that lack TK data. One critical HTTK parameter that can be measured in vitro is the unbound fraction of a chemical in plasma (Fub). However, for chemicals that bind strongly to plasma, Fub is below the limits of detection (LOD) for high throughput analytical chemistry, and therefore cannot be quantified. A novel method for quantifying Fub was implemented for 85 strategically selected chemicals: measurement of Fub was attempted at 10%, 30%, and 100% of physiological plasma concentrations using rapid equilibrium dialysis assays. Varying plasma concentrations instead of chemical concentrations makes high throughput analytical methodology more likely to be successful. Assays at 100% plasma concentration were unsuccessful for 34 chemicals. For 12 of these 34 chemicals, Fub could be quantified at 10% and/or 30% plasma concentrations; these results imply that the assay failure at 100% plasma concentration was caused by plasma protein binding for these chemicals. Assay failure for the remaining 22 chemicals may

Genome-wide RNAi Screening to Identify Host Factors That Modulate Oncolytic Virus Therapy.

PubMed

Allan, Kristina J; Mahoney, Douglas J; Baird, Stephen D; Lefebvre, Charles A; Stojdl, David F

2018-04-03

High-throughput genome-wide RNAi (RNA interference) screening technology has been widely used for discovering host factors that impact virus replication. Here we present the application of this technology to uncovering host targets that specifically modulate the replication of Maraba virus, an oncolytic rhabdovirus, and vaccinia virus with the goal of enhancing therapy. While the protocol has been tested for use with oncolytic Maraba virus and oncolytic vaccinia virus, this approach is applicable to other oncolytic viruses and can also be utilized for identifying host targets that modulate virus replication in mammalian cells in general. This protocol describes the development and validation of an assay for high-throughput RNAi screening in mammalian cells, the key considerations and preparation steps important for conducting a primary high-throughput RNAi screen, and a step-by-step guide for conducting a primary high-throughput RNAi screen; in addition, it broadly outlines the methods for conducting secondary screen validation and tertiary validation studies. The benefit of high-throughput RNAi screening is that it allows one to catalogue, in an extensive and unbiased fashion, host factors that modulate any aspect of virus replication for which one can develop an in vitro assay such as infectivity, burst size, and cytotoxicity. It has the power to uncover biotherapeutic targets unforeseen based on current knowledge.

X-ray transparent microfluidic chips for high-throughput screening and optimization of in meso membrane protein crystallization

PubMed Central

Schieferstein, Jeremy M.; Pawate, Ashtamurthy S.; Wan, Frank; Sheraden, Paige N.; Broecker, Jana; Ernst, Oliver P.; Gennis, Robert B.

2017-01-01

Elucidating and clarifying the function of membrane proteins ultimately requires atomic resolution structures as determined most commonly by X-ray crystallography. Many high impact membrane protein structures have resulted from advanced techniques such as in meso crystallization that present technical difficulties for the set-up and scale-out of high-throughput crystallization experiments. In prior work, we designed a novel, low-throughput X-ray transparent microfluidic device that automated the mixing of protein and lipid by diffusion for in meso crystallization trials. Here, we report X-ray transparent microfluidic devices for high-throughput crystallization screening and optimization that overcome the limitations of scale and demonstrate their application to the crystallization of several membrane proteins. Two complementary chips are presented: (1) a high-throughput screening chip to test 192 crystallization conditions in parallel using as little as 8 nl of membrane protein per well and (2) a crystallization optimization chip to rapidly optimize preliminary crystallization hits through fine-gradient re-screening. We screened three membrane proteins for new in meso crystallization conditions, identifying several preliminary hits that we tested for X-ray diffraction quality. Further, we identified and optimized the crystallization condition for a photosynthetic reaction center mutant and solved its structure to a resolution of 3.5 Å. PMID:28469762

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

PubMed

Liu, Guangbo; Lanham, Clayton; Buchan, J Ross; Kaplan, Matthew E

2017-01-01

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.

High-Throughput Toxicity Testing: New Strategies for ...

EPA Pesticide Factsheets

In recent years, the food industry has made progress in improving safety testing methods focused on microbial contaminants in order to promote food safety. However, food industry toxicologists must also assess the safety of food-relevant chemicals including pesticides, direct additives, and food contact substances. With the rapidly growing use of new food additives, as well as innovation in food contact substance development, an interest in exploring the use of high-throughput chemical safety testing approaches has emerged. Currently, the field of toxicology is undergoing a paradigm shift in how chemical hazards can be evaluated. Since there are tens of thousands of chemicals in use, many of which have little to no hazard information and there are limited resources (namely time and money) for testing these chemicals, it is necessary to prioritize which chemicals require further safety testing to better protect human health. Advances in biochemistry and computational toxicology have paved the way for animal-free (in vitro) high-throughput screening which can characterize chemical interactions with highly specific biological processes. Screening approaches are not novel; in fact, quantitative high-throughput screening (qHTS) methods that incorporate dose-response evaluation have been widely used in the pharmaceutical industry. For toxicological evaluation and prioritization, it is the throughput as well as the cost- and time-efficient nature of qHTS that makes it

From organized high throughput data to phenomenological theory: The example of dielectric breakdown

NASA Astrophysics Data System (ADS)

Kim, Chiho; Pilania, Ghanshyam; Ramprasad, Rampi

Understanding the behavior (and failure) of dielectric insulators experiencing extreme electric fields is critical to the operation of present and emerging electrical and electronic devices. Despite its importance, the development of a predictive theory of dielectric breakdown has remained a challenge, owing to the complex multiscale nature of this process. Here, we focus on the intrinsic dielectric breakdown field of insulators--the theoretical limit of breakdown determined purely by the chemistry of the material, i.e., the elements the material is composed of, the atomic-level structure, and the bonding. Starting from a benchmark dataset (generated from laborious first principles computations) of the intrinsic dielectric breakdown field of a variety of model insulators, simple predictive phenomenological models of dielectric breakdown are distilled using advanced statistical or machine learning schemes, revealing key correlations and analytical relationships between the breakdown field and easily accessible material properties. The models are shown to be general, and can hence guide the screening and systematic identification of high electric field tolerant materials.

New Python-based methods for data processing

PubMed Central

Sauter, Nicholas K.; Hattne, Johan; Grosse-Kunstleve, Ralf W.; Echols, Nathaniel

2013-01-01

Current pixel-array detectors produce diffraction images at extreme data rates (of up to 2 TB h−1) that make severe demands on computational resources. New multiprocessing frameworks are required to achieve rapid data analysis, as it is important to be able to inspect the data quickly in order to guide the experiment in real time. By utilizing readily available web-serving tools that interact with the Python scripting language, it was possible to implement a high-throughput Bragg-spot analyzer (cctbx.spotfinder) that is presently in use at numerous synchrotron-radiation beamlines. Similarly, Python interoperability enabled the production of a new data-reduction package (cctbx.xfel) for serial femto­second crystallography experiments at the Linac Coherent Light Source (LCLS). Future data-reduction efforts will need to focus on specialized problems such as the treatment of diffraction spots on interleaved lattices arising from multi-crystal specimens. In these challenging cases, accurate modeling of close-lying Bragg spots could benefit from the high-performance computing capabilities of graphics-processing units. PMID:23793153

Reptiles as Reservoirs of Bacterial Infections: Real Threat or Methodological Bias?

PubMed

Zancolli, Giulia; Mahsberg, Dieter; Sickel, Wiebke; Keller, Alexander

2015-10-01

Bacterial infections secondary to snakebites and human pathogens (e.g., Salmonella) have been linked to the oral microbiota of snakes and pet reptiles. Based on culture-dependent studies, it is speculated that snakes' oral microbiota reflects the fecal flora of their ingested preys. However, cultured-based techniques have been shown to be limited as they fail to identify unculturable microorganisms which represent the vast majority of the microbial diversity. Here, we used culture-independent high-throughput sequencing to identify reptile-associated pathogens and to characterize the oral microbial community of five snakes, one gecko, and two terrapins. Few potential human pathogens were detected at extremely low frequencies. Moreover, bacterial taxa represented in the snake's oral cavity bore little resemblance to their preys' fecal microbiota. Overall, we found distinct, highly diverse microbial communities with consistent, species-specific patterns contrary to previous culture-based studies. Our study does not support the widely held assumption that reptiles' oral cavity acts as pathogen reservoir and provides important insights for future research.

Lead discovery for mammalian elongation of long chain fatty acids family 6 using a combination of high-throughput fluorescent-based assay and RapidFire mass spectrometry assay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takamiya, Mari; Discovery Technology Laboratories, Sohyaku, Innovative Research Division, Mitsubishi Tanabe Pharma Corporation, Kawagishi, Toda-shi, Saitama; Sakurai, Masaaki

A high-throughput RapidFire mass spectrometry assay is described for elongation of very long-chain fatty acids family 6 (Elovl6). Elovl6 is a microsomal enzyme that regulates the elongation of C12-16 saturated and monounsaturated fatty acids. Elovl6 may be a new therapeutic target for fat metabolism disorders such as obesity, type 2 diabetes, and nonalcoholic steatohepatitis. To identify new Elovl6 inhibitors, we developed a high-throughput fluorescence screening assay in 1536-well format. However, a number of false positives caused by fluorescent interference have been identified. To pick up the real active compounds among the primary hits from the fluorescence assay, we developed amore » RapidFire mass spectrometry assay and a conventional radioisotope assay. These assays have the advantage of detecting the main products directly without using fluorescent-labeled substrates. As a result, 276 compounds (30%) of the primary hits (921 compounds) in a fluorescence ultra-high-throughput screening method were identified as common active compounds in these two assays. It is concluded that both methods are very effective to eliminate false positives. Compared with the radioisotope method using an expensive {sup 14}C-labeled substrate, the RapidFire mass spectrometry method using unlabeled substrates is a high-accuracy, high-throughput method. In addition, some of the hit compounds selected from the screening inhibited cellular fatty acid elongation in HEK293 cells expressing Elovl6 transiently. This result suggests that these compounds may be promising lead candidates for therapeutic drugs. Ultra-high-throughput fluorescence screening followed by a RapidFire mass spectrometry assay was a suitable strategy for lead discovery against Elovl6. - Highlights: • A novel assay for elongation of very-long-chain fatty acids 6 (Elovl6) is proposed. • RapidFire mass spectrometry (RF-MS) assay is useful to select real screening hits. • RF-MS assay is proved to be beneficial because of its high-throughput and accuracy. • A combination of fluorescent and RF-MS assays is effective for Elovl6 inhibitors.« less

Robust DNA Isolation and High-throughput Sequencing Library Construction for Herbarium Specimens.

PubMed

Saeidi, Saman; McKain, Michael R; Kellogg, Elizabeth A

2018-03-08

Herbaria are an invaluable source of plant material that can be used in a variety of biological studies. The use of herbarium specimens is associated with a number of challenges including sample preservation quality, degraded DNA, and destructive sampling of rare specimens. In order to more effectively use herbarium material in large sequencing projects, a dependable and scalable method of DNA isolation and library preparation is needed. This paper demonstrates a robust, beginning-to-end protocol for DNA isolation and high-throughput library construction from herbarium specimens that does not require modification for individual samples. This protocol is tailored for low quality dried plant material and takes advantage of existing methods by optimizing tissue grinding, modifying library size selection, and introducing an optional reamplification step for low yield libraries. Reamplification of low yield DNA libraries can rescue samples derived from irreplaceable and potentially valuable herbarium specimens, negating the need for additional destructive sampling and without introducing discernible sequencing bias for common phylogenetic applications. The protocol has been tested on hundreds of grass species, but is expected to be adaptable for use in other plant lineages after verification. This protocol can be limited by extremely degraded DNA, where fragments do not exist in the desired size range, and by secondary metabolites present in some plant material that inhibit clean DNA isolation. Overall, this protocol introduces a fast and comprehensive method that allows for DNA isolation and library preparation of 24 samples in less than 13 h, with only 8 h of active hands-on time with minimal modifications.

High Throughput Microplate Respiratory Measurements Using Minimal Quantities Of Isolated Mitochondria

PubMed Central

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

Evaluation of e-liquid toxicity using an open-source high-throughput screening assay

PubMed Central

Keating, James E.; Zorn, Bryan T.; Kochar, Tavleen K.; Wolfgang, Matthew C.; Glish, Gary L.; Tarran, Robert

2018-01-01

The e-liquids used in electronic cigarettes (E-cigs) consist of propylene glycol (PG), vegetable glycerin (VG), nicotine, and chemical additives for flavoring. There are currently over 7,700 e-liquid flavors available, and while some have been tested for toxicity in the laboratory, most have not. Here, we developed a 3-phase, 384-well, plate-based, high-throughput screening (HTS) assay to rapidly triage and validate the toxicity of multiple e-liquids. Our data demonstrated that the PG/VG vehicle adversely affected cell viability and that a large number of e-liquids were more toxic than PG/VG. We also performed gas chromatography–mass spectrometry (GC-MS) analysis on all tested e-liquids. Subsequent nonmetric multidimensional scaling (NMDS) analysis revealed that e-liquids are an extremely heterogeneous group. Furthermore, these data indicated that (i) the more chemicals contained in an e-liquid, the more toxic it was likely to be and (ii) the presence of vanillin was associated with higher toxicity values. Further analysis of common constituents by electron ionization revealed that the concentration of cinnamaldehyde and vanillin, but not triacetin, correlated with toxicity. We have also developed a publicly available searchable website (www.eliquidinfo.org). Given the large numbers of available e-liquids, this website will serve as a resource to facilitate dissemination of this information. Our data suggest that an HTS approach to evaluate the toxicity of multiple e-liquids is feasible. Such an approach may serve as a roadmap to enable bodies such as the Food and Drug Administration (FDA) to better regulate e-liquid composition. PMID:29584716

All solid state mid-infrared dual-comb spectroscopy platform based on QCL technology

NASA Astrophysics Data System (ADS)

Hugi, Andreas; Geiser, Markus; Villares, Gustavo; Cappelli, Francesco; Blaser, Stephane; Faist, Jérôme

2015-01-01

We develop a spectroscopy platform for industrial applications based on semiconductor quantum cascade laser (QCL) frequency combs. The platform's key features will be an unmatched combination of bandwidth of 100 cm-1, resolution of 100 kHz, speed of ten to hundreds of μs as well as size and robustness, opening doors to beforehand unreachable markets. The sensor can be built extremely compact and robust since the laser source is an all-electrically pumped semiconductor optical frequency comb and no mechanical elements are required. However, the parallel acquisition of dual-comb spectrometers comes at the price of enormous data-rates. For system scalability, robustness and optical simplicity we use free-running QCL combs. Therefore no complicated optical locking mechanisms are required. To reach high signal-to-noise ratios, we develop an algorithm, which is based on combination of coherent and non-coherent averaging. This algorithm is specifically optimized for free-running and small footprint, therefore high-repetition rate, comb sources. As a consequence, our system generates data-rates of up to 3.2 GB/sec. These data-rates need to be reduced by several orders of magnitude in real-time in order to be useful for spectral fitting algorithms. We present the development of a data-treatment solution, which reaches a single-channel throughput of 22% using a standard laptop-computer. Using a state-of-the art desktop computer, the throughput is increased to 43%. This is combined with a data-acquisition board to a stand-alone data processing unit, allowing real-time industrial process observation and continuous averaging to achieve highest signal fidelity.

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

PubMed Central

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

2016-01-01

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

High-throughput Screening of ToxCast" Phase I Chemicals in an Embryonic Stem Cell Assay Reveals Potential Disruption of a Critical Developmental Signaling Pathway

EPA Science Inventory

Little is known about the developmental toxicity of the expansive chemical landscape in existence today. Significant efforts are being made to apply novel methods to predict developmental activity of chemicals utilizing high-throughput screening (HTS) and high-content screening (...

Comparison of Human Induced Pluripotent Stem Cell-Derived Neurons and Rat Primary CorticalNeurons as In Vitro Models of Neurite Outgrowth

EPA Science Inventory

High-throughput assays that can quantify chemical-induced changes at the cellular and molecular level have been recommended for use in chemical safety assessment. High-throughput, high content imaging assays for the key cellular events of neurodevelopment have been proposed to ra...

Evaluation of sequencing approaches for high-throughput toxicogenomics (SOT)

EPA Science Inventory

Whole-genome in vitro transcriptomics has shown the capability to identify mechanisms of action and estimates of potency for chemical-mediated effects in a toxicological framework, but with limited throughput and high cost. We present the evaluation of three toxicogenomics platfo...

High Throughput Assays and Exposure Science (ISES annual meeting)

EPA Science Inventory

High throughput screening (HTS) data characterizing chemical-induced biological activity has been generated for thousands of environmentally-relevant chemicals by the US inter-agency Tox21 and the US EPA ToxCast programs. For a limited set of chemicals, bioactive concentrations r...

High Throughput Exposure Estimation Using NHANES Data (SOT)

EPA Science Inventory

In the ExpoCast project, high throughput (HT) exposure models enable rapid screening of large numbers of chemicals for exposure potential. Evaluation of these models requires empirical exposure data and due to the paucity of human metabolism/exposure data such evaluations includ...

Atlanta I-85 HOV-to-HOT conversion : analysis of vehicle and person throughput.

DOT National Transportation Integrated Search

2013-10-01

This report summarizes the vehicle and person throughput analysis for the High Occupancy Vehicle to High Occupancy Toll Lane : conversion in Atlanta, GA, undertaken by the Georgia Institute of Technology research team. The team tracked changes in : o...

EMBRYONIC VASCULAR DISRUPTION ADVERSE OUTCOMES: LINKING HIGH THROUGHPUT SIGNALING SIGNATURES WITH FUNCTIONAL CONSEQUENCES

EPA Science Inventory

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...

Accounting For Uncertainty in The Application Of High Throughput Datasets

EPA Science Inventory

The use of high throughput screening (HTS) datasets will need to adequately account for uncertainties in the data generation process and propagate these uncertainties through to ultimate use. Uncertainty arises at multiple levels in the construction of predictors using in vitro ...

Advances in High-Throughput Speed, Low-Latency Communication for Embedded Instrumentation (7th Annual SFAF Meeting, 2012)

ScienceCinema

Jordan, Scott

2018-01-24

Scott Jordan on "Advances in high-throughput speed, low-latency communication for embedded instrumentation" at the 2012 Sequencing, Finishing, Analysis in the Future Meeting held June 5-7, 2012 in Santa Fe, New Mexico.

Inter-Individual Variability in High-Throughput Risk Prioritization of Environmental Chemicals (Sot)

EPA Science Inventory

We incorporate realistic human variability into an open-source high-throughput (HT) toxicokinetics (TK) modeling framework for use in a next-generation risk prioritization approach. Risk prioritization involves rapid triage of thousands of environmental chemicals, most which have...

Inter-individual variability in high-throughput risk prioritization of environmental chemicals (IVIVE)

EPA Science Inventory

We incorporate inter-individual variability into an open-source high-throughput (HT) toxicokinetics (TK) modeling framework for use in a next-generation risk prioritization approach. Risk prioritization involves rapid triage of thousands of environmental chemicals, most which hav...

HIGH-THROUGHPUT IDENTIFICATION OF CATALYTIC REDOX-ACTIVE CYSTEINE RESIDUES

EPA Science Inventory

Cysteine (Cys) residues often play critical roles in proteins; however, identification of their specific functions has been limited to case-by-case experimental approaches. We developed a procedure for high-throughput identification of catalytic redox-active Cys in proteins by se...

Development of a thyroperoxidase inhibition assay for high-throughput screening

EPA Science Inventory

High-throughput screening (HTPS) assays to detect inhibitors of thyroperoxidase (TPO), the enzymatic catalyst for thyroid hormone (TH) synthesis, are not currently available. Herein we describe the development of a HTPS TPO inhibition assay. Rat thyroid microsomes and a fluores...

High-throughput screening, predictive modeling and computational embryology - Abstract

EPA Science Inventory

High-throughput screening (HTS) studies are providing a rich source of data that can be applied to chemical profiling to address sensitivity and specificity of molecular targets, biological pathways, cellular and developmental processes. EPA’s ToxCast project is testing 960 uniq...

Evaluating and Refining High Throughput Tools for Toxicokinetics

EPA Science Inventory

This poster summarizes efforts of the Chemical Safety for Sustainability's Rapid Exposure and Dosimetry (RED) team to facilitate the development and refinement of toxicokinetics (TK) tools to be used in conjunction with the high throughput toxicity testing data generated as a par...

Picking Cell Lines for High-Throughput Transcriptomic Toxicity Screening (SOT)

EPA Science Inventory

High throughput, whole genome transcriptomic profiling is a promising approach to comprehensively evaluate chemicals for potential biological effects. To be useful for in vitro toxicity screening, gene expression must be quantified in a set of representative cell types that captu...

20180312 - Uncertainty and Variability in High-Throughput Toxicokinetics for Risk Prioritization (SOT)

EPA Science Inventory

Streamlined approaches that use in vitro experimental data to predict chemical toxicokinetics (TK) are increasingly being used to perform risk-based prioritization based upon dosimetric adjustment of high-throughput screening (HTS) data across thousands of chemicals. However, ass...

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

PubMed Central

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

Prediction of protein-protein interactions from amino acid sequences with ensemble extreme learning machines and principal component analysis.

PubMed

You, Zhu-Hong; Lei, Ying-Ke; Zhu, Lin; Xia, Junfeng; Wang, Bing

2013-01-01

Protein-protein interactions (PPIs) play crucial roles in the execution of various cellular processes and form the basis of biological mechanisms. Although large amount of PPIs data for different species has been generated by high-throughput experimental techniques, current PPI pairs obtained with experimental methods cover only a fraction of the complete PPI networks, and further, the experimental methods for identifying PPIs are both time-consuming and expensive. Hence, it is urgent and challenging to develop automated computational methods to efficiently and accurately predict PPIs. We present here a novel hierarchical PCA-EELM (principal component analysis-ensemble extreme learning machine) model to predict protein-protein interactions only using the information of protein sequences. In the proposed method, 11188 protein pairs retrieved from the DIP database were encoded into feature vectors by using four kinds of protein sequences information. Focusing on dimension reduction, an effective feature extraction method PCA was then employed to construct the most discriminative new feature set. Finally, multiple extreme learning machines were trained and then aggregated into a consensus classifier by majority voting. The ensembling of extreme learning machine removes the dependence of results on initial random weights and improves the prediction performance. When performed on the PPI data of Saccharomyces cerevisiae, the proposed method achieved 87.00% prediction accuracy with 86.15% sensitivity at the precision of 87.59%. Extensive experiments are performed to compare our method with state-of-the-art techniques Support Vector Machine (SVM). Experimental results demonstrate that proposed PCA-EELM outperforms the SVM method by 5-fold cross-validation. Besides, PCA-EELM performs faster than PCA-SVM based method. Consequently, the proposed approach can be considered as a new promising and powerful tools for predicting PPI with excellent performance and less time.

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

PubMed

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).

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

PubMed Central

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). PMID:29432475

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

PubMed

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.

Turning tumor-promoting copper into an anti-cancer weapon via high-throughput chemistry.

PubMed

Wang, F; Jiao, P; Qi, M; Frezza, M; Dou, Q P; Yan, B

2010-01-01

Copper is an essential element for multiple biological processes. Its concentration is elevated to a very high level in cancer tissues for promoting cancer development through processes such as angiogenesis. Organic chelators of copper can passively reduce cellular copper and serve the role as inhibitors of angiogenesis. However, they can also actively attack cellular targets such as proteasome, which plays a critical role in cancer development and survival. The discovery of such molecules initially relied on a step by step synthesis followed by biological assays. Today high-throughput chemistry and high-throughput screening have significantly expedited the copper-binding molecules discovery to turn "cancer-promoting" copper into anti-cancer agents.

HTP-OligoDesigner: An Online Primer Design Tool for High-Throughput Gene Cloning and Site-Directed Mutagenesis.

PubMed

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.

A high performance hardware implementation image encryption with AES algorithm

NASA Astrophysics Data System (ADS)

Farmani, Ali; Jafari, Mohamad; Miremadi, Seyed Sohrab

2011-06-01

This paper describes implementation of a high-speed encryption algorithm with high throughput for encrypting the image. Therefore, we select a highly secured symmetric key encryption algorithm AES(Advanced Encryption Standard), in order to increase the speed and throughput using pipeline technique in four stages, control unit based on logic gates, optimal design of multiplier blocks in mixcolumn phase and simultaneous production keys and rounds. Such procedure makes AES suitable for fast image encryption. Implementation of a 128-bit AES on FPGA of Altra company has been done and the results are as follow: throughput, 6 Gbps in 471MHz. The time of encrypting in tested image with 32*32 size is 1.15ms.

High-throughput continuous hydrothermal synthesis of nanomaterials (part II): unveiling the as-prepared CexZryYzO2-δ phase diagram.

PubMed

Quesada-Cabrera, Raul; Weng, Xiaole; Hyett, Geoff; Clark, Robin J H; Wang, Xue Z; Darr, Jawwad A

2013-09-09

High-throughput continuous hydrothermal flow synthesis was used to manufacture 66 unique nanostructured oxide samples in the Ce-Zr-Y-O system. This synthesis approach resulted in a significant increase in throughput compared to that of conventional batch or continuous hydrothermal synthesis methods. The as-prepared library samples were placed into a wellplate for both automated high-throughput powder X-ray diffraction and Raman spectroscopy data collection, which allowed comprehensive structural characterization and phase mapping. The data suggested that a continuous cubic-like phase field connects all three Ce-Zr-O, Ce-Y-O, and Y-Zr-O binary systems together with a smooth and steady transition between the structures of neighboring compositions. The continuous hydrothermal process led to as-prepared crystallite sizes in the range of 2-7 nm (as determined by using the Scherrer equation).

State of the Art High-Throughput Approaches to Genotoxicity: Flow Micronucleus, Ames II, GreenScreen and Comet

EPA Pesticide Factsheets

State of the Art High-Throughput Approaches to Genotoxicity: Flow Micronucleus, Ames II, GreenScreen and Comet (Presented by Dr. Marilyn J. Aardema, Chief Scientific Advisor, Toxicology, Dr. Leon Stankowski, et. al. (6/28/2012)

Fun with High Throughput Toxicokinetics (CalEPA webinar)

EPA Science Inventory

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 ...

Incorporating Human Dosimetry and Exposure into High-Throughput In Vitro Toxicity Screening

EPA Science Inventory

Many chemicals in commerce today have undergone limited or no safety testing. To reduce the number of untested chemicals and prioritize limited testing resources, several governmental programs are using high-throughput in vitro screens for assessing chemical effects across multip...

Environmental Impact on Vascular Development Predicted by High Throughput Screening

EPA Science Inventory

Understanding health risks to embryonic development from exposure to environmental chemicals is a significant challenge given the diverse chemical landscape and paucity of data for most of these compounds. High throughput screening (HTS) in EPA’s ToxCastTM project provides vast d...

High-Throughput Dietary Exposure Predictions for Chemical Migrants from Food Packaging Materials

EPA Science Inventory

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...

AOPs and Biomarkers: Bridging High Throughput Screening and Regulatory Decision Making

EPA Science Inventory

As high throughput screening (HTS) plays a larger role in toxicity testing, camputational toxicology has emerged as a critical component in interpreting the large volume of data produced. Computational models designed to quantify potential adverse effects based on HTS data will b...

Human variability in high-throughput risk prioritization of environmental chemicals (Texas AM U. webinar)

EPA Science Inventory

We incorporate inter-individual variability into an open-source high-throughput (HT) toxicokinetics (TK) modeling framework for use in a next-generation risk prioritization approach. Risk prioritization involves rapid triage of thousands of environmental chemicals, most which hav...

HTTK: R Package for High-Throughput Toxicokinetics

EPA Science Inventory

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...

tcpl: The ToxCast Pipeline for High-Throughput Screening Data

EPA Science Inventory

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...

High-throughput screening, predictive modeling and computational embryology

EPA Science Inventory

High-throughput screening (HTS) studies are providing a rich source of data that can be applied to profile thousands of chemical compounds for biological activity and potential toxicity. EPA’s ToxCast™ project, and the broader Tox21 consortium, in addition to projects worldwide,...

In Vitro Toxicity Screening Technique for Volatile Substances Using Flow-Through System#

EPA Science Inventory

In 2007 the National Research Council envisioned the need for inexpensive, high throughput, cell based toxicity testing methods relevant to human health. High Throughput Screening (HTS) in vitro screening approaches have addressed these problems by using robotics. However the cha...

Integration of Dosimetry, Exposure and High-Throughput Screening Data in Chemical Toxicity Assessment

EPA Science Inventory

High-throughput in vitro toxicity screening can provide an efficient way to identify potential biological targets for chemicals. However, relying on nominal assay concentrations may misrepresent potential in vivo effects of these chemicals due to differences in bioavailability, c...

High-Throughput Toxicokinetics (HTTK) R package (CompTox CoP presentation)

EPA Science Inventory

Toxicokinetics (TK) provides a bridge between HTS and HTE by predicting tissue concentrations due to exposure, but traditional TK methods are resource intensive. Relatively high throughput TK (HTTK) methods have been used by the pharmaceutical industry to determine range of effic...

A review of the theory, methods and recent applications of high-throughput single-cell droplet microfluidics

NASA Astrophysics Data System (ADS)

Lagus, Todd P.; Edd, Jon F.

2013-03-01

Most cell biology experiments are performed in bulk cell suspensions where cell secretions become diluted and mixed in a contiguous sample. Confinement of single cells to small, picoliter-sized droplets within a continuous phase of oil provides chemical isolation of each cell, creating individual microreactors where rare cell qualities are highlighted and otherwise undetectable signals can be concentrated to measurable levels. Recent work in microfluidics has yielded methods for the encapsulation of cells in aqueous droplets and hydrogels at kilohertz rates, creating the potential for millions of parallel single-cell experiments. However, commercial applications of high-throughput microdroplet generation and downstream sensing and actuation methods are still emerging for cells. Using fluorescence-activated cell sorting (FACS) as a benchmark for commercially available high-throughput screening, this focused review discusses the fluid physics of droplet formation, methods for cell encapsulation in liquids and hydrogels, sensors and actuators and notable biological applications of high-throughput single-cell droplet microfluidics.

High-Throughput Cloning and Expression Library Creation for Functional Proteomics

PubMed Central

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

High-Throughput Lectin Microarray-Based Analysis of Live Cell Surface Glycosylation

PubMed Central

Li, Yu; Tao, Sheng-ce; Zhu, Heng; Schneck, Jonathan P.

2011-01-01

Lectins, plant-derived glycan-binding proteins, have long been used to detect glycans on cell surfaces. However, the techniques used to characterize serum or cells have largely been limited to mass spectrometry, blots, flow cytometry, and immunohistochemistry. While these lectin-based approaches are well established and they can discriminate a limited number of sugar isomers by concurrently using a limited number of lectins, they are not amenable for adaptation to a high-throughput platform. Fortunately, given the commercial availability of lectins with a variety of glycan specificities, lectins can be printed on a glass substrate in a microarray format to profile accessible cell-surface glycans. This method is an inviting alternative for analysis of a broad range of glycans in a high-throughput fashion and has been demonstrated to be a feasible method of identifying binding-accessible cell surface glycosylation on living cells. The current unit presents a lectin-based microarray approach for analyzing cell surface glycosylation in a high-throughput fashion. PMID:21400689

Optimization of High-Throughput Sequencing Kinetics for determining enzymatic rate constants of thousands of RNA substrates

PubMed Central

Niland, Courtney N.; Jankowsky, Eckhard; Harris, Michael E.

2016-01-01

Quantification of the specificity of RNA binding proteins and RNA processing enzymes is essential to understanding their fundamental roles in biological processes. High Throughput Sequencing Kinetics (HTS-Kin) uses high throughput sequencing and internal competition kinetics to simultaneously monitor the processing rate constants of thousands of substrates by RNA processing enzymes. This technique has provided unprecedented insight into the substrate specificity of the tRNA processing endonuclease ribonuclease P. Here, we investigate the accuracy and robustness of measurements associated with each step of the HTS-Kin procedure. We examine the effect of substrate concentration on the observed rate constant, determine the optimal kinetic parameters, and provide guidelines for reducing error in amplification of the substrate population. Importantly, we find that high-throughput sequencing, and experimental reproducibility contribute their own sources of error, and these are the main sources of imprecision in the quantified results when otherwise optimized guidelines are followed. PMID:27296633

Next-generation sequencing coupled with a cell-free display technology for high-throughput production of reliable interactome data

PubMed Central

Fujimori, Shigeo; Hirai, Naoya; Ohashi, Hiroyuki; Masuoka, Kazuyo; Nishikimi, Akihiko; Fukui, Yoshinori; Washio, Takanori; Oshikubo, Tomohiro; Yamashita, Tatsuhiro; Miyamoto-Sato, Etsuko

2012-01-01

Next-generation sequencing (NGS) has been applied to various kinds of omics studies, resulting in many biological and medical discoveries. However, high-throughput protein-protein interactome datasets derived from detection by sequencing are scarce, because protein-protein interaction analysis requires many cell manipulations to examine the interactions. The low reliability of the high-throughput data is also a problem. Here, we describe a cell-free display technology combined with NGS that can improve both the coverage and reliability of interactome datasets. The completely cell-free method gives a high-throughput and a large detection space, testing the interactions without using clones. The quantitative information provided by NGS reduces the number of false positives. The method is suitable for the in vitro detection of proteins that interact not only with the bait protein, but also with DNA, RNA and chemical compounds. Thus, it could become a universal approach for exploring the large space of protein sequences and interactome networks. PMID:23056904

NCBI GEO: archive for high-throughput functional genomic data.

PubMed

Barrett, Tanya; Troup, Dennis B; Wilhite, Stephen E; Ledoux, Pierre; Rudnev, Dmitry; Evangelista, Carlos; Kim, Irene F; Soboleva, Alexandra; Tomashevsky, Maxim; Marshall, Kimberly A; Phillippy, Katherine H; Sherman, Patti M; Muertter, Rolf N; Edgar, Ron

2009-01-01

The Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI) is the largest public repository for high-throughput gene expression data. Additionally, GEO hosts other categories of high-throughput functional genomic data, including those that examine genome copy number variations, chromatin structure, methylation status and transcription factor binding. These data are generated by the research community using high-throughput technologies like microarrays and, more recently, next-generation sequencing. The database has a flexible infrastructure that can capture fully annotated raw and processed data, enabling compliance with major community-derived scientific reporting standards such as 'Minimum Information About a Microarray Experiment' (MIAME). In addition to serving as a centralized data storage hub, GEO offers many tools and features that allow users to effectively explore, analyze and download expression data from both gene-centric and experiment-centric perspectives. This article summarizes the GEO repository structure, content and operating procedures, as well as recently introduced data mining features. GEO is freely accessible at http://www.ncbi.nlm.nih.gov/geo/.

Advanced Virus Detection Technologies Interest Group (AVDTIG): Efforts on High Throughput Sequencing (HTS) for Virus Detection.

PubMed

Khan, Arifa S; Vacante, Dominick A; Cassart, Jean-Pol; Ng, Siemon H S; Lambert, Christophe; Charlebois, Robert L; King, Kathryn E

Several nucleic-acid based technologies have recently emerged with capabilities for broad virus detection. One of these, high throughput sequencing, has the potential for novel virus detection because this method does not depend upon prior viral sequence knowledge. However, the use of high throughput sequencing for testing biologicals poses greater challenges as compared to other newly introduced tests due to its technical complexities and big data bioinformatics. Thus, the Advanced Virus Detection Technologies Users Group was formed as a joint effort by regulatory and industry scientists to facilitate discussions and provide a forum for sharing data and experiences using advanced new virus detection technologies, with a focus on high throughput sequencing technologies. The group was initiated as a task force that was coordinated by the Parenteral Drug Association and subsequently became the Advanced Virus Detection Technologies Interest Group to continue efforts for using new technologies for detection of adventitious viruses with broader participation, including international government agencies, academia, and technology service providers. © PDA, Inc. 2016.

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

PubMed

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.

Near-common-path interferometer for imaging Fourier-transform spectroscopy in wide-field microscopy

PubMed Central

Wadduwage, Dushan N.; Singh, Vijay Raj; Choi, Heejin; Yaqoob, Zahid; Heemskerk, Hans; Matsudaira, Paul; So, Peter T. C.

2017-01-01

Imaging Fourier-transform spectroscopy (IFTS) is a powerful method for biological hyperspectral analysis based on various imaging modalities, such as fluorescence or Raman. Since the measurements are taken in the Fourier space of the spectrum, it can also take advantage of compressed sensing strategies. IFTS has been readily implemented in high-throughput, high-content microscope systems based on wide-field imaging modalities. However, there are limitations in existing wide-field IFTS designs. Non-common-path approaches are less phase-stable. Alternatively, designs based on the common-path Sagnac interferometer are stable, but incompatible with high-throughput imaging. They require exhaustive sequential scanning over large interferometric path delays, making compressive strategic data acquisition impossible. In this paper, we present a novel phase-stable, near-common-path interferometer enabling high-throughput hyperspectral imaging based on strategic data acquisition. Our results suggest that this approach can improve throughput over those of many other wide-field spectral techniques by more than an order of magnitude without compromising phase stability. PMID:29392168

An improved high-throughput lipid extraction method for the analysis of human brain lipids.

PubMed

Abbott, Sarah K; Jenner, Andrew M; Mitchell, Todd W; Brown, Simon H J; Halliday, Glenda M; Garner, Brett

2013-03-01

We have developed a protocol suitable for high-throughput lipidomic analysis of human brain samples. The traditional Folch extraction (using chloroform and glass-glass homogenization) was compared to a high-throughput method combining methyl-tert-butyl ether (MTBE) extraction with mechanical homogenization utilizing ceramic beads. This high-throughput method significantly reduced sample handling time and increased efficiency compared to glass-glass homogenizing. Furthermore, replacing chloroform with MTBE is safer (less carcinogenic/toxic), with lipids dissolving in the upper phase, allowing for easier pipetting and the potential for automation (i.e., robotics). Both methods were applied to the analysis of human occipital cortex. Lipid species (including ceramides, sphingomyelins, choline glycerophospholipids, ethanolamine glycerophospholipids and phosphatidylserines) were analyzed via electrospray ionization mass spectrometry and sterol species were analyzed using gas chromatography mass spectrometry. No differences in lipid species composition were evident when the lipid extraction protocols were compared, indicating that MTBE extraction with mechanical bead homogenization provides an improved method for the lipidomic profiling of human brain tissue.

Graph-based signal integration for high-throughput phenotyping

PubMed Central

2012-01-01

Background Electronic Health Records aggregated in Clinical Data Warehouses (CDWs) promise to revolutionize Comparative Effectiveness Research and suggest new avenues of research. However, the effectiveness of CDWs is diminished by the lack of properly labeled data. We present a novel approach that integrates knowledge from the CDW, the biomedical literature, and the Unified Medical Language System (UMLS) to perform high-throughput phenotyping. In this paper, we automatically construct a graphical knowledge model and then use it to phenotype breast cancer patients. We compare the performance of this approach to using MetaMap when labeling records. Results MetaMap's overall accuracy at identifying breast cancer patients was 51.1% (n=428); recall=85.4%, precision=26.2%, and F1=40.1%. Our unsupervised graph-based high-throughput phenotyping had accuracy of 84.1%; recall=46.3%, precision=61.2%, and F1=52.8%. Conclusions We conclude that our approach is a promising alternative for unsupervised high-throughput phenotyping. PMID:23320851

High-throughput cloning and expression library creation for functional proteomics.

PubMed

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

2013-05-01

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 particularly 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 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(TM) 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). © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Genome-scale deletion screening of human long non-coding RNAs using a paired-guide RNA CRISPR library

PubMed Central

Zhu, Shiyou; Li, Wei; Liu, Jingze; Chen, Chen-Hao; Liao, Qi; Xu, Ping; Xu, Han; Xiao, Tengfei; Cao, Zhongzheng; Peng, Jingyu; Yuan, Pengfei; Brown, Myles; Liu, Xiaole Shirley; Wei, Wensheng

2017-01-01

CRISPR/Cas9 screens have been widely adopted to analyse coding gene functions, but high throughput screening of non-coding elements using this method is more challenging, because indels caused by a single cut in non-coding regions are unlikely to produce a functional knockout. A high-throughput method to produce deletions of non-coding DNA is needed. Herein, we report a high throughput genomic deletion strategy to screen for functional long non-coding RNAs (lncRNAs) that is based on a lentiviral paired-guide RNA (pgRNA) library. Applying our screening method, we identified 51 lncRNAs that can positively or negatively regulate human cancer cell growth. We individually validated 9 lncRNAs using CRISPR/Cas9-mediated genomic deletion and functional rescue, CRISPR activation or inhibition, and gene expression profiling. Our high-throughput pgRNA genome deletion method should enable rapid identification of functional mammalian non-coding elements. PMID:27798563

High-Throughput Method for Automated Colony and Cell Counting by Digital Image Analysis Based on Edge Detection

PubMed Central

Choudhry, Priya

2016-01-01

Counting cells and colonies is an integral part of high-throughput screens and quantitative cellular assays. Due to its subjective and time-intensive nature, manual counting has hindered the adoption of cellular assays such as tumor spheroid formation in high-throughput screens. The objective of this study was to develop an automated method for quick and reliable counting of cells and colonies from digital images. For this purpose, I developed an ImageJ macro Cell Colony Edge and a CellProfiler Pipeline Cell Colony Counting, and compared them to other open-source digital methods and manual counts. The ImageJ macro Cell Colony Edge is valuable in counting cells and colonies, and measuring their area, volume, morphology, and intensity. In this study, I demonstrate that Cell Colony Edge is superior to other open-source methods, in speed, accuracy and applicability to diverse cellular assays. It can fulfill the need to automate colony/cell counting in high-throughput screens, colony forming assays, and cellular assays. PMID:26848849

High-throughput determination of structural phase diagram and constituent phases using GRENDEL

NASA Astrophysics Data System (ADS)

Kusne, A. G.; Keller, D.; Anderson, A.; Zaban, A.; Takeuchi, I.

2015-11-01

Advances in high-throughput materials fabrication and characterization techniques have resulted in faster rates of data collection and rapidly growing volumes of experimental data. To convert this mass of information into actionable knowledge of material process-structure-property relationships requires high-throughput data analysis techniques. This work explores the use of the Graph-based endmember extraction and labeling (GRENDEL) algorithm as a high-throughput method for analyzing structural data from combinatorial libraries, specifically, to determine phase diagrams and constituent phases from both x-ray diffraction and Raman spectral data. The GRENDEL algorithm utilizes a set of physical constraints to optimize results and provides a framework by which additional physics-based constraints can be easily incorporated. GRENDEL also permits the integration of database data as shown by the use of critically evaluated data from the Inorganic Crystal Structure Database in the x-ray diffraction data analysis. Also the Sunburst radial tree map is demonstrated as a tool to visualize material structure-property relationships found through graph based analysis.

A simple dual online ultra-high pressure liquid chromatography system (sDO-UHPLC) for high throughput proteome analysis.

PubMed

Lee, Hangyeore; Mun, Dong-Gi; Bae, Jingi; Kim, Hokeun; Oh, Se Yeon; Park, Young Soo; Lee, Jae-Hyuk; Lee, Sang-Won

2015-08-21

We report a new and simple design of a fully automated dual-online ultra-high pressure liquid chromatography system. The system employs only two nano-volume switching valves (a two-position four port valve and a two-position ten port valve) that direct solvent flows from two binary nano-pumps for parallel operation of two analytical columns and two solid phase extraction (SPE) columns. Despite the simple design, the sDO-UHPLC offers many advantageous features that include high duty cycle, back flushing sample injection for fast and narrow zone sample injection, online desalting, high separation resolution and high intra/inter-column reproducibility. This system was applied to analyze proteome samples not only in high throughput deep proteome profiling experiments but also in high throughput MRM experiments.

Optimization and high-throughput screening of antimicrobial peptides.

PubMed

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.

Multi-step high-throughput conjugation platform for the development of antibody-drug conjugates.

PubMed

Andris, Sebastian; Wendeler, Michaela; Wang, Xiangyang; Hubbuch, Jürgen

2018-07-20

Antibody-drug conjugates (ADCs) form a rapidly growing class of biopharmaceuticals which attracts a lot of attention throughout the industry due to its high potential for cancer therapy. They combine the specificity of a monoclonal antibody (mAb) and the cell-killing capacity of highly cytotoxic small molecule drugs. Site-specific conjugation approaches involve a multi-step process for covalent linkage of antibody and drug via a linker. Despite the range of parameters that have to be investigated, high-throughput methods are scarcely used so far in ADC development. In this work an automated high-throughput platform for a site-specific multi-step conjugation process on a liquid-handling station is presented by use of a model conjugation system. A high-throughput solid-phase buffer exchange was successfully incorporated for reagent removal by utilization of a batch cation exchange step. To ensure accurate screening of conjugation parameters, an intermediate UV/Vis-based concentration determination was established including feedback to the process. For conjugate characterization, a high-throughput compatible reversed-phase chromatography method with a runtime of 7 min and no sample preparation was developed. Two case studies illustrate the efficient use for mapping the operating space of a conjugation process. Due to the degree of automation and parallelization, the platform is capable of significantly reducing process development efforts and material demands and shorten development timelines for antibody-drug conjugates. Copyright © 2018 Elsevier B.V. All rights reserved.

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

NASA Astrophysics Data System (ADS)

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.

Expected scientific performance of the three spectrometers on the extreme ultraviolet explorer

NASA Technical Reports Server (NTRS)

Vallerga, J. V.; Jelinsky, P.; Vedder, P. W.; Malina, R. F.

1990-01-01

The expected in-orbit performance of the three spectrometers included on the Extreme Ultraviolet Explorer astronomical satellite is presented. Recent calibrations of the gratings, mirrors and detectors using monochromatic and continuum EUV light sources allow the calculation of the spectral resolution and throughput of the instrument. An effective area range of 0.2 to 2.8 sq cm is achieved over the wavelength range 70-600 A with a peak spectral resolution (FWHM) of 360 assuming a spacecraft pointing knowledge of 10 arc seconds (FWHM). For a 40,000 sec observation, the average 3 sigma sensitivity to a monochromatic line source is 0.003 photons/sq cm s. Simulated observations of known classes of EUV sources, such as hot white dwarfs, and cataclysmic variables are also presented.

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

PubMed

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 the final evaluation. After the second evaluation, the final amplification curves and melting curves have been achieved.

Taxonomic and Functional Diversity of Soil and Hypolithic Microbial Communities in Miers Valley, McMurdo Dry Valleys, Antarctica.

PubMed

Wei, Sean T S; Lacap-Bugler, Donnabella C; Lau, Maggie C Y; Caruso, Tancredi; Rao, Subramanya; de Los Rios, Asunción; Archer, Stephen K; Chiu, Jill M Y; Higgins, Colleen; Van Nostrand, Joy D; Zhou, Jizhong; Hopkins, David W; Pointing, Stephen B

2016-01-01

The McMurdo Dry Valleys of Antarctica are an extreme polar desert. Mineral soils support subsurface microbial communities and translucent rocks support development of hypolithic communities on ventral surfaces in soil contact. Despite significant research attention, relatively little is known about taxonomic and functional diversity or their inter-relationships. Here we report a combined diversity and functional interrogation for soil and hypoliths of the Miers Valley in the McMurdo Dry Valleys of Antarctica. The study employed 16S rRNA fingerprinting and high throughput sequencing combined with the GeoChip functional microarray. The soil community was revealed as a highly diverse reservoir of bacterial diversity dominated by actinobacteria. Hypolithic communities were less diverse and dominated by cyanobacteria. Major differences in putative functionality were that soil communities displayed greater diversity in stress tolerance and recalcitrant substrate utilization pathways, whilst hypolithic communities supported greater diversity of nutrient limitation adaptation pathways. A relatively high level of functional redundancy in both soil and hypoliths may indicate adaptation of these communities to fluctuating environmental conditions.

Ultrasensitive Biosensors Using Enhanced Fano Resonances in Capped Gold Nanoslit Arrays

PubMed Central

Lee, Kuang-Li; Huang, Jhih-Bin; Chang, Jhih-Wei; Wu, Shu-Han; Wei, Pei-Kuen

2015-01-01

Nanostructure-based sensors are capable of sensitive and label-free detection for biomedical applications. However, plasmonic sensors capable of highly sensitive detection with high-throughput and low-cost fabrication techniques are desirable. We show that capped gold nanoslit arrays made by thermal-embossing nanoimprint method on a polymer film can produce extremely sharp asymmetric resonances for a transverse magnetic-polarized wave. An ultrasmall linewidth is formed due to the enhanced Fano coupling between the cavity resonance mode in nanoslits and surface plasmon resonance mode on periodic metallic surface. With an optimal slit length and width, the full width at half-maximum bandwidth of the Fano mode is only 3.68 nm. The wavelength sensitivity is 926 nm/RIU for 60-nm-width and 1,000-nm-period nanoslits. The figure of merit is up to 252. The obtained value is higher than the theoretically estimated upper limits of the prism-coupling SPR sensors and the previously reported record high figure-of-merit in array sensors. In addition, the structure has an ultrahigh intensity sensitivity up to 48,117%/RIU. PMID:25708955

High Throughput Transcriptomics @ USEPA (Toxicology ...

EPA Pesticide Factsheets

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.

Mobile element biology – new possibilities with high-throughput sequencing

PubMed Central

Xing, Jinchuan; Witherspoon, David J.; Jorde, Lynn B.

2014-01-01

Mobile elements compose more than half of the human genome, but until recently their large-scale detection was time-consuming and challenging. With the development of new high-throughput sequencing technologies, the complete spectrum of mobile element variation in humans can now be identified and analyzed. Thousands of new mobile element insertions have been discovered, yielding new insights into mobile element biology, evolution, and genomic variation. We review several high-throughput methods, with an emphasis on techniques that specifically target mobile element insertions in humans, and we highlight recent applications of these methods in evolutionary studies and in the analysis of somatic alterations in human cancers. PMID:23312846

Advances in high throughput DNA sequence data compression.

PubMed

Sardaraz, Muhammad; Tahir, Muhammad; Ikram, Ataul Aziz

2016-06-01

Advances in high throughput sequencing technologies and reduction in cost of sequencing have led to exponential growth in high throughput DNA sequence data. This growth has posed challenges such as storage, retrieval, and transmission of sequencing data. Data compression is used to cope with these challenges. Various methods have been developed to compress genomic and sequencing data. In this article, we present a comprehensive review of compression methods for genome and reads compression. Algorithms are categorized as referential or reference free. Experimental results and comparative analysis of various methods for data compression are presented. Finally, key challenges and research directions in DNA sequence data compression are highlighted.

Accelerating Virtual High-Throughput Ligand Docking: current technology and case study on a petascale supercomputer.

PubMed

Ellingson, Sally R; Dakshanamurthy, Sivanesan; Brown, Milton; Smith, Jeremy C; Baudry, Jerome

2014-04-25

In this paper we give the current state of high-throughput virtual screening. We describe a case study of using a task-parallel MPI (Message Passing Interface) version of Autodock4 [1], [2] to run a virtual high-throughput screen of one-million compounds on the Jaguar Cray XK6 Supercomputer at Oak Ridge National Laboratory. We include a description of scripts developed to increase the efficiency of the predocking file preparation and postdocking analysis. A detailed tutorial, scripts, and source code for this MPI version of Autodock4 are available online at http://www.bio.utk.edu/baudrylab/autodockmpi.htm.

LOCATE: a mouse protein subcellular localization database

PubMed Central

Fink, J. Lynn; Aturaliya, Rajith N.; Davis, Melissa J.; Zhang, Fasheng; Hanson, Kelly; Teasdale, Melvena S.; Kai, Chikatoshi; Kawai, Jun; Carninci, Piero; Hayashizaki, Yoshihide; Teasdale, Rohan D.

2006-01-01

We present here LOCATE, a curated, web-accessible database that houses data describing the membrane organization and subcellular localization of proteins from the FANTOM3 Isoform Protein Sequence set. Membrane organization is predicted by the high-throughput, computational pipeline MemO. The subcellular locations of selected proteins from this set were determined by a high-throughput, immunofluorescence-based assay and by manually reviewing >1700 peer-reviewed publications. LOCATE represents the first effort to catalogue the experimentally verified subcellular location and membrane organization of mammalian proteins using a high-throughput approach and provides localization data for ∼40% of the mouse proteome. It is available at . PMID:16381849

An economical and effective high-throughput DNA extraction protocol for molecular marker analysis in honey bees

USDA-ARS?s Scientific Manuscript database

Extraction of DNA from tissue samples can be expensive both in time and monetary resources and can often require handling and disposal of hazardous chemicals. We have developed a high throughput protocol for extracting DNA from honey bees that is of a high enough quality and quantity to enable hundr...

20171015 - Capabilities and Evaluation of the US EPA’s HTTK (High Throughput Toxicokinetics) R package (ISES)

EPA Science Inventory

Toxicokinetics (TK) provides a bridge between toxicity and exposure assessment by predicting tissue concentrations due to exposure, however traditional TK methods are resource intensive. Relatively high throughput TK (HTTK) methods have been used by the pharmaceutical industry to...

High-throughput dietary exposure predictions for chemical migrants from food contact substances for use in chemical prioritization

EPA Science Inventory

Under the ExpoCast program, United States Environmental Protection Agency (EPA) researchers have developed a high-throughput (HT) framework for estimating aggregate exposures to chemicals from multiple pathways to support rapid prioritization of chemicals. Here, we present method...

Environmental surveillance and monitoring. The next frontiers for high-throughput toxicology

EPA Science Inventory

High throughput toxicity testing (HTT) technologies along with the world-wide web are revolutionizing both generation and access to data regarding the bioactivities that chemicals can elicit when they interact with specific proteins, genes, or other targets in the body of an orga...

High-Throughput Models for Exposure-Based Chemical Prioritization in the ExpoCast Project

EPA Science Inventory

The United States Environmental Protection Agency (U.S. EPA) must characterize potential risks to human health and the environment associated with manufacture and use of thousands of chemicals. High-throughput screening (HTS) for biological activity allows the ToxCast research pr...

High-Throughput Exposure Potential Prioritization for ToxCast Chemicals

EPA Science Inventory

The U.S. EPA must consider lists of hundreds to thousands of chemicals when prioritizing research resources in order to identify risk to human populations and the environment. High-throughput assays to identify biological activity in vitro have allowed the ToxCastTM program to i...

Use of High-Throughput Testing and Approaches for Evaluating Chemical Risk-Relevance to Humans

EPA Science Inventory

ToxCast is profiling the bioactivity of thousands of chemicals based on high-throughput screening (HTS) and computational models that integrate knowledge of biological systems and in vivo toxicities. Many of these assays probe signaling pathways and cellular processes critical to...

High-Throughput Simulation of Environmental Chemical Fate for Exposure Prioritization

EPA Science Inventory

The U.S. EPA must consider lists of hundreds to thousands of chemicals when allocating resources to identify risk in human populations and the environment. High-throughput screening assays to characterize biological activity in vitro have allowed the ToxCastTM program to identify...

New Challenges of the Computation of Multiple Sequence Alignments in the High-Throughput Era (2010 JGI/ANL HPC Workshop)

ScienceCinema

Notredame, Cedric

2018-05-02

Cedric Notredame from the Centre for Genomic Regulation gives a presentation on New Challenges of the Computation of Multiple Sequence Alignments in the High-Throughput Era at the JGI/Argonne HPC Workshop on January 26, 2010.

Molecular characterization of a novel Nucleorhabdovirus from black currant identified by high-throughput sequencing

USDA-ARS?s Scientific Manuscript database

Contigs with sequence similarities to several nucleorhabdoviruses were identified by high-throughput sequencing analysis from a black currant (Ribes nigrum L.) cultivar. The complete genomic sequence of this new nucleorhabdovirus is 14,432 nucleotides. Its genomic organization is typical of nucleorh...

Estimating Toxicity Pathway Activating Doses for High Throughput Chemical Risk Assessments

EPA Science Inventory

Estimating a Toxicity Pathway Activating Dose (TPAD) from in vitro assays as an analog to a reference dose (RfD) derived from in vivo toxicity tests would facilitate high throughput risk assessments of thousands of data-poor environmental chemicals. Estimating a TPAD requires def...

Incorporating Population Variability and Susceptible Subpopulations into Dosimetry for High-Throughput Toxicity Testing

EPA Science Inventory

Momentum is growing worldwide to use in vitro high-throughput screening (HTS) to evaluate human health effects of chemicals. However, the integration of dosimetry into HTS assays and incorporation of population variability will be essential before its application in a risk assess...

Retrofit Strategies for Incorporating Xenobiotic Metabolism into High Throughput Screening Assays (EMGS)

EPA Science Inventory

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...

Defining the taxonomic domain of applicability for mammalian-based high-throughput screening assays

EPA Science Inventory

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...

High Throughput Plasmid Sequencing with Illumina and CLC Bio (Seventh Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting 2012)

ScienceCinema

Athavale, Ajay

2018-01-04

Ajay Athavale (Monsanto) presents "High Throughput Plasmid Sequencing with Illumina and CLC Bio" at the 7th Annual Sequencing, Finishing, Analysis in the Future (SFAF) Meeting held in June, 2012 in Santa Fe, NM.

The Impact of Data Fragmentation on High-Throughput Clinical Phenotyping

ERIC Educational Resources Information Center

Wei, Weiqi

2012-01-01

Subject selection is essential and has become the rate-limiting step for harvesting knowledge to advance healthcare through clinical research. Present manual approaches inhibit researchers from conducting deep and broad studies and drawing confident conclusions. High-throughput clinical phenotyping (HTCP), a recently proposed approach, leverages…

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

PubMed

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.

Droplet microfluidic technology for single-cell high-throughput screening.

PubMed

Brouzes, Eric; Medkova, Martina; Savenelli, Neal; Marran, Dave; Twardowski, Mariusz; Hutchison, J Brian; Rothberg, Jonathan M; Link, Darren R; Perrimon, Norbert; Samuels, Michael L

2009-08-25

We present a droplet-based microfluidic technology that enables high-throughput screening of single mammalian cells. This integrated platform allows for the encapsulation of single cells and reagents in independent aqueous microdroplets (1 pL to 10 nL volumes) dispersed in an immiscible carrier oil and enables the digital manipulation of these reactors at a very high-throughput. Here, we validate a full droplet screening workflow by conducting a droplet-based cytotoxicity screen. To perform this screen, we first developed a droplet viability assay that permits the quantitative scoring of cell viability and growth within intact droplets. Next, we demonstrated the high viability of encapsulated human monocytic U937 cells over a period of 4 days. Finally, we developed an optically-coded droplet library enabling the identification of the droplets composition during the assay read-out. Using the integrated droplet technology, we screened a drug library for its cytotoxic effect against U937 cells. Taken together our droplet microfluidic platform is modular, robust, uses no moving parts, and has a wide range of potential applications including high-throughput single-cell analyses, combinatorial screening, and facilitating small sample analyses.

Microfluidic guillotine for single-cell wound repair studies

NASA Astrophysics Data System (ADS)

Blauch, Lucas R.; Gai, Ya; Khor, Jian Wei; Sood, Pranidhi; Marshall, Wallace F.; Tang, Sindy K. Y.

2017-07-01

Wound repair is a key feature distinguishing living from nonliving matter. Single cells are increasingly recognized to be capable of healing wounds. The lack of reproducible, high-throughput wounding methods has hindered single-cell wound repair studies. This work describes a microfluidic guillotine for bisecting single Stentor coeruleus cells in a continuous-flow manner. Stentor is used as a model due to its robust repair capacity and the ability to perform gene knockdown in a high-throughput manner. Local cutting dynamics reveals two regimes under which cells are bisected, one at low viscous stress where cells are cut with small membrane ruptures and high viability and one at high viscous stress where cells are cut with extended membrane ruptures and decreased viability. A cutting throughput up to 64 cells per minute—more than 200 times faster than current methods—is achieved. The method allows the generation of more than 100 cells in a synchronized stage of their repair process. This capacity, combined with high-throughput gene knockdown in Stentor, enables time-course mechanistic studies impossible with current wounding methods.

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

PubMed

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.

Ion channel drug discovery and research: the automated Nano-Patch-Clamp technology.

PubMed

Brueggemann, A; George, M; Klau, M; Beckler, M; Steindl, J; Behrends, J C; Fertig, N

2004-01-01

Unlike the genomics revolution, which was largely enabled by a single technological advance (high throughput sequencing), rapid advancement in proteomics will require a broader effort to increase the throughput of a number of key tools for functional analysis of different types of proteins. In the case of ion channels -a class of (membrane) proteins of great physiological importance and potential as drug targets- the lack of adequate assay technologies is felt particularly strongly. The available, indirect, high throughput screening methods for ion channels clearly generate insufficient information. The best technology to study ion channel function and screen for compound interaction is the patch clamp technique, but patch clamping suffers from low throughput, which is not acceptable for drug screening. A first step towards a solution is presented here. The nano patch clamp technology, which is based on a planar, microstructured glass chip, enables automatic whole cell patch clamp measurements. The Port-a-Patch is an automated electrophysiology workstation, which uses planar patch clamp chips. This approach enables high quality and high content ion channel and compound evaluation on a one-cell-at-a-time basis. The presented automation of the patch process and its scalability to an array format are the prerequisites for any higher throughput electrophysiology instruments.

High-Reflectivity Coatings for a Vacuum Ultraviolet Spectropolarimeter

NASA Astrophysics Data System (ADS)

Narukage, Noriyuki; Kubo, Masahito; Ishikawa, Ryohko; Ishikawa, Shin-nosuke; Katsukawa, Yukio; Kobiki, Toshihiko; Giono, Gabriel; Kano, Ryouhei; Bando, Takamasa; Tsuneta, Saku; Auchère, Frédéric; Kobayashi, Ken; Winebarger, Amy; McCandless, Jim; Chen, Jianrong; Choi, Joanne

2017-03-01

Precise polarization measurements in the vacuum ultraviolet (VUV) region are expected to be a new tool for inferring the magnetic fields in the upper atmosphere of the Sun. High-reflectivity coatings are key elements to achieving high-throughput optics for precise polarization measurements. We fabricated three types of high-reflectivity coatings for a solar spectropolarimeter in the hydrogen Lyman-α (Lyα; 121.567 nm) region and evaluated their performance. The first high-reflectivity mirror coating offers a reflectivity of more than 80 % in Lyα optics. The second is a reflective narrow-band filter coating that has a peak reflectivity of 57 % in Lyα, whereas its reflectivity in the visible light range is lower than 1/10 of the peak reflectivity (˜ 5 % on average). This coating can be used to easily realize a visible light rejection system, which is indispensable for a solar telescope, while maintaining high throughput in the Lyα line. The third is a high-efficiency reflective polarizing coating that almost exclusively reflects an s-polarized beam at its Brewster angle of 68° with a reflectivity of 55 %. This coating achieves both high polarizing power and high throughput. These coatings contributed to the high-throughput solar VUV spectropolarimeter called the Chromospheric Lyman-Alpha SpectroPolarimeter (CLASP), which was launched on 3 September, 2015.

A simple and sensitive high-throughput GFP screening in woody and herbaceous plants.

PubMed

Hily, Jean-Michel; Liu, Zongrang

2009-03-01

Green fluorescent protein (GFP) has been used widely as a powerful bioluminescent reporter, but its visualization by existing methods in tissues or whole plants and its utilization for high-throughput screening remains challenging in many species. Here, we report a fluorescence image analyzer-based method for GFP detection and its utility for high-throughput screening of transformed plants. Of three detection methods tested, the Typhoon fluorescence scanner was able to detect GFP fluorescence in all Arabidopsis thaliana tissues and apple leaves, while regular fluorescence microscopy detected it only in Arabidopsis flowers and siliques but barely in the leaves of either Arabidopsis or apple. The hand-held UV illumination method failed in all tissues of both species. Additionally, the Typhoon imager was able to detect GFP fluorescence in both green and non-green tissues of Arabidopsis seedlings as well as in imbibed seeds, qualifying it as a high-throughput screening tool, which was further demonstrated by screening the seedlings of primary transformed T(0) seeds. Of the 30,000 germinating Arabidopsis seedlings screened, at least 69 GFP-positive lines were identified, accounting for an approximately 0.23% transformation efficiency. About 14,000 seedlings grown in 16 Petri plates could be screened within an hour, making the screening process significantly more efficient and robust than any other existing high-throughput screening method for transgenic plants.

Development of a high-throughput microscale cell disruption platform for Pichia pastoris in rapid bioprocess design.

PubMed

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.

High-throughput screening with nanoimprinting 3D culture for efficient drug development by mimicking the tumor environment.

PubMed

Yoshii, Yukie; Furukawa, Takako; Waki, Atsuo; Okuyama, Hiroaki; Inoue, Masahiro; Itoh, Manabu; Zhang, Ming-Rong; Wakizaka, Hidekatsu; Sogawa, Chizuru; Kiyono, Yasushi; Yoshii, Hiroshi; Fujibayashi, Yasuhisa; Saga, Tsuneo

2015-05-01

Anti-cancer drug development typically utilizes high-throughput screening with two-dimensional (2D) cell culture. However, 2D culture induces cellular characteristics different from tumors in vivo, resulting in inefficient drug development. Here, we report an innovative high-throughput screening system using nanoimprinting 3D culture to simulate in vivo conditions, thereby facilitating efficient drug development. We demonstrated that cell line-based nanoimprinting 3D screening can more efficiently select drugs that effectively inhibit cancer growth in vivo as compared to 2D culture. Metabolic responses after treatment were assessed using positron emission tomography (PET) probes, and revealed similar characteristics between the 3D spheroids and in vivo tumors. Further, we developed an advanced method to adopt cancer cells from patient tumor tissues for high-throughput drug screening with nanoimprinting 3D culture, which we termed Cancer tissue-Originated Uniformed Spheroid Assay (COUSA). This system identified drugs that were effective in xenografts of the original patient tumors. Nanoimprinting 3D spheroids showed low permeability and formation of hypoxic regions inside, similar to in vivo tumors. Collectively, the nanoimprinting 3D culture provides easy-handling high-throughput drug screening system, which allows for efficient drug development by mimicking the tumor environment. The COUSA system could be a useful platform for drug development with patient cancer cells. Copyright © 2015 Elsevier Ltd. All rights reserved.

A Multidisciplinary Approach to High Throughput Nuclear Magnetic Resonance Spectroscopy

PubMed Central

Pourmodheji, Hossein; Ghafar-Zadeh, Ebrahim; Magierowski, Sebastian

2016-01-01

Nuclear Magnetic Resonance (NMR) is a non-contact, powerful structure-elucidation technique for biochemical analysis. NMR spectroscopy is used extensively in a variety of life science applications including drug discovery. However, existing NMR technology is limited in that it cannot run a large number of experiments simultaneously in one unit. Recent advances in micro-fabrication technologies have attracted the attention of researchers to overcome these limitations and significantly accelerate the drug discovery process by developing the next generation of high-throughput NMR spectrometers using Complementary Metal Oxide Semiconductor (CMOS). In this paper, we examine this paradigm shift and explore new design strategies for the development of the next generation of high-throughput NMR spectrometers using CMOS technology. A CMOS NMR system consists of an array of high sensitivity micro-coils integrated with interfacing radio-frequency circuits on the same chip. Herein, we first discuss the key challenges and recent advances in the field of CMOS NMR technology, and then a new design strategy is put forward for the design and implementation of highly sensitive and high-throughput CMOS NMR spectrometers. We thereafter discuss the functionality and applicability of the proposed techniques by demonstrating the results. For microelectronic researchers starting to work in the field of CMOS NMR technology, this paper serves as a tutorial with comprehensive review of state-of-the-art technologies and their performance levels. Based on these levels, the CMOS NMR approach offers unique advantages for high resolution, time-sensitive and high-throughput bimolecular analysis required in a variety of life science applications including drug discovery. PMID:27294925

Break-up of droplets in a concentrated emulsion flowing through a narrow constriction

NASA Astrophysics Data System (ADS)

Kim, Minkyu; Rosenfeld, Liat; Tang, Sindy; Tang Lab Team

2014-11-01

Droplet microfluidics has enabled a wide range of high throughput screening applications. Compared with other technologies such as robotic screening technology, droplet microfluidics has 1000 times higher throughput, which makes the technology one of the most promising platforms for the ultrahigh throughput screening applications. Few studies have considered the throughput of the droplet interrogation process, however. In this research, we show that the probability of break-up increases with increasing flow rate, entrance angle to the constriction, and size of the drops. Since single drops do not break at the highest flow rate used in the system, break-ups occur primarily from the interactions between highly packed droplets close to each other. Moreover, the probabilistic nature of the break-up process arises from the stochastic variations in the packing configuration. Our results can be used to calculate the maximum throughput of the serial interrogation process. For 40 pL-drops, the highest throughput with less than 1% droplet break-up was measured to be approximately 7,000 drops per second. In addition, the results are useful for understanding the behavior of concentrated emulsions in applications such as mobility control in enhanced oil recovery.

Environmental surveillance and monitoring the next frontier for pathway-based high throughput screening

EPA Science Inventory

In response to a proposed vision and strategy for toxicity testing in the 21st century nascent high throughput toxicology (HTT) programs have tested thousands of chemicals in hundreds of pathway-based biological assays. Although, to date, use of HTT data for safety assessment of ...

Molecular characterization of a novel Luteovirus from peach identified by high-throughput sequencing

USDA-ARS?s Scientific Manuscript database

Contigs with sequence homologies to Cherry-associated luteovirus were identified by high-throughput sequencing analysis of two peach accessions undergoing quarantine testing. The complete genomic sequences of the two isolates of this virus are 5,819 and 5,814 nucleotides. Their genome organization i...

20180311 - Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells (SOT)

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

20171024 - Capabilities and Evaluation of the US EPA’s HTTK (High Throughput Toxicokinetics) R package (Webinar Presentation to European Chemical Agency

EPA Science Inventory

Toxicokinetics (TK) provides a bridge between toxicity and exposure assessment by predicting tissue concentrations due to exposure. However traditional TK methods are resource intensive. Relatively high throughput TK (HTTK) methods have been used by the pharmaceutical industry to...

Identifying Toxicity Pathways with ToxCast High-Throughput Screening and Applications to Predicting Developmental Toxicity

EPA Science Inventory

Results from rodent and non-rodent prenatal developmental toxicity tests for over 300 chemicals have been curated into the relational database ToxRefDB. These same chemicals have been run in concentration-response format through over 500 high-throughput screening assays assessin...

Discovery of viruses and virus-like pathogens in pistachio using high-throughput sequencing

USDA-ARS?s Scientific Manuscript database

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...

SeqAPASS to evaluate conservation of high-throughput screening targets across non-mammalian species

EPA Science Inventory

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...

20180312 - Applying a High-Throughput PBTK Model for IVIVE (SOT)

EPA Science Inventory

The ability to link in vitro and in vivo toxicity enables the use of high-throughput in vitro assays as an alternative to resource intensive animal studies. Toxicokinetics (TK) should help describe this link, but prior work found weak correlation when using a TK model for in vitr...

High-throughput screening of chemical effects on steroidogenesis using H295R human adrenocortical carcinoma cells

EPA Science Inventory

Disruption of steroidogenesis by environmental chemicals can result in altered hormone levels causing adverse reproductive and developmental effects. A high-throughput assay using H295R human adrenocortical carcinoma cells was used to evaluate the effect of 2,060 chemical samples...

High-Throughput Simulation of Environmental Chemical Fate for Exposure Prioritization (Annual Meeting of ISES)

EPA Science Inventory

The U.S. EPA must consider thousands of chemicals when allocating resources to assess risk in human populations and the environment. High-throughput screening assays to characterize biological activity in vitro are being implemented in the ToxCastTM program to rapidly characteri...

Integration of chemical-specific exposure and pharmacokinetic information with the chemical-agnostic AOP framework to support high throughput risk assessment

EPA Science Inventory

Application of the Adverse Outcome Pathway (AOP) framework and high throughput toxicity testing in chemical-specific risk assessment requires reconciliation of chemical concentrations sufficient to trigger a molecular initiating event measured in vitro and at the relevant target ...

Evaluation of food-relevant chemicals in the ToxCast high-throughput screening program

EPA Science Inventory

There are thousands of chemicals that are directly added to or come in contact with food, many of which have undergone little to no toxicological evaluation. The ToxCast high-throughput screening (HTS) program has evaluated over 1,800 chemicals in concentration-response across ~8...

Application of Physiologically-Based Pharmacokinetic/Pharmacodynamic Model for Interpretation of High-throughput Screening Assay for Thyroperoxidase Inhibition

EPA Science Inventory

In vitro based assays are used to identify potential endocrine disrupting chemicals. Thyroperoxidase (TPO), an enzyme essential for thyroid hormone (TH) synthesis, is a target site for disruption of the thyroid axis for which a high-throughput screening (HTPS) assay has recently ...

Incorporating High-Throughput Exposure Predictions with Dosimetry-Adjusted In Vitro Bioactivity to Inform Chemical Toxicity Testing

EPA Science Inventory

We previously integrated dosimetry and exposure with high-throughput screening (HTS) to enhance the utility of ToxCast™ HTS data by translating in vitro bioactivity concentrations to oral equivalent doses (OEDs) required to achieve these levels internally. These OEDs were compare...

An Evaluation of 25 Selected ToxCast Chemicals in Medium-Throughput Assays to Detect Genotoxicity

EPA Science Inventory

ABSTRACTToxCast is a multi-year effort to develop a cost-effective approach for the US EPA to prioritize chemicals for toxicity testing. Initial evaluation of more than 500 high-throughput (HT) microwell-based assays without metabolic activation showed that most lacked high speci...

High Throughput Assays for Exposure Science (NIEHS OHAT Staff Meeting presentation)

EPA Science Inventory

High throughput screening (HTS) data that characterize chemically induced biological activity have been generated for thousands of chemicals by the US interagency Tox21 and the US EPA ToxCast programs. In many cases there are no data available for comparing bioactivity from HTS w...

Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

Differentiating pathway-specific from nonspecific effects in high-throughput toxicity data: A foundation for prioritizing adverse outcome pathway development

EPA Science Inventory

The U.S. Environmental Protection Agency’s ToxCast program has screened thousands of chemicals for biological activity, primarily using high-throughput in vitro bioassays. Adverse outcome pathways (AOPs) offer a means to link pathway-specific biological activities with potential ...

“httk”: EPA’s Tool for High Throughput Toxicokinetics (CompTox CoP)

EPA Science Inventory

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...

Tiered High-Throughput Screening Approach to Identify Thyroperoxidase Inhibitors within the ToxCast Phase I and II Chemical Libraries

EPA Science Inventory

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 limi...

Perspectives on Validation of High-Throughput Assays Supporting 21st Century Toxicity Testing

EPA Science Inventory

In vitro high-throughput screening (HTS) assays are seeing increasing use in toxicity testing. HTS assays can simultaneously test many chemicals but have seen limited use in the regulatory arena, in part because of the need to undergo rigorous, time-consuming formal validation. ...

Evaluating the Impact of Uncertainties in Clearance and Exposure When Prioritizing Chemicals Screened in High-Throughput Assays

EPA Science Inventory

The toxicity-testing paradigm has evolved to include high-throughput (HT) methods for addressing the increasing need to screen hundreds to thousands of chemicals rapidly. Approaches that involve in vitro screening assays, in silico predictions of exposure concentrations, and phar...

Predictive Model of Rat Reproductive Toxicity from ToxCast High Throughput Screening

EPA Science Inventory

The EPA ToxCast research program uses high throughput screening for bioactivity profiling and predicting the toxicity of large numbers of chemicals. ToxCast Phase‐I tested 309 well‐characterized chemicals in over 500 assays for a wide range of molecular targets and cellular respo...

Applying a High-Throughput PBTK Model for IVIVE

EPA Science Inventory

The ability to link in vitro and in vivo toxicity enables the use of high-throughput in vitro assays as an alternative to resource intensive animal studies. Toxicokinetics (TK) should help describe this link, but prior work found weak correlation when using a TK model for in vitr...

High-throughput genotyping of hop (Humulus lupulus L.) utilising diversity arrays technology (DArT)

USDA-ARS?s Scientific Manuscript database

Implementation of molecular methods in hop breeding is dependent on the availability of sizeable numbers of polymorphic markers and a comprehensive understanding of genetic variation. Diversity Arrays Technology (DArT) is a high-throughput cost-effective method for the discovery of large numbers of...

Neural Progenitor Cells as Models for High-Throughput Screens of Developmental Neurotoxicity: State of the Science

EPA Science Inventory

In vitro, high-throughput approaches have been widely recommended as an approach to screen chemicals for the potential to cause developmental neurotoxicity and prioritize them for additional testing. The choice of cellular models for such an approach will have important ramificat...

High-throughput exposure modeling to support prioritization of chemicals in personal care products

EPA Science Inventory

We demonstrate the application of a high-throughput modeling framework to estimate exposure to chemicals used in personal care products (PCPs). As a basis for estimating exposure, we use the product intake fraction (PiF), defined as the mass of chemical taken by an individual or ...

Sensitivity of neuroprogenitor cells to chemical-induced apoptosis using a multiplexed assay suitable for high-throughput screening*

EPA Science Inventory

AbstractHigh-throughput methods are useful for rapidly screening large numbers of chemicals for biological activity, including the perturbation of pathways that may lead to adverse cellular effects. In vitro assays for the key events of neurodevelopment, including apoptosis, may ...

Using Alternative Approaches to Prioritize Testing for the Universe of Chemicals with Potential for Human Exposure (WC9)

EPA Science Inventory

One use of alternative methods is to target animal use at only those chemicals and tests that are absolutely necessary. We discuss prioritization of testing based on high-throughput screening assays (HTS), QSAR modeling, high-throughput toxicokinetics (HTTK), and exposure modelin...

Integrated Model of Chemical Perturbations of a Biological PathwayUsing 18 In Vitro High Throughput Screening Assays for the Estrogen Receptor

EPA Science Inventory

We demonstrate a computational network model that integrates 18 in vitro, high-throughput screening assays measuring estrogen receptor (ER) binding, dimerization, chromatin binding, transcriptional activation and ER-dependent cell proliferation. The network model uses activity pa...

Emory University: High-Throughput Protein-Protein Interaction Analysis for Hippo Pathway Profiling | Office of Cancer Genomics

Cancer.gov

The CTD2 Center at Emory University used high-throughput protein-protein interaction (PPI) mapping for Hippo signaling pathway profiling to rapidly unveil promising PPIs as potential therapeutic targets and advance functional understanding of signaling circuitry in cells. Read the abstract.

Harnessing High-Throughput Monitoring Methods to Strengthen 21st Century Risk-Based Evaluations (SETAC Presentation)

EPA Science Inventory

Over the past ten years, the US government has invested in high-throughput (HT) methods to screen chemicals for biological activity. Under the interagency Tox21 consortium and the US Environmental Protection Agency’s (EPA) ToxCast™ program, thousands of chemicals have...

Genomics tools available for unravelling mechanisms underlying agronomical traits in strawberry with more to come

USDA-ARS?s Scientific Manuscript database

In the last few years, high-throughput genomics promised to bridge the gap between plant physiology and plant sciences. In addition, high-throughput genotyping technologies facilitate marker-based selection for better performing genotypes. In strawberry, Fragaria vesca was the first reference sequen...

High-throughput genotoxicity assay identifies antioxidants as inducers of DNA damage response and cell death

EPA Science Inventory

Human ATAD5 is an excellent biomarker for identifying genotoxic compounds because ATADS protein levels increase post-transcriptionally following exposure to a variety of DNA damaging agents. Here we report a novel quantitative high-throughput ATAD5-Iuciferase assay that can moni...

Recent developments in software tools for high-throughput in vitro ADME support with high-resolution MS.

PubMed

Paiva, Anthony; Shou, Wilson Z

2016-08-01

The last several years have seen the rapid adoption of the high-resolution MS (HRMS) for bioanalytical support of high throughput in vitro ADME profiling. Many capable software tools have been developed and refined to process quantitative HRMS bioanalysis data for ADME samples with excellent performance. Additionally, new software applications specifically designed for quan/qual soft spot identification workflows using HRMS have greatly enhanced the quality and efficiency of the structure elucidation process for high throughput metabolite ID in early in vitro ADME profiling. Finally, novel approaches in data acquisition and compression, as well as tools for transferring, archiving and retrieving HRMS data, are being continuously refined to tackle the issue of large data file size typical for HRMS analyses.

Automated crystallographic system for high-throughput protein structure determination.

PubMed

Brunzelle, Joseph S; Shafaee, Padram; Yang, Xiaojing; Weigand, Steve; Ren, Zhong; Anderson, Wayne F

2003-07-01

High-throughput structural genomic efforts require software that is highly automated, distributive and requires minimal user intervention to determine protein structures. Preliminary experiments were set up to test whether automated scripts could utilize a minimum set of input parameters and produce a set of initial protein coordinates. From this starting point, a highly distributive system was developed that could determine macromolecular structures at a high throughput rate, warehouse and harvest the associated data. The system uses a web interface to obtain input data and display results. It utilizes a relational database to store the initial data needed to start the structure-determination process as well as generated data. A distributive program interface administers the crystallographic programs which determine protein structures. Using a test set of 19 protein targets, 79% were determined automatically.

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

NASA Astrophysics Data System (ADS)

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.

Direct assembling methodologies for high-throughput bioscreening

PubMed Central

Rodríguez-Dévora, Jorge I.; Shi, Zhi-dong; Xu, Tao

2012-01-01

Over the last few decades, high-throughput (HT) bioscreening, a technique that allows rapid screening of biochemical compound libraries against biological targets, has been widely used in drug discovery, stem cell research, development of new biomaterials, and genomics research. To achieve these ambitions, scaffold-free (or direct) assembly of biological entities of interest has become critical. Appropriate assembling methodologies are required to build an efficient HT bioscreening platform. The development of contact and non-contact assembling systems as a practical solution has been driven by a variety of essential attributes of the bioscreening system, such as miniaturization, high throughput, and high precision. The present article reviews recent progress on these assembling technologies utilized for the construction of HT bioscreening platforms. PMID:22021162

High throughput light absorber discovery, Part 2: Establishing structure–band gap energy relationships

DOE PAGES

Suram, Santosh K.; Newhouse, Paul F.; Zhou, Lan; ...

2016-09-23

Combinatorial materials science strategies have accelerated materials development in a variety of fields, and we extend these strategies to enable structure-property mapping for light absorber materials, particularly in high order composition spaces. High throughput optical spectroscopy and synchrotron X-ray diffraction are combined to identify the optical properties of Bi-V-Fe oxides, leading to the identification of Bi 4V 1.5Fe 0.5O 10.5 as a light absorber with direct band gap near 2.7 eV. Here, the strategic combination of experimental and data analysis techniques includes automated Tauc analysis to estimate band gap energies from the high throughput spectroscopy data, providing an automated platformmore » for identifying new optical materials.« less

High throughput light absorber discovery, Part 2: Establishing structure–band gap energy relationships

DOE Office of Scientific and Technical Information (OSTI.GOV)

Suram, Santosh K.; Newhouse, Paul F.; Zhou, Lan

Combinatorial materials science strategies have accelerated materials development in a variety of fields, and we extend these strategies to enable structure-property mapping for light absorber materials, particularly in high order composition spaces. High throughput optical spectroscopy and synchrotron X-ray diffraction are combined to identify the optical properties of Bi-V-Fe oxides, leading to the identification of Bi 4V 1.5Fe 0.5O 10.5 as a light absorber with direct band gap near 2.7 eV. Here, the strategic combination of experimental and data analysis techniques includes automated Tauc analysis to estimate band gap energies from the high throughput spectroscopy data, providing an automated platformmore » for identifying new optical materials.« less

High-Throughput Assay Optimization and Statistical Interpolation of Rubella-Specific Neutralizing Antibody Titers

PubMed Central

Lambert, Nathaniel D.; Pankratz, V. Shane; Larrabee, Beth R.; Ogee-Nwankwo, Adaeze; Chen, Min-hsin; Icenogle, Joseph P.

2014-01-01

Rubella remains a social and economic burden due to the high incidence of congenital rubella syndrome (CRS) in some countries. For this reason, an accurate and efficient high-throughput measure of antibody response to vaccination is an important tool. In order to measure rubella-specific neutralizing antibodies in a large cohort of vaccinated individuals, a high-throughput immunocolorimetric system was developed. Statistical interpolation models were applied to the resulting titers to refine quantitative estimates of neutralizing antibody titers relative to the assayed neutralizing antibody dilutions. This assay, including the statistical methods developed, can be used to assess the neutralizing humoral immune response to rubella virus and may be adaptable for assessing the response to other viral vaccines and infectious agents. PMID:24391140

A hybrid MAC protocol design for energy-efficient very-high-throughput millimeter wave, wireless sensor communication networks

NASA Astrophysics Data System (ADS)

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.

High Throughput Light Absorber Discovery, Part 2: Establishing Structure-Band Gap Energy Relationships.

PubMed

Suram, Santosh K; Newhouse, Paul F; Zhou, Lan; Van Campen, Douglas G; Mehta, Apurva; Gregoire, John M

2016-11-14

Combinatorial materials science strategies have accelerated materials development in a variety of fields, and we extend these strategies to enable structure-property mapping for light absorber materials, particularly in high order composition spaces. High throughput optical spectroscopy and synchrotron X-ray diffraction are combined to identify the optical properties of Bi-V-Fe oxides, leading to the identification of Bi 4 V 1.5 Fe 0.5 O 10.5 as a light absorber with direct band gap near 2.7 eV. The strategic combination of experimental and data analysis techniques includes automated Tauc analysis to estimate band gap energies from the high throughput spectroscopy data, providing an automated platform for identifying new optical materials.

Radio-over-fiber using an optical antenna based on Rydberg states of atoms

NASA Astrophysics Data System (ADS)

Deb, A. B.; Kjærgaard, N.

2018-05-01

We provide an experimental demonstration of a direct fiber-optic link for RF transmission ("radio-over-fiber") using a sensitive optical antenna based on a rubidium vapor cell. The scheme relies on measuring the transmission of laser light at an electromagnetically induced transparency resonance that involves highly excited Rydberg states. By dressing pairs of Rydberg states using microwave fields that act as local oscillators, we encoded RF signals in the optical frequency domain. The light carrying the information is linked via a virtually lossless optical fiber to a photodetector where the signal is retrieved. We demonstrate a signal bandwidth in excess of 1 MHz limited by the available coupling laser power and atomic optical density. Our sensitive, non-metallic and readily scalable optical antenna for microwaves allows extremely low-levels of optical power (˜1 μW) throughput in the fiber-optic link. It offers a promising future platform for emerging wireless network infrastructures.

Accelerating Pathology Image Data Cross-Comparison on CPU-GPU Hybrid Systems

PubMed Central

Wang, Kaibo; Huai, Yin; Lee, Rubao; Wang, Fusheng; Zhang, Xiaodong; Saltz, Joel H.

2012-01-01

As an important application of spatial databases in pathology imaging analysis, cross-comparing the spatial boundaries of a huge amount of segmented micro-anatomic objects demands extremely data- and compute-intensive operations, requiring high throughput at an affordable cost. However, the performance of spatial database systems has not been satisfactory since their implementations of spatial operations cannot fully utilize the power of modern parallel hardware. In this paper, we provide a customized software solution that exploits GPUs and multi-core CPUs to accelerate spatial cross-comparison in a cost-effective way. Our solution consists of an efficient GPU algorithm and a pipelined system framework with task migration support. Extensive experiments with real-world data sets demonstrate the effectiveness of our solution, which improves the performance of spatial cross-comparison by over 18 times compared with a parallelized spatial database approach. PMID:23355955

HITS-CLIP yields genome-wide insights into brain alternative RNA processing

NASA Astrophysics Data System (ADS)

Licatalosi, Donny D.; Mele, Aldo; Fak, John J.; Ule, Jernej; Kayikci, Melis; Chi, Sung Wook; Clark, Tyson A.; Schweitzer, Anthony C.; Blume, John E.; Wang, Xuning; Darnell, Jennifer C.; Darnell, Robert B.

2008-11-01

Protein-RNA interactions have critical roles in all aspects of gene expression. However, applying biochemical methods to understand such interactions in living tissues has been challenging. Here we develop a genome-wide means of mapping protein-RNA binding sites in vivo, by high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP). HITS-CLIP analysis of the neuron-specific splicing factor Nova revealed extremely reproducible RNA-binding maps in multiple mouse brains. These maps provide genome-wide in vivo biochemical footprints confirming the previous prediction that the position of Nova binding determines the outcome of alternative splicing; moreover, they are sufficiently powerful to predict Nova action de novo. HITS-CLIP revealed a large number of Nova-RNA interactions in 3' untranslated regions, leading to the discovery that Nova regulates alternative polyadenylation in the brain. HITS-CLIP, therefore, provides a robust, unbiased means to identify functional protein-RNA interactions in vivo.

Rapid, High-Throughput Tracking of Bacterial Motility in 3D via Phase-Contrast Holographic Video Microscopy

PubMed Central

Cheong, Fook Chiong; Wong, Chui Ching; Gao, YunFeng; Nai, Mui Hoon; Cui, Yidan; Park, Sungsu; Kenney, Linda J.; Lim, Chwee Teck

2015-01-01

Tracking fast-swimming bacteria in three dimensions can be extremely challenging with current optical techniques and a microscopic approach that can rapidly acquire volumetric information is required. Here, we introduce phase-contrast holographic video microscopy as a solution for the simultaneous tracking of multiple fast moving cells in three dimensions. This technique uses interference patterns formed between the scattered and the incident field to infer the three-dimensional (3D) position and size of bacteria. Using this optical approach, motility dynamics of multiple bacteria in three dimensions, such as speed and turn angles, can be obtained within minutes. We demonstrated the feasibility of this method by effectively tracking multiple bacteria species, including Escherichia coli, Agrobacterium tumefaciens, and Pseudomonas aeruginosa. In addition, we combined our fast 3D imaging technique with a microfluidic device to present an example of a drug/chemical assay to study effects on bacterial motility. PMID:25762336

Controlled Chemical Patterns with ThermoChemical NanoLithography (TCNL)

NASA Astrophysics Data System (ADS)

Carroll, Keith; Giordano, Anthony; Wang, Debin; Kodali, Vamsi; King, W. P.; Marder, S. R.; Riedo, E.; Curtis, J. E.

2012-02-01

Many research areas, both fundamental and applied, rely upon the ability to organize non-trivial assemblies of molecules on surfaces. In this work, we introduce a significant extension of ThermoChemical NanoLithography (TCNL), a high throughput chemical patterning technique that uses temperature-driven chemical reactions localized near the tip of a thermal cantilever. By combining a chemical kinetics based model with experiments, we have developed a protocol for varying the concentration of surface bound molecules. The result is an unprecedented ability to fabricate extremely complex patterns comprised of varying chemical concentrations, as demonstrated by sinusoidal patterns of amine groups with varying pitches (˜5-15 μm) and the replication of Leonardo da Vinci's Mona Lisa with dimensions of ˜30 x 40 μm^2. Programmed control of the chemical reaction rate should have widespread applications for a technique which has already been shown to nanopattern various substrates including graphene nanowires, piezoelectric crystals, and optoelectronic materials.

Slow Orbit Feedback at the ALS Using Matlab

DOE Office of Scientific and Technical Information (OSTI.GOV)

Portmann, G.

1999-03-25

The third generation Advanced Light Source (ALS) produces extremely bright and finely focused photon beams using undulatory, wigglers, and bend magnets. In order to position the photon beams accurately, a slow global orbit feedback system has been developed. The dominant causes of orbit motion at the ALS are temperature variation and insertion device motion. This type of motion can be removed using slow global orbit feedback with a data rate of a few Hertz. The remaining orbit motion in the ALS is only 1-3 micron rms. Slow orbit feedback does not require high computational throughput. At the ALS, the globalmore » orbit feedback algorithm, based on the singular valued decomposition method, is coded in MATLAB and runs on a control room workstation. Using the MATLAB environment to develop, test, and run the storage ring control algorithms has proven to be a fast and efficient way to operate the ALS.« less

To Demonstrate an Integrated Solution for Plasma-Material Interfaces Compatible with an Optimized Core Plasma

NASA Astrophysics Data System (ADS)

Goldston, Robert; Brooks, Jeffrey; Hubbard, Amanda; Leonard, Anthony; Lipschultz, Bruce; Maingi, Rajesh; Ulrickson, Michael; Whyte, Dennis

2009-11-01

The plasma facing components in a Demo reactor will face much more extreme boundary plasma conditions and operating requirements than any present or planned experiment. These include 1) Power density a factor of four or more greater than in ITER, 2) Continuous operation resulting in annual energy and particle throughput 100-200 times larger than ITER, 3) Elevated surface operating temperature for efficient electricity production, 4) Tritium fuel cycle control for safety and breeding requirements, and 5) Steady state plasma confinement and control. Consistent with ReNeW Thrust 12, design options are being explored for a new moderate-scale facility to assess core-edge interaction issues and solutions. Key desired features include high power density, sufficient pulse length and duty cycle, elevated wall temperature, steady-state control of an optimized core plasma, and flexibility in changing boundary components as well as access for comprehensive measurements.

Optical nano-woodpiles: large-area metallic photonic crystals and metamaterials.

PubMed

Ibbotson, Lindsey A; Demetriadou, Angela; Croxall, Stephen; Hess, Ortwin; Baumberg, Jeremy J

2015-02-09

Metallic woodpile photonic crystals and metamaterials operating across the visible spectrum are extremely difficult to construct over large areas, because of the intricate three-dimensional nanostructures and sub-50 nm features demanded. Previous routes use electron-beam lithography or direct laser writing but widespread application is restricted by their expense and low throughput. Scalable approaches including soft lithography, colloidal self-assembly, and interference holography, produce structures limited in feature size, material durability, or geometry. By multiply stacking gold nanowire flexible gratings, we demonstrate a scalable high-fidelity approach for fabricating flexible metallic woodpile photonic crystals, with features down to 10 nm produced in bulk and at low cost. Control of stacking sequence, asymmetry, and orientation elicits great control, with visible-wavelength band-gap reflections exceeding 60%, and with strong induced chirality. Such flexible and stretchable architectures can produce metamaterials with refractive index near zero, and are easily tuned across the IR and visible ranges.

Courtney E. Payne | NREL

Science.gov Websites

improved and higher throughput methods for analysis of biomass feedstocks Agronomics-using NIR spectroscopy in-house and external client training. She has also developed improved and high-throughput methods

High-throughput and automated SAXS/USAXS experiment for industrial use at BL19B2 in SPring-8

DOE Office of Scientific and Technical Information (OSTI.GOV)

Osaka, Keiichi, E-mail: k-osaka@spring8.or.jp; Inoue, Daisuke; Sato, Masugu

A highly automated system combining a sample transfer robot with focused SR beam has been established for small-angle and ultra small-angle X-ray scattering (SAXS/USAXS) measurement at BL19B2 for industrial use of SPring-8. High-throughput data collection system can be realized by means of X-ray beam of high photon flux density concentrated by a cylindrical mirror, and a two-dimensional pixel detector PILATUS-2M. For SAXS measurement, we can obtain high-quality data within 1 minute for one exposure using this system. The sample transfer robot has a capacity of 90 samples with a large variety of shapes. The fusion of high-throughput and robotic systemmore » has enhanced the usability of SAXS/USAXS capability for industrial application.« less

Characterization of the indigenous microflora in raw and pasteurized buffalo milk during storage at refrigeration temperature by high-throughput sequencing

USDA-ARS?s Scientific Manuscript database

The effect of refrigeration on bacterial communities within raw and pasteurized buffalo milk was studied using high-throughput sequencing. High quality samples of raw buffalo milk were obtained from five dairy farms in the Guangxi province of China. A sample of each milk was pasteurized, and both r...

High throughput integrated thermal characterization with non-contact optical calorimetry

NASA Astrophysics Data System (ADS)

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.

AR-NE3A, a New Macromolecular Crystallography Beamline for Pharmaceutical Applications at the Photon Factory

NASA Astrophysics Data System (ADS)

Yamada, Yusuke; Hiraki, Masahiko; Sasajima, Kumiko; Matsugaki, Naohiro; Igarashi, Noriyuki; Amano, Yasushi; Warizaya, Masaichi; Sakashita, Hitoshi; Kikuchi, Takashi; Mori, Takeharu; Toyoshima, Akio; Kishimoto, Shunji; Wakatsuki, Soichi

2010-06-01

Recent advances in high-throughput techniques for macromolecular crystallography have highlighted the importance of structure-based drug design (SBDD), and the demand for synchrotron use by pharmaceutical researchers has increased. Thus, in collaboration with Astellas Pharma Inc., we have constructed a new high-throughput macromolecular crystallography beamline, AR-NE3A, which is dedicated to SBDD. At AR-NE3A, a photon flux up to three times higher than those at existing high-throughput beams at the Photon Factory, AR-NW12A and BL-5A, can be realized at the same sample positions. Installed in the experimental hutch are a high-precision diffractometer, fast-readout, high-gain CCD detector, and sample exchange robot capable of handling more than two hundred cryo-cooled samples stored in a Dewar. To facilitate high-throughput data collection required for pharmaceutical research, fully automated data collection and processing systems have been developed. Thus, sample exchange, centering, data collection, and data processing are automatically carried out based on the user's pre-defined schedule. Although Astellas Pharma Inc. has a priority access to AR-NE3A, the remaining beam time is allocated to general academic and other industrial users.

High-Throughput Intracellular Antimicrobial Susceptibility Testing of Legionella pneumophila.

PubMed

Chiaraviglio, Lucius; Kirby, James E

2015-12-01

Legionella pneumophila is a Gram-negative opportunistic human pathogen that causes a severe pneumonia known as Legionnaires' disease. Notably, in the human host, the organism is believed to replicate solely within an intracellular compartment, predominantly within pulmonary macrophages. Consequently, successful therapy is predicated on antimicrobials penetrating into this intracellular growth niche. However, standard antimicrobial susceptibility testing methods test solely for extracellular growth inhibition. Here, we make use of a high-throughput assay to characterize intracellular growth inhibition activity of known antimicrobials. For select antimicrobials, high-resolution dose-response analysis was then performed to characterize and compare activity levels in both macrophage infection and axenic growth assays. Results support the superiority of several classes of nonpolar antimicrobials in abrogating intracellular growth. Importantly, our assay results show excellent correlations with prior clinical observations of antimicrobial efficacy. Furthermore, we also show the applicability of high-throughput automation to two- and three-dimensional synergy testing. High-resolution isocontour isobolograms provide in vitro support for specific combination antimicrobial therapy. Taken together, findings suggest that high-throughput screening technology may be successfully applied to identify and characterize antimicrobials that target bacterial pathogens that make use of an intracellular growth niche. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

High-Throughput Intracellular Antimicrobial Susceptibility Testing of Legionella pneumophila

PubMed Central

Chiaraviglio, Lucius

2015-01-01

Legionella pneumophila is a Gram-negative opportunistic human pathogen that causes a severe pneumonia known as Legionnaires' disease. Notably, in the human host, the organism is believed to replicate solely within an intracellular compartment, predominantly within pulmonary macrophages. Consequently, successful therapy is predicated on antimicrobials penetrating into this intracellular growth niche. However, standard antimicrobial susceptibility testing methods test solely for extracellular growth inhibition. Here, we make use of a high-throughput assay to characterize intracellular growth inhibition activity of known antimicrobials. For select antimicrobials, high-resolution dose-response analysis was then performed to characterize and compare activity levels in both macrophage infection and axenic growth assays. Results support the superiority of several classes of nonpolar antimicrobials in abrogating intracellular growth. Importantly, our assay results show excellent correlations with prior clinical observations of antimicrobial efficacy. Furthermore, we also show the applicability of high-throughput automation to two- and three-dimensional synergy testing. High-resolution isocontour isobolograms provide in vitro support for specific combination antimicrobial therapy. Taken together, findings suggest that high-throughput screening technology may be successfully applied to identify and characterize antimicrobials that target bacterial pathogens that make use of an intracellular growth niche. PMID:26392509

Room-temperature serial crystallography at synchrotron X-ray sources using slowly flowing free-standing high-viscosity microstreams.

PubMed

Botha, Sabine; Nass, Karol; Barends, Thomas R M; Kabsch, Wolfgang; Latz, Beatrice; Dworkowski, Florian; Foucar, Lutz; Panepucci, Ezequiel; Wang, Meitian; Shoeman, Robert L; Schlichting, Ilme; Doak, R Bruce

2015-02-01

Recent advances in synchrotron sources, beamline optics and detectors are driving a renaissance in room-temperature data collection. The underlying impetus is the recognition that conformational differences are observed in functionally important regions of structures determined using crystals kept at ambient as opposed to cryogenic temperature during data collection. In addition, room-temperature measurements enable time-resolved studies and eliminate the need to find suitable cryoprotectants. Since radiation damage limits the high-resolution data that can be obtained from a single crystal, especially at room temperature, data are typically collected in a serial fashion using a number of crystals to spread the total dose over the entire ensemble. Several approaches have been developed over the years to efficiently exchange crystals for room-temperature data collection. These include in situ collection in trays, chips and capillary mounts. Here, the use of a slowly flowing microscopic stream for crystal delivery is demonstrated, resulting in extremely high-throughput delivery of crystals into the X-ray beam. This free-stream technology, which was originally developed for serial femtosecond crystallography at X-ray free-electron lasers, is here adapted to serial crystallography at synchrotrons. By embedding the crystals in a high-viscosity carrier stream, high-resolution room-temperature studies can be conducted at atmospheric pressure using the unattenuated X-ray beam, thus permitting the analysis of small or weakly scattering crystals. The high-viscosity extrusion injector is described, as is its use to collect high-resolution serial data from native and heavy-atom-derivatized lysozyme crystals at the Swiss Light Source using less than half a milligram of protein crystals. The room-temperature serial data allow de novo structure determination. The crystal size used in this proof-of-principle experiment was dictated by the available flux density. However, upcoming developments in beamline optics, detectors and synchrotron sources will enable the use of true microcrystals. This high-throughput, high-dose-rate methodology provides a new route to investigating the structure and dynamics of macromolecules at ambient temperature.

High-throughput sequence alignment using Graphics Processing Units

PubMed Central

Schatz, Michael C; Trapnell, Cole; Delcher, Arthur L; Varshney, Amitabh

2007-01-01

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. PMID:18070356

Quartz-Seq2: a high-throughput single-cell RNA-sequencing method that effectively uses limited sequence reads.

PubMed

Sasagawa, Yohei; Danno, Hiroki; Takada, Hitomi; Ebisawa, Masashi; Tanaka, Kaori; Hayashi, Tetsutaro; Kurisaki, Akira; Nikaido, Itoshi

2018-03-09

High-throughput single-cell RNA-seq methods assign limited unique molecular identifier (UMI) counts as gene expression values to single cells from shallow sequence reads and detect limited gene counts. We thus developed a high-throughput single-cell RNA-seq method, Quartz-Seq2, to overcome these issues. Our improvements in the reaction steps make it possible to effectively convert initial reads to UMI counts, at a rate of 30-50%, and detect more genes. To demonstrate the power of Quartz-Seq2, we analyzed approximately 10,000 transcriptomes from in vitro embryonic stem cells and an in vivo stromal vascular fraction with a limited number of reads.

Perspective: Composition–structure–property mapping in high-throughput experiments: Turning data into knowledge

DOE PAGES

Hattrick-Simpers, Jason R.; Gregoire, John M.; Kusne, A. Gilad

2016-05-26

With their ability to rapidly elucidate composition-structure-property relationships, high-throughput experimental studies have revolutionized how materials are discovered, optimized, and commercialized. It is now possible to synthesize and characterize high-throughput libraries that systematically address thousands of individual cuts of fabrication parameter space. An unresolved issue remains transforming structural characterization data into phase mappings. This difficulty is related to the complex information present in diffraction and spectroscopic data and its variation with composition and processing. Here, we review the field of automated phase diagram attribution and discuss the impact that emerging computational approaches will have in the generation of phase diagrams andmore » beyond.« less

Optimizing multi-dimensional high throughput screening using zebrafish

PubMed Central

Truong, Lisa; Bugel, Sean M.; Chlebowski, Anna; Usenko, Crystal Y.; Simonich, Michael T.; Massey Simonich, Staci L.; Tanguay, Robert L.

2016-01-01

The use of zebrafish for high throughput screening (HTS) for chemical bioactivity assessments is becoming routine in the fields of drug discovery and toxicology. Here we report current recommendations from our experiences in zebrafish HTS. We compared the effects of different high throughput chemical delivery methods on nominal water concentration, chemical sorption to multi-well polystyrene plates, transcription responses, and resulting whole animal responses. We demonstrate that digital dispensing consistently yields higher data quality and reproducibility compared to standard plastic tip-based liquid handling. Additionally, we illustrate the challenges in using this sensitive model for chemical assessment when test chemicals have trace impurities. Adaptation of these better practices for zebrafish HTS should increase reproducibility across laboratories. PMID:27453428

Combinatorial and high-throughput approaches in polymer science

NASA Astrophysics Data System (ADS)

Zhang, Huiqi; Hoogenboom, Richard; Meier, Michael A. R.; Schubert, Ulrich S.

2005-01-01

Combinatorial and high-throughput approaches have become topics of great interest in the last decade due to their potential ability to significantly increase research productivity. Recent years have witnessed a rapid extension of these approaches in many areas of the discovery of new materials including pharmaceuticals, inorganic materials, catalysts and polymers. This paper mainly highlights our progress in polymer research by using an automated parallel synthesizer, microwave synthesizer and ink-jet printer. The equipment and methodologies in our experiments, the high-throughput experimentation of different polymerizations (such as atom transfer radical polymerization, cationic ring-opening polymerization and emulsion polymerization) and the automated matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS) sample preparation are described.

Sequencing the Connectome

PubMed Central

Zador, Anthony M.; Dubnau, Joshua; Oyibo, Hassana K.; Zhan, Huiqing; Cao, Gang; Peikon, Ian D.

2012-01-01

Connectivity determines the function of neural circuits. Historically, circuit mapping has usually been viewed as a problem of microscopy, but no current method can achieve high-throughput mapping of entire circuits with single neuron precision. Here we describe a novel approach to determining connectivity. We propose BOINC (“barcoding of individual neuronal connections”), a method for converting the problem of connectivity into a form that can be read out by high-throughput DNA sequencing. The appeal of using sequencing is that its scale—sequencing billions of nucleotides per day is now routine—is a natural match to the complexity of neural circuits. An inexpensive high-throughput technique for establishing circuit connectivity at single neuron resolution could transform neuroscience research. PMID:23109909

High-Throughput Thermodynamic Modeling and Uncertainty Quantification for ICME

NASA Astrophysics Data System (ADS)

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.

Extreme ultraviolet patterned mask inspection performance of advanced projection electron microscope system for 11nm half-pitch generation

NASA Astrophysics Data System (ADS)

Hirano, Ryoichi; Iida, Susumu; Amano, Tsuyoshi; Watanabe, Hidehiro; Hatakeyama, Masahiro; Murakami, Takeshi; Suematsu, Kenichi; Terao, Kenji

2016-03-01

Novel projection electron microscope optics have been developed and integrated into a new inspection system named EBEYE-V30 ("Model EBEYE" is an EBARA's model code) , and the resulting system shows promise for application to half-pitch (hp) 16-nm node extreme ultraviolet lithography (EUVL) patterned mask inspection. To improve the system's inspection throughput for 11-nm hp generation defect detection, a new electron-sensitive area image sensor with a high-speed data processing unit, a bright and stable electron source, and an image capture area deflector that operates simultaneously with the mask scanning motion have been developed. A learning system has been used for the mask inspection tool to meet the requirements of hp 11-nm node EUV patterned mask inspection. Defects are identified by the projection electron microscope system using the "defectivity" from the characteristics of the acquired image. The learning system has been developed to reduce the labor and costs associated with adjustment of the detection capability to cope with newly-defined mask defects. We describe the integration of the developed elements into the inspection tool and the verification of the designed specification. We have also verified the effectiveness of the learning system, which shows enhanced detection capability for the hp 11-nm node.

High-throughput sample processing and sample management; the functional evolution of classical cytogenetic assay towards automation.

PubMed

Ramakumar, Adarsh; Subramanian, Uma; Prasanna, Pataje G S

2015-11-01

High-throughput individual diagnostic dose assessment is essential for medical management of radiation-exposed subjects after a mass casualty. Cytogenetic assays such as the Dicentric Chromosome Assay (DCA) are recognized as the gold standard by international regulatory authorities. DCA is a multi-step and multi-day bioassay. DCA, as described in the IAEA manual, can be used to assess dose up to 4-6 weeks post-exposure quite accurately but throughput is still a major issue and automation is very essential. The throughput is limited, both in terms of sample preparation as well as analysis of chromosome aberrations. Thus, there is a need to design and develop novel solutions that could utilize extensive laboratory automation for sample preparation, and bioinformatics approaches for chromosome-aberration analysis to overcome throughput issues. We have transitioned the bench-based cytogenetic DCA to a coherent process performing high-throughput automated biodosimetry for individual dose assessment ensuring quality control (QC) and quality assurance (QA) aspects in accordance with international harmonized protocols. A Laboratory Information Management System (LIMS) is designed, implemented and adapted to manage increased sample processing capacity, develop and maintain standard operating procedures (SOP) for robotic instruments, avoid data transcription errors during processing, and automate analysis of chromosome-aberrations using an image analysis platform. Our efforts described in this paper intend to bridge the current technological gaps and enhance the potential application of DCA for a dose-based stratification of subjects following a mass casualty. This paper describes one such potential integrated automated laboratory system and functional evolution of the classical DCA towards increasing critically needed throughput. Published by Elsevier B.V.

Development of Droplet Microfluidics Enabling High-Throughput Single-Cell Analysis.

PubMed

Wen, Na; Zhao, Zhan; Fan, Beiyuan; Chen, Deyong; Men, Dong; Wang, Junbo; Chen, Jian

2016-07-05

This article reviews recent developments in droplet microfluidics enabling high-throughput single-cell analysis. Five key aspects in this field are included in this review: (1) prototype demonstration of single-cell encapsulation in microfluidic droplets; (2) technical improvements of single-cell encapsulation in microfluidic droplets; (3) microfluidic droplets enabling single-cell proteomic analysis; (4) microfluidic droplets enabling single-cell genomic analysis; and (5) integrated microfluidic droplet systems enabling single-cell screening. We examine the advantages and limitations of each technique and discuss future research opportunities by focusing on key performances of throughput, multifunctionality, and absolute quantification.

The High-Throughput Stochastic Human Exposure and Dose Simulation Model (SHEDS-HT) & The Chemical and Products Database (CPDat)

EPA Science Inventory

The Stochastic Human Exposure and Dose Simulation Model – High-Throughput (SHEDS-HT) is a U.S. Environmental Protection Agency research tool for predicting screening-level (low-tier) exposures to chemicals in consumer products. This course will present an overview of this m...

Application of Targeted Functional Assays to Assess a Putative Vascular Disruption Developmental Toxicity Pathway Informed By ToxCast High-Throughput Screening Data

EPA Science Inventory

Chemical perturbation of vascular development is a putative toxicity pathway which may result in developmental toxicity. EPA’s high-throughput screening (HTS) ToxCast program contains assays which measure cellular signals and biological processes critical for blood vessel develop...

A Framework for Linking High-Throughput Modeling and Measurement Efforts to Advance 21st Century Exposure Science

EPA Science Inventory

The past five years have witnessed a rapid shift in the exposure science and toxicology communities towards high-throughput (HT) analyses of chemicals as potential stressors of human and ecological health. Modeling efforts have largely led the charge in the exposure science field...

Forecasting Exposure in Order to Use High Throughput Hazard Data in a Risk-based Context (WC9)

EPA Science Inventory

The ToxCast program and Tox21 consortium have evaluated over 8000 chemicals using in vitro high-throughput screening (HTS) to identify potential hazards. Complementary exposure science needed to assess risk, and the U.S. Environmental Protection Agency (EPA)’s ExpoCast initiative...

An industrial engineering approach to laboratory automation for high throughput screening

PubMed Central

Menke, Karl C.

2000-01-01

Across the pharmaceutical industry, there are a variety of approaches to laboratory automation for high throughput screening. At Sphinx Pharmaceuticals, the principles of industrial engineering have been applied to systematically identify and develop those automated solutions that provide the greatest value to the scientists engaged in lead generation. PMID:18924701

Differentiating pathway-specific from non-specific effects in high-throughput toxicity data: A foundation for prioritizing adverse outcome pathway development

EPA Science Inventory

The U.S. Environmental Protection Agency’s ToxCast program has screened thousands of chemicals for biological activity, primarily using high-throughput in vitro bioassays. Adverse outcome pathways (AOPs) offer a means to link pathway-specific biological activities with pote...

Uncertainty Quantification in High Throughput Screening: Applications to Models of Endocrine Disruption, Cytotoxicity, and Zebrafish Development (GRC Drug Safety)

EPA Science Inventory

Using uncertainty quantification, we aim to improve the quality of modeling data from high throughput screening assays for use in risk assessment. ToxCast is a large-scale screening program that analyzes thousands of chemicals using over 800 assays representing hundreds of bioche...

Differentiating pathway-based toxicity from non-specific effects in high throughput data: A foundation for prioritizing targets for AOP development.

EPA Science Inventory

The Environmental Protection Agency has implemented a high throughput screening program, ToxCast, to quickly evaluate large numbers of chemicals for their effects on hundreds of different biological targets. To understand how these measurements relate to adverse effects in an or...

Development of a high-throughput SNP resource to advance genomic, genetic and breeding research in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

The rapid advancement in high-throughput SNP genotyping technologies along with next generation sequencing (NGS) platforms has decreased the cost, improved the quality of large-scale genome surveys, and allowed specialty crops with limited genomic resources such as carrot (Daucus carota) to access t...

High-Throughput Screening and Quantitative Chemical Ranking for Sodium Iodide Symporter Inhibitors in ToxCast Phase 1 Chemical Library

EPA Science Inventory

The U.S. EPA’s Endocrine Disruptor Screening Program (EDSP) and Office of Research and Development (ORD) are currently developing high throughput assays to screen chemicals that may alter the thyroid hormone pathway. One potential target in this pathway is the sodium iodide...

High Throughput PBTK: Evaluating EPA’s Open-Source Data and Tools for Dosimetry and Exposure Reconstruction

EPA Science Inventory

Thousands of chemicals have been profiled by high-throughput screening (HTS) 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 (TK). While HTS generates in vitro bioactivity d...

Inhibition of Retinoblastoma Protein Inactivation

DTIC Science & Technology

2017-11-01

SUBJECT TERMS cell cycle, Retinoblastoma protein, E2F transcription factor, high throughput screen, drug discovery, x-ray crystallography 16. SECURITY...screening by x-ray crystallography . 2.0 KEYWORDS Retinoblastoma (Rb) pathway, E2F transcription factor, cancer, cell-cycle inhibition, activation...modulation, inhibition, high throughput screening, fragment-based screening, x-ray crystallography . 3.0 ACCOMPLISHMENTS Summary: We

High Throughput Sequence Analysis for Disease Resistance in Maize

USDA-ARS?s Scientific Manuscript database

Preliminary results of a computational analysis of high throughput sequencing data from Zea mays and the fungus Aspergillus are reported. The Illumina Genome Analyzer was used to sequence RNA samples from two strains of Z. mays (Va35 and Mp313) collected over a time course as well as several specie...

High Throughput Prioritization for Integrated Toxicity Testing Based on ToxCast Chemical Profiling

EPA Science Inventory

The rational prioritization of chemicals for integrated toxicity testing is a central goal of the U.S. EPA’s ToxCast™ program (http://epa.gov/ncct/toxcast/). ToxCast includes a wide-ranging battery of over 500 in vitro high-throughput screening assays which in Phase I was used to...

Continuing Development of Alternative High-Throughput Screens to Determine Endocrine Disruption, Focusing on Androgen Receptor, Steroidogenesis, and Thyroid Pathways

EPA Science Inventory

The focus of this meeting is the SAP's review and comment on the Agency's proposed high-throughput computational model of androgen receptor pathway activity as an alternative to the current Tier 1 androgen receptor assay (OCSPP 890.1150: Androgen Receptor Binding Rat Prostate Cyt...

Evaluation of High-throughput Genotoxicity Assays Used in Profiling the US EPA ToxCast Chemicals

EPA Science Inventory

Three high-throughput screening (HTS) genotoxicity assays-GreenScreen HC GADD45a-GFP (Gentronix Ltd.), CellCiphr p53 (Cellumen Inc.) and CellSensor p53RE-bla (Invitrogen Corp.)-were used to analyze the collection of 320 predominantly pesticide active compounds being tested in Pha...

A Biologically Informed Framework for the Analysis of the PPAR Signaling Pathway using a Bayesian Network

EPA Science Inventory

The US EPA’s ToxCastTM program seeks to combine advances in high-throughput screening technology with methodologies from statistics and computer science to develop high-throughput decision support tools for assessing chemical hazard and risk. To develop new methods of analysis of...

Collaborative Core Research Program for Chemical-Biological Warfare Defense

DTIC Science & Technology

2015-01-04

Discovery through High Throughput Screening (HTS) and Fragment-Based Drug Design (FBDD...Discovery through High Throughput Screening (HTS) and Fragment-Based Drug Design (FBDD) Current pharmaceutical approaches involving drug discovery...structural analysis and docking program generally known as fragment based drug design (FBDD). The main advantage of using these approaches is that

High-Throughput Screening and Quantitative Chemical Ranking for Sodium Iodide Symporter Inhibitors in ToxCast Phase 1 Chemical Library

EPA Science Inventory

The U.S. EPA’s Endocrine Disruptor Screening Program (EDSP) and Office of Research and Development (ORD) are currently developing high throughput assays to screen chemicals that may alter the thyroid hormone pathway. One potential target in this pathway is the sodium iodide sympo...

Integrated automation for continuous high-throughput synthetic chromosome assembly and transformation to identify improved yeast strains for industrial production of peptide sweetener brazzein

USDA-ARS?s Scientific Manuscript database

Production and recycling of recombinant sweetener peptides in industrial biorefineries involves the evaluation of large numbers of genes and proteins. High-throughput integrated robotic molecular biology platforms that have the capacity to rapidly synthesize, clone, and express heterologous gene ope...

ToxCast Assay Network (TCAN) Viewer: A Visualization Tool for High-throughput Assay Chemical Data (SOT)

EPA Science Inventory

USEPA’s ToxCast program has generated high-throughput bioactivity screening (HTS) data on thousands of chemicals. The ToxCast program has described and annotated the HTS assay battery with respect to assay design and target information (e.g., gene target). Recent stakeholder and ...

PLASMA PROTEIN PROFILING AS A HIGH THROUGHPUT TOOL FOR CHEMICAL SCREENING USING A SMALL FISH MODEL

EPA Science Inventory

Hudson, R. Tod, Michael J. Hemmer, Kimberly A. Salinas, Sherry S. Wilkinson, James Watts, James T. Winstead, Peggy S. Harris, Amy Kirkpatrick and Calvin C. Walker. In press. Plasma Protein Profiling as a High Throughput Tool for Chemical Screening Using a Small Fish Model (Abstra...

Characterization and complete genome sequence of a previously uncharacterized panicovirus from Bermuda grass detected by high throughput sequencing

USDA-ARS?s Scientific Manuscript database

Bermuda grass samples were examined by transmission electron microscopy and 28-30 nm spherical virus particles were observed. Total RNA from these plants was subjected to high throughput sequencing (HTS). The nearly full genome sequence of a previously uncharacterized Panicovirus was identified from...

Multitrait, random regression, or simple repeatability model in high-throughput phenotyping data improve genomic prediction for wheat grain yield

USDA-ARS?s Scientific Manuscript database

High-throughput phenotyping (HTP) platforms can be used to measure traits that are genetically correlated with wheat (Triticum aestivum L.) grain yield across time. Incorporating such secondary traits in the multivariate pedigree and genomic prediction models would be desirable to improve indirect s...

Handheld Fluorescence Microscopy based Flow Analyzer.

PubMed

Saxena, Manish; Jayakumar, Nitin; Gorthi, Sai Siva

2016-03-01

Fluorescence microscopy has the intrinsic advantages of favourable contrast characteristics and high degree of specificity. Consequently, it has been a mainstay in modern biological inquiry and clinical diagnostics. Despite its reliable nature, fluorescence based clinical microscopy and diagnostics is a manual, labour intensive and time consuming procedure. The article outlines a cost-effective, high throughput alternative to conventional fluorescence imaging techniques. With system level integration of custom-designed microfluidics and optics, we demonstrate fluorescence microscopy based imaging flow analyzer. Using this system we have imaged more than 2900 FITC labeled fluorescent beads per minute. This demonstrates high-throughput characteristics of our flow analyzer in comparison to conventional fluorescence microscopy. The issue of motion blur at high flow rates limits the achievable throughput in image based flow analyzers. Here we address the issue by computationally deblurring the images and show that this restores the morphological features otherwise affected by motion blur. By further optimizing concentration of the sample solution and flow speeds, along with imaging multiple channels simultaneously, the system is capable of providing throughput of about 480 beads per second.

High-Throughput, Motility-Based Sorter for Microswimmers such as C. elegans

PubMed Central

Yuan, Jinzhou; Zhou, Jessie; Raizen, David M.; Bau, Haim H.

2015-01-01

Animal motility varies with genotype, disease, aging, and environmental conditions. In many studies, it is desirable to carry out high throughput motility-based sorting to isolate rare animals for, among other things, forward genetic screens to identify genetic pathways that regulate phenotypes of interest. Many commonly used screening processes are labor-intensive, lack sensitivity, and require extensive investigator training. Here, we describe a sensitive, high throughput, automated, motility-based method for sorting nematodes. Our method is implemented in a simple microfluidic device capable of sorting thousands of animals per hour per module, and is amenable to parallelism. The device successfully enriches for known C. elegans motility mutants. Furthermore, using this device, we isolate low-abundance mutants capable of suppressing the somnogenic effects of the flp-13 gene, which regulates C. elegans sleep. By performing genetic complementation tests, we demonstrate that our motility-based sorting device efficiently isolates mutants for the same gene identified by tedious visual inspection of behavior on an agar surface. Therefore, our motility-based sorter is capable of performing high throughput gene discovery approaches to investigate fundamental biological processes. PMID:26008643

A comparison of sorptive extraction techniques coupled to a new quantitative, sensitive, high throughput GC-MS/MS method for methoxypyrazine analysis in wine.

PubMed

Hjelmeland, Anna K; Wylie, Philip L; Ebeler, Susan E

2016-02-01

Methoxypyrazines are volatile compounds found in plants, microbes, and insects that have potent vegetal and earthy aromas. With sensory detection thresholds in the low ng L(-1) range, modest concentrations of these compounds can profoundly impact the aroma quality of foods and beverages, and high levels can lead to consumer rejection. The wine industry routinely analyzes the most prevalent methoxypyrazine, 2-isobutyl-3-methoxypyrazine (IBMP), to aid in harvest decisions, since concentrations decrease during berry ripening. In addition to IBMP, three other methoxypyrazines IPMP (2-isopropyl-3-methoxypyrazine), SBMP (2-sec-butyl-3-methoxypyrazine), and EMP (2-ethyl-3-methoxypyrazine) have been identified in grapes and/or wine and can impact aroma quality. Despite their routine analysis in the wine industry (mostly IBMP), accurate methoxypyrazine quantitation is hindered by two major challenges: sensitivity and resolution. With extremely low sensory detection thresholds (~8-15 ng L(-1) in wine for IBMP), highly sensitive analytical methods to quantify methoxypyrazines at trace levels are necessary. Here we were able to achieve resolution of IBMP as well as IPMP, EMP, and SBMP from co-eluting compounds using one-dimensional chromatography coupled to positive chemical ionization tandem mass spectrometry. Three extraction techniques HS-SPME (headspace-solid phase microextraction), SBSE (stirbar sorptive extraction), and HSSE (headspace sorptive extraction) were validated and compared. A 30 min extraction time was used for HS-SPME and SBSE extraction techniques, while 120 min was necessary to achieve sufficient sensitivity for HSSE extractions. All extraction methods have limits of quantitation (LOQ) at or below 1 ng L(-1) for all four methoxypyrazines analyzed, i.e., LOQ's at or below reported sensory detection limits in wine. The method is high throughput, with resolution of all compounds possible with a relatively rapid 27 min GC oven program. Copyright © 2015 Elsevier B.V. All rights reserved.

Molecular ecological network analyses.

PubMed

Deng, Ye; Jiang, Yi-Huei; Yang, Yunfeng; He, Zhili; Luo, Feng; Zhou, Jizhong

2012-05-30

Understanding the interaction among different species within a community and their responses to environmental changes is a central goal in ecology. However, defining the network structure in a microbial community is very challenging due to their extremely high diversity and as-yet uncultivated status. Although recent advance of metagenomic technologies, such as high throughout sequencing and functional gene arrays, provide revolutionary tools for analyzing microbial community structure, it is still difficult to examine network interactions in a microbial community based on high-throughput metagenomics data. Here, we describe a novel mathematical and bioinformatics framework to construct ecological association networks named molecular ecological networks (MENs) through Random Matrix Theory (RMT)-based methods. Compared to other network construction methods, this approach is remarkable in that the network is automatically defined and robust to noise, thus providing excellent solutions to several common issues associated with high-throughput metagenomics data. We applied it to determine the network structure of microbial communities subjected to long-term experimental warming based on pyrosequencing data of 16 S rRNA genes. We showed that the constructed MENs under both warming and unwarming conditions exhibited topological features of scale free, small world and modularity, which were consistent with previously described molecular ecological networks. Eigengene analysis indicated that the eigengenes represented the module profiles relatively well. In consistency with many other studies, several major environmental traits including temperature and soil pH were found to be important in determining network interactions in the microbial communities examined. To facilitate its application by the scientific community, all these methods and statistical tools have been integrated into a comprehensive Molecular Ecological Network Analysis Pipeline (MENAP), which is open-accessible now (http://ieg2.ou.edu/MENA). The RMT-based molecular ecological network analysis provides powerful tools to elucidate network interactions in microbial communities and their responses to environmental changes, which are fundamentally important for research in microbial ecology and environmental microbiology.

Bifrost: a Modular Python/C++ Framework for Development of High-Throughput Data Analysis Pipelines

NASA Astrophysics Data System (ADS)

Cranmer, Miles; Barsdell, Benjamin R.; Price, Danny C.; Garsden, Hugh; Taylor, Gregory B.; Dowell, Jayce; Schinzel, Frank; Costa, Timothy; Greenhill, Lincoln J.

2017-01-01

Large radio interferometers have data rates that render long-term storage of raw correlator data infeasible, thus motivating development of real-time processing software. For high-throughput applications, processing pipelines are challenging to design and implement. Motivated by science efforts with the Long Wavelength Array, we have developed Bifrost, a novel Python/C++ framework that eases the development of high-throughput data analysis software by packaging algorithms as black box processes in a directed graph. This strategy to modularize code allows astronomers to create parallelism without code adjustment. Bifrost uses CPU/GPU ’circular memory’ data buffers that enable ready introduction of arbitrary functions into the processing path for ’streams’ of data, and allow pipelines to automatically reconfigure in response to astrophysical transient detection or input of new observing settings. We have deployed and tested Bifrost at the latest Long Wavelength Array station, in Sevilleta National Wildlife Refuge, NM, where it handles throughput exceeding 10 Gbps per CPU core.

A high-throughput and quantitative method to assess the mutagenic potential of translesion DNA synthesis

PubMed Central

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

High throughput imaging cytometer with acoustic focussing† †Electronic supplementary information (ESI) available: High throughput imaging cytometer with acoustic focussing. See DOI: 10.1039/c5ra19497k Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.

PubMed Central

Zmijan, Robert; Jonnalagadda, Umesh S.; Carugo, Dario; Kochi, Yu; Lemm, Elizabeth; Packham, Graham; Hill, Martyn

2015-01-01

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. PMID:29456838

A high-throughput pipeline for the production of synthetic antibodies for analysis of ribonucleoprotein complexes

PubMed Central

Na, Hong; Laver, John D.; Jeon, Jouhyun; Singh, Fateh; Ancevicius, Kristin; Fan, Yujie; Cao, Wen Xi; Nie, Kun; Yang, Zhenglin; Luo, Hua; Wang, Miranda; Rissland, Olivia; Westwood, J. Timothy; Kim, Philip M.; Smibert, Craig A.; Lipshitz, Howard D.; Sidhu, Sachdev S.

2016-01-01

Post-transcriptional regulation of mRNAs plays an essential role in the control of gene expression. mRNAs are regulated in ribonucleoprotein (RNP) complexes by RNA-binding proteins (RBPs) along with associated protein and noncoding RNA (ncRNA) cofactors. A global understanding of post-transcriptional control in any cell type requires identification of the components of all of its RNP complexes. We have previously shown that these complexes can be purified by immunoprecipitation using anti-RBP synthetic antibodies produced by phage display. To develop the large number of synthetic antibodies required for a global analysis of RNP complex composition, we have established a pipeline that combines (i) a computationally aided strategy for design of antigens located outside of annotated domains, (ii) high-throughput antigen expression and purification in Escherichia coli, and (iii) high-throughput antibody selection and screening. Using this pipeline, we have produced 279 antibodies against 61 different protein components of Drosophila melanogaster RNPs. Together with those produced in our low-throughput efforts, we have a panel of 311 antibodies for 67 RNP complex proteins. Tests of a subset of our antibodies demonstrated that 89% immunoprecipitate their endogenous target from embryo lysate. This panel of antibodies will serve as a resource for global studies of RNP complexes in Drosophila. Furthermore, our high-throughput pipeline permits efficient production of synthetic antibodies against any large set of proteins. PMID:26847261

A three-dimensional in vitro ovarian cancer coculture model using a high-throughput cell patterning platform

PubMed Central

Rizvi, Imran; Moon, Sangjun; Hasan, Tayyaba; Demirci, Utkan

2013-01-01

In vitro 3D cancer models that provide a more accurate representation of disease in vivo are urgently needed to improve our understanding of cancer pathology and to develop better cancer therapies. However, development of 3D models that are based on manual ejection of cells from micropipettes suffer from inherent limitations such as poor control over cell density, limited repeatability, low throughput, and, in the case of coculture models, lack of reproducible control over spatial distance between cell types (e.g., cancer and stromal cells). In this study, we build on a recently introduced 3D model in which human ovarian cancer (OVCAR-5) cells overlaid on Matrigel™ spontaneously form multicellular acini. We introduce a high-throughput automated cell printing system to bioprint a 3D coculture model using cancer cells and normal fibroblasts micropatterned on Matrigel™. Two cell types were patterned within a spatially controlled microenvironment (e.g., cell density, cell-cell distance) in a high-throughput and reproducible manner; both cell types remained viable during printing and continued to proliferate following patterning. This approach enables the miniaturization of an established macro-scale 3D culture model and would allow systematic investigation into the multiple unknown regulatory feedback mechanisms between tumor and stromal cells and provide a tool for high-throughput drug screening. PMID:21298805

High-throughput method for the quantitation of metabolites and co-factors from homocysteine-methionine cycle for nutritional status assessment.

PubMed

Da Silva, Laeticia; Collino, Sebastiano; Cominetti, Ornella; Martin, Francois-Pierre; Montoliu, Ivan; Moreno, Sergio Oller; Corthesy, John; Kaput, Jim; Kussmann, Martin; Monteiro, Jacqueline Pontes; Guiraud, Seu Ping

2016-09-01

There is increasing interest in the profiling and quantitation of methionine pathway metabolites for health management research. Currently, several analytical approaches are required to cover metabolites and co-factors. We report the development and the validation of a method for the simultaneous detection and quantitation of 13 metabolites in red blood cells. The method, validated in a cohort of healthy human volunteers, shows a high level of accuracy and reproducibility. This high-throughput protocol provides a robust coverage of central metabolites and co-factors in one single analysis and in a high-throughput fashion. In large-scale clinical settings, the use of such an approach will significantly advance the field of nutritional research in health and disease.

High-Throughput Quantitative Lipidomics Analysis of Nonesterified Fatty Acids in Human Plasma.

PubMed

Christinat, Nicolas; Morin-Rivron, Delphine; Masoodi, Mojgan

2016-07-01

We present a high-throughput, nontargeted lipidomics approach using liquid chromatography coupled to high-resolution mass spectrometry for quantitative analysis of nonesterified fatty acids. We applied this method to screen a wide range of fatty acids from medium-chain to very long-chain (8 to 24 carbon atoms) in human plasma samples. The method enables us to chromatographically separate branched-chain species from their straight-chain isomers as well as separate biologically important ω-3 and ω-6 polyunsaturated fatty acids. We used 51 fatty acid species to demonstrate the quantitative capability of this method with quantification limits in the nanomolar range; however, this method is not limited only to these fatty acid species. High-throughput sample preparation was developed and carried out on a robotic platform that allows extraction of 96 samples simultaneously within 3 h. This high-throughput platform was used to assess the influence of different types of human plasma collection and preparation on the nonesterified fatty acid profile of healthy donors. Use of the anticoagulants EDTA and heparin has been compared with simple clotting, and only limited changes have been detected in most nonesterified fatty acid concentrations.

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

NASA Astrophysics Data System (ADS)

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.

Theory and implementation of a very high throughput true random number generator in field programmable gate array

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yonggang, E-mail: wangyg@ustc.edu.cn; Hui, Cong; Liu, Chong

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,more » 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.« less

Theory and implementation of a very high throughput true random number generator in field programmable gate array.

PubMed

Wang, Yonggang; Hui, Cong; Liu, Chong; Xu, Chao

2016-04-01

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.

A green fluorescent protein-based assay for high-throughput ligand-binding studies of a mycobacterial biotin protein ligase.

PubMed

Bond, Thomas E H; Sorenson, Alanna E; Schaeffer, Patrick M

2017-12-01

Biotin protein ligase (BirA) has been identified as an emerging drug target in Mycobacterium tuberculosis due to its essential metabolic role. Indeed, it is the only enzyme capable of covalently attaching biotin onto the biotin carboxyl carrier protein subunit of the acetyl-CoA carboxylase. Despite recent interest in this protein, there is still a gap in cost-effective high-throughput screening assays for rapid identification of mycobacterial BirA-targeting inhibitors. We present for the first time the cloning, expression, purification of mycobacterial GFP-tagged BirA and its application for the development of a high-throughput assay building on the principle of differential scanning fluorimetry of GFP-tagged proteins. The data obtained in this study reveal how biotin and ATP significantly increase the thermal stability (ΔT m =+16.5°C) of M. tuberculosis BirA and lead to formation of a high affinity holoenzyme complex (K obs =7.7nM). The new findings and mycobacterial BirA high-throughput assay presented in this work could provide an efficient platform for future anti-tubercular drug discovery campaigns. Copyright © 2017 Elsevier GmbH. All rights reserved.

An Automated Method for High-Throughput Screening of Arabidopsis Rosette Growth in Multi-Well Plates and Its Validation in Stress Conditions.

PubMed

De Diego, Nuria; Fürst, Tomáš; Humplík, Jan F; Ugena, Lydia; Podlešáková, Kateřina; Spíchal, Lukáš

2017-01-01

High-throughput plant phenotyping platforms provide new possibilities for automated, fast scoring of several plant growth and development traits, followed over time using non-invasive sensors. Using Arabidops is as a model offers important advantages for high-throughput screening with the opportunity to extrapolate the results obtained to other crops of commercial interest. In this study we describe the development of a highly reproducible high-throughput Arabidopsis in vitro bioassay established using our OloPhen platform, suitable for analysis of rosette growth in multi-well plates. This method was successfully validated on example of multivariate analysis of Arabidopsis rosette growth in different salt concentrations and the interaction with varying nutritional composition of the growth medium. Several traits such as changes in the rosette area, relative growth rate, survival rate and homogeneity of the population are scored using fully automated RGB imaging and subsequent image analysis. The assay can be used for fast screening of the biological activity of chemical libraries, phenotypes of transgenic or recombinant inbred lines, or to search for potential quantitative trait loci. It is especially valuable for selecting genotypes or growth conditions that improve plant stress tolerance.

Inertial-ordering-assisted droplet microfluidics for high-throughput single-cell RNA-sequencing.

PubMed

Moon, Hui-Sung; Je, Kwanghwi; Min, Jae-Woong; Park, Donghyun; Han, Kyung-Yeon; Shin, Seung-Ho; Park, Woong-Yang; Yoo, Chang Eun; Kim, Shin-Hyun

2018-02-27

Single-cell RNA-seq reveals the cellular heterogeneity inherent in the population of cells, which is very important in many clinical and research applications. Recent advances in droplet microfluidics have achieved the automatic isolation, lysis, and labeling of single cells in droplet compartments without complex instrumentation. However, barcoding errors occurring in the cell encapsulation process because of the multiple-beads-in-droplet and insufficient throughput because of the low concentration of beads for avoiding multiple-beads-in-a-droplet remain important challenges for precise and efficient expression profiling of single cells. In this study, we developed a new droplet-based microfluidic platform that significantly improved the throughput while reducing barcoding errors through deterministic encapsulation of inertially ordered beads. Highly concentrated beads containing oligonucleotide barcodes were spontaneously ordered in a spiral channel by an inertial effect, which were in turn encapsulated in droplets one-by-one, while cells were simultaneously encapsulated in the droplets. The deterministic encapsulation of beads resulted in a high fraction of single-bead-in-a-droplet and rare multiple-beads-in-a-droplet although the bead concentration increased to 1000 μl -1 , which diminished barcoding errors and enabled accurate high-throughput barcoding. We successfully validated our device with single-cell RNA-seq. In addition, we found that multiple-beads-in-a-droplet, generated using a normal Drop-Seq device with a high concentration of beads, underestimated transcript numbers and overestimated cell numbers. This accurate high-throughput platform can expand the capability and practicality of Drop-Seq in single-cell analysis.

RNA isolation from mammalian cells using porous polymer monoliths: an approach for high-throughput automation.

PubMed

Chatterjee, Anirban; Mirer, Paul L; Zaldivar Santamaria, Elvira; Klapperich, Catherine; Sharon, Andre; Sauer-Budge, Alexis F

2010-06-01

The life science and healthcare communities have been redefining the importance of ribonucleic acid (RNA) through the study of small molecule RNA (in RNAi/siRNA technologies), micro RNA (in cancer research and stem cell research), and mRNA (gene expression analysis for biologic drug targets). Research in this field increasingly requires efficient and high-throughput isolation techniques for RNA. Currently, several commercial kits are available for isolating RNA from cells. Although the quality and quantity of RNA yielded from these kits is sufficiently good for many purposes, limitations exist in terms of extraction efficiency from small cell populations and the ability to automate the extraction process. Traditionally, automating a process decreases the cost and personnel time while simultaneously increasing the throughput and reproducibility. As the RNA field matures, new methods for automating its extraction, especially from low cell numbers and in high throughput, are needed to achieve these improvements. The technology presented in this article is a step toward this goal. The method is based on a solid-phase extraction technology using a porous polymer monolith (PPM). A novel cell lysis approach and a larger binding surface throughout the PPM extraction column ensure a high yield from small starting samples, increasing sensitivity and reducing indirect costs in cell culture and sample storage. The method ensures a fast and simple procedure for RNA isolation from eukaryotic cells, with a high yield both in terms of quality and quantity. The technique is amenable to automation and streamlined workflow integration, with possible miniaturization of the sample handling process making it suitable for high-throughput applications.

Genomic Tools for Evolution and Conservation in the Chimpanzee: Pan troglodytes ellioti Is a Genetically Distinct Population

PubMed Central

Myers, Simon; Hellenthal, Garrett; Nerrienet, Eric; Bontrop, Ronald E.; Freeman, Colin; Donnelly, Peter; Mundy, Nicholas I.

2012-01-01

In spite of its evolutionary significance and conservation importance, the population structure of the common chimpanzee, Pan troglodytes, is still poorly understood. An issue of particular controversy is whether the proposed fourth subspecies of chimpanzee, Pan troglodytes ellioti, from parts of Nigeria and Cameroon, is genetically distinct. Although modern high-throughput SNP genotyping has had a major impact on our understanding of human population structure and demographic history, its application to ecological, demographic, or conservation questions in non-human species has been extremely limited. Here we apply these tools to chimpanzee population structure, using ∼700 autosomal SNPs derived from chimpanzee genomic data and a further ∼100 SNPs from targeted re-sequencing. We demonstrate conclusively the existence of P. t. ellioti as a genetically distinct subgroup. We show that there is clear differentiation between the verus, troglodytes, and ellioti populations at the SNP and haplotype level, on a scale that is greater than that separating continental human populations. Further, we show that only a small set of SNPs (10–20) is needed to successfully assign individuals to these populations. Tellingly, use of only mitochondrial DNA variation to classify individuals is erroneous in 4 of 54 cases, reinforcing the dangers of basing demographic inference on a single locus and implying that the demographic history of the species is more complicated than that suggested analyses based solely on mtDNA. In this study we demonstrate the feasibility of developing economical and robust tests of individual chimpanzee origin as well as in-depth studies of population structure. These findings have important implications for conservation strategies and our understanding of the evolution of chimpanzees. They also act as a proof-of-principle for the use of cheap high-throughput genomic methods for ecological questions. PMID:22396655

Visualization of small lesions in rat cartilage by means of laboratory-based x-ray phase contrast imaging

NASA Astrophysics Data System (ADS)

Marenzana, Massimo; Hagen, Charlotte K.; Das Neves Borges, Patricia; Endrizzi, Marco; Szafraniec, Magdalena B.; Ignatyev, Konstantin; Olivo, Alessandro

2012-12-01

Being able to quantitatively assess articular cartilage in three-dimensions (3D) in small rodent animal models, with a simple laboratory set-up, would prove extremely important for the development of pre-clinical research focusing on cartilage pathologies such as osteoarthritis (OA). These models are becoming essential tools for the development of new drugs for OA, a disease affecting up to 1/3 of the population older than 50 years for which there is no cure except prosthetic surgery. However, due to limitations in imaging technology, high-throughput 3D structural imaging has not been achievable in small rodent models, thereby limiting their translational potential and their efficiency as research tools. We show that a simple laboratory system based on coded-aperture x-ray phase contrast imaging (CAXPCi) can correctly visualize the cartilage layer in slices of an excised rat tibia imaged both in air and in saline solution. Moreover, we show that small, surgically induced lesions are also correctly detected by the CAXPCi system, and we support this finding with histopathology examination. Following these successful proof-of-concept results in rat cartilage, we expect that an upgrade of the system to higher resolutions (currently underway) will enable extending the method to the imaging of mouse cartilage as well. From a technological standpoint, by showing the capability of the system to detect cartilage also in water, we demonstrate phase sensitivity comparable to other lab-based phase methods (e.g. grating interferometry). In conclusion, CAXPCi holds a strong potential for being adopted as a routine laboratory tool for non-destructive, high throughput assessment of 3D structural changes in murine articular cartilage, with a possible impact in the field similar to the revolution that conventional microCT brought into bone research.

Genomic tools for evolution and conservation in the chimpanzee: Pan troglodytes ellioti is a genetically distinct population.

PubMed

Bowden, Rory; MacFie, Tammie S; Myers, Simon; Hellenthal, Garrett; Nerrienet, Eric; Bontrop, Ronald E; Freeman, Colin; Donnelly, Peter; Mundy, Nicholas I

2012-01-01

In spite of its evolutionary significance and conservation importance, the population structure of the common chimpanzee, Pan troglodytes, is still poorly understood. An issue of particular controversy is whether the proposed fourth subspecies of chimpanzee, Pan troglodytes ellioti, from parts of Nigeria and Cameroon, is genetically distinct. Although modern high-throughput SNP genotyping has had a major impact on our understanding of human population structure and demographic history, its application to ecological, demographic, or conservation questions in non-human species has been extremely limited. Here we apply these tools to chimpanzee population structure, using ∼700 autosomal SNPs derived from chimpanzee genomic data and a further ∼100 SNPs from targeted re-sequencing. We demonstrate conclusively the existence of P. t. ellioti as a genetically distinct subgroup. We show that there is clear differentiation between the verus, troglodytes, and ellioti populations at the SNP and haplotype level, on a scale that is greater than that separating continental human populations. Further, we show that only a small set of SNPs (10-20) is needed to successfully assign individuals to these populations. Tellingly, use of only mitochondrial DNA variation to classify individuals is erroneous in 4 of 54 cases, reinforcing the dangers of basing demographic inference on a single locus and implying that the demographic history of the species is more complicated than that suggested analyses based solely on mtDNA. In this study we demonstrate the feasibility of developing economical and robust tests of individual chimpanzee origin as well as in-depth studies of population structure. These findings have important implications for conservation strategies and our understanding of the evolution of chimpanzees. They also act as a proof-of-principle for the use of cheap high-throughput genomic methods for ecological questions.

Studying Host-Pathogen Interactions In 3-D: Organotypic Models For Infectious Disease And Drug Development

NASA Technical Reports Server (NTRS)

Nickerson, Cheryl A.; Richter, Emily G.; Ott, C. Mark

2006-01-01

Representative, reproducible and high-throughput models of human cells and tissues are critical for a meaningful evaluation of host-pathogen interactions and are an essential component of the research developmental pipeline. The most informative infection models - animals, organ explants and human trials - are not suited for extensive evaluation of pathogenesis mechanisms and screening of candidate drugs. At the other extreme, more cost effective and accessible infection models such as conventional cell culture and static co-culture may not capture physiological and three-dimensional aspects of tissue biology that are important in assessing pathogenesis, and effectiveness and cytotoxicity of therapeutics. Our lab has used innovative bioengineering technology to establish biologically meaningful 3-D models of human tissues that recapitulate many aspects of the differentiated structure and function of the parental tissue in vivo, and we have applied these models to study infectious disease. We have established a variety of different 3-D models that are currently being used in infection studies - including small intestine, colon, lung, placenta, bladder, periodontal ligament, and neuronal models. Published work from our lab has shown that our 3-D models respond to infection with bacterial and viral pathogens in ways that reflect the infection process in vivo. By virtue of their physiological relevance, 3-D cell cultures may also hold significant potential as models to provide insight into the neuropathogenesis of HIV infection. Furthermore, the experimental flexibility, reproducibility, cost-efficiency, and high throughput platform afforded by these 3-D models may have important implications for the design and development of drugs with which to effectively treat neurological complications of HIV infection.

"One-sample concept" micro-combinatory for high throughput TEM of binary films.

PubMed

Sáfrán, György

2018-04-01

Phases of thin films may remarkably differ from that of bulk. Unlike to the comprehensive data files of Binary Phase Diagrams [1] available for bulk, complete phase maps for thin binary layers do not exist. This is due to both the diverse metastable, non-equilibrium or instable phases feasible in thin films and the required volume of characterization work with analytical techniques like TEM, SAED and EDS. The aim of the present work was to develop a method that remarkably facilitates the TEM study of the diverse binary phases of thin films, or the creation of phase maps. A micro-combinatorial method was worked out that enables both preparation and study of a gradient two-component film within a single TEM specimen. For a demonstration of the technique thin Mn x Al 1- x binary samples with evolving concentration from x = 0 to x = 1 have been prepared so that the transition from pure Mn to pure Al covers a 1.5 mm long track within the 3 mm diameter TEM grid. The proposed method enables the preparation and study of thin combinatorial samples including all feasible phases as a function of composition or other deposition parameters. Contrary to known "combinatorial chemistry", in which a series of different samples are deposited in one run, and investigated, one at a time, the present micro-combinatorial method produces a single specimen condensing a complete library of a binary system that can be studied, efficiently, within a single TEM session. That provides extremely high throughput for TEM characterization of composition-dependent phases, exploration of new materials, or the construction of phase diagrams of binary films. Copyright © 2018 Elsevier B.V. All rights reserved.